Population abundance of potentially pathogenic organisms in intestinal microbiome of jungle crow (Corvus macrorhynchos) shown with 16S rRNA gene-based microbial community analysis.

PubMed

Maeda, Isamu; Siddiki, Mohammad Shohel Rana; Nozawa-Takeda, Tsutomu; Tsukahara, Naoki; Tani, Yuri; Naito, Taki; Sugita, Shoei

2013-01-01

Jungle Crows (Corvus macrorhynchos) prefer human habitats because of their versatility in feeding accompanied with human food consumption. Therefore, it is important from a public health viewpoint to characterize their intestinal microbiota. However, no studies have been involved in molecular characterization of the microbiota based on huge and reliable number of data acquisition. In this study, 16S rRNA gene-based microbial community analysis coupled with the next-generation DNA sequencing techniques was applied to the taxonomic classification of intestinal microbiome for three jungle crows. Clustering of the reads into 130 operational taxonomic units showed that at least 70% of analyzed sequences for each crow were highly homologous to Eimeria sp., which belongs to the protozoan phylum Apicomplexa. The microbiotas of three crows also contained potentially pathogenic bacteria with significant percentages, such as the genera Campylobacter and Brachyspira. Thus, the profiling of a large number of 16S rRNA gene sequences in crow intestinal microbiomes revealed the high-frequency existence or vestige of potentially pathogenic microorganisms.

Characterization of a novel Lactobacillus species closely related to Lactobacillus johnsonii using a combination of molecular and comparative genomics methods

PubMed Central

2010-01-01

Background Comparative genomic hybridization (CGH) constitutes a powerful tool for identification and characterization of bacterial strains. In this study we have applied this technique for the characterization of a number of Lactobacillus strains isolated from the intestinal content of rats fed with a diet supplemented with sorbitol. Results Phylogenetic analysis based on 16S rRNA gene, recA, pheS, pyrG and tuf sequences identified five bacterial strains isolated from the intestinal content of rats as belonging to the recently described Lactobacillus taiwanensis species. DNA-DNA hybridization experiments confirmed that these five strains are distinct but closely related to Lactobacillus johnsonii and Lactobacillus gasseri. A whole genome DNA microarray designed for the probiotic L. johnsonii strain NCC533 was used for CGH analysis of L. johnsonii ATCC 33200T, L. johnsonii BL261, L. gasseri ATCC 33323T and L. taiwanensis BL263. In these experiments, the fluorescence ratio distributions obtained with L. taiwanensis and L. gasseri showed characteristic inter-species profiles. The percentage of conserved L. johnsonii NCC533 genes was about 83% in the L. johnsonii strains comparisons and decreased to 51% and 47% for L. taiwanensis and L. gasseri, respectively. These results confirmed the separate status of L. taiwanensis from L. johnsonii at the level of species, and also that L. taiwanensis is closer to L. johnsonii than L. gasseri is to L. johnsonii. Conclusion Conventional taxonomic analyses and microarray-based CGH analysis have been used for the identification and characterization of the newly species L. taiwanensis. The microarray-based CGH technology has been shown as a remarkable tool for the identification and fine discrimination between phylogenetically close species, and additionally provided insight into the adaptation of the strain L. taiwanensis BL263 to its ecological niche. PMID:20849602

Gut microbiota in MS: possible influence of immunomodulators

PubMed Central

Cantarel, Brandi L.; Waubant, Emmanuelle; Chehoud, Christel; Kuczynski, Justin; DeSantis, Todd Z.; Warrington, Janet; Venkatesan, Arun; Fraser, Claire M.; Mowry, Ellen M.

2015-01-01

Objectives Differences in gut bacteria have been described in several autoimmune disorders. In this exploratory pilot study, we compared gut bacteria in multiple sclerosis patients and healthy controls and evaluated the influence of glatiramer acetate and vitamin D treatment on the microbiota. Methods Subjects were otherwise healthy white women with or without relapsing-remitting multiple sclerosis who were vitamin D insufficient. Multiple sclerosis patients were untreated or were receiving glatiramer acetate. Subjects collected stool at baseline and after 90 days of vitamin D3 (5,000 IU/day) supplementation. The abundance of operational taxonomic units was evaluated by hybridization of 16S rRNA to a DNA microarray. Results While there was overlap of gut bacterial communities, the abundance of some operational taxonomic units, including Faecalibacterium, was lower in multiple sclerosis patients. Glatiramer acetate-treated MS subjects showed differences in community composition compared to untreated subjects, including Bacteroidaceae, Faecalibacterium, Ruminococcus, Lactobacillaceae, Clostridium, and Other Clostridiales. Compared to the other groups, untreated multiple sclerosis subjects had an increase in the Akkermansia, Faecalibacterium, and Coprococcus genera after vitamin D supplementation. Conclusions While overall bacterial communities were similar, specific operational taxonomic units differed between healthy and multiple sclerosis subjects. Glatiramer acetate and vitamin D supplementation were associated with differences or changes in the microbiota. This study was exploratory, and larger studies are needed to confirm these preliminary results. PMID:25775034

Gut microbiota in multiple sclerosis: possible influence of immunomodulators.

PubMed

Cantarel, Brandi L; Waubant, Emmanuelle; Chehoud, Christel; Kuczynski, Justin; DeSantis, Todd Z; Warrington, Janet; Venkatesan, Arun; Fraser, Claire M; Mowry, Ellen M

2015-06-01

Differences in gut bacteria have been described in several autoimmune disorders. In this exploratory pilot study, we compared gut bacteria in patients with multiple sclerosis and healthy controls and evaluated the influence of glatiramer acetate and vitamin D treatment on the microbiota. Subjects were otherwise healthy white women with or without relapsing-remitting multiple sclerosis who were vitamin D insufficient. Patients with multiple sclerosis were untreated or were receiving glatiramer acetate. Subjects collected stool at baseline and after 90 days of vitamin D3 (5000 IU/d) supplementation. The abundance of operational taxonomic units was evaluated by hybridization of 16S rRNA to a DNA microarray. While there was overlap of gut bacterial communities, the abundance of some operational taxonomic units, including Faecalibacterium, was lower in patients with multiple sclerosis. Glatiramer acetate-treated patients with multiple sclerosis showed differences in community composition compared with untreated subjects, including Bacteroidaceae, Faecalibacterium, Ruminococcus, Lactobacillaceae, Clostridium, and other Clostridiales. Compared with the other groups, untreated patients with multiple sclerosis had an increase in the Akkermansia, Faecalibacterium, and Coprococcus genera after vitamin D supplementation. While overall bacterial communities were similar, specific operational taxonomic units differed between healthy controls and patients with multiple sclerosis. Glatiramer acetate and vitamin D supplementation were associated with differences or changes in the microbiota. This study was exploratory, and larger studies are needed to confirm these preliminary results.

Anaerobic Ammonium-Oxidizing Bacteria in Cow Manure Composting.

PubMed

Wang, Tingting; Cheng, Lijun; Zhang, Wenhao; Xu, Xiuhong; Meng, Qingxin; Sun, Xuewei; Liu, Huajing; Li, Hongtao; Sun, Yu

2017-07-28

Composting is widely used to transform waste into valuable agricultural organic fertilizer. Anaerobic ammonium-oxidizing (anammox) bacteria play an important role in the global nitrogen cycle, but their role in composting remains poorly understood. In the present study, the community structure, diversity, and abundance of anammox bacteria were analyzed using cloning and sequencing methods by targeting the 16S rRNA gene and the hydrazine oxidase gene ( hzo ) in samples isolated from compost produced from cow manure and rice straw. A total of 25 operational taxonomic units were classified based on 16S rRNA gene clone libraries, and 14 operational taxonomic units were classified based on hzo gene clone libraries. The phylogenetic tree analysis of the 16S rRNA gene and deduced HZO protein sequences from the corresponding encoding genes indicated that the majority of the obtained clones were related to the known anammox bacteria Candidatus "Brocadia," Candidatus "Kuenenia," and Candidatus "Scalindua." The abundances of anammox bacteria were determined by quantitative PCR, and between 2.13 × 10 5 and 1.15 × 10 6 16S rRNA gene copies per gram of compost were found. This study provides the first demonstration of the existence of anammox bacteria with limited diversity in cow manure composting.

From learning taxonomies to phylogenetic learning: integration of 16S rRNA gene data into FAME-based bacterial classification.

PubMed

Slabbinck, Bram; Waegeman, Willem; Dawyndt, Peter; De Vos, Paul; De Baets, Bernard

2010-01-30

Machine learning techniques have shown to improve bacterial species classification based on fatty acid methyl ester (FAME) data. Nonetheless, FAME analysis has a limited resolution for discrimination of bacteria at the species level. In this paper, we approach the species classification problem from a taxonomic point of view. Such a taxonomy or tree is typically obtained by applying clustering algorithms on FAME data or on 16S rRNA gene data. The knowledge gained from the tree can then be used to evaluate FAME-based classifiers, resulting in a novel framework for bacterial species classification. In view of learning in a taxonomic framework, we consider two types of trees. First, a FAME tree is constructed with a supervised divisive clustering algorithm. Subsequently, based on 16S rRNA gene sequence analysis, phylogenetic trees are inferred by the NJ and UPGMA methods. In this second approach, the species classification problem is based on the combination of two different types of data. Herein, 16S rRNA gene sequence data is used for phylogenetic tree inference and the corresponding binary tree splits are learned based on FAME data. We call this learning approach 'phylogenetic learning'. Supervised Random Forest models are developed to train the classification tasks in a stratified cross-validation setting. In this way, better classification results are obtained for species that are typically hard to distinguish by a single or flat multi-class classification model. FAME-based bacterial species classification is successfully evaluated in a taxonomic framework. Although the proposed approach does not improve the overall accuracy compared to flat multi-class classification, it has some distinct advantages. First, it has better capabilities for distinguishing species on which flat multi-class classification fails. Secondly, the hierarchical classification structure allows to easily evaluate and visualize the resolution of FAME data for the discrimination of bacterial species. Summarized, by phylogenetic learning we are able to situate and evaluate FAME-based bacterial species classification in a more informative context.

From learning taxonomies to phylogenetic learning: Integration of 16S rRNA gene data into FAME-based bacterial classification

PubMed Central

2010-01-01

Background Machine learning techniques have shown to improve bacterial species classification based on fatty acid methyl ester (FAME) data. Nonetheless, FAME analysis has a limited resolution for discrimination of bacteria at the species level. In this paper, we approach the species classification problem from a taxonomic point of view. Such a taxonomy or tree is typically obtained by applying clustering algorithms on FAME data or on 16S rRNA gene data. The knowledge gained from the tree can then be used to evaluate FAME-based classifiers, resulting in a novel framework for bacterial species classification. Results In view of learning in a taxonomic framework, we consider two types of trees. First, a FAME tree is constructed with a supervised divisive clustering algorithm. Subsequently, based on 16S rRNA gene sequence analysis, phylogenetic trees are inferred by the NJ and UPGMA methods. In this second approach, the species classification problem is based on the combination of two different types of data. Herein, 16S rRNA gene sequence data is used for phylogenetic tree inference and the corresponding binary tree splits are learned based on FAME data. We call this learning approach 'phylogenetic learning'. Supervised Random Forest models are developed to train the classification tasks in a stratified cross-validation setting. In this way, better classification results are obtained for species that are typically hard to distinguish by a single or flat multi-class classification model. Conclusions FAME-based bacterial species classification is successfully evaluated in a taxonomic framework. Although the proposed approach does not improve the overall accuracy compared to flat multi-class classification, it has some distinct advantages. First, it has better capabilities for distinguishing species on which flat multi-class classification fails. Secondly, the hierarchical classification structure allows to easily evaluate and visualize the resolution of FAME data for the discrimination of bacterial species. Summarized, by phylogenetic learning we are able to situate and evaluate FAME-based bacterial species classification in a more informative context. PMID:20113515

5S ribosomal ribonucleic acid sequences in Bacteroides and Fusobacterium: evolutionary relationships within these genera and among eubacteria in general

NASA Technical Reports Server (NTRS)

Van den Eynde, H.; De Baere, R.; Shah, H. N.; Gharbia, S. E.; Fox, G. E.; Michalik, J.; Van de Peer, Y.; De Wachter, R.

1989-01-01

The 5S ribosomal ribonucleic acid (rRNA) sequences were determined for Bacteroides fragilis, Bacteroides thetaiotaomicron, Bacteroides capillosus, Bacteroides veroralis, Porphyromonas gingivalis, Anaerorhabdus furcosus, Fusobacterium nucleatum, Fusobacterium mortiferum, and Fusobacterium varium. A dendrogram constructed by a clustering algorithm from these sequences, which were aligned with all other hitherto known eubacterial 5S rRNA sequences, showed differences as well as similarities with respect to results derived from 16S rRNA analyses. In the 5S rRNA dendrogram, Bacteroides clustered together with Cytophaga and Fusobacterium, as in 16S rRNA analyses. Intraphylum relationships deduced from 5S rRNAs suggested that Bacteroides is specifically related to Cytophaga rather than to Fusobacterium, as was suggested by 16S rRNA analyses. Previous taxonomic considerations concerning the genus Bacteroides, based on biochemical and physiological data, were confirmed by the 5S rRNA sequence analysis.

A framework for establishing predictive relationships between specific bacterial 16S rRNA sequence abundances and biotransformation rates.

PubMed

Helbling, Damian E; Johnson, David R; Lee, Tae Kwon; Scheidegger, Andreas; Fenner, Kathrin

2015-03-01

The rates at which wastewater treatment plant (WWTP) microbial communities biotransform specific substrates can differ by orders of magnitude among WWTP communities. Differences in taxonomic compositions among WWTP communities may predict differences in the rates of some types of biotransformations. In this work, we present a novel framework for establishing predictive relationships between specific bacterial 16S rRNA sequence abundances and biotransformation rates. We selected ten WWTPs with substantial variation in their environmental and operational metrics and measured the in situ ammonia biotransformation rate constants in nine of them. We isolated total RNA from samples from each WWTP and analyzed 16S rRNA sequence reads. We then developed multivariate models between the measured abundances of specific bacterial 16S rRNA sequence reads and the ammonia biotransformation rate constants. We constructed model scenarios that systematically explored the effects of model regularization, model linearity and non-linearity, and aggregation of 16S rRNA sequences into operational taxonomic units (OTUs) as a function of sequence dissimilarity threshold (SDT). A large percentage (greater than 80%) of model scenarios resulted in well-performing and significant models at intermediate SDTs of 0.13-0.14 and 0.26. The 16S rRNA sequences consistently selected into the well-performing and significant models at those SDTs were classified as Nitrosomonas and Nitrospira groups. We then extend the framework by applying it to the biotransformation rate constants of ten micropollutants measured in batch reactors seeded with the ten WWTP communities. We identified phylogenetic groups that were robustly selected into all well-performing and significant models constructed with biotransformation rates of isoproturon, propachlor, ranitidine, and venlafaxine. These phylogenetic groups can be used as predictive biomarkers of WWTP microbial community activity towards these specific micropollutants. This work is an important step towards developing tools to predict biotransformation rates in WWTPs based on taxonomic composition. Copyright © 2014 Elsevier Ltd. All rights reserved.

Investigation of Microbial Diversity in Geothermal Hot Springs in Unkeshwar, India, Based on 16S rRNA Amplicon Metagenome Sequencing

PubMed Central

Mehetre, Gajanan T.; Paranjpe, Aditi; Dastager, Syed G.

2016-01-01

Microbial diversity in geothermal waters of the Unkeshwar hot springs in Maharashtra, India, was studied using 16S rRNA amplicon metagenomic sequencing. Taxonomic analysis revealed the presence of Bacteroidetes, Proteobacteria, Cyanobacteria, Actinobacteria, Archeae, and OD1 phyla. Metabolic function prediction analysis indicated a battery of biological information systems indicating rich and novel microbial diversity, with potential biotechnological applications in this niche. PMID:26950332

Coupling Genetic and Chemical Microbiome Profiling Reveals Heterogeneity of Archaeome and Bacteriome in Subsurface Biofilms That Are Dominated by the Same Archaeal Species

PubMed Central

Holman, Hoi-Ying N.; DeSantis, Todd Z.; Wanner, Gerhard; Andersen, Gary L.; Perras, Alexandra K.; Meck, Sandra; Völkel, Jörg; Bechtel, Hans A.; Wirth, Reinhard; Moissl-Eichinger, Christine

2014-01-01

Earth harbors an enormous portion of subsurface microbial life, whose microbiome flux across geographical locations remains mainly unexplored due to difficult access to samples. Here, we investigated the microbiome relatedness of subsurface biofilms of two sulfidic springs in southeast Germany that have similar physical and chemical parameters and are fed by one deep groundwater current. Due to their unique hydrogeological setting these springs provide accessible windows to subsurface biofilms dominated by the same uncultivated archaeal species, called SM1 Euryarchaeon. Comparative analysis of infrared imaging spectra demonstrated great variations in archaeal membrane composition between biofilms of the two springs, suggesting different SM1 euryarchaeal strains of the same species at both aquifer outlets. This strain variation was supported by ultrastructural and metagenomic analyses of the archaeal biofilms, which included intergenic spacer region sequencing of the rRNA gene operon. At 16S rRNA gene level, PhyloChip G3 DNA microarray detected similar biofilm communities for archaea, but site-specific communities for bacteria. Both biofilms showed an enrichment of different deltaproteobacterial operational taxonomic units, whose families were, however, congruent as were their lipid spectra. Consequently, the function of the major proportion of the bacteriome appeared to be conserved across the geographic locations studied, which was confirmed by dsrB-directed quantitative PCR. Consequently, microbiome differences of these subsurface biofilms exist at subtle nuances for archaea (strain level variation) and at higher taxonomic levels for predominant bacteria without a substantial perturbation in bacteriome function. The results of this communication provide deep insight into the dynamics of subsurface microbial life and warrant its future investigation with regard to metabolic and genomic analyses. PMID:24971452

The Human Oral Microbiome Database: a web accessible resource for investigating oral microbe taxonomic and genomic information

PubMed Central

Chen, Tsute; Yu, Wen-Han; Izard, Jacques; Baranova, Oxana V.; Lakshmanan, Abirami; Dewhirst, Floyd E.

2010-01-01

The human oral microbiome is the most studied human microflora, but 53% of the species have not yet been validly named and 35% remain uncultivated. The uncultivated taxa are known primarily from 16S rRNA sequence information. Sequence information tied solely to obscure isolate or clone numbers, and usually lacking accurate phylogenetic placement, is a major impediment to working with human oral microbiome data. The goal of creating the Human Oral Microbiome Database (HOMD) is to provide the scientific community with a body site-specific comprehensive database for the more than 600 prokaryote species that are present in the human oral cavity based on a curated 16S rRNA gene-based provisional naming scheme. Currently, two primary types of information are provided in HOMD—taxonomic and genomic. Named oral species and taxa identified from 16S rRNA gene sequence analysis of oral isolates and cloning studies were placed into defined 16S rRNA phylotypes and each given unique Human Oral Taxon (HOT) number. The HOT interlinks phenotypic, phylogenetic, genomic, clinical and bibliographic information for each taxon. A BLAST search tool is provided to match user 16S rRNA gene sequences to a curated, full length, 16S rRNA gene reference data set. For genomic analysis, HOMD provides comprehensive set of analysis tools and maintains frequently updated annotations for all the human oral microbial genomes that have been sequenced and publicly released. Oral bacterial genome sequences, determined as part of the Human Microbiome Project, are being added to the HOMD as they become available. We provide HOMD as a conceptual model for the presentation of microbiome data for other human body sites. Database URL: http://www.homd.org PMID:20624719

Uncultivated Microbial Eukaryotic Diversity: A Method to Link ssu rRNA Gene Sequences with Morphology

PubMed Central

Hirst, Marissa B.; Kita, Kelley N.; Dawson, Scott C.

2011-01-01

Protists have traditionally been identified by cultivation and classified taxonomically based on their cellular morphologies and behavior. In the past decade, however, many novel protist taxa have been identified using cultivation independent ssu rRNA sequence surveys. New rRNA “phylotypes” from uncultivated eukaryotes have no connection to the wealth of prior morphological descriptions of protists. To link phylogenetically informative sequences with taxonomically informative morphological descriptions, we demonstrate several methods for combining whole cell rRNA-targeted fluorescent in situ hybridization (FISH) with cytoskeletal or organellar immunostaining. Either eukaryote or ciliate-specific ssu rRNA probes were combined with an anti-α-tubulin antibody or phalloidin, a common actin stain, to define cytoskeletal features of uncultivated protists in several environmental samples. The eukaryote ssu rRNA probe was also combined with Mitotracker® or a hydrogenosomal-specific anti-Hsp70 antibody to localize mitochondria and hydrogenosomes, respectively, in uncultivated protists from different environments. Using rRNA probes in combination with immunostaining, we linked ssu rRNA phylotypes with microtubule structure to describe flagellate and ciliate morphology in three diverse environments, and linked Naegleria spp. to their amoeboid morphology using actin staining in hay infusion samples. We also linked uncultivated ciliates to morphologically similar Colpoda-like ciliates using tubulin immunostaining with a ciliate-specific rRNA probe. Combining rRNA-targeted FISH with cytoskeletal immunostaining or stains targeting specific organelles provides a fast, efficient, high throughput method for linking genetic sequences with morphological features in uncultivated protists. When linked to phylotype, morphological descriptions of protists can both complement and vet the increasing number of sequences from uncultivated protists, including those of novel lineages, identified in diverse environments. PMID:22174774

Accurate, Rapid Taxonomic Classification of Fungal Large-Subunit rRNA Genes

PubMed Central

Liu, Kuan-Liang; Porras-Alfaro, Andrea; Eichorst, Stephanie A.

2012-01-01

Taxonomic and phylogenetic fingerprinting based on sequence analysis of gene fragments from the large-subunit rRNA (LSU) gene or the internal transcribed spacer (ITS) region is becoming an integral part of fungal classification. The lack of an accurate and robust classification tool trained by a validated sequence database for taxonomic placement of fungal LSU genes is a severe limitation in taxonomic analysis of fungal isolates or large data sets obtained from environmental surveys. Using a hand-curated set of 8,506 fungal LSU gene fragments, we determined the performance characteristics of a naïve Bayesian classifier across multiple taxonomic levels and compared the classifier performance to that of a sequence similarity-based (BLASTN) approach. The naïve Bayesian classifier was computationally more rapid (>460-fold with our system) than the BLASTN approach, and it provided equal or superior classification accuracy. Classifier accuracies were compared using sequence fragments of 100 bp and 400 bp and two different PCR primer anchor points to mimic sequence read lengths commonly obtained using current high-throughput sequencing technologies. Accuracy was higher with 400-bp sequence reads than with 100-bp reads. It was also significantly affected by sequence location across the 1,400-bp test region. The highest accuracy was obtained across either the D1 or D2 variable region. The naïve Bayesian classifier provides an effective and rapid means to classify fungal LSU sequences from large environmental surveys. The training set and tool are publicly available through the Ribosomal Database Project (http://rdp.cme.msu.edu/classifier/classifier.jsp). PMID:22194300

Investigation of Microbial Diversity in Geothermal Hot Springs in Unkeshwar, India, Based on 16S rRNA Amplicon Metagenome Sequencing.

PubMed

Mehetre, Gajanan T; Paranjpe, Aditi; Dastager, Syed G; Dharne, Mahesh S

2016-02-25

Microbial diversity in geothermal waters of the Unkeshwar hot springs in Maharashtra, India, was studied using 16S rRNA amplicon metagenomic sequencing. Taxonomic analysis revealed the presence of Bacteroidetes, Proteobacteria, Cyanobacteria, Actinobacteria, Archeae, and OD1 phyla. Metabolic function prediction analysis indicated a battery of biological information systems indicating rich and novel microbial diversity, with potential biotechnological applications in this niche. Copyright © 2016 Mehetre et al.

Phylogenetic screening of a bacterial, metagenomic library using homing endonuclease restriction and marker insertion

PubMed Central

Yung, Pui Yi; Burke, Catherine; Lewis, Matt; Egan, Suhelen; Kjelleberg, Staffan; Thomas, Torsten

2009-01-01

Metagenomics provides access to the uncultured majority of the microbial world. The approaches employed in this field have, however, had limited success in linking functional genes to the taxonomic or phylogenetic origin of the organism they belong to. Here we present an efficient strategy to recover environmental DNA fragments that contain phylogenetic marker genes from metagenomic libraries. Our method involves the cleavage of 23S ribsosmal RNA (rRNA) genes within pooled library clones by the homing endonuclease I-CeuI followed by the insertion and selection of an antibiotic resistance cassette. This approach was applied to screen a library of 6500 fosmid clones derived from the microbial community associated with the sponge Cymbastela concentrica. Several fosmid clones were recovered after the screen and detailed phylogenetic and taxonomic assignment based on the rRNA gene showed that they belong to previously unknown organisms. In addition, compositional features of these fosmid clones were used to classify and taxonomically assign a dataset of environmental shotgun sequences. Our approach represents a valuable tool for the analysis of rapidly increasing, environmental DNA sequencing information. PMID:19767618

Taxonomic evaluation of Streptomyces hirsutus and related species using multi-locus sequence analysis

USDA-ARS?s Scientific Manuscript database

Phylogenetic analyses of species of Streptomyces based on 16S rRNA gene sequences resulted in a statistically well-supported clade (100% bootstrap value) containing 8 species having very similar gross morphology. These species, including Streptomyces bambergiensis, Streptomyces chlorus, Streptomyces...

Bacillus Strains Most Closely Related to Bacillus nealsonii Are Not Effectively Circumscribed within the Taxonomic Species Definition

PubMed Central

Peak, K. Kealy; Duncan, Kathleen E.; Luna, Vicki A.; King, Debra S.; McCarthy, Peter J.; Cannons, Andrew C.

2011-01-01

Bacillus strains with >99.7% 16S rRNA gene sequence similarity were characterized with DNA:DNA hybridization, cellular fatty acid (CFA) analysis, and testing of 100 phenotypic traits. When paired with the most closely related type strain, percent DNA:DNA similarities (% S) for six Bacillus strains were all far below the recommended 70% threshold value for species circumscription with Bacillus nealsonii. An apparent genomic group of four Bacillus strain pairings with 94%–70% S was contradicted by the failure of the strains to cluster in CFA- and phenotype-based dendrograms as well as by their differentiation with 9–13 species level discriminators such as nitrate reduction, temperature range, and acid production from carbohydrates. The novel Bacillus strains were monophyletic and very closely related based on 16S rRNA gene sequence. Coherent genomic groups were not however supported by similarly organized phenotypic clusters. Therefore, the strains were not effectively circumscribed within the taxonomic species definition. PMID:22046187

Microbial community profiling of fresh basil and pitfalls in taxonomic assignment of enterobacterial pathogenic species based upon 16S rRNA amplicon sequencing.

PubMed

Ceuppens, Siele; De Coninck, Dieter; Bottledoorn, Nadine; Van Nieuwerburgh, Filip; Uyttendaele, Mieke

2017-09-18

Application of 16S rRNA (gene) amplicon sequencing on food samples is increasingly applied for assessing microbial diversity but may as unintended advantage also enable simultaneous detection of any human pathogens without a priori definition. In the present study high-throughput next-generation sequencing (NGS) of the V1-V2-V3 regions of the 16S rRNA gene was applied to identify the bacteria present on fresh basil leaves. However, results were strongly impacted by variations in the bioinformatics analysis pipelines (MEGAN, SILVAngs, QIIME and MG-RAST), including the database choice (Greengenes, RDP and M5RNA) and the annotation algorithm (best hit, representative hit and lowest common ancestor). The use of pipelines with default parameters will lead to discrepancies. The estimate of microbial diversity of fresh basil using 16S rRNA (gene) amplicon sequencing is thus indicative but subject to biases. Salmonella enterica was detected at low frequencies, between 0.1% and 0.4% of bacterial sequences, corresponding with 37 to 166 reads. However, this result was dependent upon the pipeline used: Salmonella was detected by MEGAN, SILVAngs and MG-RAST, but not by QIIME. Confirmation of Salmonella sequences by real-time PCR was unsuccessful. It was shown that taxonomic resolution obtained from the short (500bp) sequence reads of the 16S rRNA gene containing the hypervariable regions V1-V3 cannot allow distinction of Salmonella with closely related enterobacterial species. In conclusion 16S amplicon sequencing, getting the status of standard method in microbial ecology studies of foods, needs expertise on both bioinformatics and microbiology for analysis of results. It is a powerful tool to estimate bacterial diversity but amenable to biases. Limitations concerning taxonomic resolution for some bacterial species or its inability to detect sub-dominant (pathogenic) species should be acknowledged in order to avoid overinterpretation of results. Copyright © 2017 Elsevier B.V. All rights reserved.

Genomic insights into the taxonomic status of the Bacillus cereus group

PubMed Central

Liu, Yang; Lai, Qiliang; Göker, Markus; Meier-Kolthoff, Jan P.; Wang, Meng; Sun, Yamin; Wang, Lei; Shao, Zongze

2015-01-01

The identification and phylogenetic relationships of bacteria within the Bacillus cereus group are controversial. This study aimed at determining the taxonomic affiliations of these strains using the whole-genome sequence-based Genome BLAST Distance Phylogeny (GBDP) approach. The GBDP analysis clearly separated 224 strains into 30 clusters, representing eleven known, partially merged species and accordingly 19–20 putative novel species. Additionally, 16S rRNA gene analysis, a novel variant of multi-locus sequence analysis (nMLSA) and screening of virulence genes were performed. The 16S rRNA gene sequence was not sufficient to differentiate the bacteria within this group due to its high conservation. The nMLSA results were consistent with GBDP. Moreover, a fast typing method was proposed using the pycA gene, and where necessary, the ccpA gene. The pXO plasmids and cry genes were widely distributed, suggesting little correlation with the phylogenetic positions of the host bacteria. This might explain why classifications based on virulence characteristics proved unsatisfactory in the past. In summary, this is the first large-scale and systematic study of the taxonomic status of the bacteria within the B. cereus group using whole-genome sequences, and is likely to contribute to further insights into their pathogenicity, phylogeny and adaptation to diverse environments. PMID:26373441

Microbial community analysis of an Alabama coastal salt marsh impacted by the Deepwater Horizon Oil Spill

NASA Astrophysics Data System (ADS)

Beazley, M. J.; Martinez, R.; Rajan, S.; Powell, J.; Piceno, Y.; Tom, L.; Andersen, G. L.; Hazen, T. C.; Van Nostrand, J. D.; Zhou, J.; Mortazavi, B.; Sobecky, P. A.

2011-12-01

Microbial community responses of an Alabama coastal salt marsh environment to the Deepwater Horizon oil spill were studied by 16S rRNA (PhyloChip) and functional gene (GeoChip) microarray-based analysis. Oil and tar balls associated with the oil spill arrived along the Alabama coast in June 2010. Marsh and inlet sediment samples collected in June, July, and September 2010 from a salt marsh ecosystem at Point Aux Pines Alabama were analyzed to determine if bacterial community structure changed as a result of oil perturbation. Sediment total petroleum hydrocarbon (TPH) concentrations ranged from below detection to 189 mg kg-1 and were randomly dispersed throughout the salt marsh sediments. Total DNA extracted from sediment and particulates were used for PhyloChip and GeoChip hybridization. A total of 4000 to 8000 operational taxonomic units (OTUs) were detected in marsh and inlet samples. Distinctive changes in the number of detectable OTUs were observed between June, July, and September 2010. Surficial inlet sediments demonstrated a significant increase in the total number of OTUs between June and September that correlated with TPH concentrations. The most significant increases in bacterial abundance were observed in the phyla Actinobacteria, Firmicutes, Gemmatimonadetes, Proteobacteria, and Verrucomicrobia. Bacterial richness in marsh sediments also correlated with TPH concentrations with significant changes primarily in Acidobacteria, Actinobacteria, Firmicutes, Fusobacteria, Nitrospirae, and Proteobacteria. GeoChip microarray analysis detected 5000 to 8300 functional genes in marsh and inlet samples. Surficial inlet sediments demonstrated distinctive increases in the number of detectable genes and gene signal intensities in July samples compared to June. Signal intensities increased (> 1.5-fold) in genes associated with petroleum degradation. Genes related to metal resistance, stress, and carbon cycling also demonstrated increases in oiled sediments. This study demonstrates the value of applying phylogenetic and functional gene microarray technology to characterize the extensive microbial diversity of marsh environments. Moreover, this technology provides significant insight into bacterial community responses to anthropogenic oil events.

Nodeomics: Pathogen Detection in Vertebrate Lymph Nodes Using Meta-Transcriptomics

USGS Publications Warehouse

Wittekindt, Nicola E.; Padhi, Abinash; Schuster, Stephan C.; Qi, Ji; Zhao, Fangqing; Tomsho, Lynn P.; Kasson, Lindsay R.; Packard, Michael; Cross, Paul C.; Poss, Mary

2010-01-01

The ongoing emergence of human infections originating from wildlife highlights the need for better knowledge of the microbial community in wildlife species where traditional diagnostic approaches are limited. Here we evaluate the microbial biota in healthy mule deer (Odocoileus hemionus) by analyses of lymph node meta-transcriptomes. cDNA libraries from five individuals and two pools of samples were prepared from retropharyngeal lymph node RNA enriched for polyadenylated RNA and sequenced using Roche-454 Life Sciences technology. Protein-coding and 16S ribosomal RNA (rRNA) sequences were taxonomically profiled using protein and rRNA specific databases. Representatives of all bacterial phyla were detected in the seven libraries based on protein-coding transcripts indicating that viable microbiota were present in lymph nodes. Residents of skin and rumen, and those ubiquitous in mule deer habitat dominated classifiable bacterial species. Based on detection of both rRNA and protein-coding transcripts, we identified two new proteobacterial species; a Helicobacter closely related to Helicobacter cetorum in the Helicobacter pylori/Helicobacter acinonychis complex and an Acinetobacter related to Acinetobacter schindleri. Among viruses, a novel gamma retrovirus and other members of the Poxviridae and Retroviridae were identified. We additionally evaluated bacterial diversity by amplicon sequencing the hypervariable V6 region of 16S rRNA and demonstrate that overall taxonomic diversity is higher with the meta-transcriptomic approach. These data provide the most complete picture to date of the microbial diversity within a wildlife host. Our research advances the use of meta-transcriptomics to study microbiota in wildlife tissues, which will facilitate detection of novel organisms with pathogenic potential to human and animals.

Pyrosequencing of mcrA and Archaeal 16S rRNA Genes Reveals Diversity and Substrate Preferences of Methanogen Communities in Anaerobic Digesters

PubMed Central

Wilkins, David; Lu, Xiao-Ying; Shen, Zhiyong; Chen, Jiapeng

2014-01-01

Methanogenic archaea play a key role in biogas-producing anaerobic digestion and yet remain poorly taxonomically characterized. This is in part due to the limitations of low-throughput Sanger sequencing of a single (16S rRNA) gene, which in the past may have undersampled methanogen diversity. In this study, archaeal communities from three sludge digesters in Hong Kong and one wastewater digester in China were examined using high-throughput pyrosequencing of the methyl coenzyme M reductase (mcrA) and 16S rRNA genes. Methanobacteriales, Methanomicrobiales, and Methanosarcinales were detected in each digester, indicating that both hydrogenotrophic and acetoclastic methanogenesis was occurring. Two sludge digesters had similar community structures, likely due to their similar design and feedstock. Taxonomic classification of the mcrA genes suggested that these digesters were dominated by acetoclastic methanogens, particularly Methanosarcinales, while the other digesters were dominated by hydrogenotrophic Methanomicrobiales. The proposed euryarchaeotal order Methanomassiliicoccales and the uncultured WSA2 group were detected with the 16S rRNA gene, and potential mcrA genes for these groups were identified. 16S rRNA gene sequencing also recovered several crenarchaeotal groups potentially involved in the initial anaerobic digestion processes. Overall, the two genes produced different taxonomic profiles for the digesters, while greater methanogen richness was detected using the mcrA gene, supporting the use of this functional gene as a complement to the 16S rRNA gene to better assess methanogen diversity. A significant positive correlation was detected between methane production and the abundance of mcrA transcripts in digesters treating sludge and wastewater samples, supporting the mcrA gene as a biomarker for methane yield. PMID:25381241

Integrative analysis of environmental sequences using MEGAN4.

PubMed

Huson, Daniel H; Mitra, Suparna; Ruscheweyh, Hans-Joachim; Weber, Nico; Schuster, Stephan C

2011-09-01

A major challenge in the analysis of environmental sequences is data integration. The question is how to analyze different types of data in a unified approach, addressing both the taxonomic and functional aspects. To facilitate such analyses, we have substantially extended MEGAN, a widely used taxonomic analysis program. The new program, MEGAN4, provides an integrated approach to the taxonomic and functional analysis of metagenomic, metatranscriptomic, metaproteomic, and rRNA data. While taxonomic analysis is performed based on the NCBI taxonomy, functional analysis is performed using the SEED classification of subsystems and functional roles or the KEGG classification of pathways and enzymes. A number of examples illustrate how such analyses can be performed, and show that one can also import and compare classification results obtained using others' tools. MEGAN4 is freely available for academic purposes, and installers for all three major operating systems can be downloaded from www-ab.informatik.uni-tuebingen.de/software/megan.

Studying modification of aminoglycoside antibiotics by resistance-causing enzymes via microarray.

PubMed

Disney, Matthew D

2012-01-01

Widespread bacterial resistance to antibiotics is a significant public health concern. To remain a step ahead of evolving bacteria, new methods to study resistance to antibacterials and to uncover novel antibiotics that evade resistance are urgently needed. Herein, microarray-based methods that have been developed to study aminoglycoside modification by resistance-causing enzymes are reviewed. These arrays can also be used to study the binding of aminoglycoside antibiotics to a mimic of their therapeutic target, the rRNA aminoacyl site (A-site), and how modification by resistance-causing enzymes affects their abilities to bind RNA.

PanFP: Pangenome-based functional profiles for microbial communities

DOE PAGES

Jun, Se -Ran; Hauser, Loren John; Schadt, Christopher Warren; ...

2015-09-26

For decades there has been increasing interest in understanding the relationships between microbial communities and ecosystem functions. Current DNA sequencing technologies allows for the exploration of microbial communities in two principle ways: targeted rRNA gene surveys and shotgun metagenomics. For large study designs, it is often still prohibitively expensive to sequence metagenomes at both the breadth and depth necessary to statistically capture the true functional diversity of a community. Although rRNA gene surveys provide no direct evidence of function, they do provide a reasonable estimation of microbial diversity, while being a very cost effective way to screen samples of interestmore » for later shotgun metagenomic analyses. However, there is a great deal of 16S rRNA gene survey data currently available from diverse environments, and thus a need for tools to infer functional composition of environmental samples based on 16S rRNA gene survey data. As a result, we present a computational method called pangenome based functional profiles (PanFP), which infers functional profiles of microbial communities from 16S rRNA gene survey data for Bacteria and Archaea. PanFP is based on pangenome reconstruction of a 16S rRNA gene operational taxonomic unit (OTU) from known genes and genomes pooled from the OTU s taxonomic lineage. From this lineage, we derive an OTU functional profile by weighting a pangenome s functional profile with the OTUs abundance observed in a given sample. We validated our method by comparing PanFP to the functional profiles obtained from the direct shotgun metagenomic measurement of 65 diverse communities via Spearman correlation coefficients. These correlations improved with increasing sequencing depth, within the range of 0.8 0.9 for the most deeply sequenced Human Microbiome Project mock community samples. PanFP is very similar in performance to another recently released tool, PICRUSt, for almost all of survey data analysed here. But, our method is unique in that any OTU building method can be used, as opposed to being limited to closed reference OTU picking strategies against specific reference sequence databases. In conclusion, we developed an automated computational method, which derives an inferred functional profile based on the 16S rRNA gene surveys of microbial communities. The inferred functional profile provides a cost effective way to study complex ecosystems through predicted comparative functional metagenomes and metadata analysis. All PanFP source code and additional documentation are freely available online at GitHub.« less

PanFP: pangenome-based functional profiles for microbial communities.

PubMed

Jun, Se-Ran; Robeson, Michael S; Hauser, Loren J; Schadt, Christopher W; Gorin, Andrey A

2015-09-26

For decades there has been increasing interest in understanding the relationships between microbial communities and ecosystem functions. Current DNA sequencing technologies allows for the exploration of microbial communities in two principle ways: targeted rRNA gene surveys and shotgun metagenomics. For large study designs, it is often still prohibitively expensive to sequence metagenomes at both the breadth and depth necessary to statistically capture the true functional diversity of a community. Although rRNA gene surveys provide no direct evidence of function, they do provide a reasonable estimation of microbial diversity, while being a very cost-effective way to screen samples of interest for later shotgun metagenomic analyses. However, there is a great deal of 16S rRNA gene survey data currently available from diverse environments, and thus a need for tools to infer functional composition of environmental samples based on 16S rRNA gene survey data. We present a computational method called pangenome-based functional profiles (PanFP), which infers functional profiles of microbial communities from 16S rRNA gene survey data for Bacteria and Archaea. PanFP is based on pangenome reconstruction of a 16S rRNA gene operational taxonomic unit (OTU) from known genes and genomes pooled from the OTU's taxonomic lineage. From this lineage, we derive an OTU functional profile by weighting a pangenome's functional profile with the OTUs abundance observed in a given sample. We validated our method by comparing PanFP to the functional profiles obtained from the direct shotgun metagenomic measurement of 65 diverse communities via Spearman correlation coefficients. These correlations improved with increasing sequencing depth, within the range of 0.8-0.9 for the most deeply sequenced Human Microbiome Project mock community samples. PanFP is very similar in performance to another recently released tool, PICRUSt, for almost all of survey data analysed here. But, our method is unique in that any OTU building method can be used, as opposed to being limited to closed-reference OTU picking strategies against specific reference sequence databases. We developed an automated computational method, which derives an inferred functional profile based on the 16S rRNA gene surveys of microbial communities. The inferred functional profile provides a cost effective way to study complex ecosystems through predicted comparative functional metagenomes and metadata analysis. All PanFP source code and additional documentation are freely available online at GitHub ( https://github.com/srjun/PanFP ).

Taxonomic evaluation of species in the Streptomyces hirsutus clade using multi-locus sequence analysis and proposals to reclassify several species in this clade

USDA-ARS?s Scientific Manuscript database

Previous phylogenetic analyses of species of Streptomyces based on 16S rRNA gene sequences resulted in a statistically well-supported clade (100% bootstrap value) containing 8 species that exhibited very similar gross morphology in producing open looped (Retinaculum-Apertum) to spiral (Spira) chains...

Oligonucleotide fingerprinting of rRNA genes for analysis of fungal community composition.

PubMed

Valinsky, Lea; Della Vedova, Gianluca; Jiang, Tao; Borneman, James

2002-12-01

Thorough assessments of fungal diversity are currently hindered by technological limitations. Here we describe a new method for identifying fungi, oligonucleotide fingerprinting of rRNA genes (OFRG). ORFG sorts arrayed rRNA gene (ribosomal DNA [rDNA]) clones into taxonomic clusters through a series of hybridization experiments, each using a single oligonucleotide probe. A simulated annealing algorithm was used to design an OFRG probe set for fungal rDNA. Analysis of 1,536 fungal rDNA clones derived from soil generated 455 clusters. A pairwise sequence analysis showed that clones with average sequence identities of 99.2% were grouped into the same cluster. To examine the accuracy of the taxonomic identities produced by this OFRG experiment, we determined the nucleotide sequences for 117 clones distributed throughout the tree. For all but two of these clones, the taxonomic identities generated by this OFRG experiment were consistent with those generated by a nucleotide sequence analysis. Eighty-eight percent of the clones were affiliated with Ascomycota, while 12% belonged to BASIDIOMYCOTA: A large fraction of the clones were affiliated with the genera Fusarium (404 clones) and Raciborskiomyces (176 clones). Smaller assemblages of clones had high sequence identities to the Alternaria, Ascobolus, Chaetomium, Cryptococcus, and Rhizoctonia clades.

A Comparative Study: Taxonomic Grouping of Alkaline Protease Producing Bacilli.

PubMed

Tekin, Nilgun; Cihan, Arzu Coleri; Karaca, Basar; Cokmus, Cumhur

2017-03-30

Alkaline proteases have biotechnological importance due to their activity and stability at alkaline pH. 56 bacteria, capable of growing under alkaline conditions were isolated and their alkaline protease activities were carried out at different parameters to determine their optimum alkaline protease production conditions. Seven isolates were showed higher alkaline protease production capacity than the reference strains. The highest alkaline protease producing isolates (103125 U/g), E114 and C265, were identified as Bacillus licheniformis with 99.4% and Bacillus mojavensis 99.8% based on 16S rRNA gene sequence similarities, respectively. Interestingly, the isolates identified as Bacillus safensis were also found to be high alkaline protease producing strains. Genotypic characterizations of the isolates were also determined by using a wide range of molecular techniques (ARDRA, ITS-PCR, (GTG)5-PCR, BOX-PCR). These different techniques allowed us to differentiate the alkaliphilic isolates and the results were in concurrence with phylogenetic analyses of the 16S rRNA genes. While ITS-PCR provided the highest correlation with 16S rRNA groups, (GTG)5-PCR showed the highest differentiation at species and intra-species level. In this study, each of the biotechnologically valuable alkaline protease producing isolates was grouped into their taxonomic positions with multi-genotypic analyses.

Streptococcus bovimastitidis sp. nov., isolated from a dairy cow with mastitis.

PubMed

de Vries, Stefan P W; Hadjirin, Nazreen F; Lay, Elizabeth M; Zadoks, Ruth N; Peacock, Sharon J; Parkhill, Julian; Grant, Andrew J; McDougall, Scott; Holmes, Mark A

2018-01-01

Here we describe a new species of the genus Streptococcus that was isolated from a dairy cow with mastitis in New Zealand. Strain NZ1587 T was Gram-positive, coccus-shaped and arranged as chains, catalase and coagulase negative, γ-haemolytic and negative for Lancefield carbohydrates (A-D, F and G). The 16S rRNA sequence did not match sequences in the NCBI 16S rRNA or GreenGenes databases. Taxonomic classification of strain NZ1587 T was investigated using 16S rRNA and core genome phylogeny, genome-wide average nucleotide identity (ANI) and predicted DNA-DNA hybridisation (DDH) analyses. Phylogeny based on 16S rRNA was unable to resolve the taxonomic position of strain NZ1587 T , however NZ1587 T shared 99.4 % identity at the 16S rRNA level with a distinct branch of S. pseudoporcinus. Importantly, core genome phylogeny demonstrated that NZ1587 T grouped amongst the 'pyogenic' streptococcal species and formed a distinct branch supported by a 100 % bootstrap value. In addition, average nucleotide identity and inferred DNA-DNA hybridisation analyses showed that NZ1587 T represents a novel species. Biochemical profiling using the rapid ID 32 strep identification test enabled differentiation of strain NZ1587 T from closely related streptococcal species. In conclusion, strain NZ1587 T can be classified as a novel species, and we propose a novel taxon named Streptococcus bovimastitidis sp. nov.; the type strain is NZ1587 T . NZ1587 T has been deposited in the Culture Collection University of Gothenburg (CCUG 69277 T ) and the Belgian Co-ordinated Collections of Micro-organisms/LMG (LMG 29747).

Optimization of single-base-pair mismatch discrimination in oligonucleotide microarrays

NASA Technical Reports Server (NTRS)

Urakawa, Hidetoshi; El Fantroussi, Said; Smidt, Hauke; Smoot, James C.; Tribou, Erik H.; Kelly, John J.; Noble, Peter A.; Stahl, David A.

2003-01-01

The discrimination between perfect-match and single-base-pair-mismatched nucleic acid duplexes was investigated by using oligonucleotide DNA microarrays and nonequilibrium dissociation rates (melting profiles). DNA and RNA versions of two synthetic targets corresponding to the 16S rRNA sequences of Staphylococcus epidermidis (38 nucleotides) and Nitrosomonas eutropha (39 nucleotides) were hybridized to perfect-match probes (18-mer and 19-mer) and to a set of probes having all possible single-base-pair mismatches. The melting profiles of all probe-target duplexes were determined in parallel by using an imposed temperature step gradient. We derived an optimum wash temperature for each probe and target by using a simple formula to calculate a discrimination index for each temperature of the step gradient. This optimum corresponded to the output of an independent analysis using a customized neural network program. These results together provide an experimental and analytical framework for optimizing mismatch discrimination among all probes on a DNA microarray.

Taxonomic evaluation of Streptomyces albus and related species using multilocus sequence analysis

USDA-ARS?s Scientific Manuscript database

In phylogenetic analyses of the genus Streptomyces using 16S rRNA gene sequences, Streptomyces albus subsp. albus NRRL B-1811T formed a cluster with 5 other species having identical or nearly identical 16S rRNA gene sequences. Moreover, the morphological and physiological characteristics of these ot...

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jun, Se -Ran; Hauser, Loren John; Schadt, Christopher Warren

For decades there has been increasing interest in understanding the relationships between microbial communities and ecosystem functions. Current DNA sequencing technologies allows for the exploration of microbial communities in two principle ways: targeted rRNA gene surveys and shotgun metagenomics. For large study designs, it is often still prohibitively expensive to sequence metagenomes at both the breadth and depth necessary to statistically capture the true functional diversity of a community. Although rRNA gene surveys provide no direct evidence of function, they do provide a reasonable estimation of microbial diversity, while being a very cost effective way to screen samples of interestmore » for later shotgun metagenomic analyses. However, there is a great deal of 16S rRNA gene survey data currently available from diverse environments, and thus a need for tools to infer functional composition of environmental samples based on 16S rRNA gene survey data. As a result, we present a computational method called pangenome based functional profiles (PanFP), which infers functional profiles of microbial communities from 16S rRNA gene survey data for Bacteria and Archaea. PanFP is based on pangenome reconstruction of a 16S rRNA gene operational taxonomic unit (OTU) from known genes and genomes pooled from the OTU s taxonomic lineage. From this lineage, we derive an OTU functional profile by weighting a pangenome s functional profile with the OTUs abundance observed in a given sample. We validated our method by comparing PanFP to the functional profiles obtained from the direct shotgun metagenomic measurement of 65 diverse communities via Spearman correlation coefficients. These correlations improved with increasing sequencing depth, within the range of 0.8 0.9 for the most deeply sequenced Human Microbiome Project mock community samples. PanFP is very similar in performance to another recently released tool, PICRUSt, for almost all of survey data analysed here. But, our method is unique in that any OTU building method can be used, as opposed to being limited to closed reference OTU picking strategies against specific reference sequence databases. In conclusion, we developed an automated computational method, which derives an inferred functional profile based on the 16S rRNA gene surveys of microbial communities. The inferred functional profile provides a cost effective way to study complex ecosystems through predicted comparative functional metagenomes and metadata analysis. All PanFP source code and additional documentation are freely available online at GitHub.« less

Broad spectrum microarray for fingerprint-based bacterial species identification

PubMed Central

2010-01-01

Background Microarrays are powerful tools for DNA-based molecular diagnostics and identification of pathogens. Most target a limited range of organisms and are based on only one or a very few genes for specific identification. Such microarrays are limited to organisms for which specific probes are available, and often have difficulty discriminating closely related taxa. We have developed an alternative broad-spectrum microarray that employs hybridisation fingerprints generated by high-density anonymous markers distributed over the entire genome for identification based on comparison to a reference database. Results A high-density microarray carrying 95,000 unique 13-mer probes was designed. Optimized methods were developed to deliver reproducible hybridisation patterns that enabled confident discrimination of bacteria at the species, subspecies, and strain levels. High correlation coefficients were achieved between replicates. A sub-selection of 12,071 probes, determined by ANOVA and class prediction analysis, enabled the discrimination of all samples in our panel. Mismatch probe hybridisation was observed but was found to have no effect on the discriminatory capacity of our system. Conclusions These results indicate the potential of our genome chip for reliable identification of a wide range of bacterial taxa at the subspecies level without laborious prior sequencing and probe design. With its high resolution capacity, our proof-of-principle chip demonstrates great potential as a tool for molecular diagnostics of broad taxonomic groups. PMID:20163710

A detailed phylogeny for the Methanomicrobiales

NASA Technical Reports Server (NTRS)

Rouviere, P.; Mandelco, L.; Winker, S.; Woese, C. R.

1992-01-01

The small subunit rRNA sequence of twenty archaea, members of the Methanomicrobiales, permits a detailed phylogenetic tree to be inferred for the group. The tree confirms earlier studies, based on far fewer sequences, in showing the group to be divided into two major clusters, temporarily designated the "methanosarcina" group and the "methanogenium" group. The tree also defines phylogenetic relationships within these two groups, which in some cases do not agree with the phylogenetic relationships implied by current taxonomic names--a problem most acute for the genus Methanogenium and its relatives. The present phylogenetic characterization provides the basis for a consistent taxonomic restructuring of this major methanogenic taxon.

A taxonomic framework for cable bacteria and proposal of the candidate genera Electrothrix and Electronema.

PubMed

Trojan, Daniela; Schreiber, Lars; Bjerg, Jesper T; Bøggild, Andreas; Yang, Tingting; Kjeldsen, Kasper U; Schramm, Andreas

2016-07-01

Cable bacteria are long, multicellular filaments that can conduct electric currents over centimeter-scale distances. All cable bacteria identified to date belong to the deltaproteobacterial family Desulfobulbaceae and have not been isolated in pure culture yet. Their taxonomic delineation and exact phylogeny is uncertain, as most studies so far have reported only short partial 16S rRNA sequences or have relied on identification by a combination of filament morphology and 16S rRNA-targeted fluorescence in situ hybridization with a Desulfobulbaceae-specific probe. In this study, nearly full-length 16S rRNA gene sequences of 16 individual cable bacteria filaments from freshwater, salt marsh, and marine sites of four geographic locations are presented. These sequences formed a distinct, monophyletic sister clade to the genus Desulfobulbus and could be divided into six coherent, species-level clusters, arranged as two genus-level groups. The same grouping was retrieved by phylogenetic analysis of full or partial dsrAB genes encoding the dissimilatory sulfite reductase. Based on these results, it is proposed to accommodate cable bacteria within two novel candidate genera: the mostly marine "Candidatus Electrothrix", with four candidate species, and the mostly freshwater "Candidatus Electronema", with two candidate species. This taxonomic framework can be used to assign environmental sequences confidently to the cable bacteria clade, even without morphological information. Database searches revealed 185 16S rRNA gene sequences that affiliated within the clade formed by the proposed cable bacteria genera, of which 120 sequences could be assigned to one of the six candidate species, while the remaining 65 sequences indicated the existence of up to five additional species. Copyright © 2016 The Author(s). Published by Elsevier GmbH.. All rights reserved.

An Advanced Approach to Simultaneous Monitoring of Multiple Bacteria in Space

NASA Technical Reports Server (NTRS)

Eggers, M.

1998-01-01

The utility of a novel microarray-based microbial analyzer was demonstrated by the rapid detection, imaging, and identification of a mixture of microorganisms found in a waste water sample from the Lunar-Mars Life Support Test Project through the synergistic combination of: (1) judicious RNA probe selection via algorithms developed by University of Houston scientists; (2) tuned surface chemistries developed by Baylor College of Medicine scientists to facilitate hybridization of rRNA targets to DNA probes under very low salt conditions, thereby minimizing secondary structure; and (3) integration of the microarray printing and detection/imaging instrumentation by Genometrix to complete the quantitative analysis of microorganism mixtures.

Bacterial communities in the phylloplane of Prunus species.

PubMed

Jo, Yeonhwa; Cho, Jin Kyong; Choi, Hoseong; Chu, Hyosub; Lian, Sen; Cho, Won Kyong

2015-04-01

Bacterial populations in the phylloplane of four different Prunus species were investigated by 16 S rRNA pyrosequencing. Bioinformatic analysis identified an average of 510 operational taxonomic units belonging to 159 genera in 76 families. The two genera, Sphingomonas and Methylobacterium, were dominant in the phylloplane of four Prunus species. Twenty three genera were commonly identified in the four Prunus species, indicating a high level of bacterial diversity dependent on the plant species. Our study based on 16 S rRNA sequencing reveals the complexity of bacterial diversity in the phylloplane of Prunus species in detail. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Lineage-specific responses of microbial communities to environmental change.

PubMed

Youngblut, Nicholas D; Shade, Ashley; Read, Jordan S; McMahon, Katherine D; Whitaker, Rachel J

2013-01-01

A great challenge facing microbial ecology is how to define ecologically relevant taxonomic units. To address this challenge, we investigated how changing the definition of operational taxonomic units (OTUs) influences the perception of ecological patterns in microbial communities as they respond to a dramatic environmental change. We used pyrosequenced tags of the bacterial V2 16S rRNA region, as well as clone libraries constructed from the cytochrome oxidase C gene ccoN, to provide additional taxonomic resolution for the common freshwater genus Polynucleobacter. At the most highly resolved taxonomic scale, we show that distinct genotypes associated with the abundant Polynucleobacter lineages exhibit divergent spatial patterns and dramatic changes over time, while the also abundant Actinobacteria OTUs are highly coherent. This clearly demonstrates that different bacterial lineages demand different taxonomic definitions to capture ecological patterns. Based on the temporal distribution of highly resolved taxa in the hypolimnion, we demonstrate that change in the population structure of a single genotype can provide additional insight into the mechanisms of community-level responses. These results highlight the importance and feasibility of examining ecological change in microbial communities across taxonomic scales while also providing valuable insight into the ecological characteristics of ecologically coherent groups in this system.

Genome-Based Taxonomic Classification of Bacteroidetes

DOE PAGES

Hahnke, Richard L.; Meier-Kolthoff, Jan P.; García-López, Marina; ...

2016-12-20

The bacterial phylum Bacteroidetes, characterized by a distinct gliding motility, occurs in a broad variety of ecosystems, habitats, life styles, and physiologies. Accordingly, taxonomic classification of the phylum, based on a limited number of features, proved difficult and controversial in the past, for example, when decisions were based on unresolved phylogenetic trees of the 16S rRNA gene sequence. Here we use a large collection of type-strain genomes from Bacteroidetes and closely related phyla for assessing their taxonomy based on the principles of phylogenetic classification and trees inferred from genome-scale data. No significant conflict between 16S rRNA gene and whole-genome phylogeneticmore » analysis is found, whereas many but not all of the involved taxa are supported as monophyletic groups, particularly in the genome-scale trees. Phenotypic and phylogenomic features support the separation of Balneolaceae as new phylum Balneolaeota from Rhodothermaeota and of Saprospiraceae as new class Saprospiria from Chitinophagia. Epilithonimonas is nested within the older genus Chryseobacterium and without significant phenotypic differences; thus merging the two genera is proposed. Similarly, Vitellibacter is proposed to be included in Aequorivita. Flexibacter is confirmed as being heterogeneous and dissected, yielding six distinct genera. Hallella seregens is a later heterotypic synonym of Prevotella dentalis. Compared to values directly calculated from genome sequences, the G+C content mentioned in many species descriptions is too imprecise; moreover, corrected G+C content values have a significantly better fit to the phylogeny. Corresponding emendations of species descriptions are provided where necessary. Whereas most observed conflict with the current classification of Bacteroidetes is already visible in 16S rRNA gene trees, as expected whole-genome phylogenies are much better resolved.« less

Genome-Based Taxonomic Classification of Bacteroidetes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hahnke, Richard L.; Meier-Kolthoff, Jan P.; García-López, Marina

The bacterial phylum Bacteroidetes, characterized by a distinct gliding motility, occurs in a broad variety of ecosystems, habitats, life styles, and physiologies. Accordingly, taxonomic classification of the phylum, based on a limited number of features, proved difficult and controversial in the past, for example, when decisions were based on unresolved phylogenetic trees of the 16S rRNA gene sequence. Here we use a large collection of type-strain genomes from Bacteroidetes and closely related phyla for assessing their taxonomy based on the principles of phylogenetic classification and trees inferred from genome-scale data. No significant conflict between 16S rRNA gene and whole-genome phylogeneticmore » analysis is found, whereas many but not all of the involved taxa are supported as monophyletic groups, particularly in the genome-scale trees. Phenotypic and phylogenomic features support the separation of Balneolaceae as new phylum Balneolaeota from Rhodothermaeota and of Saprospiraceae as new class Saprospiria from Chitinophagia. Epilithonimonas is nested within the older genus Chryseobacterium and without significant phenotypic differences; thus merging the two genera is proposed. Similarly, Vitellibacter is proposed to be included in Aequorivita. Flexibacter is confirmed as being heterogeneous and dissected, yielding six distinct genera. Hallella seregens is a later heterotypic synonym of Prevotella dentalis. Compared to values directly calculated from genome sequences, the G+C content mentioned in many species descriptions is too imprecise; moreover, corrected G+C content values have a significantly better fit to the phylogeny. Corresponding emendations of species descriptions are provided where necessary. Whereas most observed conflict with the current classification of Bacteroidetes is already visible in 16S rRNA gene trees, as expected whole-genome phylogenies are much better resolved.« less

Genome-Based Taxonomic Classification of Bacteroidetes

PubMed Central

Hahnke, Richard L.; Meier-Kolthoff, Jan P.; García-López, Marina; Mukherjee, Supratim; Huntemann, Marcel; Ivanova, Natalia N.; Woyke, Tanja; Kyrpides, Nikos C.; Klenk, Hans-Peter; Göker, Markus

2016-01-01

The bacterial phylum Bacteroidetes, characterized by a distinct gliding motility, occurs in a broad variety of ecosystems, habitats, life styles, and physiologies. Accordingly, taxonomic classification of the phylum, based on a limited number of features, proved difficult and controversial in the past, for example, when decisions were based on unresolved phylogenetic trees of the 16S rRNA gene sequence. Here we use a large collection of type-strain genomes from Bacteroidetes and closely related phyla for assessing their taxonomy based on the principles of phylogenetic classification and trees inferred from genome-scale data. No significant conflict between 16S rRNA gene and whole-genome phylogenetic analysis is found, whereas many but not all of the involved taxa are supported as monophyletic groups, particularly in the genome-scale trees. Phenotypic and phylogenomic features support the separation of Balneolaceae as new phylum Balneolaeota from Rhodothermaeota and of Saprospiraceae as new class Saprospiria from Chitinophagia. Epilithonimonas is nested within the older genus Chryseobacterium and without significant phenotypic differences; thus merging the two genera is proposed. Similarly, Vitellibacter is proposed to be included in Aequorivita. Flexibacter is confirmed as being heterogeneous and dissected, yielding six distinct genera. Hallella seregens is a later heterotypic synonym of Prevotella dentalis. Compared to values directly calculated from genome sequences, the G+C content mentioned in many species descriptions is too imprecise; moreover, corrected G+C content values have a significantly better fit to the phylogeny. Corresponding emendations of species descriptions are provided where necessary. Whereas most observed conflict with the current classification of Bacteroidetes is already visible in 16S rRNA gene trees, as expected whole-genome phylogenies are much better resolved. PMID:28066339

Application of Molecular Techniques To Elucidate the Influence of Cellulosic Waste on the Bacterial Community Structure at a Simulated Low-Level-Radioactive-Waste Site▿ †

PubMed Central

Field, Erin K.; D'Imperio, Seth; Miller, Amber R.; VanEngelen, Michael R.; Gerlach, Robin; Lee, Brady D.; Apel, William A.; Peyton, Brent M.

2010-01-01

Low-level-radioactive-waste (low-level-waste) sites, including those at various U.S. Department of Energy sites, frequently contain cellulosic waste in the form of paper towels, cardboard boxes, or wood contaminated with heavy metals and radionuclides such as chromium and uranium. To understand how the soil microbial community is influenced by the presence of cellulosic waste products, multiple soil samples were obtained from a nonradioactive model low-level-waste test pit at the Idaho National Laboratory. Samples were analyzed using 16S rRNA gene clone libraries and 16S rRNA gene microarray (PhyloChip) analyses. Both methods revealed changes in the bacterial community structure with depth. In all samples, the PhyloChip detected significantly more operational taxonomic units, and therefore relative diversity, than the clone libraries. Diversity indices suggest that diversity is lowest in the fill and fill-waste interface (FW) layers and greater in the wood waste and waste-clay interface layers. Principal-coordinate analysis and lineage-specific analysis determined that the Bacteroidetes and Actinobacteria phyla account for most of the significant differences observed between the layers. The decreased diversity in the FW layer and increased members of families containing known cellulose-degrading microorganisms suggest that the FW layer is an enrichment environment for these organisms. These results suggest that the presence of the cellulosic material significantly influences the bacterial community structure in a stratified soil system. PMID:20305022

Characterization of Bacterial Communities Associated with the Tyrian Purple Producing Gland in a Marine Gastropod

PubMed Central

Ngangbam, Ajit Kumar; Baten, Abdul; Waters, Daniel L. E.; Whalan, Steve; Benkendorff, Kirsten

2015-01-01

Dicathais orbita is a marine mollusc recognised for the production of anticancer compounds that are precursors to Tyrian purple. This study aimed to assess the diversity and identity of bacteria associated with the Tyrian purple producing hypobranchial gland, in comparison with foot tissue, using a high-throughput sequencing approach. Taxonomic and phylogenetic analysis of variable region V1-V3 of 16S rRNA bacterial gene amplicons in QIIME and MEGAN were carried out. This analysis revealed a highly diverse bacterial assemblage associated with the hypobranchial gland and foot tissues of D. orbita. The dominant bacterial phylum in the 16S rRNA bacterial profiling data set was Proteobacteria followed by Bacteroidetes, Tenericutes and Spirochaetes. In comparison to the foot, the hypobranchial gland had significantly lower bacterial diversity and a different community composition, based on taxonomic assignment at the genus level. A higher abundance of indole producing Vibrio spp. and the presence of bacteria with brominating capabilities in the hypobranchial gland suggest bacteria have a potential role in biosynthesis of Tyrian purple in D. orbita. PMID:26488885

Development and validation of a microarray for the investigation of the CAZymes encoded by the human gut microbiome.

PubMed

El Kaoutari, Abdessamad; Armougom, Fabrice; Leroy, Quentin; Vialettes, Bernard; Million, Matthieu; Raoult, Didier; Henrissat, Bernard

2013-01-01

Distal gut bacteria play a pivotal role in the digestion of dietary polysaccharides by producing a large number of carbohydrate-active enzymes (CAZymes) that the host otherwise does not produce. We report here the design of a custom microarray that we used to spot non-redundant DNA probes for more than 6,500 genes encoding glycoside hydrolases and lyases selected from 174 reference genomes from distal gut bacteria. The custom microarray was tested and validated by the hybridization of bacterial DNA extracted from the stool samples of lean, obese and anorexic individuals. Our results suggest that a microarray-based study can detect genes from low-abundance bacteria better than metagenomic-based studies. A striking example was the finding that a gene encoding a GH6-family cellulase was present in all subjects examined, whereas metagenomic studies have consistently failed to detect this gene in both human and animal gut microbiomes. In addition, an examination of eight stool samples allowed the identification of a corresponding CAZome core containing 46 families of glycoside hydrolases and polysaccharide lyases, which suggests the functional stability of the gut microbiota despite large taxonomical variations between individuals.

Phylogenetic analysis of Haemaphysalis erinacei Pavesi, 1884 (Acari: Ixodidae) from China, Turkey, Italy and Romania.

PubMed

Hornok, Sándor; Wang, Yuanzhi; Otranto, Domenico; Keskin, Adem; Lia, Riccardo Paolo; Kontschán, Jenő; Takács, Nóra; Farkas, Róbert; Sándor, Attila D

2016-12-15

Haemaphysalis erinacei is one of the few ixodid tick species for which valid names of subspecies exist. Despite their disputed taxonomic status in the literature, these subspecies have not yet been compared with molecular methods. The aim of the present study was to investigate the phylogenetic relationships of H. erinacei subspecies, in the context of the first finding of this tick species in Romania. After morphological identification, DNA was extracted from five adults of H. e. taurica (from Romania and Turkey), four adults of H. e. erinacei (from Italy) and 17 adults of H. e. turanica (from China). From these samples fragments of the cytochrome c oxidase subunit 1 (cox1) and 16S rRNA genes were amplified via PCR and sequenced. Results showed that cox1 and 16S rRNA gene sequence divergences between H. e. taurica from Romania and H. e. erinacei from Italy were below 2%. However, the sequence divergences between H. e. taurica from Romania and H. e. turanica from China were high (up to 7.3% difference for the 16S rRNA gene), exceeding the reported level of sequence divergence between closely related tick species. At the same time, two adults of H. e. taurica from Turkey had higher 16S rRNA gene similarity to H. e. turanica from China (up to 97.5%) than to H. e. taurica from Romania (96.3%), but phylogenetically clustered more closely to H. e. taurica than to H. e. turanica. This is the first finding of H. erinacei in Romania, and the first (although preliminary) phylogenetic comparison of H. erinacei subspecies. Phylogenetic analyses did not support that the three H. erinacei subspecies evaluated here are of equal taxonomic rank, because the genetic divergence between H. e. turanica from China and H. e. taurica from Romania exceeded the usual level of sequence divergence between closely related tick species, suggesting that they might represent different species. Therefore, the taxonomic status of the subspecies of H. erinacei needs to be revised based on a larger number of specimens collected throughout its geographical range.

Evaluating the Detection of Hydrocarbon-Degrading Bacteria in 16S rRNA Gene Sequencing Surveys

PubMed Central

Berry, David; Gutierrez, Tony

2017-01-01

Hydrocarbonoclastic bacteria (HCB) play a key role in the biodegradation of oil hydrocarbons in marine and other environments. A small number of taxa have been identified as obligate HCB, notably the Gammaproteobacterial genera Alcanivorax, Cycloclasticus, Marinobacter, Neptumonas, Oleiphilus, Oleispira, and Thalassolituus, as well as the Alphaproteobacterial genus Thalassospira. Detection of HCB in amplicon-based sequencing surveys relies on high coverage by PCR primers and accurate taxonomic classification. In this study, we performed a phylogenetic analysis to identify 16S rRNA gene sequence regions that represent the breadth of sequence diversity within these taxa. Using validated sequences, we evaluated 449 universal 16S rRNA gene-targeted bacterial PCR primer pairs for their coverage of these taxa. The results of this analysis provide a practical framework for selection of suitable primer sets for optimal detection of HCB in sequencing surveys. PMID:28567035

Evaluating the Detection of Hydrocarbon-Degrading Bacteria in 16S rRNA Gene Sequencing Surveys.

PubMed

Berry, David; Gutierrez, Tony

2017-01-01

Hydrocarbonoclastic bacteria (HCB) play a key role in the biodegradation of oil hydrocarbons in marine and other environments. A small number of taxa have been identified as obligate HCB, notably the Gammaproteobacterial genera Alcanivorax, Cycloclasticus, Marinobacter, Neptumonas, Oleiphilus, Oleispira , and Thalassolituus , as well as the Alphaproteobacterial genus Thalassospira . Detection of HCB in amplicon-based sequencing surveys relies on high coverage by PCR primers and accurate taxonomic classification. In this study, we performed a phylogenetic analysis to identify 16S rRNA gene sequence regions that represent the breadth of sequence diversity within these taxa. Using validated sequences, we evaluated 449 universal 16S rRNA gene-targeted bacterial PCR primer pairs for their coverage of these taxa. The results of this analysis provide a practical framework for selection of suitable primer sets for optimal detection of HCB in sequencing surveys.

The Deinococcus-Thermus phylum and the effect of rRNA composition on phylogenetic tree construction

NASA Technical Reports Server (NTRS)

Weisburg, W. G.; Giovannoni, S. J.; Woese, C. R.

1989-01-01

Through comparative analysis of 16S ribosomal RNA sequences, it can be shown that two seemingly dissimilar types of eubacteria Deinococcus and the ubiquitous hot spring organism Thermus are distantly but specifically related to one another. This confirms an earlier report based upon 16S rRNA oligonucleotide cataloging studies (Hensel et al., 1986). Their two lineages form a distinctive grouping within the eubacteria that deserved the taxonomic status of a phylum. The (partial) sequence of T. aquaticus rRNA appears relatively close to those of other thermophilic eubacteria. e.g. Thermotoga maritima and Thermomicrobium roseum. However, this closeness does not reflect a true evolutionary closeness; rather it is due to a "thermophilic convergence", the result of unusually high G+C composition in the rRNAs of thermophilic bacteria. Unless such compositional biases are taken into account, the branching order and root of phylogenetic trees can be incorrectly inferred.

IMNGS: A comprehensive open resource of processed 16S rRNA microbial profiles for ecology and diversity studies.

PubMed

Lagkouvardos, Ilias; Joseph, Divya; Kapfhammer, Martin; Giritli, Sabahattin; Horn, Matthias; Haller, Dirk; Clavel, Thomas

2016-09-23

The SRA (Sequence Read Archive) serves as primary depository for massive amounts of Next Generation Sequencing data, and currently host over 100,000 16S rRNA gene amplicon-based microbial profiles from various host habitats and environments. This number is increasing rapidly and there is a dire need for approaches to utilize this pool of knowledge. Here we created IMNGS (Integrated Microbial Next Generation Sequencing), an innovative platform that uniformly and systematically screens for and processes all prokaryotic 16S rRNA gene amplicon datasets available in SRA and uses them to build sample-specific sequence databases and OTU-based profiles. Via a web interface, this integrative sequence resource can easily be queried by users. We show examples of how the approach allows testing the ecological importance of specific microorganisms in different hosts or ecosystems, and performing targeted diversity studies for selected taxonomic groups. The platform also offers a complete workflow for de novo analysis of users' own raw 16S rRNA gene amplicon datasets for the sake of comparison with existing data. IMNGS can be accessed at www.imngs.org.

Comparison of Barley Succession and Take-All Disease as Environmental Factors Shaping the Rhizobacterial Community during Take-All Decline▿

PubMed Central

Schreiner, Karin; Hagn, Alexandra; Kyselková, Martina; Moënne-Loccoz, Yvan; Welzl, Gerhard; Munch, Jean Charles; Schloter, Michael

2010-01-01

The root disease take-all, caused by Gaeumannomyces graminis var. tritici, can be managed by monoculture-induced take-all decline (TAD). This natural biocontrol mechanism typically occurs after a take-all outbreak and is believed to arise from an enrichment of antagonistic populations in the rhizosphere. However, it is not known whether these changes are induced by the monoculture or by ecological rhizosphere conditions due to a disease outbreak and subsequent attenuation. This question was addressed by comparing the rhizosphere microflora of barley, either inoculated with the pathogen or noninoculated, in a microcosm experiment in five consecutive vegetation cycles. TAD occurred in soil inoculated with the pathogen but not in noninoculated soil. Bacterial community analysis using terminal restriction fragment length polymorphism of 16S rRNA showed pronounced population shifts in the successive vegetation cycles, but pathogen inoculation had little effect. To elucidate rhizobacterial dynamics during TAD development, a 16S rRNA-based taxonomic microarray was used. Actinobacteria were the prevailing indicators in the first vegetation cycle, whereas the third cycle—affected most severely by take-all—was characterized by Proteobacteria, Bacteroidetes, Chloroflexi, Planctomycetes, and Acidobacteria. Indicator taxa for the last cycle (TAD) belonged exclusively to Proteobacteria, including several genera with known biocontrol traits. Our results suggest that TAD involves monoculture-induced enrichment of plant-beneficial taxa. PMID:20525871

Oligotyping reveals differences between gut microbiomes of free-ranging sympatric Namibian carnivores (Acinonyx jubatus, Canis mesomelas) on a bacterial species-like level

PubMed Central

Menke, Sebastian; Wasimuddin; Meier, Matthias; Melzheimer, Jörg; Mfune, John K. E.; Heinrich, Sonja; Thalwitzer, Susanne; Wachter, Bettina; Sommer, Simone

2014-01-01

Recent gut microbiome studies in model organisms emphasize the effects of intrinsic and extrinsic factors on the variation of the bacterial composition and its impact on the overall health status of the host. Species occurring in the same habitat might share a similar microbiome, especially if they overlap in ecological and behavioral traits. So far, the natural variation in microbiomes of free-ranging wildlife species has not been thoroughly investigated. The few existing studies exploring microbiomes through 16S rRNA gene reads clustered sequencing reads into operational taxonomic units (OTUs) based on a similarity threshold (e.g., 97%). This approach, in combination with the low resolution of target databases, generally limits the level of taxonomic assignments to the genus level. However, distinguishing natural variation of microbiomes in healthy individuals from “abnormal” microbial compositions that affect host health requires knowledge of the “normal” microbial flora at a high taxonomic resolution. This gap can now be addressed using the recently published oligotyping approach, which can resolve closely related organisms into distinct oligotypes by utilizing subtle nucleotide variation. Here, we used Illumina MiSeq to sequence amplicons generated from the V4 region of the 16S rRNA gene to investigate the gut microbiome of two free-ranging sympatric Namibian carnivore species, the cheetah (Acinonyx jubatus) and the black-backed jackal (Canis mesomelas). Bacterial phyla with proportions >0.2% were identical for both species and included Firmicutes, Fusobacteria, Bacteroidetes, Proteobacteria and Actinobacteria. At a finer taxonomic resolution, black-backed jackals exhibited 69 bacterial taxa with proportions ≥0.1%, whereas cheetahs had only 42. Finally, oligotyping revealed that shared bacterial taxa consisted of distinct oligotype profiles. Thus, in contrast to 3% OTUs, oligotyping can detect fine-scale taxonomic differences between microbiomes. PMID:25352837

How many novel eukaryotic 'kingdoms'? Pitfalls and limitations of environmental DNA surveys

PubMed Central

Berney, Cédric; Fahrni, José; Pawlowski, Jan

2004-01-01

Background Over the past few years, the use of molecular techniques to detect cultivation-independent, eukaryotic diversity has proven to be a powerful approach. Based on small-subunit ribosomal RNA (SSU rRNA) gene analyses, these studies have revealed the existence of an unexpected variety of new phylotypes. Some of them represent novel diversity in known eukaryotic groups, mainly stramenopiles and alveolates. Others do not seem to be related to any molecularly described lineage, and have been proposed to represent novel eukaryotic kingdoms. In order to review the evolutionary importance of this novel high-level eukaryotic diversity critically, and to test the potential technical and analytical pitfalls and limitations of eukaryotic environmental DNA surveys (EES), we analysed 484 environmental SSU rRNA gene sequences, including 81 new sequences from sediments of the small river, the Seymaz (Geneva, Switzerland). Results Based on a detailed screening of an exhaustive alignment of eukaryotic SSU rRNA gene sequences and the phylogenetic re-analysis of previously published environmental sequences using Bayesian methods, our results suggest that the number of novel higher-level taxa revealed by previously published EES was overestimated. Three main sources of errors are responsible for this situation: (1) the presence of undetected chimeric sequences; (2) the misplacement of several fast-evolving sequences; and (3) the incomplete sampling of described, but yet unsequenced eukaryotes. Additionally, EES give a biased view of the diversity present in a given biotope because of the difficult amplification of SSU rRNA genes in some taxonomic groups. Conclusions Environmental DNA surveys undoubtedly contribute to reveal many novel eukaryotic lineages, but there is no clear evidence for a spectacular increase of the diversity at the kingdom level. After re-analysis of previously published data, we found only five candidate lineages of possible novel high-level eukaryotic taxa, two of which comprise several phylotypes that were found independently in different studies. To ascertain their taxonomic status, however, the organisms themselves have now to be identified. PMID:15176975

Molecular evolution inferred from small subunit rRNA sequences: what does it tell us about phylogenetic relationships and taxonomy of the parabasalids?

PubMed

Viscogliosi, E; Edgcomb, V P; Gerbod, D; Noël, C; Delgado-Viscogliosi, P

1999-12-01

The Parabasala are a primitive group of protists divided into two classes: the trichomonads and the hypermastigids. Until recently, phylogeny and taxonomy of parabasalids were mainly based on the comparative analysis of morphological characters primarily linked to the development of their cytoskeleton. Recent use of molecular markers, such as small subunit (SSU) rRNA has led to now insights into the systematics of the Parabasala and other groups of prolists. An updated phylogeny based on SSU rRNA is provided and compared to that inferred from ultrastructural data. The SSU rRNA phylogeny contradicts the dogma equating simple characters with pumitive characters. Hypermastigids, possessing a hyperdeveloped cytoskeleton, exhibit the most basal emergence in the parabasalid lineage. Other observations emerge from the SSU rRNA analysis, such as the secondary loss of some cytoskeleton structures in all representatives of the Monocercomonadidae, the existence of secondarily free living taxa (reversibility of parasitism) and the evidence against the co-evolution of the endobiotic parabasalids and their animal hosts. According to phylogenies based on SSU rRNA, all the trichomonad families are not monophyletic groups, putting into question the validity of current taxonomic assignments. The precise branching order of some taxa remains unclear, but this issue can possibly be addressed by the molecular analysis of additional parabasalids. The goal of such additional analyses would be to propose, in a near future, a revision of the taxonomy of this group of protists that takes into account both molecular and morphological data.

Molecular evolution inferred from small subunit rRNA sequences: what does it tell us about phylogenetic relationships and taxonomy of the parabasalids?

NASA Technical Reports Server (NTRS)

Viscogliosi, E.; Edgcomb, V. P.; Gerbod, D.; Noel, C.; Delgado-Viscogliosi, P.; Sogin, M. L. (Principal Investigator)

1999-01-01

The Parabasala are a primitive group of protists divided into two classes: the trichomonads and the hypermastigids. Until recently, phylogeny and taxonomy of parabasalids were mainly based on the comparative analysis of morphological characters primarily linked to the development of their cytoskeleton. Recent use of molecular markers, such as small subunit (SSU) rRNA has led to now insights into the systematics of the Parabasala and other groups of prolists. An updated phylogeny based on SSU rRNA is provided and compared to that inferred from ultrastructural data. The SSU rRNA phylogeny contradicts the dogma equating simple characters with pumitive characters. Hypermastigids, possessing a hyperdeveloped cytoskeleton, exhibit the most basal emergence in the parabasalid lineage. Other observations emerge from the SSU rRNA analysis, such as the secondary loss of some cytoskeleton structures in all representatives of the Monocercomonadidae, the existence of secondarily free living taxa (reversibility of parasitism) and the evidence against the co-evolution of the endobiotic parabasalids and their animal hosts. According to phylogenies based on SSU rRNA, all the trichomonad families are not monophyletic groups, putting into question the validity of current taxonomic assignments. The precise branching order of some taxa remains unclear, but this issue can possibly be addressed by the molecular analysis of additional parabasalids. The goal of such additional analyses would be to propose, in a near future, a revision of the taxonomy of this group of protists that takes into account both molecular and morphological data.

Estimating the efficiency of fish cross-species cDNA microarray hybridization.

PubMed

Cohen, Raphael; Chalifa-Caspi, Vered; Williams, Timothy D; Auslander, Meirav; George, Stephen G; Chipman, James K; Tom, Moshe

2007-01-01

Using an available cross-species cDNA microarray is advantageous for examining multigene expression patterns in non-model organisms, saving the need for construction of species-specific arrays. The aim of the present study was to estimate relative efficiency of cross-species hybridizations across bony fishes, using bioinformatics tools. The methodology may serve also as a model for similar evaluations in other taxa. The theoretical evaluation was done by substituting comparative whole-transcriptome sequence similarity information into the thermodynamic hybridization equation. Complementary DNA sequence assemblages of nine fish species belonging to common families or suborders and distributed across the bony fish taxonomic branch were selected for transcriptome-wise comparisons. Actual cross-species hybridizations among fish of different taxonomic distances were used to validate and eventually to calibrate the theoretically computed relative efficiencies.

Ribosomal RNA gene detection and targeted culture of novel nitrogen-responsive fungal taxa from temperate pine forest soil.

PubMed

Hesse, Cedar N; Torres-Cruz, Terry J; Tobias, Terri Billingsley; Al-Matruk, Maryam; Porras-Alfaro, Andrea; Kuske, Cheryl R

Soil fungal communities are responsible for carbon and nitrogen (N) cycling. The high complexity of the soil fungal community and the high proportion of taxonomically unidentifiable sequences confound ecological interpretations in field studies because physiological information is lacking for many organisms known only by their rRNA sequences. This situation forces experimental comparisons to be made at broader taxonomic racks where functions become difficult to infer. The objective of this study was to determine OTU (operational taxonomic units) level responses of the soil fungal community to N enrichment in a temperate pine forest experiment and to use the sequencing data to guide culture efforts of novel N-responsive fungal taxa. Replicate samples from four soil horizons (up to 10 cm depth) were obtained from ambient, enriched CO 2 and N-fertilization plots. Through a fungal large subunit rRNA gene (LSU) sequencing survey, we identified two novel fungal clades that were abundant in our soil sampling (representing up to 27% of the sequences in some samples) and responsive to changes in soil N. The two N-responsive taxa with no predicted taxonomic association were targeted for isolation and culturing from specific soil samples where their sequences were abundant. Representatives of both OTUs were successfully cultured using a filtration approach. One taxon (OTU6) was most closely related to Saccharomycotina; the second taxon (OTU69) was most closely related to Mucoromycotina. Both taxa likely represent novel species. This study shows how observation of specific OTUs level responses to altered N status in a large rRNA gene field survey provided the impetus to design targeted culture approaches for isolation of novel N-responsive fungal taxa.

Impact of training sets on classification of high-throughput bacterial 16s rRNA gene surveys

PubMed Central

Werner, Jeffrey J; Koren, Omry; Hugenholtz, Philip; DeSantis, Todd Z; Walters, William A; Caporaso, J Gregory; Angenent, Largus T; Knight, Rob; Ley, Ruth E

2012-01-01

Taxonomic classification of the thousands–millions of 16S rRNA gene sequences generated in microbiome studies is often achieved using a naïve Bayesian classifier (for example, the Ribosomal Database Project II (RDP) classifier), due to favorable trade-offs among automation, speed and accuracy. The resulting classification depends on the reference sequences and taxonomic hierarchy used to train the model; although the influence of primer sets and classification algorithms have been explored in detail, the influence of training set has not been characterized. We compared classification results obtained using three different publicly available databases as training sets, applied to five different bacterial 16S rRNA gene pyrosequencing data sets generated (from human body, mouse gut, python gut, soil and anaerobic digester samples). We observed numerous advantages to using the largest, most diverse training set available, that we constructed from the Greengenes (GG) bacterial/archaeal 16S rRNA gene sequence database and the latest GG taxonomy. Phylogenetic clusters of previously unclassified experimental sequences were identified with notable improvements (for example, 50% reduction in reads unclassified at the phylum level in mouse gut, soil and anaerobic digester samples), especially for phylotypes belonging to specific phyla (Tenericutes, Chloroflexi, Synergistetes and Candidate phyla TM6, TM7). Trimming the reference sequences to the primer region resulted in systematic improvements in classification depth, and greatest gains at higher confidence thresholds. Phylotypes unclassified at the genus level represented a greater proportion of the total community variation than classified operational taxonomic units in mouse gut and anaerobic digester samples, underscoring the need for greater diversity in existing reference databases. PMID:21716311

Molecular taxonomy, phylogeny and evolution in the family Stichopodidae (Aspidochirotida: Holothuroidea) based on COI and 16S mitochondrial DNA.

PubMed

Byrne, Maria; Rowe, Frank; Uthicke, Sven

2010-09-01

The Stichopodidae comprise a diverse assemblage of holothuroids most of which occur in the Indo-Pacific. Phylogenetic analyses of mitochondrial gene (COI, 16S rRNA) sequence for 111 individuals (7 genera, 17 species) clarified taxonomic uncertainties, species relationships, biogeography and evolution of the family. A monophyly of the genus Stichopus was supported with the exception of Stichopus ellipes. Molecular analyses confirmed genus level taxonomy based on morphology. Most specimens harvested as S. horrens fell in the S. monotuberculatus clade, a morphologically variable assemblage with others from the S. naso clade. Taxonomic clarification of species fished as S. horrens will assist conservation measures. Evolutionary rates based on comparison of sequence from trans-ithmian Isostichopus species estimated that Stichopus and Isostichopus diverged ca. 5.5-10.7Ma (Miocene). More recent splits were estimated to be younger than 1Ma. Copyright 2010 Elsevier Inc. All rights reserved.

Seasonal dynamics of freshwater pathogens as measured by microarray at Lake Sapanca, a drinking water source in the north-eastern part of Turkey.

PubMed

Akçaalan, Reyhan; Albay, Meric; Koker, Latife; Baudart, Julia; Guillebault, Delphine; Fischer, Sabine; Weigel, Wilfried; Medlin, Linda K

2017-12-22

Monitoring drinking water quality is an important public health issue. Two objectives from the 4 years, six nations, EU Project μAqua were to develop hierarchically specific probes to detect and quantify pathogens in drinking water using a PCR-free microarray platform and to design a standardised water sampling program from different sources in Europe to obtain sufficient material for downstream analysis. Our phylochip contains barcodes (probes) that specifically identify freshwater pathogens that are human health risks in a taxonomic hierarchical fashion such that if species is present, the entire taxonomic hierarchy (genus, family, order, phylum, kingdom) leading to it must also be present, which avoids false positives. Molecular tools are more rapid, accurate and reliable than traditional methods, which means faster mitigation strategies with less harm to humans and the community. We present microarray results for the presence of freshwater pathogens from a Turkish lake used drinking water and inferred cyanobacterial cell equivalents from samples concentrated from 40 into 1 L in 45 min using hollow fibre filters. In two companion studies from the same samples, cyanobacterial toxins were analysed using chemical methods and those dates with highest toxin values also had highest cell equivalents as inferred from this microarray study.

probeBase—an online resource for rRNA-targeted oligonucleotide probes and primers: new features 2016

PubMed Central

Greuter, Daniel; Loy, Alexander; Horn, Matthias; Rattei, Thomas

2016-01-01

probeBase http://www.probebase.net is a manually maintained and curated database of rRNA-targeted oligonucleotide probes and primers. Contextual information and multiple options for evaluating in silico hybridization performance against the most recent rRNA sequence databases are provided for each oligonucleotide entry, which makes probeBase an important and frequently used resource for microbiology research and diagnostics. Here we present a major update of probeBase, which was last featured in the NAR Database Issue 2007. This update describes a complete remodeling of the database architecture and environment to accommodate computationally efficient access. Improved search functions, sequence match tools and data output now extend the opportunities for finding suitable hierarchical probe sets that target an organism or taxon at different taxonomic levels. To facilitate the identification of complementary probe sets for organisms represented by short rRNA sequence reads generated by amplicon sequencing or metagenomic analysis with next generation sequencing technologies such as Illumina and IonTorrent, we introduce a novel tool that recovers surrogate near full-length rRNA sequences for short query sequences and finds matching oligonucleotides in probeBase. PMID:26586809

Rhea: a transparent and modular R pipeline for microbial profiling based on 16S rRNA gene amplicons

PubMed Central

Fischer, Sandra; Kumar, Neeraj

2017-01-01

The importance of 16S rRNA gene amplicon profiles for understanding the influence of microbes in a variety of environments coupled with the steep reduction in sequencing costs led to a surge of microbial sequencing projects. The expanding crowd of scientists and clinicians wanting to make use of sequencing datasets can choose among a range of multipurpose software platforms, the use of which can be intimidating for non-expert users. Among available pipeline options for high-throughput 16S rRNA gene analysis, the R programming language and software environment for statistical computing stands out for its power and increased flexibility, and the possibility to adhere to most recent best practices and to adjust to individual project needs. Here we present the Rhea pipeline, a set of R scripts that encode a series of well-documented choices for the downstream analysis of Operational Taxonomic Units (OTUs) tables, including normalization steps, alpha- and beta-diversity analysis, taxonomic composition, statistical comparisons, and calculation of correlations. Rhea is primarily a straightforward starting point for beginners, but can also be a framework for advanced users who can modify and expand the tool. As the community standards evolve, Rhea will adapt to always represent the current state-of-the-art in microbial profiles analysis in the clear and comprehensive way allowed by the R language. Rhea scripts and documentation are freely available at https://lagkouvardos.github.io/Rhea. PMID:28097056

Rhea: a transparent and modular R pipeline for microbial profiling based on 16S rRNA gene amplicons.

PubMed

Lagkouvardos, Ilias; Fischer, Sandra; Kumar, Neeraj; Clavel, Thomas

2017-01-01

The importance of 16S rRNA gene amplicon profiles for understanding the influence of microbes in a variety of environments coupled with the steep reduction in sequencing costs led to a surge of microbial sequencing projects. The expanding crowd of scientists and clinicians wanting to make use of sequencing datasets can choose among a range of multipurpose software platforms, the use of which can be intimidating for non-expert users. Among available pipeline options for high-throughput 16S rRNA gene analysis, the R programming language and software environment for statistical computing stands out for its power and increased flexibility, and the possibility to adhere to most recent best practices and to adjust to individual project needs. Here we present the Rhea pipeline, a set of R scripts that encode a series of well-documented choices for the downstream analysis of Operational Taxonomic Units (OTUs) tables, including normalization steps, alpha - and beta -diversity analysis, taxonomic composition, statistical comparisons, and calculation of correlations. Rhea is primarily a straightforward starting point for beginners, but can also be a framework for advanced users who can modify and expand the tool. As the community standards evolve, Rhea will adapt to always represent the current state-of-the-art in microbial profiles analysis in the clear and comprehensive way allowed by the R language. Rhea scripts and documentation are freely available at https://lagkouvardos.github.io/Rhea.

Reclassification of Pseudomonas mephitica Claydon and Hammer 1939 as a later heterotypic synonym of Janthinobacterium lividum (Eisenberg 1891) De Ley et al. 1978.

PubMed

Kämpfer, Peter; Falsen, Enevold; Busse, Hans-Jürgen

2008-01-01

Pseudomonas mephitica CCUG 2513(T) has been reinvestigated to clarify its taxonomic position. 16S rRNA gene sequence comparisons demonstrated that this strain clusters phylogenetically closely with Janthinobacterium lividum (99.8% sequence similarity to the type strain). Investigation of fatty acid patterns, polar lipid profiles, polyamine patterns and quinone systems supported this delineation. Substrate utilization profiles and biochemical characteristics displayed no differences from the type strain of J. lividum, CCUG 2344(T). Therefore, the reclassification of Pseudomonas mephitica as a later heterotypic synonym of Janthinobacterium lividum is proposed, based upon the estimated phylogenetic position derived from 16S rRNA gene sequence data and chemotaxonomic and biochemical data.

Taxonomic evaluation of Streptomyces albus and related species using multilocus sequence analysis and proposals to emend the description of Streptomyces albus and describe Streptomyces pathocidini sp. nov

USDA-ARS?s Scientific Manuscript database

In phylogenetic analyses of the genus Streptomyces using 16S rRNA gene sequences, Streptomyces albus subsp. albus NRRL B-1811T forms a cluster with 5 other species having identical or nearly identical 16S rRNA gene sequences. Moreover, the morphological and physiological characteristics of these oth...

Molecular Phylogenetics and Systematics of the Bivalve Family Ostreidae Based on rRNA Sequence-Structure Models and Multilocus Species Tree

PubMed Central

Salvi, Daniele; Macali, Armando; Mariottini, Paolo

2014-01-01

The bivalve family Ostreidae has a worldwide distribution and includes species of high economic importance. Phylogenetics and systematic of oysters based on morphology have proved difficult because of their high phenotypic plasticity. In this study we explore the phylogenetic information of the DNA sequence and secondary structure of the nuclear, fast-evolving, ITS2 rRNA and the mitochondrial 16S rRNA genes from the Ostreidae and we implemented a multi-locus framework based on four loci for oyster phylogenetics and systematics. Sequence-structure rRNA models aid sequence alignment and improved accuracy and nodal support of phylogenetic trees. In agreement with previous molecular studies, our phylogenetic results indicate that none of the currently recognized subfamilies, Crassostreinae, Ostreinae, and Lophinae, is monophyletic. Single gene trees based on Maximum likelihood (ML) and Bayesian (BA) methods and on sequence-structure ML were congruent with multilocus trees based on a concatenated (ML and BA) and coalescent based (BA) approaches and consistently supported three main clades: (i) Crassostrea, (ii) Saccostrea, and (iii) an Ostreinae-Lophinae lineage. Therefore, the subfamily Crassotreinae (including Crassostrea), Saccostreinae subfam. nov. (including Saccostrea and tentatively Striostrea) and Ostreinae (including Ostreinae and Lophinae taxa) are recognized. Based on phylogenetic and biogeographical evidence the Asian species of Crassostrea from the Pacific Ocean are assigned to Magallana gen. nov., whereas an integrative taxonomic revision is required for the genera Ostrea and Dendostrea. This study pointed out the suitability of the ITS2 marker for DNA barcoding of oyster and the relevance of using sequence-structure rRNA models and features of the ITS2 folding in molecular phylogenetics and taxonomy. The multilocus approach allowed inferring a robust phylogeny of Ostreidae providing a broad molecular perspective on their systematics. PMID:25250663

Molecular phylogenetics and systematics of the bivalve family Ostreidae based on rRNA sequence-structure models and multilocus species tree.

PubMed

Salvi, Daniele; Macali, Armando; Mariottini, Paolo

2014-01-01

The bivalve family Ostreidae has a worldwide distribution and includes species of high economic importance. Phylogenetics and systematic of oysters based on morphology have proved difficult because of their high phenotypic plasticity. In this study we explore the phylogenetic information of the DNA sequence and secondary structure of the nuclear, fast-evolving, ITS2 rRNA and the mitochondrial 16S rRNA genes from the Ostreidae and we implemented a multi-locus framework based on four loci for oyster phylogenetics and systematics. Sequence-structure rRNA models aid sequence alignment and improved accuracy and nodal support of phylogenetic trees. In agreement with previous molecular studies, our phylogenetic results indicate that none of the currently recognized subfamilies, Crassostreinae, Ostreinae, and Lophinae, is monophyletic. Single gene trees based on Maximum likelihood (ML) and Bayesian (BA) methods and on sequence-structure ML were congruent with multilocus trees based on a concatenated (ML and BA) and coalescent based (BA) approaches and consistently supported three main clades: (i) Crassostrea, (ii) Saccostrea, and (iii) an Ostreinae-Lophinae lineage. Therefore, the subfamily Crassostreinae (including Crassostrea), Saccostreinae subfam. nov. (including Saccostrea and tentatively Striostrea) and Ostreinae (including Ostreinae and Lophinae taxa) are recognized [corrected]. Based on phylogenetic and biogeographical evidence the Asian species of Crassostrea from the Pacific Ocean are assigned to Magallana gen. nov., whereas an integrative taxonomic revision is required for the genera Ostrea and Dendostrea. This study pointed out the suitability of the ITS2 marker for DNA barcoding of oyster and the relevance of using sequence-structure rRNA models and features of the ITS2 folding in molecular phylogenetics and taxonomy. The multilocus approach allowed inferring a robust phylogeny of Ostreidae providing a broad molecular perspective on their systematics.

The Centipede Genus Scolopendra in Mainland Southeast Asia: Molecular Phylogenetics, Geometric Morphometrics and External Morphology as Tools for Species Delimitation

PubMed Central

Siriwut, Warut; Edgecombe, Gregory D.; Sutcharit, Chirasak; Panha, Somsak

2015-01-01

Seven Scolopendra species from the Southeast Asian mainland delimited based on standard external morphological characters represent monophyletic groups in phylogenetic trees inferred from concatenated sequences of three gene fragments (cytochrome c oxidase subunit 1, 16S rRNA and 28S rRNA) using Maximum likelihood and Bayesian inference. Geometric morphometric description of shape variation in the cephalic plate, forcipular coxosternite, and tergite of the ultimate leg-bearing segment provides additional criteria for distinguishing species. Colouration patterns in some Scolopendra species show a high degree of fit to phylogenetic trees at the population level. The most densely sampled species, Scolopendra dehaani Brandt, 1840, has three subclades with allopatric distributions in mainland SE Asia. The molecular phylogeny of S. pinguis Pocock, 1891, indicated ontogenetic colour variation among its populations. The taxonomic validation of S. dawydoffi Kronmüller, 2012, S. japonica Koch, 1878, and S. dehaani Brandt, 1840, each a former subspecies of S. subspinipes Leach, 1814 sensu Lewis, 2010, as full species was supported by molecular information and additional morphological data. Species delimitation in these taxonomically challenging animals is facilitated by an integrative approach that draws on both morphology and molecular phylogeny. PMID:26270342

Structure of the human gastric bacterial community in relation to Helicobacter pylori status.

PubMed

Maldonado-Contreras, Ana; Goldfarb, Kate C; Godoy-Vitorino, Filipa; Karaoz, Ulas; Contreras, Mónica; Blaser, Martin J; Brodie, Eoin L; Dominguez-Bello, Maria G

2011-04-01

The human stomach is naturally colonized by Helicobacter pylori, which, when present, dominates the gastric bacterial community. In this study, we aimed to characterize the structure of the bacterial community in the stomach of patients of differing H. pylori status. We used a high-density 16S rRNA gene microarray (PhyloChip, Affymetrix, Inc.) to hybridize 16S rRNA gene amplicons from gastric biopsy DNA of 10 rural Amerindian patients from Amazonas, Venezuela, and of two immigrants to the United States (from South Asia and Africa, respectively). H. pylori status was determined by PCR amplification of H. pylori glmM from gastric biopsy samples. Of the 12 patients, 8 (6 of the 10 Amerindians and the 2 non-Amerindians) were H. pylori glmM positive. Regardless of H. pylori status, the PhyloChip detected Helicobacteriaceae DNA in all patients, although with lower relative abundance in patients who were glmM negative. The G2-chip taxonomy analysis of PhyloChip data indicated the presence of 44 bacterial phyla (of which 16 are unclassified by the Taxonomic Outline of the Bacteria and Archaea taxonomy) in a highly uneven community dominated by only four phyla: Proteobacteria, Firmicutes, Actinobacteria and Bacteroidetes. Positive H. pylori status was associated with increased relative abundance of non-Helicobacter bacteria from the Proteobacteria, Spirochetes and Acidobacteria, and with decreased abundance of Actinobacteria, Bacteroidetes and Firmicutes. The PhyloChip detected richness of low abundance phyla, and showed marked differences in the structure of the gastric bacterial community according to H. pylori status.

Structure of the human gastric bacterial community in relation to Helicobacter pylori status

PubMed Central

Maldonado-Contreras, Ana; Goldfarb, Kate C; Godoy-Vitorino, Filipa; Karaoz, Ulas; Contreras, Mónica; Blaser, Martin J; Brodie, Eoin L; Dominguez-Bello, Maria G

2011-01-01

The human stomach is naturally colonized by Helicobacter pylori, which, when present, dominates the gastric bacterial community. In this study, we aimed to characterize the structure of the bacterial community in the stomach of patients of differing H. pylori status. We used a high-density 16S rRNA gene microarray (PhyloChip, Affymetrix, Inc.) to hybridize 16S rRNA gene amplicons from gastric biopsy DNA of 10 rural Amerindian patients from Amazonas, Venezuela, and of two immigrants to the United States (from South Asia and Africa, respectively). H. pylori status was determined by PCR amplification of H. pylori glmM from gastric biopsy samples. Of the 12 patients, 8 (6 of the 10 Amerindians and the 2 non-Amerindians) were H. pylori glmM positive. Regardless of H. pylori status, the PhyloChip detected Helicobacteriaceae DNA in all patients, although with lower relative abundance in patients who were glmM negative. The G2-chip taxonomy analysis of PhyloChip data indicated the presence of 44 bacterial phyla (of which 16 are unclassified by the Taxonomic Outline of the Bacteria and Archaea taxonomy) in a highly uneven community dominated by only four phyla: Proteobacteria, Firmicutes, Actinobacteria and Bacteroidetes. Positive H. pylori status was associated with increased relative abundance of non-Helicobacter bacteria from the Proteobacteria, Spirochetes and Acidobacteria, and with decreased abundance of Actinobacteria, Bacteroidetes and Firmicutes. The PhyloChip detected richness of low abundance phyla, and showed marked differences in the structure of the gastric bacterial community according to H. pylori status. PMID:20927139

Polyphasic taxonomic characterisation of a novel strain as Pararhizobium haloflavum sp. nov., isolated from soil samples near a sewage treatment tank.

PubMed

Shen, Xia; Li, Yu; Zhao, Zhe; Han, Yi-Fan; Zhang, Wen-Wu; Yu, Xiao-Yun; Zhang, Chong-Ya; Sun, Cong; Wu, Min

2018-04-01

A Gram-stain negative, aerobic, motile and ovoid- to rod-shaped bacteria strain, designated XC0140 T , was isolated from soil samples near the sewage treatment tank of a chemical factory in Zhejiang Province, China, and subjected to polyphasic taxonomic investigation. Strain XC0140 T grew at 10-37 °C and pH 6.0-9.0 (optimum, 35 °C and pH 7.5) and with 0-17% (w/v) NaCl (optimum, 1%). According to phylogenetic analysis based on 16S rRNA gene sequences, strain XC0140 T was assigned to the genus Pararhizobium with high 16S rRNA gene sequence similarity of 95.97% to "Pararhizobium helanshanense CCNWQTX14 T" , followed by Pararhizobium sphaerophysae CCNWGS0238 T (95.95%). Chemotaxonomic analysis showed that strain XC0140 T contains ubiquinone-10 as the predominant respiratory quinone and possessed summed feature 8 (comprising C 18: 1 ω7c and/or ω6c), 11-methyl C 18:1 ω7c, C 18: 0 and C 16: 0 as predominant forms of fatty acids. The polar lipids of strain XC0140 T consisted of seven phospholipids (PL), two aminolipids (AL), one glycolipid (GL) and three unidentified lipids (L1, L2 and L3). The DNA G+C content was 62.7 mol%. Based on the polyphasic taxonomic characterization, strain XC0140 T is considered to represent a novel species of the genus Pararhizobium, for which the name Pararhizobium haloflavum sp. nov. is proposed. (type strain XC0140 T  = MCCC 1K03228 T  = KCTC 52582 T ).

A taxonomic review of the centipede genus Scolopendra Linnaeus, 1758 (Scolopendromorpha, Scolopendridae) in mainland Southeast Asia, with description of a new species from Laos

PubMed Central

Siriwut, Warut; Edgecombe, Gregory D.; Sutcharit, Chirasak; Tongkerd, Piyoros; Panha, Somsak

2016-01-01

Abstract The centipede genus Scolopendra in mainland Southeast Asia is reviewed taxonomically based on morphological characters, informed by a molecular phylogenetic analysis using sequences from three mitochondrial and nuclear genes (COI, 16S rRNA and 28S rRNA). Eight nominal species of Scolopendra, namely Scolopendra morsitans Linnaeus, 1758, Scolopendra subspinipes Leach, 1816, Scolopendra dehaani Brandt, 1840, Scolopendra multidens Newport, 1844, Scolopendra calcarata Porat, 1876, Scolopendra japonica Koch, 1878, Scolopendra pinguis Pocock, 1891, and Scolopendra dawydoffi Kronmüller, 2012, are redescribed together with some revision of type materials. Geographical variation in each species has been compiled with reference to samples that span their distribution ranges in Southeast Asia and some parts of neighbouring areas such as East Asia, the Indian Ocean, and Africa. Comparative study of traditional taxonomic characters from external morphology provides further information to distinguish some closely related species. Scolopendra cataracta Siriwut, Edgecombe & Panha, sp. n., is described from the southern part of Laos, with additional records in Thailand and Vietnam. The phylogenetic framework for Southeast Asian Scolopendra recognizes Scolopendra calcarata + Scolopendra pinguis, Scolopendra morsitans, and a Scolopendra subspinipes group that unites the other six species as the main clades. Within the Scolopendra subspinipes group, two monophyletic groups can be distinguished by having either slender or short, thick ultimate leg prefemora and different numbers of apical spines on the coxopleuron. Scolopendra arborea Lewis, 1982, is placed in subjective synonymy with Scolopendra dehaani. A survey of external morphology of the genital segments confirms its potential for improving species identification in Scolopendra. Some observations on biology and behaviour are recorded based on field surveys in this area. PMID:27408540

Modified RNA-seq method for microbial community and diversity analysis using rRNA in different types of environmental samples

PubMed Central

Yan, Yong-Wei; Zou, Bin; Zhu, Ting; Hozzein, Wael N.

2017-01-01

RNA-seq-based SSU (small subunit) rRNA (ribosomal RNA) analysis has provided a better understanding of potentially active microbial community within environments. However, for RNA-seq library construction, high quantities of purified RNA are typically required. We propose a modified RNA-seq method for SSU rRNA-based microbial community analysis that depends on the direct ligation of a 5’ adaptor to RNA before reverse-transcription. The method requires only a low-input quantity of RNA (10–100 ng) and does not require a DNA removal step. The method was initially tested on three mock communities synthesized with enriched SSU rRNA of archaeal, bacterial and fungal isolates at different ratios, and was subsequently used for environmental samples of high or low biomass. For high-biomass salt-marsh sediments, enriched SSU rRNA and total nucleic acid-derived RNA-seq datasets revealed highly consistent community compositions for all of the SSU rRNA sequences, and as much as 46.4%-59.5% of 16S rRNA sequences were suitable for OTU (operational taxonomic unit)-based community and diversity analyses with complete coverage of V1-V2 regions. OTU-based community structures for the two datasets were also highly consistent with those determined by all of the 16S rRNA reads. For low-biomass samples, total nucleic acid-derived RNA-seq datasets were analyzed, and highly active bacterial taxa were also identified by the OTU-based method, notably including members of the previously underestimated genus Nitrospira and phylum Acidobacteria in tap water, members of the phylum Actinobacteria on a shower curtain, and members of the phylum Cyanobacteria on leaf surfaces. More than half of the bacterial 16S rRNA sequences covered the complete region of primer 8F, and non-coverage rates as high as 38.7% were obtained for phylum-unclassified sequences, providing many opportunities to identify novel bacterial taxa. This modified RNA-seq method will provide a better snapshot of diverse microbial communities, most notably by OTU-based analysis, even communities with low-biomass samples. PMID:29016661

Electrochemical detection of synthetic DNA and native 16S rRNA fragments on a microarray using a biotinylated intercalator as coupling site for an enzyme label.

PubMed

Zimdars, Andreas; Gebala, Magdalena; Hartwich, Gerhard; Neugebauer, Sebastian; Schuhmann, Wolfgang

2015-10-01

The direct electrochemical detection of synthetic DNA and native 16S rRNA fragments isolated from Escherichia coli is described. Oligonucleotides are detected via selective post-labeling of double stranded DNA and DNA-RNA duplexes with a biotinylated intercalator that enables high-specific binding of a streptavidin/alkaline phosphatase conjugate. The alkaline phosphatase catalyzes formation of p-aminophenol that is subsequently oxidized at the underlying gold electrode and hence enables the detection of complementary hybridization of the DNA capture strands due to the enzymatic signal amplification. The hybridization assay was performed on microarrays consisting of 32 individually addressable gold microelectrodes. Synthetic DNA strands with sequences representing six different pathogens which are important for the diagnosis of urinary tract infections could be detected at concentrations of 60 nM. Native 16S rRNA isolated from the different pathogens could be detected at a concentration of 30 fM. Optimization of the sensing surface is described and influences on the assay performance are discussed. Copyright © 2015 Elsevier B.V. All rights reserved.

Comparison of Species Richness Estimates Obtained Using Nearly Complete Fragments and Simulated Pyrosequencing-Generated Fragments in 16S rRNA Gene-Based Environmental Surveys▿ †

PubMed Central

Youssef, Noha; Sheik, Cody S.; Krumholz, Lee R.; Najar, Fares Z.; Roe, Bruce A.; Elshahed, Mostafa S.

2009-01-01

Pyrosequencing-based 16S rRNA gene surveys are increasingly utilized to study highly diverse bacterial communities, with special emphasis on utilizing the large number of sequences obtained (tens to hundreds of thousands) for species richness estimation. However, it is not yet clear how the number of operational taxonomic units (OTUs) and, hence, species richness estimates determined using shorter fragments at different taxonomic cutoffs correlates with the number of OTUs assigned using longer, nearly complete 16S rRNA gene fragments. We constructed a 16S rRNA clone library from an undisturbed tallgrass prairie soil (1,132 clones) and used it to compare species richness estimates obtained using eight pyrosequencing candidate fragments (99 to 361 bp in length) and the nearly full-length fragment. Fragments encompassing the V1 and V2 (V1+V2) region and the V6 region (generated using primer pairs 8F-338R and 967F-1046R) overestimated species richness; fragments encompassing the V3, V7, and V7+V8 hypervariable regions (generated using primer pairs 338F-530R, 1046F-1220R, and 1046F-1392R) underestimated species richness; and fragments encompassing the V4, V5+V6, and V6+V7 regions (generated using primer pairs 530F-805R, 805F-1046R, and 967F-1220R) provided estimates comparable to those obtained with the nearly full-length fragment. These patterns were observed regardless of the alignment method utilized or the parameter used to gauge comparative levels of species richness (number of OTUs observed, slope of scatter plots of pairwise distance values for short and nearly complete fragments, and nonparametric and parametric species richness estimates). Similar results were obtained when analyzing three other datasets derived from soil, adult Zebrafish gut, and basaltic formations in the East Pacific Rise. Regression analysis indicated that these observed discrepancies in species richness estimates within various regions could readily be explained by the proportions of hypervariable, variable, and conserved base pairs within an examined fragment. PMID:19561178

Development of a Custom-Designed, Pan Genomic DNA Microarray to Characterize Strain-Level Diversity among Cronobacter spp.

PubMed Central

Tall, Ben Davies; Gangiredla, Jayanthi; Gopinath, Gopal R.; Yan, Qiongqiong; Chase, Hannah R.; Lee, Boram; Hwang, Seongeun; Trach, Larisa; Park, Eunbi; Yoo, YeonJoo; Chung, TaeJung; Jackson, Scott A.; Patel, Isha R.; Sathyamoorthy, Venugopal; Pava-Ripoll, Monica; Kotewicz, Michael L.; Carter, Laurenda; Iversen, Carol; Pagotto, Franco; Stephan, Roger; Lehner, Angelika; Fanning, Séamus; Grim, Christopher J.

2015-01-01

Cronobacter species cause infections in all age groups; however neonates are at highest risk and remain the most susceptible age group for life-threatening invasive disease. The genus contains seven species:Cronobacter sakazakii, Cronobacter malonaticus, Cronobacter turicensis, Cronobacter muytjensii, Cronobacter dublinensis, Cronobacter universalis, and Cronobacter condimenti. Despite an abundance of published genomes of these species, genomics-based epidemiology of the genus is not well established. The gene content of a diverse group of 126 unique Cronobacter and taxonomically related isolates was determined using a pan genomic-based DNA microarray as a genotyping tool and as a means to identify outbreak isolates for food safety, environmental, and clinical surveillance purposes. The microarray constitutes 19,287 independent genes representing 15 Cronobacter genomes and 18 plasmids and 2,371 virulence factor genes of phylogenetically related Gram-negative bacteria. The Cronobacter microarray was able to distinguish the seven Cronobacter species from one another and from non-Cronobacter species; and within each species, strains grouped into distinct clusters based on their genomic diversity. These results also support the phylogenic divergence of the genus and clearly highlight the genomic diversity among each member of the genus. The current study establishes a powerful platform for further genomics research of this diverse genus, an important prerequisite toward the development of future countermeasures against this foodborne pathogen in the food safety and clinical arenas. PMID:25984509

Yersinia pekkanenii sp. nov.

PubMed

Murros-Kontiainen, Anna; Johansson, Per; Niskanen, Taina; Fredriksson-Ahomaa, Maria; Korkeala, Hannu; Björkroth, Johanna

2011-10-01

The taxonomic position of three strains from water, soil and lettuce samples was studied by using a polyphasic taxonomic approach. The strains were reported to lack the virulence-encoding genes inv and virF in a previous study. Controversially, API 20 E and some other phenotypic tests suggested that the strains belong to Yersinia pseudotuberculosis, which prompted this polyphasic taxonomic study. In both the phylogenetic analyses of four housekeeping genes (glnA, gyrB, recA and HSP60) and numerical analyses of HindIII and EcoRI ribopatterns, the strains formed a separate group within the genus Yersinia. Analysis of the 16S rRNA gene sequences showed that the strains were related to Yersinia aldovae and Yersinia mollaretii, but DNA-DNA hybridization analysis differentiated them from these species. Based on the results of the phylogenetic and DNA-DNA hybridization analyses, a novel species, Yersinia pekkanenii sp. nov., is proposed. The type strain is ÅYV7.1KOH2(T) ( = DSM 22769(T)  = LMG 25369(T)).

Shifts in soil microorganisms in response to warming are consistent across a range of Antarctic environments

PubMed Central

Yergeau, Etienne; Bokhorst, Stef; Kang, Sanghoon; Zhou, Jizhong; Greer, Charles W; Aerts, Rien; Kowalchuk, George A

2012-01-01

Because of severe abiotic limitations, Antarctic soils represent simplified systems, where microorganisms are the principal drivers of nutrient cycling. This relative simplicity makes these ecosystems particularly vulnerable to perturbations, like global warming, and the Antarctic Peninsula is among the most rapidly warming regions on the planet. However, the consequences of the ongoing warming of Antarctica on microorganisms and the processes they mediate are unknown. Here, using 16S rRNA gene pyrosequencing and qPCR, we report highly consistent responses in microbial communities across disparate sub-Antarctic and Antarctic environments in response to 3 years of experimental field warming (+0.5 to 2 °C). Specifically, we found significant increases in the abundance of fungi and bacteria and in the Alphaproteobacteria-to-Acidobacteria ratio, which could result in an increase in soil respiration. Furthermore, shifts toward generalist bacterial communities following warming weakened the linkage between the bacterial taxonomic and functional richness. GeoChip microarray analyses also revealed significant warming effects on functional communities, specifically in the N-cycling microorganisms. Our results demonstrate that soil microorganisms across a range of sub-Antarctic and Antarctic environments can respond consistently and rapidly to increasing temperatures. PMID:21938020

Weissella ghanensis sp. nov., isolated from a Ghanaian cocoa fermentation.

PubMed

De Bruyne, Katrien; Camu, Nicholas; Lefebvre, Karen; De Vuyst, Luc; Vandamme, Peter

2008-12-01

During a study on lactic acid bacteria (and their species diversity) in spontaneous heap fermentations of Ghanaian cocoa beans, two strains, designated 215(T) and 194B, were isolated. A phylogenetic analysis based on 16S rRNA gene sequences demonstrated that these strains represented a distinct lineage close to the genus Weissella and showing only 92.1 % 16S rRNA gene sequence similarity with respect to their closest neighbour, Weissella soli LMG 20113(T). Whole-cell protein electrophoresis, fluorescent amplified fragment length polymorphism fingerprinting of whole genomes and physiological and biochemical tests confirmed the unique taxonomic position of the two novel isolates. On the basis of the results of the morphological and biochemical tests and 16S rRNA gene sequence analysis, strains 215(T) and 194B represent the most peripheral lineage of the genus Weissella, for which we propose the name Weissella ghanensis sp. nov. The type strain is 215(T) (=LMG 24286(T)=DSM 19935(T)).

MetaDP: a comprehensive web server for disease prediction of 16S rRNA metagenomic datasets.

PubMed

Xu, Xilin; Wu, Aiping; Zhang, Xinlei; Su, Mingming; Jiang, Taijiao; Yuan, Zhe-Ming

2016-01-01

High-throughput sequencing-based metagenomics has garnered considerable interest in recent years. Numerous methods and tools have been developed for the analysis of metagenomic data. However, it is still a daunting task to install a large number of tools and complete a complicated analysis, especially for researchers with minimal bioinformatics backgrounds. To address this problem, we constructed an automated software named MetaDP for 16S rRNA sequencing data analysis, including data quality control, operational taxonomic unit clustering, diversity analysis, and disease risk prediction modeling. Furthermore, a support vector machine-based prediction model for intestinal bowel syndrome (IBS) was built by applying MetaDP to microbial 16S sequencing data from 108 children. The success of the IBS prediction model suggests that the platform may also be applied to other diseases related to gut microbes, such as obesity, metabolic syndrome, or intestinal cancer, among others (http://metadp.cn:7001/).

Fetal Tissues Tested for Microbial Sterility by Culture- and PCR-Based Methods Can be Safely Used in Clinics.

PubMed

Vitrenko, Yakov; Kostenko, Iryna; Kulebyakina, Kateryna; Duda, Alla; Klunnyk, Mariya; Sorochynska, Khrystyna

2017-02-16

Cell preparations to be used in clinical practice must be free of infectious agents. Safety concerns are especially elevated upon the use of human fetal tissues, which are otherwise highly advantageous in cell therapy. We demonstrate that treating fetal samples with antibiotic, extensive washing, and homogenization prior to cryoconservation efficiently removes microbes in general. Screening a large collection by an automatic culture system showed that 89.2% fetal tissue samples were sterile, while contamination was detected in 10.8% samples. Liver and chorion were contaminated more than the brain, kidney, lung, and soft tissues. Broad-range PCR from the bacterial 16s rRNA gene was adopted as a confirmatory assay; however, the concordance between the culture-based and PCR assays was weak. Taxonomic identification was done for contaminated samples by bacteriological methods and sequencing 16s rRNA PCR products. The two approaches revealed different spectra of taxonomic groups sharing only Lactobacillus, the most frequently found genus. In addition, other representatives of vaginal microbiota were detected by culture-based identification, while PCR product sequencing has also revealed a subset of nosocomial microorganisms. Importantly, species known to cause sepsis were identified by both techniques, arguing for their indispensability and mutual complementarity. We suggest that most contaminations are taken up during collection of fetal material rather than originating from an in utero infection. In conclusion, a rigorous microbiological control by culture and PCR is a prerequisite for safe clinical use of fetal tissue suspensions.

Bacterial discrimination by means of a universal array approach mediated by LDR (ligase detection reaction)

PubMed Central

Busti, Elena; Bordoni, Roberta; Castiglioni, Bianca; Monciardini, Paolo; Sosio, Margherita; Donadio, Stefano; Consolandi, Clarissa; Rossi Bernardi, Luigi; Battaglia, Cristina; De Bellis, Gianluca

2002-01-01

Background PCR amplification of bacterial 16S rRNA genes provides the most comprehensive and flexible means of sampling bacterial communities. Sequence analysis of these cloned fragments can provide a qualitative and quantitative insight of the microbial population under scrutiny although this approach is not suited to large-scale screenings. Other methods, such as denaturing gradient gel electrophoresis, heteroduplex or terminal restriction fragment analysis are rapid and therefore amenable to field-scale experiments. A very recent addition to these analytical tools is represented by microarray technology. Results Here we present our results using a Universal DNA Microarray approach as an analytical tool for bacterial discrimination. The proposed procedure is based on the properties of the DNA ligation reaction and requires the design of two probes specific for each target sequence. One oligo carries a fluorescent label and the other a unique sequence (cZipCode or complementary ZipCode) which identifies a ligation product. Ligated fragments, obtained in presence of a proper template (a PCR amplified fragment of the 16s rRNA gene) contain either the fluorescent label or the unique sequence and therefore are addressed to the location on the microarray where the ZipCode sequence has been spotted. Such an array is therefore "Universal" being unrelated to a specific molecular analysis. Here we present the design of probes specific for some groups of bacteria and their application to bacterial diagnostics. Conclusions The combined use of selective probes, ligation reaction and the Universal Array approach yielded an analytical procedure with a good power of discrimination among bacteria. PMID:12243651

High-resolution phylogenetic microbial community profiling

DOE Office of Scientific and Technical Information (OSTI.GOV)

Singer, Esther; Bushnell, Brian; Coleman-Derr, Devin

Over the past decade, high-throughput short-read 16S rRNA gene amplicon sequencing has eclipsed clone-dependent long-read Sanger sequencing for microbial community profiling. The transition to new technologies has provided more quantitative information at the expense of taxonomic resolution with implications for inferring metabolic traits in various ecosystems. We applied single-molecule real-time sequencing for microbial community profiling, generating full-length 16S rRNA gene sequences at high throughput, which we propose to name PhyloTags. We benchmarked and validated this approach using a defined microbial community. When further applied to samples from the water column of meromictic Sakinaw Lake, we show that while community structuresmore » at the phylum level are comparable between PhyloTags and Illumina V4 16S rRNA gene sequences (iTags), variance increases with community complexity at greater water depths. PhyloTags moreover allowed less ambiguous classification. Last, a platform-independent comparison of PhyloTags and in silico generated partial 16S rRNA gene sequences demonstrated significant differences in community structure and phylogenetic resolution across multiple taxonomic levels, including a severe underestimation in the abundance of specific microbial genera involved in nitrogen and methane cycling across the Lake's water column. Thus, PhyloTags provide a reliable adjunct or alternative to cost-effective iTags, enabling more accurate phylogenetic resolution of microbial communities and predictions on their metabolic potential.« less

High-resolution phylogenetic microbial community profiling

DOE PAGES

Singer, Esther; Bushnell, Brian; Coleman-Derr, Devin; ...

2016-02-09

Over the past decade, high-throughput short-read 16S rRNA gene amplicon sequencing has eclipsed clone-dependent long-read Sanger sequencing for microbial community profiling. The transition to new technologies has provided more quantitative information at the expense of taxonomic resolution with implications for inferring metabolic traits in various ecosystems. We applied single-molecule real-time sequencing for microbial community profiling, generating full-length 16S rRNA gene sequences at high throughput, which we propose to name PhyloTags. We benchmarked and validated this approach using a defined microbial community. When further applied to samples from the water column of meromictic Sakinaw Lake, we show that while community structuresmore » at the phylum level are comparable between PhyloTags and Illumina V4 16S rRNA gene sequences (iTags), variance increases with community complexity at greater water depths. PhyloTags moreover allowed less ambiguous classification. Last, a platform-independent comparison of PhyloTags and in silico generated partial 16S rRNA gene sequences demonstrated significant differences in community structure and phylogenetic resolution across multiple taxonomic levels, including a severe underestimation in the abundance of specific microbial genera involved in nitrogen and methane cycling across the Lake's water column. Thus, PhyloTags provide a reliable adjunct or alternative to cost-effective iTags, enabling more accurate phylogenetic resolution of microbial communities and predictions on their metabolic potential.« less

The sponge microbiome project.

PubMed

Moitinho-Silva, Lucas; Nielsen, Shaun; Amir, Amnon; Gonzalez, Antonio; Ackermann, Gail L; Cerrano, Carlo; Astudillo-Garcia, Carmen; Easson, Cole; Sipkema, Detmer; Liu, Fang; Steinert, Georg; Kotoulas, Giorgos; McCormack, Grace P; Feng, Guofang; Bell, James J; Vicente, Jan; Björk, Johannes R; Montoya, Jose M; Olson, Julie B; Reveillaud, Julie; Steindler, Laura; Pineda, Mari-Carmen; Marra, Maria V; Ilan, Micha; Taylor, Michael W; Polymenakou, Paraskevi; Erwin, Patrick M; Schupp, Peter J; Simister, Rachel L; Knight, Rob; Thacker, Robert W; Costa, Rodrigo; Hill, Russell T; Lopez-Legentil, Susanna; Dailianis, Thanos; Ravasi, Timothy; Hentschel, Ute; Li, Zhiyong; Webster, Nicole S; Thomas, Torsten

2017-10-01

Marine sponges (phylum Porifera) are a diverse, phylogenetically deep-branching clade known for forming intimate partnerships with complex communities of microorganisms. To date, 16S rRNA gene sequencing studies have largely utilised different extraction and amplification methodologies to target the microbial communities of a limited number of sponge species, severely limiting comparative analyses of sponge microbial diversity and structure. Here, we provide an extensive and standardised dataset that will facilitate sponge microbiome comparisons across large spatial, temporal, and environmental scales. Samples from marine sponges (n = 3569 specimens), seawater (n = 370), marine sediments (n = 65) and other environments (n = 29) were collected from different locations across the globe. This dataset incorporates at least 268 different sponge species, including several yet unidentified taxa. The V4 region of the 16S rRNA gene was amplified and sequenced from extracted DNA using standardised procedures. Raw sequences (total of 1.1 billion sequences) were processed and clustered with (i) a standard protocol using QIIME closed-reference picking resulting in 39 543 operational taxonomic units (OTU) at 97% sequence identity, (ii) a de novo clustering using Mothur resulting in 518 246 OTUs, and (iii) a new high-resolution Deblur protocol resulting in 83 908 unique bacterial sequences. Abundance tables, representative sequences, taxonomic classifications, and metadata are provided. This dataset represents a comprehensive resource of sponge-associated microbial communities based on 16S rRNA gene sequences that can be used to address overarching hypotheses regarding host-associated prokaryotes, including host specificity, convergent evolution, environmental drivers of microbiome structure, and the sponge-associated rare biosphere. © The Authors 2017. Published by Oxford University Press.

Genealogical analyses of multiple loci of litostomatean ciliates (Protista, Ciliophora, Litostomatea)

PubMed Central

Vd’ačný, Peter; Bourland, William A.; Orsi, William; Epstein, Slava S.; Foissner, Wilhelm

2012-01-01

The class Litostomatea is a highly diverse ciliate taxon comprising hundreds of free-living and endocommensal species. However, their traditional morphology-based classification conflicts with 18S rRNA gene phylogenies indicating (1) a deep bifurcation of the Litostomatea into Rhynchostomatia and Haptoria + Trichostomatia, and (2) body polarization and simplification of the oral apparatus as main evolutionary trends in the Litostomatea. To test whether 18S rRNA molecules provide a suitable proxy for litostomatean evolutionary history, we used eighteen new ITS1-5.8S rRNA-ITS2 region sequences from various free-living litostomatean orders. These single- and multiple-locus analyses are in agreement with previous 18S rRNA gene phylogenies, supporting that both 18S rRNA gene and ITS region sequences are effective tools for resolving phylogenetic relationships among the litostomateans. Despite insertions, deletions and mutational saturations in the ITS region, the present study shows that ITS1 and ITS2 molecules can be used to infer phylogenetic relationships not only at species level but also at higher taxonomic ranks when their secondary structure information is utilized to aid alignment. PMID:22789763

Genealogical analyses of multiple loci of litostomatean ciliates (Protista, Ciliophora, Litostomatea).

PubMed

Vd'ačný, Peter; Bourland, William A; Orsi, William; Epstein, Slava S; Foissner, Wilhelm

2012-11-01

The class Litostomatea is a highly diverse ciliate taxon comprising hundreds of free-living and endocommensal species. However, their traditional morphology-based classification conflicts with 18S rRNA gene phylogenies indicating (1) a deep bifurcation of the Litostomatea into Rhynchostomatia and Haptoria+Trichostomatia, and (2) body polarization and simplification of the oral apparatus as main evolutionary trends in the Litostomatea. To test whether 18S rRNA molecules provide a suitable proxy for litostomatean evolutionary history, we used eighteen new ITS1-5.8S rRNA-ITS2 region sequences from various free-living litostomatean orders. These single- and multiple-locus analyses are in agreement with previous 18S rRNA gene phylogenies, supporting that both 18S rRNA gene and ITS region sequences are effective tools for resolving phylogenetic relationships among the litostomateans. Despite insertions, deletions and mutational saturations in the ITS region, the present study shows that ITS1 and ITS2 molecules can be used to infer phylogenetic relationships not only at species level but also at higher taxonomic ranks when their secondary structure information is utilized to aid alignment. Copyright © 2012 Elsevier Inc. All rights reserved.

How close is close: 16S rRNA sequence identity may not be sufficient to guarantee species identity

NASA Technical Reports Server (NTRS)

Fox, G. E.; Wisotzkey, J. D.; Jurtshuk, P. Jr

1992-01-01

16S rRNA (genes coding for rRNA) sequence comparisons were conducted with the following three psychrophilic strains: Bacillus globisporus W25T (T = type strain) and Bacillus psychrophilus W16AT, and W5. These strains exhibited more than 99.5% sequence identity and within experimental uncertainty could be regarded as identical. Their close taxonomic relationship was further documented by phenotypic similarities. In contrast, previously published DNA-DNA hybridization results have convincingly established that these strains do not belong to the same species if current standards are used. These results emphasize the important point that effective identity of 16S rRNA sequences is not necessarily a sufficient criterion to guarantee species identity. Thus, although 16S rRNA sequences can be used routinely to distinguish and establish relationships between genera and well-resolved species, very recently diverged species may not be recognizable.

A New Integrated Approach to Taxonomy: The Fusion of Molecular and Morphological Systematics with Type Material in Benthic Foraminifera

PubMed Central

Roberts, Angela; Austin, William; Evans, Katharine; Bird, Clare; Schweizer, Magali; Darling, Kate

2016-01-01

A robust and consistent taxonomy underpins the use of fossil material in palaeoenvironmental research and long-term assessment of biodiversity. This study presents a new integrated taxonomic protocol for benthic foraminifera by unequivocally reconciling the traditional taxonomic name to a specific genetic type. To implement this protocol, a fragment of the small subunit ribosomal RNA (SSU rRNA) gene is used in combination with 16 quantitative morphometric variables to fully characterise the benthic foraminiferal species concept of Elphidium williamsoni Haynes, 1973. A combination of live contemporary topotypic specimens, original type specimens and specimens of genetic outliers were utilised in this study. Through a series of multivariate statistical tests we illustrate that genetically characterised topotype specimens are morphologically congruent with both the holotype and paratype specimens of E. williamsoni Haynes, 1973. We present the first clear link between morphologically characterised type material and the unique SSU rRNA genetic type of E. williamsoni. This example provides a standard framework for the benthic foraminifera which bridges the current discontinuity between molecular and morphological lines of evidence, allowing integration with the traditional Linnaean roots of nomenclature to offer a new prospect for taxonomic stability. PMID:27388271

Proposals for revival of Streptomyces setonii and reclassification of S. fimicarius as a later synonym of S. setonii and S. albovinaceus as a later synonym of S. globisporus based on combined 16S rRNA-gyrB gene analysis

USDA-ARS?s Scientific Manuscript database

The 16S rRNA and gyrB genes of 22 Streptomyces species belonging to the Streptomyces griseus cluster were sequenced, and their taxonomic positions were re-evaluated. For correct analysis, all of the publicly available sequences of the species were collected and compared with those obtained in this s...

Segal's Law, 16S rRNA gene sequencing, and the perils of foodborne pathogen detection within the American Gut Project.

PubMed

Pettengill, James B; Rand, Hugh

2017-01-01

Obtaining human population level estimates of the prevalence of foodborne pathogens is critical for understanding outbreaks and ameliorating such threats to public health. Estimates are difficult to obtain due to logistic and financial constraints, but citizen science initiatives like that of the American Gut Project (AGP) represent a potential source of information concerning enteric pathogens. With an emphasis on genera Listeria and Salmonella , we sought to document the prevalence of those two taxa within the AGP samples. The results provided by AGP suggest a surprising 14% and 2% of samples contained Salmonella and Listeria , respectively. However, a reanalysis of those AGP sequences described here indicated that results depend greatly on the algorithm for assigning taxonomy and differences persisted across both a range of parameter settings and different reference databases (i.e., Greengenes and HITdb). These results are perhaps to be expected given that AGP sequenced the V4 region of 16S rRNA gene, which may not provide good resolution at the lower taxonomic levels (e.g., species), but it was surprising how often methods differ in classifying reads-even at higher taxonomic ranks (e.g., family). This highlights the misleading conclusions that can be reached when relying on a single method that is not a gold standard; this is the essence of Segal's Law: an individual with one watch knows what time it is but an individual with two is never sure. Our results point to the need for an appropriate molecular marker for the taxonomic resolution of interest, and calls for the development of more conservative classification methods that are fit for purpose. Thus, with 16S rRNA gene datasets, one must be cautious regarding the detection of taxonomic groups of public health interest (e.g., culture independent identification of foodborne pathogens or taxa associated with a given phenotype).

Legionella saoudiensis sp. nov., isolated from a sewage water sample.

PubMed

Bajrai, Leena Hussein; Azhar, Esam Ibraheem; Yasir, Muhammad; Jardot, Priscilla; Barrassi, Lina; Raoult, Didier; La Scola, Bernard; Pagnier, Isabelle

2016-11-01

A Gram-stain-negative, bacilli-shaped bacterial strain, LS-1T, was isolated from a sewage water sample collected in Jeddah, Saudi Arabia. The taxonomic position of strain LS-1T was investigated using a polyphasic taxonomic approach. Phylogenetic analysis based on 16S rRNA gene sequences and those of four other genes indicated that strain LS-1T belongs to the genus Legionella in the family Legionellaceae. Regarding the 16S rRNA gene, the most closely related species are Legionella rowbothamii LLAP-6T (98.6 %) and Legionella lytica L2T (98.5 %). The mip gene sequence of strain LS-1T showed 94 % sequence similarity with that of L. lytica L2T and 93 % similarity with that of L. rowbothamii LLAP-6T. Strain LS-1T grew optimally at a temperature of 32 °C on a buffered charcoal yeast extract (BCYE) agar plate in a 5 % CO2 atmosphere and had a flagellum. The combined phylogenetic, phenotypic and genomic sequence data suggest that strain LS-1T represents a novel species of the genus Legionella, for which the name Legionella saoudiensis sp. nov. is proposed. The type strain is LS-1T (=DSM 101682T=CSUR P2101T).

Identification and characterization of rhizospheric microbial diversity by 16S ribosomal RNA gene sequencing.

PubMed

Naveed, Muhammad; Mubeen, Samavia; Khan, SamiUllah; Ahmed, Iftikhar; Khalid, Nauman; Suleria, Hafiz Ansar Rasul; Bano, Asghari; Mumtaz, Abdul Samad

2014-01-01

In the present study, samples of rhizosphere and root nodules were collected from different areas of Pakistan to isolate plant growth promoting rhizobacteria. Identification of bacterial isolates was made by 16S rRNA gene sequence analysis and taxonomical confirmation on EzTaxon Server. The identified bacterial strains were belonged to 5 genera i.e. Ensifer, Bacillus, Pseudomona, Leclercia and Rhizobium. Phylogenetic analysis inferred from 16S rRNA gene sequences showed the evolutionary relationship of bacterial strains with the respective genera. Based on phylogenetic analysis, some candidate novel species were also identified. The bacterial strains were also characterized for morphological, physiological, biochemical tests and glucose dehydrogenase (gdh) gene that involved in the phosphate solublization using cofactor pyrroloquinolone quinone (PQQ). Seven rhizoshperic and 3 root nodulating stains are positive for gdh gene. Furthermore, this study confirms a novel association between microbes and their hosts like field grown crops, leguminous and non-leguminous plants. It was concluded that a diverse group of bacterial population exist in the rhizosphere and root nodules that might be useful in evaluating the mechanisms behind plant microbial interactions and strains QAU-63 and QAU-68 have sequence similarity of 97 and 95% which might be declared as novel after further taxonomic characterization.

Identification and characterization of rhizospheric microbial diversity by 16S ribosomal RNA gene sequencing

PubMed Central

Naveed, Muhammad; Mubeen, Samavia; khan, SamiUllah; Ahmed, Iftikhar; Khalid, Nauman; Suleria, Hafiz Ansar Rasul; Bano, Asghari; Mumtaz, Abdul Samad

2014-01-01

In the present study, samples of rhizosphere and root nodules were collected from different areas of Pakistan to isolate plant growth promoting rhizobacteria. Identification of bacterial isolates was made by 16S rRNA gene sequence analysis and taxonomical confirmation on EzTaxon Server. The identified bacterial strains were belonged to 5 genera i.e. Ensifer, Bacillus, Pseudomona, Leclercia and Rhizobium. Phylogenetic analysis inferred from 16S rRNA gene sequences showed the evolutionary relationship of bacterial strains with the respective genera. Based on phylogenetic analysis, some candidate novel species were also identified. The bacterial strains were also characterized for morphological, physiological, biochemical tests and glucose dehydrogenase (gdh) gene that involved in the phosphate solublization using cofactor pyrroloquinolone quinone (PQQ). Seven rhizoshperic and 3 root nodulating stains are positive for gdh gene. Furthermore, this study confirms a novel association between microbes and their hosts like field grown crops, leguminous and non-leguminous plants. It was concluded that a diverse group of bacterial population exist in the rhizosphere and root nodules that might be useful in evaluating the mechanisms behind plant microbial interactions and strains QAU-63 and QAU-68 have sequence similarity of 97 and 95% which might be declared as novel after further taxonomic characterization. PMID:25477935

Predicting taxonomic and functional structure of microbial communities in acid mine drainage

PubMed Central

Kuang, Jialiang; Huang, Linan; He, Zhili; Chen, Linxing; Hua, Zhengshuang; Jia, Pu; Li, Shengjin; Liu, Jun; Li, Jintian; Zhou, Jizhong; Shu, Wensheng

2016-01-01

Predicting the dynamics of community composition and functional attributes responding to environmental changes is an essential goal in community ecology but remains a major challenge, particularly in microbial ecology. Here, by targeting a model system with low species richness, we explore the spatial distribution of taxonomic and functional structure of 40 acid mine drainage (AMD) microbial communities across Southeast China profiled by 16S ribosomal RNA pyrosequencing and a comprehensive microarray (GeoChip). Similar environmentally dependent patterns of dominant microbial lineages and key functional genes were observed regardless of the large-scale geographical isolation. Functional and phylogenetic β-diversities were significantly correlated, whereas functional metabolic potentials were strongly influenced by environmental conditions and community taxonomic structure. Using advanced modeling approaches based on artificial neural networks, we successfully predicted the taxonomic and functional dynamics with significantly higher prediction accuracies of metabolic potentials (average Bray–Curtis similarity 87.8) as compared with relative microbial abundances (similarity 66.8), implying that natural AMD microbial assemblages may be better predicted at the functional genes level rather than at taxonomic level. Furthermore, relative metabolic potentials of genes involved in many key ecological functions (for example, nitrogen and phosphate utilization, metals resistance and stress response) were extrapolated to increase under more acidic and metal-rich conditions, indicating a critical strategy of stress adaptation in these extraordinary communities. Collectively, our findings indicate that natural selection rather than geographic distance has a more crucial role in shaping the taxonomic and functional patterns of AMD microbial community that readily predicted by modeling methods and suggest that the model-based approach is essential to better understand natural acidophilic microbial communities. PMID:26943622

Predicting taxonomic and functional structure of microbial communities in acid mine drainage.

PubMed

Kuang, Jialiang; Huang, Linan; He, Zhili; Chen, Linxing; Hua, Zhengshuang; Jia, Pu; Li, Shengjin; Liu, Jun; Li, Jintian; Zhou, Jizhong; Shu, Wensheng

2016-06-01

Predicting the dynamics of community composition and functional attributes responding to environmental changes is an essential goal in community ecology but remains a major challenge, particularly in microbial ecology. Here, by targeting a model system with low species richness, we explore the spatial distribution of taxonomic and functional structure of 40 acid mine drainage (AMD) microbial communities across Southeast China profiled by 16S ribosomal RNA pyrosequencing and a comprehensive microarray (GeoChip). Similar environmentally dependent patterns of dominant microbial lineages and key functional genes were observed regardless of the large-scale geographical isolation. Functional and phylogenetic β-diversities were significantly correlated, whereas functional metabolic potentials were strongly influenced by environmental conditions and community taxonomic structure. Using advanced modeling approaches based on artificial neural networks, we successfully predicted the taxonomic and functional dynamics with significantly higher prediction accuracies of metabolic potentials (average Bray-Curtis similarity 87.8) as compared with relative microbial abundances (similarity 66.8), implying that natural AMD microbial assemblages may be better predicted at the functional genes level rather than at taxonomic level. Furthermore, relative metabolic potentials of genes involved in many key ecological functions (for example, nitrogen and phosphate utilization, metals resistance and stress response) were extrapolated to increase under more acidic and metal-rich conditions, indicating a critical strategy of stress adaptation in these extraordinary communities. Collectively, our findings indicate that natural selection rather than geographic distance has a more crucial role in shaping the taxonomic and functional patterns of AMD microbial community that readily predicted by modeling methods and suggest that the model-based approach is essential to better understand natural acidophilic microbial communities.

Direct profiling of environmental microbial populations by thermal dissociation analysis of native rRNAs hybridized to oligonucleotide microarrays

NASA Technical Reports Server (NTRS)

El Fantroussi, Said; Urakawa, Hidetoshi; Bernhard, Anne E.; Kelly, John J.; Noble, Peter A.; Smidt, H.; Yershov, G. M.; Stahl, David A.

2003-01-01

Oligonucleotide microarrays were used to profile directly extracted rRNA from environmental microbial populations without PCR amplification. In our initial inspection of two distinct estuarine study sites, the hybridization patterns were reproducible and varied between estuarine sediments of differing salinities. The determination of a thermal dissociation curve (i.e., melting profile) for each probe-target duplex provided information on hybridization specificity, which is essential for confirming adequate discrimination between target and nontarget sequences.

Development of the human infant intestinal microbiota.

PubMed

Palmer, Chana; Bik, Elisabeth M; DiGiulio, Daniel B; Relman, David A; Brown, Patrick O

2007-07-01

Almost immediately after a human being is born, so too is a new microbial ecosystem, one that resides in that person's gastrointestinal tract. Although it is a universal and integral part of human biology, the temporal progression of this process, the sources of the microbes that make up the ecosystem, how and why it varies from one infant to another, and how the composition of this ecosystem influences human physiology, development, and disease are still poorly understood. As a step toward systematically investigating these questions, we designed a microarray to detect and quantitate the small subunit ribosomal RNA (SSU rRNA) gene sequences of most currently recognized species and taxonomic groups of bacteria. We used this microarray, along with sequencing of cloned libraries of PCR-amplified SSU rDNA, to profile the microbial communities in an average of 26 stool samples each from 14 healthy, full-term human infants, including a pair of dizygotic twins, beginning with the first stool after birth and continuing at defined intervals throughout the first year of life. To investigate possible origins of the infant microbiota, we also profiled vaginal and milk samples from most of the mothers, and stool samples from all of the mothers, most of the fathers, and two siblings. The composition and temporal patterns of the microbial communities varied widely from baby to baby. Despite considerable temporal variation, the distinct features of each baby's microbial community were recognizable for intervals of weeks to months. The strikingly parallel temporal patterns of the twins suggested that incidental environmental exposures play a major role in determining the distinctive characteristics of the microbial community in each baby. By the end of the first year of life, the idiosyncratic microbial ecosystems in each baby, although still distinct, had converged toward a profile characteristic of the adult gastrointestinal tract.

Winter-summer succession of unicellular eukaryotes in a meso-eutrophic coastal system.

PubMed

Christaki, Urania; Kormas, Konstantinos A; Genitsaris, Savvas; Georges, Clément; Sime-Ngando, Télesphore; Viscogliosi, Eric; Monchy, Sébastien

2014-01-01

The objective of this study was to explore the succession of planktonic unicellular eukaryotes by means of 18S rRNA gene tag pyrosequencing in the eastern English Channel (EEC) during the winter to summer transition. The 59 most representative (>0.1%, representing altogether 95% of total reads), unique operational taxonomic units (OTUs) from all samples belonged to 18 known high-level taxonomic groups and 1 unaffiliated clade. The five most abundant OTUs (69.2% of total reads) belonged to Dinophyceae, Cercozoa, Haptophyceae, marine alveolate group I, and Fungi. Cluster and network analysis between samples distinguished the winter, the pre-bloom, the Phaeocystis globosa bloom and the post-bloom early summer conditions. The OTUs-based network revealed that P. globosa showed a relatively low number of connections-most of them negative-with all other OTUs. Fungi were linked to all major taxonomic groups, except Dinophyceae. Cercozoa mostly co-occurred with the Fungi, the Bacillariophyceae and several of the miscellaneous OTUs. This study provided a more detailed exploration into the planktonic succession pattern of the EEC due to its increased depth of taxonomic sampling over previous efforts based on classical monitoring observations. Data analysis implied that the food web concept in a coastal system based on predator-prey (e.g. grazer-phytoplankton) relationships is just a part of the ecological picture; and those organisms exploiting a variety of strategies, such as saprotrophy and parasitism, are persistent and abundant members of the community.

16S rRNA based microarray analysis of ten periodontal bacteria in patients with different forms of periodontitis.

PubMed

Topcuoglu, Nursen; Kulekci, Guven

2015-10-01

DNA microarray analysis is a computer based technology, that a reverse capture, which targets 10 periodontal bacteria (ParoCheck) is available for rapid semi-quantitative determination. The aim of this three-year retrospective study was to display the microarray analysis results for the subgingival biofilm samples taken from patient cases diagnosed with different forms of periodontitis. A total of 84 patients with generalized aggressive periodontitis (GAP,n:29), generalized chronic periodontitis (GCP, n:25), peri-implantitis (PI,n:14), localized aggressive periodontitis (LAP,n:8) and refractory chronic periodontitis (RP,n:8) were consecutively selected from the archives of the Oral Microbiological Diagnostic Laboratory. The subgingival biofilm samples were analyzed by the microarray-based identification of 10 selected species. All the tested species were detected in the samples. The red complex bacteria were the most prevalent with very high levels in all groups. Fusobacterium nucleatum was detected in all samples at high levels. The green and blue complex bacteria were less prevalent compared with red and orange complex, except Aggregatibacter actinomycetemcomitas was detected in all LAP group. Positive correlations were found within all the red complex bacteria and between red and orange complex bacteria especially in GCP and GAP groups. Parocheck enables to monitoring of periodontal pathogens in all forms of periodontal disease and can be alternative to other guiding and reliable microbiologic tests. Copyright © 2015 Elsevier Ltd. All rights reserved.

Differential growth responses of soil bacterial taxa to carbon substrates of varying chemical recalcitrance

DOE Office of Scientific and Technical Information (OSTI.GOV)

Goldfarb, K.C.; Karaoz, U.; Hanson, C.A.

2011-04-18

Soils are immensely diverse microbial habitats with thousands of co-existing bacterial, archaeal, and fungal species. Across broad spatial scales, factors such as pH and soil moisture appear to determine the diversity and structure of soil bacterial communities. Within any one site however, bacterial taxon diversity is high and factors maintaining this diversity are poorly resolved. Candidate factors include organic substrate availability and chemical recalcitrance, and given that they appear to structure bacterial communities at the phylum level, we examine whether these factors might structure bacterial communities at finer levels of taxonomic resolution. Analyzing 16S rRNA gene composition of nucleotide analog-labeledmore » DNA by PhyloChip microarrays, we compare relative growth rates on organic substrates of increasing chemical recalcitrance of >2,200 bacterial taxa across 43 divisions/phyla. Taxa that increase in relative abundance with labile organic substrates (i.e., glycine, sucrose) are numerous (>500), phylogenetically clustered, and occur predominantly in two phyla (Proteobacteria and Actinobacteria) including orders Actinomycetales, Enterobacteriales, Burkholderiales, Rhodocyclales, Alteromonadales, and Pseudomonadales. Taxa increasing in relative abundance with more chemically recalcitrant substrates (i.e., cellulose, lignin, or tannin-protein) are fewer (168) but more phylogenetically dispersed, occurring across eight phyla and including Clostridiales, Sphingomonadalaes, Desulfovibrionales. Just over 6% of detected taxa, including many Burkholderiales increase in relative abundance with both labile and chemically recalcitrant substrates. Estimates of median rRNA copy number per genome of responding taxa demonstrate that these patterns are broadly consistent with bacterial growth strategies. Taken together, these data suggest that changes in availability of intrinsically labile substrates may result in predictable shifts in soil bacterial composition.« less

Comparative analysis of bacteria associated with different mosses by 16S rRNA and 16S rDNA sequencing.

PubMed

Tian, Yang; Li, Yan Hong

2017-01-01

To understand the differences of the bacteria associated with different mosses, a phylogenetic study of bacterial communities in three mosses was carried out based on 16S rDNA and 16S rRNA sequencing. The mosses used were Hygroamblystegium noterophilum, Entodon compressus and Grimmia montana, representing hygrophyte, shady plant and xerophyte, respectively. In total, the operational taxonomic units (OTUs), richness and diversity were different regardless of the moss species and the library level. All the examined 1183 clones were assigned to 248 OTUs, 56 genera were assigned in rDNA libraries and 23 genera were determined at the rRNA level. Proteobacteria and Bacteroidetes were considered as the most dominant phyla in all the libraries, whereas abundant Actinobacteria and Acidobacteria were detected in the rDNA library of Entodon compressus and approximately 24.7% clones were assigned to Candidate division TM7 in Grimmia montana at rRNA level. The heatmap showed the bacterial profiles derived from rRNA and rDNA were partly overlapping. However, the principle component analysis of all the profiles derived from rDNA showed sharper differences between the different mosses than that of rRNA-based profiles. This suggests that the metabolically active bacterial compositions in different mosses were more phylogenetically similar and the differences of the bacteria associated with different mosses were mainly detected at the rDNA level. Obtained results clearly demonstrate that combination of 16S rDNA and 16S rRNA sequencing is preferred approach to have a good understanding on the constitution of the microbial communities in mosses. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Seahorses of the Hippocampus coronatus complex: taxonomic revision, and description of Hippocampus haema, a new species from Korea and Japan (Teleostei, Syngnathidae).

PubMed

Han, Sang-Yun; Kim, Jin-Koo; Kai, Yoshiaki; Senou, Hiroshi

2017-01-01

Morphological and molecular analyses were conducted on 182 specimens belonging to the Hippocampus coronatus complex ( H. coronatus sensu lato), collected in Korea and Japan 1933-2015, in order to clarify the taxonomic status of the species within this complex. Three species are recognized based on the shape of the coronet, the number of trunk rings (TrR) and tail rings (TaR), and presence or absence of a wing-tip spine (WS) at the dorsal fin base. Hippocampus coronatus Temminck & Schlegel, 1850 ( H. coronatus sensu stricto), is diagnosed by 10 TrR, 37-40 TaR, an extremely high coronet (55.7-79.0 % head length) with four tips on the corona flat (CoT), and one WS. Hippocampus sindonis Jordan & Snyder, 1901 is diagnosed by 10 TrR, 35-38 TaR, a moderately high coronet (36.3-55.4 % HL) with five CoT, and no WS. A new species, H. haema is described on the basis of 140 specimens, characterized by 10 TrR, 35-38 TaR, a moderately high coronet (34.1-54.9 % head length) with four CoT, and two WS. Hippocampus haema is only known from the Korea Strait, western Kyushu, and East/Japan Sea. Recognition of the three species is supported by differences in mitochondrial DNA fragments (cytochrome b , 16S rRNA, and 12S rRNA).

Soil biochar amendment shapes the composition of N2O-reducing microbial communities.

PubMed

Harter, Johannes; Weigold, Pascal; El-Hadidi, Mohamed; Huson, Daniel H; Kappler, Andreas; Behrens, Sebastian

2016-08-15

Soil biochar amendment has been described as a promising tool to improve soil quality, sequester carbon, and mitigate nitrous oxide (N2O) emissions. N2O is a potent greenhouse gas. The main sources of N2O in soils are microbially-mediated nitrogen transformation processes such as nitrification and denitrification. While previous studies have focused on the link between N2O emission mitigation and the abundance and activity of N2O-reducing microorganisms in biochar-amended soils, the impact of biochar on the taxonomic composition of the nosZ gene carrying soil microbial community has not been subject of systematic study to date. We used 454 pyrosequencing in order to study the microbial diversity in biochar-amended and biochar-free soil microcosms. We sequenced bacterial 16S rRNA gene amplicons as well as fragments of common (typical) nosZ genes and the recently described 'atypical' nosZ genes. The aim was to describe biochar-induced shifts in general bacterial community diversity and taxonomic variations among the nosZ gene containing N2O-reducing microbial communities. While soil biochar amendment significantly altered the 16S rRNA gene-based community composition and structure, it also led to the development of distinct functional traits capable of N2O reduction containing typical and atypical nosZ genes related to nosZ genes found in Pseudomonas stutzeri and Pedobacter saltans, respectively. Our results showed that biochar amendment can affect the relative abundance and taxonomic composition of N2O-reducing functional microbial traits in soil. Thus these findings broaden our knowledge on the impact of biochar on soil microbial community composition and nitrogen cycling. Copyright © 2016 Elsevier B.V. All rights reserved.

CloVR-ITS: Automated internal transcribed spacer amplicon sequence analysis pipeline for the characterization of fungal microbiota

PubMed Central

2013-01-01

Background Besides the development of comprehensive tools for high-throughput 16S ribosomal RNA amplicon sequence analysis, there exists a growing need for protocols emphasizing alternative phylogenetic markers such as those representing eukaryotic organisms. Results Here we introduce CloVR-ITS, an automated pipeline for comparative analysis of internal transcribed spacer (ITS) pyrosequences amplified from metagenomic DNA isolates and representing fungal species. This pipeline performs a variety of steps similar to those commonly used for 16S rRNA amplicon sequence analysis, including preprocessing for quality, chimera detection, clustering of sequences into operational taxonomic units (OTUs), taxonomic assignment (at class, order, family, genus, and species levels) and statistical analysis of sample groups of interest based on user-provided information. Using ITS amplicon pyrosequencing data from a previous human gastric fluid study, we demonstrate the utility of CloVR-ITS for fungal microbiota analysis and provide runtime and cost examples, including analysis of extremely large datasets on the cloud. We show that the largest fractions of reads from the stomach fluid samples were assigned to Dothideomycetes, Saccharomycetes, Agaricomycetes and Sordariomycetes but that all samples were dominated by sequences that could not be taxonomically classified. Representatives of the Candida genus were identified in all samples, most notably C. quercitrusa, while sequence reads assigned to the Aspergillus genus were only identified in a subset of samples. CloVR-ITS is made available as a pre-installed, automated, and portable software pipeline for cloud-friendly execution as part of the CloVR virtual machine package (http://clovr.org). Conclusion The CloVR-ITS pipeline provides fungal microbiota analysis that can be complementary to bacterial 16S rRNA and total metagenome sequence analysis allowing for more comprehensive studies of environmental and host-associated microbial communities. PMID:24451270

Detection of Porphyromonas endodontalis, Porphyromonas gingivalis and Prevotella intermedia in primary endodontic infections in a Chinese population.

PubMed

Cao, H; Qi, Z; Jiang, H; Zhao, J; Liu, Z; Tang, Z

2012-08-01

To assess the prevalence of three black-pigmented bacterial species (Porphyromonas endodontalis, Porphyromonas gingivalis and Prevotella intermedia) using microarray technology in root canals of teeth associated with primary endodontic infections in a Chinese population. Microbial samples were taken from root canals of 80 teeth with pulp necrosis and primary endodontic infections in a Chinese population. DNA extracted from the samples was amplified by PCR with universal bacterial primers based on 16S rRNA gene sequences, and the products hybridized with the microarrays in which the specific oligonucleotide probes were added. The results of hybridization were screened by a confocal laser scanner. Pearson chi-square test and the two-sided Fisher exact test were used to analyse whether a significant association existed between the species and symptoms as well as in co-existence of two target organisms by a statistical software package (SAS 8.02). The 16S rRNA gene microarray detected at least one of the three test species in 76% of the study teeth. P. endodontalis, P. gingivalis and P. intermedia were found in 50%, 33% and 45%, respectively. A significant association was found in the presence of the pair P. endodontalis / P. gingivalis (P < 0.005). Both P. endodontalis (P <0.05) and P. gingivalis (P <0.005) had a statistically significant association with the presence of a sinus tract. The simultaneous presence of P. endodontalis and P. gingivalis was also associated with the presence of a sinus tract (P<0.005) and abscess formation (P<0.05). The three black-pigmented bacteria were prevalent in teeth with pulp necrosis and primary endodontic infections in a Chinese population. P. gingivalis and P. endodontalis were associated with the presence of sinus tract and abscess formation. © 2012 International Endodontic Journal.

Diversity and potential activity patterns of planktonic eukaryotic microbes in a mesoeutrophic coastal area (eastern English Channel)

PubMed Central

Rachik, Sara; Christaki, Urania; Li, Luen Luen; Genitsaris, Savvas; Breton, Elsa

2018-01-01

The diversity of planktonic eukaryotic microbes was studied at a coastal station of the eastern English Channel (EEC) from March 2011 to July 2015 (77 samples) using high throughput sequencing (454-pyrosequencing and Illumina) of the V2-V3 hypervariable region of the 18S SSU rDNA gene. Similar estimations of OTU relative abundance and taxonomic distribution for the dominant higher taxonomic groups (contributing >1% of the total number of OTUs) were observed with the two methods (Kolmogorov-Smirnov p-value = 0.22). Eight super-groups were identified throughout all samples: Alveolata, Stramenopiles, Opisthokonta, Hacrobia, Archeaplastida, Apusozoa, Rhizaria, and Amoebozoa (ordered by decreasing OTU richness). To gain further insight into microbial activity in the EEC, ribosomal RNA was extracted for samples from 2013–2015 (30 samples). Analysis of 18S rDNA and rRNA sequences led to the detection of 696 and 700 OTUs, respectively. Cluster analysis based on OTUs’ abundance indicated three major seasonal groups that were associated to spring, winter/autumn, and summer conditions. The clusters inferred from rRNA data showed a clearer seasonal representation of the community succession than the one based on rDNA. The rRNA/rDNA ratio was used as a proxy for relative cell activity. When all OTUs were considered, the average rRNA:rDNA ratio showed a linear trend around the 1:1 line, suggesting a linear relation between OTU abundance (rDNA) and activity (rRNA). However, this ratio was highly variable over time when considering individual OTUs. Interestingly, the OTU affiliated with P. globosa displayed rRNA:rDNA ratio that allowed to delimit high vs low abundance and high vs low activity periods. It unveiled quite well the Phaeocystis bloom dynamic regarding cell proliferation and activity, and could even be used as early indicator of an upcoming bloom. PMID:29746519

Diversity and potential activity patterns of planktonic eukaryotic microbes in a mesoeutrophic coastal area (eastern English Channel).

PubMed

Rachik, Sara; Christaki, Urania; Li, Luen Luen; Genitsaris, Savvas; Breton, Elsa; Monchy, Sébastien

2018-01-01

The diversity of planktonic eukaryotic microbes was studied at a coastal station of the eastern English Channel (EEC) from March 2011 to July 2015 (77 samples) using high throughput sequencing (454-pyrosequencing and Illumina) of the V2-V3 hypervariable region of the 18S SSU rDNA gene. Similar estimations of OTU relative abundance and taxonomic distribution for the dominant higher taxonomic groups (contributing >1% of the total number of OTUs) were observed with the two methods (Kolmogorov-Smirnov p-value = 0.22). Eight super-groups were identified throughout all samples: Alveolata, Stramenopiles, Opisthokonta, Hacrobia, Archeaplastida, Apusozoa, Rhizaria, and Amoebozoa (ordered by decreasing OTU richness). To gain further insight into microbial activity in the EEC, ribosomal RNA was extracted for samples from 2013-2015 (30 samples). Analysis of 18S rDNA and rRNA sequences led to the detection of 696 and 700 OTUs, respectively. Cluster analysis based on OTUs' abundance indicated three major seasonal groups that were associated to spring, winter/autumn, and summer conditions. The clusters inferred from rRNA data showed a clearer seasonal representation of the community succession than the one based on rDNA. The rRNA/rDNA ratio was used as a proxy for relative cell activity. When all OTUs were considered, the average rRNA:rDNA ratio showed a linear trend around the 1:1 line, suggesting a linear relation between OTU abundance (rDNA) and activity (rRNA). However, this ratio was highly variable over time when considering individual OTUs. Interestingly, the OTU affiliated with P. globosa displayed rRNA:rDNA ratio that allowed to delimit high vs low abundance and high vs low activity periods. It unveiled quite well the Phaeocystis bloom dynamic regarding cell proliferation and activity, and could even be used as early indicator of an upcoming bloom.

Parallel changes of taxonomic interaction networks in lacustrine bacterial communities induced by a polymetallic perturbation

PubMed Central

Laplante, Karine; Sébastien, Boutin; Derome, Nicolas

2013-01-01

Heavy metals released by anthropogenic activities such as mining trigger profound changes to bacterial communities. In this study we used 16S SSU rRNA gene high-throughput sequencing to characterize the impact of a polymetallic perturbation and other environmental parameters on taxonomic networks within five lacustrine bacterial communities from sites located near Rouyn-Noranda, Quebec, Canada. The results showed that community equilibrium was disturbed in terms of both diversity and structure. Moreover, heavy metals, especially cadmium combined with water acidity, induced parallel changes among sites via the selection of resistant OTUs (Operational Taxonomic Unit) and taxonomic dominance perturbations favoring the Alphaproteobacteria. Furthermore, under a similar selective pressure, covariation trends between phyla revealed conservation and parallelism within interphylum interactions. Our study sheds light on the importance of analyzing communities not only from a phylogenetic perspective but also including a quantitative approach to provide significant insights into the evolutionary forces that shape the dynamic of the taxonomic interaction networks in bacterial communities. PMID:23789031

DNA microarray-based detection and identification of Burkholderia mallei, Burkholderia pseudomallei and Burkholderia spp.

PubMed

Schmoock, Gernot; Ehricht, Ralf; Melzer, Falk; Rassbach, Astrid; Scholz, Holger C; Neubauer, Heinrich; Sachse, Konrad; Mota, Rinaldo Aparecido; Saqib, Muhammad; Elschner, Mandy

2009-01-01

We developed a rapid oligonucleotide microarray assay based on genetic markers for the accurate identification and differentiation of Burkholderia (B.) mallei and Burkholderia pseudomallei, the agents of glanders and melioidosis, respectively. These two agents were clearly identified using at least 4 independent genetic markers including 16S rRNA gene, fliC, motB and also by novel species-specific target genes, identified by in silico sequence analysis. Specific hybridization signal profiles allowed the detection and differentiation of up to 10 further Burkholderia spp., including the closely related species Burkholderia thailandensis and Burkholderia-like agents, such as Burkholderia cepacia, Burkholderia cenocepacia, Burkholderia vietnamiensis, Burkholderia ambifaria, and Burkholderia gladioli, which are often associated with cystic fibrosis (CF) lung disease. The assay was developed using the easy-to-handle and economical ArrayTube (AT) platform. A representative strain panel comprising 44 B. mallei, 32 B. pseudomallei isolates, and various Burkholderia type strains were examined to validate the test. Assay specificity was determined by examination of 40 non-Burkholderia strains.

High-resolution microbiome profiling uncovers Fusobacterium nucleatum, Lactobacillus gasseri/johnsonii, and Lactobacillus vaginalis associated to oral and oropharyngeal cancer in saliva from HPV positive and HPV negative patients treated with surgery and chemo-radiation

PubMed Central

Guerrero-Preston, Rafael; White, James Robert; Godoy-Vitorino, Filipa; Rodríguez-Hilario, Arnold; Navarro, Kelvin; González, Herminio; Michailidi, Christina; Jedlicka, Anne; Canapp, Sierra; Bondy, Jessica; Dziedzic, Amanda; Mora-Lagos, Barbara; Rivera-Alvarez, Gustavo; Ili-Gangas, Carmen; Brebi-Mieville, Priscilla; Westra, William; Koch, Wayne; Kang, Hyunseok; Marchionni, Luigi; Kim, Young; Sidransky, David

2017-01-01

Microbiome studies show altered microbiota in head and neck squamous cell carcinoma (HNSCC), both in terms of taxonomic composition and metabolic capacity. These studies utilized a traditional bioinformatics methodology, which allows for accurate taxonomic assignment down to the genus level, but cannot accurately resolve species level membership. We applied Resphera Insight, a high-resolution methodology for 16S rRNA taxonomic assignment that is able to provide species-level context in its assignments of 16S rRNA next generation sequencing (NGS) data. Resphera Insight applied to saliva samples from HNSCC patients and healthy controls led to the discovery that a subset of HNSCC saliva samples is significantly enriched with commensal species from the vaginal flora, including Lactobacillus gasseri/johnsonii (710x higher in saliva) and Lactobacillus vaginalis (52x higher in saliva). These species were not observed in normal saliva from Johns Hopkins patients, nor in 16S rRNA NGS saliva samples from the Human Microbiome Project (HMP). Interestingly, both species were only observed in saliva from Human Papilloma Virus (HPV) positive and HPV negative oropharyngeal cancer patients. We confirmed the representation of both species in HMP data obtained from mid-vagina (n=128) and vaginal introitus (n=121) samples. Resphera Insight also led to the discovery that Fusobacterium nucleatum, an oral cavity flora commensal bacterium linked to colon cancer, is enriched (600x higher) in saliva from a subset of HNSCC patients with advanced tumors stages. Together, these high-resolution analyses on 583 samples suggest a possible role for bacterial species in the therapeutic outcome of HPV positive and HPV negative HNSCC patients. PMID:29340028

High-resolution microbiome profiling uncovers Fusobacterium nucleatum, Lactobacillus gasseri/johnsonii, and Lactobacillus vaginalis associated to oral and oropharyngeal cancer in saliva from HPV positive and HPV negative patients treated with surgery and chemo-radiation.

PubMed

Guerrero-Preston, Rafael; White, James Robert; Godoy-Vitorino, Filipa; Rodríguez-Hilario, Arnold; Navarro, Kelvin; González, Herminio; Michailidi, Christina; Jedlicka, Anne; Canapp, Sierra; Bondy, Jessica; Dziedzic, Amanda; Mora-Lagos, Barbara; Rivera-Alvarez, Gustavo; Ili-Gangas, Carmen; Brebi-Mieville, Priscilla; Westra, William; Koch, Wayne; Kang, Hyunseok; Marchionni, Luigi; Kim, Young; Sidransky, David

2017-12-19

Microbiome studies show altered microbiota in head and neck squamous cell carcinoma (HNSCC), both in terms of taxonomic composition and metabolic capacity. These studies utilized a traditional bioinformatics methodology, which allows for accurate taxonomic assignment down to the genus level, but cannot accurately resolve species level membership. We applied Resphera Insight, a high-resolution methodology for 16S rRNA taxonomic assignment that is able to provide species-level context in its assignments of 16S rRNA next generation sequencing (NGS) data. Resphera Insight applied to saliva samples from HNSCC patients and healthy controls led to the discovery that a subset of HNSCC saliva samples is significantly enriched with commensal species from the vaginal flora, including Lactobacillus gasseri/johnsonii (710x higher in saliva) and Lactobacillus vaginalis (52x higher in saliva). These species were not observed in normal saliva from Johns Hopkins patients, nor in 16S rRNA NGS saliva samples from the Human Microbiome Project (HMP). Interestingly, both species were only observed in saliva from Human Papilloma Virus (HPV) positive and HPV negative oropharyngeal cancer patients. We confirmed the representation of both species in HMP data obtained from mid-vagina (n=128) and vaginal introitus (n=121) samples. Resphera Insight also led to the discovery that Fusobacterium nucleatum , an oral cavity flora commensal bacterium linked to colon cancer, is enriched (600x higher) in saliva from a subset of HNSCC patients with advanced tumors stages. Together, these high-resolution analyses on 583 samples suggest a possible role for bacterial species in the therapeutic outcome of HPV positive and HPV negative HNSCC patients.

Use of rpoB gene analysis for identification of nitrogen-fixing Paenibacillus species as an alternative to the 16S rRNA gene.

PubMed

da Mota, F F; Gomes, E A; Paiva, E; Rosado, A S; Seldin, L

2004-01-01

To avoid the limitations of 16S rRNA-based phylogenetic analysis for Paenibacillus species, the usefulness of the RNA polymerase beta-subunit encoding gene (rpoB) was investigated as an alternative to the 16S rRNA gene for taxonomic studies. Partial rpoB sequences were generated for the type strains of eight nitrogen-fixing Paenibacillus species. The presence of only one copy of rpoB in the genome of P. graminis strain RSA19(T) was demonstrated by denaturing gradient gel electrophoresis and hybridization assays. A comparative analysis of the sequences of the 16S rRNA and rpoB genes was performed and the eight species showed between 91.6-99.1% (16S rRNA) and 77.9-97.3% (rpoB) similarity, allowing a more accurate discrimination between the different species using the rpoB gene. Finally, 24 isolates from the rhizosphere of different cultivars of maize previously identified as Paenibacillus spp. were assigned correctly to one of the nitrogen-fixing species. The data obtained in this study indicate that rpoB is a powerful identification tool, which can be used for the correct discrimination of the nitrogen-fixing species of agricultural and industrial importance within the genus Paenibacillus.

Rare but active taxa contribute to community dynamics of benthic biofilms in glacier-fed streams.

PubMed

Wilhelm, Linda; Besemer, Katharina; Fasching, Christina; Urich, Tim; Singer, Gabriel A; Quince, Christopher; Battin, Tom J

2014-08-01

Glaciers harbour diverse microorganisms, which upon ice melt can be released downstream. In glacier-fed streams microorganisms can attach to stones or sediments to form benthic biofilms. We used 454-pyrosequencing to explore the bulk (16S rDNA) and putatively active (16S rRNA) microbial communities of stone and sediment biofilms across 26 glacier-fed streams. We found differences in community composition between bulk and active communities among streams and a stronger congruence between biofilm types. Relative abundances of rRNA and rDNA were positively correlated across different taxa and taxonomic levels, but at lower taxonomic levels, the higher abundance in either the active or the bulk communities became more apparent. Here, environmental variables played a minor role in structuring active communities. However, we found a large number of rare taxa with higher relative abundances in rRNA compared with rDNA. This suggests that rare taxa contribute disproportionately to microbial community dynamics in glacier-fed streams. Our findings propose that high community turnover, where taxa repeatedly enter and leave the 'seed bank', contributes to the maintenance of microbial biodiversity in harsh ecosystems with continuous environmental perturbations, such as glacier-fed streams. © 2014 The Authors. Environmental Microbiology published by Society for Applied Microbiology and John Wiley & Sons Ltd.

Phylogenetic and Functional Diversity of Microbial Communities Associated with Subsurface Sediments of the Sonora Margin, Guaymas Basin

PubMed Central

Vigneron, Adrien; Cruaud, Perrine; Roussel, Erwan G.; Pignet, Patricia; Caprais, Jean-Claude; Callac, Nolwenn; Ciobanu, Maria-Cristina; Godfroy, Anne; Cragg, Barry A.; Parkes, John R.; Van Nostrand, Joy D.; He, Zhili; Zhou, Jizhong; Toffin, Laurent

2014-01-01

Subsurface sediments of the Sonora Margin (Guaymas Basin), located in proximity of active cold seep sites were explored. The taxonomic and functional diversity of bacterial and archaeal communities were investigated from 1 to 10 meters below the seafloor. Microbial community structure and abundance and distribution of dominant populations were assessed using complementary molecular approaches (Ribosomal Intergenic Spacer Analysis, 16S rRNA libraries and quantitative PCR with an extensive primers set) and correlated to comprehensive geochemical data. Moreover the metabolic potentials and functional traits of the microbial community were also identified using the GeoChip functional gene microarray and metabolic rates. The active microbial community structure in the Sonora Margin sediments was related to deep subsurface ecosystems (Marine Benthic Groups B and D, Miscellaneous Crenarchaeotal Group, Chloroflexi and Candidate divisions) and remained relatively similar throughout the sediment section, despite defined biogeochemical gradients. However, relative abundances of bacterial and archaeal dominant lineages were significantly correlated with organic carbon quantity and origin. Consistently, metabolic pathways for the degradation and assimilation of this organic carbon as well as genetic potentials for the transformation of detrital organic matters, hydrocarbons and recalcitrant substrates were detected, suggesting that chemoorganotrophic microorganisms may dominate the microbial community of the Sonora Margin subsurface sediments. PMID:25099369

How Much Do rRNA Gene Surveys Underestimate Extant Bacterial Diversity?

PubMed

Rodriguez-R, Luis M; Castro, Juan C; Kyrpides, Nikos C; Cole, James R; Tiedje, James M; Konstantinidis, Konstantinos T

2018-03-15

The most common practice in studying and cataloguing prokaryotic diversity involves the grouping of sequences into operational taxonomic units (OTUs) at the 97% 16S rRNA gene sequence identity level, often using partial gene sequences, such as PCR-generated amplicons. Due to the high sequence conservation of rRNA genes, organisms belonging to closely related yet distinct species may be grouped under the same OTU. However, it remains unclear how much diversity has been underestimated by this practice. To address this question, we compared the OTUs of genomes defined at the 97% or 98.5% 16S rRNA gene identity level against OTUs of the same genomes defined at the 95% whole-genome average nucleotide identity (ANI), which is a much more accurate proxy for species. Our results show that OTUs resulting from a 98.5% 16S rRNA gene identity cutoff are more accurate than 97% compared to 95% ANI (90.5% versus 89.9% accuracy) but indistinguishable from any other threshold in the 98.29 to 98.78% range. Even with the more stringent thresholds, however, the 16S rRNA gene-based approach commonly underestimates the number of OTUs by ∼12%, on average, compared to the ANI-based approach (∼14% underestimation when using the 97% identity threshold). More importantly, the degree of underestimation can become 50% or more for certain taxa, such as the genera Pseudomonas , Burkholderia , Escherichia , Campylobacter , and Citrobacter These results provide a quantitative view of the degree of underestimation of extant prokaryotic diversity by 16S rRNA gene-defined OTUs and suggest that genomic resolution is often necessary. IMPORTANCE Species diversity is one of the most fundamental pieces of information for community ecology and conservational biology. Therefore, employing accurate proxies for what a species or the unit of diversity is are cornerstones for a large set of microbial ecology and diversity studies. The most common proxies currently used rely on the clustering of 16S rRNA gene sequences at some threshold of nucleotide identity, typically 97% or 98.5%. Here, we explore how well this strategy reflects the more accurate whole-genome-based proxies and determine the frequency with which the high conservation of 16S rRNA sequences masks substantial species-level diversity. Copyright © 2018 American Society for Microbiology.

Activity and bacterial diversity of snow around Russian Antarctic stations.

PubMed

Lopatina, Anna; Krylenkov, Vjacheslav; Severinov, Konstantin

2013-11-01

The diversity and temporal dynamics of bacterial communities in pristine snow around two Russian Antarctic stations was investigated. Taxonomic analysis of rDNA libraries revealed that snow communities were dominated by bacteria from a small number of operational taxonomic units (OTUs) that underwent dramatic swings in abundance between the 54th (2008-2009) and 55th (2009-2010) Russian Antarctic expeditions. Moreover, analysis of the 55th expedition samples indicated that there was very little, if any, correspondence in abundance of clones belonging to the same OTU present in rDNA and rRNA libraries. The latter result suggests that most rDNA clones originate from bacteria that are not alive and/or active and may have been deposited on the snow surface from the atmosphere. In contrast, clones most abundant in rRNA libraries (mostly belonging to Variovorax, Janthinobacterium, Pseudomonas, and Sphingomonas genera) may be considered as endogenous Antarctic snow inhabitants. Copyright © 2013 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

Re-examination of the taxonomic position of Bacillus silvestris Rheims et al. 1999 and proposal to transfer it to Solibacillus gen. nov. as Solibacillus silvestris comb. nov.

PubMed

Krishnamurthi, Srinivasan; Chakrabarti, Tapan; Stackebrandt, Erko

2009-05-01

Following the transfer of three of the six species enclosed in the original definition of rRNA group 2 of Bacillus to the genus Sporosarcina and two to Lysinibacillus, other species of this group, some of which were added later, still await taxonomic revision. In a recent publication, a set of 'core' characteristics was proposed for species to be included in the genus Bacillus (Kämpfer et al., 2006). Except for Bacillus silvestris, however, several or none of these properties are available for members of rRNA group 2. According to our analysis of data including the 'core' characteristics, Bacillus silvestris should not be a member of the genus Bacillus. We therefore propose the establishment of a new genus, Solibacillus gen. nov., and transfer Bacillus silvestris to this genus as Solibacillus silvestris comb. nov., with the type strain HR3-23(T) (=DSM 12223(T)=ATCC BAA-269(T)=CIP 106059(T)).

25S ribosomal RNA homologies of basidiomycetous yeasts: taxonomic and phylogenetic implications

NASA Technical Reports Server (NTRS)

Baharaeen, S.; Vishniac, H. S.

1984-01-01

Genera, families, and possibly orders of basidiomycetous yeasts can be defined by 25S rRNA homology and correlated phenotypic characters. The teleomorphic genera Filobasidium, Leucosporidium, and Rhodosporidium have greater than 96 relative binding percent (rb%) intrageneric 25S rRNA homology and significant intergeneric separation from each other and from Filobasidiella. The anamorphic genus Cryptococcus can be defined by morphology (monopolar budding), colony color, and greater than 75 rb% intrageneric homology; Vanrija is heterogeneous. Agaricostilbum (Phragmobasidiomycetes, Auriculariales), Hansenula (Ascomycotera, Endomycota), Tremella (Phragmobasidiomycetes, Tremellales), and Ustilago (Ustomycota, Ustilaginales) appear equally unrelated to the Cryptococcus, Filobasidiella, and Rhodosporidium spp. used as probes. The Filobasidiaceae and Sporidiaceae, Filobasidiales and Sporidiales, form coherent homology groups which appear to have undergone convergent 25S rRNA evolution, since their relatedness is much greater than that indicated by 5S rRNA homology. Ribosomal RNA homologies do not appear to measure evolutionary distance.

Taxonomic and functional metagenomic profiling of gastrointestinal tract microbiome of the farmed adult turbot (Scophthalmus maximus).

PubMed

Xing, Mengxin; Hou, Zhanhui; Yuan, Jianbo; Liu, Yuan; Qu, Yanmei; Liu, Bin

2013-12-01

Metagenomics combined with 16S rRNA gene sequence analyses was applied to unveil the taxonomic composition and functional diversity of the farmed adult turbot gastrointestinal (GI) microbiome. Proteobacteria and Firmicutes which existed in both GI content and mucus were dominated in the turbot GI microbiome. 16S rRNA gene sequence analyses also indicated that the turbot GI tract may harbor some bacteria which originated from associated seawater. Functional analyses indicated that the clustering-based subsystem and many metabolic subsystems were dominant in the turbot GI metagenome. Compared with other gut metagenomes, quorum sensing and biofilm formation was overabundant in the turbot GI metagenome. Genes associated with quorum sensing and biofilm formation were found in species within Vibrio, including Vibrio vulnificus, Vibrio cholerae and Vibrio parahaemolyticus. In farmed fish gut metagenomes, the stress response and protein folding subsystems were over-represented and several genes concerning antibiotic and heavy metal resistance were also detected. These data suggested that the turbot GI microbiome may be affected by human factors in aquaculture. Additionally, iron acquisition and the metabolism subsystem were more abundant in the turbot GI metagenome when compared with freshwater fish gut metagenome, suggesting that unique metabolic potential may be observed in marine animal GI microbiomes. © 2013 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

Bacillus nanhaiisediminis sp. nov., an alkalitolerant member of Bacillus rRNA group 6.

PubMed

Zhang, Jianli; Wang, Jiewei; Song, Fei; Fang, Caiyuan; Xin, Yuhua; Zhang, Yabo

2011-05-01

A Gram-stain-positive, rod-shaped bacterium, designated strain NH3(T), was isolated from a sediment sample from the South China Sea and was subjected to a polyphasic taxonomic study. The isolate grew optimally at 37 °C and pH 9. Strain NH3(T) had cell-wall peptidoglycan based on meso-diaminopimelic acid and MK-7 as the predominant menaquinone. The cellular fatty acid profile included significant amounts of iso-C(15 : 0) and iso-C(14 : 0). The major polar lipids were phosphatidylethanolamine, phosphatidylglycerol and diphosphatidylglycerol. The DNA G+C content of strain NH3(T) was 40.3 mol%. Phylogenetic analysis of the 16S rRNA gene sequence revealed that strain NH3(T) was a member of rRNA group 6 of the genus Bacillus, which includes alkalitolerant, alkaliphilic and halotolerant species. The closest phylogenetic relatives were Bacillus akibai 1139(T) (96.82 % 16S rRNA gene sequence similarity), B. pseudofirmus DSM 8715(T) (96.76 %), B. okhensis Kh10-101(T) (96.76 %) and B. alkalidiazotrophicus MS 6(T) (96.47 %). Strain NH3(T) could be distinguished from these phylogenetically close neighbours based on a number of phenotypic properties. On the basis of phenotypic and chemotaxonomic characteristics and phylogenetic data, we conclude that strain NH3(T) ( = CGMCC 1.10116(T)  = JCM 16507(T)) merits classification as the type strain of a novel species, for which the name Bacillus nanhaiisediminis sp. nov. is proposed.

Classifying the bacterial gut microbiota of termites and cockroaches: A curated phylogenetic reference database (DictDb).

PubMed

Mikaelyan, Aram; Köhler, Tim; Lampert, Niclas; Rohland, Jeffrey; Boga, Hamadi; Meuser, Katja; Brune, Andreas

2015-10-01

Recent developments in sequencing technology have given rise to a large number of studies that assess bacterial diversity and community structure in termite and cockroach guts based on large amplicon libraries of 16S rRNA genes. Although these studies have revealed important ecological and evolutionary patterns in the gut microbiota, classification of the short sequence reads is limited by the taxonomic depth and resolution of the reference databases used in the respective studies. Here, we present a curated reference database for accurate taxonomic analysis of the bacterial gut microbiota of dictyopteran insects. The Dictyopteran gut microbiota reference Database (DictDb) is based on the Silva database but was significantly expanded by the addition of clones from 11 mostly unexplored termite and cockroach groups, which increased the inventory of bacterial sequences from dictyopteran guts by 26%. The taxonomic depth and resolution of DictDb was significantly improved by a general revision of the taxonomic guide tree for all important lineages, including a detailed phylogenetic analysis of the Treponema and Alistipes complexes, the Fibrobacteres, and the TG3 phylum. The performance of this first documented version of DictDb (v. 3.0) using the revised taxonomic guide tree in the classification of short-read libraries obtained from termites and cockroaches was highly superior to that of the current Silva and RDP databases. DictDb uses an informative nomenclature that is consistent with the literature also for clades of uncultured bacteria and provides an invaluable tool for anyone exploring the gut community structure of termites and cockroaches. Copyright © 2015 Elsevier GmbH. All rights reserved.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Andersen, G.L.; He, Z.; DeSantis, T.Z.

Microarrays have proven to be a useful and high-throughput method to provide targeted DNA sequence information for up to many thousands of specific genetic regions in a single test. A microarray consists of multiple DNA oligonucleotide probes that, under high stringency conditions, hybridize only to specific complementary nucleic acid sequences (targets). A fluorescent signal indicates the presence and, in many cases, the abundance of genetic regions of interest. In this chapter we will look at how microarrays are used in microbial ecology, especially with the recent increase in microbial community DNA sequence data. Of particular interest to microbial ecologists, phylogeneticmore » microarrays are used for the analysis of phylotypes in a community and functional gene arrays are used for the analysis of functional genes, and, by inference, phylotypes in environmental samples. A phylogenetic microarray that has been developed by the Andersen laboratory, the PhyloChip, will be discussed as an example of a microarray that targets the known diversity within the 16S rRNA gene to determine microbial community composition. Using multiple, confirmatory probes to increase the confidence of detection and a mismatch probe for every perfect match probe to minimize the effect of cross-hybridization by non-target regions, the PhyloChip is able to simultaneously identify any of thousands of taxa present in an environmental sample. The PhyloChip is shown to reveal greater diversity within a community than rRNA gene sequencing due to the placement of the entire gene product on the microarray compared with the analysis of up to thousands of individual molecules by traditional sequencing methods. A functional gene array that has been developed by the Zhou laboratory, the GeoChip, will be discussed as an example of a microarray that dynamically identifies functional activities of multiple members within a community. The recent version of GeoChip contains more than 24,000 50mer oligonucleotide probes and covers more than 10,000 gene sequences in 150 gene categories involved in carbon, nitrogen, sulfur, and phosphorus cycling, metal resistance and reduction, and organic contaminant degradation. GeoChip can be used as a generic tool for microbial community analysis, and also link microbial community structure to ecosystem functioning. Examples of the application of both arrays in different environmental samples will be described in the two subsequent sections.« less

Metagenome Analyses of Corroded Concrete Wastewater Pipe Biofilms Reveals a Complex Microbial System

EPA Science Inventory

Analysis of whole-metagenome pyrosequencing data and 16S rRNA gene clone libraries was used to determine microbial composition and functional genes associated with biomass harvested from crown (top) and invert (bottom) sections of a corroded wastewater pipe. Taxonomic and functio...

[Identification of Hydrocarbon-Oxidizing Dietzia Bacteria from Petroleum Reservoirs Based on Phenotypic Properties and Analysis of the 16S rRNA and gyrB Genes].

PubMed

Nazina, T N; Shumkova, E S; Sokolova, D Sh; Babich, T L; Zhurina, M V; Xue, Yan-Fen; Osipov, G A; Poltaraus, A B; Tourova, T P

2015-01-01

The taxonomic position of hydrocarbon-oxidizing bacterial strains 263 and 32d isolated from formation water of the Daqing petroleum reservoir (PRC) was determined by polyphasic taxonomy techniques, including analysis of the 16S rRNA and the gyrB genes. The major chemotaxonomic characteristics of both strains, including the IV type cell wall, composition of cell wall fatty acids, mycolic acids, and menaquinones, agreed with those typical of Dietzia strains. The DNA G+C content of strains 263 and 32d were 67.8 and 67.6 mol%, respectively. Phylogenetic analysis of the 16S rRNA gene of strain 32d revealed 99.7% similarity to the gene of D. maris, making it possible to identify strain 32d as belonging to this species. The 16S rRNA gene sequence of strain 263 exhibited 99.7 and 99.9% similarity to those of D. natronolimnaea and D. cercidiphylli YIM65002(T), respectively. Analysis of the gyrB genes of the subterranean isolates and of a number of Dietzia type strains confirmed classiffication of strain 32d as a D. maris strain and of strain 263, as a D. natronolimnaea strain. A conclusion was made concerning higher resolving power of phylogenetic analysis of the gyrB gene compared to the 16S rRNA gene analysis in the case of determination of the species position of Dietzia isolates.

A taxonomic framework for emerging groups of ecologically important marine gammaproteobacteria based on the reconstruction of evolutionary relationships using genome-scale data

PubMed Central

Spring, Stefan; Scheuner, Carmen; Göker, Markus; Klenk, Hans-Peter

2015-01-01

In recent years a large number of isolates were obtained from saline environments that are phylogenetically related to distinct clades of oligotrophic marine gammaproteobacteria, which were originally identified in seawater samples using cultivation independent methods and are characterized by high seasonal abundances in coastal environments. To date a sound taxonomic framework for the classification of these ecologically important isolates and related species in accordance with their evolutionary relationships is missing. In this study we demonstrate that a reliable allocation of members of the oligotrophic marine gammaproteobacteria (OMG) group and related species to higher taxonomic ranks is possible by phylogenetic analyses of whole proteomes but also of the RNA polymerase beta subunit, whereas phylogenetic reconstructions based on 16S rRNA genes alone resulted in unstable tree topologies with only insignificant bootstrap support. The identified clades could be correlated with distinct phenotypic traits illustrating an adaptation to common environmental factors in their evolutionary history. Genome wide gene-content analyses revealed the existence of two distinct ecological guilds within the analyzed lineage of marine gammaproteobacteria which can be distinguished by their trophic strategies. Based on our results a novel order within the class Gammaproteobacteria is proposed, which is designated Cellvibrionales ord. nov. and comprises the five novel families Cellvibrionaceae fam. nov., Halieaceae fam. nov., Microbulbiferaceae fam. nov., Porticoccaceae fam. nov., and Spongiibacteraceae fam. nov. PMID:25914684

Comparison of Sewage and Animal Fecal Microbiomes by Using Oligotyping Reveals Potential Human Fecal Indicators in Multiple Taxonomic Groups

PubMed Central

Fisher, Jenny C.; Eren, A. Murat; Green, Hyatt C.; Shanks, Orin C.; Morrison, Hilary G.; Vineis, Joseph H.; Sogin, Mitchell L.

2015-01-01

Most DNA-based microbial source tracking (MST) approaches target host-associated organisms within the order Bacteroidales, but the gut microbiota of humans and other animals contain organisms from an array of other taxonomic groups that might provide indicators of fecal pollution sources. To discern between human and nonhuman fecal sources, we compared the V6 regions of the 16S rRNA genes detected in fecal samples from six animal hosts to those found in sewage (as a proxy for humans). We focused on 10 abundant genera and used oligotyping, which can detect subtle differences between rRNA gene sequences from ecologically distinct organisms. Our analysis showed clear patterns of differential oligotype distributions between sewage and animal samples. Over 100 oligotypes of human origin occurred preferentially in sewage samples, and 99 human oligotypes were sewage specific. Sequences represented by the sewage-specific oligotypes can be used individually for development of PCR-based assays or together with the oligotypes preferentially associated with sewage to implement a signature-based approach. Analysis of sewage from Spain and Brazil showed that the sewage-specific oligotypes identified in U.S. sewage have the potential to be used as global alternative indicators of human fecal pollution. Environmental samples with evidence of prior human fecal contamination had consistent ratios of sewage signature oligotypes that corresponded to the trends observed for sewage. Our methodology represents a promising approach to identifying new bacterial taxa for MST applications and further highlights the potential of the family Lachnospiraceae to provide human-specific markers. In addition to source tracking applications, the patterns of the fine-scale population structure within fecal taxa suggest a fundamental relationship between bacteria and their hosts. PMID:26231648

Hybrid-denovo: a de novo OTU-picking pipeline integrating single-end and paired-end 16S sequence tags.

PubMed

Chen, Xianfeng; Johnson, Stephen; Jeraldo, Patricio; Wang, Junwen; Chia, Nicholas; Kocher, Jean-Pierre A; Chen, Jun

2018-03-01

Illumina paired-end sequencing has been increasingly popular for 16S rRNA gene-based microbiota profiling. It provides higher phylogenetic resolution than single-end reads due to a longer read length. However, the reverse read (R2) often has significant low base quality, and a large proportion of R2s will be discarded after quality control, resulting in a mixture of paired-end and single-end reads. A typical 16S analysis pipeline usually processes either paired-end or single-end reads but not a mixture. Thus, the quantification accuracy and statistical power will be reduced due to the loss of a large amount of reads. As a result, rare taxa may not be detectable with the paired-end approach, or low taxonomic resolution will result in a single-end approach. To have both the higher phylogenetic resolution provided by paired-end reads and the higher sequence coverage by single-end reads, we propose a novel OTU-picking pipeline, hybrid-denovo, that can process a hybrid of single-end and paired-end reads. Using high-quality paired-end reads as a gold standard, we show that hybrid-denovo achieved the highest correlation with the gold standard and performed better than the approaches based on paired-end or single-end reads in terms of quantifying the microbial diversity and taxonomic abundances. By applying our method to a rheumatoid arthritis (RA) data set, we demonstrated that hybrid-denovo captured more microbial diversity and identified more RA-associated taxa than a paired-end or single-end approach. Hybrid-denovo utilizes both paired-end and single-end 16S sequencing reads and is recommended for 16S rRNA gene targeted paired-end sequencing data.

Analyses of the Stability and Core Taxonomic Memberships of the Human Microbiome

PubMed Central

Li, Kelvin; Bihan, Monika; Methé, Barbara A.

2013-01-01

Analyses of the taxonomic diversity associated with the human microbiome continue to be an area of great importance. The study of the nature and extent of the commonly shared taxa (“core”), versus those less prevalent, establishes a baseline for comparing healthy and diseased groups by quantifying the variation among people, across body habitats and over time. The National Institutes of Health (NIH) sponsored Human Microbiome Project (HMP) has provided an unprecedented opportunity to examine and better define what constitutes the taxonomic core within and across body habitats and individuals through pyrosequencing-based profiling of 16S rRNA gene sequences from oral, skin, distal gut (stool), and vaginal body habitats from over 200 healthy individuals. A two-parameter model is introduced to quantitatively identify the core taxonomic members of each body habitat’s microbiota across the healthy cohort. Using only cutoffs for taxonomic ubiquity and abundance, core taxonomic members were identified for each of the 18 body habitats and also for the 4 higher-level body regions. Although many microbes were shared at low abundance, they exhibited a relatively continuous spread in both their abundance and ubiquity, as opposed to a more discretized separation. The numbers of core taxa members in the body regions are comparatively small and stable, reflecting the relatively high, but conserved, interpersonal variability within the cohort. Core sizes increased across the body regions in the order of: vagina, skin, stool, and oral cavity. A number of “minor” oral taxonomic core were also identified by their majority presence across the cohort, but with relatively low and stable abundances. A method for quantifying the difference between two cohorts was introduced and applied to samples collected on a second visit, revealing that over time, the oral, skin, and stool body regions tended to be more transient in their taxonomic structure than the vaginal body region. PMID:23671663

Drinking Water Microbiome as a Screening Tool for Nitrification in Chloraminated Drinking Water Distribution Systems (abstract)

EPA Science Inventory

Many water utilities in the US using chloramine as disinfectant treatment in their distribution systems have experienced nitrification episodes, which detrimentally impact the water quality. Here, we used 16S rRNA sequencing data to generate high-resolution taxonomic profiles of...

Identifying Fishes through DNA Barcodes and Microarrays.

PubMed

Kochzius, Marc; Seidel, Christian; Antoniou, Aglaia; Botla, Sandeep Kumar; Campo, Daniel; Cariani, Alessia; Vazquez, Eva Garcia; Hauschild, Janet; Hervet, Caroline; Hjörleifsdottir, Sigridur; Hreggvidsson, Gudmundur; Kappel, Kristina; Landi, Monica; Magoulas, Antonios; Marteinsson, Viggo; Nölte, Manfred; Planes, Serge; Tinti, Fausto; Turan, Cemal; Venugopal, Moleyur N; Weber, Hannes; Blohm, Dietmar

2010-09-07

International fish trade reached an import value of 62.8 billion Euro in 2006, of which 44.6% are covered by the European Union. Species identification is a key problem throughout the life cycle of fishes: from eggs and larvae to adults in fisheries research and control, as well as processed fish products in consumer protection. This study aims to evaluate the applicability of the three mitochondrial genes 16S rRNA (16S), cytochrome b (cyt b), and cytochrome oxidase subunit I (COI) for the identification of 50 European marine fish species by combining techniques of "DNA barcoding" and microarrays. In a DNA barcoding approach, neighbour Joining (NJ) phylogenetic trees of 369 16S, 212 cyt b, and 447 COI sequences indicated that cyt b and COI are suitable for unambiguous identification, whereas 16S failed to discriminate closely related flatfish and gurnard species. In course of probe design for DNA microarray development, each of the markers yielded a high number of potentially species-specific probes in silico, although many of them were rejected based on microarray hybridisation experiments. None of the markers provided probes to discriminate the sibling flatfish and gurnard species. However, since 16S-probes were less negatively influenced by the "position of label" effect and showed the lowest rejection rate and the highest mean signal intensity, 16S is more suitable for DNA microarray probe design than cty b and COI. The large portion of rejected COI-probes after hybridisation experiments (>90%) renders the DNA barcoding marker as rather unsuitable for this high-throughput technology. Based on these data, a DNA microarray containing 64 functional oligonucleotide probes for the identification of 30 out of the 50 fish species investigated was developed. It represents the next step towards an automated and easy-to-handle method to identify fish, ichthyoplankton, and fish products.

Anaerococcus degenerii sp. nov., isolated from human clinical specimens.

PubMed

Veloo, A C M; Elgersma, P E; van Winkelhoff, A J

2015-06-01

Four clinical isolates of gram-positive strict anaerobic cocci were isolated from four different human mixed anaerobic infections. The taxonomical status of the four strains could not be established using standard identification techniques. The biochemical features of the strains were established and their taxonomic position was determined using 16S rRNA sequencing. The four strains form a homogeneous phenotypical and genotypical cluster. A new Anaerococcus species is proposed for these isolates: Anaerococcus degenerii sp. nov. The type strain is UMCG-104(T) = DSM29674(T) (accession number AM176528). Copyright © 2015 Elsevier Ltd. All rights reserved.

5S ribosomal RNA database Y2K

PubMed Central

Szymanski, Maciej; Barciszewska, Miroslawa Z.; Barciszewski, Jan; Erdmann, Volker A.

2000-01-01

This paper presents the updated version (Y2K) of the database of ribosomal 5S ribonucleic acids (5S rRNA) and their genes (5S rDNA), http://rose.man/poznan. pl/5SData/index.html . This edition of the database contains 1985 primary structures of 5S rRNA and 5S rDNA. They include 60 archaebacterial, 470 eubacterial, 63 plastid, nine mitochondrial and 1383 eukaryotic sequences. The nucleotide sequences of the 5S rRNAs or 5S rDNAs are divided according to the taxonomic position of the source organisms. PMID:10592212

5S ribosomal RNA database Y2K.

PubMed

Szymanski, M; Barciszewska, M Z; Barciszewski, J; Erdmann, V A

2000-01-01

This paper presents the updated version (Y2K) of the database of ribosomal 5S ribonucleic acids (5S rRNA) and their genes (5S rDNA), http://rose.man/poznan.pl/5SData/index.html. This edition of the database contains 1985primary structures of 5S rRNA and 5S rDNA. They include 60 archaebacterial, 470 eubacterial, 63 plastid, nine mitochondrial and 1383 eukaryotic sequences. The nucleotide sequences of the 5S rRNAs or 5S rDNAs are divided according to the taxonomic position of the source organisms.

Phylogenetic relatedness determined between antibiotic resistance and 16S rRNA genes in actinobacteria.

PubMed

Sagova-Mareckova, Marketa; Ulanova, Dana; Sanderova, Petra; Omelka, Marek; Kamenik, Zdenek; Olsovska, Jana; Kopecky, Jan

2015-04-01

Distribution and evolutionary history of resistance genes in environmental actinobacteria provide information on intensity of antibiosis and evolution of specific secondary metabolic pathways at a given site. To this day, actinobacteria producing biologically active compounds were isolated mostly from soil but only a limited range of soil environments were commonly sampled. Consequently, soil remains an unexplored environment in search for novel producers and related evolutionary questions. Ninety actinobacteria strains isolated at contrasting soil sites were characterized phylogenetically by 16S rRNA gene, for presence of erm and ABC transporter resistance genes and antibiotic production. An analogous analysis was performed in silico with 246 and 31 strains from Integrated Microbial Genomes (JGI_IMG) database selected by the presence of ABC transporter genes and erm genes, respectively. In the isolates, distances of erm gene sequences were significantly correlated to phylogenetic distances based on 16S rRNA genes, while ABC transporter gene distances were not. The phylogenetic distance of isolates was significantly correlated to soil pH and organic matter content of isolation sites. In the analysis of JGI_IMG datasets the correlation between phylogeny of resistance genes and the strain phylogeny based on 16S rRNA genes or five housekeeping genes was observed for both the erm genes and ABC transporter genes in both actinobacteria and streptomycetes. However, in the analysis of sequences from genomes where both resistance genes occurred together the correlation was observed for both ABC transporter and erm genes in actinobacteria but in streptomycetes only in the erm gene. The type of erm resistance gene sequences was influenced by linkage to 16S rRNA gene sequences and site characteristics. The phylogeny of ABC transporter gene was correlated to 16S rRNA genes mainly above the genus level. The results support the concept of new specific secondary metabolite scaffolds occurring more likely in taxonomically distant producers but suggest that the antibiotic selection of gene pools is also influenced by site conditions.

Rapid and reliable detection and identification of GM events using multiplex PCR coupled with oligonucleotide microarray.

PubMed

Xu, Xiaodan; Li, Yingcong; Zhao, Heng; Wen, Si-yuan; Wang, Sheng-qi; Huang, Jian; Huang, Kun-lun; Luo, Yun-bo

2005-05-18

To devise a rapid and reliable method for the detection and identification of genetically modified (GM) events, we developed a multiplex polymerase chain reaction (PCR) coupled with a DNA microarray system simultaneously aiming at many targets in a single reaction. The system included probes for screening gene, species reference gene, specific gene, construct-specific gene, event-specific gene, and internal and negative control genes. 18S rRNA was combined with species reference genes as internal controls to assess the efficiency of all reactions and to eliminate false negatives. Two sets of the multiplex PCR system were used to amplify four and five targets, respectively. Eight different structure genes could be detected and identified simultaneously for Roundup Ready soybean in a single microarray. The microarray specificity was validated by its ability to discriminate two GM maizes Bt176 and Bt11. The advantages of this method are its high specificity and greatly reduced false-positives and -negatives. The multiplex PCR coupled with microarray technology presented here is a rapid and reliable tool for the simultaneous detection of GM organism ingredients.

The gene coding for small ribosomal subunit RNA in the basidiomycete Ustilago maydis contains a group I intron.

PubMed Central

De Wachter, R; Neefs, J M; Goris, A; Van de Peer, Y

1992-01-01

The nucleotide sequence of the gene coding for small ribosomal subunit RNA in the basidiomycete Ustilago maydis was determined. It revealed the presence of a group I intron with a length of 411 nucleotides. This is the third occurrence of such an intron discovered in a small subunit rRNA gene encoded by a eukaryotic nuclear genome. The other two occurrences are in Pneumocystis carinii, a fungus of uncertain taxonomic status, and Ankistrodesmus stipitatus, a green alga. The nucleotides of the conserved core structure of 101 group I intron sequences present in different genes and genome types were aligned and their evolutionary relatedness was examined. This revealed a cluster including all group I introns hitherto found in eukaryotic nuclear genes coding for small and large subunit rRNAs. A secondary structure model was designed for the area of the Ustilago maydis small ribosomal subunit RNA precursor where the intron is situated. It shows that the internal guide sequence pairing with the intron boundaries fits between two helices of the small subunit rRNA, and that minimal rearrangement of base pairs suffices to achieve the definitive secondary structure of the 18S rRNA upon splicing. PMID:1561081

Protistan diversity and activity inferred from RNA and DNA at a coastal ocean site in the eastern North Pacific.

PubMed

Hu, Sarah K; Campbell, Victoria; Connell, Paige; Gellene, Alyssa G; Liu, Zhenfeng; Terrado, Ramon; Caron, David A

2016-04-01

Microbial eukaryotes fulfill key ecological positions in marine food webs. Molecular approaches that connect protistan diversity and biogeography to their diverse metabolisms will greatly improve our understanding of marine ecosystem function. The majority of molecular-based studies to date use 18S rRNA gene sequencing to characterize natural microbial assemblages, but this approach does not necessarily discriminate between active and non-active cells. We incorporated RNA sequencing into standard 18S rRNA gene sequence surveys with the purpose of assessing those members of the protistan community contributing to biogeochemical cycling (active organisms), using the ratio of cDNA (reverse transcribed from total RNA) to 18S rRNA gene sequences within major protistan taxonomic groups. Trophically important phytoplankton, such as diatoms and chlorophytes exhibited seasonal trends in relative activity. Additionally, both radiolaria and ciliates displayed previously unreported high relative activities below the euphotic zone. This study sheds new light on the relative metabolic activity of specific protistan groups and how microbial communities respond to changing environmental conditions. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Prediction of functional profiles of gut microbiota from 16S rRNA metagenomic data provides a more robust evaluation of gut dysbiosis occurring in Japanese type 2 diabetic patients.

PubMed

Inoue, Ryo; Ohue-Kitano, Ryuji; Tsukahara, Takamitsu; Tanaka, Masashi; Masuda, Shinya; Inoue, Takayuki; Yamakage, Hajime; Kusakabe, Toru; Hasegawa, Koji; Shimatsu, Akira; Satoh-Asahara, Noriko

2017-11-01

We assessed whether gut microbial functional profiles predicted from 16S rRNA metagenomics differed in Japanese type 2 diabetic patients. A total of 22 Japanese subjects were recruited from our outpatient clinic in an observational study. Fecal samples were obtained from 12 control and 10 type 2 diabetic subjects. 16S rRNA metagenomic data were generated and functional profiles predicted using "Phylogenetic Investigation of Communities by Reconstruction of Unobserved States" software. We measured the parameters of glucose metabolism, gut bacterial taxonomy and functional profile, and examined the associations in a cross-sectional manner. Eleven of 288 "Kyoto Encyclopedia of Genes and Genomes" pathways were significantly enriched in diabetic patients compared with control subjects ( p <0.05, q<0.1). The relative abundance of almost all pathways, including the Insulin signaling pathway and Glycolysis/Gluconeogenesis , showed strong, positive correlations with hemoglobin A 1c (HbA 1c ) and fasting plasma glucose (FPG) levels. Bacterial taxonomic analysis showed that genus Blautia significantly differed between groups and had negative correlations with HbA 1c and FPG levels. Our findings suggest a novel pathophysiological relationship between gut microbial communities and diabetes, further highlighting the significance and utility of combining prediction of functional profiles with ordinal bacterial taxonomic analysis (UMIN Clinical Trails Registry number: UMIN000026592).

Classification of thermophilic actinobacteria isolated from arid desert soils, including the description of Amycolatopsis deserti sp. nov.

PubMed

Busarakam, Kanungnid; Brown, Ros; Bull, Alan T; Tan, Geok Yuan Annie; Zucchi, Tiago D; da Silva, Leonardo José; de Souza, Wallace Rafael; Goodfellow, Michael

2016-02-01

The taxonomic position of 26 filamentous actinobacteria isolated from a hyper-arid Atacama Desert soil and 2 from an arid Australian composite soil was established using a polyphasic approach. All of the isolates gave the diagnostic amplification product using 16S rRNA oligonucleotide primers specific for the genus Amycolatopsis. Representative isolates had chemotaxonomic and morphological properties typical of members of the genus Amycolatopsis. 16S rRNA gene analyses showed that all of the isolates belong to the Amycolatopsis methanolica 16S rRNA gene clade. The Atacama Desert isolates were assigned to one or other of two recognised species, namely Amycolatopsis ruanii and Amycolatopsis thermalba, based on 16S rRNA gene sequence, DNA:DNA relatedness and phenotypic data; emended descriptions are given for these species. In contrast, the two strains from the arid Australian composite soil, isolates GY024(T) and GY142, formed a distinct branch at the periphery of the A. methanolica 16S rRNA phyletic line, a taxon that was supported by all of the tree-making algorithms and by a 100 % bootstrap value. These strains shared a high degree of DNA:DNA relatedness and have many phenotypic properties in common, some of which distinguished them from all of the constituent species classified in the A. methanolica 16S rRNA clade. Isolates GY024(T) and GY142 merit recognition as a new species within the A. methanolica group of thermophilic strains. The name proposed for the new species is Amycolatopsis deserti sp. nov.; the type strain is GY024(T) (=NCIMB 14972(T) = NRRL B-65266(T)).

Ecological Consistency of SSU rRNA-Based Operational Taxonomic Units at a Global Scale

PubMed Central

Schmidt, Thomas S. B.; Matias Rodrigues, João F.; von Mering, Christian

2014-01-01

Operational Taxonomic Units (OTUs), usually defined as clusters of similar 16S/18S rRNA sequences, are the most widely used basic diversity units in large-scale characterizations of microbial communities. However, it remains unclear how well the various proposed OTU clustering algorithms approximate ‘true’ microbial taxa. Here, we explore the ecological consistency of OTUs – based on the assumption that, like true microbial taxa, they should show measurable habitat preferences (niche conservatism). In a global and comprehensive survey of available microbial sequence data, we systematically parse sequence annotations to obtain broad ecological descriptions of sampling sites. Based on these, we observe that sequence-based microbial OTUs generally show high levels of ecological consistency. However, different OTU clustering methods result in marked differences in the strength of this signal. Assuming that ecological consistency can serve as an objective external benchmark for cluster quality, we conclude that hierarchical complete linkage clustering, which provided the most ecologically consistent partitions, should be the default choice for OTU clustering. To our knowledge, this is the first approach to assess cluster quality using an external, biologically meaningful parameter as a benchmark, on a global scale. PMID:24763141

Marker genes that are less conserved in their sequences are useful for predicting genome-wide similarity levels between closely related prokaryotic strains

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lan, Yemin; Rosen, Gail; Hershberg, Ruth

The 16s rRNA gene is so far the most widely used marker for taxonomical classification and separation of prokaryotes. Since it is universally conserved among prokaryotes, it is possible to use this gene to classify a broad range of prokaryotic organisms. At the same time, it has often been noted that the 16s rRNA gene is too conserved to separate between prokaryotes at finer taxonomic levels. In this paper, we examine how well levels of similarity of 16s rRNA and 73 additional universal or nearly universal marker genes correlate with genome-wide levels of gene sequence similarity. We demonstrate that themore » percent identity of 16s rRNA predicts genome-wide levels of similarity very well for distantly related prokaryotes, but not for closely related ones. In closely related prokaryotes, we find that there are many other marker genes for which levels of similarity are much more predictive of genome-wide levels of gene sequence similarity. Finally, we show that the identities of the markers that are most useful for predicting genome-wide levels of similarity within closely related prokaryotic lineages vary greatly between lineages. However, the most useful markers are always those that are least conserved in their sequences within each lineage. In conclusion, our results show that by choosing markers that are less conserved in their sequences within a lineage of interest, it is possible to better predict genome-wide gene sequence similarity between closely related prokaryotes than is possible using the 16s rRNA gene. We point readers towards a database we have created (POGO-DB) that can be used to easily establish which markers show lowest levels of sequence conservation within different prokaryotic lineages.« less

Marker genes that are less conserved in their sequences are useful for predicting genome-wide similarity levels between closely related prokaryotic strains

DOE PAGES

Lan, Yemin; Rosen, Gail; Hershberg, Ruth

2016-05-03

The 16s rRNA gene is so far the most widely used marker for taxonomical classification and separation of prokaryotes. Since it is universally conserved among prokaryotes, it is possible to use this gene to classify a broad range of prokaryotic organisms. At the same time, it has often been noted that the 16s rRNA gene is too conserved to separate between prokaryotes at finer taxonomic levels. In this paper, we examine how well levels of similarity of 16s rRNA and 73 additional universal or nearly universal marker genes correlate with genome-wide levels of gene sequence similarity. We demonstrate that themore » percent identity of 16s rRNA predicts genome-wide levels of similarity very well for distantly related prokaryotes, but not for closely related ones. In closely related prokaryotes, we find that there are many other marker genes for which levels of similarity are much more predictive of genome-wide levels of gene sequence similarity. Finally, we show that the identities of the markers that are most useful for predicting genome-wide levels of similarity within closely related prokaryotic lineages vary greatly between lineages. However, the most useful markers are always those that are least conserved in their sequences within each lineage. In conclusion, our results show that by choosing markers that are less conserved in their sequences within a lineage of interest, it is possible to better predict genome-wide gene sequence similarity between closely related prokaryotes than is possible using the 16s rRNA gene. We point readers towards a database we have created (POGO-DB) that can be used to easily establish which markers show lowest levels of sequence conservation within different prokaryotic lineages.« less

[Characterization of Black and Dichothrix Cyanobacteria Based on the 16S Ribosomal RNA Gene Sequence

NASA Technical Reports Server (NTRS)

Ortega, Maya

2010-01-01

My project focuses on characterizing different cyanobacteria in thrombolitic mats found on the island of Highborn Cay, Bahamas. Thrombolites are interesting ecosystems because of the ability of bacteria in these mats to remove carbon dioxide from the atmosphere and mineralize it as calcium carbonate. In the future they may be used as models to develop carbon sequestration technologies, which could be used as part of regenerative life systems in space. These thrombolitic communities are also significant because of their similarities to early communities of life on Earth. I targeted two cyanobacteria in my research, Dichothrix spp. and whatever black is, since they are believed to be important to carbon sequestration in these thrombolitic mats. The goal of my summer research project was to molecularly identify these two cyanobacteria. DNA was isolated from each organism through mat dissections and DNA extractions. I ran Polymerase Chain Reactions (PCR) to amplify the 16S ribosomal RNA (rRNA) gene in each cyanobacteria. This specific gene is found in almost all bacteria and is highly conserved, meaning any changes in the sequence are most likely due to evolution. As a result, the 16S rRNA gene can be used for bacterial identification of different species based on the sequence of their 16S rRNA gene. Since the exact sequence of the Dichothrix gene was unknown, I designed different primers that flanked the gene based on the known sequences from other taxonomically similar cyanobacteria. Once the 16S rRNA gene was amplified, I cloned the gene into specialized Escherichia coli cells and sent the gene products for sequencing. Once the sequence is obtained, it will be added to a genetic database for future reference to and classification of other Dichothrix sp.

Phylogenetically Structured Differences in rRNA Gene Sequence Variation among Species of Arbuscular Mycorrhizal Fungi and Their Implications for Sequence Clustering

PubMed Central

Ekanayake, Saliya; Ruan, Yang; Schütte, Ursel M. E.; Kaonongbua, Wittaya; Fox, Geoffrey; Ye, Yuzhen; Bever, James D.

2016-01-01

ABSTRACT Arbuscular mycorrhizal (AM) fungi form mutualisms with plant roots that increase plant growth and shape plant communities. Each AM fungal cell contains a large amount of genetic diversity, but it is unclear if this diversity varies across evolutionary lineages. We found that sequence variation in the nuclear large-subunit (LSU) rRNA gene from 29 isolates representing 21 AM fungal species generally assorted into genus- and species-level clades, with the exception of species of the genera Claroideoglomus and Entrophospora. However, there were significant differences in the levels of sequence variation across the phylogeny and between genera, indicating that it is an evolutionarily constrained trait in AM fungi. These consistent patterns of sequence variation across both phylogenetic and taxonomic groups pose challenges to interpreting operational taxonomic units (OTUs) as approximations of species-level groups of AM fungi. We demonstrate that the OTUs produced by five sequence clustering methods using 97% or equivalent sequence similarity thresholds failed to match the expected species of AM fungi, although OTUs from AbundantOTU, CD-HIT-OTU, and CROP corresponded better to species than did OTUs from mothur or UPARSE. This lack of OTU-to-species correspondence resulted both from sequences of one species being split into multiple OTUs and from sequences of multiple species being lumped into the same OTU. The OTU richness therefore will not reliably correspond to the AM fungal species richness in environmental samples. Conservatively, this error can overestimate species richness by 4-fold or underestimate richness by one-half, and the direction of this error will depend on the genera represented in the sample. IMPORTANCE Arbuscular mycorrhizal (AM) fungi form important mutualisms with the roots of most plant species. Individual AM fungi are genetically diverse, but it is unclear whether the level of this diversity differs among evolutionary lineages. We found that the amount of sequence variation in an rRNA gene that is commonly used to identify AM fungal species varied significantly between evolutionary groups that correspond to different genera, with the exception of two genera that are genetically indistinguishable from each other. When we clustered groups of similar sequences into operational taxonomic units (OTUs) using five different clustering methods, these patterns of sequence variation caused the number of OTUs to either over- or underestimate the actual number of AM fungal species, depending on the genus. Our results indicate that OTU-based inferences about AM fungal species composition from environmental sequences can be improved if they take these taxonomically structured patterns of sequence variation into account. PMID:27260357

Comparison of reduced metagenome and 16S rRNA gene sequencing for determination of genetic diversity and mother-child overlap of the gut associated microbiota.

PubMed

Ravi, Anuradha; Avershina, Ekaterina; Angell, Inga Leena; Ludvigsen, Jane; Manohar, Prasanth; Padmanaban, Sumathi; Nachimuthu, Ramesh; Snipen, Lars; Rudi, Knut

2018-06-01

Use of the 16S rRNA gene in microbiota studies is limited by the lack of taxonomic and functional resolution. High resolution analyses are particularly important for understanding transmission and persistence of bacteria. The aim of our work was therefore to compare a novel reduced metagenome sequencing (RMS) approach with 16S rRNA gene sequencing to determine both the metagenome genetic diversity and the mother-to-child sharing of the microbiota in a cohort of 17 mother-child pairs. We found that although both approaches gave comparable results with respect to sample separation and taxonomy, RMS gave higher resolution and the potential for genomic-/functional assignment. Using RMS we estimated that the metagenome size increased from about 60 Mbp for 4-day-old children to about 225 Mbp for mothers. The 4-day-old children shared 7% of the metagenome sequences with the mothers, while the metagenome sequence sharing was >30% among the mothers. We found 15 genomes shared across >50% of the mothers, of which 10 belonged to Clostridia. Only Bacteroides showed a direct mother-child association, with B. vulgatus being abundant in both 4-day-old children and mothers. For the functional assignments, we identified a significant association between antibiotic usage during labor, and quantity of Fosfomycin resistance genes. In conclusion, our results show a higher functional and taxonomic resolution for RMS compared to 16S rRNA gene sequencing, where RMS enabled a detailed description of mother to child gut microbiota transmission - supporting a late recruitment of most gut bacteria and an effect of antibiotic treatment during labor on infant antibiotic resistance gene patterns. Copyright © 2018. Published by Elsevier B.V.

Biogeographic patterns of bacterial microdiversity in Arctic deep-sea sediments (HAUSGARTEN, Fram Strait)

PubMed Central

Buttigieg, Pier Luigi; Ramette, Alban

2015-01-01

Marine bacteria colonizing deep-sea sediments beneath the Arctic ocean, a rapidly changing ecosystem, have been shown to exhibit significant biogeographic patterns along transects spanning tens of kilometers and across water depths of several thousand meters (Jacob et al., 2013). Jacob et al. (2013) adopted what has become a classical view of microbial diversity – based on operational taxonomic units clustered at the 97% sequence identity level of the 16S rRNA gene – and observed a very large microbial community replacement at the HAUSGARTEN Long Term Ecological Research station (Eastern Fram Strait). Here, we revisited these data using the oligotyping approach and aimed to obtain new insight into ecological and biogeographic patterns associated with bacterial microdiversity in marine sediments. We also assessed the level of concordance of these insights with previously obtained results. Variation in oligotype dispersal range, relative abundance, co-occurrence, and taxonomic identity were related to environmental parameters such as water depth, biomass, and sedimentary pigment concentration. This study assesses ecological implications of the new microdiversity-based technique using a well-characterized dataset of high relevance for global change biology. PMID:25601856

Marine Actinobacteria as a source of compounds for phytopathogen control: An integrative metabolic-profiling / bioactivity and taxonomical approach

PubMed Central

Betancur, Luz A.; Naranjo-Gaybor, Sandra J.; Vinchira-Villarraga, Diana M.; Moreno-Sarmiento, Nubia C.; Maldonado, Luis A.; Suarez-Moreno, Zulma R.; Acosta-González, Alejandro; Padilla-Gonzalez, Gillermo F.; Puyana, Mónica; Castellanos, Leonardo; Ramos, Freddy A.

2017-01-01

Marine bacteria are considered as promising sources for the discovery of novel biologically active compounds. In this study, samples of sediment, invertebrate and algae were collected from the Providencia and Santa Catalina coral reef (Colombian Caribbean Sea) with the aim of isolating Actinobateria-like strain able to produce antimicrobial and quorum quenching compounds against pathogens. Several approaches were used to select actinobacterial isolates, obtaining 203 strains from all samples. According to their 16S rRNA gene sequencing, a total of 24 strains was classified within Actinobacteria represented by three genera: Streptomyces, Micromonospora, and Gordonia. In order to assess their metabolic profiles, the actinobacterial strains were grown in liquid cultures, and LC-MS-based analyses from ethyl acetate fractions were performed. Based on taxonomical classification, screening information of activity against phytopathogenic strains and quorum quenching activity, as well as metabolic profiling, six out of the 24 isolates were selected for follow-up with chemical isolation and structure identification analyses of putative metabolites involved in antimicrobial activities. PMID:28225766

Candidatus Phytoplasma malaysianum, a novel taxon associated with virescence and phyllody of Madagascar periwinkle (Catharanthus roseus)

USDA-ARS?s Scientific Manuscript database

This study addressed the taxonomic position and group classification of a phytoplasma responsible for virescence and phyllody symptoms in naturally diseased Madagascar periwinkle plants in western Malaysia. Unique regions in the 16S rRNA gene from the Malaysian periwinkle virescence (MaPV) phytopla...

Polaribacter gen. nov., with three new species, P. irgensii sp. nov., P. franzmannii sp. nov. and P. filamentus sp. nov., gas vacuolate polar marine bacteria of the Cytophaga-Flavobacterium-Bacteroides group and reclassification of 'Flectobacillus glomeratus' as Polaribacter glomeratus comb. nov

NASA Technical Reports Server (NTRS)

Gosink, J. J.; Woese, C. R.; Staley, J. T.

1998-01-01

Several psychrophilic, gas vacuolate strains of the Cytophage-Flavobacterium-Bacteroides (CFB) phylogenetic group were isolated from sea ice and water from the Arctic and the Antarctic. The closest taxonomically defined species by 16S rRNA sequence analysis is 'Flectobacillus glomeratus'. However, 'Flc. glomeratus' is phylogenetically distant from the Flectobacillus type species, Flc. major. On the basis of phenotypic, genotypic and 16S rRNA sequence analyses we propose a new genus, Polaribacter, with three new species, Polaribacter irgensii strain 23-P (ATCC 700398), Polaribacter franzmannii strain 301 (ATCC 700399) and Polaribacter filamentus strain 215 (ATCC 700397). P. filamentus is the type species of the genus. None of these species exhibits a cosmopolitan or bipolar distribution. This is the first taxonomic description of gas vacuolate bacteria in the CFB group. Additionally, we propose that 'Flc. glomeratus' be reclassified to the genus Polaribacter as P. glomeratus, comb. nov.

Taxonomic revision of the Chinese Limnonectes (Anura, Dicroglossidae) with the description of a new species from China and Myanmar.

PubMed

Suwannapoom, Chatmongkon; Yuan, Zhi-Yong; Chen, Jin-Min; Hou, Mian; Zhao, Hai-Peng; Wang, Li-Jun; Nguyen, Truong Son; Nguyen, Truong Q; Murphy, Robert W; Sullivan, Jaqueline; Mcleod, David S; Che, Jing

2016-03-21

Phylogenetic reconstructions derived from DNA sequence data play a central role in documenting the number of species in a complex. Such analyses are pointing to the existence of many cryptic species, especially in poorly understood groups such as the genus Limnonectes, and the L. kuhlii species complex in particular. To understand the Limnonectes frogs of China, we reconstruct the major matrilineal genealogy of Limnonectes from China and Southeast Asia based on 12S rRNA, tRNA^Val and 16S rRNA gene sequences. Based on new data we recognize five species of Limnonectes in China including L. bannaensis, L. fujianensis, L. fragilis, L. taylori (new record), and a new species from southern China and Myanmar. Phylogenetically, the new species is more closely related to the clade comprising L. taylori, L. megastomias, L. isanensis, L. nguyenorum, and L. jarujini from Thailand than to other Chinese species. This study supports previous findings of sympatric members of a species complex that are not each other's closest relatives.

Skin lesion-associated pathogens from Octopus vulgaris: first detection of Photobacterium swingsii, Lactococcus garvieae and betanodavirus.

PubMed

Fichi, G; Cardeti, G; Perrucci, S; Vanni, A; Cersini, A; Lenzi, C; De Wolf, T; Fronte, B; Guarducci, M; Susini, F

2015-07-23

The common octopus Octopus vulgaris Cuvier, 1798 is extremely important in fisheries and is a useful protein source in most Mediterranean countries. Here we investigated pathogens associated with skin lesions in 9 naturally deceased specimens that included both cultured and wild common octopus. Within 30 min after death, each octopus was stored at 4°C and microbiologically examined within 24 h. Bacterial colonies, cultured from swabs taken from the lesions, were examined using taxonomical and biochemical analyses. Vibrio alginolyticus and V. parahaemolyticus were only isolated from cultured animals. A conventional PCR targeting the 16S ribosomal RNA (rRNA) gene and sequencing were performed on 2 bacterial isolates that remained unidentified after taxonomical and biochemical analysis. The sequence results indicated that the bacteria had a 99% identity with Lactococcus garvieae and Photobacterium swingsii. L. garvieae was confirmed using a specific PCR based on the 16S-23S rRNA internal transcribed spacer region, while P. swingsii was confirmed by phylogenetic analyses. Although all animals examined were found to be infected by the protozoan species Aggregata octopiana localised in the intestines, it was also present in skin lesions of 2 of the animals. Betanodavirus was detected in both cultured and wild individuals by cell culture, PCR and electron microscopy. These findings are the first report of L. garvieae and betanodavirus from skin lesions of common octopus and the first identification of P. swingsii both in octopus skin lesions and in marine invertebrates in Italy.

The Microbial Genomes Atlas (MiGA) webserver: taxonomic and gene diversity analysis of Archaea and Bacteria at the whole genome level.

PubMed

Rodriguez-R, Luis M; Gunturu, Santosh; Harvey, William T; Rosselló-Mora, Ramon; Tiedje, James M; Cole, James R; Konstantinidis, Konstantinos T

2018-06-14

The small subunit ribosomal RNA gene (16S rRNA) has been successfully used to catalogue and study the diversity of prokaryotic species and communities but it offers limited resolution at the species and finer levels, and cannot represent the whole-genome diversity and fluidity. To overcome these limitations, we introduced the Microbial Genomes Atlas (MiGA), a webserver that allows the classification of an unknown query genomic sequence, complete or partial, against all taxonomically classified taxa with available genome sequences, as well as comparisons to other related genomes including uncultivated ones, based on the genome-aggregate Average Nucleotide and Amino Acid Identity (ANI/AAI) concepts. MiGA integrates best practices in sequence quality trimming and assembly and allows input to be raw reads or assemblies from isolate genomes, single-cell sequences, and metagenome-assembled genomes (MAGs). Further, MiGA can take as input hundreds of closely related genomes of the same or closely related species (a so-called 'Clade Project') to assess their gene content diversity and evolutionary relationships, and calculate important clade properties such as the pangenome and core gene sets. Therefore, MiGA is expected to facilitate a range of genome-based taxonomic and diversity studies, and quality assessment across environmental and clinical settings. MiGA is available at http://microbial-genomes.org/.

Land scale biogeography of arsenic biotransformation genes in estuarine wetland.

PubMed

Zhang, Si-Yu; Su, Jian-Qiang; Sun, Guo-Xin; Yang, Yunfeng; Zhao, Yi; Ding, Junjun; Chen, Yong-Shan; Shen, Yu; Zhu, Guibing; Rensing, Christopher; Zhu, Yong-Guan

2017-06-01

As an analogue of phosphorus, arsenic (As) has a biogeochemical cycle coupled closely with other key elements on the Earth, such as iron, sulfate and phosphate. It has been documented that microbial genes associated with As biotransformation are widely present in As-rich environments. Nonetheless, their presence in natural environment with low As levels remains unclear. To address this issue, we investigated the abundance levels and diversities of aioA, arrA, arsC and arsM genes in estuarine sediments at low As levels across Southeastern China to uncover biogeographic patterns at a large spatial scale. Unexpectedly, genes involved in As biotransformation were characterized by high abundance and diversity. The functional microbial communities showed a significant decrease in similarity along the geographic distance, with higher turnover rates than taxonomic microbial communities based on the similarities of 16S rRNA genes. Further investigation with niche-based models showed that deterministic processes played primary roles in shaping both functional and taxonomic microbial communities. Temperature, pH, total nitrogen concentration, carbon/nitrogen ratio and ferric iron concentration rather than As content in these sediments were significantly linked to functional microbial communities, while sediment temperature and pH were linked to taxonomic microbial communities. We proposed several possible mechanisms to explain these results. © 2017 Society for Applied Microbiology and John Wiley & Sons Ltd.

CVTree3 Web Server for Whole-genome-based and Alignment-free Prokaryotic Phylogeny and Taxonomy.

PubMed

Zuo, Guanghong; Hao, Bailin

2015-10-01

A faithful phylogeny and an objective taxonomy for prokaryotes should agree with each other and ultimately follow the genome data. With the number of sequenced genomes reaching tens of thousands, both tree inference and detailed comparison with taxonomy are great challenges. We now provide one solution in the latest Release 3.0 of the alignment-free and whole-genome-based web server CVTree3. The server resides in a cluster of 64 cores and is equipped with an interactive, collapsible, and expandable tree display. It is capable of comparing the tree branching order with prokaryotic classification at all taxonomic ranks from domains down to species and strains. CVTree3 allows for inquiry by taxon names and trial on lineage modifications. In addition, it reports a summary of monophyletic and non-monophyletic taxa at all ranks as well as produces print-quality subtree figures. After giving an overview of retrospective verification of the CVTree approach, the power of the new server is described for the mega-classification of prokaryotes and determination of taxonomic placement of some newly-sequenced genomes. A few discrepancies between CVTree and 16S rRNA analyses are also summarized with regard to possible taxonomic revisions. CVTree3 is freely accessible to all users at http://tlife.fudan.edu.cn/cvtree3/ without login requirements. Copyright © 2015 The Authors. Production and hosting by Elsevier Ltd.. All rights reserved.

CVTree3 Web Server for Whole-genome-based and Alignment-free Prokaryotic Phylogeny and Taxonomy

PubMed Central

Zuo, Guanghong; Hao, Bailin

2015-01-01

A faithful phylogeny and an objective taxonomy for prokaryotes should agree with each other and ultimately follow the genome data. With the number of sequenced genomes reaching tens of thousands, both tree inference and detailed comparison with taxonomy are great challenges. We now provide one solution in the latest Release 3.0 of the alignment-free and whole-genome-based web server CVTree3. The server resides in a cluster of 64 cores and is equipped with an interactive, collapsible, and expandable tree display. It is capable of comparing the tree branching order with prokaryotic classification at all taxonomic ranks from domains down to species and strains. CVTree3 allows for inquiry by taxon names and trial on lineage modifications. In addition, it reports a summary of monophyletic and non-monophyletic taxa at all ranks as well as produces print-quality subtree figures. After giving an overview of retrospective verification of the CVTree approach, the power of the new server is described for the mega-classification of prokaryotes and determination of taxonomic placement of some newly-sequenced genomes. A few discrepancies between CVTree and 16S rRNA analyses are also summarized with regard to possible taxonomic revisions. CVTree3 is freely accessible to all users at http://tlife.fudan.edu.cn/cvtree3/ without login requirements. PMID:26563468

High-Density 16S Microarray and Clone Library-Based Microbial Community Composition of the Phoenix Spacecraft Assembly Clean Room

NASA Astrophysics Data System (ADS)

Vaishampayan, Parag; Osman, Shariff; Andersen, Gary; Venkateswaran, Kasthuri

2010-06-01

The bacterial diversity and comparative community structure of a clean room used for assembling the Phoenix spacecraft was characterized throughout the spacecraft assembly process by using 16S rRNA gene cloning/sequencing and DNA microarray (PhyloChip) technologies. Samples were collected from several locations of the clean room at three time points: before Phoenix's arrival (PHX-B), during hardware assembly (PHX-D), and after the spacecraft was removed for launch (PHX-A). Bacterial diversity comprised of all major bacterial phyla of PHX-B was found to be statistically different from PHX-D and PHX-A samples. Due to stringent cleaning and decontamination protocols during assembly, PHX-D bacterial diversity was dramatically reduced when compared to PHX-B and PHX-A samples. Comparative community analysis based on PhyloChip results revealed similar overall trends as were seen in clone libraries, but the high-density phylogenetic microarray detected larger diversity in all sampling events. The decrease in community complexity in PHX-D compared to PHX-B, and the subsequent recurrence of these organisms in PHX-A, speaks to the effectiveness of NASA cleaning protocols. However, the persistence of a subset of bacterial signatures throughout all spacecraft assembly phases underscores the need for continued refinement of sterilization technologies and the implementation of safeguards that monitor and inventory microbial contaminants.

High-density 16S microarray and clone library-based microbial community composition of the Phoenix spacecraft assembly clean room.

PubMed

Vaishampayan, Parag; Osman, Shariff; Andersen, Gary; Venkateswaran, Kasthuri

2010-06-01

The bacterial diversity and comparative community structure of a clean room used for assembling the Phoenix spacecraft was characterized throughout the spacecraft assembly process by using 16S rRNA gene cloning/sequencing and DNA microarray (PhyloChip) technologies. Samples were collected from several locations of the clean room at three time points: before Phoenix's arrival (PHX-B), during hardware assembly (PHX-D), and after the spacecraft was removed for launch (PHX-A). Bacterial diversity comprised of all major bacterial phyla of PHX-B was found to be statistically different from PHX-D and PHX-A samples. Due to stringent cleaning and decontamination protocols during assembly, PHX-D bacterial diversity was dramatically reduced when compared to PHX-B and PHX-A samples. Comparative community analysis based on PhyloChip results revealed similar overall trends as were seen in clone libraries, but the high-density phylogenetic microarray detected larger diversity in all sampling events. The decrease in community complexity in PHX-D compared to PHX-B, and the subsequent recurrence of these organisms in PHX-A, speaks to the effectiveness of NASA cleaning protocols. However, the persistence of a subset of bacterial signatures throughout all spacecraft assembly phases underscores the need for continued refinement of sterilization technologies and the implementation of safeguards that monitor and inventory microbial contaminants.

Assessing macroinvertebrate biodiversity in freshwater ecosystems: Advances and challenges in dna-based approaches

USGS Publications Warehouse

Pfrender, M.E.; Ferrington, L.C.; Hawkins, C.P.; Hartzell, P.L.; Bagley, M.; Jackson, S.; Courtney, G.W.; Larsen, D.P.; Creutzburg, B.R.; Levesque, C.A.; Epler, J.H.; Morse, J.C.; Fend, S.; Petersen, M.J.; Ruiter, D.; Schindel, D.; Whiting, M.

2010-01-01

Assessing the biodiversity of macroinvertebrate fauna in freshwater ecosystems is an essential component of both basic ecological inquiry and applied ecological assessments. Aspects of taxonomic diversity and composition in freshwater communities are widely used to quantify water quality and measure the efficacy of remediation and restoration efforts. The accuracy and precision of biodiversity assessments based on standard morphological identifications are often limited by taxonomic resolution and sample size. Morphologically based identifications are laborious and costly, significantly constraining the sample sizes that can be processed. We suggest that the development of an assay platform based on DNA signatures will increase the precision and ease of quantifying biodiversity in freshwater ecosystems. Advances in this area will be particularly relevant for benthic and planktonic invertebrates, which are often monitored by regulatory agencies. Adopting a genetic assessment platform will alleviate some of the current limitations to biodiversity assessment strategies. We discuss the benefits and challenges associated with DNA-based assessments and the methods that are currently available. As recent advances in microarray and next-generation sequencing technologies will facilitate a transition to DNA-based assessment approaches, future research efforts should focus on methods for data collection, assay platform development, establishing linkages between DNA signatures and well-resolved taxonomies, and bioinformatics. ?? 2010 by The University of Chicago Press.

Cryptic Species in Tropic Sands - Interactive 3D Anatomy, Molecular Phylogeny and Evolution of Meiofaunal Pseudunelidae (Gastropoda, Acochlidia)

PubMed Central

Neusser, Timea P.; Jörger, Katharina M.; Schrödl, Michael

2011-01-01

Background Towards realistic estimations of the diversity of marine animals, tiny meiofaunal species usually are underrepresented. Since the biological species concept is hardly applicable on exotic and elusive animals, it is even more important to apply a morphospecies concept on the best level of information possible, using accurate and efficient methodology such as 3D modelling from histological sections. Molecular approaches such as sequence analyses may reveal further, cryptic species. This is the first case study on meiofaunal gastropods to test diversity estimations from traditional taxonomy against results from modern microanatomical methodology and molecular systematics. Results The examined meiofaunal Pseudunela specimens from several Indo-Pacific islands cannot be distinguished by external features. Their 3D microanatomy shows differences in the organ systems and allows for taxonomic separation in some cases. Additional molecular analyses based on partial mitochondrial cytochrome c oxidase subunit I (COI) and 16S rRNA markers revealed considerable genetic structure that is largely congruent with anatomical or geographical patterns. Two new species (Pseudunela viatoris and P. marteli spp. nov.) are formally described integrating morphological and genetic analyses. Phylogenetic analysis using partial 16S rRNA, COI and the nuclear 18S rRNA markers shows a clade of Pseudunelidae species as the sister group to limnic Acochlidiidae. Within Pseudunela, two subtypes of complex excretory systems occur. A complex kidney already evolved in the ancestor of Hedylopsacea. Several habitat shifts occurred during hedylopsacean evolution. Conclusions Cryptic species occur in tropical meiofaunal Pseudunela gastropods, and likely in other meiofaunal groups with poor dispersal abilities, boosting current diversity estimations. Only a combined 3D microanatomical and molecular approach revealed actual species diversity within Pseudunela reliably. Such integrative methods are recommended for all taxonomic approaches and biodiversity surveys on soft-bodied and small-sized invertebrates. With increasing taxon sampling and details studied, the evolution of acochlidian panpulmonates is even more complex than expected. PMID:21912592

Metagenomic and metabolic profiling of nonlithifying and lithifying stromatolitic mats of Highborne Cay, The Bahamas.

PubMed

Khodadad, Christina L M; Foster, Jamie S

2012-01-01

Stromatolites are laminated carbonate build-ups formed by the metabolic activity of microbial mats and represent one of the oldest known ecosystems on Earth. In this study, we examined a living stromatolite located within the Exuma Sound, The Bahamas and profiled the metagenome and metabolic potential underlying these complex microbial communities. The metagenomes of the two dominant stromatolitic mat types, a nonlithifying (Type 1) and lithifying (Type 3) microbial mat, were partially sequenced and compared. This deep-sequencing approach was complemented by profiling the substrate utilization patterns of the mats using metabolic microarrays. Taxonomic assessment of the protein-encoding genes confirmed previous SSU rRNA analyses that bacteria dominate the metagenome of both mat types. Eukaryotes comprised less than 13% of the metagenomes and were rich in sequences associated with nematodes and heterotrophic protists. Comparative genomic analyses of the functional genes revealed extensive similarities in most of the subsystems between the nonlithifying and lithifying mat types. The one exception was an increase in the relative abundance of certain genes associated with carbohydrate metabolism in the lithifying Type 3 mats. Specifically, genes associated with the degradation of carbohydrates commonly found in exopolymeric substances, such as hexoses, deoxy- and acidic sugars were found. The genetic differences in carbohydrate metabolisms between the two mat types were confirmed using metabolic microarrays. Lithifying mats had a significant increase in diversity and utilization of carbon, nitrogen, phosphorus and sulfur substrates. The two stromatolitic mat types retained similar microbial communities, functional diversity and many genetic components within their metagenomes. However, there were major differences detected in the activity and genetic pathways of organic carbon utilization. These differences provide a strong link between the metagenome and the physiology of the mats, as well as new insights into the biological processes associated with carbonate precipitation in modern marine stromatolites.

Nocardiopsis potens sp. nov., isolated from household waste.

PubMed

Yassin, A F; Spröer, C; Hupfer, H; Siering, C; Klenk, H-P

2009-11-01

The taxonomic position of an actinomycete, designated strain IMMIB L-21(T), was determined using a polyphasic taxonomic approach. The organism, which had phenotypic properties consistent with its classification in the genus Nocardiopsis, formed a distinct clade in the 16S rRNA gene sequence tree together with the type strain of Nocardiopsis composta, but was readily distinguished from this species using DNA-DNA relatedness and phenotypic data. The genotypic and phenotypic data show that the organism represents a novel species of the genus Nocardiopsis, for which the name Nocardiopsis potens sp. nov. is proposed. The type strain is IMMIB L-21(T) (=DSM 45234(T)=CCUG 56587(T)).

Correcting names of bacteria deposited in National Microbial Repositories: an analysed sequence data necessary for taxonomic re-categorization of misclassified bacteria-ONE example, genus Lysinibacillus.

PubMed

Rekadwad, Bhagwan N; Gonzalez, Juan M

2017-08-01

A report on 16S rRNA gene sequence re-analysis and digitalization is presented using Lysinibacillus species (one example) deposited in National Microbial Repositories in India. Lysinibacillus species 16S rRNA gene sequences were digitalized to provide quick response (QR) codes, Chaose Game Representation (CGR) and Frequency of Chaose Game Representation (FCGR). GC percentage, phylogenetic analysis, and principal component analysis (PCA) are tools used for the differentiation and reclassification of the strains under investigation. The seven reasons supporting the statements made by us as misclassified Lysinibacillus species deposited in National Microbial Depositories are given in this paper. Based on seven reasons, bacteria deposited in National Microbial Repositories such as Lysinibacillus and many other needs reanalyses for their exact identity. Leaves of identity with type strains of related species shows difference 2 to 8 % suggesting that reclassification is needed to correctly assign species names to the analyzed Lysinibacillus strains available in National Microbial Repositories.

Detection of microbial communities in continuous and discontinuous membrane bioreactor using high-density oligonucleotide Microarray

NASA Astrophysics Data System (ADS)

Duan, Liang; Song, Yonghui; Xia, Siqing; Hermanowicz, Slawomir W.

2010-11-01

This study compared the whole composition of microbial communities in continuous-flow (MBR) and batch-fed (discontinuous) (MSBR) aerobic membrane bioreactors using high-density universal 16S rRNA Microarray. The array includes 506,944 probes targeted to 8935 clusters in 16S rRNA gene sequences. The Microarray results showed that both MBR and MSBR had high microbial diversity. 1126 and 1002 bacterial subfamilies were detected and can separate as 37 and 32 phyla in MBR and MSBR, respectively. Proteobacteria was the predominant phylum, 703 and 597 subfamilies were found in two systems, which constituted 62.4% and 59.6% of the whole bacteria. Gamma- and Alpha-were the dominant classes in Proteobacteria. It occupied 38.1% and 26.3%, 31.2% and 39.2% for MBR and MSBR, respectively. Bacteroidetes, Firmicutes and Actinobacteria were the subdominant groups, occupying around 9.4% and 7.6%, 6.1% and 6.5%, 6.0% and 9.0% of the total bacteria in two reactors. Some bacterial groups such as Acidobacteria, Chloroflexi, Cyanobacteria, Verrucomicrobia and Spirochaetes also found more than 15 subfamilies. All the results indicated that the MBR system had more bacteria community diversity than MSBR's. Moreover, it was very interested that MBR and MSBR had almost the same bacterial composition except Enterobacteriaceae. 63 OTUs of Enterobacteriaceae were detected in MBR, while just 10 OTUs were found in MSBR. That's one of the reasons leading to the difference of the bacterial diversity between two bioreactors.

Protein relative abundance patterns associated with sucrose-induced dysbiosis are conserved across taxonomically diverse oral microcosm biofilm models of dental caries.

PubMed

Rudney, Joel D; Jagtap, Pratik D; Reilly, Cavan S; Chen, Ruoqiong; Markowski, Todd W; Higgins, LeeAnn; Johnson, James E; Griffin, Timothy J

2015-12-19

The etiology of dental caries is multifactorial, but frequent consumption of free sugars, notably sucrose, appears to be a major factor driving the supragingival microbiota in the direction of dysbiosis. Recent 16S rRNA-based studies indicated that caries-associated communities were less diverse than healthy supragingival plaque but still displayed considerable taxonomic diversity between individuals. Metagenomic studies likewise have found that healthy oral sites from different people were broadly similar with respect to gene function, even though there was an extensive individual variation in their taxonomic profiles. That pattern may also extend to dysbiotic communities. In that case, shifts in community-wide protein relative abundance might provide better biomarkers of dysbiosis that can be achieved through taxonomy alone. In this study, we used a paired oral microcosm biofilm model of dental caries to investigate differences in community composition and protein relative abundance in the presence and absence of sucrose. This approach provided large quantities of protein, which facilitated deep metaproteomic analysis. Community composition was evaluated using 16S rRNA sequencing and metaproteomic approaches. Although taxonomic diversity was reduced by sucrose pulsing, considerable inter-subject variation in community composition remained. By contrast, functional analysis using the SEED ontology found that sucrose induced changes in protein relative abundance patterns for pathways involving glycolysis, lactate production, aciduricity, and ammonia/glutamate metabolism that were conserved across taxonomically diverse dysbiotic oral microcosm biofilm communities. Our findings support the concept of using function-based changes in protein relative abundance as indicators of dysbiosis. Our microcosm model cannot replicate all aspects of the oral environment, but the deep level of metaproteomic analysis it allows makes it suitable for discovering which proteins are most consistently abundant during dysbiosis. It then may be possible to define biomarkers that could be used to detect at-risk tooth surfaces before the development of overt carious lesions.

A polyphasic taxonomic approach in isolated strains of Cyanobacteria from thermal springs of Greece.

PubMed

Bravakos, Panos; Kotoulas, Georgios; Skaraki, Katerina; Pantazidou, Adriani; Economou-Amilli, Athena

2016-05-01

Strains of Cyanobacteria isolated from mats of 9 thermal springs of Greece have been studied for their taxonomic evaluation. A polyphasic taxonomic approach was employed which included: morphological observations by light microscopy and scanning electron microscopy, maximum parsimony, maximum likelihood and Bayesian analysis of 16S rDNA sequences, secondary structural comparisons of 16S-23S rRNA Internal Transcribed Spacer sequences, and finally environmental data. The 17 cyanobacterial isolates formed a diverse group that contained filamentous, coccoid and heterocytous strains. These included representatives of the polyphyletic genera of Synechococcus and Phormidium, and the orders Oscillatoriales, Spirulinales, Chroococcales and Nostocales. After analysis, at least 6 new taxa at the genus level provide new evidence in the taxonomy of Cyanobacteria and highlight the abundant diversity of thermal spring environments with many potential endemic species or ecotypes. Copyright © 2016 Elsevier Inc. All rights reserved.

The effect of column purification on cDNA indirect labelling for microarrays

PubMed Central

Molas, M Lia; Kiss, John Z

2007-01-01

Background The success of the microarray reproducibility is dependent upon the performance of standardized procedures. Since the introduction of microarray technology for the analysis of global gene expression, reproducibility of results among different laboratories has been a major problem. Two of the main contributors to this variability are the use of different microarray platforms and different laboratory practices. In this paper, we address the latter question in terms of how variation in one of the steps of a labelling procedure affects the cDNA product prior to microarray hybridization. Results We used a standard procedure to label cDNA for microarray hybridization and employed different types of column chromatography for cDNA purification. After purifying labelled cDNA, we used the Agilent 2100 Bioanalyzer and agarose gel electrophoresis to assess the quality of the labelled cDNA before its hybridization onto a microarray platform. There were major differences in the cDNA profile (i.e. cDNA fragment lengths and abundance) as a result of using four different columns for purification. In addition, different columns have different efficiencies to remove rRNA contamination. This study indicates that the appropriate column to use in this type of protocol has to be experimentally determined. Finally, we present new evidence establishing the importance of testing the method of purification used during an indirect labelling procedure. Our results confirm the importance of assessing the quality of the sample in the labelling procedure prior to hybridization onto a microarray platform. Conclusion Standardization of column purification systems to be used in labelling procedures will improve the reproducibility of microarray results among different laboratories. In addition, implementation of a quality control check point of the labelled samples prior to microarray hybridization will prevent hybridizing a poor quality sample to expensive micorarrays. PMID:17597522

The effect of column purification on cDNA indirect labelling for microarrays.

PubMed

Molas, M Lia; Kiss, John Z

2007-06-27

The success of the microarray reproducibility is dependent upon the performance of standardized procedures. Since the introduction of microarray technology for the analysis of global gene expression, reproducibility of results among different laboratories has been a major problem. Two of the main contributors to this variability are the use of different microarray platforms and different laboratory practices. In this paper, we address the latter question in terms of how variation in one of the steps of a labelling procedure affects the cDNA product prior to microarray hybridization. We used a standard procedure to label cDNA for microarray hybridization and employed different types of column chromatography for cDNA purification. After purifying labelled cDNA, we used the Agilent 2100 Bioanalyzer and agarose gel electrophoresis to assess the quality of the labelled cDNA before its hybridization onto a microarray platform. There were major differences in the cDNA profile (i.e. cDNA fragment lengths and abundance) as a result of using four different columns for purification. In addition, different columns have different efficiencies to remove rRNA contamination. This study indicates that the appropriate column to use in this type of protocol has to be experimentally determined. Finally, we present new evidence establishing the importance of testing the method of purification used during an indirect labelling procedure. Our results confirm the importance of assessing the quality of the sample in the labelling procedure prior to hybridization onto a microarray platform. Standardization of column purification systems to be used in labelling procedures will improve the reproducibility of microarray results among different laboratories. In addition, implementation of a quality control check point of the labelled samples prior to microarray hybridization will prevent hybridizing a poor quality sample to expensive micorarrays.

[Effect of fluoride on gut microflora of silkworm (Bombyx mori)].

PubMed

Li, Guannan; Xia, Xuejuan; Sendegeya, Parfait; Zhao, Huanhuan; Long, Yaohang; Zhu, Yong

2015-07-04

We examined the effect of fluoride on gut microflora of silkworm. After DNA extraction and PCR amplification, clone libraries of 16S rRNA gene fragment were constructed. Amplified ribosomal DNA restriction analysis (ARDRA) was performed by digestion of the 16S rRNA gene, and each unique restriction fragment polymorphism pattern was designated as an operational taxonomic unit (OTU). A total of 14 OTUs were identified from intestinal samples of both T6 and 734. Phylogenetic trees of bacterial 16S rRNA nucleotide sequences were constructed and analyzed. Furthermore, the dominant bacteria were studied by the nested polymerase chain reaction denaturing gradient gel electrophoresis (PCR-DDGE) technology. After fluorosis, the flora of Enterococcus and Bacillus reduced. However, the flora of Staphylococcus increased. Fluoride can destroy the balance of microflora in the gut of silkworm by changing the bacteria diversity and proportion, which has bigger effect to 734 than T6.

Diversity of anaerobic microbes in spacecraft assembly clean rooms.

PubMed

Probst, Alexander; Vaishampayan, Parag; Osman, Shariff; Moissl-Eichinger, Christine; Andersen, Gary L; Venkateswaran, Kasthuri

2010-05-01

Although the cultivable and noncultivable microbial diversity of spacecraft assembly clean rooms has been previously documented using conventional and state-of-the-art molecular techniques, the occurrence of obligate anaerobes within these clean rooms is still uncertain. Therefore, anaerobic bacterial communities of three clean-room facilities were analyzed during assembly of the Mars Science Laboratory rover. Anaerobic bacteria were cultured on several media, and DNA was extracted from suitable anaerobic enrichments and examined with conventional 16S rRNA gene clone library, as well as high-density phylogenetic 16S rRNA gene microarray (PhyloChip) technologies. The culture-dependent analyses predominantly showed the presence of clostridial and propionibacterial strains. The 16S rRNA gene sequences retrieved from clone libraries revealed distinct microbial populations associated with each clean-room facility, clustered exclusively within gram-positive organisms. PhyloChip analysis detected a greater microbial diversity, spanning many phyla of bacteria, and provided a deeper insight into the microbial community structure of the clean-room facilities. This study presents an integrated approach for assessing the anaerobic microbial population within clean-room facilities, using both molecular and cultivation-based analyses. The results reveal that highly diverse anaerobic bacterial populations persist in the clean rooms even after the imposition of rigorous maintenance programs and will pose a challenge to planetary protection implementation activities.

CLUSTOM-CLOUD: In-Memory Data Grid-Based Software for Clustering 16S rRNA Sequence Data in the Cloud Environment.

PubMed

Oh, Jeongsu; Choi, Chi-Hwan; Park, Min-Kyu; Kim, Byung Kwon; Hwang, Kyuin; Lee, Sang-Heon; Hong, Soon Gyu; Nasir, Arshan; Cho, Wan-Sup; Kim, Kyung Mo

2016-01-01

High-throughput sequencing can produce hundreds of thousands of 16S rRNA sequence reads corresponding to different organisms present in the environmental samples. Typically, analysis of microbial diversity in bioinformatics starts from pre-processing followed by clustering 16S rRNA reads into relatively fewer operational taxonomic units (OTUs). The OTUs are reliable indicators of microbial diversity and greatly accelerate the downstream analysis time. However, existing hierarchical clustering algorithms that are generally more accurate than greedy heuristic algorithms struggle with large sequence datasets. To keep pace with the rapid rise in sequencing data, we present CLUSTOM-CLOUD, which is the first distributed sequence clustering program based on In-Memory Data Grid (IMDG) technology-a distributed data structure to store all data in the main memory of multiple computing nodes. The IMDG technology helps CLUSTOM-CLOUD to enhance both its capability of handling larger datasets and its computational scalability better than its ancestor, CLUSTOM, while maintaining high accuracy. Clustering speed of CLUSTOM-CLOUD was evaluated on published 16S rRNA human microbiome sequence datasets using the small laboratory cluster (10 nodes) and under the Amazon EC2 cloud-computing environments. Under the laboratory environment, it required only ~3 hours to process dataset of size 200 K reads regardless of the complexity of the human microbiome data. In turn, one million reads were processed in approximately 20, 14, and 11 hours when utilizing 20, 30, and 40 nodes on the Amazon EC2 cloud-computing environment. The running time evaluation indicates that CLUSTOM-CLOUD can handle much larger sequence datasets than CLUSTOM and is also a scalable distributed processing system. The comparative accuracy test using 16S rRNA pyrosequences of a mock community shows that CLUSTOM-CLOUD achieves higher accuracy than DOTUR, mothur, ESPRIT-Tree, UCLUST and Swarm. CLUSTOM-CLOUD is written in JAVA and is freely available at http://clustomcloud.kopri.re.kr.

CLUSTOM-CLOUD: In-Memory Data Grid-Based Software for Clustering 16S rRNA Sequence Data in the Cloud Environment

PubMed Central

Park, Min-Kyu; Kim, Byung Kwon; Hwang, Kyuin; Lee, Sang-Heon; Hong, Soon Gyu; Nasir, Arshan; Cho, Wan-Sup; Kim, Kyung Mo

2016-01-01

High-throughput sequencing can produce hundreds of thousands of 16S rRNA sequence reads corresponding to different organisms present in the environmental samples. Typically, analysis of microbial diversity in bioinformatics starts from pre-processing followed by clustering 16S rRNA reads into relatively fewer operational taxonomic units (OTUs). The OTUs are reliable indicators of microbial diversity and greatly accelerate the downstream analysis time. However, existing hierarchical clustering algorithms that are generally more accurate than greedy heuristic algorithms struggle with large sequence datasets. To keep pace with the rapid rise in sequencing data, we present CLUSTOM-CLOUD, which is the first distributed sequence clustering program based on In-Memory Data Grid (IMDG) technology–a distributed data structure to store all data in the main memory of multiple computing nodes. The IMDG technology helps CLUSTOM-CLOUD to enhance both its capability of handling larger datasets and its computational scalability better than its ancestor, CLUSTOM, while maintaining high accuracy. Clustering speed of CLUSTOM-CLOUD was evaluated on published 16S rRNA human microbiome sequence datasets using the small laboratory cluster (10 nodes) and under the Amazon EC2 cloud-computing environments. Under the laboratory environment, it required only ~3 hours to process dataset of size 200 K reads regardless of the complexity of the human microbiome data. In turn, one million reads were processed in approximately 20, 14, and 11 hours when utilizing 20, 30, and 40 nodes on the Amazon EC2 cloud-computing environment. The running time evaluation indicates that CLUSTOM-CLOUD can handle much larger sequence datasets than CLUSTOM and is also a scalable distributed processing system. The comparative accuracy test using 16S rRNA pyrosequences of a mock community shows that CLUSTOM-CLOUD achieves higher accuracy than DOTUR, mothur, ESPRIT-Tree, UCLUST and Swarm. CLUSTOM-CLOUD is written in JAVA and is freely available at http://clustomcloud.kopri.re.kr. PMID:26954507

'Candidatus Phytoplasma palmicola', associated with a lethal yellowing-type disease of coconut (Cocos nucifera L.) in Mozambique.

PubMed

Harrison, Nigel A; Davis, Robert E; Oropeza, Carlos; Helmick, Ericka E; Narváez, María; Eden-Green, Simon; Dollet, Michel; Dickinson, Matthew

2014-06-01

In this study, the taxonomic position and group classification of the phytoplasma associated with a lethal yellowing-type disease (LYD) of coconut (Cocos nucifera L.) in Mozambique were addressed. Pairwise similarity values based on alignment of nearly full-length 16S rRNA gene sequences (1530 bp) revealed that the Mozambique coconut phytoplasma (LYDM) shared 100% identity with a comparable sequence derived from a phytoplasma strain (LDN) responsible for Awka wilt disease of coconut in Nigeria, and shared 99.0-99.6% identity with 16S rRNA gene sequences from strains associated with Cape St Paul wilt (CSPW) disease of coconut in Ghana and Côte d'Ivoire. Similarity scores further determined that the 16S rRNA gene of the LYDM phytoplasma shared <97.5% sequence identity with all previously described members of 'Candidatus Phytoplasma'. The presence of unique regions in the 16S rRNA gene sequence distinguished the LYDM phytoplasma from all currently described members of 'Candidatus Phytoplasma', justifying its recognition as the reference strain of a novel taxon, 'Candidatus Phytoplasma palmicola'. Virtual RFLP profiles of the F2n/R2 portion (1251 bp) of the 16S rRNA gene and pattern similarity coefficients delineated coconut LYDM phytoplasma strains from Mozambique as novel members of established group 16SrXXII, subgroup A (16SrXXII-A). Similarity coefficients of 0.97 were obtained for comparisons between subgroup 16SrXXII-A strains and CSPW phytoplasmas from Ghana and Côte d'Ivoire. On this basis, the CSPW phytoplasma strains were designated members of a novel subgroup, 16SrXXII-B.

Dynamic of active microorganisms inhabiting a bioleaching industrial heap of low‐grade copper sulfide ore monitored by real‐time PCR and oligonucleotide prokaryotic acidophile microarray

PubMed Central

Remonsellez, Francisco; Galleguillos, Felipe; Moreno‐Paz, Mercedes; Parro, Víctor; Acosta, Mauricio; Demergasso, Cecilia

2009-01-01

Summary The bioleaching of metal sulfide has developed into a very important industrial process and understanding the microbial dynamic is key to advancing commercial bioleaching operations. Here we report the first quantitative description of the dynamic of active communities in an industrial bioleaching heap. Acidithiobacillus ferrooxidans was the most abundant during the first part of the leaching cycle, while the abundance of Leptospirillum ferriphilum and Ferroplasma acidiphilum increased with age of the heap. Acidithiobacillus thiooxidans kept constant throughout the leaching cycle, and Firmicutes group showed a low and a patchy distribution in the heap. The Acidiphilium‐like bacteria reached their highest abundance corresponding to the amount of autotrophs. The active microorganisms in the leaching system were determined using two RNA‐based sensitive techniques. In most cases, the 16S rRNA copy numbers of At. ferrooxidans, L. ferriphilum, At. thiooxidans and F. acidiphilum, was concomitant with the DNA copy numbers, whereas Acidiphilium‐like bacteria and some Firmicutes members did not show a clear correlation between 16S rRNA accumulation and DNA copy numbers. However, the prokaryotic acidophile microarray (PAM) analysis showed active members of Alphaproteobacteria in all samples and of Sulfobacillus genus in older ones. Also, new active groups such as Actinobacteria and Acidobacterium genus were detected by PAM. The results suggest that changes during the leaching cycle in chemical and physical conditions, such as pH and Fe3+/Fe2+ ion rate, are primary factors shaping the microbial dynamic in the heap. PMID:21255296

Diversity surveys and evolutionary relationships of aoxB genes in aerobic arsenite-oxidizing bacteria.

PubMed

Quéméneur, Marianne; Heinrich-Salmeron, Audrey; Muller, Daniel; Lièvremont, Didier; Jauzein, Michel; Bertin, Philippe N; Garrido, Francis; Joulian, Catherine

2008-07-01

A new primer set was designed to specifically amplify ca. 1,100 bp of aoxB genes encoding the As(III) oxidase catalytic subunit from taxonomically diverse aerobic As(III)-oxidizing bacteria. Comparative analysis of AoxB protein sequences showed variable conservation levels and highlighted the conservation of essential amino acids and structural motifs. AoxB phylogeny of pure strains showed well-discriminated taxonomic groups and was similar to 16S rRNA phylogeny. Alphaproteobacteria-, Betaproteobacteria-, and Gammaproteobacteria-related sequences were retrieved from environmental surveys, demonstrating their prevalence in mesophilic As-contaminated soils. Our study underlines the usefulness of the aoxB gene as a functional marker of aerobic As(III) oxidizers.

Cellulolytic Bacteria in the Foregut of the Dromedary Camel (Camelus dromedarius)

PubMed Central

Samsudin, Anjas A.; Wright, André-Denis G.

2012-01-01

Foregut digesta from five feral dromedary camels were inoculated into three different enrichment media: cotton thread, filter paper, and neutral detergent fiber. A total of 283 16S rRNA gene sequences were assigned to 33 operational taxonomic units by using 99% species-level identity. LIBSHUFF revealed significant differences in the community composition across all three libraries. PMID:23042173

Cellulolytic bacteria in the foregut of the dromedary camel (Camelus dromedarius).

PubMed

Samsudin, Anjas A; Wright, André-Denis G; Al Jassim, Rafat

2012-12-01

Foregut digesta from five feral dromedary camels were inoculated into three different enrichment media: cotton thread, filter paper, and neutral detergent fiber. A total of 283 16S rRNA gene sequences were assigned to 33 operational taxonomic units by using 99% species-level identity. LIBSHUFF revealed significant differences in the community composition across all three libraries.

Streptomyces asenjonii sp. nov., isolated from arid Atacama Desert soils and emended description of Streptomyces viridosporus Pridham et al. 1958

USDA-ARS?s Scientific Manuscript database

A polyphasic study was undertaken to establish the taxonomic status of Streptomyces strains isolated from arid Atacama Desert soils. Analysis of the 16S rRNA gene sequences of the isolates showed that they formed a well-defined lineage that was loosely associated with the type strains of several Str...

Metagenomic Evaluation of Bacterial and Archaeal Diversity in the Geothermal Hot Springs of Manikaran, India

PubMed Central

Pathak, Ashish; Green, Stefan J.; Joshi, Amit; Chauhan, Ashvini

2015-01-01

Bacterial and archaeal diversity in geothermal spring water were investigated using 16S rRNA gene amplicon metagenomic sequencing. This revealed the dominance of Firmicutes, Aquificae, and the Deinococcus-Thermus group in this thermophilic environment. A number of sequences remained taxonomically unresolved, indicating the presence of potentially novel microbes in this unique habitat. PMID:25700403

Animal Viruses Probe dataset (AVPDS) for microarray-based diagnosis and identification of viruses.

PubMed

Yadav, Brijesh S; Pokhriyal, Mayank; Vasishtha, Dinesh P; Sharma, Bhaskar

2014-03-01

AVPDS (Animal Viruses Probe dataset) is a dataset of virus-specific and conserve oligonucleotides for identification and diagnosis of viruses infecting animals. The current dataset contain 20,619 virus specific probes for 833 viruses and their subtypes and 3,988 conserved probes for 146 viral genera. Dataset of virus specific probe has been divided into two fields namely virus name and probe sequence. Similarly conserved probes for virus genera table have genus, and subgroup within genus name and probe sequence. The subgroup within genus is artificially divided subgroups with no taxonomic significance and contains probes which identifies viruses in that specific subgroup of the genus. Using this dataset we have successfully diagnosed the first case of Newcastle disease virus in sheep and reported a mixed infection of Bovine viral diarrhea and Bovine herpesvirus in cattle. These dataset also contains probes which cross reacts across species experimentally though computationally they meet specifications. These probes have been marked. We hope that this dataset will be useful in microarray-based detection of viruses. The dataset can be accessed through the link https://dl.dropboxusercontent.com/u/94060831/avpds/HOME.html.

Taxonomic structure and stability of the bacterial community in belgian sourdough ecosystems as assessed by culture and population fingerprinting.

PubMed

Scheirlinck, Ilse; Van der Meulen, Roel; Van Schoor, Ann; Vancanneyt, Marc; De Vuyst, Luc; Vandamme, Peter; Huys, Geert

2008-04-01

A total of 39 traditional sourdoughs were sampled at 11 bakeries located throughout Belgium which were visited twice with a 1-year interval. The taxonomic structure and stability of the bacterial communities occurring in these traditional sourdoughs were assessed using both culture-dependent and culture-independent methods. A total of 1,194 potential lactic acid bacterium (LAB) isolates were tentatively grouped and identified by repetitive element sequence-based PCR, followed by sequence-based identification using 16S rRNA and pheS genes from a selection of genotypically unique LAB isolates. In parallel, all samples were analyzed by denaturing gradient gel electrophoresis (DGGE) of V3-16S rRNA gene amplicons. In addition, extensive metabolite target analysis of more than 100 different compounds was performed. Both culturing and DGGE analysis showed that the species Lactobacillus sanfranciscensis, Lactobacillus paralimentarius, Lactobacillus plantarum, and Lactobacillus pontis dominated the LAB population of Belgian type I sourdoughs. In addition, DGGE band sequence analysis demonstrated the presence of Acetobacter sp. and a member of the Erwinia/Enterobacter/Pantoea group in some samples. Overall, the culture-dependent and culture-independent approaches each exhibited intrinsic limitations in assessing bacterial LAB diversity in Belgian sourdoughs. Irrespective of the LAB biodiversity, a large majority of the sugar and amino acid metabolites were detected in all sourdough samples. Principal component-based analysis of biodiversity and metabolic data revealed only little variation among the two samples of the sourdoughs produced at the same bakery. The rare cases of instability observed could generally be linked with variations in technological parameters or differences in detection capacity between culture-dependent and culture-independent approaches. Within a sampling interval of 1 year, this study reinforces previous observations that the bakery environment rather than the type or batch of flour largely determines the development of a stable LAB population in sourdoughs.

Taxonomic Structure and Stability of the Bacterial Community in Belgian Sourdough Ecosystems as Assessed by Culture and Population Fingerprinting▿ †

PubMed Central

Scheirlinck, Ilse; Van der Meulen, Roel; Van Schoor, Ann; Vancanneyt, Marc; De Vuyst, Luc; Vandamme, Peter; Huys, Geert

2008-01-01

A total of 39 traditional sourdoughs were sampled at 11 bakeries located throughout Belgium which were visited twice with a 1-year interval. The taxonomic structure and stability of the bacterial communities occurring in these traditional sourdoughs were assessed using both culture-dependent and culture-independent methods. A total of 1,194 potential lactic acid bacterium (LAB) isolates were tentatively grouped and identified by repetitive element sequence-based PCR, followed by sequence-based identification using 16S rRNA and pheS genes from a selection of genotypically unique LAB isolates. In parallel, all samples were analyzed by denaturing gradient gel electrophoresis (DGGE) of V3-16S rRNA gene amplicons. In addition, extensive metabolite target analysis of more than 100 different compounds was performed. Both culturing and DGGE analysis showed that the species Lactobacillus sanfranciscensis, Lactobacillus paralimentarius, Lactobacillus plantarum, and Lactobacillus pontis dominated the LAB population of Belgian type I sourdoughs. In addition, DGGE band sequence analysis demonstrated the presence of Acetobacter sp. and a member of the Erwinia/Enterobacter/Pantoea group in some samples. Overall, the culture-dependent and culture-independent approaches each exhibited intrinsic limitations in assessing bacterial LAB diversity in Belgian sourdoughs. Irrespective of the LAB biodiversity, a large majority of the sugar and amino acid metabolites were detected in all sourdough samples. Principal component-based analysis of biodiversity and metabolic data revealed only little variation among the two samples of the sourdoughs produced at the same bakery. The rare cases of instability observed could generally be linked with variations in technological parameters or differences in detection capacity between culture-dependent and culture-independent approaches. Within a sampling interval of 1 year, this study reinforces previous observations that the bakery environment rather than the type or batch of flour largely determines the development of a stable LAB population in sourdoughs. PMID:18310426

Fiber-optic microarray for simultaneous detection of multiple harmful algal bloom species.

PubMed

Ahn, Soohyoun; Kulis, David M; Erdner, Deana L; Anderson, Donald M; Walt, David R

2006-09-01

Harmful algal blooms (HABs) are a serious threat to coastal resources, causing a variety of impacts on public health, regional economies, and ecosystems. Plankton analysis is a valuable component of many HAB monitoring and research programs, but the diversity of plankton poses a problem in discriminating toxic from nontoxic species using conventional detection methods. Here we describe a sensitive and specific sandwich hybridization assay that combines fiber-optic microarrays with oligonucleotide probes to detect and enumerate the HAB species Alexandrium fundyense, Alexandrium ostenfeldii, and Pseudo-nitzschia australis. Microarrays were prepared by loading oligonucleotide probe-coupled microspheres (diameter, 3 mum) onto the distal ends of chemically etched imaging fiber bundles. Hybridization of target rRNA from HAB cells to immobilized probes on the microspheres was visualized using Cy3-labeled secondary probes in a sandwich-type assay format. We applied these microarrays to the detection and enumeration of HAB cells in both cultured and field samples. Our study demonstrated a detection limit of approximately 5 cells for all three target organisms within 45 min, without a separate amplification step, in both sample types. We also developed a multiplexed microarray to detect the three HAB species simultaneously, which successfully detected the target organisms, alone and in combination, without cross-reactivity. Our study suggests that fiber-optic microarrays can be used for rapid and sensitive detection and potential enumeration of HAB species in the environment.

Diversity of Ptychadena in Rwanda and taxonomic status of P. chrysogaster Laurent, 1954 (Amphibia, Anura, Ptychadenidae)

PubMed Central

Dehling, J. Maximilian; Sinsch, Ulrich

2013-01-01

Abstract We assess the diversity of Ptychadena species in Rwanda based on re-examination of voucher specimens in museum collections and our own data from recent assessment of the species composition of amphibian communities in Rwanda. We recognize five species which we allocate to the following available names: P. anchietae, P. chrysogaster, P. nilotica, P. porosissima, and P. uzungwensis. We did not find evidence for the presence of P. grandisonae and P. oxyrhynchus which have been listed for the country. The five species can be distinguished by quantitative morphometrics (discriminant analysis, success rate: 100 %) and a number of qualitative characters of external morphology. We provide an identification key to the Rwandan species and describe the morphology of each species in detail. The taxonomic status and the phylogenetic position of Ptychadena chrysogaster are further assessed based on the partial sequence of the mitochondrial 16S rRNA. The species differs genetically from available homologous sequences from congeners by an uncorrected p distance of at least 4.2 % and appears to be most closely related to specimens assigned to P. porosissima, P. mahnerti, “P. aff. uzungwensis” and “P. aff. bibroni”. PMID:24363574

Taxonomic research on deep-sea macrofauna in the South China Sea using the Chinese deep-sea submersible Jiaolong.

PubMed

Li, Xinzheng

2017-07-01

This paper reviews the taxonomic and biodiversity studies of deep-sea invertebrates in the South China Sea based on the samples collected by the Chinese manned deep-sea submersible Jiaolong. To date, 6 new species have been described, including the sponges Lophophysema eversa, Saccocalyx microhexactin and Semperella jiaolongae as well as the crustaceans Uroptychus jiaolongae, Uroptychus spinulosus and Globospongicola jiaolongi; some newly recorded species from the South China Sea have also been reported. The Bathymodiolus platifrons-Shinkaia crosnieri deep-sea cold seep community has been reported by Li (2015), as has the mitochondrial genome of the glass sponge L. eversa by Zhang et al. (2016). The population structures of two dominant species, the shrimp Shinkaia crosnieri and the mussel Bathymodiolus platifrons, from the cold seep Bathymodiolus platifrons-Shinkaia crosnieri community in the South China Sea and the hydrothermal vents in the Okinawa Trough, were compared using molecular analysis. The systematic position of the shrimp genus Globospongicola was discussed based on 16S rRNA gene sequences. © 2017 International Society of Zoological Sciences, Institute of Zoology/Chinese Academy of Sciences and John Wiley & Sons Australia, Ltd.

CATCh, an Ensemble Classifier for Chimera Detection in 16S rRNA Sequencing Studies

PubMed Central

Mysara, Mohamed; Saeys, Yvan; Leys, Natalie; Raes, Jeroen

2014-01-01

In ecological studies, microbial diversity is nowadays mostly assessed via the detection of phylogenetic marker genes, such as 16S rRNA. However, PCR amplification of these marker genes produces a significant amount of artificial sequences, often referred to as chimeras. Different algorithms have been developed to remove these chimeras, but efforts to combine different methodologies are limited. Therefore, two machine learning classifiers (reference-based and de novo CATCh) were developed by integrating the output of existing chimera detection tools into a new, more powerful method. When comparing our classifiers with existing tools in either the reference-based or de novo mode, a higher performance of our ensemble method was observed on a wide range of sequencing data, including simulated, 454 pyrosequencing, and Illumina MiSeq data sets. Since our algorithm combines the advantages of different individual chimera detection tools, our approach produces more robust results when challenged with chimeric sequences having a low parent divergence, short length of the chimeric range, and various numbers of parents. Additionally, it could be shown that integrating CATCh in the preprocessing pipeline has a beneficial effect on the quality of the clustering in operational taxonomic units. PMID:25527546

Optimizing taxonomic classification of marker-gene amplicon sequences with QIIME 2's q2-feature-classifier plugin.

PubMed

Bokulich, Nicholas A; Kaehler, Benjamin D; Rideout, Jai Ram; Dillon, Matthew; Bolyen, Evan; Knight, Rob; Huttley, Gavin A; Gregory Caporaso, J

2018-05-17

Taxonomic classification of marker-gene sequences is an important step in microbiome analysis. We present q2-feature-classifier ( https://github.com/qiime2/q2-feature-classifier ), a QIIME 2 plugin containing several novel machine-learning and alignment-based methods for taxonomy classification. We evaluated and optimized several commonly used classification methods implemented in QIIME 1 (RDP, BLAST, UCLUST, and SortMeRNA) and several new methods implemented in QIIME 2 (a scikit-learn naive Bayes machine-learning classifier, and alignment-based taxonomy consensus methods based on VSEARCH, and BLAST+) for classification of bacterial 16S rRNA and fungal ITS marker-gene amplicon sequence data. The naive-Bayes, BLAST+-based, and VSEARCH-based classifiers implemented in QIIME 2 meet or exceed the species-level accuracy of other commonly used methods designed for classification of marker gene sequences that were evaluated in this work. These evaluations, based on 19 mock communities and error-free sequence simulations, including classification of simulated "novel" marker-gene sequences, are available in our extensible benchmarking framework, tax-credit ( https://github.com/caporaso-lab/tax-credit-data ). Our results illustrate the importance of parameter tuning for optimizing classifier performance, and we make recommendations regarding parameter choices for these classifiers under a range of standard operating conditions. q2-feature-classifier and tax-credit are both free, open-source, BSD-licensed packages available on GitHub.

High-throughput amplicon sequencing and stream benthic bacteria: identifying the best taxonomic level for multiple-stressor research

PubMed Central

Salis, R. K.; Bruder, A.; Piggott, J. J.; Summerfield, T. C.; Matthaei, C. D.

2017-01-01

Disentangling the individual and interactive effects of multiple stressors on microbial communities is a key challenge to our understanding and management of ecosystems. Advances in molecular techniques allow studying microbial communities in situ and with high taxonomic resolution. However, the taxonomic level which provides the best trade-off between our ability to detect multiple-stressor effects versus the goal of studying entire communities remains unknown. We used outdoor mesocosms simulating small streams to investigate the effects of four agricultural stressors (nutrient enrichment, the nitrification inhibitor dicyandiamide (DCD), fine sediment and flow velocity reduction) on stream bacteria (phyla, orders, genera, and species represented by Operational Taxonomic Units with 97% sequence similarity). Community composition was assessed using amplicon sequencing (16S rRNA gene, V3-V4 region). DCD was the most pervasive stressor, affecting evenness and most abundant taxa, followed by sediment and flow velocity. Stressor pervasiveness was similar across taxonomic levels and lower levels did not perform better in detecting stressor effects. Community coverage decreased from 96% of all sequences for abundant phyla to 28% for species. Order-level responses were generally representative of responses of corresponding genera and species, suggesting that this level may represent the best compromise between stressor sensitivity and coverage of bacterial communities. PMID:28327636

[Isolation and phylogenetic analysis of one actinomycete strain YIM 90022 exhibiting anticancer activity].

PubMed

Chen, Yi-Guang; Li, Wen-Jun; Cui, Xiao-Long; Jiang, Cheng-Lin; Xu, Li-Hua

2006-10-01

One facultative alkaliphilic actinomycete strain YIM 90022 was isolated from hypersaline alkaline soil in Qinghai province, China. An almost-complete 16S rRNA gene sequence (1500 bp) for strain YIM 90022 was obtained. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain YIM 90022 was closely related to four members of the genus Nocardiopsis with 16S rRNA gene sequence similarity values of 98.8% (N. exhalans DSM 44407T), 98.5% (N. prasina DSM 43845T), 98.4% (N. metallicus DSM 44598T) and 97.8% (N. listeri DSM 40297T), but represented a distinct phylogenetic lineage. Repetitive element sequence-based PCR (rep-PCR) genomic fingerprinting was evaluated on strain YIM 90022 and its closest relatives to investigate their genetic relatedness. The analysis of the rep-PCR genomic fingerprints showed that strain YIM 90022 was distinguishable from its closest relatives. The polyphasic taxonomic data presented in this study, including its morphology, physiological and biochemical characteristics, chemotaxonomy, 16S rRNA gene sequence-based phylogenetic analysis and rep-PCR genomic fingerprinting, supported the view that strain YIM 90022 represented a potential new species of the genus Nocardiopsis. The fermentation broth of strain YIM 90022 strongly inhibited growth of cell series of gastric cancer, lung cancer, mammary cancer, melanoma cancer, renal cancer and uterus cancer. Strain YIM 90022 grew well on most tested media, producing exuberant vegetative hyphae and aerial hyphae. The vegetative hyphae are long and fragmented. Light yellow to deep brown diffusible pigments were produced on ISP 2, ISP 3 and ISP 6. Growth of the strain occurred in the pH range 6.0-12.0, with optimal pH8.5. The NaCl tolerate range was 0-15% (W/V). Cell walls contain meso-diaminopimelic acid and have no diagnostic sugars. Polar lipids are phosphatidylcholine, phosphatidylglycerol, diphosphatidylglycerol, phosphatidylmethylethanolamine. Major menaquinones are MK-10 (H4, H6). The DNA G + C content is 71.5 mol %.

In-silico Taxonomic Classification of 373 Genomes Reveals Species Misidentification and New Genospecies within the Genus Pseudomonas.

PubMed

Tran, Phuong N; Savka, Michael A; Gan, Han Ming

2017-01-01

The genus Pseudomonas has one of the largest diversity of species within the Bacteria kingdom. To date, its taxonomy is still being revised and updated. Due to the non-standardized procedure and ambiguous thresholds at species level, largely based on 16S rRNA gene or conventional biochemical assay, species identification of publicly available Pseudomonas genomes remains questionable. In this study, we performed a large-scale analysis of all Pseudomonas genomes with species designation (excluding the well-defined P. aeruginosa ) and re-evaluated their taxonomic assignment via in silico genome-genome hybridization and/or genetic comparison with valid type species. Three-hundred and seventy-three pseudomonad genomes were analyzed and subsequently clustered into 145 distinct genospecies. We detected 207 erroneous labels and corrected 43 to the proper species based on Average Nucleotide Identity Multilocus Sequence Typing (MLST) sequence similarity to the type strain. Surprisingly, more than half of the genomes initially designated as Pseudomonas syringae and Pseudomonas fluorescens should be classified either to a previously described species or to a new genospecies. Notably, high pairwise average nucleotide identity (>95%) indicating species-level similarity was observed between P. synxantha-P. libanensis, P. psychrotolerans - P. oryzihabitans , and P. kilonensis- P. brassicacearum , that were previously differentiated based on conventional biochemical tests and/or genome-genome hybridization techniques.

prepare_taxa_charts.py: A Python program to automate generation of publication ready taxonomic pie chart images from QIIME.

PubMed

Lakhujani, Vijay; Badapanda, Chandan

2017-06-01

QIIME (Quantitative Insights Into Microbial Ecology) is one of the most popular open-source bioinformatics suite for performing metagenome, 16S rRNA amplicon and Internal Transcribed Spacer (ITS) data analysis. Although, it is very comprehensive and powerful tool, it lacks a method to provide publication ready taxonomic pie charts. The script plot_taxa_summary . py bundled with QIIME generate a html file and a folder containing taxonomic pie chart and legend as separate images. The images have randomly generated alphanumeric names. Therefore, it is difficult to associate the pie chart with the legend and the corresponding sample identifier. Even if the option to have the legend within the html file is selected while executing plot_taxa_summary . py , it is very tedious to crop a complete image (having both the pie chart and the legend) due to unequal image sizes. It requires a lot of time to manually prepare the pie charts for multiple samples for publication purpose. Moreover, there are chances of error while identifying the pie chart and legend pair due to random alphanumeric names of the images. To bypass all these bottlenecks and make this process efficient, we have developed a python based program, prepare_taxa_charts . py , to automate the renaming, cropping and merging of taxonomic pie chart and corresponding legend image into a single, good quality publication ready image. This program not only augments the functionality of plot_taxa_summary . py but is also very fast in terms of CPU time and user friendly.

Development of a multilocus-based approach for sponge (phylum Porifera) identification: refinement and limitations.

PubMed

Yang, Qi; Franco, Christopher M M; Sorokin, Shirley J; Zhang, Wei

2017-02-02

For sponges (phylum Porifera), there is no reliable molecular protocol available for species identification. To address this gap, we developed a multilocus-based Sponge Identification Protocol (SIP) validated by a sample of 37 sponge species belonging to 10 orders from South Australia. The universal barcode COI mtDNA, 28S rRNA gene (D3-D5), and the nuclear ITS1-5.8S-ITS2 region were evaluated for their suitability and capacity for sponge identification. The highest Bit Score was applied to infer the identity. The reliability of SIP was validated by phylogenetic analysis. The 28S rRNA gene and COI mtDNA performed better than the ITS region in classifying sponges at various taxonomic levels. A major limitation is that the databases are not well populated and possess low diversity, making it difficult to conduct the molecular identification protocol. The identification is also impacted by the accuracy of the morphological classification of the sponges whose sequences have been submitted to the database. Re-examination of the morphological identification further demonstrated and improved the reliability of sponge identification by SIP. Integrated with morphological identification, the multilocus-based SIP offers an improved protocol for more reliable and effective sponge identification, by coupling the accuracy of different DNA markers.

Development of a multilocus-based approach for sponge (phylum Porifera) identification: refinement and limitations

PubMed Central

Yang, Qi; Franco, Christopher M. M.; Sorokin, Shirley J.; Zhang, Wei

2017-01-01

For sponges (phylum Porifera), there is no reliable molecular protocol available for species identification. To address this gap, we developed a multilocus-based Sponge Identification Protocol (SIP) validated by a sample of 37 sponge species belonging to 10 orders from South Australia. The universal barcode COI mtDNA, 28S rRNA gene (D3–D5), and the nuclear ITS1-5.8S-ITS2 region were evaluated for their suitability and capacity for sponge identification. The highest Bit Score was applied to infer the identity. The reliability of SIP was validated by phylogenetic analysis. The 28S rRNA gene and COI mtDNA performed better than the ITS region in classifying sponges at various taxonomic levels. A major limitation is that the databases are not well populated and possess low diversity, making it difficult to conduct the molecular identification protocol. The identification is also impacted by the accuracy of the morphological classification of the sponges whose sequences have been submitted to the database. Re-examination of the morphological identification further demonstrated and improved the reliability of sponge identification by SIP. Integrated with morphological identification, the multilocus-based SIP offers an improved protocol for more reliable and effective sponge identification, by coupling the accuracy of different DNA markers. PMID:28150727

Phylogenic inference using alignment-free methods for applications in microbial community surveys using 16s rRNA gene

PubMed Central

2017-01-01

The diversity of microbiota is best explored by understanding the phylogenetic structure of the microbial communities. Traditionally, sequence alignment has been used for phylogenetic inference. However, alignment-based approaches come with significant challenges and limitations when massive amounts of data are analyzed. In the recent decade, alignment-free approaches have enabled genome-scale phylogenetic inference. Here we evaluate three alignment-free methods: ACS, CVTree, and Kr for phylogenetic inference with 16s rRNA gene data. We use a taxonomic gold standard to compare the accuracy of alignment-free phylogenetic inference with that of common microbiome-wide phylogenetic inference pipelines based on PyNAST and MUSCLE alignments with FastTree and RAxML. We re-simulate fecal communities from Human Microbiome Project data to evaluate the performance of the methods on datasets with properties of real data. Our comparisons show that alignment-free methods are not inferior to alignment-based methods in giving accurate and robust phylogenic trees. Moreover, consensus ensembles of alignment-free phylogenies are superior to those built from alignment-based methods in their ability to highlight community differences in low power settings. In addition, the overall running times of alignment-based and alignment-free phylogenetic inference are comparable. Taken together our empirical results suggest that alignment-free methods provide a viable approach for microbiome-wide phylogenetic inference. PMID:29136663

Soil organic matter quantity and quality shape microbial community compositions of subtropical broadleaved forests.

PubMed

Ding, Junjun; Zhang, Yuguang; Wang, Mengmeng; Sun, Xin; Cong, Jing; Deng, Ye; Lu, Hui; Yuan, Tong; Van Nostrand, Joy D; Li, Diqiang; Zhou, Jizhong; Yang, Yunfeng

2015-10-01

As two major forest types in the subtropics, broadleaved evergreen and broadleaved deciduous forests have long interested ecologists. However, little is known about their belowground ecosystems despite their ecological importance in driving biogeochemical cycling. Here, we used Illumina MiSeq sequencing targeting 16S rRNA gene and a microarray named GeoChip targeting functional genes to analyse microbial communities in broadleaved evergreen and deciduous forest soils of Shennongjia Mountain of Central China, a region known as 'The Oriental Botanic Garden' for its extraordinarily rich biodiversity. We observed higher plant diversity and relatively richer nutrients in the broadleaved evergreen forest than the deciduous forest. In odds to our expectation that plant communities shaped soil microbial communities, we found that soil organic matter quantity and quality, but not plant community parameters, were the best predictors of microbial communities. Actinobacteria, a copiotrophic phylum, was more abundant in the broadleaved evergreen forest, while Verrucomicrobia, an oligotrophic phylum, was more abundant in the broadleaved deciduous forest. The density of the correlation network of microbial OTUs was higher in the broadleaved deciduous forest but its modularity was smaller, reflecting lower resistance to environment changes. In addition, keystone OTUs of the broadleaved deciduous forest were mainly oligotrophic. Microbial functional genes associated with recalcitrant carbon degradation were also more abundant in the broadleaved deciduous forests, resulting in low accumulation of organic matters. Collectively, these findings revealed the important role of soil organic matter in shaping microbial taxonomic and functional traits. © 2015 John Wiley & Sons Ltd.

Impact of ethnicity, geography, and disease on the microbiota in health and inflammatory bowel disease.

PubMed

Prideaux, Lani; Kang, Seungha; Wagner, Josef; Buckley, Michael; Mahar, Jackie E; De Cruz, Peter; Wen, Zhonghui; Chen, Liping; Xia, Bing; van Langenberg, Daniel R; Lockett, Trevor; Ng, Siew C; Sung, Joseph J Y; Desmond, Paul; McSweeney, Chris; Morrison, Mark; Kirkwood, Carl D; Kamm, Michael A

2013-12-01

The gut microbiota is central to health and disorders such as inflammatory bowel disease. Differences in microbiota related to geography and ethnicity may hold the key to recent changes in the incidence of microbiota-related disorders. Gut mucosal microbiota was analyzed in 190 samples from 87 Caucasian and Chinese subjects, from Australia and Hong Kong, comprising 22 patients with Crohn's disease, 30 patients with ulcerative colitis, 29 healthy controls, and 6 healthy relatives of patients with Crohn's disease. Bacterial 16S rRNA microarray and 454 pyrosequencing were performed. The microbiota was diverse in health, regardless of ethnicity or geography (operational taxonomic unit number and Shannon diversity index). Ethnicity and geography, however, did affect microbial composition. Crohn's disease resulted in reduced bacterial diversity, regardless of ethnicity or geography, and was the strongest determinant of composition. In ulcerative colitis, diversity was reduced in Chinese subjects only, suggesting that ethnicity is a determinant of bacterial diversity, whereas composition was determined by disease and ethnicity. Specific phylotypes were different between health and disease. Chinese patients with inflammatory bowel disease more often than healthy Chinese tended to have had a Western diet in childhood, in the East and West. The healthy microbiota is diverse but compositionally affected by geographical and ethnic factors. The microbiota is substantially altered in inflammatory bowel disease, but ethnicity may also play an important role. This may be key to the changing epidemiology in developing countries, and emigrants to the West.

Seasonal variation of plankton communities influenced by environmental factors in an artificial lake

NASA Astrophysics Data System (ADS)

Li, Xuemei; Yu, Yuhe; Zhang, Tanglin; Feng, Weisong; Ao, Hongyi; Yan, Qingyun

2012-05-01

We evaluated the seasonal variation in plankton community composition in an artificial lake. We conducted microscopic analysis and denaturing gradient gel electrophoresis (DGGE) of PCR-amplified partial 16S rRNA and 18S rRNA genes to characterize the plankton community. The clustering of unweighted pair group method with arithmetic mean (UPGMA) was then used to investigate the similarity of these plankton communities. DGGE fingerprinting revealed that samples collected at the different sites within a season shared high similarity and were generally grouped together. In contrast, we did not observe any seasonal variation based on microscopic analysis. Redundancy analysis (RDA) of the plankton operational taxonomic units (OTUs) in relation to environmental factors revealed that transparency was negatively correlated with the first axis ( R=-0.931), and temperature and total phosphorus (TP) were positively correlated with the first axis ( R=0.736 and R=0.660, respectively). In conclusion, plankton communities in the artificial lake exhibited significant seasonal variation. Transparency, phosphorus and temperature appear to be the major factors driving the differences in plankton composition.

Microbial community structure in a full-scale anaerobic treatment plant during start-up and first year of operation revealed by high-throughput 16S rRNA gene amplicon sequencing.

PubMed

Fykse, Else Marie; Aarskaug, Tone; Madslien, Elisabeth H; Dybwad, Marius

2016-12-01

High-throughput amplicon sequencing of six biomass samples from a full-scale anaerobic reactor at a Norwegian wood and pulp factory using Biothane Biobed Expanded Granular Sludge Bed (EGSB) technology during start-up and first year of operation was performed. A total of 106,166 16S rRNA gene sequences (V3-V5 region) were obtained. The number of operational taxonomic units (OTUs) ranged from 595 to 2472, and a total of 38 different phyla and 143 families were observed. The predominant phyla were Bacteroidetes, Chloroflexi, Firmicutes, Proteobacteria, and Spirochaetes. A more diverse microbial community was observed in the inoculum biomass coming from an Upflow Anaerobic Sludge Blanket (USAB) reactor, reflecting an adaptation of the inoculum diversity to the specific conditions of the new reactor. In addition, no taxa classified as obligate pathogens were identified and potentially opportunistic pathogens were absent or observed in low abundances. No Legionella bacteria were identified by traditional culture-based and molecular methods. Copyright © 2016 Elsevier Ltd. All rights reserved.

Complete mitochondrial genome of the Tyto longimembris (Strigiformes: Tytonidae).

PubMed

Xu, Peng; Li, Yankuo; Miao, Lujun; Xie, Guangyong; Huang, Yan

2016-07-01

The complete mitochondrial genome of Tyto longimembris has been determined in this study. It is 18,466 bp in length and consists of 13 protein-coding genes, 22 transfer RNA (tRNA) genes, 2 ribosomal RNA (rRNA) genes and a non-coding control region (D-loop). The overall base composition of the heavy strand of the T. longimembris mitochondrial genome is A: 30.1%, T: 23.5%, C: 31.8% and G: 14.6%. The structure of control region should be characterized by a region containing tandem repeats as two definitely separated clusters of tandem repeats were found. This study provided an important data set for phylogenetic and taxonomic analyses of Tyto species.

Preliminary study on 2 colour patterns in Ochlerotatus caspius (Pallas, 1771) (Diptera, Culicidae).

PubMed

Toma, Luciano; Severini, Francesco; Romi, Roberto; Di Luca, Marco

2016-08-30

Ochlerotatus caspius is a mosquito of medical and veterinary relevance both for its synanthropy and for its potential role in transmission of viruses and nematodes in the areas that it inhabits. Due to its wide range and the marked variability in the adult colour pattern, some authors have recognized Ochlerotatus caspius as a complex of species. In this study, we purposed to evaluate the possible taxonomic heterogeneity between 2 chromatic forms by using both morphological and molecular approaches. The preliminary results based on the identity of the rRNA internal transcribed spacer 2 (ITS-2) lead us to believe the 2 forms as a single species with a chromatic polymorphism.

Composition and Dynamics of Bacterial Communities of a Drinking Water Supply System as Assessed by RNA- and DNA-Based 16S rRNA Gene Fingerprinting

PubMed Central

Eichler, Stefan; Christen, Richard; Höltje, Claudia; Westphal, Petra; Bötel, Julia; Brettar, Ingrid; Mehling, Arndt; Höfle, Manfred G.

2006-01-01

Bacterial community dynamics of a whole drinking water supply system (DWSS) were studied from source to tap. Raw water for this DWSS is provided by two reservoirs with different water characteristics in the Harz mountains of Northern Germany. Samples were taken after different steps of treatment of raw water (i.e., flocculation, sand filtration, and chlorination) and at different points along the supply system to the tap. RNA and DNA were extracted from the sampled water. The 16S rRNA or its genes were partially amplified by reverse transcription-PCR or PCR and analyzed by single-strand conformation polymorphism community fingerprints. The bacterial community structures of the raw water samples from the two reservoirs were very different, but no major changes of these structures occurred after flocculation and sand filtration. Chlorination of the processed raw water strongly affected bacterial community structure, as reflected by the RNA-based fingerprints. This effect was less pronounced for the DNA-based fingerprints. After chlorination, the bacterial community remained rather constant from the storage containers to the tap. Furthermore, the community structure of the tap water did not change substantially for several months. Community composition was assessed by sequencing of abundant bands and phylogenetic analysis of the sequences obtained. The taxonomic compositions of the bacterial communities from both reservoirs were very different at the species level due to their different limnologies. On the other hand, major taxonomic groups, well known to occur in freshwater, such as Alphaproteobacteria, Betaproteobacteria, and Bacteroidetes, were found in both reservoirs. Significant differences in the detection of the major groups were observed between DNA-based and RNA-based fingerprints irrespective of the reservoir. Chlorination of the drinking water seemed to promote growth of nitrifying bacteria. Detailed analysis of the community dynamics of the whole DWSS revealed a significant influence of both source waters on the overall composition of the drinking water microflora and demonstrated the relevance of the raw water microflora for the drinking water microflora provided to the end user. PMID:16517632

Diversity Surveys and Evolutionary Relationships of aoxB Genes in Aerobic Arsenite-Oxidizing Bacteria▿ †

PubMed Central

Quéméneur, Marianne; Heinrich-Salmeron, Audrey; Muller, Daniel; Lièvremont, Didier; Jauzein, Michel; Bertin, Philippe N.; Garrido, Francis; Joulian, Catherine

2008-01-01

A new primer set was designed to specifically amplify ca. 1,100 bp of aoxB genes encoding the As(III) oxidase catalytic subunit from taxonomically diverse aerobic As(III)-oxidizing bacteria. Comparative analysis of AoxB protein sequences showed variable conservation levels and highlighted the conservation of essential amino acids and structural motifs. AoxB phylogeny of pure strains showed well-discriminated taxonomic groups and was similar to 16S rRNA phylogeny. Alphaproteobacteria-, Betaproteobacteria-, and Gammaproteobacteria-related sequences were retrieved from environmental surveys, demonstrating their prevalence in mesophilic As-contaminated soils. Our study underlines the usefulness of the aoxB gene as a functional marker of aerobic As(III) oxidizers. PMID:18502920

DNA extraction for streamlined metagenomics of diverse environmental samples.

PubMed

Marotz, Clarisse; Amir, Amnon; Humphrey, Greg; Gaffney, James; Gogul, Grant; Knight, Rob

2017-06-01

A major bottleneck for metagenomic sequencing is rapid and efficient DNA extraction. Here, we compare the extraction efficiencies of three magnetic bead-based platforms (KingFisher, epMotion, and Tecan) to a standardized column-based extraction platform across a variety of sample types, including feces, oral, skin, soil, and water. Replicate sample plates were extracted and prepared for 16S rRNA gene amplicon sequencing in parallel to assess extraction bias and DNA quality. The data demonstrate that any effect of extraction method on sequencing results was small compared with the variability across samples; however, the KingFisher platform produced the largest number of high-quality reads in the shortest amount of time. Based on these results, we have identified an extraction pipeline that dramatically reduces sample processing time without sacrificing bacterial taxonomic or abundance information.

Low Maternal Microbiota Sharing across Gut, Breast Milk and Vagina, as Revealed by 16S rRNA Gene and Reduced Metagenomic Sequencing.

PubMed

Avershina, Ekaterina; Angell, Inga Leena; Simpson, Melanie; Storrø, Ola; Øien, Torbjørn; Johnsen, Roar; Rudi, Knut

2018-05-01

The maternal microbiota plays an important role in infant gut colonization. In this work we have investigated which bacterial species are shared across the breast milk, vaginal and stool microbiotas of 109 women shortly before and after giving birth using 16S rRNA gene sequencing and a novel reduced metagenomic sequencing (RMS) approach in a subgroup of 16 women. All the species predicted by the 16S rRNA gene sequencing were also detected by RMS analysis and there was good correspondence between their relative abundances estimated by both approaches. Both approaches also demonstrate a low level of maternal microbiota sharing across the population and RMS analysis identified only two species common to most women and in all sample types ( Bifidobacterium longum and Enterococcus faecalis ). Breast milk was the only sample type that had significantly higher intra- than inter- individual similarity towards both vaginal and stool samples. We also searched our RMS dataset against an in silico generated reference database derived from bacterial isolates in the Human Microbiome Project. The use of this reference-based search enabled further separation of Bifidobacterium longum into Bifidobacterium longum ssp. longum and Bifidobacterium longum ssp. infantis . We also detected the Lactobacillus rhamnosus GG strain, which was used as a probiotic supplement by some women, demonstrating the potential of RMS approach for deeper taxonomic delineation and estimation.

Comparison of community structures of Candidatus Methylomirabilis oxyfera-like bacteria of NC10 phylum in different freshwater habitats.

PubMed

Shen, Li-Dong; Wu, Hong-Sheng; Gao, Zhi-Qiu; Liu, Xu; Li, Ji

2016-05-09

Methane oxidation coupled to nitrite reduction is mediated by 'Candidatus Methylomirabilis oxyfera' (M. oxyfera), which belongs to the NC10 phylum. In this study, the community composition and diversity of M. oxyfera-like bacteria of NC10 phylum were examined and compared in four different freshwater habitats, including reservoir sediments (RS), pond sediments (PS), wetland sediments (WS) and paddy soils (PAS), by using Illumina-based 16S rRNA gene sequencing. The recovered NC10-related sequences accounted for 0.4-2.5% of the 16S rRNA pool in the examined habitats, and the highest percentage was found in WS. The diversity of NC10 bacteria were the highest in RS, medium in WS, and lowest in PS and PAS. The observed number of OTUs (operational taxonomic unit; at 3% cut-off) were 97, 46, 61 and 40, respectively, in RS, PS, WS and PAS. A heterogeneous distribution of NC10 bacterial communities was observed in the examined habitats, though group B members were the dominant bacteria in each habitat. The copy numbers of NC10 bacterial 16S rRNA genes ranged between 5.8 × 10(6) and 3.2 × 10(7) copies g(-1) sediment/soil in the examined habitats. These results are helpful for a systematic understanding of NC10 bacterial communities in different types of freshwater habitats.

Low Maternal Microbiota Sharing across Gut, Breast Milk and Vagina, as Revealed by 16S rRNA Gene and Reduced Metagenomic Sequencing

PubMed Central

Angell, Inga Leena; Storrø, Ola; Øien, Torbjørn; Johnsen, Roar; Rudi, Knut

2018-01-01

The maternal microbiota plays an important role in infant gut colonization. In this work we have investigated which bacterial species are shared across the breast milk, vaginal and stool microbiotas of 109 women shortly before and after giving birth using 16S rRNA gene sequencing and a novel reduced metagenomic sequencing (RMS) approach in a subgroup of 16 women. All the species predicted by the 16S rRNA gene sequencing were also detected by RMS analysis and there was good correspondence between their relative abundances estimated by both approaches. Both approaches also demonstrate a low level of maternal microbiota sharing across the population and RMS analysis identified only two species common to most women and in all sample types (Bifidobacterium longum and Enterococcus faecalis). Breast milk was the only sample type that had significantly higher intra- than inter- individual similarity towards both vaginal and stool samples. We also searched our RMS dataset against an in silico generated reference database derived from bacterial isolates in the Human Microbiome Project. The use of this reference-based search enabled further separation of Bifidobacterium longum into Bifidobacterium longum ssp. longum and Bifidobacterium longum ssp. infantis. We also detected the Lactobacillus rhamnosus GG strain, which was used as a probiotic supplement by some women, demonstrating the potential of RMS approach for deeper taxonomic delineation and estimation. PMID:29724017

Diversity of Anaerobic Microbes in Spacecraft Assembly Clean Rooms ▿ †

PubMed Central

Probst, Alexander; Vaishampayan, Parag; Osman, Shariff; Moissl-Eichinger, Christine; Andersen, Gary L.; Venkateswaran, Kasthuri

2010-01-01

Although the cultivable and noncultivable microbial diversity of spacecraft assembly clean rooms has been previously documented using conventional and state-of-the-art molecular techniques, the occurrence of obligate anaerobes within these clean rooms is still uncertain. Therefore, anaerobic bacterial communities of three clean-room facilities were analyzed during assembly of the Mars Science Laboratory rover. Anaerobic bacteria were cultured on several media, and DNA was extracted from suitable anaerobic enrichments and examined with conventional 16S rRNA gene clone library, as well as high-density phylogenetic 16S rRNA gene microarray (PhyloChip) technologies. The culture-dependent analyses predominantly showed the presence of clostridial and propionibacterial strains. The 16S rRNA gene sequences retrieved from clone libraries revealed distinct microbial populations associated with each clean-room facility, clustered exclusively within gram-positive organisms. PhyloChip analysis detected a greater microbial diversity, spanning many phyla of bacteria, and provided a deeper insight into the microbial community structure of the clean-room facilities. This study presents an integrated approach for assessing the anaerobic microbial population within clean-room facilities, using both molecular and cultivation-based analyses. The results reveal that highly diverse anaerobic bacterial populations persist in the clean rooms even after the imposition of rigorous maintenance programs and will pose a challenge to planetary protection implementation activities. PMID:20228115

PhylArray: phylogenetic probe design algorithm for microarray.

PubMed

Militon, Cécile; Rimour, Sébastien; Missaoui, Mohieddine; Biderre, Corinne; Barra, Vincent; Hill, David; Moné, Anne; Gagne, Geneviève; Meier, Harald; Peyretaillade, Eric; Peyret, Pierre

2007-10-01

Microbial diversity is still largely unknown in most environments, such as soils. In order to get access to this microbial 'black-box', the development of powerful tools such as microarrays are necessary. However, the reliability of this approach relies on probe efficiency, in particular sensitivity, specificity and explorative power, in order to obtain an image of the microbial communities that is close to reality. We propose a new probe design algorithm that is able to select microarray probes targeting SSU rRNA at any phylogenetic level. This original approach, implemented in a program called 'PhylArray', designs a combination of degenerate and non-degenerate probes for each target taxon. Comparative experimental evaluations indicate that probes designed with PhylArray yield a higher sensitivity and specificity than those designed by conventional approaches. Applying the combined PhyArray/GoArrays strategy helps to optimize the hybridization performance of short probes. Finally, hybridizations with environmental targets have shown that the use of the PhylArray strategy can draw attention to even previously unknown bacteria.

Development of microbial genome-probing microarrays using digital multiple displacement amplification of uncultivated microbial single cells.

PubMed

Chang, Ho-Won; Sung, Youlboong; Kim, Kyoung-Ho; Nam, Young-Do; Roh, Seong Woon; Kim, Min-Soo; Jeon, Che Ok; Bae, Jin-Woo

2008-08-15

A crucial problem in the use of previously developed genome-probing microarrays (GPM) has been the inability to use uncultivated bacterial genomes to take advantage of the high sensitivity and specificity of GPM in microbial detection and monitoring. We show here a method, digital multiple displacement amplification (MDA), to amplify and analyze various genomes obtained from single uncultivated bacterial cells. We used 15 genomes from key microbes involved in dichloromethane (DCM)-dechlorinating enrichment as microarray probes to uncover the bacterial population dynamics of samples without PCR amplification. Genomic DNA amplified from single cells originating from uncultured bacteria with 80.3-99.4% similarity to 16S rRNA genes of cultivated bacteria. The digital MDA-GPM method successfully monitored the dynamics of DCM-dechlorinating communities from different phases of enrichment status. Without a priori knowledge of microbial diversity, the digital MDA-GPM method could be designed to monitor most microbial populations in a given environmental sample.

Aestuariispira insulae gen. nov., sp. nov., a lipolytic bacterium isolated from a tidal flat.

PubMed

Park, Sooyeon; Park, Ji-Min; Kang, Chul-Hyung; Yoon, Jung-Hoon

2014-06-01

A Gram-stain-negative, non-motile, aerobic, curved-to-spiral-rod-shaped bacterium, designated AH-MY2(T), was isolated from a tidal flat on Aphae island in the sea to the south-west of South Korea, and its taxonomic position was investigated using a polyphasic taxonomic approach. Strain AH-MY2(T) grew optimally at 30 °C, at pH 7.0-8.0 and in the presence of 2.0% (w/v) NaCl. Neighbour-joining, maximum-likelihood and maximum-parsimony phylogenetic trees based on 16S rRNA gene sequences showed that strain AH-MY2(T) clustered with the type strain of Terasakiella pusilla and that this cluster joined the clade comprising the type strains of species of the genus Thalassospira. Strain AH-MY2(T) exhibited 16S rRNA gene sequence similarity values of 90.6% to the type strain of Terasakiella pusilla and of less than 91.0% to the type strains of other species with validly published names. Strain AH-MY2(T) contained Q-10 as the predominant ubiquinone and C(18 : 1)ω7c as the major fatty acid. The major polar lipids detected in strain AH-MY2(T) were phosphatidylglycerol, phosphatidylethanolamine, two unidentified aminolipids and one unidentified glycolipid. The DNA G+C content of strain AH-MY2(T) was 56.0 mol%. The phylogenetic data and differential chemotaxonomic and other phenotypic properties revealed that strain AH-MY2(T) represented a novel genus and species within the family Rhodospirillaceae of the class Alphaproteobacteria, for which the name Aestuariispira insulae gen. nov., sp. nov. is proposed. The type strain of Aestuariispira insulae is AH-MY2(T) ( = KCTC 32577(T) = CECT 8488(T)). © 2014 IUMS.

Streptomyces atlanticus sp. nov., a novel actinomycete isolated from marine sponge Aplysina fulva (Pallas, 1766).

PubMed

Silva, Fábio Sérgio Paulino; Souza, Danilo Tosta; Zucchi, Tiago Domingues; Pansa, Camila Cristiane; de Figueiredo Vasconcellos, Rafael Leandro; Crevelin, Eduardo José; de Moraes, Luiz Alberto Beraldo; Melo, Itamar Soares

2016-11-01

The taxonomic position of a novel marine actinomycete isolated from a marine sponge, Aplysina fulva, which had been collected in the Archipelago of Saint Peter and Saint Paul (Equatorial Atlantic Ocean), was determined by using a polyphasic approach. The organism showed a combination of morphological and chemotaxonomic characteristics consistent with its classification in the genus Streptomyces and forms a distinct branch within the Streptomyces somaliensis 16S rRNA gene tree subclade. It is closely related to Streptomyces violascens ISP 5183 T (97.27 % 16S rRNA gene sequence similarity) and Streptomyces hydrogenans NBRC 13475 T (97.15 % 16S rRNA gene sequence similarity). The 16S rRNA gene similarities between the isolate and the remaining members of the subclade are lower than 96.77 %. The organism can be distinguished readily from other members of the S. violacens subclade using a combination of phenotypic properties. On the basis of these results, it is proposed that isolate 103 T (=NRRL B-65309 T  = CMAA 1378 T ) merits recognition as the type strain of a new Streptomyces species, namely Streptomyces atlanticus sp. nov.

Fine-scale analysis of 16S rRNA sequences reveals a high level of taxonomic diversity among vaginal Atopobium spp.

PubMed Central

Mendes-Soares, Helena; Krishnan, Vandhana; Settles, Matthew L.; Ravel, Jacques; Brown, Celeste J.; Forney, Larry J.

2015-01-01

Although vaginal microbial communities of some healthy women have high proportions of Atopobium vaginae, the genus Atopobium is more commonly associated with bacterial vaginosis, a syndrome associated with an increased risk of adverse pregnancy outcomes and the transmission of sexually transmitted diseases. Genetic differences within Atopobium species may explain why single species can be associated with both health and disease. We used 16S rRNA gene sequences from previously published studies to explore the taxonomic diversity of the genus Atopobium in vaginal microbial communities of healthy women. Although A. vaginae was the species most commonly found, we also observed three other Atopobium species in the vaginal microbiota, one of which, A. parvulum, was not previously known to reside in the human vagina. Furthermore, we found several potential novel species of the genus Atopobium and multiple phylogenetic clades of A. vaginae. The diversity of Atopobium found in our study, which focused only on samples from healthy women, is greater than previously recognized, suggesting that analysis of samples from women with BV would yield even more diversity. Classification of microbes only to the genus level may thus obfuscate differences that might be important to better understand health or disease. PMID:25778779

Lactobacillus gorillae sp. nov., isolated from the faeces of captive and wild western lowland gorillas (Gorilla gorilla gorilla).

PubMed

Tsuchida, Sayaka; Kitahara, Maki; Nguema, Pierre Philippe Mbehang; Norimitsu, Saeko; Fujita, Shiho; Yamagiwa, Juichi; Ngomanda, Alfred; Ohkuma, Moriya; Ushida, Kazunari

2014-12-01

Four strains of Gram-staining-positive, anaerobic rods were isolated from the faeces of western lowland gorillas (Gorilla gorilla gorilla). Three strains, KZ01(T), KZ02 and KZ03, were isolated at the Kyoto City Zoo, Japan, and one strain, GG02, was isolated in the Moukalaba-Doudou National Park, Gabon. These strains were investigated taxonomically. These strains belonged to the Lactobacillus reuteri phylogenetic group according to phylogenetic analysis based on 16S rRNA gene sequences and specific phenotypic characteristics. Phylogenetic analysis of their 16S rRNA gene sequences revealed that strains KZ01(T), KZ02, KZ03 and GG02 formed a single monophyletic cluster and had a distinct line of descent. Based on sequence similarity of the 16S rRNA gene, Lactobacillus fermentum JCM 1173(T) (96.6 %) was the closest neighbour to these novel strains, although it was clear that these strains belonged to a different species. Partial pheS sequences also supported these relationships. DNA-DNA relatedness between strain KZ01(T) and L. fermentum JCM 1173(T) was less than 22 % and the DNA G+C content of strain KZ01(T) was 50.7 mol%. The cell-wall peptidoglycan type was A4β (l-Orn-d-Asp) and the major fatty acids were C16 : 0, C18 : 1ω9c and C19 : 1 cyclo 9,10. Therefore, based on phylogenetic, phenotypic and physiological evidence, these strains represent a novel species of the genus Lactobacillus, for which the name Lactobacillus gorillae sp. nov. is proposed. The type strain is KZ01(T) ( = JCM 19575(T) = DSM 28356(T)). © 2014 IUMS.

Identification of a microsporidian isolate from Cnaphalocrocis Medinalis and its pathogenicity to Bombyx mori.

PubMed

Huang, Xuhua; Qi, Guangjun; Pan, Zhixin; Zhu, Fangrong; Huang, Yuanjiao; Wu, Yonghu

2014-11-01

A microsporidian, CmM2, was isolated from Cnaphalocrocis medinalis. The biological characters, molecular analysis and pathogenicity of CmM2 were studied. The spore of CmM2 is long oval in shape and 3.45 ± 0.25 × 1.68 ± 0.18 µm in size, the life cycle includes meronts, sporonts, sporoblasts, and spores, with typical diplokaryon in each stage, propagated in binary fission. There is positive coagulation reaction between CmM2 and the polyclonal antibody of Nosema bombycis (N.b.). CmM2 spores is binuclear, and has 10-12 polar filament coils. The small subunit ribosomal RNA (SSU rRNA) gene sequence of CmM2 was obtained by PCR amplification and sequencing, the phylogenetic tree based on SSU rRNA sequences had been constructed, and the similarity and genetic distance of SSU rRNA sequences were analyzed, showed that CmM2 was grouped in the Nosema clade. The 50% infectious concentration of CmM2 to Bombyx mori is 4.72 × 10(4)  spores ml(-1) , and the germinative infection rate is 12.33%. The results showed that CmM2 is classified into genus Nosema, as Nosema sp. CmM2, and has a heavy infectivity to B. mori. The result indicated as well that it is valuable taxonomic determination for microsporidian isolates based on both biological characters and molecular evidence. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Optimisation of 16S rRNA gut microbiota profiling of extremely low birth weight infants.

PubMed

Alcon-Giner, Cristina; Caim, Shabhonam; Mitra, Suparna; Ketskemety, Jennifer; Wegmann, Udo; Wain, John; Belteki, Gusztav; Clarke, Paul; Hall, Lindsay J

2017-11-02

Infants born prematurely, particularly extremely low birth weight infants (ELBW) have altered gut microbial communities. Factors such as maternal health, gut immaturity, delivery mode, and antibiotic treatments are associated with microbiota disturbances, and are linked to an increased risk of certain diseases such as necrotising enterocolitis. Therefore, there is a requirement to optimally characterise microbial profiles in this at-risk cohort, via standardisation of methods, particularly for studying the influence of microbiota therapies (e.g. probiotic supplementation) on community profiles and health outcomes. Profiling of faecal samples using the 16S rRNA gene is a cost-efficient method for large-scale clinical studies to gain insights into the gut microbiota and additionally allows characterisation of cohorts were sample quantities are compromised (e.g. ELBW infants). However, DNA extraction method, and the 16S rRNA region targeted can significantly change bacterial community profiles obtained, and so confound comparisons between studies. Thus, we sought to optimise a 16S rRNA profiling protocol to allow standardisation for studying ELBW infant faecal samples, with or without probiotic supplementation. Using ELBW faecal samples, we compared three different DNA extraction methods, and subsequently PCR amplified and sequenced three hypervariable regions of the 16S rRNA gene (V1 + V2 + V3), (V4 + V5) and (V6 + V7 + V8), and compared two bioinformatics approaches to analyse results (OTU and paired end). Paired shotgun metagenomics was used as a 'gold-standard'. Results indicated a longer bead-beating step was required for optimal bacterial DNA extraction and that sequencing regions (V1 + V2 + V3) and (V6 + V7 + V8) provided the most representative taxonomic profiles, which was confirmed via shotgun analysis. Samples sequenced using the (V4 + V5) region were found to be underrepresented in specific taxa including Bifidobacterium, and had altered diversity profiles. Both bioinformatics 16S rRNA pipelines used in this study (OTU and paired end) presented similar taxonomic profiles at genus level. We determined that DNA extraction from ELBW faecal samples, particularly those infants receiving probiotic supplementation, should include a prolonged beat-beating step. Furthermore, use of the 16S rRNA (V1 + V2 + V3) and (V6 + V7 + V8) regions provides reliable representation of ELBW microbiota profiles, while inclusion of the (V4 + V5) region may not be appropriate for studies where Bifidobacterium constitutes a resident microbiota member.

Novel approaches for the taxonomic and metabolic characterization of lactobacilli: Integration of 16S rRNA gene sequencing with MALDI-TOF MS and 1H-NMR

PubMed Central

Parolin, Carola; Giordani, Barbara; Compri, Monica; Cevenini, Roberto; Vitali, Beatrice

2017-01-01

Lactobacilli represent a wide range of bacterial species with several implications for the human host. They play a crucial role in maintaining the ecological equilibrium of different biological niches and are essential for fermented food production and probiotic formulation. Despite the consensus about the ‘health-promoting’ significance of Lactobacillus genus, its genotypic and phenotypic characterization still poses several difficulties. The aim of this study was to assess the integration of different approaches, genotypic (16S rRNA gene sequencing), proteomic (MALDI-TOF MS) and metabolomic (1H-NMR), for the taxonomic and metabolic characterization of Lactobacillus species. For this purpose we analyzed 40 strains of various origin (intestinal, vaginal, food, probiotics), belonging to different species. The high discriminatory power of MALDI-TOF for species identification was underlined by the excellent agreement with the genotypic analysis. Indeed, MALDI-TOF allowed to correctly identify 39 out of 40 Lactobacillus strains at the species level, with an overall concordance of 97.5%. In the perspective to simplify the MALDI TOF sample preparation, especially for routine practice, we demonstrated the perfect agreement of the colony-picking from agar plates with the protein extraction protocol. 1H-NMR analysis, applied to both culture supernatants and bacterial lysates, identified a panel of metabolites whose variations in concentration were associated with the taxonomy, but also revealed a high intra-species variability that did not allow a species-level identification. Therefore, despite not suitable for mere taxonomic purposes, metabolomics can be useful to correlate particular biological activities with taxonomy and to understand the mechanisms related to the antimicrobial effect shown by some Lactobacillus species. PMID:28207855

Automated detection and quantitation of bacterial RNA by using electrical microarrays.

PubMed

Elsholz, B; Wörl, R; Blohm, L; Albers, J; Feucht, H; Grunwald, T; Jürgen, B; Schweder, T; Hintsche, Rainer

2006-07-15

Low-density electrical 16S rRNA specific oligonucleotide microarrays and an automated analysis system have been developed for the identification and quantitation of pathogens. The pathogens are Escherichia coli, Pseudomonas aeruginosa, Enterococcus faecalis, Staphylococcus aureus, and Staphylococcus epidermidis, which are typically involved in urinary tract infections. Interdigitated gold array electrodes (IDA-electrodes), which have structures in the nanometer range, have been used for very sensitive analysis. Thiol-modified oligonucleotides are immobilized on the gold IDA as capture probes. They mediate the specific recognition of the target 16S rRNA by hybridization. Additionally three unlabeled oligonucleotides are hybridized in close proximity to the capturing site. They are supporting molecules, because they improve the RNA hybridization at the capturing site. A biotin labeled detector oligonucleotide is also allowed to hybridize to the captured RNA sequence. The biotin labels enable the binding of avidin alkaline phophatase conjugates. The phosphatase liberates the electrochemical mediator p-aminophenol from its electrically inactive phosphate derivative. The electrical signals were generated by amperometric redox cycling and detected by a unique multipotentiostat. The read out signals of the microarray are position specific current and change over time in proportion to the analyte concentration. If two additional biotins are introduced into the affinity binding complex via the supporting oligonucleotides, the sensitivity of the assays increase more than 60%. The limit of detection of Escherichia coli total RNA has been determined to be 0.5 ng/microL. The control of fluidics for variable assay formats as well as the multichannel electrical read out and data handling have all been fully automated. The fast and easy procedure does not require any amplification of the targeted nucleic acids by PCR.

Evaluation of composition and individual variability of rumen microbiota in yaks by 16S rRNA high-throughput sequencing technology.

PubMed

Guo, Wei; Li, Ying; Wang, Lizhi; Wang, Jiwen; Xu, Qin; Yan, Tianhai; Xue, Bai

2015-08-01

The Yak (Bos grunniens) is a unique species of ruminant animals that is important to agriculture of the Tibetan plateau, and has a complex intestinal microbial community. The objective of the present study was to characterize the composition and individual variability of microbiota in the rumen of yaks using 16S rRNA gene high-throughput sequencing technique. Rumen samples used in the present study were obtained from grazing adult male yaks (n = 6) in a commercial farm in Ganzi Autonomous Prefecture of Sichuan Province, China. Universal prokaryote primers were used to target the V4-V5 hypervariable region of 16S rRNA gene. A total of 7200 operational taxonomic units (OTUs) were obtained after sequence filtering and chimera removal. Within these OTUs, 0.56% belonged to Archaea (40 OTUs), 7.19% to unassigned species (518 OTUs), and the remaining OTUs (6642) in all samples were of bacterial origin. When examining the community structure of bacteria, we identified 23 phyla within 159 families after taxonomic summarization. Bacteroidetes and Firmicutes were the predominant phyla accounting for 39.68% (SD = 0.05) and 45.90% (SD = 0.06), respectively. Moreover, 3764 OTUs were identified as shared OTUs (i.e. represented in all yaks) and belonged to 35 genera, exhibiting highly variable abundance across individual samples. Phylogenetic placement of these genera across individual samples was examined. In addition, we evaluated the distance among the 6 rumen samples by adding taxon phylogeny using UniFrac, representing 24.1% of average distance. In summary, the current study reveals a shared rumen microbiome and phylogenetic lineage and presents novel information on composition and individual variability of the bacterial community in the rumen of yaks. Copyright © 2015. Published by Elsevier Ltd.

Molecular diversity of the rumen microbiome of Norwegian reindeer on natural summer pasture.

PubMed

Sundset, Monica A; Edwards, Joan E; Cheng, Yan Fen; Senosiain, Roberto S; Fraile, Maria N; Northwood, Korinne S; Praesteng, Kirsti E; Glad, Trine; Mathiesen, Svein D; Wright, André-Denis G

2009-02-01

The molecular diversity of the rumen microbiome was investigated in five semi-domesticated adult female Norwegian reindeer (Rangifer tarandus tarandus) grazing on natural summer pastures on the coast of northern Norway (71.00 degrees N, 25.30 degrees E). Mean population densities (numbers per gram wet weight) of methanogenic archaea, rumen bacteria and ciliate protozoa, estimated using quantitative real-time polymerase chain reaction (PCR), were 3.17x10(9), 5.17x10(11) and 4.02x10(7), respectively. Molecular diversity of rumen methanogens was revealed using a 16S rRNA gene library (54 clones) constructed using pooled PCR products from the whole rumen contents of the five individual reindeer. Based upon a similarity criterion of <97%, a total of 19 distinct operational taxonomic units (OTUs) were identified, nine of which are potential new species. The 16S rRNA sequences generated from the reindeer rumen exhibited a high degree of sequence similarity to methanogens affiliated with the families Methanobacteriaceae (14 OTUs) and Methanosarcinaceae (one OTU). Four of the OTUs detected belonged to a group of uncultivated archaea previously found in domestic ruminants and thought to be dominant in the rumen together with Methanobrevibacter spp. Denaturing gradient gel electrophoresis profiling of the rumen bacterial 16S rRNA gene and the protozoal 18S rRNA gene indicated a high degree of animal variation, although some bands were common to all individuals. Automated ribosomal intergenic spacer analysis (ARISA) profiling of the ruminal Neocallimastigales population indicated that the reindeer are likely to contain more than one type of anaerobic fungus. The ARISA profile from one animal was distinct from the other four. This is the first molecular investigation of the ruminal methanogenic archaea in reindeer, revealing higher numbers than expected based on methane emission data available. Also, many of the reindeer archaeal 16S rRNA gene sequences were similar to those reported in domesticated ruminants in Australia, Canada, China, New Zealand and Venezuela, supporting previous findings that there seems to be no host type or geographical effect on the methanogenic archaea community structure in ruminants.

Parallel characterization of anaerobic toluene- and ethylbenzene-degrading microbial consortia by PCR-denaturing gradient gel electrophoresis, RNA-DNA membrane hybridization, and DNA microarray technology

NASA Technical Reports Server (NTRS)

Koizumi, Yoshikazu; Kelly, John J.; Nakagawa, Tatsunori; Urakawa, Hidetoshi; El-Fantroussi, Said; Al-Muzaini, Saleh; Fukui, Manabu; Urushigawa, Yoshikuni; Stahl, David A.

2002-01-01

A mesophilic toluene-degrading consortium (TDC) and an ethylbenzene-degrading consortium (EDC) were established under sulfate-reducing conditions. These consortia were first characterized by denaturing gradient gel electrophoresis (DGGE) fingerprinting of PCR-amplified 16S rRNA gene fragments, followed by sequencing. The sequences of the major bands (T-1 and E-2) belonging to TDC and EDC, respectively, were affiliated with the family Desulfobacteriaceae. Another major band from EDC (E-1) was related to an uncultured non-sulfate-reducing soil bacterium. Oligonucleotide probes specific for the 16S rRNAs of target organisms corresponding to T-1, E-1, and E-2 were designed, and hybridization conditions were optimized for two analytical formats, membrane and DNA microarray hybridization. Both formats were used to characterize the TDC and EDC, and the results of both were consistent with DGGE analysis. In order to assess the utility of the microarray format for analysis of environmental samples, oil-contaminated sediments from the coast of Kuwait were analyzed. The DNA microarray successfully detected bacterial nucleic acids from these samples, but probes targeting specific groups of sulfate-reducing bacteria did not give positive signals. The results of this study demonstrate the limitations and the potential utility of DNA microarrays for microbial community analysis.

Parallel Characterization of Anaerobic Toluene- and Ethylbenzene-Degrading Microbial Consortia by PCR-Denaturing Gradient Gel Electrophoresis, RNA-DNA Membrane Hybridization, and DNA Microarray Technology

PubMed Central

Koizumi, Yoshikazu; Kelly, John J.; Nakagawa, Tatsunori; Urakawa, Hidetoshi; El-Fantroussi, Saïd; Al-Muzaini, Saleh; Fukui, Manabu; Urushigawa, Yoshikuni; Stahl, David A.

2002-01-01

A mesophilic toluene-degrading consortium (TDC) and an ethylbenzene-degrading consortium (EDC) were established under sulfate-reducing conditions. These consortia were first characterized by denaturing gradient gel electrophoresis (DGGE) fingerprinting of PCR-amplified 16S rRNA gene fragments, followed by sequencing. The sequences of the major bands (T-1 and E-2) belonging to TDC and EDC, respectively, were affiliated with the family Desulfobacteriaceae. Another major band from EDC (E-1) was related to an uncultured non-sulfate-reducing soil bacterium. Oligonucleotide probes specific for the 16S rRNAs of target organisms corresponding to T-1, E-1, and E-2 were designed, and hybridization conditions were optimized for two analytical formats, membrane and DNA microarray hybridization. Both formats were used to characterize the TDC and EDC, and the results of both were consistent with DGGE analysis. In order to assess the utility of the microarray format for analysis of environmental samples, oil-contaminated sediments from the coast of Kuwait were analyzed. The DNA microarray successfully detected bacterial nucleic acids from these samples, but probes targeting specific groups of sulfate-reducing bacteria did not give positive signals. The results of this study demonstrate the limitations and the potential utility of DNA microarrays for microbial community analysis. PMID:12088997

Distinct classical and molecular cytogenetics of Astyanax marionae and A. fasciatus (Characiformes: Characidae): a comparative study of the organization of heterochromatin and repetitive genes.

PubMed

Piscor, Diovani; Centofante, Liano; Parise-Maltempi, Patricia Pasquali

2017-09-01

Genus Astyanax is well distributed in Neotropical freshwater environments and its taxonomic position is uncertain, as is the case with other Characidae genera allocated in the group incertae sedis. This study aimed to analyse the karyotype of different populations of Astyanax fasciatus (Corumbataí River basin) using Giemsa staining, C-band technique, and fluorescence in situ hybridization for the H3 histone and 5S rRNA genes, in addition we describe for the first time the chromosomal organization of H3 histone and 5S rRNAgenes in A. marionae (ParaguayRiver basin). Chromosomes of three A. fasciatus populations were analysed (two with 2n = 50 and one with 2n = 48) and the heterochromatin was organized in two forms (blocks with blurred boundaries and distinct blocks). H3 histone and 5S rRNA genes were observed in all the three populations of A. fasciatus on two chromosome pairs (one metacentric chromosome showing H3 histone and 5S rRNA gene clusters). In A. marionae (2n = 48), H3 histone and 5S rRNA genes were observed in one acrocentric chromosome pair (different pairs). Further, differences between karyotypes and heterochromatin, as well as the chromosomal organization of H3 histone and 5S rRNA genes in Astyanax species, focussing on chromosome evolution in the group are discussed.

A New Oligonucleotide Microarray for Detection of Pathogenic and Non-Pathogenic Legionella spp.

PubMed Central

Cao, Boyang; Liu, Xiangqian; Yu, Xiang; Chen, Min; Feng, Lu; Wang, Lei

2014-01-01

Legionella pneumophila has been recognized as the major cause of legionellosis since the discovery of the deadly disease. Legionella spp. other than L. pneumophila were later found to be responsible to many non-pneumophila infections. The non-L. pneumophila infections are likely under-detected because of a lack of effective diagnosis. In this report, we have sequenced the 16S-23S rRNA gene internal transcribed spacer (ITS) of 10 Legionella species and subspecies, including L. anisa, L. bozemanii, L. dumoffii, L. fairfieldensis, L. gormanii, L. jordanis, L. maceachernii, L. micdadei, L. pneumophila subspp. fraseri and L. pneumophila subspp. pasculleii, and developed a rapid oligonucleotide microarray detection technique accordingly to identify 12 most common Legionella spp., which consist of 11 pathogenic species of L. anisa, L. bozemanii, L. dumoffii, L. gormanii, L. jordanis, L. longbeachae, L. maceachernii, L. micdadei, and L. pneumophila (including subspp. pneumophila, subspp. fraseri, and subspp. pasculleii) and one non-pathogenic species, L. fairfieldensis. Twenty-nine probes that reproducibly detected multiple Legionella species with high specificity were included in the array. A total of 52 strains, including 30 target pathogens and 22 non-target bacteria, were used to verify the oligonucleotide microarray assay. The sensitivity of the detection was at 1.0 ng with genomic DNA or 13 CFU/100 mL with Legionella cultures. The microarray detected seven samples of air conditioner-condensed water with 100% accuracy, validating the technique as a promising method for applications in basic microbiology, clinical diagnosis, food safety, and epidemiological surveillance. The phylogenetic study based on the ITS has also revealed that the non-pathogenic L. fairfieldensis is the closest to L. pneumophila than the nine other pathogenic Legionella spp. PMID:25469776

A new oligonucleotide microarray for detection of pathogenic and non-pathogenic Legionella spp.

PubMed

Cao, Boyang; Liu, Xiangqian; Yu, Xiang; Chen, Min; Feng, Lu; Wang, Lei

2014-01-01

Legionella pneumophila has been recognized as the major cause of legionellosis since the discovery of the deadly disease. Legionella spp. other than L. pneumophila were later found to be responsible to many non-pneumophila infections. The non-L. pneumophila infections are likely under-detected because of a lack of effective diagnosis. In this report, we have sequenced the 16S-23S rRNA gene internal transcribed spacer (ITS) of 10 Legionella species and subspecies, including L. anisa, L. bozemanii, L. dumoffii, L. fairfieldensis, L. gormanii, L. jordanis, L. maceachernii, L. micdadei, L. pneumophila subspp. fraseri and L. pneumophila subspp. pasculleii, and developed a rapid oligonucleotide microarray detection technique accordingly to identify 12 most common Legionella spp., which consist of 11 pathogenic species of L. anisa, L. bozemanii, L. dumoffii, L. gormanii, L. jordanis, L. longbeachae, L. maceachernii, L. micdadei, and L. pneumophila (including subspp. pneumophila, subspp. fraseri, and subspp. pasculleii) and one non-pathogenic species, L. fairfieldensis. Twenty-nine probes that reproducibly detected multiple Legionella species with high specificity were included in the array. A total of 52 strains, including 30 target pathogens and 22 non-target bacteria, were used to verify the oligonucleotide microarray assay. The sensitivity of the detection was at 1.0 ng with genomic DNA or 13 CFU/100 mL with Legionella cultures. The microarray detected seven samples of air conditioner-condensed water with 100% accuracy, validating the technique as a promising method for applications in basic microbiology, clinical diagnosis, food safety, and epidemiological surveillance. The phylogenetic study based on the ITS has also revealed that the non-pathogenic L. fairfieldensis is the closest to L. pneumophila than the nine other pathogenic Legionella spp.

First Record of Raillietina celebensis (Cestoda: Cyclophyllidea) in South America: Redescription and Phylogeny.

PubMed

de Oliveira Simões, Raquel; Simões, Susana Balmant Enrique; Luque, José Luis; Iñiguez, Alena Mayo; Júnior, Arnaldo Maldonado

2017-08-01

Raillietina celebensis is a cestode that parasitizes the small intestine of rats and humans. Here, we detail the morphology and morphometry of R. celebensis based on specimens collected from Rattus norvegicus in the municipality of São Gonçalo, state of Rio de Janeiro, Brazil, by light and confocal scanning laser microscopies and also report the results of molecular phylogenetic analyses to determine its relationships within the family Davaineidae. Analysis of the number and size of testes, number and shape of rostellar hooks, cirrus sac length, capsules and eggs per capsule, and morphology of the mature proglottid allowed concluding that the present specimens constitute a new record of R. celebensis in South America. Our genetic and phylogenetic analyses, based on the partial small subunit 18S rRNA gene, revealed R. celebensis to be in the family Davaineidae within the genus Raillietina, in agreement with the morphological taxonomy. Phylogenetic trees obtained by neighbor-joining and maximum likelihood methods demonstrated R. celebensis as a unique taxonomic unit, and also demonstrated some taxonomic inconsistences. The incorporation of Brazilian R. celebensis sequences derived from mammals in the phylogeny of davaineids is consistent with the assertion that neither Raillietina nor Fuhrmannetta can be supported as distinct genera.

Drylands soil bacterial community is affected by land use change and different irrigation practices in the Mezquital Valley, Mexico.

PubMed

Lüneberg, Kathia; Schneider, Dominik; Siebe, Christina; Daniel, Rolf

2018-01-23

Dryland agriculture nourishes one third of global population, although crop irrigation is often mandatory. As freshwater sources are scarce, treated and untreated wastewater is increasingly used for irrigation. Here, we investigated how the transformation of semiarid shrubland into rainfed farming or irrigated agriculture with freshwater, dam-stored or untreated wastewater affects the total (DNA-based) and active (RNA-based) soil bacterial community composition, diversity, and functionality. To do this we collected soil samples during the dry and rainy seasons and isolated DNA and RNA. Soil moisture, sodium content and pH were the strongest drivers of the bacterial community composition. We found lineage-specific adaptations to drought and sodium content in specific land use systems. Predicted functionality profiles revealed gene abundances involved in nitrogen, carbon and phosphorous cycles differed among land use systems and season. Freshwater irrigated bacterial community is taxonomically and functionally susceptible to seasonal environmental changes, while wastewater irrigated ones are taxonomically susceptible but functionally resistant to them. Additionally, we identified potentially harmful human and phytopathogens. The analyses of 16 S rRNA genes, its transcripts and deduced functional profiles provided extensive understanding of the short-term and long-term responses of bacterial communities associated to land use, seasonality, and water quality used for irrigation in drylands.

Suitability and setup of next-generation sequencing-based method for taxonomic characterization of aquatic microbial biofilm.

PubMed

Bakal, Tomas; Janata, Jiri; Sabova, Lenka; Grabic, Roman; Zlabek, Vladimir; Najmanova, Lucie

2018-06-16

A robust and widely applicable method for sampling of aquatic microbial biofilm and further sample processing is presented. The method is based on next-generation sequencing of V4-V5 variable regions of 16S rRNA gene and further statistical analysis of sequencing data, which could be useful not only to investigate taxonomic composition of biofilm bacterial consortia but also to assess aquatic ecosystem health. Five artificial materials commonly used for biofilm growth (glass, stainless steel, aluminum, polypropylene, polyethylene) were tested to determine the one giving most robust and reproducible results. The effect of used sampler material on total microbial composition was not statistically significant; however, the non-plastic materials (glass, metal) gave more stable outputs without irregularities among sample parallels. The bias of the method is assessed with respect to the employment of a non-quantitative step (PCR amplification) to obtain quantitative results (relative abundance of identified taxa). This aspect is often overlooked in ecological and medical studies. We document that sequencing of a mixture of three merged primary PCR reactions for each sample and further evaluation of median values from three technical replicates for each sample enables to overcome this bias and gives robust and repeatable results well distinguishing among sampling localities and seasons.

Micromonospora halotolerans sp. nov., isolated from the rhizosphere of a Pisum sativum plant.

PubMed

Carro, Lorena; Pukall, Rüdiger; Spröer, Cathrin; Kroppenstedt, Reiner M; Trujillo, Martha E

2013-06-01

A filamentous actinomycete strain designated CR18(T) was isolated on humic acid agar from the rhizosphere of a Pisum sativum plant collected in Spain. This isolate was observed to grow optimally at 28 °C, pH 7.0 and in the presence of 5 % NaCl. Phylogenetic analyses based on the 16S rRNA gene sequence indicated a close relationship with the type strains of Micromonospora chersina and Micromonospora endolithica. A further analysis based on a concatenated DNA sequence stretch of 4,523 bp that included partial sequences of the atpD, gyrB, recA, rpoB and 16S rRNA genes clearly differentiated the new strain from recognized Micromonospora species compared. DNA-DNA hybridization studies further supported the taxonomic position of strain CR18(T) as a novel genomic species. Chemotaxonomic analyses which included whole cell sugars, polar lipids, fatty acid profiles and menaquinone composition confirmed the affiliation of the new strain to the genus Micromonospora and also highlighted differences at the species level. These studies were finally complemented with an array of physiological tests to help differentiate between the new strain and its phylogenetic neighbours. Consequently, strain CR18(T) (= CECT 7890(T) = DSM 45598(T)) is proposed as the type strain of a novel species, Micromonospora halotolerans sp. nov.

Phylogenetic analyses of bacteria associated with the processing of iru and ogiri condiments.

PubMed

Ademola, Oluwatoyin M; Adeyemi, Taiwo E; Ezeokoli, Obinna T; Ayeni, Kolawole I; Obadina, Adewale O; Somorin, Yinka M; Omemu, Adebukola M; Adeleke, Rasheed A; Nwangburuka, Cyril C; Oluwafemi, Flora; Oyewole, Olusola B; Ezekiel, Chibundu N

2018-06-27

Analysis of the bacterial community dynamics during the production of traditional fermented condiments is important for food safety assessment, quality control and development of starter culture technology. In this study, bacteria isolated during the processing of iru and ogiri, two commonly consumed condiments in Nigeria, were characterised based on phylogenetic analyses of the bacterial 16S rRNA gene. A total of 227 isolates were obtained and clustered into 12 operational taxonomic units (OTUs) based on 97% 16S rRNA gene similarity. The OTUs spanned three phyla (Firmicutes, Actinobacteria and Proteobacteria), and nine genera: Acinetobacter, Aerococcus, Bacillus, Enterococcus, Enterobacter, Lysinibacillus, Micrococcus, Proteus and Staphylococcus. OTUs closely related to species of Bacillus dominated the processing stages of both condiments. Although no single OTU occurred throughout iru processing stages, an OTU (mostly related to B. safensis) dominated the ogiri processing stages indicating potentials for the development of starter culture. However, other isolates such as those of Enterococcus spp. and Lysinibacillus spp. may be potential starters for iru fermentation. Presumptive foodborne pathogens were also detected at some stages of the condiments' processing, possibly due to poor hygienic practices. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Streptococcus ovuberis sp. nov., isolated from a subcutaneous abscess in the udder of a sheep.

PubMed

Zamora, Leydis; Pérez-Sancho, Marta; Fernández-Garayzábal, Jose Francisco; Orden, Jose Antonio; Domínguez-Bernal, Gustavo; de la Fuente, Ricardo; Domínguez, Lucas; Vela, Ana Isabel

2017-11-01

One unidentified, Gram-stain-positive, catalase-negative coccus-shaped organism was recovered from a subcutaneous abscess of the udder of a sheep and subjected to a polyphasic taxonomic analysis. Based on cellular morphology and biochemical criteria, the isolate was tentatively assigned to the genus Streptococcus, although the organism did not appear to match any recognized species. 16S rRNA gene sequence comparison studies confirmed its identification as a member of the genus Streptococcus and showed that the nearest phylogenetic relatives of the unknown coccus corresponded to Streptococcus moroccensis and Streptococcus cameli (95.9 % 16S rRNA gene sequence similarity). The sodA sequence analysis showed less than 89.3 % sequence similarity with the currently recognized species of the genus Streptococcus. The novel bacterial isolate was distinguished from close relatives of the genus Streptococcusby using biochemical tests. A mass spectrometry profile was also obtained for the novel isolate using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Based on both phenotypic and phylogenetic findings, it is proposed that the unknown bacterium be classified as a representative of a novel species of the genus Streptococcus, Streptococcus ovuberis sp. nov. The type strain of Streptococcus ovuberissp. nov. is VB15-00779 T (=CECT 9179 T =CCUG 69612 T ).

Jonquetella anthropi gen. nov., sp. nov., the first member of the candidate phylum 'Synergistetes' isolated from man.

PubMed

Jumas-Bilak, Estelle; Carlier, Jean-Philippe; Jean-Pierre, Hélène; Citron, Diane; Bernard, Kathryn; Damay, Audrey; Gay, Bernard; Teyssier, Corinne; Campos, Josiane; Marchandin, Hélène

2007-12-01

Six clinical isolates of a hitherto unknown, strictly anaerobic, Gram-negative rod showing fastidious growth were subjected to a polyphasic taxonomic study, including phenotypic, genomic and phylogenetic feature analyses. 16S rRNA gene sequenced-based phylogeny revealed that the novel strains represent a homogeneous group distant from any recognized species in the candidate phylum 'Synergistetes'. The novel isolates were most closely related to species of the genus Dethiosulfovibrio, with 88.2-88.7 % 16S rRNA gene sequence similarity. Large-scale chromosome structure and DNA G+C content also differentiated the novel strains from members of the genus Dethiosulfovibrio. The novel strains were asaccharolytic. Major metabolic end products in trypticase/glucose/yeast extract broth were acetic, lactic, succinic and isovaleric acids and the major cellular fatty acids iso-C(15 : 0) and C(16 : 0). Based on the data presented here, a new genus, Jonquetella gen. nov., is proposed with one novel species, Jonquetella anthropi sp. nov. J. anthropi is the first characterized species of the candidate phylum 'Synergistetes' that includes human isolates. The G+C content of the DNA of the type strain of J. anthropi ADV 126(T) (=AIP 136.05(T)=CIP 109408(T)=CCUG 53819(T)) is 59.4 mol%.

Diversity and population structure of Marine Group A bacteria in the Northeast subarctic Pacific Ocean.

PubMed

Allers, Elke; Wright, Jody J; Konwar, Kishori M; Howes, Charles G; Beneze, Erica; Hallam, Steven J; Sullivan, Matthew B

2013-02-01

Marine Group A (MGA) is a candidate phylum of Bacteria that is ubiquitous and abundant in the ocean. Despite being prevalent, the structural and functional properties of MGA populations remain poorly constrained. Here, we quantified MGA diversity and population structure in relation to nutrients and O(2) concentrations in the oxygen minimum zone (OMZ) of the Northeast subarctic Pacific Ocean using a combination of catalyzed reporter deposition fluorescence in situ hybridization (CARD-FISH) and 16S small subunit ribosomal RNA (16S rRNA) gene sequencing (clone libraries and 454-pyrotags). Estimates of MGA abundance as a proportion of total bacteria were similar across all three methods although estimates based on CARD-FISH were consistently lower in the OMZ (5.6%±1.9%) than estimates based on 16S rRNA gene clone libraries (11.0%±3.9%) or pyrotags (9.9%±1.8%). Five previously defined MGA subgroups were recovered in 16S rRNA gene clone libraries and five novel subgroups were defined (HF770D10, P262000D03, P41300E03, P262000N21 and A714018). Rarefaction analysis of pyrotag data indicated that the ultimate richness of MGA was very nearly sampled. Spearman's rank analysis of MGA abundances by CARD-FISH and O(2) concentrations resulted in significant correlation. Analyzed in more detail by 16S rRNA pyrotag sequencing, MGA operational taxonomic units affiliated with subgroups Arctic95A-2 and A714018 comprised 0.3-2.4% of total bacterial sequences and displayed strong correlations with decreasing O(2) concentration. This study is the first comprehensive description of MGA diversity using complementary techniques. These results provide a phylogenetic framework for interpreting future studies on ecotype selection among MGA subgroups, and suggest a potentially important role for MGA in the ecology and biogeochemistry of OMZs.

New Insights into the RNA-Based Mechanism of Action of the Anticancer Drug 5′-Fluorouracil in Eukaryotic Cells

PubMed Central

Mojardín, Laura; Botet, Javier; Quintales, Luis; Moreno, Sergio; Salas, Margarita

2013-01-01

5-Fluorouracil (5FU) is a chemotherapeutic drug widely used in treating a range of advanced, solid tumours and, in particular, colorectal cancer. Here, we used high-density tiling DNA microarray technology to obtain the specific transcriptome-wide response induced by 5FU in the eukaryotic model Schizosaccharomyces pombe. This approach combined with real-time quantitative PCR analysis allowed us to detect splicing defects of a significant number of intron-containing mRNA, in addition to identify some rRNA and tRNA processing defects after 5FU treatment. Interestingly, our studies also revealed that 5FU specifically induced the expression of certain genes implicated in the processing of mRNA, tRNA and rRNA precursors, and in the post-transcriptional modification of uracil residues in RNA. The transcription of several tRNA genes was also significantly induced after drug exposure. These transcriptional changes might represent a cellular response mechanism to counteract 5FU damage since deletion strains for some of these up-regulated genes were hypersensitive to 5FU. Moreover, most of these RNA processing genes have human orthologs that participate in conserved pathways, suggesting that they could be novel targets to improve the efficacy of 5FU-based treatments. PMID:24223771

The complete mitochondrial genome sequence of the Datong yak (Bos grunniens).

PubMed

Wu, Xiaoyun; Chu, Min; Liang, Chunnian; Ding, Xuezhi; Guo, Xian; Bao, Pengjia; Yan, Ping

2016-01-01

Datong yak is a famous artificially cultivated breed in China. In the present work, we report the complete mitochondrial genome sequence of Datong yak for the first time. The total length of the mitogenome is 16,323 bp long, containing 13 protein-coding genes, 22 tRNA genes, two rRNA genes and one non-coding region (D-loop region). The gene order of Datong yak mitogenome is identical to that observed in most other vertebrates. The overall base composition is 33.71% A, 25.8.0% C, 13.21% G and 27.27% T, with an A + T content of 60.98%. The complete mitogenome sequence information of Datong yak can provide useful data for further studies on molecular breeding and taxonomic status.

In-silico Taxonomic Classification of 373 Genomes Reveals Species Misidentification and New Genospecies within the Genus Pseudomonas

PubMed Central

Tran, Phuong N.; Savka, Michael A.; Gan, Han Ming

2017-01-01

The genus Pseudomonas has one of the largest diversity of species within the Bacteria kingdom. To date, its taxonomy is still being revised and updated. Due to the non-standardized procedure and ambiguous thresholds at species level, largely based on 16S rRNA gene or conventional biochemical assay, species identification of publicly available Pseudomonas genomes remains questionable. In this study, we performed a large-scale analysis of all Pseudomonas genomes with species designation (excluding the well-defined P. aeruginosa) and re-evaluated their taxonomic assignment via in silico genome-genome hybridization and/or genetic comparison with valid type species. Three-hundred and seventy-three pseudomonad genomes were analyzed and subsequently clustered into 145 distinct genospecies. We detected 207 erroneous labels and corrected 43 to the proper species based on Average Nucleotide Identity Multilocus Sequence Typing (MLST) sequence similarity to the type strain. Surprisingly, more than half of the genomes initially designated as Pseudomonas syringae and Pseudomonas fluorescens should be classified either to a previously described species or to a new genospecies. Notably, high pairwise average nucleotide identity (>95%) indicating species-level similarity was observed between P. synxantha-P. libanensis, P. psychrotolerans–P. oryzihabitans, and P. kilonensis- P. brassicacearum, that were previously differentiated based on conventional biochemical tests and/or genome-genome hybridization techniques. PMID:28747902

Molecular phylogeny of the Cricetinae subfamily based on the mitochondrial cytochrome b and 12S rRNA genes and the nuclear vWF gene.

PubMed

Neumann, Karsten; Michaux, Johan; Lebedev, Vladimir; Yigit, Nuri; Colak, Ercument; Ivanova, Natalia; Poltoraus, Andrey; Surov, Alexei; Markov, Georgi; Maak, Steffen; Neumann, Sabine; Gattermann, Rolf

2006-04-01

Despite some popularity of hamsters as pets and laboratory animals there is no reliable phylogeny of the subfamily Cricetinae available so far. Contradicting views exist not only about the actual number of species but also concerning the validity of several genera. We used partial DNA sequences of two mitochondrial (cytochrome b, 12S rRNA) and one partial nuclear gene (von Willebrand Factor exon 28) to provide a first gene tree of the Cricetinae based on 15 taxa comprising six genera. According to our data, Palaearctic hamsters fall into three distinct phylogenetic groups: Phodopus, Mesocricetus, and Cricetus-related species which evolved during the late Miocene about 7-12MY ago. Surprisingly, the genus Phodopus, which was previously thought to have appeared during the Pleistocene, forms the oldest clade. The largest number of extant hamster genera is found in a group of Cricetus-related hamsters. The genus Cricetulus itself proved to be not truly monophyletic with Cricetulus migratorius appearing more closely related to Tscherskia, Cricetus, and Allocricetulus. We propose to place the species within a new monotypic genus. Molecular clock calculations are not always in line with the dating of fossil records. DNA based divergence time estimates as well as taxonomic relationships demand a reevaluation of morphological characters previously used to identify fossils and extant hamsters.

Peatland succession induces a shift in the community composition of Sphagnum-associated active methanotrophs.

PubMed

Putkinen, Anuliina; Larmola, Tuula; Tuomivirta, Tero; Siljanen, Henri M P; Bodrossy, Levente; Tuittila, Eeva-Stiina; Fritze, Hannu

2014-06-01

Sphagnum-associated methanotrophs (SAM) are an important sink for the methane (CH4) formed in boreal peatlands. We aimed to reveal how peatland succession, which entails a directional change in several environmental variables, affects SAM and their activity. Based on the pmoA microarray results, SAM community structure changes when a peatland develops from a minerotrophic fen to an ombrotrophic bog. Methanotroph subtypes Ia, Ib, and II showed slightly contrasting patterns during succession, suggesting differences in their ecological niche adaptation. Although the direct DNA-based analysis revealed a high diversity of type Ib and II methanotrophs throughout the studied peatland chronosequence, stable isotope probing (SIP) of the pmoA gene indicated they were active mainly during the later stages of succession. In contrast, type Ia methanotrophs showed active CH4 consumption in all analyzed samples. SIP-derived (13)C-labeled 16S rRNA gene clone libraries revealed a high diversity of SAM in every succession stage including some putative Methylocella/Methyloferula methanotrophs that are not detectable with the pmoA-based approach. In addition, a high diversity of 16S rRNA gene sequences likely representing cross-labeled nonmethanotrophs was discovered, including a significant proportion of Verrucomicrobia-related sequences. These results help to predict the effects of changing environmental conditions on SAM communities and activity. © 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

Aestuariicola saemankumensis gen. nov., sp. nov., a member of the family Flavobacteriaceae, isolated from tidal flat sediment.

PubMed

Yoon, Jung-Hoon; Kang, So-Jung; Jung, Yong-Taek; Oh, Tae-Kwang

2008-09-01

A Gram-negative, non-motile, pleomorphic bacterial strain, designated SMK-142(T), was isolated from a tidal flat of the Yellow Sea, Korea, and was subjected to a polyphasic taxonomic study. Strain SMK-142(T) grew optimally at pH 7.0-8.0, 25 degrees C and in the presence of 2% (w/v) NaCl. Phylogenetic analyses based on 16S rRNA gene sequences showed that strain SMK-142(T) clustered with Lutibacter litoralis with which it exhibited a 16S rRNA gene sequence similarity value of 91.2%. This cluster joined the clade comprising the genera Tenacibaculum and Polaribacter at a high bootstrap resampling value. Strain SMK-142(T) contained MK-6 as the predominant menaquinone and iso-C(15:0), iso-C(15:1) and iso-C(17:0) 3-OH as the major fatty acids. The DNA G+C content was 37.2 mol%. Strain SMK-142(T) was differentiated from three phylogenetically related genera, Lutibacter, Tenacibaculum and Polaribacter, on the basis of low 16S rRNA gene sequence similarity values and differences in fatty acid profiles and in some phenotypic properties. On the basis of phenotypic, chemotaxonomic and phylogenetic data, strain SMK-142(T) represents a novel genus and species for which the name Aestuariicola saemankumensis gen. nov., sp. nov. is proposed (phylum Bacteroidetes, family Flavobacteriaceae). The type strain of the type species, Aestuariicola saemankumensis sp. nov., is SMK-142(T) (=KCTC 22171(T)=CCUG 55329(T)).

No need to replace an "anomalous" primate (Primates) with an "anomalous" bear (Carnivora, Ursidae).

PubMed

Gutiérrez, Eliécer E; Pine, Ronald H

2015-01-01

By means of mitochondrial 12S rRNA sequencing of putative "yeti", "bigfoot", and other "anomalous primate" hair samples, a recent study concluded that two samples, presented as from the Himalayas, do not belong to an "anomalous primate", but to an unknown, anomalous type of ursid. That is, that they match 12S rRNA sequences of a fossil Polar Bear (Ursusmaritimus), but neither of modern Polar Bears, nor of Brown Bears (Ursusarctos), the closest relative of Polar Bears, and one that occurs today in the Himalayas. We have undertaken direct comparison of sequences; replication of the original comparative study; inference of phylogenetic relationships of the two samples with respect to those from all extant species of Ursidae (except for the Giant Panda, Ailuropodamelanoleuca) and two extinct Pleistocene species; and application of a non-tree-based population aggregation approach for species diagnosis and identification. Our results demonstrate that the very short fragment of the 12S rRNA gene sequenced by Sykes et al. is not sufficiently informative to support the hypotheses provided by these authors with respect to the taxonomic identity of the individuals from which these sequences were obtained. We have concluded that there is no reason to believe that the two samples came from anything other than Brown Bears. These analyses afforded an opportunity to test the monophyly of morphologically defined species and to comment on both their phylogenetic relationships and future efforts necessary to advance our understanding of ursid systematics.

Pseudoclavibacter caeni sp. nov., isolated from sludge of a sewage disposal plant.

PubMed

Srinivasan, Sathiyaraj; Kim, Hyun Sook; Kim, Myung Kyum; Lee, Myungjin

2012-04-01

A Gram-positive, strictly aerobic, rod-shaped, non-motile bacterial strain, designated MJ28T, was isolated from a sludge sample from the Daejeon sewage disposal plant in South Korea. A polyphasic approach was applied to study the taxonomic position of strain MJ28T. Strain MJ28T showed highest 16S rRNA gene sequence similarity to Pseudoclavibacter soli KP02T (95.2 %). Levels of 16S rRNA gene sequence similarity to the type strains of other Pseudoclavibacter species were less than 94.0 %. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain MJ28T belonged to the clade formed by members of the genus Pseudoclavibacter in the family Microbacteriaceae. The G+C content of the genomic DNA of strain MJ28T was 65.8 mol%. The chemotaxonomic characteristics of strain MJ28T showed features typical of the genus Pseudoclavibacter, with MK-9 as the predominant respiratory quinone, 2,4-diaminobutryic acid as the diamino acid in the peptidoglycan, and anteiso-C17:0 (44.6 %), anteiso-C15:0 (35.7 %) and C16:0 (9.5 %) as the major fatty acids. On the basis of phylogenetic inference, fatty acid profile and other phenotypic properties, strain MJ28T is considered to represent a novel species of the genus Pseudoclavibacter, for which the name Pseudoclavibacter caeni sp. nov. is proposed. The type strain is MJ28T (=KCTC 19773T=JCM 16921T).

Microbial analysis in primary and persistent endodontic infections by using pyrosequencing.

PubMed

Hong, Bo-Young; Lee, Tae-Kwon; Lim, Sang-Min; Chang, Seok Woo; Park, Joonhong; Han, Seung Hyun; Zhu, Qiang; Safavi, Kamran E; Fouad, Ashraf F; Kum, Kee Yeon

2013-09-01

The aim of this study was to investigate the bacterial community profile of intracanal microbiota in primary and persistent endodontic infections associated with asymptomatic chronic apical periodontitis by using GS-FLX Titanium pyrosequencing. The null hypothesis was that there is no difference in diversity of overall bacterial community profiles between primary and persistent infections. Pyrosequencing analysis from 10 untreated and 8 root-filled samples was conducted. Analysis from 18 samples yielded total of 124,767 16S rRNA gene sequences (with a mean of 6932 reads per sample) that were taxonomically assigned into 803 operational taxonomic units (3% distinction), 148 genera, and 10 phyla including unclassified. Bacteroidetes was the most abundant phylum in both primary and persistent infections. There were no significant differences in bacterial diversity between the 2 infection groups (P > .05). The bacterial community profile that was based on dendrogram showed that bacterial population in both infections was not significantly different in their structure and composition (P > .05). The present pyrosequencing study demonstrates that persistent infections have as diverse bacterial community as primary infections. Copyright © 2013 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

Neofusicoccum ribis Associated with Leaf Blight on Rubber (Hevea brasiliensis) in Peninsular Malaysia

PubMed Central

Nyaka Ngobisa, A. I. C.; Zainal Abidin, M. A.; Wong, M. Y.; Wan Noordin, M. W. D.

2013-01-01

Hevea brasiliensis is a natural source of rubber and an important plantation tree species in Malaysia. Leaf blight disease caused by Fusicoccum substantially reduces the growth and performance of H. brasiliensis. The aim of this study was to use a combination of both morphological characteristics and molecular data to clarify the taxonomic position of the fungus associated with leaf blight disease. Fusicoccum species were isolated from infected leaves collected from plantations at 3 widely separated locations – Selangor, Perak, and Johor states – in Peninsular Malaysia in 2010. All the isolates were identified according to their conidial patterns and DNA sequences generated from internal transcribed spacers (ITS1 and ITS2), the 5.8S rRNA, and an unknown locus (BotF15) containing microsatellite repeats. Based on taxonomic and sequence data, Neofusicoccum ribis was identified as the main cause of leaf blight disease in H. brasiliensis in commercial plantations in Malaysia. A pathogenicity trial on detached leaves further confirmed that N. ribis causes leaf blight disease. N. ribis is an important leaf pathogen, and its detection in Malaysia has important implications for future planting of H. brasiliensis. PMID:25288924

Neofusicoccum ribis Associated with Leaf Blight on Rubber (Hevea brasiliensis) in Peninsular Malaysia.

PubMed

Nyaka Ngobisa, A I C; Zainal Abidin, M A; Wong, M Y; Wan Noordin, M W D

2013-03-01

Hevea brasiliensis is a natural source of rubber and an important plantation tree species in Malaysia. Leaf blight disease caused by Fusicoccum substantially reduces the growth and performance of H. brasiliensis. The aim of this study was to use a combination of both morphological characteristics and molecular data to clarify the taxonomic position of the fungus associated with leaf blight disease. Fusicoccum species were isolated from infected leaves collected from plantations at 3 widely separated locations - Selangor, Perak, and Johor states - in Peninsular Malaysia in 2010. All the isolates were identified according to their conidial patterns and DNA sequences generated from internal transcribed spacers (ITS1 and ITS2), the 5.8S rRNA, and an unknown locus (BotF15) containing microsatellite repeats. Based on taxonomic and sequence data, Neofusicoccum ribis was identified as the main cause of leaf blight disease in H. brasiliensis in commercial plantations in Malaysia. A pathogenicity trial on detached leaves further confirmed that N. ribis causes leaf blight disease. N. ribis is an important leaf pathogen, and its detection in Malaysia has important implications for future planting of H. brasiliensis.

Increased diversity of egg-associated bacteria on brown trout (Salmo trutta) at elevated temperatures.

PubMed

Wilkins, Laetitia G E; Rogivue, Aude; Schütz, Frédéric; Fumagalli, Luca; Wedekind, Claus

2015-11-27

The taxonomic composition of egg-associated microbial communities can play a crucial role in the development of fish embryos. In response, hosts increasingly influence the composition of their associated microbial communities during embryogenesis, as concluded from recent field studies and laboratory experiments. However, little is known about the taxonomic composition and the diversity of egg-associated microbial communities within ecosystems; e.g., river networks. We sampled late embryonic stages of naturally spawned brown trout at nine locations within two different river networks and applied 16S rRNA pyrosequencing to describe their bacterial communities. We found no evidence for a significant isolation-by-distance effect on the composition of bacterial communities, and no association between neutral genetic divergence of fish host (based on 11 microsatellites) and phylogenetic distances of the composition of their associated bacterial communities. We characterized core bacterial communities on brown trout eggs and compared them to corresponding water samples with regard to bacterial composition and its presumptive function. Bacterial diversity was positively correlated with water temperature at the spawning locations. We discuss this finding in the context of the increased water temperatures that have been recorded during the last 25 years in the study area.

Increased diversity of egg-associated bacteria on brown trout (Salmo trutta) at elevated temperatures

PubMed Central

Wilkins, Laetitia G. E.; Rogivue, Aude; Schütz, Frédéric; Fumagalli, Luca; Wedekind, Claus

2015-01-01

The taxonomic composition of egg-associated microbial communities can play a crucial role in the development of fish embryos. In response, hosts increasingly influence the composition of their associated microbial communities during embryogenesis, as concluded from recent field studies and laboratory experiments. However, little is known about the taxonomic composition and the diversity of egg-associated microbial communities within ecosystems; e.g., river networks. We sampled late embryonic stages of naturally spawned brown trout at nine locations within two different river networks and applied 16S rRNA pyrosequencing to describe their bacterial communities. We found no evidence for a significant isolation-by-distance effect on the composition of bacterial communities, and no association between neutral genetic divergence of fish host (based on 11 microsatellites) and phylogenetic distances of the composition of their associated bacterial communities. We characterized core bacterial communities on brown trout eggs and compared them to corresponding water samples with regard to bacterial composition and its presumptive function. Bacterial diversity was positively correlated with water temperature at the spawning locations. We discuss this finding in the context of the increased water temperatures that have been recorded during the last 25 years in the study area. PMID:26611640

Dynamics of bacterial communities during manufacture and ripening of traditional Caciocavallo of Castelfranco cheese in relation to cows' feeding.

PubMed

Giello, Marina; La Storia, Antonietta; Masucci, Felicia; Di Francia, Antonio; Ercolini, Danilo; Villani, Francesco

2017-05-01

Traditional Caciocavallo of Castelfranco is a semi-hard "pasta-filata" cheese produced from raw cows' milk in Campania region. The aim of the present research is mainly focused on the study, by 16S rRNA gene pyrosequencing and viable counts, of the dynamics of bacterial communities during manufacture and ripening of traditional Caciocavallo cheese. Moreover, the possible correlation between cheese microbiota and cows' feeding based on silage or hay was also evaluated. In general, except for enterococci, the technological process significantly affected all the microbial groups. According to 16S rRNA, raw cows' milk was dominated by Streptococcus thermophilus, L. lactis and Pseudomonas sp. in hay cheese production, whereas Lactococcus lactis and Acinetobacter sp. dominated silage production. Differences in the taxonomic structure of the milk's microbiota within diet groups were not related to silage and hay cows' feeding. Moreover, S. thermophilus was the unique species that dominate from raw milks to fermented intermediates and cheese in both hay and silage cheese productions. Feeding and ripening time influenced significantly sensory characteristics of the cheeses. Copyright © 2016 Elsevier Ltd. All rights reserved.

Identification, characterization and description of Arcobacter faecis sp. nov., isolated from a human waste septic tank.

PubMed

Whiteduck-Léveillée, Kerri; Whiteduck-Léveillée, Jenni; Cloutier, Michel; Tambong, James T; Xu, Renlin; Topp, Edward; Arts, Michael T; Chao, Jerry; Adam, Zaky; Lévesque, C André; Lapen, David R; Villemur, Richard; Khan, Izhar U H

2016-03-01

A study on the taxonomic classification of Arcobacter species was performed on the cultures isolated from various fecal sources where an Arcobacter strain AF1078(T) from human waste septic tank near Ottawa, Ontario, Canada was characterized using a polyphasic approach. Genetic investigations including 16S rRNA, atpA, cpn60, gyrA, gyrB and rpoB gene sequences of strain AF1078(T) are unique in comparison with other arcobacters. Phylogenetic analysis based on the 16S rRNA gene sequence revealed that the strain is most closely related to Arcobacter lanthieri and Arcobacter cibarius. Analyses of atpA, cpn60, gyrA, gyrB and rpoB gene sequences suggested that strain AF1078(T) formed a phylogenetic lineage independent of other species in the genus. Whole-genome sequence, DNA-DNA hybridization, fatty acid profile and phenotypic analysis further supported the conclusion that strain AF1078(T) represents a novel Arcobacter species, for which the name Arcobacter faecis sp. nov. is proposed, with type strain AF1078(T) (=LMG 28519(T); CCUG 66484(T)). Crown Copyright © 2015. Published by Elsevier GmbH. All rights reserved.

Diversity of bacteria and glycosyl hydrolase family 48 genes in cellulolytic consortia enriched from thermophilic biocompost.

PubMed

Izquierdo, Javier A; Sizova, Maria V; Lynd, Lee R

2010-06-01

The enrichment from nature of novel microbial communities with high cellulolytic activity is useful in the identification of novel organisms and novel functions that enhance the fundamental understanding of microbial cellulose degradation. In this work we identify predominant organisms in three cellulolytic enrichment cultures with thermophilic compost as an inoculum. Community structure based on 16S rRNA gene clone libraries featured extensive representation of clostridia from cluster III, with minor representation of clostridial clusters I and XIV and a novel Lutispora species cluster. Our studies reveal different levels of 16S rRNA gene diversity, ranging from 3 to 18 operational taxonomic units (OTUs), as well as variability in community membership across the three enrichment cultures. By comparison, glycosyl hydrolase family 48 (GHF48) diversity analyses revealed a narrower breadth of novel clostridial genes associated with cultured and uncultured cellulose degraders. The novel GHF48 genes identified in this study were related to the novel clostridia Clostridium straminisolvens and Clostridium clariflavum, with one cluster sharing as little as 73% sequence similarity with the closest known relative. In all, 14 new GHF48 gene sequences were added to the known diversity of 35 genes from cultured species.

Comparison of the Microbial Community Structures of Untreated Wastewaters from Different Geographic Locales

PubMed Central

Shanks, Orin C.; Newton, Ryan J.; Kelty, Catherine A.; Huse, Susan M.; Sogin, Mitchell L.

2013-01-01

Microbial sewage communities consist of a combination of human fecal microorganisms and nonfecal microorganisms, which may be residents of urban sewer infrastructure or flowthrough originating from gray water or rainwater inputs. Together, these different microorganism sources form an identifiable community structure that may serve as a signature for sewage discharges and as candidates for alternative indicators specific for human fecal pollution. However, the structure and variability of this community across geographic space remains uncharacterized. We used massively parallel 454 pyrosequencing of the V6 region in 16S rRNA genes to profile microbial communities from 13 untreated sewage influent samples collected from a wide range of geographic locations in the United States. We obtained a total of 380,175 high-quality sequences for sequence-based clustering, taxonomic analyses, and profile comparisons. The sewage profile included a discernible core human fecal signature made up of several abundant taxonomic groups within Firmicutes, Bacteroidetes, Actinobacteria, and Proteobacteria. DNA sequences were also classified into fecal, sewage infrastructure (i.e., nonfecal), and transient groups based on data comparisons with fecal samples. Across all sewage samples, an estimated 12.1% of sequences were fecal in origin, while 81.4% were consistently associated with the sewage infrastructure. The composition of feces-derived operational taxonomic units remained congruent across all sewage samples regardless of geographic locale; however, the sewage infrastructure community composition varied among cities, with city latitude best explaining this variation. Together, these results suggest that untreated sewage microbial communities harbor a core group of fecal bacteria across geographically dispersed wastewater sewage lines and that ambient water quality indicators targeting these select core microorganisms may perform well across the United States. PMID:23435885

Characterization of the methanogen community in a household anaerobic digester fed with swine manure in China.

PubMed

Qin, Huibin; Lang, Huihua; Yang, Hongjiang

2013-09-01

Household anaerobic digesters have been installed across rural China for biogas production, but information on methanogen community structure in these small biogas units is sparsely available. By creating clone libraries for 16S rRNA and methyl coenzyme M reductase alpha subunit (mcrA) genes, we investigated the methanogenic consortia in a household biogas digester treating swine manure. Operational taxonomic units (OTUs) were defined by comparative sequence analysis, seven OTUs were identified in the 16S rRNA gene library, and ten OTUs were identified in the mcrA gene library. Both libraries were dominated by clones highly related to the type strain Methanocorpusculum labreanum Z, 64.0 % for 16S rRNA gene clones and 64.3 % for mcrA gene clones. Additionally, gas chromatography assays showed that formic acid was 84.54 % of the total volatile fatty acids and methane was 57.20 % of the biogas composition. Our results may help further isolation and characterization of methanogenic starter strains for industrial biogas production.

Characterization of Microbial Communities Associated With Deep-Sea Hydrothermal Vent Animals of the East Pacific Rise and the Galápagos Rift

NASA Astrophysics Data System (ADS)

Ward, N.; Page, S.; Heidelberg, J.; Eisen, J. A.; Fraser, C. M.

2002-12-01

The composition of microbial communities associated with deep-sea hydrothermal vent animals is of interest because of the key role of bacterial symbionts in driving the chemosynthetic food chain of the vent system, and also because bacterial biofilms attached to animal exterior surfaces may play a part in settlement of larval forms. Sequence analysis of 16S ribosomal RNA (rRNA) genes from such communities provides a snapshot of community structure, as this gene is present in all Bacteria and Archaea, and a useful phylogenetic marker for both cultivated microbial species, and uncultivated species such as many of those found in the deep-sea environment. Specimens of giant tube worms (Riftia pachyptila), mussels (Bathymodiolus thermophilus), and clams (Calyptogena magnifica) were collected during the 2002 R/V Atlantis research cruises to the East Pacific Rise (9N) and Galápagos Rift. Microbial biofilms attached to the exterior surfaces of individual animals were sampled, as were tissues known to harbor chemosynthetic bacterial endosymbionts. Genomic DNA was extracted from the samples using a commercially available kit, and 16S rRNA genes amplified from the mixed bacterial communities using the polymerase chain reaction (PCR) and oligonucleotide primers targeting conserved terminal regions of the 16S rRNA gene. The PCR products obtained were cloned into a plasmid vector and the recombinant plasmids transformed into cells of Escherichia coli. Individual cloned 16S rRNA genes were sequenced at the 5' end of the gene (the most phylogenetically informative region in most taxa) and the sequence data compared to publicly available gene sequence databases, to allow a preliminary assignment of clones to taxonomic groups within the Bacteria and Archaea, and to determine the overall composition and phylogenetic diversity of the animal-associated microbial communities. Analysis of Riftia pachyptila exterior biofilm samples revealed the presence of members of the delta and epsilon proteobacteria, low GC Gram positive bacteria (firmicutes), spirochetes, CFB (Cytophaga-Flavobacterium-Bacteroides) group, green nonsulfur bacteria, acidobacteria, verrucomicrobia, and planctomycetes. The presence of the latter three taxonomic groups is of special interest, as they represent phylogenetically distinct groups within the Bacteria for which specific ecological functions have not yet been identified, but which have been found to be widely distributed and often numerically significant in diverse terrestrial and aquatic habitats. Although further sequencing is required to demonstrate the presence of a Riftia-associated microbial population distinct from that of the surrounding seawater, results available from three Riftia individuals from the East Pacific Rise suggest this to be the case. Analysis of microbial communities associated with the gill tissue of the mussel Bathymodiolus thermophilus shows a population dominated by gamma-Proteobacterial chemoautotrophic symbionts, although lower frequency novel phylotypes have been detected. Representatives of specific taxonomic groups have been selected for sequencing of the complete 16S rRNA gene, and the sequences used to reconstruct phylogenetic trees to more accurately determine the evolutionary relationships between the novel sequences, and available sequences for both cultured and non-cultured bacteria.

Variable Copy Number, Intra-Genomic Heterogeneities and Lateral Transfers of the 16S rRNA Gene in Pseudomonas

PubMed Central

Bodilis, Josselin; Nsigue-Meilo, Sandrine; Besaury, Ludovic; Quillet, Laurent

2012-01-01

Even though the 16S rRNA gene is the most commonly used taxonomic marker in microbial ecology, its poor resolution is still not fully understood at the intra-genus level. In this work, the number of rRNA gene operons, intra-genomic heterogeneities and lateral transfers were investigated at a fine-scale resolution, throughout the Pseudomonas genus. In addition to nineteen sequenced Pseudomonas strains, we determined the 16S rRNA copy number in four other Pseudomonas strains by Southern hybridization and Pulsed-Field Gel Electrophoresis, and studied the intra-genomic heterogeneities by Denaturing Gradient Gel Electrophoresis and sequencing. Although the variable copy number (from four to seven) seems to be correlated with the evolutionary distance, some close strains in the P. fluorescens lineage showed a different number of 16S rRNA genes, whereas all the strains in the P. aeruginosa lineage displayed the same number of genes (four copies). Further study of the intra-genomic heterogeneities revealed that most of the Pseudomonas strains (15 out of 19 strains) had at least two different 16S rRNA alleles. A great difference (5 or 19 nucleotides, essentially grouped near the V1 hypervariable region) was observed only in two sequenced strains. In one of our strains studied (MFY30 strain), we found a difference of 12 nucleotides (grouped in the V3 hypervariable region) between copies of the 16S rRNA gene. Finally, occurrence of partial lateral transfers of the 16S rRNA gene was further investigated in 1803 full-length sequences of Pseudomonas available in the databases. Remarkably, we found that the two most variable regions (the V1 and V3 hypervariable regions) had probably been laterally transferred from another evolutionary distant Pseudomonas strain for at least 48.3 and 41.6% of the 16S rRNA sequences, respectively. In conclusion, we strongly recommend removing these regions of the 16S rRNA gene during the intra-genus diversity studies. PMID:22545126

Cellulosilyticum ruminicola gen. nov., sp. nov., isolated from the rumen of yak, and reclassification of Clostridium lentocellum as Cellulosilyticum lentocellum comb. nov.

PubMed

Cai, Shichun; Dong, Xiuzhu

2010-04-01

An obligate anaerobic, Gram-staining-negative, mesophilic, cellulolytic bacterium, strain H1(T), was isolated from the rumen content of yak. Cells were straight to slightly curved rods, 0.8-1.0 x 3.0-4.0 microm in size, non-motile and encapsulated with mucous materials. Elliptical and terminal spores that swelled the cells were produced occasionally. The strain grew at 25-45 degrees C (optimum, 38 degrees C) and pH 6.0-7.8 (optimum, pH 6.7). Cellulose, cellobiose, xylan, xylose and maltose were used as carbon and energy sources, but not glucose. Products from cellulose and cellobiose fermentation were formic acid, acetic acid, carbon dioxide and trace amounts of ethanol, lactic acid and succinic acid. The genomic DNA G+C content was 33.7+/-1.2 mol%. The predominant fatty acids were C(16 : 0) (27.1 %), C(14 : 0) (9.2 %) and iso-C( 16 : 0) (6.4%). Based on the 16S rRNA gene sequence analysis, strain H1(T) was affiliated to the clostridial rRNA cluster XIVb and showed the highest 16S rRNA gene sequence similarity to Clostridium lentocellum DSM 5427(T) (96.0 %). These two strains formed a distinct lineage of the family 'Lachnospiraceae '. Based on data from this polyphasic taxonomic study, a new genus, Cellulosilyticum gen. nov., is proposed. Cellulosilyticum ruminicola sp. nov. is proposed for strain H1(T). The type strain of Cellulosilyticum ruminicola sp. nov. is strain H1(T) (=CGMCC 1.5065(T)=JCM 14822(T)). Clostridium lentocellum was reclassified in the new genus as Cellulosilyticum lentocellum comb. nov. (type strain RHM5(T)=ATCC 49066( T)=DSM 5427(T)=NCIMB 11756(T)).

Emergence of Cryptosporidium hominis Monkey Genotype II and Novel Subtype Family Ik in the Squirrel Monkey (Saimiri sciureus) in China.

PubMed

Liu, Xuehan; Xie, Na; Li, Wei; Zhou, Ziyao; Zhong, Zhijun; Shen, Liuhong; Cao, Suizhong; Yu, Xingming; Hu, Yanchuan; Chen, Weigang; Peng, Gangneng

2015-01-01

A single Cryptosporidium isolate from a squirrel monkey with no clinical symptoms was obtained from a zoo in Ya'an city, China, and was genotyped by PCR amplification and DNA sequencing of the small-subunit ribosomal RNA (SSU rRNA), 70-kDa heat shock protein (HSP70), Cryptosporidium oocyst wall protein, and actin genes. This multilocus genetic characterization determined that the isolate was Cryptosporidium hominis, but carried 2, 10, and 6 nucleotide differences in the SSU rRNA, HSP70, and actin loci, respectively, which is comparable to the variations at these loci between C. hominis and the previously reported monkey genotype (2, 3, and 3 nucleotide differences). Phylogenetic studies, based on neighbor-joining and maximum likelihood methods, showed that the isolate identified in the current study had a distinctly discordant taxonomic status, distinct from known C. hominis and also from the monkey genotype, with respect to the three loci. Restriction fragment length polymorphisms of the SSU rRNA gene obtained from this study were similar to those of known C. hominis but clearly differentiated from the monkey genotype. Further subtyping was performed by sequence analysis of the gene encoding the 60-kDa glycoprotein (gp60). Maximum homology of only 88.3% to C. hominis subtype IdA10G4 was observed for the current isolate, and phylogenetic analysis demonstrated that this particular isolate belonged to a novel C. hominis subtype family, IkA7G4. This study is the first to report C. hominis infection in the squirrel monkey and, based on the observed genetic characteristics, confirms a new C. hominis genotype, monkey genotype II. Thus, these results provide novel insights into genotypic variation in C. hominis.

Resolving prokaryotic taxonomy without rRNA: longer oligonucleotide word lengths improve genome and metagenome taxonomic classification.

PubMed

Alsop, Eric B; Raymond, Jason

2013-01-01

Oligonucleotide signatures, especially tetranucleotide signatures, have been used as method for homology binning by exploiting an organism's inherent biases towards the use of specific oligonucleotide words. Tetranucleotide signatures have been especially useful in environmental metagenomics samples as many of these samples contain organisms from poorly classified phyla which cannot be easily identified using traditional homology methods, including NCBI BLAST. This study examines oligonucleotide signatures across 1,424 completed genomes from across the tree of life, substantially expanding upon previous work. A comprehensive analysis of mononucleotide through nonanucleotide word lengths suggests that longer word lengths substantially improve the classification of DNA fragments across a range of sizes of relevance to high throughput sequencing. We find that, at present, heptanucleotide signatures represent an optimal balance between prediction accuracy and computational time for resolving taxonomy using both genomic and metagenomic fragments. We directly compare the ability of tetranucleotide and heptanucleotide world lengths (tetranucleotide signatures are the current standard for oligonucleotide word usage analyses) for taxonomic binning of metagenome reads. We present evidence that heptanucleotide word lengths consistently provide more taxonomic resolving power, particularly in distinguishing between closely related organisms that are often present in metagenomic samples. This implies that longer oligonucleotide word lengths should replace tetranucleotide signatures for most analyses. Finally, we show that the application of longer word lengths to metagenomic datasets leads to more accurate taxonomic binning of DNA scaffolds and have the potential to substantially improve taxonomic assignment and assembly of metagenomic data.

Taxonomic and Functional Differences between Microbial Communities in Qinghai Lake and Its Input Streams

PubMed Central

Ren, Ze; Wang, Fang; Qu, Xiaodong; Elser, James J.; Liu, Yang; Chu, Limin

2017-01-01

Understanding microbial communities in terms of taxon and function is essential to decipher the biogeochemical cycling in aquatic ecosystems. Lakes and their input streams are highly linked. However, the differences between microbial assemblages in streams and lakes are still unclear. In this study, we conducted an intensive field sampling of microbial communities from lake water and stream biofilms in the Qinghai Lake watershed, the largest lake in China. We determined bacterial communities using high-throughput 16S rRNA gene sequencing and predicted functional profiles using PICRUSt to determine the taxonomic and functional differences between microbial communities in stream biofilms and lake water. The results showed that stream biofilms and lake water harbored distinct microbial communities. The microbial communities were different taxonomically and functionally between stream and lake. Moreover, streams biofilms had a microbial network with higher connectivity and modularity than lake water. Functional beta diversity was strongly correlated with taxonomic beta diversity in both the stream and lake microbial communities. Lake microbial assemblages displayed greater predicted metabolic potentials of many metabolism pathways while the microbial assemblages in stream biofilms were more abundant in xenobiotic biodegradation and metabolism and lipid metabolism. Furthermore, lake microbial assemblages had stronger predicted metabolic potentials in amino acid metabolism, carbon fixation, and photosynthesis while stream microbial assemblages were higher in carbohydrate metabolism, oxidative phosphorylation, and nitrogen metabolism. This study adds to our knowledge of stream-lake linkages from the functional and taxonomic composition of microbial assemblages. PMID:29213266

Taxonomic differences of gut microbiomes drive cellulolytic enzymatic potential within hind-gut fermenting mammals.

PubMed

Finlayson-Trick, Emma C L; Getz, Landon J; Slaine, Patrick D; Thornbury, Mackenzie; Lamoureux, Emily; Cook, Jamie; Langille, Morgan G I; Murray, Lois E; McCormick, Craig; Rohde, John R; Cheng, Zhenyu

2017-01-01

Host diet influences the diversity and metabolic activities of the gut microbiome. Previous studies have shown that the gut microbiome provides a wide array of enzymes that enable processing of diverse dietary components. Because the primary diet of the porcupine, Erethizon dorsatum, is lignified plant material, we reasoned that the porcupine microbiome would be replete with enzymes required to degrade lignocellulose. Here, we report on the bacterial composition in the porcupine microbiome using 16S rRNA sequencing and bioinformatics analysis. We extended this analysis to the microbiomes of 20 additional mammals located in Shubenacadie Wildlife Park (Nova Scotia, Canada), enabling the comparison of bacterial diversity amongst three mammalian taxonomic orders (Rodentia, Carnivora, and Artiodactyla). 16S rRNA sequencing was validated using metagenomic shotgun sequencing on selected herbivores (porcupine, beaver) and carnivores (coyote, Arctic wolf). In the microbiome, functionality is more conserved than bacterial composition, thus we mined microbiome data sets to identify conserved microbial functions across species in each order. We measured the relative gene abundances for cellobiose phosphorylase, endoglucanase, and beta-glucosidase to evaluate the cellulose-degrading potential of select mammals. The porcupine and beaver had higher proportions of genes encoding cellulose-degrading enzymes than the Artic wolf and coyote. These findings provide further evidence that gut microbiome diversity and metabolic capacity are influenced by host diet.

Taxonomic differences of gut microbiomes drive cellulolytic enzymatic potential within hind-gut fermenting mammals

PubMed Central

Thornbury, Mackenzie; Lamoureux, Emily; Cook, Jamie; Langille, Morgan G. I.; Murray, Lois E.; McCormick, Craig; Rohde, John R.

2017-01-01

Host diet influences the diversity and metabolic activities of the gut microbiome. Previous studies have shown that the gut microbiome provides a wide array of enzymes that enable processing of diverse dietary components. Because the primary diet of the porcupine, Erethizon dorsatum, is lignified plant material, we reasoned that the porcupine microbiome would be replete with enzymes required to degrade lignocellulose. Here, we report on the bacterial composition in the porcupine microbiome using 16S rRNA sequencing and bioinformatics analysis. We extended this analysis to the microbiomes of 20 additional mammals located in Shubenacadie Wildlife Park (Nova Scotia, Canada), enabling the comparison of bacterial diversity amongst three mammalian taxonomic orders (Rodentia, Carnivora, and Artiodactyla). 16S rRNA sequencing was validated using metagenomic shotgun sequencing on selected herbivores (porcupine, beaver) and carnivores (coyote, Arctic wolf). In the microbiome, functionality is more conserved than bacterial composition, thus we mined microbiome data sets to identify conserved microbial functions across species in each order. We measured the relative gene abundances for cellobiose phosphorylase, endoglucanase, and beta-glucosidase to evaluate the cellulose-degrading potential of select mammals. The porcupine and beaver had higher proportions of genes encoding cellulose-degrading enzymes than the Artic wolf and coyote. These findings provide further evidence that gut microbiome diversity and metabolic capacity are influenced by host diet. PMID:29281673

The molecular systematics of blowflies and screwworm flies (Diptera: Calliphoridae) using 28S rRNA, COX1 and EF-1α: insights into the evolution of dipteran parasitism.

PubMed

McDonagh, Laura M; Stevens, Jamie R

2011-11-01

The Calliphoridae include some of the most economically significant myiasis-causing flies in the world - blowflies and screwworm flies - with many being notorious for their parasitism of livestock. However, despite more than 50 years of research, key taxonomic relationships within the family remain unresolved. This study utilizes nucleotide sequence data from the protein-coding genes COX1 (mitochondrial) and EF1α (nuclear), and the 28S rRNA (nuclear) gene, from 57 blowfly taxa to improve resolution of key evolutionary relationships within the family Calliphoridae. Bayesian phylogenetic inference was carried out for each single-gene data set, demonstrating significant topological difference between the three gene trees. Nevertheless, all gene trees supported a Calliphorinae-Luciliinae subfamily sister-lineage, with respect to Chrysomyinae. In addition, this study also elucidates the taxonomic and evolutionary status of several less well-studied groups, including the genus Bengalia (either within Calliphoridae or as a separate sister-family), genus Onesia (as a sister-genera to, or sub-genera within, Calliphora), genus Dyscritomyia and Lucilia bufonivora, a specialised parasite of frogs and toads. The occurrence of cross-species hybridisation within Calliphoridae is also further explored, focusing on the two economically significant species Lucilia cuprina and Lucilia sericata. In summary, this study represents the most comprehensive molecular phylogenetic analysis of family Calliphoridae undertaken to date.

Lactobacillus curtus sp. nov., isolated from beer in Finland.

PubMed

Asakawa, Yuki; Takesue, Nobuchika; Asano, Shizuka; Shimotsu, Satoshi; Iijima, Kazumaru; Suzuki, Koji; Motoyama, Yasuo; Aizawa, Masayuki

2017-10-01

A Gram-stain-positive, catalase-negative and short-rod-shaped organism, designated VTT E-94560, was isolated from beer in Finland and deposited in the VTT culture collection as a strain of Lactobacillus rossiae. However, the results of 16S rRNA gene sequence analysis showed that VTT E-94560 was only related to Lactobacillus rossiae JCM 16176 T with 97.0 % sequence similarity, lower than the 98.7 % regarded as the boundary for the species differentiation. Additional phylogenetic studies on the pheS gene, rpoA gene and 16S-23S rRNA internally transcribed spacer region further reinforced the taxonomically independent status of VTT E-94560 and its related Lactobacillus species including L. rossiae and Lactobacillus siliginis. Strain VTT E-94560 also exhibited several differences in its carbohydrate fermentation profiles from those related Lactobacillus species. In addition, DNA-DNA relatedness between VTT E-94560 and these two type strains was 4 % (L. rossiae JCM 16176 T ) and 12 % (L. siliginins JCM 16155 T ), respectively, which were lower than the 70 % cut-off for general species delineation, indicating that these three strains are not taxonomically identical at the species level. These studies revealed that VTT E-94560 represents a novel species, for which the name Lactobacillus curtus sp. nov. is proposed. The type strain is VTT E-94560 T (=JCM 31185 T ).

Development of a Genome-Proxy Microarray for Profiling Marine Microbial Communities and its Application to a Time Series in Monterey Bay, California

DTIC Science & Technology

2008-09-01

community representation. 12 survey a complex microbial community. Community DNA or rRNA extracted from a sample may require amplification before...restricted to cultivated clades, since not only do many clades have sufficient database representation due to 16S environmental surveys , but such...well developed for standard and comprehensive surveys . Depending on the population being targeted and the identification method, FCM can be a

Weissella fabaria sp. nov., from a Ghanaian cocoa fermentation.

PubMed

De Bruyne, Katrien; Camu, Nicholas; De Vuyst, Luc; Vandamme, Peter

2010-09-01

Two lactic acid bacteria, strains 257(T) and 252, were isolated from traditional heap fermentations of Ghanaian cocoa beans. 16S rRNA gene sequence analysis of these strains allocated them to the genus Weissella, showing 99.5 % 16S rRNA gene sequence similarity towards Weissella ghanensis LMG 24286(T). Whole-cell protein electrophoresis, fluorescent amplified fragment length polymorphism fingerprinting of whole genomes and biochemical tests confirmed their unique taxonomic position. DNA-DNA hybridization experiments towards their nearest phylogenetic neighbour demonstrated that the two strains represent a novel species, for which we propose the name Weissella fabaria sp. nov., with strain 257(T) (=LMG 24289(T) =DSM 21416(T)) as the type strain. Additional sequence analysis using pheS gene sequences proved useful for identification of all Weissella-Leuconostoc-Oenococcus species and for the recognition of the novel species.

Microbial Contaminants of Cord Blood Units Identified by 16S rRNA Sequencing and by API Test System, and Antibiotic Sensitivity Profiling

PubMed Central

França, Luís; Simões, Catarina; Taborda, Marco; Diogo, Catarina; da Costa, Milton S.

2015-01-01

Over a period of ten months a total of 5618 cord blood units (CBU) were screened for microbial contamination under routine conditions. The antibiotic resistance profile for all isolates was also examined using ATB strips. The detection rate for culture positive units was 7.5%, corresponding to 422 samples.16S rRNA sequence analysis and identification with API test system were used to identify the culturable aerobic, microaerophilic and anaerobic bacteria from CBUs. From these samples we recovered 485 isolates (84 operational taxonomic units, OTUs) assigned to the classes Bacteroidia, Actinobacteria, Clostridia, Bacilli, Betaproteobacteria and primarily to the Gammaproteobacteria. Sixty-nine OTUs, corresponding to 447 isolates, showed 16S rRNA sequence similarities above 99.0% with known cultured bacteria. However, 14 OTUs had 16S rRNA sequence similarities between 95 and 99% in support of genus level identification and one OTU with 16S rRNA sequence similarity of 90.3% supporting a family level identification only. The phenotypic identification formed 29 OTUs that could be identified to the species level and 9 OTUs that could be identified to the genus level by API test system. We failed to obtain identification for 14 OTUs, while 32 OTUs comprised organisms producing mixed identifications. Forty-two OTUs covered species not included in the API system databases. The API test system Rapid ID 32 Strep and Rapid ID 32 E showed the highest proportion of identifications to the species level, the lowest ratio of unidentified results and the highest agreement to the results of 16S rRNA assignments. Isolates affiliated to the Bacilli and Bacteroidia showed the highest antibiotic multi-resistance indices and microorganisms of the Clostridia displayed the most antibiotic sensitive phenotypes. PMID:26512991

Microbial Contaminants of Cord Blood Units Identified by 16S rRNA Sequencing and by API Test System, and Antibiotic Sensitivity Profiling.

PubMed

França, Luís; Simões, Catarina; Taborda, Marco; Diogo, Catarina; da Costa, Milton S

2015-01-01

Over a period of ten months a total of 5618 cord blood units (CBU) were screened for microbial contamination under routine conditions. The antibiotic resistance profile for all isolates was also examined using ATB strips. The detection rate for culture positive units was 7.5%, corresponding to 422 samples.16S rRNA sequence analysis and identification with API test system were used to identify the culturable aerobic, microaerophilic and anaerobic bacteria from CBUs. From these samples we recovered 485 isolates (84 operational taxonomic units, OTUs) assigned to the classes Bacteroidia, Actinobacteria, Clostridia, Bacilli, Betaproteobacteria and primarily to the Gammaproteobacteria. Sixty-nine OTUs, corresponding to 447 isolates, showed 16S rRNA sequence similarities above 99.0% with known cultured bacteria. However, 14 OTUs had 16S rRNA sequence similarities between 95 and 99% in support of genus level identification and one OTU with 16S rRNA sequence similarity of 90.3% supporting a family level identification only. The phenotypic identification formed 29 OTUs that could be identified to the species level and 9 OTUs that could be identified to the genus level by API test system. We failed to obtain identification for 14 OTUs, while 32 OTUs comprised organisms producing mixed identifications. Forty-two OTUs covered species not included in the API system databases. The API test system Rapid ID 32 Strep and Rapid ID 32 E showed the highest proportion of identifications to the species level, the lowest ratio of unidentified results and the highest agreement to the results of 16S rRNA assignments. Isolates affiliated to the Bacilli and Bacteroidia showed the highest antibiotic multi-resistance indices and microorganisms of the Clostridia displayed the most antibiotic sensitive phenotypes.

Censusing marine eukaryotic diversity in the twenty-first century

PubMed Central

Knowlton, Nancy

2016-01-01

The ocean constitutes one of the vastest and richest biomes on our planet. Most recent estimations, all based on indirect approaches, suggest that there are millions of marine eukaryotic species. Moreover, a large majority of these are small (less than 1 mm), cryptic and still unknown to science. However, this knowledge gap, caused by the lack of diagnostic morphological features in small organisms and the limited sampling of the global ocean, is currently being filled, thanks to new DNA-based approaches. The molecular technique of PCR amplification of homologous gene regions combined with high-throughput sequencing, routinely used to census unculturable prokaryotes, is now also being used to characterize whole communities of marine eukaryotes. Here, we review how this methodological advancement has helped to better quantify the magnitude and patterns of marine eukaryotic diversity, with an emphasis on taxonomic groups previously largely overlooked. We then discuss obstacles remaining to achieve a global understanding of marine eukaryotic diversity. In particular, we argue that 18S variable regions do not provide sufficient taxonomic resolution to census marine life, and suggest combining broad eukaryotic surveys targeting the 18S rRNA region with more taxon-focused analyses of hypervariable regions to improve our understanding of the diversity of species, the functional units of marine ecosystems. This article is part of the themed issue ‘From DNA barcodes to biomes’. PMID:27481783

Rumen microbial diversity in Svalbard reindeer, with particular emphasis on methanogenic archaea.

PubMed

Sundset, Monica A; Edwards, Joan E; Cheng, Yan Fen; Senosiain, Roberto S; Fraile, Maria N; Northwood, Korinne S; Praesteng, Kirsti E; Glad, Trine; Mathiesen, Svein D; Wright, André-Denis G

2009-12-01

Ruminal methanogens, bacteria and ciliate protozoa of Svalbard reindeer grazing natural pastures in October (late fall) and April (late winter) were investigated using molecular-based approaches. The appetite of the Svalbard reindeer peaks in August (summer) and is at its lowest in March (winter). Microbial numbers, quantified by real-time PCR, did not change significantly between October and April, when food intakes are at similar levels, although the numbers of methanogens tended to be higher in October (P=0.074), and ciliate numbers tended to be higher in April (P=0.055). Similarly, no change was detected in the bacterial and protozoal population composition by rRNA gene-based denaturing gradient gel electrophoresis analysis. Dominant methanogens were identified using a 16S rRNA gene library (97 clones) prepared from pooled PCR products from reindeer on October pasture (n=5). Eleven of the 22 distinct operational taxonomic units (OTUs) generated exhibited a high degree of sequence similarity to methanogens affiliated with Methanobacteriales (eight OTUs), Methanomicrobiales (one OTU) and Methanosarcinales (two OTUs). The remaining 11 OTUs (53% of the clones) were associated with a cluster of uncultivated ruminal archaea. This study has provided important insights into the rumen microbiome of a high-arctic herbivorous animal living under harsh nutritional conditions, and evidence suggesting that host type affects the population size of ruminal methanogens.

Methylobacterium phyllosphaerae sp. nov., a pink-pigmented, facultative methylotroph from the phyllosphere of rice.

PubMed

Madhaiyan, Munusamy; Poonguzhali, Selvaraj; Kwon, Soon-Wo; Sa, Tong-Min

2009-01-01

A pink-pigmented, aerobic, facultatively methylotrophic bacterial strain, CBMB27T, isolated from leaf tissues of rice (Oryza sativa L. 'Dong-Jin'), was analysed using a polyphasic taxonomic approach. Comparative 16S rRNA gene sequence-based phylogenetic analysis placed the strain in a clade with the species Methylobacterium oryzae, Methylobacterium fujisawaense and Methylobacterium mesophilicum; strain CBMB27T showed sequence similarities of 98.3, 98.5 and 97.3 %, respectively, to the type strains of these three species. DNA-DNA hybridization experiments revealed low levels (<38 %) of DNA-DNA relatedness between strain CBMB27T and its closest relatives. The sequence of the 1-aminocyclopropane-1-carboxylate deaminase gene (acdS) in strain CBMB27T differed from those of close relatives. The major fatty acid of the isolate was C(18 : 1)omega7c and the G+C content of the genomic DNA was 66.8 mol%. Based on the results of 16S rRNA gene sequence analysis, DNA-DNA hybridization, and physiological and biochemical characterization, which enabled the isolate to be differentiated from all recognized species of the genus Methylobacterium, it was concluded that strain CBMB27T represents a novel species in the genus Methylobacterium for which the name Methylobacterium phyllosphaerae sp. nov. is proposed (type strain CBMB27T =LMG 24361T =KACC 11716T =DSM 19779T).

Feasibility of nuclear ribosomal region ITS1 over ITS2 in barcoding taxonomically challenging genera of subtribe Cassiinae (Fabaceae).

PubMed

Mishra, Priyanka; Kumar, Amit; Rodrigues, Vereena; Shukla, Ashutosh K; Sundaresan, Velusamy

2016-01-01

The internal transcribed spacer (ITS) region is situated between 18S and 26S in a polycistronic rRNA precursor transcript. It had been proved to be the most commonly sequenced region across plant species to resolve phylogenetic relationships ranging from shallow to deep taxonomic levels. Despite several taxonomical revisions in Cassiinae, a stable phylogeny remains elusive at the molecular level, particularly concerning the delineation of species in the genera Cassia, Senna and Chamaecrista . This study addresses the comparative potential of ITS datasets (ITS1, ITS2 and concatenated) in resolving the underlying morphological disparity in the highly complex genera, to assess their discriminatory power as potential barcode candidates in Cassiinae. A combination of experimental data and an in-silico approach based on threshold genetic distances, sequence similarity based and hierarchical tree-based methods was performed to decipher the discriminating power of ITS datasets on 18 different species of Cassiinae complex. Lab-generated s equences were compared against those available in the GenBank using BLAST and were aligned through MUSCLE 3.8.31 and analysed in PAUP 4.0 and BEAST1.8 using parsimony ratchet, maximum likelihood and Bayesian inference (BI) methods of gene and species tree reconciliation with bootstrapping. DNA barcoding gap was realized based on the Kimura two-parameter distance model (K2P) in TaxonDNA and MEGA. Based on the K2P distance, significant divergences between the inter- and intra-specific genetic distances were observed, while the presence of a DNA barcoding gap was obvious. The ITS1 region efficiently identified 81.63% and 90% of species using TaxonDNA and BI methods, respectively. The PWG-distance method based on simple pairwise matching indicated the significance of ITS1 whereby highest number of variable (210) and informative sites (206) were obtained. The BI tree-based methods outperformed the similarity-based methods producing well-resolved phylogenetic trees with many nodes well supported by bootstrap analyses. The reticulated phylogenetic hypothesis using the ITS1 region mainly supported the relationship between the species of Cassiinae established by traditional morphological methods. The ITS1 region showed a higher discrimination power and desirable characteristics as compared to ITS2 and ITS1 + 2, thereby concluding to be the locus of choice. Considering the complexity of the group and the underlying biological ambiguities, the results presented here are encouraging for developing DNA barcoding as a useful tool for resolving taxonomical challenges in corroboration with morphological framework.

Allopatric Speciation within a Cryptic Species Complex of Australasian Octopuses

PubMed Central

Amor, Michael D.; Norman, Mark D.; Cameron, Hayley E.; Strugnell, Jan M.

2014-01-01

Despite extensive revisions over recent decades, the taxonomy of benthic octopuses (Family Octopodidae) remains in a considerable flux. Among groups of unresolved status is a species complex of morphologically similar shallow-water octopods from subtropical Australasia, including: Allopatric populations of Octopus tetricus on the eastern and western coasts of Australia, of which the Western Australian form is speculated to be a distinct or sub-species; and Octopus gibbsi from New Zealand, a proposed synonym of Australian forms. This study employed a combination of molecular and morphological techniques to resolve the taxonomic status of the ‘tetricus complex’. Phylogenetic analyses (based on five mitochondrial genes: 12S rRNA, 16S rRNA, COI, COIII and Cytb) and Generalised Mixed Yule Coalescent (GMYC) analysis (based on COI, COIII and Cytb) distinguished eastern and Western Australian O. tetricus as distinct species, while O. gibbsi was found to be synonymous with the east Australian form (BS = >97, PP = 1; GMYC p = 0.01). Discrete morphological differences in mature male octopuses (based on sixteen morphological traits) provided further evidence of cryptic speciation between east (including New Zealand) and west coast populations; although females proved less useful in morphological distinction among members of the tetricus complex. In addition, phylogenetic analyses suggested populations of octopuses currently treated under the name Octopus vulgaris are paraphyletic; providing evidence of cryptic speciation among global populations of O. vulgaris, the most commercially valuable octopus species worldwide. PMID:24964133

Rarimicrobium hominis gen. nov., sp. nov., representing the fifth genus in the phylum Synergistetes that includes human clinical isolates.

PubMed

Jumas-Bilak, Estelle; Bouvet, Philippe; Allen-Vercoe, Emma; Aujoulat, Fabien; Lawson, Paul A; Jean-Pierre, Hélène; Marchandin, Hélène

2015-11-01

Five human clinical isolates of an unknown, strictly anaerobic, slow-growing, Gram-stain-negative, rod-shaped micro-organism were subjected to a polyphasic taxonomic study. Comparative 16S rRNA gene sequence-based phylogeny showed that the isolates grouped in a clade that included members of the genera Pyramidobacter, Jonquetella, and Dethiosulfovibrio; the type strain of Pyramidobacter piscolens was the closest relative with 91.5-91.7 % 16S rRNA gene sequence similarity. The novel strains were mainly asaccharolytic and unreactive in most conventional biochemical tests. Major metabolic end products in trypticase/glucose/yeast extract broth were acetic acid and propionic acid and the major cellular fatty acids were C13 : 0 and C16 : 0, each of which could be used to differentiate the strains from P. piscolens. The DNA G+C content based on whole genome sequencing for the reference strain 22-5-S 12D6FAA was 57 mol%. Based on these data, a new genus, Rarimicrobium gen. nov., is proposed with one novel species, Rarimicrobium hominis sp. nov., named after the exclusive and rare finding of the taxon in human samples. Rarimicrobium is the fifth genus of the 14 currently characterized in the phylum Synergistetes and the third one in subdivision B that includes human isolates. The type strain of Rarimicrobium hominis is ADV70T ( = LMG 28163T = CCUG 65426T).

Phylogenetic Analysis of Shewanella Strains by DNA Relatedness Derived from Whole Genome Microarray DNA-DNA Hybridization and Comparison with Other Methods

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wu, Liyou; Yi, T. Y.; Van Nostrand, Joy

Phylogenetic analyses were done for the Shewanella strains isolated from Baltic Sea (38 strains), US DOE Hanford Uranium bioremediation site [Hanford Reach of the Columbia River (HRCR), 11 strains], Pacific Ocean and Hawaiian sediments (8 strains), and strains from other resources (16 strains) with three out group strains, Rhodopseudomonas palustris, Clostridium cellulolyticum, and Thermoanaerobacter ethanolicus X514, using DNA relatedness derived from WCGA-based DNA-DNA hybridizations, sequence similarities of 16S rRNA gene and gyrB gene, and sequence similarities of 6 loci of Shewanella genome selected from a shared gene list of the Shewanella strains with whole genome sequenced based on the averagemore » nucleotide identity of them (ANI). The phylogenetic trees based on 16S rRNA and gyrB gene sequences, and DNA relatedness derived from WCGA hybridizations of the tested Shewanella strains share exactly the same sub-clusters with very few exceptions, in which the strains were basically grouped by species. However, the phylogenetic analysis based on DNA relatedness derived from WCGA hybridizations dramatically increased the differentiation resolution at species and strains level within Shewanella genus. When the tree based on DNA relatedness derived from WCGA hybridizations was compared to the tree based on the combined sequences of the selected functional genes (6 loci), we found that the resolutions of both methods are similar, but the clustering of the tree based on DNA relatedness derived from WMGA hybridizations was clearer. These results indicate that WCGA-based DNA-DNA hybridization is an idea alternative of conventional DNA-DNA hybridization methods and it is superior to the phylogenetics methods based on sequence similarities of single genes. Detailed analysis is being performed for the re-classification of the strains examined.« less

High taxonomic diversity of cultivation-recalcitrant endophytic bacteria in grapevine field shoots, their in vitro introduction, and unsuspected persistence.

PubMed

Thomas, Pious; Sekhar, Aparna C; Shaik, Sadiq Pasha

2017-11-01

Molecular and microscopic analyses reveal enormous non-cultivable endophytic bacteria in grapevine field shoots with functional significance. Diverse bacteria enter tissue cultures through surface-sterilized tissues and survive surreptitiously with varying taxonomic realignments. The study was envisaged to assess the extent of endophytic bacterial association with field shoot tissues of grapevine and the likelihood of introduction of such internally colonizing bacteria in vitro adopting molecular techniques targeting the non-cultivable bacterial community. PowerFood ® -kit derived DNA from surface-sterilized field shoot tips of grapevine Flame Seedless was employed in a preliminary bacterial class-specific PCR screening proving positive for major prokaryotic taxa including Archaea. Taxonomic and functional diversity were analyzed through whole metagenome profiling (WMG) which revealed predominantly phylum Actinobacteria, Proteobacteria, and minor shares of Firmicutes, Bacteroidetes, and Deinococcus-Thermus with varying functional roles ascribable to the whole bacterial community. Field shoot tip tissues and callus derived from stem segments were further employed in 16S rRNA V3-V4 amplicon taxonomic profiling. This revealed elevated taxonomic diversity in field shoots over WMG, predominantly Proteobacteria succeeded by Actinobacteria, Firmicutes, Bacteroidetes, and 15 other phyla including several candidate phyla (135 families, 179 genera). Callus stocks also displayed broad bacterial diversity (16 phyla; 96 families; 141 genera) bearing resemblance to field tissues with Proteobacterial dominance but a reduction in its share, enrichment of Actinobacteria and Firmicutes, disappearance of some field-associated phyla and detection of a few additional taxonomic groups over field community. Similar results were documented during 16S V3-V4 amplicon taxonomic profiling on Thompson Seedless field shoot tip and callus tissues. Video microscopy on tissue homogenates corroborated enormous endophytic bacteria. This study elucidates a vast diversity of cultivation-recalcitrant endophytic bacteria prevailing in grapevine field shoots, their in vitro introduction, and unsuspecting sustenance with possible silent participation in tissue culture processes.

Oligonucleotide microarray for the identification of potential mycotoxigenic fungi

PubMed Central

2010-01-01

Background Mycotoxins are secondary metabolites which are produced by numerous fungi and pose a continuous challenge to the safety and quality of food commodities in South Africa. These toxins have toxicologically relevant effects on humans and animals that eat contaminated foods. In this study, a diagnostic DNA microarray was developed for the identification of the most common food-borne fungi, as well as the genes leading to toxin production. Results A total of 40 potentially mycotoxigenic fungi isolated from different food commodities, as well as the genes that are involved in the mycotoxin synthetic pathways, were analyzed. For fungal identification, oligonucleotide probes were designed by exploiting the sequence variations of the elongation factor 1-alpha (EF-1 α) coding regions and the internal transcribed spacer (ITS) regions of the rRNA gene cassette. For the detection of fungi able to produce mycotoxins, oligonucleotide probes directed towards genes leading to toxin production from different fungal strains were identified in data available in the public domain. The probes selected for fungal identification and the probes specific for toxin producing genes were spotted onto microarray slides. Conclusions The diagnostic microarray developed can be used to identify single pure strains or cultures of potentially mycotoxigenic fungi as well as genes leading to toxin production in both laboratory samples and maize-derived foods offering an interesting potential for microbiological laboratories. PMID:20307326

Design and verification of a pangenome microarray oligonucleotide probe set for Dehalococcoides spp.

PubMed

Hug, Laura A; Salehi, Maryam; Nuin, Paulo; Tillier, Elisabeth R; Edwards, Elizabeth A

2011-08-01

Dehalococcoides spp. are an industrially relevant group of Chloroflexi bacteria capable of reductively dechlorinating contaminants in groundwater environments. Existing Dehalococcoides genomes revealed a high level of sequence identity within this group, including 98 to 100% 16S rRNA sequence identity between strains with diverse substrate specificities. Common molecular techniques for identification of microbial populations are often not applicable for distinguishing Dehalococcoides strains. Here we describe an oligonucleotide microarray probe set designed based on clustered Dehalococcoides genes from five different sources (strain DET195, CBDB1, BAV1, and VS genomes and the KB-1 metagenome). This "pangenome" probe set provides coverage of core Dehalococcoides genes as well as strain-specific genes while optimizing the potential for hybridization to closely related, previously unknown Dehalococcoides strains. The pangenome probe set was compared to probe sets designed independently for each of the five Dehalococcoides strains. The pangenome probe set demonstrated better predictability and higher detection of Dehalococcoides genes than strain-specific probe sets on nontarget strains with <99% average nucleotide identity. An in silico analysis of the expected probe hybridization against the recently released Dehalococcoides strain GT genome and additional KB-1 metagenome sequence data indicated that the pangenome probe set performs more robustly than the combined strain-specific probe sets in the detection of genes not included in the original design. The pangenome probe set represents a highly specific, universal tool for the detection and characterization of Dehalococcoides from contaminated sites. It has the potential to become a common platform for Dehalococcoides-focused research, allowing meaningful comparisons between microarray experiments regardless of the strain examined.

[Phylogenetic diversity of microorganisms associated with the deep-water sponge Baikalospongia intermedia].

PubMed

Kalyzhnaya, O V; Itskovich, V B

2014-07-01

The diversity of bacteria associated with deep-water sponge Baikalospongia intermedia was evaluated by sequence analysis of 16S rRNA genes from two sponge samples collected in Lake Baikal from depths of 550 and 1204 m. A total of 64 operational taxonomic units, belonging to nine bacterial phyla, Proteobacteria (classes Alphaproteobacteria,. Betaproteobacteria, Gammaproteobacteria, and Deltaproteobacteria), Actinobacteria, Planctomycetes, Cloroflexi, Verrucomicrobia, Acidobacteria, Chlorobi, and Nitrospirae, including candidate phylum WS5, were identified. Phylogenetic analysis showed that the examined communities contained phylotypes exhibiting homology to uncultured bacteria from different lake ecosystems, freshwater sediments, soil and geological formations. Moreover, a number of phylotypes were relative to psychrophilic, methane-oxidizing, sulfate-reducing bacteria, and to microorganisms resistant to the influence of heavy metals. It seems likely that the unusual habitation conditions of deep-water sponges contribute to the taxonomic diversity of associated bacteria and have an influence on the presence of functionally important microorganisms in bacterial communities.

Two distinct extracellular RNA signatures released by a single cell type identified by microarray and next-generation sequencing

PubMed Central

Lässer, Cecilia; Shelke, Ganesh Vilas; Yeri, Ashish; Kim, Dae-Kyum; Crescitelli, Rossella; Raimondo, Stefania; Sjöstrand, Margareta; Gho, Yong Song; Van Keuren Jensen, Kendall; Lötvall, Jan

2017-01-01

ABSTRACT Cells secrete extracellular RNA (exRNA) to their surrounding environment and exRNA has been found in many body fluids such as blood, breast milk and cerebrospinal fluid. However, there are conflicting results regarding the nature of exRNA. Here, we have separated 2 distinct exRNA profiles released by mast cells, here termed high-density (HD) and low-density (LD) exRNA. The exRNA in both fractions was characterized by microarray and next-generation sequencing. Both exRNA fractions contained mRNA and miRNA, and the mRNAs in the LD exRNA correlated closely with the cellular mRNA, whereas the HD mRNA did not. Furthermore, the HD exRNA was enriched in lincRNA, antisense RNA, vault RNA, snoRNA, and snRNA with little or no evidence of full-length 18S and 28S rRNA. The LD exRNA was enriched in mitochondrial rRNA, mitochondrial tRNA, tRNA, piRNA, Y RNA, and full-length 18S and 28S rRNA. The proteomes of the HD and LD exRNA-containing fractions were determined with LC-MS/MS and analyzed with Gene Ontology term finder, which showed that both proteomes were associated with the term extracellular vesicles and electron microscopy suggests that at least a part of the exRNA is associated with exosome-like extracellular vesicles. Additionally, the proteins in the HD fractions tended to be associated with the nucleus and ribosomes, whereas the LD fraction proteome tended to be associated with the mitochondrion. We show that the 2 exRNA signatures released by a single cell type can be separated by floatation on a density gradient. These results show that cells can release multiple types of exRNA with substantial differences in RNA species content. This is important for any future studies determining the nature and function of exRNA released from different cells under different conditions. PMID:27791479

Description of Mycobacterium chelonae subsp. bovis subsp. nov., isolated from cattle (Bos taurus coreanae), emended description of Mycobacterium chelonae and creation of Mycobacterium chelonae subsp. chelonae subsp. nov.

PubMed

Kim, Byoung-Jun; Kim, Ga-Na; Kim, Bo-Ram; Jeon, Che Ok; Jeong, Joseph; Lee, Seon Ho; Lim, Ji-Hun; Lee, Seung-Heon; Kim, Chang Ki; Kook, Yoon-Hoh; Kim, Bum-Joon

2017-10-01

Three rapidly growing mycobacterial strains, QIA-37 T , QIA-40 and QIA-41, were isolated from the lymph nodes of three separate Korean native cattle, Hanwoo (Bos taurus coreanae). These strains were previously shown to be phylogenetically distinct but closely related to Mycobacterium chelonae ATCC 35752 T by taxonomic approaches targeting three genes (16S rRNA, hsp6 and rpoB) and were further characterized using a polyphasic approach in this study. The 16S rRNA gene sequences of all three strains showed 99.7 % sequence similarity with that of the M. chelonae type strain. A multilocus sequence typing analysis targeting 10 housekeeping genes, including hsp65 and rpoB, revealed a phylogenetic cluster of these strains with M. chelonae. DNA-DNA hybridization values of 78.2 % between QIA-37 T and M. chelonae indicated that it belongs to M. chelonae but is a novel subspecies distinct from M. chelonae. Phylogenetic analysis based on whole-genome sequences revealed a 95.44±0.06 % average nucleotide identity (ANI) value with M. chelonae, slightly higher than the 95.0 % ANI criterion for determining a novel species. In addition, distinct phenotypic characteristics such as positive growth at 37 °C, at which temperature M. chelonae does not grow, further support the taxonomic status of these strains as representatives of a novel subspecies of M. chelonae. Therefore, we propose an emended description of Mycobacterium chelonae, and descriptions of M. chelonae subsp. chelonae subsp. nov. and M. chelonae subsp. bovis subsp. nov. are presented; strains ATCC 35752 T (=CCUG 47445 T =CIP 104535 T =DSM 43804 T =JCM 6388 T =NCTC 946 T ) and QIA-37 T (=KCTC 39630 T =JCM 30986 T ) are the type strains of the two novel subspecies.

Multilocus Phylogeny of the Afrotropical Freshwater Crab Fauna Reveals Historical Drainage Connectivity and Transoceanic Dispersal Since the Eocene.

PubMed

Daniels, Savel R; Phiri, Ethel E; Klaus, Sebastian; Albrecht, Christian; Cumberlidge, Neil

2015-07-01

Phylogenetic reconstruction, divergence time estimations and ancestral range estimation were undertaken for 66% of the Afrotropical freshwater crab fauna (Potamonautidae) based on four partial DNA loci (12S rRNA, 16S rRNA, cytochrome oxidase one [COI], and histone 3). The present study represents the most comprehensive taxonomic sampling of any freshwater crab family globally, and explores the impact of paleodrainage interconnectivity on cladogenesis among freshwater crabs. Phylogenetic analyses of the total evidence data using maximum-likelihood (ML), maximum parsimony (MP), and Bayesian inference (BI) produced a robust statistically well-supported tree topology that reaffirmed the monophyly of the Afrotropical freshwater crab fauna. The estimated divergence times suggest that the Afrotropical Potamonautidae diverged during the Eocene. Cladogenesis within and among several genera occurred predominantly during the Miocene, which was associated with major tectonic and climatic ameliorations throughout the region. Paleodrainage connectivity was observed with specimens from the Nilo-Sudan and East African coast proving to be sister to specimens from the Upper Guinea Forests in West Africa. In addition, we observed strong sister taxon affinity between specimens from East Africa and the Congo basin, including specimens from Lake Tanganyika, while the southern African fauna was retrieved as sister to the Angolan taxa. Within the East African clade we observed two independent transoceanic dispersal events, one to the Seychelles Archipelago and a second to Madagascar, while we observe a single transoceanic dispersal event from West Africa to São Tomé. The ancestral area estimation suggested a West African/East African ancestral range for the family with multiple dispersal events between southern Africa and East Africa, and between East Africa and Central Africa The taxonomic implications of our results are discussed in light of the widespread paraphyly evident among a number of genera. © The Author(s) 2015. Published by Oxford University Press, on behalf of the Society of Systematic Biologists. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Paenibacillus oenotherae sp. nov. and Paenibacillus hemerocallicola sp. nov., isolated from the roots of herbaceous plants.

PubMed

Kim, Tae-Su; Han, Ji-Hye; Joung, Yochan; Kim, Seung Bum

2015-08-01

Two Gram-staining-positive, aerobic, endospore-forming, motile bacteria, strains DT7-4T and DLE-12T, were isolated from roots of evening primrose (Oenothera biennis) and day lily (Hemerocallis fulva), respectively, and subjected to taxonomic characterization. Analysis of 16S rRNA gene sequences indicated that the two strains fell into two distinct phylogenetic clusters belonging to the genus Paenibacillus. Strain DT7-4T was most closely related to Paenibacillus phyllosphaerae PALXIL04T and Paenibacillus taihuensis THMBG22T, with 96.3% 16S rRNA gene sequence similarity to each, and strain DLE-12T was most closely related to Paenibacillus ginsengarvi Gsoil 139T and Paenibacillus hodogayensis SGT, with 96.6 and 93.3% sequence similarity, respectively. Both isolates contained anteiso-C15 : 0 as the dominant fatty acid, meso-diaminopimelic acid as the diagnostic diamino acid in the cell-wall peptidoglycan and MK-7 as the respiratory menaquinone. The cellular polar lipids were composed of diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol and unidentified polar lipids. The DNA G+C contents of strains DT7-4T and DLE-12T were 50.1 ± 0.7 and 55.2 ± 0.5 mol%, respectively. The chemotaxonomic properties of both isolates were typical of members of the genus Paenibacillus. However, our biochemical and phylogenetic analyses distinguished each isolate from related species. Based on our polyphasic taxonomic analysis, strains DT7-4T and DLE-12T should be recognized as representatives of novel species of Paenibacillus, for which the names Paenibacillus oenotherae sp. nov. (type strain DT7-4T = KCTC 33186T = JCM 19573T) and Paenibacillus hemerocallicola sp. nov. (type strain DLE-12T = KCTC 33185T = JCM 19572T) are proposed.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Walters, William; Hyde, Embriette R.; Berg-Lyons, Donna

ABSTRACT Designing primers for PCR-based taxonomic surveys that amplify a broad range of phylotypes in varied community samples is a difficult challenge, and the comparability of data sets amplified with varied primers requires attention. Here, we examined the performance of modified 16S rRNA gene and internal transcribed spacer (ITS) primers for archaea/bacteria and fungi, respectively, with nonaquatic samples. We moved primer bar codes to the 5′ end, allowing for a range of different 3′ primer pairings, such as the 515f/926r primer pair, which amplifies variable regions 4 and 5 of the 16S rRNA gene. We additionally demonstrated that modifications tomore » the 515f/806r (variable region 4) 16S primer pair, which improves detection ofThaumarchaeotaand clade SAR11 in marine samples, do not degrade performance on taxa already amplified effectively by the original primer set. Alterations to the fungal ITS primers did result in differential but overall improved performance compared to the original primers. In both cases, the improved primers should be widely adopted for amplicon studies. ImportanceWe continue to uncover a wealth of information connecting microbes in important ways to human and environmental ecology. As our scientific knowledge and technical abilities improve, the tools used for microbiome surveys can be modified to improve the accuracy of our techniques, ensuring that we can continue to identify groundbreaking connections between microbes and the ecosystems they populate, from ice caps to the human body. It is important to confirm that modifications to these tools do not cause new, detrimental biases that would inhibit the field rather than continue to move it forward. We therefore demonstrated that two recently modified primer pairs that target taxonomically discriminatory regions of bacterial and fungal genomic DNA do not introduce new biases when used on a variety of sample types, from soil to human skin. This confirms the utility of these primers for maintaining currently recommended microbiome research techniques as the state of the art.« less

No need to replace an “anomalous” primate (Primates) with an “anomalous” bear (Carnivora, Ursidae)

PubMed Central

Gutiérrez, Eliécer E.; Pine, Ronald H.

2015-01-01

Abstract By means of mitochondrial 12S rRNA sequencing of putative “yeti”, “bigfoot”, and other “anomalous primate” hair samples, a recent study concluded that two samples, presented as from the Himalayas, do not belong to an “anomalous primate”, but to an unknown, anomalous type of ursid. That is, that they match 12S rRNA sequences of a fossil Polar Bear (Ursus maritimus), but neither of modern Polar Bears, nor of Brown Bears (Ursus arctos), the closest relative of Polar Bears, and one that occurs today in the Himalayas. We have undertaken direct comparison of sequences; replication of the original comparative study; inference of phylogenetic relationships of the two samples with respect to those from all extant species of Ursidae (except for the Giant Panda, Ailuropoda melanoleuca) and two extinct Pleistocene species; and application of a non-tree-based population aggregation approach for species diagnosis and identification. Our results demonstrate that the very short fragment of the 12S rRNA gene sequenced by Sykes et al. is not sufficiently informative to support the hypotheses provided by these authors with respect to the taxonomic identity of the individuals from which these sequences were obtained. We have concluded that there is no reason to believe that the two samples came from anything other than Brown Bears. These analyses afforded an opportunity to test the monophyly of morphologically defined species and to comment on both their phylogenetic relationships and future efforts necessary to advance our understanding of ursid systematics. PMID:25829853

Bacillus horneckiae sp. nov., isolated from a spacecraft-assembly clean room.

PubMed

Vaishampayan, Parag; Probst, Alexander; Krishnamurthi, Srinivasan; Ghosh, Sudeshna; Osman, Shariff; McDowall, Alasdair; Ruckmani, Arunachalam; Mayilraj, Shanmugam; Venkateswaran, Kasthuri

2010-05-01

Five Gram-stain-positive, motile, aerobic strains were isolated from a clean room of the Kennedy Space Center where the Phoenix spacecraft was assembled. All strains are rod-shaped, spore-forming bacteria, whose spores were resistant to UV radiation up to 1000 J m(-2). The spores were subterminally positioned and produced an external layer. A polyphasic taxonomic study including traditional biochemical tests, fatty acid analysis, cell-wall typing, lipid analyses, 16S rRNA gene sequencing and DNA-DNA hybridization studies was performed to characterize these novel strains. 16S rRNA gene sequencing and lipid analyses convincingly grouped these novel strains within the genus Bacillus as a cluster separate from already described species. The similarity of 16S rRNA gene sequences among the novel strains was >99 %, but the similarity was only about 97 % with their nearest neighbours Bacillus pocheonensis, Bacillus firmus and Bacillus bataviensis. DNA-DNA hybridization dissociation values were <24 % to the closest related type strains. The novel strains had a G+C content 35.6+/-0.5 mol% and could liquefy gelatin but did not utilize or produce acids from any of the carbon substrates tested. The major fatty acids were iso-C(15 : 0) and anteiso-C(15 : 0) and the cell-wall diamino acid was meso-diaminopimelic acid. Based on phylogenetic and phenotypic results, it is concluded that these strains represent a novel species of the genus Bacillus, for which the name Bacillus horneckiae sp. nov. is proposed. The type strain is 1P01SC(T) (=NRRL B-59162(T) =MTCC 9535(T)).

Natural revegetation of a semiarid habitat alters taxonomic and functional diversity of soil microbial communities.

PubMed

Guo, Yanqing; Chen, Xiaotian; Wu, Yuanyuan; Zhang, Lu; Cheng, Jimin; Wei, Gehong; Lin, Yanbing

2018-04-18

Revegetation of degraded lands has a profound impact on the maintenance and stability of ecosystem processes. However, the impacts of this land use change on functional diversity of soil microbial communities are poorly understood. Here, using 16S rRNA gene amplicon and shotgun metagenomic sequencing, we compared the taxonomic and functional communities of soil microbiome, and analyzed the effects of plant diversity and soil chemical properties, in a chronosequence of restored ex-farmland that had been naturally revegetated to grassland over periods of 5, 15 and 30years with adjacent farmland, on the Loess Plateau, China. We found that microbial taxonomic diversity was positively correlated with plant diversity and was higher in the revegetated sites. Functional diversity increased significantly in the oldest grassland. Actinobacteria, commonly considered a copiotrophic phylum, was more abundant in the revegetated sites, while Acidobacteria, an oligotrophic phylum, was more abundant in farmland. Furthermore, the structure of taxonomic and functional communities was significantly different between revegetated sites and farmland, and organic matter was the best environmental predictor in determining these microbial communities. Compared with the farmland, revegetation increased the proportion of genes associated with energy metabolism, carbohydrate metabolism and xenobiotics biodegradation and metabolism. Notably, the higher proportion of carbohydrate degradation gene subfamilies in the revegetated sites indicated higher levels of soil nutrient cycling. These results elucidate the significant shifts in belowground microbial taxonomic and functional diversity following vegetation restoration and have implications for ecological restoration programs in arid and semi-arid ecosystems. Copyright © 2018. Published by Elsevier B.V.

Metagenome, metatranscriptome, and metaproteome approaches unraveled compositions and functional relationships of microbial communities residing in biogas plants.

PubMed

Hassa, Julia; Maus, Irena; Off, Sandra; Pühler, Alfred; Scherer, Paul; Klocke, Michael; Schlüter, Andreas

2018-06-01

The production of biogas by anaerobic digestion (AD) of agricultural residues, organic wastes, animal excrements, municipal sludge, and energy crops has a firm place in sustainable energy production and bio-economy strategies. Focusing on the microbial community involved in biomass conversion offers the opportunity to control and engineer the biogas process with the objective to optimize its efficiency. Taxonomic profiling of biogas producing communities by means of high-throughput 16S rRNA gene amplicon sequencing provided high-resolution insights into bacterial and archaeal structures of AD assemblages and their linkages to fed substrates and process parameters. Commonly, the bacterial phyla Firmicutes and Bacteroidetes appeared to dominate biogas communities in varying abundances depending on the apparent process conditions. Regarding the community of methanogenic Archaea, their diversity was mainly affected by the nature and composition of the substrates, availability of nutrients and ammonium/ammonia contents, but not by the temperature. It also appeared that a high proportion of 16S rRNA sequences can only be classified on higher taxonomic ranks indicating that many community members and their participation in AD within functional networks are still unknown. Although cultivation-based approaches to isolate microorganisms from biogas fermentation samples yielded hundreds of novel species and strains, this approach intrinsically is limited to the cultivable fraction of the community. To obtain genome sequence information of non-cultivable biogas community members, metagenome sequencing including assembly and binning strategies was highly valuable. Corresponding research has led to the compilation of hundreds of metagenome-assembled genomes (MAGs) frequently representing novel taxa whose metabolism and lifestyle could be reconstructed based on nucleotide sequence information. In contrast to metagenome analyses revealing the genetic potential of microbial communities, metatranscriptome sequencing provided insights into the metabolically active community. Taking advantage of genome sequence information, transcriptional activities were evaluated considering the microorganism's genetic background. Metaproteome studies uncovered enzyme profiles expressed by biogas community members. Enzymes involved in cellulose and hemicellulose decomposition and utilization of other complex biopolymers were identified. Future studies on biogas functional microbial networks will increasingly involve integrated multi-omics analyses evaluating metagenome, transcriptome, proteome, and metabolome datasets.

Diversity and community composition of methanogenic archaea in the rumen of Scottish upland sheep assessed by different methods.

PubMed

Snelling, Timothy J; Genç, Buğra; McKain, Nest; Watson, Mick; Waters, Sinéad M; Creevey, Christopher J; Wallace, R John

2014-01-01

Ruminal archaeomes of two mature sheep grazing in the Scottish uplands were analysed by different sequencing and analysis methods in order to compare the apparent archaeal communities. All methods revealed that the majority of methanogens belonged to the Methanobacteriales order containing the Methanobrevibacter, Methanosphaera and Methanobacteria genera. Sanger sequenced 1.3 kb 16S rRNA gene amplicons identified the main species of Methanobrevibacter present to be a SGMT Clade member Mbb. millerae (≥ 91% of OTUs); Methanosphaera comprised the remainder of the OTUs. The primers did not amplify ruminal Thermoplasmatales-related 16S rRNA genes. Illumina sequenced V6-V8 16S rRNA gene amplicons identified similar Methanobrevibacter spp. and Methanosphaera clades and also identified the Thermoplasmatales-related order as 13% of total archaea. Unusually, both methods concluded that Mbb. ruminantium and relatives from the same clade (RO) were almost absent. Sequences mapping to rumen 16S rRNA and mcrA gene references were extracted from Illumina metagenome data. Mapping of the metagenome data to 16S rRNA gene references produced taxonomic identification to Order level including 2-3% Thermoplasmatales, but was unable to discriminate to species level. Mapping of the metagenome data to mcrA gene references resolved 69% to unclassified Methanobacteriales. Only 30% of sequences were assigned to species level clades: of the sequences assigned to Methanobrevibacter, most mapped to SGMT (16%) and RO (10%) clades. The Sanger 16S amplicon and Illumina metagenome mcrA analyses showed similar species richness (Chao1 Index 19-35), while Illumina metagenome and amplicon 16S rRNA analysis gave lower richness estimates (10-18). The values of the Shannon Index were low in all methods, indicating low richness and uneven species distribution. Thus, although much information may be extracted from the other methods, Illumina amplicon sequencing of the V6-V8 16S rRNA gene would be the method of choice for studying rumen archaeal communities.

Validation of the Hirst-Type Spore Trap for Simultaneous Monitoring of Prokaryotic and Eukaryotic Biodiversities in Urban Air Samples by Next-Generation Sequencing.

PubMed

Núñez, Andrés; Amo de Paz, Guillermo; Ferencova, Zuzana; Rastrojo, Alberto; Guantes, Raúl; García, Ana M; Alcamí, Antonio; Gutiérrez-Bustillo, A Montserrat; Moreno, Diego A

2017-07-01

Pollen, fungi, and bacteria are the main microscopic biological entities present in outdoor air, causing allergy symptoms and disease transmission and having a significant role in atmosphere dynamics. Despite their relevance, a method for monitoring simultaneously these biological particles in metropolitan environments has not yet been developed. Here, we assessed the use of the Hirst-type spore trap to characterize the global airborne biota by high-throughput DNA sequencing, selecting regions of the 16S rRNA gene and internal transcribed spacer for the taxonomic assignment. We showed that aerobiological communities are well represented by this approach. The operational taxonomic units (OTUs) of two traps working synchronically compiled >87% of the total relative abundance for bacterial diversity collected in each sampler, >89% for fungi, and >97% for pollen. We found a good correspondence between traditional characterization by microscopy and genetic identification, obtaining more-accurate taxonomic assignments and detecting a greater diversity using the latter. We also demonstrated that DNA sequencing accurately detects differences in biodiversity between samples. We concluded that high-throughput DNA sequencing applied to aerobiological samples obtained with Hirst spore traps provides reliable results and can be easily implemented for monitoring prokaryotic and eukaryotic entities present in the air of urban areas. IMPORTANCE Detection, monitoring, and characterization of the wide diversity of biological entities present in the air are difficult tasks that require time and expertise in different disciplines. We have evaluated the use of the Hirst spore trap (an instrument broadly employed in aerobiological studies) to detect and identify these organisms by DNA-based analyses. Our results showed a consistent collection of DNA and a good concordance with traditional methods for identification, suggesting that these devices can be used as a tool for continuous monitoring of the airborne biodiversity, improving taxonomic resolution and characterization together. They are also suitable for acquiring novel DNA amplicon-based information in order to gain a better understanding of the biological particles present in a scarcely known environment such as the air. Copyright © 2017 American Society for Microbiology.

Validation of the Hirst-Type Spore Trap for Simultaneous Monitoring of Prokaryotic and Eukaryotic Biodiversities in Urban Air Samples by Next-Generation Sequencing

PubMed Central

Núñez, Andrés; Amo de Paz, Guillermo; Ferencova, Zuzana; Rastrojo, Alberto; Guantes, Raúl; García, Ana M.; Alcamí, Antonio; Gutiérrez-Bustillo, A. Montserrat

2017-01-01

ABSTRACT Pollen, fungi, and bacteria are the main microscopic biological entities present in outdoor air, causing allergy symptoms and disease transmission and having a significant role in atmosphere dynamics. Despite their relevance, a method for monitoring simultaneously these biological particles in metropolitan environments has not yet been developed. Here, we assessed the use of the Hirst-type spore trap to characterize the global airborne biota by high-throughput DNA sequencing, selecting regions of the 16S rRNA gene and internal transcribed spacer for the taxonomic assignment. We showed that aerobiological communities are well represented by this approach. The operational taxonomic units (OTUs) of two traps working synchronically compiled >87% of the total relative abundance for bacterial diversity collected in each sampler, >89% for fungi, and >97% for pollen. We found a good correspondence between traditional characterization by microscopy and genetic identification, obtaining more-accurate taxonomic assignments and detecting a greater diversity using the latter. We also demonstrated that DNA sequencing accurately detects differences in biodiversity between samples. We concluded that high-throughput DNA sequencing applied to aerobiological samples obtained with Hirst spore traps provides reliable results and can be easily implemented for monitoring prokaryotic and eukaryotic entities present in the air of urban areas. IMPORTANCE Detection, monitoring, and characterization of the wide diversity of biological entities present in the air are difficult tasks that require time and expertise in different disciplines. We have evaluated the use of the Hirst spore trap (an instrument broadly employed in aerobiological studies) to detect and identify these organisms by DNA-based analyses. Our results showed a consistent collection of DNA and a good concordance with traditional methods for identification, suggesting that these devices can be used as a tool for continuous monitoring of the airborne biodiversity, improving taxonomic resolution and characterization together. They are also suitable for acquiring novel DNA amplicon-based information in order to gain a better understanding of the biological particles present in a scarcely known environment such as the air. PMID:28455334

Common and distinguishing features of the bacterial and fungal communities in biological soil crusts and shrub root zone soils

USGS Publications Warehouse

Steven, Blaire; Gallegos-Graves, La Verne; Yeager, Chris; Belnap, Jayne; Kuske, Cheryl R.

2013-01-01

Soil microbial communities in dryland ecosystems play important roles as root associates of the widely spaced plants and as the dominant members of biological soil crusts (biocrusts) colonizing the plant interspaces. We employed rRNA gene sequencing (bacterial 16S/fungal large subunit) and shotgun metagenomic sequencing to compare the microbial communities inhabiting the root zones of the dominant shrub, Larrea tridentata (creosote bush), and the interspace biocrusts in a Mojave desert shrubland within the Nevada Free Air CO2 Enrichment (FACE) experiment. Most of the numerically abundant bacteria and fungi were present in both the biocrusts and root zones, although the proportional abundance of those members differed significantly between habitats. Biocrust bacteria were predominantly Cyanobacteria while root zones harbored significantly more Actinobacteria and Proteobacteria. Pezizomycetes fungi dominated the biocrusts while Dothideomycetes were highest in root zones. Functional gene abundances in metagenome sequence datasets reflected the taxonomic differences noted in the 16S rRNA datasets. For example, functional categories related to photosynthesis, circadian clock proteins, and heterocyst-associated genes were enriched in the biocrusts, where populations of Cyanobacteria were larger. Genes related to potassium metabolism were also more abundant in the biocrusts, suggesting differences in nutrient cycling between biocrusts and root zones. Finally, ten years of elevated atmospheric CO2 did not result in large shifts in taxonomic composition of the bacterial or fungal communities or the functional gene inventories in the shotgun metagenomes.

Genetic characterization of Rhipicephalus sanguineus (sensu lato) ticks from dogs in Portugal.

PubMed

Dantas-Torres, Filipe; Maia, Carla; Latrofa, Maria Stefania; Annoscia, Giada; Cardoso, Luís; Otranto, Domenico

2017-03-13

The taxonomic status of the brown dog tick Rhipicephalus sanguineus (sensu stricto) is a subject of on-going debate; there is a consensus that populations of this tick species should be referred to as R. sanguineus (sensu lato) until its taxonomic status is resolved. Recent genetic studies revealed the existence of more than one lineage of R. sanguineus (s.l.) in temperate countries. In this study, we assessed the genetic identity of ticks collected from rural dogs living in several areas located in all major geographical regions of Portugal. A total of 347 ticks were collected from rural dogs living in different regions of Portugal. These ticks were morphologically identified and partial mitochondrial 16S rRNA gene sequences (~300 bp) were obtained from representative specimens. The ticks were morphologically identified as Ixodes ricinus (seven males and 27 females), Rhipicephalus bursa (one male), Rhipicephalus pusillus (one female) and R. sanguineus (s.l.) (two larvae, 101 nymphs, 108 males and 100 females). Partial mitochondrial 16S rRNA gene sequences were obtained from 58 R. sanguineus (s.l.) specimens, and all of them were genetically identified as belonging to the so-called temperate lineage of R. sanguineus (s.l.) CONCLUSIONS: These results strongly suggest that the temperate species of R. sanguineus (s.l.) is the only representative of this tick group found on dogs in Portugal. It also adds weight to the hypothesis that Rhipicephalus turanicus is not present in this country, although further investigations are necessary to confirm this.

Taxonomic and predicted metabolic profiles of the human gut microbiome in pre-Columbian mummies.

PubMed

Santiago-Rodriguez, Tasha M; Fornaciari, Gino; Luciani, Stefania; Dowd, Scot E; Toranzos, Gary A; Marota, Isolina; Cano, Raul J

2016-11-01

Characterization of naturally mummified human gut remains could potentially provide insights into the preservation and evolution of commensal and pathogenic microorganisms, and metabolic profiles. We characterized the gut microbiome of two pre-Columbian Andean mummies dating to the 10-15th centuries using 16S rRNA gene high-throughput sequencing and metagenomics, and compared them to a previously characterized gut microbiome of an 11th century AD pre-Columbian Andean mummy. Our previous study showed that the Clostridiales represented the majority of the bacterial communities in the mummified gut remains, but that other microbial communities were also preserved during the process of natural mummification, as shown with the metagenomics analyses. The gut microbiome of the other two mummies were mainly comprised by Clostridiales or Bacillales, as demonstrated with 16S rRNA gene amplicon sequencing, many of which are facultative anaerobes, possibly consistent with the process of natural mummification requiring low oxygen levels. Metagenome analyses showed the presence of other microbial groups that were positively or negatively correlated with specific metabolic profiles. The presence of sequences similar to both Trypanosoma cruzi and Leishmania donovani could suggest that these pathogens were prevalent in pre-Columbian individuals. Taxonomic and functional profiling of mummified human gut remains will aid in the understanding of the microbial ecology of the process of natural mummification. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Saccharothrix ghardaiensis sp. nov., an actinobacterium isolated from Saharan soil.

PubMed

Bouznada, Khaoula; Bouras, Noureddine; Mokrane, Salim; Chaabane Chaouch, Fawzia; Zitouni, Abdelghani; Pötter, Gabriele; Spröer, Cathrin; Klenk, Hans-Peter; Sabaou, Nasserdine

2017-03-01

The taxonomic position of a new Saccharothrix strain, designated MB46 T , isolated from a Saharan soil sample collected in Mzab region (Ghardaïa province, South Algeria) was established following a polyphasic approach. The novel microorganism has morphological and chemical characteristics typical of the members of the genus Saccharothrix and formed a phyletic line at the periphery of the Saccharothrix espanaensis subcluster in the 16S rRNA gene dendrograms. Results of the 16S rRNA gene sequence comparisons revealed that strain MB46 T shares high degrees of similarity with S. espanaensis DSM 44229 T (99.2%), Saccharothrix variisporea DSM 43911 T (98.7%) and Saccharothrix texasensis NRRL B-16134 T (98.6%). However, the new strain exhibited only 12.5-17.5% DNA relatedness to the neighbouring Saccharothrix spp. On the basis of phenotypic characteristics, 16S rRNA gene sequence comparisons and DNA-DNA hybridizations, strain MB46 T is concluded to represent a novel species of the genus Saccharothrix, for which the name Saccharothrix ghardaiensis sp. nov. (type strain MB46 T  = DSM 46886 T  = CECT 9046 T ) is proposed.

Composition and Activity of Microbial Communities along the Redox Gradient of an Alkaline, Hypersaline, Lake

PubMed Central

Edwardson, Christian F.; Hollibaugh, James T.

2018-01-01

We compared the composition of microbial communities obtained by sequencing 16S rRNA gene amplicons with taxonomy derived from metatranscriptomes from the same samples. Samples were collected from alkaline, hypersaline Mono Lake, California, USA at five depths that captured the major redox zones of the lake during the onset of meromixis. The prokaryotic community was dominated by bacteria from the phyla Proteobacteria, Firmicutes, and Bacteroidetes, while the picoeukaryotic chlorophyte Picocystis dominated the eukaryotes. Most (80%) of the abundant (>1% relative abundance) OTUs recovered as amplicons of 16S rRNA genes have been reported in previous surveys, indicating that Mono Lake's microbial community has remained stable over 12 years that have included periods of regular, annual overturn interspersed by episodes of prolonged meromixis that result in extremely reducing conditions in bottom water. Metatranscriptomic sequences binned predominately to the Gammaproteobacteria genera Thioalkalivibrio (4–13%) and Thioalkalimicrobium (0–14%); and to the Firmicutes genera Dethiobacter (0–5%) and Clostridium (1–4%), which were also abundant in the 16S rRNA gene amplicon libraries. This study provides insight into the taxonomic affiliations of transcriptionally active communities of the lake's water column under different redox conditions. PMID:29445359

Reclassification of Lactobacillus kimchii and Lactobacillus bobalius as later subjective synonyms of Lactobacillus paralimentarius.

PubMed

Pang, Huili; Kitahara, Maki; Tan, Zhongfang; Wang, Yanping; Qin, Guangyong; Ohkuma, Moriya; Cai, Yimin

2012-10-01

Characterization and identification of strain CW 1 ( = JCM 17161) isolated from corn silage were performed. Strain CW 1 was a Gram-positive, catalase-negative and homofermentative rod that produced the DL-form of lactic acid. This strain exhibited more than 99.6% 16S rRNA gene sequence similarity and greater than 82% DNA-DNA reassociation with type strains of Lactobacillus kimchii, L. bobalius and L. paralimentarius. To clarify the taxonomic positions of these type strains, phenotypic characterization, 16S rRNA gene sequencing, ribotyping and DNA-DNA relatedness were examined. The three type strains displayed different L-arabinose, lactose, melibiose, melezitose, raffinose and N-acetyl-β-glucosaminidase fermentation patterns. Phylogenetic analysis showed that L. paralimentarius is a closer neighbour of L. kimchii and L. bobalius, sharing 99.5-99.9% 16S rRNA gene sequence similarity, which was confirmed by the high DNA-DNA relatedness (≥82%) between L. paralimentarius JCM 10415(T), L. bobalius JCM 16180(T) and L. kimchii JCM 10707(T). Therefore, it is proposed that L. kimchii and L. bobalius should be reclassified as later synonyms of L. paralimentarius.

Bacterial community and arsenic functional genes diversity in arsenic contaminated soils from different geographic locations

PubMed Central

Gu, Yunfu; D. Van Nostrand, Joy; Wu, Liyou; He, Zhili; Qin, Yujia; Zhao, Fang-Jie; Zhou, Jizhong

2017-01-01

To understand how soil microbial communities and arsenic (As) functional genes respond to soil arsenic (As) contamination, five soils contaminated with As at different levels were collected from diverse geographic locations, incubated for 54 days under flooded conditions, and examined by both MiSeq sequencing of 16S rRNA gene amplicons and functional gene microarray (GeoChip 4.0). The results showed that both bacterial community structure and As functional gene structure differed among geographical locations. The diversity of As functional genes correlated positively with the diversity of 16S rRNA genes (P< 0.05). Higher diversities of As functional genes and 16S rRNA genes were observed in the soils with higher available As. Soil pH, phosphate-extractable As, and amorphous Fe content were the most important factors in shaping the bacterial community structure and As transformation functional genes. Geographic location was also important in controlling both the bacterial community and As transformation functional potential. These findings provide insights into the variation of As transformation functional genes in soils contaminated with different levels of As at different geographic locations, and the impact of environmental As contamination on the soil bacterial community. PMID:28475654

DIVERSITY AND HOST SPECIFICITY OF AZOLLA CYANOBIONTS(1).

PubMed

Papaefthimiou, Dimitra; Van Hove, Charles; Lejeune, André; Rasmussen, Ulla; Wilmotte, Annick

2008-02-01

A unique, hereditary symbiosis exists between the water fern Azolla and cyanobacteria that reside within a cavity in the dorsal leaf-lobe of the plant. This association has been studied extensively, and questions have frequently been raised regarding the number and diversity of cyanobionts (cyanobacterial symbionts) among the different Azolla strains and species. In this work, denaturating gradient gel electrophoresis (DGGE) and a clone library based on the 16S rRNA gene were used to study the genetic diversity and host specificity of the cyanobionts in 35 Azolla strains covering a wide taxonomic and geographic range. DNA was extracted directly from the cyanobacterial packets, isolated after enzymatic digestion of the Azolla leaves. Our results indicated the existence of different cyanobiont strains among Azolla species, and diversity within a single Azolla species, independent of the geographic origin of the host. Furthermore, the cyanobiont exhibited host-species specificity and showed most divergence between the two sections of genus Azolla, Azolla and Rhizosperma. These findings are in agreement with the recent redefinition of the taxon Azolla cristata within the section Azolla. With regard to the taxonomic status of the cyanobiont, the genus Anabaena of the Nostocaceae family was identified as the closest relative by this work. © 2008 Phycological Society of America.

Diversity of Phylogenetic Information According to the Locus and the Taxonomic Level: An Example from a Parasitic Mesostigmatid Mite Genus

PubMed Central

Roy, Lise; Dowling, Ashley P.G.; Chauve, Claude Marie; Buronfosse, Thierry

2010-01-01

Molecular markers for cladistic analyses may perform differently according to the taxonomic group considered and the historical level under investigation. Here we evaluate the phylogenetic potential of five different markers for resolving evolutionary relationships within the ectoparasitic genus Dermanyssus at the species level, and their ability to address questions about the evolution of specialization. COI provided 9–18% divergence between species (up to 9% within species), 16S rRNA 10–16% (up to 4% within species), ITS1 and 2 2–9% (up to 1% within species) and Tropomyosin intron n 8–20% (up to 6% within species). EF-1α revealed different non-orthologous copies within individuals of Dermanyssus and Ornithonyssus. Tropomyosin intron n was shown containing consistent phylogenetic signal at the specific level within Dermanyssus and represents a promising marker for future prospects in phylogenetics of Acari. Phylogenetic analyses revealed that the generalist condition is apomorphic and D. gallinae might represent a complex of hybridized lineages. The split into hirsutus-group and gallinae-group in Dermanyssus does not seem to be appropriate based upon these results and D. longipes appears to be composed of two different entities. PMID:20480038

PhyloChip™ microarray comparison of sampling methods used for coral microbial ecology

USGS Publications Warehouse

Kellogg, Christina A.; Piceno, Yvette M.; Tom, Lauren M.; DeSantis, Todd Z.; Zawada, David G.; Andersen, Gary L.

2012-01-01

Interest in coral microbial ecology has been increasing steadily over the last decade, yet standardized methods of sample collection still have not been defined. Two methods were compared for their ability to sample coral-associated microbial communities: tissue punches and foam swabs, the latter being less invasive and preferred by reef managers. Four colonies of star coral, Montastraea annularis, were sampled in the Dry Tortugas National Park (two healthy and two with white plague disease). The PhyloChip™ G3 microarray was used to assess microbial community structure of amplified 16S rRNA gene sequences. Samples clustered based on methodology rather than coral colony. Punch samples from healthy and diseased corals were distinct. All swab samples clustered closely together with the seawater control and did not group according to the health state of the corals. Although more microbial taxa were detected by the swab method, there is a much larger overlap between the water control and swab samples than punch samples, suggesting some of the additional diversity is due to contamination from water absorbed by the swab. While swabs are useful for noninvasive studies of the coral surface mucus layer, these results show that they are not optimal for studies of coral disease.

PhyloChip™ microarray comparison of sampling methods used for coral microbial ecology.

PubMed

Kellogg, Christina A; Piceno, Yvette M; Tom, Lauren M; DeSantis, Todd Z; Zawada, David G; Andersen, Gary L

2012-01-01

Interest in coral microbial ecology has been increasing steadily over the last decade, yet standardized methods of sample collection still have not been defined. Two methods were compared for their ability to sample coral-associated microbial communities: tissue punches and foam swabs, the latter being less invasive and preferred by reef managers. Four colonies of star coral, Montastraea annularis, were sampled in the Dry Tortugas National Park (two healthy and two with white plague disease). The PhyloChip™ G3 microarray was used to assess microbial community structure of amplified 16S rRNA gene sequences. Samples clustered based on methodology rather than coral colony. Punch samples from healthy and diseased corals were distinct. All swab samples clustered closely together with the seawater control and did not group according to the health state of the corals. Although more microbial taxa were detected by the swab method, there is a much larger overlap between the water control and swab samples than punch samples, suggesting some of the additional diversity is due to contamination from water absorbed by the swab. While swabs are useful for noninvasive studies of the coral surface mucus layer, these results show that they are not optimal for studies of coral disease. Published by Elsevier B.V.

Microbial Community Structure of Subglacial Lake Whillans, West Antarctica

PubMed Central

Achberger, Amanda M.; Christner, Brent C.; Michaud, Alexander B.; Priscu, John C.; Skidmore, Mark L.; Vick-Majors, Trista J.; Adkins, W.

2016-01-01

Subglacial Lake Whillans (SLW) is located beneath ∼800 m of ice on the Whillans Ice Stream in West Antarctica and was sampled in January of 2013, providing the first opportunity to directly examine water and sediments from an Antarctic subglacial lake. To minimize the introduction of surface contaminants to SLW during its exploration, an access borehole was created using a microbiologically clean hot water drill designed to reduce the number and viability of microorganisms in the drilling water. Analysis of 16S rRNA genes (rDNA) amplified from samples of the drilling and borehole water allowed an evaluation of the efficacy of this approach and enabled a confident assessment of the SLW ecosystem inhabitants. Based on an analysis of 16S rDNA and rRNA (i.e., reverse-transcribed rRNA molecules) data, the SLW community was found to be bacterially dominated and compositionally distinct from the assemblages identified in the drill system. The abundance of bacteria (e.g., Candidatus Nitrotoga, Sideroxydans, Thiobacillus, and Albidiferax) and archaea (Candidatus Nitrosoarchaeum) related to chemolithoautotrophs was consistent with the oxidation of reduced iron, sulfur, and nitrogen compounds having important roles as pathways for primary production in this permanently dark ecosystem. Further, the prevalence of Methylobacter in surficial lake sediments combined with the detection of methanogenic taxa in the deepest sediment horizons analyzed (34–36 cm) supported the hypothesis that methane cycling occurs beneath the West Antarctic Ice Sheet. Large ratios of rRNA to rDNA were observed for several operational taxonomic units abundant in the water column and sediments (e.g., Albidiferax, Methylobacter, Candidatus Nitrotoga, Sideroxydans, and Smithella), suggesting a potentially active role for these taxa in the SLW ecosystem. Our findings are consistent with chemosynthetic microorganisms serving as the ecological foundation in this dark subsurface environment, providing new organic matter that sustains a microbial ecosystem beneath the West Antarctic Ice Sheet. PMID:27713727

Paenibacillus phoenicis sp. nov., isolated from the Phoenix Lander assembly facility and a subsurface molybdenum mine.

PubMed

Benardini, James N; Vaishampayan, Parag A; Schwendner, Petra; Swanner, Elizabeth; Fukui, Youhei; Osman, Sharif; Satomi, Masakata; Venkateswaran, Kasthuri

2011-06-01

A novel Gram-positive, motile, endospore-forming, aerobic bacterium was isolated from the NASA Phoenix Lander assembly clean room that exhibits 100 % 16S rRNA gene sequence similarity to two strains isolated from a deep subsurface environment. All strains are rod-shaped, endospore-forming bacteria, whose endospores are resistant to UV radiation up to 500 J m(-2). A polyphasic taxonomic study including traditional phenotypic tests, fatty acid analysis, 16S rRNA gene sequencing and DNA-DNA hybridization analysis was performed to characterize these novel strains. The 16S rRNA gene sequencing convincingly grouped these novel strains within the genus Paenibacillus as a separate cluster from previously described species. The similarity of 16S rRNA gene sequences among the novel strains was identical but only 98.1 to 98.5 % with their nearest neighbours Paenibacillus barengoltzii ATCC BAA-1209(T) and Paenibacillus timonensis CIP 108005(T). The menaquinone MK-7 was dominant in these novel strains as shown in other species of the genus Paenibacillus. The DNA-DNA hybridization dissociation value was <45 % with the closest related species. The novel strains had DNA G+C contents of 51.9 to 52.8 mol%. Phenotypically, the novel strains can be readily differentiated from closely related species by the absence of urease and gelatinase and the production of acids from a variety of sugars including l-arabinose. The major fatty acid was anteiso-C(15 : 0) as seen in P. barengoltzii and P. timonensis whereas the proportion of C(16 : 0) was significantly different from the closely related species. Based on phylogenetic and phenotypic results, it was concluded that these strains represent a novel species of the genus Paenibacillus, for which the name Paenibacillus phoenicis sp. nov. is proposed. The type strain is 3PO2SA(T) ( = NRRL B-59348(T)  = NBRC 106274(T)).

Culture-dependent and independent studies of microbial diversity in highly copper-contaminated Chilean marine sediments.

PubMed

Besaury, Ludovic; Marty, Florence; Buquet, Sylvaine; Mesnage, Valérie; Muyzer, Gerard; Quillet, Laurent

2013-02-01

Cultivation and molecular-based approaches were used to study microbial diversity in two Chilean marine sediments contaminated with high (835 ppm) and very high concentrations of copper (1,533 ppm). The diversity of cultivable bacteria resistant to copper was studied at oxic and anoxic conditions, focusing on sulfate-, thiosulfate-, and iron-reducing bacteria. For both sediments, the cultivable bacteria isolated at oxic conditions were mostly affiliated to the genus Bacillus, while at anoxic conditions the majority of the cultivable bacteria found were closely related to members of the genera Desulfovibrio, Sphingomonas, and Virgibacillus. Copper resistance was between 100 and 400 ppm, with the exception of a strain affiliated to members of the genus Desulfuromonas, which was resistant up to 1,000 ppm of copper. In parallel, cloning and sequencing of 16S rRNA was performed to study the total bacterial diversity in the sediments. A weak correlation was observed between the isolated strains and the 16S rRNA operational taxonomic units detected. The presence of copper resistance genes (copA, cusA, and pcoA) was tested for all the strains isolated; only copA was detected in a few isolates, suggesting that other copper resistance mechanisms could be used by the bacteria in those highly copper-contaminated sediments.

Colwellia aestuarii sp. nov., isolated from a tidal flat sediment in Korea.

PubMed

Jung, Seo-Youn; Oh, Tae-Kwang; Yoon, Jung-Hoon

2006-01-01

A novel Colwellia-like bacterial strain, SMK-10T, was isolated from a tidal flat sediment in Korea and subjected to a polyphasic taxonomic analysis. Cells of strain SMK-10T were Gram-negative, motile, greyish yellow-pigmented, curved rods. Optimal growth occurred at 25-30 degrees C and in the presence of 2-3 % (w/v) NaCl. Strain SMK-10T contained Q-8 as the predominant ubiquinone and C(16 : 1)omega7c and/or iso-C(15 : 0) 2-OH, C(17 : 1), C(15 : 1) and iso-C(16 : 0) as major fatty acids. The DNA G+C content was 39.3 mol%. Phylogenetic trees based on 16S rRNA gene sequence analysis showed that strain SMK-10T belonged to the genus Colwellia. 16S rRNA gene sequence similarity values (94.7-96.7 %) to the type strains of all other Colwellia species and various differential phenotypic properties were sufficient to distinguish strain SMK-10T from recognized Colwellia species. On the basis of its phenotypic properties and phylogenetic distinctiveness, strain SMK-10T (= KCTC 12480T = DSM 17314T) is classified as the type strain of a novel Colwellia species, for which the name Colwellia aestuarii sp. nov. is proposed.

Phylogeography and systematics of the westernmost Italian Dolichopoda species (Orthoptera, Rhaphidophoridae)

PubMed Central

Allegrucci, Giuliana; Rampini, Mauro; Di Russo, Claudio; Lana, Enrico; Cocchi, Sara; Sbordoni, Valerio

2014-01-01

Abstract The genus Dolichopoda (Orthoptera; Rhaphidopohoridae) is present in Italy with 9 species distributed from northwestern Italy (Piedmont and Liguria) to the southernmost Apennines (Calabria), occurring also in the Tyrrhenian coastal areas and in Sardinia. Three morphologically very close taxa have been described in Piedmont and Liguria, i.e., D. ligustica ligustica, D. ligustica septentrionalis and D. azami azami. To investigate the delimitation of the northwestern species of Dolichopoda, we performed both morphological and molecular analyses. Morphological analysis was carried out by considering diagnostic characters generally used to distinguish different taxa, as the shape of epiphallus in males and the subgenital fig in females. Molecular analysis was performed by sequencing three mitochondrial genes, 12S rRNA, 16S rRNA, partially sequenced and the entire gene of COI. Results from both morphological and molecular analyses highlighted a very homogeneous group of populations, although genetically structured. Three haplogroups geographically distributed could be distinguished and based on these results we suggest a new taxonomic arrangement. All populations, due to the priority of description, should be assigned to D. azami azami Saulcy, 1893 and to preserve the names ligustica and septentrionalis, corresponding to different genetic haplogroups, we assign them to D. azami ligustica stat. n. Baccetti & Capra, 1959 and to D. azami septentrionalis stat. n. Baccetti & Capra, 1959. PMID:25197209

Detailed investigation of the microbial community in foaming activated sludge reveals novel foam formers

PubMed Central

Guo, Feng; Wang, Zhi-Ping; Yu, Ke; Zhang, T.

2015-01-01

Foaming of activated sludge (AS) causes adverse impacts on wastewater treatment operation and hygiene. In this study, we investigated the microbial communities of foam, foaming AS and non-foaming AS in a sewage treatment plant via deep-sequencing of the taxonomic marker genes 16S rRNA and mycobacterial rpoB and a metagenomic approach. In addition to Actinobacteria, many genera (e.g., Clostridium XI, Arcobacter, Flavobacterium) were more abundant in the foam than in the AS. On the other hand, deep-sequencing of rpoB did not detect any obligate pathogenic mycobacteria in the foam. We found that unknown factors other than the abundance of Gordonia sp. could determine the foaming process, because abundance of the same species was stable before and after a foaming event over six months. More interestingly, although the dominant Gordonia foam former was the closest with G. amarae, it was identified as an undescribed Gordonia species by referring to the 16S rRNA gene, gyrB and, most convincingly, the reconstructed draft genome from metagenomic reads. Our results, based on metagenomics and deep sequencing, reveal that foams are derived from diverse taxa, which expands previous understanding and provides new insight into the underlying complications of the foaming phenomenon in AS. PMID:25560234

Molecular tools for the selective detection of nine diatom species biomarkers of various water quality levels.

PubMed

Cimarelli, Lucia; Singh, Kumar Saurabh; Mai, Nguyen Thi Nhu; Dhar, Bidhan Chandra; Brandi, Anna; Brandi, Letizia; Spurio, Roberto

2015-05-22

Our understanding of the composition of diatom communities and their response to environmental changes is currently limited by laborious taxonomic identification procedures. Advances in molecular technologies are expected to contribute more efficient, robust and sensitive tools for the detection of these ecologically relevant microorganisms. There is a need to explore and test phylogenetic markers as an alternative to the use of rRNA genes, whose limited sequence divergence does not allow the accurate discrimination of diatoms at the species level. In this work, nine diatom species belonging to eight genera, isolated from epylithic environmental samples collected in central Italy, were chosen to implement a panel of diatoms covering the full range of ecological status of freshwaters. The procedure described in this work relies on the PCR amplification of specific regions in two conserved diatom genes, elongation factor 1-a (eEF1-a) and silicic acid transporter (SIT), as a first step to narrow down the complexity of the targets, followed by microarray hybridization experiments. Oligonucleotide probes with the potential to discriminate closely related species were designed taking into account the genetic polymorphisms found in target genes. These probes were tested, refined and validated on a small-scale prototype DNA chip. Overall, we obtained 17 highly specific probes targeting eEF1-a and SIT, along with 19 probes having lower discriminatory power recognizing at the same time two or three species. This basic array was validated in a laboratory setting and is ready for tests with crude environmental samples eventually to be scaled-up to include a larger panel of diatoms. Its possible use for the simultaneous detection of diatoms selected from the classes of water quality identified by the European Water Framework Directive is discussed.

Bacterial profiling of White Plague Disease across corals and oceans indicates a conserved and distinct disease microbiome

PubMed Central

Roder, Cornelia; Arif, Chatchanit; Daniels, Camille; Weil, Ernesto; Voolstra, Christian R

2014-01-01

Coral diseases are characterized by microbial community shifts in coral mucus and tissue, but causes and consequences of these changes are vaguely understood due to the complexity and dynamics of coral-associated bacteria. We used 16S rRNA gene microarrays to assay differences in bacterial assemblages of healthy and diseased colonies displaying White Plague Disease (WPD) signs from two closely related Caribbean coral species, Orbicella faveolata and Orbicella franksi. Analysis of differentially abundant operational taxonomic units (OTUs) revealed strong differences between healthy and diseased specimens, but not between coral species. A subsequent comparison to data from two Indo-Pacific coral species (Pavona duerdeni and Porites lutea) revealed distinct microbial community patterns associated with ocean basin, coral species and health state. Coral species were clearly separated by site, but also, the relatedness of the underlying bacterial community structures resembled the phylogenetic relationship of the coral hosts. In diseased samples, bacterial richness increased and putatively opportunistic bacteria were consistently more abundant highlighting the role of opportunistic conditions in structuring microbial community patterns during disease. Our comparative analysis shows that it is possible to derive conserved bacterial footprints of diseased coral holobionts that might help in identifying key bacterial species related to the underlying etiopathology. Furthermore, our data demonstrate that similar-appearing disease phenotypes produce microbial community patterns that are consistent over coral species and oceans, irrespective of the putative underlying pathogen. Consequently, profiling coral diseases by microbial community structure over multiple coral species might allow the development of a comparative disease framework that can inform on cause and relatedness of coral diseases. PMID:24350609

Complete mitochondrial genome of the a rare subspecies of genus Bos, Tianzhu white yak from Tibetan area in China.

PubMed

E, Guangxin; Na, Ri-Su; Zhao, Yong-Ju; Gao, Hui-Jiang; An, Tian-Wu; Huang, Yong-Fu

2016-01-01

The population of domestic yak, Tianzhu white yak, from Tibetan area in China is considered as a rare Bos grunniens species. We first determined and annotated its complete mitochondrial genome. The mitogenome is 16,319 bp in length, consisting of 13 protein-coding genes, 22 transfer RNA (tRNA) genes, 2 ribosomal RNA (rRNA) genes and a control region. As in other mammals, most mitochondrial genes are encoded on the heavy strand, except for ND6 and eight tRNA genes, which are encoded on the light strand. Its overall base composition is A: 33.7%, T: 27.2%, C: 25.8% and G: 13.2%. The complete mitogenome of the new subspecies of Bos grunniens could provide an important data to further explore the taxonomic status of the subspecies.

Metagenomic analysis of bacterial and archaeal assemblages in the soil-mousse surrounding a geothermal spring.

PubMed

Bhatia, Sonu; Batra, Navneet; Pathak, Ashish; Joshi, Amit; Souza, Leila; Almeida, Paulo; Chauhan, Ashvini

2015-09-01

The soil-mousse surrounding a geothermal spring was analyzed for bacterial and archaeal diversity using 16S rRNA gene amplicon metagenomic sequencing which revealed the presence of 18 bacterial phyla distributed across 109 families and 219 genera. Firmicutes, Actinobacteria, and the Deinococcus-Thermus group were the predominant bacterial assemblages with Crenarchaeota and Thaumarchaeota as the main archaeal assemblages in this largely understudied geothermal habitat. Several metagenome sequences remained taxonomically unassigned suggesting the presence of a repertoire of hitherto undescribed microbes in this geothermal soil-mousse econiche.

Stability of operational taxonomic units: an important but neglected property for analyzing microbial diversity.

PubMed

He, Yan; Caporaso, J Gregory; Jiang, Xiao-Tao; Sheng, Hua-Fang; Huse, Susan M; Rideout, Jai Ram; Edgar, Robert C; Kopylova, Evguenia; Walters, William A; Knight, Rob; Zhou, Hong-Wei

2015-01-01

The operational taxonomic unit (OTU) is widely used in microbial ecology. Reproducibility in microbial ecology research depends on the reliability of OTU-based 16S ribosomal subunit RNA (rRNA) analyses. Here, we report that many hierarchical and greedy clustering methods produce unstable OTUs, with membership that depends on the number of sequences clustered. If OTUs are regenerated with additional sequences or samples, sequences originally assigned to a given OTU can be split into different OTUs. Alternatively, sequences assigned to different OTUs can be merged into a single OTU. This OTU instability affects alpha-diversity analyses such as rarefaction curves, beta-diversity analyses such as distance-based ordination (for example, Principal Coordinate Analysis (PCoA)), and the identification of differentially represented OTUs. Our results show that the proportion of unstable OTUs varies for different clustering methods. We found that the closed-reference method is the only one that produces completely stable OTUs, with the caveat that sequences that do not match a pre-existing reference sequence collection are discarded. As a compromise to the factors listed above, we propose using an open-reference method to enhance OTU stability. This type of method clusters sequences against a database and includes unmatched sequences by clustering them via a relatively stable de novo clustering method. OTU stability is an important consideration when analyzing microbial diversity and is a feature that should be taken into account during the development of novel OTU clustering methods.

Comparative and Evolutionary Analyses of Meloidogyne spp. Based on Mitochondrial Genome Sequences

PubMed Central

García, Laura Evangelina; Sánchez-Puerta, M. Virginia

2015-01-01

Molecular taxonomy and evolution of nematodes have been recently the focus of several studies. Mitochondrial sequences were proposed as an alternative for precise identification of Meloidogyne species, to study intraspecific variability and to follow maternal lineages. We characterized the mitochondrial genomes (mtDNAs) of the root knot nematodes M. floridensis, M. hapla and M. incognita. These were AT rich (81–83%) and highly compact, encoding 12 proteins, 2 rRNAs, and 22 tRNAs. Comparisons with published mtDNAs of M. chitwoodi, M. incognita (another strain) and M. graminicola revealed that they share protein and rRNA gene order but differ in the order of tRNAs. The mtDNAs of M. floridensis and M. incognita were strikingly similar (97–100% identity for all coding regions). In contrast, M. floridensis, M. chitwoodi, M. hapla and M. graminicola showed 65–84% nucleotide identity for coding regions. Variable mitochondrial sequences are potentially useful for evolutionary and taxonomic studies. We developed a molecular taxonomic marker by sequencing a highly-variable ~2 kb mitochondrial region, nad5-cox1, from 36 populations of root-knot nematodes to elucidate relationships within the genus Meloidogyne. Isolates of five species formed monophyletic groups and showed little intraspecific variability. We also present a thorough analysis of the mitochondrial region cox2-rrnS. Phylogenies based on either mitochondrial region had good discrimination power but could not discriminate between M. arenaria, M. incognita and M. floridensis. PMID:25799071

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ruvindy, Rendy; White III, Richard Allen; Neilan, Brett Anthony

Modern microbial mats are potential analogues of some of Earth’s earliest ecosystems. Excellent examples can be found in Shark Bay, Australia, with mats of various morphologies. To further our understanding of the functional genetic potential of these complex microbial ecosystems, we conducted for the first time shotgun metagenomic analyses. We assembled metagenomic nextgeneration sequencing data to classify the taxonomic and metabolic potential across diverse morphologies of marine mats in Shark Bay. The microbial community across taxonomic classifications using protein-coding and small subunit rRNA genes directly extracted from the metagenomes suggests that three phyla Proteobacteria, Cyanobacteria and Bacteriodetes dominate all marinemore » mats. However, the microbial community structure between Shark Bay and Highbourne Cay (Bahamas) marine systems appears to be distinct from each other. The metabolic potential (based on SEED subsystem classifications) of the Shark Bay and Highbourne Cay microbial communities were also distinct. Shark Bay metagenomes have a metabolic pathway profile consisting of both heterotrophic and photosynthetic pathways, whereas Highbourne Cay appears to be dominated almost exclusively by photosynthetic pathways. Alternative non-rubisco-based carbon metabolism including reductive TCA cycle and 3-hydroxypropionate/4-hydroxybutyrate pathways is highly represented in Shark Bay metagenomes while not represented in Highbourne Cay microbial mats or any other mat forming ecosystems investigated to date. Potentially novel aspects of nitrogen cycling were also observed, as well as putative heavy metal cycling (arsenic, mercury, copper and cadmium). Finally, archaea are highly represented in Shark Bay and may have critical roles in overall ecosystem function in these modern microbial mats.« less

Cyanobacterial diversity and halotolerance in a variable hypersaline environment.

PubMed

Kirkwood, Andrea E; Buchheim, Julie A; Buchheim, Mark A; Henley, William J

2008-04-01

The Great Salt Plains (GSP) in north-central Oklahoma, USA is an expansive salt flat (approximately 65 km(2)) that is part of the federally protected Salt Plains National Wildlife Refuge. The GSP serves as an ideal environment to study the microbial diversity of a terrestrial, hypersaline system that experiences wide fluctuations in freshwater influx and diel temperature. Our study assessed cyanobacterial diversity at the GSP by focusing on the taxonomic and physiological diversity of GSP isolates, and the 16S rRNA phylogenetic diversity of isolates and environmental clones from three sites (north, central, and south). Taxonomic diversity of isolates was limited to a few genera (mostly Phormidium and Geitlerinema), but physiological diversity based on halotolerance ranges was strikingly more diverse, even between strains of the same phylotype. The phylogenetic tree revealed diversity that spanned a number of cyanobacterial lineages, although diversity at each site was dominated by only a few phylotypes. Unlike other hypersaline systems, a number of environmental clones from the GSP were members of the heterocystous lineage. Although a number of cyanobacterial isolates were close matches with prevalent environmental clones, it is not certain if these clones reflect the same halotolerance ranges of their matching isolates. This caveat is based on the notable disparities we found between strains of the same phylotype and their inherent halotolerance. Our findings support the hypothesis that variable or poikilotrophic environments promote diversification, and in particular, select for variation in ecotype more than phylotype.

Bacterial diversity characterization in petroleum samples from Brazilian reservoirs

PubMed Central

de Oliveira, Valéria Maia; Sette, Lara Durães; Simioni, Karen Christina Marques; dos Santos Neto, Eugênio Vaz

2008-01-01

This study aimed at evaluating potential differences among the bacterial communities from formation water and oil samples originated from biodegraded and non-biodegraded Brazilian petroleum reservoirs by using a PCR-DGGE based approach. Environmental DNA was isolated and used in PCR reactions with bacterial primers, followed by separation of 16S rDNA fragments in the DGGE. PCR products were also cloned and sequenced, aiming at the taxonomic affiliation of the community members. The fingerprints obtained allowed the direct comparison among the bacterial communities from oil samples presenting distinct degrees of biodegradation, as well as between the communities of formation water and oil sample from the non-biodegraded reservoir. Very similar DGGE band profiles were observed for all samples, and the diversity of the predominant bacterial phylotypes was shown to be low. Cloning and sequencing results revealed major differences between formation water and oil samples from the non-biodegraded reservoir. Bacillus sp. and Halanaerobium sp. were shown to be the predominant components of the bacterial community from the formation water sample, whereas the oil sample also included Alicyclobacillus acidoterrestris, Rhodococcus sp., Streptomyces sp. and Acidithiobacillus ferrooxidans. The PCR-DGGE technique, combined with cloning and sequencing of PCR products, revealed the presence of taxonomic groups not found previously in these samples when using cultivation-based methods and 16S rRNA gene library assembly, confirming the need of a polyphasic study in order to improve the knowledge of the extent of microbial diversity in such extreme environments. PMID:24031244

Systematics of the grey mullets (Teleostei: Mugiliformes: Mugilidae): molecular phylogenetic evidence challenges two centuries of morphology-based taxonomy.

PubMed

Durand, J-D; Shen, K-N; Chen, W-J; Jamandre, B W; Blel, H; Diop, K; Nirchio, M; Garcia de León, F J; Whitfield, A K; Chang, C-W; Borsa, P

2012-07-01

The family Mugilidae comprises mainly coastal marine species that are widely distributed in all tropical, subtropical and temperate seas. Mugilid species are generally considered to be ecologically important and they are a major food resource for human populations in certain parts of the world. The taxonomy and systematics of the Mugilidae are still much debated and based primarily on morphological characters. In this study, we provide the first comprehensive molecular systematic account of the Mugilidae using phylogenetic analyses of nucleotide sequence variation at three mitochondrial loci (16S rRNA, cytochrome oxidase I, and cytochrome b) for 257 individuals from 55 currently recognized species. The study covers all 20 mugilid genera currently recognized as being valid. The family comprises seven major lineages that radiated early on from the ancestor to all current forms. All genera that were represented by two species or more, except Cestraeus, turned out to be paraphyletic or polyphyletic. Thus, the present phylogenetic results generally disagree with the current taxonomy at the genus level and imply that the anatomical characters used for the systematics of the Mugilidae may be poorly informative phylogenetically. The present results should provide a sound basis for a taxonomic revision of the mugilid genera. A proportion of the species with large distribution ranges (including Moolgarda seheli, Mugil cephalus and M. curema) appear to consist of cryptic species, thus warranting further taxonomic and genetic work at the infra-generic level. Copyright © 2012 Elsevier Inc. All rights reserved.

Sensitive and substrate-specific detection of metabolically active microorganisms in natural microbial consortia using community isotope arrays.

PubMed

Tourlousse, Dieter M; Kurisu, Futoshi; Tobino, Tomohiro; Furumai, Hiroaki

2013-05-01

The goal of this study was to develop and validate a novel fosmid-clone-based metagenome isotope array approach - termed the community isotope array (CIArray) - for sensitive detection and identification of microorganisms assimilating a radiolabeled substrate within complex microbial communities. More specifically, a sample-specific CIArray was used to identify anoxic phenol-degrading microorganisms in activated sludge treating synthetic coke-oven wastewater in a single-sludge predenitrification-nitrification process. Hybridization of the CIArray with DNA from the (14) C-phenol-amended sample indicated that bacteria assimilating (14) C-atoms, presumably directly from phenol, under nitrate-reducing conditions were abundant in the reactor, and taxonomic assignment of the fosmid clone end sequences suggested that they belonged to the Gammaproteobacteria. The specificity of the CIArray was validated by quantification of fosmid-clone-specific DNA in density-resolved DNA fractions from samples incubated with (13) C-phenol, which verified that all CIArray-positive probes stemmed from microorganisms that assimilated isotopically labeled carbon. This also demonstrated that the CIArray was more sensitive than DNA-SIP, as the former enabled positive detection at a phenol concentration that failed to yield a 'heavy' DNA fraction. Finally, two operational taxonomic units distantly related to marine Gammaproteobacteria were identified to account for more than half of 16S rRNA gene clones in the 'heavy' DNA library, corroborating the CIArray-based identification. © 2013 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

The phylogenetic composition and structure of soil microbial communities shifts in response to elevated carbon dioxide.

PubMed

He, Zhili; Piceno, Yvette; Deng, Ye; Xu, Meiying; Lu, Zhenmei; Desantis, Todd; Andersen, Gary; Hobbie, Sarah E; Reich, Peter B; Zhou, Jizhong

2012-02-01

One of the major factors associated with global change is the ever-increasing concentration of atmospheric CO(2). Although the stimulating effects of elevated CO(2) (eCO(2)) on plant growth and primary productivity have been established, its impacts on the diversity and function of soil microbial communities are poorly understood. In this study, phylogenetic microarrays (PhyloChip) were used to comprehensively survey the richness, composition and structure of soil microbial communities in a grassland experiment subjected to two CO(2) conditions (ambient, 368 p.p.m., versus elevated, 560 p.p.m.) for 10 years. The richness based on the detected number of operational taxonomic units (OTUs) significantly decreased under eCO(2). PhyloChip detected 2269 OTUs derived from 45 phyla (including two from Archaea), 55 classes, 99 orders, 164 families and 190 subfamilies. Also, the signal intensity of five phyla (Crenarchaeota, Chloroflexi, OP10, OP9/JS1, Verrucomicrobia) significantly decreased at eCO(2), and such significant effects of eCO(2) on microbial composition were also observed at the class or lower taxonomic levels for most abundant phyla, such as Proteobacteria, Firmicutes, Actinobacteria, Bacteroidetes and Acidobacteria, suggesting a shift in microbial community composition at eCO(2). Additionally, statistical analyses showed that the overall taxonomic structure of soil microbial communities was altered at eCO(2). Mantel tests indicated that such changes in species richness, composition and structure of soil microbial communities were closely correlated with soil and plant properties. This study provides insights into our understanding of shifts in the richness, composition and structure of soil microbial communities under eCO(2) and environmental factors shaping the microbial community structure.

Comparing bacterial community composition between healthy and white plague-like disease states in Orbicella annularis using PhyloChip™ G3 microarrays

USGS Publications Warehouse

Kellogg, Christina A.; Piceno, Yvette M.; Tom, Lauren M.; DeSantis, Todd Z.; Gray, Michael A.; Zawada, David G.; Andersen, Gary L.

2013-01-01

Coral disease is a global problem. Diseases are typically named or described based on macroscopic changes, but broad signs of coral distress such as tissue loss or discoloration are unlikely to be specific to a particular pathogen. For example, there appear to be multiple diseases that manifest the rapid tissue loss that characterizes ‘white plague.’ PhyloChip™ G3 microarrays were used to compare the bacterial community composition of both healthy and white plague-like diseased corals. Samples of lobed star coral (Orbicella annularis, formerly of the genus Montastraea [1]) were collected from two geographically distinct areas, Dry Tortugas National Park and Virgin Islands National Park, to determine if there were biogeographic differences between the diseases. In fact, all diseased samples clustered together, however there was no consistent link to Aurantimonas coralicida, which has been described as the causative agent of white plague type II. The microarrays revealed a large amount of bacterial heterogeneity within the healthy corals and less diversity in the diseased corals. Gram-positive bacterial groups (Actinobacteria, Firmicutes) comprised a greater proportion of the operational taxonomic units (OTUs) unique to healthy samples. Diseased samples were enriched in OTUs from the families Corynebacteriaceae, Lachnospiraceae, Rhodobacteraceae, and Streptococcaceae. Much previous coral disease work has used clone libraries, which seem to be methodologically biased toward recovery of Gram-negative bacterial sequences and may therefore have missed the importance of Gram-positive groups. The PhyloChip™ data presented here provide a broader characterization of the bacterial community changes that occur within Orbicella annularis during the shift from a healthy to diseased state.

Comparing Bacterial Community Composition between Healthy and White Plague-Like Disease States in Orbicella annularis Using PhyloChip™ G3 Microarrays

PubMed Central

Kellogg, Christina A.; Piceno, Yvette M.; Tom, Lauren M.; DeSantis, Todd Z.; Gray, Michael A.; Zawada, David G.; Andersen, Gary L.

2013-01-01

Coral disease is a global problem. Diseases are typically named or described based on macroscopic changes, but broad signs of coral distress such as tissue loss or discoloration are unlikely to be specific to a particular pathogen. For example, there appear to be multiple diseases that manifest the rapid tissue loss that characterizes ‘white plague.’ PhyloChip™ G3 microarrays were used to compare the bacterial community composition of both healthy and white plague-like diseased corals. Samples of lobed star coral (Orbicella annularis, formerly of the genus Montastraea [1]) were collected from two geographically distinct areas, Dry Tortugas National Park and Virgin Islands National Park, to determine if there were biogeographic differences between the diseases. In fact, all diseased samples clustered together, however there was no consistent link to Aurantimonas coralicida, which has been described as the causative agent of white plague type II. The microarrays revealed a large amount of bacterial heterogeneity within the healthy corals and less diversity in the diseased corals. Gram-positive bacterial groups (Actinobacteria, Firmicutes) comprised a greater proportion of the operational taxonomic units (OTUs) unique to healthy samples. Diseased samples were enriched in OTUs from the families Corynebacteriaceae, Lachnospiraceae, Rhodobacteraceae, and Streptococcaceae. Much previous coral disease work has used clone libraries, which seem to be methodologically biased toward recovery of Gram-negative bacterial sequences and may therefore have missed the importance of Gram-positive groups. The PhyloChip™data presented here provide a broader characterization of the bacterial community changes that occur within Orbicella annularis during the shift from a healthy to diseased state. PMID:24278181

Diversity of Bacterial Communities in Container Habitats of Mosquitoes

PubMed Central

Ponnusamy, Loganathan; Xu, Ning; Stav, Gil; Wesson, Dawn M.; Schal, Coby

2010-01-01

We investigated the bacterial diversity of microbial communities in water-filled, human-made and natural container habitats of the mosquitoes Aedes aegypti and Aedes albopictus in suburban landscapes of New Orleans, Louisiana in 2003. We collected water samples from three classes of containers, including tires (n=12), cemetery urns (n=23), and miscellaneous containers that included two tree holes (n=19). Total genomic DNA was extracted from water samples, and 16S ribosomal DNA fragments (operational taxonomic units, OTUs) were amplified by PCR and separated by denaturing gradient gel electrophoresis (DGGE). The bacterial communities in containers represented diverse DGGE-DNA banding patterns that were not related to the class of container or to the local spatial distribution of containers. Mean richness and evenness of OTUs were highest in water samples from tires. Bacterial phylotypes were identified by comparative sequence analysis of 90 16S rDNA DGGE band amplicons. The majority of sequences were placed in five major taxa: Alpha-, Beta- and Gammaproteobacteria, Actinobacteria, Bacteroidetes, Cyanobacteria, Firmicutes, and an unclassified group; Proteobacteria and Bacteroidetes were the predominant heterotrophic bacteria in containers. The bacterial communities in human-made containers consisted mainly of undescribed species, and a phylogenetic analysis based on 16S rRNA sequences suggested that species composition was independent of both container type and the spatial distribution of containers. Comparative PCR-based, cultivation-independent rRNA surveys of microbial communities associated with mosquito habitats can provide significant insight into community organization and dynamics of bacterial species. PMID:18373113

Lacinutrix chionocetis sp. nov., isolated from gut of a red snow crab.

PubMed

Kim, Hyangmi; Yoon, Sang-Chul; Choi, Kwang-Ho; Kim, Sung-Tae; Lee, Jae-Bong; Kim, Dong-Sun; Le Han, Ho; Bae, Kyung Sook; Park, Doo-Sang

2017-05-01

A Gram-negative, aerobic, non-motile, rod-shaped bacterial strain, designated MAB-07 T , was isolated from the gut of a red snow crab. The novel strain grew optimally at 20 °C, pH 7.0-8.0, and in the presence of 3% (w/v) NaCl. A phylogenetic analysis based on the 16S rRNA gene sequence indicated that the strain MAB-07 T belongs to the type strains of species of the genus Lacinutrix. Strain MAB-07 T exhibited 16S rRNA gene sequence similarity values of 95.5-97.8% with the type strains of species of the genus Lacinutrix. The predominant cellular fatty acids of strain MAB-07 T were iso-C 15:1 G (27.5%) and iso-C 15:0 (21.7%). The major respiratory quinine was identified as MK-6. The polar lipids consisted of phosphatidylethanolamine, four unidentified aminolipids, and two unidentified lipids. The genomic DNA G + C content was determined to be 33.3%, and its DNA-DNA relatedness values with the type strains of L. venerupis, L. mariniflava, L. jangbogonensis, L. algicola, and Olleya aquimaris were 28-32%. Based on the data from this polyphasic taxonomic study, strain MAB-07 T is considered to represent a novel species of the genus Lacinutrix, for which the name L. chionocetis sp. nov. is proposed. The type strain is MAB-07 T (=KCTC 42767 T  = JCM 30988 T ).

Vector soup: high-throughput identification of Neotropical phlebotomine sand flies using metabarcoding.

PubMed

Kocher, Arthur; Gantier, Jean-Charles; Gaborit, Pascal; Zinger, Lucie; Holota, Helene; Valiere, Sophie; Dusfour, Isabelle; Girod, Romain; Bañuls, Anne-Laure; Murienne, Jerome

2017-03-01

Phlebotomine sand flies are haematophagous dipterans of primary medical importance. They represent the only proven vectors of leishmaniasis worldwide and are involved in the transmission of various other pathogens. Studying the ecology of sand flies is crucial to understand the epidemiology of leishmaniasis and further control this disease. A major limitation in this regard is that traditional morphological-based methods for sand fly species identifications are time-consuming and require taxonomic expertise. DNA metabarcoding holds great promise in overcoming this issue by allowing the identification of multiple species from a single bulk sample. Here, we assessed the reliability of a short insect metabarcode located in the mitochondrial 16S rRNA for the identification of Neotropical sand flies, and constructed a reference database for 40 species found in French Guiana. Then, we conducted a metabarcoding experiment on sand flies mixtures of known content and showed that the method allows an accurate identification of specimens in pools. Finally, we applied metabarcoding to field samples caught in a 1-ha forest plot in French Guiana. Besides providing reliable molecular data for species-level assignations of phlebotomine sand flies, our study proves the efficiency of metabarcoding based on the mitochondrial 16S rRNA for studying sand fly diversity from bulk samples. The application of this high-throughput identification procedure to field samples can provide great opportunities for vector monitoring and eco-epidemiological studies. © 2016 John Wiley & Sons Ltd.

Molecular detection and phylogenetic analysis of Hepatozoon spp. in questing Ixodes ricinus ticks and rodents from Slovakia and Czech Republic.

PubMed

Hamšíková, Zuzana; Silaghi, Cornelia; Rudolf, Ivo; Venclíková, Kristýna; Mahríková, Lenka; Slovák, Mirko; Mendel, Jan; Blažejová, Hana; Berthová, Lenka; Kocianová, Elena; Hubálek, Zdeněk; Schnittger, Leonhard; Kazimírová, Mária

2016-10-01

By amplification and sequencing of 18S rRNA gene fragments, Hepatozoon spp. DNA was detected in 0.08 % (4/5057) and 0.04 % (1/2473) of questing Ixodes ricinus ticks from Slovakia and Czech Republic, respectively. Hepatozoon spp. DNA was also detected in spleen and/or lungs of 4.45 % (27/606) of rodents from Slovakia. Prevalence of infection was significantly higher in Myodes glareolus (11.45 %) than in Apodemus spp. (0.28 %) (P < 0.001). Sequencing of 18S rRNA Hepatozoon spp. gene amplicons from I. ricinus showed 100 % identity with Hepatozoon canis isolates from red foxes or dogs in Europe. Phylogenetic analysis showed that at least two H. canis 18S rRNA genotypes exist in Slovakia of which one was identified also in the Czech Republic. The finding of H. canis in questing I. ricinus suggests the geographical spread of the parasite and a potential role of other ticks as its vectors in areas where Rhipicephalus sanguineus is not endemic. Sequencing of 18S rRNA gene amplicons from M. glareolus revealed the presence of two closely related genetic variants, Hepatozoon sp. SK1 and Hepatozoon sp. SK2, showing 99-100 % identity with isolates from M. glareolus from other European countries. Phylogenetic analysis demonstrates that 18S rRNA variants SK1 and SK2 correspond to previously described genotypes UR1 and UR2 of H. erhardovae, respectively. The isolate from Apodemus flavicollis (Hepatozoon sp. SK3b) was 99 % identical with isolates from reptiles in Africa and Asia. Further studies are necessary to identify the taxonomic status of Hepatozoon spp. parasitizing rodents in Europe and the host-parasite interactions in natural foci.

Assessing hog lagoon waste contamination in the Cape Fear Watershed using Bacteroidetes 16S rRNA gene pyrosequencing.

PubMed

Arfken, Ann M; Song, Bongkeun; Mallin, Michael A

2015-09-01

Hog lagoons can be major sources of waste and nutrient contamination to watersheds adjacent to pig farms. Fecal source tracking methods targeting Bacteroidetes 16S rRNA genes in pig fecal matter may underestimate or fail to detect hog lagoon contamination in riverine environments. In order to detect hog lagoon wastewater contamination in the Cape Fear Watershed, where a large number of hog farms are present, we conducted pyrosequencing analyses of Bacteroidetes 16S rRNA genes in hog lagoon waste and identified new hog lagoon-specific marker sequences. Additional pyrosequencing analyses of Bacteroidetes 16S rRNA genes were conducted with surface water samples collected at 4 sites during 5 months in the Cape Fear Watershed. Using an operational taxonomic unit (OTU) identity cutoff value of 97 %, these newly identified hog lagoon markers were found in 3 of the river samples, while only 1 sample contained the pig fecal marker. In the sample containing the pig fecal marker, there was a relatively high percentage (14.1 %) of the hog lagoon markers and a low pig fecal marker relative abundance of 0.4 % in the Bacteroidetes 16S rRNA gene sequences. This suggests that hog lagoon contamination must be somewhat significant in order for pig fecal markers to be detected, and low levels of hog lagoon contamination cannot be detected targeting only pig-specific fecal markers. Thus, new hog lagoon markers have a better detection capacity for lagoon waste contamination, and in conjunction with a pig fecal marker, provide a more comprehensive and accurate detection of hog lagoon waste contamination in susceptible watersheds.

Examination into the taxonomic position of Bacillus thermotolerans Yang et al., 2013, proposal for its reclassification into a new genus and species Quasibacillus thermotolerans gen. nov., comb. nov. and reclassification of B. encimensis Dastager et al., 2015 as a later heterotypic synonym of B. badius.

PubMed

Verma, Ashish; Pal, Yash; Khatri, Indu; Ojha, Anup Kumar; Gruber-Vodicka, Harald; Schumann, Peter; Dastager, Syed; Subramanian, Srikrishna; Mayilraj, Shanmugam; Krishnamurthi, Srinivasan

2017-10-01

Two novel Gram-staining positive, rod-shaped, moderately halotolerant, endospore forming bacterial strains 5.5LF 38TD and 5.5LF 48TD were isolated and taxonomically characterized from a landfill in Chandigarh, India. The analysis of 16S rRNA gene sequences of the strains confirmed their closest identity to Bacillus thermotolerans SgZ-8T with 99.9% sequence similarity. A comparative phylogenetic analysis of strains 5.5LF 38TD, 5.5LF 48TD and B. thermotolerans SgZ-8 T confirmed their separation into a novel genus with B. badius and genus Domibacillus as the closest phylogenetic relatives. The major fatty acids of the strains are iso-C 15:0 and iso-C 16:0 and MK-7 is the only quinone. The major polar lipids are diphosphatidylglycerol, phosphatidylglycerol and phosphatidylethanolamine. The digital DNA-DNA hybridization (DDH) and ortho average nucleotide identity (ANI) values calculated through whole genome sequences indicated that the three strains showed low relatedness with their phylogenetic neighbours. Based on evidences from phylogenomic analyses and polyphasic taxonomic characterization we propose reclassification of the species B. thermotolerans into a novel genus named Quasibacillus thermotolerans gen. nov., comb. nov with the type strain SgZ-8 T (=CCTCC AB2012108 T =KACC 16706 T ). Further our analyses also revealed that B. encimensis SGD-V-25 T is a later heterotypic synonym of Bacillus badius DSM 23 T . Copyright © 2017 Elsevier GmbH. All rights reserved.

Construction and Validation of the Rhodobacter sphaeroides 2.4.1 DNA Microarray: Transcriptome Flexibility at Diverse Growth Modes

PubMed Central

Pappas, Christopher T.; Sram, Jakub; Moskvin, Oleg V.; Ivanov, Pavel S.; Mackenzie, R. Christopher; Choudhary, Madhusudan; Land, Miriam L.; Larimer, Frank W.; Kaplan, Samuel; Gomelsky, Mark

2004-01-01

A high-density oligonucleotide DNA microarray, a genechip, representing the 4.6-Mb genome of the facultative phototrophic proteobacterium, Rhodobacter sphaeroides 2.4.1, was custom-designed and manufactured by Affymetrix, Santa Clara, Calif. The genechip contains probe sets for 4,292 open reading frames (ORFs), 47 rRNA and tRNA genes, and 394 intergenic regions. The probe set sequences were derived from the genome annotation generated by Oak Ridge National Laboratory after extensive revision, which was based primarily upon codon usage characteristic of this GC-rich bacterium. As a result of the revision, numerous missing ORFs were uncovered, nonexistent ORFs were deleted, and misidentified start codons were corrected. To evaluate R. sphaeroides transcriptome flexibility, expression profiles for three diverse growth modes—aerobic respiration, anaerobic respiration in the dark, and anaerobic photosynthesis—were generated. Expression levels of one-fifth to one-third of the R. sphaeroides ORFs were significantly different in cells under any two growth modes. Pathways involved in energy generation and redox balance maintenance under three growth modes were reconstructed. Expression patterns of genes involved in these pathways mirrored known functional changes, suggesting that massive changes in gene expression are the major means used by R. sphaeroides in adaptation to diverse conditions. Differential expression was observed for genes encoding putative new participants in these pathways (additional photosystem genes, duplicate NADH dehydrogenase, ATP synthases), whose functionality has yet to be investigated. The DNA microarray data correlated well with data derived from quantitative reverse transcription-PCR, as well as with data from the literature, thus validating the R. sphaeroides genechip as a powerful and reliable tool for studying unprecedented metabolic versatility of this bacterium. PMID:15231807

Use of a Hierarchical Oligonucleotide Primer Extension Approach for Multiplexed Relative Abundance Analysis of Methanogens in Anaerobic Digestion Systems

PubMed Central

Chuang, Hui-Ping; Hsu, Mao-Hsuan; Chen, Wei-Yu

2013-01-01

In this study, we established a rapid multiplex method to detect the relative abundances of amplified 16S rRNA genes from known cultivatable methanogens at hierarchical specificities in anaerobic digestion systems treating industrial wastewater and sewage sludge. The method was based on the hierarchical oligonucleotide primer extension (HOPE) technique and combined with a set of 27 primers designed to target the total archaeal populations and methanogens from 22 genera within 4 taxonomic orders. After optimization for their specificities and detection sensitivity under the conditions of multiple single-nucleotide primer extension reactions, the HOPE approach was applied to analyze the methanogens in 19 consortium samples from 7 anaerobic treatment systems (i.e., 513 reactions). Among the samples, the methanogen populations detected with order-level primers accounted for >77.2% of the PCR-amplified 16S rRNA genes detected using an Archaea-specific primer. The archaeal communities typically consisted of 2 to 7 known methanogen genera within the Methanobacteriales, Methanomicrobiales, and Methanosarcinales and displayed population dynamic and spatial distributions in anaerobic reactor operations. Principal component analysis of the HOPE data further showed that the methanogen communities could be clustered into 3 distinctive groups, in accordance with the distribution of the Methanosaeta, Methanolinea, and Methanomethylovorans, respectively. This finding suggested that in addition to acetotrophic and hydrogenotrophic methanogens, the methylotrophic methanogens might play a key role in the anaerobic treatment of industrial wastewater. Overall, the results demonstrated that the HOPE approach is a specific, rapid, and multiplexing platform to determine the relative abundances of targeted methanogens in PCR-amplified 16S rRNA gene products. PMID:24077716

Mucilaginibacter antarcticus sp. nov., isolated from tundra soil.

PubMed

Zheng, Ruichen; Zhao, Yiming; Wang, Liqiu; Chang, Xulu; Zhang, Yumin; Da, Xuyang; Peng, Fang

2016-12-01

The novel, pale yellow bacterial strain, designated S14-88T, was isolated from a tundra soil near Antarctic Peninsula, South Shetland Islands, and its taxonomic position was investigated by a genotypic and phenotypic analysis. Cells were facultatively anaerobic, Gram-stain-negative, non-motile and rod-shaped. Growth occurred at 4-28 °C (optimum at 15 °C), at pH 7.0-8.0 (optimum at 7.0) and with 0-0.6 % (w/v) NaCl (optimum, no NaCl). Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain S14-88T formed a lineage within the genus Mucilaginibacter. The 16S rRNA gene sequence similarity between strain S14-88T and the type strains of related species ranged from 92.2 to 96.5 %, and the 16S rRNA gene sequence of S14-88T showed highest similarity of 96.5 % to Mucilaginibacter soyangensis HME6664T. The major cellular fatty acids of strain S14-88T were iso-C15 : 0 and summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c). The major respiratory quinone was menaquinone MK-7, and the main polar lipid was phosphatidylethanolamine. The DNA G+C content of strain S14-88T was 42.3 mol%. On the basis of the evidence presented in this study, strain S14-88T is considered to represent a novel species of the genus Mucilaginibacter, for which the name Mucilaginibacter antarcticus sp. nov. is proposed. The type strain is S14-88T (=CCTCC AB 2015321T=KCTC 52232T).

Bacterial communities in soil samples from the Mingyong Glacier of southwestern China.

PubMed

Li, Haoyu; Taj, Muhammad Kamran; Ji, Xiuling; Zhang, Qi; Lin, Liangbing; Zhou, Zhimei; Wei, Yunlin

2017-05-01

The present study was an effort to determine the bacterial diversity of soils in Mingyong Glacier located at the Meili Snow Mountains of southwestern China. Mingyong Glacier has different climatic zones within a very narrow area, and bacterial community diversity in this low temperature area remains largely unknown. In this study, soil samples were collected from four different climatic zones: M11A (dry warm valley), M14 (forest), M15 (grass land), and M16 (glacier zones). Phylogenetic analysis based on 16S rRNA gene V6 hypervariable region showed high bacterial abundance in the glacier. The number of Operational Taxonomic Units ranged from 2.24×10 3 to 5.56×10 3 in soil samples. Statistical analysis of 16S rRNA gene clone libraries results showed that bacterial diversity in zones M11A，M14 and M16 are higher than in zone M15. The bacterial community structures are clearly distinguishable, and phylogenetic analysis showed that the predominant phyla were Proteobacteria, Deinococcus-Thermus, Firmicutes, Actinobacteria, and Nitrospirae in Mingyong Glacier. Seventy-nine different orders from four zones have been isolated. Bacterial diversity and distribution of bacterial communities related to the anthropogenic perturbations in zone (M15) were confirmed by diversity index analysis, and the diversity index of other three zones was satisfactory through this analysis software. The results suggest that bacterial diversity and distribution analyses using bacterial 16S rRNA gene V6 hypervariable region were successful, and bacterial communities in this area not only had the same bacterial phyla compared to other glaciers but also had their own rare species.

Luteibacter jiangsuensis sp. nov.: a methamidophos-degrading bacterium isolated from a methamidophos-manufacturing factory.

PubMed

Wang, Li; Wang, Guang-li; Li, Shun-peng; Jiang, Jian-dong

2011-01-01

A Gram-stain-negative, non-motile, rod-shaped bacterial strain, JW-64-1(T), capable of degrading methamidophos was isolated from a methamidophos-manufacturing factory in China, and was subjected to a polyphasic taxonomic investigation. Strain JW-64-1(T) produced circular, smooth, transparent, yellow-colored colonies (1.0-2.0 mm) on LB agar after 2 days incubation. It grew optimally at 25-30°C and pH 7.0 without the presence of NaCl. The G+C content of the total DNA was 63.6 mol%. A phylogenetic analysis based on 16S rRNA gene sequences showed that strain JW-64-1(T) fell within the cluster comprising Luteibacter species. The 16S rRNA gene sequence of strain JW-64-1(T) was most closely related to Luteibacter rhizovicinus DSM 16549(T) (98.6%), followed by Luteibacter yeojuensis DSM 17673(T) (98.4%) and L. anthropi CCUG 25036(T) (98.2%). The major cellular fatty acids of strain JW-64-1(T) were iso-C(15:0) (24.1%), iso-C(17:0) (20.2%) and summed feature 9 comprising iso-C(17:1) ω9c and/or C(16:0) 10-methyl (20.3%). The major isoprenoid quinine was Q-8 (98%), and the major polar lipids were diphosphatidylglycerol, phosphatidylglycerol, phosphoaminolipid, aminolipids-1, aminolipids-2, and phospholipids. The values for DNA-DNA relatedness between strain JW-64-1(T) and the closest phylogenetic relatives of L. rhizovicinus and Luteibacter yeojuensis were 34.8 ± 2.6 and 25.6 ± 3.1%, respectively. On the basis of the phenotypic, chemotaxonomic, DNA-DNA relatedness and phylogenetic analysis based on the 16S rRNA gene sequences, strain JW-64-1(T) represents a novel species of the genus Luteibacter, for which the name Luteibacter jiangsuensis sp. nov. is proposed. The type strain is JW-64-1(T) (=CGMCC 1.10133(T) = DSM 22396(T)).

The Functional Implications of Bottom Up and Top Down Controls on Marine Bacteria in Arthur Harbor, a Highly Productive Coastal Setting on the West Antarctic Peninsula

NASA Astrophysics Data System (ADS)

Bowman, J. S.; Amaral-Zettler, L. A.; Rich, J. J.; Luria, C.; Ducklow, H. W.

2016-02-01

Marine bacteria can be broadly classified into two groups based on their ecology; slow growing oligotrophic specialists and fast growing copiotrophs. These ecological strategies are associated with specific taxonomic and functional groups, making it possible to use 16S rRNA gene amplicon and shotgun metagenomic data to qualitatively, and possibly quantitatively, identify the contribution of each strategy to marine biogeochemical cycles. We leveraged a 5-year (2009 to 2014) time series of 16S rRNA gene amplicon data for Arthur Harbor, located near Palmer Station, Antarctica, to identify trends in the abundance of taxa associated with each ecological strategy. Using emergent self-organizing maps, we identified four recurring "modes" in bacterial community structure based on the relative abundance of the ubiquitous SAR11 clade. A different bacterial genus was dominant in each mode; Pelagibacter, Polaribacter, Roseobacter, and Colwellia. To explore the functional implications of these different modes we applied shotgun metagenomics and functional predictions using the newly available tool PAPRICA, in combination with flow cytometry and estimates of bacterial production. Our annotation and assembly of binned contigs corresponding to the dominant genera illuminate the succession of metabolic functions across the 2013-2014 austral summer and inform the timing of autotrophic and mixotrophic (putatively bacterivorous) phytoplankton blooms. Surprisingly, while the abundance of Pelagibacter 16S rRNA gene reads was negatively correlated with the concentration of chlorophyll a, the ratio of Pelagibacter to Polaribacter and Roseobacter was poorly correlated with the ratio of high nucleic acid (HNA) to low nucleic acid (LNA) bacteria as determined by flow cytometry, and the relative size of the HNA population at times contrasted sharply with chlorophyll a. These findings suggest that the physiological state of bacterial cells and top down controls play a strong role in HNA: LNA dynamics.

Bacterial microbiome in the nose of healthy cats and in cats with nasal disease

PubMed Central

Tress, Barbara; Suchodolski, Jan S.; Nisar, Tariq; Ravindran, Prajesh; Weber, Karin; Hartmann, Katrin; Schulz, Bianka S.

2017-01-01

Background Traditionally, changes in the microbial population of the nose have been assessed using conventional culture techniques. Sequencing of bacterial 16S rRNA genes demonstrated that the human nose is inhabited by a rich and diverse bacterial microbiome that cannot be detected using culture-based methods. The goal of this study was to describe the nasal microbiome of healthy cats, cats with nasal neoplasia, and cats with feline upper respiratory tract disease (FURTD). Methodology/Principal findings DNA was extracted from nasal swabs of healthy cats (n = 28), cats with nasal neoplasia (n = 16), and cats with FURTD (n = 15), and 16S rRNA genes were sequenced. High species richness was observed in all samples. Rarefaction analysis revealed that healthy cats living indoors had greater species richness (observed species p = 0.042) and Shannon diversity (p = 0.003) compared with healthy cats living outdoors. Higher species richness (observed species p = 0.001) and Shannon diversity (p<0.001) were found in middle-aged cats in comparison to healthy cats in different age groups. Principal coordinate analysis revealed separate clustering based on similarities in bacterial molecular phylogenetic trees of 16S rRNA genes for indoor and outdoor cats. In all groups examined, the most abundant phyla identified were Proteobacteria, Firmicutes, and Bacteroidetes. At the genus level, 375 operational taxonomic units (OTUs) were identified. In healthy cats and cats with FURTD, Moraxella spp. was the most common genus, while it was unclassified Bradyrhizobiaceae in cats with nasal neoplasia. High individual variability was observed. Conclusion This study demonstrates that the nose of cats is inhabited by much more variable and diverse microbial communities than previously shown. Future research in this field might help to develop new diagnostic tools to easily identify nasal microbial changes, relate them to certain disease processes, and help clinicians in the decision process of antibiotic selection for individual patients. PMID:28662139

Natrinema gari sp. nov., a halophilic archaeon isolated from fish sauce in Thailand.

PubMed

Tapingkae, Wanaporn; Tanasupawat, Somboon; Itoh, Takashi; Parkin, Kirk L; Benjakul, Soottawat; Visessanguan, Wonnop; Valyasevi, Ruud

2008-10-01

Two Gram-negative, rod-shaped, halophilic archaea, designated strains HIS40-3(T) and HDS3-1, were isolated from anchovy fish sauce (nam-pla) collected from two different locations in Thailand. The two strains were able to grow at 20-60 degrees C (optimum 37-40 degrees C), at 1.7-5.1 M NaCl (optimum 2.6-3.4 M NaCl) and at pH 5.5-8.5 (optimum pH 6.0-6.5). Hypotonic treatment with less than 1.7 M NaCl caused cell lysis. The major polar lipids of the isolates were C(20)C(20) and C(20)C(25) derivatives of phosphatidylglycerol, phosphatidylglycerol phosphate methyl ester, phosphatidylglycerol sulfate, two glycolipids and one unidentified lipid. The DNA G+C contents were 64.0-65.4 mol%. In addition to phenotypic and chemotaxonomic characteristics, phylogenetic analysis based on 16S rRNA gene sequence similarities showed that strains HIS40-3(T) and HDS3-1 were related most closely to species of the genus Natrinema. Levels of 16S rRNA gene sequence similarity between strains HIS40-3(T) and HDS3-1 and the type strains of recognized Natrinema species were 99.1-96.6 %. The two novel strains could be distinguished from recognized Natrinema species on the basis of low levels of DNA-DNA relatedness and differences in whole-cell protein patterns and phenotypic properties. Levels of 16S rRNA gene sequence similarity and DNA-DNA relatedness between the two strains were 99.7 and 77.7 %, respectively, suggesting that they should be classified as representing a single species. Based on these taxonomic data, strains HIS40-3(T) and HDS3-1 are considered to represent a novel species of the genus Natrinema, for which the name Natrinema gari sp. nov. is proposed. The type strain is HIS40-3(T) (=BCC 24370(T) =JCM 14663(T) =PCU 303(T)).

Colorectal Cancer and the Human Gut Microbiome: Reproducibility with Whole-Genome Shotgun Sequencing

PubMed Central

Hua, Xing; Zeller, Georg; Sunagawa, Shinichi; Voigt, Anita Y.; Hercog, Rajna; Goedert, James J.; Shi, Jianxin; Bork, Peer; Sinha, Rashmi

2016-01-01

Accumulating evidence indicates that the gut microbiota affects colorectal cancer development, but previous studies have varied in population, technical methods, and associations with cancer. Understanding these variations is needed for comparisons and for potential pooling across studies. Therefore, we performed whole-genome shotgun sequencing on fecal samples from 52 pre-treatment colorectal cancer cases and 52 matched controls from Washington, DC. We compared findings from a previously published 16S rRNA study to the metagenomics-derived taxonomy within the same population. In addition, metagenome-predicted genes, modules, and pathways in the Washington, DC cases and controls were compared to cases and controls recruited in France whose specimens were processed using the same platform. Associations between the presence of fecal Fusobacteria, Fusobacterium, and Porphyromonas with colorectal cancer detected by 16S rRNA were reproduced by metagenomics, whereas higher relative abundance of Clostridia in cancer cases based on 16S rRNA was merely borderline based on metagenomics. This demonstrated that within the same sample set, most, but not all taxonomic associations were seen with both methods. Considering significant cancer associations with the relative abundance of genes, modules, and pathways in a recently published French metagenomics dataset, statistically significant associations in the Washington, DC population were detected for four out of 10 genes, three out of nine modules, and seven out of 17 pathways. In total, colorectal cancer status in the Washington, DC study was associated with 39% of the metagenome-predicted genes, modules, and pathways identified in the French study. More within and between population comparisons are needed to identify sources of variation and disease associations that can be reproduced despite these variations. Future studies should have larger sample sizes or pool data across studies to have sufficient power to detect associations that are reproducible and significant after correction for multiple testing. PMID:27171425

Phylogenetic analysis of nitrite, nitric oxide, and nitrous oxide respiratory enzymes reveal a complex evolutionary history for denitrification.

PubMed

Jones, Christopher M; Stres, Blaz; Rosenquist, Magnus; Hallin, Sara

2008-09-01

Denitrification is a facultative respiratory pathway in which nitrite (NO2(-)), nitric oxide (NO), and nitrous oxide (N2O) are successively reduced to nitrogen gas (N(2)), effectively closing the nitrogen cycle. The ability to denitrify is widely dispersed among prokaryotes, and this polyphyletic distribution has raised the possibility of horizontal gene transfer (HGT) having a substantial role in the evolution of denitrification. Comparisons of 16S rRNA and denitrification gene phylogenies in recent studies support this possibility; however, these results remain speculative as they are based on visual comparisons of phylogenies from partial sequences. We reanalyzed publicly available nirS, nirK, norB, and nosZ partial sequences using Bayesian and maximum likelihood phylogenetic inference. Concomitant analysis of denitrification genes with 16S rRNA sequences from the same organisms showed substantial differences between the trees, which were supported by examining the posterior probability of monophyletic constraints at different taxonomic levels. Although these differences suggest HGT of denitrification genes, the presence of structural variants for nirK, norB, and nosZ makes it difficult to determine HGT from other evolutionary events. Additional analysis using phylogenetic networks and likelihood ratio tests of phylogenies based on full-length sequences retrieved from genomes also revealed significant differences in tree topologies among denitrification and 16S rRNA gene phylogenies, with the exception of the nosZ gene phylogeny within the data set of the nirK-harboring genomes. However, inspection of codon usage and G + C content plots from complete genomes gave no evidence for recent HGT. Instead, the close proximity of denitrification gene copies in the genomes of several denitrifying bacteria suggests duplication. Although HGT cannot be ruled out as a factor in the evolution of denitrification genes, our analysis suggests that other phenomena, such gene duplication/divergence and lineage sorting, may have differently influenced the evolution of each denitrification gene.

Electing a candidate: a speculative history of the bacterial phylum OP10.

PubMed

Dunfield, Peter F; Tamas, Ivica; Lee, Kevin C; Morgan, Xochitl C; McDonald, Ian R; Stott, Matthew B

2012-12-01

In 1998, a cultivation-independent survey of the microbial community in Obsidian Pool, Yellowstone National Park, detected 12 new phyla within the Domain Bacteria. These were dubbed 'candidate divisions' OP1 to OP12. Since that time the OP10 candidate division has been commonly detected in various environments, usually as part of the rare biosphere, but occasionally as a predominant community component. Based on 16S rRNA gene phylogeny, OP10 comprises at least 12 class-level subdivisions. However, despite this broad ecological and evolutionary diversity, all OP10 bacteria have eluded cultivation until recently. In 2011, two reference species of OP10 were taxonomically validated, removing the phylum from its 'candidate' status. Construction of a highly resolved phylogeny based on 29 universally conserved genes verifies its standing as a unique bacterial phylum. In the following paper we summarize what is known and what is suspected about the newest described bacterial phylum, the Armatimonadetes. © 2012 Society for Applied Microbiology and Blackwell Publishing Ltd.

Staphylococcus petrasii subsp. pragensis subsp. nov., occurring in human clinical material.

PubMed

Švec, Pavel; De Bel, Annelies; Sedláček, Ivo; Petráš, Petr; Gelbíčová, Tereza; Černohlávková, Jitka; Mašlanˇová, Ivana; Cnockaert, Margo; Varbanovová, Ivana; Echahidi, Fedoua; Vandamme, Peter; Pantuček, Roman

2015-07-01

Seven coagulase-negative, oxidase-negative and novobiocin-susceptible staphylococci assigned tentatively as Staphylococcus petrasii were investigated in this study in order to elucidate their taxonomic position. All strains were initially shown to form a genetically homogeneous group separated from remaining species of the genus Staphylococcus by using a repetitive sequence-based PCR fingerprinting with the (GTG)5 primer. Phylogenetic analysis based on 16S rRNA gene, hsp60, rpoB, dnaJ, gap and tuf sequences showed that the group is closely related to Staphylococcus petrasii but separated from the three hitherto known subspecies, S. petrasii subsp. petrasii, S. petrasii subsp. croceilyticus and S. petrasii subsp. jettensis. Further investigation using automated ribotyping, MALDI-TOF mass spectrometry, fatty acid methyl ester analysis, DNA-DNA hybridization and extensive biotyping confirmed that the analysed group represents a novel subspecies within S. petrasii, for which the name Staphylococcus petrasii subsp. pragensis subsp. nov. is proposed. The type strain is NRL/St 12/356(T) ( = CCM 8529(T) = LMG 28327(T)).

Effect of land use and soil organic matter quality on the structure and function of microbial communities in pastoral soils: Implications for disease suppression

PubMed Central

O’Callaghan, Maureen; Condron, Leo M.; Kowalchuk, George A.; Van Nostrand, Joy D.; Zhou, Jizhong; Wakelin, Steven A.

2018-01-01

Cropping soils vary in extent of natural suppression of soil-borne plant diseases. However, it is unknown whether similar variation occurs across pastoral agricultural systems. We examined soil microbial community properties known to be associated with disease suppression across 50 pastoral fields varying in management intensity. The composition and abundance of the disease-suppressive community were assessed from both taxonomic and functional perspectives. Pseudomonas bacteria were selected as a general taxonomic indicator of disease suppressive potential, while genes associated with the biosynthesis of a suite of secondary metabolites provided functional markers (GeoChip 5.0 microarray analysis). The composition of both the Pseudomonas communities and disease suppressive functional genes were responsive to land use. Underlying soil properties explained 37% of the variation in Pseudomonas community structure and up to 61% of the variation in the abundance of disease suppressive functional genes. Notably, measures of soil organic matter quality, C:P ratio, and aromaticity of the dissolved organic matter content (carbon recalcitrance), influenced both the taxonomic and functional disease suppressive potential of the pasture soils. Our results suggest that key components of the soil microbial community may be managed on-farm to enhance disease suppression and plant productivity. PMID:29734390

Effect of land use and soil organic matter quality on the structure and function of microbial communities in pastoral soils: Implications for disease suppression.

PubMed

Dignam, Bryony E A; O'Callaghan, Maureen; Condron, Leo M; Kowalchuk, George A; Van Nostrand, Joy D; Zhou, Jizhong; Wakelin, Steven A

2018-01-01

Cropping soils vary in extent of natural suppression of soil-borne plant diseases. However, it is unknown whether similar variation occurs across pastoral agricultural systems. We examined soil microbial community properties known to be associated with disease suppression across 50 pastoral fields varying in management intensity. The composition and abundance of the disease-suppressive community were assessed from both taxonomic and functional perspectives. Pseudomonas bacteria were selected as a general taxonomic indicator of disease suppressive potential, while genes associated with the biosynthesis of a suite of secondary metabolites provided functional markers (GeoChip 5.0 microarray analysis). The composition of both the Pseudomonas communities and disease suppressive functional genes were responsive to land use. Underlying soil properties explained 37% of the variation in Pseudomonas community structure and up to 61% of the variation in the abundance of disease suppressive functional genes. Notably, measures of soil organic matter quality, C:P ratio, and aromaticity of the dissolved organic matter content (carbon recalcitrance), influenced both the taxonomic and functional disease suppressive potential of the pasture soils. Our results suggest that key components of the soil microbial community may be managed on-farm to enhance disease suppression and plant productivity.

Application of Stochastic Labeling with Random-Sequence Barcodes for Simultaneous Quantification and Sequencing of Environmental 16S rRNA Genes.

PubMed

Hoshino, Tatsuhiko; Inagaki, Fumio

2017-01-01

Next-generation sequencing (NGS) is a powerful tool for analyzing environmental DNA and provides the comprehensive molecular view of microbial communities. For obtaining the copy number of particular sequences in the NGS library, however, additional quantitative analysis as quantitative PCR (qPCR) or digital PCR (dPCR) is required. Furthermore, number of sequences in a sequence library does not always reflect the original copy number of a target gene because of biases caused by PCR amplification, making it difficult to convert the proportion of particular sequences in the NGS library to the copy number using the mass of input DNA. To address this issue, we applied stochastic labeling approach with random-tag sequences and developed a NGS-based quantification protocol, which enables simultaneous sequencing and quantification of the targeted DNA. This quantitative sequencing (qSeq) is initiated from single-primer extension (SPE) using a primer with random tag adjacent to the 5' end of target-specific sequence. During SPE, each DNA molecule is stochastically labeled with the random tag. Subsequently, first-round PCR is conducted, specifically targeting the SPE product, followed by second-round PCR to index for NGS. The number of random tags is only determined during the SPE step and is therefore not affected by the two rounds of PCR that may introduce amplification biases. In the case of 16S rRNA genes, after NGS sequencing and taxonomic classification, the absolute number of target phylotypes 16S rRNA gene can be estimated by Poisson statistics by counting random tags incorporated at the end of sequence. To test the feasibility of this approach, the 16S rRNA gene of Sulfolobus tokodaii was subjected to qSeq, which resulted in accurate quantification of 5.0 × 103 to 5.0 × 104 copies of the 16S rRNA gene. Furthermore, qSeq was applied to mock microbial communities and environmental samples, and the results were comparable to those obtained using digital PCR and relative abundance based on a standard sequence library. We demonstrated that the qSeq protocol proposed here is advantageous for providing less-biased absolute copy numbers of each target DNA with NGS sequencing at one time. By this new experiment scheme in microbial ecology, microbial community compositions can be explored in more quantitative manner, thus expanding our knowledge of microbial ecosystems in natural environments.

Nitrification of archaeal ammonia oxidizers in a high- temperature hot spring

NASA Astrophysics Data System (ADS)

Chen, Shun; Peng, Xiaotong; Xu, Hengchao; Ta, Kaiwen

2016-04-01

The oxidation of ammonia by microbes has been shown to occur in diverse natural environments. However, the link of in situ nitrification activity to taxonomic identities of ammonia oxidizers in high-temperature environments remains poorly understood. Here, we studied in situ ammonia oxidation rates and the diversity of ammonia-oxidizing Archaea (AOA) in surface and bottom sediments at 77 °C in the Gongxiaoshe hot spring, Tengchong, Yunnan, China. The in situ ammonia oxidation rates measured by the 15N-NO3- pool dilution technique in the surface and bottom sediments were 4.80 and 5.30 nmol N g-1 h-1, respectively. Real-time quantitative polymerase chain reaction (qPCR) indicated that the archaeal 16S rRNA genes and amoA genes were present in the range of 0.128 to 1.96 × 108 and 2.75 to 9.80 × 105 gene copies g-1 sediment, respectively, while bacterial amoA was not detected. Phylogenetic analysis of 16S rRNA genes showed high sequence similarity to thermophilic Candidatus Nitrosocaldus yellowstonii, which represented the most abundant operational taxonomic units (OTU) in both surface and bottom sediments. The archaeal predominance was further supported by fluorescence in situ hybridization (FISH) visualization. The cell-specific rate of ammonia oxidation was estimated to range from 0.410 to 0.790 fmol N archaeal cell-1 h-1, higher than those in the two US Great Basin hot springs. These results suggest the importance of archaeal rather than bacterial ammonia oxidation in driving the nitrogen cycle in terrestrial geothermal environments.

Streptococcal Diversity of Human Milk and Comparison of Different Methods for the Taxonomic Identification of Streptococci.

PubMed

Martín, Virginia; Mediano, Pilar; Del Campo, Rosa; Rodríguez, Juan M; Marín, María

2016-11-01

The genus Streptococcus is 1 of the dominant bacterial groups in human milk, but the taxonomic identification of some species remains difficult. The objective of this study was to investigate the discriminatory ability of different methods to identify streptococcal species in order to perform an assessment of the streptococcal diversity of human milk microbiota as accurately as possible. The identification of 105 streptococcal strains from human milk was performed by 16S rRNA, tuf, and sodA gene sequencing, phylogenetic analysis, and Matrix Assisted Laser Desorption Ionization-Time of Flight (MALDI-TOF) mass spectrometry. Streptococcus salivarius, Streptococcus mitis, and Streptococcus parasanguinis were the streptococcal dominant species in the human milk microbiota. Sequencing of housekeeping genes allowed the classification of 96.2% (16S rRNA), 84.8% ( sodA), and 88.6% ( tuf) of the isolates. Phylogenetic analysis showed 3 main streptococcal clusters corresponding with the mitis (73 isolates), salivarius (29), mutans (1)-pyogenic (2) groups, but many of the mitis group isolates (36) could not be assigned to any species. The application of the MALDI-TOF Bruker Biotyper system resulted in the identification of 56 isolates (53.33%) at the species level, but it could not discriminate between S pneumoniae and S mitis isolates, in contrast to the Vitek-MS system. There was a good agreement among the different methods assessed in this study to identify those isolates of the salivarius, mutans, and pyogenic groups, whereas unambiguous discrimination could not be achieved concerning some species of the mitis group ( S mitis, S pneumoniae, S pseudopneumoniae, S oralis).

Reappraisal of the taxonomy of Streptococcus suis serotypes 20, 22 and 26: Streptococcus parasuis sp. nov.

PubMed

Nomoto, R; Maruyama, F; Ishida, S; Tohya, M; Sekizaki, T; Osawa, Ro

2015-02-01

In order to clarify the taxonomic position of serotypes 20, 22 and 26 of Streptococcus suis, biochemical and molecular genetic studies were performed on isolates (SUT-7, SUT-286(T), SUT-319, SUT-328 and SUT-380) reacted with specific antisera of serotypes 20, 22 or 26 from the saliva of healthy pigs as well as reference strains of serotypes 20, 22 and 26. Comparative recN gene sequencing showed high genetic relatedness among our isolates, but marked differences from the type strain S. suis NCTC 10234(T), i.e. 74.8-75.7 % sequence similarity. The genomic relatedness between the isolates and other strains of species of the genus Streptococcus, including S. suis, was calculated using the average nucleotide identity values of whole genome sequences, which indicated that serotypes 20, 22 and 26 should be removed taxonomically from S. suis and treated as a novel genomic species. Comparative sequence analysis revealed 99.0-100 % sequence similarities for the 16S rRNA genes between the reference strains of serotypes 20, 22 and 26, and our isolates. Isolate STU-286(T) had relatively high 16S rRNA gene sequence similarity with S. suis NCTC 10234(T) (98.8 %). SUT-286(T) could be distinguished from S. suis and other closely related species of the genus Streptococcus using biochemical tests. Due to its phylogenetic and phenotypic similarities to S. suis we propose naming the novel species Streptococcus parasuis sp. nov., with SUT-286(T) ( = JCM 30273(T) = DSM 29126(T)) as the type strain. © 2015 IUMS.

Pyrosequencing analysis of free-living and attached bacterial communities in Meiliang Bay, Lake Taihu, a large eutrophic shallow lake in China.

PubMed

Tang, Xiangming; Li, Linlin; Shao, Keqiang; Wang, Boweng; Cai, Xianlei; Zhang, Lei; Chao, Jianying; Gao, Guang

2015-01-01

To elucidate the relationship between particle-attached (PA, ≥ 5.0 μm) and free-living (FL, 0.2-5.0 μm) bacterial communities, samplings were collected seasonally from November 2011 to August 2012 in Meiliang Bay, Lake Taihu, China. We used 454 pyrosequencing of 16S rRNA genes to study bacterial diversity and structure of PA and FL communities. The analysis rendered 37,985 highly qualified reads, subsequently assigned to 1755 operational taxonomic units (97% similarity) for the 8 samples. Although 27 high-level taxonomic groups were obtained, the 3 dominant phyla (Proteobacteria, Actinobacteria, and Bacteroidetes) comprised about 75.9% and 82.4% of the PA and FL fractions, respectively. Overall, we found no significant differences between community types, as indicated by ANOSIM R statistics (R = 0.063, P > 0.05) and the Parsimony test (P = 0.222). Dynamics of bacterial communities were correlated with changes in concentrations of total suspended solids (TSS) and total phosphorus (TP). In summer, a significant taxonomic overlap in the 2 size fractions was observed when Cyanobacteria, a major contributor of TSS and TP, dominated in the water, highlighting the potential rapid exchange between PA and FL bacterial populations in large shallow eutrophic lakes.

Microbial biogeography of arctic streams: exploring influences of lithology and habitat.

PubMed

Larouche, Julia R; Bowden, William B; Giordano, Rosanna; Flinn, Michael B; Crump, Byron C

2012-01-01

Terminal restriction fragment length polymorphism and 16S rRNA gene sequencing were used to explore the community composition of bacterial communities in biofilms on sediments (epipssamon) and rocks (epilithon) in stream reaches that drain watersheds with contrasting lithologies in the Noatak National Preserve, Alaska. Bacterial community composition varied primarily by stream habitat and secondarily by lithology. Positive correlations were detected between bacterial community structure and nutrients, base cations, and dissolved organic carbon. Our results showed significant differences at the stream habitat, between epipssamon and epilithon bacterial communities, which we expected. Our results also showed significant differences at the landscape scale that could be related to different lithologies and associated stream biogeochemistry. These results provide insight into the bacterial community composition of little known and pristine arctic stream ecosystems and illustrate how differences in the lithology, soils, and vegetation community of the terrestrial environment interact to influence stream bacterial taxonomic richness and composition.

Microbial Biogeography of Arctic Streams: Exploring Influences of Lithology and Habitat

PubMed Central

Larouche, Julia R.; Bowden, William B.; Giordano, Rosanna; Flinn, Michael B.; Crump, Byron C.

2012-01-01

Terminal restriction fragment length polymorphism and 16S rRNA gene sequencing were used to explore the community composition of bacterial communities in biofilms on sediments (epipssamon) and rocks (epilithon) in stream reaches that drain watersheds with contrasting lithologies in the Noatak National Preserve, Alaska. Bacterial community composition varied primarily by stream habitat and secondarily by lithology. Positive correlations were detected between bacterial community structure and nutrients, base cations, and dissolved organic carbon. Our results showed significant differences at the stream habitat, between epipssamon and epilithon bacterial communities, which we expected. Our results also showed significant differences at the landscape scale that could be related to different lithologies and associated stream biogeochemistry. These results provide insight into the bacterial community composition of little known and pristine arctic stream ecosystems and illustrate how differences in the lithology, soils, and vegetation community of the terrestrial environment interact to influence stream bacterial taxonomic richness and composition. PMID:22936932

Taxonomic study of extreme halophilic archaea isolated from the "Salar de Atacama", Chile.

PubMed

Lizama, C; Monteoliva-Sánchez, M; Prado, B; Ramos-Cormenzana, A; Weckesser, J; Campos, V

2001-11-01

A large number of halophilic bacteria were isolated in 1984-1992 from the Atacama Saltern (North of Chile). For this study 82 strains of extreme halophilic archaea were selected. The characterization was performed by using the phenotypic characters including morphological, physiological, biochemical, nutritional and antimicrobial susceptibility test. The results, together with those from reference strains, were subjected to numerical analysis, using the Simple Matching (S(SM)) coefficient and clustered by the unweighted pair group method of association (UPGMA). Fifteen phena were obtained at an 70% similarity level. The results obtained reveal a high diversity among the halophilic archaea isolated. Representative strains from the phena were chosen to determine their DNA base composition and the percentage of DNA-DNA similarity compared to reference strains. The 16S rRNA studies showed that some of these strains constitutes a new taxa of extreme halophilic archaea.

Methods for understanding microbial community structures and functions in microbial fuel cells: a review.

PubMed

Zhi, Wei; Ge, Zheng; He, Zhen; Zhang, Husen

2014-11-01

Microbial fuel cells (MFCs) employ microorganisms to recover electric energy from organic matter. However, fundamental knowledge of electrochemically active bacteria is still required to maximize MFCs power output for practical applications. This review presents microbiological and electrochemical techniques to help researchers choose the appropriate methods for the MFCs study. Pre-genomic and genomic techniques such as 16S rRNA based phylogeny and metagenomics have provided important information in the structure and genetic potential of electrode-colonizing microbial communities. Post-genomic techniques such as metatranscriptomics allow functional characterizations of electrode biofilm communities by quantifying gene expression levels. Isotope-assisted phylogenetic analysis can further link taxonomic information to microbial metabolisms. A combination of electrochemical, phylogenetic, metagenomic, and post-metagenomic techniques offers opportunities to a better understanding of the extracellular electron transfer process, which in turn can lead to process optimization for power output. Copyright © 2014 Elsevier Ltd. All rights reserved.

Reclassification of Xuhuaishuia manganoxidans Wang et al. 2015 as a later heterotypic synonym of Brevirhabdus pacifica Wu et al. 2015 and emendation of the species description.

PubMed

Liu, Yang; Lai, Qiliang; Xu, Xue-Wei; Wu, Yue-Hong; Cheng, Hong; Zhang, Xiao-Hua; Wang, Long; Shao, Zongze

2017-08-01

A polyphasic taxonomic study was undertaken to clarify the exact position of type strain DY6-4T of Xuhuaishuia manganoxidans. A combination of physiological properties of X. manganoxidans DY6-4T was consistent with those of type strain 22DY15T of Brevirhabdus pacifica. The 16S rRNA gene sequence analyses indicated that X. manganoxidans DY6-4T and B. pacifica 22DY15T shared 100 % similarity and formed a monophyletic group. The close relationship between the two strains was underpinned by the results of chemotaxonomic characteristics, including the fatty acids, quinone and polar lipids. The digital DNA-DNA hybridization and average nucleotide identity values between the two strains were 99.90 and 99.98 %, respectively. Based on these results, we propose that Xuhuaishuia manganoxidans is a later heterotypic synonym of Brevirhabdus pacifica.

Enhancement of the microbial community biomass and diversity during air sparging bioremediation of a soil highly contaminated with kerosene and BTEX.

PubMed

Kabelitz, Nadja; Machackova, Jirina; Imfeld, Gwenaël; Brennerova, Maria; Pieper, Dietmar H; Heipieper, Hermann J; Junca, Howard

2009-03-01

In order to obtain insights in complexity shifts taking place in natural microbial communities under strong selective pressure, soils from a former air force base in the Czech Republic, highly contaminated with jet fuel and at different stages of a bioremediation air sparging treatment, were analyzed. By tracking phospholipid fatty acids and 16S rRNA genes, a detailed monitoring of the changes in quantities and composition of the microbial communities developed at different stages of the bioventing treatment progress was performed. Depending on the length of the air sparging treatment that led to a significant reduction in the contamination level, we observed a clear shift in the soil microbial community being dominated by Pseudomonads under the harsh conditions of high aromatic contamination to a status of low aromatic concentrations, increased biomass content, and a complex composition with diverse bacterial taxonomical branches.

The status of the species Enterobacter siamensisKhunthongpan et al. 2014. Request for an Opinion.

PubMed

Kämpfer, Peter; Doijad, Swapnil; Chakraborty, Trinad; Glaeser, Stefanie P

2016-01-01

In the course of a taxonomic study describing novel species of the genus Enterobacter it was found that the 16S rRNA gene sequence of the type strain of Enterobacter siamensis, obtained both directly from the authors of the publication on Enterobacter siamensis and from the Korean Collection for Type Cultures (C2361T and KCTC 23282T, respectively), was not congruent with the 16S rRNA gene sequence deposited in the GenBank database under the accession number HQ888848, which was applied for phylogenetic analysis in the species proposal. The remaining deposit in the Japanese type culture collection, NBRC 107138T, showed an identical 16S rRNA gene sequence to the other two cultures and overall, this sequence differed at 35 positions in comparison with the 1429 bp sequence published under the accession number HQ888848.Therefore, the type strain of this species cannot be included in any further scientific comparative study. It is proposed that the Judicial Commission of the International Committee on Systematics of Prokaryotes place the name Enterobacter siamensis on the list of rejected names, if a suitable replacement for the type strain is not found or a neotype strain is not proposed within two years following the publication of this Request for an Opinion.

Isolation and genetic characterization of Aurantimonas and Methylobacterium strains from stems of hypernodulated soybeans.

PubMed

Anda, Mizue; Ikeda, Seishi; Eda, Shima; Okubo, Takashi; Sato, Shusei; Tabata, Satoshi; Mitsui, Hisayuki; Minamisawa, Kiwamu

2011-01-01

The aims of this study were to isolate Aurantimonas and Methylobacterium strains that responded to soybean nodulation phenotypes and nitrogen fertilization rates in a previous culture-independent analysis (Ikeda et al. ISME J. 4:315-326, 2010). Two strategies were adopted for isolation from enriched bacterial cells prepared from stems of field-grown, hypernodulated soybeans: PCR-assisted isolation for Aurantimonas and selective cultivation for Methylobacterium. Thirteen of 768 isolates cultivated on Nutrient Agar medium were identified as Aurantimonas by colony PCR specific for Aurantimonas and 16S rRNA gene sequencing. Meanwhile, among 187 isolates on methanol-containing agar media, 126 were identified by 16S rRNA gene sequences as Methylobacterium. A clustering analysis (>99% identity) of the 16S rRNA gene sequences for the combined datasets of the present and previous studies revealed 4 and 8 operational taxonomic units (OTUs) for Aurantimonas and Methylobacterium, respectively, and showed the successful isolation of target bacteria for these two groups. ERIC- and BOX-PCR showed the genomic uniformity of the target isolates. In addition, phylogenetic analyses of Aurantimonas revealed a phyllosphere-specific cluster in the genus. The isolates obtained in the present study will be useful for revealing unknown legume-microbe interactions in relation to the autoregulation of nodulation.

Endophytic bacterial diversity in grapevine (Vitis vinifera L.) leaves described by 16S rRNA gene sequence analysis and length heterogeneity-PCR.

PubMed

Bulgari, Daniela; Casati, Paola; Brusetti, Lorenzo; Quaglino, Fabio; Brasca, Milena; Daffonchio, Daniele; Bianco, Piero Attilio

2009-08-01

Diversity of bacterial endophytes associated with grapevine leaf tissues was analyzed by cultivation and cultivation-independent methods. In order to identify bacterial endophytes directly from metagenome, a protocol for bacteria enrichment and DNA extraction was optimized. Sequence analysis of 16S rRNA gene libraries underscored five diverse Operational Taxonomic Units (OTUs), showing best sequence matches with gamma-Proteobacteria, family Enterobacteriaceae, with a dominance of the genus Pantoea. Bacteria isolation through cultivation revealed the presence of six OTUs, showing best sequence matches with Actinobacteria, genus Curtobacterium, and with Firmicutes genera Bacillus and Enterococcus. Length Heterogeneity-PCR (LH-PCR) electrophoretic peaks from single bacterial clones were used to setup a database representing the bacterial endophytes identified in association with grapevine tissues. Analysis of healthy and phytoplasma-infected grapevine plants showed that LH-PCR could be a useful complementary tool for examining the diversity of bacterial endophytes especially for diversity survey on a large number of samples.

Co-occurrence Networks Among Bacteria and Microbial Eukaryotes of Lake Baikal During a Spring Phytoplankton Bloom.

PubMed

Mikhailov, Ivan S; Zakharova, Yulia R; Bukin, Yuri S; Galachyants, Yuri P; Petrova, Darya P; Sakirko, Maria V; Likhoshway, Yelena V

2018-06-07

The pelagic zone of Lake Baikal is an ecological niche where phytoplankton bloom causes increasing microbial abundance in spring which plays a key role in carbon turnover in the freshwater lake. Co-occurrence patterns revealed among different microbes can be applied to predict interactions between the microbes and environmental conditions in the ecosystem. We used 454 pyrosequencing of 16S rRNA and 18S rRNA genes to study bacterial and microbial eukaryotic communities and their co-occurrence patterns at the pelagic zone of Lake Baikal during a spring phytoplankton bloom. We found that microbes within one domain mostly correlated positively with each other and are highly interconnected. The highly connected taxa in co-occurrence networks were operational taxonomic units (OTUs) of Actinobacteria, Bacteroidetes, Alphaproteobacteria, and autotrophic and unclassified Eukaryota which might be analogous to microbial keystone taxa. Constrained correspondence analysis revealed the relationships of bacterial and microbial eukaryotic communities with geographical location.

ELISA-BASE: An Integrated Bioinformatics Tool for Analyzing and Tracking ELISA Microarray Data

DOE Office of Scientific and Technical Information (OSTI.GOV)

White, Amanda M.; Collett, James L.; Seurynck-Servoss, Shannon L.

ELISA-BASE is an open-source database for capturing, organizing and analyzing protein enzyme-linked immunosorbent assay (ELISA) microarray data. ELISA-BASE is an extension of the BioArray Soft-ware Environment (BASE) database system, which was developed for DNA microarrays. In order to make BASE suitable for protein microarray experiments, we developed several plugins for importing and analyzing quantitative ELISA microarray data. Most notably, our Protein Microarray Analysis Tool (ProMAT) for processing quantita-tive ELISA data is now available as a plugin to the database.

A close-up view on ITS2 evolution and speciation - a case study in the Ulvophyceae (Chlorophyta, Viridiplantae)

PubMed Central

2011-01-01

Background The second Internal Transcriber Spacer (ITS2) is a fast evolving part of the nuclear-encoded rRNA operon located between the 5.8S and 28S rRNA genes. Based on crossing experiments it has been proposed that even a single Compensatory Base Change (CBC) in helices 2 and 3 of the ITS2 indicates sexual incompatibility and thus separates biological species. Taxa without any CBC in these ITS2 regions were designated as a 'CBC clade'. However, in depth comparative analyses of ITS2 secondary structures, ITS2 phylogeny, the origin of CBCs, and their relationship to biological species have rarely been performed. To gain 'close-up' insights into ITS2 evolution, (1) 86 sequences of ITS2 including secondary structures have been investigated in the green algal order Ulvales (Chlorophyta, Viridiplantae), (2) after recording all existing substitutions, CBCs and hemi-CBCs (hCBCs) were mapped upon the ITS2 phylogeny, rather than merely comparing ITS2 characters among pairs of taxa, and (3) the relation between CBCs, hCBCs, CBC clades, and the taxonomic level of organisms was investigated in detail. Results High sequence and length conservation allowed the generation of an ITS2 consensus secondary structure, and introduction of a novel numbering system of ITS2 nucleotides and base pairs. Alignments and analyses were based on this structural information, leading to the following results: (1) in the Ulvales, the presence of a CBC is not linked to any particular taxonomic level, (2) most CBC 'clades' sensu Coleman are paraphyletic, and should rather be termed CBC grades. (3) the phenetic approach of pairwise comparison of sequences can be misleading, and thus, CBCs/hCBCs must be investigated in their evolutionary context, including homoplasy events (4) CBCs and hCBCs in ITS2 helices evolved independently, and we found no evidence for a CBC that originated via a two-fold hCBC substitution. Conclusions Our case study revealed several discrepancies between ITS2 evolution in the Ulvales and generally accepted assumptions underlying ITS2 evolution as e.g. the CBC clade concept. Therefore, we developed a suite of methods providing a critical 'close-up' view into ITS2 evolution by directly tracing the evolutionary history of individual positions, and we caution against a non-critical use of the ITS2 CBC clade concept for species delimitation. PMID:21933414

Impact of water heater temperature setting and water use frequency on the building plumbing microbiome

PubMed Central

Ji, Pan; Rhoads, William J; Edwards, Marc A; Pruden, Amy

2017-01-01

Hot water plumbing is an important conduit of microbes into the indoor environment and can increase risk of opportunistic pathogens (for example, Legionella pneumophila). We examined the combined effects of water heater temperature (39, 42, 48, 51 and 58 °C), pipe orientation (upward/downward), and water use frequency (21, 3 and 1 flush per week) on the microbial composition at the tap using a pilot-scale pipe rig. 16S rRNA gene amplicon sequencing indicated that bulk water and corresponding biofilm typically had distinct taxonomic compositions (R2Adonis=0.246, PAdonis=0.001), yet similar predicted functions based on PICRUSt analysis (R2Adonis=0.087, PAdonis=0.001). Although a prior study had identified 51 °C under low water use frequency to enrich Legionella at the tap, here we reveal that 51 °C is also a threshold above which there are marked effects of the combined influences of temperature, pipe orientation, and use frequency on taxonomic and functional composition. A positive association was noted between relative abundances of Legionella and mitochondrial DNA of Vermamoeba, a genus of amoebae that can enhance virulence and facilitate replication of some pathogens. This study takes a step towards intentional control of the plumbing microbiome and highlights the importance of microbial ecology in governing pathogen proliferation. PMID:28282040

Community Structures of Fecal Bacteria in Cattle from Different Animal Feeding Operations▿†

PubMed Central

Shanks, Orin C.; Kelty, Catherine A.; Archibeque, Shawn; Jenkins, Michael; Newton, Ryan J.; McLellan, Sandra L.; Huse, Susan M.; Sogin, Mitchell L.

2011-01-01

The fecal microbiome of cattle plays a critical role not only in animal health and productivity but also in food safety, pathogen shedding, and the performance of fecal pollution detection methods. Unfortunately, most published molecular surveys fail to provide adequate detail about variability in the community structures of fecal bacteria within and across cattle populations. Using massively parallel pyrosequencing of a hypervariable region of the rRNA coding region, we profiled the fecal microbial communities of cattle from six different feeding operations where cattle were subjected to consistent management practices for a minimum of 90 days. We obtained a total of 633,877 high-quality sequences from the fecal samples of 30 adult beef cattle (5 individuals per operation). Sequence-based clustering and taxonomic analyses indicate less variability within a population than between populations. Overall, bacterial community composition correlated significantly with fecal starch concentrations, largely reflected in changes in the Bacteroidetes, Proteobacteria, and Firmicutes populations. In addition, network analysis demonstrated that annotated sequences clustered by management practice and fecal starch concentration, suggesting that the structures of bovine fecal bacterial communities can be dramatically different in different animal feeding operations, even at the phylum and family taxonomic levels, and that the feeding operation is a more important determinant of the cattle microbiome than is the geographic location of the feedlot. PMID:21378055

Genome sequence of the thermophilic fresh-water bacterium Spirochaeta caldaria type strain (H1T), reclassification of Spirochaeta caldaria, Spirochaeta stenostrepta, and Spirochaeta zuelzerae in the genus Treponema as Treponema caldaria comb. nov., Treponema stenostrepta comb. nov., and Treponema zuelzerae comb. nov., and emendation of the genus Tr

DOE Office of Scientific and Technical Information (OSTI.GOV)

Abt, Birte; Goker, Markus; Scheuner, Carmen

2013-01-01

Spirochaeta caldaria Pohlschroeder et al. 1995 is an obligately anaerobic, spiral-shaped bac- terium that is motile via periplasmic flagella. The type strain, H1T, was isolated in 1990 from cyanobacterial mat samples collected at a freshwater hot spring in Oregon, USA, and is of in- terest because it enhances the degradation of cellulose when grown in co-culture with Clos- tridium thermocellum. Here we provide a taxonomic re-evaluation for S. caldaria based on phylogenetic analyses of 16S rRNA sequences and whole genomes, and propose the reclassi- fication of S. caldaria and two other Spirochaeta species as members of the emended genus Treponema.more » Whereas genera such as Borrelia and Sphaerochaeta possess well-distinguished genomic features related to their divergent lifestyles, the physiological and functional ge- nomic characteristics of Spirochaeta and Treponema appear to be intermixed and are of little taxonomic value. The 3,239,340 bp long genome of strain H1T with its 2,869 protein-coding and 59 RNA genes is a part of the Genomic Encyclopedia of Bacteria and Archaea project.« less

Molecular identification of Amazonian stingless bees using polymerase chain reaction single-strand conformation polymorphism.

PubMed

Souza, M T; Carvalho-Zilse, G A

2014-07-25

In countries containing a mega diversity of wildlife, such as Brazil, identifying and characterizing biological diversity is a continuous process for the scientific community, even in face of technological and scientific advances. This activity demands initiatives for the taxonomic identification of highly diverse groups, such as stingless bees, including molecular analysis strategies. This type of bee is distributed in all of the Brazilian states, with the highest species diversity being found in the State of Amazônia. However, the estimated number of species diverges among taxonomists. These bees are considered the main pollinators in the Amazon rainforest, in which they obtain food and shelter; however, their persistence is constantly threatened by deforestation pressure. Hence, it is important to classify the number and abundance of bee specie, to measure their decline and implement meaningful, priority conservation strategies. This study aims to maximize the implementation of more direct, economic and successful techniques for the taxonomic identification of stingless bees. Specifically, the genes 16S rRNA and COI from mitochondrial DNA were used as molecular markers to differentiate 9 species of Amazonian stingless bees based on DNA polymorphism, using the polymerase chain reaction-single-strand conformation polymorphism technique. We registered different, exclusive SSCP haplotypes for both genes in all species analyzed. These results demonstrate that SSCP is a simple and cost-effective technique that is applicable to the molecular identification of stingless bee species.

‘ Candidatus Dichloromethanomonas elyunquensis’ gen. nov., sp. nov., a dichloromethane-degrading anaerobe of the Peptococcaceae family

DOE PAGES

Kleindienst, Sara; Higgins, Steven A.; Tsementzi, Despina; ...

2016-12-21

Taxonomic assignments of anaerobic dichloromethane (DCM)-degrading bacteria remain poorly constrained but are important for understanding the microbial diversity of organisms contributing to DCM turnover in environmental systems. Here, we describe the taxonomic classification of a novel DCM degrader in consortium RM obtained from pristine Rio Mameyes sediment. Phylogenetic analysis of full-length 16S rRNA gene sequences demonstrated that the DCM degrader was most closely related to members of the genera Dehalobacter and Syntrophobotulus, but sequence similarities did not exceed 94% and 93%, respectively. Genome-aggregate average amino acid identities against Peptococcaceae members did not exceed 66%, suggesting that the DCM degrader doesmore » not affiliate with any described genus. Phylogenetic analysis of conserved single-copy functional genes supported that the DCM degrader represents a novel clade. Growth strictly depended on the presence of DCM, which was consumed at a rate of 160 ± 3 μmol L -1 d -1. The DCM degrader attained 5.25 × 10 7 ± 1.0 × 10 7 cells per μmol DCM consumed. Fluorescence in situ hybridization revealed rod-shaped cells 4 ± 0.8 μm long and 0.4 ± 0.1 μm wide. Furthermore, based on the unique phylogenetic, genomic, and physiological characteristics, we propose that the DCM degrader represents a new genus and species, ‘Candidatus Dichloromethanomonas elyunquensis’.« less

Conservation genetics of North American freshwater mussels Amblema and Megalonaias

USGS Publications Warehouse

Mulvey, M.; Lydeard, C.; Pyer, D.L.; Hicks, K.M.; Brim-Box, J.; Williams, J.D.; Butler, R.S.

1997-01-01

Freshwater bivalves are among the most endangered groups of organisms in North America. Efforts to protect the declining mussel fauna are confounded by ambiguities associated with recognition of distinct evolutionary entities or species. This, in part, is due to the paucity of reliable morphological characters for differentiating taxa. We have employed allozymes and DNA sequence data to search for diagnosably distinct evolutionary entities within two problematic genera of unionid mussels, Amblema and Megalonaias. Within the genus Amblema three species are recognized based on our DNA sequence data for the mitochondrial 16S rRNA and allozyme data (Amblema neislerii, A. plicata, and A. elliotti). Only one taxonomically distinct entity is recognized within the genus Megalonaias—M. nervosa. Megalonaias boykiniana of the Apalachicolan Region is not diagnosable and does not warrant specific taxonomic status. Interestingly, Megalonaias from west of the Mississippi River, including the Mississippi, exhibited an allozyme and mtDNA haplotype frequency shift suggestive of an east-west dichotomy. The results of this study eliminate one subspecies of Amblema and increase the range of A. plicata. This should not affect the conservation status of “currently stable” assigned to A. plicata by Williams et al. (1993). The conservation status of A. elliotti needs to be reexamined because its distribution appears to be limited to the Coosa River System in Alabama and Georgia.

‘ Candidatus Dichloromethanomonas elyunquensis’ gen. nov., sp. nov., a dichloromethane-degrading anaerobe of the Peptococcaceae family

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kleindienst, Sara; Higgins, Steven A.; Tsementzi, Despina

Taxonomic assignments of anaerobic dichloromethane (DCM)-degrading bacteria remain poorly constrained but are important for understanding the microbial diversity of organisms contributing to DCM turnover in environmental systems. Here, we describe the taxonomic classification of a novel DCM degrader in consortium RM obtained from pristine Rio Mameyes sediment. Phylogenetic analysis of full-length 16S rRNA gene sequences demonstrated that the DCM degrader was most closely related to members of the genera Dehalobacter and Syntrophobotulus, but sequence similarities did not exceed 94% and 93%, respectively. Genome-aggregate average amino acid identities against Peptococcaceae members did not exceed 66%, suggesting that the DCM degrader doesmore » not affiliate with any described genus. Phylogenetic analysis of conserved single-copy functional genes supported that the DCM degrader represents a novel clade. Growth strictly depended on the presence of DCM, which was consumed at a rate of 160 ± 3 μmol L -1 d -1. The DCM degrader attained 5.25 × 10 7 ± 1.0 × 10 7 cells per μmol DCM consumed. Fluorescence in situ hybridization revealed rod-shaped cells 4 ± 0.8 μm long and 0.4 ± 0.1 μm wide. Furthermore, based on the unique phylogenetic, genomic, and physiological characteristics, we propose that the DCM degrader represents a new genus and species, ‘Candidatus Dichloromethanomonas elyunquensis’.« less

Expanding the World of Marine Bacterial and Archaeal Clades

PubMed Central

Yilmaz, Pelin; Yarza, Pablo; Rapp, Josephine Z.; Glöckner, Frank O.

2016-01-01

Determining which microbial taxa are out there, where they live, and what they are doing is a driving approach in marine microbial ecology. The importance of these questions is underlined by concerted, large-scale, and global ocean sampling initiatives, for example the International Census of Marine Microbes, Ocean Sampling Day, or Tara Oceans. Given decades of effort, we know that the large majority of marine Bacteria and Archaea belong to about a dozen phyla. In addition to the classically culturable Bacteria and Archaea, at least 50 “clades,” at different taxonomic depths, exist. These account for the majority of marine microbial diversity, but there is still an underexplored and less abundant portion remaining. We refer to these hitherto unrecognized clades as unknown, as their boundaries, names, and classifications are not available. In this work, we were able to characterize up to 92 of these unknown clades found within the bacterial and archaeal phylogenetic diversity currently reported for marine water column environments. We mined the SILVA 16S rRNA gene datasets for sequences originating from the marine water column. Instead of the usual subjective taxa delineation and nomenclature methods, we applied the candidate taxonomic unit (CTU) circumscription system, along with a standardized nomenclature to the sequences in newly constructed phylogenetic trees. With this new phylogenetic and taxonomic framework, we performed an analysis of ICoMM rRNA gene amplicon datasets to gain insights into the global distribution of the new marine clades, their ecology, biogeography, and interaction with oceanographic variables. Most of the new clades we identified were interspersed by known taxa with cultivated members, whose genome sequences are available. This result encouraged us to perform metabolic predictions for the novel marine clades using the PICRUSt approach. Our work also provides an update on the taxonomy of several phyla and widely known marine clades as our CTU approach breaks down these randomly lumped clades into smaller objectively calculated subgroups. Finally, all taxa were classified and named following standards compatible with the Bacteriological Code rules, enhancing their digitization, and comparability with future microbial ecological and taxonomy studies. PMID:26779174

Strain diversity and host specificity in bee gut symbionts revealed by deep sampling of single copy protein-coding sequences

PubMed Central

Powell, J. Elijah; Ratnayeke, Nalin; Moran, Nancy A.

2017-01-01

High throughput rRNA amplicon surveys of bacterial communities provide a rapid snapshot of taxonomic composition. But strains with nearly identical rRNA sequences often differ in gene repertoires and metabolic capabilities. To assess strain-level variation within Snodgrassella alvi, a gut symbiont of corbiculate bees, we performed deep sequencing on amplicons of a single copy coding gene (minD) as well as the 16S rDNA V4 region. We surveyed honey bees (Apis mellifera) sampled globally and 12 bumble bee species (Bombus) sampled from two regions of the USA. The minD analyses reveal that S. alvi contains far more strain diversity than is evident from 16S rDNA analysis. Many taxa inferred on the basis of 16S rDNA are shared between A. mellifera and Bombus species, but taxa inferred on the basis of minD are never shared and often are restricted to particular Bombus species. Clustering based on minD revealed that gut communities often reflect host species and geographic location. Both minD and 16S rDNA analyses indicate that strain diversity is higher in A. mellifera than in Bombus species. The minD locus flanks a 16S gene, enabling development of strain-specific 16S fluorescent probes to illuminate the spatial relationship of strains within the bee gut. PMID:27482856

Status of the Microbial Census

PubMed Central

Schloss, Patrick D.; Handelsman, Jo

2004-01-01

Over the past 20 years, more than 78,000 16S rRNA gene sequences have been deposited in GenBank and the Ribosomal Database Project, making the 16S rRNA gene the most widely studied gene for reconstructing bacterial phylogeny. While there is a general appreciation that these sequences are largely unique and derived from diverse species of bacteria, there has not been a quantitative attempt to describe the extent of sequencing efforts to date. We constructed rarefaction curves for each bacterial phylum and for the entire bacterial domain to assess the current state of sampling and the relative taxonomic richness of each phylum. This analysis quantifies the general sense among microbiologists that we are a long way from a complete census of the bacteria on Earth. Moreover, the analysis indicates that current sampling strategies might not be the most effective ones to describe novel diversity because there remain numerous phyla that are globally distributed yet poorly sampled. Based on the current level of sampling, it is not possible to estimate the total number of bacterial species on Earth, but the minimum species richness is 35,498. Considering previous global species richness estimates of 107 to 109, we are certain that this estimate will increase with additional sequencing efforts. The data support previous calls for extensive surveys of multiple chemically disparate environments and of specific phylogenetic groups to advance the census most rapidly. PMID:15590780

Bacterial community composition in the gut content of Lampetra japonica revealed by 16S rRNA gene pyrosequencing.

PubMed

Zuo, Yu; Xie, Wenfang; Pang, Yue; Li, Tiesong; Li, Qingwei; Li, Yingying

2017-01-01

The composition of the bacterial communities in the hindgut contents of Lampetrs japonica was surveyed by Illumina MiSeq of the 16S rRNA gene. An average of 32385 optimized reads was obtained from three samples. The rarefaction curve based on the operational taxonomic units tended to approach the asymptote. The rank abundance curve representing the species richness and evenness was calculated. The composition of microbe in six classification levels was also analyzed. Top 20 members in genera level were displayed as the classification tree. The abundance of microorganisms in different individuals was displayed as the pie charts at the branch nodes in the classification tree. The differences of top 50 genera in abundance between individuals of lamprey are displayed as a heatmap. The pairwise comparison of bacterial taxa abundance revealed that there are no significant differences of gut microbiota between three individuals of lamprey at a given rarefied depth. Also, the gut microbiota derived from L. japonica displays little similarity with other aquatic organism of Vertebrata after UPGMA analysis. The metabolic function of the bacterial communities was predicted through KEGG analysis. This study represents the first analysis of the bacterial community composition in the gut content of L. japonica. The investigation of the gut microbiota associated with L. japonica will broaden our understanding of this unique organism.

Zooming-in on floral nectar: a first exploration of nectar-associated bacteria in wild plant communities.

PubMed

Alvarez-Pérez, Sergio; Herrera, Carlos M; de Vega, Clara

2012-06-01

Floral nectar of some animal-pollinated plants usually harbours highly adapted yeast communities which can profoundly alter nectar characteristics and, therefore, potentially have significant impacts on plant reproduction through their effects on insect foraging behaviour. Bacteria have also been occasionally observed in floral nectar, but their prevalence, phylogenetic diversity and ecological role within plant-pollinator-yeast systems remains unclear. Here we present the first reported survey of bacteria in floral nectar from a natural plant community. Culturable bacteria occurring in a total of 71 nectar samples collected from 27 South African plant species were isolated and identified by 16S rRNA gene sequencing. Rarefaction-based analyses were used to assess operational taxonomic units (OTUs) richness at the plant community level using nectar drops as sampling units. Our results showed that bacteria are common inhabitants of floral nectar of South African plants (53.5% of samples yielded growth), and their communities are characterized by low species richness (18 OTUs at a 16S rRNA gene sequence dissimilarity cut-off of 3%) and moderate phylogenetic diversity, with most isolates belonging to the Gammaproteobacteria. Furthermore, isolates showed osmotolerance, catalase activity and the ability to grow under microaerobiosis, three traits that might help bacteria to overcome important factors limiting their survival and/or growth in nectar. © 2012 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

Penicillium simile sp. nov. revealed by morphological and phylogenetic analysis.

PubMed

Davolos, Domenico; Pietrangeli, Biancamaria; Persiani, Anna Maria; Maggi, Oriana

2012-02-01

The morphology of three phenetically identical Penicillium isolates, collected from the bioaerosol in a restoration laboratory in Italy, displayed macro- and microscopic characteristics that were similar though not completely ascribable to Penicillium raistrickii. For this reason, a phylogenetic approach based on DNA sequencing analysis was performed to establish both the taxonomic status and the evolutionary relationships of these three peculiar isolates in relation to previously described species of the genus Penicillium. We used four nuclear loci (both rRNA and protein coding genes) that have previously proved useful for the molecular investigation of taxa belonging to the genus Penicillium at various evolutionary levels. The internal transcribed spacer region (ITS1-5.8S-ITS2), domains D1 and D2 of the 28S rDNA, a region of the tubulin beta chain gene (benA) and part of the calmodulin gene (cmd) were amplified by PCR and sequenced. Analysis of the rRNA genes and of the benA and cmd sequence data indicates the presence of three isogenic isolates belonging to a genetically distinct species of the genus Penicillium, here described and named Penicillium simile sp. nov. (ATCC MYA-4591(T)  = CBS 129191(T)). This novel species is phylogenetically different from P. raistrickii and other related species of the genus Penicillium (e.g. Penicillium scabrosum), from which it can be distinguished on the basis of morphological trait analysis.

Gaetbulimicrobium brevivitae gen. nov., sp. nov., a novel member of the family Flavobacteriaceae isolated from a tidal flat of the Yellow Sea in Korea.

PubMed

Yoon, Jung-Hoon; Kang, So-Jung; Jung, Seo-Youn; Oh, Hyun Woo; Oh, Tae-Kwang

2006-01-01

A Gram-negative, non-spore-forming and rod-shaped gliding bacterium, designated strain SMK-19T, was isolated from a tidal flat sediment of the Yellow Sea, Korea, and its taxonomic position was investigated by using a polyphasic approach. Strain SMK-19T grew optimally at 37 degrees C, in the presence of 2-3 % (w/v) NaCl and at pH 7.0-8.0. It contained MK-6 as the predominant menaquinone, and iso-C(17 : 0) 3-OH and iso-C(15 : 0) as the major fatty acids. Major polar lipids were phosphatidylethanolamine, unidentified phospholipids and an amino-group-containing lipid that is ninhydrin-positive. The DNA G+C content was 36.0 mol%. Phylogenetic analyses based on 16S rRNA gene sequences demonstrated that strain SMK-19T formed a distinct evolutionary lineage within the family Flavobacteriaceae. The 16S rRNA gene sequence of strain SMK-19T exhibited similarity values of <94.4 % to those of other members of the family Flavobacteriaceae. Strain SMK-19T was distinguished from phylogenetically related genera by differences in several phenotypic properties. On the basis of phenotypic, phylogenetic and chemotaxonomic data, SMK-19T (= KCTC 12390T = DSM 17196T) was classified at the type strain of a novel genus and species, Gaetbulimicrobium brevivitae gen. nov., sp. nov.

Jannaschia seohaensis sp. nov., isolated from a tidal flat sediment.

PubMed

Yoon, Jung-Hoon; Kang, So-Jung; Park, Sooyeon; Oh, Ki-Hoon; Oh, Tae-Kwang

2010-01-01

A Gram-negative, motile and pleomorphic bacterial strain, SMK-146(T), was isolated from a tidal flat sediment of the Yellow Sea, Korea, and its taxonomic position was investigated. Strain SMK-146(T) grew optimally at pH 7.0-8.0 and 30 degrees C. It contained Q-10 as the predominant ubiquinone and C(18 : 1)omega7c and 11-methyl C(18 : 1)omega7c as the major fatty acids. The major polar lipids were phosphatidylcholine, phosphatidylglycerol and phosphatidylethanolamine. The DNA G+C content was 68.4 mol%. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain SMK-146(T) belongs to the genus Jannaschia. Strain SMK-146(T) exhibited 16S rRNA gene sequence similarity values of 95.3-97.0 % to the type strains of the five recognized Jannaschia species. The mean DNA-DNA relatedness value between strain SMK-146(T) and Jannaschia seosinensis KCCM 42114(T), the closest phylogenetic neighbour, was 17 %. Differential phenotypic properties also revealed that strain SMK-146(T) differs from the recognized Jannaschia species. On the basis of phenotypic, phylogenetic and genetic data, strain SMK-146(T) represents a novel species of the genus Jannaschia, for which the name Jannaschia seohaensis sp. nov. is proposed. The type strain is SMK-146(T) (=KCTC 22172(T) =CCUG 55326(T)).

Pedobacter insulae sp. nov., isolated from soil.

PubMed

Yoon, Jung-Hoon; Kang, So-Jung; Oh, Hyun Woo; Oh, Tae-Kwang

2007-09-01

A Gram-negative, non-motile, rod-shaped bacterium, DS-139(T), was isolated from a soil sample collected from Dokdo, Korea, and subjected to a polyphasic taxonomic analysis. Strain DS-139(T) grew optimally at 25 degrees C and pH 6.5-7.5 in the presence of 0-0.5 % (w/v) NaCl. It contained MK-7 as the predominant menaquinone and iso-C(15 : 0), C(16 : 1)omega7c and/or iso-C(15 : 0) 2-OH and iso-C(17 : 0) 3-OH as the major fatty acids. The DNA G+C content was 39.4 mol%. Phylogenetic analyses based on 16S rRNA gene sequences revealed that strain DS-39(T) belongs to the genus Pedobacter in the family Sphingobacteriaceae. The similarity values between the 16S rRNA gene sequence of strain DS-139(T) and those of the type strains of recognized Pedobacter species, except Pedobacter saltans, were in the range 93.9-96.7 %. The differential phenotypic properties, together with the phylogenetic distinctiveness, were sufficient to assign strain DS-139(T) to a species that is separate from recognized Pedobacter species. On the basis of the phenotypic and phylogenetic data, therefore, strain DS-139(T) represents a novel species of the genus Pedobacter, for which the name Pedobacter insulae sp. nov. is proposed. The type strain is DS-139(T) (=KCTC 12820(T) =DSM 18684(T)).

Phylogenetic relationships and species circumscription in Trentepohlia and Printzina (Trentepohliales, Chlorophyta).

PubMed

Rindi, Fabio; Lam, Daryl W; López-Bautista, Juan M

2009-08-01

Subaerial green microalgae represent a polyphyletic complex of organisms, whose genetic diversity is much higher than their simple morphologies suggest. The order Trentepohliales is the only species-rich group of subaerial algae belonging to the class Ulvophyceae and represents an ideal model taxon to investigate evolutionary patterns of these organisms. We studied phylogenetic relationships in two common genera of Trentepohliales (Trentepohlia and Printzina) by separate and combined analyses of the rbcL and 18S rRNA genes. Trentepohlia and Printzina were not resolved as monophyletic groups. Three main clades were recovered in all analyses, but none corresponded to any trentepohlialean genus as defined based on morphological grounds. The rbcL and 18S rRNA datasets provided congruent phylogenetic signals and similar topologies were recovered in single-gene analyses. Analyses performed on the combined 2-gene dataset inferred generally higher nodal support. The results clarified several taxonomic problems and showed that the evolution of these algae has been characterized by considerable morphological convergence. Trentepohlia abietina and T. flava were shown to be separate species from T. aurea; Printzina lagenifera, T. arborum and T. umbrina were resolved as polyphyletic taxa, whose vegetative morphology appears to have evolved independently in separate lineages. Incongruence between phylogenetic relationships and traditional morphological classification was demonstrated, showing that the morphological characters commonly used in the taxonomy of the Trentepohliales are phylogenetically irrelevant.

Tetrahymena australis (Protozoa, Ciliophora): A Well-Known But "Non-Existing" Taxon - Consideration of Its Identification, Definition and Systematic Position.

PubMed

Liu, Mingjian; Fan, Xinpeng; Gao, Feng; Gao, Shan; Yu, Yuhe; Warren, Alan; Huang, Jie

2016-11-01

A cryptic species of the Tetrahymena pyriformis complex, Tetrahymena australis, has been known for a long time but never properly diagnosed based on taxonomic methods. The species name is thus invalid according to the International Code of Zoological Nomenclature. Recently, a population isolated from a freshwater lake in Wuhan, China was investigated using live observations, silver staining methods and gene sequence data. This organism can be separated from other described species of the T. pyriformis complex by its relatively small body size, the number of somatic kineties and differences in sequences of two genes, namely the small subunit ribosomal RNA (SSU rRNA) and the mitochondrial cytochrome c oxidase subunit I (cox1). We compared the SSU rRNA gene sequences of all available Tetrahymena species to reveal the nucleotide differences within this genus. The sequence of the Wuhan population is identical to two sequences of a previously isolated strain of T. australis (ATCC #30831). Phylogenetic analyses indicate that these three sequences (X56167, M98015, KT334373) cluster with Tetrahymena shanghaiensis (EF070256) in a polytomy. However, sequence divergence of the cox1 gene between the Wuhan population and another strain of T. australis (ATCC #30271) is 1.4%, suggesting that these may represent different subspecies. © 2016 The Author(s) Journal of Eukaryotic Microbiology © 2016 International Society of Protistologists.

A new species of Proctoeces and reinstatement of Proctoeces humboldti George-Nascimento and Quiroga 1983 (Digenea: Fellodistomidae) based on molecular and morphological evidence.

PubMed

Oliva, Marcelo E; Valdivia, Isabel M; Cárdenas, Leyla; Muñoz, Gabriela; Escribano, Ruben; George-Nascimento, Mario

2018-04-01

The most studied digenean of marine organisms in Chile is by far Proctoeces humboldti, a parasite of the intestine of the clingfish Sicyases sanguineus and gonad of the keyhole limpet Fissurella spp. (progenetic metacercariae). The mussel Perumytilus purpuratus has been suggested as the first intermediate host for this digenean. In a study examining the parasites of S. sanguineus from central Chile, we found specimens of Proctoeces showing significant morphological differences with P. humboldti. To assist in the resolution of the taxonomic identification of these specimens, as well sporocysts obtained from the mussel P. purpuratus from central and northern Chile, phylogenetic studies using DNA sequences from the SSU rRNA, as well the LSU rRNA and Cox 1 gene were performed. Results showed that the clingfish S. sanguineus is a host for two species of Proctoeces (P. humboldti and P. syciases n. sp.) along the northern and central Chilean coast, without geographic separation; the mussel P. purpuratus is the first intermediate host for P. syciases n. sp. but not for P. humboldti in central and northern Chile. Fissurellids (Archaeogastropoda) along the Chilean coast harbor only progenetic stages of P. humboldti, but there is no evidence of progenesis for P. syciases. The reinstatement of Proctoeces humboldti is strongly suggested. Copyright © 2017 Elsevier B.V. All rights reserved.

Widespread occurrence of organelle genome-encoded 5S rRNAs including permuted molecules

PubMed Central

Valach, Matus; Burger, Gertraud; Gray, Michael W.; Lang, B. Franz

2014-01-01

5S Ribosomal RNA (5S rRNA) is a universal component of ribosomes, and the corresponding gene is easily identified in archaeal, bacterial and nuclear genome sequences. However, organelle gene homologs (rrn5) appear to be absent from most mitochondrial and several chloroplast genomes. Here, we re-examine the distribution of organelle rrn5 by building mitochondrion- and plastid-specific covariance models (CMs) with which we screened organelle genome sequences. We not only recover all organelle rrn5 genes annotated in GenBank records, but also identify more than 50 previously unrecognized homologs in mitochondrial genomes of various stramenopiles, red algae, cryptomonads, malawimonads and apusozoans, and surprisingly, in the apicoplast (highly derived plastid) genomes of the coccidian pathogens Toxoplasma gondii and Eimeria tenella. Comparative modeling of RNA secondary structure reveals that mitochondrial 5S rRNAs from brown algae adopt a permuted triskelion shape that has not been seen elsewhere. Expression of the newly predicted rrn5 genes is confirmed experimentally in 10 instances, based on our own and published RNA-Seq data. This study establishes that particularly mitochondrial 5S rRNA has a much broader taxonomic distribution and a much larger structural variability than previously thought. The newly developed CMs will be made available via the Rfam database and the MFannot organelle genome annotator. PMID:25429974

Widespread occurrence of organelle genome-encoded 5S rRNAs including permuted molecules.

PubMed

Valach, Matus; Burger, Gertraud; Gray, Michael W; Lang, B Franz

2014-12-16

5S Ribosomal RNA (5S rRNA) is a universal component of ribosomes, and the corresponding gene is easily identified in archaeal, bacterial and nuclear genome sequences. However, organelle gene homologs (rrn5) appear to be absent from most mitochondrial and several chloroplast genomes. Here, we re-examine the distribution of organelle rrn5 by building mitochondrion- and plastid-specific covariance models (CMs) with which we screened organelle genome sequences. We not only recover all organelle rrn5 genes annotated in GenBank records, but also identify more than 50 previously unrecognized homologs in mitochondrial genomes of various stramenopiles, red algae, cryptomonads, malawimonads and apusozoans, and surprisingly, in the apicoplast (highly derived plastid) genomes of the coccidian pathogens Toxoplasma gondii and Eimeria tenella. Comparative modeling of RNA secondary structure reveals that mitochondrial 5S rRNAs from brown algae adopt a permuted triskelion shape that has not been seen elsewhere. Expression of the newly predicted rrn5 genes is confirmed experimentally in 10 instances, based on our own and published RNA-Seq data. This study establishes that particularly mitochondrial 5S rRNA has a much broader taxonomic distribution and a much larger structural variability than previously thought. The newly developed CMs will be made available via the Rfam database and the MFannot organelle genome annotator. © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.

Plantactinospora sonchi sp. nov., an actinobacterium isolated from the leaves of common sowthistle (Sonchus oleraceus L.).

PubMed

Ma, Zhaoxu; Liu, Chongxi; Fan, Jianlong; He, Hairong; Li, Chuang; Li, Jiansong; Zhao, Shanshan; Xiang, Wensheng; Wang, Xiangjing

2015-12-01

A novel actinobacterium, designated strain NEAU-QY2T, was isolated from the leaves of Sonchus oleraceus L. specimen, collected from Wuchang, Heilongjiang Province, China. A polyphasic study was carried out to establish the taxonomic position of this strain. The organism formed single spores with rough surfaces on substrate mycelia. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain NEAU-QY2T belonged to the genus Plantactinospora and formed a monophyletic clade with its closest related strains Plantactinospora endophytica YIM 68255T (99.2 % 16S rRNA gene sequence similarity), Plantactinospora veratri NEAU-FHS4T (98.8 %) and Plantactinospora mayteni YIM 61359T(98.7 %), an association that was supported by a bootstrap value of 90 % in the neighbor-joining tree and also recovered with the maximum-likelihood algorithm. However, DNA-DNA hybridization values between strain NEAU-QY2T and the three closely related strains were below 70 %. With reference to phenotypic characteristics, phylogenetic data and DNA-DNA hybridization results, strain NEAU-QY2T was distinguished from closely related strains and is classified as representing a novel species of the genus Plantactinospora, for which the name Plantactinospora sonchi sp. nov. is proposed. The type strain is NEAU-QY2T (=CGMCC4.7216T=JCM 30345T).

Census of bacterial microbiota associated with the glacier ice worm Mesenchytraeus solifugus.

PubMed

Murakami, Takumi; Segawa, Takahiro; Bodington, Dylan; Dial, Roman; Takeuchi, Nozomu; Kohshima, Shiro; Hongoh, Yuichi

2015-03-01

The glacier ice worm, Mesenchytraeus solifugus, is a unique annelid, inhabiting only snow and ice in North American glaciers. Here, we analyzed the taxonomic composition of bacteria associated with M. solifugus based on the 16S rRNA gene. We analyzed four fixed-on-site and 10 starved ice worm individuals, along with glacier surface samples. In total, 1341 clones of 16S rRNA genes were analyzed for the ice worm samples, from which 65 bacterial phylotypes (99.0% cut-off) were identified. Of these, 35 phylotypes were closely related to sequences obtained from their habitat glacier and/or other components of cryosphere; whereas three dominant phylotypes were affiliated with animal-associated lineages of the class Mollicutes. Among the three, phylotype Ms-13 shared less than 89% similarity with database sequences and was closest to a gut symbiont of a terrestrial earthworm. Using fluorescence in situ hybridization, Ms-13 was located on the gut wall surface of the ice worms. We propose a novel genus and species, 'Candidatus Vermiplasma glacialis', for this bacterium. Our results raise the possibility that the ice worm has exploited indigenous glacier bacteria, while several symbiotic bacterial lineages have maintained their association with the ice worm during the course of adaptive evolution to the permanently cold environment. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Microbiology of aquatic environments: Characterizations of the microbiotas of municipal water supplies, the International Space Station Internal Active Thermal Control System's heat transport fluid, and US Space Shuttle drinking water

NASA Astrophysics Data System (ADS)

Bernardini, James Nicholas, III

An understanding of the microbiota within life support systems is essential for the prolonged presence of humans in space. This is because microbes may cause disease or induce biofouling and/or corrosion within spacecraft water systems. It is imperative that we develop effective high-throughput technologies for characterizing microbial populations that can eventually be used in the space environment. This dissertation describes testing and development of such methodologies, targeting both bacteria and viruses in water, and examines the bacterial and viral diversity within two spacecraft life support systems. The bacterial community of the International Space Station Internal Active Thermal Control System (IATCS) was examined using conventional culture-based and advanced molecular techniques including adenosine triphosphate (ATP) and Limulus Amebocyte Lysate (LAL) assays, direct microscopic examination, and analyses of 16S rRNA gene libraries from the community metagenome. The cultivable heterotrophs of the IATCS fluids ranged from below detection limit to 1.1x10 5/100 ml, and viable cells, measured by ATP, ranged from 1.4x10 3/100 ml to 7.7x105/100 ml. DNA extraction, cloning, sequencing, and bioinformatic analysis of the clones from 16S RNA gene libraries showed members of the firmicutes, alpha, beta, and gamma-proteobacteria to be present in the fluids. This persistent microbial bioburden and the presence of probable metal reducers, biofilm formers, and opportunistic pathogens illustrate the need for better characterization of bacterial communities present within spacecraft fluids. A new methodology was developed for detection of viruses in water using microarrays. Samples were concentrated by lyophilization, resuspended and filtered (0.22microm). Viral nucleic acids were then extracted, amplified, fluorescently labeled and hybridized onto a custom microarray with probes for ˜1000 known viruses. Numerous virus signatures were observed. Human Adenovirus C and Influenza A viruses were used to verify positive microarray hybridizations by quantitative polymerase chain reaction (PCR), reverse transcriptase PCR, and conventional PCR. Experiments were performed using municipal drinking water, IATCS fluids, and Shuttle drinking water. Thus, this dissertation describes what we believe is the first molecular analysis of the IATCS bacterial ecology and the first use and validation of a microarray-based assay for the detection of viral genetic signatures within drinking waters.

Metagenomic analysis of bacterial and archaeal assemblages in the soil-mousse surrounding a geothermal spring

PubMed Central

Bhatia, Sonu; Batra, Navneet; Pathak, Ashish; Joshi, Amit; Souza, Leila; Almeida, Paulo; Chauhan, Ashvini

2015-01-01

The soil-mousse surrounding a geothermal spring was analyzed for bacterial and archaeal diversity using 16S rRNA gene amplicon metagenomic sequencing which revealed the presence of 18 bacterial phyla distributed across 109 families and 219 genera. Firmicutes, Actinobacteria, and the Deinococcus-Thermus group were the predominant bacterial assemblages with Crenarchaeota and Thaumarchaeota as the main archaeal assemblages in this largely understudied geothermal habitat. Several metagenome sequences remained taxonomically unassigned suggesting the presence of a repertoire of hitherto undescribed microbes in this geothermal soil-mousse econiche. PMID:26484255

Spatial and temporal dynamics of the microbial community in the Hanford unconfined aquifer

PubMed Central

Lin, Xueju; McKinley, James; Resch, Charles T; Kaluzny, Rachael; Lauber, Christian L; Fredrickson, James; Knight, Rob; Konopka, Allan

2012-01-01

Pyrosequencing analysis of 16S rRNA genes was used to study temporal dynamics of groundwater bacteria and archaea over 10 months within three well clusters separated by ∼30 m and located 250 m from the Columbia River on the Hanford Site, WA. Each cluster contained three wells screened at different depths ranging from 10 to 17 m that differed in hydraulic conductivities. Representative samples were selected for analyses of prokaryotic 16S and eukaryotic 18S rRNA gene copy numbers. Temporal changes in community composition occurred in all nine wells over the 10-month sampling period. However, there were particularly strong effects near the top of the water table when the seasonal rise in the Columbia River caused river water intrusion at the top of the aquifer. The occurrence and disappearance of some microbial assemblages (such as Actinobacteria ACK-M1) were correlated with river water intrusion. This seasonal impact on microbial community structure was greater in the shallow saturated zone than deeper zone in the aquifer. Spatial and temporal patterns for several 16S rRNA gene operational taxonomic units associated with particular physiological functions (for example, methane oxidizers and metal reducers) suggests dynamic changes in fluxes of electron donors and acceptors over an annual cycle. In addition, temporal dynamics in eukaryotic 18S rRNA gene copies and the dominance of protozoa in 18S clone libraries suggest that bacterial community dynamics could be affected not only by the physical and chemical environment but also by top-down biological control. PMID:22456444

Spatial and temporal dynamics of the microbial community in the Hanford unconfined aquifer.

PubMed

Lin, Xueju; McKinley, James; Resch, Charles T; Kaluzny, Rachael; Lauber, Christian L; Fredrickson, James; Knight, Rob; Konopka, Allan

2012-09-01

Pyrosequencing analysis of 16S rRNA genes was used to study temporal dynamics of groundwater bacteria and archaea over 10 months within three well clusters separated by ~30 m and located 250 m from the Columbia River on the Hanford Site, WA. Each cluster contained three wells screened at different depths ranging from 10 to 17 m that differed in hydraulic conductivities. Representative samples were selected for analyses of prokaryotic 16S and eukaryotic 18S rRNA gene copy numbers. Temporal changes in community composition occurred in all nine wells over the 10-month sampling period. However, there were particularly strong effects near the top of the water table when the seasonal rise in the Columbia River caused river water intrusion at the top of the aquifer. The occurrence and disappearance of some microbial assemblages (such as Actinobacteria ACK-M1) were correlated with river water intrusion. This seasonal impact on microbial community structure was greater in the shallow saturated zone than deeper zone in the aquifer. Spatial and temporal patterns for several 16S rRNA gene operational taxonomic units associated with particular physiological functions (for example, methane oxidizers and metal reducers) suggests dynamic changes in fluxes of electron donors and acceptors over an annual cycle. In addition, temporal dynamics in eukaryotic 18S rRNA gene copies and the dominance of protozoa in 18S clone libraries suggest that bacterial community dynamics could be affected not only by the physical and chemical environment but also by top-down biological control.

Halomonas titanicae sp. nov., a halophilic bacterium isolated from the RMS Titanic.

PubMed

Sánchez-Porro, Cristina; Kaur, Bhavleen; Mann, Henrietta; Ventosa, Antonio

2010-12-01

A Gram-negative, heterotrophic, aerobic, non-endospore-forming, peritrichously flagellated and motile bacterial strain, designated BH1(T), was isolated from samples of rusticles, which are formed in part by a consortium of micro-organisms, collected from the RMS Titanic wreck site. The strain grew optimally at 30-37°C, pH 7.0-7.5 and in the presence of 2-8 % (w/v) NaCl. We carried out a polyphasic taxonomic study in order to characterize the strain in detail. Phylogenetic analyses based on 16S rRNA gene sequence comparison indicated that strain BH1(T) clustered within the branch consisting of species of Halomonas. The most closely related type strains were Halomonas neptunia (98.6 % 16S rRNA sequence similarity), Halomonas variabilis (98.4 %), Halomonas boliviensis (98.3 %) and Halomonas sulfidaeris (97.5 %). Other closely related species were Halomonas alkaliphila (96.5 % sequence similarity), Halomonas hydrothermalis (96.3 %), Halomonas gomseomensis (96.3 %), Halomonas venusta (96.3 %) and Halomonas meridiana (96.2 %). The major fatty acids of strain BH1(T) were C(18 : 1)ω7c (36.3 %), C(16 : 0) (18.4 %) and C(19 : 0) cyclo ω8c (17.9 %). The DNA G+C content was 60.0 mol% (T(m)). Ubiquinone 9 (Q-9) was the major lipoquinone. The phenotypic features, fatty acid profile and DNA G+C content further supported the placement of strain BH1(T) in the genus Halomonas. DNA-DNA hybridization values between strain BH1(T) and H. neptunia CECT 5815(T), H. variabilis DSM 3051(T), H. boliviensis DSM 15516(T) and H. sulfidaeris CECT 5817(T) were 19, 17, 30 and 29 %, respectively, supporting the differential taxonomic status of BH1(T). On the basis of the phenotypic, chemotaxonomic and phylogenetic data, strain BH1(T) is considered to represent a novel species, for which the name Halomonas titanicae sp. nov. is proposed. The type strain is BH1(T) (=ATCC BAA-1257(T) =CECT 7585(T) =JCM 16411(T) =LMG 25388(T)).

Micromonospora schwarzwaldensis sp. nov., a producer of telomycin, isolated from soil.

PubMed

Vela Gurovic, Maria Soledad; Müller, Sebastian; Domin, Nicole; Seccareccia, Ivana; Nietzsche, Sandor; Martin, Karin; Nett, Markus

2013-10-01

A Gram-stain-positive, spore-forming actinomycete strain (HKI0641(T)) was isolated from a soil sample collected in the Black Forest, Germany. During screening for antimicrobial natural products this bacterium was identified as a producer of the antibiotic telomycin. Morphological characteristics and chemotaxonomic data indicated that the strain belonged to the genus Micromonospora. The peptidoglycan of strain HKI0641(T) contained meso-diaminopimelic acid, and the fatty acid profile consisted predominantly of anteiso-C15 : 0, iso-C15 : 0, iso-C16 : 0 and C16 : 0. MK-10(H4), MK-10(H2) and MK-10 were identified as the major menaquinones. To determine the taxonomic positioning of strain HKI0641(T), we computed a binary tanglegram of two rooted phylogenetic trees that were based upon 16S rRNA and gyrB gene sequences. The comparative analysis of the two common classification methods strongly supported the phylogenetic affiliation with the genus Micromonospora, but it also revealed discrepancies in the assignment at the level of the genomic species. 16S rRNA gene sequence analysis identified Micromonospora coxensis DSM 45161(T) (99.1 % sequence similarity) and Micromonospora marina DSM 45555(T) (99.0 %) as the nearest taxonomic neighbours, whereas the gyrB sequence of strain HKI0641(T) indicated a closer relationship to Micromonospora aurantiaca DSM 43813(T) (95.1 %). By means of DNA-DNA hybridization experiments, it was possible to resolve this issue and to clearly differentiate strain HKI0641(T) from other species of the genus Micromonospora. The type strains of the aforementioned species of the genus Micromonospora could be further distinguished from strain HKI0641(T) by several phenotypic properties, such as colony colour, NaCl tolerance and the utilization of carbon sources. The isolate was therefore assigned to a novel species of the genus Micromonospora, for which the name Micromonospora schwarzwaldensis sp. nov. is proposed. The type strain is HKI0641(T) ( = DSM 45708(T) = CIP 110415(T)).

The phylogenetic relationships and molecular systematics of scincid lizards of the genus Heremites (Sauria, Scincidae) in the Middle East based on mtDNA sequences.

PubMed

Bahmani, Zahed; Rastegar-Pouyani, Eskandar; Rastegar-Pouyani, Nasrullah

2017-09-08

The taxonomic status of species included in the genus Heremites in Iran and Iraq is uncertain. Three of these species have been assigned to the genus based on morphology: Heremites auratus transcaucasica, H. vittatus, and H. septemtaeniatus. We examined the phylogenetic relationships and taxonomic status of the Iranian and Iraqi species of Heremites by performing phylogenetic analyses using mitochondrial DNA sequences (cytochrome b and 16S rRNA). Phylogenetic relationships and estimated genetic distances indicated that the Heremites populations of the area (Iran and Iraq) form five distinct clades. Three of these clades are found only in Iran, specifically in: (1) Fars and Hormozgan provinces; (2) Northeastern Khuzestan; and (3) Khorasan and Isfahan provinces. The fourth clade (H. septemtaeniatus) is found in west and Mahshahr in Iran as well as in eastern and northern parts of Iraq. The fifth clade, Heremites vittatus, is found in Iran and Iraq. We also confirm the absence of H. auratus in Iran and Iraq. It also indicated that H. vittatus is sister taxon to the other groups that our analyses estimate the divergence of this clade in the Middle Miocene (15.9 Mya). The clade containing the Fars-Hormozgan and Khuzestan populations diverged at the end of the Miocene (8.5 Mya). The Isfahan and Khorasan populations separated at the Pliocene (4.2 Mya) from the western Iranian group, the group in Mahshahr, Iran and the groups in northern and eastern Iraq.

Unravelling core microbial metabolisms in the hypersaline microbial mats of Shark Bay using high-throughput metagenomics

PubMed Central

Ruvindy, Rendy; White III, Richard Allen; Neilan, Brett Anthony; Burns, Brendan Paul

2016-01-01

Modern microbial mats are potential analogues of some of Earth's earliest ecosystems. Excellent examples can be found in Shark Bay, Australia, with mats of various morphologies. To further our understanding of the functional genetic potential of these complex microbial ecosystems, we conducted for the first time shotgun metagenomic analyses. We assembled metagenomic next-generation sequencing data to classify the taxonomic and metabolic potential across diverse morphologies of marine mats in Shark Bay. The microbial community across taxonomic classifications using protein-coding and small subunit rRNA genes directly extracted from the metagenomes suggests that three phyla Proteobacteria, Cyanobacteria and Bacteriodetes dominate all marine mats. However, the microbial community structure between Shark Bay and Highbourne Cay (Bahamas) marine systems appears to be distinct from each other. The metabolic potential (based on SEED subsystem classifications) of the Shark Bay and Highbourne Cay microbial communities were also distinct. Shark Bay metagenomes have a metabolic pathway profile consisting of both heterotrophic and photosynthetic pathways, whereas Highbourne Cay appears to be dominated almost exclusively by photosynthetic pathways. Alternative non-rubisco-based carbon metabolism including reductive TCA cycle and 3-hydroxypropionate/4-hydroxybutyrate pathways is highly represented in Shark Bay metagenomes while not represented in Highbourne Cay microbial mats or any other mat forming ecosystems investigated to date. Potentially novel aspects of nitrogen cycling were also observed, as well as putative heavy metal cycling (arsenic, mercury, copper and cadmium). Finally, archaea are highly represented in Shark Bay and may have critical roles in overall ecosystem function in these modern microbial mats. PMID:26023869

Effects of antibiotic growth promoter and characterization of ecological succession in Swine gut microbiota.

PubMed

Unno, Tatsuya; Kim, Jung-Man; Guevarra, Robin B; Nguyen, Son G

2015-04-01

Ever since the ban on antibiotic growth promoters (AGPs), the livestock death rate has increased owing to pathogenic bacterial infections. There is a need of developing AGP alternatives; however, the mechanisms by which AGP enhances livestock growth performance are not clearly understood. In this study, we fed 3-week-old swine for 9 weeks with and without AGPs containing chlortetracycline, sulfathiazole, and penicillin to investigate the effects of AGPs on swine gut microbiota. Microbial community analysis was done based on bacterial 16S rRNA genes using MiSeq. The use of AGP showed no growth promoting effect, but inhibited the growth of potential pathogens during the early growth stage. Our results showed the significant increase in species richness after the stabilization of gut microbiota during the post-weaning period (4-week-old). Moreover, the swine gut microbiota was divided into four clusters based on the distribution of operational taxonomic units, which was significantly correlated to the swine weight regardless of AGP treatments. Taxonomic abundance analysis indicated a negative correlation between host weight and the abundance of the family Prevotellaceae species, but showed positive correlation to the abundance of the family Spirochaetaceae, Clostridiaceae_1, and Peptostreptococcaeae species. Although no growth performance enhancement was observed, the use of AGP inhibited the potential pathogens in the early growth stage of swine. In addition, our results indicated the ecological succession of swine gut microbiota according to swine weight. Here, we present a characterization of swine gut microbiota with respect to the effects of AGPs on growth performance.

Molecular phylogeny of Pholadoidea Lamarck, 1809 supports a single origin for xylotrophy (wood feeding) and xylotrophic bacterial endosymbiosis in Bivalvia.

PubMed

Distel, Daniel L; Amin, Mehwish; Burgoyne, Adam; Linton, Eric; Mamangkey, Gustaf; Morrill, Wendy; Nove, John; Wood, Nicole; Yang, Joyce

2011-11-01

The ability to consume wood as food (xylotrophy) is unusual among animals. In terrestrial environments, termites and other xylotrophic insects are the principle wood consumers while in marine environments wood-boring bivalves fulfill this role. However, the evolutionary origin of wood feeding in bivalves has remained largely unexplored. Here we provide data indicating that xylotrophy has arisen just once in Bivalvia in a single wood-feeding bivalve lineage that subsequently diversified into distinct shallow- and deep-water branches, both of which have been broadly successful in colonizing the world's oceans. These data also suggest that the appearance of this remarkable life habit was approximately coincident with the acquisition of bacterial endosymbionts. Here we generate a robust phylogeny for xylotrophic bivalves and related species based on sequences of small and large subunit nuclear rRNA genes. We then trace the distribution among the modern taxa of morphological characters and character states associated with xylotrophy and xylotrepesis (wood-boring) and use a parsimony-based method to infer their ancestral states. Based on these ancestral state reconstructions we propose a set of plausible hypotheses describing the evolution of symbiotic xylotrophy in Bivalvia. Within this context, we reinterpret one of the most remarkable progressions in bivalve evolution, the transformation of the "typical" myoid body plan to create a unique lineage of worm-like, tube-forming, wood-feeding clams. The well-supported phylogeny presented here is inconsistent with most taxonomic treatments for xylotrophic bivalves, indicating that the bivalve family Pholadidae and the subfamilies Teredininae and Bankiinae of the family Teredinidae are non-monophyletic, and that the principle traits used for their taxonomic diagnosis are phylogenetically misleading. Copyright © 2011 Elsevier Inc. All rights reserved.

Single-base-pair discrimination of terminal mismatches by using oligonucleotide microarrays and neural network analyses

NASA Technical Reports Server (NTRS)

Urakawa, Hidetoshi; Noble, Peter A.; El Fantroussi, Said; Kelly, John J.; Stahl, David A.

2002-01-01

The effects of single-base-pair near-terminal and terminal mismatches on the dissociation temperature (T(d)) and signal intensity of short DNA duplexes were determined by using oligonucleotide microarrays and neural network (NN) analyses. Two perfect-match probes and 29 probes having a single-base-pair mismatch at positions 1 to 5 from the 5' terminus of the probe were designed to target one of two short sequences representing 16S rRNA. Nonequilibrium dissociation rates (i.e., melting profiles) of all probe-target duplexes were determined simultaneously. Analysis of variance revealed that position of the mismatch, type of mismatch, and formamide concentration significantly affected the T(d) and signal intensity. Increasing the concentration of formamide in the washing buffer decreased the T(d) and signal intensity, and it decreased the variability of the signal. Although T(d)s of probe-target duplexes with mismatches in the first or second position were not significantly different from one another, duplexes with mismatches in the third to fifth positions had significantly lower T(d)s than those with mismatches in the first or second position. The trained NNs predicted the T(d) with high accuracies (R(2) = 0.93). However, the NNs predicted the signal intensity only moderately accurately (R(2) = 0.67), presumably due to increased noise in the signal intensity at low formamide concentrations. Sensitivity analysis revealed that the concentration of formamide explained most (75%) of the variability in T(d)s, followed by position of the mismatch (19%) and type of mismatch (6%). The results suggest that position of the mismatch at or near the 5' terminus plays a greater role in determining the T(d) and signal intensity of duplexes than the type of mismatch.

Thermus caliditerrae sp. nov., a novel thermophilic species isolated from a geothermal area.

PubMed

Ming, Hong; Yin, Yi-Rui; Li, Shuai; Nie, Guo-Xing; Yu, Tian-Tian; Zhou, En-Min; Liu, Lan; Dong, Lei; Li, Wen-Jun

2014-02-01

Two thermophilic bacterial strains, designated YIM 77925(T) and YIM 77777, were isolated from two hot springs, one in the Hydrothermal Explosion (Shuirebaozhaqu) area and Frog Mouth Spring in Tengchong county, Yunnan province, south-western China. The taxonomic positions of the two isolates were investigated by a polyphasic approach. Cells of the two strains were Gram-stain-negative, aerobic and rod-shaped. They were able to grow at 50-70 °C, pH 6.0-8.0 and with a NaCl tolerance up to 0.5% (w/v). Colonies are circular, convex, non-transparent and produce yellow pigment. Phylogenetic analyses based on 16S rRNA gene sequences comparison clearly demonstrated that strains YIM 77925(T) and YIM 77777 represent members of the genus Thermus, and they also detected low-level similarities of 16S rRNA gene sequences (below 97%) compared with all other species in this genus. Their predominant menaquinone was MK-8. The genomic DNA G+C contents of strains YIM 77925(T) and YIM 77777 were 65.6 mol% and 67.2 mol%, respectively. Based on the results of physiological and biochemical tests and phylogenetic analyses, strains YIM 77925(T) and YIM 77777 could not be classified as representing any species of the genus Thermus with a validly published name. Thus the two strains are considered to represent a novel species of the genus Thermus, for which the name Thermus caliditerrae sp. nov. is proposed. The type strain is YIM 77925(T) ( = DSM 25901(T) = CCTCC 2012061(T)).

Effect of physical sediments reworking on hydrocarbon degradation and bacterial community structure in marine coastal sediments.

PubMed

Duran, Robert; Bonin, Patricia; Jezequel, Ronan; Dubosc, Karine; Gassie, Claire; Terrisse, Fanny; Abella, Justine; Cagnon, Christine; Militon, Cecile; Michotey, Valérie; Gilbert, Franck; Cuny, Philippe; Cravo-Laureau, Cristiana

2015-10-01

The present study aimed to examine whether the physical reworking of sediments by harrowing would be suitable for favouring the hydrocarbon degradation in coastal marine sediments. Mudflat sediments were maintained in mesocosms under conditions as closer as possible to those prevailing in natural environments with tidal cycles. Sediments were contaminated with Ural blend crude oil, and in half of them, harrowing treatment was applied in order to mimic physical reworking of surface sediments. Hydrocarbon distribution within the sediment and its removal was followed during 286 days. The harrowing treatment allowed hydrocarbon compounds to penetrate the first 6 cm of the sediments, and biodegradation indexes (such as n-C18/phytane) indicated that biodegradation started 90 days before that observed in untreated control mesocosms. However, the harrowing treatment had a severe impact on benthic organisms reducing drastically the macrofaunal abundance and diversity. In the harrowing-treated mesocosms, the bacterial abundance, determined by 16S rRNA gene Q-PCR, was slightly increased; and terminal restriction fragment length polymorphism (T-RFLP) analyses of 16S rRNA genes showed distinct and specific bacterial community structure. Co-occurrence network and canonical correspondence analyses (CCA) based on T-RFLP data indicated the main correlations between bacterial operational taxonomic units (OTUs) as well as the associations between OTUs and hydrocarbon compound contents further supported by clustered correlation (ClusCor) analysis. The analyses highlighted the OTUs constituting the network structural bases involved in hydrocarbon degradation. Negative correlations indicated the possible shifts in bacterial communities that occurred during the ecological succession.

Biochemical characterization and phylogenetic analysis based on 16S rRNA sequences for V-factor dependent members of Pasteurellaceae derived from laboratory rats.

PubMed

Hayashimoto, Nobuhito; Ueno, Masami; Tkakura, Akira; Itoh, Toshio

2007-06-01

Phylogenetic analysis based on 16S rRNA sequences with sequence data of some bacterial species of Pasteurellaceae related to rodents deposited in GenBank was performed along with biochemical characterization for the 20 strains of V-factor dependent members of Pasteurellaceae derived from laboratory rats to obtain basic information and to investigate the taxonomic positions. The results of biochemical tests for all strains were identical except for three tests, the ornithine decarboxylase test, and fermentation tests of D(+) mannose and D(+) xylose. The biochemical properties of 8 of 20 strains that showed negative results for the fermentation test of D(+) xylose agreed with those of Haemophilus parainfluenzae complex. By phylogenetic analysis, the strains were divided into two clusters that agreed with the results of the fermentation test of xylose (group I: negative reaction for xylose, group II: positive reaction for xylose). The clusters were independent of other bacterial species of Pasteurellaceae tested. The sequences of the strains in group I showed 99.7-99.8% similarity and the strains in group II showed 99.3-99.7% similarity. None of the strains in group I had a close relation with Haemophilus parainfluenzae by phylogenetic analysis, although they showed the same biochemical properties. In conclusion, the strains had characteristic biochemical properties and formed two independent groups within the "rodent cluster" of Pasteurellaceae that differed in the results of the fermentation test of xylose. Therefore, they seemed to be hitherto undescribed taxa in Pasteurellaceae.

A novel genus of the class Actinobacteria, Longivirga aurantiaca gen. nov., sp. nov., isolated from lake sediment.

PubMed

Qu, Jian-Hang; Zhang, Lu-Jie; Fu, Yun-Hui; Li, Xiao-Dan; Li, Hai-Feng; Tian, Hai-Long

2018-03-01

A novel actinobacterial strain, designated X5 T , was isolated from the sediment of Taihu Lake in China and was subjected to a polyphasic taxonomic characterization. The strain formed orange-red colonies comprising aerobic, Gram-stain-negative, rod-shaped cells on R2A agar. Phylogenetic analysis based on the 16S rRNA gene sequences revealed that the organism was closely related to the genus Sporichthya and consistently formed a distinct clade along with the members of this genus. The closest phylogenetic neighbour was Sporichthya polymorpha NBRC 12702 T with 93.7 % 16S rRNA gene sequence similarity. The major fatty acids (>10 %) were iso-C16 : 0 (18.7 %), C18 : 1ω9c (18.6 %) and C17 : 1ω8c (14.0 %). The genomic DNA G+C content was 74.4 mol%. The organism contained menaquinone MK-8(H2), MK-9(H4) and an unidentified menaquinone. Polar lipids were composed of phosphatidylglycerol, an unidentified lipid, two unidentified phospholipids and two unidentified aminolipids. The whole-cell sugars contained ribose, xylose, mannose, glucose and galactose. The cell-wall peptidoglycan contained ll-diaminopimelic acid. Based on the physiological, biochemical and chemotaxonomic data, the organism is proposed to represent a novel genus and species, for which the name Longivirga aurantiaca gen. nov., sp. nov. is proposed. The type strain is X5 T (=CGMCC 4.7317 T =NBRC 112237 T ).

Halobacillus alkaliphilus sp. nov., a halophilic bacterium isolated from a salt lake in Fuente de Piedra, southern Spain.

PubMed

Romano, Ida; Finore, Ilaria; Nicolaus, Giancarlo; Huertas, F Javier; Lama, Licia; Nicolaus, Barbara; Poli, Annarita

2008-04-01

A Gram-positive, spore-forming, halophilic bacterial strain, FP5T, was isolated from a salt lake in southern Spain and subjected to a polyphasic taxonomic study. Strain FP5T was strictly aerobic. Cells were coccoidal, occurring singly or in clusters. The cell-wall peptidoglycan type of strain FP5T was A4 beta based on l-Orn-d-Asp. Strain FP5T was characterized chemotaxonomically by having MK-7 as the major menaquinone and anteiso-C15 : 0, anteiso-C17 : 0, iso-C15 : 0 and iso-C16 : 0 as the main fatty acids. The isolate grew optimally at 37 degrees C and in presence of 10 % NaCl; no growth was observed in the absence of NaCl. The DNA G+C content was 43.5 mol%. Phylogenetic analyses based on 16S rRNA gene sequences showed that strain FP5T falls within the evolutionary radiation of species of the genus Halobacillus. Levels of 16S rRNA gene sequence similarity between strain FP5T and the type strains of nine recognized Halobacillus species were in the range 97.0-99.0 %. Levels of DNA-DNA relatedness indicated that strain FP5T represents a genomic species that is distinct from recognized Halobacillus species. Strain FP5T could be differentiated from recognized Halobacillus species based on several phenotypic characteristics. On the basis of phenotypic, phylogenetic and genomic data, strain FP5T is considered to represent a novel species of the genus Halobacillus, for which the name Halobacillus alkaliphilus sp. nov. is proposed. The type strain is FP5T (=DSM 18525T =ATCC BAA-1361T).

Re-evaluation of the taxonomy of the Mitis group of the genus Streptococcus based on whole genome phylogenetic analyses, and proposed reclassification of Streptococcus dentisani as Streptococcus oralis subsp. dentisani comb. nov., Streptococcus tigurinus as Streptococcus oralis subsp. tigurinus comb. nov., and Streptococcus oligofermentans as a later synonym of Streptococcus cristatus.

PubMed

Jensen, Anders; Scholz, Christian F P; Kilian, Mogens

2016-11-01

The Mitis group of the genus Streptococcus currently comprises 20 species with validly published names, including the pathogen S. pneumoniae. They have been the subject of much taxonomic confusion, due to phenotypic overlap and genetic heterogeneity, which has hampered a full appreciation of their clinical significance. The purpose of this study was to critically re-examine the taxonomy of the Mitis group using 195 publicly available genomes, including designated type strains for phylogenetic analyses based on core genomes, multilocus sequences and 16S rRNA gene sequences, combined with estimates of average nucleotide identity (ANI) and in silico and in vitro analyses of specific phenotypic characteristics. Our core genomic phylogenetic analyses revealed distinct clades that, to some extent, and from the clustering of type strains represent known species. However, many of the genomes have been incorrectly identified adding to the current confusion. Furthermore, our data show that 16S rRNA gene sequences and ANI are unsuitable for identifying and circumscribing new species of the Mitis group of the genus Streptococci. Based on the clustering patterns resulting from core genome phylogenetic analysis, we conclude that S. oligofermentans is a later synonym of S. cristatus. The recently described strains of the species Streptococcus dentisani includes one previously referred to as 'S. mitis biovar 2'. Together with S. oralis, S. dentisani and S. tigurinus form subclusters within a coherent phylogenetic clade. We propose that the species S. oralis consists of three subspecies: S. oralis subsp. oralis subsp. nov., S. oralis subsp. tigurinus comb. nov., and S. oralis subsp. dentisani comb. nov.

Integrative taxonomy helps to reveal the mask of the genus Gynandropaa (Amphibia: Anura: Dicroglossidae).

PubMed

Huang, Yan; Hu, Junhua; Wang, Bin; Song, Zhaobin; Zhou, Caiquan; Jiang, Jianping

2016-03-01

Species of the genus Gynandropaa within the family Dicroglossidae are typical spiny frogs whose taxonomic status has long been in doubt. We used integrative methods, involving morphological and molecular analyses, to elucidate the phylogenetic relationships, and to determine identities and the geographic distribution of each valid species. We obtained DNA sequence data of 5 species of Gynandropaa (complete sequences of the mitochondrial NADH dehydrogenase subunit 2 [ND2] gene, and 890 bp of 12S rRNA and 16S rRNA partial sequences) from 37 localities (including the topotypes of 5 described species) and constructed Bayesian and maximum-likelihood trees to examine the patterns of phylogeography. A total of 28 morphological variables were taken on 624 specimens. Three clades with clear geographic patterns were recognized: clade C (from south-western Sichuan Province and central Yunnan Province), clade E (western Guizhou Province and eastern to central Yunnan Province) and clade W (western to southern Yunnan Province). Integrating morphological characteristics and distribution information, the clades W, E and C represent Gynandropaa yunnanensis, G. phrynoides and G. sichuanensis, respectively. We draw the following conclusions: (i) the taxon G. phrynoides, formerly evaluated as a junior synonym of G. yunnanensis, is revalidated herein at the rank of species; (ii) G. liui is a junior synonym of G. sichuanensis; and (iii) G. yunnanensis is a valid species while G. bourreti is probably a subspecies of G. yunnanensis, with the distribution range from Vietnam to southern Yunnan Province. This study clears up the taxonomic status of Gynandropaa and provides important information for understanding the evolution and conservation of these spiny frogs. © 2015 International Society of Zoological Sciences, Institute of Zoology/Chinese Academy of Sciences and John Wiley & Sons Australia, Ltd.

From genus to phylum: large-subunit and internal transcribed spacer rRNA operon regions show similar classification accuracies influenced by database composition.

PubMed

Porras-Alfaro, Andrea; Liu, Kuan-Liang; Kuske, Cheryl R; Xie, Gary

2014-02-01

We compared the classification accuracy of two sections of the fungal internal transcribed spacer (ITS) region, individually and combined, and the 5' section (about 600 bp) of the large-subunit rRNA (LSU), using a naive Bayesian classifier and BLASTN. A hand-curated ITS-LSU training set of 1,091 sequences and a larger training set of 8,967 ITS region sequences were used. Of the factors evaluated, database composition and quality had the largest effect on classification accuracy, followed by fragment size and use of a bootstrap cutoff to improve classification confidence. The naive Bayesian classifier and BLASTN gave similar results at higher taxonomic levels, but the classifier was faster and more accurate at the genus level when a bootstrap cutoff was used. All of the ITS and LSU sections performed well (>97.7% accuracy) at higher taxonomic ranks from kingdom to family, and differences between them were small at the genus level (within 0.66 to 1.23%). When full-length sequence sections were used, the LSU outperformed the ITS1 and ITS2 fragments at the genus level, but the ITS1 and ITS2 showed higher accuracy when smaller fragment sizes of the same length and a 50% bootstrap cutoff were used. In a comparison using the larger ITS training set, ITS1 and ITS2 had very similar accuracy classification for fragments between 100 and 200 bp. Collectively, the results show that any of the ITS or LSU sections we tested provided comparable classification accuracy to the genus level and underscore the need for larger and more diverse classification training sets.

From Genus to Phylum: Large-Subunit and Internal Transcribed Spacer rRNA Operon Regions Show Similar Classification Accuracies Influenced by Database Composition

PubMed Central

Liu, Kuan-Liang; Kuske, Cheryl R.

2014-01-01

We compared the classification accuracy of two sections of the fungal internal transcribed spacer (ITS) region, individually and combined, and the 5′ section (about 600 bp) of the large-subunit rRNA (LSU), using a naive Bayesian classifier and BLASTN. A hand-curated ITS-LSU training set of 1,091 sequences and a larger training set of 8,967 ITS region sequences were used. Of the factors evaluated, database composition and quality had the largest effect on classification accuracy, followed by fragment size and use of a bootstrap cutoff to improve classification confidence. The naive Bayesian classifier and BLASTN gave similar results at higher taxonomic levels, but the classifier was faster and more accurate at the genus level when a bootstrap cutoff was used. All of the ITS and LSU sections performed well (>97.7% accuracy) at higher taxonomic ranks from kingdom to family, and differences between them were small at the genus level (within 0.66 to 1.23%). When full-length sequence sections were used, the LSU outperformed the ITS1 and ITS2 fragments at the genus level, but the ITS1 and ITS2 showed higher accuracy when smaller fragment sizes of the same length and a 50% bootstrap cutoff were used. In a comparison using the larger ITS training set, ITS1 and ITS2 had very similar accuracy classification for fragments between 100 and 200 bp. Collectively, the results show that any of the ITS or LSU sections we tested provided comparable classification accuracy to the genus level and underscore the need for larger and more diverse classification training sets. PMID:24242255

An integrative description of Macrobiotus paulinae sp. nov. (Tardigrada: Eutardigrada: Macrobiotidae: hufelandi group) from Kenya.

PubMed

Stec, Daniel; Smolak, Radoslav; Kaczmarek, Łukasz; Michalczyk, Łukasz

2015-12-07

In this paper we describe Macrobiotus paulinae, a new species of the hufelandi group from the Kenyan highlands. In addition to the traditional taxonomic description, aided with morphometrics as well as light and scanning microscopy imaging, we also provide nucleotide sequences of three nuclear (18S rRNA, 28S rRNA, ITS-2) and one mitochondrial (COI) DNA fragment of the new species. The sequences allowed not only a more accurate description but also provided an independent verification of the taxonomic status of Ma. paulinae sp. nov. Such integrative approach requires a considerable number of individuals and eggs, which we have partially subsidised by employing an in vitro culture of the new species. Our analyses revealed that Ma. paulinae sp. nov. is most similar to Macrobiotus madegassus Maucci, 1993 and Macrobiotus modestus Pilato & Lisi, 2009, however it differs from these species, as well as from all other known species of the hufelandi group, by the presence of seven paired dorso-lateral patches of cuticular granulation and the presence of chorionic filaments growing out of terminal discs of egg processes. Macrobiotus paulinae sp. nov. is an example of a species with a miniaturised buccal apparatus (i.e. with reduced peribuccal lamellae and oral cavity armature, and stylet supports inserted on the buccal tube more anteriorly than in typical Ma. hufelandi group species), and it therefore resembles two recently described two-macroplacoided Minibiotus species: Mi. acadianus Meyer & Domingue, 2011 and Mi. julianae Meyer, 2012. The re-examination of the type material for these two species confirmed that they are equipped with peribuccal lamellae and therefore we transfer them to the genus Macrobiotus, specifically to the hufelandi group.

16S rRNA amplicon sequencing identifies microbiota associated with oral cancer, human papilloma virus infection and surgical treatment.

PubMed

Guerrero-Preston, Rafael; Godoy-Vitorino, Filipa; Jedlicka, Anne; Rodríguez-Hilario, Arnold; González, Herminio; Bondy, Jessica; Lawson, Fahcina; Folawiyo, Oluwasina; Michailidi, Christina; Dziedzic, Amanda; Thangavel, Rajagowthamee; Hadar, Tal; Noordhuis, Maartje G; Westra, William; Koch, Wayne; Sidransky, David

2016-08-09

Systemic inflammatory events and localized disease, mediated by the microbiome, may be measured in saliva as head and neck squamous cell carcinoma (HNSCC) diagnostic and prognostic biomonitors. We used a 16S rRNA V3-V5 marker gene approach to compare the saliva microbiome in DNA isolated from Oropharyngeal (OPSCC), Oral Cavity Squamous Cell Carcinoma (OCSCC) patients and normal epithelium controls, to characterize the HNSCC saliva microbiota and examine their abundance before and after surgical resection.The analyses identified a predominance of Firmicutes, Proteobacteria and Bacteroidetes, with less frequent presence of Actinobacteria and Fusobacteria before surgery. At lower taxonomic levels, the most abundant genera were Streptococcus, Prevotella, Haemophilus, Lactobacillus and Veillonella, with lower numbers of Citrobacter and Neisseraceae genus Kingella. HNSCC patients had a significant loss in richness and diversity of microbiota species (p<0.05) compared to the controls. Overall, the Operational Taxonomic Units network shows that the relative abundance of OTU's within genus Streptococcus, Dialister, and Veillonella can be used to discriminate tumor from control samples (p<0.05). Tumor samples lost Neisseria, Aggregatibacter (Proteobacteria), Haemophillus (Firmicutes) and Leptotrichia (Fusobacteria). Paired taxa within family Enterobacteriaceae, together with genus Oribacterium, distinguish OCSCC samples from OPSCC and normal samples (p<0.05). Similarly, only HPV positive samples have an abundance of genus Gemellaceae and Leuconostoc (p<0.05). Longitudinal analyses of samples taken before and after surgery, revealed a reduction in the alpha diversity measure after surgery, together with an increase of this measure in patients that recurred (p<0.05). These results suggest that microbiota may be used as HNSCC diagnostic and prognostic biomonitors.

16S rRNA amplicon sequencing identifies microbiota associated with oral cancer, human papilloma virus infection and surgical treatment

PubMed Central

Guerrero-Preston, Rafael; Godoy-Vitorino, Filipa; Jedlicka, Anne; Rodríguez-Hilario, Arnold; González, Herminio; Bondy, Jessica; Lawson, Fahcina; Folawiyo, Oluwasina; Michailidi, Christina; Dziedzic, Amanda; Thangavel, Rajagowthamee; Hadar, Tal; Noordhuis, Maartje G.; Westra, William; Koch, Wayne; Sidransky, David

2016-01-01

Systemic inflammatory events and localized disease, mediated by the microbiome, may be measured in saliva as head and neck squamous cell carcinoma (HNSCC) diagnostic and prognostic biomonitors. We used a 16S rRNA V3-V5 marker gene approach to compare the saliva microbiome in DNA isolated from Oropharyngeal (OPSCC), Oral Cavity Squamous Cell Carcinoma (OCSCC) patients and normal epithelium controls, to characterize the HNSCC saliva microbiota and examine their abundance before and after surgical resection. The analyses identified a predominance of Firmicutes, Proteobacteria and Bacteroidetes, with less frequent presence of Actinobacteria and Fusobacteria before surgery. At lower taxonomic levels, the most abundant genera were Streptococcus, Prevotella, Haemophilus, Lactobacillus and Veillonella, with lower numbers of Citrobacter and Neisseraceae genus Kingella. HNSCC patients had a significant loss in richness and diversity of microbiota species (p<0.05) compared to the controls. Overall, the Operational Taxonomic Units network shows that the relative abundance of OTU's within genus Streptococcus, Dialister, and Veillonella can be used to discriminate tumor from control samples (p<0.05). Tumor samples lost Neisseria, Aggregatibacter (Proteobacteria), Haemophillus (Firmicutes) and Leptotrichia (Fusobacteria). Paired taxa within family Enterobacteriaceae, together with genus Oribacterium, distinguish OCSCC samples from OPSCC and normal samples (p<0.05). Similarly, only HPV positive samples have an abundance of genus Gemellaceae and Leuconostoc (p<0.05). Longitudinal analyses of samples taken before and after surgery, revealed a reduction in the alpha diversity measure after surgery, together with an increase of this measure in patients that recurred (p<0.05). These results suggest that microbiota may be used as HNSCC diagnostic and prognostic biomonitors. PMID:27259999

An accurate and efficient experimental approach for characterization of the complex oral microbiota.

PubMed

Zheng, Wei; Tsompana, Maria; Ruscitto, Angela; Sharma, Ashu; Genco, Robert; Sun, Yijun; Buck, Michael J

2015-10-05

Currently, taxonomic interrogation of microbiota is based on amplification of 16S rRNA gene sequences in clinical and scientific settings. Accurate evaluation of the microbiota depends heavily on the primers used, and genus/species resolution bias can arise with amplification of non-representative genomic regions. The latest Illumina MiSeq sequencing chemistry has extended the read length to 300 bp, enabling deep profiling of large number of samples in a single paired-end reaction at a fraction of the cost. An increasingly large number of researchers have adopted this technology for various microbiome studies targeting the 16S rRNA V3-V4 hypervariable region. To expand the applicability of this powerful platform for further descriptive and functional microbiome studies, we standardized and tested an efficient, reliable, and straightforward workflow for the amplification, library construction, and sequencing of the 16S V1-V3 hypervariable region using the new 2 × 300 MiSeq platform. Our analysis involved 11 subgingival plaque samples from diabetic and non-diabetic human subjects suffering from periodontitis. The efficiency and reliability of our experimental protocol was compared to 16S V3-V4 sequencing data from the same samples. Comparisons were based on measures of observed taxonomic richness and species evenness, along with Procrustes analyses using beta(β)-diversity distance metrics. As an experimental control, we also analyzed a total of eight technical replicates for the V1-V3 and V3-V4 regions from a synthetic community with known bacterial species operon counts. We show that our experimental protocol accurately measures true bacterial community composition. Procrustes analyses based on unweighted UniFrac β-diversity metrics depicted significant correlation between oral bacterial composition for the V1-V3 and V3-V4 regions. However, measures of phylotype richness were higher for the V1-V3 region, suggesting that V1-V3 offers a deeper assessment of population diversity and community ecology for the complex oral microbiota. This study provides researchers with valuable experimental evidence for the selection of appropriate 16S amplicons for future human oral microbiome studies. We expect that the tested 16S V1-V3 framework will be widely applicable to other types of microbiota, allowing robust, time-efficient, and inexpensive examination of thousands of samples for population, phylogenetic, and functional crossectional and longitutidal studies.

Consistent changes in the taxonomic structure and functional attributes of bacterial communities during primary succession.

PubMed

Ortiz-Álvarez, Rüdiger; Fierer, Noah; de Los Ríos, Asunción; Casamayor, Emilio O; Barberán, Albert

2018-02-20

Ecologists have long studied primary succession, the changes that occur in biological communities after initial colonization of an environment. Most of this work has focused on succession in plant communities, laying the conceptual foundation for much of what we currently know about community assembly patterns over time. Because of their prevalence and importance in ecosystems, an increasing number of studies have focused on microbial community dynamics during succession. Here, we conducted a meta-analysis of bacterial primary succession patterns across a range of distinct habitats, including the infant gut, plant surfaces, soil chronosequences, and aquatic environments, to determine whether consistent changes in bacterial diversity, community composition, and functional traits are evident over the course of succession. Although these distinct habitats harbor unique bacterial communities, we were able to identify patterns in community assembly that were shared across habitat types. We found an increase in taxonomic and functional diversity with time while the taxonomic composition and functional profiles of communities became less variable (lower beta diversity) in late successional stages. In addition, we found consistent decreases in the rRNA operon copy number and in the high-efficient phosphate assimilation process (Pst system) suggesting that reductions in resource availability during succession select for taxa adapted to low-resource conditions. Together, these results highlight that, like many plant communities, microbial communities also exhibit predictable patterns during primary succession.

The sRNAome mining revealed existence of unique signature small RNAs derived from 5.8SrRNA from Piper nigrum and other plant lineages.

PubMed

Asha, Srinivasan; Soniya, E V

2017-02-01

Small RNAs derived from ribosomal RNAs (srRNAs) are rarely explored in the high-throughput data of plant systems. Here, we analyzed srRNAs from the deep-sequenced small RNA libraries of Piper nigrum, a unique magnoliid plant. The 5' end of the putative long form of 5.8S rRNA (5.8S L rRNA) was identified as the site for biogenesis of highly abundant srRNAs that are unique among the Piperaceae family of plants. A subsequent comparative analysis of the ninety-seven sRNAomes of diverse plants successfully uncovered the abundant existence and precise cleavage of unique rRF signature small RNAs upstream of a novel 5' consensus sequence of the 5.8S rRNA. The major cleavage process mapped identically among the different tissues of the same plant. The differential expression and cleavage of 5'5.8S srRNAs in Phytophthora capsici infected P. nigrum tissues indicated the critical biological functions of these srRNAs during stress response. The non-canonical short hairpin precursor structure, the association with Argonaute proteins, and the potential targets of 5'5.8S srRNAs reinforced their regulatory role in the RNAi pathway in plants. In addition, this novel lineage specific small RNAs may have tremendous biological potential in the taxonomic profiling of plants.

Comparing K-mer based methods for improved classification of 16S sequences.

PubMed

Vinje, Hilde; Liland, Kristian Hovde; Almøy, Trygve; Snipen, Lars

2015-07-01

The need for precise and stable taxonomic classification is highly relevant in modern microbiology. Parallel to the explosion in the amount of sequence data accessible, there has also been a shift in focus for classification methods. Previously, alignment-based methods were the most applicable tools. Now, methods based on counting K-mers by sliding windows are the most interesting classification approach with respect to both speed and accuracy. Here, we present a systematic comparison on five different K-mer based classification methods for the 16S rRNA gene. The methods differ from each other both in data usage and modelling strategies. We have based our study on the commonly known and well-used naïve Bayes classifier from the RDP project, and four other methods were implemented and tested on two different data sets, on full-length sequences as well as fragments of typical read-length. The difference in classification error obtained by the methods seemed to be small, but they were stable and for both data sets tested. The Preprocessed nearest-neighbour (PLSNN) method performed best for full-length 16S rRNA sequences, significantly better than the naïve Bayes RDP method. On fragmented sequences the naïve Bayes Multinomial method performed best, significantly better than all other methods. For both data sets explored, and on both full-length and fragmented sequences, all the five methods reached an error-plateau. We conclude that no K-mer based method is universally best for classifying both full-length sequences and fragments (reads). All methods approach an error plateau indicating improved training data is needed to improve classification from here. Classification errors occur most frequent for genera with few sequences present. For improving the taxonomy and testing new classification methods, the need for a better and more universal and robust training data set is crucial.

[Study of generational risk in deafness inflicted couples using deafness gene microarray technique].

PubMed

Wang, Ping; Zhao, Jia; Yu, Shu-yuan; Jin, Peng; Zhu, Wei; DU, Bo

2011-06-01

To explored the significance of screening the gene mutations of deafness related in deaf-mute (deaf & dumb) family using DNA microarray. Total of 52 couples of deaf-mute were recruited from Changchun deaf-mute community. With an average age of (58.3 ± 6.7) years old (x(-) ± s). Blood samples were obtained with informed consent. Their genomic DNA was extracted from peripheral blood and PCR was performed. Nine of hot spot mutations in four most common deafness pathologic gene were examined with the DNA microarray, including GJB2, GJB3, PDS and mtDNA 12S rRNA genes. At the same time, the results were verified with the traditional methods of sequencing. Fifty of normal people served as a control group. All patients were diagnosed non-syndromic sensorineural hearing loss by subjective pure tone audiometry. Thirty-two of 104 cases appeared GJB2 gene mutation (30.7%), the mutation sites included 35delG, 176del16, 235delC and 299delAT. Eighteen of 32 cases of GJB2 mutations were 235delC (59.1%). Seven of 104 cases appeared SLC26A4 gene IVS7-2 A > G mutation. Questionnaire survey and gene diagnosis revealed that four of 52 families have deaf offspring (7.6%). When a couple carries the same gene mutation, the risk of their children deafness was 100%. The results were confirmed with the traditional methods of sequencing. There is a high risk of deafness if a deaf-mute family is planning to have a new baby. It is very important and helpful to avoid deaf newborns again in deaf-mute family by DNA microarray.

Molecular characterization of viable Legionella spp. in cooling tower water samples by combined use of ethidium monoazide and PCR.

PubMed

Inoue, Hiroaki; Fujimura, Reiko; Agata, Kunio; Ohta, Hiroyuki

2015-01-01

Viable Legionella spp. in environmental water samples were characterized phylogenetically by a clone library analysis combining the use of ethidium monoazide and quantitative PCR. To examine the diversity of Legionella spp., six cooling tower water samples and three bath water samples were collected and analyzed. A total of 617 clones were analyzed for their 16S rRNA gene sequences and classified into 99 operational taxonomic units (OTUs). The majority of OTUs were not clustered with currently described Legionella spp., suggesting the wide diversity of not-yet-cultured Legionella groups harbored in cooling tower water environments.

Phylogenetic relationships of leopard frogs (Rana pipiens complex) from an isolated coastal mountain range in southern Sonora, Mexico.

PubMed

Pfeiler, E; Markow, T A

2008-10-01

Mitochondrial DNA sequence data from the control region and 12S rRNA in leopard frogs from the Sierra El Aguaje of southern Sonora, Mexico, together with GenBank sequences, were used to infer taxonomic identity and provide phylogenetic hypotheses for relationships with other members of the Rana pipiens complex. We show that frogs from the Sierra El Aguaje belong to the Rana berlandieri subgroup, or Scurrilirana clade, of the R. pipiens group, and are most closely related to Rana magnaocularis from Nayarit, Mexico. We also provide further evidence that Rana magnaocularis and R. yavapaiensis are close relatives.

Molecular Characterization of Viable Legionella spp. in Cooling Tower Water Samples by Combined Use of Ethidium Monoazide and PCR

PubMed Central

Inoue, Hiroaki; Fujimura, Reiko; Agata, Kunio; Ohta, Hiroyuki

2015-01-01

Viable Legionella spp. in environmental water samples were characterized phylogenetically by a clone library analysis combining the use of ethidium monoazide and quantitative PCR. To examine the diversity of Legionella spp., six cooling tower water samples and three bath water samples were collected and analyzed. A total of 617 clones were analyzed for their 16S rRNA gene sequences and classified into 99 operational taxonomic units (OTUs). The majority of OTUs were not clustered with currently described Legionella spp., suggesting the wide diversity of not-yet-cultured Legionella groups harbored in cooling tower water environments. PMID:25736979

Staphylococcus muscae, a new species isolated from flies.

PubMed

Hájek, V; Ludwig, W; Schleifer, K H; Springer, N; Zitzelsberger, W; Kroppenstedt, R M; Kocur, M

1992-01-01

A new coagulase-negative species of the genus Staphylococcus, Staphylococcus muscae, is described on the basis of the results of a study of four strains that were isolated from flies. 16S rRNA sequences of the type strains of S. muscae, Staphylococcus schleiferi, and Staphylococcus sciuri were determined and used, together with the corresponding sequences of Staphylococcus aureus and Staphylococcus epidermidis, for a comparative analysis. The new species is characterized taxonomically; this species is differentiated from the other novobiocin-susceptible staphylococci by its physiological and biochemical activities, cell wall composition, and levels of genetic relatedness. The type strain of this species is strain MB4 (= CCM 4175).

Fluorescence-based bioassays for the detection and evaluation of food materials.

PubMed

Nishi, Kentaro; Isobe, Shin-Ichiro; Zhu, Yun; Kiyama, Ryoiti

2015-10-13

We summarize here the recent progress in fluorescence-based bioassays for the detection and evaluation of food materials by focusing on fluorescent dyes used in bioassays and applications of these assays for food safety, quality and efficacy. Fluorescent dyes have been used in various bioassays, such as biosensing, cell assay, energy transfer-based assay, probing, protein/immunological assay and microarray/biochip assay. Among the arrays used in microarray/biochip assay, fluorescence-based microarrays/biochips, such as antibody/protein microarrays, bead/suspension arrays, capillary/sensor arrays, DNA microarrays/polymerase chain reaction (PCR)-based arrays, glycan/lectin arrays, immunoassay/enzyme-linked immunosorbent assay (ELISA)-based arrays, microfluidic chips and tissue arrays, have been developed and used for the assessment of allergy/poisoning/toxicity, contamination and efficacy/mechanism, and quality control/safety. DNA microarray assays have been used widely for food safety and quality as well as searches for active components. DNA microarray-based gene expression profiling may be useful for such purposes due to its advantages in the evaluation of pathway-based intracellular signaling in response to food materials.

Fluorescence-Based Bioassays for the Detection and Evaluation of Food Materials

PubMed Central

Nishi, Kentaro; Isobe, Shin-Ichiro; Zhu, Yun; Kiyama, Ryoiti

2015-01-01

We summarize here the recent progress in fluorescence-based bioassays for the detection and evaluation of food materials by focusing on fluorescent dyes used in bioassays and applications of these assays for food safety, quality and efficacy. Fluorescent dyes have been used in various bioassays, such as biosensing, cell assay, energy transfer-based assay, probing, protein/immunological assay and microarray/biochip assay. Among the arrays used in microarray/biochip assay, fluorescence-based microarrays/biochips, such as antibody/protein microarrays, bead/suspension arrays, capillary/sensor arrays, DNA microarrays/polymerase chain reaction (PCR)-based arrays, glycan/lectin arrays, immunoassay/enzyme-linked immunosorbent assay (ELISA)-based arrays, microfluidic chips and tissue arrays, have been developed and used for the assessment of allergy/poisoning/toxicity, contamination and efficacy/mechanism, and quality control/safety. DNA microarray assays have been used widely for food safety and quality as well as searches for active components. DNA microarray-based gene expression profiling may be useful for such purposes due to its advantages in the evaluation of pathway-based intracellular signaling in response to food materials. PMID:26473869

Insect Gut Bacterial Diversity Determined by Environmental Habitat, Diet, Developmental Stage, and Phylogeny of Host

PubMed Central

Yun, Ji-Hyun; Roh, Seong Woon; Whon, Tae Woong; Jung, Mi-Ja; Kim, Min-Soo; Park, Doo-Sang; Yoon, Changmann; Nam, Young-Do; Kim, Yun-Ji; Choi, Jung-Hye; Kim, Joon-Yong; Shin, Na-Ri; Kim, Sung-Hee; Lee, Won-Jae

2014-01-01

Insects are the most abundant animals on Earth, and the microbiota within their guts play important roles by engaging in beneficial and pathological interactions with these hosts. In this study, we comprehensively characterized insect-associated gut bacteria of 305 individuals belonging to 218 species in 21 taxonomic orders, using 454 pyrosequencing of 16S rRNA genes. In total, 174,374 sequence reads were obtained, identifying 9,301 bacterial operational taxonomic units (OTUs) at the 3% distance level from all samples, with an average of 84.3 (±97.7) OTUs per sample. The insect gut microbiota were dominated by Proteobacteria (62.1% of the total reads, including 14.1% Wolbachia sequences) and Firmicutes (20.7%). Significant differences were found in the relative abundances of anaerobes in insects and were classified according to the criteria of host environmental habitat, diet, developmental stage, and phylogeny. Gut bacterial diversity was significantly higher in omnivorous insects than in stenophagous (carnivorous and herbivorous) insects. This insect-order-spanning investigation of the gut microbiota provides insights into the relationships between insects and their gut bacterial communities. PMID:24928884

Productivity and salinity structuring of the microplankton revealed by comparative freshwater metagenomics

PubMed Central

Eiler, Alexander; Zaremba-Niedzwiedzka, Katarzyna; Martínez-García, Manuel; McMahon, Katherine D; Stepanauskas, Ramunas; Andersson, Siv G E; Bertilsson, Stefan

2014-01-01

Little is known about the diversity and structuring of freshwater microbial communities beyond the patterns revealed by tracing their distribution in the landscape with common taxonomic markers such as the ribosomal RNA. To address this gap in knowledge, metagenomes from temperate lakes were compared to selected marine metagenomes. Taxonomic analyses of rRNA genes in these freshwater metagenomes confirm the previously reported dominance of a limited subset of uncultured lineages of freshwater bacteria, whereas Archaea were rare. Diversification into marine and freshwater microbial lineages was also reflected in phylogenies of functional genes, and there were also significant differences in functional beta-diversity. The pathways and functions that accounted for these differences are involved in osmoregulation, active transport, carbohydrate and amino acid metabolism. Moreover, predicted genes orthologous to active transporters and recalcitrant organic matter degradation were more common in microbial genomes from oligotrophic versus eutrophic lakes. This comparative metagenomic analysis allowed us to formulate a general hypothesis that oceanic- compared with freshwater-dwelling microorganisms, invest more in metabolism of amino acids and that strategies of carbohydrate metabolism differ significantly between marine and freshwater microbial communities. PMID:24118837

Long-term organic-inorganic fertilization ensures great soil productivity and bacterial diversity after natural-to-agricultural ecosystem conversion.

PubMed

Xun, Weibing; Xu, Zhihui; Li, Wei; Ren, Yi; Huang, Ting; Ran, Wei; Wang, Boren; Shen, Qirong; Zhang, Ruifu

2016-09-01

Natural ecosystems comprise the planet's wild plant and animal resources, but large tracts of land have been converted to agroecosystems to support the demand for agricultural products. This conversion limits the number of plant species and decreases the soil biological diversity. Here we used high-throughput 16S rRNA gene sequencing to evaluate the responses of soil bacterial communities in long-term converted and fertilized red soils (a type of Ferralic Cambisol). We observed that soil bacterial diversity was strongly affected by different types of fertilization management. Oligotrophic bacterial taxa demonstrated large relative abundances in chemically fertilized soil, whereas copiotrophic bacterial taxa were found in large relative abundances in organically fertilized and fallow management soils. Only organic-inorganic fertilization exhibited the same local taxonomic and phylogenetic diversity as that of a natural ecosystem. However, the independent use of organic or inorganic fertilizer reduced local taxonomic and phylogenetic diversity and caused biotic homogenization. This study demonstrated that the homogenization of bacterial communities caused by natural-to-agricultural ecosystem conversion can be mitigated by employing rational organic-inorganic fertilization management.

Evaluation of the Systematic Status of Geographical Variations in Arcuphantes hibanus (Arachnida: Araneae: Linyphiidae), with Descriptions of Two New Species.

PubMed

Nakano, Takafumi; Ihara, Yoh; Kumasaki, Yusuke; Baba, Yuki G; Tomikawa, Ko

2017-08-01

The systematic status of geographical variants of Arcuphantes hibanus Saito, 1992 belonging to the A. longiscapus species group, indigenous to western Honshu and Shikoku, Japan, was evaluated using morphological and molecular data. Two species, A. enmusubi Ihara, Nakano and Tomikawa, sp. nov. and A. occidentalis Ihara, Nakano and Tomikawa, sp. nov., are described, and A. hibanus is redescribed with redefinition of its taxonomic status. These three species are diagnosed by the characteristics of paracymbium, pseudolamella, and epigynal basal part. Phylogenetic trees obtained with mitochondrial cytochrome c oxidase subunit I and 16S rRNA markers showed that the variants are mutually genetically highly diverged. However, the mtDNA phylogenies failed to recover the monophyly of A. hibanus redefined herein. Contrary to the mtDNA phylogenetic analyses, a neighbor-network analysis of nuclear internal transcribed spacer 1 sequences of A. hibanus, A. enmusubi and A. occidentalis spiders showed that each of them forms a cluster. The results of mitochondrial and nuclear DNA analyses in each of the three species are briefly discussed, along with their taxonomic identities.

Metaproteogenomics Reveals Taxonomic and Functional Changes between Cecal and Fecal Microbiota in Mouse

PubMed Central

Tanca, Alessandro; Manghina, Valeria; Fraumene, Cristina; Palomba, Antonio; Abbondio, Marcello; Deligios, Massimo; Silverman, Michael; Uzzau, Sergio

2017-01-01

Previous studies on mouse models report that cecal and fecal microbial communities may differ in the taxonomic structure, but little is known about their respective functional activities. Here, we employed a metaproteogenomic approach, including 16S rRNA gene sequencing, shotgun metagenomics and shotgun metaproteomics, to analyze the microbiota of paired mouse cecal contents (CCs) and feces, with the aim of identifying changes in taxon-specific functions. As a result, Gram-positive anaerobes were observed as considerably higher in CCs, while several key enzymes, involved in oxalate degradation, glutamate/glutamine metabolism, and redox homeostasis, and most actively expressed by Bacteroidetes, were clearly more represented in feces. On the whole, taxon and function abundance appeared to vary consistently with environmental changes expected to occur throughout the transit from the cecum to outside the intestine, especially when considering metaproteomic data. The results of this study indicate that functional and metabolic differences exist between CC and stool samples, paving the way to further metaproteogenomic investigations aimed at elucidating the functional dynamics of the intestinal microbiota. PMID:28352255

Taxonomic profiling of bacterial community structure from coastal sediment of Alang-Sosiya shipbreaking yard near Bhavnagar, India.

PubMed

Patel, Vilas; Munot, Hitendra; Shah, Varun; Shouche, Yogesh S; Madamwar, Datta

2015-12-30

The Alang-Sosiya shipbreaking yard (ASSBY) is considered the largest of its kind in the world, and a major source of anthropogenic pollutants. The aim of this study was to investigate the impact of shipbreaking activities on the bacterial community structure with a combination of culture-dependent and culture-independent approaches. In the culture-dependent approach, 200 bacterial cultures were isolated and analyzed by molecular fingerprinting and 16S ribosomal RNA (r-RNA) gene sequencing, as well as being studied for degradation of polycyclic aromatic hydrocarbons (PAHs). In the culture-independent approach, operational taxonomic units (OTUs) were related to eight major phyla, of which Betaproteobacteria (especially Acidovorax) was predominantly found in the polluted sediments of ASSBY and Gammaproteobacteria in the pristine sediment sample. The statistical approaches showed a significant difference in the bacterial community structure between the pristine and polluted sediments. To the best of our knowledge, this is the first study investigating the effect of shipbreaking activity on the bacterial community structure of the coastal sediment at ASSBY. Copyright © 2015. Published by Elsevier Ltd.

Novel chytrid lineages dominate fungal sequences in diverse marine and freshwater habitats

NASA Astrophysics Data System (ADS)

Comeau, André M.; Vincent, Warwick F.; Bernier, Louis; Lovejoy, Connie

2016-07-01

In aquatic environments, fungal communities remain little studied despite their taxonomic and functional diversity. To extend the ecological coverage of this group, we conducted an in-depth analysis of fungal sequences within our collection of 3.6 million V4 18S rRNA pyrosequences originating from 319 individual marine (including sea-ice) and freshwater samples from libraries generated within diverse projects studying Arctic and temperate biomes in the past decade. Among the ~1.7 million post-filtered reads of highest taxonomic and phylogenetic quality, 23,263 fungal sequences were identified. The overall mean proportion was 1.35%, but with large variability; for example, from 0.01 to 59% of total sequences for Arctic seawater samples. Almost all sample types were dominated by Chytridiomycota-like sequences, followed by moderate-to-minor contributions of Ascomycota, Cryptomycota and Basidiomycota. Species and/or strain richness was high, with many novel sequences and high niche separation. The affinity of the most common reads to phytoplankton parasites suggests that aquatic fungi deserve renewed attention for their role in algal succession and carbon cycling.

Towards a phylogenetic classification of Leptothecata (Cnidaria, Hydrozoa)

PubMed Central

Maronna, Maximiliano M.; Miranda, Thaís P.; Peña Cantero, Álvaro L.; Barbeitos, Marcos S.; Marques, Antonio C.

2016-01-01

Leptothecata are hydrozoans whose hydranths are covered by perisarc and gonophores and whose medusae bear gonads on their radial canals. They develop complex polypoid colonies and exhibit considerable morphological variation among species with respect to growth, defensive structures and mode of development. For instance, several lineages within this order have lost the medusa stage. Depending on the author, traditional taxonomy in hydrozoans may be either polyp- or medusa-oriented. Therefore, the absence of the latter stage in some lineages may lead to very different classification schemes. Molecular data have proved useful in elucidating this taxonomic challenge. We analyzed a super matrix of new and published rRNA gene sequences (16S, 18S and 28S), employing newly proposed methods to measure branch support and improve phylogenetic signal. Our analysis recovered new clades not recognized by traditional taxonomy and corroborated some recently proposed taxa. We offer a thorough taxonomic revision of the Leptothecata, erecting new orders, suborders, infraorders and families. We also discuss the origination and diversification dynamics of the group from a macroevolutionary perspective. PMID:26821567

Microarray Analysis of Microbiota of Gingival Lesions in Noma Patients

PubMed Central

Huyghe, Antoine; François, Patrice; Mombelli, Andrea; Tangomo, Manuela; Girard, Myriam; Baratti-Mayer, Denise; Bolivar, Ignacio; Pittet, Didier; Schrenzel, Jacques

2013-01-01

Noma (cancrum oris) is a gangrenous disease of unknown etiology affecting the maxillo-facial region of young children in extremely limited resource countries. In an attempt to better understand the microbiological events occurring during this disease, we used phylogenetic and low-density microarrays targeting the 16S rRNA gene to characterize the gingival flora of acute noma and acute necrotizing gingivitis (ANG) lesions, and compared them to healthy control subjects of the same geographical and social background. Our observations raise doubts about Fusobacterium necrophorum, a previously suspected causative agent of noma, as this species was not associated with noma lesions. Various oral pathogens were more abundant in noma lesions, notably Atopobium spp., Prevotella intermedia, Peptostreptococcus spp., Streptococcus pyogenes and Streptococcus anginosus. On the other hand, pathogens associated with periodontal diseases such as Aggregatibacter actinomycetemcomitans, Capnocytophaga spp., Porphyromonas spp. and Fusobacteriales were more abundant in healthy controls. Importantly, the overall loss of bacterial diversity observed in noma samples as well as its homology to that of ANG microbiota supports the hypothesis that ANG might be the immediate step preceding noma. PMID:24086784

Fast, accurate and easy-to-pipeline methods for amplicon sequence processing

NASA Astrophysics Data System (ADS)

Antonielli, Livio; Sessitsch, Angela

2016-04-01

Next generation sequencing (NGS) technologies established since years as an essential resource in microbiology. While on the one hand metagenomic studies can benefit from the continuously increasing throughput of the Illumina (Solexa) technology, on the other hand the spreading of third generation sequencing technologies (PacBio, Oxford Nanopore) are getting whole genome sequencing beyond the assembly of fragmented draft genomes, making it now possible to finish bacterial genomes even without short read correction. Besides (meta)genomic analysis next-gen amplicon sequencing is still fundamental for microbial studies. Amplicon sequencing of the 16S rRNA gene and ITS (Internal Transcribed Spacer) remains a well-established widespread method for a multitude of different purposes concerning the identification and comparison of archaeal/bacterial (16S rRNA gene) and fungal (ITS) communities occurring in diverse environments. Numerous different pipelines have been developed in order to process NGS-derived amplicon sequences, among which Mothur, QIIME and USEARCH are the most well-known and cited ones. The entire process from initial raw sequence data through read error correction, paired-end read assembly, primer stripping, quality filtering, clustering, OTU taxonomic classification and BIOM table rarefaction as well as alternative "normalization" methods will be addressed. An effective and accurate strategy will be presented using the state-of-the-art bioinformatic tools and the example of a straightforward one-script pipeline for 16S rRNA gene or ITS MiSeq amplicon sequencing will be provided. Finally, instructions on how to automatically retrieve nucleotide sequences from NCBI and therefore apply the pipeline to targets other than 16S rRNA gene (Greengenes, SILVA) and ITS (UNITE) will be discussed.

The chaperonin-60 universal target is a barcode for bacteria that enables de novo assembly of metagenomic sequence data.

PubMed

Links, Matthew G; Dumonceaux, Tim J; Hemmingsen, Sean M; Hill, Janet E

2012-01-01

Barcoding with molecular sequences is widely used to catalogue eukaryotic biodiversity. Studies investigating the community dynamics of microbes have relied heavily on gene-centric metagenomic profiling using two genes (16S rRNA and cpn60) to identify and track Bacteria. While there have been criteria formalized for barcoding of eukaryotes, these criteria have not been used to evaluate gene targets for other domains of life. Using the framework of the International Barcode of Life we evaluated DNA barcodes for Bacteria. Candidates from the 16S rRNA gene and the protein coding cpn60 gene were evaluated. Within complete bacterial genomes in the public domain representing 983 species from 21 phyla, the largest difference between median pairwise inter- and intra-specific distances ("barcode gap") was found from cpn60. Distribution of sequence diversity along the ∼555 bp cpn60 target region was remarkably uniform. The barcode gap of the cpn60 universal target facilitated the faithful de novo assembly of full-length operational taxonomic units from pyrosequencing data from a synthetic microbial community. Analysis supported the recognition of both 16S rRNA and cpn60 as DNA barcodes for Bacteria. The cpn60 universal target was found to have a much larger barcode gap than 16S rRNA suggesting cpn60 as a preferred barcode for Bacteria. A large barcode gap for cpn60 provided a robust target for species-level characterization of data. The assembly of consensus sequences for barcodes was shown to be a reliable method for the identification and tracking of novel microbes in metagenomic studies.

Different bacterial communities in ectomycorrhizae and surrounding soil

PubMed Central

Vik, Unni; Logares, Ramiro; Blaalid, Rakel; Halvorsen, Rune; Carlsen, Tor; Bakke, Ingrid; Kolstø, Anne-Brit; Økstad, Ole Andreas; Kauserud, Håvard

2013-01-01

Several eukaryotic symbioses have shown to host a rich diversity of prokaryotes that interact with their hosts. Here, we study bacterial communities associated with ectomycorrhizal root systems of Bistorta vivipara compared to bacterial communities in bulk soil using pyrosequencing of 16S rRNA amplicons. A high richness of Operational Taxonomic Units (OTUs) was found in plant roots (3,571 OTUs) and surrounding soil (3,476 OTUs). The community composition differed markedly between these two environments. Actinobacteria, Armatimonadetes, Chloroflexi and OTUs unclassified at phylum level were significantly more abundant in plant roots than in soil. A large proportion of the OTUs, especially those in plant roots, presented low similarity to Sanger 16S rRNA reference sequences, suggesting novel bacterial diversity in ectomycorrhizae. Furthermore, the bacterial communities of the plant roots were spatially structured up to a distance of 60 cm, which may be explained by bacteria using fungal hyphae as a transport vector. The analyzed ectomycorrhizae presents a distinct microbiome, which likely influence the functioning of the plant-fungus symbiosis. PMID:24326907

The impact of tropical forest logging and oil palm agriculture on the soil microbiome.

PubMed

Tripathi, Binu M; Edwards, David P; Mendes, Lucas William; Kim, Mincheol; Dong, Ke; Kim, Hyoki; Adams, Jonathan M

2016-05-01

Selective logging and forest conversion to oil palm agriculture are rapidly altering tropical forests. However, functional responses of the soil microbiome to these land-use changes are poorly understood. Using 16S rRNA gene and shotgun metagenomic sequencing, we compared composition and functional attributes of soil biota between unlogged, once-logged and twice-logged rainforest, and areas converted to oil palm plantations in Sabah, Borneo. Although there was no significant effect of logging history, we found a significant difference between the taxonomic and functional composition of both primary and logged forests and oil palm. Oil palm had greater abundances of genes associated with DNA, RNA, protein metabolism and other core metabolic functions, but conversely, lower abundance of genes associated with secondary metabolism and cell-cell interactions, indicating less importance of antagonism or mutualism in the more oligotrophic oil palm environment. Overall, these results show a striking difference in taxonomic composition and functional gene diversity of soil microorganisms between oil palm and forest, but no significant difference between primary forest and forest areas with differing logging history. This reinforces the view that logged forest retains most features and functions of the original soil community. However, networks based on strong correlations between taxonomy and functions showed that network complexity is unexpectedly increased due to both logging and oil palm agriculture, which suggests a pervasive effect of both land-use changes on the interaction of soil microbes. © 2016 John Wiley & Sons Ltd.

Morphological, Physiological, and Taxonomic Characterization of Actinobacterial Isolates Living as Endophytes of Cacao Pods and Cacao Seeds

PubMed Central

Tchinda, Romaric Armel Mouafo; Boudjeko, Thaddée; Simao-Beaunoir, Anne-Marie; Lerat, Sylvain; Tsala, Éric; Monga, Ernest; Beaulieu, Carole

2016-01-01

Vascular plants are commonly colonized by endophytic actinobacteria. However, very little is known about the relationship between these microorganisms and cacao fruits. In order to determine the physiological and taxonomic relationships between the members of this community, actinobacteria were isolated from cacao fruits and seeds. Among the 49 isolates recovered, 11 morphologically distinct isolates were selected for further characterization. Sequencing of the 16S rRNA gene allowed the partition of the selected isolates into three phylogenetic clades. Most of the selected endophytic isolates belonged to the Streptomyces violaceusniger clade. Physiological characterization was carried out and a similarity index was used to cluster the isolates. However, clustering based on physiological properties did not match phylogenetic lineages. Isolates were also characterized for traits commonly associated with plant growth-promoting bacteria, including antibiosis and auxin biosynthesis. All isolates exhibited resistance to geldanamycin, whereas only two isolates were shown to produce this antibiotic. Endophytes were inoculated on radish seedlings and most isolates were found to possess plant growth-promoting abilities. These endophytic actinobacteria inhibited the growth of various plant pathogenic fungi and/or bacteria. The present study showed that S. violaceusniger clade members represent a significant part of the actinobacterial community living as endophytes in cacao fruits and seeds. While several members of this clade are known to be geldanamycin producers and efficient biocontrol agents of plant diseases, we herein established the endophytic lifestyle of some of these microorganisms, demonstrating their potential as plant health agents. PMID:26947442

Cellulose-Enriched Microbial Communities from Leaf-Cutter Ant (Atta colombica) Refuse Dumps Vary in Taxonomic Composition and Degradation Ability

DOE PAGES

Lewin, Gina R.; Johnson, Amanda L.; Soto, Rolando D. Moreira; ...

2016-03-21

Deconstruction of the cellulose in plant cell walls is critical for carbon flow through ecosystems and for the production of sustainable cellulosic biofuels. Our understanding of cellulose deconstruction is largely limited to the study of microbes in isolation, but in nature, this process is driven by microbes within complex communities. In Neotropical forests, microbes in leaf-cutter ant refuse dumps are important for carbon turnover. These dumps consist of decaying plant material and a diverse bacterial community, as shown here by electron microscopy. To study the portion of the community capable of cellulose degradation, we performed enrichments on cellulose using materialmore » from five Atta colombica refuse dumps. The ability of enriched communities to degrade cellulose varied significantly across refuse dumps. 16S rRNA gene amplicon sequencing of enriched samples identified that the community structure correlated with refuse dump and with degradation ability. Overall, samples were dominated by Bacteroidetes, Gammaproteobacteria, and Betaproteobacteria. Half of abundant operational taxonomic units (OTUs) across samples were classified within general containing known cellulose degraders, including Acidovorax, the most abundant OTU detected across samples, which was positively correlated with cellulolytic ability. Lastly, a representative Acidovorax strain was isolated, but did not grow on cellulose alone. Phenotypic and compositional analyses of enrichment cultures, such as those presented here, help link community composition with cellulolytic ability and provide insight into the complexity of community-based cellulose degradation.« less

Morphological, Physiological, and Taxonomic Characterization of Actinobacterial Isolates Living as Endophytes of Cacao Pods and Cacao Seeds.

PubMed

Tchinda, Romaric Armel Mouafo; Boudjeko, Thaddée; Simao-Beaunoir, Anne-Marie; Lerat, Sylvain; Tsala, Éric; Monga, Ernest; Beaulieu, Carole

2016-01-01

Vascular plants are commonly colonized by endophytic actinobacteria. However, very little is known about the relationship between these microorganisms and cacao fruits. In order to determine the physiological and taxonomic relationships between the members of this community, actinobacteria were isolated from cacao fruits and seeds. Among the 49 isolates recovered, 11 morphologically distinct isolates were selected for further characterization. Sequencing of the 16S rRNA gene allowed the partition of the selected isolates into three phylogenetic clades. Most of the selected endophytic isolates belonged to the Streptomyces violaceusniger clade. Physiological characterization was carried out and a similarity index was used to cluster the isolates. However, clustering based on physiological properties did not match phylogenetic lineages. Isolates were also characterized for traits commonly associated with plant growth-promoting bacteria, including antibiosis and auxin biosynthesis. All isolates exhibited resistance to geldanamycin, whereas only two isolates were shown to produce this antibiotic. Endophytes were inoculated on radish seedlings and most isolates were found to possess plant growth-promoting abilities. These endophytic actinobacteria inhibited the growth of various plant pathogenic fungi and/or bacteria. The present study showed that S. violaceusniger clade members represent a significant part of the actinobacterial community living as endophytes in cacao fruits and seeds. While several members of this clade are known to be geldanamycin producers and efficient biocontrol agents of plant diseases, we herein established the endophytic lifestyle of some of these microorganisms, demonstrating their potential as plant health agents.

Cellulose-Enriched Microbial Communities from Leaf-Cutter Ant (Atta colombica) Refuse Dumps Vary in Taxonomic Composition and Degradation Ability

PubMed Central

Lewin, Gina R.; Johnson, Amanda L.; Soto, Rolando D. Moreira; Perry, Kailene; Book, Adam J.; Horn, Heidi A.; Pinto-Tomás, Adrián A.; Currie, Cameron R.

2016-01-01

Deconstruction of the cellulose in plant cell walls is critical for carbon flow through ecosystems and for the production of sustainable cellulosic biofuels. Our understanding of cellulose deconstruction is largely limited to the study of microbes in isolation, but in nature, this process is driven by microbes within complex communities. In Neotropical forests, microbes in leaf-cutter ant refuse dumps are important for carbon turnover. These dumps consist of decaying plant material and a diverse bacterial community, as shown here by electron microscopy. To study the portion of the community capable of cellulose degradation, we performed enrichments on cellulose using material from five Atta colombica refuse dumps. The ability of enriched communities to degrade cellulose varied significantly across refuse dumps. 16S rRNA gene amplicon sequencing of enriched samples identified that the community structure correlated with refuse dump and with degradation ability. Overall, samples were dominated by Bacteroidetes, Gammaproteobacteria, and Betaproteobacteria. Half of abundant operational taxonomic units (OTUs) across samples were classified within genera containing known cellulose degraders, including Acidovorax, the most abundant OTU detected across samples, which was positively correlated with cellulolytic ability. A representative Acidovorax strain was isolated, but did not grow on cellulose alone. Phenotypic and compositional analyses of enrichment cultures, such as those presented here, help link community composition with cellulolytic ability and provide insight into the complexity of community-based cellulose degradation. PMID:26999749

Cellulose-Enriched Microbial Communities from Leaf-Cutter Ant (Atta colombica) Refuse Dumps Vary in Taxonomic Composition and Degradation Ability

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lewin, Gina R.; Johnson, Amanda L.; Soto, Rolando D. Moreira

Deconstruction of the cellulose in plant cell walls is critical for carbon flow through ecosystems and for the production of sustainable cellulosic biofuels. Our understanding of cellulose deconstruction is largely limited to the study of microbes in isolation, but in nature, this process is driven by microbes within complex communities. In Neotropical forests, microbes in leaf-cutter ant refuse dumps are important for carbon turnover. These dumps consist of decaying plant material and a diverse bacterial community, as shown here by electron microscopy. To study the portion of the community capable of cellulose degradation, we performed enrichments on cellulose using materialmore » from five Atta colombica refuse dumps. The ability of enriched communities to degrade cellulose varied significantly across refuse dumps. 16S rRNA gene amplicon sequencing of enriched samples identified that the community structure correlated with refuse dump and with degradation ability. Overall, samples were dominated by Bacteroidetes, Gammaproteobacteria, and Betaproteobacteria. Half of abundant operational taxonomic units (OTUs) across samples were classified within general containing known cellulose degraders, including Acidovorax, the most abundant OTU detected across samples, which was positively correlated with cellulolytic ability. Lastly, a representative Acidovorax strain was isolated, but did not grow on cellulose alone. Phenotypic and compositional analyses of enrichment cultures, such as those presented here, help link community composition with cellulolytic ability and provide insight into the complexity of community-based cellulose degradation.« less

Pseudomonas gallaeciensis sp. nov., isolated from crude-oil-contaminated intertidal sand samples after the Prestige oil spill.

PubMed

Mulet, Magdalena; Sánchez, David; Rodríguez, Ana C; Nogales, Balbina; Bosch, Rafael; Busquets, Antonio; Gomila, Margarita; Lalucat, Jorge; García-Valdés, Elena

2018-04-11

Strains V113 T , V92 and V120 have been isolated from sand samples taken at the Atlantic intertidal shore in Galicia, Spain, after the Prestige oil spill. A preliminary analysis of the 16S rRNA and the partial rpoD gene sequences indicated that these strains belonged to the Pseudomonas genus, but they were distinct from any known Pseudomonas species. They were extensively characterized by a polyphasic taxonomic approach and phylogenetic data that confirmed that these strains belonged to the Pseudomonas pertucinogena group. Phylogenetic analysis of 16S rRNA, gyrB and rpoD gene sequences showed that the three strains were 99% similar and were closely related to members of the P. pertucinogena group, with less than 94% similarity to strains of established species; Pseudomonas pachastrellae was the closest relative. The Average Nucleotide Index based on blast values was 89.0% between V113 T and the P. pachastrellae type strain, below the accepted species level (95%). The predominant cellular fatty acid contents and whole cell protein profiles determined by MALDI-TOF mass spectrometry also differentiated the studied strains from known Pseudomonas species. We therefore conclude that strains V113 T , V92 and V120 represent a novel species of Pseudomonas, for which the name Pseudomonas gallaeciensis is proposed; the type strain is V113 T (=CCUG 67583 T =LMG 29038 T ). Copyright © 2018 Elsevier GmbH. All rights reserved.

Microbiota and Metatranscriptome Changes Accompanying the Onset of Gingivitis

PubMed Central

2018-01-01

ABSTRACT Over half of adults experience gingivitis, a mild yet treatable form of periodontal disease caused by the overgrowth of oral microbes. Left untreated, gingivitis can progress to a more severe and irreversible disease, most commonly chronic periodontitis. While periodontal diseases are associated with a shift in the oral microbiota composition, it remains unclear how this shift impacts microbiota function early in disease progression. Here, we analyzed the transition from health to gingivitis through both 16S v4-v5 rRNA amplicon and metatranscriptome sequencing of subgingival plaque samples from individuals undergoing an experimental gingivitis treatment. Beta-diversity analysis of 16S rRNA reveals that samples cluster based on disease severity and patient but not by oral hygiene status. Significant shifts in the abundance of several genera occurred during disease transition, suggesting a dysbiosis due to development of gingivitis. Comparing taxonomic abundance with transcriptomic activity revealed concordance of bacterial diversity composition between the two quantification assays in samples originating from both healthy and diseased teeth. Metatranscriptome sequencing analysis indicates that during the early stages of transition to gingivitis, a number of virulence-related transcripts were significantly differentially expressed in individual and across pooled patient samples. Upregulated genes include those involved in proteolytic and nucleolytic processes, while expression levels of those involved in surface structure assembly and other general virulence functions leading to colonization or adaptation within the host are more dynamic. These findings help characterize the transition from health to periodontal disease and identify genes associated with early disease. PMID:29666288

The Egyptian Red Sea coastal microbiome: A study revealing differential microbial responses to diverse anthropogenic pollutants.

PubMed

Mustafa, Ghada A; Abd-Elgawad, Amr; Ouf, Amged; Siam, Rania

2016-07-01

The Red Sea is considered one of the youngest oceanic systems, with unique physical, geochemical and biological characteristics. Tourism, industrialization, extensive fishing, oil processing and shipping are extensive sources of pollution in the Red Sea. We analyzed the geochemical characteristics and microbial community of sediments along the Egyptian coast of the Red Sea. Our sites mainly included 1) four ports used for shipping aluminum, ilmenite and phosphate; 2) a site previously reported to have suffered extensive oil spills; and 3) a site impacted by tourism. Two major datasets for the sediment of ten Red Sea coastal sites were generated; i) a chemical dataset included measurements of carbon, hydrogen, nitrogen and sulfur, metals and selected semi-volatile oil; and ii) a 16S rRNA Pyrotags bacterial metagenomic dataset. Based on the taxonomic assignments of the 16S rRNA Pyrotags to major bacterial groups, we report 30 taxa constituting an Egyptian Red Sea Coastal Microbiome. Bacteria that degrade hydrocarbons were predominant in the majority of the sites, particularly in two ports where they reached up to 76% of the total identified genera. In contrast, sulfate-reducing and sulfate-oxidizing bacteria dominated two lakes at the expense of other hydrocarbon metabolizers. Despite the reported "Egyptian Red Sea Coastal Microbiome," sites with similar anthropogenic pollutants showed unique microbial community abundances. This suggests that the abundance of a specific bacterial community is an evolutionary mechanism induced in response to selected anthropogenic pollutants. Copyright © 2016 Elsevier Ltd. All rights reserved.

Streptomyces aridus sp. nov., isolated from a high altitude Atacama Desert soil and emended description of Streptomyces noboritoensis Isono et al. 1957.

PubMed

Idris, Hamidah; Labeda, David P; Nouioui, Imen; Castro, Jean Franco; Del Carmen Montero-Calasanz, Maria; Bull, Alan T; Asenjo, Juan A; Goodfellow, Michael

2017-05-01

A polyphasic study was undertaken to determine the taxonomic status of a Streptomyces strain which had been isolated from a high altitude Atacama Desert soil and shown to have bioactive properties. The strain, isolate H9 T , was found to have chemotaxonomic, cultural and morphological properties that place it in the genus Streptomyces. 16S rRNA gene sequence analyses showed that the isolate forms a distinct branch at the periphery of a well-delineated subclade in the Streptomyces 16S rRNA gene tree together with the type strains of Streptomyces crystallinus, Streptomyces melanogenes and Streptomyces noboritoensis. Multi-locus sequence analysis (MLSA) based on five house-keeping gene alleles showed that isolate H9 T is closely related to the latter two type strains and to Streptomyces polyantibioticus NRRL B-24448 T . The isolate was distinguished readily from the type strains of S. melanogenes, S. noboritoensis and S. polyantibioticus using a combination of phenotypic properties. Consequently, the isolate is considered to represent a new species of Streptomyces for which the name Streptomyces aridus sp. nov. is proposed; the type strain is H9 T (=NCIMB 14965 T =NRRL B65268 T ). In addition, the MLSA and phenotypic data show that the S. melanogenes and S. noboritoensis type strains belong to a single species, it is proposed that S. melanogenes be recognised as a heterotypic synonym of S. noboritoensis for which an emended description is given.

Molecular techniques revealed highly diverse microbial communities in natural marine biofilms on polystyrene dishes for invertebrate larval settlement.

PubMed

Lee, On On; Chung, Hong Chun; Yang, Jiangke; Wang, Yong; Dash, Swagatika; Wang, Hao; Qian, Pei-Yuan

2014-07-01

Biofilm microbial communities play an important role in the larval settlement response of marine invertebrates. However, the underlying mechanism has yet to be resolved, mainly because of the uncertainties in characterizing members in the communities using traditional 16S rRNA gene-based molecular methods and in identifying the chemical signals involved. In this study, pyrosequencing was used to characterize the bacterial communities in intertidal and subtidal marine biofilms developed during two seasons. We revealed highly diverse biofilm bacterial communities that varied with season and tidal level. Over 3,000 operational taxonomic units with estimates of up to 8,000 species were recovered in a biofilm sample, which is by far the highest number recorded in subtropical marine biofilms. Nineteen phyla were found, of which Cyanobacteria and Proteobacteria were the most dominant one in the intertidal and subtidal biofilms, respectively. Apart from these, Actinobacteria, Bacteroidetes, and Planctomycetes were the major groups recovered in both intertidal and subtidal biofilms, although their relative abundance varied among samples. Full-length 16S rRNA gene clone libraries were constructed for the four biofilm samples and showed similar bacterial compositions at the phylum level to those revealed by pyrosequencing. Laboratory assays confirmed that cyrids of the barnacle Balanus amphitrite preferred to settle on the intertidal rather than subtidal biofilms. This preference was independent of the biofilm bacterial density or biomass but was probably related to the biofilm community structure, particularly, the Proteobacterial and Cyanobacterial groups.

Hexavalent chromium reduction by bacterial consortia and pure strains from an alkaline industrial effluent.

PubMed

Piñón-Castillo, H A; Brito, E M S; Goñi-Urriza, M; Guyoneaud, R; Duran, R; Nevarez-Moorillon, G V; Gutiérrez-Corona, J F; Caretta, C A; Reyna-López, G E

2010-12-01

To characterize the bacterial consortia and isolates selected for their role in hexavalent chromium removal by adsorption and reduction. Bacterial consortia from industrial wastes revealed significant Cr(VI) removal after 15 days when incubated in medium M9 at pH 6·5 and 8·0. The results suggested chromium reduction. The bacterial consortia diversity (T-RFLP based on 16S rRNA gene) indicated a highest number of operational taxonomic units in an alkaline carbonate medium mimicking in situ conditions. However, incubations under such conditions revealed low Cr(VI) removal. Genomic libraries were obtained for the consortia exhibiting optimal Cr(VI) removal (M9 medium at pH 6·5 and 8·0). They revealed the dominance of 16S rRNA gene sequences related to the genera Pseudomonas/Stenotrophomonas or Enterobacter/Halomonas, respectively. Isolates related to Pseudomonas fluorescens and Enterobacter aerogenes were efficient in Cr(VI) reduction and adsorption to the biomass. Cr(VI) reduction was better at neutral pH rather than under in situ conditions (alkaline pH with carbonate). Isolated strains exhibited significant capacity for Cr(VI) reduction and adsorption. Bacterial communities from chromium-contaminated industrial wastes as well as isolates were able to remove Cr(VI). The results suggest a good potential for bioremediation of industrial wastes when optimal conditions are applied. Journal of Applied Microbiology © 2010 The Society for Applied Microbiology. No claim to Mexican Government works.

Spatial and temporal changes in microbial community structure associated with recharge-influenced chemical gradients in a contaminated aquifer

USGS Publications Warehouse

Haack, S.K.; Fogarty, L.R.; West, T.G.; Alm, E.W.; McGuire, J.T.; Long, D.T.; Hyndman, D.W.; Forney, L.J.

2004-01-01

In a contaminated water-table aquifer, we related microbial community structure on aquifer sediments to gradients in 24 geochemical and contaminant variables at five depths, under three recharge conditions. Community amplified ribsosomal DNA restriction analysis (ARDRA) using universal 16S rDNA primers and denaturing gradient gel electrophoresis (DGGE) using bacterial 16S rDNA primers indicated: (i) communities in the anoxic, contaminated central zone were similar regardless of recharge; (ii) after recharge, communities at greatest depth were similar to those in uncontaminated zones; and (iii) after extended lack of recharge, communities at upper and lower aquifer margins differed from communities at the same depths on other dates. General aquifer geochemistry was as important as contaminant or terminal electron accepting process (TEAP) chemistry in discriminant analysis of community groups. The Shannon index of diversity (H) and the evenness index (E), based on DGGE operational taxonomic units (OTUs), were statistically different across community groups and aquifer depths. Archaea or sulphate-reducing bacteria 16S rRNA abundance was not clearly correlated with TEAP chemistry indicative of methanogenesis or sulphate reduction. Eukarya rRNA abundance varied by depth and date from 0 to 13% of the microbial community. This contaminated aquifer is a dynamic ecosystem, with complex interactions between physical, chemical and biotic components, which should be considered in the interpretation of aquifer geochemistry and in the development of conceptual or predictive models for natural attenuation or remediation.

Microbial Signatures of Cadaver Gravesoil During Decomposition.

PubMed

Finley, Sheree J; Pechal, Jennifer L; Benbow, M Eric; Robertson, B K; Javan, Gulnaz T

2016-04-01

Genomic studies have estimated there are approximately 10(3)-10(6) bacterial species per gram of soil. The microbial species found in soil associated with decomposing human remains (gravesoil) have been investigated and recognized as potential molecular determinants for estimates of time since death. The nascent era of high-throughput amplicon sequencing of the conserved 16S ribosomal RNA (rRNA) gene region of gravesoil microbes is allowing research to expand beyond more subjective empirical methods used in forensic microbiology. The goal of the present study was to evaluate microbial communities and identify taxonomic signatures associated with the gravesoil human cadavers. Using 16S rRNA gene amplicon-based sequencing, soil microbial communities were surveyed from 18 cadavers placed on the surface or buried that were allowed to decompose over a range of decomposition time periods (3-303 days). Surface soil microbial communities showed a decreasing trend in taxon richness, diversity, and evenness over decomposition, while buried cadaver-soil microbial communities demonstrated increasing taxon richness, consistent diversity, and decreasing evenness. The results show that ubiquitous Proteobacteria was confirmed as the most abundant phylum in all gravesoil samples. Surface cadaver-soil communities demonstrated a decrease in Acidobacteria and an increase in Firmicutes relative abundance over decomposition, while buried soil communities were consistent in their community composition throughout decomposition. Better understanding of microbial community structure and its shifts over time may be important for advancing general knowledge of decomposition soil ecology and its potential use during forensic investigations.

Phylogeny of 54 representative strains of species in the family Pasteurellaceae as determined by comparison of 16S rRNA sequences.

PubMed Central

Dewhirst, F E; Paster, B J; Olsen, I; Fraser, G J

1992-01-01

Virtually complete 16S rRNA sequences were determined for 54 representative strains of species in the family Pasteurellaceae. Of these strains, 15 were Pasteurella, 16 were Actinobacillus, and 23 were Haemophilus. A phylogenetic tree was constructed based on sequence similarity, using the Neighbor-Joining method. Fifty-three of the strains fell within four large clusters. The first cluster included the type strains of Haemophilus influenzae, H. aegyptius, H. aphrophilus, H. haemolyticus, H. paraphrophilus, H. segnis, and Actinobacillus actinomycetemcomitans. This cluster also contained A. actinomycetemcomitans FDC Y4, ATCC 29522, ATCC 29523, and ATCC 29524 and H. aphrophilus NCTC 7901. The second cluster included the type strains of A. seminis and Pasteurella aerogenes and H. somnus OVCG 43826. The third cluster was composed of the type strains of Pasteurella multocida, P. anatis, P. avium, P. canis, P. dagmatis, P. gallinarum, P. langaa, P. stomatis, P. volantium, H. haemoglobinophilus, H. parasuis, H. paracuniculus, H. paragallinarum, and A. capsulatus. This cluster also contained Pasteurella species A CCUG 18782, Pasteurella species B CCUG 19974, Haemophilus taxon C CAPM 5111, H. parasuis type 5 Nagasaki, P. volantium (H. parainfluenzae) NCTC 4101, and P. trehalosi NCTC 10624. The fourth cluster included the type strains of Actinobacillus lignieresii, A. equuli, A. pleuropneumoniae, A. suis, A. ureae, H. parahaemolyticus, H. parainfluenzae, H. paraphrohaemolyticus, H. ducreyi, and P. haemolytica. This cluster also contained Actinobacillus species strain CCUG 19799 (Bisgaard taxon 11), A. suis ATCC 15557, H. ducreyi ATCC 27722 and HD 35000, Haemophilus minor group strain 202, and H. parainfluenzae ATCC 29242. The type strain of P. pneumotropica branched alone to form a fifth group. The branching of the Pasteurellaceae family tree was quite complex. The four major clusters contained multiple subclusters. The clusters contained both rapidly and slowly evolving strains (indicated by differing numbers of base changes incorporated into the 16S rRNA sequence relative to outgroup organisms). While the results presented a clear picture of the phylogenetic relationships, the complexity of the branching will make division of the family into genera a difficult and somewhat subjective task. We do not suggest any taxonomic changes at this time. PMID:1548238

Fast discovery and visualization of conserved regions in DNA sequences using quasi-alignment

PubMed Central

2013-01-01

Background Next Generation Sequencing techniques are producing enormous amounts of biological sequence data and analysis becomes a major computational problem. Currently, most analysis, especially the identification of conserved regions, relies heavily on Multiple Sequence Alignment and its various heuristics such as progressive alignment, whose run time grows with the square of the number and the length of the aligned sequences and requires significant computational resources. In this work, we present a method to efficiently discover regions of high similarity across multiple sequences without performing expensive sequence alignment. The method is based on approximating edit distance between segments of sequences using p-mer frequency counts. Then, efficient high-throughput data stream clustering is used to group highly similar segments into so called quasi-alignments. Quasi-alignments have numerous applications such as identifying species and their taxonomic class from sequences, comparing sequences for similarities, and, as in this paper, discovering conserved regions across related sequences. Results In this paper, we show that quasi-alignments can be used to discover highly similar segments across multiple sequences from related or different genomes efficiently and accurately. Experiments on a large number of unaligned 16S rRNA sequences obtained from the Greengenes database show that the method is able to identify conserved regions which agree with known hypervariable regions in 16S rRNA. Furthermore, the experiments show that the proposed method scales well for large data sets with a run time that grows only linearly with the number and length of sequences, whereas for existing multiple sequence alignment heuristics the run time grows super-linearly. Conclusion Quasi-alignment-based algorithms can detect highly similar regions and conserved areas across multiple sequences. Since the run time is linear and the sequences are converted into a compact clustering model, we are able to identify conserved regions fast or even interactively using a standard PC. Our method has many potential applications such as finding characteristic signature sequences for families of organisms and studying conserved and variable regions in, for example, 16S rRNA. PMID:24564200

Fast discovery and visualization of conserved regions in DNA sequences using quasi-alignment.

PubMed

Nagar, Anurag; Hahsler, Michael

2013-01-01

Next Generation Sequencing techniques are producing enormous amounts of biological sequence data and analysis becomes a major computational problem. Currently, most analysis, especially the identification of conserved regions, relies heavily on Multiple Sequence Alignment and its various heuristics such as progressive alignment, whose run time grows with the square of the number and the length of the aligned sequences and requires significant computational resources. In this work, we present a method to efficiently discover regions of high similarity across multiple sequences without performing expensive sequence alignment. The method is based on approximating edit distance between segments of sequences using p-mer frequency counts. Then, efficient high-throughput data stream clustering is used to group highly similar segments into so called quasi-alignments. Quasi-alignments have numerous applications such as identifying species and their taxonomic class from sequences, comparing sequences for similarities, and, as in this paper, discovering conserved regions across related sequences. In this paper, we show that quasi-alignments can be used to discover highly similar segments across multiple sequences from related or different genomes efficiently and accurately. Experiments on a large number of unaligned 16S rRNA sequences obtained from the Greengenes database show that the method is able to identify conserved regions which agree with known hypervariable regions in 16S rRNA. Furthermore, the experiments show that the proposed method scales well for large data sets with a run time that grows only linearly with the number and length of sequences, whereas for existing multiple sequence alignment heuristics the run time grows super-linearly. Quasi-alignment-based algorithms can detect highly similar regions and conserved areas across multiple sequences. Since the run time is linear and the sequences are converted into a compact clustering model, we are able to identify conserved regions fast or even interactively using a standard PC. Our method has many potential applications such as finding characteristic signature sequences for families of organisms and studying conserved and variable regions in, for example, 16S rRNA.

Photobacterium damselae ssp. piscicida: detection by direct amplification of 16S rRNA gene sequences and genotypic variation as determined by amplified fragment length polymorphism (AFLP).

PubMed

Kvitt, H; Ucko, M; Colorni, A; Batargias, C; Zlotkin, A; Knibb, W

2002-04-05

A PCR protocol for the rapid diagnosis of fish 'pasteurellosis' based on 16S rRNA gene sequences was developed. The procedure combines low annealing temperature that detects low titers of Photobacterium damselae but also related species, and high annealing temperature for the specific identification of P. damselae directly from infected fish. The PCR protocol was validated on 19 piscine isolates of P. damselae ssp. piscicida from different geographic regions (Japan, Italy, Spain, Greece and Israel), on spontaneously infected sea bream Sparus aurata and sea bass Dicentrarchus labrax, and on closely related American Type Culture Collection (ATCC) reference strains. PCR using high annealing temperature (64 degrees C) discriminated between P. damselae and closely related reference strains, including P. histaminum. Sixteen isolates of P. damselae ssp. piscicida, 2 P. damselae ssp. piscicida reference strains and 1 P. damselae ssp. damselae reference strain were subjected to Amplified Fragment Length Polymorphism (AFLP) analysis, and a similarity matrix was produced. Accordingly, the Japanese isolates of P. damselae ssp. piscicida were distinguished from the Mediterranean/European isolates at a cut-off value of 83% similarity. A further subclustering at a cut-off value of 97% allowed discrimination between the Israeli P. damselae ssp. piscicida isolates and the other Mediterranean/European isolates. The combination of PCR direct amplification and AFLP provides a 2-step procedure, where P. damselae is rapidly identified at genus level on the basis of its 16S rRNA gene sequence and then grouped into distinct clusters on the basis of AFLP polymorphisms. The first step of direct amplification is highly sensitive and has immediate practical consequences, offering fish farmers a rapid diagnosis, while the AFLP is more specific and detects intraspecific variation which, in our study, also reflected geographic correspondence. Because of its superior discriminative properties, AFLP can be an important tool for epidemiological and taxonomic studies of this highly homogeneous genus.

New rRNA Gene-Based Phylogenies of the Alphaproteobacteria Provide Perspective on Major Groups, Mitochondrial Ancestry and Phylogenetic Instability

PubMed Central

Ferla, Matteo P.; Thrash, J. Cameron; Giovannoni, Stephen J.; Patrick, Wayne M.

2013-01-01

Bacteria in the class Alphaproteobacteria have a wide variety of lifestyles and physiologies. They include pathogens of humans and livestock, agriculturally valuable strains, and several highly abundant marine groups. The ancestor of mitochondria also originated in this clade. Despite significant effort to investigate the phylogeny of the Alphaproteobacteria with a variety of methods, there remains considerable disparity in the placement of several groups. Recent emphasis on phylogenies derived from multiple protein-coding genes remains contentious due to disagreement over appropriate gene selection and the potential influences of systematic error. We revisited previous investigations in this area using concatenated alignments of the small and large subunit (SSU and LSU) rRNA genes, as we show here that these loci have much lower GC bias than whole genomes. This approach has allowed us to update the canonical 16S rRNA gene tree of the Alphaproteobacteria with additional important taxa that were not previously included, and with added resolution provided by concatenating the SSU and LSU genes. We investigated the topological stability of the Alphaproteobacteria by varying alignment methods, rate models, taxon selection and RY-recoding to circumvent GC content bias. We also introduce RYMK-recoding and show that it avoids some of the information loss in RY-recoding. We demonstrate that the topology of the Alphaproteobacteria is sensitive to inclusion of several groups of taxa, but it is less affected by the choice of alignment and rate methods. The majority of topologies and comparative results from Approximately Unbiased tests provide support for positioning the Rickettsiales and the mitochondrial branch within a clade. This composite clade is a sister group to the abundant marine SAR11 clade (Pelagibacterales). Furthermore, we add support for taxonomic assignment of several recently sequenced taxa. Accordingly, we propose three subclasses within the Alphaproteobacteria: the Caulobacteridae, the Rickettsidae, and the Magnetococcidae. PMID:24349502

Characterization of a Wild, Novel Nisin A-Producing Lactococcus Strain with an L. lactis subsp. cremoris Genotype and an L. lactis subsp. lactis Phenotype, Isolated from Greek Raw Milk

PubMed Central

Parapouli, Maria; Delbès-Paus, Céline; Kakouri, Athanasia; Koukkou, Anna-Irini; Montel, Marie-Christine

2013-01-01

Several molecular taxonomic studies have revealed that many natural (wild) Lactococcus lactis strains of dairy origin which are phenotypically representative of the L. lactis subspecies lactis cluster genotypically within subspecies cremoris and vice versa. Recently, we isolated two wild nisin-producing (Nis+) L. lactis strains, M78 and M104, of the lactis phenotype from Greek raw milk (J. Samelis, A. Lianou, A. Kakouri, C. Delbès, I. Rogelj, B. B. Matijašic, and M. C. Montel, J. Food Prot. 72:783–790, 2009); strain M78 possess a novel nisin A sequence (GenBank accession number HM219853). In this study, the actual subspecies identity of M78 and M104 isolates was elucidated, using 16S rRNA and acmA (encoding lactococcal N-acetylmuramidase) gene and histidine biosynthesis operon polymorphisms and 16S rRNA and ldh (encoding lactate dehydrogenase) gene phylogenies. Except the acmA gene analysis, molecular tools revealed that isolates M78 and M104 clustered with strains of the cremoris genotype, including the LMG 6897T strain, while they were distant from strains of the lactis genotype, including the LMG 6890T strain. The two wild isolates had identical repetitive sequence-based PCR (rep-PCR), randomly amplified polymorphic DNA (RAPD), plasmid, and whole-cell protein profiles and shared high 16S rRNA (99.9%) and ldh (100%) gene sequence homologies. In contrast, they exhibited identical sugar fermentation and enzymatic patterns which were similar to those of the subspecies lactis LMG 6890T strain. To our knowledge, this is the first complete identification report on a wild L. lactis subsp. cremoris genotype of the lactis phenotype which is capable of nisin A production and, thus, has strong potential for use as a novel dairy starter and/or protective culture. PMID:23542625

Bacterial community succession during pig manure and wheat straw aerobic composting covered with a semi-permeable membrane under slight positive pressure.

PubMed

Ma, Shuangshuang; Fang, Chen; Sun, Xiaoxi; Han, Lujia; He, Xueqin; Huang, Guangqun

2018-07-01

Bacteria play an important role in organic matter degradation and maturity during aerobic composting. This study analyzed composting with or without a membrane cover in laboratory-scale aerobic composting reactor systems. 16S rRNA gene analysis was used to study the bacterial community succession during composting. The richness of the bacterial community decreased and the diversity increased after covering with a semi-permeable membrane and applying a slight positive pressure. Principal components analysis based on operational taxonomic units could distinguish the main composting phases. Linear Discriminant Analysis Effect Size analysis indicated that covering with a semi-permeable membrane reduced the relative abundance of anaerobic Clostridiales and pathogenic Pseudomonas and increased the abundance of Cellvibrionales. In membrane-covered aerobic composting systems, the relative abundance of some bacteria could be affected, especially anaerobic bacteria. Covering could effectively promote fermentation, reduce emissions and ensure organic fertilizer quality. Copyright © 2018 Elsevier Ltd. All rights reserved.

Molecular phylogeny of the Achatinoidea (Mollusca: Gastropoda).

PubMed

Fontanilla, Ian Kendrich; Naggs, Fred; Wade, Christopher Mark

2017-09-01

This study presents a multi-gene phylogenetic analysis of the Achatinoidea and provides an initial basis for a taxonomic re-evaluation of family level groups within the superfamily. A total of 5028 nucleotides from the nuclear rRNA, actin and histone 3 genes and the 1st and 2nd codon positions of the mitochondrial cytochrome c oxidase subunit I gene were sequenced from 24 species, representing six currently recognised families. Results from maximum likelihood, neighbour joining, maximum parsimony and Bayesian inference trees revealed that, of currently recognised families, only the Achatinidae are monophyletic. For the Ferussaciidae, Ferussacia folliculus fell separately to Cecilioides gokweanus and formed a sister taxon to the rest of the Achatinoidea. For the Coeliaxidae, Coeliaxis blandii and Pyrgina umbilicata did not group together. The Subulinidae was not resolved, with some subulinids clustering with the Coeliaxidae and Thyrophorellidae. Three subfamilies currently included within the Subulinidae based on current taxonomy likewise did not form monophyletic groups. Copyright © 2017 Elsevier Inc. All rights reserved.

The complete mitochondrial genome of the desert darkling beetle Asbolus verrucosus (Coleoptera, Tenebrionidae).

PubMed

Rider, Stanley Dean

2016-07-01

The complete mitochondrial genome of the desert darkling beetle Asbolus verrucosus (LeConte, 1851) was sequenced using paired-end technology to an average depth of 42,111× and assembled using De Bruijn graph-based methods. The genome is 15,828 bp in length and conforms to the basal arthropod mitochondrial gene composition with the same gene orders and orientations as other darkling beetle mitochondria. This arrangement includes a control region, 22 tRNA genes, 2 rRNA genes and 13 protein-coding genes. The main coding strand is probably replicated as the lagging strand (GC skew of -0.36 and AT skew of +0.19). Phylogenomics analyses are consistent with taxonomic classifications and indicate that Tenebrio molitor is the closest relative that has a completely sequenced mitochondrial genome available for analysis. This is the first fully assembled mitogenome sequence for a darkling beetle in the subfamily Pimeliinae and will be useful for population studies on members of this ecologically important group of beetles.

Complete mitochondrial genome of the brown alga Sargassum fusiforme (Sargassaceae, Phaeophyceae): genome architecture and taxonomic consideration.

PubMed

Liu, Feng; Pang, Shaojun; Luo, Minbo

2016-01-01

Sargassum fusiforme (Harvey) Setchell (=Hizikia fusiformis (Harvey) Okamura) is one of the most important economic seaweeds for mariculture in China. In this study, we present the complete mitochondrial genome of S. fusiforme. The genome is 34,696 bp in length with circular organization, encoding the standard set of three ribosomal RNA genes (rRNA), 25 transfer RNA genes (tRNA), 35 protein-coding genes, and two conserved open reading frames (ORFs). Its total AT content is 62.47%, lower than other brown algae except Pylaiella littoralis. The mitogenome carries 1571 bp of intergenic region constituting 4.53% of the genome, and 13 pairs of overlapping genes with the overlap size from 1 to 90 bp. The phylogenetic analyses based on 35 protein-coding genes reveal that S. fusiforme has a closer evolutionary relationship with Sargassum muticum than Sargassum horneri, indicating Hizikia are not distinct evolutionary entity and should be reduced to synonymy with Sargassum.

Comparison of the Rhizosphere Bacterial Communities of Zigongdongdou Soybean and a High-Methionine Transgenic Line of This Cultivar

PubMed Central

Ji, Jun; Wu, Haiying; Meng, Fang; Zhang, Mingrong; Zheng, Xiaobo; Wu, Cunxiang; Zhang, Zhengguang

2014-01-01

Previous studies have shown that methionine from root exudates affects the rhizosphere bacterial population involved in soil nitrogen fixation. A transgenic line of Zigongdongdou soybean cultivar (ZD91) that expresses Arabidopsis cystathionine γ-synthase resulting in an increased methionine production was examined for its influence to the rhizosphere bacterial population. Using 16S rRNA gene-based pyrosequencing analysis of the V4 region and DNA extracted from bacterial consortia collected from the rhizosphere of soybean plants grown in an agricultural field at the pod-setting stage, we characterized the populational structure of the bacterial community involved. In total, 87,267 sequences (approximately 10,908 per sample) were analyzed. We found that Acidobacteria, Proteobacteria, Bacteroidetes, Actinobacteria, Chloroflexi, Planctomycetes, Gemmatimonadetes, Firmicutes, and Verrucomicrobia constitute the dominant taxonomic groups in either the ZD91 transgenic line or parental cultivar ZD, and that there was no statistically significant difference in the rhizosphere bacterial community structure between the two cultivars. PMID:25079947

Seasonally variable intestinal metagenomes of the red palm weevil (Rhynchophorus ferrugineus)

PubMed Central

Jia, Shangang; Zhang, Xiaowei; Zhang, Guangyu; Yin, An; Zhang, Sun; Li, Fusen; Wang, Lei; Zhao, Duojun; Yun, Quanzheng; Tala; Wang, Jixiang; Sun, Gaoyuan; Baabdullah, Mohammed; Yu, Xiaoguang; Hu, Songnian; Al-Mssallem, Ibrahim S; Yu, Jun

2013-01-01

The intestinal microbes residing in the red palm weevil (RPW, Rhynchophorus ferrugineus) larva consume tender interior fibrous tissues of date palm trunks. The understanding of such microbiota at molecular level provides vital clues for the biological control of this devastating pest. Using pyrosequencing and shotgun strategy, we first study taxonomic profiles of the microbiota sampled at different months (March, July and November), and then confirm the impact of high-temperature stress on the microbial populations based on data from 16S rRNA amplicons using both field and laboratory samples. We further identify Klebsiella pneumoniae in November and Lactococcus lactis in July as the dominant species of the microbiota. We find that the RPW gut microbiota degrades polysaccharides and sucrose with hydrolases and that different active bacterial species in November and July are responsible for the symbiotic relationship between the microbiota and the host. Our results provide vital information for pest control and cellulolytic bacterial species characterization. PMID:24102776

Phylogenetic relationships of Paradiclybothrium pacificum and Diclybothrium armatum (Monogenoidea: Diclybothriidae) inferred from 18S rDNA sequence data.

PubMed

Rozhkovan, Konstantin V; Shedko, Marina B

2015-10-01

The Diclybothriidae (Monogenoidea: Oligonchoinea) includes specific parasites of fishes assigned to the ancient order Acipenseriformes. Phylogeny of the Diclybothriidae is still unclear despite several systematic studies based on morphological characters. Together with the closely related Hexabothriidae represented by parasites of sharks and ray-fishes, the position of Diclybothriidae in different taxonomical systems has been matter of discussion. Here, we present the first molecular data on Diclybothriidae. The SSU rRNA gene was used to investigate the phylogenetic position of Paradiclybothrium pacificum and Diclybothrium armatum among the other Oligonchoinea. Complete nucleotide sequences of P. pacificum and D. armatum demonstrated high identity (98.53%) with no intraspecific sequence variability. Specimens of D. armatum were obtained from different hosts (Acipenser schrenckii and Huso dauricus); however, variation by host was not detected. The sequence divergence and phylogenetic trees data show that Diclybothriidae and Hexabothriidae are more closely related to each other than with other representatives of Oligonchoinea. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Bacterial communities in commercial aircraft high-efficiency particulate air (HEPA) filters assessed by PhyloChip analysis.

PubMed

Korves, T M; Piceno, Y M; Tom, L M; Desantis, T Z; Jones, B W; Andersen, G L; Hwang, G M

2013-02-01

Air travel can rapidly transport infectious diseases globally. To facilitate the design of biosensors for infectious organisms in commercial aircraft, we characterized bacterial diversity in aircraft air. Samples from 61 aircraft high-efficiency particulate air (HEPA) filters were analyzed with a custom microarray of 16S rRNA gene sequences (PhyloChip), representing bacterial lineages. A total of 606 subfamilies from 41 phyla were detected. The most abundant bacterial subfamilies included bacteria associated with humans, especially skin, gastrointestinal and respiratory tracts, and with water and soil habitats. Operational taxonomic units that contain important human pathogens as well as their close, more benign relatives were detected. When compared to 43 samples of urban outdoor air, aircraft samples differed in composition, with higher relative abundance of Firmicutes and Gammaproteobacteria lineages in aircraft samples, and higher relative abundance of Actinobacteria and Betaproteobacteria lineages in outdoor air samples. In addition, aircraft and outdoor air samples differed in the incidence of taxa containing human pathogens. Overall, these results demonstrate that HEPA filter samples can be used to deeply characterize bacterial diversity in aircraft air and suggest that the presence of close relatives of certain pathogens must be taken into account in probe design for aircraft biosensors. A biosensor that could be deployed in commercial aircraft would be required to function at an extremely low false alarm rate, making an understanding of microbial background important. This study reveals a diverse bacterial background present on aircraft, including bacteria closely related to pathogens of public health concern. Furthermore, this aircraft background is different from outdoor air, suggesting different probes may be needed to detect airborne contaminants to achieve minimal false alarm rates. This study also indicates that aircraft HEPA filters could be used with other molecular techniques to further characterize background bacteria and in investigations in the wake of a disease outbreak. © 2012 John Wiley & Sons A/S.

Characterization of three plant biomass-degrading microbial consortia by metagenomics- and metasecretomics-based approaches.

PubMed

Jiménez, Diego Javier; de Lima Brossi, Maria Julia; Schückel, Julia; Kračun, Stjepan Krešimir; Willats, William George Tycho; van Elsas, Jan Dirk

2016-12-01

The selection of microbes by enrichment on plant biomass has been proposed as an efficient way to develop new strategies for lignocellulose saccharification. Here, we report an in-depth analysis of soil-derived microbial consortia that were trained to degrade once-used wheat straw (WS1-M), switchgrass (SG-M) and corn stover (CS-M) under aerobic and mesophilic conditions. Molecular fingerprintings, bacterial 16S ribosomal RNA (rRNA) gene amplicon sequencing and metagenomic analyses showed that the three microbial consortia were taxonomically distinct. Based on the taxonomic affiliation of protein-encoding sequences, members of the Bacteroidetes (e.g. Chryseobacterium, Weeksella, Flavobacterium and Sphingobacterium) were preferentially selected on WS1-M, whereas SG-M and CS-M favoured members of the Proteobacteria (e.g. Caulobacter, Brevundimonas, Stenotrophomonas and Xanthomonas). The highest degradation rates of lignin (~59 %) were observed with SG-M, whereas CS-M showed a high consumption of cellulose and hemicellulose. Analyses of the carbohydrate-active enzymes in the three microbial consortia showed the dominance of glycosyl hydrolases (e.g. of families GH3, GH43, GH13, GH10, GH29, GH28, GH16, GH4 and GH92). In addition, proteins of families AA6, AA10 and AA2 were detected. Analysis of secreted protein fractions (metasecretome) for each selected microbial consortium mainly showed the presence of enzymes able to degrade arabinan, arabinoxylan, xylan, β-glucan, galactomannan and rhamnogalacturonan. Notably, these metasecretomes contain enzymes that enable us to produce oligosaccharides directly from wheat straw, sugarcane bagasse and willow. Thus, the underlying microbial consortia constitute valuable resources for the production of enzyme cocktails for the efficient saccharification of plant biomass.

Microbiome Analysis of Stool Samples from African Americans with Colon Polyps

PubMed Central

Brim, Hassan; Yooseph, Shibu; Zoetendal, Erwin G.; Lee, Edward; Torralbo, Manolito; Laiyemo, Adeyinka O.; Shokrani, Babak; Nelson, Karen; Ashktorab, Hassan

2013-01-01

Background Colonic polyps are common tumors occurring in ~50% of Western populations with ~10% risk of malignant progression. Dietary agents have been considered the primary environmental exposure to promote colorectal cancer (CRC) development. However, the colonic mucosa is permanently in contact with the microbiota and its metabolic products including toxins that also have the potential to trigger oncogenic transformation. Aim To analyze fecal DNA for microbiota composition and functional potential in African Americans with pre-neoplastic lesions. Materials & Methods We analyzed the bacterial composition of stool samples from 6 healthy individuals and 6 patients with colon polyps using 16S ribosomal RNA-based phylogenetic microarray; the Human intestinal Tract Chip (HITChip) and 16S rRNA gene barcoded 454 pyrosequencing. The functional potential was determined by sequence-based metagenomics using 454 pyrosequencing. Results Fecal microbiota profiling of samples from the healthy and polyp patients using both a phylogenetic microarraying (HITChip) and barcoded 454 pyrosequencing generated similar results. A distinction between both sets of samples was only obtained when the analysis was performed at the sub-genus level. Most of the species leading to the dissociation were from the Bacteroides group. The metagenomic analysis did not reveal major differences in bacterial gene prevalence/abundances between the two groups even when the analysis and comparisons were restricted to available Bacteroides genomes. Conclusion This study reveals that at the pre-neoplastic stages, there is a trend showing microbiota changes between healthy and colon polyp patients at the sub-genus level. These differences were not reflected at the genome/functions levels. Bacteria and associated functions within the Bacteroides group need to be further analyzed and dissected to pinpoint potential actors in the early colon oncogenic transformation in a large sample size. PMID:24376500

[Comparison of gut microbiotal compositional analysis of patients with irritable bowel syndrome through different bioinformatics pipelines].

PubMed

Zhu, S W; Liu, Z J; Li, M; Zhu, H Q; Duan, L P

2018-04-18

To assess whether the same biological conclusion, diagnostic or curative effects regarding microbial composition of irritable bowel syndrome (IBS) patients could be reached through different bioinformatics pipelines, we used two common bioinformatics pipelines (Uparse V2.0 and Mothur V1.39.5)to analyze the same fecal microbial 16S rRNA high-throughput sequencing data. The two pipelines were used to analyze the diversity and richness of fecal microbial 16S rRNA high-throughput sequencing data of 27 samples, including 9 healthy controls (HC group), 9 diarrhea IBS patients before (IBS group) and after Rifaximin treatment (IBS-treatment, IBSt group). Analyses such as microbial diversity, principal co-ordinates analysis (PCoA), nonmetric multidimensional scaling (NMDS) and linear discriminant analysis effect size (LEfSe) were used to find out the microbial differences among HC group vs. IBS group and IBS group vs. IBSt group. (1) Microbial composition comparison of the 27 samples in the two pipelines showed significant variations at both family and genera levels while no significant variations at phylum level; (2) There was no significant difference in the comparison of HC vs. IBS or IBS vs. IBSt (Uparse: HC vs. IBS, F=0.98, P=0.445; IBS vs. IBSt, F=0.47,P=0.926; Mothur: HC vs.IBS, F=0.82, P=0.646; IBS vs. IBSt, F=0.37, P=0.961). The Shannon index was significantly decreased in IBSt; (3) Both workshops distinguished the significantly enriched genera between HC and IBS groups. For example, Nitrosomonas and Paraprevotella increased while Pseudoalteromonadaceae and Anaerotruncus decreased in HC group through Uparse pipeline, nevertheless Roseburia 62 increased while Butyricicoccus and Moraxellaceae decreased in HC group through Mothur pipeline.Only Uparse pipeline could pick out significant genera between IBS and IBSt, such as Pseudobutyricibrio, Clostridiaceae 1 and Clostridiumsensustricto 1. There were taxonomic and phylogenetic diversity differences between the two pipelines, Mothur can get more taxonomic details because the count number of each taxonomic level is higher. Both pipelines could distinguish the significantly enriched genera between HC and IBS groups, but Uparse was more capable to identity the difference between IBS and IBSt groups. To increase the reproducibility and reliability and to retain the consistency among similar studies, it is very important to consider the impact on different pipelines.

Microarray-integrated optoelectrofluidic immunoassay system

PubMed Central

Han, Dongsik

2016-01-01

A microarray-based analytical platform has been utilized as a powerful tool in biological assay fields. However, an analyte depletion problem due to the slow mass transport based on molecular diffusion causes low reaction efficiency, resulting in a limitation for practical applications. This paper presents a novel method to improve the efficiency of microarray-based immunoassay via an optically induced electrokinetic phenomenon by integrating an optoelectrofluidic device with a conventional glass slide-based microarray format. A sample droplet was loaded between the microarray slide and the optoelectrofluidic device on which a photoconductive layer was deposited. Under the application of an AC voltage, optically induced AC electroosmotic flows caused by a microarray-patterned light actively enhanced the mass transport of target molecules at the multiple assay spots of the microarray simultaneously, which reduced tedious reaction time from more than 30 min to 10 min. Based on this enhancing effect, a heterogeneous immunoassay with a tiny volume of sample (5 μl) was successfully performed in the microarray-integrated optoelectrofluidic system using immunoglobulin G (IgG) and anti-IgG, resulting in improved efficiency compared to the static environment. Furthermore, the application of multiplex assays was also demonstrated by multiple protein detection. PMID:27190571

Microarray-integrated optoelectrofluidic immunoassay system.

PubMed

Han, Dongsik; Park, Je-Kyun

2016-05-01

A microarray-based analytical platform has been utilized as a powerful tool in biological assay fields. However, an analyte depletion problem due to the slow mass transport based on molecular diffusion causes low reaction efficiency, resulting in a limitation for practical applications. This paper presents a novel method to improve the efficiency of microarray-based immunoassay via an optically induced electrokinetic phenomenon by integrating an optoelectrofluidic device with a conventional glass slide-based microarray format. A sample droplet was loaded between the microarray slide and the optoelectrofluidic device on which a photoconductive layer was deposited. Under the application of an AC voltage, optically induced AC electroosmotic flows caused by a microarray-patterned light actively enhanced the mass transport of target molecules at the multiple assay spots of the microarray simultaneously, which reduced tedious reaction time from more than 30 min to 10 min. Based on this enhancing effect, a heterogeneous immunoassay with a tiny volume of sample (5 μl) was successfully performed in the microarray-integrated optoelectrofluidic system using immunoglobulin G (IgG) and anti-IgG, resulting in improved efficiency compared to the static environment. Furthermore, the application of multiplex assays was also demonstrated by multiple protein detection.

The MicroArray Quality Control (MAQC)-II study of common practices for the development and validation of microarray-based predictive models

EPA Science Inventory

The second phase of the MicroArray Quality Control (MAQC-II) project evaluated common practices for developing and validating microarray-based models aimed at predicting toxicological and clinical endpoints. Thirty-six teams developed classifiers for 13 endpoints - some easy, som...

Qingshengfania soli gen. nov., sp. nov., a member of the order Rhizobiales isolated from the soil of a pesticide factory.

PubMed

Zhang, Long; Zhou, Qing-Xin; Song, Man; Chen, Xiao-Long; Xu, Xi-Hui; Chen, Kai; Li, Shun-Peng; Jiang, Jian-Dong

2015-12-01

Two Gram-stain negative, coccoid to oval-shaped, non-spore-forming bacteria (LR4T and LR4-1), isolated from the soil of a pesticide factory in Nanjing, China, were investigated for their taxonomic allocation by using a polyphasic approach. Both strains grew optimally at pH 7.0, 30 °C and in the absence of NaCl. Both strains were positive for catalase and oxidase activities. Q-10 was the predominant respiratory ubiquinone. The major polar lipids were phosphatidylmonomethylethanolamine, diphosphatidylglycerol, phosphatidylglycerol, phosphatidylcholine and two unknown aminolipids. The major fatty acids (>10 % of the total fatty acids) were C18:1ω7c/C18:1ω6c (summed feature 8) and C17:1 iso I/C17:1 anteiso B (summed feature 4). Phylogenetic analysis based on 16S rRNA gene sequence comparisons showed that the two isolates formed a distinct line within a clade containing the genera Chelatococcus, Bosea, Camelimonas, Salinarimonas, Psychroglaciecola, Microvirga, Methylobacterium, Albibacter, Hansschlegelia and Methylopila in the order Rhizobiales, with the highest 16S rRNA gene sequence similarity to Chelatococcus asaccharovorans TE2T (94.12 %), followed by Bosea thiooxidans DSM 9653T (93.25 %). Strains LR4T and LR4-1 were closely related on the basis of DNA-DNA reassociation and therefore represent a single novel species. Based on phenotypic, chemotaxonomic and phylogenetic data, strains LR4T and LR4-1 represent a novel species of a new genus in the order Rhizobiales, for which the name Qingshengfania soli gen. nov., sp. nov. is proposed. The type strain of the type species is LR4T ( = CCTCC AB 2015036T = KCTC 42463T).

Streptomonospora tuzyakensis sp. nov., a halophilic actinomycete isolated from saline soil.

PubMed

Tatar, Demet; Guven, Kiymet; Inan, Kadriye; Cetin, Demet; Belduz, Ali Osman; Sahin, Nevzat

2016-01-01

A novel actinobacterium, designated strain BN506(T), was isolated from soil collected from Tuz (Salt) Lake, Konya, Turkey, and was characterised to determine its taxonomic position. The isolate was found to have morphological and chemotaxonomic properties associated with members of the genus Streptomonospora. The isolate was found to grow optimally at 37 °C and in the presence of 10 % (w/v) NaCl but not in the absence of NaCl. Phylogenetic analyses based on an almost-complete 16S rRNA gene sequences indicated that isolate is closely related to members of the genus Streptomonospora and forms a distinct phyletic line in the Streptomonospora phylogenetic tree. Strain BN506(T) is closely related to Streptomonospora halophila YIM 91355(T) (98.1 % sequence similarity). Sequence similarities with other type strains of the genus Streptomyces were lower than 98.0 %. The cell wall of the novel strain was found to contain meso-diaminopimelic acid. Whole cell hydrolysates were found to contain galactose, glucose and ribose. The predominant menaquinone was identified as MK-10(H8) (57.0 %). The polar lipids detected were identified as diphosphatidylglycerol, phosphatidylglycerol and phosphatidylcholine. The major fatty acids were found to be anteiso-C17:0, iso-C16:0 and 10 methyl C18:0. Based on 16S rRNA gene sequence analysis, DNA-DNA relatedness, phenotypic characteristics and chemotaxonomic data, strain BN506(T) was identified as a member of a novel species of the genus Streptomonospora, for which the name Streptomonospora tuzyakensis sp. nov. (type strain BN506(T) = DSM 45930(T) = KCTC 29210(T)) is proposed.

Diversity of Bradyrhizobium strains nodulating Lupinus micranthus on both sides of the Western Mediterranean: Algeria and Spain.

PubMed

Bourebaba, Yasmina; Durán, David; Boulila, Farida; Ahnia, Hadjira; Boulila, Abdelghani; Temprano, Francisco; Palacios, José M; Imperial, Juan; Ruiz-Argüeso, Tomás; Rey, Luis

2016-06-01

Lupinus micranthus is a lupine distributed in the Mediterranean basin whose nitrogen fixing symbiosis has not been described in detail. In this study, 101 slow-growing nodule isolates were obtained from L. micranthus thriving in soils on both sides of the Western Mediterranean. The diversity of the isolates, 60 from Algeria and 41 from Spain, was addressed by multilocus sequence analysis of housekeeping genes (16S rRNA, atpD, glnII and recA) and one symbiotic gene (nodC). Using genomic fingerprints from BOX elements, 37 different profiles were obtained (22 from Algeria and 15 from Spain). Phylogenetic analysis based on 16S rRNA and concatenated atpD, glnII and recA sequences of a representative isolate of each BOX profile displayed a homogeneous distribution of profiles in six different phylogenetic clusters. All isolates were taxonomically ascribed to the genus Bradyrhizobium. Three clusters comprising 24, 6, and 4 isolates, respectively, accounted for most of the profiles. The largest cluster was close to the Bradyrhizobium canariense lineage, while the other two were related to B. cytisi/B. rifense. The three remaining clusters included only one isolate each, and were close to B. canariense, B. japonicum and B. elkanii species, respectively. In contrast, phylogenetic clustering of BOX profiles based on nodC sequences yielded only two phylogenetic groups. One of them included all the profiles except one, and belonged to symbiovar genistearum. The remaining profile, constituted by a strain related to B. elkanii, was not related to any well-defined symbiotic lineage, and may constitute both a new symbiovar and a new genospecies. Copyright © 2016 Elsevier GmbH. All rights reserved.

Parvularcula flava sp. nov., an alphaproteobacterium isolated from surface seawater of the South China Sea.

PubMed

Zhang, Xin-Qi; Wu, Yue-Hong; Zhou, Xiang; Zhang, Xin; Xu, Xue-Wei; Wu, Min

2016-09-01

An aerobic, coccoid to short rod, yellow-pigmented, non-sporulating and Gram-staining-negative bacterium, designated NH6-79T, was isolated from surface seawater of the South China Sea. The isolate was motile with a polar flagellum. Growth was observed at 4-42 °C (optimum 37 °C), at pH 6.0-8.5 (optimum pH 7.0), and with 0.5-11 % (w/v) NaCl (optimum 4.5 %) and 1.5-17 % (w/v) sea salt (optimum 3.5-5 %). Strain NH6-79T could decompose peptone to produce H2S, but could not hydrolyse skimmed milk. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain NH6-79T had the closest affinity to the genus Parvularcula, sharing the highest 16S rRNA gene sequence similarity with 'Parvularcula oceanus' JLT2013 (94.1 %), Parvularcula lutaonensis CC-MMS-1T (93.4 %), Parvularcula dongshanensis SH25T (92.9 %) and Parvularcula bermudensis HTCC2503T (92.7 %), and lower sequence similarities (<90 %) with all other genera. The dominant fatty acids were C18 : 1ω7c and C16 : 0. The polar lipid profile was mainly composed of three unidentified glycolipids. The predominant isoprenoid quinone was ubiquinone-10. The DNA G+C content was 60.7 mol%. Based on the polyphasic taxonomic characterization, strain NH6-79T is considered to represent a novel species of the genus Parvularcula, for which the name Parvularcula flava sp. nov. is proposed. The type strain is NH6-79T (=CGMCC 1.14984T=JCM 30557T=MCCC 1K00277T).

Horizon-Specific Bacterial Community Composition of German Grassland Soils, as Revealed by Pyrosequencing-Based Analysis of 16S rRNA Genes ▿ †

PubMed Central

Will, Christiane; Thürmer, Andrea; Wollherr, Antje; Nacke, Heiko; Herold, Nadine; Schrumpf, Marion; Gutknecht, Jessica; Wubet, Tesfaye; Buscot, François; Daniel, Rolf

2010-01-01

The diversity of bacteria in soil is enormous, and soil bacterial communities can vary greatly in structure. Here, we employed a pyrosequencing-based analysis of the V2-V3 16S rRNA gene region to characterize the overall and horizon-specific (A and B horizons) bacterial community compositions in nine grassland soils, which covered three different land use types. The entire data set comprised 752,838 sequences, 600,544 of which could be classified below the domain level. The average number of sequences per horizon was 41,824. The dominant taxonomic groups present in all samples and horizons were the Acidobacteria, Betaproteobacteria, Actinobacteria, Gammaproteobacteria, Alphaproteobacteria, Deltaproteobacteria, Chloroflexi, Firmicutes, and Bacteroidetes. Despite these overarching dominant taxa, the abundance, diversity, and composition of bacterial communities were horizon specific. In almost all cases, the estimated bacterial diversity (H′) was higher in the A horizons than in the corresponding B horizons. In addition, the H′ was positively correlated with the organic carbon content, the total nitrogen content, and the C-to-N ratio, which decreased with soil depth. It appeared that lower land use intensity results in higher bacterial diversity. The majority of sequences affiliated with the Actinobacteria, Bacteroidetes, Cyanobacteria, Fibrobacteres, Firmicutes, Spirochaetes, Verrucomicrobia, Alphaproteobacteria, Betaproteobacteria, and Gammaproteobacteria were derived from A horizons, whereas the majority of the sequences related to Acidobacteria, Chloroflexi, Gemmatimonadetes, Nitrospira, TM7, and WS3 originated from B horizons. The distribution of some bacterial phylogenetic groups and subgroups in the different horizons correlated with soil properties such as organic carbon content, total nitrogen content, or microbial biomass. PMID:20729324

A novel marine nitrite-oxidizing Nitrospira species from Dutch coastal North Sea water

PubMed Central

Haaijer, Suzanne C. M.; Ji, Ke; van Niftrik, Laura; Hoischen, Alexander; Speth, Daan; Jetten, Mike S. M.; Damsté, Jaap S. Sinninghe; Op den Camp, Huub J. M.

2013-01-01

Marine microorganisms are important for the global nitrogen cycle, but marine nitrifiers, especially aerobic nitrite oxidizers, remain largely unexplored. To increase the number of cultured representatives of marine nitrite-oxidizing bacteria (NOB), a bioreactor cultivation approach was adopted to first enrich nitrifiers and ultimately nitrite oxidizers from Dutch coastal North Sea water. With solely ammonia as the substrate an active nitrifying community consisting of novel marine Nitrosomonas aerobic ammonia oxidizers (ammonia-oxidizing bacteria) and Nitrospina and Nitrospira NOB was obtained which converted a maximum of 2 mmol of ammonia per liter per day. Switching the feed of the culture to nitrite as a sole substrate resulted in a Nitrospira NOB dominated community (approximately 80% of the total microbial community based on fluorescence in situ hybridization and metagenomic data) converting a maximum of 3 mmol of nitrite per liter per day. Phylogenetic analyses based on the 16S rRNA gene indicated that the Nitrospira enriched from the North Sea is a novel Nitrospira species with Nitrospira marina as the next taxonomically described relative (94% 16S rRNA sequence identity). Transmission electron microscopy analysis revealed a cell plan typical for Nitrospira species. The cytoplasm contained electron light particles that might represent glycogen storage. A large periplasmic space was present which was filled with electron dense particles. Nitrospira-targeted polymerase chain reaction analyses demonstrated the presence of the enriched Nitrospira species in a time series of North Sea genomic DNA samples. The availability of this new Nitrospira species enrichment culture facilitates further in-depth studies such as determination of physiological constraints, and comparison to other NOB species. PMID:23515432

Verrucosispora sonchi sp. nov., a novel endophytic actinobacterium isolated from the leaves of common sowthistle (Sonchus oleraceus L.).

PubMed

Ma, Zhaoxu; Zhao, Shanshan; Cao, Tingting; Liu, Chongxi; Huang, Ying; Gao, Yuhang; Yan, Kai; Xiang, Wensheng; Wang, Xiangjing

2016-12-01

A novel actinobacterium, designated strain NEAU-QY3T, was isolated from the leaves of Sonchus oleraceus L. and examined using a polyphasic taxonomic approach. The organism formed single spores with smooth surface on substrate mycelia. Phylogenetic analysis based on the 16S rRNA gene sequence indicated that the strain had a close association with the genus Verrucosispora and shared the highest sequence similarity with Verrucosispora qiuiae RtIII47T (99.17 %), an association that was supported by a bootstrap value of 94 % in the neighbour-joining tree and also recovered with the maximum-likelihood algorithm. The strain also showed high 16S rRNA gene sequence similarities to Xiangella phaseoli NEAU-J5T (98.78 %), Jishengella endophytica 202201T (98.51 %), Micromonospora eburnea LK2-10T (98.28 %), Verrucosispora lutea YIM 013T (98.23 %) and Salinispora pacifica CNR-114T (98.23 %). Furthermore, phylogenetic analysis based on the gyrB gene sequences supported the conclusion that strain NEAU-QY3T should be assigned to the genus Verrucosispora. However, the DNA-DNA hybridization relatedness values between strain NEAU-QY3T and V. qiuiae RtIII47T and V. lutea YIM 013T were below 70 %. With reference to phenotypic characteristics, phylogenetic data and DNA-DNA hybridization results, strain NEAU-QY3T was readily distinguished from its most closely related strains and classified as a new species, for which the name Verrucosispora sonchi sp. nov. is proposed. The type strain is NEAU-QY3T (=CGMCC 4.7312T=DSM 101530T).

Gordonia caeni sp. nov., isolated from sludge of a sewage disposal plant.

PubMed

Srinivasan, Sathiyaraj; Park, Giho; Yang, Hyejin; Hwang, Supyong; Bae, Yoonjung; Jung, Yong-An; Kim, Myung Kyum; Lee, Myungjin

2012-11-01

A Gram-stain-positive, strictly aerobic, short-rod-shaped, non-motile strain (designated MJ32(T)) was isolated from a sludge sample of the Daejeon sewage disposal plant in South Korea. A polyphasic approach was applied to study the taxonomic position of strain MJ32(T). Strain MJ32(T) showed highest 16S rRNA gene sequence similarity to Gordonia hirsuta DSM 44140(T) (98.1%) and Gordonia hydrophobica DSM 44015(T) (97.0%); levels of sequence similarity to the type strains of other recognized Gordonia species were less than 97.0%. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain MJ32(T) belonged to the clade formed by members of the genus Gordonia in the family Gordoniaceae. The G+C content of the genomic DNA of strain MJ32(T) was 69.2 mol%. Chemotaxonomically, strain MJ32(T) showed features typical of the genus Gordonia. The predominant respiratory quinone was MK-9(H(2)), the mycolic acids present had C(56)-C(60) carbon atoms, and the major fatty acids were C(16:0) (34.6%), tuberculostearic acid (21.8%), C(16:1)ω7c (19.5%) and C(18:1)ω9c (12.7%). The peptidoglycan type was based on meso-2,6-diaminopimelic acid as the diagnostic diamino acid with glycolated sugars. On the basis of phylogenetic inference, fatty acid profile and other phenotypic properties, strain MJ32(T) is considered to represent a novel species of the genus Gordonia, for which the name Gordonia caeni sp. nov. is proposed. The type strain is MJ32(T) (=KCTC 19771(T)=JCM 16923(T)).

Porcine intestinal microbiota is shaped by diet composition based on rye or triticale.

PubMed

Burbach, K; Strang, E J P; Mosenthin, R; Camarinha-Silva, A; Seifert, J

2017-12-01

The present study aimed to compare the microbiota composition from pigs fed different cereal grain types, either rye or triticale, as sole energy source. Ileal digesta and faeces were sampled from eight pigs of each experiment. Illumina amplicon sequencing of the 16S rRNA gene was used to analyse the microbiota. Concentrations of short-chain fatty acids and ammonia were determined from faecal samples. The grain type revealed significant alterations in the overall microbiota structure. The rye-based diet was associated with an increased abundance of Lactobacillus in ileal digesta and Streptococcus in faeces and significantly higher concentrations of faecal short-chain fatty acids and ammonia compared to triticale. However, triticale significantly promoted the abundance of Streptococcus in ileal digesta and Clostridium sensu stricto in faeces. Diets based on rye or triticale affect varying intestinal microbiota, both of taxonomical and metabolic structure, with rye indicating an enhanced saccharolytic potential and triticale a more cellulolytic potential. Nutrient composition of rye and triticale are attractive for porcine nutrition. Both cereal grains show varying stimuli on the microbiota composition and microbial products of the ileum and faeces. © 2017 The Society for Applied Microbiology.

A microarray for assessing transcription from pelagic marine microbial taxa

PubMed Central

Shilova, Irina N; Robidart, Julie C; James Tripp, H; Turk-Kubo, Kendra; Wawrik, Boris; Post, Anton F; Thompson, Anne W; Ward, Bess; Hollibaugh, James T; Millard, Andy; Ostrowski, Martin; J Scanlan, David; Paerl, Ryan W; Stuart, Rhona; Zehr, Jonathan P

2014-01-01

Metagenomic approaches have revealed unprecedented genetic diversity within microbial communities across vast expanses of the world's oceans. Linking this genetic diversity with key metabolic and cellular activities of microbial assemblages is a fundamental challenge. Here we report on a collaborative effort to design MicroTOOLs (Microbiological Targets for Ocean Observing Laboratories), a high-density oligonucleotide microarray that targets functional genes of diverse taxa in pelagic and coastal marine microbial communities. MicroTOOLs integrates nucleotide sequence information from disparate data types: genomes, PCR-amplicons, metagenomes, and metatranscriptomes. It targets 19 400 unique sequences over 145 different genes that are relevant to stress responses and microbial metabolism across the three domains of life and viruses. MicroTOOLs was used in a proof-of-concept experiment that compared the functional responses of microbial communities following Fe and P enrichments of surface water samples from the North Pacific Subtropical Gyre. We detected transcription of 68% of the gene targets across major taxonomic groups, and the pattern of transcription indicated relief from Fe limitation and transition to N limitation in some taxa. Prochlorococcus (eHLI), Synechococcus (sub-cluster 5.3) and Alphaproteobacteria SAR11 clade (HIMB59) showed the strongest responses to the Fe enrichment. In addition, members of uncharacterized lineages also responded. The MicroTOOLs microarray provides a robust tool for comprehensive characterization of major functional groups of microbes in the open ocean, and the design can be easily amended for specific environments and research questions. PMID:24477198

Microbial communities associated with ferromanganese nodules and the surrounding sediments

PubMed Central

Tully, Benjamin J.; Heidelberg, John F.

2013-01-01

The formation and maintenance of deep-sea ferromanganese/polymetallic nodules still remains a mystery 140 years after their discovery. The wealth of rare metals concentrated in these nodules has spurred global interest in exploring the mining potential of these resources. The prevailing theory of abiotic formation has been called into question and the role of microbial metabolisms in nodule development is now an area of active research. To understand the community structure of microbes associated with nodules and their surrounding sediment, we performed targeted sequencing of the V4 hypervariable region of the 16S rRNA gene from three nodules collected from the central South Pacific. Results have shown that the microbial communities of the nodules are significantly distinct from the communities in the surrounding sediments, and that the interiors of the nodules harbor communities different from the exterior. This suggests not only differences in potential metabolisms between the nodule and sediment communities, but also differences in the dominant metabolisms of interior and exterior communities. We identified several operational taxonomic units (OTUs) unique to both the nodule and sediment environments. The identified OTUs were assigned putative taxonomic identifications, including two OTUs only found associated with the nodules, which were assigned to the α-Proteobacteria. Finally, we explored the diversity of the most assigned taxonomic group, the Thaumarchaea MG-1, which revealed novel OTUs compared to previous research from the region and suggests a potential role as a source of fixed carbon for ammonia oxidizing archaea in the environment. PMID:23805131

Size-fractionated diversity of eukaryotic microbial communities in the Eastern Tropical North Pacific oxygen minimum zone.

PubMed

Duret, Manon T; Pachiadaki, Maria G; Stewart, Frank J; Sarode, Neha; Christaki, Urania; Monchy, Sébastien; Srivastava, Ankita; Edgcomb, Virginia P

2015-05-01

Oxygen minimum zones (OMZs) caused by water column stratification appear to expand in parts of the world's ocean, with consequences for marine biogeochemical cycles. OMZ formation is often fueled by high surface primary production, and sinking organic particles can be hotspots of interactions and activity within microbial communities. This study investigated the diversity of OMZ protist communities in two biomass size fractions (>30 and 30-1.6 μm filters) from the world's largest permanent OMZ in the Eastern Tropical North Pacific. Diversity was quantified via Illumina MiSeq sequencing of V4 region of 18S SSU rRNA genes in samples spanning oxygen gradients at two stations. Alveolata and Rhizaria dominated the two size fractions at both sites along the oxygen gradient. Community composition at finer taxonomic levels was partially shaped by oxygen concentration, as communities associated with versus anoxic waters shared only ∼32% of operational taxonomic unit (OTU) (97% sequence identity) composition. Overall, only 9.7% of total OTUs were recovered at both stations and under all oxygen conditions sampled, implying structuring of the eukaryotic community in this area. Size-fractionated communities exhibited different taxonomical features (e.g. Syndiniales Group I in the 1.6-30 μm fraction) that could be explained by the microniches created on the surface-originated sinking particles. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

RubisCO Gene Clusters Found in a Metagenome Microarray from Acid Mine Drainage

PubMed Central

Guo, Xue; Yin, Huaqun; Cong, Jing; Dai, Zhimin; Liang, Yili

2013-01-01

The enzyme responsible for carbon dioxide fixation in the Calvin cycle, ribulose-1,5-bisphosphate carboxylase/oxygenase (RubisCO), is always detected as a phylogenetic marker to analyze the distribution and activity of autotrophic bacteria. However, such an approach provides no indication as to the significance of genomic content and organization. Horizontal transfers of RubisCO genes occurring in eubacteria and plastids may seriously affect the credibility of this approach. Here, we presented a new method to analyze the diversity and genomic content of RubisCO genes in acid mine drainage (AMD). A metagenome microarray containing 7,776 large-insertion fosmids was constructed to quickly screen genome fragments containing RubisCO form I large-subunit genes (cbbL). Forty-six cbbL-containing fosmids were detected, and six fosmids were fully sequenced. To evaluate the reliability of the metagenome microarray and understand the microbial community in AMD, the diversities of cbbL and the 16S rRNA gene were analyzed. Fosmid sequences revealed that the form I RubisCO gene cluster could be subdivided into form IA and IB RubisCO gene clusters in AMD, because of significant divergences in molecular phylogenetics and conservative genomic organization. Interestingly, the form I RubisCO gene cluster coexisted with the form II RubisCO gene cluster in one fosmid genomic fragment. Phylogenetic analyses revealed that horizontal transfers of RubisCO genes may occur widely in AMD, which makes the evolutionary history of RubisCO difficult to reconcile with organismal phylogeny. PMID:23335778

Mineral Type and Solution Chemistry Affect the Structure and Composition of Actively Growing Bacterial Communities as Revealed by Bromodeoxyuridine Immunocapture and 16S rRNA Pyrosequencing.

PubMed

Kelly, L C; Colin, Y; Turpault, M-P; Uroz, S

2016-08-01

Understanding how minerals affect bacterial communities and their in situ activities in relation to environmental conditions are central issues in soil microbial ecology, as minerals represent essential reservoirs of inorganic nutrients for the biosphere. To determine the impact of mineral type and solution chemistry on soil bacterial communities, we compared the diversity, composition, and functional abilities of a soil bacterial community incubated in presence/absence of different mineral types (apatite, biotite, obsidian). Microcosms were prepared containing different liquid culture media devoid of particular essential nutrients, the nutrients provided only in the introduced minerals and therefore only available to the microbial community through mineral dissolution by biotic and/or abiotic processes. By combining functional screening of bacterial isolates and community analysis by bromodeoxyuridine DNA immunocapture and 16S rRNA gene pyrosequencing, we demonstrated that bacterial communities were mainly impacted by the solution chemistry at the taxonomic level and by the mineral type at the functional level. Metabolically active bacterial communities varied with solution chemistry and mineral type. Burkholderia were significantly enriched in the obsidian treatment compared to the biotite treatment and were the most effective isolates at solubilizing phosphorous or mobilizing iron, in all the treatments. A detailed analysis revealed that the 16S rRNA gene sequences of the OTUs or isolated strains assigned as Burkholderia in our study showed high homology with effective mineral-weathering bacteria previously recovered from the same experimental site.

Analysis of 16S libraries of mouse gastrointestinal microflora reveals a large new group of mouse intestinal bacteria.

PubMed

Salzman, Nita H; de Jong, Hendrik; Paterson, Yvonne; Harmsen, Hermie J M; Welling, Gjalt W; Bos, Nicolaas A

2002-11-01

Total genomic DNA from samples of intact mouse small intestine, large intestine, caecum and faeces was used as template for PCR amplification of 16S rRNA gene sequences with conserved bacterial primers. Phylogenetic analysis of the amplification products revealed 40 unique 16S rDNA sequences. Of these sequences, 25% (10/40) corresponded to described intestinal organisms of the mouse, including Lactobacillus spp., Helicobacter spp., segmented filamentous bacteria and members of the altered Schaedler flora (ASF360, ASF361, ASF502 and ASF519); 75% (30/40) represented novel sequences. A large number (11/40) of the novel sequences revealed a new operational taxonomic unit (OTU) belonging to the Cytophaga-Flavobacter-Bacteroides phylum, which the authors named 'mouse intestinal bacteria'. 16S rRNA probes were developed for this new OTU. Upon analysis of the novel sequences, eight were found to cluster within the Eubacterium rectale-Clostridium coccoides group and three clustered within the Bacteroides group. One of the novel sequences was distantly related to Verrucomicrobium spinosum and one was distantly related to Bacillus mycoides. Oligonucleotide probes specific for the 16S rRNA of these novel clones were generated. Using a combination of four previously described and four newly designed probes, approximately 80% of bacteria recovered from the murine large intestine and 71% of bacteria recovered from the murine caecum could be identified by fluorescence in situ hybridization (FISH).

Pyrosequencing-based assessment of the bacteria diversity in surface and subsurface peat layers of a northern wetland, with focus on poorly studied phyla and candidate divisions.

PubMed

Serkebaeva, Yulia M; Kim, Yongkyu; Liesack, Werner; Dedysh, Svetlana N

2013-01-01

Northern peatlands play a key role in the global carbon and water budget, but the bacterial diversity in these ecosystems remains poorly described. Here, we compared the bacterial community composition in the surface (0-5 cm depth) and subsurface (45-50 cm) peat layers of an acidic (pH 4.0) Sphagnum-dominated wetland, using pyrosequencing of 16S rRNA genes. The denoised sequences (37,229 reads, average length ∼430 bp) were affiliated with 27 bacterial phyla and corresponded to 1,269 operational taxonomic units (OTUs) determined at 97% sequence identity. Abundant OTUs were affiliated with the Acidobacteria (35.5±2.4% and 39.2±1.2% of all classified sequences in surface and subsurface peat, respectively), Alphaproteobacteria (15.9±1.7% and 25.8±1.4%), Actinobacteria (9.5±2.0% and 10.7±0.5%), Verrucomicrobia (8.5±1.4% and 0.6±0.2%), Planctomycetes (5.8±0.4% and 9.7±0.6%), Deltaproteobacteria (7.1±0.4% and 4.4%±0.3%), and Gammaproteobacteria (6.6±0.4% and 2.1±0.1%). The taxonomic patterns of the abundant OTUs were uniform across all the subsamples taken from each peat layer. In contrast, the taxonomic patterns of rare OTUs were different from those of the abundant OTUs and varied greatly among subsamples, in both surface and subsurface peat. In addition to the bacterial taxa listed above, rare OTUs represented the following groups: Armatimonadetes, Bacteroidetes, Chlamydia, Chloroflexi, Cyanobacteria, Elusimicrobia, Fibrobacteres, Firmicutes, Gemmatimonadetes, Spirochaetes, AD3, WS1, WS4, WS5, WYO, OD1, OP3, BRC1, TM6, TM7, WPS-2, and FCPU426. OTU richness was notably higher in the surface layer (882 OTUs) than in the anoxic subsurface peat (483 OTUs), with only 96 OTUs common to both data sets. Most members of poorly studied phyla, such as the Acidobacteria, Verrucomicrobia, Planctomycetes and the candidate division TM6, showed a clear preference for growth in either oxic or anoxic conditions. Apparently, the bacterial communities in surface and subsurface layers of northern peatlands are highly diverse and taxonomically distinct, reflecting the different abiotic conditions in microhabitats within the peat profile.

A Web-Based Multi-Database System Supporting Distributed Collaborative Management and Sharing of Microarray Experiment Information

PubMed Central

Burgarella, Sarah; Cattaneo, Dario; Masseroli, Marco

2006-01-01

We developed MicroGen, a multi-database Web based system for managing all the information characterizing spotted microarray experiments. It supports information gathering and storing according to the Minimum Information About Microarray Experiments (MIAME) standard. It also allows easy sharing of information and data among all multidisciplinary actors involved in spotted microarray experiments. PMID:17238488

Carbonate-Dissolving Bacteria from ‘Miliolite’, a Bioclastic Limestone, from Gopnath, Gujarat, Western India

PubMed Central

Subrahmanyam, Gangavarapu; Vaghela, Ravi; Bhatt, Nilesh Pinakinprasad; Archana, Gattupalli

2012-01-01

In the present investigation, the abundance and molecular phylogeny of part of the culturable bacterial population involved in the dissolution of “miliolite”, a bioclastic limestone, from Gopnath, India, was studied. Carbonate-dissolving bacteria were isolated, enumerated and screened for their ability to dissolve miliolite. Amplified ribosomal DNA restriction analysis (ARDRA) indicated 14 operational taxonomic units (OTUs) to be distributed in 5 different clades at a similarity coefficient of 0.85. Then, 16S rRNA sequence analysis helped to decipher that the majority of carbonate-dissolving bacteria were affiliated to phyla Firmicutes (Families Bacillaceae and Staphylococcaceae) and Actinobacteria (Family Promicromonosporaceae) indicating their role in miliolite weathering. PMID:22446314

Phylogenetic relationships of Sonoran Desert cactus beetles in the tribe Hololeptini (Coleoptera: Histeridae: Histerinae), with comments on the taxonomic status of Iliotona beyeri.

PubMed

Pfeiler, Edward; Vergara-Quintanar, Joel E; Castrezana, Sergio; Caterino, Michael S; Markow, Therese A

2010-07-01

Nucleotide sequences from 16S rRNA and cytochrome c oxidase subunit I (COI) were used to examine phylogenetic relationships and evolution of beetles from the tribe Hololeptini (Coleoptera: Histeridae: Histerinae) that inhabit necrotic tissue of columnar cacti in the Sonoran Desert. Phylogenetic and morphological analyses revealed the presence of seven separate lineages, three representing species in the genus Iliotona, including I. beyeri stat. nov., and four species belonging to the genus Hololepta (sensu lato). The possible roles of historical vicariance and host plant associations on the evolution of the Hololeptini from the Sonoran Desert are discussed. Copyright 2010 Elsevier Inc. All rights reserved.

The complete chloroplast genomes of two Wisteria species, W. floribunda and W. sinensis (Fabaceae).

PubMed

Kim, Na-Rae; Kim, Kyunghee; Lee, Sang-Choon; Lee, Jung-Hoon; Cho, Seong-Hyun; Yu, Yeisoo; Kim, Young-Dong; Yang, Tae-Jin

2016-11-01

Wisteria floribunda and Wisteria sinensis are ornamental woody vines in the Fabaceae. The complete chloroplast genome sequences of the two species were generated by de novo assembly using whole genome next generation sequences. The chloroplast genomes of W. floribunda and W. sinensis were 130 960 bp and 130 561 bp long, respectively, and showed inverted repeat (IR)-lacking structures as those reported in IRLC in the Fabaceae. The chloroplast genomes of both species contained same number of protein-coding sequences (77), tRNA genes (30), and rRNA genes (4). The phylogenetic analysis with the reported chloroplast genomes confirmed close taxonomical relationship of W. floribunda and W. sinensis.

Rhizobium capsici sp. nov., isolated from root tumor of a green bell pepper (Capsicum annuum var. grossum) plant.

PubMed

Lin, Shih-Yao; Hung, Mei-Hua; Hameed, Asif; Liu, You-Cheng; Hsu, Yi-Han; Wen, Cheng-Zhe; Arun, A B; Busse, Hans-Jürgen; Glaeser, Stefanie P; Kämpfer, Peter; Young, Chiu-Chung

2015-03-01

A novel, Gram-staining-negative, rod-shaped, aerobic and motile bacterium, designated strain CC-SKC2(T), was isolated from the root tumor of a green bell pepper (Capsicum annuum var. grossum) plant in Taiwan. Cells were positive for oxidase and catalase activities and exhibited growth at 25-37 °C, pH 4.0-9.0 and tolerated NaCl concentrations up to 4.0 % (w/v). Strain CC-SKC2(T) is able to trigger nodulation in soybean (Glycine max Merr.), but not in Capsicum annuum var. grossum, red bean (Vigna angularis), sesbania (Sesbania roxburghii Merr.) or alfalfa (Medicago varia Martin.). The novel strain shared highest 16S rRNA gene sequence similarity to Rhizobium rhizoryzae KCTC 23652(T) and Rhizobium straminoryzae CC-LY845(T) (both 97.5 %) followed by Rhizobium lemnae L6-16(T) (97.3 %), Rhizobium pseudoryzae KCTC 23294(T) (97.1 %), and Rhizobium paknamense NBRC 109338(T) (97.0 %), whereas other Rhizobium species shared <96.7 % similarity. The DNA-DNA relatedness values of strain CC-SKC2(T) with R. rhizoryzae KCTC 23652(T), R. pseudoryzae KCTC 23294(T) and R. paknamense NBRC 109338(T) were 11.4, 17.2 and 17.0 %, respectively (reciprocal values were 11.1, 28.3 and 24.0 %, respectively). Phylogenetic analysis based on 16S rRNA, atpD and recA genes revealed a distinct taxonomic position attained by strain CC-SKC2(T) with respect to other Rhizobium species. The major fatty acids in strain CC-SKC2(T) were C16:0, C19:0 cyclo ω8c, C14:0 3-OH and/or C16:1 iso I and C18:1 ω7c and/or C18:1 ω6c. The polyamine pattern showed predominance of spermidine and moderate amounts of sym-homospermidine. The predominant quinone system was ubiquinone (Q-10) and the DNA G+C content was 60.5 mol%. On the basis of polyphasic taxonomic evidence presented here, strain CC-SKC2(T) is proposed to represent a novel species within the genus Rhizobium, for which the name Rhizobium capsici sp. nov. is proposed. The type strain is CC-SKC2(T) (=BCRC 80699(T) = JCM 19535(T)).