21 CFR 866.2330 - Enriched culture medium.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Enriched culture medium. 866.2330 Section 866.2330 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Microbiology Devices § 866.2330 Enriched culture...

21 CFR 866.2330 - Enriched culture medium.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Enriched culture medium. 866.2330 Section 866.2330 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Microbiology Devices § 866.2330 Enriched culture...

21 CFR 866.2330 - Enriched culture medium.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Enriched culture medium. 866.2330 Section 866.2330 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Microbiology Devices § 866.2330 Enriched culture...

21 CFR 866.2330 - Enriched culture medium.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Enriched culture medium. 866.2330 Section 866.2330 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Microbiology Devices § 866.2330 Enriched culture...

21 CFR 866.2330 - Enriched culture medium.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Enriched culture medium. 866.2330 Section 866.2330 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Microbiology Devices § 866.2330 Enriched culture...

TOWARD AN ENRICHED CULTURAL FUTURE.

ERIC Educational Resources Information Center

New York City Board of Education, Brooklyn, NY.

DEVELOPMENT OF INTEREST IN WORTHWHILE CULTURAL ACTIVITIES HAS BEEN ONE OF IMPORTANT EMPHASES OF THE HIGH HORIZONS PROGRAM. THE CULTURAL ENRICHMENT TEACHER HAS HAD THE RESPONSIBILITY FOR TEACHING MUSIC AND/OR ART IN ADDITION TO CORRELATING CULTURAL ACTIVITIES WITH ALL FACETS OF THE CURRICULUM TO PROVIDE INSPIRATION AND NEW EXPERIENCES AS WELL AS TO…

Characterization of bacterial diversity in an atrazine degrading enrichment culture and degradation of atrazine, cyanuric acid and biuret in industrial wastewater.

PubMed

Dutta, Anirban; Vasudevan, Venugopal; Nain, Lata; Singh, Neera

2016-01-01

An enrichment culture was used to study atrazine degradation in mineral salt medium (MSM) (T1), MSM+soil extract (1:1, v/v) (T2) and soil extract (T3). Results suggested that enrichment culture required soil extract to degrade atrazine, as after second sequential transfer only partial atrazine degradation was observed in T1 treatment while atrazine was completely degraded in T2 and T3 treatments even after fourth transfer. Culture independent polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) technique confirmed selective enrichment of genus Bacillus along with Pseudomonas and Burkholderia. Degradation of atrazine/metabolites in the industrial wastewater was studied at different initial concentrations of the contaminants [wastewater-water (v/v) ratio: T1, 1:9; T2, 2:8; T3, 3:7; T4, 5:5 and T5, undiluted effluent]. The initial concentrations of atrazine, cyanuric acid and biuret ranged between 5.32 and 53.92 µg mL(-1), 265.6 and 1805.2 µg mL(-1) and 1.85 and 16.12 µg mL(-1), respectively. The enrichment culture was able to completely degrade atrazine, cyanuric acid and biuret up to T4 treatment, while no appreciable degradation of contaminants was observed in the undiluted effluent (T5). Inability of enrichment culture to degrade atrazine/metabolites might be due to high concentrations of cyanuric acid. Therefore, a separate study on cyanuric acid degradation suggested: (i) no appreciable cyanuric acid degradation with accumulation of an unidentified metabolite in the medium where cyanuric acid was supplemented as the sole source of carbon and nitrogen; (ii) partial cyanuric acid degradation with accumulation of unidentified metabolite in the medium containing additional nitrogen source; and (iii) complete cyanuric acid degradation in the medium supplemented with an additional carbon source. This unidentified metabolite observed during cyanuric acid degradation and also detected in the enrichment culture inoculated wastewater samples

Differential Response of High-Elevation Planktonic Bacterial Community Structure and Metabolism to Experimental Nutrient Enrichment

PubMed Central

Nelson, Craig E.; Carlson, Craig A.

2011-01-01

Nutrient enrichment of high-elevation freshwater ecosystems by atmospheric deposition is increasing worldwide, and bacteria are a key conduit for the metabolism of organic matter in these oligotrophic environments. We conducted two distinct in situ microcosm experiments in a high-elevation lake (Emerald Lake, Sierra Nevada, California, USA) to evaluate responses in bacterioplankton growth, carbon utilization, and community structure to short-term enrichment by nitrate and phosphate. The first experiment, conducted just following ice-off, employed dark dilution culture to directly assess the impact of nutrients on bacterioplankton growth and consumption of terrigenous dissolved organic matter during snowmelt. The second experiment, conducted in transparent microcosms during autumn overturn, examined how bacterioplankton in unmanipulated microbial communities responded to nutrients concomitant with increasing phytoplankton-derived organic matter. In both experiments, phosphate enrichment (but not nitrate) caused significant increases in bacterioplankton growth, changed particulate organic stoichiometry, and induced shifts in bacterial community composition, including consistent declines in the relative abundance of Actinobacteria. The dark dilution culture showed a significant increase in dissolved organic carbon removal in response to phosphate enrichment. In transparent microcosms nutrient enrichment had no effect on concentrations of chlorophyll, carbon, or the fluorescence characteristics of dissolved organic matter, suggesting that bacterioplankton responses were independent of phytoplankton responses. These results demonstrate that bacterioplankton communities in unproductive high-elevation habitats can rapidly alter their taxonomic composition and metabolism in response to short-term phosphate enrichment. Our results reinforce the key role that phosphorus plays in oligotrophic lake ecosystems, clarify the nature of bacterioplankton nutrient limitation, and

THE ANAEROBIC BIODEGRADATION OF O-,M- AND P-CRESOL BY SULFATE-REDUCING BACTERIAL ENRICHMENT CULTURES OBTAINED FROM A SHALLOW ANOXIC AQUIFER

EPA Science Inventory

Sulfate-reducing bacterial enrichments were obtained from a shallow anoxic aquifer for their ability to metabolize either o-, m-, orp-cresol. GC/MS and simultaneous adaptation experiments suggested that the anaerobic decomposition of p-cresol proceeds ...

Evaluation of culture techniques and bacterial cultures from uroliths.

PubMed

Perry, Leigh A; Kass, Philip H; Johnson, Dee L; Ruby, Annette L; Shiraki, Ryoji; Westropp, Jodi L

2013-03-01

The association between urolithiasis and growth of bacteria in the urine or urolith has not been recently evaluated in the past 15 years, and the effects of antimicrobial administration on urolith cultures have not been reported. As well, laboratory techniques for urolith cultures have not been critically evaluated. The objectives of the current study were to 1) report bacterial isolates from uroliths and their association with signalment, urolith composition, antimicrobial use, and urine cultures and 2) evaluate laboratory techniques for urolith cultures. For the first objective, a retrospective search of bacterial isolates cultured from uroliths submitted to the laboratory as well as the signalment, urine culture results, and antimicrobial use were recorded. For the second objective, 50 urolith pairs were cultured by washing each urolith either 1or 4 times and culturing the core. Five hundred twenty canine and 168 feline uroliths were reviewed. Struvite-containing uroliths had an increased prevalence of a positive culture compared to nonstruvite-containing uroliths (P < 0.0001, odds ratio [OR] = 5.4), as did uroliths from female dogs (P < 0.0001, OR = 2.9). No significant difference between culture results and previous antimicrobial administration was found (P = 0.41). Eighteen percent of cases with negative urine cultures had positive urolith cultures. There was no significant difference in core culture results whether the urolith was washed 1 or 4 times (P = 0.07). Urolith culture outcome was not always influenced by previous antimicrobial administration, and bacterial culture of a urolith may not yield the same results as those obtained from the urine. The modified protocol, which requires less time and expense for urolith cultures, may be an acceptable alternative.

In-Vessel Co-Composting of Food Waste Employing Enriched Bacterial Consortium

PubMed Central

2018-01-01

Summary The aim of the present study is to develop a good initial composting mix using a bacterial consortium and 2% lime for effective co-composting of food waste in a 60-litre in-vessel composter. In the experiment that lasted for 42 days, the food waste was first mixed with sawdust and 2% lime (by dry mass), then one of the reactors was inoculated with an enriched bacterial consortium, while the other served as control. The results show that inoculation of the enriched natural bacterial consortium effectively overcame the oil-laden co-composting mass in the composter and increased the rate of mineralization. In addition, CO2 evolution rate of (0.81±0.2) g/(kg·day), seed germination index of (105±3) %, extractable ammonium mass fraction of 305.78 mg/kg, C/N ratio of 16.18, pH=7.6 and electrical conductivity of 3.12 mS/cm clearly indicate that the compost was well matured and met the composting standard requirements. In contrast, control treatment exhibited a delayed thermophilic phase and did not mature after 42 days, as evidenced by the maturity parameters. Therefore, a good composting mix and potential bacterial inoculum to degrade the oil are essential for food waste co-composting systems. PMID:29796000

In-Vessel Co-Composting of Food Waste Employing Enriched Bacterial Consortium.

PubMed

Awasthi, Mukesh Kumar; Wang, Quan; Wang, Meijing; Chen, Hongyu; Ren, Xiuna; Zhao, Junchao; Zhang, Zengqiang

2018-03-01

The aim of the present study is to develop a good initial composting mix using a bacterial consortium and 2% lime for effective co-composting of food waste in a 60-litre in-vessel composter. In the experiment that lasted for 42 days, the food waste was first mixed with sawdust and 2% lime (by dry mass), then one of the reactors was inoculated with an enriched bacterial consortium, while the other served as control. The results show that inoculation of the enriched natural bacterial consortium effectively overcame the oil-laden co-composting mass in the composter and increased the rate of mineralization. In addition, CO 2 evolution rate of (0.81±0.2) g/(kg·day), seed germination index of (105±3) %, extractable ammonium mass fraction of 305.78 mg/kg, C/N ratio of 16.18, pH=7.6 and electrical conductivity of 3.12 mS/cm clearly indicate that the compost was well matured and met the composting standard requirements. In contrast, control treatment exhibited a delayed thermophilic phase and did not mature after 42 days, as evidenced by the maturity parameters. Therefore, a good composting mix and potential bacterial inoculum to degrade the oil are essential for food waste co-composting systems.

Bacterial diversity of Taxus rhizosphere: culture-independent and culture-dependent approaches.

PubMed

Hao, Da Cheng; Ge, Guang Bo; Yang, Ling

2008-07-01

The regional variability of Taxus rhizosphere bacterial community composition and diversity was studied by comparative analysis of three large 16S rRNA gene clone libraries from the Taxus rhizosphere in different regions of China (subtropical and temperate regions). One hundred and forty-six clones were screened for three libraries. Phylogenetic analysis of 16S rRNA gene sequences demonstrated that the abundance of sequences affiliated with Gammaproteobacteria, Betaproteobacteria, and Actinobacteria was higher in the library from the T. xmedia rhizosphere of the temperate region compared with the subtropical Taxus mairei rhizosphere. On the other hand, Acidobacteria was more abundant in libraries from the subtropical Taxus mairei rhizosphere. Richness estimates and diversity indices of three libraries revealed major differences, indicating a higher richness in the Taxus rhizosphere bacterial communities of the subtropical region and considerable variability in the bacterial community composition within this region. By enrichment culture, a novel Actinobacteria strain DICP16 was isolated from the T. xmedia rhizosphere of the temperate region and was identified as Leifsonia shinshuensis sp. via 16S rRNA gene and gyrase B sequence analyses. DICP16 was able to remove the xylosyl group from 7-xylosyl-10-deacetylbaccatin III and 7-xylosyl-10-deacetylpaclitaxel, thereby making the xylosyltaxanes available as sources of 10-deacetylbaccatin III and the anticancer drug paclitaxel. Taken together, the present studies provide, for the first time, the knowledge of the biodiversity of microorganisms populating Taxus rhizospheres.

Determining the culturability of the rumen bacterial microbiome

PubMed Central

Creevey, Christopher J; Kelly, William J; Henderson, Gemma; Leahy, Sinead C

2014-01-01

The goal of the Hungate1000 project is to generate a reference set of rumen microbial genome sequences. Toward this goal we have carried out a meta-analysis using information from culture collections, scientific literature, and the NCBI and RDP databases and linked this with a comparative study of several rumen 16S rRNA gene-based surveys. In this way we have attempted to capture a snapshot of rumen bacterial diversity to examine the culturable fraction of the rumen bacterial microbiome. Our analyses have revealed that for cultured rumen bacteria, there are many genera without a reference genome sequence. Our examination of culture-independent studies highlights that there are few novel but many uncultured taxa within the rumen bacterial microbiome. Taken together these results have allowed us to compile a list of cultured rumen isolates that are representative of abundant, novel and core bacterial species in the rumen. In addition, we have identified taxa, particularly within the phylum Bacteroidetes, where further cultivation efforts are clearly required. This information is being used to guide the isolation efforts and selection of bacteria from the rumen microbiota for sequencing through the Hungate1000. PMID:24986151

Use of Nitrogen-15-Enriched Escherichia coli as a Bacterial Tracer in Karst Aquifers.

PubMed

Ward, James W; Warden, John G; Bandy, Ashley M; Fryar, Alan E; Brion, Gail M; Macko, Stephen A; Romanek, Christopher S; Coyne, Mark S

2016-11-01

Karst aquifers are susceptible to contamination by microorganisms, but relatively few studies have used bacteria as tracers. We demonstrate the utility of Escherichia coli enriched in the stable isotope nitrogen-15 ( 15 N) as a novel bacterial tracer. Nonpathogenic E. coli from two springs in central Kentucky were grown on 15 N-enriched media. Survival of E. coli and persistence of the isotopic signal were assessed in two sets of laboratory experiments conducted with sterilized spring water in dark microcosms at 14 °C. First, isotopically labeled bacteria survived for 130 d at concentrations within one log unit of the average initial value, and there was no significant difference in δ 15 N values from Day 1 to Day 130. Second, water samples with E. coli were inoculated with either of two different species of protozoa (Tetrahymena pyriformis or Colpoda steinii). During 7 d, δ 15 N values increased in T. pyriformis while bacterial populations decreased. In a field test, following a 2.1-cm rainfall, 15 N-labeled E. coli, solutes (rhodamine WT dye and bromide), and latex microspheres were injected into a sinkhole approximately 530 m upgradient of a spring. Breakthrough of all tracers coincided, but microspheres were remobilized by subsequent storms, unlike other tracers. Enriched E. coli exhibited more tailing than solute tracers during the initial storm-flow recession. These results indicate that 15 N-enriched E. coli is a viable tracer of bacterial transport in karst aquifers, although predation may attenuate the isotopic signal in systems that are not rapidly flushed. © 2016, National Ground Water Association.

Arsenic uptake in bacterial calcite

NASA Astrophysics Data System (ADS)

Catelani, Tiziano; Perito, Brunella; Bellucci, Francesco; Lee, Sang Soo; Fenter, Paul; Newville, Matthew; Rimondi, Valentina; Pratesi, Giovanni; Costagliola, Pilario

2018-02-01

Bio-mediated processes for arsenic (As) uptake in calcite were investigated by means of X-ray Diffraction (XRD) and X-ray Absorption Spectroscopy (XAS) coupled with X-ray Fluorescence measurements. The environmental bacterial strain Bacillus licheniformis BD5, sampled at the Bullicame Hot Springs (Viterbo, Central Italy), was used to synthesize calcite from As-enriched growth media. Both liquid and solid cultures were applied to simulate planktonic and biofilm community environments, respectively. Bacterial calcite samples cultured in liquid media had an As enrichment factor (Kd) 50 times higher than that from solid media. The XRD analysis revealed an elongation of the crystal lattice along the c axis (by 0.03 Å) for biogenic calcite, which likely resulted from the substitution of larger arsenate for carbonate in the crystal. The XAS data also showed a clear difference in the oxidation state of sorbed As between bacterial and abiotic calcite. Abiotic chemical processes yielded predominantly As(V) uptake whereas bacterial precipitation processes led to the uptake of both As(III) and As(V). The presence of As(III) in bacterial calcite is proposed to result from subsequent reduction of arsenate to arsenite by bacterial activities. To the best of our knowledge, this is the first experimental observation of the incorporation of As(III) in the calcite crystal lattice, revealing a critical role of biochemical processes for the As cycling in nature.

Bacterial Cell Production from Hexadecane at High Temperatures

PubMed Central

Sukatsch, Dieter A.; Johnson, Marvin J.

1972-01-01

On mineral medium with hexadecane as the sole carbon source, stable mixed bacterial enrichment cultures were obtained from soil inoculum at 25, 35, 45, 55, and 65 C. Cell yields (grams of dry cells per gram of hexadecane) were determined for each of the enrichment cultures grown at the temperature at which they were enriched, and also for the 55 and 65 C cultures grown at various temperatures. In all cases, cell yields decreased with increasing growth temperature. The highest yield obtained at 65 C was 0.26, and the lowest yield obtained at 25 or 35 C was 1.02. Slower growth was observed at higher temperatures. PMID:5021971

Multiplexed Single Intact Cell Droplet Digital PCR (MuSIC ddPCR) Method for Specific Detection of Enterohemorrhagic E. coli (EHEC) in Food Enrichment Cultures

PubMed Central

McMahon, Tanis C.; Blais, Burton W.; Wong, Alex; Carrillo, Catherine D.

2017-01-01

Foodborne illness attributed to enterohemorrhagic E. coli (EHEC), a highly pathogenic subset of Shiga toxin-producing E. coli (STEC), is increasingly recognized as a significant public health issue. Current microbiological methods for identification of EHEC in foods often use PCR-based approaches to screen enrichment broth cultures for characteristic gene markers [i.e., Shiga toxin (stx) and intimin (eae)]. However, false positives arise when complex food matrices, such as beef, contain mixtures of eae-negative STEC and eae-positive E. coli, but no EHEC with both markers in a single cell. To reduce false-positive detection of EHEC in food enrichment samples, a Multiplexed, Single Intact Cell droplet digital PCR (MuSIC ddPCR) assay capable of detecting the co-occurrence of the stx and eae genes in a single bacterial cell was developed. This method requires: (1) dispersal of intact bacteria into droplets; (2) release of genomic DNA (gDNA) by heat lysis; and (3) amplification and detection of genetic targets (stx and eae) using standard TaqMan chemistries with ddPCR. Performance of the method was tested with panels of EHEC and non-target E. coli. By determining the linkage (i.e., the proportion of droplets in which stx and eae targets were both amplified), samples containing EHEC (typically greater than 20% linkage) could be distinguished from samples containing mixtures of eae-negative STEC and eae-positive E. coli (0–2% linkage). The use of intact cells was necessary as this linkage was not observed with gDNA extracts. EHEC could be accurately identified in enrichment broth cultures containing excess amounts of background E. coli and in enrichment cultures derived from ground beef/pork and leafy-green produce samples. To our knowledge, this is the first report of dual-target detection in single bacterial cells using ddPCR. The application of MuSIC ddPCR to enrichment-culture screening would reduce false-positives, thereby improving the cost, speed, and accuracy of

Multiplexed Single Intact Cell Droplet Digital PCR (MuSIC ddPCR) Method for Specific Detection of Enterohemorrhagic E. coli (EHEC) in Food Enrichment Cultures.

PubMed

McMahon, Tanis C; Blais, Burton W; Wong, Alex; Carrillo, Catherine D

2017-01-01

Foodborne illness attributed to enterohemorrhagic E. coli (EHEC), a highly pathogenic subset of Shiga toxin-producing E. coli (STEC), is increasingly recognized as a significant public health issue. Current microbiological methods for identification of EHEC in foods often use PCR-based approaches to screen enrichment broth cultures for characteristic gene markers [i.e., Shiga toxin ( stx ) and intimin ( eae )]. However, false positives arise when complex food matrices, such as beef, contain mixtures of eae -negative STEC and eae -positive E. coli , but no EHEC with both markers in a single cell. To reduce false-positive detection of EHEC in food enrichment samples, a Multiplexed, Single Intact Cell droplet digital PCR (MuSIC ddPCR) assay capable of detecting the co-occurrence of the stx and eae genes in a single bacterial cell was developed. This method requires: (1) dispersal of intact bacteria into droplets; (2) release of genomic DNA (gDNA) by heat lysis; and (3) amplification and detection of genetic targets ( stx and eae ) using standard TaqMan chemistries with ddPCR. Performance of the method was tested with panels of EHEC and non-target E. coli . By determining the linkage (i.e., the proportion of droplets in which stx and eae targets were both amplified), samples containing EHEC (typically greater than 20% linkage) could be distinguished from samples containing mixtures of eae -negative STEC and eae -positive E. coli (0-2% linkage). The use of intact cells was necessary as this linkage was not observed with gDNA extracts. EHEC could be accurately identified in enrichment broth cultures containing excess amounts of background E. coli and in enrichment cultures derived from ground beef/pork and leafy-green produce samples. To our knowledge, this is the first report of dual-target detection in single bacterial cells using ddPCR. The application of MuSIC ddPCR to enrichment-culture screening would reduce false-positives, thereby improving the cost, speed, and

CUE (CULTURE, UNDERSTANDING, ENRICHMENT)--SOCIAL STUDIES.

ERIC Educational Resources Information Center

BROWN, ROBERT M.; AND OTHERS

THIS PUBLICATION IS A TEACHING GUIDE TO PROVIDE GUIDANCE FOR INTEGRATING CAREFULLY SELECTED AUDIOVISUAL ITEMS INTO EXISTING NINTH-GRADE CURRICULUMS IN SOCIAL STUDIES. IT IS ONE OF FIVE GUIDES PREPARED FOR USE IN PROJECT CUE. AN EXPERIMENTAL PROGRAM DESIGNED TO INCREASE CULTURAL UNDERSTANDING AND ENRICHMENT IN THE EDUCATIONAL PROGRAMS OF HIGH…

Initial nitrogen enrichment conditions determines variations in nitrogen substrate utilization by heterotrophic bacterial isolates.

PubMed

Ghosh, Suchismita; Ayayee, Paul A; Valverde-Barrantes, Oscar J; Blackwood, Christopher B; Royer, Todd V; Leff, Laura G

2017-04-04

The nitrogen (N) cycle consists of complex microbe-mediated transformations driven by a variety of factors, including diversity and concentrations of N compounds. In this study, we examined taxonomic diversity and N substrate utilization by heterotrophic bacteria isolated from streams under complex and simple N-enrichment conditions. Diversity estimates differed among isolates from the enrichments, but no significant composition were detected. Substrate utilization and substrate range of bacterial assemblages differed within and among enrichments types, and not simply between simple and complex N-enrichments. N substrate use patterns differed between isolates from some complex and simple N-enrichments while others were unexpectedly similar. Taxonomic composition of isolates did not differ among enrichments and was unrelated to N use suggesting strong functional redundancy. Ultimately, our results imply that the available N pool influences physiology and selects for bacteria with various abilities that are unrelated to their taxonomic affiliation.

Bacterial communities and metabolic activity of faecal cultures from equol producer and non-producer menopausal women under treatment with soy isoflavones.

PubMed

Guadamuro, Lucía; Dohrmann, Anja B; Tebbe, Christoph C; Mayo, Baltasar; Delgado, Susana

2017-04-17

Isoflavones are polyphenols with estrogenic activity found mainly in soy and soy-derived products that need to be metabolised in the intestine by the gut bacteria to be fully active. There is little knowledge about isoflavone bioconversion and equol production in the human intestine. In this work, we developed an in vitro anaerobic culture model based on faecal slurries to assess the impact of isoflavone supplementation on the overall intestinal bacterial composition changes and associated metabolic transformations. In the faecal anaerobic batch cultures of this study bioconversion of isoflavones into equol was possible, suggesting the presence of viable equol-producing bacterial taxa within the faeces of menopausal women with an equol producer phenotype. The application of high-throughput DNA sequencing of 16S rRNA gene amplicons revealed the composition of the faecal cultures to be modified by the addition of isoflavones, with enrichment of some bacterial gut members associated with the metabolism of phenolics and/or equol production, such as Collinsella, Faecalibacterium and members of the Clostridium clusters IV and XIVa. In addition, the concentration of short-chain fatty acids (SCFAs) detected in the isoflavone-containing faecal cultures was higher in those inoculated with faecal slurries from equol-producing women. This study constitutes the first step in the development of a faecal culturing system with isoflavones that would further allow the selection and isolation of intestinal bacterial types able to metabolize these compounds and produce equol in vitro. Although limited by the low number of faecal cultures analysed and the inter-individual bacterial diversity, the in vitro results obtained in this work tend to indicate that soy isoflavones might provide an alternative energy source for the increase of equol-producing taxa and enhancement of SCFAs production. SCFAs and equol are both considered pivotal bacterial metabolites in the triggering of

Arsenic uptake in bacterial calcite

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

Catelani, Tiziano; Perito, Brunella; Bellucci, Francesco

Bio-mediated processes for arsenic (As) uptake in calcite were investigated by means of X-ray Diffraction (XRD) and Xray Absorption Spectroscopy (XAS) coupled with X-ray Fluorescence measurements. The environmental bacterial strain Bacillus licheniformis BD5, sampled at the Bullicame Hot Springs (Viterbo, Central Italy), was used to synthesize calcite from As-enriched growth media. Both liquid and solid cultures were applied to simulate planktonic and biofilm community environments, respectively. Bacterial calcite samples cultured in liquid media had an As enrichment factor (Kd) 50 times higher than that from solid media. The XRD analysis revealed an elongation of the crystal lattice along the cmore » axis (by 0.03Å) for biogenic calcite, which likely resulted from the substitution of larger arsenate for carbonate in the crystal. The XAS data also showed a clear difference in the oxidation state of sorbed As between bacterial and abiotic calcite. Abiotic chemical processes yielded predominantly As(V) uptake whereas bacterial precipitation processes led to the uptake of both As(III) and As(V). The presence of As(III) in bacterial calcite is proposed to result from subsequent reduction of arsenate to arsenite by bacterial activities. To the best of our knowledge, this is the first experimental observation of the incorporation of As(III) in the calcite crystal lattice, revealing a critical role of biochemical processes for the As cycling in nature.« less

Culture Media and Individual Hosts Affect the Recovery of Culturable Bacterial Diversity from Amphibian Skin

PubMed Central

Medina, Daniel; Walke, Jenifer B.; Gajewski, Zachary; Becker, Matthew H.; Swartwout, Meredith C.; Belden, Lisa K.

2017-01-01

One current challenge in microbial ecology is elucidating the functional roles of the large diversity of free-living and host-associated bacteria identified by culture-independent molecular methods. Importantly, the characterization of this immense bacterial diversity will likely require merging data from culture-independent approaches with work on bacterial isolates in culture. Amphibian skin bacterial communities have become a recent focus of work in host-associated microbial systems due to the potential role of these skin bacteria in host defense against the pathogenic fungus Batrachochytrium dendrobatidis (Bd), which is associated with global amphibian population declines and extinctions. As there is evidence that some skin bacteria may inhibit growth of Bd and prevent infection in some cases, there is interest in using these bacteria as probiotic therapy for conservation of at-risk amphibians. In this study, we used skin swabs from American toads (Anaxyrus americanus) to: (1) assess the diversity and community structure of culturable amphibian skin bacteria grown on high and low nutrient culture media, (2) determine which culture media recover the highest proportion of the total skin bacterial community of individual toads relative to culture-independent data, and (3) assess whether the plated communities from the distinct media types vary in their ability to inhibit Bd growth in in-vitro assays. Overall, we found that culture media with low nutrient concentrations facilitated the growth of more diverse bacterial taxa and grew distinct communities relative to media with higher nutrient concentrations. Use of low nutrient media also resulted in culturing proportionally more of the bacterial diversity on individual toads relative to the overall community defined using culture-independent methods. However, while there were differences in diversity among media types, the variation among individual hosts was greater than variation among media types, suggesting that

Culture Media and Individual Hosts Affect the Recovery of Culturable Bacterial Diversity from Amphibian Skin.

PubMed

Medina, Daniel; Walke, Jenifer B; Gajewski, Zachary; Becker, Matthew H; Swartwout, Meredith C; Belden, Lisa K

2017-01-01

One current challenge in microbial ecology is elucidating the functional roles of the large diversity of free-living and host-associated bacteria identified by culture-independent molecular methods. Importantly, the characterization of this immense bacterial diversity will likely require merging data from culture-independent approaches with work on bacterial isolates in culture. Amphibian skin bacterial communities have become a recent focus of work in host-associated microbial systems due to the potential role of these skin bacteria in host defense against the pathogenic fungus Batrachochytrium dendrobatidis (Bd), which is associated with global amphibian population declines and extinctions. As there is evidence that some skin bacteria may inhibit growth of Bd and prevent infection in some cases, there is interest in using these bacteria as probiotic therapy for conservation of at-risk amphibians. In this study, we used skin swabs from American toads ( Anaxyrus americanus ) to: (1) assess the diversity and community structure of culturable amphibian skin bacteria grown on high and low nutrient culture media, (2) determine which culture media recover the highest proportion of the total skin bacterial community of individual toads relative to culture-independent data, and (3) assess whether the plated communities from the distinct media types vary in their ability to inhibit Bd growth in in-vitro assays. Overall, we found that culture media with low nutrient concentrations facilitated the growth of more diverse bacterial taxa and grew distinct communities relative to media with higher nutrient concentrations. Use of low nutrient media also resulted in culturing proportionally more of the bacterial diversity on individual toads relative to the overall community defined using culture-independent methods. However, while there were differences in diversity among media types, the variation among individual hosts was greater than variation among media types, suggesting

Protein Chips for Detection of Salmonella spp. from Enrichment Culture

PubMed Central

Poltronieri, Palmiro; Cimaglia, Fabio; De Lorenzis, Enrico; Chiesa, Maurizio; Mezzolla, Valeria; Reca, Ida Barbara

2016-01-01

Food pathogens are the cause of foodborne epidemics, therefore there is a need to detect the pathogens in food productions rapidly. A pre-enrichment culture followed by selective agar plating are standard detection methods. Molecular methods such as qPCR have provided a first rapid protocol for detection of pathogens within 24 h of enrichment culture. Biosensors also may provide a rapid tool to individuate a source of Salmonella contamination at early times of pre-enrichment culture. Forty mL of Salmonella spp. enrichment culture were processed by immunoseparation using the Pathatrix, as in AFNOR validated qPCR protocols. The Salmonella biosensor combined with immunoseparation showed a limit of detection of 100 bacteria/40 mL, with a 400 fold increase to previous results. qPCR analysis requires processing of bead-bound bacteria with lysis buffer and DNA clean up, with a limit of detection of 2 cfu/50 μL. Finally, a protein chip was developed and tested in screening and identification of 5 common pathogen species, Salmonella spp., E. coli, S. aureus, Campylobacter spp. and Listeria spp. The protein chip, with high specificity in species identification, is proposed to be integrated into a Lab-on-Chip system, for rapid and reproducible screening of Salmonella spp. and other pathogen species contaminating food productions. PMID:27110786

Phylogenetic analysis of anaerobic psychrophilic enrichment cultures obtained from a greenland glacier ice core

NASA Technical Reports Server (NTRS)

Sheridan, Peter P.; Miteva, Vanya I.; Brenchley, Jean E.

2003-01-01

The examination of microorganisms in glacial ice cores allows the phylogenetic relationships of organisms frozen for thousands of years to be compared with those of current isolates. We developed a method for aseptically sampling a sediment-containing portion of a Greenland ice core that had remained at -9 degrees C for over 100,000 years. Epifluorescence microscopy and flow cytometry results showed that the ice sample contained over 6 x 10(7) cells/ml. Anaerobic enrichment cultures inoculated with melted ice were grown and maintained at -2 degrees C. Genomic DNA extracted from these enrichments was used for the PCR amplification of 16S rRNA genes with bacterial and archaeal primers and the preparation of clone libraries. Approximately 60 bacterial inserts were screened by restriction endonuclease analysis and grouped into 27 unique restriction fragment length polymorphism types, and 24 representative sequences were compared phylogenetically. Diverse sequences representing major phylogenetic groups including alpha, beta, and gamma Proteobacteria as well as relatives of the Thermus, Bacteroides, Eubacterium, and Clostridium groups were found. Sixteen clone sequences were closely related to those from known organisms, with four possibly representing new species. Seven sequences may reflect new genera and were most closely related to sequences obtained only by PCR amplification. One sequence was over 12% distant from its closest relative and may represent a novel order or family. These results show that phylogenetically diverse microorganisms have remained viable within the Greenland ice core for at least 100,000 years.

Increased detection of mastitis pathogens by real-time PCR compared to bacterial culture.

PubMed

Keane, O M; Budd, K E; Flynn, J; McCoy, F

2013-09-21

Rapid and accurate identification of mastitis pathogens is important for disease control. Bacterial culture and isolate identification is considered the gold standard in mastitis diagnosis but is time consuming and results in many culture-negative samples. Identification of mastitis pathogens by PCR has been proposed as a fast and sensitive alternative to bacterial culture. The results of bacterial culture and PCR for the identification of the aetiological agent of clinical mastitis were compared. The pathogen identified by traditional culture methods was also detected by PCR in 98 per cent of cases indicating good agreement between the positive results of bacterial culture and PCR. A mastitis pathogen could not be recovered from approximately 30 per cent of samples by bacterial culture, however, an aetiological agent was identified by PCR in 79 per cent of these samples. Therefore, a mastitis pathogen was detected in significantly more milk samples by PCR than by bacterial culture (92 per cent and 70 per cent, respectively) although the clinical relevance of PCR-positive culture-negative results remains controversial. A mixed infection of two or more mastitis pathogens was also detected more commonly by PCR. Culture-negative samples due to undetected Staphylococcus aureus infections were rare. The use of PCR technology may assist in rapid mastitis diagnosis, however, accurate interpretation of PCR results in the absence of bacterial culture remains problematic.

Atrazine and its metabolites degradation in mineral salts medium and soil using an enrichment culture.

PubMed

Kumar, Anup; Singh, Neera

2016-03-01

An atrazine-degrading enrichment culture was used to study degradation of atrazine metabolites viz. hydroxyatrazine, deethylatrazine, and deisopropylatrazine in mineral salts medium. Results suggested that the enrichment culture was able to degrade only hydroxyatrazine, and it was used as the sole source of carbon and nitrogen. Hydroxyatrazine degradation slowed down when sucrose and/or ammonium hydrogen phosphate were supplemented as the additional sources of carbon and nitrogen, respectively. The enrichment culture could degrade high concentrations of atrazine (up to 110 μg/mL) in mineral salts medium, and neutral pH was optimum for atrazine degradation. Further, except in an acidic soil, enrichment culture was able to degrade atrazine in three soil types having different physico-chemical properties. Raising the pH of acidic soil to neutral or alkaline enabled the enrichment culture to degrade atrazine suggesting that acidic pH inhibited atrazine-degrading ability. The study suggested that the enrichment culture can be successfully utilized to achieve complete degradation of atrazine and its persistent metabolite hydroxyatrazine in the contaminated soil and water.

Salmonella testing of pooled pre-enrichment broth cultures for screening multiple food samples.

PubMed

Price, W R; Olsen, R A; Hunter, J E

1972-04-01

A method has been described for testing multiple food samples for Salmonella without loss in sensitivity. The method pools multiple pre-enrichment broth cultures into single enrichment broths. The subsequent stages of the Salmonella analysis are not altered. The method was found applicable to several dry food materials including nonfat dry milk, dried egg albumin, cocoa, cottonseed flour, wheat flour, and shredded coconut. As many as 25 pre-enrichment broth cultures were pooled without apparent loss in the sensitivity of Salmonella detection as compared to individual sample analysis. The procedure offers a simple, yet effective, way to increase sample capacity in the Salmonella testing of foods, particularly where a large proportion of samples ordinarily is negative. It also permits small portions of pre-enrichment broth cultures to be retained for subsequent individual analysis if positive tests are found. Salmonella testing of pooled pre-enrichment broths provides increased consumer protection for a given amount of analytical effort as compared to individual sample analysis.

Phylogenetic Analysis of Anaerobic Psychrophilic Enrichment Cultures Obtained from a Greenland Glacier Ice Core

PubMed Central

Sheridan, Peter P.; Miteva, Vanya I.; Brenchley, Jean E.

2003-01-01

The examination of microorganisms in glacial ice cores allows the phylogenetic relationships of organisms frozen for thousands of years to be compared with those of current isolates. We developed a method for aseptically sampling a sediment-containing portion of a Greenland ice core that had remained at −9°C for over 100,000 years. Epifluorescence microscopy and flow cytometry results showed that the ice sample contained over 6 × 107 cells/ml. Anaerobic enrichment cultures inoculated with melted ice were grown and maintained at −2°C. Genomic DNA extracted from these enrichments was used for the PCR amplification of 16S rRNA genes with bacterial and archaeal primers and the preparation of clone libraries. Approximately 60 bacterial inserts were screened by restriction endonuclease analysis and grouped into 27 unique restriction fragment length polymorphism types, and 24 representative sequences were compared phylogenetically. Diverse sequences representing major phylogenetic groups including alpha, beta, and gamma Proteobacteria as well as relatives of the Thermus, Bacteroides, Eubacterium, and Clostridium groups were found. Sixteen clone sequences were closely related to those from known organisms, with four possibly representing new species. Seven sequences may reflect new genera and were most closely related to sequences obtained only by PCR amplification. One sequence was over 12% distant from its closest relative and may represent a novel order or family. These results show that phylogenetically diverse microorganisms have remained viable within the Greenland ice core for at least 100,000 years. PMID:12676695

Enrichment of lignocellulose-degrading microbial communities from natural and engineered methanogenic environments.

PubMed

Ozbayram, Emine Gozde; Kleinsteuber, Sabine; Nikolausz, Marcell; Ince, Bahar; Ince, Orhan

2018-01-01

The aim of this study was to develop an effective bioaugmentation concept for anaerobic digesters treating lignocellulosic biomass such as straw. For that purpose, lignocellulose-degrading methanogenic communities were enriched on wheat straw from cow and goat rumen fluid as well as from a biogas reactor acclimated to lignocellulosic biomass (sorghum as mono-substrate). The bacterial communities of the enriched cultures and the different inocula were examined by 454 amplicon sequencing of 16S rRNA genes while the methanogenic archaeal communities were analyzed by terminal restriction fragment length polymorphism (T-RFLP) fingerprinting of the mcrA gene. Bacteroidetes was the most abundant phylum in all samples. Within the Bacteroidetes phylum, Bacteroidaceae was the most abundant family in the rumen-derived enrichment cultures, whereas Porphyromonadaceae was the predominant one in the reactor-derived culture. Additionally, the enrichment procedure increased the relative abundance of Ruminococcaceae (phylum: Firmicutes) in all cultures. T-RFLP profiles of the mcrA gene amplicons highlighted that the ruminal methanogenic communities were composed of hydrogenotrophic methanogens dominated by the order Methanobacteriales regardless of the host species. The methanogenic communities changed significantly during the enrichment procedure, but still the strict hydrogenotrophic Methanobacteriales and Methanomicrobiales were the predominant orders in the enrichment cultures. The bioaugmentation potential of the enriched methanogenic cultures will be evaluated in further studies.

Culturable heterotrophic bacteria from the marine sponge Dendrilla nigra: isolation and phylogenetic diversity of actinobacteria

NASA Astrophysics Data System (ADS)

Selvin, Joseph; Gandhimathi, R.; Kiran, G. Seghal; Priya, S. Shanmugha; Ravji, T. Rajeetha; Hema, T. A.

2009-09-01

Culturable heterotrophic bacterial composition of marine sponge Dendrilla nigra was analysed using different enrichments. Five media compositions including without enrichment (control), enriched with sponge extract, with growth regulator (antibiotics), with autoinducers, and complete enrichment containing sponge extract, antibiotics, and autoinducers were developed. DNA hybridization assay was performed to explore host specific bacteria and ecotypes of culturable sponge-associated bacteria. Enrichment with selective inducers (AHLs and sponge extract) and regulators (antibiotics) considerably enhanced the cultivation potential of sponge-associated bacteria. It was found that Marinobacter (MSI032), Micromonospora (MSI033), Streptomyces (MSI051), and Pseudomonas (MSI057) were sponge-associated obligate symbionts. The present findings envisaged that “ Micromonospora-Saccharomonospora-Streptomyces” group was the major culturable actinobacteria in the marine sponge D. nigra. The DNA hybridization assay was a reliable method for the analysis of culturable bacterial community in marine sponges. Based on the culturable community structure, the sponge-associated bacteria can be grouped (ecotypes) as general symbionts, specific symbionts, habitat flora, and antagonists.

Direct detection of various pathogens by loop-mediated isothermal amplification assays on bacterial culture and bacterial colony.

PubMed

Yan, Muxia; Li, Weidong; Zhou, Zhenwen; Peng, Hongxia; Luo, Ziyan; Xu, Ling

2017-01-01

In this work, loop-mediated isothermal amplification based detection assay using bacterial culture and bacterial colony for various common pathogens direct detection had been established, evaluated and further applied. A total of five species of common pathogens and nine detection targets (tlh, tdh and trh for V. Parahaemolyticus, rfbE, stx1 and stx2 for E. coli, oprI for P. aeruginosa, invA for Salmonella and hylA for L. monocytogenes) were performed on bacterial culture and bacterial colony LAMP. To evaluate and optimize this assay, a total of 116 standard strains were included. Then, for each detected targets, 20 random selected strains were applied. Results were determined through both visual observation of the changed color by naked eye and electrophoresis, which increased the accuracy of survey. The minimum adding quantity of each primer had been confirmed, and the optimal amplification was obtained under 65 °C for 45 min with 25 μl reaction volume. The detection limit of bacterial culture LAMP and PCR assay were determined to be 10 2 and 10 4 or 10 5  CFU/reaction, respectively. No false positive amplification was observed when subjecting the bacterial -LAMP assay to 116 reference strains. This was the first report of colony-LAMP and culture-LAMP assay, which had been demonstrated to be a fast, reliable, cost-effective and simple method on detection of various common pathogens. Copyright © 2016 Elsevier Ltd. All rights reserved.

Bacterial diversity and reductive dehalogenase redundancy in a 1,2-dichloroethane-degrading bacterial consortium enriched from a contaminated aquifer

PubMed Central

2010-01-01

Background Bacteria possess a reservoir of metabolic functionalities ready to be exploited for multiple purposes. The use of microorganisms to clean up xenobiotics from polluted ecosystems (e.g. soil and water) represents an eco-sustainable and powerful alternative to traditional remediation processes. Recent developments in molecular-biology-based techniques have led to rapid and accurate strategies for monitoring and identification of bacteria and catabolic genes involved in the degradation of xenobiotics, key processes to follow up the activities in situ. Results We report the characterization of the response of an enriched bacterial community of a 1,2-dichloroethane (1,2-DCA) contaminated aquifer to the spiking with 5 mM lactate as electron donor in microcosm studies. After 15 days of incubation, the microbial community structure was analyzed. The bacterial 16S rRNA gene clone library showed that the most represented phylogenetic group within the consortium was affiliated with the phylum Firmicutes. Among them, known degraders of chlorinated compounds were identified. A reductive dehalogenase genes clone library showed that the community held four phylogenetically-distinct catalytic enzymes, all conserving signature residues previously shown to be linked to 1,2-DCA dehalogenation. Conclusions The overall data indicate that the enriched bacterial consortium shares the metabolic functionality between different members of the microbial community and is characterized by a high functional redundancy. These are fundamental features for the maintenance of the community's functionality, especially under stress conditions and suggest the feasibility of a bioremediation treatment with a potential prompt dehalogenation and a process stability over time. PMID:20170484

Anaerobic biotransformation of hexachlorocyclohexane isomers by Dehalococcoides species and an enrichment culture.

PubMed

Bashir, Safdar; Kuntze, Kevin; Vogt, Carsten; Nijenhuis, Ivonne

2018-06-18

The biotransformation of hexachlorocyclohexane isomers (HCH) by two Dehalococcoides mccartyi strains (195 and BTF08) and an enrichment culture was investigated and compared to conversion by the obligate anaerobic strain Clostridium pasteurianum strain DSMZ 525. The D. mccartyi strains preferentially transformed γ-HCH over α-HCH and δ-HCH isomers while β-HCH biotransformation was not significant. In case of the enrichment culture, γ-HCH was preferentially transformed over the δ-HCH, β-HCH and α-HCH isomers. Major observed metabolites in both cases were tetrachlorocyclohexene and as end products monochlorobenzene (MCB) and benzene. Dechlorination of the γ-HCH isomer was linked to an increase in cell numbers for strain 195. γ-HCH transformation was linked to considerable carbon stable isotope fractionation with the enrichment factor ε c  = - 5.5 ± 0.8‰ for D. mccartyi strain 195, ε c  = - 3.1 ± 0.4‰ for the enrichment culture and ε c  = - 4.1 ± 0.6‰ for co-metabolic transformation by C. pasteurianum.

Comparative evaluation of rRNA depletion procedures for the improved analysis of bacterial biofilm and mixed pathogen culture transcriptomes

PubMed Central

Petrova, Olga E.; Garcia-Alcalde, Fernando; Zampaloni, Claudia; Sauer, Karin

2017-01-01

Global transcriptomic analysis via RNA-seq is often hampered by the high abundance of ribosomal (r)RNA in bacterial cells. To remove rRNA and enrich coding sequences, subtractive hybridization procedures have become the approach of choice prior to RNA-seq, with their efficiency varying in a manner dependent on sample type and composition. Yet, despite an increasing number of RNA-seq studies, comparative evaluation of bacterial rRNA depletion methods has remained limited. Moreover, no such study has utilized RNA derived from bacterial biofilms, which have potentially higher rRNA:mRNA ratios and higher rRNA carryover during RNA-seq analysis. Presently, we evaluated the efficiency of three subtractive hybridization-based kits in depleting rRNA from samples derived from biofilm, as well as planktonic cells of the opportunistic human pathogen Pseudomonas aeruginosa. Our results indicated different rRNA removal efficiency for the three procedures, with the Ribo-Zero kit yielding the highest degree of rRNA depletion, which translated into enhanced enrichment of non-rRNA transcripts and increased depth of RNA-seq coverage. The results indicated that, in addition to improving RNA-seq sensitivity, efficient rRNA removal enhanced detection of low abundance transcripts via qPCR. Finally, we demonstrate that the Ribo-Zero kit also exhibited the highest efficiency when P. aeruginosa/Staphylococcus aureus co-culture RNA samples were tested. PMID:28117413

Environmental Selection, Dispersal, and Organism Interactions Shape Community Assembly in High-Throughput Enrichment Culturing

DOE PAGES

Justice, N. B.; Sczesnak, A.; Hazen, T. C.; ...

2017-08-04

A central goal of microbial ecology is to identify and quantify the forces that lead to observed population distributions and dynamics. However, these forces, which include environmental selection, dispersal, and organism interactions, are often difficult to assess in natural environments. Here in this paper, we present a method that links microbial community structures with selective and stochastic forces through highly replicated subsampling and enrichment of a single environmental inoculum. Specifically, groundwater from a well-studied natural aquifer was serially diluted and inoculated into nearly 1,000 aerobic and anaerobic nitrate-reducing cultures, and the final community structures were evaluated with 16S rRNA genemore » amplicon sequencing. We analyzed the frequency and abundance of individual operational taxonomic units (OTUs) to understand how probabilistic immigration, relative fitness differences, environmental factors, and organismal interactions contributed to divergent distributions of community structures. We further used a most probable number (MPN) method to estimate the natural condition-dependent cultivable abundance of each of the nearly 400 OTU cultivated in our study and infer the relative fitness of each. Additionally, we infer condition-specific organism interactions and discuss how this high-replicate culturing approach is essential in dissecting the interplay between overlapping ecological forces and taxon-specific attributes that underpin microbial community assembly. IMPORTANCEThrough highly replicated culturing, in which inocula are subsampled from a single environmental sample, we empirically determine how selective forces, interspecific interactions, relative fitness, and probabilistic dispersal shape bacterial communities. These methods offer a novel approach to untangle not only interspecific interactions but also taxon-specific fitness differences that manifest across different cultivation conditions and lead to the selection

Environmental Selection, Dispersal, and Organism Interactions Shape Community Assembly in High-Throughput Enrichment Culturing

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

Justice, N. B.; Sczesnak, A.; Hazen, T. C.

A central goal of microbial ecology is to identify and quantify the forces that lead to observed population distributions and dynamics. However, these forces, which include environmental selection, dispersal, and organism interactions, are often difficult to assess in natural environments. Here in this paper, we present a method that links microbial community structures with selective and stochastic forces through highly replicated subsampling and enrichment of a single environmental inoculum. Specifically, groundwater from a well-studied natural aquifer was serially diluted and inoculated into nearly 1,000 aerobic and anaerobic nitrate-reducing cultures, and the final community structures were evaluated with 16S rRNA genemore » amplicon sequencing. We analyzed the frequency and abundance of individual operational taxonomic units (OTUs) to understand how probabilistic immigration, relative fitness differences, environmental factors, and organismal interactions contributed to divergent distributions of community structures. We further used a most probable number (MPN) method to estimate the natural condition-dependent cultivable abundance of each of the nearly 400 OTU cultivated in our study and infer the relative fitness of each. Additionally, we infer condition-specific organism interactions and discuss how this high-replicate culturing approach is essential in dissecting the interplay between overlapping ecological forces and taxon-specific attributes that underpin microbial community assembly. IMPORTANCEThrough highly replicated culturing, in which inocula are subsampled from a single environmental sample, we empirically determine how selective forces, interspecific interactions, relative fitness, and probabilistic dispersal shape bacterial communities. These methods offer a novel approach to untangle not only interspecific interactions but also taxon-specific fitness differences that manifest across different cultivation conditions and lead to the selection

Demonstration of bacterial biofilms in culture-negative silicone stent and jones tube.

PubMed

Parsa, Kami; Schaudinn, Christoph; Gorur, Amita; Sedghizadeh, Parish P; Johnson, Thomas; Tse, David T; Costerton, John W

2010-01-01

To demonstrate the presence of bacterial biofilms on a dacryocystorhinostomy silicone stent and a Jones tube. One dacryocystorhinostomy silicone stent and one Jones tube were removed from 2 patients who presented with an infection of their respective nasolacrimal system. Cultures were obtained, and the implants were processed for scanning electron microscopy and confocal laser scanning microscopy, advanced microscopic methods that are applicable for detection of uncultivable biofilm organisms. Routine bacterial cultures revealed no growth, but bacterial biofilms on outer and inner surfaces of both implants were confirmed by advanced microscopic techniques. To the authors' knowledge, this is the first article that documents the presence of biofilms on a Crawford stent or a Jones tube on patients who presented with infections involving the nasolacrimal system. Although initial cultures revealed absence of any bacterial growth, confocal laser scanning microscopy and scanning electron microscopy documented bacterial colonization. Clinicians should consider the role of biofilms and the limitation of our standard culturing techniques while treating patients with device- or implant-related infections.

Persistence of pentolite (PETN and TNT) in soil microcosms and microbial enrichment cultures.

PubMed

Arbeli, Ziv; Garcia-Bonilla, Erika; Pardo, Cindy; Hidalgo, Kelly; Velásquez, Trigal; Peña, Luis; C, Eliana Ramos; Avila-Arias, Helena; Molano-Gonzalez, Nicolás; Brandão, Pedro F B; Roldan, Fabio

2016-05-01

Pentolite is a mixture (1:1) of 2,4,6-trinitrotoluene (TNT) and pentaerythritol tetranitrate (PETN), and little is known about its fate in the environment. This study was aimed to determine the dissipation of pentolite in soils under laboratory conditions. Microcosm experiments conducted with two soils demonstrated that dissipation rate of PETN was significantly slower than that of TNT. Interestingly, the dissipation of PETN was enhanced by the presence of TNT, while PETN did not enhanced the dissipation of TNT. Pentolite dissipation rate was significantly faster under biostimulation treatment (addition of carbon source) in soil from the artificial wetland, while no such stimulation was observed in soil from detonation field. In addition, the dissipation rate of TNT and PETN in soil from artificial wetland under biostimulation was significantly faster than the equivalent abiotic control, although it seems that non-biological processes might also be important for the dissipation of TNT and PETN. Transformation of PETN was also slower during establishment of enrichment culture using pentolite as the sole nitrogen source. In addition, transformation of these explosives was gradually reduced and practically stopped after the forth cultures transfer (80 days). DGGE analysis of bacterial communities from these cultures indicates that all consortia were dominated by bacteria from the order Burkholderiales and Rhodanobacter. In conclusion, our results suggest that PETN might be more persistent than TNT.

Cholera Rapid Test with Enrichment Step Has Diagnostic Performance Equivalent to Culture

PubMed Central

Ontweka, Lameck N.; Deng, Lul O.; Rauzier, Jean; Debes, Amanda K.; Tadesse, Fisseha; Parker, Lucy A.; Wamala, Joseph F.; Bior, Bior K.; Lasuba, Michael; But, Abiem Bona; Grandesso, Francesco; Jamet, Christine; Cohuet, Sandra; Ciglenecki, Iza; Serafini, Micaela; Sack, David A.; Quilici, Marie-Laure; Azman, Andrew S.; Luquero, Francisco J.

2016-01-01

Cholera rapid diagnostic tests (RDT) could play a central role in outbreak detection and surveillance in low-resource settings, but their modest performance has hindered their broad adoption. The addition of an enrichment step may improve test specificity. We describe the results of a prospective diagnostic evaluation of the Crystal VC RDT (Span Diagnostics, India) with enrichment step and of culture, each compared to polymerase chain reaction (PCR), during a cholera outbreak in South Sudan. RDTs were performed on alkaline peptone water inoculated with stool and incubated for 4–6 hours at ambient temperature. Cholera culture was performed from wet filter paper inoculated with stool. Molecular detection of Vibrio cholerae O1 by PCR was done from dry Whatman 903 filter papers inoculated with stool, and from wet filter paper supernatant. In August and September 2015, 101 consecutive suspected cholera cases were enrolled, of which 36 were confirmed by PCR. The enriched RDT had 86.1% (95% CI: 70.5–95.3) sensitivity and 100% (95% CI: 94.4–100) specificity compared to PCR as the reference standard. The sensitivity of culture versus PCR was 83.3% (95% CI: 67.2–93.6) for culture performed on site and 72.2% (95% CI: 54.8–85.8) at the international reference laboratory, where samples were tested after an average delay of two months after sample collection, and specificity was 98.5% (95% CI: 91.7–100) and 100% (95% CI: 94.5–100), respectively. The RDT with enrichment showed performance comparable to that of culture and could be a sustainable alternative to culture confirmation where laboratory capacity is limited. PMID:27992488

Effect of bioaugmentation by cellulolytic bacteria enriched from sheep rumen on methane production from wheat straw.

PubMed

Ozbayram, E Gozde; Kleinsteuber, Sabine; Nikolausz, Marcell; Ince, Bahar; Ince, Orhan

2017-08-01

The aim of this study was to determine the potential of bioaugmentation with cellulolytic rumen microbiota to enhance the anaerobic digestion of lignocellulosic feedstock. An anaerobic cellulolytic culture was enriched from sheep rumen fluid using wheat straw as substrate under mesophilic conditions. To investigate the effects of bioaugmentation on methane production from straw, the enrichment culture was added to batch reactors in proportions of 2% (Set-1) and 4% (Set-2) of the microbial cell number of the standard inoculum slurry. The methane production in the bioaugmented reactors was higher than in the control reactors. After 30 days of batch incubation, the average methane yield was 154 mL N CH 4 g VS -1 in the control reactors. Addition of 2% enrichment culture did not enhance methane production, whereas in Set-2 the methane yield was increased by 27%. The bacterial communities were examined by 454 amplicon sequencing of 16S rRNA genes, while terminal restriction fragment length polymorphism (T-RFLP) fingerprinting of mcrA genes was applied to analyze the methanogenic communities. The results highlighted that relative abundances of Ruminococcaceae and Lachnospiraceae increased during the enrichment. However, Cloacamonaceae, which were abundant in the standard inoculum, dominated the bacterial communities of all batch reactors. T-RFLP profiles revealed that Methanobacteriales were predominant in the rumen fluid, whereas the enrichment culture was dominated by Methanosarcinales. In the batch rectors, the most abundant methanogens were affiliated to Methanobacteriales and Methanomicrobiales. Our results suggest that bioaugmentation with sheep rumen enrichment cultures can enhance the performance of digesters treating lignocellulosic feedstock. Copyright © 2017 Elsevier Ltd. All rights reserved.

Characteristics of enriched cultures for bio-huff-`n`-puff tests at Jilin oil field

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

Xiu-Yuan Wang; Gang Dai; Yan-Fen Xue

1995-12-31

Three enriched cultures (48, 15a, and 26a), selected from more than 80 soil and water samples, could grow anaerobically in the presence of crude oil at 30{degrees}C and could ferment molasses to gases and organic acids. Oil recovery by culture 48 in the laboratory model experiment was enhanced by 25.2% over the original reserves and by 53.7% over the residual reserves. Enriched culture 48 was composed of at least 4 species belonging to the genera Eubacterium, Fusobacterium, and Bacteroides. This enriched culture was used as inoculum for MEOR field trials at Jilin oil field with satisfactory results. The importance ofmore » the role of these isolates in EOR was confirmed by their presence and behavior in the fluids produced from the microbiologically treated reservoir.« less

Improved enrichment culture technique for methane-oxidizing bacteria from marine ecosystems: the effect of adhesion material and gas composition.

PubMed

Vekeman, Bram; Dumolin, Charles; De Vos, Paul; Heylen, Kim

2017-02-01

Cultivation of microbial representatives of specific functional guilds from environmental samples depends largely on the suitability of the applied growth conditions. Especially the cultivation of marine methanotrophs has received little attention, resulting in only a limited number of ex situ cultures available. In this study we investigated the effect of adhesion material and headspace composition on the methane oxidation activity in methanotrophic enrichments obtained from marine sediment. Addition of sterilized natural sediment or alternatively the addition of acid-washed silicon dioxide significantly increased methane oxidation. This positive effect was attributed to bacterial adhesion on the particles via extracellular compounds, with a minimum amount of particles required for effect. As a result, the particles were immobilized, thus creating a stratified environment in which a limited diffusive gas gradients could build up and various microniches were formed. Such diffusive gas gradient might necessitate high headspace concentrations of CH 4 and CO 2 for sufficient concentrations to reach the methane-oxidizing bacteria in the enrichment culture technique. Therefore, high concentrations of methane and carbon dioxide, in addition to the addition of adhesion material, were tested and indeed further stimulated methane oxidation. Use of adhesion material in combination with high concentrations of methane and carbon dioxide might thus facilitate the cultivation and subsequent enrichment of environmentally important members of this functional guild. The exact mechanism of the observed positive effects on methane oxidation and the differential effect on methanotrophic diversity still needs to be explored.

Comparison of culture and PCR methods in the diagnosis of bacterial meningitis.

PubMed

Başpınar, Emel Ödemiş; Dayan, Saim; Bekçibaşı, Muhammed; Tekin, Recep; Ayaz, Celal; Deveci, Özcan; Hoşoğlu, Salih

Our aim in this study is to compare the standard culture method with the multiplex PCR and the Speed-Oligo ® Bacterial Meningitis Test (SO-BMT) - a hybridization-based molecular test method - during the CSF examination of the patients with the pre-diagnosis of acute bacterial meningitis. For the purposes of this study, patients with acute bacterial meningitis treated at the Dicle University Medical Faculty Hospital, Infectious Diseases and Clinical Microbiology Clinic between December 2009 and April 2012 were retrospectively evaluated. The diagnosis of bacterial meningitis was made based on the clinical findings, laboratory test anomalies, CSF analysis results, and the radiological images. Growth was observed in the CSF cultures of 10 out of the 57 patients included in the study (17.5%) and Streptococcus pneumoniae was isolated in all of them. The CSF samples of 34 patients (59.6%) were positive according to the SO-BMT and S. pneumoniae was detected in 33 of the samples (97.05%), while Neisseria meningitidis was found in 1 sample (2.95%). In a total of 10 patients, S. pneumoniae was both isolated in the CSF culture and detected in the SO-BMT. The culture and the SO-BMT were negative in 23 of the CSF samples. There was no sample in which the CSF culture was positive although the SO-BMT was negative. While SO-BMT seems to be a more efficient method than bacterial culturing to determine the pathogens that most commonly cause bacterial meningitis in adults, further studies conducted on larger populations are needed in order to assess its efficiency and uses. Copyright © 2016 Sociedade Brasileira de Microbiologia. Published by Elsevier Editora Ltda. All rights reserved.

Co-metabolic degradation of diazo dye- reactive blue 160 by enriched mixed cultures BDN.

PubMed

Balapure, Kshama H; Jain, Kunal; Chattaraj, Sananda; Bhatt, Nikhil S; Madamwar, Datta

2014-08-30

Mixed cultures BDN (BDN) proficient in decolourizing diazo dye-reactive blue 160 (RB160) consist of eight bacterial strains, was developed through culture enrichment method from soil samples contaminated with anthropogenic activities. The synthrophic interactions of BDN have led to complete decolourization and degradation of RB160 (100mg/L) within 4h along with co-metabolism of yeast extract (0.5%) in minimal medium. BDN microaerophilicaly decolourized even 1500mg/L of RB160 under high saline conditions (20g/L NaCl) at 37°C and pH 7.0. BDN exhibited broad substrate specificity and decolourized 27 structurally different dyes. The reductase enzymes symmetrically cleaved RB160 and oxidative enzymes further metabolised the degraded products and five different intermediates were identified using FTIR, (1)HNMR and GC-MS. The phytotoxicity assay confirmed that intact RB160 was more toxic than dye degraded intermediates. The BDN was able to colonize and decolourized RB160 in soil model system in presence of indigenous miocroflora as well as in sterile soil without any amendment of additional nutrients, which signifies it useful and potential application in bioremediation. Copyright © 2014 Elsevier B.V. All rights reserved.

Enrichment of denitrifying methane-oxidizing microorganisms using up-flow continuous reactors and batch cultures.

PubMed

Hatamoto, Masashi; Kimura, Masafumi; Sato, Takafumi; Koizumi, Masato; Takahashi, Masanobu; Kawakami, Shuji; Araki, Nobuo; Yamaguchi, Takashi

2014-01-01

Denitrifying anaerobic methane oxidizing (DAMO) microorganisms were enriched from paddy field soils using continuous-flow and batch cultures fed with nitrate or nitrite as a sole electron acceptor. After several months of cultivation, the continuous-flow cultures using nitrite showed remarkable simultaneous methane oxidation and nitrite reduction and DAMO bacteria belonging to phylum NC10 were enriched. A maximum volumetric nitrite consumption rate of 70.4±3.4 mg-N·L(-1)·day(-1) was achieved with very short hydraulic retention time of 2.1 hour. In the culture, about 68% of total microbial cells were bacteria and no archaeal cells were detected by fluorescence in situ hybridization. In the nitrate-fed continuous-flow cultures, 58% of total microbial cells were bacteria while archaeal cells accounted for 7% of total cell numbers. Phylogenetic analysis of pmoA gene sequence showed that enriched DAMO bacteria in the continuous-flow cultivation had over 98% sequence similarity to DAMO bacteria in the inoculum. In contrast, for batch culture, the enriched pmoA gene sequences had 89-91% sequence similarity to DAMO bacteria in the inoculum. These results indicate that electron acceptor and cultivation method strongly affect the microbial community structures of DAMO consortia.

Biodegradation of tributyl phosphate by novel bacteria isolated from enrichment cultures.

PubMed

Ahire, Kedar C; Kapadnis, Balu P; Kulkarni, Girish J; Shouche, Yogesh S; Deopurkar, Rajendra L

2012-02-01

Tributyl phosphate (TBP) is an organophosphorous compound, used extensively (3000-5000 tonnes/annum) as a solvent for nuclear fuel processing and as a base stock in the formulation of fire-resistant aircraft hydraulic fluids and other applications. Because of its wide applications and relative stability in the natural environment TBP poses the problem of pollution and health hazards. In the present study, fifteen potent bacterial strains capable of using tributyl phosphate (TBP) as sole carbon and phosphorus source were isolated from enrichment cultures. These isolates were identified on the basis of biochemical and morphological characteristics and 16S rRNA gene sequence analysis. Phylogenetic analysis of 16S rRNA gene sequences revealed that two isolates belonged to class Bacilli and thirteen to β and γ-Proteobacteria. All these isolates were found to be members of genera Alcaligenes, Providencia, Delftia, Ralstonia, and Bacillus. These isolates were able to tolerate and degrade up to 5 mM TBP, the highest concentration reported to date. The GC-MS method was developed to monitor TBP degradation. Two strains, Providencia sp. BGW4 and Delftia sp. BGW1 showed respectively, 61.0 ± 2.8% and 57.0 ± 2.0% TBP degradation within 4 days. The degradation rate constants, calculated by first order kinetic model were between 0.0024 and 0.0099 h(-1). These bacterial strains are novel for TBP degradation and could be used as an important bioresource for efficient decontamination of TBP polluted waste streams.

Detection of Salmonella invA gene in shrimp enrichment culture by polymerase chain reaction.

PubMed

Upadhyay, Bishnu Prasad; Utrarachkij, Fuangfa; Thongshoob, Jarinee; Mahakunkijcharoen, Yuvadee; Wongchinda, Niracha; Suthienkul, Orasa; Khusmith, Srisin

2010-03-01

Contamination of seafood with salmonellae is a major public health concern. Detection of Salmonella by standard culture methods is time consuming. In this study, an enrichment culture step prior to polymerase chain reaction (PCR) was applied to detect 284 bp fragment of Salmonella invA in comparison with the conventional culture method in 100 shrimp samples collected from four different shrimp farms and fresh food markets around Bangkok. Samples were pre-enriched in non-selective lactose broth (LB) and selective tetrathionate broth (TTB). PCR detection limit was 10 pg and 10(4) cfu/ml of viable salmonellae with 100% specificity. PCR assay detected 19 different Salmonella serovars belonging to 8 serogroups (B, C1, C2-C3, D1, E1, E4 and K) commonly found in clinical and environmental samples in Thailand. The detection rate of PCR following TTB enrichment (24%) was higher than conventional culture method (19%). PCR following TTB, but not in LB enrichment allowed salmonella detection with 84% sensitivity, 90% specificity and 89% accuracy. Shrimp samples collected from fresh food markets had higher levels of contaminated salmonellae than those from shrimp farms. The results indicated that incorporation of an enrichment step prior to PCR has the potential to be applied for detection of naturally contaminated salmonellae in food, environment and clinical samples.

Bacterial growth and substrate degradation by BTX-oxidizing culture in response to salt stress.

PubMed

Lee, Chi-Yuan; Lin, Ching-Hsing

2006-01-01

Interactions between microbial growth and substrate degradation are important in determining the performance of trickle-bed bioreactors (TBB), especially when salt is added to reduce biomass formation in order to alleviate media clogging. This study was aimed at quantifying salinity effects on bacterial growth and substrate degradation, and at acquiring kinetic information in order to improve the design and operation of TBB. Experiment works began by cultivating a mixed culture in a chemostat reactor receiving artificial influent containing a mixture of benzene, toluene, and xylene (BTX), followed by using the enrichment culture to degrade the individual BTX substrates under a particular salinity, which ranged 0-50 g l(-1) in batch mode. Then, the measured concentrations of biomass and residual substrate versus time were analyzed with the microbial kinetics; moreover, the obtained microbial kinetic constants under various salinities were modeled using noncompetitive inhibition kinetics. For the three substrates the observed bacterial yields appeared to be decreased from 0.51-0.74 to 0.20-0.22 mg mg(-1) and the maximum specific rate of substrate utilization, q, declined from 0.25-0.42 to 0.07-0.11 h(-1), as the salinity increased from 0 to 50 NaCl g l(-1). The NaCl acted as noncompetitive inhibitor, where the modeling inhibitions of the coefficients, K ( T(S)), were 22.7-29.7 g l(-1) for substrate degradation and K ( T(mu)), 13.0-19.0 g l(-1), for biomass formation. The calculated ratios for the bacterial maintenance rate, m (S), to q, further indicated that the percentage energy spent on maintenance increased from 19-24 to 86-91% as salinity level increased from 0 to 50 g l(-1). These results revealed that the bacterial growth was more inhibited than substrate degradation by the BTX oxidizers under the tested salinity levels. The findings from this study demonstrate the potential of applying NaCl salt to control excessive biomass formation in biotrickling filters.

Successful enrichment of the ubiquitous freshwater acI Actinobacteria.

PubMed

Garcia, Sarahi L; McMahon, Katherine D; Grossart, Hans-Peter; Warnecke, Falk

2014-02-01

Actinobacteria of the acI lineage are often the numerically dominant bacterial phylum in surface freshwaters, where they can account for > 50% of total bacteria. Despite their abundance, there are no described isolates. In an effort to obtain enrichment of these ubiquitous freshwater Actinobacteria, diluted freshwater samples from Lake Grosse Fuchskuhle, Germany, were incubated in 96-well culture plates. With this method, a successful enrichment containing high abundances of a member of the lineage acI was established. Phylogenetic classification showed that the acI Actinobacteria of the enrichment belonged to the acI-B2 tribe, which seems to prefer acidic lakes. This enrichment grows to low cell densities and thus the oligotrophic nature of acI-B2 was confirmed. © 2013 Society for Applied Microbiology and John Wiley & Sons Ltd.

Current and Past Strategies for Bacterial Culture in Clinical Microbiology

PubMed Central

Lagier, Jean-Christophe; Edouard, Sophie; Pagnier, Isabelle; Mediannikov, Oleg; Drancourt, Michel

2015-01-01

SUMMARY A pure bacterial culture remains essential for the study of its virulence, its antibiotic susceptibility, and its genome sequence in order to facilitate the understanding and treatment of caused diseases. The first culture conditions empirically varied incubation time, nutrients, atmosphere, and temperature; culture was then gradually abandoned in favor of molecular methods. The rebirth of culture in clinical microbiology was prompted by microbiologists specializing in intracellular bacteria. The shell vial procedure allowed the culture of new species of Rickettsia. The design of axenic media for growing fastidious bacteria such as Tropheryma whipplei and Coxiella burnetii and the ability of amoebal coculture to discover new bacteria constituted major advances. Strong efforts associating optimized culture media, detection methods, and a microaerophilic atmosphere allowed a dramatic decrease of the time of Mycobacterium tuberculosis culture. The use of a new versatile medium allowed an extension of the repertoire of archaea. Finally, to optimize the culture of anaerobes in routine bacteriology laboratories, the addition of antioxidants in culture media under an aerobic atmosphere allowed the growth of strictly anaerobic species. Nevertheless, among usual bacterial pathogens, the development of axenic media for the culture of Treponema pallidum or Mycobacterium leprae remains an important challenge that the patience and innovations of cultivators will enable them to overcome. PMID:25567228

Culture dependent and independent analysis of bacterial communities associated with commercial salad leaf vegetables.

PubMed

Jackson, Colin R; Randolph, Kevin C; Osborn, Shelly L; Tyler, Heather L

2013-12-01

Plants harbor a diverse bacterial community, both as epiphytes on the plant surface and as endophytes within plant tissue. While some plant-associated bacteria act as plant pathogens or promote plant growth, others may be human pathogens. The aim of the current study was to determine the bacterial community composition of organic and conventionally grown leafy salad vegetables at the point of consumption using both culture-dependent and culture-independent methods. Total culturable bacteria on salad vegetables ranged from 8.0 × 10(3) to 5.5 × 10(8) CFU g(-1). The number of culturable endophytic bacteria from surface sterilized plants was significantly lower, ranging from 2.2 × 10(3) to 5.8 × 10(5) CFU g(-1). Cultured isolates belonged to six major bacterial phyla, and included representatives of Pseudomonas, Pantoea, Chryseobacterium, and Flavobacterium. Eleven different phyla and subphyla were identified by culture-independent pyrosequencing, with Gammaproteobacteria, Betaproteobacteria, and Bacteroidetes being the most dominant lineages. Other bacterial lineages identified (e.g. Firmicutes, Alphaproteobacteria, Acidobacteria, and Actinobacteria) typically represented less than 1% of sequences obtained. At the genus level, sequences classified as Pseudomonas were identified in all samples and this was often the most prevalent genus. Ralstonia sequences made up a greater portion of the community in surface sterilized than non-surface sterilized samples, indicating that it was largely endophytic, while Acinetobacter sequences appeared to be primarily associated with the leaf surface. Analysis of molecular variance indicated there were no significant differences in bacterial community composition between organic versus conventionally grown, or surface-sterilized versus non-sterilized leaf vegetables. While culture-independent pyrosequencing identified significantly more bacterial taxa, the dominant taxa from pyrosequence data were also detected by traditional

Culture dependent and independent analysis of bacterial communities associated with commercial salad leaf vegetables

PubMed Central

2013-01-01

Background Plants harbor a diverse bacterial community, both as epiphytes on the plant surface and as endophytes within plant tissue. While some plant-associated bacteria act as plant pathogens or promote plant growth, others may be human pathogens. The aim of the current study was to determine the bacterial community composition of organic and conventionally grown leafy salad vegetables at the point of consumption using both culture-dependent and culture-independent methods. Results Total culturable bacteria on salad vegetables ranged from 8.0 × 103 to 5.5 × 108 CFU g-1. The number of culturable endophytic bacteria from surface sterilized plants was significantly lower, ranging from 2.2 × 103 to 5.8 × 105 CFU g-1. Cultured isolates belonged to six major bacterial phyla, and included representatives of Pseudomonas, Pantoea, Chryseobacterium, and Flavobacterium. Eleven different phyla and subphyla were identified by culture-independent pyrosequencing, with Gammaproteobacteria, Betaproteobacteria, and Bacteroidetes being the most dominant lineages. Other bacterial lineages identified (e.g. Firmicutes, Alphaproteobacteria, Acidobacteria, and Actinobacteria) typically represented less than 1% of sequences obtained. At the genus level, sequences classified as Pseudomonas were identified in all samples and this was often the most prevalent genus. Ralstonia sequences made up a greater portion of the community in surface sterilized than non-surface sterilized samples, indicating that it was largely endophytic, while Acinetobacter sequences appeared to be primarily associated with the leaf surface. Analysis of molecular variance indicated there were no significant differences in bacterial community composition between organic versus conventionally grown, or surface-sterilized versus non-sterilized leaf vegetables. While culture-independent pyrosequencing identified significantly more bacterial taxa, the dominant taxa from pyrosequence data were also detected by

Enrichment of skin-derived neural precursor cells from dermal cell populations by altering culture conditions.

PubMed

Bayati, Vahid; Gazor, Rohoullah; Nejatbakhsh, Reza; Negad Dehbashi, Fereshteh

2016-01-01

As stem cells play a critical role in tissue repair, their manipulation for being applied in regenerative medicine is of great importance. Skin-derived precursors (SKPs) may be good candidates for use in cell-based therapy as the only neural stem cells which can be isolated from an accessible tissue, skin. Herein, we presented a simple protocol to enrich neural SKPs by monolayer adherent cultivation to prove the efficacy of this method. To enrich neural SKPs from dermal cell populations, we have found that a monolayer adherent cultivation helps to increase the numbers of neural precursor cells. Indeed, we have cultured dermal cells as monolayer under serum-supplemented (control) and serum-supplemented culture, followed by serum free cultivation (test) and compared. Finally, protein markers of SKPs were assessed and compared in both experimental groups and differentiation potential was evaluated in enriched culture. The cells of enriched culture concurrently expressed fibronectin, vimentin and nestin, an intermediate filament protein expressed in neural and skeletal muscle precursors as compared to control culture. In addition, they possessed a multipotential capacity to differentiate into neurogenic, glial, adipogenic, osteogenic and skeletal myogenic cell lineages. It was concluded that serum-free adherent culture reinforced by growth factors have been shown to be effective on proliferation of skin-derived neural precursor cells (skin-NPCs) and drive their selective and rapid expansion.

Anti-protozoal and anti-bacterial antibiotics that inhibit protein synthesis kill cancer subtypes enriched for stem cell-like properties.

PubMed

Cuyàs, Elisabet; Martin-Castillo, Begoña; Corominas-Faja, Bruna; Massaguer, Anna; Bosch-Barrera, Joaquim; Menendez, Javier A

2015-01-01

Key players in translational regulation such as ribosomes might represent powerful, but hitherto largely unexplored, targets to eliminate drug-refractory cancer stem cells (CSCs). A recent study by the Lisanti group has documented how puromycin, an old antibiotic derived from Streptomyces alboniger that inhibits ribosomal protein translation, can efficiently suppress CSC states in tumorspheres and monolayer cultures. We have used a closely related approach based on Biolog Phenotype Microarrays (PM), which contain tens of lyophilized antimicrobial drugs, to assess the chemosensitivity profiles of breast cancer cell lines enriched for stem cell-like properties. Antibiotics directly targeting active sites of the ribosome including emetine, puromycin and cycloheximide, inhibitors of ribosome biogenesis such as dactinomycin, ribotoxic stress agents such as daunorubicin, and indirect inhibitors of protein synthesis such as acriflavine, had the largest cytotoxic impact against claudin-low and basal-like breast cancer cells. Thus, biologically aggressive, treatment-resistant breast cancer subtypes enriched for stem cell-like properties exhibit exacerbated chemosensitivities to anti-protozoal and anti-bacterial antibiotics targeting protein synthesis. These results suggest that old/existing microbicides might be repurposed not only as new cancer therapeutics, but also might provide the tools and molecular understanding needed to develop second-generation inhibitors of ribosomal translation to eradicate CSC traits in tumor tissues.

Anti-protozoal and anti-bacterial antibiotics that inhibit protein synthesis kill cancer subtypes enriched for stem cell-like properties

PubMed Central

Cuyàs, Elisabet; Martin-Castillo, Begoña; Corominas-Faja, Bruna; Massaguer, Anna; Bosch-Barrera, Joaquim; Menendez, Javier A

2015-01-01

Key players in translational regulation such as ribosomes might represent powerful, but hitherto largely unexplored, targets to eliminate drug-refractory cancer stem cells (CSCs). A recent study by the Lisanti group has documented how puromycin, an old antibiotic derived from Streptomyces alboniger that inhibits ribosomal protein translation, can efficiently suppress CSC states in tumorspheres and monolayer cultures. We have used a closely related approach based on Biolog Phenotype Microarrays (PM), which contain tens of lyophilized antimicrobial drugs, to assess the chemosensitivity profiles of breast cancer cell lines enriched for stem cell-like properties. Antibiotics directly targeting active sites of the ribosome including emetine, puromycin and cycloheximide, inhibitors of ribosome biogenesis such as dactinomycin, ribotoxic stress agents such as daunorubicin, and indirect inhibitors of protein synthesis such as acriflavine, had the largest cytotoxic impact against claudin-low and basal-like breast cancer cells. Thus, biologically aggressive, treatment-resistant breast cancer subtypes enriched for stem cell-like properties exhibit exacerbated chemosensitivities to anti-protozoal and anti-bacterial antibiotics targeting protein synthesis. These results suggest that old/existing microbicides might be repurposed not only as new cancer therapeutics, but also might provide the tools and molecular understanding needed to develop second-generation inhibitors of ribosomal translation to eradicate CSC traits in tumor tissues. PMID:25970790

Bacterial diversity of autotrophic enriched cultures from remote, glacial Antarctic, Alpine and Andean aerosol, snow and soil samples

NASA Astrophysics Data System (ADS)

González-Toril, E.; Amils, R.; Delmas, R. J.; Petit, J.-R.; Komárek, J.; Elster, J.

2009-01-01

Four different communities and one culture of autotrophic microbial assemblages were obtained by incubation of samples collected from high elevation snow in the Alps (Mt. Blanc area) and the Andes (Nevado Illimani summit, Bolivia), from Antarctic aerosol (French station Dumont d'Urville) and a maritime Antarctic soil (King George Island, South Shetlands, Uruguay Station Artigas), in a minimal mineral (oligotrophic) media. Molecular analysis of more than 200 16S rRNA gene sequences showed that all cultured cells belong to the Bacteria domain. Phylogenetic comparison with the currently available rDNA database allowed sequences belonging to Proteobacteria Alpha-, Beta- and Gamma-proteobacteria), Actinobacteria and Bacteroidetes phyla to be identified. The Andes snow culture was the richest in bacterial diversity (eight microorganisms identified) and the marine Antarctic soil the poorest (only one). Snow samples from Col du Midi (Alps) and the Andes shared the highest number of identified microorganisms (Agrobacterium, Limnobacter, Aquiflexus and two uncultured Alphaproteobacteria clones). These two sampling sites also shared four sequences with the Antarctic aerosol sample (Limnobacter, Pseudonocardia and an uncultured Alphaproteobacteriaclone). The only microorganism identified in the Antarctica soil (Brevundimonas sp.) was also detected in the Antarctic aerosol. Most of the identified microorganisms had been detected previously in cold environments, marine sediments soils and rocks. Air current dispersal is the best model to explain the presence of very specific microorganisms, like those identified in this work, in environments very distant and very different from each other.

Effect of red clay on diesel bioremediation and soil bacterial community.

PubMed

Jung, Jaejoon; Choi, Sungjong; Hong, Hyerim; Sung, Jung-Suk; Park, Woojun

2014-08-01

Red clay is a type of soil, the red color of which results from the presence of iron oxide. It is considered an eco-friendly material, with many industrial, cosmetic, and architectural uses. A patented method was applied to red clay in order to change its chemical composition and mineral bioavailability. The resulting product was designated processed red clay. This study evaluates the novel use of red clay and processed red clay as biostimulation agents in diesel-contaminated soils. Diesel biodegradation was enhanced in the presence of red clay and processed red clay by 4.9- and 6.7-fold, respectively, and the number of culturable bacterial cells was correlated with the amount of diesel biodegradation. The growth of Acinetobacter oleivorans DR1, Pseudomonas putida KT2440, and Cupriavidus necator was promoted by both types of red clays. Culture-independent community analysis determined via barcoded pyrosequencing indicated that Nocardioidaceae, Xanthomonadaceae, Pseudomonadaceae, and Caulobacteraceae were enriched by diesel contamination. Bacterial strain isolation from naphthalene- and liquid paraffin-amended media was affiliated with enriched taxa based on 16S rRNA gene sequence identity. We suggest that the biostimulating mechanism of red clay and processed red clay is able to support bacterial growth without apparent selection for specific bacterial species.

Current and past strategies for bacterial culture in clinical microbiology.

PubMed

Lagier, Jean-Christophe; Edouard, Sophie; Pagnier, Isabelle; Mediannikov, Oleg; Drancourt, Michel; Raoult, Didier

2015-01-01

A pure bacterial culture remains essential for the study of its virulence, its antibiotic susceptibility, and its genome sequence in order to facilitate the understanding and treatment of caused diseases. The first culture conditions empirically varied incubation time, nutrients, atmosphere, and temperature; culture was then gradually abandoned in favor of molecular methods. The rebirth of culture in clinical microbiology was prompted by microbiologists specializing in intracellular bacteria. The shell vial procedure allowed the culture of new species of Rickettsia. The design of axenic media for growing fastidious bacteria such as Tropheryma whipplei and Coxiella burnetii and the ability of amoebal coculture to discover new bacteria constituted major advances. Strong efforts associating optimized culture media, detection methods, and a microaerophilic atmosphere allowed a dramatic decrease of the time of Mycobacterium tuberculosis culture. The use of a new versatile medium allowed an extension of the repertoire of archaea. Finally, to optimize the culture of anaerobes in routine bacteriology laboratories, the addition of antioxidants in culture media under an aerobic atmosphere allowed the growth of strictly anaerobic species. Nevertheless, among usual bacterial pathogens, the development of axenic media for the culture of Treponema pallidum or Mycobacterium leprae remains an important challenge that the patience and innovations of cultivators will enable them to overcome. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Biotransformation of acyclovir by an enriched nitrifying culture.

PubMed

Xu, Yifeng; Yuan, Zhiguo; Ni, Bing-Jie

2017-03-01

This work evaluates the biodegradation of the antiviral drug acyclovir by an enriched nitrifying culture during ammonia oxidation and without the addition of ammonium. The study on kinetics was accompanied with the structural elucidation of biotransformation products through batch biodegradation experiments at two different initial levels of acyclovir (15 mg L -1 and 15 μg L -1 ). The pseudo first order kinetic studies of acyclovir in the presence of ammonium indicated the higher degradation rates under higher ammonia oxidation rates than those constant degradation rates in the absence of ammonium. The positive correlation was found between acyclovir degradation rate and ammonia oxidation rate, confirming the cometabolism of acyclovir by the enriched nitrifying culture in the presence of ammonium. Formation of the product carboxy-acyclovir (P239) indicated the main biotransformation pathway was aerobic oxidation of the terminal hydroxyl group, which was independent on the metabolic type (i.e. cometabolism or metabolism). This enzyme-linked reaction might be catalyzed by monooxygenase from ammonia oxidizing bacteria or heterotrophs. The formation of carboxy-acyclovir was demonstrated to be irrelevant to the acyclovir concentrations applied, indicating the revealed biotransformation pathway might be the dominant removal pathway of acyclovir in wastewater treatment. Copyright © 2016 Elsevier Ltd. All rights reserved.

Effect of dissolved oxygen concentration (microaerobic and aerobic) on selective enrichment culture for bioaugmentation of acidic industrial wastewater.

PubMed

Quan, Ying; Han, Hui; Zheng, Shaokui

2012-09-01

The successful application of bioaugmentation is largely dependent on the selective enrichment of culture with regards to pH, temperature, salt, or specific toxic organic pollutants. In this study, we investigated the effect of dissolved oxygen (DO) concentrations (aerobic, >2 mg L(-1); microaerobic, <1 mg L(-1)) on yeast enrichment culture for bioaugmentation of acidic industrial wastewater (pH 3.9-4.7). Clone library analyses revealed that the yeast community shifted in response to different DO levels, and that Candida humilis and Candida pseudolambica were individually dominant in the aerobic and microaerobic enrichment cultures. This would significantly influence the isolation results, and further hinder bioaugmentation due to differences in DO environments during the enrichment and application periods. However, differences in the selective enrichment culture cannot be predicted based on differences in pollutant removal performance. Thus, DO concentrations (aerobic/microaerobic) should be considered a secondary selective pressure to achieve successful bioaugmentation. Copyright © 2012 Elsevier Ltd. All rights reserved.

Study of bacterial meningitis in children below 5 years with comparative evaluation of gram staining, culture and bacterial antigen detection.

PubMed

Yadhav Ml, Kala

2014-04-01

Bacterial meningitis is one of the most serious infections seen in infants and children, which is associated with acute complications and chronic morbidity. Infections of Central Nervous System (CNS) still dominate the scene of childhood neurological disorders in most of the developing tropical countries. To isolate, identify and determine the antibiotic susceptibility patterns of pathogens associated with bacterial meningitis. We also aimed to comparatively evaluate of Gram staining, culture and bacterial antigen detection in cerebrospinal fluid samples. Present comparative study included 100 CSF samples of children below the age of 5 years, who were clinically suspected meningitis cases. The samples were subjected to Gram staining, culture and Latex agglutination test (LAT). The organisms isolated in the study were characterized and antibiotic susceptibility test was done according to standard guidelines. It was done by using Gaussian test. Of the 100 cases, 24 were diagnosed as Acute bacterial meningitis (ABM) cases by. Gram staining, culture and latex agglutination test. 21 (87.5%) cases were culture positive, with 2 cases being positive for polymicrobial isolates. Gram staining was positive in 17 (70.53%) cases and LAT was positive in 18 (33.33%) cases. Streptococcus pneumoniae was the predominant organism which was isolated and it was sensitive to antibiotics. In the present study, male to female ratio was 1.27:1, which showed a male preponderance. With the combination of Gram staining, culture, and LAT, 100% sensitivity and specificity can be achieved (p < 0.001). Gram staining and LAT can detect 85% of cases of ABM. Bacterial meningitis is a medical emergency and making an early diagnosis and providing treatment early are life saving and they reduce chronic morbidity.

Identification of Uncultured Bacterial Species from Firmicutes, Bacteroidetes and CANDIDATUS Saccharibacteria as Candidate Cellulose Utilizers from the Rumen of Beef Cows

PubMed Central

Opdahl, Lee James; Gonda, Michael G.

2018-01-01

The ability of ruminants to utilize cellulosic biomass is a result of the metabolic activities of symbiotic microbial communities that reside in the rumen. To gain further insight into this complex microbial ecosystem, a selection-based batch culturing approach was used to identify candidate cellulose-utilizing bacterial consortia. Prior to culturing with cellulose, rumen contents sampled from three beef cows maintained on a forage diet shared 252 Operational Taxonomic Units (OTUs), accounting for 41.6–50.0% of bacterial 16S rRNA gene sequences in their respective samples. Despite this high level of overlap, only one OTU was enriched in cellulose-supplemented cultures from all rumen samples. Otherwise, each set of replicate cellulose supplemented cultures originating from a sampled rumen environment was found to have a distinct bacterial composition. Two of the seven most enriched OTUs were closely matched to well-established rumen cellulose utilizers (Ruminococcus flavefaciens and Fibrobacter succinogenes), while the others did not show high nucleotide sequence identity to currently defined bacterial species. The latter were affiliated to Prevotella (1 OTU), Ruminococcaceae (3 OTUs), and the candidate phylum Saccharibacteria (1 OTU), respectively. While further investigations will be necessary to elucidate the metabolic function(s) of each enriched OTU, these results together further support cellulose utilization as a ruminal metabolic trait shared across vast phylogenetic distances, and that the rumen is an environment conducive to the selection of a broad range of microbial adaptations for the digestion of plant structural polysaccharides. PMID:29495256

Identification of Uncultured Bacterial Species from Firmicutes, Bacteroidetes and CANDIDATUS Saccharibacteria as Candidate Cellulose Utilizers from the Rumen of Beef Cows.

PubMed

Opdahl, Lee James; Gonda, Michael G; St-Pierre, Benoit

2018-02-24

The ability of ruminants to utilize cellulosic biomass is a result of the metabolic activities of symbiotic microbial communities that reside in the rumen. To gain further insight into this complex microbial ecosystem, a selection-based batch culturing approach was used to identify candidate cellulose-utilizing bacterial consortia. Prior to culturing with cellulose, rumen contents sampled from three beef cows maintained on a forage diet shared 252 Operational Taxonomic Units (OTUs), accounting for 41.6-50.0% of bacterial 16S rRNA gene sequences in their respective samples. Despite this high level of overlap, only one OTU was enriched in cellulose-supplemented cultures from all rumen samples. Otherwise, each set of replicate cellulose supplemented cultures originating from a sampled rumen environment was found to have a distinct bacterial composition. Two of the seven most enriched OTUs were closely matched to well-established rumen cellulose utilizers ( Ruminococcus flavefaciens and Fibrobacter succinogenes ), while the others did not show high nucleotide sequence identity to currently defined bacterial species. The latter were affiliated to Prevotella (1 OTU), Ruminococcaceae (3 OTUs), and the candidate phylum Saccharibacteria (1 OTU), respectively. While further investigations will be necessary to elucidate the metabolic function(s) of each enriched OTU, these results together further support cellulose utilization as a ruminal metabolic trait shared across vast phylogenetic distances, and that the rumen is an environment conducive to the selection of a broad range of microbial adaptations for the digestion of plant structural polysaccharides.

Association between Gallbladder Ultrasound Findings and Bacterial Culture of Bile in 70 Cats and 202 Dogs.

PubMed

Policelli Smith, R; Gookin, J L; Smolski, W; Di Cicco, M F; Correa, M; Seiler, G S

2017-09-01

Bacterial cholecystitis often is diagnosed by combination of gallbladder ultrasound (US) findings and positive results of bile culture. The value of gallbladder US in determining the likelihood of bile bacterial infection in cats and dogs with suspected biliary disease is unknown. To determine the value of gallbladder US in predicting bile bacterial culture results, identify most common bacterial isolates from bile, and describe complications after cholecystocentesis in cats and dogs with suspected hepatobiliary disease. Cats (70) and dogs (202) that underwent an abdominal US and submission of bile for culture were included in the study. A cross-sectional study design was used to determine the association of gallbladder US abnormalities and the results of bile cultures, and complications of cholecystocentesis. Abnormal gallbladder US had high sensitivity (96%) but low specificity (49%) in cats with positive and negative results of bile bacterial culture, respectively. Cats with normal gallbladder US findings were unlikely to have positive bile bacterial culture (negative predictive value of 96%). Gallbladder US had lower sensitivity (81%), specificity (31%), positive predictive value (20%), and negative predictive value (88%) in dogs. The most common bacterial isolates were of enteric origin, the prevalence being higher in cats. Incidence of complications after cholecystocentesis was 3.4%. Gallbladder US has a high negative predictive value for bile culture results in cats. This modality is less predictive of infection in dogs. Percutaneous US-guided cholecystocentesis has a low complication rate. Copyright © 2017 The Authors. Journal of Veterinary Internal Medicine published by Wiley Periodicals, Inc. on behalf of the American College of Veterinary Internal Medicine.

Marine bacterial degradation of brominated methanes

USGS Publications Warehouse

Goodwin, K.D.; Lidstrom, M.E.; Oremland, R.S.

1997-01-01

Brominated methanes are ozone-depleting compounds whose natural sources include marine algae such as kelp. Brominated methane degradation by bacteria was investigated to address whether bacterial processes might effect net emission of these compounds to the atmosphere. Bacteria in seawater collected from California kelp beds degraded CH2Br2 but not CHBr3. Specific inhibitors showed that methanotrophs and nitrifiers did not significantly contribute to CH2Br2 removal. A seawater enrichment culture oxidized 14CH2Br2 to 14CO2 as well as 14CH3Br to 14CO2. The rates of CH2Br2 degradation in laboratory experiments suggest that bacterial degradation of CH2Br2 in a kelp bed accounts for <1% of the CH2Br2 produced by the kelp. However, the half-life of CH2Br2 due to bacterial removal appears faster than hydrolysis and within an order of magnitude of volatilization to the atmosphere.Brominated methanes are ozone-depleting compounds whose natural sources include marine algae such as kelp. Brominated methane degradation by bacteria was investigated to address whether bacterial processes might effect net emission of these compounds to the atmosphere. Bacteria in seawater collected from California kelp beds degraded CH2Br2 but not CHBr3. Specific inhibitors showed that methanotrophs and nitrifiers did not significantly contribute to CH2Br2 removal. A seawater enrichment culture oxidized 14CH2Br2 to 14CO2 as well as 14CH3Br to 14CO2. The rates of CH2Br2 degradation in laboratory experiments suggest that bacterial degradation of CH2Br2 in a kelp bed accounts for <1% of the CH2Br2 produced by the kelp. However, the half-life of CH2Br2 due to bacterial removal appears faster than hydrolysis and within an order of magnitude of volatilization to the atmosphere.

Enrichments of methanotrophic-heterotrophic cultures with high poly-β-hydroxybutyrate (PHB) accumulation capacities.

PubMed

Zhang, Tingting; Wang, Xiaowei; Zhou, Jiti; Zhang, Yu

2018-03-01

Methanotrophic-heterotrophic communities were selectively enriched from sewage sludge to obtain a mixed culture with high levels of poly-β-hydroxybutyrate (PHB) accumulation capacity from methane. Methane was used as the carbon source, N 2 as sole nitrogen source, and oxygen and Cu content were varied. Copper proved essential for PHB synthesis. All cultures enriched with Cu could accumulate high content of PHB (43.2%-45.9%), while only small amounts of PHB were accumulated by cultures enriched without Cu (11.9%-17.5%). Batch assays revealed that communities grown with Cu and a higher O 2 content synthesized more PHB, which had a wider optimal CH 4 :O 2 range and produced a high PHB content (48.7%) even though in the presence of N 2 . In all methanotrophic-heterotrophic communities, both methanotrophic and heterotrophic populations showed the ability to accumulate PHB. Although methane was added as the sole carbon source, heterotrophs dominated with abundances between 77.2% and 85.6%. All methanotrophs detected belonged to type II genera, which formed stable communities with heterotrophs of different PHB production capacities. Copyright © 2017. Published by Elsevier B.V.

Electrical response of culture media during bacterial growth on a paper-based device

NASA Astrophysics Data System (ADS)

Srimongkon, Tithimanan; Buerkle, Marius; Nakamura, Akira; Enomae, Toshiharu; Ushijima, Hirobumi; Fukuda, Nobuko

2017-05-01

In this work, we evaluated the feasibility of a paper-based bacterial detection system. The paper served as a substrate for the measurement electrodes and the culture medium. Using a printing technique, we patterned gold electrodes onto the paper substrate and applied Luria broth (LB) agar gel as a culture medium on top of the electrodes. As the first step towards the development of a bacterial detection system, we determined changes in the surface potential during bacterial growth and monitored these changes over 24 h. This allowed us to correlate changes in the surface potential with the different growth phases of the bacteria.

Bacterial diversity associated with the rotifer Brachionus plicatilis sp. complex determined by culture-dependent and -independent methods.

PubMed

Ishino, Ryota; Iehata, Shunpei; Nakano, Miyo; Tanaka, Reiji; Yoshimatsu, Takao; Maeda, Hiroto

2012-03-01

The bacterial communities associated with rotifers (Brachionus plicatilis sp. complex) and their culture water were determined using culture-dependent and -independent methods (16S rRNA gene clone library). The bacterial communities determined by the culture-independent method were more diverse than those determined by the culture-dependent method. Although the culture-dependent method indicated the bacterial community of rotifers was relatively similar to that of the culture water, 16S rRNA gene clone library analyses revealed a great difference between the two microbiotas. Our results suggest that most bacteria associated with rotifers are not easily cultured using conventional methods, and that the microbiota of rotifers do not correspond with that of the culture water completely.

Microbial biomass, community structure and metal tolerance of a naturally Pb-enriched forest soil.

PubMed

Bååth, E; Díaz-Raviña, M; Bakken, L R

2005-11-01

The effect of long-term elevated soil Pb levels on soil microbiota was studied at a forest site in Norway, where the soil has been severely contaminated with Pb since the last period of glaciation (several thousand years). Up to 10% Pb (total amount, w/w) has been found in the top layer. The microbial community was drastically affected, as judged from changes in the phospholipid fatty acid (PLFA) pattern. Specific PLFAs that were high in Pb-enriched soil were branched (especially br17:0 and br18:0), whereas PLFAs common in eukaryotic organisms such as fungi (18:2omega6,9 and 20:4) were low compared with levels at adjacent, uncontaminated sites. Congruent changes in the PLFA pattern were found upon analyzing the culturable part of the bacterial community. The high Pb concentrations in the soil resulted in increased tolerance to Pb of the bacterial community, measured using both thymidine incorporation and plate counts. Furthermore, changes in tolerance were correlated to changes in the community structure. The bacterial community of the most contaminated soils showed higher specific activity (thymidine and leucine incorporation rates) and higher culturability than that of control soils. Fungal colony forming units (CFUs) were 10 times lower in the most Pb-enriched soils, the species composition was widely different from that in control soils, and the isolated fungi had high Pb tolerance. The most commonly isolated fungus in Pb-enriched soils was Tolypocladium inflatum. Comparison of isolates from Pb-enriched soil and isolates from unpolluted soils showed that T. inflatum was intrinsically Pb-tolerant, and that the prolonged conditions with high Pb had not selected for any increased tolerance.

An Effectiveness Study of a Culturally Enriched School-Based CBT Anxiety Prevention Program

ERIC Educational Resources Information Center

Miller, Lynn D.; Laye-Gindhu, Aviva; Bennett, Joanna L.; Liu, Yan; Gold, Stephenie; March, John S.; Olson, Brent F.; Waechtler, Vanessa E.

2011-01-01

Anxiety disorders are prevalent in the school-aged population and are present across cultural groups. Scant research exists on culturally relevant prevention and intervention programs for mental health problems in the Aboriginal populations. An established cognitive behavioral program, FRIENDS for Life, was enriched to include content that was…

Enrichment and Molecular Detection of Denitrifying Methanotrophic Bacteria of the NC10 Phylum▿

PubMed Central

Ettwig, Katharina F.; van Alen, Theo; van de Pas-Schoonen, Katinka T.; Jetten, Mike S. M.; Strous, Marc

2009-01-01

Anaerobic methane oxidation coupled to denitrification was recently assigned to bacteria belonging to the uncultured phylum NC10. In this study, we incubated sediment from a eutrophic ditch harboring a diverse community of NC10 bacteria in a bioreactor with a constant supply of methane and nitrite. After 6 months, fluorescence in situ hybridization showed that NC10 bacteria dominated the resulting population. The enrichment culture oxidized methane and reduced nitrite to dinitrogen gas. We assessed NC10 phylum diversity in the inoculum and the enrichment culture, compiled the sequences currently available for this bacterial phylum, and showed that of the initial diversity, only members of one subgroup had been enriched. The growth of this subgroup was monitored by quantitative PCR and correlated to nitrite-reducing activity and the total biomass of the culture. Together, the results indicate that the enriched subgroup of NC10 bacteria is responsible for anaerobic methane oxidation coupled to nitrite reduction. Due to methodological limitations (a strong bias against NC10 bacteria in 16S rRNA gene clone libraries and inhibition by commonly used stopper material) the environmental distribution and importance of these bacteria could be largely underestimated at present. PMID:19329658

Corexit 9500 Enhances Oil Biodegradation and Changes Active Bacterial Community Structure of Oil-Enriched Microcosms.

PubMed

Techtmann, Stephen M; Zhuang, Mobing; Campo, Pablo; Holder, Edith; Elk, Michael; Hazen, Terry C; Conmy, Robyn; Santo Domingo, Jorge W

2017-05-15

To better understand the impacts of Corexit 9500 on the structure and activity levels of hydrocarbon-degrading microbial communities, we analyzed next-generation 16S rRNA gene sequencing libraries of hydrocarbon enrichments grown at 5 and 25°C using both DNA and RNA extracts as the sequencing templates. Oil biodegradation patterns in both 5 and 25°C enrichments were consistent with those reported in the literature (i.e., aliphatics were degraded faster than aromatics). Slight increases in biodegradation were observed in the presence of Corexit at both temperatures. Differences in community structure were observed between treatment conditions in the DNA-based libraries. The 25°C consortia were dominated by Vibrio , Idiomarina , Marinobacter , Alcanivorax , and Thalassospira species, while the 5°C consortia were dominated by several species of the genera Flavobacterium , Alcanivorax , and Oleispira Most of these genera have been linked to hydrocarbon degradation and have been observed after oil spills. Colwellia and Cycloclasticus , known aromatic degraders, were also found in these enrichments. The addition of Corexit did not have an effect on the active bacterial community structure of the 5°C consortia, while at 25°C, a decrease in the relative abundance of Marinobacter was observed. At 25°C, Thalassospira , Marinobacter , and Idiomarina were present at higher relative abundances in the RNA than DNA libraries, suggesting that they were active in degradation. Similarly, Oleispira was greatly stimulated by the addition of oil at 5°C. IMPORTANCE While dispersants such as Corexit 9500 can be used to treat oil spills, there is still debate on the effectiveness on enhancing oil biodegradation and its potential toxic effect on oil-degrading microbial communities. The results of this study provide some insights on the microbial dynamics of hydrocarbon-degrading bacterial populations in the presence of Corexit 9500. Operational taxonomic unit (OTU) analyses

Corexit 9500 Enhances Oil Biodegradation and Changes Active Bacterial Community Structure of Oil-Enriched Microcosms

PubMed Central

Zhuang, Mobing; Campo, Pablo; Holder, Edith; Elk, Michael; Conmy, Robyn

2017-01-01

ABSTRACT To better understand the impacts of Corexit 9500 on the structure and activity levels of hydrocarbon-degrading microbial communities, we analyzed next-generation 16S rRNA gene sequencing libraries of hydrocarbon enrichments grown at 5 and 25°C using both DNA and RNA extracts as the sequencing templates. Oil biodegradation patterns in both 5 and 25°C enrichments were consistent with those reported in the literature (i.e., aliphatics were degraded faster than aromatics). Slight increases in biodegradation were observed in the presence of Corexit at both temperatures. Differences in community structure were observed between treatment conditions in the DNA-based libraries. The 25°C consortia were dominated by Vibrio, Idiomarina, Marinobacter, Alcanivorax, and Thalassospira species, while the 5°C consortia were dominated by several species of the genera Flavobacterium, Alcanivorax, and Oleispira. Most of these genera have been linked to hydrocarbon degradation and have been observed after oil spills. Colwellia and Cycloclasticus, known aromatic degraders, were also found in these enrichments. The addition of Corexit did not have an effect on the active bacterial community structure of the 5°C consortia, while at 25°C, a decrease in the relative abundance of Marinobacter was observed. At 25°C, Thalassospira, Marinobacter, and Idiomarina were present at higher relative abundances in the RNA than DNA libraries, suggesting that they were active in degradation. Similarly, Oleispira was greatly stimulated by the addition of oil at 5°C. IMPORTANCE While dispersants such as Corexit 9500 can be used to treat oil spills, there is still debate on the effectiveness on enhancing oil biodegradation and its potential toxic effect on oil-degrading microbial communities. The results of this study provide some insights on the microbial dynamics of hydrocarbon-degrading bacterial populations in the presence of Corexit 9500. Operational taxonomic unit (OTU) analyses

Aerobic Growth on Nitroglycerin as the Sole Carbon, Nitrogen, and Energy Source by a Mixed Bacterial Culture

PubMed Central

Accashian, John V.; Vinopal, Robert T.; Kim, Byung-Joon; Smets, Barth F.

1998-01-01

Nitroglycerin (glycerol trinitrate [GTN]), an explosive and vasodilatory compound, was metabolized by mixed microbial cultures from aeration tank sludge previously exposed to GTN. Aerobic enrichment cultures removed GTN rapidly in the absence of a supplemental carbon source. Complete denitration of GTN, provided as the sole C and N source, was observed in aerobic batch cultures and proceeded stepwise via the dinitrate and mononitrate isomers, with successive steps occurring at lower rates. The denitration of all glycerol nitrate esters was found to be concomitant, and 1,2-glycerol dinitrate (1,2-GDN) and 2-glycerol mononitrate (2-GMN) were the primary GDN and GMN isomers observed. Denitration of GTN resulted in release of primarily nitrite-N, indicating a reductive denitration mechanism. Biomass growth at the expense of GTN was verified by optical density and plate count measurements. The kinetics of GTN biotransformation were 10-fold faster than reported for complete GTN denitration under anaerobic conditions. A maximum specific growth rate of 0.048 ± 0.005 h−1 (mean ± standard deviation) was estimated for the mixed culture at 25°C. Evidence of GTN toxicity was observed at GTN concentrations above 0.3 mM. To our knowledge, this is the first report of complete denitration of GTN used as a primary growth substrate by a bacterial culture under aerobic conditions. PMID:9726874

Bacterial isolates from polysaccharide enrichments cluster by host origin for Firmicutes but not Bacteroidetes.

USDA-ARS?s Scientific Manuscript database

The intestinal microbiota allows mammals to recover energy stored in plant biomass through fermentation of plant cell walls, primarily cellulose and hemicellulose. Bacteria were isolated from 8 week continuous culture enrichments with cellulose and xylan/pectin from cow (C, n=4), goat (G, n=4), huma...

Visual Estimation of Bacterial Growth Level in Microfluidic Culture Systems.

PubMed

Kim, Kyukwang; Kim, Seunggyu; Jeon, Jessie S

2018-02-03

Microfluidic devices are an emerging platform for a variety of experiments involving bacterial cell culture, and has advantages including cost and convenience. One inevitable step during bacterial cell culture is the measurement of cell concentration in the channel. The optical density measurement technique is generally used for bacterial growth estimation, but it is not applicable to microfluidic devices due to the small sample volumes in microfluidics. Alternately, cell counting or colony-forming unit methods may be applied, but these do not work in situ; nor do these methods show measurement results immediately. To this end, we present a new vision-based method to estimate the growth level of the bacteria in microfluidic channels. We use Fast Fourier transform (FFT) to detect the frequency level change of the microscopic image, focusing on the fact that the microscopic image becomes rough as the number of cells in the field of view increases, adding high frequencies to the spectrum of the image. Two types of microfluidic devices are used to culture bacteria in liquid and agar gel medium, and time-lapsed images are captured. The images obtained are analyzed using FFT, resulting in an increase in high-frequency noise proportional to the time passed. Furthermore, we apply the developed method in the microfluidic antibiotics susceptibility test by recognizing the regional concentration change of the bacteria that are cultured in the antibiotics gradient. Finally, a deep learning-based data regression is performed on the data obtained by the proposed vision-based method for robust reporting of data.

Characterisation of culture-independent and -dependent microbial communities in a high-temperature offshore chalk petroleum reservoir.

PubMed

Kaster, Krista M; Bonaunet, Kristin; Berland, Harald; Kjeilen-Eilertsen, Grethe; Brakstad, Odd Gunnar

2009-11-01

Recent studies have indicated that oil reservoirs harbour diverse microbial communities. Culture-dependent and culture-independent methods were used to evaluate the microbial diversity in produced water samples of the Ekofisk oil field, a high temperature, and fractured chalk reservoir in the North Sea. DGGE analyses of 16S rRNA gene fragments were used to assess the microbial diversity of both archaeal and bacterial communities in produced water samples and enrichment cultures from 4 different wells (B-08, X-08, X-18 and X-25). Low diversity communities were found when 16S rDNA libraries of bacterial and archaeal assemblages were generated from total community DNA obtained from produced water samples and enrichment cultures. Sequence analysis of the clones indicated close matches to microbes associated with high-temperature oil reservoirs or other similar environments. Sequences were found to be similar to members of the genera Thermotoga, Caminicella, Thermoanaerobacter, Archaeoglobus, Thermococcus, and Methanobulbus. Enrichment cultures obtained from the produced water samples were dominated by sheathed rods. Sequence analyses of the cultures indicated predominance of the genera Petrotoga, Arcobacter, Archaeoglobus and Thermococcus. The communities of both produced water and enrichment cultures appeared to be dominated by thermophilic fermenters capable of reducing sulphur compounds. These results suggest that the biochemical processes in the Ekofisk chalk reservoir are similar to those observed in high-temperature sandstone reservoirs.

Comparison of methods for identifying causative bacterial microorganisms in presumed acute endophthalmitis: conventional culture, blood culture, and PCR.

PubMed

Pongsachareonnont, Pear; Honglertnapakul, Worawalun; Chatsuwan, Tanittha

2017-02-21

Identification of bacterial pathogens in endophthalmitis is important to inform antibiotic selection and treatment decisions. Hemoculture bottles and polymerase chain reaction (PCR) analysis have been proposed to offer good detection sensitivity. This study compared the sensitivity and accuracy of a blood culture system, a PCR approach, and conventional culture methods for identification of causative bacteria in cases of acute endophthalmitis. Twenty-nine patients with a diagnosis of presumed acute bacterial endophthalmitis who underwent vitreous specimen collection at King Chulalongkorn Memorial Hospital were enrolled in this study. Forty-one specimens were collected. Each specimen was divided into three parts, and each part was analyzed using one of three microbial identification techniques: conventional plate culture, blood culture, and polymerase chain reaction and sequencing. The results of the three methods were then compared. Bacteria were identified in 15 of the 41 specimens (36.5%). Five (12.2%) specimens were positive by conventional culture methods, 11 (26.8%) were positive by hemoculture, and 11 (26.8%) were positive by PCR. Cohen's kappa analysis revealed p-values for conventional methods vs. hemoculture, conventional methods vs. PCR, and hemoculture vs. PCR of 0.057, 0.33, and 0.009, respectively. Higher detection rates of Enterococcus faecalis were observed for hemoculture and PCR than for conventional methods. Blood culture bottles and PCR detection may facilitate bacterial identification in cases of presumed acute endophthalmitis. These techniques should be used in addition to conventional plate culture methods because they provide a greater degree of sensitivity than conventional plate culture alone for the detection of specific microorganisms such as E. faecalis. Thai Clinical Trial Register No. TCTR20110000024 .

Culturable endophytic bacterial communities associated with field-grown soybean.

PubMed

de Almeida Lopes, K B; Carpentieri-Pipolo, V; Oro, T H; Stefani Pagliosa, E; Degrassi, G

2016-03-01

Assess the diversity of the culturable endophytic bacterial population associated with transgenic and nontransgenic soybean grown in field trial sites in Brazil and characterize them phenotypically and genotypically focusing on characteristics related to plant growth promotion. Endophytic bacteria were isolated from roots, stems and leaves of soybean cultivars (nontransgenic (C) and glyphosate-resistant (GR) transgenic soybean), including the isogenic BRS133 and BRS245RR. Significant differences were observed in bacterial densities in relation to genotype and tissue from which the isolates were obtained. The highest number of bacteria was observed in roots and in GR soybean. Based on characteristics related to plant growth promotion, 54 strains were identified by partial 16S rRNA sequence analysis, with most of the isolates belonging to the species Enterobacter ludwigii and Variovorax paradoxus. Among the isolates, 44·4% were able to either produce indoleacetic acid (IAA) or solubilize phosphates, and 9·2% (all from GR soybean) presented both plant growth-promoting activities. The results from this study indicate that the abundance of endophytic bacterial communities of soybean differs between cultivars and in general it was higher in the transgenic cultivars than in nontransgenic cultivars. BRS 245 RR exhibited no significant difference in abundance compared to nontransgenic BRS133. This suggests that the impact of the management used in the GR soybean fields was comparable with the impacts of some enviromental factors. However, the bacterial endophytes associated to GR and nontransgenic soybean were different. The soybean-associated bacteria showing characteristics related to plant growth promotion were identified as belonging to the species Pantoea agglomerans and Variovorax paradoxus. Our study demonstrated differences concerning compostion of culturable endophytic bacterial population in nontransgenic and transgenic soybean. © 2016 The Society for Applied

Hydrogenotrophic culture enrichment reveals rumen Lachnospiraceae and Ruminococcaceae acetogens and hydrogen-responsive Bacteroidetes from pasture-fed cattle.

PubMed

Gagen, Emma J; Padmanabha, Jagadish; Denman, Stuart E; McSweeney, Christopher S

2015-07-01

Molecular information suggests that there is a broad diversity of acetogens in the rumen, distinct from any currently isolated acetogens. We combined molecular analysis with enrichment culture techniques to investigate this diversity further. Methane-inhibited, hydrogenotrophic enrichment cultures produced acetate as the dominant end product. Acetyl-CoA synthase gene analysis revealed putative acetogens in the cultures affiliated with the Lachnospiraceae and Ruminococcaceae as has been found in other rumen studies. No formyltetrahydrofolate synthetase genes affiliating with acetogens or with 'homoacetogen similarity' scores >90% were identified. To further investigate the hydrogenotrophic populations in these cultures and link functional gene information with 16S rRNA gene identity, cultures were subcultured quickly, twice, through medium without exogenous hydrogen, followed by incubation without exogenous hydrogen. Comparison of cultures lacking hydrogen and their parent cultures revealed novel Lachnospiraceae and Ruminococcaceae that diminished in the absence of hydrogen, supporting the hypothesis that they were likely the predominant acetogens in the enrichments. Interestingly, a range of Bacteroidetes rrs sequences that demonstrated <86% identity to any named isolate also diminished in cultures lacking hydrogen. Acetogens or sulphate reducers from the Bacteroidetes have not been reported previously; therefore this observation requires further investigation. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Enrichment and identification of polycyclic aromatic compound-degrading bacteria enriched from sediment samples.

PubMed

Long, Rachel M; Lappin-Scott, Hilary M; Stevens, Jamie R

2009-07-01

The degradation of polycyclic aromatic compounds (PACs) has been widely studied. Knowledge of the degradation of PACs by microbial populations can be utilized in the remediation of contaminated sites. To isolate and identify PAC-degrading bacteria for potential use in future bioremediation programmes, we established a series of PAC enrichments under the same experimental conditions from a single sediment sample taken from a highly polluted estuarine site. Enrichment cultures were established using the pollutants: anthracene, phenanthrene and dibenzothiophene as a sole carbon source. The shift in microbial community structure on each of these carbon sources was monitored by analysis of a time series of samples from each culture using 16S rRNA polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE). Significantly, our findings demonstrate that shifts in the constituent species within each degradative community are directly attributable to enrichment with different PACs. Subsequently, we characterized the microorganisms comprising the degradative communities within each enrichment using 16S rRNA sequence data. Our findings demonstrate that the ability to degrade PACs is present in five divisions of the Proteobacteria and Actinobacteria. By determining the precise identity of the PAC-degrading bacterial species isolated from a single sediment sample, and by comparing our findings with previously published research, we demonstrate how bacteria with similar PAC degrading capabilities and 16S rRNA signatures are found in similarly polluted environments in geographically very distant locations, e.g., China, Italy, Japan and Hawaii. Such a finding suggests that geographical barriers do not limit the distribution of key PAC-degrading bacteria; this finding is in accordance with the Baas-Becking hypothesis "everything is everywhere; the environment selects" and may have significant consequences for the global distribution of PAC-degrading bacteria and

Simplifying Collection of Corneal Specimens in Cases of Suspected Bacterial Keratitis

PubMed Central

Kaye, Stephen B.; Rao, Prasad G.; Smith, Godfrey; Scott, John A.; Hoyles, Sharon; Morton, Clare E.; Willoughby, Colin; Batterbury, Mark; Harvey, Graham

2003-01-01

Identification of the causative organisms in suspected bacterial keratitis traditionally involves collecting multiple corneal scrapes, which are plated directly onto different solid agar culture media. Difficulties have been reported with this practice, so the development of a simpler diagnostic method in suspected bacterial keratitis would be useful. It is unclear whether a single corneal scrape sent to the microbiology laboratory in a liquid transport culture medium (indirect method) is as reliable for the diagnosis of bacterial keratitis as inoculation of multiple scrapes directly onto agar plates (direct method). To investigate this, bacterial recovery was assessed following transfer and transport of different concentrations and types of bacteria from an artificially contaminated surgical blade into brain heart infusion (BHI). Bacterial recovery rates between the proposed (indirect) and standard (direct) method were then compared after the in vitro inoculation of pig corneas and following specimen collection in patients with presumed bacterial ulcerative keratitis. Recovery of bacteria from contaminated surgical blades was found to be the same from both solid and liquid culture media. There was no significant difference in the numbers of positive cultures from solid (direct) and liquid (indirect) culture media, both in the experimental pig cornea inoculation study (P = 0.34) and in experiments with patients with clinical infections (P = 0.4), with an 85.2% agreement between methods (kappa = 0.61, P < 0.0001). In conclusion, therefore, the collection of two corneal scrapes, one used for Gram staining and the other transported in BHI followed by plating and subculturing in an enrichment medium, provides a simple method for the investigation of presumed bacterial keratitis. PMID:12843063

Effect of organic acids production and bacterial community on the possible mechanism of phosphorus solubilization during composting with enriched phosphate-solubilizing bacteria inoculation.

PubMed

Wei, Yuquan; Zhao, Yue; Shi, Mingzi; Cao, Zhenyu; Lu, Qian; Yang, Tianxue; Fan, Yuying; Wei, Zimin

2018-01-01

Enriched phosphate-solubilizing bacteria (PSB) agent were acquired by domesticated cultivation, and inoculated into kitchen waste composting in different stages. The effect of different treatments on organic acids production, tricalcium phosphate (TCP) solubilization and their relationship with bacterial community were investigated during composting. Our results pointed out that inoculation affected pH, total acidity and the production of oxalic, lactic, citric, succinic, acetic and formic acids. We also found a strong advantage in the solubilization of TCP and phosphorus (P) availability for PSB inoculation especially in the cooling stage. Redundancy analysis and structural equation models demonstrated inoculation by different methods changed the correlation of the bacterial community composition with P fractions as well as organic acids, and strengthened the cooperative function related to P transformation among species during composting. Finally, we proposed a possible mechanism of P solubilization with enriched PSB inoculation, which was induced by bacterial community and organic acids production. Copyright © 2017 Elsevier Ltd. All rights reserved.

Measuring the Level of Agreement Between Cloacal Gram's Stains and Bacterial Cultures in Hispaniolan Amazon Parrots ( Amazona ventralis ).

PubMed

Evans, Erika E; Mitchell, Mark A; Whittington, Julia K; Roy, Alma; Tully, Thomas N

2014-12-01

Cloacal or fecal Gram's stains and bacterial cultures are routinely performed during avian physical examinations to assess the microbial flora of the gastrointestinal tract. Although cloacal or fecal Gram's stains and bacterial cultures are considered routine diagnostic procedures, the level of agreement between the individual tests has not been determined. To investigate the level of agreement between results from Gram's stain and bacterial culture when used to assess cloacal or fecal samples from psittacine birds, samples were taken from 21 clinically healthy Hispaniolan Amazon parrots ( Amazona ventralis ) and tested by Gram's stain cytology and bacterial culture. Most bacteria (97.2%) identified by Gram's stain were gram positive. However, gram-negative organisms were identified in 7 of 21 (33.3%; 95% confidence interval: 13.3%-53.3%) birds. Escherichia coli was the only gram-negative organism identified on culture. Agreement between results of Gram's stain and culture was fair (weighted κ = 0.27). The results of this study suggest that Gram's stains and bacterial culture may need to be performed with a parallel testing strategy to limit the likelihood of misclassifying the microbial flora of psittacine patients.

Yersinia enterocolitica in slaughter pig tonsils: enumeration and detection by enrichment versus direct plating culture.

PubMed

Van Damme, Inge; Habib, Ihab; De Zutter, Lieven

2010-02-01

Tonsil samples from 139 slaughter pigs were examined for the presence of pathogenic Yersinia enterocolitica by enrichment procedures based on the standard method ISO 10273:2003. In addition, samples were tested by direct plating method to evaluate its efficiency compared to the enrichment culture methods and to quantify the level of contamination in porcine tonsils. In total, 52 samples (37.4%) were positive for pathogenic Y. enterocolitica, all belonging to bioserotype 4/O:3. Fifty out of the 52 positive samples (96.2%) were detected by direct plating. Enumeration showed an average concentration of 4.5 log(10) CFU g(-1) and 4.4 log(10) CFU g(-1) tonsil on Salmonella-Shigella-desoxycholate-calcium chloride (SSDC) and cefsulodin-irgasan-novobiocin (CIN) agar plates, respectively. The enrichment procedures recommended by the ISO 10273:2003 method were not optimal for the isolation of pathogenic Y. enterocolitica from pig tonsils: two days enrichment in irgasan-ticarcillin-potassium chlorate (ITC) broth resulted in an isolation rate of 84.6%, while 5 days enrichment in peptone-sorbitol-bile (PSB) broth recovered only 59.6% of positive samples. Reducing the enrichment time in PSB from 5 to 2 days resulted in a significantly higher recovery rate (94.2%) and might serve as an appropriate enrichment protocol for the isolation of pathogenic Y. enterocolitica from pig tonsils. Compared to enrichment culture methods, results based on direct plating can be obtained in a shorter time course and provide quantitative data that might be needed for further risk assessment studies.

A quasi-universal medium to break the aerobic/anaerobic bacterial culture dichotomy in clinical microbiology.

PubMed

Dione, N; Khelaifia, S; La Scola, B; Lagier, J C; Raoult, D

2016-01-01

In the mid-19th century, the dichotomy between aerobic and anaerobic bacteria was introduced. Nevertheless, the aerobic growth of strictly anaerobic bacterial species such as Ruminococcus gnavus and Fusobacterium necrophorum, in a culture medium containing antioxidants, was recently demonstrated. We tested aerobically the culture of 623 bacterial strains from 276 bacterial species including 82 strictly anaerobic, 154 facultative anaerobic, 31 aerobic and nine microaerophilic bacterial species as well as ten fungi. The basic culture medium was based on Schaedler agar supplemented with 1 g/L ascorbic acid and 0.1 g/L glutathione (R-medium). We successively optimized this media, adding 0.4 g/L uric acid, using separate autoclaving of the component, or adding haemin 0.1 g/L or α-ketoglutarate 2 g/L. In the basic medium, 237 bacterial species and ten fungal species grew but with no growth of 36 bacterial species, including 22 strict anaerobes. Adding uric acid allowed the growth of 14 further species including eight strict anaerobes, while separate autoclaving allowed the growth of all tested bacterial strains. To extend its potential use for fastidious bacteria, we added haemin for Haemophilus influenzae, Haemophilus parainfluenzae and Eikenella corrodens and α-ketoglutarate for Legionella pneumophila. This medium allowed the growth of all tested strains with the exception of Mycobacterium tuberculosis and Mycobacterium bovis. Testing primoculture and more fastidious species will constitute the main work to be done, but R-medium coupled with a rapid identification method (matrix-assisted laser desorption/ionization time-of-flight mass spectrometry) will facilitate the anaerobic culture in clinical microbiology laboratories. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

Absence of bacterial DNA in culture-negative urine from cats with and without lower urinary tract disease.

PubMed

Lund, Heidi Sjetne; Skogtun, Gaute; Sørum, Henning; Eggertsdóttir, Anna Vigdís

2015-10-01

A diagnosis of bacterial cystitis commonly relies on a positive microbiological culture demonstrating the presence of a significant number of colony-forming units/ml urine, as urine within the upper urinary tract, bladder and proximal urethra generally is considered sterile. Recent studies from human and veterinary medicine indicate the presence of non-culturable bacteria in culture-negative urine samples. The aim of the present study was to determine the occurrence of bacterial DNA in culture-negative urine samples from cats with signs of feline lower urinary tract disease (FLUTD) and healthy control cats by 16S ribosomal DNA PCR and subsequent sequencing. The study sample included 38 culture-negative urine samples from cats with FLUTD and 43 culture-negative samples from control cats. Eight culture-positive urine samples from cats with FLUTD were included as external positive controls in addition to negative reaction controls. Of possible methodological limitations, degradation of DNA due to storage, the use of non-sedimented urine for DNA isolation and lack of internal positive reaction controls should be mentioned. The positive controls were recognised, but occurrence of bacterial DNA in culture-negative urine from cats with or without signs of lower urinary tract disease was not demonstrated. However, considering the possible methodological limitations, the presence of bacterial DNA in the urine of culture-negative FLUTD cats cannot be excluded based on the present results alone. Therefore, a prospective study reducing the possibility of degradation of DNA due to storage, in combination with modifications enhancing the chance of detecting even lower levels of bacterial DNA in culture-negative samples, seems warranted. © ISFM and AAFP 2014.

An integrated microfluidic analysis microsystems with bacterial capture enrichment and in-situ impedance detection

NASA Astrophysics Data System (ADS)

Liu, Hai-Tao; Wen, Zhi-Yu; Xu, Yi; Shang, Zheng-Guo; Peng, Jin-Lan; Tian, Peng

2017-09-01

In this paper, an integrated microfluidic analysis microsystems with bacterial capture enrichment and in-situ impedance detection was purposed based on microfluidic chips dielectrophoresis technique and electrochemical impedance detection principle. The microsystems include microfluidic chip, main control module, and drive and control module, and signal detection and processing modulet and result display unit. The main control module produce the work sequence of impedance detection system parts and achieve data communication functions, the drive and control circuit generate AC signal which amplitude and frequency adjustable, and it was applied on the foodborne pathogens impedance analysis microsystems to realize the capture enrichment and impedance detection. The signal detection and processing circuit translate the current signal into impendence of bacteria, and transfer to computer, the last detection result is displayed on the computer. The experiment sample was prepared by adding Escherichia coli standard sample into chicken sample solution, and the samples were tested on the dielectrophoresis chip capture enrichment and in-situ impedance detection microsystems with micro-array electrode microfluidic chips. The experiments show that the Escherichia coli detection limit of microsystems is 5 × 104 CFU/mL and the detection time is within 6 min in the optimization of voltage detection 10 V and detection frequency 500 KHz operating conditions. The integrated microfluidic analysis microsystems laid the solid foundation for rapid real-time in-situ detection of bacteria.

Mountain pine beetles colonizing historical and naive host trees are associated with a bacterial community highly enriched in genes contributing to terpene metabolism.

PubMed

Adams, Aaron S; Aylward, Frank O; Adams, Sandye M; Erbilgin, Nadir; Aukema, Brian H; Currie, Cameron R; Suen, Garret; Raffa, Kenneth F

2013-06-01

The mountain pine beetle, Dendroctonus ponderosae, is a subcortical herbivore native to western North America that can kill healthy conifers by overcoming host tree defenses, which consist largely of high terpene concentrations. The mechanisms by which these beetles contend with toxic compounds are not well understood. Here, we explore a component of the hypothesis that beetle-associated bacterial symbionts contribute to the ability of D. ponderosae to overcome tree defenses by assisting with terpene detoxification. Such symbionts may facilitate host tree transitions during range expansions currently being driven by climate change. For example, this insect has recently breached the historical geophysical barrier of the Canadian Rocky Mountains, providing access to näive tree hosts and unprecedented connectivity to eastern forests. We use culture-independent techniques to describe the bacterial community associated with D. ponderosae beetles and their galleries from their historical host, Pinus contorta, and their more recent host, hybrid P. contorta-Pinus banksiana. We show that these communities are enriched with genes involved in terpene degradation compared with other plant biomass-processing microbial communities. These pine beetle microbial communities are dominated by members of the genera Pseudomonas, Rahnella, Serratia, and Burkholderia, and the majority of genes involved in terpene degradation belong to these genera. Our work provides the first metagenome of bacterial communities associated with a bark beetle and is consistent with a potential microbial contribution to detoxification of tree defenses needed to survive the subcortical environment.

Erythrocyte Enrichment in Hematopoietic Progenitor Cell Cultures Based on Magnetic Susceptibility of the Hemoglobin

PubMed Central

Jin, Xiaoxia; Abbot, Stewart; Zhang, Xiaokui; Kang, Lin; Voskinarian-Berse, Vanessa; Zhao, Rui; Kameneva, Marina V.; Moore, Lee R.; Chalmers, Jeffrey J.; Zborowski, Maciej

2012-01-01

Using novel media formulations, it has been demonstrated that human placenta and umbilical cord blood-derived CD34+ cells can be expanded and differentiated into erythroid cells with high efficiency. However, obtaining mature and functional erythrocytes from the immature cell cultures with high purity and in an efficient manner remains a significant challenge. A distinguishing feature of a reticulocyte and maturing erythrocyte is the increasing concentration of hemoglobin and decreasing cell volume that results in increased cell magnetophoretic mobility (MM) when exposed to high magnetic fields and gradients, under anoxic conditions. Taking advantage of these initial observations, we studied a noninvasive (label-free) magnetic separation and analysis process to enrich and identify cultured functional erythrocytes. In addition to the magnetic cell separation and cell motion analysis in the magnetic field, the cell cultures were characterized for cell sedimentation rate, cell volume distributions using differential interference microscopy, immunophenotyping (glycophorin A), hemoglobin concentration and shear-induced deformability (elongation index, EI, by ektacytometry) to test for mature erythrocyte attributes. A commercial, packed column high-gradient magnetic separator (HGMS) was used for magnetic separation. The magnetically enriched fraction comprised 80% of the maturing cells (predominantly reticulocytes) that showed near 70% overlap of EI with the reference cord blood-derived RBC and over 50% overlap with the adult donor RBCs. The results demonstrate feasibility of label-free magnetic enrichment of erythrocyte fraction of CD34+ progenitor-derived cultures based on the presence of paramagnetic hemoglobin in the maturing erythrocytes. PMID:22952572

Engaging High School Girls in Native American Culturally Responsive STEAM Enrichment Activities

ERIC Educational Resources Information Center

Kant, Joanita M.; Burckhard, Suzette R.; Meyers, Richard T.

2018-01-01

Providing science, technology, engineering, art, and mathematics (STEAM) culturally responsive enrichment activities is one way of promoting more interest in science, technology, engineering, and mathematics (STEM) studies and careers among indigenous students. The purpose of the study was to explore the impact, if any, of STEAM culturally…

Anaerobic Degradation of Benzene, Toluene, Ethylbenzene, and o-Xylene in Sediment-Free Iron-Reducing Enrichment Cultures

PubMed Central

Jahn, Michael K.; Haderlein, Stefan B.; Meckenstock, Rainer U.

2005-01-01

Monoaromatic hydrocarbons such as benzene, toluene, ethylbenzene, and xylene (BTEX) are widespread contaminants in groundwater. We examined the anaerobic degradation of BTEX compounds with amorphous ferric oxide as electron acceptor. Successful enrichment cultures were obtained for all BTEX substrates both in the presence and absence of AQDS (9,10-anthraquinone-2,6-disulfonic acid). The electron balances showed a complete anaerobic oxidation of the aromatic compounds to CO2. This is the first report on the anaerobic degradation of o-xylene and ethylbenzene in sediment-free iron-reducing enrichment cultures. PMID:15933041

Vision Marker-Based In Situ Examination of Bacterial Growth in Liquid Culture Media.

PubMed

Kim, Kyukwang; Choi, Duckyu; Lim, Hwijoon; Kim, Hyeongkeun; Jeon, Jessie S

2016-12-18

The detection of bacterial growth in liquid media is an essential process in determining antibiotic susceptibility or the level of bacterial presence for clinical or research purposes. We have developed a system, which enables simplified and automated detection using a camera and a striped pattern marker. The quantification of bacterial growth is possible as the bacterial growth in the culturing vessel blurs the marker image, which is placed on the back of the vessel, and the blurring results in a decrease in the high-frequency spectrum region of the marker image. The experiment results show that the FFT (fast Fourier transform)-based growth detection method is robust to the variations in the type of bacterial carrier and vessels ranging from the culture tubes to the microfluidic devices. Moreover, the automated incubator and image acquisition system are developed to be used as a comprehensive in situ detection system. We expect that this result can be applied in the automation of biological experiments, such as the Antibiotics Susceptibility Test or toxicity measurement. Furthermore, the simple framework of the proposed growth measurement method may be further utilized as an effective and convenient method for building point-of-care devices for developing countries.

A new protocol to detect multiple foodborne pathogens with PCR dipstick DNA chromatography after a six-hour enrichment culture in a broad-range food pathogen enrichment broth.

PubMed

Hayashi, Masahiro; Natori, Tatsuya; Kubota-Hayashi, Sayoko; Miyata, Machiko; Ohkusu, Kiyofumi; Kawamoto, Keiko; Kurazono, Hisao; Makino, Souichi; Ezaki, Takayuki

2013-01-01

A quick foodborne pathogen screening method after six-hour enrichment culture with a broad-range food pathogen enrichment broth is described. Pathogenic factors of Salmonella enterica, Shigella spp., enteroinvasive Escherichia coli, and enterohemorrhagic E. coli are amplified with a cocktail primer and rapid polymerase chain reaction (PCR), which finishes amplification in 30 min. The PCR amplicon was differentiated with a dipstick DNA chromatography assay in 5-10 min. Starting from a four- to six-hour enrichment culture, this assay was finished within 45 min. Detection sensitivity of this protocol was less than 2.5 CFU/25 g for S. enterica and 3.3 CFU/25 g for enterohemorrhagic E. coli in spiked ground meat experiments.

A New Protocol to Detect Multiple Foodborne Pathogens with PCR Dipstick DNA Chromatography after a Six-Hour Enrichment Culture in a Broad-Range Food Pathogen Enrichment Broth

PubMed Central

Hayashi, Masahiro; Natori, Tatsuya; Kubota-Hayashi, Sayoko; Miyata, Machiko; Ohkusu, Kiyofumi; Kurazono, Hisao; Makino, Souichi; Ezaki, Takayuki

2013-01-01

A quick foodborne pathogen screening method after six-hour enrichment culture with a broad-range food pathogen enrichment broth is described. Pathogenic factors of Salmonella enterica, Shigella spp., enteroinvasive Escherichia coli, and enterohemorrhagic E. coli are amplified with a cocktail primer and rapid polymerase chain reaction (PCR), which finishes amplification in 30 min. The PCR amplicon was differentiated with a dipstick DNA chromatography assay in 5–10 min. Starting from a four- to six-hour enrichment culture, this assay was finished within 45 min. Detection sensitivity of this protocol was less than 2.5 CFU/25 g for S. enterica and 3.3 CFU/25 g for enterohemorrhagic E. coli in spiked ground meat experiments. PMID:24364031

Effect of different mixing methods on the bacterial microleakage of calcium-enriched mixture cement.

PubMed

Shahi, Shahriar; Jeddi Khajeh, Soniya; Rahimi, Saeed; Yavari, Hamid R; Jafari, Farnaz; Samiei, Mohammad; Ghasemi, Negin; Milani, Amin S

2016-10-01

Calcium-enriched mixture (CEM) cement is used in the field of endodontics. It is similar to mineral trioxide aggregate in its main ingredients. The present study investigated the effect of different mixing methods on the bacterial microleakage of CEM cement. A total of 55 human single-rooted human permanent teeth were decoronated so that 14-mm-long samples were obtained and obturated with AH26 sealer and gutta-percha using lateral condensation technique. Three millimeters of the root end were cut off and randomly divided into 3 groups of 15 each (3 mixing methods of amalgamator, ultrasonic and conventional) and 2 negative and positive control groups (each containing 5 samples). BHI (brain-heart infusion agar) suspension containing Enterococcus faecalis was used for bacterial leakage assessment. Statistical analysis was carried out using descriptive statistics, Kaplan-Meier survival analysis with censored data and log rank test. Statistical significance was set at P<0.05. The survival means for conventional, amalgamator and ultrasonic methods were 62.13±12.44, 68.87±12.79 and 77.53±12.52 days, respectively. The log rank test showed no significant differences between the groups. Based on the results of the present study it can be concluded that different mixing methods had no significant effect on the bacterial microleakage of CEM cement.

Bacterial stimulation of adventitious rooting on in vitro cultured slash pine (Pinus elliottii Engelm.) seedling explants.

PubMed

Burns, J A; Schwarz, O J

1996-02-01

A bacterium has been isolated that initiates adventitious rooting when co-cultured under in vitro conditions with seedling-produced hypocotylary explants of slash pine (Pinus elliottii). Rooting efficiencies produced through bacterial-explant co-culture range from approximately 15% to greater than 90% over non-treated controls. Explant exposure to the root inducing bacterium has produced no obvious pathology in the regenerated plantlets. Seedling explants rooted by bacterial-explant co-culture have been successfully transitioned to ambient greenhouse conditions.

The molecular bacterial load assay replaces solid culture for measuring early bactericidal response to antituberculosis treatment.

PubMed

Honeyborne, Isobella; Mtafya, Bariki; Phillips, Patrick P J; Hoelscher, Michael; Ntinginya, Elias N; Kohlenberg, Anke; Rachow, Andrea; Rojas-Ponce, Gabriel; McHugh, Timothy D; Heinrich, Norbert

2014-08-01

We evaluated the use of the molecular bacterial load (MBL) assay, for measuring viable Mycobacterium tuberculosis in sputum, in comparison with solid agar and liquid culture. The MBL assay provides early information on the rate of decline in bacterial load and has technical advantages over culture in either form. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Enrichment of spinal cord cell cultures with motoneurons

PubMed Central

1978-01-01

Spinal cord cell cultures contain several types of neurons. Two methods are described for enriching such cultures with motoneurons (defined here simply as cholinergic cells that are capable of innervating muscle). In the first method, 7-day embryonic chick spinal cord neurons were separated according to size by 1 g velocity sedimentation. It is assumed that cholinergic motoneurons are among the largest cells present at this stage. The spinal cords were dissociated vigorously so that 95-98% of the cells in the initial suspension were isolated from one another. Cells in leading fractions (large cell fractions: LCFs) contain about seven times as much choline acetyltransferase (CAT) activity per unit cytoplasm as do cells in trailing fractions (small cell fractions: SCFs). Muscle cultures seeded with LCFs develop 10-70 times as much CAT as cultures seeded with SCFs and six times as much CAT as cultures seeded with control (unfractionated) spinal cord cells. More than 20% of the large neurons in LCF-muscle cultures innervate nearby myotubes. In the second method, neurons were gently dissociated from 4-day embryonic spinal cords and maintained in vitro. This approach is based on earlier observations that cholinergic neurons are among the first cells to withdraw form the mitotic cycle in the developing chick embryo (Hamburger, V. 1948. J. Comp. Neurol. 88:221- 283; and Levi-Montalcini, R. 1950. J. Morphol. 86:253-283). 4-Day spinal cord-muscle cultures develop three times as much CAT as do 7-day spinal cord-muscle plates, prepared in the same (gentle) manner. More than 50% of the relatively large 4-day neurons innervate nearby myotubes. Thus, both methods are useful first steps toward the complete isolation of motoneurons. Both methods should facilitate study of the development of cholinergic neurons and of nerve-muscle synapse formation. PMID:566275

DEPENDENCE OF A HIGH-RATE, PCE-DECHLORINATING ENRICHMENT CULTURE ON METHANOGENIC ACTIVITY. (R825549C053)

EPA Science Inventory

The role served by the presence of methanogenic activity within a tetrachloroethene (PCE)-dechlorinating culture was investigated through a series of supplementation experiments. An acclimated lactate-enrichment culture (LEC 1) capable of rapidly converting PCE to ethene was s...

Diamagnetic levitation enhances growth of liquid bacterial cultures by increasing oxygen availability

PubMed Central

Dijkstra, Camelia E.; Larkin, Oliver J.; Anthony, Paul; Davey, Michael R.; Eaves, Laurence; Rees, Catherine E. D.; Hill, Richard J. A.

2011-01-01

Diamagnetic levitation is a technique that uses a strong, spatially varying magnetic field to reproduce aspects of weightlessness, on the Earth. We used a superconducting magnet to levitate growing bacterial cultures for up to 18 h, to determine the effect of diamagnetic levitation on all phases of the bacterial growth cycle. We find that diamagnetic levitation increases the rate of population growth in a liquid culture and reduces the sedimentation rate of the cells. Further experiments and microarray gene analysis show that the increase in growth rate is owing to enhanced oxygen availability. We also demonstrate that the magnetic field that levitates the cells also induces convective stirring in the liquid. We present a simple theoretical model, showing how the paramagnetic force on dissolved oxygen can cause convection during the aerobic phases of bacterial growth. We propose that this convection enhances oxygen availability by transporting oxygen around the liquid culture. Since this process results from the strong magnetic field, it is not present in other weightless environments, e.g. in Earth orbit. Hence, these results are of significance and timely to researchers considering the use of diamagnetic levitation to explore effects of weightlessness on living organisms and on physical phenomena. PMID:20667843

Diamagnetic levitation enhances growth of liquid bacterial cultures by increasing oxygen availability.

PubMed

Dijkstra, Camelia E; Larkin, Oliver J; Anthony, Paul; Davey, Michael R; Eaves, Laurence; Rees, Catherine E D; Hill, Richard J A

2011-03-06

Diamagnetic levitation is a technique that uses a strong, spatially varying magnetic field to reproduce aspects of weightlessness, on the Earth. We used a superconducting magnet to levitate growing bacterial cultures for up to 18 h, to determine the effect of diamagnetic levitation on all phases of the bacterial growth cycle. We find that diamagnetic levitation increases the rate of population growth in a liquid culture and reduces the sedimentation rate of the cells. Further experiments and microarray gene analysis show that the increase in growth rate is owing to enhanced oxygen availability. We also demonstrate that the magnetic field that levitates the cells also induces convective stirring in the liquid. We present a simple theoretical model, showing how the paramagnetic force on dissolved oxygen can cause convection during the aerobic phases of bacterial growth. We propose that this convection enhances oxygen availability by transporting oxygen around the liquid culture. Since this process results from the strong magnetic field, it is not present in other weightless environments, e.g. in Earth orbit. Hence, these results are of significance and timely to researchers considering the use of diamagnetic levitation to explore effects of weightlessness on living organisms and on physical phenomena.

Population Structure of Manganese-Oxidizing Bacteria in Stratified Soils and Properties of Manganese Oxide Aggregates under Manganese–Complex Medium Enrichment

PubMed Central

Zhang, Zhongming; Chen, Hong; Liu, Jin; Ali, Muhammad; Liu, Fan; Li, Lin

2013-01-01

Manganese-oxidizing bacteria in the aquatic environment have been comprehensively investigated. However, little information is available about the distribution and biogeochemical significance of these bacteria in terrestrial soil environments. In this study, stratified soils were initially examined to investigate the community structure and diversity of manganese-oxidizing bacteria. Total 344 culturable bacterial isolates from all substrata exhibited Mn(II)-oxidizing activities at the range of 1 µM to 240 µM of the equivalent MnO2. The high Mn(II)-oxidizing isolates (>50 mM MnO2) were identified as the species of phyla Actinobacteria, Firmicutes and Proteobacteria. Seven novel Mn(II)-oxidizing bacterial genera (species), namely, Escherichia, Agromyces, Cellulomonas, Cupriavidus, Microbacterium, Ralstonia, and Variovorax, were revealed via comparative phylogenetic analysis. Moreover, an increase in the diversity of soil bacterial community was observed after the combined enrichment of Mn(II) and carbon-rich complex. The phylogenetic classification of the enriched bacteria represented by predominant denaturing gradient gel electrophoresis bands, was apparently similar to culturable Mn(II)-oxidizing bacteria. The experiments were further undertaken to investigate the properties of the Mn oxide aggregates formed by the bacterial isolates with high Mn(II)-oxidizing activity. Results showed that these bacteria were closely encrusted with their Mn oxides and formed regular microspherical aggregates under prolonged Mn(II) and carbon-rich medium enrichment for three weeks. The biotic oxidation of Mn(II) to Mn(III/IV) by these isolates was confirmed by kinetic examinations. X-ray diffraction assays showed the characteristic peaks of several Mn oxides and rhodochrosite from these aggregates. Leucoberbelin blue tests also verified the Mn(II)-oxidizing activity of these aggregates. These results demonstrated that Mn oxides were formed at certain amounts under the enrichment

Direct cloning from enrichment cultures, a reliable strategy for isolation of complete operons and genes from microbial consortia.

PubMed

Entcheva, P; Liebl, W; Johann, A; Hartsch, T; Streit, W R

2001-01-01

Enrichment cultures of microbial consortia enable the diverse metabolic and catabolic activities of these populations to be studied on a molecular level and to be explored as potential sources for biotechnology processes. We have used a combined approach of enrichment culture and direct cloning to construct cosmid libraries with large (>30-kb) inserts from microbial consortia. Enrichment cultures were inoculated with samples from five environments, and high amounts of avidin were added to the cultures to favor growth of biotin-producing microbes. DNA was extracted from three of these enrichment cultures and used to construct cosmid libraries; each library consisted of between 6,000 and 35,000 clones, with an average insert size of 30 to 40 kb. The inserts contained a diverse population of genomic DNA fragments isolated from the consortia organisms. These three libraries were used to complement the Escherichia coli biotin auxotrophic strain ATCC 33767 Delta(bio-uvrB). Initial screens resulted in the isolation of seven different complementing cosmid clones, carrying biotin biosynthesis operons. Biotin biosynthesis capabilities and growth under defined conditions of four of these clones were studied. Biotin measured in the different culture supernatants ranged from 42 to 3,800 pg/ml/optical density unit. Sequencing the identified biotin synthesis genes revealed high similarities to bio operons from gram-negative bacteria. In addition, random sequencing identified other interesting open reading frames, as well as two operons, the histidine utilization operon (hut), and the cluster of genes involved in biosynthesis of molybdopterin cofactors in bacteria (moaABCDE).

Evaluating the effect of intraoperative peritoneal lavage on bacterial culture in dogs with suspected septic peritonitis.

PubMed

Swayne, Seanna L; Brisson, Brigitte; Weese, J Scott; Sears, William

2012-09-01

This pilot study describes the effect of intraoperative peritoneal lavage (IOPL) on bacterial counts and outcome in clinical cases of septic peritonitis. Intraoperative samples were cultured before and after IOPL. Thirty-three dogs with presumed septic peritonitis on the basis of cytology were managed surgically during the study period. Positive pre-lavage bacterial cultures were found in 14 cases, 13 of which were a result of intestinal leakage. The post-lavage cultures showed fewer isolates in 9 cases and in 1 case became negative. The number of dogs with a decrease in the concentration of bacteria cultured from pre-lavage to post-lavage samples was not statistically significant. There was no significant effect of the change in pre- to post-lavage culture, single versus multiple types of bacteria, selection of an appropriate empiric antimicrobial on survival or the need for subsequent surgery. This pilot study describes the effect of intraoperative peritoneal lavage (IOPL) on bacterial counts and outcome in clinical cases of septic peritonitis. Intraoperative samples were cultured before and after IOPL. Thirty-three dogs with presumed septic peritonitis on the basis of cytology were managed surgically during the study period. Positive pre-lavage bacterial cultures were found in 14 cases, 13 of which were a result of intestinal leakage. The post-lavage cultures showed fewer isolates in 9 cases and in 1 case became negative. The number of dogs with a decrease in the concentration of bacteria cultured from pre-lavage to post-lavage samples was not statistically significant. There was no significant effect of the change in pre- to post-lavage culture, single versus multiple types of bacteria, selection of an appropriate empiric antimicrobial on survival or the need for subsequent surgery.

Effect of biostimulants on 2,4,6-trinitrotoluene (TNT) degradation and bacterial community composition in contaminated aquifer sediment enrichments.

PubMed

Fahrenfeld, Nicole; Zoeckler, Jeffrey; Widdowson, Mark A; Pruden, Amy

2013-04-01

2,4,6-Trinitrotoluene (TNT) is a toxic and persistent explosive compound occurring as a contaminant at numerous sites worldwide. Knowledge of the microbial dynamics driving TNT biodegradation is limited, particularly in native aquifer sediments where it poses a threat to water resources. The purpose of this study was to quantify the effect of organic amendments on anaerobic TNT biodegradation rate and pathway in an enrichment culture obtained from historically contaminated aquifer sediment and to compare the bacterial community dynamics. TNT readily biodegraded in all microcosms, with the highest biodegradation rate obtained under the lactate amended condition followed by ethanol amended and naturally occurring organic matter (extracted from site sediment) amended conditions. Although a reductive pathway of TNT degradation was observed across all conditions, denaturing gradient gel electrophoresis (DGGE) analysis revealed distinct bacterial community compositions. In all microcosms, Gram-negative γ- or β-Proteobacteria and Gram-positive Negativicutes or Clostridia were observed. A Pseudomonas sp. in particular was observed to be stimulated under all conditions. According to non-metric multidimensional scaling analysis of DGGE profiles, the microcosm communities were most similar to heavily TNT-contaminated field site sediment, relative to moderately and uncontaminated sediments, suggesting that TNT contamination itself is a major driver of microbial community structure. Overall these results provide a new line of evidence of the key bacteria driving TNT degradation in aquifer sediments and their dynamics in response to organic carbon amendment, supporting this approach as a promising technology for stimulating in situ TNT bioremediation in the subsurface.

Identification of the bacterial etiology of culture-negative endocarditis by amplification and sequencing of a small ribosomal RNA gene.

PubMed

Khulordava, Irakli; Miller, Geraldine; Haas, David; Li, Haijing; McKinsey, Joel; Vanderende, Daniel; Tang, Yi-Wei

2003-05-01

We report two cases of culture-negative bacterial endocarditis in which the organisms were identified by amplification and sequencing of the bacterial 16S rRNA gene. These results support an important role for polymerase chain reaction followed by direct sequencing to determine the etiology of culture-negative bacterial endocarditis and to guide appropriate antimicrobial therapy.

Local environmental pollution strongly influences culturable bacterial aerosols at an urban aquatic superfund site.

PubMed

Dueker, M Elias; O'Mullan, Gregory D; Juhl, Andrew R; Weathers, Kathleen C; Uriarte, Maria

2012-10-16

In polluted environments, when microbial aerosols originate locally, species composition of the aerosols should reflect the polluted source. To test the connection between local environmental pollution and microbial aerosols near an urban waterfront, we characterized bacterial aerosols at Newtown Creek (NTC), a public waterway and Superfund site in a densely populated area of New York, NY, USA. Culturable bacterial aerosol fallout rate and surface water bacterial concentrations were at least an order of magnitude greater at NTC than at a neighboring, less polluted waterfront and a nonurban coastal site in Maine. The NTC culturable bacterial aerosol community was significantly different in taxonomic structure from previous urban and coastal aerosol studies, particularly in relative abundances of Actinobacteria and Proteobacteria. Twenty-four percent of the operational taxonomic units in the NTC overall (air + water) bacterial isolate library were most similar to bacterial 16S rRNA gene sequences previously described in terrestrial or aquatic environments contaminated with sewage, hydrocarbons, heavy metals, and other industrial waste. This study is the first to examine the community composition and local deposition of bacterial aerosols from an aquatic Superfund site. The findings have important implications for the use of aeration remediation in polluted aquatic environments and suggest a novel pathway of microbial exposure in densely populated urban communities containing contaminated soil and water.

N-acyl homoserine lactone-degrading microbial enrichment cultures isolated from Penaeus vannamei shrimp gut and their probiotic properties in Brachionus plicatilis cultures.

PubMed

Tinh, Nguyen Thi Ngoc; Asanka Gunasekara, R A Y S; Boon, Nico; Dierckens, Kristof; Sorgeloos, Patrick; Bossier, Peter

2007-10-01

Three bacterial enrichment cultures (ECs) were isolated from the digestive tract of Pacific white shrimp Penaeus vannamei, by growing the shrimp microbial communities in a mixture of N-acyl homoserine lactone (AHL) molecules. The ECs, characterized by denaturing gradient gel electrophoresis analysis and subsequent rRNA sequencing, degraded AHL molecules in the degradation assays. Apparently, the resting cells of the ECs also degraded one of the three types of quorum-sensing signal molecules produced by Vibrio harveyi in vitro [i.e. harveyi autoinducer 1 (HAI-1)]. The most efficient AHL-degrading ECs, EC5, was tested in Brachionus experiments. EC5 degraded the V. harveyi HAI-1 autoinducer in vivo, neutralizing the negative effect of V. harveyi autoinducer 2 (AI-2) mutant, in which only the HAI-1- and CAI-1-mediated components of the quorum-sensing system are functional on the growth of Brachionus. This suggests that EC5 interferes with HAI-1-regulated metabolism in V. harveyi. These AHL-degrading ECs need to be tested in other aquatic systems for their probiotic properties, preferably in combination with specific AI-2-degrading bacteria.

Vaginal lactobacilli inhibiting growth of Gardnerella vaginalis, Mobiluncus and other bacterial species cultured from vaginal content of women with bacterial vaginosis.

PubMed

Skarin, A; Sylwan, J

1986-12-01

On a solid agar medium the growth-inhibitory effect of 9 Lactobacillus strains cultured from vaginal content was tested on bacteria cultured from vaginal content of women with bacterial vaginosis: Mobiluncus, Gardnerella vaginalis, Bacteroides and anaerobic cocci. Inhibition zones were observed in the growth of all of the strains isolated from women with bacterial vaginosis around all lactobacilli. The inhibitory effect of the lactobacilli was further tested on various anaerobic and facultatively anaerobic species, both type strains and fresh extragenitally cultured strains. Four Bacteroides fragilis strains as well as 2 out of 4 Staphylococcus aureus strains were clearly inhibited by the lactobacilli. The inhibition zones were generally wider at pH 5.5 than at 6.0. For all inhibited strains, (the S. aureus excepted) a low pH on the agar around the lactobacilli correlated to wider growth-inhibition zones.

Innovative Approaches Using Lichen Enriched Media to Improve Isolation and Culturability of Lichen Associated Bacteria

PubMed Central

Biosca, Elena G.; Flores, Raquel; Santander, Ricardo D.; Díez-Gil, José Luis; Barreno, Eva

2016-01-01

Lichens, self-supporting mutualistic associations between a fungal partner and one or more photosynthetic partners, also harbor non-photosynthetic bacteria. The diversity and contribution of these bacteria to the functioning of lichen symbiosis have recently begun to be studied, often by culture-independent techniques due to difficulties in their isolation and culture. However, culturing as yet unculturable lichenic bacteria is critical to unravel their potential functional roles in lichen symbiogenesis, to explore and exploit their biotechnological potential and for the description of new taxa. Our objective was to improve the recovery of lichen associated bacteria by developing novel isolation and culture approaches, initially using the lichen Pseudevernia furfuracea. We evaluated the effect of newly developed media enriched with novel lichen extracts, as well as the influence of thalli washing time and different disinfection and processing protocols of thalli. The developed methodology included: i) the use of lichen enriched media to mimic lichen nutrients, supplemented with the fungicide natamycin; ii) an extended washing of thalli to increase the recovery of ectolichenic bacteria, thus allowing the disinfection of thalli to be discarded, hence enhancing endolichenic bacteria recovery; and iii) the use of an antioxidant buffer to prevent or reduce oxidative stress during thalli disruption. The optimized methodology allowed significant increases in the number and diversity of culturable bacteria associated with P. furfuracea, and it was also successfully applied to the lichens Ramalina farinacea and Parmotrema pseudotinctorum. Furthermore, we provide, for the first time, data on the abundance of culturable ecto- and endolichenic bacteria that naturally colonize P. furfuracea, R. farinacea and P. pseudotinctorum, some of which were only able to grow on lichen enriched media. This innovative methodology is also applicable to other microorganisms inhabiting these

Jellyfish modulate bacterial dynamic and community structure.

PubMed

Tinta, Tinkara; Kogovšek, Tjaša; Malej, Alenka; Turk, Valentina

2012-01-01

Jellyfish blooms have increased in coastal areas around the world and the outbreaks have become longer and more frequent over the past few decades. The Mediterranean Sea is among the heavily affected regions and the common bloom-forming taxa are scyphozoans Aurelia aurita s.l., Pelagia noctiluca, and Rhizostoma pulmo. Jellyfish have few natural predators, therefore their carcasses at the termination of a bloom represent an organic-rich substrate that supports rapid bacterial growth, and may have a large impact on the surrounding environment. The focus of this study was to explore whether jellyfish substrate have an impact on bacterial community phylotype selection. We conducted in situ jellyfish-enrichment experiment with three different jellyfish species. Bacterial dynamic together with nutrients were monitored to assess decaying jellyfish-bacteria dynamics. Our results show that jellyfish biomass is characterized by protein rich organic matter, which is highly bioavailable to 'jellyfish-associated' and 'free-living' bacteria, and triggers rapid shifts in bacterial population dynamics and composition. Based on 16S rRNA clone libraries and denaturing gradient gel electrophoresis (DGGE) analysis, we observed a rapid shift in community composition from unculturable Alphaproteobacteria to culturable species of Gammaproteobacteria and Flavobacteria. The results of sequence analyses of bacterial isolates and of total bacterial community determined by culture independent genetic analysis showed the dominance of the Pseudoalteromonadaceae and the Vibrionaceae families. Elevated levels of dissolved proteins, dissolved organic and inorganic nutrient release, bacterial abundance and carbon production as well as ammonium concentrations characterized the degradation process. The biochemical composition of jellyfish species may influence changes in the amount of accumulated dissolved organic and inorganic nutrients. Our results can contribute insights into possible changes in

Antarctic ice core samples: culturable bacterial diversity.

PubMed

Shivaji, Sisinthy; Begum, Zareena; Shiva Nageswara Rao, Singireesu Soma; Vishnu Vardhan Reddy, Puram V; Manasa, Poorna; Sailaja, Buddi; Prathiba, Mambatta S; Thamban, Meloth; Krishnan, Kottekkatu P; Singh, Shiv M; Srinivas, Tanuku N R

2013-01-01

Culturable bacterial abundance at 11 different depths of a 50.26 m ice core from the Tallaksenvarden Nunatak, Antarctica, varied from 0.02 to 5.8 × 10(3) CFU ml(-1) of the melt water. A total of 138 bacterial strains were recovered from the 11 different depths of the ice core. Based on 16S rRNA gene sequence analyses, the 138 isolates could be categorized into 25 phylotypes belonging to phyla Actinobacteria, Bacteroidetes, Firmicutes and Proteobacteria. All isolates had 16S rRNA sequences similar to previously determined sequences (97.2-100%). No correlation was observed in the distribution of the isolates at the various depths either at the phylum, genus or species level. The 25 phylotypes varied in growth temperature range, tolerance to NaCl, growth pH range and ability to produce eight different extracellular enzymes at either 4 or 18 °C. Iso-, anteiso-, unsaturated and saturated fatty acids together constituted a significant proportion of the total fatty acid composition. Copyright © 2012 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

Oil removal from petroleum sludge using bacterial culture with molasses substrate at temperature variation

NASA Astrophysics Data System (ADS)

Ni'matuzahroh, Puspitasari, Alvin Oktaviana; Pratiwi, Intan Ayu; Fatimah, Sumarsih, Sri; Surtiningsih, Tini; Salamun

2016-03-01

The study aims to reveal the potency of biosurfactant-producing bacterial culture with molasses as substrate growth in releasing oil from the petroleum sludge at temperature variations. Bacteria used consisted of (Acinetobacter sp. P2(1), Pseudomonas putida T1(8), Bacillus subtilis 3KP and Micrococcus sp. L II 61). The treatments were tested at 40°C, 50°C and 60 °C for 7 days of incubation. Synthetic surfactant (Tween 20) was used as a positive control and molasses as a negative control. Release of petroleum hydrocarbons from oil sludge was expressed in percentage of oil removal from oil sludge (%). Data were analyzed statistically using the Analysis of Variance (α = 0.05) and continued with Games-Howell test. The kinds of bacterial cultures, incubation temperature and combination of both affected the percentage of oil removal. The abilities of Bacillus subtilis 3KP and Micrococcus sp. LII 61cultures in oil removal from oil sludge at the temperature exposure of 60°C were higher than Tween 20. Both of bacterial cultures grown on molasses can be proposed as a replacement for synthetic surfactant to clean up the accumulation of oil sludge in a bottom of oil refinery tank.

In vitro effect of Reiki treatment on bacterial cultures: Role of experimental context and practitioner well-being.

PubMed

Rubik, Beverly; Brooks, Audrey J; Schwartz, Gary E

2006-01-01

To measure effects of Reiki treatments on growth of heat-shocked bacteria, and to determine the influence of healing context and practitioner well-being. Overnight cultures of Escherichia coli K12 in fresh medium were used. Culture samples were paired with controls to minimize any ordering effects. Samples were heat-shocked prior to Reiki treatment, which was performed by Reiki practitioners for up to 15 minutes, with untreated controls. Plate-count assay using an automated colony counter determined the number of viable bacteria. Fourteen Reiki practitioners each completed 3 runs (n = 42 runs) without healing context, and another 2 runs (n = 28 runs) in which they first treated a pain patient for 30 minutes (healing context). Well-being questionnaires were administered to practitioners pre-post all sessions. No overall difference was found between the Reiki and control plates in the nonhealing context. In the healing context, the Reiki treated cultures overall exhibited significantly more bacteria than controls (p < 0.05). Practitioner social (p < 0.013) and emotional well-being (p < 0.021) correlated with Reiki treatment outcome on bacterial cultures in the nonhealing context. Practitioner social (p < 0.031), physical (p < 0.030), and emotional (p < 0.026) well-being correlated with Reiki treatment outcome on the bacterial cultures in the healing context. For practitioners starting with diminished well-being, control counts were likely to be higher than Reiki-treated bacterial counts. For practitioners starting with a higher level of well-being, Reiki counts were likely to be higher than control counts. Reiki improved growth of heat-shocked bacterial cultures in a healing context. The initial level of well-being of the Reiki practitioners correlates with the outcome of Reiki on bacterial culture growth and is key to the results obtained.

Using Enrichment Clusters to Address the Needs of Culturally and Linguistically Diverse Learners

ERIC Educational Resources Information Center

Allen, Jennifer K.; Robbins, Margaret A.; Payne, Yolanda Denise; Brown, Katherine Backes

2016-01-01

Using data from teacher interviews, classroom observations, and a professional development workshop, this article explains how one component of the schoolwide enrichment model (SEM) has been implemented at a culturally diverse elementary school serving primarily Latina/o and African American students. Based on a broadened conception of giftedness,…

Testosterone-mineralizing culture enriched from swine manure: characterization of degradation pathways and microbial community composition.

PubMed

Yang, Yun-Ya; Pereyra, Luciana P; Young, Robert B; Reardon, Kenneth F; Borch, Thomas

2011-08-15

Environmental releases and fate of steroid sex hormones from livestock and wastewater treatment plants are of increasing regulatory concern. Despite the detection of these hormones in manures, biosolids, and the environment, little attention has been paid to characterization of fecal bacteria capable of hormone degradation. The enrichments of (swine) manure-borne bacteria capable of aerobic testosterone degradation were prepared and the testosterone mineralization pathway was elucidated. Six DNA sequences of bacteria from the Proteobacteria phylum distributed among the genera Acinetobacter, Brevundimonas, Comamonas, Sphingomonas, Stenotrophomonas, and Rhodobacter were identified in a testosterone-degrading enriched culture with testosterone as the sole carbon source. Three degradation products of testosterone were identified as androstenedione, androstadienedione, and dehydrotestosterone using commercially available reference standards, liquid chromatography-UV diode array detection, and liquid chromatography-time-of-flight mass spectrometry (LC-TOF/MS). Three additional degradation products of testosterone were tentatively identified as 9α-hydroxytestosterone, 9α-hydroxyandrostadienedione or 3-hydroxy-9,10-secoandrosta-1,3,5(10)-triene-9,17-dione, and 9α-hydroxydehydrotestosterone or 9α-hydroxyandrostenedione using LC-TOF/MS. When (14)C-testosterone was introduced to the enriched culture, 49-68% of the added (14)C-testosterone was mineralized to (14)CO(2) within 8 days of incubation. The mineralization of (14)C-testosterone followed pseudo-first-order reaction kinetics in the enriched culture with half-lives (t(1/2)) of 10-143 h. This work suggests that Proteobacteria play an important environmental role in degradation of steroid sex hormones and that androgens have the potential to be mineralized during aerobic manure treatment or after land application to agricultural fields by manure-borne bacteria.

Characterization of Cultures Enriched from Acidic Polycyclic Aromatic Hydrocarbon-Contaminated Soil for Growth on Pyrene at Low pH▿

PubMed Central

Uyttebroek, Maarten; Vermeir, Steven; Wattiau, Pierre; Ryngaert, Annemie; Springael, Dirk

2007-01-01

Two polycyclic aromatic hydrocarbon (PAH)-contaminated soils of pH 2 were successfully used as inoculum to enrich cultures growing on phenanthrene and pyrene at different pHs, including pH 3. Selected pyrene-utilizing cultures obtained at pH 3, pH 5, and pH 7 were further characterized. All showed rapid [14C]pyrene mineralization at pH 3 and pH 5 and grew on pyrene at pH values ranging from 2 to 6. Eubacterial and mycobacterial 16S rRNA gene denaturing gradient gel electrophoresis fingerprinting and sequencing indicated that the cultures were dominated by a single bacterium closely related to Mycobacterium montefiorense, belonging to the slow-growing Mycobacterium sp. In contrast, a culture enriched on pyrene at pH 7 from a slightly alkaline soil sampled at the same site was dominated by Pseudomonas putida and a fast-growing Mycobacterium sp. The M. montefiorense-related species dominating the pyrene-utilizing cultures enriched from the acidic soils was also the dominant Mycobacterium species in the acidic soils. Our data indicate that a slow-growing Mycobacterium species is involved in PAH degradation in that culture and show that bacteria able to degrade high-molecular-weight PAHs at low pH are present in acidic PAH-contaminated soil. PMID:17369339

The importance of the viable but non-culturable state in human bacterial pathogens

PubMed Central

Li, Laam; Mendis, Nilmini; Trigui, Hana; Oliver, James D.; Faucher, Sebastien P.

2014-01-01

Many bacterial species have been found to exist in a viable but non-culturable (VBNC) state since its discovery in 1982. VBNC cells are characterized by a loss of culturability on routine agar, which impairs their detection by conventional plate count techniques. This leads to an underestimation of total viable cells in environmental or clinical samples, and thus poses a risk to public health. In this review, we present recent findings on the VBNC state of human bacterial pathogens. The characteristics of VBNC cells, including the similarities and differences to viable, culturable cells and dead cells, and different detection methods are discussed. Exposure to various stresses can induce the VBNC state, and VBNC cells may be resuscitated back to culturable cells under suitable stimuli. The conditions that trigger the induction of the VBNC state and resuscitation from it are summarized and the mechanisms underlying these two processes are discussed. Last but not least, the significance of VBNC cells and their potential influence on human health are also reviewed. PMID:24917854

Bacterial Diversity Associated with Wild Caught Anopheles Mosquitoes from Dak Nong Province, Vietnam Using Culture and DNA Fingerprint

PubMed Central

Ngo, Chung Thuy; Aujoulat, Fabien; Veas, Francisco; Jumas-Bilak, Estelle; Manguin, Sylvie

2015-01-01

Background Microbiota of Anopheles midgut can modulate vector immunity and block Plasmodium development. Investigation on the bacterial biodiversity in Anopheles, and specifically on the identification of bacteria that might be used in malaria transmission blocking approaches, has been mainly conducted on malaria vectors of Africa. Vietnam is an endemic country for both malaria and Bancroftian filariasis whose parasitic agents can be transmitted by the same Anopheles species. No information on the microbiota of Anopheles mosquitoes in Vietnam was available previous to this study. Method The culture dependent approach, using different mediums, and culture independent (16S rRNA PCR – TTGE) method were used to investigate the bacterial biodiversity in the abdomen of 5 Anopheles species collected from Dak Nong Province, central-south Vietnam. Molecular methods, sequencing and phylogenetic analysis were used to characterize the microbiota. Results and Discussion The microbiota in wild-caught Anopheles was diverse with the presence of 47 bacterial OTUs belonging to 30 genera, including bacterial genera impacting Plasmodium development. The bacteria were affiliated with 4 phyla, Actinobacteria, Bacteroidetes, Firmicutes and Proteobacteria, the latter being the dominant phylum. Four bacterial genera are newly described in Anopheles mosquitoes including Coxiella, Yersinia, Xanthomonas, and Knoellia. The bacterial diversity per specimen was low ranging from 1 to 4. The results show the importance of pairing culture and fingerprint methods to better screen the bacterial community in Anopheles mosquitoes. Conclusion Sampled Anopheles species from central-south Vietnam contained a diverse bacterial microbiota that needs to be investigated further in order to develop new malaria control approaches. The combination of both culture and DNA fingerprint methods allowed a thorough and complementary screening of the bacterial community in Anopheles mosquitoes. PMID:25747513

Predominance of Viable Spore-Forming Piezophilic Bacteria in High-Pressure Enrichment Cultures from ~1.5 to 2.4 km-Deep Coal-Bearing Sediments below the Ocean Floor

PubMed Central

Fang, Jiasong; Kato, Chiaki; Runko, Gabriella M.; Nogi, Yuichi; Hori, Tomoyuki; Li, Jiangtao; Morono, Yuki; Inagaki, Fumio

2017-01-01

Phylogenetically diverse microorganisms have been observed in marine subsurface sediments down to ~2.5 km below the seafloor (kmbsf). However, very little is known about the pressure-adapted and/or pressure-loving microorganisms, the so called piezophiles, in the deep subseafloor biosphere, despite that pressure directly affects microbial physiology, metabolism, and biogeochemical processes of carbon and other elements in situ. In this study, we studied taxonomic compositions of microbial communities in high-pressure incubated sediment, obtained during the Integrated Ocean Drilling Program (IODP) Expedition 337 off the Shimokita Peninsula, Japan. Analysis of 16S rRNA gene-tagged sequences showed that members of spore-forming bacteria within Firmicutes and Actinobacteria were predominantly detected in all enrichment cultures from ~1.5 to 2.4 km-deep sediment samples, followed by members of Proteobacteria, Acidobacteria, and Bacteroidetes according to the sequence frequency. To further study the physiology of the deep subseafloor sedimentary piezophilic bacteria, we isolated and characterized two bacterial strains, 19R1-5 and 29R7-12, from 1.9 and 2.4 km-deep sediment samples, respectively. The isolates were both low G+C content, gram-positive, endospore-forming and facultative anaerobic piezophilic bacteria, closely related to Virgibacillus pantothenticus and Bacillus subtilis within the phylum Firmicutes, respectively. The optimal pressure and temperature conditions for growth were 20 MPa and 42°C for strain 19R1-5, and 10 MPa and 43°C for strain 29R7-12. Bacterial (endo)spores were observed in both the enrichment and pure cultures examined, suggesting that these piezophilic members were derived from microbial communities buried in the ~20 million-year-old coal-bearing sediments after the long-term survival as spores and that the deep biosphere may host more abundant gram-positive spore-forming bacteria and their spores than hitherto recognized. PMID:28220112

Predominance of Viable Spore-Forming Piezophilic Bacteria in High-Pressure Enrichment Cultures from ~1.5 to 2.4 km-Deep Coal-Bearing Sediments below the Ocean Floor.

PubMed

Fang, Jiasong; Kato, Chiaki; Runko, Gabriella M; Nogi, Yuichi; Hori, Tomoyuki; Li, Jiangtao; Morono, Yuki; Inagaki, Fumio

2017-01-01

Phylogenetically diverse microorganisms have been observed in marine subsurface sediments down to ~2.5 km below the seafloor (kmbsf). However, very little is known about the pressure-adapted and/or pressure-loving microorganisms, the so called piezophiles, in the deep subseafloor biosphere, despite that pressure directly affects microbial physiology, metabolism, and biogeochemical processes of carbon and other elements in situ . In this study, we studied taxonomic compositions of microbial communities in high-pressure incubated sediment, obtained during the Integrated Ocean Drilling Program (IODP) Expedition 337 off the Shimokita Peninsula, Japan. Analysis of 16S rRNA gene-tagged sequences showed that members of spore-forming bacteria within Firmicutes and Actinobacteria were predominantly detected in all enrichment cultures from ~1.5 to 2.4 km-deep sediment samples, followed by members of Proteobacteria, Acidobacteria, and Bacteroidetes according to the sequence frequency. To further study the physiology of the deep subseafloor sedimentary piezophilic bacteria, we isolated and characterized two bacterial strains, 19R1-5 and 29R7-12, from 1.9 and 2.4 km-deep sediment samples, respectively. The isolates were both low G+C content, gram-positive, endospore-forming and facultative anaerobic piezophilic bacteria, closely related to Virgibacillus pantothenticus and Bacillus subtilis within the phylum Firmicutes, respectively. The optimal pressure and temperature conditions for growth were 20 MPa and 42°C for strain 19R1-5, and 10 MPa and 43°C for strain 29R7-12. Bacterial (endo)spores were observed in both the enrichment and pure cultures examined, suggesting that these piezophilic members were derived from microbial communities buried in the ~20 million-year-old coal-bearing sediments after the long-term survival as spores and that the deep biosphere may host more abundant gram-positive spore-forming bacteria and their spores than hitherto recognized.

Effect of inoculum sources on the enrichment of nitrite-dependent anaerobic methane-oxidizing bacteria.

PubMed

He, Zhanfei; Cai, Chen; Shen, Lidong; Lou, Liping; Zheng, Ping; Xu, Xinhua; Hu, Baolan

2015-01-01

Nitrite-dependent anaerobic methane oxidation (n-damo) is a newly discovered biological process that couples anaerobic oxidation of methane (AOM) to nitrite reduction. In this study, three different inocula, methanogenic sludge, paddy soil, and freshwater sediment were used to enrich n-damo bacteria in three sequencing batch reactors (SBRs), and three n-damo enrichment cultures, C1, C2 and C3, were obtained, respectively. After 500 days of incubation, Methylomirabilis oxyfera-like bacteria and n-damo activities were observed in cultures C1, C2, and C3, and the specific activities were 0.8 ± 0.1, 1.4 ± 0.1, and 1.0 ± 0.1 μmol CH4 h(-1) g(-1) VSS, respectively. The copy numbers of 16S rRNA genes from cultures C1, C2, and C3 were 5.0 ± 0.4 × 10(8), 6.1 ± 0.1 × 10(9), and 1.0 ± 0.2 × 10(9) copies g(-1) dry weight, respectively. The results indicated that paddy soil is an excellent inoculum for n-damo bacterial enrichment. This work expanded the alternative source of n-damo inoculum and benefited the further research of n-damo process.

Comparative analysis of metagenomes from three methanogenic hydrocarbon-degrading enrichment cultures with 41 environmental samples.

PubMed

Tan, Boonfei; Fowler, S Jane; Abu Laban, Nidal; Dong, Xiaoli; Sensen, Christoph W; Foght, Julia; Gieg, Lisa M

2015-09-01

Methanogenic hydrocarbon metabolism is a key process in subsurface oil reservoirs and hydrocarbon-contaminated environments and thus warrants greater understanding to improve current technologies for fossil fuel extraction and bioremediation. In this study, three hydrocarbon-degrading methanogenic cultures established from two geographically distinct environments and incubated with different hydrocarbon substrates (added as single hydrocarbons or as mixtures) were subjected to metagenomic and 16S rRNA gene pyrosequencing to test whether these differences affect the genetic potential and composition of the communities. Enrichment of different putative hydrocarbon-degrading bacteria in each culture appeared to be substrate dependent, though all cultures contained both acetate- and H2-utilizing methanogens. Despite differing hydrocarbon substrates and inoculum sources, all three cultures harbored genes for hydrocarbon activation by fumarate addition (bssA, assA, nmsA) and carboxylation (abcA, ancA), along with those for associated downstream pathways (bbs, bcr, bam), though the cultures incubated with hydrocarbon mixtures contained a broader diversity of fumarate addition genes. A comparative metagenomic analysis of the three cultures showed that they were functionally redundant despite their enrichment backgrounds, sharing multiple features associated with syntrophic hydrocarbon conversion to methane. In addition, a comparative analysis of the culture metagenomes with those of 41 environmental samples (containing varying proportions of methanogens) showed that the three cultures were functionally most similar to each other but distinct from other environments, including hydrocarbon-impacted environments (for example, oil sands tailings ponds and oil-affected marine sediments). This study provides a basis for understanding key functions and environmental selection in methanogenic hydrocarbon-associated communities.

Methanogenic degradation of acetone by an enrichment culture.

PubMed

Platen, H; Schink, B

1987-01-01

An anaerobic enrichment culture degraded 1 mol of acetone to 2 mol of methane and 1 mol of carbon dioxide. Two microorganisms were involved in this process, a filament-forming rod similar to Methanothrix sp. and an unknown rod with round to slightly pointed ends. Both organisms formed aggregates up to 300 micron in diameter. No fluorescing bacteria were observed indicating that hydrogen or formate-utilizing methanogens are not involved in this process. Acetate was utilized in this culture by the Methanothrix sp. Inhibition of methanogenesis by bromoethanesulfonic acid or acetylene decreased the acetone degradation rate drastically and led to the formation of 2 mol acetate per mol of acetone. Streptomycin completely inhibited acetone degradation, and neither acetate nor methane was formed. 14CO2 was incorporated exclusively into the C-1 atom of acetate indicating that acetone is degraded via carboxylation to an acetoacetate residue. It is concluded that acetone is degraded by a coculture of an eubacterium and an acetate-utilizing methanogen and that acetate is the only intermediate transferred between both. The energetical problems of the eubacterium converting acetone to acetate are discussed.

Characterization of wheat straw-degrading anaerobic alkali-tolerant mixed cultures from soda lake sediments by molecular and cultivation techniques

PubMed Central

Porsch, Katharina; Wirth, Balázs; Tóth, Erika M; Schattenberg, Florian; Nikolausz, Marcell

2015-01-01

Alkaline pretreatment has the potential to enhance the anaerobic digestion of lignocellulosic biomass to biogas. However, the elevated pH of the substrate may require alkalitolerant microbial communities for an effective digestion. Three mixed anaerobic lignocellulolytic cultures were enriched from sediments from two soda lakes with wheat straw as substrate under alkaline (pH 9) mesophilic (37°C) and thermophilic (55°C) conditions. The gas production of the three cultures ceased after 4 to 5 weeks, and the produced gas was composed of carbon dioxide and methane. The main liquid intermediates were acetate and propionate. The physiological behavior of the cultures was stable even after several transfers. The enrichment process was also followed by molecular fingerprinting (terminal restriction fragment length polymorphism) of the bacterial 16S rRNA gene and of the mcrA/mrtA functional gene for methanogens. The main shift in the microbial community composition occurred between the sediment samples and the first enrichment, whereas the structure was stable in the following transfers. The bacterial communities mainly consisted of Sphingobacteriales, Clostridiales and Spirochaeta, but differed at genus level. Methanothermobacter and Methanosarcina genera and the order Methanomicrobiales were predominant methanogenes in the obtained cultures. Additionally, single cellulolytic microorganisms were isolated from enrichment cultures and identified as members of the alkaliphilic or alkalitolerant genera. The results show that anaerobic alkaline habitats harbor diverse microbial communities, which can degrade lignocellulose effectively and are therefore a potential resource for improving anaerobic digestion. PMID:25737100

Diagnostic utility and cost-effectiveness of reflex bacterial culture for the detection of urinary tract infection in dogs with low urine specific gravity.

PubMed

Tivapasi, Musavenga T; Hodges, Joanne; Byrne, Barbara A; Christopher, Mary M

2009-09-01

Urinary tract infections (UTIs) may be subclinical or difficult to detect in dilute urine as sediment abnormalities may not be observed. In our laboratory, bacterial culture is automatically performed (reflex culture) on samples with urine specific gravity (USG)< or =1.013 to increase the likelihood of detecting infection. The value of routine culture of dilute urine, however, has not been fully assessed. The purpose of this retrospective study was to evaluate the frequency of positive bacterial cultures and analyze the diagnostic utility and cost-effectiveness of culture compared with routine sediment examination for detecting UTI in dilute urine specimens from dogs. Urinalysis and concurrent aerobic bacterial culture results were obtained from the electronic medical record system at the University of California-Davis Veterinary Medical Teaching Hospital for samples with USG< or =1.013 analyzed from July 1998 through January 2005. Urine collection method, presence of leukocytes and bacteria, bacterial culture results, and clinical diagnosis were recorded. Cost-effectiveness of reflex culture, based on low USG as the sole criterion, was evaluated. Of 1264 urine specimens, 106 (8.4%) had positive bacterial cultures. Using culture as the gold standard, sediment evaluation had a diagnostic sensitivity of 58.5% and specificity of 98.3% (diagnostic accuracy 94.9%). An additional cost of $60 per patient was incurred, leading to average annual costs of $11,668 for reflex bacterial cultures of all samples with low USG, regardless of collection method. Within our study population, 10 urine samples needed to be cultured for each true positive result. The sensitivity of urine sediment evaluation is low for UTI in dilute urine samples; however, reflex bacterial culture does not appear to be cost-effective in dogs with USG< or =1.013 in the absence of active urine sediment or high clinical suspicion for UTI.

Nitrogen source effects on the denitrifying anaerobic methane oxidation culture and anaerobic ammonium oxidation bacteria enrichment process.

PubMed

Fu, Liang; Ding, Jing; Lu, Yong-Ze; Ding, Zhao-Wei; Zeng, Raymond J

2017-05-01

The co-culture system of denitrifying anaerobic methane oxidation (DAMO) and anaerobic ammonium oxidation (Anammox) has a potential application in wastewater treatment plant. This study explored the effects of permutation and combination of nitrate, nitrite, and ammonium on the culture enrichment from freshwater sediments. The co-existence of NO 3 - , NO 2 - , and NH 4 + shortened the enrichment time from 75 to 30 days and achieved a total nitrogen removal rate of 106.5 mg/L/day on day 132. Even though ammonium addition led to Anammox bacteria increase and a higher nitrogen removal rate, DAMO bacteria still dominated in different reactors with the highest proportion of 64.7% and the maximum abundance was 3.07 ± 0.25 × 10 8 copies/L (increased by five orders of magnitude) in the nitrite reactor. DAMO bacteria showed greater diversity in the nitrate reactor, and one was similar to M. oxyfera; DAMO bacteria in the nitrite reactor were relatively unified and similar to M. sinica. Interestingly, no DAMO archaea were found in the nitrate reactor. This study will improve the understanding of the impact of nitrogen source on DAMO and Anammox co-culture enrichment.

Enrichment and identification of cellulolytic bacteria from the gastrointestinal tract of Giant African snail, Achatina fulica.

PubMed

Pawar, Kiran D; Dar, Mudasir A; Rajput, Bharati P; Kulkarni, Girish J

2015-02-01

The cellulolytic bacterial community structure in gastrointestinal (GI) tract of Achatina fulica was studied using culture-independent and -dependent methods by enrichment in carboxymethyl cellulose (CMC). Culture-dependent method indicated that GI tract of snail was dominated by Enterobacteriaceae members. When tested for cellulase activities, all isolates obtained by culture-dependent method showed both or either of CMCase or avicelase activity. Isolate identified as Citrobacter freundii showed highest CMCase and medium avicelase activity. Sequencing of clones from the 16S rRNA gene clone library identified ten operational taxonomic units (OTUs), which were affiliated to Enterobacteriaceae of phylum Gammaproteobacteria. Of these ten OTUs, eight OTUs closely matched with Enterobacter and Klebsiella genera. The most abundant OTU allied to Klebsiella oxytoca accounted for 70 % of the total sequences. The members of Klebsiella and Enterobacter were observed by both methods indicating their dominance among the cellulolytic bacterial community in the GI tract of the snail.

Faculty Use of Culturally Mediated Instruction in a Community College Academic Enrichment Program

ERIC Educational Resources Information Center

Lacey, Charna L.

2009-01-01

The purpose of this study was to examine faculty use of Culturally Mediated Instructional (CMI) practices in a community college-based academic enrichment program. The intent of the study was two-fold: (a) to search for evidence that instructional practices were reflective of Hollins' (1996) theory of CMI, and (b) to explore faculty perceptions of…

The Learning Effects of an Ecology Enrichment Summer Program on Gifted Students from Mainstream and Diverse Cultural Backgrounds: A Preliminary Study

ERIC Educational Resources Information Center

Wang, Wen-Ling; Wu, Jiun-Wei; Lin, Yu-Chin

2006-01-01

Enrichment is one of the important educational models for gifted students. However, the research on gifted enrichment programs rarely leads to instructional interventions for culturally diverse students. The purposes of this study were: (a) to propose an ecology enrichment summer program for gifted students from mainstream and diverse cultural…

Effects of space flight and mixing on bacterial growth in low volume cultures

NASA Technical Reports Server (NTRS)

Kacena, M. A.; Manfredi, B.; Todd, P.

1999-01-01

Previous investigations have shown that liquid suspension bacterial cultures grow to higher cell concentrations in spaceflight than on Earth. None of these studies included ground-control experiments designed to evaluate the fluid effects potentially responsible for the reported increases. Therefore, the emphasis of this research was to both confirm differences in final cell concentration between 1g and microgravity cultures, and to examine the effects of mixing as a partial explanation for this difference. Flight experiments were performed in the Fluid Processing Apparatus (FPA), aboard Space Shuttle Missions STS-63 and STS-69, with simultaneous 1g static and agitated controls. Additional static 1g, agitated, and clino-rotated controls were performed in 9-ml culture tubes. This research revealed that both E. coli and B. subtilis samples cultured in space flight grew to higher final cell densities (120-345% increase) than simultaneous static 1g controls. The final cell concentration of E. coli cells cultured under agitation was 43% higher than in static 1g cultures and was 102% higher with clino-rotation. However, for B. subtilis cultures grown while being agitated on a shaker or clino-rotated, the final cell concentrations were nearly identical to those of the simultaneous static 1g controls. Therefore, these data suggest that the unique fluid quiescence in the microgravity environment (lack of sedimentation, creating unique transfer of nutrients and waste products), was responsible for the enhanced bacterial proliferation reported in this and other studies.

Enrichment of prostate cancer stem cells from primary prostate cancer cultures of biopsy samples

PubMed Central

Wang, Shunqi; Huang, Shengsong; Zhao, Xin; Zhang, Qimin; Wu, Min; Sun, Feng; Han, Gang; Wu, Denglong

2014-01-01

This study was to enrich prostate cancer stem cells (PrCSC) from primary prostate cancer cultures (PPrCC). Primary prostate cancer cells were amplified in keratinocyte serum-free medium with epidermal growth factor (EGF) and bovine pituitary extract (BPE), supplemented with leukemia inhibitory factor (LIF), stem cell factor (SCF) and cholera toxin. After amplification, cells were transferred into ultra-low attachment dishes with serum-free DMEM/F12 medium, supplemented with EGF, basic fibroblast growth factor (bFGF), bovine serum albumin (BSA), insulin, and N2 nutrition. Expression of cell-type-specific markers was determined by RT-qPCR and immunostaining. Tumorigenicity of enriched PrCSC was determined by soft agar assay and xenograft assay in NOD/SCID mice. Biopsy samples from 19 confirmed prostate cancer patients were used for establishing PPrCC, and 18 cases (95%) succeeded. Both basal marker (CK5) and luminal markers (androgen receptor and CK8) strongly co-expressed in most of PPrCC, indicating their basal epithelial origin. After amplification under adherent culture condition in vitro, transient amplifying cells were the dominant cells. Sphere formation efficiency (SFE) of passaged PPrCC was about 0.5%, which was 27 times lower than SFE of LNCaP (13.67%) in the same condition. Compared with adherent cells from PPrCC, prostasphere from PPrCC showed up regulated stem cell markers and increased tumorigenic potential in soft-agar assay. However, spheroid cells from PPrCC prostasphere failed to initiate tumor in xenograft assay in 6 months. Thus, PPrCC can be established and amplified from prostate cancer biopsy samples. Our modified sphere culture system can enrich PrCSC from PPrCC. PMID:24427338

Agreement between microscopic examination and bacterial culture of bile samples for detection of bactibilia in dogs and cats with hepatobiliary disease.

PubMed

Pashmakova, Medora B; Piccione, Julie; Bishop, Micah A; Nelson, Whitney R; Lawhon, Sara D

2017-05-01

OBJECTIVE To evaluate the agreement between results of microscopic examination and bacterial culture of bile samples from dogs and cats with hepatobiliary disease for detection of bactibilia. DESIGN Cross-sectional study. ANIMALS 31 dogs and 21 cats with hepatobiliary disease for which subsequent microscopic examination and bacterial culture of bile samples was performed from 2004 through 2014. PROCEDURES Electronic medical records of included dogs and cats were reviewed to extract data regarding diagnosis, antimicrobials administered, and results of microscopic examination and bacterial culture of bile samples. Agreement between these 2 diagnostic tests was assessed by calculation of the Cohen κ value. RESULTS 17 (33%) dogs and cats had bactibilia identified by microscopic examination of bile samples, and 11 (21%) had bactibilia identified via bacterial culture. Agreement between these 2 tests was substantial (percentage agreement [positive and negative results], 85%; κ = 0.62; 95% confidence interval, 0.38 to 0.89) and improved to almost perfect when calculated for only animals that received no antimicrobials within 24 hours prior to sample collection (percentage agreement, 94%; κ = 0.84; 95% confidence interval, 0.61 to 1.00). CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that agreement between microscopic examination and bacterial culture of bile samples for detection of bactibilia is optimized when dogs and cats are not receiving antimicrobials at the time of sample collection. Concurrent bacterial culture and microscopic examination of bile samples are recommended for all cats and dogs evaluated for hepatobiliary disease.

Combination of culture-independent and culture-dependent molecular methods for the determination of bacterial community of iru, a fermented Parkia biglobosa seeds.

PubMed

Adewumi, Gbenga A; Oguntoyinbo, Folarin A; Keisam, Santosh; Romi, Wahengbam; Jeyaram, Kumaraswamy

2012-01-01

In this study, bacterial composition of iru produced by natural, uncontrolled fermentation of Parkia biglobosa seeds was assessed using culture-independent method in combination with culture-based genotypic typing techniques. PCR-denaturing gradient gel electrophoresis (DGGE) revealed similarity in DNA fragments with the two DNA extraction methods used and confirmed bacterial diversity in the 16 iru samples from different production regions. DNA sequencing of the highly variable V3 region of the 16S rRNA genes obtained from PCR-DGGE identified species related to Bacillus subtilis as consistent bacterial species in the fermented samples, while other major bands were identified as close relatives of Staphylococcus vitulinus, Morganella morganii, B. thuringiensis, S. saprophyticus, Tetragenococcus halophilus, Ureibacillus thermosphaericus, Brevibacillus parabrevis, Salinicoccus jeotgali, Brevibacterium sp. and uncultured bacteria clones. Bacillus species were cultured as potential starter cultures and clonal relationship of different isolates determined using amplified ribosomal DNA restriction analysis (ARDRA) combined with 16S-23S rRNA gene internal transcribed spacer (ITS) PCR amplification, restriction analysis (ITS-PCR-RFLP), and randomly amplified polymorphic DNA (RAPD-PCR). This further discriminated B. subtilis and its variants from food-borne pathogens such as B. cereus and suggested the need for development of controlled fermentation processes and good manufacturing practices (GMP) for iru production to achieve product consistency, safety quality, and improved shelf life.

Metagenomic Analyses of the Autotrophic Fe(II)-Oxidizing, Nitrate-Reducing Enrichment Culture KS

PubMed Central

Tominski, Claudia; Kappler, Andreas; Behrens, Sebastian

2016-01-01

Nitrate-dependent ferrous iron [Fe(II)] oxidation (NDFO) is a well-recognized chemolithotrophic pathway in anoxic sediments. The neutrophilic chemolithoautotrophic enrichment culture KS originally obtained from a freshwater sediment (K. L. Straub, M. Benz, B. Schink, and F. Widdel, Appl Environ Microbiol 62:1458–1460, 1996) has been used as a model system to study NDFO. However, the primary Fe(II) oxidizer in this culture has not been isolated, despite extensive efforts to do so. Here, we present a metagenomic analysis of this enrichment culture in order to gain insight into electron transfer pathways and the roles of different bacteria in the culture. We obtained a near-complete genome of the primary Fe(II) oxidizer, a species in the family Gallionellaceae, and draft genomes from its flanking community members. A search of the putative extracellular electron transfer pathways in these genomes led to the identification of a homolog of the MtoAB complex [a porin-multiheme cytochrome c system identified in neutrophilic microaerobic Fe(II)-oxidizing Sideroxydans lithotrophicus ES-1] in a Gallionellaceae sp., and findings of other putative genes involving cytochrome c and multicopper oxidases, such as Cyc2 and OmpB. Genome-enabled metabolic reconstruction revealed that this Gallionellaceae sp. lacks nitric oxide and nitrous oxide reductase genes and may partner with flanking populations capable of complete denitrification to avoid toxic metabolite accumulation, which may explain its resistance to growth in pure culture. This and other revealed interspecies interactions and metabolic interdependencies in nitrogen and carbon metabolisms may allow these organisms to cooperate effectively to achieve robust chemolithoautotrophic NDFO. Overall, the results significantly expand our knowledge of NDFO and suggest a range of genetic targets for further exploration. PMID:26896135

Application of real-time PCR for total airborne bacterial assessment: Comparison with epifluorescence microscopy and culture-dependent methods

NASA Astrophysics Data System (ADS)

Rinsoz, Thomas; Duquenne, Philippe; Greff-Mirguet, Guylaine; Oppliger, Anne

Traditional culture-dependent methods to quantify and identify airborne microorganisms are limited by factors such as short-duration sampling times and inability to count non-culturable or non-viable bacteria. Consequently, the quantitative assessment of bioaerosols is often underestimated. Use of the real-time quantitative polymerase chain reaction (Q-PCR) to quantify bacteria in environmental samples presents an alternative method, which should overcome this problem. The aim of this study was to evaluate the performance of a real-time Q-PCR assay as a simple and reliable way to quantify the airborne bacterial load within poultry houses and sewage treatment plants, in comparison with epifluorescence microscopy and culture-dependent methods. The estimates of bacterial load that we obtained from real-time PCR and epifluorescence methods, are comparable, however, our analysis of sewage treatment plants indicate these methods give values 270-290 fold greater than those obtained by the "impaction on nutrient agar" method. The culture-dependent method of air impaction on nutrient agar was also inadequate in poultry houses, as was the impinger-culture method, which gave a bacterial load estimate 32-fold lower than obtained by Q-PCR. Real-time quantitative PCR thus proves to be a reliable, discerning, and simple method that could be used to estimate airborne bacterial load in a broad variety of other environments expected to carry high numbers of airborne bacteria.

Census of the bacterial community of the gypsy moth larval midgut by using culturing and culture-independent methods.

PubMed

Broderick, Nichole A; Raffa, Kenneth F; Goodman, Robert M; Handelsman, Jo

2004-01-01

Little is known about bacteria associated with Lepidoptera, the large group of mostly phytophagous insects comprising the moths and butterflies. We inventoried the larval midgut bacteria of a polyphagous foliivore, the gypsy moth (Lymantria dispar L.), whose gut is highly alkaline, by using traditional culturing and culture-independent methods. We also examined the effects of diet on microbial composition. Analysis of individual third-instar larvae revealed a high degree of similarity of microbial composition among insects fed on the same diet. DNA sequence analysis indicated that most of the PCR-amplified 16S rRNA genes belong to the gamma-Proteobacteria and low G+C gram-positive divisions and that the cultured members represented more than half of the phylotypes identified. Less frequently detected taxa included members of the alpha-Proteobacterium, Actinobacterium, and Cytophaga/Flexibacter/Bacteroides divisions. The 16S rRNA gene sequences from 7 of the 15 cultured organisms and 8 of the 9 sequences identified by PCR amplification diverged from previously reported bacterial sequences. The microbial composition of midguts differed substantially among larvae feeding on a sterilized artificial diet, aspen, larch, white oak, or willow. 16S rRNA analysis of cultured isolates indicated that an Enterococcus species and culture-independent analysis indicated that an Entbacter sp. were both present in all larvae, regardless of the feeding substrate; the sequences of these two phylotypes varied less than 1% among individual insects. These results provide the first comprehensive description of the microbial diversity of a lepidopteran midgut and demonstrate that the plant species in the diet influences the composition of the gut bacterial community.

Use of Natural Antimicrobial Peptides and Bacterial Biopolymers for Cultured Pearl Production

PubMed Central

Simon-Colin, Christelle; Gueguen, Yannick; Bachere, Evelyne; Kouzayha, Achraf; Saulnier, Denis; Gayet, Nicolas; Guezennec, Jean

2015-01-01

Cultured pearls are the product of grafting and rearing of Pinctada margaritifera pearl oysters in their natural environment. Nucleus rejections and oyster mortality appear to result from bacterial infections or from an inappropriate grafting practice. To reduce the impact of bacterial infections, synthetic antibiotics have been applied during the grafting practice. However, the use of such antibiotics presents a number of problems associated with their incomplete biodegradability, limited efficacy in some cases, and an increased risk of selecting for antimicrobial resistant bacteria. We investigated the application of a marine antimicrobial peptide, tachyplesin, which is present in the Japanese horseshoe crab Tachypleus tridentatus, in combination with two marine bacterial exopolymers as alternative treatment agents. In field studies, the combination treatment resulted in a significant reduction in graft failures vs. untreated controls. The combination of tachyplesin (73 mg/L) with two bacterial exopolysaccharides (0.5% w/w) acting as filming agents, reduces graft-associated bacterial contamination. The survival data were similar to that reported for antibiotic treatments. These data suggest that non-antibiotic treatments of pearl oysters may provide an effective means of improving oyster survival following grafting procedures. PMID:26110895

Impact of eye bank lamellar tissue cutting for endothelial keratoplasty on bacterial and fungal corneoscleral donor rim cultures after corneal transplantation.

PubMed

Rauen, Matthew P; Goins, Kenneth M; Sutphin, John E; Kitzmann, Anna S; Schmidt, Gregory A; Wagoner, Michael D

2012-04-01

To determine if the lamellar cut of donor tissue for endothelial keratoplasty (EK) by an eye bank facility is associated with a change in the prevalence of positive bacterial or fungal donor rim cultures after corneal transplantation. A retrospective review was conducted of bacterial and fungal cultures of donor rims used for corneal transplantation at a tertiary eye care center from January 1, 2003, to December 31, 2008, with tissue provided by a single eye bank. The cases were divided into 2 groups. Group 1 ("no-cut") included keratoplasty procedures in which a lamellar cut was not performed. Group 2 ("precut") included EK procedures in which a 4-hour period of prewarming of tissue followed by a lamellar cut was performed in the eye bank before tissue delivery to the operating surgeon. There were 351 donor rim cultures in group 1 and 278 in group 2. Bacterial cultures were positive in 30 donor rims (8.5%) in group 1 and 13 (4.7%) in group 2 (P = 0.058). Positive bacterial cultures were not associated with any postoperative infections. Fungal cultures were positive in 8 donor rims (2.3%) in group 1 and 7 (2.5%) in group 2 (P = 1.0). Positive fungal cultures were associated with 2 cases (13.3%) of postoperative fungal infections. Corneal donor tissue can be precut for EK by trained eye bank personnel without an increased risk of bacterial or fungal contamination.

Impact of estuarine gradients on reductive dechlorination of 1,2,3,4-tetrachlorodibenzo-p-dioxin in river sediment enrichment cultures.

PubMed

Dam, Hang T; Häggblom, Max M

2017-02-01

Polychlorinated dibenzo-p-dioxins (PCDDs) are among the most persistent organic pollutants. Although the total input of PCDDs into the environment has decreased substantially over the past four decades, their input via non-point sources is still increasing, especially in estuarine metropolitan areas. Here we report on the microbially mediated reductive dechlorination of PCDDs in anaerobic enrichment cultures established from sediments collected from five locations along the Hackensack River, NJ and investigate the impacts of sediment physicochemical characteristics on dechlorination activity. Dechlorination of 1,2,3,4-tetrachlorodibenzo-p-dioxin (1,2,3,4-TeCDD) and abundance of Dehalococcoides spp. negatively correlated with salinity and sulfate concentration in sediments used to establish the cultures. 1,2,3,4-TeCDD was dechlorinated to a lesser extent in cultures established from sediments from the tidally influenced estuarine mouth of the river. In cultures established from low salinity sediments, 1,2,3,4-TeCDD was reductively dechlorinated with the accumulation of 2-monochlorodibenzo-p-dioxin as the major product. Sulfate concentrations above 2 mM inhibited 1,2,3,4-TecDD dechlorination activity. Consecutive lateral- and peri- dechlorination took place in enrichment cultures with a minimal accumulation of 2,3-dichlorodibenzo-p-dioxin in active cultures. A Dehalococcoides spp. community was enriched and accounted for up to 64% of Chloroflexi detected in these sediment cultures. Copyright © 2016 Elsevier Ltd. All rights reserved.

Enumeration of the contaminating bacterial microbiota in unfermented pasteurized milks enriched with probiotic bacteria.

PubMed

Champagne, C P; Raymond, Y; Gonthier, J; Audet, P

2009-04-01

Pasteurized and unfermented milks supplemented with probiotic bacteria are appearing on the market. It then becomes a challenge to ascertain the undesirable contamination microbiota in the presence of a largely superior population of probiotic bacteria. A method to enumerate the contaminating microbial microbiota in such probiotic-enriched milks was developed. The probiotic cultures, Lactobacillus rhamnosus Lb-Immuni-T and Bifidobacterium animalis subsp. lactis BB-12(R), were added to a pasteurized unfermented milk to reach a minimum of 1 billion CFU per 250 mL portion, as ascertained by plating on de Man - Rogosa - Sharpe (MRS) agar in anaerobic conditions. No growth of B. animalis subsp. lactis BB-12 was noted on plate count agar (PCA) or Petrifilm plates, and the presence of this culture did not affect standard plate counts (SPC) of contaminating bacteria. However, L. rhamnosus formed colonies on PCA and Petrifilm plates. Attempts were thus made to inhibit the growth of the probiotic lactobacilli in PCA. The addition of 2% sodium phosphate (SP) or 5% glycerophosphate (GP) inhibited the growth of the lactobacilli in broths, but pin-point colonies of L. rhamnosus Lb-Immuni-T nevertheless appeared on PCA supplemented with phosphates. SPC could be obtained on PCA + 2% SP by only counting the large colonies, but this resulted in a significant (4.4 fold) underestimation of SPC values. On Petrifilm AC, at dilutions 0 to 2, all colonies were considered as being contaminants, while at dilutions 3 and 4, only large colonies were counted for SPC determinations. There was a direct correlation (R2 = 0.99) between SPC values with Petrifilm in uninoculated milks and those obtained on probiotic-enriched milks. The high correlation obtained over the 102 to 106 CFU/mL range of SPC values show that this Petrifilm method is appropriate to evaluate the microbiological quality of pasteurized milks enriched with L. rhamnosus Lb-Immuni-T and B. animalis subsp. lactis BB-12.

Polyhydroxyalkanoate accumulation ability and associated microbial community in activated sludge-derived acetate-fed microbial cultures enriched under different temperature and pH conditions.

PubMed

Inoue, Daisuke; Suzuki, Yuta; Sawada, Kazuko; Sei, Kazunari

2018-03-01

The influence of temperature and pH during enrichment on the polyhydroxyalkanoate (PHA) accumulation ability and composition of PHA-accumulating microorganisms (PHAAMOs) in enrichment cultures was investigated. Enrichment of PHAAMOs from activated sludge was conducted in acetate-fed sequencing batch reactors using a feast-famine regime under different temperature (20°C, 28°C, and 36°C) and pH (controlled at 7.2 or not) conditions. PHA accumulation ability, which was evaluated in nitrogen- and phosphorus-deficient 24-h single-batch cultures, was greatly enhanced by enrichment, irrespective of the temperature and pH. Enrichment at 20°C or 28°C and without pH control seemed most appropriate for strong PHA accumulation. Analyses of the PHAAMO composition by the clone library method targeting phaC genes, which encode the class I and II PHA synthases, revealed that Burkholderiales were the dominant PHAAMOs in the seed sludge, while Rhodocyclales, specifically Azoarcus spp. and Thauera spp., were dominant after enrichment without pH control, showing a strong ability to accumulate PHA. The results indicated that Azoarcus spp. and Thauera spp. are key PHAAMOs in an enrichment culture based on the feast-famine method, with high PHA accumulation ability. Copyright © 2017 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Culture-dependent and culture-independent characterization of potentially functional biphenyl-degrading bacterial community in response to extracellular organic matter from Micrococcus luteus.

PubMed

Su, Xiao-Mei; Liu, Yin-Dong; Hashmi, Muhammad Zaffar; Ding, Lin-Xian; Shen, Chao-Feng

2015-05-01

Biphenyl (BP)-degrading bacteria were identified to degrade various polychlorinated BP (PCB) congers in long-term PCB-contaminated sites. Exploring BP-degrading capability of potentially useful bacteria was performed for enhancing PCB bioremediation. In the present study, the bacterial composition of the PCB-contaminated sediment sample was first investigated. Then extracellular organic matter (EOM) from Micrococcus luteus was used to enhance BP biodegradation. The effect of the EOM on the composition of bacterial community was investigated by combining with culture-dependent and culture-independent methods. The obtained results indicate that Proteobacteria and Actinobacteria were predominant community in the PCB-contaminated sediment. EOM from M. luteus could stimulate the activity of some potentially difficult-to-culture BP degraders, which contribute to significant enhancement of BP biodegradation. The potentially difficult-to-culture bacteria in response to EOM addition were mainly Rhodococcus and Pseudomonas belonging to Gammaproteobacteria and Actinobacteria respectively. This study provides new insights into exploration of functional difficult-to-culture bacteria with EOM addition and points out broader BP/PCB degrading, which could be employed for enhancing PCB-bioremediation processes. © 2015 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.

Dichloromethane Fermentation by a Dehalobacter sp. in an Enrichment Culture Derived from Pristine River Sediment

PubMed Central

Justicia-Leon, Shandra D.; Ritalahti, Kirsti M.; Mack, E. Erin

2012-01-01

Dichloromethane (DCM) as the sole substrate supported growth of a Dehalobacter sp. in an enrichment culture derived from noncontaminated river sediment. DCM was not reductively dechlorinated, and acetate was produced, indicating DCM fermentation and further suggesting Dehalobacter growth is not limited to organohalide respiration. PMID:22179245

Anaerobic oxidation of acetylene by estuarine sediments and enrichment cultures

USGS Publications Warehouse

Culbertson, Charles W.; Zehnder, Alexander J. B.; Oremland, Ronald S.

1981-01-01

Acetylene disappeared from the gas phase of anaerobically incubated estuarine sediment slurries, and loss was accompanied by increased levels of carbon dioxide. Acetylene loss was inhibited by chloramphenicol, air, and autoclaving. Addition of 14C2H2 to slurries resulted in the formation of 14CO2 and the transient appearance of 14C-soluble intermediates, of which acetate was a major component. Acetylene oxidation stimulated sulfate reduction; however, sulfate reduction was not required for the loss of C2H2 to occur. Enrichment cultures were obtained which grew anaerobically at the expense of C2H2.

Use of bromodeoxyuridine immunocapture to identify psychrotolerant phenanthrene-degrading bacteria in phenanthrene-enriched polluted Baltic Sea sediments

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

Edlund, A.; Jansson, J.

The aim of this study was to enrich and identify psychrotolerant phenanthrenedegrading bacteria from polluted Baltic Sea sediments. Polyaromatic hydrocarbon (PAH)-contaminated sediments were spiked with phenanthrene and incubated for 2 months in the presence of bromodeoxyuridine that is incorporated into the DNA of replicating cells. The bromodeoxyuridine-incorporated DNA was extracted by immunocapture and analyzed by terminal-restriction fragment length polymorphism and 16S rRNA gene cloning and sequencing to identify bacterial populations that were growing. In addition, degradation genes were quantified in the bromodeoxyuridine-incorporated DNA by real-time PCR. Phenanthrene concentrations decreased after 2 months of incubation in the phenanthrene-enriched sediments and thismore » reduction correlated to increases in copy numbers of xylE and phnAc dioxygenase genes. Representatives of Exiguobacterium, Schewanella,Methylomonas, Pseudomonas, Bacteroides and an uncultured Deltaproteobacterium and a Gammaproteobacterium dominated the growing community in the phenanthrene spiked sediments. Isolates that were closely related to three of these bacteria (two pseudomonads and an Exiguobacterium sp.) could reduce phenanthrene concentrations in pure cultures and they all harbored phnAc dioxygenase genes. These results confirm that this combination of culture-based and molecular approaches was useful for identification of actively growing bacterial species with a high potential for phenanthrene degradation.« less

Conversion of crude oil to methane by a microbial consortium enriched from oil reservoir production waters

PubMed Central

Berdugo-Clavijo, Carolina; Gieg, Lisa M.

2014-01-01

The methanogenic biodegradation of crude oil is an important process occurring in petroleum reservoirs and other oil-containing environments such as contaminated aquifers. In this process, syntrophic bacteria degrade hydrocarbon substrates to products such as acetate, and/or H2 and CO2 that are then used by methanogens to produce methane in a thermodynamically dependent manner. We enriched a methanogenic crude oil-degrading consortium from production waters sampled from a low temperature heavy oil reservoir. Alkylsuccinates indicative of fumarate addition to C5 and C6 n-alkanes were identified in the culture (above levels found in controls), corresponding to the detection of an alkyl succinate synthase encoding gene (assA/masA) in the culture. In addition, the enrichment culture was tested for its ability to produce methane from residual oil in a sandstone-packed column system simulating a mature field. Methane production rates of up to 5.8 μmol CH4/g of oil/day were measured in the column system. Amounts of produced methane were in relatively good agreement with hydrocarbon loss showing depletion of more than 50% of saturate and aromatic hydrocarbons. Microbial community analysis revealed that the enrichment culture was dominated by members of the genus Smithella, Methanosaeta, and Methanoculleus. However, a shift in microbial community occurred following incubation of the enrichment in the sandstone columns. Here, Methanobacterium sp. were most abundant, as were bacterial members of the genus Pseudomonas and other known biofilm forming organisms. Our findings show that microorganisms enriched from petroleum reservoir waters can bioconvert crude oil components to methane both planktonically and in sandstone-packed columns as test systems. Further, the results suggest that different organisms may contribute to oil biodegradation within different phases (e.g., planktonic vs. sessile) within a subsurface crude oil reservoir. PMID:24829563

Culture Bottle Investigations of Nutrient Enriched Oligotrophic Phytoplankton Communities Challenge Contemporary Beliefs

NASA Astrophysics Data System (ADS)

Harrison, D. P.

2016-02-01

Humankind has fundamentally altered the global nitrogen cycle, such that today as much nitrogen is fixed from the atmosphere anthropogenically, as is fixed naturally by terrestrial and aquatic systems. 70% of this alteration is in the form of nitrogenous fertilizers, and Haber-Bosh production of urea now accounts for 20% of total global nitrogen fixation (anthropogenic and natural). Cultural eutrophication has long been implicated in an apparent increase in the number and severity of harmful algal blooms (HAB). More recently the form of introduced nitrogen has been receiving attention, with urea in particular singled out as a potential causative agent, yet this deduction seems to largely rely on observed correlations rather than establishment of a direct link. An alternative hypothesis is that environmental factors rather than the form of nitrogen exert a controlling influence on the nature of phytoplankton response to nutrient enrichment. Here I present the results of a series of eight repeated experiments conducted over an annual cycle in 2013-2014 using oligotrophic coastal phytoplankton assemblages to asses the effect of urea and nitrate enrichment on size distribution, speciation, and biochemistry. Experiments were conducted at one location offshore Sydney, Australia but had very different oceanographic starting conditions. The result of enrichment (+8 μM N & +0.5 μM P) using both nitrate and urea was a greater abundance of diatoms than dinoflagellates in all cases. Overall species composition was not significantly different (at 0.05 level) for nitrate and urea as revealed by multidimensional scaling and permutational ANOVA. However in some cases, contrary to published speculation, nitrate rather than urea resulted in increased abundance of dinoflagellates. A generalized mixed modeling approach identified aspects of the water column which appear to be associated with the presence of the East Australian Current as being influential. These results imply that

An investigation of total bacterial communities, culturable antibiotic-resistant bacterial communities and integrons in the river water environments of Taipei city.

PubMed

Yang, Chu-Wen; Chang, Yi-Tang; Chao, Wei-Liang; Shiung, Iau-Iun; Lin, Han-Sheng; Chen, Hsuan; Ho, Szu-Han; Lu, Min-Jheng; Lee, Pin-Hsuan; Fan, Shao-Ning

2014-07-30

The intensive use of antibiotics may accelerate the development of antibiotic-resistant bacteria (ARB). The global geographical distribution of environmental ARB has been indicated by many studies. However, the ARB in the water environments of Taiwan has not been extensively investigated. The objective of this study was to investigate the communities of ARB in Huanghsi Stream, which presents a natural acidic (pH 4) water environment. Waishuanghsi Stream provides a neutral (pH 7) water environment and was thus also monitored to allow comparison. The plate counts of culturable bacteria in eight antibiotics indicate that the numbers of culturable carbenicillin- and vancomycin-resistant bacteria in both Huanghsi and Waishuanghsi Streams are greater than the numbers of culturable bacteria resistant to the other antibiotics tested. Using a 16S rDNA sequencing approach, both the antibiotic-resistant bacterial communities (culture-based) and the total bacterial communities (metagenome-based) in Waishuanghsi Stream exhibit a higher diversity than those in Huanghsi Stream were observed. Of the three classes of integron, only class I integrons were identified in Waishuanghsi Stream. Our results suggest that an acidic (pH 4) water environment may not only affect the community composition of antibiotic-resistant bacteria but also the horizontal gene transfer mediated by integrons. Copyright © 2013 Elsevier B.V. All rights reserved.

Wheat bran promotes enrichment within the human colonic microbiota of butyrate-producing bacteria that release ferulic acid.

PubMed

Duncan, Sylvia H; Russell, Wendy R; Quartieri, Andrea; Rossi, Maddalena; Parkhill, Julian; Walker, Alan W; Flint, Harry J

2016-07-01

Cereal fibres such as wheat bran are considered to offer human health benefits via their impact on the intestinal microbiota. We show here by 16S rRNA gene-based community analysis that providing amylase-pretreated wheat bran as the sole added energy source to human intestinal microbial communities in anaerobic fermentors leads to the selective and progressive enrichment of a small number of bacterial species. In particular, OTUs corresponding to uncultured Lachnospiraceae (Firmicutes) related to Eubacterium xylanophilum and Butyrivibrio spp. were strongly enriched (by five to 160 fold) over 48 h in four independent experiments performed with different faecal inocula, while nine other Firmicutes OTUs showed > 5-fold enrichment in at least one experiment. Ferulic acid was released from the wheat bran during degradation but was rapidly converted to phenylpropionic acid derivatives via hydrogenation, demethylation and dehydroxylation to give metabolites that are detected in human faecal samples. Pure culture work using bacterial isolates related to the enriched OTUs, including several butyrate-producers, demonstrated that the strains caused substrate weight loss and released ferulic acid, but with limited further conversion. We conclude that breakdown of wheat bran involves specialist primary degraders while the conversion of released ferulic acid is likely to involve a multi-species pathway. © 2015 The Authors. Environmental Microbiology published by Society for Applied Microbiology and John Wiley & Sons Ltd.

Rapid Identification of Shiga Toxin-Producing Escherichia coli O Serogroups from Fresh Produce and Raw Milk Enrichment Cultures by Luminex Bead-Based Suspension Array.

PubMed

Kase, Julie A; Maounounen-Laasri, Anna; Lin, Andrew

2016-09-01

The U.S. Food and Drug Administration's Bacteriological Analytical Manual (BAM) Chapter 4a describes a Luminex microbead-based suspension array used to screen colonies for 11 clinically relevant Shiga toxin-producing Escherichia coli (STEC) serogroups: O26, O45, O91, O103, O104, O111, O113, O121, O128, O145, and O157. We evaluated the usefulness of this method to identify STEC-positive enrichment samples before agar plating. Twelve E. coli strains were added to three types of fresh produce (bagged baby spinach, alfalfa sprouts, and cilantro) at levels near the detection limit of the test. A subset of these strains (six O serogroups) was similarly evaluated in raw milk. For comparison, portions of each of the 168 enrichment cultures were analyzed for serogroup by a real-time PCR assay and a Bio-Plex 200 assay with the bead-based suspensions. No false-positive results were obtained. Of the 112 samples with a reported cycle threshold (C T ) value, 101 undiluted, diluted, or extracted enrichment cultures also produced ratios above 5.0 in the Bio-Plex assay. When PCR C T values approached or were greater than 35, Bio-Plex detection became less reliable. Using undiluted or extracted enrichment cultures resulted in a significantly larger number of positive results. With the same enrichment material prepared for real-time PCR analysis as described in the BAM Chapter 4a, the STEC microbead-based suspension array can accurately screen food enrichment cultures.

Analyses of n-alkanes degrading community dynamics of a high-temperature methanogenic consortium enriched from production water of a petroleum reservoir by a combination of molecular techniques.

PubMed

Zhou, Lei; Li, Kai-Ping; Mbadinga, Serge Maurice; Yang, Shi-Zhong; Gu, Ji-Dong; Mu, Bo-Zhong

2012-08-01

Despite the knowledge on anaerobic degradation of hydrocarbons and signature metabolites in the oil reservoirs, little is known about the functioning microbes and the related biochemical pathways involved, especially about the methanogenic communities. In the present study, a methanogenic consortium enriched from high-temperature oil reservoir production water and incubated at 55 °C with a mixture of long chain n-alkanes (C(15)-C(20)) as the sole carbon and energy sources was characterized. Biodegradation of n-alkanes was observed as methane production in the alkanes-amended methanogenic enrichment reached 141.47 μmol above the controls after 749 days of incubation, corresponding to 17 % of the theoretical total. GC-MS analysis confirmed the presence of putative downstream metabolites probably from the anaerobic biodegradation of n-alkanes and indicating an incomplete conversion of the n-alkanes to methane. Enrichment cultures taken at different incubation times were subjected to microbial community analysis. Both 16S rRNA gene clone libraries and DGGE profiles showed that alkanes-degrading community was dynamic during incubation. The dominant bacterial species in the enrichment cultures were affiliated with Firmicutes members clustering with thermophilic syntrophic bacteria of the genera Moorella sp. and Gelria sp. Other represented within the bacterial community were members of the Leptospiraceae, Thermodesulfobiaceae, Thermotogaceae, Chloroflexi, Bacteroidetes and Candidate Division OP1. The archaeal community was predominantly represented by members of the phyla Crenarchaeota and Euryarchaeota. Corresponding sequences within the Euryarchaeota were associated with methanogens clustering with orders Methanomicrobiales, Methanosarcinales and Methanobacteriales. On the other hand, PCR amplification for detection of functional genes encoding the alkylsuccinate synthase α-subunit (assA) was positive in the enrichment cultures. Moreover, the appearance of a new ass

Biotreatment of refinery spent-sulfidic caustic using an enrichment culture immobilized in a novel support matrix.

PubMed

Conner, J A; Beitle, R R; Duncan, K; Kolhatkar, R; Sublette, K L

2000-01-01

Sodium hydroxide solutions are used in petroleum refining to remove hydrogen sulfide (H2S) and mercaptans from various hydrocarbon streams. The resulting sulfide-laden waste stream is called spent-sulfidic caustic. An aerobic enrichment culture was previously developed using a gas mixture of H2S and methyl-mercaptan (MeSH) as the sole energy source. This culture has now been immobilized in a novel support matrix, DuPont BIO-SEP beads, and is used to bio-treat a refinery spent-sulfidic caustic containing both inorganic sulfide and mercaptans in a continuous flow, fluidized-bed column bioreactor. Complete oxidation of both inorganic and organic sulfur to sulfate was observed with no breakthrough of H2S and < 2 ppmv of MeSH produced in the bioreactor outlet gas. Excessive buildup of sulfate (> 12 g/L) in the bioreactor medium resulted in an upset condition evidenced by excessive MeSH breakthrough. Therefore, bioreactor performance was limited by the steady-state sulfate concentration. Further improvement in volumetric productivity of a bioreactor system based on this enrichment culture will be dependent on maintenance of sulfate concentrations below inhibitory levels.

A duplex PCR-based assay for measuring the amount of bacterial contamination in a nucleic acid extract from a culture of free-living protists.

PubMed

Marron, Alan O; Akam, Michael; Walker, Giselle

2013-01-01

Cultures of heterotrophic protists often require co-culturing with bacteria to act as a source of nutrition. Such cultures will contain varying levels of intrinsic bacterial contamination that can interfere with molecular research and cause problems with the collection of sufficient material for sequencing. Measuring the levels of bacterial contamination for the purposes of molecular biology research is non-trivial, and can be complicated by the presence of a diverse bacterial flora, or by differences in the relative nucleic acid yield per bacterial or eukaryotic cell. Here we describe a duplex PCR-based assay that can be used to measure the levels of contamination from marine bacteria in a culture of loricate choanoflagellates. By comparison to a standard culture of known target sequence content, the assay can be used to quantify the relative proportions of bacterial and choanoflagellate material in DNA or RNA samples extracted from a culture. We apply the assay to compare methods of purifying choanoflagellate cultures prior to DNA extraction, to determine their effectiveness in reducing bacterial contamination. Together with measurements of the total nucleic acid concentration, the assay can then be used as the basis for determining the absolute amounts of choanoflagellate DNA or RNA present in a sample. The assay protocol we describe here is a simple and relatively inexpensive method of measuring contamination levels in nucleic acid samples. This provides a new way to establish quantification and purification protocols for molecular biology and genomics in novel heterotrophic protist species. Guidelines are provided to develop a similar protocol for use with any protistan culture. This assay method is recommended where qPCR equipment is unavailable, where qPCR is not viable because of the nature of the bacterial contamination or starting material, or where prior sequence information is insufficient to develop qPCR protocols.

Characterization of Fe (III)-reducing enrichment culture and isolation of Fe (III)-reducing bacterium Enterobacter sp. L6 from marine sediment.

PubMed

Liu, Hongyan; Wang, Hongyu

2016-07-01

To enrich the Fe (III)-reducing bacteria, sludge from marine sediment was inoculated into the medium using Fe (OH)3 as the sole electron acceptor. Efficiency of Fe (III) reduction and composition of Fe (III)-reducing enrichment culture were analyzed. The results indicated that the Fe (III)-reducing enrichment culture with the dominant bacteria relating to Clostridium and Enterobacter sp. had high Fe (III) reduction of (2.73 ± 0.13) mmol/L-Fe (II). A new Fe (III)-reducing bacterium was isolated from the Fe (III)-reducing enrichment culture and identified as Enterobacter sp. L6 by 16S rRNA gene sequence analysis. The Fe (III)-reducing ability of strain L6 under different culture conditions was investigated. The results indicated that strain L6 had high Fe (III)-reducing activity using glucose and pyruvate as carbon sources. Strain L6 could reduce Fe (III) at the range of NaCl concentrations tested and had the highest Fe (III) reduction of (4.63 ± 0.27) mmol/L Fe (II) at the NaCl concentration of 4 g/L. This strain L6 could reduce Fe (III) with unique properties in adaptability to salt variation, which indicated that it can be used as a model organism to study Fe (III)-reducing activity isolated from marine environment. Copyright © 2015 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Long-Term Enrichment on Cellulose or Xylan Causes Functional and Taxonomic Convergence of Microbial Communities from Anaerobic Digesters

PubMed Central

Jia, Yangyang; Wilkins, David; Lu, Hongyuan; Cai, Mingwei

2015-01-01

Cellulose and xylan are two major components of lignocellulosic biomass, which represents a potentially important energy source, as it is abundant and can be converted to methane by microbial action. However, it is recalcitrant to hydrolysis, and the establishment of a complete anaerobic digestion system requires a specific repertoire of microbial functions. In this study, we maintained 2-year enrichment cultures of anaerobic digestion sludge amended with cellulose or xylan to investigate whether a cellulose- or xylan-digesting microbial system could be assembled from sludge previously used to treat neither of them. While efficient methane-producing communities developed under mesophilic (35°C) incubation, they did not under thermophilic (55°C) conditions. Illumina amplicon sequencing results of the archaeal and bacterial 16S rRNA genes revealed that the mature cultures were much lower in richness than the inocula and were dominated by single archaeal (genus Methanobacterium) and bacterial (order Clostridiales) groups, although at finer taxonomic levels the bacteria were differentiated by substrates. Methanogenesis was primarily via the hydrogenotrophic pathway under all conditions, although the identity and growth requirements of syntrophic acetate-oxidizing bacteria were unclear. Incubation conditions (substrate and temperature) had a much greater effect than inoculum source in shaping the mature microbial community, although analysis based on unweighted UniFrac distance found that the inoculum still determined the pool from which microbes could be enriched. Overall, this study confirmed that anaerobic digestion sludge treating nonlignocellulosic material is a potential source of microbial cellulose- and xylan-digesting functions given appropriate enrichment conditions. PMID:26712547

MICROBIAL REDUCTIVE DECHLORINATION OF HEXACHLORO-1,3-BUTADIENE IN A METHANOGENIC ENRICHMENT CULTURE. (R825513C007)

EPA Science Inventory

Sequential reductive dechlorination of hexachloro-1,3-butadiene (HCBD) was achieved by a mixed, methanogenic culture enriched from a contaminated estuarine sediment. Both methanol and lactate served as carbon and electron sources. Methanol was stoichiometrically converted to m...

Integrated biogas upgrading and hydrogen utilization in an anaerobic reactor containing enriched hydrogenotrophic methanogenic culture.

PubMed

Luo, Gang; Angelidaki, Irini

2012-11-01

Biogas produced by anaerobic digestion, is mainly used in a gas motor for heat and electricity production. However, after removal of CO(2) , biogas can be upgraded to natural gas quality, giving more utilization possibilities, such as utilization as autogas, or distant utilization by using the existing natural gas grid. The current study presents a new biological method for biogas upgrading in a separate biogas reactor, containing enriched hydrogenotrophic methanogens and fed with biogas and hydrogen. Both mesophilic- and thermophilic anaerobic cultures were enriched to convert CO(2) to CH(4) by addition of H(2) . Enrichment at thermophilic temperature (55°C) resulted in CO(2) and H(2) bioconversion rate of 320 mL CH(4) /(gVSS h), which was more than 60% higher than that under mesophilic temperature (37°C). Different dominant species were found at mesophilic- and thermophilic-enriched cultures, as revealed by PCR-DGGE. Nonetheless, they all belonged to the order Methanobacteriales, which can mediate hydrogenotrophic methanogenesis. Biogas upgrading was then tested in a thermophilic anaerobic reactor under various operation conditions. By continuous addition of hydrogen in the biogas reactor, high degree of biogas upgrading was achieved. The produced biogas had a CH(4) content, around 95% at steady-state, at gas (mixture of biogas and hydrogen) injection rate of 6 L/(L day). The increase of gas injection rate to 12 L/(L day) resulted in the decrease of CH(4) content to around 90%. Further study showed that by decreasing the gas-liquid mass transfer by increasing the stirring speed of the mixture the CH(4) content was increased to around 95%. Finally, the CH(4) content around 90% was achieved in this study with the gas injection rate as high as 24 L/(L day). Copyright © 2012 Wiley Periodicals, Inc.

Culture-independent analysis of hydrocarbonoclastic bacterial communities in environmental samples during oil-bioremediation.

PubMed

Dashti, Narjes; Ali, Nedaa; Salamah, Samar; Khanafer, Majida; Al-Shamy, Ghada; Al-Awadhi, Husain; Radwan, Samir S

2018-04-15

To analyze microbial communities in environmental samples, this study combined Denaturing Gradient Gel Electrophoresis of amplified 16S rRNA-genes in total genomic DNA extracts from those samples with gene sequencing. The environmental samples studied were oily seawater and soil samples, that had been bioaugmented with natural materials rich in hydrocarbonoclastic bacteria. This molecular approach revealed much more diverse bacterial taxa than the culture-dependent method we had used in an earlier study for the analysis of the same samples. The study described the dynamics of bacterial communities during bioremediation. The main limitation associated with this molecular approach, namely of not distinguishing hydrocarbonoclastic taxa from others, was overcome by consulting the literature for the hydrocarbonoclastic potential of taxa related to those identified in this study. By doing so, it was concluded that the hydrocarbonoclastic bacterial taxa were much more diverse than those captured by the culture-dependent approach. The molecular analysis also revealed the frequent occurrence of nifH-genes in the total genomic DNA extracts of all the studied environmental samples, which reflects a nitrogen-fixation potential. Nitrogen fertilization is long known to enhance microbial oil-bioremediation. The study revealed that bioaugmentation using plant rhizospheres or soil with long history of oil-pollution was more effective in oil-removal in the desert soil than in seawater microcosms. © 2018 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

Evaluation of bacterial diversity recovered from petroleum samples using different physical matrices.

PubMed

Dellagnezze, Bruna Martins; Vasconcellos, Suzan Pantaroto de; Melo, Itamar Soares de; Santos Neto, Eugênio Vaz Dos; Oliveira, Valéria Maia de

2016-01-01

Unraveling the microbial diversity and its complexity in petroleum reservoir environments has been a challenge throughout the years. Despite the techniques developed in order to improve methodologies involving DNA extraction from crude oil, microbial enrichments using different culture conditions can be applied as a way to increase the recovery of DNA from environments with low cellular density for further microbiological analyses. This work aimed at the evaluation of different matrices (arenite, shale and polyurethane foam) as support materials for microbial growth and biofilm formation in enrichments using a biodegraded petroleum sample as inoculum in sulfate reducing condition. Subsequent microbial diversity characterization was carried out using Scanning Electronic Microscopy (SEM), Denaturing Gradient Gel Electrophoresis (DGGE) and 16S rRNA gene libraries in order to compare the microbial biomass yield, DNA recovery efficiency and diversity among the enrichments. The DNA from microbial communities in petroleum enrichments was purified according to a protocol established in this work and used for 16S rRNA amplification with bacterial generic primers. The PCR products were cloned, and positive clones were screened by Amplified Ribosomal DNA Restriction Analysis (ARDRA). Sequencing and phylogenetic analyses revealed that the bacterial community was mostly represented by members of the genera Petrotoga, Bacillus, Pseudomonas, Geobacillus and Rahnella. The use of different support materials in the enrichments yielded an increase in microbial biomass and biofilm formation, indicating that these materials may be employed for efficient biomass recovery from petroleum reservoir samples. Nonetheless, the most diverse microbiota were recovered from the biodegraded petroleum sample using polyurethane foam cubes as support material. Copyright © 2016 Sociedade Brasileira de Microbiologia. Published by Elsevier Editora Ltda. All rights reserved.

Ultrastructure and Viral Metagenome of Bacteriophages from an Anaerobic Methane Oxidizing Methylomirabilis Bioreactor Enrichment Culture

PubMed Central

Gambelli, Lavinia; Cremers, Geert; Mesman, Rob; Guerrero, Simon; Dutilh, Bas E.; Jetten, Mike S. M.; Op den Camp, Huub J. M.; van Niftrik, Laura

2016-01-01

With its capacity for anaerobic methane oxidation and denitrification, the bacterium Methylomirabilis oxyfera plays an important role in natural ecosystems. Its unique physiology can be exploited for more sustainable wastewater treatment technologies. However, operational stability of full-scale bioreactors can experience setbacks due to, for example, bacteriophage blooms. By shaping microbial communities through mortality, horizontal gene transfer, and metabolic reprogramming, bacteriophages are important players in most ecosystems. Here, we analyzed an infected Methylomirabilis sp. bioreactor enrichment culture using (advanced) electron microscopy, viral metagenomics and bioinformatics. Electron micrographs revealed four different viral morphotypes, one of which was observed to infect Methylomirabilis cells. The infected cells contained densely packed ~55 nm icosahedral bacteriophage particles with a putative internal membrane. Various stages of virion assembly were observed. Moreover, during the bacteriophage replication, the host cytoplasmic membrane appeared extremely patchy, which suggests that the bacteriophages may use host bacterial lipids to build their own putative internal membrane. The viral metagenome contained 1.87 million base pairs of assembled viral sequences, from which five putative complete viral genomes were assembled and manually annotated. Using bioinformatics analyses, we could not identify which viral genome belonged to the Methylomirabilis- infecting bacteriophage, in part because the obtained viral genome sequences were novel and unique to this reactor system. Taken together these results show that new bacteriophages can be detected in anaerobic cultivation systems and that the effect of bacteriophages on the microbial community in these systems is a topic for further study. PMID:27877158

Bacterial respiration of arsenic and selenium

USGS Publications Warehouse

Stolz, J.F.; Oremland, R.S.

1999-01-01

Oxyanions of arsenic and selenium can be used in microbial anaerobic respiration as terminal electron acceptors. The detection of arsenate and selenate respiring bacteria in numerous pristine and contaminated environments and their rapid appearance in enrichment culture suggest that they are widespread and metabolically active in nature. Although the bacterial species that have been isolated and characterized are still few in number, they are scattered throughout the bacterial domain and include Gram- positive bacteria, beta, gamma and epsilon Proteobacteria and the sole member of a deeply branching lineage of the bacteria, Chrysiogenes arsenatus. The oxidation of a number of organic substrates (i.e. acetate, lactate, pyruvate, glycerol, ethanol) or hydrogen can be coupled to the reduction of arsenate and selenate, but the actual donor used varies from species to species. Both periplasmic and membrane-associated arsenate and selenate reductases have been characterized. Although the number of subunits and molecular masses differs, they all contain molybdenum. The extent of the environmental impact on the transformation and mobilization of arsenic and selenium by microbial dissimilatory processes is only now being fully appreciated.

Biotransformation of macrolide antibiotics using enriched activated sludge culture: Kinetics, transformation routes and ecotoxicological evaluation.

PubMed

Terzic, Senka; Udikovic-Kolic, Nikolina; Jurina, Tamara; Krizman-Matasic, Ivona; Senta, Ivan; Mihaljevic, Ivan; Loncar, Jovica; Smital, Tvrtko; Ahel, Marijan

2018-05-05

The biotransformation of three prominent macrolide antibiotics (azithromycin, clarithromycin and erythromycin) by an activated sludge culture, which was adapted to high concentrations of azithromycin (10 mg/L) was investigated. The study included determination of removal kinetics of the parent compounds, identification of their major biotransformation products (TPs) and assessment of ecotoxicological effects of biotransformation. The chemical analyses were performed by ultra-performance liquid chromatography/quadrupole-time-of-flight mass spectrometry, which enabled a tentative identification of TPs formed during the experiments. The ecotoxicological evaluation included two end-points, residual antibiotic activity and toxicity to freshwater algae. The enriched activated sludge culture was capable of degrading all studied macrolide compounds with high removal efficiencies (>99%) of the parent compounds at elevated concentrations (10 mg/L). The elimination of all three macrolide antibiotics was associated with the formation of different TPs, including several novel compounds previously unreported in the literature. Some of the TPs were rather abundant and contributed significantly to the overall mass balance at the end of the biodegradation experiments. Biodegradation of all investigated macrolides was associated with a pronounced reduction of the residual antibiotic activity and algal toxicity, indicating a rather positive ecotoxicological outcome of the biotransformation processes achieved by the enriched sludge culture. Copyright © 2018 Elsevier B.V. All rights reserved.

Coral and macroalgal exudates vary in neutral sugar composition and differentially enrich reef bacterioplankton lineages

PubMed Central

Nelson, Craig E; Goldberg, Stuart J; Wegley Kelly, Linda; Haas, Andreas F; Smith, Jennifer E; Rohwer, Forest; Carlson, Craig A

2013-01-01

Increasing algal cover on tropical reefs worldwide may be maintained through feedbacks whereby algae outcompete coral by altering microbial activity. We hypothesized that algae and coral release compositionally distinct exudates that differentially alter bacterioplankton growth and community structure. We collected exudates from the dominant hermatypic coral holobiont Porites spp. and three dominant macroalgae (one each Ochrophyta, Rhodophyta and Chlorophyta) from reefs of Mo'orea, French Polynesia. We characterized exudates by measuring dissolved organic carbon (DOC) and fractional dissolved combined neutral sugars (DCNSs) and subsequently tracked bacterioplankton responses to each exudate over 48 h, assessing cellular growth, DOC/DCNS utilization and changes in taxonomic composition (via 16S rRNA amplicon pyrosequencing). Fleshy macroalgal exudates were enriched in the DCNS components fucose (Ochrophyta) and galactose (Rhodophyta); coral and calcareous algal exudates were enriched in total DCNS but in the same component proportions as ambient seawater. Rates of bacterioplankton growth and DOC utilization were significantly higher in algal exudate treatments than in coral exudate and control incubations with each community selectively removing different DCNS components. Coral exudates engendered the smallest shift in overall bacterioplankton community structure, maintained high diversity and enriched taxa from Alphaproteobacteria lineages containing cultured representatives with relatively few virulence factors (VFs) (Hyphomonadaceae and Erythrobacteraceae). In contrast, macroalgal exudates selected for less diverse communities heavily enriched in copiotrophic Gammaproteobacteria lineages containing cultured pathogens with increased VFs (Vibrionaceae and Pseudoalteromonadaceae). Our results demonstrate that algal exudates are enriched in DCNS components, foster rapid growth of bacterioplankton and select for bacterial populations with more potential VFs than

Bacterial cellulose production by Gluconacetobacter xylinus by employing alternative culture media.

PubMed

Jozala, Angela Faustino; Pértile, Renata Aparecida Nedel; dos Santos, Carolina Alves; de Carvalho Santos-Ebinuma, Valéria; Seckler, Marcelo Martins; Gama, Francisco Miguel; Pessoa, Adalberto

2015-02-01

Bacterial cellulose (BC) is used in different fields as a biological material due to its unique properties. Despite there being many BC applications, there still remain many problems associated with bioprocess technology, such as increasing productivity and decreasing production cost. New technologies that use waste from the food industry as raw materials for culture media promote economic advantages because they reduce environmental pollution and stimulate new research for science sustainability. For this reason, BC production requires optimized conditions to increase its application. The main objective of this study was to evaluate BC production by Gluconacetobacter xylinus using industry waste, namely, rotten fruits and milk whey, as culture media. Furthermore, the structure of BC produced at different conditions was also determined. The culture media employed in this study were composed of rotten fruit collected from the disposal of free markets, milk whey from a local industrial disposal, and their combination, and Hestrin and Schramm media was used as standard culture media. Although all culture media studied produced BC, the highest BC yield-60 mg/mL-was achieved with the rotten fruit culture. Thus, the results showed that rotten fruit can be used for BC production. This culture media can be considered as a profitable alternative to generate high-value products. In addition, it combines environmental concern with sustainable processes that can promote also the reduction of production cost.

Enrichment and characterization of anaerobic TNT-degrading bacteria

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

Roberts, D.J.; Pendharkar, S.

1995-12-31

Three media constitutions were used to enrich for mixed cultures capable of degrading 2,4,6-trinitrotoluene (TNT) under strictly anaerobic conditions. The media were derived from a mineral salts solution buffered to pH 7 with CO{sub 2}/bicarbonate and all contained TNT. The cultures were enriched in the TNT mineral salts medium or the TNT mineral salts medium supplemented with glucose, yeast extract, or ammonia in various combinations. Inocula were obtained from a treated soil, previously contaminated with dinoseb and then treated using anaerobic procedures, or from a bench-top aqueous culture, maintained with an extract from a munitions-contaminated soil for more than 4more » years. Several cultures reduced TNT, producing 4-amino-2,6-dinitrotoluene and 2,4-diamino-6-nitrotoluene as the major products. The cultures were unable to effectively remove TNT when cross-transferred to the media they were not enriched on, suggesting that different media had enriched different subcultures form the original inoculum. The treated soil provided the most successful inoculum. Two media were chosen for further studies. Medium 1 contained TNT and glucose and produced a culture that might have used TNT as a nitrogen source. Medium 2, containing TNT and yeast extract, enriched cultures that degraded TNT, accumulating small amounts of p-cresol during the degradation.« less

Limiting and detecting bacterial contamination of apheresis platelets: inlet-line diversion and increased culture volume improve component safety.

PubMed

Eder, Anne F; Kennedy, Jean M; Dy, Beth A; Notari, Edward P; Skeate, Robert; Bachowski, Gary; Mair, David C; Webb, Jonathan S; Wagner, Stephen J; Dodd, Roger Y; Benjamin, Richard J

2009-08-01

Septic transfusion reactions to apheresis platelets (PLTs) continue to occur despite preventive measures. This study evaluated the effect of two operational changes designed to reduce bacterial risk: 1) introducing inlet-line sample diversion on two-arm procedures and 2) increasing the sample volume cultured from 4 to 8 mL from all donations. Aerobic culture results and septic transfusion reactions reported between December 1, 2006, and July 31, 2008 (Period 2), were compared to March 1, 2004, to May 31, 2006 (Period 1). During Period 2, a total of 781,936 apheresis PLT collections were cultured, of which 130 donations (1:6015) were confirmed positive and 9 (1:86,882) had negative culture results but were associated with 11 septic reactions. Confirmed-positive cultures from two-arm procedures decreased (27.2 to 14.7 per 105 collections; odds ratio [OR], 0.54; 95% confidence interval [CI], 0.41-0.70) in Period 2, owing to a lower rate of skin flora contamination. Detection of contamination of one-arm collections significantly increased by 54% in Period 2 (13.7 vs. 21.1 per 105 collections; OR, 1.54; 95% CI, 1.05-2.27). Fewer septic transfusion reactions occurred in Period 2, but the difference did not reach significance (1.7 vs. 1.2 per 105 donations; OR, 0.68; 95% CI, 0.30-1.53). Inlet-line diversion decreased bacterial contamination during two-arm collections by more than 46%. Concurrently, doubling the sample volume was associated with a 54% relative increase in culture sensitivity. These interventions act cooperatively to decrease bacterial risk.

Enrichment of a mixed microbial culture for polyhydroxyalkanoates production: Effect of pH and N and P concentrations.

PubMed

Montiel-Jarillo, Gabriela; Carrera, Julián; Suárez-Ojeda, María Eugenia

2017-04-01

Polyhydroxyalkanoates (PHA) are biopolymers that can be an alternative against conventional plastics. The study reported herein evaluated the enrichment of a mixed microbial culture (MMC) operated under feast/famine regime and different pHs in a sequencing batch reactor (SBR) using acetate as sole carbon source to produce polyhydroxyalkanoates (PHAs). The enrichment step was evaluated at controlled pH of 7.5 and also without pH control (averaged value of 9.0). The acetate uptake rate (-q S ) of both enrichments at the end of the experimental period exhibited similar behaviour being about 0.18CmolAcCmolX -1 h -1 and 0.19CmolAcCmolX -1 h -1 for SBR-A and SBR-B, respectively. However, the PHA-storing capacity of the biomass enriched without pH control was better, exhibiting a maximum PHA content of 36% (gPHAg -1 VSS) with a PHA production rate (q PHA ) of 0.16CmolPHACmolX -1 h -1 . Batch experiments were performed to evaluate PHA-storing capacity of the enriched culture at different pHs and nutrients concentrations. In the pH experiments (without nutrient limitation), it was found that in the absence of controlled pH, the enriched biomass exhibited a PHA content of 44% gPHAg -1 VSS with -q S and PHA to substrate yield (Y PHA/Ac ) of 0.57CmolAcCmolX -1 h -1 and 0.33CmolPHACmolAc -1 , respectively. Regarding the experiments at variable nutrients concentration (pH ranging 8.8 to 9.2), the results indicate that the PHA content in the enriched biomass is significantly higher being around 51% gPHAg -1 VSS under nitrogen limitation. This work demonstrated the feasibility of the enrichment of a MMC with PHA storage ability without pH control. Results also suggest that better PHAs contents and substrate uptake rates are obtained without controlling the pH in the accumulation step. Finally, this work also highlights the importance of understanding the role of nutrients concentration during the accumulation step. Copyright © 2017 Elsevier B.V. All rights reserved.

Draft Genome Sequence of a Novel Thermofilum sp. Strain from a New Zealand Hot Spring Enrichment Culture

DOE PAGES

Reysenbach, Anna-Louise; Donaho, John; Hinsch, Todd; ...

2018-02-22

A draft genome of a newThermofilumsp. strain was obtained from an enrichment culture metagenome. Like its relatives,Thermofilumsp. strain NZ13 is adapted to organic-rich thermal environments and has to depend on other organisms and the environment for some key amino acids, purines, and cofactors.

Draft Genome Sequence of a Novel Thermofilum sp. Strain from a New Zealand Hot Spring Enrichment Culture

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

Reysenbach, Anna-Louise; Donaho, John; Hinsch, Todd

A draft genome of a newThermofilumsp. strain was obtained from an enrichment culture metagenome. Like its relatives,Thermofilumsp. strain NZ13 is adapted to organic-rich thermal environments and has to depend on other organisms and the environment for some key amino acids, purines, and cofactors.

Implementation of secondary bacterial culture testing of platelets to mitigate residual risk of septic transfusion reactions.

PubMed

Bloch, Evan M; Marshall, Christi E; Boyd, Joan S; Shifflett, Lisa; Tobian, Aaron A R; Gehrie, Eric A; Ness, Paul M

2018-04-01

Bacterial contamination of platelets remains a major transfusion-associated risk despite long-standing safety measures in the United States. We evaluated an approach using secondary bacterial culture (SBC) to contend with residual risk of bacterial contamination. Phased implementation of SBC was initiated in October 2016 for platelets (all apheresis collected) received at our institution from the blood donor center (Day 3 post collection). Platelet products were sampled aseptically (5 mL inoculated into an aerobic bottle [BacT/ALERT BPA, BioMerieux, Inc.]) by the blood bank staff upon receipt, using a sterile connection device and sampling kit. The platelet sample was inoculated into an aerobic blood culture bottle and incubated at 35°C for 3 days. The cost of SBC was calculated on the basis of consumables and labor costs at time of implementation. In the 13 months following implementation (October 6, 2016, to November 30, 2017), 23,044/24,653 (93.47%) platelet products underwent SBC. A total of eight positive cultures were detected (incidence 1 in 2881 platelet products), seven of which were positive within 24 hours of SBC. Coagulase negative Staphyloccus spp. were identified in four cases. Five of the eight cases were probable true positive (repeat reactive) and interdicted (cost per averted case was US$77,935). The remaining three cases were indeterminate. No septic transfusion reactions were reported during the observation period. We demonstrate the feasibility of SBC of apheresis platelets to mitigate bacterial risk. SBC is lower cost than alternative measures (e.g., pathogen reduction and point-of-release testing) and can be integrated into workflow at hospital transfusion services. © 2018 AABB.

Looking Beyond Respiratory Cultures: Microbiome-Cytokine Signatures of Bacterial Pneumonia and Tracheobronchitis in Lung Transplant Recipients.

PubMed

Shankar, J; Nguyen, M H; Crespo, M M; Kwak, E J; Lucas, S K; McHugh, K J; Mounaud, S; Alcorn, J F; Pilewski, J M; Shigemura, N; Kolls, J K; Nierman, W C; Clancy, C J

2016-06-01

Bacterial pneumonia and tracheobronchitis are diagnosed frequently following lung transplantation. The diseases share clinical signs of inflammation and are often difficult to differentiate based on culture results. Microbiome and host immune-response signatures that distinguish between pneumonia and tracheobronchitis are undefined. Using a retrospective study design, we selected 49 bronchoalveolar lavage fluid samples from 16 lung transplant recipients associated with pneumonia (n = 8), tracheobronchitis (n = 12) or colonization without respiratory infection (n = 29). We ensured an even distribution of Pseudomonas aeruginosa or Staphylococcus aureus culture-positive samples across the groups. Bayesian regression analysis identified non-culture-based signatures comprising 16S ribosomal RNA microbiome profiles, cytokine levels and clinical variables that characterized the three diagnoses. Relative to samples associated with colonization, those from pneumonia had significantly lower microbial diversity, decreased levels of several bacterial genera and prominent multifunctional cytokine responses. In contrast, tracheobronchitis was characterized by high microbial diversity and multifunctional cytokine responses that differed from those of pneumonia-colonization comparisons. The dissimilar microbiomes and cytokine responses underlying bacterial pneumonia and tracheobronchitis following lung transplantation suggest that the diseases result from different pathogenic processes. Microbiomes and cytokine responses had complementary features, suggesting that they are closely interconnected in the pathogenesis of both diseases. © Copyright 2016 The American Society of Transplantation and the American Society of Transplant Surgeons.

Gene and transcript abundances of bacterial type III secretion systems from the rumen microbiome are correlated with methane yield in sheep.

PubMed

Kamke, Janine; Soni, Priya; Li, Yang; Ganesh, Siva; Kelly, William J; Leahy, Sinead C; Shi, Weibing; Froula, Jeff; Rubin, Edward M; Attwood, Graeme T

2017-08-08

Ruminants are important contributors to global methane emissions via microbial fermentation in their reticulo-rumens. This study is part of a larger program, characterising the rumen microbiomes of sheep which vary naturally in methane yield (g CH 4 /kg DM/day) and aims to define differences in microbial communities, and in gene and transcript abundances that can explain the animal methane phenotype. Rumen microbiome metagenomic and metatranscriptomic data were analysed by Gene Set Enrichment, sparse partial least squares regression and the Wilcoxon Rank Sum test to estimate correlations between specific KEGG bacterial pathways/genes and high methane yield in sheep. KEGG genes enriched in high methane yield sheep were reassembled from raw reads and existing contigs and analysed by MEGAN to predict their phylogenetic origin. Protein coding sequences from Succinivibrio dextrinosolvens strains were analysed using Effective DB to predict bacterial type III secreted proteins. The effect of S. dextrinosolvens strain H5 growth on methane formation by rumen methanogens was explored using co-cultures. Detailed analysis of the rumen microbiomes of high methane yield sheep shows that gene and transcript abundances of bacterial type III secretion system genes are positively correlated with methane yield in sheep. Most of the bacterial type III secretion system genes could not be assigned to a particular bacterial group, but several genes were affiliated with the genus Succinivibrio, and searches of bacterial genome sequences found that strains of S. dextrinosolvens were part of a small group of rumen bacteria that encode this type of secretion system. In co-culture experiments, S. dextrinosolvens strain H5 showed a growth-enhancing effect on a methanogen belonging to the order Methanomassiliicoccales, and inhibition of a representative of the Methanobrevibacter gottschalkii clade. This is the first report of bacterial type III secretion system genes being associated with high

Investigation of Endophytic Bacterial Community in Supposedly Axenic Cultures of Pineapple and Orchids with Evidence on Abundant Intracellular Bacteria.

PubMed

Esposito-Polesi, Natalia Pimentel; de Abreu-Tarazi, Monita Fiori; de Almeida, Cristina Vieira; Tsai, Siu Mui; de Almeida, Marcílio

2017-01-01

Asepsis, defined as the absence of microbial contamination, is one of the most important requirements of plant micropropagation. In long-term micropropagated cultures, there may occasionally occur scattered microorganism growth in the culture medium. These microorganisms are common plant components and are known as latent endophytes. Thus, the aim of this research was to investigate the presence of endophytic bacteria in asymptomatic pineapple and orchid microplants, which were cultivated in three laboratories for 1 year. Isolation and characterization of bacterial isolates, PCR-DGGE from total genomic DNA of microplants and ultrastructural analysis of leaves were performed. In the culture-dependent technique, it was only possible to obtain bacterial isolates from pineapple microplants. In this case, the bacteria genera identified in the isolation technique were Bacillus, Acinetobacter, and Methylobacterium. The scanning electron microscopy and transmission electron microscopy (SEM and TEM) analyses revealed the presence of endophytic bacteria in intracellular spaces in the leaves of pineapple and orchid microplants, independent of the laboratory or cultivation protocol. Our results strongly indicate that there are endophytic bacterial communities inhabiting the microplants before initiation of the in vitro culture and that some of these endophytes persist in their latent form and can also grow in the culture medium even after long-term micropropagation, thus discarding the concept of "truly axenic plants."

Biodegradation of γ-hexachlorocyclohexane by transgenic hairy root cultures of Cucurbita moschata that accumulate recombinant bacterial LinA.

PubMed

Nanasato, Yoshihiko; Namiki, Sayuri; Oshima, Masao; Moriuchi, Ryota; Konagaya, Ken-Ichi; Seike, Nobuyasu; Otani, Takashi; Nagata, Yuji; Tsuda, Masataka; Tabei, Yutaka

2016-09-01

γ-HCH was successfully degraded using LinA-expressed transgenic hairy root cultures of Cucurbita moschata . Fusing an endoplasmic reticulum-targeting signal peptide to LinA was essential for stable accumulation in the hairy roots. The pesticide γ-hexachlorocyclohexane (γ-HCH) is a persistent organic pollutant (POP) that raises public health and environmental pollution concerns worldwide. Although several isolates of γ-HCH-degrading bacteria are available, inoculating them directly into γ-HCH-contaminated soil is ineffective because of the bacterial survival rate. Cucurbita species incorporate significant amounts of POPs from soils compared with other plant species. Here, we describe a novel bioremediation strategy that combines the bacterial degradation of γ-HCH and the efficient uptake of γ-HCH by Cucurbita species. We produced transgenic hairy root cultures of Cucurbita moschata that expressed recombinant bacterial linA, isolated from the bacterium Sphingobium japonicum UT26. The LinA protein was accumulated stably in the hairy root cultures by fusing an endoplasmic reticulum (ER)-targeting signal peptide to LinA. Then, we demonstrated that the cultures degraded more than 90 % of γ-HCH (1 ppm) overnight and produced the γ-HCH metabolite 1,2,4-trichlorobenzene, indicating that LinA degraded γ-HCH. These results indicate that the gene linA has high potential for phytoremediation of environmental γ-HCH.

Formulation of Biocides Increases Antimicrobial Potency and Mitigates the Enrichment of Nonsusceptible Bacteria in Multispecies Biofilms

PubMed Central

Forbes, Sarah; Cowley, Nicola; Mistry, Hitesh; Amézquita, Alejandro

2017-01-01

ABSTRACT The current investigation aimed to generate data to inform the development of risk assessments of biocide usage. Stabilized domestic drain biofilm microcosms were exposed daily over 6 months to increasing concentrations (0.01% to 1%) of the biocide benzalkonium chloride (BAC) in a simple aqueous solution (BAC-s) or in a complex formulation (BAC-f) representative of a domestic cleaning agent. Biofilms were analyzed by culture, differentiating by bacterial functional group and by BAC or antibiotic susceptibility. Bacterial isolates were identified by 16S rRNA sequencing, and changes in biofilm composition were assessed by high-throughput sequencing. Exposure to BAC-f resulted in significantly larger reductions in levels of viable bacteria than exposure to BAC-s, while bacterial diversity greatly decreased during exposure to both BAC-s and BAC-f, as evidenced by sequencing and viable counts. Increases in the abundance of bacteria exhibiting reduced antibiotic or BAC susceptibility following exposure to BAC at 0.1% were significantly greater for BAC-s than BAC-f. Bacteria with reduced BAC and antibiotic susceptibility were generally suppressed by higher BAC concentrations, and formulation significantly enhanced this effect. Significant decreases in the antimicrobial susceptibility of bacteria isolated from the systems before and after long-term BAC exposure were not detected. In summary, dose-dependent suppression of bacterial viability by BAC was enhanced by formulation. Biocide exposure decreased bacterial diversity and transiently enriched populations of organisms with lower antimicrobial susceptibility, and the effects were subsequently suppressed by exposure to 1% BAC-f, the concentration most closely reflecting deployment in formulated products. IMPORTANCE Assessment of the risks of biocide use has been based mainly on the exposure of axenic cultures of bacteria to biocides in simple aqueous solutions. The current investigation aimed to assess the

Effects of carbon sources on the enrichment of halophilic polyhydroxyalkanoate-storing mixed microbial culture in an aerobic dynamic feeding process

PubMed Central

Cui, You-Wei; Zhang, Hong-Yu; Lu, Peng-Fei; Peng, Yong-Zhen

2016-01-01

Microbial polyhydroxyalkanoate (PHA) production serves as a substitute for petroleum-based plastics. Enriching mixed microbial cultures (MMCs) with the capacity to store PHA is a key precursor for low-cost PHA production. This study investigated the impact of carbon types on enrichment outcomes. Three MMCs were separately fed by acetate sodium, glucose, and starch as an enriching carbon source, and were exposed to long-term aerobic dynamic feeding (ADF) periods. The PHA production capacity, kinetics and stoichiometry of the enrichments, the PHA composition, and the microbial diversity and community composition were explored to determine carbon and enrichment correlations. After 350-cycle enriching periods under feast-famine (F-F) regimes, the MMCs enriched by acetate sodium and glucose contained a maximum PHA content of 64.7% and 60.5% cell dry weight (CDW). The starch-enriched MMC only had 27.3% CDW of PHA. High-throughput sequencing revealed that non-PHA bacteria survived alongside PHA storing bacteria, even under severe F-F selective pressure. Genus of Pseudomonas and Stappia were the possible PHA accumulating bacteria in acetate-enriched MMC. Genus of Oceanicella, Piscicoccus and Vibrio were found as PHA accumulating bacteria in glucose-enriched MMC. Vibrio genus was the only PHA accumulating bacteria in starch-enriched MMC. The community diversity and composition were regulated by the substrate types. PMID:27485896

Effects of carbon sources on the enrichment of halophilic polyhydroxyalkanoate-storing mixed microbial culture in an aerobic dynamic feeding process

NASA Astrophysics Data System (ADS)

Cui, You-Wei; Zhang, Hong-Yu; Lu, Peng-Fei; Peng, Yong-Zhen

2016-08-01

Microbial polyhydroxyalkanoate (PHA) production serves as a substitute for petroleum-based plastics. Enriching mixed microbial cultures (MMCs) with the capacity to store PHA is a key precursor for low-cost PHA production. This study investigated the impact of carbon types on enrichment outcomes. Three MMCs were separately fed by acetate sodium, glucose, and starch as an enriching carbon source, and were exposed to long-term aerobic dynamic feeding (ADF) periods. The PHA production capacity, kinetics and stoichiometry of the enrichments, the PHA composition, and the microbial diversity and community composition were explored to determine carbon and enrichment correlations. After 350-cycle enriching periods under feast-famine (F-F) regimes, the MMCs enriched by acetate sodium and glucose contained a maximum PHA content of 64.7% and 60.5% cell dry weight (CDW). The starch-enriched MMC only had 27.3% CDW of PHA. High-throughput sequencing revealed that non-PHA bacteria survived alongside PHA storing bacteria, even under severe F-F selective pressure. Genus of Pseudomonas and Stappia were the possible PHA accumulating bacteria in acetate-enriched MMC. Genus of Oceanicella, Piscicoccus and Vibrio were found as PHA accumulating bacteria in glucose-enriched MMC. Vibrio genus was the only PHA accumulating bacteria in starch-enriched MMC. The community diversity and composition were regulated by the substrate types.

Effects of carbon sources on the enrichment of halophilic polyhydroxyalkanoate-storing mixed microbial culture in an aerobic dynamic feeding process.

PubMed

Cui, You-Wei; Zhang, Hong-Yu; Lu, Peng-Fei; Peng, Yong-Zhen

2016-08-03

Microbial polyhydroxyalkanoate (PHA) production serves as a substitute for petroleum-based plastics. Enriching mixed microbial cultures (MMCs) with the capacity to store PHA is a key precursor for low-cost PHA production. This study investigated the impact of carbon types on enrichment outcomes. Three MMCs were separately fed by acetate sodium, glucose, and starch as an enriching carbon source, and were exposed to long-term aerobic dynamic feeding (ADF) periods. The PHA production capacity, kinetics and stoichiometry of the enrichments, the PHA composition, and the microbial diversity and community composition were explored to determine carbon and enrichment correlations. After 350-cycle enriching periods under feast-famine (F-F) regimes, the MMCs enriched by acetate sodium and glucose contained a maximum PHA content of 64.7% and 60.5% cell dry weight (CDW). The starch-enriched MMC only had 27.3% CDW of PHA. High-throughput sequencing revealed that non-PHA bacteria survived alongside PHA storing bacteria, even under severe F-F selective pressure. Genus of Pseudomonas and Stappia were the possible PHA accumulating bacteria in acetate-enriched MMC. Genus of Oceanicella, Piscicoccus and Vibrio were found as PHA accumulating bacteria in glucose-enriched MMC. Vibrio genus was the only PHA accumulating bacteria in starch-enriched MMC. The community diversity and composition were regulated by the substrate types.

Short-term in-vitro culture of goat enriched spermatogonial stem cells using different serum concentrations.

PubMed

Bahadorani, M; Hosseini, S M; Abedi, P; Hajian, M; Hosseini, S E; Vahdati, A; Baharvand, H; Nasr-Esfahani, Mohammad H

2012-01-01

To investigate the effect of serum supplementing on short-term culture, fate determination and gene expression of goat spermatogonial stem cells (SSCs). Crude testicular cells were plated over Datura-Stramonium Agglutinin (DSA) for 1 h, and non-adhering cells were cultured in the presence of different serum concentrations (1, 5, 10, and 15%) for 7 days in a highly enriched medium initially developed in mice. Colonies developed in each group were used for the assessment of morphology, immunocytochemistry, and gene expression. Brief incubation of testicular cells with DSA resulted in a significant increase in the number of cells that expressed the germ cell marker (VASA). The expression of THY1, a specific marker of undifferentiated spermatogonia, was significantly higher in colonies developed in the presence of 1% rather than 5, 10 and 15% serum. Goat SSCs could proliferate and maintain in SSC culture media for 1 week at serum concentrations as low as 1%, while higher concentrations had detrimental effects on SSC culture/expansion.

[Comparative studies of methods of salmonella enrichment (author's transl)].

PubMed

Pietzsch, O; Kretschmer, F J; Bulling, E

1975-07-01

Eight different methods of salmonella enrichment were compared in two series of experiments involving 100 samples of whole-egg powder and 80 samples of frozen whole liquid egg, respectively. 66 out of a total of 100 samples of whole-egg powder had been artificially infected with varying numbers of S. typhi-murium; 60 out of 80 samples of frozen whole liquid egg were found to be naturally infected with various salmonella species. 3 of the 8 methods (Table 1) were compared within an international collaborative study with 14 laboratories in 11 countries participating. A reduction of the pre-enrichment period from 18 to 6 hours and of volumes used in pre-enrichment and selective enrichment from 10 and 100 ml, respectively to 1 and 10 ml, respectively were found to have adverse influence upon the result of isolations, in particular in the case of weakly infected samples. In contrast, extended incubation over 48 hours as well as preparation of two sub-cultures on solid selective media following incubation of enrichment cultures over 18-24 hours and 42-48 hours, respectively always resulted in a certain increase of salmonella yield which, however, exhibited gradual differences for the individual methods examined. Preparation of a 2nd sub-culture meant, in particular, a decisive improvement of the result of isolations from artificially infected samples if selenite-cystine enrichment volumes were 10 and 100 ml, respectively. The best results could be obtained by means of the following methods of enrichment: Pre-enrichment of material in buffered peptone water at 37 degrees C over 18 hours; pipetting of 10 ml inoculated and incubated pre-enriched material into 100 ml selenite-cystine or tetrathionate enrichment medium according to MULLER-KAUFFMANN; onward incubation of the enrichment culture at 43 degrees C over 48 hours; and preparation of sub-cultures on solid selective media after 24 and 48 hours. The method using tetrathionate enrichment medium was found to be most

Bacterial siderophores efficiently provide iron to iron-starved tomato plants in hydroponics culture.

PubMed

Radzki, W; Gutierrez Mañero, F J; Algar, E; Lucas García, J A; García-Villaraco, A; Ramos Solano, B

2013-09-01

Iron is one of the essential elements for a proper plant development. Providing plants with an accessible form of iron is crucial when it is scant or unavailable in soils. Chemical chelates are the only current alternative and are highly stable in soils, therefore, posing a threat to drinking water. The aim of this investigation was to quantify siderophores produced by two bacterial strains and to determine if these bacterial siderophores would palliate chlorotic symptoms of iron-starved tomato plants. For this purpose, siderophore production in MM9 medium by two selected bacterial strains was quantified, and the best was used for biological assay. Bacterial culture media free of bacteria (S) and with bacterial cells (BS), both supplemented with Fe were delivered to 12-week-old plants grown under iron starvation in hydroponic conditions; controls with full Hoagland solution, iron-free Hoagland solution and water were also conducted. Treatments were applied twice along the experiment, with a week in between. At harvest, plant yield, chlorophyll content and nutritional status in leaves were measured. Both the bacterial siderophore treatments significantly increased plant yield, chlorophyll and iron content over the positive controls with full Hoagland solution, indicating that siderophores are effective in providing Fe to the plant, either with or without the presence of bacteria. In summary, siderophores from strain Chryseobacterium C138 are effective in supplying Fe to iron-starved tomato plants by the roots, either with or without the presence of bacteria. Based on the amount of siderophores produced, an effective and economically feasible organic Fe chelator could be developed.

Clinical features, cytology and bacterial culture results in dogs with and without cheilitis and comparison of three sampling techniques.

PubMed

Doelle, Maren; Loeffler, Anette; Wolf, Katharina; Kostka, Veit; Linek, Monika

2016-06-01

Cheilitis is a common presentation in dogs associated with a variety of skin diseases and often complicated by microbial infections. To describe and compare clinical and cytological features and bacterial culture results from the lower lips of dogs with cheilitis (as compared to healthy controls), and to evaluate three cytology sampling techniques for their abilities to differentiate between the groups. Fifty six dogs with cheilitis and 54 controls. Anatomy and clinical signs of the lower lip were recorded. Cytology samples taken by tape strip, direct impression and swabs rolled over skin were scored semiquantitatively for microorganisms, inflammatory cells and keratinocytes. Cytology scores were correlated with semiquantitative bacterial culture scores. Pure breeds, frequency of lip folds and all cytology scores except keratinocytes were higher in dogs with cheilitis than in controls, but a substantial overlap was seen in all microorganisms between the groups. Hypersensitivity disorders were diagnosed in 40 of 56 dogs with cheilitis. The tape strip technique yielded the greatest differences between groups. Bacterial growth was reported in 100% of dogs with cheilitis and in 93% of the controls. Pathogens such as Staphylococcus pseudintermedius, Escherichia coli and Pseudomonas spp were found more frequently in dogs with cheilitis. Cytology and bacterial culture were poorly correlated. Cheilitis was associated with primary hypersensitivity disorders and the presence of a lip fold was a predisposing factor. Results of aerobic culture were similar to prior studies on pyoderma of other body sites, except for higher rates of Pseudomonas spp. isolation. © 2016 ESVD and ACVD.

Comparison of microbial DNA enrichment tools for metagenomic whole genome sequencing.

PubMed

Thoendel, Matthew; Jeraldo, Patricio R; Greenwood-Quaintance, Kerryl E; Yao, Janet Z; Chia, Nicholas; Hanssen, Arlen D; Abdel, Matthew P; Patel, Robin

2016-08-01

Metagenomic whole genome sequencing for detection of pathogens in clinical samples is an exciting new area for discovery and clinical testing. A major barrier to this approach is the overwhelming ratio of human to pathogen DNA in samples with low pathogen abundance, which is typical of most clinical specimens. Microbial DNA enrichment methods offer the potential to relieve this limitation by improving this ratio. Two commercially available enrichment kits, the NEBNext Microbiome DNA Enrichment Kit and the Molzym MolYsis Basic kit, were tested for their ability to enrich for microbial DNA from resected arthroplasty component sonicate fluids from prosthetic joint infections or uninfected sonicate fluids spiked with Staphylococcus aureus. Using spiked uninfected sonicate fluid there was a 6-fold enrichment of bacterial DNA with the NEBNext kit and 76-fold enrichment with the MolYsis kit. Metagenomic whole genome sequencing of sonicate fluid revealed 13- to 85-fold enrichment of bacterial DNA using the NEBNext enrichment kit. The MolYsis approach achieved 481- to 9580-fold enrichment, resulting in 7 to 59% of sequencing reads being from the pathogens known to be present in the samples. These results demonstrate the usefulness of these tools when testing clinical samples with low microbial burden using next generation sequencing. Copyright © 2016 Elsevier B.V. All rights reserved.

Metatranscriptome of an Anaerobic Benzene-Degrading, Nitrate-Reducing Enrichment Culture Reveals Involvement of Carboxylation in Benzene Ring Activation

PubMed Central

Luo, Fei; Gitiafroz, Roya; Devine, Cheryl E.; Gong, Yunchen; Hug, Laura A.; Raskin, Lutgarde

2014-01-01

The enzymes involved in the initial steps of anaerobic benzene catabolism are not known. To try to elucidate this critical step, a metatranscriptomic analysis was conducted to compare the genes transcribed during the metabolism of benzene and benzoate by an anaerobic benzene-degrading, nitrate-reducing enrichment culture. RNA was extracted from the mixed culture and sequenced without prior mRNA enrichment, allowing simultaneous examination of the active community composition and the differential gene expression between the two treatments. Ribosomal and mRNA sequences attributed to a member of the family Peptococcaceae from the order Clostridiales were essentially only detected in the benzene-amended culture samples, implicating this group in the initial catabolism of benzene. Genes similar to each of two subunits of a proposed benzene-carboxylating enzyme were transcribed when the culture was amended with benzene. Anaerobic benzoate degradation genes from strict anaerobes were transcribed only when the culture was amended with benzene. Genes for other benzoate catabolic enzymes and for nitrate respiration were transcribed in both samples, with those attributed to an Azoarcus species being most abundant. These findings indicate that the mineralization of benzene starts with its activation by a strict anaerobe belonging to the Peptococcaceae, involving a carboxylation step to form benzoate. These data confirm the previously hypothesized syntrophic association between a benzene-degrading Peptococcaceae strain and a benzoate-degrading denitrifying Azoarcus strain for the complete catabolism of benzene with nitrate as the terminal electron acceptor. PMID:24795366

The Importance of Bacterial Culture to Food Microbiology in the Age of Genomics.

PubMed

Gill, Alexander

2017-01-01

Culture-based and genomics methods provide different insights into the nature and behavior of bacteria. Maximizing the usefulness of both approaches requires recognizing their limitations and employing them appropriately. Genomic analysis excels at identifying bacteria and establishing the relatedness of isolates. Culture-based methods remain necessary for detection and enumeration, to determine viability, and to validate phenotype predictions made on the bias of genomic analysis. The purpose of this short paper is to discuss the application of culture-based analysis and genomics to the questions food microbiologists routinely need to ask regarding bacteria to ensure the safety of food and its economic production and distribution. To address these issues appropriate tools are required for the detection and enumeration of specific bacterial populations and the characterization of isolates for, identification, phylogenetics, and phenotype prediction.

Bacterial communities in sediment of a Mediterranean marine protected area.

PubMed

Catania, Valentina; Sarà, Gianluca; Settanni, Luca; Quatrini, Paola

2017-04-01

Biodiversity is crucial in preservation of ecosystems, and bacterial communities play an indispensable role for the functioning of marine ecosystems. The Mediterranean marine protected area (MPA) "Capo Gallo-Isola delle Femmine" was instituted to preserve marine biodiversity. The bacterial diversity associated with MPA sediment was compared with that from sediment of an adjacent harbour exposed to intense nautical traffic. The MPA sediment showed higher diversity with respect to the impacted site. A 16S rDNA clone library of the MPA sediment allowed the identification of 7 phyla: Proteobacteria (78%), Firmicutes (11%), Acidobacteria (3%), Actinobacteria (3%), Bacteroidetes (2%), Planctomycetes (2%), and Cyanobacteria (1%). Analysis of the hydrocarbon (HC)-degrading bacteria was performed using enrichment cultures. Most of the MPA sediment isolates were affiliated with Gram-positive G+C rich bacteria, whereas the majority of taxa in the harbour sediment clustered with Alpha- and Gammaproteobacteria; no Gram-positive HC degraders were isolated from the harbour sediment. Our results show that protection probably has an influence on bacterial diversity, and suggest the importance of monitoring the effects of protection at microbial level as well. This study creates a baseline of data that can be used to assess changes over time in bacterial communities associated with a Mediterranean MPA.

Effect of copper treatment on the composition and function of the bacterial community in the sponge Haliclona cymaeformis.

PubMed

Tian, Ren-Mao; Wang, Yong; Bougouffa, Salim; Gao, Zhao-Ming; Cai, Lin; Zhang, Wei-Peng; Bajic, Vladimir; Qian, Pei-Yuan

2014-11-04

Marine sponges are the most primitive metazoan and host symbiotic microorganisms. They are crucial components of the marine ecological system and play an essential role in pelagic processes. Copper pollution is currently a widespread problem and poses a threat to marine organisms. Here, we examined the effects of copper treatment on the composition of the sponge-associated bacterial community and the genetic features that facilitate the survival of enriched bacteria under copper stress. The 16S rRNA gene sequencing results showed that the sponge Haliclona cymaeformis harbored symbiotic sulfur-oxidizing Ectothiorhodospiraceae and photosynthetic Cyanobacteria as dominant species. However, these autotrophic bacteria decreased substantially after treatment with a high copper concentration, which enriched for a heterotrophic-bacterium-dominated community. Metagenomic comparison revealed a varied profile of functional genes and enriched functions, including bacterial motility and chemotaxis, extracellular polysaccharide and capsule synthesis, virulence-associated genes, and genes involved in cell signaling and regulation, suggesting short-period mechanisms of the enriched bacterial community for surviving copper stress in the microenvironment of the sponge. Microscopic observation and comparison revealed dynamic bacterial aggregation within the matrix and lysis of sponge cells. The bacteriophage community was also enriched, and the complete genome of a dominant phage was determined, implying that a lytic phage cycle was stimulated by the high copper concentration. This study demonstrated a copper-induced shift in the composition of functional genes of the sponge-associated bacterial community, revealing the selective effect of copper treatment on the functions of the bacterial community in the microenvironment of the sponge. This study determined the bacterial community structure of the common sponge Haliclona cymaeformis and examined the effect of copper treatment on

Simplified Protocol for Carba NP Test for Enhanced Detection of Carbapenemase Producers Directly from Bacterial Cultures

PubMed Central

Pasteran, Fernando; Tijet, Nathalie; Melano, Roberto G.

2015-01-01

We compared carbapenemase detection among 266 Gram-negative bacilli (161 carbapenemase producers) using the Carba NP tests issued by the CLSI (CNPt-CLSI) and a novel protocol (CNPt-direct) designed for carbapenemase detection direct from bacterial cultures (instead of bacterial extracts required by the CLSI tests). The specificities were comparable (100%), but the CNPt-direct was more sensitive (98% versus 84%). The CNPt-direct was easier to perform due to the direct use of colonies and offered a more robust detection of carbapenemase producers. PMID:26424841

Bacterial degradation of synthetic and kraft lignin by axenic and mixed culture and their metabolic products.

PubMed

Chandra, Ram; Bharagava, Ram Naresh

2013-11-01

Pulp paper mill effluent has high pollution load due to presence of lignin and its derivatives as major colouring and polluting constituents. In this study, two lignin degrading bacteria IITRL1 and IITRSU7 were isolated and identified as Citrobacter freundii (FJ581026) and Citrobacter sp. (FJ581023), respectively. In degradation study by axenic and mixed culture, mixed bacterial culture was found more effective compared to axenic culture as it decolourized 85 and 62% of synthetic and kraft lignin whereas in axenic conditions, bacterium IITRL1 and IITRSU7 decolourized 61 and 64% synthetic and 49 and 54% kraft lignin, respectively. Further, the mixed bacterial culture also showed the removal of 71, 58% TOC; 78, 53% AOX; 70, 58% COD and 74, 58% lignin from synthetic and kraft lignin, respectively. The ligninolytic enzyme was characterized as manganese peroxidase by SDS-PAGE yielding a single band of 43 KDa. The HPLC analysis of degraded samples showed reduction as well as shifting of peaks compared to control indicating the degradation as well as transformation of compounds. Further, in GC-MS analysis of synthetic and kraft lignin degraded samples, hexadecanoic acid was found as recalcitrant compounds while 2,4,6-trichloro-phenol, 2,3,4,5-tetrachloro-phenol and pentachloro-phenol were detected as new metabolites.

Enrichment of hexachlorobenzene and 1,3,5-trichlorobenzene transforming bacteria from sediments in Germany and Vietnam.

PubMed

Duan, Tran Hoa; Adrian, Lorenz

2013-07-01

Bacterial cultures were enriched from sediments in Germany and Vietnam reductively dechlorinating hexachlorobenzene and the highly persistent 1,3,5-trichlorobenzene to monochlorobenzene. The main products of the reductive dechlorination of hexachlorobenzene were monochlorobenzene and dichlorobenzenes (1,2-; 1,3- and 1,4-dichlorobenzene) while no trichlorobenzenes accumulated. For the reductive dechlorination of 1,3,5-trichlorobenzene with the mixed culture from Vietnam sediment, 1,3- dichlorobenzene and monochlorobenzene were produced as intermediate and final end-product, respectively. The pattern of dechlorination did not change when the cultures were repeatedly exposed to oxygen over seven transfers demonstrating oxygen tolerance of the dechlorinating bacteria. However, reductive dechlorination of 1,3,5-trichlorobenzene was inhibited by vancomycin at a concentration of 5 mg L(-1). Vancomycin delayed reductive dechlorination of hexachlorobenzene in mixed cultures by about 6 months. When repeatedly applied, vancomycin completely abolished the ability of the mixed culture to transform hexachlorobenzene. Sensitivity to vancomycin and insensitivity to brief exposure of oxygen indicates that the dechlorinating bacteria in the mixed cultures did not belong to the genus Dehalococcoides.

Draft Genome Sequence of a Dictyoglomus sp. from an Enrichment Culture of a New Zealand Geothermal Spring

DOE PAGES

Reysenbach, Anna-Louise; Donaho, John; Kelley, John; ...

2018-03-15

A draft genome of a novelDictyoglomussp., NZ13-RE01, was obtained from a New Zealand hot spring enrichment culture. The 1,927,012-bp genome is similar in both size and G+C content to otherDictyoglomusspp. Like its relatives,Dictyoglomussp. NZ13-RE01 encodes many genes involved in complex carbohydrate metabolism.

Draft Genome Sequence of a Dictyoglomus sp. from an Enrichment Culture of a New Zealand Geothermal Spring

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

Reysenbach, Anna-Louise; Donaho, John; Kelley, John

A draft genome of a novelDictyoglomussp., NZ13-RE01, was obtained from a New Zealand hot spring enrichment culture. The 1,927,012-bp genome is similar in both size and G+C content to otherDictyoglomusspp. Like its relatives,Dictyoglomussp. NZ13-RE01 encodes many genes involved in complex carbohydrate metabolism.

Effect of signal compounds and incubation conditions on the culturability of freshwater bacterioplankton.

PubMed

Bruns, Alke; Nübel, Ulrich; Cypionka, Heribert; Overmann, Jörg

2003-04-01

The effect of signal compounds and of different incubation conditions on the culturability (i.e., the fraction of all cells capable of growth) of natural bacterioplankton from the eutrophic lake Zwischenahner Meer was investigated over a period of 20 months. Numbers of growing cells were determined by the most-probable-number technique in liquid media containing low concentrations (10 micro M) of the signal compounds N-(oxohexanoyl)-DL-homoserine lactone, N-(butyryl)-DL-homoserine lactone, cyclic AMP (cAMP), or ATP. cAMP was the most effective signal compound, leading to significantly increased cultivation efficiencies of up to 10% of the total bacterial counts. Microautoradiography with [2,8-(3)H]cAMP, combined with fluorescence in situ hybridization, demonstrated that cAMP was taken up by 18% of all cells. The bacterial cAMP uptake systems had a very low K(m) value of cultured bacteria by 16S rRNA gene fingerprinting showed that different bacterial phylotypes were recovered in the presence and in the absence of cAMP. Consequently, the addition of cAMP caused a stimulation of otherwise nonculturable bacteria. Phylogenetically different bacteria were also recovered at different temperatures and oxygen partial pressures. Throughout the study period, mainly members of the beta-subclass of the Proteobacteria were cultivated. In addition, some members of the Actinomycetales were enriched. Quantification by culture-independent fluorescence in situ hybridization demonstrated that beta-Proteobacteria and Actinomycetales also dominated the natural bacterioplankton assemblage. Sequence comparison revealed that two members of the Actinomycetales which reached high numbers in the natural bacterioplankton assemblage could actually be enriched by our cultivation approach.

Maize Inoculation with Azospirillum brasilense Ab-V5 Cells Enriched with Exopolysaccharides and Polyhydroxybutyrate Results in High Productivity under Low N Fertilizer Input

PubMed Central

Oliveira, André L. M.; Santos, Odair J. A. P.; Marcelino, Paulo R. F.; Milani, Karina M. L.; Zuluaga, Mónica Y. A.; Zucareli, Claudemir; Gonçalves, Leandro S. A.

2017-01-01

Although Azospirillum strains used in commercial inoculant formulations presents diazotrophic activity, it has been reported that their ability to produce phytohormones plays a pivotal role in plant growth-promotion, leading to a general recommendation of its use in association with regular N-fertilizer doses. In addition, a high variability in the effectiveness of Azospirillum inoculants is still reported under field conditions, contributing to the adoption of the inoculation technology as an additional management practice rather than its use as an alternative practice to the use of chemical inputs in agriculture. To investigate whether the content of stress-resistance biopolymers would improve the viability and performance of Azospirillum inoculants when used as substitute of N-fertilizers, biomass of A. brasilense strain Ab-V5 enriched in exopolysaccharides (EPS) and polyhydroxybutirate (PHB) was produced using a new culture medium developed by factorial mixture design, and the effectiveness of resulting inoculants was evaluated under field conditions. The culture medium formulation extended the log phase of A. brasilense cultures, which presented higher cell counts and increased EPS and PHB contents than observed in the cultures grown in the OAB medium used as control. An inoculation trial with maize conducted under greenhouse conditions and using the biopolymers-enriched Ab-V5 cells demonstrated the importance of EPS and PHB to the long term bacterial viability in soil and to the effectiveness of inoculation. The effectiveness of liquid and peat inoculants prepared with Ab-V5 cells enriched with EPS and PHB was also evaluated under field conditions, using maize as target crop along different seasons, with the inoculants applied directly over seeds or at topdressing under limiting levels of N-fertilization. No additive effect on yield resulted from inoculation under high N fertilizer input, while inoculated plants grown under 80% reduction in N fertilizer

Field comparison of real-time polymerase chain reaction and bacterial culture for identification of bovine mastitis bacteria.

PubMed

Koskinen, M T; Wellenberg, G J; Sampimon, O C; Holopainen, J; Rothkamp, A; Salmikivi, L; van Haeringen, W A; Lam, T J G M; Pyörälä, S

2010-12-01

Fast and reliable identification of the microorganisms causing mastitis is important for management of the disease and for targeting antimicrobial treatment. Methods based on PCR are being used increasingly in mastitis diagnostics. Comprehensive field comparisons of PCR and traditional milk bacteriology have not been available. The results of a PCR kit capable of detecting 11 important etiological agents of mastitis directly from milk in 4h were compared with those of conventional bacterial culture (48h). In total, 1,000 quarter milk samples were taken from cows with clinical or subclinical mastitis, or from clinically healthy quarters with low somatic cell count (SCC). Bacterial culture identified udder pathogens in 600/780 (77%) of the clinical samples, whereas PCR identified bacteria in 691/780 (89%) of the clinical samples. The PCR analysis detected major pathogens in a large number of clinical samples that were negative for the species in culture. These included 53 samples positive for Staphylococcus aureus by PCR, but negative by culture. A total of 137 samples from clinical mastitis, 5 samples from subclinical mastitis, and 1 sample from a healthy quarter were positive for 3 or more bacterial species in PCR, whereas culture identified 3 or more species in 60 samples from clinical mastitis. Culture identified a species not targeted by the PCR test in 44 samples from clinical mastitis and in 9 samples from subclinical mastitis. Low SCC samples provided a small number of positive results both in culture (4/93; 4.3%) and by PCR (7/93; 7.5%). In conclusion, the PCR kit provided several benefits over conventional culture, including speed, automated interpretation of results, and increased sensitivity. This kit holds much promise as a tool to complement traditional methods in identification of pathogens. In conventional mastitis bacteriology, a sample with 3 or more species is considered contaminated, and resampling of the cow is recommended. Further study is

Improved detection of Burkholderia pseudomallei from non-blood clinical specimens using enrichment culture and PCR: narrowing diagnostic gap in resource-constrained settings.

PubMed

Tellapragada, Chaitanya; Shaw, Tushar; D'Souza, Annet; Eshwara, Vandana Kalwaje; Mukhopadhyay, Chiranjay

2017-07-01

To evaluate the diagnostic utility of enrichment culture and PCR for improved case detection rates of non-bacteraemic form of melioidosis in limited resource settings. Clinical specimens (n = 525) obtained from patients presenting at a tertiary care hospital of South India with clinical symptoms suggestive of community-acquired pneumonia, lower respiratory tract infections, superficial or internal abscesses, chronic skin ulcers and bone or joint infections were tested for the presence of Burkholderia pseudomallei using conventional culture (CC), enrichment culture (EC) and PCR. Sensitivity, specificity, positive and negative predictive values of CC and PCR were initially deduced using EC as the gold standard method. Further, diagnostic accuracies of all the three methods were analysed using Bayesian latent class modelling (BLCM). Detection rates of B. pseudomallei using CC, EC and PCR were 3.8%, 5.3% and 6%, respectively. Diagnostic sensitivities and specificities of CC and PCR were 71.4, 98.4% and 100 and 99.4%, respectively in comparison with EC as the gold standard test. With Bayesian latent class modelling, EC and PCR demonstrated sensitivities of 98.7 and 99.3%, respectively, while CC showed a sensitivity of 70.3% for detection of B. pseudomallei. An increase of 1.6% (95% CI: 1.08-4.32%) in the case detection rate of melioidosis was observed in the study population when EC and/or PCR were used in adjunct to the conventional culture technique. Our study findings underscore the diagnostic superiority of enrichment culture and/or PCR over conventional microbiological culture for improved case detection of melioidosis from non-blood clinical specimens. © 2017 John Wiley & Sons Ltd.

Aerobic De-Epoxydation of Trichothecene Mycotoxins by a Soil Bacterial Consortium Isolated Using In Situ Soil Enrichment.

PubMed

He, Wei-Jie; Yuan, Qing-Song; Zhang, You-Bing; Guo, Mao-Wei; Gong, An-Dong; Zhang, Jing-Bo; Wu, Ai-Bo; Huang, Tao; Qu, Bo; Li, He-Ping; Liao, Yu-Cai

2016-09-24

Globally, the trichothecene mycotoxins deoxynivalenol (DON) and nivalenol (NIV) are among the most widely distributed mycotoxins that contaminate small grain cereals. In this study, a bacterial consortium, PGC-3, with de-epoxydation activity was isolated from soil by an in situ soil enrichment method. Screening of 14 soil samples that were sprayed with DON revealed that 4 samples were able to biotransform DON into de-epoxydized DON (dE-DON). Among these, the PGC-3 consortium showed the highest and most stable activity to biotransform DON into dE-DON and NIV into dE-NIV. PGC-3 exhibited de-epoxydation activity at a wide range of pH (5-10) and temperatures (20-37 °C) values under aerobic conditions. Sequential subculturing with a continued exposure to DON substantially reduced the microbial population diversity of this consortium. Analyses of the 16S rDNA sequences indicated that PGC-3 comprised 10 bacterial genera. Among these, one species, Desulfitobacterium, showed a steady increase in relative abundance, from 0.03% to 1.55% (a 52-fold increase), as higher concentrations of DON were used in the subculture media, from 0 to 500 μg/mL. This study establishes the foundation to further develop bioactive agents that can detoxify trichothecene mycotoxins in cereals and enables for the characterization of detoxifying genes and their regulation.

Aerobic De-Epoxydation of Trichothecene Mycotoxins by a Soil Bacterial Consortium Isolated Using In Situ Soil Enrichment

PubMed Central

He, Wei-Jie; Yuan, Qing-Song; Zhang, You-Bing; Guo, Mao-Wei; Gong, An-Dong; Zhang, Jing-Bo; Wu, Ai-Bo; Huang, Tao; Qu, Bo; Li, He-Ping; Liao, Yu-Cai

2016-01-01

Globally, the trichothecene mycotoxins deoxynivalenol (DON) and nivalenol (NIV) are among the most widely distributed mycotoxins that contaminate small grain cereals. In this study, a bacterial consortium, PGC-3, with de-epoxydation activity was isolated from soil by an in situ soil enrichment method. Screening of 14 soil samples that were sprayed with DON revealed that 4 samples were able to biotransform DON into de-epoxydized DON (dE-DON). Among these, the PGC-3 consortium showed the highest and most stable activity to biotransform DON into dE-DON and NIV into dE-NIV. PGC-3 exhibited de-epoxydation activity at a wide range of pH (5–10) and temperatures (20–37 °C) values under aerobic conditions. Sequential subculturing with a continued exposure to DON substantially reduced the microbial population diversity of this consortium. Analyses of the 16S rDNA sequences indicated that PGC-3 comprised 10 bacterial genera. Among these, one species, Desulfitobacterium, showed a steady increase in relative abundance, from 0.03% to 1.55% (a 52-fold increase), as higher concentrations of DON were used in the subculture media, from 0 to 500 μg/mL. This study establishes the foundation to further develop bioactive agents that can detoxify trichothecene mycotoxins in cereals and enables for the characterization of detoxifying genes and their regulation. PMID:27669304

Aerobic cyanide degradation by bacterial isolates from cassava factory wastewater

PubMed Central

Kandasamy, Sujatha; Dananjeyan, Balachandar; Krishnamurthy, Kumar; Benckiser, Gero

2015-01-01

Ten bacterial strains that utilize cyanide (CN) as a nitrogen source were isolated from cassava factory wastewater after enrichment in a liquid media containing sodium cyanide (1 mM) and glucose (0.2% w/v). The strains could tolerate and grow in cyanide concentrations of up to 5 mM. Increased cyanide levels in the media caused an extension of lag phase in the bacterial growth indicating that they need some period of acclimatisation. The rate of cyanide removal by the strains depends on the initial cyanide and glucose concentrations. When initial cyanide and glucose concentrations were increased up to 5 mM, cyanide removal rate increased up to 63 and 61 per cent by Bacillus pumilus and Pseudomonas putida. Metabolic products such as ammonia and formate were detected in culture supernatants, suggesting a direct hydrolytic pathway without an intermediate formamide. The study clearly demonstrates the potential of aerobic treatment with cyanide degrading bacteria for cyanide removal in cassava factory wastewaters. PMID:26413045

Aerobic cyanide degradation by bacterial isolates from cassava factory wastewater.

PubMed

Kandasamy, Sujatha; Dananjeyan, Balachandar; Krishnamurthy, Kumar; Benckiser, Gero

2015-01-01

Ten bacterial strains that utilize cyanide (CN) as a nitrogen source were isolated from cassava factory wastewater after enrichment in a liquid media containing sodium cyanide (1 mM) and glucose (0.2% w/v). The strains could tolerate and grow in cyanide concentrations of up to 5 mM. Increased cyanide levels in the media caused an extension of lag phase in the bacterial growth indicating that they need some period of acclimatisation. The rate of cyanide removal by the strains depends on the initial cyanide and glucose concentrations. When initial cyanide and glucose concentrations were increased up to 5 mM, cyanide removal rate increased up to 63 and 61 per cent by Bacillus pumilus and Pseudomonas putida. Metabolic products such as ammonia and formate were detected in culture supernatants, suggesting a direct hydrolytic pathway without an intermediate formamide. The study clearly demonstrates the potential of aerobic treatment with cyanide degrading bacteria for cyanide removal in cassava factory wastewaters.

Comparative study with two different enrichments in the culture media used in the disinfectant efficacy assay.

PubMed

Sabagh, Bruna Peres; Souto, Aline da Silva Soares; Reis, Louise Moreira; Silva, Sérgio Alves da; Pereira, Daniella Cristina Rodrigues; Neves, Marta de Campos; Pinheiro, Rodrigo Rollin; Duarte, Rafael Silva; Miyazaki, Neide Hiromi Tokumaru; Bôas, Maria Helena Simões Villas

2012-02-01

Recent changes in Brazilian legislation for commercial disinfectants have been published due to the recent epidemic of nosocomial infections caused by rapidly growing mycobacteria (RGM) in many states of Brazil over the last 8years. One of these documents requires that all the manufacturers provide evidence of efficacy of sterilizing and disinfectant products, used for semi critical medical devices, against the Mycobacterium bovis BCG Moreau and Mycobacterium abscessus subsp. bolletii INCQS 00594 strains by using the Confirmative in vitro Test for Determining Tuberculocidal Activity of Disinfectants recommended by the Association of Official Analytical Chemists. These changes have caused additional costs and increased problems for importation of enrichment products at national laboratories where disinfectant efficacy assay service is performed. Middlebrook ADC Enrichment (ADC) is provided by a unique manufacturer and used in the official protocol. The aim of the present study was to evaluate an alternative in house low-cost enrichment composed of fetal bovine serum and glucose (FBSG) with ADC for performance of disinfectant efficacy assay against mycobacteria. After obtaining the growth curves for M. abscessus ATCC 19977, M. abscessus subsp. bolletii INCQS 00594, Mycobacterium chelonae ATCC 35752, and Mycobacterium fortuitum ATCC 6841 by using ADC enrichment and FBSG in Kirchners and 7H9 culture media. Through statistical analysis via the Kruskal-Wallis test on the evaluation of microorganism growth rate, it was observed that there was no inhibition of RGM growth by any of the enrichments used. These results suggest that low-cost enrichment FBSG may be used as a potential substitute of ADC for composition of media for mycobacterial growth, including in disinfectant tests. Copyright © 2011 Elsevier B.V. All rights reserved.

Simplified Protocol for Carba NP Test for Enhanced Detection of Carbapenemase Producers Directly from Bacterial Cultures.

PubMed

Pasteran, Fernando; Tijet, Nathalie; Melano, Roberto G; Corso, Alejandra

2015-12-01

We compared carbapenemase detection among 266 Gram-negative bacilli (161 carbapenemase producers) using the Carba NP tests issued by the CLSI (CNPt-CLSI) and a novel protocol (CNPt-direct) designed for carbapenemase detection direct from bacterial cultures (instead of bacterial extracts required by the CLSI tests). The specificities were comparable (100%), but the CNPt-direct was more sensitive (98% versus 84%). The CNPt-direct was easier to perform due to the direct use of colonies and offered a more robust detection of carbapenemase producers. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Detection of Prosthetic Hip Infection at Revision Arthroplasty by Immunofluorescence Microscopy and PCR Amplification of the Bacterial 16S rRNA Gene

PubMed Central

Tunney, Michael M.; Patrick, Sheila; Curran, Martin D.; Ramage, Gordon; Hanna, Donna; Nixon, James R.; Gorman, Sean P.; Davis, Richard I.; Anderson, Neil

1999-01-01

In this study the detection rates of bacterial infection of hip prostheses by culture and nonculture methods were compared for 120 patients with total hip revision surgery. By use of strict anaerobic bacteriological practice during the processing of samples and without enrichment, the incidence of infection by culture of material dislodged from retrieved prostheses after ultrasonication (sonicate) was 22%. Bacteria were observed by immunofluorescence microscopy in 63% of sonicate samples with a monoclonal antibody specific for Propionibacterium acnes and polyclonal antiserum specific for Staphylococcus spp. The bacteria were present either as single cells or in aggregates of up to 300 bacterial cells. These aggregates were not observed without sonication to dislodge the biofilm. Bacteria were observed in all of the culture-positive samples, and in some cases in which only one type of bacterium was identified by culture, both coccoid and coryneform bacteria were observed by immunofluorescence microscopy. Bacteria from skin-flake contamination were readily distinguishable from infecting bacteria by immunofluorescence microscopy. Examination of skin scrapings did not reveal large aggregates of bacteria but did reveal skin cells. These were not observed in the sonicates. Bacterial DNA was detected in 72% of sonicate samples by PCR amplification of a region of the bacterial 16S rRNA gene with universal primers. All of the culture-positive samples were also positive for bacterial DNA. Evidence of high-level infiltration either of neutrophils or of lymphocytes or macrophages into associated tissue was observed in 73% of patients. Our results indicate that the incidence of prosthetic joint infection is grossly underestimated by current culture detection methods. It is therefore imperative that current clinical practice with regard to the detection and subsequent treatment of prosthetic joint infection be reassessed in the light of these results. PMID:10488193

Enrichment of arsenic transforming and resistant heterotrophic bacteria from sediments of two salt lakes in Northern Chile.

PubMed

Lara, José; Escudero González, Lorena; Ferrero, Marcela; Chong Díaz, Guillermo; Pedrós-Alió, Carlos; Demergasso, Cecilia

2012-05-01

Microbial populations are involved in the arsenic biogeochemical cycle in catalyzing arsenic transformations and playing indirect roles. To investigate which ecotypes among the diverse microbial communities could have a role in cycling arsenic in salt lakes in Northern Chile and to obtain clues to facilitate their isolation in pure culture, sediment samples from Salar de Ascotán and Salar de Atacama were cultured in diluted LB medium amended with NaCl and arsenic, at different incubation conditions. The samples and the cultures were analyzed by nucleic acid extraction, fingerprinting analysis, and sequencing. Microbial reduction of As was evidenced in all the enrichments carried out in anaerobiosis. The results revealed that the incubation factors were more important for determining the microbial community structure than arsenic species and concentrations. The predominant microorganisms in enrichments from both sediments belonged to the Firmicutes and Proteobacteria phyla, but most of the bacterial ecotypes were confined to only one system. The occurrence of an active arsenic biogeochemical cycle was suggested in the system with the highest arsenic content that included populations compatible with microorganisms able to transform arsenic for energy conservation, accumulate arsenic, produce H(2), H(2)S and acetic acid (potential sources of electrons for arsenic reduction) and tolerate high arsenic levels.

Quantification of Growth of Campylobacter and Extended Spectrum β-Lactamase Producing Bacteria Sheds Light on Black Box of Enrichment Procedures.

PubMed

Hazeleger, Wilma C; Jacobs-Reitsma, Wilma F; den Besten, Heidy M W

2016-01-01

Campylobacter is well recognized as the leading cause of bacterial foodborne diarrheal disease worldwide, and is routinely found in meat originating from poultry, sheep, pigs, and cattle. Effective monitoring of Campylobacter contamination is dependent on the availability of reliable detection methods. The method of the International Organization for Standardization for the detection of Campylobacter spp. in food (ISO 10272-1:2006) recommends the use of Bolton broth (BB) as selective enrichment medium, including a pre-enrichment step of 4-6 h at 37°C to revive sublethally damaged cells prior to incubation for 2 days at 41.5°C. Recently the presence of abundantly growing extended spectrum β-lactamase producing Enterobacteriaceae (ESBL bacteria) has become one of the most important factors that interfere with the isolation of Campylobacter, resulting in false-negative detection. However, detailed growth dynamics of Campylobacter and its competitors remain unclear, where these would provide a solid base for further improvement of the enrichment procedure for Campylobacter. Other enrichment broths, such as Preston broth (PB) and BB plus clavulanic acid (BBc) have been suggested to inhibit competitive flora. Therefore, these different broths were used as enrichments to measure the growth kinetics of several strains of Campylobacter jejuni and ESBL bacteria separately, in co-culture and of strains in chicken samples. The maximum cell numbers and often the growth rates of Campylobacter in mixed culture with ESBL bacteria were significantly lower than in single cultures, indicating severe suppression of Campylobacter by ESBL bacteria, also in naturally contaminated samples. PB and BBc successfully diminished ESBL bacteria and might therefore be a better choice as enrichment medium in possibly ESBL-bacteria contaminated samples. The efficacy of a pre-enrichment step in the BB ISO-procedure was not supported for cold-stressed and non-stressed cells. Therefore, omission of

Nodulation-dependent communities of culturable bacterial endophytes from stems of field-grown soybeans.

PubMed

Okubo, Takashi; Ikeda, Seishi; Kaneko, Takakazu; Eda, Shima; Mitsui, Hisayuki; Sato, Shusei; Tabata, Satoshi; Minamisawa, Kiwamu

2009-01-01

Endophytic bacteria (247 isolates) were randomly isolated from surface-sterilized stems of non-nodulated (Nod(-)), wild-type nodulated (Nod(+)), and hypernodulated (Nod(++)) soybeans (Glycine max [L.] Merr) on three agar media (R2A, nutrient agar, and potato dextrose agar). Their diversity was compared on the basis of 16S rRNA gene sequences. The phylogenetic composition depended on the soybean nodulation phenotype, although diversity indexes were not correlated with nodulation phenotype. The most abundant phylum throughout soybean lines tested was Proteobacteria (58-79%). Gammaproteobacteria was the dominant class (21-72%) with a group of Pseudomonas sp. significantly abundant in Nod(+) soybeans. A high abundance of Alphaproteobacteria was observed in Nod(-) soybeans, which was explained by the increase in bacterial isolates of the families Rhizobiaceae and Sphingomonadaceae. A far greater abundance of Firmicutes was observed in Nod(-) and Nod(++) mutant soybeans than in Nod(+) soybeans. An impact of culture media on the diversity of isolated endophytic bacteria was also observed: The highest diversity indexes were obtained on the R2A medium, which enabled us to access Alphaproteobacteria and other phyla more frequently. The above results indicated that the extent of nodulation changes the phylogenetic composition of culturable bacterial endophytes in soybean stems.

Culture-dependent and culture-independent characterization of microbial assemblages associated with high-temperature petroleum reservoirs.

PubMed

Orphan, V J; Taylor, L T; Hafenbradl, D; Delong, E F

2000-02-01

Recent investigations of oil reservoirs in a variety of locales have indicated that these habitats may harbor active thermophilic prokaryotic assemblages. In this study, we used both molecular and culture-based methods to characterize prokaryotic consortia associated with high-temperature, sulfur-rich oil reservoirs in California. Enrichment cultures designed for anaerobic thermophiles, both autotrophic and heterotrophic, were successful at temperatures ranging from 60 to 90 degrees C. Heterotrophic enrichments from all sites yielded sheathed rods (Thermotogales), pleomorphic rods resembling Thermoanaerobacter, and Thermococcus-like isolates. The predominant autotrophic microorganisms recovered from inorganic enrichments using H(2), acetate, and CO(2) as energy and carbon sources were methanogens, including isolates closely related to Methanobacterium, Methanococcus, and Methanoculleus species. Two 16S rRNA gene (rDNA) libraries were generated from total community DNA collected from production wellheads, using either archaeal or universal oligonucleotide primer sets. Sequence analysis of the universal library indicated that a large percentage of clones were highly similar to known bacterial and archaeal isolates recovered from similar habitats. Represented genera in rDNA clone libraries included Thermoanaerobacter, Thermococcus, Desulfothiovibrio, Aminobacterium, Acidaminococcus, Pseudomonas, Halomonas, Acinetobacter, Sphingomonas, Methylobacterium, and Desulfomicrobium. The archaeal library was dominated by methanogen-like rDNAs, with a lower percentage of clones belonging to the Thermococcales. Our results strongly support the hypothesis that sulfur-utilizing and methane-producing thermophilic microorganisms have a widespread distribution in oil reservoirs and the potential to actively participate in the biogeochemical transformation of carbon, hydrogen, and sulfur in situ.

Cultivable Bacterial Microbiota of Northern Bobwhite (Colinus virginianus): A New Reservoir of Antimicrobial Resistance?

PubMed Central

Su, Hongwen; McKelvey, Jessica; Rollins, Dale; Zhang, Michael; Brightsmith, Donald J.; Derr, James; Zhang, Shuping

2014-01-01

The northern bobwhite (Colinus virginianus) is an ecologically and economically important avian species. At the present time, little is known about the microbial communities associated with these birds. As the first step to create a quail microbiology knowledge base, the current study conducted an inventory of cultivable quail tracheal, crop, cecal, and cloacal microbiota and associated antimicrobial resistance using a combined bacteriology and DNA sequencing approach. A total of 414 morphologically unique bacterial colonies were selected from nonselective aerobic and anaerobic cultures, as well as selective and enrichment cultures. Analysis of the first 500-bp 16S rRNA gene sequences in conjunction with biochemical identifications revealed 190 non-redundant species-level taxonomic units, representing 160 known bacterial species and 30 novel species. The bacterial species were classified into 4 phyla, 14 orders, 37 families, and 59 or more genera. Firmicutes was the most commonly encountered phylum (57%) followed by Actinobacteria (24%), Proteobacteria (17%) and Bacteroidetes (0.02%). Extensive diversity in the species composition of quail microbiota was observed among individual birds and anatomical locations. Quail microbiota harbored several opportunistic pathogens, such as E. coli and Ps. aeruginosa, as well as human commensal organisms, including Neisseria species. Phenotypic characterization of selected bacterial species demonstrated a high prevalence of resistance to the following classes of antimicrobials: phenicol, macrolide, lincosamide, quinolone, and sulphate. Data from the current investigation warrant further investigation on the source, transmission, pathology, and control of antimicrobial resistance in wild quail populations. PMID:24937705

Characterization of Bacterial Communities in Venous Insufficiency Wounds by Use of Conventional Culture and Molecular Diagnostic Methods▿

PubMed Central

Tuttle, Marie S.; Mostow, Eliot; Mukherjee, Pranab; Hu, Fen Z.; Melton-Kreft, Rachael; Ehrlich, Garth D.; Dowd, Scot E.; Ghannoum, Mahmoud A.

2011-01-01

Microbial infections delay wound healing, but the effect of the composition of the wound microbiome on healing parameters is unknown. To better understand bacterial communities in chronic wounds, we analyzed debridement samples from lower-extremity venous insufficiency ulcers using the following: conventional anaerobic and aerobic bacterial cultures; the Ibis T5000 universal biosensor (Abbott Molecular); and 16S 454 FLX titanium series pyrosequencing (Roche). Wound debridement samples were obtained from 10 patients monitored clinically for at least 6 months, at which point 5 of the 10 sampled wounds had healed. Pyrosequencing data revealed significantly higher bacterial abundance and diversity in wounds that had not healed at 6 months. Additionally, Actinomycetales was increased in wounds that had not healed, and Pseudomonadaceae was increased in wounds that had healed by the 6-month follow-up. Baseline wound surface area, duration, or analysis by Ibis or conventional culture did not reveal significant differences between wounds that healed after 6 months and those that did not. Thus, pyrosequencing identified distinctive baseline characteristics of wounds that did not heal by the 6-month follow-up, furthering our understanding of potentially unique microbiome characteristics of chronic wounds. PMID:21880958

Draft Genome Sequence of a Dictyoglomus sp. from an Enrichment Culture of a New Zealand Geothermal Spring

PubMed Central

Donaho, John A.; Kelley, John F.; St. John, Emily; Turner, Christina; Podar, Mircea; Stott, Matthew B.

2018-01-01

ABSTRACT A draft genome of a novel Dictyoglomus sp., NZ13-RE01, was obtained from a New Zealand hot spring enrichment culture. The 1,927,012-bp genome is similar in both size and G+C content to other Dictyoglomus spp. Like its relatives, Dictyoglomus sp. NZ13-RE01 encodes many genes involved in complex carbohydrate metabolism. PMID:29545298

Study of selenocompounds from selenium-enriched culture of edible sprouts.

PubMed

Funes-Collado, Virginia; Morell-Garcia, Albert; Rubio, Roser; López-Sánchez, José Fermín

2013-12-15

Selenium is recognised as an essential micronutrient for humans and animals. One of the main sources of selenocompounds in the human diet is vegetables. Therefore, this study deals with the Se species present in different edible sprouts grown in Se-enriched media. We grew alfalfa, lentil and soy in a hydroponic system amended with soluble salts, containing the same proportion of Se, in the form of Se(VI) and Se(IV). Total Se in the sprouts was determined by acidic digestion in a microwave system and by ICP/MS. Se speciation was carried out by enzymatic extraction (Protease XIV) and measured by LC-ICP/MS. The study shows that the Se content of plants depends on the content in the growth culture, and that part of the inorganic Se was biotransformed mainly into SeMet. These results contribute to our understanding of the uptake of inorganic Se and its biotransformation by edible plants. Copyright © 2013 Elsevier Ltd. All rights reserved.

Exploring bacterial infections: theoretical and experimental studies of the bacterial population dynamics and antibiotic treatment

NASA Astrophysics Data System (ADS)

Shao, Xinxian

Bacterial infections are very common in human society. Thus extensive research has been conducted to reveal the molecular mechanisms of the pathogenesis and to evaluate the antibiotics' efficacy against bacteria. Little is known, however, about the population dynamics of bacterial populations and their interactions with the host's immune system. In this dissertation, a stochatic model is developed featuring stochastic phenotypic switching of bacterial individuals to explain the single-variant bottleneck discovered in multi strain bacterial infections. I explored early events in a bacterial infection establishment using classical experiments of Moxon and Murphy on neonatal rats. I showed that the minimal model and its simple variants do not work. I proposed modifications to the model that could explain the data quantitatively. The bacterial infections are also commonly established in physical structures, as biofilms or 3-d colonies. In contrast, most research on antibiotic treatment of bacterial infections has been conducted in well-mixed liquid cultures. I explored the efficacy of antibiotics to treat such bacterial colonies, a broadly applicable method is designed and evaluated where discrete bacterial colonies on 2-d surfaces were exposed to antibiotics. I discuss possible explanations and hypotheses for the experimental results. To verify these hypotheses, we investigated the dynamics of bacterial population as 3-d colonies. We showed that a minimal mathematical model of bacterial colony growth in 3-d was able to account for the experimentally observed presence of a diffusion-limited regime. The model further revealed highly loose packing of the cells in 3-d colonies and smaller cell sizes in colonies than plancktonic cells in corresponding liquid culture. Further experimental tests of the model predictions have revealed that the ratio of the cell size in liquid culture to that in colony cultures was consistent with the model prediction, that the dead cells

Comparative study on treatment of kitchen wastewater using a mixed microalgal culture and an aerobic bacterial culture: kinetic evaluation and FAME analysis.

PubMed

Katam, Keerthi; Bhattacharyya, Debraj

2018-05-12

Microalgae-based treatment systems have been successfully used for the polishing of domestic wastewater. Research is underway in studying the suitability of using these systems as main treatment units. This study focuses on comparing the performances of a mixed microalgal culture and an aerobic bacterial culture, based on the kinetic evaluation, in removing organic carbon from a kitchen wastewater. The two systems were operated at six different solid retention times (SRTs)-2, 4, 6, 8, 10, and 12 days in continuous mode. The influent and effluent samples were analyzed for chemical oxygen demand (COD), total organic carbon (TOC), total nitrogen (TN), phosphates, and surfactants. Steady-state kinetics (k, K s , Y, and k d ) for organic carbon removal were obtained by fitting experimental data in linearized Michaelis-Menten and Monod equations. The mixed microalgal system showed similar or better performance in COD and TN removal (88 and 85%, respectively) when compared with the COD and TN removal by the aerobic bacterial system (89 and 48%). A maximum lipid yield of 40% (w/w of dry biomass) was observed in the microalgal system. Saturated fatty acids accounted for 50% of the total observed FAME species. The study indicates that the mixed microalgal culture is capable of treating kitchen wastewater and has the potential to replace aerobic bacteria in biological treatment systems in certain cases.

Enrichment dynamics of Listeria monocytogenes and the associated microbiome from naturally contaminated ice cream linked to a listeriosis outbreak.

PubMed

Ottesen, Andrea; Ramachandran, Padmini; Reed, Elizabeth; White, James R; Hasan, Nur; Subramanian, Poorani; Ryan, Gina; Jarvis, Karen; Grim, Christopher; Daquiqan, Ninalynn; Hanes, Darcy; Allard, Marc; Colwell, Rita; Brown, Eric; Chen, Yi

2016-11-16

Microbiota that co-enrich during efforts to recover pathogens from foodborne outbreaks interfere with efficient detection and recovery. Here, dynamics of co-enriching microbiota during recovery of Listeria monocytogenes from naturally contaminated ice cream samples linked to an outbreak are described for three different initial enrichment formulations used by the Food and Drug Administration (FDA), the International Organization of Standardization (ISO), and the United States Department of Agriculture (USDA). Enrichment cultures were analyzed using DNA extraction and sequencing from samples taken every 4 h throughout 48 h of enrichment. Resphera Insight and CosmosID analysis tools were employed for high-resolution profiling of 16S rRNA amplicons and whole genome shotgun data, respectively. During enrichment, other bacterial taxa were identified, including Anoxybacillus, Geobacillus, Serratia, Pseudomonas, Erwinia, and Streptococcus spp. Surprisingly, incidence of L. monocytogenes was proportionally greater at hour 0 than when tested 4, 8, and 12 h later with all three enrichment schemes. The corresponding increase in Anoxybacillus and Geobacillus spp.indicated these taxa co-enriched in competition with L. monocytogenes during early enrichment hours. L. monocytogenes became dominant after 24 h in all three enrichments. DNA sequences obtained from shotgun metagenomic data of Listeria monocytogenes at 48 h were assembled to produce a consensus draft genome which appeared to have a similar tracking utility to pure culture isolates of L. monocytogenes. All three methods performed equally well for enrichment of Listeria monocytogenes. The observation of potential competitive exclusion of L. mono by Anoxybacillus and Geobacillus in early enrichment hours provided novel information that may be used to further optimize enrichment formulations. Application of Resphera Insight for high-resolution analysis of 16S amplicon sequences accurately identified L. monocytogenes

A rapid, highly sensitive and culture-free detection of pathogens from blood by positive enrichment.

PubMed

Vutukuru, Manjula Ramya; Sharma, Divya Khandige; Ragavendar, M S; Schmolke, Susanne; Huang, Yiwei; Gumbrecht, Walter; Mitra, Nivedita

2016-12-01

Molecular diagnostics is a promising alternative to culture based methods for the detection of bloodstream infections, notably due to its overall lower turnaround time when starting directly from patient samples. Whole blood is usually the starting diagnostic sample in suspected bloodstream infections. The detection of low concentrations of pathogens in blood using a molecular assay necessitates a fairly high starting volume of blood sample in the range of 5-10mL. This large volume of blood sample has a substantial accompanying human genomic content that interferes with pathogen detection. In this study, we have established a workflow using magnetic beads coated with Apolipoprotein H that makes it possible to concentrate pathogens from a 5.0mL whole blood sample, thereby enriching pathogens from whole blood background and also reducing the sample volume to ~200μL or less. We have also demonstrated that this method of enrichment allows detection of 1CFU/mL of Escherichia coli, Enterococcus gallinarum and Candida tropicalis from 5mL blood using quantitative PCR; a detection limit that is not possible in unenriched samples. The enrichment method demonstrated here took 30min to complete and can be easily integrated with various downstream molecular and microbiological techniques. Copyright © 2016 Elsevier B.V. All rights reserved.

Influence of in vitro gastrointestinal digestion of fruit juices enriched with pine bark extract on intestinal microflora.

PubMed

López-Nicolás, Rubén; González-Bermúdez, Carlos A; Ros-Berruezo, Gaspar; Frontela-Saseta, Carmen

2014-08-15

The selective antimicrobial effect of fruit juices enriched with pine bark extract (PBE) (0.5 g/L) has been studied before and after in vitro gastrointestinal digestion. PBE (a concentrate of water-soluble bioflavonoids, mainly including phenolic compounds) has been proven to have high stability to the digestion process. Pure phenolic compounds such as gallic acid had a high antimicrobial effect on Staphylococcus aureus and Escherichia coli, maintaining the lactic acid bacteria population (≈100%). Otherwise, E. coli O157:H7 only growth 50% when PBE was added to the culture media, while a slight increase on the growth of lactobacilli and bifidobacteria was observed after exposition to the bark extract. Fresh fruit juices enriched with PBE showed the highest inhibitory effect on pathogenic intestinal bacterial growth, mainly E. coli and Enterococcus faecalis. The in vitro digestion process reduced the antibacterial effect of juices against most pathogenic bacteria in approximately 10%. However, the beneficial effect of fruit juices enriched with PBE (0.5 g/L) on gut microbiota is still considerable after digestion. Copyright © 2014 Elsevier Ltd. All rights reserved.

Stable Carbon Isotope Fractionation during Bacterial Acetylene Fermentation: Potential for Life Detection in Hydrocarbon-Rich Volatiles of Icy Planet(oid)s.

PubMed

Miller, Laurence G; Baesman, Shaun M; Oremland, Ronald S

2015-11-01

We report the first study of stable carbon isotope fractionation during microbial fermentation of acetylene (C2H2) in sediments, sediment enrichments, and bacterial cultures. Kinetic isotope effects (KIEs) averaged 3.7 ± 0.5‰ for slurries prepared with sediment collected at an intertidal mudflat in San Francisco Bay and 2.7 ± 0.2‰ for a pure culture of Pelobacter sp. isolated from these sediments. A similar KIE of 1.8 ± 0.7‰ was obtained for methanogenic enrichments derived from sediment collected at freshwater Searsville Lake, California. However, C2H2 uptake by a highly enriched mixed culture (strain SV7) obtained from Searsville Lake sediments resulted in a larger KIE of 9.0 ± 0.7‰. These are modest KIEs when compared with fractionation observed during oxidation of C1 compounds such as methane and methyl halides but are comparable to results obtained with other C2 compounds. These observations may be useful in distinguishing biologically active processes operating at distant locales in the Solar System where C2H2 is present. These locales include the surface of Saturn's largest moon Titan and the vaporous water- and hydrocarbon-rich jets emanating from Enceladus. Acetylene-Fermentation-Isotope fractionation-Enceladus-Life detection.

Bacterial strain changes during chronic otitis media surgery.

PubMed

Kim, G J; Yoo, S; Han, S; Bu, J; Hong, Y; Kim, D-K

2017-09-01

Cultures obtained from pre-operative middle-ear swabs from patients with chronic otitis media have traditionally been used to guide antibiotic selection. This study investigated changes in the bacterial strains of the middle ear during chronic otitis media surgery. Pre-operative bacterial cultures of otorrhoea, and peri-operative cultures of the granulation tissue in either the middle ear or mastoid cavity, were obtained. Post-operative cultures were selectively obtained when otorrhoea developed after surgery. Bacterial growth was observed in 45.5 per cent of pre-operative cultures, 13.5 per cent of peri-operative cultures and 4.5 per cent of post-operative cultures. Methicillin-resistant Staphylococcus aureus was identified as the most common bacteria in all pre-operative (32.4 per cent), peri-operative (52.4 per cent) and post-operative (71.4 per cent) tests, and the percentage of Methicillin-resistant S aureus increased from the pre- to the post-operative period. The bacterial culture results for post-operative otorrhoea showed low agreement with those for pre-operative or peri-operative culture, and strain re-identification was required.

Gravimetric enrichment of high lipid and starch accumulating microalgae.

PubMed

Hassanpour, Morteza; Abbasabadi, Mahsa; Ebrahimi, Sirous; Hosseini, Maryam; Sheikhbaglou, Ahmad

2015-11-01

This study presents gravimetric enrichment of mixed culture to screen starch and lipid producing species separately in a sequencing batch reactor. In the enriched starch-producing mixed culture photobioreactor, the starch content at the end of steady state batch became 3.42 times the beginning of depletion. Whereas in the enriched lipid-producing photobioreactor, the lipid content at the end of steady state batch became 3 times the beginning of famine phase. The obtained results revealed that the gravimetric enrichment is a suitable screening method for specific production of storage compounds in none-sterile large-scaled condition. Copyright © 2015 Elsevier Ltd. All rights reserved.

Draft Genome Sequence of a Dictyoglomus sp. from an Enrichment Culture of a New Zealand Geothermal Spring.

PubMed

Reysenbach, Anna-Louise; Donaho, John A; Kelley, John F; St John, Emily; Turner, Christina; Podar, Mircea; Stott, Matthew B

2018-03-15

A draft genome of a novel Dictyoglomus sp., NZ13-RE01, was obtained from a New Zealand hot spring enrichment culture. The 1,927,012-bp genome is similar in both size and G+C content to other Dictyoglomus spp. Like its relatives, Dictyoglomus sp. NZ13-RE01 encodes many genes involved in complex carbohydrate metabolism. Copyright © 2018 Reysenbach et al.

Evaluation of performance of bacterial culture of feces and serum ELISA across stages of Johne's disease in cattle using a Bayesian latent class model.

PubMed

Espejo, L A; Zagmutt, F J; Groenendaal, H; Muñoz-Zanzi, C; Wells, S J

2015-11-01

The objective of this study was to evaluate the performance of bacterial culture of feces and serum ELISA to correctly identify cows with Mycobacterium avium ssp. paratuberculosis (MAP) at heavy, light, and non-fecal-shedding levels. A total of 29,785 parallel test results from bacterial culture of feces and serum ELISA were collected from 17 dairy herds in Minnesota, Pennsylvania, and Colorado. Samples were obtained from adult cows from dairy herds enrolled for up to 10 yr in the National Johne's Disease Demonstration Herd Project. A Bayesian latent class model was fitted to estimate the probabilities that bacterial culture of feces (using 72-h sedimentation or 30-min centrifugation methods) and serum ELISA results correctly identified cows as high positive, low positive, or negative given that cows were heavy, light, and non-shedders, respectively. The model assumed that no gold standard test was available and conditional independency existed between diagnostic tests. The estimated conditional probabilities that bacterial culture of feces correctly identified heavy shedders, light shedders, and non-shedders were 70.9, 32.0, and 98.5%, respectively. The same values for the serum ELISA were 60.6, 18.7, and 99.5%, respectively. Differences in diagnostic test performance were observed among states. These results improve the interpretation of results from bacterial culture of feces and serum ELISA for detection of MAP and MAP antibody (respectively), which can support on-farm infection control decisions and can be used to evaluate disease-testing strategies, taking into account the accuracy of these tests. Copyright © 2015 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Integrative approach to produce hydrogen and polyhydroxybutyrate from biowaste using defined bacterial cultures.

PubMed

Patel, Sanjay K S; Kumar, Prasun; Singh, Mamtesh; Lee, Jung-Kul; Kalia, Vipin C

2015-01-01

Biological production of hydrogen (H2) and polyhydroxybutyrate (PHB) from pea-shell slurry (PSS) was investigated using defined mixed culture (MMC4, composed of Enterobacter, Proteus, Bacillus spp.). Under batch culture, 19.0LH2/kg of PSS (total solid, TS, 2%w/v) was evolved. Using effluent from the H2 producing stage, Bacillus cereus EGU43 could produce 12.4% (w/w) PHB. Dilutions of PSS hydrolysate containing glucose (0.5%, w/v) resulted in 45-75LH2/kg TS fed and 19.1% (w/w) of PHB content. Under continuous culture, MMC4 immobilized on coconut coir (CC) lead to an H2 yield of 54L/kg TS fed and a PHB content of 64.7% (w/w). An improvement of 2- and 3.7-fold in H2 and PHB yields were achieved in comparison to control. This integrative approach using defined set of bacterial strains can prove effective in producing biomolecules from biowastes. Copyright © 2014. Published by Elsevier Ltd.

Anaerobic biodegradation of partially hydrolyzed polyacrylamide in long-term methanogenic enrichment cultures from production water of oil reservoirs.

PubMed

Hu, Hao; Liu, Jin-Feng; Li, Cai-Yun; Yang, Shi-Zhong; Gu, Ji-Dong; Mu, Bo-Zhong

2018-06-01

The increasing usage of partially hydrolyzed polyacrylamide (HPAM) in oilfields as a flooding agent to enhance oil recovery at so large quantities is an ecological hazard to the subsurface ecosystem due to persistence and inertness. Biodegradation of HPAM is a potentially promising strategy for dealing with this problem among many other methods available. To understand the responsible microorganisms and mechanism of HPAM biodegradation under anaerobic conditions, an enrichment culture from production waters of oil reservoirs were established with HPAM as the sole source of carbon and nitrogen incubated for over 328 days, and analyzed using both molecular microbiology and chemical characterization methods. Gel permeation chromatography, High-pressure liquid chromatography and Fourier-transformed infrared spectroscopy results indicated that, after 328 days of anaerobic incubation, some of the amide groups on HPAM were removed and released as ammonia/ammonium and carboxylic groups, while the carbon backbone of HPAM was converted to smaller polymeric fragments, including oligomers and various fatty acids. Based on these results, the biochemical process of anaerobic biodegradation of HPAM was proposed. The phylogenetic analysis of 16S rRNA gene sequences retrieved from the enrichments showed that Proteobacteria and Planctomycetes were the dominant bacteria in the culture with HPAM as the source of carbon and nitrogen, respectively. For archaea, Methanofollis was more abundant in the anaerobic enrichment. These results are helpful for understanding the process of HPAM biodegradation and provide significant insights to the fate of HPAM in subsurface environment and for possible bioremediation.

Degradation of trimethylbenzene isomers by an enrichment culture under N{sub 2}O-reducing conditions

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

Haener, A.; Hoehener, P.; Zeyer, J.

1997-03-01

In mineral oil-contaminated soils and aquifers, monoaromatic hydrocarbons are often a major concern because of high water solubility and toxicity. Under aerobic conditions, these compounds are rapidly mineralized. However, only limited data are available on the anaerobic degradation. This study reports on the growth of an enrichment culture on 1,3,5-trimethylbenzene (TMB) and 1,2,4-TMB under N2O-reducing conditions, and provides carbon mass and electron balances for the biodegradation of 1,3,5-TMB. 43 refs., 3 figs.

Adherence of Escherichia coli O157:H7 to epithelial cells in vitro and in pig gut loops is affected by bacterial culture conditions

PubMed Central

Yin, Xianhua; Feng, Yanni; Wheatcroft, Roger; Chambers, James; Gong, Joshua; Gyles, Carlton L.

2011-01-01

The objectives of this study were to determine the effect of bacterial culture conditions on adherence of enterohemorrhagic Escherichia coli (EHEC) O157:H7 strain 86-24 in vivo to pig enterocytes and to compare the results with adherence in vitro to cultured HEp-2 and IPEC-J2 cells. Growth of O157:H7 in MacConkey broth (MB) resulted in almost no adherence to both HEp-2 and IPEC-J2 cells; prior exposure of the bacteria to pH 2.5 reduced adherence. There was greater adherence by bacteria from static cultures than by those from shaken cultures and by bacteria cultured in brain–heart infusion (BHI) plus NaHCO3 (BHIN) than by bacteria cultured in BHI. In contrast, in pig ileal loops, bacteria cultured in MB adhered well to enterocytes, and prior exposure to pH 2.5 had no effect on adherence. Among several media tested for their effect on bacterial adherence in the pig intestine, MB and BHIN proved to be the best. Bacterial adherence was dose-dependent and was more extensive in the ileum than in the colon. This study demonstrated that there are remarkable differences between culture conditions that promote adherence of an EHEC O157:H7 strain in vitro and in vivo, that culture conditions profoundly affect adherence to epithelial cells in vitro and in vivo, and that pig ileal loops are better suited to adherence studies than are colon loops. PMID:21731177

Biosynthesis of highly enriched 13C-lycopene for human metabolic studies using repeated batch tomato cell culturing with 13C-glucose

PubMed Central

Moran, Nancy E.; Rogers, Randy B.; Lu, Chi-Hua; Conlon, Lauren E.; Lila, Mary Ann; Clinton, Steven K.; Erdman, John W.

2013-01-01

While putative disease-preventing lycopene metabolites are found in both tomato (Solanum lycopersicum) products and in their consumers, mammalian lycopene metabolism is poorly understood. Advances in tomato cell culturing techniques offer an economical tool for generation of highly-enriched 13C-lycopene for human bioavailability and metabolism studies. To enhance the 13C-enrichment and yields of labeled lycopene from the hp-1 tomato cell line, cultures were first grown in 13C-glucose media for three serial batches and produced increasing proportions of uniformly labeled lycopene (14.3 +/− 1.2 %, 39.6 +/− 0.5 %, and 48.9 +/− 1.5% with consistent yields (from 5.8 to 9 mg/L). An optimized 9-day-long 13C-loading and 18-day-long labeling strategy developed based on glucose utilization and lycopene yields, yielded 13C-lycopene with 93% 13C isotopic purity, and 55% of isotopomers were uniformly labeled. Furthermore, an optimized acetone and hexane extraction led to a four-fold increase in lycopene recovery from cultures compared to a standard extraction. PMID:23561155

Effective bioleaching of chromium in tannery sludge with an enriched sulfur-oxidizing bacterial community.

PubMed

Zeng, Jing; Gou, Min; Tang, Yue-Qin; Li, Guo-Ying; Sun, Zhao-Yong; Kida, Kenji

2016-10-01

In this study, a sulfur-oxidizing community was enriched from activated sludge generated in tannery wastewater treatment plants. Bioleaching of tannery sludge containing 0.9-1.2% chromium was investigated to evaluate the effectiveness of the enriched community, the effect of chromium binding forms on bioleaching efficiency, and the dominant microbes contributing to chromium bioleaching. Sludge samples inoculated with the enriched community presented 79.9-96.8% of chromium leaching efficiencies, much higher than those without the enriched community. High bioleaching efficiencies of over 95% were achieved for chromium in reducible fraction, while 60.9-97.9% were observed for chromium in oxidizable and residual fractions. Acidithiobacillus thiooxidans, the predominant bacteria in the enriched community, played an important role in bioleaching, whereas some indigenous heterotrophic species in sludge might have had a supporting role. The results indicated that A. thiooxidans-dominant enriched microbial community had high chromium bioleaching efficiency, and chromium binding forms affected the bioleaching performance. Copyright © 2016 Elsevier Ltd. All rights reserved.

Development of bioconcrete material using an enrichment culture of novel thermophilic anaerobic bacteria.

PubMed

Ghosh, P; Mandal, S; Pal, S; Bandyopadhyaya, G; Chattopadhyay, B D

2006-04-01

In the biosphere, bacteria can function as geo-chemical agents, promoting the dispersion, fractionation and/or concentration of materials. Microbial mineral precipitation is resulted from metabolic activities of microorganisms. Based on this biomineralogy concept, an attempt has been made to develop bioconcrete material incorporating of an enrichment culture of thermophilic and anaerobic bacteria within cement-sand mortar/concrete. The results showed a significant increase in compressive strength of both cement-sand mortar and concrete due to the development of filler material within the pores of cement sand matrix. Maximum strength was observed at concentration 10(5)cell/ml of water used in mortar/concrete. Addition of Escherichia coil or media composition on mortar showed no such improvement in strength.

Direct PCR - A rapid method for multiplexed detection of different serotypes of Salmonella in enriched pork meat samples.

PubMed

Chin, Wai Hoe; Sun, Yi; Høgberg, Jonas; Quyen, Than Linh; Engelsmann, Pia; Wolff, Anders; Bang, Dang Duong

2017-04-01

Salmonellosis, an infectious disease caused by Salmonella spp., is one of the most common foodborne diseases. Isolation and identification of Salmonella by conventional bacterial culture method is time consuming. In response to the demand for rapid on line or at site detection of pathogens, in this study, we developed a multiplex Direct PCR method for rapid detection of different Salmonella serotypes directly from pork meat samples without any DNA purification steps. An inhibitor-resistant Phusion Pfu DNA polymerase was used to overcome PCR inhibition. Four pairs of primers including a pair of newly designed primers targeting Salmonella spp. at subtype level were incorporated in the multiplex Direct PCR. To maximize the efficiency of the Direct PCR, the ratio between sample and dilution buffer was optimized. The sensitivity and specificity of the multiplex Direct PCR were tested using naturally contaminated pork meat samples for detecting and subtyping of Salmonella spp. Conventional bacterial culture methods were used as reference to evaluate the performance of the multiplex Direct PCR. Relative accuracy, sensitivity and specificity of 98.8%; 97.6% and 100%, respectively, were achieved by the method. Application of the multiplex Direct PCR to detect Salmonella in pork meat at slaughter reduces the time of detection from 5 to 6 days by conventional bacterial culture and serotyping methods to 14 h (including 12 h enrichment time). Furthermore, the method poses a possibility of miniaturization and integration into a point-of-need Lab-on-a-chip system for rapid online pathogen detection. Copyright © 2016 Elsevier Ltd. All rights reserved.

Adherent culture conditions enrich the side population obtained from the cochlear modiolus-derived stem/progenitor cells.

PubMed

Chao, Ting-Ting; Wang, Chih-Hung; Chen, Hsin-Chien; Shih, Cheng-Ping; Sytwu, Huey-Kang; Huang, Kun-Lun; Chen, Shao-Yuan

2013-05-01

Previously, our group reported that sphere-forming cells derived from the organ of Corti represent the stem/progenitor cells (SPCs) of the cochlea due to their properties of self-renewal and multipotency. However, long-term propagation of sphere-forming cells under suspension culture conditions may fail to maintain the characteristic stemness of these cells. Therefore, this study investigated whether an adherent culture system would be beneficial in terms of preserving more stem-like cells for long-term manipulations in vitro. Isolated modiolus-derived SPCs were placed on poly-d-lysine-coated petri dishes to form the so-called "adherent" culture system. Modiolus SPCs cultured under adherent conditions exhibited a significantly increased percentage of cells with the side population (SP) phenotype (18.6%) compared with cells cultured under conventional suspension culture conditions (0.8%). Even after repeated passages, modiolus SPCs cultured under adherent culture conditions preserved more SP phenotype cells. In comparison with the non-SP phenotype cells, the sorted SP cells exhibited more stem-like but less differentiated properties, with an upregulated expression of the ATP-binding cassette subfamily G member 2 (ABCG2), Nestin, Sox2, and Nanog proteins. Furthermore, Retinoic acid (RA) treatment confirmed the expression of the multipotent differentiation markers in the SP cells, including TUJ1, pancytokeratin, glial fibrillary acidic protein (GFAP), and p27(Kip1). Employment of an adherent culture system, instead of a suspension culture system, resulted in the enrichment of the SP cells from SPCs while retaining their stemness and multipotency. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Effects of processing delay, temperature, and transport tube type on results of quantitative bacterial culture of canine urine.

PubMed

Patterson, Carly A; Bishop, Micah A; Pack, Julie D; Cook, Audrey K; Lawhon, Sara D

2016-01-15

To determine the impact of processing delay, temperature, and transport tube type on results of quantitative bacterial culture (QBC) of canine urine. Diagnostic test evaluation. 60 mL of pooled urine from 4 dogs, divided into six 10-mL aliquots. Urine aliquots were spiked with bacteria from 1 of 6 independent Escherichia coli cultures to achieve a target bacterial concentration of 10(5) CFUs/mL. One milliliter from each aliquot was transferred into 5 silicone-coated clot tubes (SCTs) and 5 urine transport tubes (UTTs). Samples were stored at 4°C (39°F) and 25°C (77°F) for 0, 8, and 24 hours, and then standard QBCs were performed. Median bacterial concentration for urine samples stored in a UTT for 24 hours at 4°C was lower than that for samples stored in an SCT under the same conditions. Conversely, a substantial decrease in median bacterial concentration was identified for samples stored for 24 hours in an SCT at 25°C, compared with the median concentration for samples stored in a UTT under the same conditions. Median bacterial concentration in samples stored in an SCT at 25°C for 24 hours (275 CFUs/mL) was less than the cutoff typically used to define clinically important bacteriuria by use of urine samples obtained via cystocentesis (ie, > 1,000 CFUs/mL). Canine urine samples submitted for immediate QBC should be transported in plain sterile tubes such as SCTs. When prolonged (24-hour) storage at room temperature is anticipated, urine samples should be transported in UTTs.

Effects of FSH addition to an enriched medium containing insulin and EGF after long-term culture on functionality of equine ovarian biopsy tissue.

PubMed

Aguiar, F L N; Gastal, G D A; Ishak, G M; Gastal, M O; Teixeira, D I A; Feugang, J M; Figueiredo, J R; Gastal, E L

2017-09-01

The effect of FSH supplementation on an enriched cultured medium containing insulin (10 ng/mL) and EGF (50 ng/mL) was investigated on in vitro culture of equine ovarian biopsy tissue. Ovarian tissue fragments were collected from mares (n = 10) and distributed in the following treatments: noncultured control, cultured control, and cultured + FSH. Both treated groups were cultured for 7 or 15 days. The end points evaluated were: follicular morphology, estradiol levels in the culture medium, fluorescence intensity for TUNEL, EGFR and Ki-67 detection, and gene expression of GDF-9, BMP-15, and Cyclin-D2 in the ovarian tissue. After seven days of culture, medium supplemented with FSH had a similar (P > 0.05) percentage of morphologically normal follicles compared to the noncultured control group. Estradiol levels increased (P < 0.05) from Day 7 to Day 15 of culture for both treated groups. No difference (P > 0.05) was observed for TUNEL and EGFR intensity between the noncultured control group and the treated groups after 15 days of culture. Ki-67 intensity did not differ (P > 0.05) between treated groups after 15 days of culture, but decreased (P < 0.05) when compared with the noncultured control group. Similar (P > 0.05) mRNA expression for GDF-9, BMP-15, and Cyclin-D2 was observed among all treatments after 15 days of culture. In conclusion, an enriched medium supplemented or not with FSH was able to maintain the functionality of equine ovarian biopsy tissue after a long-term in vitro culture. Copyright © 2017 Elsevier Inc. All rights reserved.

Neonicotinoid Insecticides Alter the Gene Expression Profile of Neuron-Enriched Cultures from Neonatal Rat Cerebellum

PubMed Central

Kimura-Kuroda, Junko; Nishito, Yasumasa; Yanagisawa, Hiroko; Kuroda, Yoichiro; Komuta, Yukari; Kawano, Hitoshi; Hayashi, Masaharu

2016-01-01

Neonicotinoids are considered safe because of their low affinities to mammalian nicotinic acetylcholine receptors (nAChRs) relative to insect nAChRs. However, because of importance of nAChRs in mammalian brain development, there remains a need to establish the safety of chronic neonicotinoid exposures with regards to children’s health. Here we examined the effects of long-term (14 days) and low dose (1 μM) exposure of neuron-enriched cultures from neonatal rat cerebellum to nicotine and two neonicotinoids: acetamiprid and imidacloprid. Immunocytochemistry revealed no differences in the number or morphology of immature neurons or glial cells in any group versus untreated control cultures. However, a slight disturbance in Purkinje cell dendritic arborization was observed in the exposed cultures. Next we performed transcriptome analysis on total RNAs using microarrays, and identified significant differential expression (p < 0.05, q < 0.05, ≥1.5 fold) between control cultures versus nicotine-, acetamiprid-, or imidacloprid-exposed cultures in 34, 48, and 67 genes, respectively. Common to all exposed groups were nine genes essential for neurodevelopment, suggesting that chronic neonicotinoid exposure alters the transcriptome of the developing mammalian brain in a similar way to nicotine exposure. Our results highlight the need for further careful investigations into the effects of neonicotinoids in the developing mammalian brain. PMID:27782041

Enrichment of prostate cancer stem-like cells from human prostate cancer cell lines by culture in serum-free medium and chemoradiotherapy.

PubMed

Wang, Lei; Huang, Xing; Zheng, Xinmin; Wang, Xinghuan; Li, Shiwen; Zhang, Lin; Yang, Zhonghua; Xia, Zhiping

2013-01-01

The discovery of rare subpopulations of cancer stem cells (CSCs) has created a new focus in cancer research. As CSCs demonstrate resistance to chemoradiation therapy relative to other cancer cells, this allows the enrichment of CSC populations by killing apoptosis-susceptible cancer cells. In this study, three commonly used human prostate cancer (PCa) cell lines (DU145, PC-3 and LNCaP) were examined for their expression of the putative stem cell markers CD133 and CD44 via flow cytometric analysis. Under normal culture conditions, CD133(+)/CD44(+) cells were only present in the DU145 cell line, and comprised only a minor percentage (0.1% ± 0.01%) of the total population. However, the proportion of these CD133(+)/CD44(+) prostate CSCs could be increased in these cell lines via culture in serum-free medium (SFM), or through chemotherapy or radiotherapy. Indeed, after culture in SFM, the proportion of CD133(+)/CD44(+) cells in DU145 and PC-3 had increased to 10.3% and 3.0%, respectively. Moreover, the proportion had increased to 9.8% enriched by chemotherapy and 3.5% by radiotherapy in DU145. Colony-formation tests, cell invasion assays, and tumor xenografts in BALB/c nude mice were used to evaluate the stem cell properties of CD133(+)/CD44(+) PCa cells that were isolated via fluorescence-activated cell sorting (FACS). CD133(+)/CD44(+) cells had an enhanced colony-formation capability and invasive ability in vitro, and displayed greater tumorigenic properties in vivo. These results demonstrate the presence of CD133(+)/CD44(+) prostate CSCs in established PCa cell lines and that populations of these cells can be enriched by culture in SFM or chemoradiotherapy. Finding novel therapies to override chemoradiation resistance in the prostate CSCs is the key to improve long-term results in PCa management.

Bacterial Shifts in Nutrient Solutions Flowing Through Biofilters Used in Tomato Soilless Culture.

PubMed

Renault, David; Déniel, Franck; Vallance, Jessica; Bruez, Emilie; Godon, Jean-Jacques; Rey, Patrice

2017-11-25

In soilless culture, slow filtration is used to eliminate plant pathogenic microorganisms from nutrient solutions. The present study focused on the characterization and the potential functions of microbial communities colonizing the nutrient solutions recycled on slow filters during a whole cultivation season of 7 months in a tomato growing system. Bacterial microflora colonizing the solutions before and after they flew through the columns were studied. Two filters were amended with Pseudomonas putida (P-filter) or Bacillus cereus strains (B-filter), and a third filter was a control (C-filter). Biological activation of filter unit through bacterial amendment enhanced very significantly filter efficacy against plant potential pathogens Pythium spp. and Fusarium oxysporum. However, numerous bacteria (10 3 -10 4  CFU/mL) were detected in the effluent solutions. The community-level physiological profiling indicated a temporal shift of bacterial microflora, and the metabolism of nutrient solutions originally oriented towards carbohydrates progressively shifted towards degradation of amino acids and carboxylic acids over the 7-month period of experiment. Single-strand conformation polymorphism fingerprinting profiles showed that a shift between bacterial communities colonizing influent and effluent solutions of slow filters occurred. In comparison with influent, 16S rDNA sequencing revealed that phylotype diversity was low in the effluent of P- and C-filters, but no reduction was observed in the effluent of the B-filter. Suppressive potential of solutions filtered on a natural filter (C-filter), where the proportion of Proteobacteria (α- and β-) increased, whereas the proportion of uncultured candidate phyla rose in P- and B-filters, is discussed.

Bacterial community composition in Brazilian Anthrosols and adjacent soils characterized using culturing and molecular identification.

PubMed

O'Neill, B; Grossman, J; Tsai, M T; Gomes, J E; Lehmann, J; Peterson, J; Neves, E; Thies, J E

2009-07-01

Microbial community composition was examined in two soil types, Anthrosols and adjacent soils, sampled from three locations in the Brazilian Amazon. The Anthrosols, also known as Amazonian dark earths, are highly fertile soils that are a legacy of pre-Columbian settlement. Both Anthrosols and adjacent soils are derived from the same parent material and subject to the same environmental conditions, including rainfall and temperature; however, the Anthrosols contain high levels of charcoal-like black carbon from which they derive their dark color. The Anthrosols typically have higher cation exchange capacity, higher pH, and higher phosphorus and calcium contents. We used culture media prepared from soil extracts to isolate bacteria unique to the two soil types and then sequenced their 16S rRNA genes to determine their phylogenetic placement. Higher numbers of culturable bacteria, by over two orders of magnitude at the deepest sampling depths, were counted in the Anthrosols. Sequences of bacteria isolated on soil extract media yielded five possible new bacterial families. Also, a higher number of families in the bacteria were represented by isolates from the deeper soil depths in the Anthrosols. Higher bacterial populations and a greater diversity of isolates were found in all of the Anthrosols, to a depth of up to 1 m, compared to adjacent soils located within 50-500 m of their associated Anthrosols. Compared to standard culture media, soil extract media revealed diverse soil microbial populations adapted to the unique biochemistry and physiological ecology of these Anthrosols.

Bacterial community dynamics and activity in relation to dissolved organic matter availability during sea-ice formation in a mesocosm experiment.

PubMed

Eronen-Rasimus, Eeva; Kaartokallio, Hermanni; Lyra, Christina; Autio, Riitta; Kuosa, Harri; Dieckmann, Gerhard S; Thomas, David N

2014-02-01

The structure of sea-ice bacterial communities is frequently different from that in seawater. Bacterial entrainment in sea ice has been studied with traditional microbiological, bacterial abundance, and bacterial production methods. However, the dynamics of the changes in bacterial communities during the transition from open water to frozen sea ice is largely unknown. Given previous evidence that the nutritional status of the parent water may affect bacterial communities during ice formation, bacterial succession was studied in under ice water and sea ice in two series of mesocosms: the first containing seawater from the North Sea and the second containing seawater enriched with algal-derived dissolved organic matter (DOM). The composition and dynamics of bacterial communities were investigated with terminal restriction fragment length polymorphism (T-RFLP), and cloning alongside bacterial production (thymidine and leucine uptake) and abundance measurements (measured by flow cytometry). Enriched and active sea-ice bacterial communities developed in ice formed in both unenriched and DOM-enriched seawater (0-6 days). γ-Proteobacteria dominated in the DOM-enriched samples, indicative of their capability for opportunistic growth in sea ice. The bacterial communities in the unenriched waters and ice consisted of the classes Flavobacteria, α- and γ-Proteobacteria, which are frequently found in natural sea ice in polar regions. Furthermore, the results indicate that seawater bacterial communities are able to adapt rapidly to sudden environmental changes when facing considerable physicochemical stress such as the changes in temperature, salinity, nutrient status, and organic matter supply during ice formation. © 2014 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

Continous application of bioorganic fertilizer induced resilient culturable bacteria community associated with banana Fusarium wilt suppression

PubMed Central

Fu, Lin; Ruan, Yunze; Tao, Chengyuan; Li, Rong; Shen, Qirong

2016-01-01

Fusarium wilt of banana always drives farmers to find new land for banana cultivation due to the comeback of the disease after a few cropping years. A novel idea for solving this problem is the continuous application of bioorganic fertilizer (BIO), which should be practiced from the beginning of banana planting. In this study, BIO was applied in newly reclaimed fields to pre-control banana Fusarium wilt and the culturable rhizobacteria community were evaluated using Biolog Ecoplates and culture-dependent denaturing gradient gel electrophoresis (CD-DGGE). The results showed that BIO application significantly reduced disease incidences and increased crop yields, respectivly. And the stabilized general bacterial metabolic potential, especially for the utilization of carbohydrates, carboxylic acids and phenolic compounds, was induced by BIO application. DGGE profiles demonstrated that resilient community structure of culturable rhizobacteria with higher richness and diversity were observed in BIO treated soils. Morever, enriched culturable bacteria affiliated with Firmicutes, Gammaproteobacteria and Actinobacteria were also detected. In total, continuous application of BIO effectively suppressed Fusarium wilt disease by stabilizing culturable bacterial metabolic potential and community structure. This study revealed a new method to control Fusarium wilt of banana for long term banana cultivation. PMID:27306096

Continous application of bioorganic fertilizer induced resilient culturable bacteria community associated with banana Fusarium wilt suppression

NASA Astrophysics Data System (ADS)

Fu, Lin; Ruan, Yunze; Tao, Chengyuan; Li, Rong; Shen, Qirong

2016-06-01

Fusarium wilt of banana always drives farmers to find new land for banana cultivation due to the comeback of the disease after a few cropping years. A novel idea for solving this problem is the continuous application of bioorganic fertilizer (BIO), which should be practiced from the beginning of banana planting. In this study, BIO was applied in newly reclaimed fields to pre-control banana Fusarium wilt and the culturable rhizobacteria community were evaluated using Biolog Ecoplates and culture-dependent denaturing gradient gel electrophoresis (CD-DGGE). The results showed that BIO application significantly reduced disease incidences and increased crop yields, respectivly. And the stabilized general bacterial metabolic potential, especially for the utilization of carbohydrates, carboxylic acids and phenolic compounds, was induced by BIO application. DGGE profiles demonstrated that resilient community structure of culturable rhizobacteria with higher richness and diversity were observed in BIO treated soils. Morever, enriched culturable bacteria affiliated with Firmicutes, Gammaproteobacteria and Actinobacteria were also detected. In total, continuous application of BIO effectively suppressed Fusarium wilt disease by stabilizing culturable bacterial metabolic potential and community structure. This study revealed a new method to control Fusarium wilt of banana for long term banana cultivation.

Protozoa Drive the Dynamics of Culturable Biocontrol Bacterial Communities.

PubMed

Müller, Maren Stella; Scheu, Stefan; Jousset, Alexandre

2013-01-01

Some soil bacteria protect plants against soil-borne diseases by producing toxic secondary metabolites. Such beneficial biocontrol bacteria can be used in agricultural systems as alternative to agrochemicals. The broad spectrum toxins responsible for plant protection also inhibit predation by protozoa and nematodes, the main consumers of bacteria in soil. Therefore, predation pressure may favour biocontrol bacteria and contribute to plant health. We analyzed the effect of Acanthamoeba castellanii on semi-natural soil bacterial communities in a microcosm experiment. We determined the frequency of culturable bacteria carrying genes responsible for the production of the antifungal compounds 2,4-diacetylphloroglucinol (DAPG), pyrrolnitrin (PRN) and hydrogen cyanide (HCN) in presence and absence of A. castellanii. We then measured if amoebae affected soil suppressiveness in a bioassay with sugar beet seedlings confronted to the fungal pathogen Rhizoctonia solani. Amoebae increased the frequency of both DAPG and HCN positive bacteria in later plant growth phases (2 and 3 weeks), as well as the average number of biocontrol genes per bacterium. The abundance of DAPG positive bacteria correlated with disease suppression, suggesting that their promotion by amoebae may enhance soil health. However, the net effect of amoebae on soil suppressiveness was neutral to slightly negative, possibly because amoebae slow down the establishment of biocontrol bacteria on the recently emerged seedlings used in the assay. The results indicate that microfaunal predators foster biocontrol bacterial communities. Understanding interactions between biocontrol bacteria and their predators may thus help developing environmentally friendly management practices of agricultural systems.

Protozoa Drive the Dynamics of Culturable Biocontrol Bacterial Communities

PubMed Central

Müller, Maren Stella; Scheu, Stefan; Jousset, Alexandre

2013-01-01

Some soil bacteria protect plants against soil-borne diseases by producing toxic secondary metabolites. Such beneficial biocontrol bacteria can be used in agricultural systems as alternative to agrochemicals. The broad spectrum toxins responsible for plant protection also inhibit predation by protozoa and nematodes, the main consumers of bacteria in soil. Therefore, predation pressure may favour biocontrol bacteria and contribute to plant health. We analyzed the effect of Acanthamoeba castellanii on semi-natural soil bacterial communities in a microcosm experiment. We determined the frequency of culturable bacteria carrying genes responsible for the production of the antifungal compounds 2,4-diacetylphloroglucinol (DAPG), pyrrolnitrin (PRN) and hydrogen cyanide (HCN) in presence and absence of A. castellanii. We then measured if amoebae affected soil suppressiveness in a bioassay with sugar beet seedlings confronted to the fungal pathogen Rhizoctonia solani. Amoebae increased the frequency of both DAPG and HCN positive bacteria in later plant growth phases (2 and 3 weeks), as well as the average number of biocontrol genes per bacterium. The abundance of DAPG positive bacteria correlated with disease suppression, suggesting that their promotion by amoebae may enhance soil health. However, the net effect of amoebae on soil suppressiveness was neutral to slightly negative, possibly because amoebae slow down the establishment of biocontrol bacteria on the recently emerged seedlings used in the assay. The results indicate that microfaunal predators foster biocontrol bacterial communities. Understanding interactions between biocontrol bacteria and their predators may thus help developing environmentally friendly management practices of agricultural systems. PMID:23840423

Anaerobic Oxidization of Methane in a Minerotrophic Peatland: Enrichment of Nitrite-Dependent Methane-Oxidizing Bacteria

PubMed Central

Zhu, Baoli; van Dijk, Gijs; Fritz, Christian; Smolders, Alfons J. P.; Pol, Arjan; Jetten, Mike S. M.

2012-01-01

The importance of anaerobic oxidation of methane (AOM) as a methane sink in freshwater systems is largely unexplored, particularly in peat ecosystems. Nitrite-dependent anaerobic methane oxidation (n-damo) was recently discovered and reported to be catalyzed by the bacterium “Candidatus Methylomirabilis oxyfera,” which is affiliated with the NC10 phylum. So far, several “Ca. Methylomirabilis oxyfera” enrichment cultures have been obtained using a limited number of freshwater sediments or wastewater treatment sludge as the inoculum. In this study, using stable isotope measurements and porewater profiles, we investigated the potential of n-damo in a minerotrophic peatland in the south of the Netherlands that is infiltrated by nitrate-rich ground water. Methane and nitrate profiles suggested that all methane produced was oxidized before reaching the oxic layer, and NC10 bacteria could be active in the transition zone where countergradients of methane and nitrate occur. Quantitative PCR showed high NC10 bacterial cell numbers at this methane-nitrate transition zone. This soil section was used to enrich the prevalent NC10 bacteria in a continuous culture supplied with methane and nitrite at an in situ pH of 6.2. An enrichment of nitrite-reducing methanotrophic NC10 bacteria was successfully obtained. Phylogenetic analysis of retrieved 16S rRNA and pmoA genes showed that the enriched bacteria were very similar to the ones found in situ and constituted a new branch of NC10 bacteria with an identity of less than 96 and 90% to the 16S rRNA and pmoA genes of “Ca. Methylomirabilis oxyfera,” respectively. The results of this study expand our knowledge of the diversity and distribution of NC10 bacteria in the environment and highlight their potential contribution to nitrogen and methane cycles. PMID:23042166

Anaerobic oxidation of ethene coupled to sulfate reduction in microcosms and enrichment cultures.

PubMed

Fullerton, Heather; Crawford, Michael; Bakenne, Ademola; Freedman, David L; Zinder, Stephen H

2013-01-01

Ethene is considered recalcitrant under anaerobic conditions, but biological reduction to ethane and oxidation to CO2 have been reported; however, little is known about these processes or the organisms carrying them out. In this report we describe sulfate dependent ethene consumption in microcosms prepared with sediments from a freshwater canal. A first dose of 0.6 mmol/L ethene was consumed within 77 days, and a second dose was largely consumed twelve days later. Material from this microcosm was transferred into growth medium with ethene as the only electron donor (except for trace amounts of vitamins) and sulfate as the electron acceptor. Four doses of ethene were consumed at increasing rates, and the cultures have been transferred at least eight times in this medium. Conversion of [(14)C]ethene primarily to (14)CO2 was demonstrated in fifth and sixth generation cultures, as well as production of sulfide in other cultures, confirming the ethene/sulfate couple. Ovoid cells 1-2 μm in diameter were found in cultures containing ethene and sulfate, and quantitative PCR showed large increases in bacterial 16S rRNA gene copy number. Over half of a 16S rRNA gene clone library from a sixth-generation culture was a phylotype with a sequence ca. 90% identical with a clade of Deltaproteobacteria that includes Desulfovirga adipica and several Syntrophobacter spp. These studies have solidified the concept that deficits in mass balances for chloroethene fate in sulfate reducing zones of contaminated groundwater sites may be due to ethene oxidation, and suggest a unique phylotype is involved in this process.

Cerebrospinal fluid cytokines in the diagnosis of bacterial meningitis in infants.

PubMed

Srinivasan, Lakshmi; Kilpatrick, Laurie; Shah, Samir S; Abbasi, Soraya; Harris, Mary C

2016-10-01

Bacterial meningitis poses diagnostic challenges in infants. Antibiotic pretreatment and low bacterial density diminish cerebrospinal fluid (CSF) culture yield, while laboratory parameters do not reliably identify bacterial meningitis. Pro and anti-inflammatory cytokines are elevated in bacterial meningitis and may be useful diagnostic adjuncts when CSF cultures are negative. In a prospective cohort study of infants, we used cytometric bead arrays to measure tumor necrosis factor alpha (TNF-α), interleukin 1 (IL-1), IL-6, IL-8, IL-10, and IL-12 in CSF. Receiver operating characteristic (ROC) analyses and Principal component analysis (PCA) were used to determine cytokine combinations that identified bacterial meningitis. Six hundred and eighty four infants < 6 mo were included; 11 had culture-proven bacterial meningitis. IL-6 and IL-10 were the individual cytokines possessing greatest accuracy in diagnosis of culture proven bacterial meningitis (ROC analyses; area under the concentration-time curve (AUC) 0.91; 0.9103 respectively), and performed as well as, or better than combinations identified using ROC and PCA. CSF cytokines were highly correlated with each other and with CSF white blood cell count (WBC) counts in infants with meningitis. A subset of antibiotic pretreated culture-negative subjects demonstrated cytokine patterns similar to culture positive subjects. CSF cytokine levels may aid diagnosis of bacterial meningitis, and facilitate decision-making regarding treatment for culture negative meningitis.

Efficacy of a commercial probiotic relative to oxytetracycline as Gram-negative bacterial control agents in a rotifer (Brachionus plicatilis) batch culture

USDA-ARS?s Scientific Manuscript database

Two trials were conducted to evaluate two gram-negative bacterial control strategies in batch cultures of the rotifer Brachionus plicatilis. In the first trial, rotifers at an initial density of 47/mL were cultured for 5 d and dosed with a 10-mg/L solution of either oxytetracycline or a commercial p...

Stable Carbon Isotope Fractionation during Bacterial Acetylene Fermentation: Potential for Life Detection in Hydrocarbon-Rich Volatiles of Icy Planet(oid)s

PubMed Central

Baesman, Shaun M.; Oremland, Ronald S.

2015-01-01

Abstract We report the first study of stable carbon isotope fractionation during microbial fermentation of acetylene (C2H2) in sediments, sediment enrichments, and bacterial cultures. Kinetic isotope effects (KIEs) averaged 3.7 ± 0.5‰ for slurries prepared with sediment collected at an intertidal mudflat in San Francisco Bay and 2.7 ± 0.2‰ for a pure culture of Pelobacter sp. isolated from these sediments. A similar KIE of 1.8 ± 0.7‰ was obtained for methanogenic enrichments derived from sediment collected at freshwater Searsville Lake, California. However, C2H2 uptake by a highly enriched mixed culture (strain SV7) obtained from Searsville Lake sediments resulted in a larger KIE of 9.0 ± 0.7‰. These are modest KIEs when compared with fractionation observed during oxidation of C1 compounds such as methane and methyl halides but are comparable to results obtained with other C2 compounds. These observations may be useful in distinguishing biologically active processes operating at distant locales in the Solar System where C2H2 is present. These locales include the surface of Saturn's largest moon Titan and the vaporous water- and hydrocarbon-rich jets emanating from Enceladus. Key Words: Acetylene—Fermentation—Isotope fractionation—Enceladus—Life detection. Astrobiology 15, 977–986. PMID:26539733

Stable carbon isotope fractionation during bacterial acetylene fermentation: Potential for life detection in hydrocarbon-rich volatiles of icy planet(oid)s

USGS Publications Warehouse

Miller, Laurence; Baesman, Shaun; Oremland, Ron

2015-01-01

We report the first study of stable carbon isotope fractionation during microbial fermentation of acetylene (C2H2) in sediments, sediment enrichments, and bacterial cultures. Kinetic isotope effects (KIEs) averaged 3.7 ± 0.5‰ for slurries prepared with sediment collected at an intertidal mudflat in San Francisco Bay and 2.7 ± 0.2‰ for a pure culture of Pelobacter sp. isolated from these sediments. A similar KIE of 1.8 ± 0.7‰ was obtained for methanogenic enrichments derived from sediment collected at freshwater Searsville Lake, California. However, C2H2 uptake by a highly enriched mixed culture (strain SV7) obtained from Searsville Lake sediments resulted in a larger KIE of 9.0 ± 0.7‰. These are modest KIEs when compared with fractionation observed during oxidation of C1 compounds such as methane and methyl halides but are comparable to results obtained with other C2compounds. These observations may be useful in distinguishing biologically active processes operating at distant locales in the Solar System where C2H2 is present. These locales include the surface of Saturn's largest moon Titan and the vaporous water- and hydrocarbon-rich jets emanating from Enceladus.

Molecular Phylogenetic Exploration of Bacterial Diversity in a Bakreshwar (India) Hot Spring and Culture of Shewanella-Related Thermophiles

PubMed Central

Ghosh, Dhritiman; Bal, Bijay; Kashyap, V. K.; Pal, Subrata

2003-01-01

The bacterial diversity of a hot spring in Bakreshwar, India, was investigated by a culture-independent approach. 16S ribosomal DNA clones derived from the sediment samples were found to be associated with gamma-Proteobacteria, cyanobacteria, and green nonsulfur and low-GC gram-positive bacteria. The first of the above phylotypes cobranches with Shewanella, a well-known iron reducer. This phylogenetic correlation has been exploited to develop culture conditions for thermophilic iron-reducing microorganisms. PMID:12839826

Bacterial community dynamics during start-up of a trickle-bed bioreactor degrading aromatic compounds.

PubMed

Stoffels, M; Amann, R; Ludwig, W; Hekmat, D; Schleifer, K H

1998-03-01

This study was performed with a laboratory-scale fixed-bed bioreactor degrading a mixture of aromatic compounds (Solvesso100). The starter culture for the bioreactor was prepared in a fermentor with a wastewater sample of a care painting facility as the inoculum and Solvesso100 as the sole carbon source. The bacterial community dynamics in the fermentor and the bioreactor were examined by a conventional isolation procedure and in situ hybridization with fluorescently labeled rRNA-targeted oligonucleotides. Two significant shifts in the bacterial community structure could be demonstrated. The original inoculum from the wastewater of the car factory was rich in proteobacteria of the alpha and beta subclasses, while the final fermentor enrichment was dominated by bacteria closely related to Pseudomonas putida or Pseudomonas mendocina, which both belong to the gamma subclass of the class Proteobacteria. A second significant shift was observed when the fermentor culture was transferred as inoculum to the trickle-bed bioreactor. The community structure in the bioreactor gradually returned to a higher complexity, with the dominance of beta and alpha subclass proteobacteria, whereas the gamma subclass proteobacteria sharply declined. Obviously, the preceded pollutant adaptant did not lead to a significant enrichment of bacteria that finally dominated in the trickle-bed bioreactor. In the course of experiments, three new 16S as well as 23S rRNA-targeted probes for beta subclass proteobacteria were designed, probe SUBU1237 for the genera Burkholderia and Sutterella, probe ALBO34a for the genera Alcaligenes and Bordetella, and probe Bcv13b for Burkholderia cepacia and Burkholderia vietnamiensis. Bacteria hybridizing with the probe Bcv13b represented the main Solvesso100-degrading population in the reactor.

Proteomics as a Quality Control Tool of Pharmaceutical Probiotic Bacterial Lysate Products

PubMed Central

Klein, Günter; Schanstra, Joost P.; Hoffmann, Janosch; Mischak, Harald; Siwy, Justyna; Zimmermann, Kurt

2013-01-01

Probiotic bacteria have a wide range of applications in veterinary and human therapeutics. Inactivated probiotics are complex samples and quality control (QC) should measure as many molecular features as possible. Capillary electrophoresis coupled to mass spectrometry (CE/MS) has been used as a multidimensional and high throughput method for the identification and validation of biomarkers of disease in complex biological samples such as biofluids. In this study we evaluate the suitability of CE/MS to measure the consistency of different lots of the probiotic formulation Pro-Symbioflor which is a bacterial lysate of heat-inactivated Escherichia coli and Enterococcus faecalis. Over 5000 peptides were detected by CE/MS in 5 different lots of the bacterial lysate and in a sample of culture medium. 71 to 75% of the total peptide content was identical in all lots. This percentage increased to 87–89% when allowing the absence of a peptide in one of the 5 samples. These results, based on over 2000 peptides, suggest high similarity of the 5 different lots. Sequence analysis identified peptides of both E. coli and E. faecalis and peptides originating from the culture medium, thus confirming the presence of the strains in the formulation. Ontology analysis suggested that the majority of the peptides identified for E. coli originated from the cell membrane or the fimbrium, while peptides identified for E. faecalis were enriched for peptides originating from the cytoplasm. The bacterial lysate peptides as a whole are recognised as highly conserved molecular patterns by the innate immune system as microbe associated molecular pattern (MAMP). Sequence analysis also identified the presence of soybean, yeast and casein protein fragments that are part of the formulation of the culture medium. In conclusion CE/MS seems an appropriate QC tool to analyze complex biological products such as inactivated probiotic formulations and allows determining the similarity between lots. PMID

Epidemiology of Salmonella sp. in California cull dairy cattle: prevalence of fecal shedding and diagnostic accuracy of pooled enriched broth culture of fecal samples

PubMed Central

Abu Aboud, Omran A.; Adaska, John M.; Williams, Deniece R.; Rossitto, Paul V.; Champagne, John D.; Lehenbauer, Terry W.; Atwill, Robert; Li, Xunde

2016-01-01

Background The primary objective of this cross-sectional study was to estimate the crude, seasonal and cull-reason stratified prevalence of Salmonella fecal shedding in cull dairy cattle on seven California dairies. A secondary objective was to estimate and compare the relative sensitivity (Se) and specificity (Sp) for pools of 5 and 10 enriched broth cultures of fecal samples for Salmonella sp. detection. Methods Seven dairy farms located in the San Joaquin Valley of California were identified and enrolled in the study as a convenience sample. Cull cows were identified for fecal sampling once during each season between 2014 and 2015, specifically during spring, summer, fall, and winter, and 10 cows were randomly selected for fecal sampling at the day of their sale. In addition, study personnel completed a survey based on responses of the herd manager to questions related to the previous four month’s herd management. Fecal samples were frozen until testing for Salmonella. After overnight enrichment in liquid broth, pools of enrichment broth (EBP) were created for 5 and 10 samples. All individual and pooled broths were cultured on selective media with putative Salmonella colonies confirmed by biochemical testing before being serogrouped and serotyped. Results A total of 249 cull cows were enrolled into the study and their fecal samples tested for Salmonella. The survey-weighted period prevalence of fecal shedding of all Salmonella sp. in the cull cow samples across all study herds and the entire study period was 3.42% (N = 249; SE 1.07). The within herd prevalence of Salmonella shed in feces did not differ over the four study seasons (P = 0.074). The Se of culture of EBP of five samples was 62.5% (SE = 17.12), which was not statistically different from the Se of culture of EBP of 10 (37.5%, SE = 17.12, P = 0.48). The Sp of culture of EBP of five samples was 95.24% (SE = 3.29) and for pools of 10 samples was 100.00% (SE = 0). There was no statistical difference

Epidemiology of Salmonella sp. in California cull dairy cattle: prevalence of fecal shedding and diagnostic accuracy of pooled enriched broth culture of fecal samples.

PubMed

Abu Aboud, Omran A; Adaska, John M; Williams, Deniece R; Rossitto, Paul V; Champagne, John D; Lehenbauer, Terry W; Atwill, Robert; Li, Xunde; Aly, Sharif S

2016-01-01

The primary objective of this cross-sectional study was to estimate the crude, seasonal and cull-reason stratified prevalence of Salmonella fecal shedding in cull dairy cattle on seven California dairies. A secondary objective was to estimate and compare the relative sensitivity (Se) and specificity (Sp) for pools of 5 and 10 enriched broth cultures of fecal samples for Salmonella sp. detection. Seven dairy farms located in the San Joaquin Valley of California were identified and enrolled in the study as a convenience sample. Cull cows were identified for fecal sampling once during each season between 2014 and 2015, specifically during spring, summer, fall, and winter, and 10 cows were randomly selected for fecal sampling at the day of their sale. In addition, study personnel completed a survey based on responses of the herd manager to questions related to the previous four month's herd management. Fecal samples were frozen until testing for Salmonella. After overnight enrichment in liquid broth, pools of enrichment broth (EBP) were created for 5 and 10 samples. All individual and pooled broths were cultured on selective media with putative Salmonella colonies confirmed by biochemical testing before being serogrouped and serotyped. A total of 249 cull cows were enrolled into the study and their fecal samples tested for Salmonella. The survey-weighted period prevalence of fecal shedding of all Salmonella sp. in the cull cow samples across all study herds and the entire study period was 3.42% (N = 249; SE 1.07). The within herd prevalence of Salmonella shed in feces did not differ over the four study seasons (P = 0.074). The Se of culture of EBP of five samples was 62.5% (SE = 17.12), which was not statistically different from the Se of culture of EBP of 10 (37.5%, SE = 17.12, P = 0.48). The Sp of culture of EBP of five samples was 95.24% (SE = 3.29) and for pools of 10 samples was 100.00% (SE = 0). There was no statistical difference between the culture relative

Identifying the bacterial community on the surface of Intralox belting in a meat boning room by culture-dependent and culture-independent 16S rDNA sequence analysis.

PubMed

Brightwell, Gale; Boerema, Jackie; Mills, John; Mowat, Eilidh; Pulford, David

2006-05-25

We examined the bacterial community present on an Intralox conveyor belt system in an operating lamb boning room by sequencing the 16S ribosomal DNA (rDNA) of bacteria extracted in the presence or absence of cultivation. RFLP patterns for 16S rDNA clone library and cultures were generated using HaeIII and MspI restriction endonucleases. 16S rDNA amplicons produced 8 distinct RFLP pattern groups. RFLP groups I-IV were represented in the clone library and RFLP groups I and V-VIII were represented amongst the cultured isolates. Partial DNA sequences from each RFLP group revealed that all group I, II and VIII representatives were Pseudomonas spp., group III were Sphingomonas spp., group IV clones were most similar to an uncultured alpha proteobacterium, group V was similar to a Serratia spp., group VI with an Alcaligenes spp., and group VII with Microbacterium spp. Sphingomonads were numerically dominant in the culture-independent clone library and along with the group IV alpha proteobacterium were not represented amongst the cultured isolates. Serratia, Alcaligenes and Microbacterium spp. were only represented with cultured isolates. Pseudomonads were detected by both culture-dependent (84% of isolates) and culture-independent (12.5% of clones) methods and their presence at high frequency does pose the risk of product spoilage if transferred onto meat stored under aerobic conditions. The detection of sphingomonads in large numbers by the culture-independent method demands further analysis because sphingomonads may represent a new source of meat spoilage that has not been previously recognised in the meat processing environment. The 16S rDNA collections generated by both methods were important at representing the diversity of the bacterial population associated with an Intralox conveyor belt system.

Bacterial diversity in three distinct sub-habitats within the pitchers of the northern pitcher plant, Sarracenia purpurea.

PubMed

Krieger, Joseph R; Kourtev, Peter S

2012-03-01

Pitcher plants have been widely used in ecological studies of food webs; however, their bacterial communities are poorly characterized. Pitchers of Sarracenia purpurea contain several distinct sub-habitats, namely the bottom sediment, the liquid, and the internal pitcher wall. We hypothesized that those three sub-habitats within pitcher plants are inhabited by distinct bacterial populations. We used denaturing gradient gel electrophoresis and 16S rRNA gene sequencing to characterize bacterial populations in pitchers from three bogs. DGGE and sequencing revealed that in any given pitcher, the three sub-habitats contain significantly different bacterial populations. However, there was significant variability between bacterial populations inhabiting the same type of habitat in different pitchers, even at the same site. Therefore, no consistent set of bacterial populations was enriched in any of the three sub-habitats. All sub-habitats appeared to be dominated by alpha- and betaproteobacteria in differing proportions. In addition, sequences from the Bacteroidetes and Firmicutes were obtained from all three sub-habitats. We conclude that container aquatic habitats such as the pitchers of S. purpurea possess a very high bacterial diversity, with many unique bacterial populations enriched in individual pitchers. Within an individual pitcher, populations of certain bacterial families may be enriched in one of the three studied sub-habitats. © 2011 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

Distributions of Bacterial Generalists among the Guts of Birds ...

EPA Pesticide Factsheets

Complex distributions of bacterial taxa within diverse animal microbiomes have inspired ecological and biogeographical approaches to revealing the functions of taxa that may be most important for host health. Of particular interest are bacteria that find many diverse habitats suitable for growth and remain competitive amongst finely-tuned host specialists. While previous work has focused on identifying these specialists, here our aims were to 1) identify generalist taxa, 2) identify taxonomic clades with enriched generalist diversity, and 3) describe the distribution of the largest generalist groups among hosts. We analyzed existing bacterial rRNA tag-sequencing data (v6) available on VAMPs (vamps.mbl.edu) from the microbiomes of 12 host species (106 samples total) spanning birds, mammals, and fish for generalist taxa using the CLAM test. OTUs with approximately equal abundance and a minimum of 10 reads in two hosts were classified as generalists. Generalist OTUs (n=2,982) were found in all hosts tested. Bacterial families Alcaligenaceae and Burkholderiaceae were significantly enriched with generalists OTUs compared to other families. Bacterial families such as Bacteroidaceae and Lachnospiraceae significantly lacked generalists OTUs compared to other families. Enterobacteriaceae, Peptostreptococcaceae, and Erysipelotrichaceae more so than other bacterial families were widely distributed and abundant in birds, mammals, and fish suggesting that these taxa mainta

Effect of Copper Treatment on the Composition and Function of the Bacterial Community in the Sponge Haliclona cymaeformis

PubMed Central

Tian, Ren-Mao; Wang, Yong; Bougouffa, Salim; Gao, Zhao-Ming; Cai, Lin; Zhang, Wei-Peng; Bajic, Vladimir

2014-01-01

ABSTRACT Marine sponges are the most primitive metazoan and host symbiotic microorganisms. They are crucial components of the marine ecological system and play an essential role in pelagic processes. Copper pollution is currently a widespread problem and poses a threat to marine organisms. Here, we examined the effects of copper treatment on the composition of the sponge-associated bacterial community and the genetic features that facilitate the survival of enriched bacteria under copper stress. The 16S rRNA gene sequencing results showed that the sponge Haliclona cymaeformis harbored symbiotic sulfur-oxidizing Ectothiorhodospiraceae and photosynthetic Cyanobacteria as dominant species. However, these autotrophic bacteria decreased substantially after treatment with a high copper concentration, which enriched for a heterotrophic-bacterium-dominated community. Metagenomic comparison revealed a varied profile of functional genes and enriched functions, including bacterial motility and chemotaxis, extracellular polysaccharide and capsule synthesis, virulence-associated genes, and genes involved in cell signaling and regulation, suggesting short-period mechanisms of the enriched bacterial community for surviving copper stress in the microenvironment of the sponge. Microscopic observation and comparison revealed dynamic bacterial aggregation within the matrix and lysis of sponge cells. The bacteriophage community was also enriched, and the complete genome of a dominant phage was determined, implying that a lytic phage cycle was stimulated by the high copper concentration. This study demonstrated a copper-induced shift in the composition of functional genes of the sponge-associated bacterial community, revealing the selective effect of copper treatment on the functions of the bacterial community in the microenvironment of the sponge. PMID:25370493

Bacterial meningitis.

PubMed

Heckenberg, Sebastiaan G B; Brouwer, Matthijs C; van de Beek, Diederik

2014-01-01

Bacterial meningitis is a neurologic emergency. Vaccination against common pathogens has decreased the burden of disease. Early diagnosis and rapid initiation of empiric antimicrobial and adjunctive therapy are vital. Therapy should be initiated as soon as blood cultures have been obtained, preceding any imaging studies. Clinical signs suggestive of bacterial meningitis include fever, headache, meningismus, and an altered level of consciousness but signs may be scarce in children, in the elderly, and in meningococcal disease. Host genetic factors are major determinants of susceptibility to meningococcal and pneumococcal disease. Dexamethasone therapy has been implemented as adjunctive treatment of adults with pneumococcal meningitis. Adequate and prompt treatment of bacterial meningitis is critical to outcome. In this chapter we review the epidemiology, pathophysiology, and management of bacterial meningitis. © 2014 Elsevier B.V. All rights reserved.

Delignification of disposable wooden chopsticks waste for fermentative hydrogen production by an enriched culture from a hot spring.

PubMed

Phummala, Kanthima; Imai, Tsuyoshi; Reungsang, Alissara; Chairattanamanokorn, Prapaipid; Sekine, Masahiko; Higuchi, Takaya; Yamamoto, Koichi; Kanno, Ariyo

2014-06-01

Hydrogen (H2) production from lignocellulosic materials may be enhanced by removing lignin and increasing the porosity of the material prior to enzymatic hydrolysis. Alkaline pretreatment conditions, used to delignify disposable wooden chopsticks (DWC) waste, were investigated. The effects of NaOH concentration, temperature and retention time were examined and it was found that retention time had no effect on lignin removal or carbohydrate released in enzymatic hydrolysate. The highest percentage of lignin removal (41%) was obtained with 2% NaOH at 100°C, correlated with the highest carbohydrate released (67 mg/g pretreated DWC) in the hydrolysate. An enriched culture from a hot spring was used as inoculum for fermentative H2 production, and its optimum initial pH and temperature were determined to be 7.0 and 50°C, respectively. Furthermore, enzymatic hydrolysate from pretreated DWC was successfully demonstrated as a substrate for fermentative H2 production by the enriched culture. The maximum H2 yield and production rate were achieved at 195 mL H2/g total sugars consumed and 116 mL H2/(L·day), respectively. Copyright © 2014 The Research Centre for Eco-Environmental Sciences, Chinese Academy of Sciences. Published by Elsevier B.V. All rights reserved.

Effect of bacterial components of mixed culture supernatants of planktonic and biofilm Pseudomonas aeruginosa with commensal Escherichia coli on the neutrophil response in vitro.

PubMed

Maslennikova, Irina L; Kuznetsova, Marina V; Nekrasova, Irina V; Shirshev, Sergei V

2017-11-30

Pseudomonas aeruginosa (PA) responsible for acute and chronic infections often forms a well-organized bacterial population with different microbial species including commensal strains of Escherichia coli. Bacterial extracellular components of mixed culture can modulate the influence of bacteria on the neutrophil functions. The objective of this study was to compare the effect of pyocyanin, pyoverdine, LPS, exopolysaccharide of single species and mixed culture supernatants of PA strains and E. coli K12 on microbicidal, secretory activity of human neutrophils in vitro. Bacterial components of E. coli K12 in mixed supernatants with 'biofilm' PA strains (PA ATCC, PA BALG) enhanced short-term microbicidal mechanisms and inhibited neutrophil secretion delayed in time. The influence of 'planktonic' PA (PA 9-3) exometabolites in mixed culture is almost mimicked by E. coli K12 effect on functional neutrophil changes. This investigation may help to understand some of the mechanisms of neutrophil response to mixed infections of different PA with other bacteria species. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Aerobic biological treatment of low-strength synthetic wastewater in membrane-coupled bioreactors: the structure and function of bacterial enrichment cultures as the net growth rate approaches zero.

PubMed

Chen, Ruoyu; LaPara, Timothy M

2006-01-01

The goal of the current research was to determine if the stringent nutrient limitation imposed by membrane-coupled bioreactors (MBRs) could be used to force mixed bacterial communities to exhibit a zero net growth rate over an extended time period. Mechanistically, this zero net growth rate could be achieved when the amount of energy available for growth is balanced by the maintenance requirements of the bacterial community. Bench-scale MBRs were fed synthetic feed medium containing gelatin as the major organic substrate. Biomass concentrations initially increased rapidly, but subsequently declined until an asymptote was reached. Leucine aminopeptidase activities concomitantly increased by at least 10-fold, suggesting that bacterial catabolic activity remained high even while growth rates became negligible. In contrast, alpha-glucosidase and heptanoate esterase activities decreased, indicating that the bacterial community specifically adapted to the carbon source in the feed medium. Bacterial community analysis by denaturing gradient gel electrophoresis of PCR-amplified 16S rRNA gene fragments (PCR-DGGE) suggested that the bacterial community structure completely changed from the beginning to the end of each MBR. Excision and nucleotide sequence analysis of prominent PCR-DGGE bands suggested that many of the dominant populations were similar to novel bacterial strains that were previously uncultivated or recently cultivated during studies specifically targeting these novel populations. This research demonstrates that MBRs have substantial practical applications for biological wastewater treatment; in addition, MBRs are a useful tool to study the ecology of slow-growing bacteria.

Analysis of Bacterial Community Structure in Sulfurous-Oil-Containing Soils and Detection of Species Carrying Dibenzothiophene Desulfurization (dsz) Genes

PubMed Central

Duarte, Gabriela Frois; Rosado, Alexandre Soares; Seldin, Lucy; de Araujo, Welington; van Elsas, Jan Dirk

2001-01-01

The selective effects of sulfur-containing hydrocarbons, with respect to changes in bacterial community structure and selection of desulfurizing organisms and genes, were studied in soil. Samples taken from a polluted field soil (A) along a concentration gradient of sulfurous oil and from soil microcosms treated with dibenzothiophene (DBT)-containing petroleum (FSL soil) were analyzed. Analyses included plate counts of total bacteria and of DBT utilizers, molecular community profiling via soil DNA-based PCR-denaturing gradient gel electrophoresis (PCR-DGGE), and detection of genes that encode enzymes involved in the desulfurization of hydrocarbons, i.e., dszA, dszB, and dszC.Data obtained from the A soil showed no discriminating effects of oil levels on the culturable bacterial numbers on either medium used. Generally, counts of DBT degraders were 10- to 100-fold lower than the total culturable counts. However, PCR-DGGE showed that the numbers of bands detected in the molecular community profiles decreased with increasing oil content of the soil. Analysis of the sequences of three prominent bands of the profiles generated with the highly polluted soil samples suggested that the underlying organisms were related to Actinomyces sp., Arthrobacter sp., and a bacterium of uncertain affiliation. dszA, dszB, and dszC genes were present in all A soil samples, whereas a range of unpolluted soils gave negative results in this analysis. Results from the study of FSL soil revealed minor effects of the petroleum-DBT treatment on culturable bacterial numbers and clear effects on the DBT-utilizing communities. The molecular community profiles were largely stable over time in the untreated soil, whereas they showed a progressive change over time following treatment with DBT-containing petroleum. Direct PCR assessment revealed the presence of dszB-related signals in the untreated FSL soil and the apparent selection of dszA- and dszC-related sequences by the petroleum-DBT treatment

Microbial Diversity in Sulfate-Reducing Marine Sediment Enrichment Cultures Associated with Anaerobic Biotransformation of Coastal Stockpiled Phosphogypsum (Sfax, Tunisia)

PubMed Central

Zouch, Hana; Karray, Fatma; Armougom, Fabrice; Chifflet, Sandrine; Hirschler-Réa, Agnès; Kharrat, Hanen; Kamoun, Lotfi; Ben Hania, Wajdi; Ollivier, Bernard; Sayadi, Sami; Quéméneur, Marianne

2017-01-01

Anaerobic biotechnology using sulfate-reducing bacteria (SRB) is a promising alternative for reducing long-term stockpiling of phosphogypsum (PG), an acidic (pH ~3) by-product of the phosphate fertilizer industries containing high amounts of sulfate. The main objective of this study was to evaluate, for the first time, the diversity and ability of anaerobic marine microorganisms to convert sulfate from PG into sulfide, in order to look for marine SRB of biotechnological interest. A series of sulfate-reducing enrichment cultures were performed using different electron donors (i.e., acetate, formate, or lactate) and sulfate sources (i.e., sodium sulfate or PG) as electron acceptors. Significant sulfide production was observed from enrichment cultures inoculated with marine sediments, collected near the effluent discharge point of a Tunisian fertilizer industry (Sfax, Tunisia). Sulfate sources impacted sulfide production rates from marine sediments as well as the diversity of SRB species belonging to Deltaproteobacteria. When PG was used as sulfate source, Desulfovibrio species dominated microbial communities of marine sediments, while Desulfobacter species were mainly detected using sodium sulfate. Sulfide production was also affected depending on the electron donor used, with the highest production obtained using formate. In contrast, low sulfide production (acetate-containing cultures) was associated with an increase in the population of Firmicutes. These results suggested that marine Desulfovibrio species, to be further isolated, are potential candidates for bioremediation of PG by immobilizing metals and metalloids thanks to sulfide production by these SRB. PMID:28871244

Microbiome and Culture Based Analysis of Chronic Rhinosinusitis Compared to Healthy Sinus Mucosa

PubMed Central

Koeller, Kerstin; Herlemann, Daniel P. R.; Schuldt, Tobias; Ovari, Attila; Guder, Ellen; Podbielski, Andreas; Kreikemeyer, Bernd; Olzowy, Bernhard

2018-01-01

The role of bacteria in chronic rhinosinusitis (CRS) is still not well understood. Whole microbiome analysis adds new aspects to our current understanding that is mainly based on isolated bacteria. It is still unclear how the results of microbiome analysis and the classical culture based approaches interrelate. To address this, middle meatus swabs and tissue samples were obtained during sinus surgery in 5 patients with CRS with nasal polyps (CRSwNP), 5 patients with diffuse CRS without nasal polyps (CRSsNP), 5 patients with unilateral purulent maxillary CRS (upm CRS) and 3 patients with healthy sinus mucosa. Swabs were cultured, and associated bacteria were identified. Additionally, parts of each tissue sample also underwent culture approaches, and in parallel DNA was extracted for 16S rRNA gene amplicon-based microbiome analysis. From tissue samples 4.2 ± 1.2 distinct species per patient were cultured, from swabs 5.4 ± 1.6. The most frequently cultured species from the swabs were Propionibacterium acnes, Staphylococcus epidermidis, Corynebacterium spp. and Staphylococcus aureus. The 16S-RNA gene analysis revealed no clear differentiation of the bacterial community of healthy compared to CRS samples of unilateral purulent maxillary CRS and CRSwNP. However, the bacterial community of CRSsNP differed significantly from the healthy controls. In the CRSsNP samples Flavobacterium, Pseudomonas, Pedobacter, Porphyromonas, Stenotrophomonas, and Brevundimonas were significantly enriched compared to the healthy controls. Species isolated from culture did not generally correspond with the most abundant genera in microbiome analysis. Only Fusobacteria, Parvimonas, and Prevotella found in 2 unilateral purulent maxillary CRS samples by the cultivation dependent approach were also found in the cultivation independent approach in high abundance, suggesting a classic infectious pathogenesis of odontogenic origin in these two specific cases. Alterations of the bacterial community

Microbiome and Culture Based Analysis of Chronic Rhinosinusitis Compared to Healthy Sinus Mucosa.

PubMed

Koeller, Kerstin; Herlemann, Daniel P R; Schuldt, Tobias; Ovari, Attila; Guder, Ellen; Podbielski, Andreas; Kreikemeyer, Bernd; Olzowy, Bernhard

2018-01-01

The role of bacteria in chronic rhinosinusitis (CRS) is still not well understood. Whole microbiome analysis adds new aspects to our current understanding that is mainly based on isolated bacteria. It is still unclear how the results of microbiome analysis and the classical culture based approaches interrelate. To address this, middle meatus swabs and tissue samples were obtained during sinus surgery in 5 patients with CRS with nasal polyps (CRSwNP), 5 patients with diffuse CRS without nasal polyps (CRSsNP), 5 patients with unilateral purulent maxillary CRS (upm CRS) and 3 patients with healthy sinus mucosa. Swabs were cultured, and associated bacteria were identified. Additionally, parts of each tissue sample also underwent culture approaches, and in parallel DNA was extracted for 16S rRNA gene amplicon-based microbiome analysis. From tissue samples 4.2 ± 1.2 distinct species per patient were cultured, from swabs 5.4 ± 1.6. The most frequently cultured species from the swabs were Propionibacterium acnes, Staphylococcus epidermidis, Corynebacterium spp. and Staphylococcus aureus . The 16S-RNA gene analysis revealed no clear differentiation of the bacterial community of healthy compared to CRS samples of unilateral purulent maxillary CRS and CRSwNP. However, the bacterial community of CRSsNP differed significantly from the healthy controls. In the CRSsNP samples Flavobacterium, Pseudomonas, Pedobacter, Porphyromonas, Stenotrophomonas , and Brevundimonas were significantly enriched compared to the healthy controls. Species isolated from culture did not generally correspond with the most abundant genera in microbiome analysis. Only Fusobacteria, Parvimonas , and Prevotella found in 2 unilateral purulent maxillary CRS samples by the cultivation dependent approach were also found in the cultivation independent approach in high abundance, suggesting a classic infectious pathogenesis of odontogenic origin in these two specific cases. Alterations of the bacterial

Evaluation of different selective media and culturing techniques for the quantification of Campylobacter ssp. from broiler litter.

PubMed

Kiess, A S; Parker, H M; McDaniel, C D

2010-08-01

Poultry is a major reservoir for Campylobacter, the leading cause of foodborne illness in the United States, but how broilers become initially colonized is still under debate. Broiler litter is a potential source, but the best technique for quantifying Campylobacter from litter is still unknown. Therefore, our objectives were to determine if certain media are more selective for quantifying Campylobacter and if enrichment allows for the detection of stressed or viable but nonculturable cells from broiler litter samples. In this trial, 5 media and 2 culturing techniques were used to enumerate Campylobacter from broiler litter. The media used were campy-Line agar (CLA), campy-cefex agar (CCA), modified CCA, Campylobacter agar plates (CAP), and modified charcoal cefoperazone deoxycholate agar. Litter samples were obtained from a commercial broiler house. Each sample was equally divided and diluted 10-fold into peptone, for direct plating, or 4-fold into Campylobacter enrichment broth. Samples diluted in peptone were direct-plated onto each media and incubated under microaerophilic conditions for 48 h at 42 degrees C. Samples diluted in enrichment broth were incubated under the same conditions for 24 h, then further diluted to 10-fold before plating. Plates from enriched samples were incubated for an additional 24 h after plating. After incubation, all plates (direct and enriched) were counted and presumptive positive colonies were confirmed using a Campylobacter latex agglutination kit. Results indicated that there was no difference in the ability of any of the selective media tested to grow Campylobacter. Direct-plated samples had a higher Campylobacter isolation rate compared with enriched samples. The CLA and CAP were able to suppress total bacterial growth better than modified charcoal cefoperazone deoxycholate, modified CCA, and CCA. The CLA and CAP were the only media able to detect total bacterial population shifts over time. In conclusion, it is important

Phylogenetic and functional diversity within toluene-degrading, sulphate-reducing consortia enriched from a contaminated aquifer.

PubMed

Kuppardt, Anke; Kleinsteuber, Sabine; Vogt, Carsten; Lüders, Tillmann; Harms, Hauke; Chatzinotas, Antonis

2014-08-01

Three toluene-degrading microbial consortia were enriched under sulphate-reducing conditions from different zones of a benzene, toluene, ethylbenzene and xylenes (BTEX) plume of two connected contaminated aquifers. Two cultures were obtained from a weakly contaminated zone of the lower aquifer, while one culture originated from the highly contaminated upper aquifer. We hypothesised that the different habitat characteristics are reflected by distinct degrader populations. Degradation of toluene with concomitant production of sulphide was demonstrated in laboratory microcosms and the enrichment cultures were phylogenetically characterised. The benzylsuccinate synthase alpha-subunit (bssA) marker gene, encoding the enzyme initiating anaerobic toluene degradation, was targeted to characterise the catabolic diversity within the enrichment cultures. It was shown that the hydrogeochemical parameters in the different zones of the plume determined the microbial composition of the enrichment cultures. Both enrichment cultures from the weakly contaminated zone were of a very similar composition, dominated by Deltaproteobacteria with the Desulfobulbaceae (a Desulfopila-related phylotype) as key players. Two different bssA sequence types were found, which were both affiliated to genes from sulphate-reducing Deltaproteobacteria. In contrast, the enrichment culture from the highly contaminated zone was dominated by Clostridia with a Desulfosporosinus-related phylotype as presumed key player. A distinct bssA sequence type with high similarity to other recently detected sequences from clostridial toluene degraders was dominant in this culture. This work contributes to our understanding of the niche partitioning between degrader populations in distinct compartments of BTEX-contaminated aquifers.

A comparative study on sealing ability of mineral trioxide aggregate, calcium enriched cement and bone cement in furcal perforations.

PubMed

Nazari Moghadam, K; Aghili, H; Rashed Mohassel, A; Zahedpasha, S; Moghadamnia, A A

2014-06-01

The aim of this study was to compare the bacterial leakage of mineral trioxide aggregate (MTA), calcium enriched cement (CEM), and bone cement (BC) as repair materials in furcal perforations. The pulp chambers of 57 human mandibular molar teeth were accessed and the root canal orifices were located. The roots were horizontally sectioned in the middle third. Composite resin was used to fill the root canal orifices and the apical end of the roots. The 1 mm furcation perforations were performed in the center of the pulp chamber floor, using diamond fissure burs. Fifty one teeth were divided into 3 groups. Six teeth were used as controls. Perforation defects were repaired with either MTA, CEM, or BC. A bacterial leakage model utilizing phenol red with 3% lactose broth was used for evaluation. The upper pulp chambers were subsequently filled with 5μL bacterial suspension containing Enterococcus faecalis. Then the top of the assembly was covered with aluminum foil to avoid unintentional contamination. The entire apparatus was incubated at 37°C, and bacterial leakage was evaluated daily by checking the turbidity in the culture medium of the lower part of the chamber. The bacterial inoculation was renewed every day, for 30 days. Leakage was noted when color conversion of the culture media was observed and was statistically analyzed using the Chi-square test with significance set at P< 0.05. Sixteen (94%) of the 17 samples of the MTA group, thirteen (81%) of the 17 samples of the CEM group and sixteen (94%) of the 17 samples in BC group were fully contaminated at 30 days. There was no statistically significant difference between the three study groups (P>0.05). According to the present study, in teeth with furcation perforations, the coronal seal produced by MTA preparations was equally to that produced by CEM cement and Bone cement.

Coupling Bacterial Activity Measurements with Cell Sorting by Flow Cytometry.

PubMed

Servais; Courties; Lebaron; Troussellier

1999-08-01

> Abstract A new procedure to investigate the relationship between bacterial cell size and activity at the cellular level has been developed; it is based on the coupling of radioactive labeling of bacterial cells and cell sorting by flow cytometry after SYTO 13 staining. Before sorting, bacterial cells were incubated in the presence of tritiated leucine using a procedure similar to that used for measuring bacterial production by leucine incorporation and then stained with SYTO 13. Subpopulations of bacterial cells were sorted according to their average right-angle light scatter (RALS) and fluorescence. Average RALS was shown to be significantly related to the average biovolume. Experiments were performed on samples collected at different times in a Mediterranean seawater mesocosm enriched with nitrogen and phosphorus. At four sampling times, bacteria were sorted in two subpopulations (cells smaller and larger than 0.25 µm(3)). The results indicate that, at each sampling time, the growth rate of larger cells was higher than that of smaller cells. In order to confirm this tendency, cell sorting was performed on six subpopulations differing in average biovolume during the mesocosm follow-up. A clear increase of the bacterial growth rates was observed with increasing cell size for the conditions met in this enriched mesocosm.http://link.springer-ny.com/link/service/journals/00248/bibs/38n2p180.html

Bacterial Sepsis in Patients with Visceral Leishmaniasis in Northwest Ethiopia

PubMed Central

Takele, Yegnasew; Woldeyohannes, Desalegn; Tiruneh, Moges; Mohammed, Rezika; Lynen, Lutgarde; van Griensven, Johan

2014-01-01

Background and Objectives. Visceral leishmaniasis (VL) is one of the neglected diseases affecting the poorest segment of world populations. Sepsis is one of the predictors for death of patients with VL. This study aimed to assess the prevalence and factors associated with bacterial sepsis, causative agents, and their antimicrobial susceptibility patterns among patients with VL. Methods. A cross-sectional study was conducted among parasitologically confirmed VL patients suspected of sepsis admitted to the University of Gondar Hospital, Northwest Ethiopia, from February 2012 to May 2012. Blood cultures and other clinical samples were collected and cultured following the standard procedures. Results. Among 83 sepsis suspected VL patients 16 (19.3%) had culture confirmed bacterial sepsis. The most frequently isolated organism was Staphylococcus aureus (68.8%; 11/16), including two methicillin-resistant isolates (MRSA). Patients with focal bacterial infection were more likely to have bacterial sepsis (P < 0.001). Conclusions. The prevalence of culture confirmed bacterial sepsis was high, predominantly due to S. aureus. Concurrent focal bacterial infection was associated with bacterial sepsis, suggesting that focal infections could serve as sources for bacterial sepsis among VL patients. Careful clinical evaluation for focal infections and prompt initiation of empiric antibiotic treatment appears warranted in VL patients. PMID:24895569

Long-Term Nitrogen Amendment Alters the Diversity and Assemblage of Soil Bacterial Communities in Tallgrass Prairie

PubMed Central

Todd, Timothy C.; Blair, John M.; Herman, Michael A.

2013-01-01

Anthropogenic changes are altering the environmental conditions and the biota of ecosystems worldwide. In many temperate grasslands, such as North American tallgrass prairie, these changes include alteration in historically important disturbance regimes (e.g., frequency of fires) and enhanced availability of potentially limiting nutrients, particularly nitrogen. Such anthropogenically-driven changes in the environment are known to elicit substantial changes in plant and consumer communities aboveground, but much less is known about their effects on soil microbial communities. Due to the high diversity of soil microbes and methodological challenges associated with assessing microbial community composition, relatively few studies have addressed specific taxonomic changes underlying microbial community-level responses to different fire regimes or nutrient amendments in tallgrass prairie. We used deep sequencing of the V3 region of the 16S rRNA gene to explore the effects of contrasting fire regimes and nutrient enrichment on soil bacterial communities in a long-term (20 yrs) experiment in native tallgrass prairie in the eastern Central Plains. We focused on responses to nutrient amendments coupled with two extreme fire regimes (annual prescribed spring burning and complete fire exclusion). The dominant bacterial phyla identified were Proteobacteria, Verrucomicrobia, Bacteriodetes, Acidobacteria, Firmicutes, and Actinobacteria and made up 80% of all taxa quantified. Chronic nitrogen enrichment significantly impacted bacterial community diversity and community structure varied according to nitrogen treatment, but not phosphorus enrichment or fire regime. We also found significant responses of individual bacterial groups including Nitrospira and Gammaproteobacteria to long-term nitrogen enrichment. Our results show that soil nitrogen enrichment can significantly alter bacterial community diversity, structure, and individual taxa abundance, which have important

Influence of oyster culture on biogeochemistry and bacterial community structure at the sediment-water interface.

PubMed

Azandégbé, Afi; Poly, Franck; Andrieux-Loyer, Françoise; Kérouel, Roger; Philippon, Xavier; Nicolas, Jean-Louis

2012-10-01

Bacterial community structure and some biogeochemical parameters were studied in the sediment of two Pacific oyster farming sites, Aber Benoît (AB) and Rivière d'Auray (RA) in Brittany (France), to examine the ecological impact of oysters and to evaluate the emission of sulfide and ammonia from sediment. At AB, the organic matter accumulated in the sediment beneath the oyster tables was rapidly mineralized, with strong fluxes of ammonia and sulfide that reached 1014 and 215 μmol m(-2) h(-1), respectively, in June 2007. At RA, the fluxes were about half as strong on average and better distributed through the year. The ammonia and sulfide concentrations in the overlying water never reached levels that would be toxic to oysters in either site, nor did hypoxia occur. Total culturable bacteria (TCB) varied greatly according to the temperature: from 1.6 × 10(4) to 9.4 × 10(7) cell g(-1) sediment. Inversely, the bacterial community structure remained surprising stable through the seasons, marginally influenced by the presence of oysters and by temperature. Bacterial communities appeared to be characteristic of the sites, with only one common phylotype, Vibrio aestuarianus, a potential oyster pathogen. These data refine the hypothesis of seawater toxicity to oysters because of ammonia and sulfide fluxes and show that the measured environmental factors had only a weak influence on bacterial community structure. © 2012 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

Bacterial Disproportionation of Elemental Sulfur Coupled to Chemical Reduction of Iron or Manganese

PubMed Central

Thamdrup, Bo; Finster, Kai; Hansen, Jens Würgler; Bak, Friedhelm

1993-01-01

A new chemolithotrophic bacterial metabolism was discovered in anaerobic marine enrichment cultures. Cultures in defined medium with elemental sulfur (S0) and amorphous ferric hydroxide (FeOOH) as sole substrates showed intense formation of sulfate. Furthermore, precipitation of ferrous sulfide and pyrite was observed. The transformations were accompanied by growth of slightly curved, rod-shaped bacteria. The quantification of the products revealed that S0 was microbially disproportionated to sulfate and sulfide, as follows: 4S0 + 4H2O → SO42- + 3H2S + 2H+. Subsequent chemical reactions between the formed sulfide and the added FeOOH led to the observed precipitation of iron sulfides. Sulfate and iron sulfides were also produced when FeOOH was replaced by FeCO3. Further enrichment with manganese oxide, MnO2, instead of FeOOH yielded stable cultures which formed sulfate during concomitant reduction of MnO2 to Mn2+. Growth of small rod-shaped bacteria was observed. When incubated without MnO2, the culture did not grow but produced small amounts of SO42- and H2S at a ratio of 1:3, indicating again a disproportionation of S0. The observed microbial disproportionation of S0 only proceeds significantly in the presence of sulfide-scavenging agents such as iron and manganese compounds. The population density of bacteria capable of S0 disproportionation in the presence of FeOOH or MnO2 was high, > 104 cm-3 in coastal sediments. The metabolism offers an explanation for recent observations of anaerobic sulfide oxidation to sulfate in anoxic sediments. PMID:16348835

Bacterial Community Dynamics during Start-Up of a Trickle-Bed Bioreactor Degrading Aromatic Compounds

PubMed Central

Stoffels, Marion; Amann, Rudolf; Ludwig, Wolfgang; Hekmat, Dariusch; Schleifer, Karl-Heinz

1998-01-01

This study was performed with a laboratory-scale fixed-bed bioreactor degrading a mixture of aromatic compounds (Solvesso100). The starter culture for the bioreactor was prepared in a fermentor with a wastewater sample of a car painting facility as the inoculum and Solvesso100 as the sole carbon source. The bacterial community dynamics in the fermentor and the bioreactor were examined by a conventional isolation procedure and in situ hybridization with fluorescently labeled rRNA-targeted oligonucleotides. Two significant shifts in the bacterial community structure could be demonstrated. The original inoculum from the wastewater of the car factory was rich in proteobacteria of the alpha and beta subclasses, while the final fermentor enrichment was dominated by bacteria closely related to Pseudomonas putida or Pseudomonas mendocina, which both belong to the gamma subclass of the class Proteobacteria. A second significant shift was observed when the fermentor culture was transferred as inoculum to the trickle-bed bioreactor. The community structure in the bioreactor gradually returned to a higher complexity, with the dominance of beta and alpha subclass proteobacteria, whereas the gamma subclass proteobacteria sharply declined. Obviously, the preceded pollutant adaptant did not lead to a significant enrichment of bacteria that finally dominated in the trickle-bed bioreactor. In the course of experiments, three new 16S as well as 23S rRNA-targeted probes for beta subclass proteobacteria were designed, probe SUBU1237 for the genera Burkholderia and Sutterella, probe ALBO34a for the genera Alcaligenes and Bordetella, and probe Bcv13b for Burkholderia cepacia and Burkholderia vietnamiensis. Bacteria hybridizing with the probe Bcv13b represented the main Solvesso100-degrading population in the reactor. PMID:9501433

Enrichment of Thermophilic Ammonia-Oxidizing Archaea from an Alkaline Hot Spring in the Great Basin, USA

NASA Astrophysics Data System (ADS)

Zhang, C.; Huang, Z.; Jiang, H.; Wiegel, J.; Li, W.; Dong, H.

2010-12-01

One of the major advances in the nitrogen cycle is the recent discovery of ammonia oxidation by archaea. While culture-independent studies have revealed occurrence of ammonia-oxidizing archaea (AOA) in nearly every surface niche on earth, most of these microorganisms have resisted isolation and so far only a few species have been identified. The Great Basin contains numerous hot springs, which are characterized by moderately high temperature (40-65 degree C) and circumneutral or alkaline pH. Unique thermophilic archaea have been identified based on molecular DNA and lipid biomarkers; some of which may be ammonia oxidizers. This study aims to isolate some of these archaea from a California hot spring that has pH around 9.0 and temperature around 42 degree C. Mat material was collected from the spring and transported on ice to the laboratory. A synthetic medium (SCM-5) was inoculated with the mat material and the culture was incubated under varying temperature (35-65 degree C) and pH (7.0-10.0) conditions using antibiotics to suppress bacterial growth. Growth of the culture was monitored by microscopy, decrease in ammonium and increase in nitrite, and increases in Crenarchaeota and AOA abundances over time. Clone libraries were constructed to compare archaeal community structures before and after the enrichment experiment. Temperature and pH profiles indicated that the culture grew optimally at pH 9.0 and temperature 45 degree C, which are consistent with the geochemical conditions of the natural environment. Phylogenetic analysis showed that the final OTU was distantly related to all known hyperthermophilic archaea. Analysis of the amoA genes showed two OTUs in the final culture; one of them was closely related to Candidatus Nitrososphaera gargensis. However, the enrichment culture always contained bacteria and attempts to separate them from archaea have failed. This highlights the difficulty in bringing AOA into pure culture and suggests that some of the AOA may

Municipal wastewater treatment and biomass accumulation with a wastewater-born and settleable algal-bacterial culture.

PubMed

Su, Yanyan; Mennerich, Artur; Urban, Brigitte

2011-05-01

A wastewater-born and settleable algal-bacterial culture, cultivated in a stirred tank photobioreactor under lab conditions, was used to remove the carbon and nutrients in municipal wastewater and accumulate biomass simultaneously. The algal-bacterial culture showed good settleable property and could totally settle down over 20 min, resulting in a reduction of total suspended solids from an initial 1.84 to 0.016 g/l. The average removal efficiencies of chemical oxygen demand, total kjeldahl nitrogen and phosphate were 98.2 ± 1.3%, 88.3 ± 1.6% and 64.8 ± 1.0% within 8 days, respectively, while the average biomass productivity was 10.9 ± 1.1 g/m(2) · d. Accumulation into biomass, identified as the main nitrogen and phosphorus removal mechanism, accounted for 44.9 ± 0.4% and 61.6 ± 0.5% of total inlet nitrogen and phosphorus, respectively. Microscopic analysis showed the main algae species in the bioreactor were filamentous blue-green algae. Furthermore, denaturing gradient gel electrophoresis and 16S rDNA gene sequencing revealed that the main bacteria present in the photobioreactor were consortia with sequences similar to those of Flavobacteria, Gammaproteobacteria, Bacteroidia and Betaproteobacteria. This study explores a better understanding of an algae-bacteria system and offers new information on further usage of biomass accumulated during treatment. Copyright © 2011 Elsevier Ltd. All rights reserved.

Drought and host selection influence bacterial community dynamics in the grass root microbiome

PubMed Central

Naylor, Dan; DeGraaf, Stephanie; Purdom, Elizabeth; Coleman-Derr, Devin

2017-01-01

Root endophytes have been shown to have important roles in determining host fitness under periods of drought stress, and yet the effect of drought on the broader root endosphere bacterial community remains largely uncharacterized. In this study, we present phylogenetic profiles of bacterial communities associated with drought-treated root and rhizosphere tissues of 18 species of plants with varying degrees of drought tolerance belonging to the Poaceae family, including important crop plants. Through 16S rRNA gene profiling across two distinct watering regimes and two developmental time points, we demonstrate that there is a strong correlation between host phylogenetic distance and the microbiome dissimilarity within root tissues, and that drought weakens this correlation by inducing conserved shifts in bacterial community composition. We identify a significant enrichment in a wide variety of Actinobacteria during drought within the roots of all hosts, and demonstrate that this enrichment is higher within the root than it is in the surrounding environments. Furthermore, we show that this observed enrichment is the result of an absolute increase in Actinobacterial abundance and that previously hypothesized mechanisms for observed enrichments in Actinobacteria in drought-treated soils are unlikely to fully account for the phenomena observed here within the plant root. PMID:28753209

Drought and host selection influence bacterial community dynamics in the grass root microbiome.

PubMed

Naylor, Dan; DeGraaf, Stephanie; Purdom, Elizabeth; Coleman-Derr, Devin

2017-12-01

Root endophytes have been shown to have important roles in determining host fitness under periods of drought stress, and yet the effect of drought on the broader root endosphere bacterial community remains largely uncharacterized. In this study, we present phylogenetic profiles of bacterial communities associated with drought-treated root and rhizosphere tissues of 18 species of plants with varying degrees of drought tolerance belonging to the Poaceae family, including important crop plants. Through 16S rRNA gene profiling across two distinct watering regimes and two developmental time points, we demonstrate that there is a strong correlation between host phylogenetic distance and the microbiome dissimilarity within root tissues, and that drought weakens this correlation by inducing conserved shifts in bacterial community composition. We identify a significant enrichment in a wide variety of Actinobacteria during drought within the roots of all hosts, and demonstrate that this enrichment is higher within the root than it is in the surrounding environments. Furthermore, we show that this observed enrichment is the result of an absolute increase in Actinobacterial abundance and that previously hypothesized mechanisms for observed enrichments in Actinobacteria in drought-treated soils are unlikely to fully account for the phenomena observed here within the plant root.

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

Enrichment and Physiological Characterization of a Cold-Adapted Nitrite-Oxidizing Nitrotoga sp. from an Eelgrass Sediment

PubMed Central

Ishii, Kento; Fujitani, Hirotsugu; Soh, Kentaro; Nakagawa, Tatsunori; Takahashi, Reiji

2017-01-01

ABSTRACT Nitrite-oxidizing bacteria (NOB) are responsible for the second step of nitrification in natural and engineered ecosystems. The recently discovered genus Nitrotoga belongs to the Betaproteobacteria and potentially has high environmental importance. Although environmental clones affiliated with Nitrotoga are widely distributed, the limited number of cultivated Nitrotoga spp. results in a poor understanding of their ecophysiological features. In this study, we successfully enriched the nonmarine cold-adapted Nitrotoga sp. strain AM1 from coastal sand in an eelgrass zone and investigated its physiological characteristics. Multistep-enrichment approaches led to an increase in the abundance of AM1 to approximately 80% of the total bacterial population. AM1 was the only detectable NOB in the bacterial community. The 16S rRNA gene sequence of AM1 was 99.6% identical to that of “Candidatus Nitrotoga arctica,” which was enriched from permafrost-affected soil. The highest nitrogen oxidation rate of AM1 was observed at 16°C. The half-saturation constant (Km) and the generation time were determined to be 25 μM NO2− and 54 h, respectively. The nitrite oxidation rate of AM1 was stimulated at concentrations of <30 mM NH4Cl but completely inhibited at 50 mM NH4Cl. AM1 can grow well under specific environmental conditions, such as low temperature and in the presence of a relatively high concentration of free ammonia. These results help improve our comprehension of the functional importance of Nitrotoga. IMPORTANCE Nitrite-oxidizing bacteria (NOB) are key players in the second step of nitrification, which is an important process of the nitrogen cycle. Recent studies have suggested that the organisms of the novel NOB genus Nitrotoga were widely distributed and played a functional role in natural and engineered ecosystems. However, only a few Nitrotoga enrichments have been obtained, and little is known about their ecology and physiology. In this study, we

Enrichment and Physiological Characterization of a Cold-Adapted Nitrite-Oxidizing Nitrotoga sp. from an Eelgrass Sediment.

PubMed

Ishii, Kento; Fujitani, Hirotsugu; Soh, Kentaro; Nakagawa, Tatsunori; Takahashi, Reiji; Tsuneda, Satoshi

2017-07-15

Nitrite-oxidizing bacteria (NOB) are responsible for the second step of nitrification in natural and engineered ecosystems. The recently discovered genus Nitrotoga belongs to the Betaproteobacteria and potentially has high environmental importance. Although environmental clones affiliated with Nitrotoga are widely distributed, the limited number of cultivated Nitrotoga spp. results in a poor understanding of their ecophysiological features. In this study, we successfully enriched the nonmarine cold-adapted Nitrotoga sp. strain AM1 from coastal sand in an eelgrass zone and investigated its physiological characteristics. Multistep-enrichment approaches led to an increase in the abundance of AM1 to approximately 80% of the total bacterial population. AM1 was the only detectable NOB in the bacterial community. The 16S rRNA gene sequence of AM1 was 99.6% identical to that of " Candidatus Nitrotoga arctica," which was enriched from permafrost-affected soil. The highest nitrogen oxidation rate of AM1 was observed at 16°C. The half-saturation constant ( K m ) and the generation time were determined to be 25 μM NO 2 - and 54 h, respectively. The nitrite oxidation rate of AM1 was stimulated at concentrations of <30 mM NH 4 Cl but completely inhibited at 50 mM NH 4 Cl. AM1 can grow well under specific environmental conditions, such as low temperature and in the presence of a relatively high concentration of free ammonia. These results help improve our comprehension of the functional importance of Nitrotoga IMPORTANCE Nitrite-oxidizing bacteria (NOB) are key players in the second step of nitrification, which is an important process of the nitrogen cycle. Recent studies have suggested that the organisms of the novel NOB genus Nitrotoga were widely distributed and played a functional role in natural and engineered ecosystems. However, only a few Nitrotoga enrichments have been obtained, and little is known about their ecology and physiology. In this study, we successfully

Composting-Like Conditions Are More Efficient for Enrichment and Diversity of Organisms Containing Cellulase-Encoding Genes than Submerged Cultures

PubMed Central

Fayolle-Guichard, Françoise; Lombard, Vincent; Hébert, Agnès; Coutinho, Pedro M.; Groppi, Alexis; Barre, Aurélien; Henrissat, Bernard

2016-01-01

Cost-effective biofuel production from lignocellulosic biomass depends on efficient degradation of the plant cell wall. One of the major obstacles for the development of a cost-efficient process is the lack of resistance of currently used fungal enzymes to harsh conditions such as high temperature. Adapted, thermophilic microbial communities provide a huge reservoir of potentially interesting lignocellulose-degrading enzymes for improvement of the cellulose hydrolysis step. In order to identify such enzymes, a leaf and wood chip compost was enriched on a mixture of thermo-chemically pretreated wheat straw, poplar and Miscanthus under thermophile conditions, but in two different set-ups. Unexpectedly, metagenome sequencing revealed that incubation of the lignocellulosic substrate with compost as inoculum in a suspension culture resulted in an impoverishment of putative cellulase- and hemicellulase-encoding genes. However, mimicking composting conditions without liquid phase yielded a high number and diversity of glycoside hydrolase genes and an enrichment of genes encoding cellulose binding domains. These identified genes were most closely related to species from Actinobacteria, which seem to constitute important players of lignocellulose degradation under the applied conditions. The study highlights that subtle changes in an enrichment set-up can have an important impact on composition and functions of the microcosm. Composting-like conditions were found to be the most successful method for enrichment in species with high biomass degrading capacity. PMID:27936240

Identification of Cellulose-Responsive Bacterial and Fungal Communities in Geographically and Edaphically Different Soils by Using Stable Isotope Probing

PubMed Central

Eichorst, Stephanie A.

2012-01-01

Many bacteria and fungi are known to degrade cellulose in culture, but their combined response to cellulose in different soils is unknown. Replicate soil microcosms amended with [13C]cellulose were used to identify bacterial and fungal communities responsive to cellulose in five geographically and edaphically different soils. The diversity and composition of the cellulose-responsive communities were assessed by DNA-stable isotope probing combined with Sanger sequencing of small-subunit and large-subunit rRNA genes for the bacterial and fungal communities, respectively. In each soil, the 13C-enriched, cellulose-responsive communities were of distinct composition compared to the original soil community or 12C-nonenriched communities. The composition of cellulose-responsive taxa, as identified by sequence operational taxonomic unit (OTU) similarity, differed in each soil. When OTUs were grouped at the bacterial order level, we found that members of the Burkholderiales, Caulobacteriales, Rhizobiales, Sphingobacteriales, Xanthomonadales, and the subdivision 1 Acidobacteria were prevalent in the 13C-enriched DNA in at least three of the soils. The cellulose-responsive fungi were identified as members of the Trichocladium, Chaetomium, Dactylaria, and Arthrobotrys genera, along with two novel Ascomycota clusters, unique to one soil. Although similarities were identified in higher-level taxa among some soils, the composition of cellulose-responsive bacteria and fungi was generally unique to a certain soil type, suggesting a strong potential influence of multiple edaphic factors in shaping the community. PMID:22287013

Biodegradation of di-n-butyl phthalate by bacterial consortium LV-1 enriched from river sludge

PubMed Central

Li, Fangfang; Ruan, Xinling; Song, Jian; Lv, Lv; Chai, Liyuan; Yang, Zhihui; Luo, Lin

2017-01-01

A stable bacterial consortium (LV-1) capable of degrading di-n-butyl phthalate (DBP) was enriched from river sludge. Community analysis revealed that the main families of LV-1 are Brucellaceae (62.78%) and Sinobacteraceae (14.83%), and the main genera of LV-1 are Brucella spp. (62.78%) and Sinobacter spp. (14.83%). The optimal pH and temperature for LV-1 to degrade DBP were pH 6.0 and 30°C, respectively. Inoculum size influenced the degradation ratio when the incubation time was < 24 h. The initial concentration of DBP also influenced the degradation rates of DBP by LV-1, and the degradation rates ranged from 69.0–775.0 mg/l/d in the first 24 h. Degradation of DBP was best fitted by first-order kinetics when the initial concentration was < 300 mg/l. In addition, Cd2+, Cr6+, and Zn2+ inhibited DBP degradation by LV-1 at all considered concentrations, but low concentrations of Pb2+, Cu2+, and Mn2+ enhanced DBP degradation. The main intermediates (mono-ethyl phthalate [MEP], mono-butyl phthalate [MBP], and phthalic acid [PA]) were identified in the DBP degradation process, thus a new biochemical pathway of DBP degradation is proposed. Furthermore, LV-1 also degraded other phthalates with shorter ester chains (DMP, DEP, and PA). PMID:28542471

Bacterial Community Profiling of H2/CO2 or Formate-Utilizing Acetogens Enriched from Diverse Ecosystems

NASA Astrophysics Data System (ADS)

Han, R.; Zhang, L.; Fu, B.; Liu, H.

2014-12-01

Synthetic gases are usually generated from either cellulosic agricultural waste combustion or industrial release and could be subsequently transformed into acetate, ethanol, and/or butyrate by homoacetogenic bacteria, which commonly possess reductive acetyl-CoA synthesis pathway. Homoacetogen-based syngas fermentation technology provides an alternative solution to link greenhouse gas emission control and cellulosic solid waste treatment with biofuels production. The objective of our current project is to hunt for homoacetogens with capabilities of highly efficiently converting syngases to chemical solvents. In this study, we evaluated homoacetogens population dynamics during enrichments and pinpointed dominant homoacetogens representing diverse ecosystems enriched by different substrates. We enriched homoacetogens from four different samples including waste activate sludge, freshwater sediment, anaerobic methanogenic sludge, and cow manure using H2/CO2 (4:1) or formate as substrate for homoacetogen enrichment. Along with the formyltetrahydrofolate synthetase (FTHFS) gene (fhs gene)-specific real time qPCR assay and Terminal Restriction Fragment Length Polymorphism (T-RFLP) analysis, 16S rRNA based 454 high-throughput pyrosequencing was applied to reveal the population dynamic and community structure during enrichment from different origins. Enrichment of homoacetogenic populations coincided with accumulations of short chain fatty acids such as acetate and butyrate. 454 high-throughput pyrosequencing revealed Firmicutes and Spirochaetes populations became dominant while the overall microbial diversity decreased after enrichment. The most abundant sequences among the four origins belonged to the following phyla: Firmicutes, Spirochaetes, Proteobacteria, and Bacteroidetes, accounting for 62.1%-99.1% of the total reads. The major putative homoacetogenic species enriched on H2/CO2 or formate belonged to Clostridium spp., Acetobacterium spp., Acetoanaerobium spp

Value of bacterial culture of vaginal swabs in diagnosis of vaginal infections.

PubMed

Nenadić, Dane; Pavlović, Miloš D

2015-06-01

Vaginal and cervical swab culture is still very common procedure in our country's everyday practice whereas simple and rapid diagnostic methods have been very rarely used. The aim of this study was to show that the employment of simple and rapid diagnostic tools [vaginal fluid wet mount microscopy (VFWMM), vaginal pH and potassium hydroxide (KOH) test] offers better assessment of vaginal environment than standard microbiologic culture commonly used in Serbia. This prospective study included 505 asymptomatic pregnant women undergoing VFWMM, test with 10% KOH, determination of vaginal pH and standard culture of cervicovaginal swabs. Combining findings from the procedures was used to make diagnoses of bacterial vaginosis (BV) and vaginitis. In addition, the number of polymorphonuclear leukocytes (PMN) was determined in each sample and analyzed along with other findings. Infections with Candida albicans and Trichomonas vaginalis were confirmed or excluded by microscopic examination. In 36 (6%) patients cervicovaginal swab cultures retrieved several aerobes and facultative anaerobes, whereas in 52 (11%) women Candida albicans was isolated. Based on VFWMM findings and clinical criteria 96 (19%) women had BV, 19 (4%) vaginitis, and 72 (14%) candidiasis. Of 115 women with BV and vaginitis, pH 4.5 was found in 5, and of 390 with normal findings 83 (21%) had vaginal pH 4.5. Elevated numbers of PMN were found in 154 (30%) women--in 83 (54%) of them VFWMM was normal. Specificity and sensitivity of KOH test and vaginal pH determination in defining pathological vaginal flora were 95% and 81%, and 79% and 91%, respectively. Cervicovaginal swab culture is expensive but almost non-informative test in clinical practice. The use of simpler and rapid methods as vaginal fluid wet mount microscopy, KOH test and vaginal pH offers better results in diagnosis, and probably in the treatment and prevention of sequels of vaginal infections.

Comparative usefulness of inflammatory markers to indicate bacterial infection-analyzed according to blood culture results and related clinical factors.

PubMed

Nishikawa, Hirokazu; Shirano, Michinori; Kasamatsu, Yu; Morimura, Ayumi; Iida, Ko; Kishi, Tomomi; Goto, Tetsushi; Okamoto, Saki; Ehara, Eiji

2016-01-01

To assess relationships of inflammatory markers and 2 related clinical factors with blood culture results, we retrospectively investigated inpatients' blood culture and blood chemistry findings that were recorded from January to December 2014 using electronic medical records and analyzed the data of 852 subjects (426 culture-positive and 426 culture-negative). Results suggested that the risk of positive blood culture statistically increased as inflammatory marker levels and the number of related factors increased. Concerning the effectiveness of inflammatory markers, when the outcome definition was also changed for C-reactive protein (CRP), the odds ratio had a similar value, whereas when the outcome definition of blood culture positivity was used for procalcitonin (PCT), the greatest effectiveness of that was detected. Therefore, the current results suggest that PCT is more useful than CRP as an auxiliary indication of bacterial infection. Copyright © 2016 Elsevier Inc. All rights reserved.

[Clinical, epidemiological and microbiological aspects of Mobiluncus sp. in bacterial vaginosis].

PubMed

Menolascina, A; Nieves, B; Velazco, E; Rivero, N; Calderas, Z

1999-05-01

In this paper, our goal was to determine the optimal isolation conditions, biochemical characterization, and preservation of species of the genus Mobiluncus, associated with bacterial vaginosis in patients attending the family planning clinic. Also, we tried to relate its presence with demographic variables and criteria used in the clinical diagnosis of bacterial diagnosis. The specimen from the posterior fornix were collected and transported to the laboratory in a Stuart medium, one at room temperature and the other at 4 degrees C. These samples were inoculated in anaerobic culture media. Of a total of 92 patients studied, 61 (66.3%) were normal, 28 (30.4%) bacterial vaginosis, and 3 (3.3%) had intermediate vaginosis. There was statistically significant relationship only with intrauterine device use (p = 0.00499). The presence of curved rod, using Gram's method, was significantly related with pH (p = 0.00000) positive amines test (p = 0.00000), and the presence of clue cells (p = 0.00000). Mobiluncus was observed in 23 samples (82%), and the majority (15) using RLK agar (cold enrichment technique). With conventional techniques, we identified 12 strains as Mobiluncus curtisii and 3 strains as Mobiluncus mulieris. The strains of Mobiluncus sp. grew better from litmus milk conserved at -30 degrees C. Isolating Mobiluncus sp. is fairly easy, if the right media and the techniques are used.

Degradation of paracetamol by pure bacterial cultures and their microbial consortium.

PubMed

Zhang, Lili; Hu, Jun; Zhu, Runye; Zhou, Qingwei; Chen, Jianmeng

2013-04-01

Three bacterial strains utilizing paracetamol as the sole carbon, nitrogen, and energy source were isolated from a paracetamol-degrading aerobic aggregate, and assigned to species of the genera Stenotrophomonas and Pseudomonas. The Stenotrophomonas species have not included any known paracetamol degraders until now. In batch cultures, the organisms f1, f2, and fg-2 could perform complete degradation of paracetamol at concentrations of 400, 2,500, and 2,000 mg/L or below, respectively. A combination of three microbial strains resulted in significantly improved degradation and mineralization of paracetamol. The co-culture was able to use paracetamol up to concentrations of 4,000 mg/L, and mineralized 87.1 % of the added paracetamol at the initial of 2,000 mg/L. Two key metabolites of the biodegradation pathway of paracetamol, 4-aminophenol, and hydroquinone were detected. Paracetamol was degraded predominantly via 4-aminophenol to hydroquinone with subsequent ring fission, suggesting new pathways for paracetamol-degrading bacteria. The degradation of paracetamol could thus be performed by the single isolates, but is stimulated by a synergistic interaction of the three-member consortium, suggesting a possible complementary interaction among the various isolates. The exact roles of each of the strains in the consortium need to be further elucidated.

Predictive value of bacterial analysis of laparotomy wounds.

PubMed

Minutolo, Mario; Blandino, Giovanna; Arena, Manuel; Licciardello, Alessio; Di Stefano, Biagio; Lanteri, Raffaele; Puleo, Stefano; Licata, Antonio; Minutolo, Vincenzo

2014-01-01

Despite improvements in antibiotic prophylaxis, surgical site infections represent the most common postoperative complication with important clinical consequences for patients. The hypothesis that a bacterial analysis of the surgical wound in the operating room could predict the likelihood of developing a clinical infection, and might allow a tailored and preemptive approach, aimed to reduce the consequences of an infection, seems appealing. We would like to present a prospective study on the predictive value of the bacterial analysis of laparotomy wounds. Seventy eight prospective patients undergoing surgery were included in the study. To evaluate the risk factors associated with increased rate of wound infection, we performed a bacterial analysis of the wound. 48 patients out of 78 (61%) had positive cultures. 23 patients out of 32 patients (72%) who didn't receive antibiotic prophylaxis were positive to the wound culture whereas 25 patients out of 46 patients (54%) grew positive cultures in the group of patients that received antibiotic prophylaxis. None of the 30 patients with negative cultures developed clinical infection. Only 6 patients out of 48 patients who had positive cultures (12.5%) developed wound infection. Clinical infection occurred in 5 patients who had gram-negative contamination of the wound. No clinical infection occurred in patients who had gram-positive contamination. Wound cultures and their positivity are predictive tools to identify the patients that are at risk to develop wound infection. The positive predictive value of the bacterial analysis of the wound was 12.5%. Abdominal surgery, Bacterial analysis, Wound infection.

Microbial oxidation of elemental selenium in soil slurries and bacterial cultures

USGS Publications Warehouse

Dowdle, P.R.; Oremland, R.S.

1998-01-01

The microbial oxidation of elemental selenium [Se(O)] was studied by employing 75Se(O) as a tracer. Live, oxic soil slurries demonstrated a linear production of mostly Se(IV), with the formation of smaller quantities of Se(VI). Production of both Se(IV) and Se(VI) was inhibited by autoclaving, formalin, antibiotics, azide, and 2,4-dinitrophenol, thereby indicating the involvement of microbes. Oxidation of Se(O) in slurries was enhanced by addition of acetate, glucose, or sulfide, which implied involvement of chemoheterotrophs as well as chemoautotrophic thiobacilli. Cultures of Thiobacillus ASN-1, Leptothrix MnB1, and a heterotrophic soil enrichment all oxidized Se(O) with Se(VI) observed as the major product rather than Se(IV). This indicated that microbial oxidation in soils is partly constrained by the adsorption of Se(IV) onto soil surfaces. Rate constants for unamended soil slurry Se(O) oxidation ranged from 0.0009 to 0.0117 day-1 which were 3-4 orders of magnitude lower than those reported for dissimilatory Se(VI) reduction in organic-rich, anoxic sediments.The microbial oxidation of elemental selenium [Se(0)] was studied by employing 75Se(0) as a tracer. Live, oxic soil slurries demonstrated a linear production of mostly Se(IV), with the formation of smaller quantities of Se(VI). Production of both Se(IV) and Se(VI) was inhibited by autoclaving, formalin, antibiotics, azide, and 2,4-dinitrophenol, thereby indicating the involvement of microbes. Oxidation of Se(O) in slurries was enhanced by addition of acetate, glucose, or sulfide, which implied involvement of chemoheterotrophs as well as chemoautotrophic thiobacilli. Cultures of Thiobacillus ASN-1, Leptothrix MnB1, and a heterotrophic soil enrichment all oxidized Se(O) with Se(VI) observed as the major product rather than Se(IV). This indicated that microbial oxidation in soils is partly constrained by the adsorption of Se(IV) onto soil surfaces. Rate constants for unamended soil slurry Se(O) oxidation

Label-free isolation and deposition of single bacterial cells from heterogeneous samples for clonal culturing

NASA Astrophysics Data System (ADS)

Riba, J.; Gleichmann, T.; Zimmermann, S.; Zengerle, R.; Koltay, P.

2016-09-01

The isolation and analysis of single prokaryotic cells down to 1 μm and less in size poses a special challenge and requires micro-engineered devices to handle volumes in the picoliter to nanoliter range. Here, an advanced Single-Cell Printer (SCP) was applied for automated and label-free isolation and deposition of bacterial cells encapsulated in 35 pl droplets by inkjet-like printing. To achieve this, dispenser chips to generate micro droplets have been fabricated with nozzles 20 μm in size. Further, the magnification of the optical system used for cell detection was increased. Redesign of the optical path allows for collision-free addressing of any flat substrate since no compartment protrudes below the nozzle of the dispenser chip anymore. The improved system allows for deterministic isolation of individual bacterial cells. A single-cell printing efficiency of 93% was obtained as shown by printing fluorescent labeled E. coli. A 96-well plate filled with growth medium is inoculated with single bacteria cells on average within about 8 min. Finally, individual bacterial cells from a heterogeneous sample of E. coli and E. faecalis were isolated for clonal culturing directly on agar plates in user-defined array geometry.

Temperature effect on nitrogen removal performance and bacterial community in culture of marine anammox bacteria derived from sea-based waste disposal site.

PubMed

Kawagoshi, Yasunori; Fujisaki, Koichiro; Tomoshige, Yuki; Yamashiro, Kento; Wei, Qiaoyan; Qiao, Yanwei

2012-04-01

Anaerobic ammonium oxidation (anammox) bacteria have been detected in variety of marine environment in recent years, however, there have been only a few studies on their characteristics in the culture. The aim of this study is to reveal the effect of temperature on nitrogen removal ability and bacterial community in a culture of marine anammox bacteria (MAAOB). The MAAOB were cultured from the sediment of a sea-based waste disposal site at the North Port of Osaka Bay in Japan. The maximum nitrogen removal rate (NRR) was observed at 25°C in the MAAOB culture, and it decreased both at below 20°C and over 33°C. The activation energy of the MAAOB culture was calculated to be 54.6 kJ mol(-1) in the 5°C to 30°C range. No significant change in bacterial community according with temperature (5-37°C) was confirmed in the results of polymerase chain reaction and denaturing gradient gel electrophoresis (PCR-DGGE). Meanwhile, a number of bacteria related to the oxidation-reduction reaction of sulfur were confirmed and it is speculated that they involved in the activity of MAAOB and nitrogen removal ability in the culture. Copyright © 2011 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Short-term incubation of positive blood cultures in brain-heart infusion broth accelerates identification of bacteria by matrix-assisted laser desorption/ionization time-of-flight mass-spectrometry.

PubMed

Torres, Ignacio; Gimenez, Estela; Pascual, Tania; Bueno, Felipe; Huntley, Dixie; Martínez, Mireia; Navarro, David

2017-12-01

Fast identification of bacteria directly from positive blood cultures (BCs) by matrix-assisted laser desorption/ionization time-of-flight mass-spectrometry (MALDI-TOF MS) can be achieved either using the MALDI Sepsityper kit (protein extraction method) or after a short-term pre-cultivation step on solid medium. We developed a new method that involves short-term enrichment of positive BCs in brain-heart infusion broth (BHI) prior to MALDI-TOF MS analysis. Eighty-four BCs flagged as positive were included in this study; these were processed in parallel either directly using the MALDI Sepsityper kit or following a short-term culture either in BHI or on Columbia blood agar with 5 % sheep blood (CBA). Bacterial species were successfully identified in 91.6, 89.2 and 65.4 % of cases after pre-cultivation for 4 h in BHI, on CBA, or by using the MALDI Sepsityper kit, respectively. Overall, the mean incubation time to correct identification was shorter when pre-cultures were performed in BHI; the mean time for Gram-negative rods was 78.2 min in BHI and 108.2 min on CBA (P=0.045), and the mean time for Gram-positive cocci was 128.5 min in BHI and 169.6 min on CBA (P=0.013). Short-term enrichment of BCs in BHI accelerates identification of a number of bacterial species by MALDI-TOF MS. Further prospective studies are needed to validate our method and gauge its potential clinical impact on the management of bloodstream bacterial infections.

Short communication: Repeatability of differential goat bulk milk culture and associations with somatic cell count, total bacterial count, and standard plate count.

PubMed

Koop, G; Dik, N; Nielen, M; Lipman, L J A

2010-06-01

The aims of this study were to assess how different bacterial groups in bulk milk are related to bulk milk somatic cell count (SCC), bulk milk total bacterial count (TBC), and bulk milk standard plate count (SPC) and to measure the repeatability of bulk milk culturing. On 53 Dutch dairy goat farms, 3 bulk milk samples were collected at intervals of 2 wk. The samples were cultured for SPC, coliform count, and staphylococcal count and for the presence of Staphylococcus aureus. Furthermore, SCC (Fossomatic 5000, Foss, Hillerød, Denmark) and TBC (BactoScan FC 150, Foss) were measured. Staphylococcal count was correlated to SCC (r=0.40), TBC (r=0.51), and SPC (r=0.53). Coliform count was correlated to TBC (r=0.33), but not to any of the other variables. Staphylococcus aureus did not correlate to SCC. The contribution of the staphylococcal count to the SPC was 31%, whereas the coliform count comprised only 1% of the SPC. The agreement of the repeated measurements was low. This study indicates that staphylococci in goat bulk milk are related to SCC and make a significant contribution to SPC. Because of the high variation in bacterial counts, repeated sampling is necessary to draw valid conclusions from bulk milk culturing. 2010 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Comprehensive Analysis of Bacterial Flora in Postoperative Maxillary Cyst Fluid by 16S rRNA Gene and Culture Methods

PubMed Central

Sano, Naoto; Yamashita, Yoshio; Fukuda, Kazumasa; Taniguchi, Hatsumi; Goto, Masaaki; Miyamoto, Hiroshi

2012-01-01

Intracystic fluid was aseptically collected from 11 patients with postoperative maxillary cyst (POMC), and DNA was extracted from the POMC fluid. Bacterial species were identified by sequencing after cloning of approximately 580 bp of the 16S rRNA gene. Identification of pathogenic bacteria was also performed by culture methods. The phylogenetic identity was determined by sequencing 517–596 bp in each of the 1139 16S rRNA gene clones. A total of 1114 clones were classified while the remaining 25 clones were unclassified. A total of 103 bacterial species belonging to 42 genera were identified in POMC fluid samples by 16S rRNA gene analysis. Species of Prevotella (91%), Neisseria (73%), Fusobacterium (73%), Porphyromonas (73%), and Propionibacterium (73%) were found to be highly prevalent in all patients. Streptococcus mitis (64%), Fusobacterium nucleatum (55%), Propionibacterium acnes (55%), Staphylococcus capitis (55%), and Streptococcus salivarius (55%) were detected in more than 6 of the 11 patients. The results obtained by the culture method were different from those obtained by 16S rRNA gene analysis, but both approaches may be necessary for the identification of pathogens, especially of bacteria that are difficult to detect by culture methods, and the development of rational treatments for patients with POMC. PMID:22685668

In vitro bacterial isolate susceptibility to empirically selected antimicrobials in 111 dogs with bacterial pneumonia.

PubMed

Proulx, Alexandre; Hume, Daniel Z; Drobatz, Kenneth J; Reineke, Erica L

2014-01-01

To determine the proportion of airway bacterial isolates resistant to both empirically selected and recently administered antimicrobials, and to assess the impact of inappropriate initial empiric antimicrobials selection on length of hospital stay and survival to discharge in dogs with bacterial pneumonia. Retrospective study. University veterinary teaching hospital. One hundred and eleven dogs with a clinical diagnosis of bacterial pneumonia that had aerobic bacterial culture and susceptibility testing performed from a tracheal wash sample. None. Overall, 26% (29/111) of the dogs had at least 1 bacterial isolate that was resistant to empirically selected antimicrobials. In dogs with a history of antimicrobial administration within the preceding 4 weeks, a high incidence (57.4%, 31/54) of in vitro bacterial resistance to those antimicrobials was found: 64.7% (11/17) in the community-acquired pneumonia group, 55.2% (16/29) in the aspiration pneumonia group, and 50.0% (4/8) in the other causes of bacterial pneumonia group. No statistically significant association was found between bacterial isolate resistance to empirically selected antimicrobials and length of hospital stay or mortality. The high proportion of in vitro airway bacterial resistance to empiric antimicrobials would suggest that airway sampling for bacterial culture and susceptibility testing may be helpful in guiding antimicrobial therapy and recently administered antimicrobials should be avoided when empirically selecting antimicrobials. Although no relationship was found between inappropriate initial empiric antimicrobial selection and length of hospital stay or mortality, future prospective studies using standardized airway-sampling techniques, treatment modalities, and stratification of disease severity based on objective values, such as arterial blood gas analysis in all dogs with pneumonia, would be needed to determine if a clinical effect of in vitro bacterial resistance to empirically

Rapid and Specific Detection of Salmonella spp. in Animal Feed Samples by PCR after Culture Enrichment

PubMed Central

Löfström, Charlotta; Knutsson, Rickard; Axelsson, Charlotta Engdahl; Rådström, Peter

2004-01-01

A PCR procedure has been developed for routine analysis of viable Salmonella spp. in feed samples. The objective was to develop a simple PCR-compatible enrichment procedure to enable DNA amplification without any sample pretreatment such as DNA extraction or cell lysis. PCR inhibition by 14 different feed samples and natural background flora was circumvented by the use of the DNA polymerase Tth. This DNA polymerase was found to exhibit a high level of resistance to PCR inhibitors present in these feed samples compared to DyNAzyme II, FastStart Taq, Platinum Taq, Pwo, rTth, Taq, and Tfl. The specificity of the Tth assay was confirmed by testing 101 Salmonella and 43 non-Salmonella strains isolated from feed and food samples. A sample preparation method based on culture enrichment in buffered peptone water and DNA amplification with Tth DNA polymerase was developed. The probability of detecting small numbers of salmonellae in feed, in the presence of natural background flora, was accurately determined and found to follow a logistic regression model. From this model, the probability of detecting 1 CFU per 25 g of feed in artificially contaminated soy samples was calculated and found to be 0.81. The PCR protocol was evaluated on 155 naturally contaminated feed samples and compared to an established culture-based method, NMKL-71. Eight percent of the samples were positive by PCR, compared with 3% with the conventional method. The reasons for the differences in sensitivity are discussed. Use of this method in the routine analysis of animal feed samples would improve safety in the food chain. PMID:14711627

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

Environmental proteomics reveals taxonomic and functional changes in an enriched aquatic ecosystem.

PubMed

Northrop, Amanda C; Brooks, Rachel; Ellison, Aaron M; Gotelli, Nicholas J; Ballif, Bryan A

2017-10-01

Aquatic ecosystem enrichment can lead to distinct and irreversible changes to undesirable states. Understanding changes in active microbial community function and composition following organic-matter loading in enriched ecosystems can help identify biomarkers of such state changes. In a field experiment, we enriched replicate aquatic ecosystems in the pitchers of the northern pitcher plant, Sarracenia purpurea . Shotgun metaproteomics using a custom metagenomic database identified proteins, molecular pathways, and contributing microbial taxa that differentiated control ecosystems from those that were enriched. The number of microbial taxa contributing to protein expression was comparable between treatments; however, taxonomic evenness was higher in controls. Functionally active bacterial composition differed significantly among treatments and was more divergent in control pitchers than enriched pitchers. Aerobic and facultative anaerobic bacteria contributed most to identified proteins in control and enriched ecosystems, respectively. The molecular pathways and contributing taxa in enriched pitcher ecosystems were similar to those found in larger enriched aquatic ecosystems and are consistent with microbial processes occurring at the base of detrital food webs. Detectable differences between protein profiles of enriched and control ecosystems suggest that a time series of environmental proteomics data may identify protein biomarkers of impending state changes to enriched states.

Bridging Culture On-Line: Strategies for Teaching Cultural Sensitivity.

ERIC Educational Resources Information Center

Wendler, M. Cecilia; Struthers, Roxanne

2002-01-01

An online cross-cultural health course for nurses sought to provide access to cultural experiences by culturally congruent use of a minority visiting scholar and required participation in cultural enrichment activities. Course and faculty evaluations were designed to be appropriate for the asynchronous environment. (Contains 25 references.) (SK)

Camelina seed supplementation at two dietary fat levels changes ruminal bacterial community composition in a dual-flow continuous culture system

USDA-ARS?s Scientific Manuscript database

This study sought to determine the effects of camelina seed (CS) supplementation at different dietary fat levels on the ruminal bacterial community composition in dairy cows, and how it relates to changes in ruminal fermentation and metabolism in a dual-flow continuous culture system. Diets were ran...

The prophylactic and therapeutic effects of glutamine- and arginine-enriched diets on radiation-induced enteritis in rats.

PubMed

Ersin, S; Tuncyurek, P; Esassolak, M; Alkanat, M; Buke, C; Yilmaz, M; Telefoncu, A; Kose, T

2000-04-01

Recent studies indicated that glutamine and arginine support the mucosal barrier in several ways. This experimental study hypothesized that administration of glutamine- and arginine-enriched diets before abdominal radiation therapy would provide a radioprotective effect on intestinal mucosa, and this would augment the therapeutic effectiveness provided by postirradiation administration. A rat model of radiation enteritis was designed with a single dose of 1100 cGy to the abdomen. Thirty-five rats were randomized into five groups of seven. A 7-day glutamine-enriched diet for Group I and a 7-day arginine-enriched diet for Group II were administered both pre- and postradiation. For Groups III and IV, the same glutamine and arginine diets were given, respectively, postradiation only. Group V was fed a glutamine- and arginine-free diet and was the control group. The rats underwent laparotomy for culture of mesenteric lymph nodes and removal of segments of ileum, jejenum, and colon for microscopic examination. Bacterial translocation was significantly higher in Group V (P < 0.05), while intestinal villus count and villus height were significantly higher in all of the groups fed glutamine and arginine when compared with the control group (P < 0.0001 and P < 0.05, respectively). Both arginine- and glutamine-enriched diets have protective effects on gut mucosa in the postirradiation state; however, pre- and postirradiation administration together does not provide superior protection versus postradiation administration alone. Copyright 2000 Academic Press.

An evaluation of bacterial contamination of barriers used in periapical tissue regeneration: Part 2--Bacterial penetration.

PubMed

Sharma, Priya; Mickel, André K; Chogle, Sami; Sharma, Prem Nath; Han, Yiping W; Jones, Jefferson J

2008-03-01

To compare the relative penetration of Prevotella melaninogenica and Enterococcus faecalis through 3 guided tissue regeneration membranes: Atrisorb, Lambone, and OsseoQuest. It was hypothesized that OsseoQuest would show increased bacterial penetration when compared to Lambone and Atrisorb. Centrifuge tubes containing trypticase soy broth were sealed with circular sections of membranes and placed in test tubes containing culture media. The bacterial penetration was assessed by passage of bacteria from the outer tube culture media to the inner centrifuge tube media through the membrane. After incubation for 4 and 48 hours, the media from the outer and inner tubes were compared for bacterial count. P melaninogenica exhibited 91% penetration for Lambone in 2 days, while OsseoQuest displayed 87% penetration with E faecalis in the same time. Atrisorb displayed a minimal penetration with both bacteria (2%). Atrisorb displayed the least bacterial penetration, which may be attributed to membrane structure, chemical configuration, hydrophobicity, and porosity of tested membranes.

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

The Pattern of Change in the Abundances of Specific Bacterioplankton Groups Is Consistent across Different Nutrient-Enriched Habitats in Crete

PubMed Central

Fodelianakis, Stilianos; Papageorgiou, Nafsika; Pitta, Paraskevi; Kasapidis, Panagiotis; Karakassis, Ioannis

2014-01-01

A common source of disturbance for coastal aquatic habitats is nutrient enrichment through anthropogenic activities. Although the water column bacterioplankton communities in these environments have been characterized in some cases, changes in α-diversity and/or the abundances of specific taxonomic groups across enriched habitats remain unclear. Here, we investigated the bacterial community changes at three different nutrient-enriched and adjacent undisturbed habitats along the north coast of Crete, Greece: a fish farm, a closed bay within a town with low water renewal rates, and a city port where the level of nutrient enrichment and the trophic status of the habitat were different. Even though changes in α-diversity were different at each site, we observed across the sites a common change pattern accounting for most of the community variation for five of the most abundant bacterial groups: a decrease in the abundance of the Pelagibacteraceae and SAR86 and an increase in the abundance of the Alteromonadaceae, Rhodobacteraceae, and Cryomorphaceae in the impacted sites. The abundances of the groups that increased and decreased in the impacted sites were significantly correlated (positively and negatively, respectively) with the total heterotrophic bacterial counts and the concentrations of dissolved organic carbon and/or dissolved nitrogen and chlorophyll α, indicating that the common change pattern was associated with nutrient enrichment. Our results provide an in situ indication concerning the association of specific bacterioplankton groups with nutrient enrichment. These groups could potentially be used as indicators for nutrient enrichment if the pattern is confirmed over a broader spatial and temporal scale by future studies. PMID:24747897

Quantitative chemical biosensing by bacterial chemotaxis in microfluidic chips.

PubMed

Roggo, Clémence; Picioreanu, Cristian; Richard, Xavier; Mazza, Christian; van Lintel, Harald; van der Meer, Jan Roelof

2018-01-01

Whole-cell bacterial bioreporters are proposed as alternatives to chemical analysis of, for example, pollutants in environmental compartments. Commonly based on reporter gene induction, bioreporters produce a detectable signal within 30 min to a few hours after exposure to the chemical target, which is impractical for applications aiming at a fast response. In an attempt to attain faster readout but maintain flexibility of chemical targeting, we explored the concept for quantitative chemical sensing by bacterial chemotaxis. Chemotaxis was quantified from enrichment of cells across a 600 µm-wide chemical gradient stabilized by parallel flow in a microfluidic chip, further supported by transport and chemotaxis steady state and kinetic modelling. As proof-of-concept, we quantified Escherichia coli chemotaxis towards serine, aspartate and methylaspartate as a function of attractant concentration and exposure time. E. coli chemotaxis enrichment increased sharply between 0 and 10 µM serine, before saturating at 100 µM. The chemotaxis accumulation rate was maximal at 10 µM serine, leading to observable cell enrichment within 5 min. The potential application for biosensing of environmental toxicants was investigated by quantifying chemotaxis of Cupriavidus pinatubonensis JMP134 towards the herbicide 2,4-dichlorophenoxyacetate. Our results show that bacterial chemotaxis can be quantified on a scale of minutes and may be used for developing faster bioreporter assays. © 2017 The Authors. Environmental Microbiology published by Society for Applied Microbiology and John Wiley & Sons Ltd.

The effect of boric acid on bacterial culture of canine and feline urine.

PubMed

Rowlands, M; Blackwood, L; Mas, A; Cripps, P; Crompton, C; Burrow, R

2011-10-01

To identify the optimal method of submission of canine and feline urine for bacterial culture. Cystocentesis samples from 250 animals (200 dogs, 50 cats) suspected of having urinary tract infections were collected. The reference aliquot, without preservative, was processed on site within 2 hours. Two further aliquots (one without preservative, one with boric acid) were stored at room temperature for up to 7 hours and then posted by guaranteed next day delivery to a commercial laboratory for analysis. Forty-seven of the samples were positive on culture in the reference test. There was no significant difference between reference test results and those of samples posted without preservative (P=0·39), but samples posted in boric acid were significantly less likely to give a positive result (P=0·01). Samples posted without preservative had a sensitivity of 82% and a specificity of 98%; for boric acid, sensitivity was 73% and specificity 99%. Postal urine samples should be submitted to the laboratory in a plain sterile tube. © 2011 British Small Animal Veterinary Association.

The Unculturables: targeted isolation of bacterial species associated with canine periodontal health or disease from dental plaque.

PubMed

Davis, Ian J; Bull, Christopher; Horsfall, Alexander; Morley, Ian; Harris, Stephen

2014-08-01

The current inability to culture the entirety of observed bacteria is well known and with the advent of ever more powerful molecular tools, that can survey bacterial communities at previously unattainable depth, the gap in our capacity to culture and define all of these species increases exponentially. This gap has essentially become the rate limiting step in determining how the knowledge of which species are present in a sample can be applied to understand the role of these species in an ecosystem or disease process. A case in point is periodontal disease, which is the most widespread oral disease in dogs. If untreated the disease results in significant pain, eventual loss of the dentition and potentially an increased risk of systemic diseases. Previous molecular based studies have identified the bacterial species associated with periodontal disease in dogs; however without cultured strains from many of these species it has not been possible to study whether they play a role in the disease process. Using a quantitative polymerase chain reaction (qPCR) directed approach a range of microbiological media were screened and optimized to enrich for previously uncultivated target species. A systematic screening methodology was then employed to isolate the species of interest. In cases where the target species were not cultivable in isolation, helper strains grown underneath a nitrocellulose membrane were used to provide the necessary growth factors. This guided media optimization approach enabled the purification of 14 species, 8 of which we had previously been unable to cultivate in isolation. It is also applicable to the targeted isolation of isolates from species that have previously been cultured (for example to study intra-species variation) as demonstrated by the successful isolation of 6 targeted isolates of already cultured species. To our knowledge this is the first time this combination of qPCR guided media optimization, strategic screening and helper strain

Characterization of a Highly Enriched Microbial Consortium Reductively Dechlorinating 2,3-Dichlorophenol and 2,4,6-Trichlorophenol and the Corresponding cprA Genes from River Sediment.

PubMed

El-Sayed, Wael S

2016-08-26

Anaerobic reductive dechlorination of 2,3-dichlorophenol (2,3DCP) and 2,4,6-trichlorophenol (2,4,6TCP) was investigated in microcosms from River Nile sediment. A stable sediment-free anaerobic microbial consortium reductively dechlorinating 2,3DCP and 2,4,6TCP was established. Defined sediment-free cultures showing stable dechlorination were restricted to ortho chlorine when enriched with hydrogen as the electron donor, acetate as the carbon source, and either 2,3-DCP or 2,4,6-TCP as electron acceptors. When acetate, formate, or pyruvate were used as electron donors, dechlorination activity was lost. Only lactate can replace dihydrogen as an electron donor. However, the dechlorination potential was decreased after successive transfers. To reveal chlororespiring species, the microbial community structure of chlorophenol-reductive dechlorinating enrichment cultures was analyzed by PCR-denaturing gradient gel electrophoresis (DGGE) of 16S rRNA gene fragments. Eight dominant bacteria were detected in the dechlorinating microcosms including members of the genera Citrobacter, Geobacter, Pseudomonas, Desulfitobacterium, Desulfovibrio and Clostridium. Highly enriched dechlorinating cultures were dominated by four bacterial species belonging to the genera Pseudomonas, Desulfitobacterium, and Clostridium. Desulfitobacterium represented the major fraction in DGGE profiles indicating its importance in dechlorination activity, which was further confirmed by its absence resulting in complete loss of dechlorination. Reductive dechlorination was confirmed by the stoichiometric dechlorination of 2,3DCP and 2,4,6TCP to metabolites with less chloride groups and by the detection of chlorophenol RD cprA gene fragments in dechlorinating cultures. PCR amplified cprA gene fragments were cloned and sequenced and found to cluster with the cprA/pceA type genes of Dehalobacter restrictus.

Limitation of Bacterial Growth by Dissolved Organic Matter and Iron in the Southern Ocean†

PubMed Central

Church, Matthew J.; Hutchins, David A.; Ducklow, Hugh W.

2000-01-01

The importance of resource limitation in controlling bacterial growth in the high-nutrient, low-chlorophyll (HNLC) region of the Southern Ocean was experimentally determined during February and March 1998. Organic- and inorganic-nutrient enrichment experiments were performed between 42°S and 55°S along 141°E. Bacterial abundance, mean cell volume, and [3H]thymidine and [3H]leucine incorporation were measured during 4- to 5-day incubations. Bacterial biomass, production, and rates of growth all responded to organic enrichments in three of the four experiments. These results indicate that bacterial growth was constrained primarily by the availability of dissolved organic matter. Bacterial growth in the subtropical front, subantarctic zone, and subantarctic front responded most favorably to additions of dissolved free amino acids or glucose plus ammonium. Bacterial growth in these regions may be limited by input of both organic matter and reduced nitrogen. Unlike similar experimental results in other HNLC regions (subarctic and equatorial Pacific), growth stimulation of bacteria in the Southern Ocean resulted in significant biomass accumulation, apparently by stimulating bacterial growth in excess of removal processes. Bacterial growth was relatively unchanged by additions of iron alone; however, additions of glucose plus iron resulted in substantial increases in rates of bacterial growth and biomass accumulation. These results imply that bacterial growth efficiency and nitrogen utilization may be partly constrained by iron availability in the HNLC Southern Ocean. PMID:10653704

Differential resistance of drinking water bacterial populations to monochloramine disinfection.

PubMed

Chiao, Tzu-Hsin; Clancy, Tara M; Pinto, Ameet; Xi, Chuanwu; Raskin, Lutgarde

2014-04-01

The impact of monochloramine disinfection on the complex bacterial community structure in drinking water systems was investigated using culture-dependent and culture-independent methods. Changes in viable bacterial diversity were monitored using culture-independent methods that distinguish between live and dead cells based on membrane integrity, providing a highly conservative measure of viability. Samples were collected from lab-scale and full-scale drinking water filters exposed to monochloramine for a range of contact times. Culture-independent detection of live cells was based on propidium monoazide (PMA) treatment to selectively remove DNA from membrane-compromised cells. Quantitative PCR (qPCR) and pyrosequencing of 16S rRNA genes was used to quantify the DNA of live bacteria and characterize the bacterial communities, respectively. The inactivation rate determined by the culture-independent PMA-qPCR method (1.5-log removal at 664 mg·min/L) was lower than the inactivation rate measured by the culture-based methods (4-log removal at 66 mg·min/L). Moreover, drastic changes in the live bacterial community structure were detected during monochloramine disinfection using PMA-pyrosequencing, while the community structure appeared to remain stable when pyrosequencing was performed on samples that were not subject to PMA treatment. Genera that increased in relative abundance during monochloramine treatment include Legionella, Escherichia, and Geobacter in the lab-scale system and Mycobacterium, Sphingomonas, and Coxiella in the full-scale system. These results demonstrate that bacterial populations in drinking water exhibit differential resistance to monochloramine, and that the disinfection process selects for resistant bacterial populations.

Sensitivity of Direct Culture, Enrichment and PCR for Detection of Campylobacter jejuni and C. coli in Broiler Flocks at Slaughter.

PubMed

Rodgers, J D; Simpkin, E; Lee, R; Clifton-Hadley, F A; Vidal, A B

2017-06-01

Broiler chicken flocks are a significant source of Campylobacter jejuni and Campylobacter coli that result in the major public health problem of campylobacteriosis. Accurate estimates of the prevalence of both C. coli and C. jejuni in flocks would enhance epidemiological understanding, risk assessment and control options. This study combined results from a panel of 10 detection tests (direct culture, enrichment and PCR) on caecal samples from flocks at slaughter. A parallel interpretation approach was used to determine the presence of Campylobacter spp. and for C. jejuni and C. coli individually. The sample was considered positive if at least one method detected the target and this interpretation was taken to represent a 'proxy gold standard' for detection in the absence of a gold standard reference test. The sensitivity of each individual method to detect Campylobacter spp., C. jejuni and C. coli was then estimated relative to the proxy gold standard. Enrichment in adapted Exeter broth (deficient in polymyxin B) with a resuscitation step was 100% sensitive, whilst direct culture on modified charcoal cefoperazone deoxycholate agar (mCCDA) was highly sensitive (97.9%). Enrichment methods using Preston broth and Bolton broth were significantly less sensitive. Enrichment in Exeter broth promoted the recovery of C. jejuni, whilst enrichment in Bolton broth favoured C. coli. A RT-PCR detection test could identify 80% of flocks that were co-colonised with both species. This study found that 76.3% (n = 127) of flocks were colonised with Campylobacter spp. The majority (95.9%) of Campylobacter-positive flocks were colonised with C. jejuni; however, approximately one-third of positive flocks were simultaneously colonised with both C. jejuni and C. coli. The findings highlight the impact of different detection methodologies on the accuracy of the estimated incidence of both C. jejuni and C. coli entering the abattoir within broiler flocks and the associated

Profiling bacterial communities associated with sediment-based aquaculture bioremediation systems under contrasting redox regimes

NASA Astrophysics Data System (ADS)

Robinson, Georgina; Caldwell, Gary S.; Wade, Matthew J.; Free, Andrew; Jones, Clifford L. W.; Stead, Selina M.

2016-12-01

Deposit-feeding invertebrates are proposed bioremediators in microbial-driven sediment-based aquaculture effluent treatment systems. We elucidate the role of the sediment reduction-oxidation (redox) regime in structuring benthic bacterial communities, having direct implications for bioremediation potential and deposit-feeder nutrition. The sea cucumber Holothuria scabra was cultured on sediments under contrasting redox regimes; fully oxygenated (oxic) and redox stratified (oxic-anoxic). Taxonomically, metabolically and functionally distinct bacterial communities developed between the redox treatments with the oxic treatment supporting the greater diversity; redox regime and dissolved oxygen levels were the main environmental drivers. Oxic sediments were colonised by nitrifying bacteria with the potential to remediate nitrogenous wastes. Percolation of oxygenated water prevented the proliferation of anaerobic sulphate-reducing bacteria, which were prevalent in the oxic-anoxic sediments. At the predictive functional level, bacteria within the oxic treatment were enriched with genes associated with xenobiotics metabolism. Oxic sediments showed the greater bioremediation potential; however, the oxic-anoxic sediments supported a greater sea cucumber biomass. Overall, the results indicate that bacterial communities present in fully oxic sediments may enhance the metabolic capacity and bioremediation potential of deposit-feeder microbial systems. This study highlights the benefits of incorporating deposit-feeding invertebrates into effluent treatment systems, particularly when the sediment is oxygenated.

Profiling bacterial communities associated with sediment-based aquaculture bioremediation systems under contrasting redox regimes

PubMed Central

Robinson, Georgina; Caldwell, Gary S.; Wade, Matthew J.; Free, Andrew; Jones, Clifford L. W.; Stead, Selina M.

2016-01-01

Deposit-feeding invertebrates are proposed bioremediators in microbial-driven sediment-based aquaculture effluent treatment systems. We elucidate the role of the sediment reduction-oxidation (redox) regime in structuring benthic bacterial communities, having direct implications for bioremediation potential and deposit-feeder nutrition. The sea cucumber Holothuria scabra was cultured on sediments under contrasting redox regimes; fully oxygenated (oxic) and redox stratified (oxic-anoxic). Taxonomically, metabolically and functionally distinct bacterial communities developed between the redox treatments with the oxic treatment supporting the greater diversity; redox regime and dissolved oxygen levels were the main environmental drivers. Oxic sediments were colonised by nitrifying bacteria with the potential to remediate nitrogenous wastes. Percolation of oxygenated water prevented the proliferation of anaerobic sulphate-reducing bacteria, which were prevalent in the oxic-anoxic sediments. At the predictive functional level, bacteria within the oxic treatment were enriched with genes associated with xenobiotics metabolism. Oxic sediments showed the greater bioremediation potential; however, the oxic-anoxic sediments supported a greater sea cucumber biomass. Overall, the results indicate that bacterial communities present in fully oxic sediments may enhance the metabolic capacity and bioremediation potential of deposit-feeder microbial systems. This study highlights the benefits of incorporating deposit-feeding invertebrates into effluent treatment systems, particularly when the sediment is oxygenated. PMID:27941918

Mastitis diagnosis in dairy cows using PathoProof real-time polymerase chain reaction assay in comparison with conventional bacterial culture in a Northern German field study.

PubMed

Spittel, Susanne; Hoedemaker, Martina

2012-01-01

In the following field study, the commercial PathoProof Mastitis PCR Assay, a real-time PCR for identifying eleven mastitis pathogens and the staphylococcal beta-lactamase gene, was compared with conventional bacterial culture. For this purpose, 681 udder quarter samples from 173 clinically healthy cows with varying somatic cell count from four dairy herds in the region of Osnabrück, Lower Saxony, Germany, were collected between July 2010 and February 2011 and subjected to PCR and bacterial culture. The frequency of positive pathogen signals was markedly higher with PCR compared with culture (70.6% vs. 32.2%). This was accompanied by a substantial higher percentage of multiple pathogen identifications and a lower percentage of single identifications in the PCR compared with bacterial culture. Using bacterial culture as gold standard, moderate to high sensitivities (76.9-100%) and specificities (63.3-98.7%) were calculated for six out of seven pathogens with sufficient detection numbers. For Enterococcus spp, the sensitivity was only 9.1%. When the PCR results of pooled udder quarter samples of the 173 cows were compared with the single udder quarter samples, in 72% of the cases, major pathogen DNA was either not found in both types of samples, or in the case of a positive pool sample, the respective pathogens were found in at least one udder quarter sample. With both methods, the most frequently detected mastitis pathogens were coryneform bacteria (PCR: Corynebacterium bovis), coagulase-negative staphylococci (CNS) and Staphylococcus (S.) aureus, followed by Arcanobacterium pyogenes/Peptoniphilus indolicus with PCR, and then with both methods, Streptococcus uberis. The staphylococcal beta-lactamase gene was found in 27.7% of the S. aureus and in 37.0% of the CNS identifications.

Enrichment of the Cancer Stem Phenotype in Sphere Cultures of Prostate Cancer Cell Lines Occurs through Activation of Developmental Pathways Mediated by the Transcriptional Regulator ΔNp63α

PubMed Central

Portillo-Lara, Roberto; Alvarez, Mario Moisés

2015-01-01

Background Cancer stem cells (CSC) drive prostate cancer tumor survival and metastasis. Nevertheless, the development of specific therapies against CSCs is hindered by the scarcity of these cells in prostate tissues. Suspension culture systems have been reported to enrich CSCs in primary cultures and cell lines. However, the molecular mechanisms underlying this phenomenon have not been fully explored. Methodology/Principal Findings We describe a prostasphere assay for the enrichment of CD133+ CSCs in four commercial PCa cell lines: 22Rv1, DU145, LNCaP, and PC3. Overexpression of CD133, as determined by flow cytometric analysis, correlated with an increased clonogenic, chemoresistant, and invasive potential in vitro. This phenotype is concordant to that of CSCs in vivo. Gene expression profiling was then carried out using the Cancer Reference panel and the nCounter system from NanoString Technologies. This analysis revealed several upregulated transcripts that can be further explored as potential diagnostic markers or therapeutic targets. Furthermore, functional annotation analysis suggests that ΔNp63α modulates the activation of developmental pathways responsible for the increased stem identity of cells growing in suspension cultures. Conclusions/Significance We conclude that profiling the genetic mechanisms involved in CSC enrichment will help us to better understand the molecular pathways that underlie CSC pathophysiology. This platform can be readily adapted to enrich and assay actual patient samples, in order to design patient-specific therapies that are aimed particularly against CSCs. PMID:26110651

Effect of organic phosphorus and nitrogen enrichment of mesotrophic lake water on dynamics and diversity of planktonic microbial communities--DNA and protein case studies (mesocosm experiments).

PubMed

Chróst, Ryszard J; Adamczewski, Tomasz; Kalinowska, Krystyna; Skowrońska, Agnieszka

2009-01-01

Effects of mesotrophic lake water enrichment with organic phosphorus and nitrogen substrates (DNA and model protein, bovine serum albumin--BSA) on dynamics and diversity of natural microbial communities (bacteria, heterotrophic nanoflagellates, ciliates) were studied in mesocosm experiments. Simultaneous enrichment with DNA and BSA strongly increased the abundance and biomass of all studied groups of microorganisms and induced changes in their morphological and taxonomic structure. The increased participation of large heterotrophic nanoflagellates cells (larger than 10 microm) in their total numbers and shifts in taxonomic and trophic structure of the ciliates, from algivorous to small bacterivorous, species were observed. Grazing caused changes in bacterial size distribution in all enriched mesocosms. Large (10-50 microm) filamentous bacteria significantly contributed to the total bacterial numbers and biomass. Pronounced increase in populations of beta- and gamma-Proteobacteria was found in lake water enriched with organic P and N sources, whereas alpha-Proteobacteria did not change markedly in the studied mesocosms. DNA additions stimulated the rates of bacterial secondary production. BSA shortened the rates of bacterial biomass turnover in lake water. Relatively high and constant (approximately 30%) percentage contribution of active bacteria (MEM+) in two mesocosms enriched with DNA and DNA+BSA suggested the important role of nucleic acids as a source of phosphorus for bacterial growth, activity and production. Numerous and statistically significant correlations between bacteria and protists indicated the direct and selective predator-prey relationship.

Yeast and bacterial diversity along a transect in an acidic, As-Fe rich environment revealed by cultural approaches.

PubMed

Delavat, François; Lett, Marie-Claire; Lièvremont, Didier

2013-10-01

Acid mine drainages (AMDs) are often thought to harbour low biodiversity, yet little is known about the diversity distribution along the drainages. Using culture-dependent approaches, the microbial diversity from the Carnoulès AMD sediment was investigated for the first time along a transect showing progressive environmental stringency decrease. In total, 20 bacterial genera were detected, highlighting a higher bacterial diversity than previously thought. Moreover, this approach led to the discovery of 16 yeast species, demonstrating for the first time the presence of this important phylogenetic group in this AMD. All in all, the location of the microbes along the transect helps to better understand their distribution in a pollution gradient. Copyright © 2013 Elsevier B.V. All rights reserved.

Polymerase Chain Reaction-Electrospray-Time-of-Flight Mass Spectrometry Versus Culture for Bacterial Detection in Septic Arthritis and Osteoarthritis.

PubMed

Palmer, Michael P; Melton-Kreft, Rachael; Nistico, Laura; Hiller, N Louisa; Kim, Leon H J; Altman, Gregory T; Altman, Daniel T; Sotereanos, Nicholas G; Hu, Fen Z; De Meo, Patrick J; Ehrlich, Garth D

2016-12-01

Preliminary studies have identified known bacterial pathogens in the knees of patients with osteoarthritis (OA) before arthroplasty. The current study was designed to determine the incidence and types of bacteria present in the synovial fluid of native knee joints from adult patients with diagnoses of septic arthritis and OA. Patients were enrolled between October 2010 and January 2013. Synovial fluid samples from the affected knee were collected and evaluated with both traditional microbial culture and polymerase chain reaction-electrospray ionization-time-of-flight mass spectrometry (molecular diagnostics [MDx]) to prospectively characterize the microbial content. Patients were grouped by diagnosis into one of two cohorts, those with clinical suspicion of septic arthritis (n = 44) and those undergoing primary arthroplasty of the knee for OA (n = 21). In all cases where discrepant culture and MDx results were obtained, we performed species-specific 16S rRNA fluorescence in situ hybridization (FISH) as a confirmatory test. MDx testing identified bacteria in 50% of the suspected septic arthritis cases and 29% of the arthroplasty cases, whereas culture detected bacteria in only 16% of the former and 0% of the latter group. The overall difference in detection rates for culture and MDx was very highly significant, p-value = 2.384 × 10 -7 . All of the culture-positive cases were typed as Staphylococcus aureus. Two of the septic arthritis cases were polymicrobial as was one of the OA cases by MDx. FISH testing of the specimens with discordant results supported the MDx findings in 91% (19/21) of the cases, including one case where culture detected S. aureus and MDx detected Streptococcus agalactiae. MDx were more sensitive than culture, as confirmed by FISH. FISH only identifies bacteria that are embedded or infiltrated within the tissue and is thus not susceptible to contamination. Not all suspected cases of septic arthritis contain bacteria, but a

Production of bacterial cellulose using different carbon sources and culture media.

PubMed

Mohammadkazemi, Faranak; Azin, Mehrdad; Ashori, Alireza

2015-03-06

In this work, the effects of carbon sources and culture media on the production and structural properties of bacterial cellulose (BC) have been studied. BC nanofibers were synthesized using Gluconacetobacter xylinus strain PTCC 1734. Media used were Hestrin-Schramm (H), Yamanaka (Y), and Zhou (Z). Five different carbon sources, namely date syrup, glucose, mannitol, sucrose, and food-grade sucrose were used in these media. All the produced BC pellicles were characterized in terms of dry weight production, biomass yield, thermal stability, crystallinity and morphology by thermogravimetric analysis (TGA), x-ray diffraction (XRD), and field emission scanning electron microscopy (FE-SEM). The obtained results showed that mannitol lead to the highest yield, followed by sucrose. The highest production efficiency of mannitol might be due to the nitrogen source, which plays an important role. The maximum improvement on the thermal stability of the composites was achieved when mannitol was used in H medium. In addition, the crystallinity was higher in BC formed in H medium compared to other media. FE-SEM micrographs illustrated that the BC pellicles, synthesized in the culture media H and Z, were stable, unlike those in medium Y that were unstable. The micrographs of BC produced in media containing mannitol and sucrose provided evidence of the strong interfacial adhesion between the BC fibers without noticeable aggregates. Copyright © 2014 Elsevier Ltd. All rights reserved.

Acute bacterial meningitis cases diagnosed by culture and PCR in a children's hospital throughout a 9-Year period (2000-2008) in Athens, Greece.

PubMed

Papavasileiou, Konstantina; Papavasileiou, Eleni; Tzanakaki, Georgina; Voyatzi, Aliki; Kremastinou, Jenny; Chatzipanagiotou, Stylianos

2011-04-01

Acute bacterial meningitis is one of the most severe infectious diseases, affecting mainly infants and, secondarily, older children and adolescents. Diagnosis in the early stages is often difficult and despite treatment with appropriate antibiotic therapy, the case fatality rate remains high. In the present study, the incidence of bacterial meningitis was registered in a general pediatric hospital in Athens, Greece, during a 9-year period (2000-2008), and the use of molecular methods in the diagnosis of bacterial meningitis versus the conventional cultural methods was evaluated. The impact of vaccination against meningitis-causing bacteria on the incidence of bacterial meningitis was also assessed. From a total of 1833 children hospitalized with suspected clinical symptoms and signs of meningitis, all cerebrospinal fluid (CSF) and blood samples were analyzed by white blood cell (WBC) count, measurement of glucose, protein, and C-reactive protein (CRP) levels, as well as by conventional bacteriologic culture methods. If samples showed altered CSF markers that were consistent with meningitis in general, they were further investigated by PCR for bacterial pathogens. Of the 1833 patients, 289 (15.76%) were found to be positive for meningitis after CSF examination, based on white blood cell count and differentiation, glucose, protein, and CRP. Fifty-six of the 289 (19.37%) had confirmed bacterial meningitis, as diagnosed by either culture and/or PCR. Of these 56 cases, 44 (78.6%) were detected only by PCR, and 12 cases (21.4%) were confirmed by PCR and culture. The predominant microorganism was Neisseria meningitidis serogroup B (n = 40; 71.4%), followed by Streptococcus pneumoniae not typed [NT] (n = 7; 12.5%), Streptococcus spp. (n =4; 7.1%), Haemophilus influenzae NT (n = 2; 3.6%), and S. pneumoniae serotype 3, Streptococcus group B, and S. pneumoniae serotype 18C (each n = 1; 1.8%). In Greece, according to data from the National Meningitis Reference

Enrichment of Root Endophytic Bacteria from Populus deltoides and Single-Cell-Genomics Analysis

DOE PAGES

Utturkar, Sagar M.; Cude, W. Nathan; Robeson, Jr., Michael S.; ...

2016-07-15

Bacterial endophytes that colonize Populus trees contribute to nutrient acquisition, prime immunity responses, and directly or indirectly increase both above- and below-ground biomasses. Endophytes are embedded within plant material, so physical separation and isolation are difficult tasks. Application of culture-independent methods, such as metagenome or bacterial transcriptome sequencing, has been limited due to the predominance of DNA from the plant biomass. In this paper, we present a modified differential and density gradient centrifugation-based protocol for the separation of endophytic bacteria from Populus roots. This protocol achieved substantial reduction in contaminating plant DNA, allowed enrichment of endophytic bacteria away from themore » plant material, and enabled single-cell genomics analysis. Four single-cell genomes were selected for whole-genome amplification based on their rarity in the microbiome (potentially uncultured taxa) as well as their inferred abilities to form associations with plants. Bioinformatics analyses, including assembly, contamination removal, and completeness estimation, were performed to obtain single-amplified genomes (SAGs) of organisms from the phyla Armatimonadetes, Verrucomicrobia, and Planctomycetes, which were unrepresented in our previous cultivation efforts. Finally, comparative genomic analysis revealed unique characteristics of each SAG that could facilitate future cultivation efforts for these bacteria.« less

Polymicrobial airway bacterial communities in adult bronchiectasis patients

PubMed Central

2014-01-01

Background Chronic airway infection contributes to the underlying pathogenesis of non-cystic fibrosis bronchiectasis (NCFBr). In contrast to other chronic airway infections, associated with COPD and CF bronchiectasis, where polymicrobial communities have been implicated in lung damage due to the vicious circle of recurrent bacterial infections and inflammation, there is sparse information on the composition of bacterial communities in NCFBr. Seventy consecutive patients were recruited from an outpatient adult NCFBr clinic. Bacterial communities in sputum samples were analysed by culture and pyrosequencing approaches. Bacterial sequences were analysed using partial least square discrimination analyses to investigate trends in community composition and identify those taxa that contribute most to community variation. Results The lower airway in NCFBr is dominated by three bacterial taxa Pasteurellaceae, Streptococcaceae and Pseudomonadaceae. Moreover, the bacterial community is much more diverse than indicated by culture and contains significant numbers of other genera including anaerobic Prevotellaceae, Veillonellaceae and Actinomycetaceae. We found particular taxa are correlated with different clinical states, 27 taxa were associated with acute exacerbations, whereas 11 taxa correlated with stable clinical states. We were unable to demonstrate a significant effect of antibiotic therapy, gender, or lung function on the diversity of the bacterial community. However, presence of clinically significant culturable taxa; particularly Pseudomonas aeruginosa and Haemophilus influenzae correlated with a significant change in the diversity of the bacterial community in the lung. Conclusions We have demonstrated that acute exacerbations, the frequency of exacerbation and episodes of clinical stability are correlated, in some patients, with a significantly different bacterial community structure, that are associated with a presence of particular taxa in the NCFBr lung. Moreover

Neonatal blood cultures: effect of delayed entry into the blood culture machine and bacterial concentration on the time to positive growth in a simulated model.

PubMed

Jardine, Luke Anthony; Sturgess, Barbara Ruth; Inglis, Garry Donald Trevor; Davies, Mark William

2009-04-01

To determine if: time from blood culture inoculation to positive growth (total time to positive) and time from blood culture machine entry to positive growth (machine time to positive) is altered by delayed entry into the automated blood culture machine, and if the total time to positive differs by the concentration of organisms inoculated into blood culture bottles. Staphylococcus epidermidis, Escherichia coli and group B beta-haemolytic streptococci were chosen as clinically significant representative organisms. Two concentrations (> or =10 colony-forming units per millilitre and <1 colony-forming units per millilitre) were inoculated into PEDS BacT/Alert blood culture bottles and randomly allocated to one of three delayed automated blood culture machine entry times (30 min/8.5 h/15.5 h). For all organisms at all concentrations, except the Staphylococcus epidermidis, the machine time to positive was significantly decreased by delayed entry. For all organisms at all concentrations, the mean total time to positive significantly increased with increasing delayed entry into the blood culture machine. Higher concentrations of group B beta-haemolytic streptococci and Escherichia coli grew significantly faster than lower concentrations. Bacterial growth in inoculated bottles, stored at room temperature, continues although at a slower rate than in those blood culture bottles immediately entered into the machine. If a blood culture specimen has been stored at room temperature for greater than 15.5 h, the currently allowed safety margin of 36 h (before declaring a result negative) may be insufficient.

Simple and Versatile Turbidimetric Monitoring of Bacterial Growth in Liquid Cultures Using a Customized 3D Printed Culture Tube Holder and a Miniaturized Spectrophotometer: Application to Facultative and Strictly Anaerobic Bacteria.

PubMed

Maia, Margarida R G; Marques, Sara; Cabrita, Ana R J; Wallace, R John; Thompson, Gertrude; Fonseca, António J M; Oliveira, Hugo M

2016-01-01

Here we introduce a novel strategy for turbidimetric monitoring of bacterial growth in liquid culture. The instrumentation comprises a light source, a customized 3D printed culture tube holder and a miniaturized spectrophotometer, connected through optical cables. Due to its small footprint and the possibility to operate with external light, bacterial growth was directly monitored from culture tubes in a simple and versatile fashion. This new portable measurement technique was used to monitor the growth of facultative (Escherichia coli ATCC/25922, and Staphylococcus aureus ATCC/29213) and strictly (Butyrivibrio fibrisolvens JW11, Butyrivibrio proteoclasticus P18, and Propionibacterium acnes DSMZ 1897) anaerobic bacteria. For E. coli and S. aureus, the growth rates calculated from normalized optical density values were compared with those ones obtained using a benchtop spectrophotometer without significant differences (P = 0.256). For the strictly anaerobic species, a high precision (relative standard deviation < 3.5%) was observed between replicates up to 48 h. Regarding its potential for customization, this manifold could accommodate further developments for customized turbidimetric monitoring, such as the use of light-emitting diodes as a light source or flow cells.

Simple and Versatile Turbidimetric Monitoring of Bacterial Growth in Liquid Cultures Using a Customized 3D Printed Culture Tube Holder and a Miniaturized Spectrophotometer: Application to Facultative and Strictly Anaerobic Bacteria

PubMed Central

Maia, Margarida R. G.; Marques, Sara; Cabrita, Ana R. J.; Wallace, R. John; Thompson, Gertrude; Fonseca, António J. M.; Oliveira, Hugo M.

2016-01-01

Here we introduce a novel strategy for turbidimetric monitoring of bacterial growth in liquid culture. The instrumentation comprises a light source, a customized 3D printed culture tube holder and a miniaturized spectrophotometer, connected through optical cables. Due to its small footprint and the possibility to operate with external light, bacterial growth was directly monitored from culture tubes in a simple and versatile fashion. This new portable measurement technique was used to monitor the growth of facultative (Escherichia coli ATCC/25922, and Staphylococcus aureus ATCC/29213) and strictly (Butyrivibrio fibrisolvens JW11, Butyrivibrio proteoclasticus P18, and Propionibacterium acnes DSMZ 1897) anaerobic bacteria. For E. coli and S. aureus, the growth rates calculated from normalized optical density values were compared with those ones obtained using a benchtop spectrophotometer without significant differences (P = 0.256). For the strictly anaerobic species, a high precision (relative standard deviation < 3.5%) was observed between replicates up to 48 h. Regarding its potential for customization, this manifold could accommodate further developments for customized turbidimetric monitoring, such as the use of light-emitting diodes as a light source or flow cells. PMID:27630632

Bacterial Associates Modify Growth Dynamics of the Dinoflagellate Gymnodinium catenatum

PubMed Central

Bolch, Christopher J. S.; Bejoy, Thaila A.; Green, David H.

2017-01-01

Marine phytoplankton cells grow in close association with a complex microbial associate community known to affect the growth, behavior, and physiology of the algal host. The relative scale and importance these effects compared to other major factors governing algal cell growth remain unclear. Using algal-bacteria co-culture models based on the toxic dinoflagellate Gymnodinium catenatum, we tested the hypothesis that associate bacteria exert an independent effect on host algal cell growth. Batch co-cultures of G. catenatum were grown under identical environmental conditions with simplified bacterial communities composed of one-, two-, or three-bacterial associates. Modification of the associate community membership and complexity induced up to four-fold changes in dinoflagellate growth rate, equivalent to the effect of a 5°C change in temperature or an almost six-fold change in light intensity (20–115 moles photons PAR m-2 s-1). Almost three-fold changes in both stationary phase cell concentration and death rate were also observed. Co-culture with Roseobacter sp. DG874 reduced dinoflagellate exponential growth rate and led to a more rapid death rate compared with mixed associate community controls or co-culture with either Marinobacter sp. DG879, Alcanivorax sp. DG881. In contrast, associate bacteria concentration was positively correlated with dinoflagellate cell concentration during the exponential growth phase, indicating growth was limited by supply of dinoflagellate-derived carbon. Bacterial growth increased rapidly at the onset of declining and stationary phases due to either increasing availability of algal-derived carbon induced by nutrient stress and autolysis, or at mid-log phase in Roseobacter co-cultures potentially due to the onset of bacterial-mediated cell lysis. Co-cultures with the three bacterial associates resulted in dinoflagellate and bacterial growth dynamics very similar to more complex mixed bacterial community controls, suggesting that

Comparison of Bacterial Community Composition of Primary and Persistent Endodontic Infections Using Pyrosequencing.

PubMed

Tzanetakis, Giorgos N; Azcarate-Peril, M Andrea; Zachaki, Sophia; Panopoulos, Panos; Kontakiotis, Evangelos G; Madianos, Phoebus N; Divaris, Kimon

2015-08-01

Elucidating the microbial ecology of endodontic infections (EIs) is a necessary step in developing effective intracanal antimicrobials. The aim of the present study was to investigate the bacterial composition of symptomatic and asymptomatic primary and persistent infections in a Greek population using high-throughput sequencing methods. 16S amplicon pyrosequencing of 48 root canal bacterial samples was conducted, and sequencing data were analyzed using an oral microbiome-specific and a generic (Greengenes) database. Bacterial abundance and diversity were examined by EI type (primary or persistent), and statistical analysis was performed by using non-parametric and parametric tests accounting for clustered data. Bacteroidetes was the most abundant phylum in both infection groups. Significant, albeit weak associations of bacterial diversity were found, as measured by UniFrac distances with infection type (analyses of similarity, R = 0.087, P = .005) and symptoms (analyses of similarity, R = 0.055, P = .047). Persistent infections were significantly enriched for Proteobacteria and Tenericutes compared with primary ones; at the genus level, significant differences were noted for 14 taxa, including increased enrichment of persistent infections for Lactobacillus, Streptococcus, and Sphingomonas. More but less abundant phyla were identified using the Greengenes database; among those, Cyanobacteria (0.018%) and Acidobacteria (0.007%) were significantly enriched among persistent infections. Persistent infections showed higher phylogenetic diversity (PD) (asymptomatic: PD = 9.2, standard error [SE] = 1.3; symptomatic: PD = 8.2, SE = 0.7) compared with primary infections (asymptomatic: PD = 5.9, SE = 0.8; symptomatic: PD = 7.4, SE = 1.0). The present study revealed a high bacterial diversity of EI and suggests that persistent infections may have more diverse bacterial communities than primary infections. Copyright © 2015 American Association of Endodontists. Published by

Investigation of the biotransformation of pentachlorophenol and pulp paper mill effluent decolorisation by the bacterial strains in a mixed culture.

PubMed

Singh, Shail; Chandra, R; Patel, D K; Reddy, M M K; Rai, Vibhuti

2008-09-01

Mixed culture of two bacterial strains Bacillus sp. and Serratia marcescens showed potential pentachlorophenol (PCP) degradation and decolorisation of pulp paper mill effluent. The physico-chemical quality of pulp paper mill effluent has been analyzed after 168 h incubation period degraded by mixed culture. The study revealed that it has decreased high load of BOD, COD, TS, TDS, TSS, sulphate, phosphate, total nitrogen, total phenols, metals and different salts (i.e. chloride, sodium, nitrate, potassium) at 168 h incubation period. PCP degradation in pulp paper mill effluent was confirmed by HPLC analysis. Mixed culture was found to degrade PCP up to (94%) present in pulp paper mill effluent with 1% glucose and 0.5% peptone (w/v) at 30+/-1 degrees C, pH 8.0+/-0.2 at 120 rpm in 168 h incubation period. The simultaneous release of chloride ion up to 1,200 mg/l at 168 h emphasized the bacterial dechlorination in the medium. The pulp paper mill effluent degradation was also supported by decline in pH, AOX (absorbable organic halides), color, D.O., BOD, COD and PCP. The analysis of pulp paper mill effluent degradation products by GC-MS analysis revealed the formation of low molecular weight compound like 2-chlorophenol (RT=3.8 min) and tetrachlorohydroquinone (RT=11.86 min) from PCP extracted degraded sample. Further, mixed culture may be used for bioremediation of PCP containing pulp paper mill waste in the environment.

Detection limits of Legionella pneumophila in environmental samples after co-culture with Acanthamoeba polyphaga

PubMed Central

2013-01-01

Background The efficiency of recovery and the detection limit of Legionella after co-culture with Acanthamoeba polyphaga are not known and so far no investigations have been carried out to determine the efficiency of the recovery of Legionella spp. by co-culture and compare it with that of conventional culturing methods. This study aimed to assess the detection limits of co-culture compared to culture for Legionella pneumophila in compost and air samples. Compost and air samples were spiked with known concentrations of L. pneumophila. Direct culturing and co-culture with amoebae were used in parallel to isolate L. pneumophila and recovery standard curves for both methods were produced for each sample. Results The co-culture proved to be more sensitive than the reference method, detecting 102-103 L. pneumophila cells in 1 g of spiked compost or 1 m3 of spiked air, as compared to 105-106 cells in 1 g of spiked compost and 1 m3 of spiked air. Conclusions Co-culture with amoebae is a useful, sensitive and reliable technique to enrich L. pneumophila in environmental samples that contain only low amounts of bacterial cells. PMID:23442526

Books to Expand and Enrich Experiences.

ERIC Educational Resources Information Center

Winfield, Evelyn T.

1983-01-01

Books are briefly described that parents can read and discuss with their children to enrich travel and cultural activities. Books on the nation's capital city, the Liberty Bell, the Statue of Liberty, architecture, zoos, dinosaurs, and other subjects are included. (PP)

Enantioselective degradation of ofloxacin and levofloxacin by the bacterial strains Labrys portucalensis F11 and Rhodococcus sp. FP1.

PubMed

Maia, Alexandra S; Tiritan, Maria Elizabeth; Castro, Paula M L

2018-07-15

Fluoroquinolones are a class of antibiotics widely prescribed in both human and veterinary medicine of high environmental concern and characterized as environmental micropollutants due to their ecotoxicity and persistence and antibacterial resistance potential. Ofloxacin and levofloxacin are chiral fluoroquinolones commercialized as racemate and in enantiomerically pure form, respectively. Since the pharmacological properties and toxicity of the enantiomers may be very different, understanding the stereochemistry of these compounds should be a priority in environmental monitoring. This work presents the biodegradation of racemic ofloxacin and its (S)-enantiomer levofloxacin by the bacterial strains Labrys portucalensis F11 and Rhodococcus sp. FP1 at a laboratory-scale microcosm following the removal and the behavior of the enantiomers. Strain F11 could degrade both antibiotics almost completely when acetate was supplied regularly to the cultures. Enrichment of the (R)-enantiomer was observed in FP1 and F11 cultures supplied with ofloxacin. Racemization was observed in the biodegradation of the pure (S)-ofloxacin (levofloxacin) by strain F11, which was confirmed by liquid chromatography - exact mass spectrometry. Biodegradation of ofloxacin at 450 µg L -1 by both bacterial strains expressed good linear fits (R 2 > 0.98) according to the Rayleigh equation. The enantiomeric enrichment factors were comprised between - 22.5% to - 9.1%, and - 18.7% to - 9.0% in the biodegradation of ofloxacin by strains F11 and FP1, respectively, with no significant differences for the two bacteria under the same conditions. This is the first time that enantioselective biodegradation of ofloxacin and levofloxacin by single bacteria is reported. Copyright © 2018 Elsevier Inc. All rights reserved.

Analysis of Culture-Dependent versus Culture-Independent Techniques for Identification of Bacteria in Clinically Obtained Bronchoalveolar Lavage Fluid

PubMed Central

Dickson, Robert P.; Erb-Downward, John R.; Prescott, Hallie C.; Martinez, Fernando J.; Curtis, Jeffrey L.; Lama, Vibha N.

2014-01-01

The diagnosis and management of pneumonia are limited by the use of culture-based techniques of microbial identification, which may fail to identify unculturable, fastidious, and metabolically active viable but unculturable bacteria. Novel high-throughput culture-independent techniques hold promise but have not been systematically compared to conventional culture. We analyzed 46 clinically obtained bronchoalveolar lavage (BAL) fluid specimens from symptomatic and asymptomatic lung transplant recipients both by culture (using a clinical microbiology laboratory protocol) and by bacterial 16S rRNA gene pyrosequencing. Bacteria were identified in 44 of 46 (95.7%) BAL fluid specimens by culture-independent sequencing, significantly more than the number of specimens in which bacteria were detected (37 of 46, 80.4%, P ≤ 0.05) or “pathogen” species reported (18 of 46, 39.1%, P ≤ 0.0001) via culture. Identification of bacteria by culture was positively associated with culture-independent indices of infection (total bacterial DNA burden and low bacterial community diversity) (P ≤ 0.01). In BAL fluid specimens with no culture growth, the amount of bacterial DNA was greater than that in reagent and rinse controls, and communities were markedly dominated by select Gammaproteobacteria, notably Escherichia species and Pseudomonas fluorescens. Culture growth above the threshold of 104 CFU/ml was correlated with increased bacterial DNA burden (P < 0.01), decreased community diversity (P < 0.05), and increased relative abundance of Pseudomonas aeruginosa (P < 0.001). We present two case studies in which culture-independent techniques identified a respiratory pathogen missed by culture and clarified whether a cultured “oral flora” species represented a state of acute infection. In summary, we found that bacterial culture of BAL fluid is largely effective in discriminating acute infection from its absence and identified some specific limitations of BAL fluid culture in

Analysis of culture-dependent versus culture-independent techniques for identification of bacteria in clinically obtained bronchoalveolar lavage fluid.

PubMed

Dickson, Robert P; Erb-Downward, John R; Prescott, Hallie C; Martinez, Fernando J; Curtis, Jeffrey L; Lama, Vibha N; Huffnagle, Gary B

2014-10-01

The diagnosis and management of pneumonia are limited by the use of culture-based techniques of microbial identification, which may fail to identify unculturable, fastidious, and metabolically active viable but unculturable bacteria. Novel high-throughput culture-independent techniques hold promise but have not been systematically compared to conventional culture. We analyzed 46 clinically obtained bronchoalveolar lavage (BAL) fluid specimens from symptomatic and asymptomatic lung transplant recipients both by culture (using a clinical microbiology laboratory protocol) and by bacterial 16S rRNA gene pyrosequencing. Bacteria were identified in 44 of 46 (95.7%) BAL fluid specimens by culture-independent sequencing, significantly more than the number of specimens in which bacteria were detected (37 of 46, 80.4%, P ≤ 0.05) or "pathogen" species reported (18 of 46, 39.1%, P ≤ 0.0001) via culture. Identification of bacteria by culture was positively associated with culture-independent indices of infection (total bacterial DNA burden and low bacterial community diversity) (P ≤ 0.01). In BAL fluid specimens with no culture growth, the amount of bacterial DNA was greater than that in reagent and rinse controls, and communities were markedly dominated by select Gammaproteobacteria, notably Escherichia species and Pseudomonas fluorescens. Culture growth above the threshold of 10(4) CFU/ml was correlated with increased bacterial DNA burden (P < 0.01), decreased community diversity (P < 0.05), and increased relative abundance of Pseudomonas aeruginosa (P < 0.001). We present two case studies in which culture-independent techniques identified a respiratory pathogen missed by culture and clarified whether a cultured "oral flora" species represented a state of acute infection. In summary, we found that bacterial culture of BAL fluid is largely effective in discriminating acute infection from its absence and identified some specific limitations of BAL fluid culture in the

Effect of different enrichment strategies on microbial community structure in petroleum-contaminated marine sediment in Dalian, China.

PubMed

Chen, Chao; Liu, Qiu; Liu, Changjian; Yu, Jicheng

2017-04-15

An oil spill occurred at Xingang Port, Dalian, China in 2010. Four years after this spill, oil contamination was still detected in samples collected nearby. In this study, the strains that evolved in the sediment were screened by high-throughput sequencing technology. Most of these strains were genera reported to have functions associated with crude oil biodegradation. The diversities and numbers of microbes were monitored through enrichment culturing; the dominant strains propagated at first, but the enrichment could not be continued, which indicated that the prolonged culture was not effective in the enrichment of the micro-consortium. Oxygen was also observed to affect the propagation of the dominant microbes. The results showed the role of culture strategies and oxygen in the enrichment of the petroleum-degrading microbes. Therefore, dominant strains could be screened by optimizing both the enrichment time and oxygen concentration used for culturing to facilitate oil biodegradation in the marine ecosystem. Copyright © 2017 Elsevier Ltd. All rights reserved.

Morphomechanics of bacterial biofilms undergoing anisotropic differential growth

NASA Astrophysics Data System (ADS)

Zhang, Cheng; Li, Bo; Huang, Xiao; Ni, Yong; Feng, Xi-Qiao

2016-10-01

Growing bacterial biofilms exhibit a number of surface morphologies, e.g., concentric wrinkles, radial ridges, and labyrinthine networks, depending on their physiological status and nutrient access. We explore the mechanisms underlying the emergence of these greatly different morphologies. Ginzburg-Landau kinetic method and Fourier spectral method are integrated to simulate the morphological evolution of bacterial biofilms. It is shown that the morphological instability of biofilms is triggered by the stresses induced by anisotropic and heterogeneous bacterial expansion, and involves the competition between membrane energy and bending energy. Local interfacial delamination further enriches the morphologies of biofilms. Phase diagrams are established to reveal how the anisotropy and spatial heterogeneity of growth modulate the surface patterns. The mechanics of three-dimensional microbial morphogenesis may also underpin self-organization in other development systems and provide a potential strategy for engineering microscopic structures from bacterial aggregates.

Generalist hydrocarbon-degrading bacterial communities in the oil-polluted water column of the North Sea

PubMed Central

Chronopoulou, Panagiota-Myrsini; Sanni, Gbemisola O; Silas-Olu, Daniel I; van der Meer, Jan Roelof; Timmis, Kenneth N; Brussaard, Corina P D; McGenity, Terry J

2015-01-01

The aim of this work was to determine the effect of light crude oil on bacterial communities during an experimental oil spill in the North Sea and in mesocosms (simulating a heavy, enclosed oil spill), and to isolate and characterize hydrocarbon-degrading bacteria from the water column. No oil-induced changes in bacterial community (3 m below the sea surface) were observed 32 h after the experimental spill at sea. In contrast, there was a decrease in the dominant SAR11 phylotype and an increase in Pseudoalteromonas spp. in the oiled mesocosms (investigated by 16S rRNA gene analysis using denaturing gradient gel electrophoresis), as a consequence of the longer incubation, closer proximity of the samples to oil, and the lack of replenishment with seawater. A total of 216 strains were isolated from hydrocarbon enrichment cultures, predominantly belonging to the genus Pseudoaltero monas; most strains grew on PAHs, branched and straight-chain alkanes, as well as many other carbon sources. No obligate hydrocarbonoclastic bacteria were isolated or detected, highlighting the potential importance of cosmopolitan marine generalists like Pseudoalteromonas spp. in degrading hydrocarbons in the water column beneath an oil slick, and revealing the susceptibility to oil pollution of SAR11, the most abundant bacterial clade in the surface ocean. PMID:25251384

Gamma-irradiated bacterial preparation having anti-tumor activity

DOEpatents

Vass, Arpad A.; Tyndall, Richard L.; Terzaghi-Howe, Peggy

1999-01-01

A bacterial preparation from Pseudomonas species isolated #15 ATCC 55638 that has been exposed to gamma radiation exhibits cytotoxicity that is specific for neoplastic carcinoma cells. A method for obtaining a bacterial preparation having antitumor activity consists of suspending a bacterial isolate in media and exposing the suspension to gamma radiation. A bacterial preparation of an aged culture of an amoeba-associated bacteria exhibits anti-reverse transcriptase activity. A method for obtaining a bacterial preparation having anti-reverse transcriptase activity from an amoeba-associated bacterial isolate grown to stationary phase is disclosed.

Building a Morbidostat: An automated continuous-culture device for studying bacterial drug resistance under dynamically sustained drug inhibition

PubMed Central

Toprak, Erdal; Veres, Adrian; Yildiz, Sadik; Pedraza, Juan M.; Chait, Remy; Paulsson, Johan; Kishony, Roy

2013-01-01

We present a protocol for building and operating an automated fluidic system for continuous culture that we call the “morbidostat”. The morbidostat is used to follow evolution of microbial drug resistance in real time. Instead of exposing bacteria to predetermined drug environments, the morbidostat constantly measures the growth rates of evolving microbial populations and dynamically adjusts drug concentrations inside culture vials in order to maintain a constant drug induced inhibition. The growth rate measurements are done using an optical detection system that is based on measuring the intensity of back-scattered light from bacterial cells suspended in the liquid culture. The morbidostat can additionally be used as a chemostat or a turbidostat. The whole system can be built from readily available components within two to three weeks, by biologists with some electronics experience or engineers familiar with basic microbiology. PMID:23429717

Biodiversity within hot spring microbial mat communities: molecular monitoring of enrichment cultures

NASA Technical Reports Server (NTRS)

Ward, D. M.; Santegoeds, C. M.; Nold, S. C.; Ramsing, N. B.; Ferris, M. J.; Bateson, M. M.

1997-01-01

We have begun to examine the basis for incongruence between hot spring microbial mat populations detected by cultivation or by 16S rRNA methods. We used denaturing gradient gel electrophoresis (DGGE) to monitor enrichments and isolates plated therefrom. At near extincting inoculum dilutions we observed Chloroflexus-like and cyanobacterial populations whose 16S rRNA sequences have been detected in the 'New Pit' Spring Chloroflexus mat and the Octopus Spring cyanobacterial mat. Cyanobacterial populations enriched from 44 to 54 degrees C and 56 to 63 degrees C samples at near habitat temperatures were similar to those previously detected in mat samples of comparable temperatures. However, a lower temperature enrichment from the higher temperature sample selected for the populations found in the lower temperature sample. Three Thermus populations detected by both DGGE and isolation exemplify even more how enrichment may bias our view of community structure. The most abundant population was adapted to the habitat temperature (50 degrees C), while populations adapted to 65 degrees C and 70 degrees C were 10(2)- and 10(4)-fold less abundant, respectively. However, enrichment at 70 degrees C favored the least abundant strain. Inoculum dilution and incubation at the habitat temperature favored the more numerically relevant populations. We enriched many other aerobic chemoorganotrophic populations at various inoculum dilutions and substrate concentrations, most of whose 16S rRNA sequences have not been detected in mats. A common feature of numerically relevant cyanobacterial, Chloroflexus-like and aerobic chemorganotrophic populations, is that they grow poorly and resist cultivation on solidified medium, suggesting plating bias, and that the medium composition and incubation conditions may not reflect the natural microenvironments these populations inhabit.

Cultural Literacy: Negotiating Language, Culture, and Thought

ERIC Educational Resources Information Center

Clark, Ellen Riojas; Flores, Belinda Bustos

2007-01-01

Our schools see increasing numbers of students who reflect the wide diversity of this country's population, but too often these differences--culture, language, socioeconomic backgrounds, ethnicity are viewed from negative or deficit perspectives when they are, in fact, the cultural capital that enriches discussion, broadens viewpoints, and…

Cultural Enrichment through Community Action.

ERIC Educational Resources Information Center

Wilson, O. J.

This project was conceived as a technique for helping to eliminate a cultural void in the areas of art, music, and theatre in the service area of Western Kentucky University. To implement this concept, demonstrations were conducted in art, music, theatre, and in library and lecture resources in 16 counties over a four-year period. The attendance…

Bioleaching of electronic scrap by mixed culture of moderately thermophilic microorganisms

NASA Astrophysics Data System (ADS)

Ivǎnuş, D.; ǎnuş, R. C., IV; Cǎlmuc, F.

2010-06-01

A process for the metal recovery from electronic scrap using bacterial leaching was investigated. A mixed culture of moderately thermophilic microorganisms was enriched from acid mine drainages (AMDs) samples collected from several sulphide mines in Romania, and the bioleaching of electronic scrap was conducted both in shake flask and bioreactor. The results show that in the shake flask, the mixture can tolerate 50 g/L scrap after being acclimated to gradually increased concentrations of scrap. The copper extraction increases obviously in bioleaching of scrap with moderately thermophilic microorganisms supplemented with 0.4 g/L yeast extract at 180 r/min, 74% copper can be extracted in the pulp of 50 g/L scrap after 20 d. Compared with copper extractions of mesophilic culture, unacclimated culture and acclimated culture without addition of yeast extract, that of accliniated culture with addition of yeast extract is increased by 53%, 44% and 16%, respectively. In a completely stirred tank reactor, the mass fraction of copper and total iron extraction reach up to 81% and 56%, respectively. The results also indicate that it is necessary to add a large amount of acid to the pulp to extract copper from electronic scrap effectively.

Effect of NO2(-) on stable isotope fractionation during bacterial sulfate reduction.

PubMed

Einsiedl, Florian

2009-01-01

The effects of low NO2(-) concentrations on stable isotope fractionation during dissimilatory sulfate reduction by strain Desulfovibrio desulfuricans were investigated. Nitrite, formed as an intermediate during nitrification and denitrification processes in marine and freshwater habitats, inhibits the reduction of the sulfuroxy intermediate SO3(2-) to H2S even at low concentrations. To gain an understanding of the inhibition effect of the reduction of the sulfuroxy intermediate on stable isotope fractionation in sulfur and oxygen during bacterial sulfate reduction, nitrite was added in the form of short pulses. In the batch experiments that contained 0.02, 0.05, and 0.1 mM nitrite, sulfur enrichment factors epsilon of -12 +/- 1.6, -15 +/- 1.1, and -26 +/- 1.3 per thousand, respectively were observed. In the control experiment (no addition of nitrite) a sulfur enrichment factor epsilon of around -11 per thousand was calculated. In the experiments that contained no 18O enriched water (delta18O: -10 per thousand) and nitrite concentrations of 0.02, 0.05, and 0.1 mM, delta18O values in the remaining sulfate were fairly constant during the experiments (delta18O sulfate: approximately equal to 10 per thousand) and were similar to those obtained from the control experiment (no nitrite and no enriched water). However, in the batch experiments that contained 18O enriched water (+700 per thousand) and nitrite concentrations of 0.05 and 0.1 mM increasing delta18O values in the remaining sulfate from around 15 per thousand to approximately 65 and 85 per thousand, respectively, were found. Our experiments that contained isotopic enriched water and nitrite show clear evidence that the ratio of forward and backward fluxes regulated by adenosine-5'-phosphosulfate reductase (APSR) controls the extent of sulfur isotope fractionation during bacterial sulfate reduction in strain Desulfovibrio desulfuricans. Since the metabolic sulfuroxy intermediate SO3(2-) exchanges with water

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

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

Direct molecular testing to assess the incidence of meningococcal and other bacterial causes of meningitis among persons reported with unspecified bacterial meningitis.

PubMed

Ramautar, Arianne E; Halse, Tanya A; Arakaki, Lola; Antwi, Mike; Del Rosso, Paula; Dorsinville, Marie; Nazarian, Elizabeth; Steiner-Sichel, Linda; Lee, Lillian; Dickinson, Michelle; Wroblewski, Danielle; Dumas, Nellie; Musser, Kimberlee; Isaac, Beth; Rakeman, Jennifer; Weiss, Don

2015-11-01

Confirmed and probable cases of invasive Neisseria meningitidis (Nm) infection are reportable in New York City. We conducted a study to identify Nm among culture-negative reports of bacterial and viral meningitis. During the study period, 262 reports of suspected meningitis were eligible. Cerebrospinal fluid (CSF) specimens from 138 patients were obtained for testing. No Nm cases were detected. Results from real-time polymerase chain reaction and 16S on CSF specimens were concordant with hospital microbiology findings in 80%; however, other pathogenic organisms were detected in 14 culture-negative specimens. New York City's surveillance system appears to be effective at capturing cases of Nm meningitis. Nucleic acid testing is useful for detecting the presence of bacterial DNA when antibiotic therapy precedes lumbar puncture or bacterial cultures are negative. It remains unanswered whether culture-negative cases of Nm bacteremia are being missed by reportable disease surveillance. Copyright © 2015 Elsevier Inc. All rights reserved.

Enrichment and activity of methanotrophic microorganisms from municipal wastewater sludge.

PubMed

Siniscalchi, Luciene Alves Batista; Vale, Isabel Campante; Dell'Isola, Jéssica; Chernicharo, Carlos Augusto; Calabria Araujo, Juliana

2015-01-01

In this study, methanotrophic microorganisms were enriched from a municipal wastewater sludge taken from an Upflow Anaerobic Sludge Blanket reactor. The enrichment was performed in a sequencing batch reactor (SBR) with an autotrophic medium containing nitrite and nitrate. The microbial community composition of the inoculum and of the enrichment culture after 100 days of SBR operation was investigated and compared with the help of data obtained from 454 pyrosequencing analyses. The nitrite and nitrate removal efficiencies were 68% and 53%, respectively, probably due to heterotrophic denitrification. Archaeal cells of the anaerobic methanotrophic Archaic (ANME)-I and ANME-II groups were detected by polymerase chain reaction throughout the whole cultivation period. Pyrosequencing analysis showed that community composition was different among the two samples analysed. The dominant phyla found in the inoculum were Synergistestes, Firmicutes and Euryarchaeota, while Planctomycetes, Verrucomicrobia, Chloroflexi and Proteobacteria prevailed in the enriched biomass. The cultivation conditions decreased Methanobacterium abundance from 8% to 1%, and enriched for methanotrophic bacteria such as Methylocaldum, Methylocistis and Methylosinus. Sequences of Methylocaldum sp. accounted for 2.5% of the total reads. The presence and high predominance of Verrucomicrobia in the enriched biomass suggested that other unknown methanotrophic species related to that phylum might also have occurred in the reactor. Anaerobic methane oxidation activity was measured for both samples, and showed that the activity of the enrichment culture was nearly three times higher than the activity of the inoculum. Taken together, these results showed that the inoculum type and cultivation conditions were properly suited for methanotrophic enrichment.

Bacterial communities in floral nectar.

PubMed

Fridman, Svetlana; Izhaki, Ido; Gerchman, Yoram; Halpern, Malka

2012-02-01

Floral nectar is regarded as the most important reward available to animal-pollinated plants to attract pollinators. Despite the vast amount of publications on nectar properties, the role of nectar as a natural bacterial habitat is yet unexplored. To gain a better understanding of bacterial communities inhabiting floral nectar, culture-dependent and -independent (454-pyrosequencing) methods were used. Our findings demonstrate that bacterial communities in nectar are abundant and diverse. Using culture-dependent method we showed that bacterial communities of nectar displayed significant variation among three plant species: Amygdalus communis, Citrus paradisi and Nicotiana glauca. The dominant class in the nectar bacterial communities was Gammaproteobacteria. About half of the isolates were novel species (< 97% similarities of the 16S rRNA gene with known species). Using 454-pyrosequencing we demonstrated that nectar microbial community are distinct for each of the plant species while there are no significant differences between nectar microbial communities within nectars taken from different plants of the same species. Primary selection of the nectar bacteria is unclear; it may be affected by variations in the chemical composition of the nectar in each plant. The role of the rich and diverse nectar microflora in the attraction-repulsion relationships between the plant and its nectar consumers has yet to be explored. © 2011 Society for Applied Microbiology and Blackwell Publishing Ltd.

Enhanced green fluorescent protein in optofluidic Fabry-Perot microcavity to detect laser induced temperature changes in a bacterial culture

NASA Astrophysics Data System (ADS)

Lahoz, F.; Martín, I. R.; Walo, D.; Freire, R.; Gil-Rostra, J.; Yubero, F.; Gonzalez-Elipe, A. R.

2017-09-01

Thermal therapy using laser sources can be used in combination with other cancer therapies to eliminate tumors. However, high precision temperature control is required to avoid damage in healthy surrounding tissues. Therefore, in order to detect laser induced temperature changes, we have used the fluorescence signal of the enhanced Green Fluorescent Protein (eGFP) over-expressed in an E. coli bacterial culture. For that purpose, the bacteria expressing eGFP are injected in a Fabry-Perot (FP) optofluidic planar microcavity. In order to locally heat the bacterial culture, external infrared or ultraviolet lasers were used. Shifts in the wavelengths of the resonant FP modes are used to determine the temperature increase as a function of the heating laser pump power. Laser induced local temperature increments up to 6-7 °C were measured. These results show a relatively easy way to measure laser induced local temperature changes using a FP microcavity and using eGFP as a molecular probe instead of external nanoparticles, which could damage/alter the cell. Therefore, we believe that this approach can be of interest for the study of thermal effects in laser induced thermal therapies.

Formation of bacterial nanocells

NASA Astrophysics Data System (ADS)

Vainshtein, Mikhail; Kudryashova, Ekaterina; Suzina, Natalia; Ariskina, Elena; Voronkov, Vadim

1998-07-01

Existence of nanobacteria received increasing attention both in environmental microbiology/geomicro-biology and in medical microbiology. In order to study a production of nanoforms by typical bacterial cells. Effects of different physical factors were investigated. Treatment of bacterial cultures with microwave radiation, or culturing in field of electric current resulted in formation a few types of nanocells. The number and type of nanoforms were determined with type and dose of the treatment. The produced nanoforms were: i) globules, ii) clusters of the globules--probably produced by liaison, iii) nanocells coated with membrane. The viability of the globules is an object opened for doubts. The nanocells discovered multiplication and growth on solidified nutrient media. The authors suggest that formation of nanocells is a common response of bacteria to stress-actions produced by different agents.

Statistical optimization of culture conditions for bacterial cellulose production using Box-Behnken design.

PubMed

Bae, Sangok; Shoda, Makoto

2005-04-05

Culture conditions in a jar fermentor for bacterial cellulose (BC) production from A. xylinum BPR2001 were optimized by statistical analysis using Box-Behnken design. Response surface methodology was used to predict the levels of the factors, fructose (X1), corn steep liquor (CSL) (X2), dissolved oxygen (DO) (X3), and agar concentration (X4). Total 27 experimental runs by combination of each factor were carried out in a 10-L jar fermentor, and a three-dimensional response surface was generated to determine the effect of the factors and to find out the optimum concentration of each factor for maximum BC production and BC yield. The fructose and agar concentration highly influenced the BC production and BC yield. However, the optimum conditions according to changes in CSL and DO concentrations were predicted at almost central values of tested ranges. The predicted results showed that BC production was 14.3 g/L under the condition of 4.99% fructose, 2.85% CSL, 28.33% DO, and 0.38% agar concentration. On the other hand, BC yield was predicted in 0.34 g/g under the condition of 3.63% fructose, 2.90% CSL, 31.14% DO, and 0.42% agar concentration. Under optimized culture conditions, improvement of BC production and BC yield were experimentally confirmed, which increased 76% and 57%, respectively, compared to BC production and BC yield before optimizing the culture conditions. Copyright (c) 2005 Wiley Periodicals, Inc.

Microbial succession in response to pollutants in batch-enrichment culture

PubMed Central

Jiao, Shuo; Chen, Weimin; Wang, Entao; Wang, Junman; Liu, Zhenshan; Li, Yining; Wei, Gehong

2016-01-01

As a global problem, environmental pollution is an important factor to shape the microbial communities. The elucidation of the succession of microbial communities in response to pollutants is essential for developing bioremediation procedures. In the present study, ten batches of soil-enrichment subcultures were subjected to four treatments: phenanthrene, n-octadecane, phenanthrene + n-octadecane, or phenanthrene + n-octadecane + CdCl2. Forty pollutant-degrading consortia, corresponding to each batch of the four treatments were obtained. High-throughput sequencing of the 16S rRNA gene revealed that the diversity, richness and evenness of the consortia decreased throughout the subculturing procedure. The well-known hydrocarbon degraders Acinetobacter, Gordonia, Sphingobium, Sphingopyxis, and Castellaniella and several other genera, including Niabella and Naxibacter, were detected in the enriched consortia. The predominant microbes varied and the microbial community in the consortia gradually changed during the successive subculturing depending on the treatment, indicating that the pollutants influenced the microbial successions. Comparison of the networks in the treatments indicated that organic pollutants and CdCl2 affected the co-occurrence patterns in enriched consortia. In conclusion, single environmental factors, such as the addition of nutrients or selection pressure, can shape microbial communities and partially explain the extensive differences in microbial community structures among diverse environments. PMID:26905741

Bacterial mobilization and transport through manure enriched soils: Experiment and modeling.

PubMed

Sepehrnia, N; Memarianfard, L; Moosavi, A A; Bachmann, J; Guggenberger, G; Rezanezhad, F

2017-10-01

A precise evaluation of bacteria transport and mathematical investigations are useful for best management practices in agroecosystems. In this study, using laboratory experiments and modeling approaches, we assess the transport of bacteria released from three types of manure (cow, sheep, and poultry) to find the importance of the common manures in agricultural activities in soil and water pollution. Thirty six intact soil columns with different textures (sandy, loamy, and silty clay loam) were sampled. Fecal coliform leaching from layers of the manures on the soil surface was conducted under steady-state saturated flow conditions at 20 °C for up to four Pore Volumes (PVs). Separate leaching experiments were conducted to obtain the initial concentrations of bacteria released from the manures (Co). Influent (Co) and effluent (C) bacteria concentrations were measured by the plate-count method and the normalized concentrations (C/C0) were plotted versus PV representing the breakthrough curves (BTCs). Transport parameters were predicted using the attachment/detachment model (two-kinetic site) in HYDRUS-1D. Simulations fitted well the experimental data (R 2  = 0.50-0.96). The attachment, detachment, and straining coefficients of bacteria were more influenced by the soils treated with cow manure compared to the sheep and poultry manures. Influent curves of fecal coliforms from the manures (leached without soil) illustrated that the poultry manure had the highest potential to pollute the effluent water from the soils in term of concentration, but the BTCs and simulated data related to the treated soils illustrated that the physical shape of cow manure was more important to both straining and detachment of bacteria back into the soil solution. Detachment trends of bacteria were observed through loam and silty clay loam soils treated with cow manure compared to the cow manure enriched sandy soil. We conclude that management strategies must specifically minimize the

Dynamics of indigenous bacterial communities associated with crude oil degradation in soil microcosms during nutrient-enhanced bioremediation.

PubMed

Chikere, Chioma B; Surridge, Karen; Okpokwasili, Gideon C; Cloete, Thomas E

2012-03-01

Bacterial population dynamics were examined during bioremediation of an African soil contaminated with Arabian light crude oil and nutrient enrichment (biostimulation). Polymerase chain reaction followed by denaturing gradient gel electrophoresis (DGGE) were used to generate bacterial community fingerprints of the different treatments employing the 16S ribosomal ribonucleic acid (rRNA) gene as molecular marker. The DGGE patterns of the nutrient-amended soils indicated the presence of distinguishable bands corresponding to the oil-contaminated-nutrient-enriched soils, which were not present in the oil-contaminated and pristine control soils. Further characterization of the dominant DGGE bands after excision, reamplification and sequencing revealed that Corynebacterium spp., Dietzia spp., Rhodococcus erythropolis sp., Nocardioides sp., Low G+C (guanine plus cytosine) Gram positive bacterial clones and several uncultured bacterial clones were the dominant bacterial groups after biostimulation. Prominent Corynebacterium sp. IC10 sequence was detected across all nutrient-amended soils but not in oil-contaminated control soil. Total heterotrophic and hydrocarbon utilizing bacterial counts increased significantly in the nutrient-amended soils 2 weeks post contamination whereas oil-contaminated and pristine control soils remained fairly stable throughout the experimental period. Gas chromatographic analysis of residual hydrocarbons in biostimulated soils showed marked attenuation of contaminants starting from the second to the sixth week after contamination whereas no significant reduction in hydrocarbon peaks were seen in the oil-contaminated control soil throughout the 6-week experimental period. Results obtained indicated that nutrient amendment of oil-contaminated soil selected and enriched the bacterial communities mainly of the Actinobacteria phylogenetic group capable of surviving in toxic contamination with concomitant biodegradation of the hydrocarbons. The

Degradation of petroleum hydrocarbons and treatment of refinery wastewater under saline condition by a halophilic bacterial consortium enriched from marine environment (Red Sea), Jeddah, Saudi Arabia.

PubMed

Jamal, Mamdoh T; Pugazhendi, Arulazhagan

2018-06-01

A halophilic bacterial consortium was enriched from Red Sea saline water and sediment samples collected from Abhor, Jeddah, Saudi Arabia. The consortium potentially degraded different low (above 90% for phenanthrene and fluorene) and high (69 ± 1.4 and 56 ± 1.8% at 50 and 100 mg/L of pyrene) molecular weight polycyclic aromatic hydrocarbons (PAHs) at different concentrations under saline condition (40 g/L NaCl concentration). The cell hydrophobicity (91° ± 1°) and biosurfactant production (30 mN/m) confirmed potential bacterial cell interaction with PAHs to facilitate biodegradation process. Co-metabolic study with phenanthrene as co-substrate during pyrene degradation recorded 90% degradation in 12 days. The consortium in continuous stirred tank reactor with petroleum refinery wastewater showed complete and 90% degradation of low and high molecular weight PAHs, respectively. The reactor study also revealed 94 ± 1.8% chemical oxygen demand removal by the halophilic consortium under saline condition (40 g/L NaCl concentration). The halophilic bacterial strains present in the consortium were identified as Ochrobactrum halosaudis strain CEES1 (KX377976), Stenotrophomonas maltophilia strain CEES2 (KX377977), Achromobacter xylosoxidans strain CEES3 (KX377978) and Mesorhizobium halosaudis strain CEES4 (KX377979). Thus, the promising halophilic consortium was highly recommended to be employed in petroleum saline wastewater treatment process.

Matrix-assisted laser desorption ionization-time of flight mass spectrometry for direct bacterial identification from positive blood culture pellets.

PubMed

Prod'hom, Guy; Bizzini, Alain; Durussel, Christian; Bille, Jacques; Greub, Gilbert

2010-04-01

An ammonium chloride erythrocyte-lysing procedure was used to prepare a bacterial pellet from positive blood cultures for direct matrix-assisted laser desorption-ionization time of flight (MALDI-TOF) mass spectrometry analysis. Identification was obtained for 78.7% of the pellets tested. Moreover, 99% of the MALDI-TOF identifications were congruent at the species level when considering valid scores. This fast and accurate method is promising.

Gamma-irradiated bacterial preparation having anti-tumor activity

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

Vass, A.A.; Tyndall, R.L.; Terzaghi-Howe, P.

1999-11-16

This application describes a bacterial preparation from Pseudomonas species isolated {number{underscore}sign}15 ATCC 55638 that has been exposed to gamma radiation exhibits cytotoxicity that is specific for neoplastic carcinoma cells. A method for obtaining a bacterial preparation having antitumor activity consists of suspending a bacterial isolate in media and exposing the suspension to gamma radiation. A bacterial preparation of an aged culture of an amoeba-associated bacteria exhibits anti-reverse transcriptase activity. A method for obtaining a bacterial preparation having anti-reverse transcriptase activity from an amoeba-associated bacterial isolate grown to stationary phase is disclosed.

Enhanced methane production via repeated batch bioaugmentation pattern of enriched microbial consortia.

PubMed

Yang, Zhiman; Guo, Rongbo; Xu, Xiaohui; Wang, Lin; Dai, Meng

2016-09-01

Using batch and repeated batch cultivations, this study investigated the effects of bioaugmentation with enriched microbial consortia (named as EMC) on methane production from effluents of hydrogen-producing stage of potato slurry, as well as on the indigenous bacterial community. The results demonstrated that the improved methane production and shift of the indigenous bacterial community structure were dependent on the EMC/sludge ratio and bioaugmentation patterns. The methane yield and production rate in repeated batch bioaugmentation pattern of EMC were, respectively, average 15% and 10% higher than in one-time bioaugmentation pattern of EMC. DNA-sequencing approach showed that the enhanced methane production in the repeated batch bioaugmentation pattern of EMC mainly resulted from the enriched iron-reducing bacteria and the persistence of the introduced Syntrophomonas, which led to a rapid degradation of individual VFAs to methane. The findings contributed to understanding the correlation between the bioaugmentation of microbial consortia, community shift, and methane production. Copyright © 2016 Elsevier Ltd. All rights reserved.

Early Recovery of Salmonella from Food Using a 6-Hour Non-selective Pre-enrichment and Reformulation of Tetrathionate Broth.

PubMed

Daquigan, Ninalynn; Grim, Christopher J; White, James R; Hanes, Darcy E; Jarvis, Karen G

2016-01-01

Culture based methods are commonly employed to detect pathogens in food and environmental samples. These methods are time consuming and complex, requiring multiple non-selective and selective enrichment broths, and usually take at least 1 week to recover and identify pathogens. Improving pathogen detection in foods is a primary goal for regulatory agencies and industry. Salmonella detection in food relies on a series of culture steps in broth formulations optimized to resuscitate Salmonella and reduce the abundance of competitive bacteria. Examples of non-selective pre-enrichment broths used to isolate Salmonella from food include Lactose, Universal Pre-enrichment, BPW, and Trypticase Soy broths. Tetrathionate (TT) and Rappaport-Vassiliadis (RV) broths are employed after a 24-h non-selective enrichment to select for Salmonella and hamper the growth of competitive bacteria. In this study, we tested a new formulation of TT broth that lacks brilliant green dye and has lower levels of TT . We employed this TT broth formulation in conjunction with a 6-h non-selective pre-enrichment period and determined that Salmonella recovery was possible one day earlier than standard food culture methods. We tested the shortened culture method in different non-selective enrichment broths, enumerated Salmonella in the non-selective enrichments, and used 16S rRNA gene sequencing to determine the proportional abundances of Salmonella in the TT and RV selective enrichments. Together these data revealed that a 6-h non-selective pre-enrichment reduces the levels of competitive bacteria inoculated into the selective TT and RV broths, enabling the recovery of Salmonella 1 day earlier than standard culture enrichment methods.

Two decades of warming increases diversity of a potentially lignolytic bacterial community

PubMed Central

Pold, Grace; Melillo, Jerry M.; DeAngelis, Kristen M.

2015-01-01

As Earth's climate warms, the massive stores of carbon found in soil are predicted to become depleted, and leave behind a smaller carbon pool that is less accessible to microbes. At a long-term forest soil-warming experiment in central Massachusetts, soil respiration and bacterial diversity have increased, while fungal biomass and microbially-accessible soil carbon have decreased. Here, we evaluate how warming has affected the microbial community's capability to degrade chemically-complex soil carbon using lignin-amended BioSep beads. We profiled the bacterial and fungal communities using PCR-based methods and completed extracellular enzyme assays as a proxy for potential community function. We found that lignin-amended beads selected for a distinct community containing bacterial taxa closely related to known lignin degraders, as well as members of many genera not previously noted as capable of degrading lignin. Warming tended to drive bacterial community structure more strongly in the lignin beads, while the effect on the fungal community was limited to unamended beads. Of those bacterial operational taxonomic units (OTUs) enriched by the warming treatment, many were enriched uniquely on lignin-amended beads. These taxa may be contributing to enhanced soil respiration under warming despite reduced readily available C availability. In aggregate, these results suggest that there is genetic potential for chemically complex soil carbon degradation that may lead to extended elevated soil respiration with long-term warming. PMID:26042112

Trace Elements Affect Methanogenic Activity and Diversity in Enrichments from Subsurface Coal Bed Produced Water

PubMed Central

Ünal, Burcu; Perry, Verlin Ryan; Sheth, Mili; Gomez-Alvarez, Vicente; Chin, Kuk-Jeong; Nüsslein, Klaus

2012-01-01

Microbial methane from coal beds accounts for a significant and growing percentage of natural gas worldwide. Our knowledge of physical and geochemical factors regulating methanogenesis is still in its infancy. We hypothesized that in these closed systems, trace elements (as micronutrients) are a limiting factor for methanogenic growth and activity. Trace elements are essential components of enzymes or cofactors of metabolic pathways associated with methanogenesis. This study examined the effects of eight trace elements (iron, nickel, cobalt, molybdenum, zinc, manganese, boron, and copper) on methane production, on mcrA transcript levels, and on methanogenic community structure in enrichment cultures obtained from coal bed methane (CBM) well produced water samples from the Powder River Basin, Wyoming. Methane production was shown to be limited both by a lack of additional trace elements as well as by the addition of an overly concentrated trace element mixture. Addition of trace elements at concentrations optimized for standard media enhanced methane production by 37%. After 7 days of incubation, the levels of mcrA transcripts in enrichment cultures with trace element amendment were much higher than in cultures without amendment. Transcript levels of mcrA correlated positively with elevated rates of methane production in supplemented enrichments (R2 = 0.95). Metabolically active methanogens, identified by clone sequences of mcrA mRNA retrieved from enrichment cultures, were closely related to Methanobacterium subterraneum and Methanobacterium formicicum. Enrichment cultures were dominated by M. subterraneum and had slightly higher predicted methanogenic richness, but less diversity than enrichment cultures without amendments. These results suggest that varying concentrations of trace elements in produced water from different subsurface coal wells may cause changing levels of CBM production and alter the composition of the active methanogenic community. PMID

Direct identification of bacteria in blood culture by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry: a new methodological approach.

PubMed

Kroumova, Vesselina; Gobbato, Elisa; Basso, Elisa; Mucedola, Luca; Giani, Tommaso; Fortina, Giacomo

2011-08-15

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) has recently been demonstrated to be a powerful tool for the rapid identification of bacteria from growing colonies. In order to speed up the identification of bacteria, several authors have evaluated the usefulness of this MALDI-TOF MS technology for the direct and quick identification bacteria from positive blood cultures. The results obtained so far have been encouraging but have also shown some limitations, mainly related to the bacterial growth and to the presence of interference substances belonging to the blood cultures. In this paper, we present a new methodological approach that we have developed to overcome these limitations, based mainly on an enrichment of the sample into a growing medium before the extraction process, prior to mass spectrometric analysis. The proposed method shows important advantages for the identification of bacterial strains, yielding an increased identification score, which gives higher confidence in the results. Copyright © 2011 John Wiley & Sons, Ltd.

Identification of Viable Helicobacter pylori in Drinking Water Supplies by Cultural and Molecular Techniques.

PubMed

Santiago, Paula; Moreno, Yolanda; Ferrús, M Antonía

2015-08-01

Helicobacter pylori is one of the most common causes of chronic bacterial infection in humans, directly related to peptic ulcer and gastric cancer. It has been suggested that H. pylori can be acquired through different transmission routes, including water. In this study, culture and qPCR were used to detect and identify the presence of H. pylori in drinking water. Furthermore, the combined techniques PMA-qPCR and DVC-FISH were applied for detection of viable cells of H. pylori. Among 24 drinking water samples, 16 samples were positive for the presence of H. pylori, but viable cells were only detected in six samples. Characteristic colonies, covered by a mass of bacterial unspecific growth, were observed on selective agar plates from an only sample, after enrichment. The mixed culture was submitted to DVC-FISH and qPCR analysis, followed by sequencing of the amplicons. Molecular techniques confirmed the growth of H. pylori on the agar plate. Our results demonstrate for the first time that H. pylori can survive and be potentially infective in drinking water, showing that water distribution systems could be a potential route for H. pylori transmission. © 2015 John Wiley & Sons Ltd.

Characterization of rumen bacterial strains isolated from enrichments of rumen content in the presence of propolis.

PubMed

de Aguiar, Sílvia Cristina; Zeoula, Lucia Maria; do Prado, Odimari Pricila Pires; Arcuri, Pedro Braga; Forano, Evelyne

2014-11-01

Propolis presents many biological properties, including antibacterial activities, and has been proposed as an additive in ruminant nutrition. Twenty bacterial strains, previously isolated from enrichments of Brazilian cow rumen contents in the presence of different propolis extracts (LLOS), were characterized using phenotyping and 16S rRNA identification. Seven strains were assigned to Streptococcus sp., most likely S. bovis, and were all degrading starch. One amylolytic lactate-utilizing strain of Selenomonas ruminantium was also found. Two strains of Clostridium bifermentans were identified and showed proteolytic activity. Two strains were assigned to Mitsuokella jalaludinii and were saccharolytic. One strain belonged to a Bacillus species and seven strains were affiliated with Escherichia coli. All of the 20 strains were able to use many sugars, but none of them were able to degrade the polysaccharides carboxymethylcellulose and xylans. The effect of three propolis extracts (LLOS B1, C1 and C3) was tested on the in vitro growth of four representative isolates of S. bovis, E. coli, M. jalaludinii and C. bifermentans. The growth of S. bovis, E. coli and M. jalaludinii was not affected by the three propolis extracts at 1 mg ml(-1). C. bifermentans growth was completely inhibited at this LLOS concentration, but this bacterium was partially resistant at lower concentrations. LLOS C3, with the lower concentration of phenolic compounds, was a little less inhibitory than B1 and C1 on this strain.

How to use: bacterial cultures in diagnosing lower respiratory tract infections in cystic fibrosis.

PubMed

Ahmed, Bushra; Bush, Andrew; Davies, Jane C

2014-10-01

Respiratory infections are the leading cause of morbidity and mortality in cystic fibrosis. Certain bacteria, such as Pseudomonas aeruginosa, are associated with a worse clinical outcome than others, but can be completely eradicated if identified and treated early. The diagnosis of lower respiratory tract infections can be challenging in the non-expectorating patient, in whom upper airway samples, such as cough swabs, are a surrogate for lower airway sampling. However, the results of these often do not fit with the clinical picture, presenting a management dilemma. Frequently, clinicians are faced with a negative culture result in a progressively symptomatic patient and vice versa. When judging the clinical significance of a positive upper airway culture result in an asymptomatic patient, it is important to consider the prognostic significance of the organism cultured. Given that the reported sensitivity of upper airway swabs (which includes throat swabs) is variable, ranging from 35.7% to 71% for Pseudomonas aeruginosa, 50% to 86% for Staphylococcus aureus and 11% to 92% for Haemophilus influenza, upper airway samples may fail to identify lower airway infections. Therefore, in symptomatic children, a repeatedly negative upper airway swab should not be considered as reassuring, and alternative sampling methods, such as induced sputum or bronchoalveolar lavage, should be considered. Here we use some examples of common scenarios to illustrate how best to use bacterial cultures to aid management decisions in cystic fibrosis. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.

Coexisting bacterial populations responsible for multiphasic mineralization kinetics in soil. [Janthinobacterium sp. Rhodococcus sp

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

Schmidt, S.K.; Gier, M.J.

1990-09-01

Experiments were conducted to study populations of indigenous microorganisms capable of mineralizing 2,4-dinitrophenol (DNP) in two soils. Previous kinetic analyses indicated the presence of two coexisting populations of DNP-mineralizing microorganisms in a forest soil (soil 1). Studies in which eucaryotic and procaryotic inhibitors were added to this soil indicated that both populations were bacterial. Most-probable-number counts with media containing different concentrations of DNP indicated that more bacteria could mineralize low concentrations of DNP than could metabolize high concentrations of it. Enrichments with varying concentrations of DNP and various combinations of inhibitors consistently resulted in the isolation of the same twomore » species of bacteria from soil 1. This soil contained a large number and variety of fungi, but no fungi capable of mineralizing DNP were isolated. The two bacterial isolates were identified as a Janthinobacterium sp. and a Rhodococcus sp. The Janthinobacterium sp. had a low {mu}{sub max} and a low K{sub m} for DNP mineralization, whereas the Rhodococcus sp. had much higher values for both parameters. These differences between the two species of bacteria were similar to differences seen when soil was incubated with different concentrations of DNP. Values for {mu}{sub max} from soil incubations were similar to {mu}{sub max} values obtained in pure culture studies. In contrast, K{sub s} and K{sub m} values showed greater variation between soil and pure culture studies.« less

Comprehensive Evaluation of the MBT STAR-BL Module for Simultaneous Bacterial Identification and β-Lactamase-Mediated Resistance Detection in Gram-Negative Rods from Cultured Isolates and Positive Blood Cultures.

PubMed

Lee, Annie W T; Lam, Johnson K S; Lam, Ricky K W; Ng, Wan H; Lee, Ella N L; Lee, Vicky T Y; Sze, Po P; Rajwani, Rahim; Fung, Kitty S C; To, Wing K; Lee, Rodney A; Tsang, Dominic N C; Siu, Gilman K H

2018-01-01

Objective: This study evaluated the capability of a MALDI Biotyper system equipped with the newly introduced MBT STAR-BL module to simultaneously perform species identification and β-lactamase-mediated resistance detection in bacteremia -causing bacteria isolated from cultured isolates and patient-derived blood cultures (BCs). Methods: Two hundred retrospective cultured isolates and 153 prospective BCs containing Gram-negative rods (GNR) were collected and subjected to direct bacterial identification, followed by the measurement of β-lactamase activities against ampicillin, piperacillin, cefotaxime, ceftazidime, and meropenem using the MBT STAR-BL module. The results and turnaround times were compared with those of routine microbiological processing. All strains were also characterized by beta-lactamase PCR and sequencing. Results: Using the saponin-based extraction method, MALDI-TOF MS correctly identified bacteria in 116/134 (86.6%) monomicrobial BCs. The detection sensitivities for β-lactamase activities against ampicillin, piperacillin, third-generation cephalosporin and meropenem were 91.3, 100, 97.9, and 100% for cultured isolates, and 80.4, 100, 68.8, and 40% for monomicrobial BCs ( n = 134) respectively. The overall specificities ranged from 91.5 to 100%. Furthermore, the MBT STAR-BL and conventional drug susceptibility test results were concordant in 14/19 (73.7%) polymicrobial cultures. Reducing the logRQ cut-off value from 0.4 to 0.2 increased the direct detection sensitivities for β-lactamase activities against ampicillin, cefotaxime and meropenem in BCs to 85.7, 87.5, and 100% respectively. The MBT STAR-BL test enabled the reporting of β-lactamase-producing GNR at 14.16 and 47.64 h before the interim and final reports of routine BCs processing, respectively, were available. Conclusion: The MALDI Biotyper system equipped with the MBT STAR-BL module enables the simultaneous rapid identification of bacterial species and �

Comprehensive Evaluation of the MBT STAR-BL Module for Simultaneous Bacterial Identification and β-Lactamase-Mediated Resistance Detection in Gram-Negative Rods from Cultured Isolates and Positive Blood Cultures

PubMed Central

Lee, Annie W. T.; Lam, Johnson K. S.; Lam, Ricky K. W.; Ng, Wan H.; Lee, Ella N. L.; Lee, Vicky T. Y.; Sze, Po P.; Rajwani, Rahim; Fung, Kitty S. C.; To, Wing K.; Lee, Rodney A.; Tsang, Dominic N. C.; Siu, Gilman K. H.

2018-01-01

Objective: This study evaluated the capability of a MALDI Biotyper system equipped with the newly introduced MBT STAR-BL module to simultaneously perform species identification and β-lactamase-mediated resistance detection in bacteremia -causing bacteria isolated from cultured isolates and patient-derived blood cultures (BCs). Methods: Two hundred retrospective cultured isolates and 153 prospective BCs containing Gram-negative rods (GNR) were collected and subjected to direct bacterial identification, followed by the measurement of β-lactamase activities against ampicillin, piperacillin, cefotaxime, ceftazidime, and meropenem using the MBT STAR-BL module. The results and turnaround times were compared with those of routine microbiological processing. All strains were also characterized by beta-lactamase PCR and sequencing. Results: Using the saponin-based extraction method, MALDI-TOF MS correctly identified bacteria in 116/134 (86.6%) monomicrobial BCs. The detection sensitivities for β-lactamase activities against ampicillin, piperacillin, third-generation cephalosporin and meropenem were 91.3, 100, 97.9, and 100% for cultured isolates, and 80.4, 100, 68.8, and 40% for monomicrobial BCs (n = 134) respectively. The overall specificities ranged from 91.5 to 100%. Furthermore, the MBT STAR-BL and conventional drug susceptibility test results were concordant in 14/19 (73.7%) polymicrobial cultures. Reducing the logRQ cut-off value from 0.4 to 0.2 increased the direct detection sensitivities for β-lactamase activities against ampicillin, cefotaxime and meropenem in BCs to 85.7, 87.5, and 100% respectively. The MBT STAR-BL test enabled the reporting of β-lactamase-producing GNR at 14.16 and 47.64 h before the interim and final reports of routine BCs processing, respectively, were available. Conclusion: The MALDI Biotyper system equipped with the MBT STAR-BL module enables the simultaneous rapid identification of bacterial species and �

Culturable bacterial communities associated to Brazilian Oscarella species (Porifera: Homoscleromorpha) and their antagonistic interactions.

PubMed

Laport, Marinella Silva; Bauwens, Mathieu; de Oliveira Nunes, Suzanne; Willenz, Philippe; George, Isabelle; Muricy, Guilherme

2017-04-01

Sponges offer an excellent model to investigate invertebrate-microorganism interactions. Furthermore, bacteria associated with marine sponges represent a rich source of bioactive metabolites. The aim of this study was to characterize the bacteria inhabiting a genus of sponges, Oscarella, and their potentiality for antimicrobial production. Bacterial isolates were recovered from different Oscarella specimens, among which 337 were phylogenetically identified. The culturable community was dominated by Proteobacteria and Firmicutes, and Vibrio was the most frequently isolated genus, followed by Shewanella. When tested for antimicrobial production, bacteria of the 12 genera isolated were capable of producing antimicrobial substances. The majority of strains were involved in antagonistic interactions and inhibitory activities were also observed against bacteria of medical importance. It was more pronounced in some isolated genera (Acinetobacter, Bacillus, Photobacterium, Shewanella and Vibrio). These findings suggest that chemical antagonism could play a significant role in shaping bacterial communities within Oscarella, a genus classified as low-microbial abundance sponge. Moreover, the identified strains may contribute to the search for new sources of antimicrobial substances, an important strategy for developing therapies to treat infections caused by multidrug-resistant bacteria. This study was the first to investigate the diversity and antagonistic activity of bacteria isolated from Oscarella spp. It highlights the biotechnological potential of sponge-associated bacteria.

Integrated analysis of bacterial and microeukaryotic communities from differentially active mud volcanoes in the Gulf of Cadiz

NASA Astrophysics Data System (ADS)

Coelho, Francisco J. R. C.; Louvado, António; Domingues, Patrícia M.; Cleary, Daniel F. R.; Ferreira, Marina; Almeida, Adelaide; Cunha, Marina R.; Cunha, Ângela; Gomes, Newton C. M.

2016-10-01

The present study assesses the diversity and composition of sediment bacterial and microeukaryotic communities from deep-sea mud volcanoes (MVs) associated with strike-slip faults in the South-West Iberian Margin (SWIM). We used a 16S/18S rRNA gene based pyrosequencing approach to characterize and correlate the sediment bacterial and microeukaryotic communities from MVs with differing gas seep regimes and from an additional site with no apparent seeping activity. In general, our results showed significant compositional changes of bacterial and microeukaryotic communities in sampling sites with different seepage regimes. Sediment bacterial communities were enriched with Methylococcales (putative methanotrophs) but had lower abundances of Rhodospirillales, Nitrospirales and SAR202 in the more active MVs. Within microeukaryotic communities, members of the Lobosa (lobose amoebae) were enriched in more active MVs. We also showed a strong correlation between Methylococcales populations and lobose amoeba in active MVs. This study provides baseline information on the diversity and composition of bacterial and microeukaryotic communities in deep-sea MVs associated with strike-slip faults.

Integrated analysis of bacterial and microeukaryotic communities from differentially active mud volcanoes in the Gulf of Cadiz

PubMed Central

Coelho, Francisco J. R. C.; Louvado, António; Domingues, Patrícia M.; Cleary, Daniel F. R.; Ferreira, Marina; Almeida, Adelaide; Cunha, Marina R.; Cunha, Ângela; Gomes, Newton C. M.