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Sample records for monomorphic bacteria dna

  1. Phylogenetic Analysis of Geographically Diverse Radopholus similis via rDNA Sequence Reveals a Monomorphic Motif

    PubMed Central

    Kaplan, D. T.; Thomas, W. K.; Frisse, L. M.; Sarah, J. L.; Stanton, J. M.; Speijer, P. R.; Marin, D. H.; Opperman, C. H.

    2000-01-01

    The nucleic acid sequences of rDNA ITS1 and the rDNA D2/D3 expansion segment were compared for 57 burrowing nematode isolates collected from Australia, Cameroon, Central America, Cuba, Dominican Republic, Florida, Guadeloupe, Hawaii, Nigeria, Honduras, Indonesia, Ivory Coast, Puerto Rico, South Africa, and Uganda. Of the 57 isolates, 55 were morphologically similar to Radopholus similis and seven were citrus-parasitic. The nucleic acid sequences for PCR-amplified ITS1 and for the D2/D3 expansion segment of the 28S rDNA gene were each identical for all putative R. similis. Sequence divergence for both the ITS1 and the D2/D3 was concordant with morphological differences that distinguish R. similis from other burrowing nematode species. This result substantiates previous observations that the R. similis genome is highly conserved across geographic regions. Autapomorphies that would delimit phylogenetic lineages of non-citrus-parasitic R. similis from those that parasitize citrus were not observed. The data presented herein support the concept that R. similis is comprised of two pathotypes-one that parasitizes citrus and one that does not. PMID:19270959

  2. DNA Uptake by Transformable Bacteria

    SciTech Connect

    Lacks, Sanford A.

    1999-03-31

    The various processes of DNA uptake by cells can be categorized as: viral DNA entry, conjugation, or transformation. Within each category, a variety of mechanisms have been found. However, considerable similarities occur among the different mechanisms of conjugation and, especially, transformation. All of these natural mechanisms of DNA transfer are quite elaborate and involve multiple protein components, as the case may be, of the virus, the donor cell, and the recipient cell. The mechanisms of viral infection and conjugation will be discussed mainly with respect to their relevance to transformation.

  3. DNA UPTAKE BY TRANSFORMABLE BACTERIA

    SciTech Connect

    LACKS,S.A.

    1999-09-07

    The various processes of DNA uptake by cells can be categorized as: viral DNA entry, conjugation, or transformation. Within each category, a variety of mechanisms have been found. However, considerable similarities occur among the different mechanisms of conjugation and, especially, transformation. All of these natural mechanisms of DNA transfer are quite elaborate and involve multiple protein components, as the case may be, of the virus, the donor cell, and the recipient cell. The mechanisms of viral infection and conjugation will be discussed mainly with respect to their relevance to transformation.

  4. Carcinomas ex monomorphic adenoma of salivary glands.

    PubMed

    Luna, M A; Batsakis, J G; Tortoledo, M E; del Junco, G W

    1989-08-01

    A clinicopathological analysis of eight examples of carcinomas arising from salivary gland monomorphic adenomas, carcinomas ex monomorphic adenoma, is presented. These uncommon to rare neoplasms have a predilection for the parotid glands, are diagnosed about a decade later than their benign precursors, and most often arise from the dermal analogue type of monomorphic adenoma. As judged by follow-up periods of two to 16 years, carcinomas ex monomorphic adenoma are locally aggressive neoplasms with the clinical course marred by recurrences but without regional or distant metastases.

  5. DNA methyltransferases and epigenetic regulation in bacteria.

    PubMed

    Adhikari, Satish; Curtis, Patrick D

    2016-09-01

    Epigenetics is a change in gene expression that is heritable without a change in DNA sequence itself. This phenomenon is well studied in eukaryotes, particularly in humans for its role in cellular differentiation, X chromosome inactivation and diseases like cancer. However, comparatively little is known about epigenetic regulation in bacteria. Bacterial epigenetics is mainly present in the form of DNA methylation where DNA methyltransferases add methyl groups to nucleotides. This review focuses on two methyltransferases well characterized for their roles in gene regulation: Dam and CcrM. Dam methyltransferase in Escherichia coli is important for expression of certain genes such as the pap operon, as well as other cellular processes like DNA replication initiation and DNA repair. In Caulobacter crescentus and other Alphaproteobacteria, the methyltransferase CcrM is cell cycle regulated and is involved in the cell-cycle-dependent regulation of several genes. The diversity of regulatory targets as well as regulatory mechanisms suggests that gene regulation by methylation could be a widespread and potent method of regulation in bacteria. PMID:27476077

  6. Transfer of DNA from Bacteria to Eukaryotes.

    PubMed

    Lacroix, Benoît; Citovsky, Vitaly

    2016-01-01

    Historically, the members of the Agrobacterium genus have been considered the only bacterial species naturally able to transfer and integrate DNA into the genomes of their eukaryotic hosts. Yet, increasing evidence suggests that this ability to genetically transform eukaryotic host cells might be more widespread in the bacterial world. Indeed, analyses of accumulating genomic data reveal cases of horizontal gene transfer from bacteria to eukaryotes and suggest that it represents a significant force in adaptive evolution of eukaryotic species. Specifically, recent reports indicate that bacteria other than Agrobacterium, such as Bartonella henselae (a zoonotic pathogen), Rhizobium etli (a plant-symbiotic bacterium related to Agrobacterium), or even Escherichia coli, have the ability to genetically transform their host cells under laboratory conditions. This DNA transfer relies on type IV secretion systems (T4SSs), the molecular machines that transport macromolecules during conjugative plasmid transfer and also during transport of proteins and/or DNA to the eukaryotic recipient cells. In this review article, we explore the extent of possible transfer of genetic information from bacteria to eukaryotic cells as well as the evolutionary implications and potential applications of this transfer. PMID:27406565

  7. Transfer of DNA from Bacteria to Eukaryotes

    PubMed Central

    2016-01-01

    ABSTRACT Historically, the members of the Agrobacterium genus have been considered the only bacterial species naturally able to transfer and integrate DNA into the genomes of their eukaryotic hosts. Yet, increasing evidence suggests that this ability to genetically transform eukaryotic host cells might be more widespread in the bacterial world. Indeed, analyses of accumulating genomic data reveal cases of horizontal gene transfer from bacteria to eukaryotes and suggest that it represents a significant force in adaptive evolution of eukaryotic species. Specifically, recent reports indicate that bacteria other than Agrobacterium, such as Bartonella henselae (a zoonotic pathogen), Rhizobium etli (a plant-symbiotic bacterium related to Agrobacterium), or even Escherichia coli, have the ability to genetically transform their host cells under laboratory conditions. This DNA transfer relies on type IV secretion systems (T4SSs), the molecular machines that transport macromolecules during conjugative plasmid transfer and also during transport of proteins and/or DNA to the eukaryotic recipient cells. In this review article, we explore the extent of possible transfer of genetic information from bacteria to eukaryotic cells as well as the evolutionary implications and potential applications of this transfer. PMID:27406565

  8. [Monomorphic post-transplant T-lymphoproliferative disorder after autologous stem cell transplantation for multiple myeloma].

    PubMed

    Ishikawa, Tetsuya; Shimizu, Hiroaki; Takei, Toshifumi; Koya, Hiroko; Iriuchishima, Hirono; Hosiho, Takumi; Hirato, Junko; Kojima, Masaru; Handa, Hiroshi; Nojima, Yoshihisa; Murakami, Hirokazu

    2016-01-01

    We report a rare case of T cell type monomorphic post-transplant lymphoproliferative disorders (PTLD) after autologous stem cell transplantation. A 53-year-old man with multiple myeloma received autologous stem cell transplantation and achieved a very good partial response. Nine months later, he developed a high fever and consciousness disturbance, and had multiple swollen lymph nodes and a high titer of Epstein-Barr (EB) virus DNA in his peripheral blood. Neither CT nor MRI of the brain revealed any abnormalities. Cerebrospinal fluid contained no malignant cells, but the EB virus DNA titer was high. Lymph node biopsy revealed T cell type monomorphic PTLD. Soon after high-dose treatment with methotrexate and cytosine arabinoside, the high fever and consciousness disturbance subsided, and the lymph node swelling and EB virus DNA disappeared. Given the efficacy of chemotherapy in this case, we concluded that the consciousness disturbance had been induced by central nervous system involvement of monomorphic PTLD. PMID:26861102

  9. [Monomorphic post-transplant T-lymphoproliferative disorder after autologous stem cell transplantation for multiple myeloma].

    PubMed

    Ishikawa, Tetsuya; Shimizu, Hiroaki; Takei, Toshifumi; Koya, Hiroko; Iriuchishima, Hirono; Hosiho, Takumi; Hirato, Junko; Kojima, Masaru; Handa, Hiroshi; Nojima, Yoshihisa; Murakami, Hirokazu

    2016-01-01

    We report a rare case of T cell type monomorphic post-transplant lymphoproliferative disorders (PTLD) after autologous stem cell transplantation. A 53-year-old man with multiple myeloma received autologous stem cell transplantation and achieved a very good partial response. Nine months later, he developed a high fever and consciousness disturbance, and had multiple swollen lymph nodes and a high titer of Epstein-Barr (EB) virus DNA in his peripheral blood. Neither CT nor MRI of the brain revealed any abnormalities. Cerebrospinal fluid contained no malignant cells, but the EB virus DNA titer was high. Lymph node biopsy revealed T cell type monomorphic PTLD. Soon after high-dose treatment with methotrexate and cytosine arabinoside, the high fever and consciousness disturbance subsided, and the lymph node swelling and EB virus DNA disappeared. Given the efficacy of chemotherapy in this case, we concluded that the consciousness disturbance had been induced by central nervous system involvement of monomorphic PTLD.

  10. Monomorphic adenoma, canalicular variant: report of case.

    PubMed

    Wiener, A P; Meadows, F

    1977-05-01

    A case of monomorphic adenoma, canalicular variant, has been presented. This lesion is a rare benign neoplasm most often found in the minor salivary glands of the upper lip. This appears to be the first reported case of the lesion in a non-Caucasian.

  11. Liquid-Crystalline Mesophases of Plasmid DNA in Bacteria

    NASA Astrophysics Data System (ADS)

    Reich, Ziv; Wachtel, Ellen J.; Minsky, Abraham

    1994-06-01

    Bacterial plasmids may often reach a copy number larger than 1000 per cell, corresponding to a total amount of DNA that may exceed the amount of DNA within the bacterial chromosome. This observation highlights the problem of cellular accommodation of large amounts of closed-circular nucleic acids, whose interwound conformation offers negligible DNA compaction. As determined by x-ray scattering experiments conducted on intact bacteria, supercoiled plasmids segregate within the cells into dense clusters characterized by a long-range order. In vitro studies performed at physiological DNA concentrations indicated that interwound DNA spontaneously forms liquid crystalline phases whose macroscopic structural properties are determined by the features of the molecular supercoiling. Because these features respond to cellular factors, DNA supercoiling may provide a sensitive regulatory link between cellular parameters and the packaging modes of interwound DNA in vivo.

  12. Ultrasound-mediated DNA transfer for bacteria.

    PubMed

    Song, Yizhi; Hahn, Thomas; Thompson, Ian P; Mason, Timothy J; Preston, Gail M; Li, Guanghe; Paniwnyk, Larysa; Huang, Wei E

    2007-01-01

    In environmental microbiology, the most commonly used methods of bacterial DNA transfer are conjugation and electroporation. However, conjugation requires physical contact and cell-pilus-cell interactions; electroporation requires low-ionic strength medium and high voltage. These limitations have hampered broad applications of bacterial DNA delivery. We have employed a standard low frequency 40 kHz ultrasound bath to successfully transfer plasmid pBBR1MCS2 into Pseudomonas putida UWC1, Escherichia coli DH5alpha and Pseudomonas fluorescens SBW25 with high efficiency. Under optimal conditions: ultrasound exposure time of 10 s, 50 mM CaCl(2), temperature of 22 degrees C, plasmid concentration of 0.8 ng/microl, P. putida UWC1 cell concentration of 2.5 x 10(9) CFU (colony forming unit)/ml and reaction volume of 500 microl, the efficiency of ultrasound DNA delivery (UDD) was 9.8 +/- 2.3 x 10(-6) transformants per cell, which was nine times more efficient than conjugation, and even four times greater than electroporation. We have also transferred pBBR1MCS2 into E. coli DH5alpha and P. fluorescens SBW25 with efficiencies of 1.16 +/- 0.13 x 10(-6) and 4.33 +/- 0.78 x 10(-6) transformants per cell, respectively. Low frequency UDD can be readily scaled up, allowing for the application of UDD not only in laboratory conditions but also on an industrial scale. PMID:17890732

  13. Failsafe mechanisms couple division and DNA replication in bacteria

    PubMed Central

    Arjes, Heidi A.; Kriel, Allison; Sorto, Nohemy A.; Shaw, Jared T.; Wang, Jue D.; Levin, Petra Anne

    2014-01-01

    Summary The past twenty years have seen tremendous advances in our understanding of the mechanisms underlying bacterial cytokinesis, particularly the composition of the division machinery and the factors controlling its assembly [1]. At the same time, we understand very little about the relationship between cell division and other cell cycle events in bacteria. Here we report that inhibiting division in Bacillus subtilis and Staphylococcus aureus quickly leads to an arrest in the initiation of new rounds of DNA replication followed by a complete arrest in cell growth. Arrested cells are metabolically active but unable to initiate new rounds of either DNA replication or division when shifted to permissive conditions. Inhibiting DNA replication results in entry into a similar quiescent state, in which cells are unable to resume growth or division when returned to permissive conditions. Our data suggest the presence of two failsafe mechanisms: one linking division to the initiation of DNA replication and another linking the initiation of DNA replication to division. These findings contradict the prevailing view of the bacterial cell cycle as a series of coordinated but uncoupled events. Importantly, the terminal nature of the cell cycle arrest validates the bacterial cell cycle machinery as an effective target for antimicrobial development. PMID:25176632

  14. DNA Integrity and Shock Wave Transformation Efficiency of Bacteria and Fungi

    NASA Astrophysics Data System (ADS)

    Loske, Achim M.; Campos-Guillén, Juan; Fernández, Francisco; Pastrana, Xóchitl; Magaña-Ortíz, Denis; Coconi-Linares, Nancy; Ortíz-Vázquez, Elizabeth; Gómez-Lim, Miguel

    Delivery of DNA into bacteria and fungi is essential in medicine and biotechnology to produce metabolites, enzymes, antibiotics and proteins. So far, protocols to genetically transform bacteria and fungi are inefficient and have low reproducibility.

  15. Social amoebae trap and kill bacteria by casting DNA nets

    PubMed Central

    Zhang, Xuezhi; Zhuchenko, Olga; Kuspa, Adam; Soldati, Thierry

    2016-01-01

    Extracellular traps (ETs) from neutrophils are reticulated nets of DNA decorated with anti-microbial granules, and are capable of trapping and killing extracellular pathogens. Various phagocytes of mammals and invertebrates produce ETs, however, the evolutionary history of this DNA-based host defence strategy is unclear. Here we report that Sentinel (S) cells of the multicellular slug stage of the social amoeba Dictyostelium discoideum produce ETs upon stimulation with bacteria or lipopolysaccharide in a reactive oxygen species-dependent manner. The production of ETs by S cells requires a Toll/Interleukin-1 receptor domain-containing protein TirA and reactive oxygen species-generating NADPH oxidases. Disruption of these genes results in decreased clearance of bacterial infections. Our results demonstrate that D. discoideum is a powerful model organism to study the evolution and conservation of mechanisms of cell-intrinsic immunity, and suggest that the origin of DNA-based ETs as an innate immune defence predates the emergence of metazoans. PMID:26927887

  16. Social amoebae trap and kill bacteria by casting DNA nets.

    PubMed

    Zhang, Xuezhi; Zhuchenko, Olga; Kuspa, Adam; Soldati, Thierry

    2016-01-01

    Extracellular traps (ETs) from neutrophils are reticulated nets of DNA decorated with anti-microbial granules, and are capable of trapping and killing extracellular pathogens. Various phagocytes of mammals and invertebrates produce ETs, however, the evolutionary history of this DNA-based host defence strategy is unclear. Here we report that Sentinel (S) cells of the multicellular slug stage of the social amoeba Dictyostelium discoideum produce ETs upon stimulation with bacteria or lipopolysaccharide in a reactive oxygen species-dependent manner. The production of ETs by S cells requires a Toll/Interleukin-1 receptor domain-containing protein TirA and reactive oxygen species-generating NADPH oxidases. Disruption of these genes results in decreased clearance of bacterial infections. Our results demonstrate that D. discoideum is a powerful model organism to study the evolution and conservation of mechanisms of cell-intrinsic immunity, and suggest that the origin of DNA-based ETs as an innate immune defence predates the emergence of metazoans. PMID:26927887

  17. Survival of phosphate-solubilizing bacteria against DNA damaging agents.

    PubMed

    Shrivastava, Manoj; Rajpurohit, Yogendra S; Misra, Hari S; D'Souza, S F

    2010-10-01

    Phosphate-solubilizing bacteria (PSBs) were isolated from different plant rhizosphere soils of various agroecological regions of India. These isolates showed synthesis of pyrroloquinoline quinone (PQQ), production of gluconic acid, and release of phosphorus from insoluble tricalcium phosphate. The bacterial isolates synthesizing PQQ also showed higher tolerance to ultraviolet C radiation and mitomycin C as compared to Escherichia coli but were less tolerant than Deinococcus radiodurans. Unlike E. coli, PSB isolates showed higher tolerance to DNA damage when grown in the absence of inorganic phosphate. Higher tolerance to ultraviolet C radiation and oxidative stress in these PSBs grown under PQQ synthesis inducible conditions, namely phosphate starvation, might suggest the possible additional role of this redox cofactor in the survival of these isolates under extreme abiotic stress conditions. PMID:20962905

  18. A single protocol for extraction of gDNA from bacteria and yeast.

    PubMed

    Vingataramin, Laurie; Frost, Eric H

    2015-03-01

    Guanidine thiocyanate breakage of microorganisms has been the standard initial step in genomic DNA (gDNA) extraction of microbial DNA for two decades, despite the requirement for pretreatments to extract DNA from microorganisms other than Gram-negative bacteria. We report a quick and low-cost gDNA extraction protocol called EtNa that is efficient for bacteria and yeast over a broad range of concentrations. EtNa is based on a hot alkaline ethanol lysis. The solution can be immediately centrifuged to yield a crude gDNA extract suitable for PCR, or it can be directly applied to a silica column for purification. PMID:25757544

  19. A single protocol for extraction of gDNA from bacteria and yeast.

    PubMed

    Vingataramin, Laurie; Frost, Eric H

    2015-03-01

    Guanidine thiocyanate breakage of microorganisms has been the standard initial step in genomic DNA (gDNA) extraction of microbial DNA for two decades, despite the requirement for pretreatments to extract DNA from microorganisms other than Gram-negative bacteria. We report a quick and low-cost gDNA extraction protocol called EtNa that is efficient for bacteria and yeast over a broad range of concentrations. EtNa is based on a hot alkaline ethanol lysis. The solution can be immediately centrifuged to yield a crude gDNA extract suitable for PCR, or it can be directly applied to a silica column for purification.

  20. Working with DNA & Bacteria in Precollege Science Classrooms.

    ERIC Educational Resources Information Center

    Horn, Toby Mogollon; Frame, Kathy, Ed.

    This document describes ways to work with DNA and host organisms in precollege classrooms. The guidelines are intended to assist the teacher who already has training in working with microbes, DNA, and associated chemicals. The contents of the guidelines include: (1) Permitted DNA molecules, vectors, and recommended host organisms for constructing…

  1. Salivary gland monomorphic adenoma. Ultrastructural, immunoperoxidase, and histogenetic aspects.

    PubMed Central

    Dardick, I.; Kahn, H. J.; Van Nostrand, A. W.; Baumal, R.

    1984-01-01

    Monomorphic adenoma of basal cell type is a salivary gland tumor believed to result from a proliferation of a single type of cell. However, ultrastructural and immunocytochemical investigations of 6 monomorphic adenomas (5 from parotid and 1 from intraoral minor salivary gland) indicate that there are two classes of these lesions, one composed of two types of tumor cells and the other wholly or predominantly made up of one type of cell (isomorphic). In the former group, the organization of the tumor cells closely mimicked that of normal and hyperplastic salivary gland intercalated ducts. Aggregates of tumor cells were arranged as an inner layer of luminal epithelial cells which were surrounded by an outer layer of cells that, in some cases, had ultrastructural and immunohistochemical features indicating myoepithelial cell differentiation. In some adenomas formed by two types of tumor cells, basal-lamina-lined extracellular spaces were identified ultrastructurally in relation to modified myoepithelial cells; such spaces had the same fine-structural features as those reported in pleomorphic adenoma and adenoid cystic carcinoma. Predominantly isomorphic adenomas were composed exclusively of luminal epithelial cells. These results indicate that despite the varied histologic patterns in the numerous subtypes of monomorphic adenoma, there is a central theme of differentiation and organization in this type of neoplasm which recapitulates the ductoacinar unit of normal salivary gland parenchyma. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 9 Figure 10 Figure 11 Figure 12 Figure 13 Figure 14 Figure 15 PMID:6375388

  2. DNA fingerprinting of lactic acid bacteria in sauerkraut fermentations

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Previous studies using traditional biochemical methods to study the ecology of commercial sauerkraut fermentations revealed that four lactic acid bacteria species, Leuconostoc mesenteroides, Lactobacillus plantarum, Pediococcus pentosaceus, and Lactobacillus brevis were the primary microorganisms in...

  3. How-to-Do-It: A Simple DNA Isolation Technique Using Halophilic Bacteria.

    ERIC Educational Resources Information Center

    Guilfoile, Patrick

    1989-01-01

    Described is a simple technique for isolating DNA from halophilic bacteria. Materials, procedure, and additional experiments are outlined. It is stated that the DNA obtained will be somewhat contaminated with cellular proteins and RNA. Offers a procedure for greater purification. (RT)

  4. Engineering nanoparticle-coated bacteria as oral DNA vaccines for cancer immunotherapy.

    PubMed

    Hu, Qinglian; Wu, Min; Fang, Chun; Cheng, Changyong; Zhao, Mengmeng; Fang, Weihuan; Chu, Paul K; Ping, Yuan; Tang, Guping

    2015-04-01

    Live attenuated bacteria are of increasing importance in biotechnology and medicine in the emerging field of cancer immunotherapy. Oral DNA vaccination mediated by live attenuated bacteria often suffers from low infection efficiency due to various biological barriers during the infection process. To this end, we herein report, for the first time, a new strategy to engineer cationic nanoparticle-coated bacterial vectors that can efficiently deliver oral DNA vaccine for efficacious cancer immunotherapy. By coating live attenuated bacteria with synthetic nanoparticles self-assembled from cationic polymers and plasmid DNA, the protective nanoparticle coating layer is able to facilitate bacteria to effectively escape phagosomes, significantly enhance the acid tolerance of bacteria in stomach and intestines, and greatly promote dissemination of bacteria into blood circulation after oral administration. Most importantly, oral delivery of DNA vaccines encoding autologous vascular endothelial growth factor receptor 2 (VEGFR2) by this hybrid vector showed remarkable T cell activation and cytokine production. Successful inhibition of tumor growth was also achieved by efficient oral delivery of VEGFR2 with nanoparticle-coated bacterial vectors due to angiogenesis suppression in the tumor vasculature and tumor necrosis. This proof-of-concept work demonstrates that coating live bacterial cells with synthetic nanoparticles represents a promising strategy to engineer efficient and versatile DNA vaccines for the era of immunotherapy.

  5. Isolation of high molecular weight DNA from marine sponge bacteria for BAC library construction.

    PubMed

    Ouyang, Yongchang; Dai, Shikun; Xie, Lianwu; Ravi Kumar, M S; Sun, Wei; Sun, Huimin; Tang, Danling; Li, Xiang

    2010-06-01

    Metagenomics is a powerful tool for mining the genetic repositories from environmental microorganisms. Bacteria associated with marine sponges (phylum Porifera) are rich sources of biologically active natural products. However, to date, few compounds are discovered from the sponge metagenomic libraries, and the main reason might be the difficulties in recovery of high molecular weight (HMW) DNA from sponge symbionts to construct large insert libraries. Here, we describe a method to recover HMW bacterial DNA from diverse sponges with high quality for bacterial artificial chromosome (BAC) library construction. Microorganisms concentrated from sponges by differential centrifugation were embedded in agarose plugs to lyse out the HMW DNA for recovery. DNA fragments over 436 kb size were recovered from three different types of sponges, Halichondria sp., Haliclona sp., and Xestospongia sp. To evaluate the recovered DNA quality, the diversity of bacterial DNA comprised in the HMW DNA derived from sponge Halichondria sp. was analyzed, and this HMW DNA sample was also cloned into a shuttle BAC vector between Escherichia coli and Streptomyces sp. The results showed that more than five types of bacterial DNA, i.e., Proteobacteria, Nitrospirae, Cyanobacteria, Planctomycetes, and unidentified bacteria, had been recovered by this method, and an average 100 kb size insert DNA in a constructed BAC library demonstrated that the recovered HMW DNA is suitable for metagenomic library construction.

  6. Molecular detection of bacteria in plant tissues, using universal 16S ribosomal DNA degenerated primers

    PubMed Central

    Tsoktouridis, Georgios; Tsiamis, George; Koutinas, Nikolaos; Mantell, Sinclair

    2014-01-01

    Highly specific, sensitive and rapid tests are required for the detection and identification of covert bacterial contaminations in plant tissue cultures. Current methods available for this purpose are tedious, time consuming, highly error prone, expensive, require advanced technical expertise and are sometimes ineffective. We report here the development of a sensitive polymerase chain reaction (PCR) based method for the rapid detection and identification of bacteria occurring in plant tissue cultures. A total of 121 16S ribosomal DNA (rDNA) coding regions from 14 different groups of bacteria, algae and plants, available in the Gene Bank/European Molecular Biology Laboratory databases, were aligned and several conserved DNA sequences of bacterial origin were identified. From those, five degenerated primers were designed in order to amplify only the bacterial DNA present in mixed plant/bacteria genomic DNA extracts. A known amount of bacterial suspension of either covert Pseudomonas or covert Bacillus were added to in vitro plant leaves and total plant/bacterial DNA extracted using three different methods to determine the lowest number of bacteria required to be present in order to allow their detection. The highest sensitivity of the bacterial cell detection was 2.5 × 106 cells of both Bacillus and Pseudomonas inoculums, using template DNA prepared by the MiniPrep method. Generation of PCR amplification fragments was achieved only for the 16S rDNA bacterial gene by using four combinations of degenerated primers. Successive sequence analysis of these amplified fragments led to the rapid detection and molecular identification of bacteria covertly associated with plants. PMID:26019546

  7. Antimicrobials targeted to the replication-specific DNA polymerases of gram-positive bacteria: target potential of dnaE.

    PubMed

    Barnes, Marjorie H; Butler, Michelle M; Wright, George E; Brown, Neal C

    2012-10-01

    DNA polymerases pol IIIC and dnaE [i.e. pol IIIE] are essential for replicative DNA synthesis in low G:C Gram-positive eubacteria. Therefore, they have strong potential as targets for development of Gram-positive-selective antibacterial agents. This work has sought to extend to dnaE the recent discovery of antimicrobial agents based on pol IIIC-specific dGTP analogs. Compound 324C, a member of the same dGTP analog family, was found to be a potent and selective inhibitor of isolated dnaE in vitro. Surprisingly, 324C had no inhibitory effect in either intact Bacillus subtilis cells or in permeabilized cell preparations used to assess replicative DNA synthesis directly. It is proposed that the failure of 324C in the intact cell is a consequence of two major factors: (i) its template-dependent base pairing mechanism, and (ii) a specific subordinate role which dnaE apparently plays to pol IIIC. To generate an effective dnaE-selective inhibitor of replicative DNA synthesis in Gram-positive bacteria, it will likely be necessary to develop a molecule that attacks the enzyme's active site directly, without binding to template DNA.

  8. Isolation and characterization of yeast monomorphic mutants of Candida albicans.

    PubMed Central

    Elorza, M V; Sentandreu, R; Ruiz-Herrera, J

    1994-01-01

    A method was devised for the isolation of yeast monomorphic (LEV) mutants of Candida albicans. By this procedure, about 20 stable yeast-like mutants were isolated after mutagenesis with ethyl methane sulfonate. The growth rate of the mutants in different carbon sources, both fermentable and not, was indistinguishable from that of the parental strain, but they were unable to grow as mycelial forms after application of any of the common effective inducers, i.e., heat shock, pH alterations, proline addition, or use of GlcNAc as the carbon source. Studies performed with one selected strain demonstrated that it had severe alterations in the chemical composition of the cell wall, mainly in the levels of chitin and glucans, and in specific mannoproteins, some of them recognizable by specific polyclonal and monoclonal antibodies. It is suggested that these structural alterations hinder the construction of a normal hyphal wall. Images PMID:8157600

  9. Utility of Filter Paper for Preserving Insects, Bacteria, and Host Reservoir DNA for Molecular Testing

    PubMed Central

    Karimian, F; Sedaghat, MM; Oshaghi, MA; Mohtarami, F; Dehkordi, A Sanei; Koosha, M; Akbari, S; Hashemi-Aghdam, SS

    2011-01-01

    Background: Appropriate methodology for storage biological materials, extraction of DNA, and proper DNA preservation is vital for studies involving genetic analysis of insects, bacteria, and reservoir hosts as well as for molecular diagnostics of pathogens carried by vectors and reservoirs. Here we tried to evaluate the utility of a simple filter paper-based for storage of insects, bacteria, rodent, and human DNAs using PCR assays. Methods: Total body or haemolymph of individual mosquitoes, sand flies or cockroaches squashed or placed on the paper respectively. Extracted DNA of five different bacteria species as well as blood specimens of human and great gerbil Rhombomys opimus was pipetted directly onto filter paper. The papers were stored in room temperature up to 12 months during 2009 until 2011. At monthly intervals, PCR was conducted using a 1-mm disk from the DNA impregnated filter paper as target DNA. PCR amplification was performed against different target genes of the organisms including the ITS2-rDNA of mosquitoes, mtDNA-COI of the sand flies and cockroaches, 16SrRNA gene of the bacteria, and the mtDNA-CytB of the vertebrates. Results: Successful PCR amplification was observed for all of the specimens regardless of the loci, taxon, or time of storage. The PCR amplification were ranged from 462 to 1500 bp and worked well for the specified target gene/s. Time of storage did not affect the amplification up to one year. Conclusion: The filter paper method is a simple and economical way to store, to preserve, and to distribute DNA samples for PCR analysis. PMID:22808417

  10. Cultivation-independent detection of autotrophic hydrogen-oxidizing bacteria by DNA stable-isotope probing.

    PubMed

    Pumphrey, Graham M; Ranchou-Peyruse, Anthony; Spain, Jim C

    2011-07-01

    Knallgas bacteria are a physiologically defined group that is primarily studied using cultivation-dependent techniques. Given that current cultivation techniques fail to grow most bacteria, cultivation-independent techniques that selectively detect and identify knallgas bacteria will improve our ability to study their diversity and distribution. We used stable-isotope probing (SIP) to identify knallgas bacteria in rhizosphere soil of legumes and in a microbial mat from Obsidian Pool in Yellowstone National Park. When samples were incubated in the dark, incorporation of (13)CO(2) was H(2) dependent. SIP enabled the detection of knallgas bacteria that were not detected by cultivation, and the majority of bacteria identified in the rhizosphere soils were betaproteobacteria predominantly related to genera previously known to oxidize hydrogen. Bacteria in soil grew on hydrogen at concentrations as low as 100 ppm. A hydB homolog encoding a putative high-affinity NiFe hydrogenase was amplified from (13)C-labeled DNA from both vetch and clover rhizosphere soil. The results indicate that knallgas bacteria can be detected by SIP and populations that respond to different H(2) concentrations can be distinguished. The methods described here should be applicable to a variety of ecosystems and will enable the discovery of additional knallgas bacteria that are resistant to cultivation. PMID:21622787

  11. Cultivation-independent detection of autotrophic hydrogen-oxidizing bacteria by DNA stable-isotope probing.

    PubMed

    Pumphrey, Graham M; Ranchou-Peyruse, Anthony; Spain, Jim C

    2011-07-01

    Knallgas bacteria are a physiologically defined group that is primarily studied using cultivation-dependent techniques. Given that current cultivation techniques fail to grow most bacteria, cultivation-independent techniques that selectively detect and identify knallgas bacteria will improve our ability to study their diversity and distribution. We used stable-isotope probing (SIP) to identify knallgas bacteria in rhizosphere soil of legumes and in a microbial mat from Obsidian Pool in Yellowstone National Park. When samples were incubated in the dark, incorporation of (13)CO(2) was H(2) dependent. SIP enabled the detection of knallgas bacteria that were not detected by cultivation, and the majority of bacteria identified in the rhizosphere soils were betaproteobacteria predominantly related to genera previously known to oxidize hydrogen. Bacteria in soil grew on hydrogen at concentrations as low as 100 ppm. A hydB homolog encoding a putative high-affinity NiFe hydrogenase was amplified from (13)C-labeled DNA from both vetch and clover rhizosphere soil. The results indicate that knallgas bacteria can be detected by SIP and populations that respond to different H(2) concentrations can be distinguished. The methods described here should be applicable to a variety of ecosystems and will enable the discovery of additional knallgas bacteria that are resistant to cultivation.

  12. DNA from oral bacteria by sodium hydroxide-paper method suitable for polymerase chain reaction.

    PubMed

    Lefimil, Claudia; Lozano, Carla; Morales-Bozo, Irene; Plaza, Anita; Maturana, Cristian; Urzúa, Blanca

    2013-02-15

    In the oral cavity, we can find a complex mixture of microorganisms, commensals, and pathogens. The studies of normal oral microbiota, as well as the studies of much oral pathology (e.g., caries, periodontitis), involve the isolation and cultivation of these microorganisms and their molecular analysis. The aim of this study was to validate a quick, easy, efficient, and inexpensive DNA extraction method for the recovery of genomic DNA from gram-positive and gram-negative oral bacteria to be used in polymerase chain reaction amplification. This method worked great with all samples analyzed, providing an approach to extract DNA for different microorganisms.

  13. 40 CFR 798.5500 - Differential growth inhibition of repair proficient and repair deficient bacteria: “Bacterial DNA...

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... recommendations as specified under 40 CFR part 792, subpart J the following specific information should be... repair proficient and repair deficient bacteria: âBacterial DNA damage or repair tests.â 798.5500 Section... inhibition of repair proficient and repair deficient bacteria: “Bacterial DNA damage or repair tests.”...

  14. 40 CFR 798.5500 - Differential growth inhibition of repair proficient and repair deficient bacteria: “Bacterial DNA...

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... recommendations as specified under 40 CFR part 792, subpart J the following specific information should be... repair proficient and repair deficient bacteria: âBacterial DNA damage or repair tests.â 798.5500 Section... inhibition of repair proficient and repair deficient bacteria: “Bacterial DNA damage or repair tests.”...

  15. 40 CFR 798.5500 - Differential growth inhibition of repair proficient and repair deficient bacteria: “Bacterial DNA...

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... recommendations as specified under 40 CFR part 792, subpart J the following specific information should be... repair proficient and repair deficient bacteria: âBacterial DNA damage or repair tests.â 798.5500 Section... inhibition of repair proficient and repair deficient bacteria: “Bacterial DNA damage or repair tests.”...

  16. 40 CFR 798.5500 - Differential growth inhibition of repair proficient and repair deficient bacteria: “Bacterial DNA...

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... recommendations as specified under 40 CFR part 792, subpart J the following specific information should be... repair proficient and repair deficient bacteria: âBacterial DNA damage or repair tests.â 798.5500 Section... inhibition of repair proficient and repair deficient bacteria: “Bacterial DNA damage or repair tests.”...

  17. Genomewide expression analysis in amino acid-producing bacteria using DNA microarrays.

    PubMed

    Polen, Tino; Wendisch, Volker F

    2004-01-01

    DNA microarray technology has become an important research tool for biotechnology and microbiology. It is now possible to characterize genetic diversity and gene expression in a genomewide manner. DNA microarrays have been applied extensively to study the biology of many bacteria including Escherichia coli, but only recently have they been developed for the Gram-positive Corynebacterium glutamicum. Both bacteria are widely used for biotechnological amino acid production. In this article, in addition to the design and generation of microarrays as well as their use in hybridization experiments and subsequent data analysis, we describe recent applications of DNA microarray technology regarding amino acid production in C. glutamicum and E. coli. We also discuss the impact of functional genomics studies on fundamental as well as applied aspects of amino acid production with C. glutamicum and E. coli. PMID:15304751

  18. LL37:DNA complexes provide antimicrobial activity against intracellular bacteria in human macrophages.

    PubMed

    Stephan, Alexander; Batinica, Marina; Steiger, Julia; Hartmann, Pia; Zaucke, Frank; Bloch, Wilhelm; Fabri, Mario

    2016-08-01

    As part of the innate host response neutrophils release neutrophil extracellular traps (NETs), protein:DNA complexes that contain a number of antimicrobial peptides (AMPs), such as cathelicidin. Human cathelicidin in its active form, LL37, has potent antimicrobial activity against bacteria. However, whether LL37 derived from NETs contributes to antimicrobial activity against intracellular pathogens remains unclear. Here, we report that NETs induced by mycobacteria contain cathelicidin. Human macrophages internalized NET-bound cathelicidin, which is transported to lysosomal compartments. Furthermore, using a model of in vitro-generated LL37:DNA complexes we found that LL37 derived from such complexes attacks mycobacteria in macrophage phagolysosomes resulting in antimicrobial activity. Taken together, our results suggest a mechanism by which LL37 in complex with DNA contributes to host defence against intracellular bacteria in human macrophages.

  19. Identification of DNA Methyltransferase Genes in Human Pathogenic Bacteria by Comparative Genomics.

    PubMed

    Brambila-Tapia, Aniel Jessica Leticia; Poot-Hernández, Augusto Cesar; Perez-Rueda, Ernesto; Rodríguez-Vázquez, Katya

    2016-06-01

    DNA methylation plays an important role in gene expression and virulence in some pathogenic bacteria. In this report, we describe DNA methyltransferases (MTases) present in human pathogenic bacteria and compared them with related species, which are not pathogenic or less pathogenic, based in comparative genomics. We performed a search in the KEGG database of the KEGG database orthology groups associated with adenine and cytosine DNA MTase activities (EC: 2.1.1.37, EC: 2.1.1.113 and EC: 2.1.1.72) in 37 human pathogenic species and 18 non/less pathogenic relatives and performed comparisons of the number of these MTases sequences according to their genome size, the DNA MTase type and with their non-less pathogenic relatives. We observed that Helicobacter pylori and Neisseria spp. presented the highest number of MTases while ten different species did not present a predicted DNA MTase. We also detected a significant increase of adenine MTases over cytosine MTases (2.19 vs. 1.06, respectively, p < 0.001). Adenine MTases were the only MTases associated with restriction modification systems and DNA MTases associated with type I restriction modification systems were more numerous than those associated with type III restriction modification systems (0.84 vs. 0.17, p < 0.001); additionally, there was no correlation with the genome size and the total number of DNA MTases, indicating that the number of DNA MTases is related to the particular evolution and lifestyle of specific species, regulating the expression of virulence genes in some pathogenic bacteria.

  20. Identification of DNA Methyltransferase Genes in Human Pathogenic Bacteria by Comparative Genomics.

    PubMed

    Brambila-Tapia, Aniel Jessica Leticia; Poot-Hernández, Augusto Cesar; Perez-Rueda, Ernesto; Rodríguez-Vázquez, Katya

    2016-06-01

    DNA methylation plays an important role in gene expression and virulence in some pathogenic bacteria. In this report, we describe DNA methyltransferases (MTases) present in human pathogenic bacteria and compared them with related species, which are not pathogenic or less pathogenic, based in comparative genomics. We performed a search in the KEGG database of the KEGG database orthology groups associated with adenine and cytosine DNA MTase activities (EC: 2.1.1.37, EC: 2.1.1.113 and EC: 2.1.1.72) in 37 human pathogenic species and 18 non/less pathogenic relatives and performed comparisons of the number of these MTases sequences according to their genome size, the DNA MTase type and with their non-less pathogenic relatives. We observed that Helicobacter pylori and Neisseria spp. presented the highest number of MTases while ten different species did not present a predicted DNA MTase. We also detected a significant increase of adenine MTases over cytosine MTases (2.19 vs. 1.06, respectively, p < 0.001). Adenine MTases were the only MTases associated with restriction modification systems and DNA MTases associated with type I restriction modification systems were more numerous than those associated with type III restriction modification systems (0.84 vs. 0.17, p < 0.001); additionally, there was no correlation with the genome size and the total number of DNA MTases, indicating that the number of DNA MTases is related to the particular evolution and lifestyle of specific species, regulating the expression of virulence genes in some pathogenic bacteria. PMID:27570304

  1. Nonchromosomal Antibiotic Resistance in Bacteria: Genetic Transformation of Escherichia coli by R-Factor DNA*

    PubMed Central

    Cohen, Stanley N.; Chang, Annie C. Y.; Hsu, Leslie

    1972-01-01

    Transformation of E. coli cells treated with CaCl2 to multiple antibiotic resistance by purified R-factor DNA is reported. Drug resistance is expressed in a small fraction of the recipient bacterial population almost immediately after uptake of DNA, but full genetic expression of resistance requires subsequent incubation in drugfree medium before antibiotic challenge. Transformed bacteria acquire a closed circular, transferable DNA species having the resistance, fertility, and sedimentation characteristics of the parent R factor. Covalently-closed, catenated, and open (nicked) circular forms of R-factor DNA are all effective in transformation, but denaturation and sonication abolish the transforming ability of R-factor DNA in this system. PMID:4559594

  2. Principles and Concepts of DNA Replication in Bacteria, Archaea, and Eukarya

    PubMed Central

    O’Donnell, Michael; Langston, Lance; Stillman, Bruce

    2013-01-01

    The accurate copying of genetic information in the double helix of DNA is essential for inheritance of traits that define the phenotype of cells and the organism. The core machineries that copy DNA are conserved in all three domains of life: bacteria, archaea, and eukaryotes. This article outlines the general nature of the DNA replication machinery, but also points out important and key differences. The most complex organisms, eukaryotes, have to coordinate the initiation of DNA replication from many origins in each genome and impose regulation that maintains genomic integrity, not only for the sake of each cell, but for the organism as a whole. In addition, DNA replication in eukaryotes needs to be coordinated with inheritance of chromatin, developmental patterning of tissues, and cell division to ensure that the genome replicates once per cell division cycle. PMID:23818497

  3. Principles and concepts of DNA replication in bacteria, archaea, and eukarya.

    PubMed

    O'Donnell, Michael; Langston, Lance; Stillman, Bruce

    2013-07-01

    The accurate copying of genetic information in the double helix of DNA is essential for inheritance of traits that define the phenotype of cells and the organism. The core machineries that copy DNA are conserved in all three domains of life: bacteria, archaea, and eukaryotes. This article outlines the general nature of the DNA replication machinery, but also points out important and key differences. The most complex organisms, eukaryotes, have to coordinate the initiation of DNA replication from many origins in each genome and impose regulation that maintains genomic integrity, not only for the sake of each cell, but for the organism as a whole. In addition, DNA replication in eukaryotes needs to be coordinated with inheritance of chromatin, developmental patterning of tissues, and cell division to ensure that the genome replicates once per cell division cycle. PMID:23818497

  4. Sex-specific foraging behaviour in a monomorphic seabird.

    PubMed Central

    Lewis, S; Benvenuti, S; Dall'Antonia, L; Griffiths, R; Money, L; Sherratt, T N; Wanless, S; Hamer, K C

    2002-01-01

    Sexual differences in the foraging behaviour of parents have been observed in a number of sexually sizedimorphic birds, particularly seabirds, and the usual inference has been that these sex-specific differences are mediated primarily by differences in body size. To test this explanation, we compared the foraging behaviour of parents in a monomorphic seabird species, the northern gannet Morus bassanus. Using specially designed instruments and radio telemetry we found that individuals of both sexes were consistent in the directions and durations of their foraging trips. However, there were significant differences in the foraging behaviour of males and females. Female gannets were not only more selective than males in the areas where they foraged, but they also made longer, deeper dives and spent more time on the sea surface than males. As the sexes are morphologically similar in this species, then these differences are unlikely to have been mediated by body size. Our work highlights the need to investigate sexual differences in the foraging behaviour of seabirds and other species more closely, in order to test alternative theories that do not rely on differences in body size. PMID:12204129

  5. Horizontal DNA transfer from bacteria to eukaryotes and a lesson from experimental transfers.

    PubMed

    Suzuki, Katsunori; Moriguchi, Kazuki; Yamamoto, Shinji

    2015-12-01

    Horizontal gene transfer (HGT) is widespread among bacteria and plays a key role in genome dynamics. HGT is much less common in eukaryotes, but is being reported with increasing frequency in eukaryotes. The mechanism as to how eukaryotes acquired genes from distantly related organisms remains obscure yet. This paper cites examples of bacteria-derived genes found in eukaryotic organisms, and then describes experimental DNA transports to eukaryotes by bacterial type 4 secretion systems in optimized conditions. The mechanisms of the latter are efficient, quite reproducible in vitro and predictable, and thereby would provide insight into natural HGT and to the development of new research tools.

  6. Atomic Resolution Structure of Monomorphic Aβ42 Amyloid Fibrils.

    PubMed

    Colvin, Michael T; Silvers, Robert; Ni, Qing Zhe; Can, Thach V; Sergeyev, Ivan; Rosay, Melanie; Donovan, Kevin J; Michael, Brian; Wall, Joseph; Linse, Sara; Griffin, Robert G

    2016-08-01

    Amyloid-β (Aβ) is a 39-42 residue protein produced by the cleavage of the amyloid precursor protein (APP), which subsequently aggregates to form cross-β amyloid fibrils that are a hallmark of Alzheimer's disease (AD). The most prominent forms of Aβ are Aβ1-40 and Aβ1-42, which differ by two amino acids (I and A) at the C-terminus. However, Aβ42 is more neurotoxic and essential to the etiology of AD. Here, we present an atomic resolution structure of a monomorphic form of AβM01-42 amyloid fibrils derived from over 500 (13)C-(13)C, (13)C-(15)N distance and backbone angle structural constraints obtained from high field magic angle spinning NMR spectra. The structure (PDB ID: 5KK3 ) shows that the fibril core consists of a dimer of Aβ42 molecules, each containing four β-strands in a S-shaped amyloid fold, and arranged in a manner that generates two hydrophobic cores that are capped at the end of the chain by a salt bridge. The outer surface of the monomers presents hydrophilic side chains to the solvent. The interface between the monomers of the dimer shows clear contacts between M35 of one molecule and L17 and Q15 of the second. Intermolecular (13)C-(15)N constraints demonstrate that the amyloid fibrils are parallel in register. The RMSD of the backbone structure (Q15-A42) is 0.71 ± 0.12 Å and of all heavy atoms is 1.07 ± 0.08 Å. The structure provides a point of departure for the design of drugs that bind to the fibril surface and therefore interfere with secondary nucleation and for other therapeutic approaches to mitigate Aβ42 aggregation. PMID:27355699

  7. Atomic Resolution Structure of Monomorphic Aβ42 Amyloid Fibrils.

    PubMed

    Colvin, Michael T; Silvers, Robert; Ni, Qing Zhe; Can, Thach V; Sergeyev, Ivan; Rosay, Melanie; Donovan, Kevin J; Michael, Brian; Wall, Joseph; Linse, Sara; Griffin, Robert G

    2016-08-01

    Amyloid-β (Aβ) is a 39-42 residue protein produced by the cleavage of the amyloid precursor protein (APP), which subsequently aggregates to form cross-β amyloid fibrils that are a hallmark of Alzheimer's disease (AD). The most prominent forms of Aβ are Aβ1-40 and Aβ1-42, which differ by two amino acids (I and A) at the C-terminus. However, Aβ42 is more neurotoxic and essential to the etiology of AD. Here, we present an atomic resolution structure of a monomorphic form of AβM01-42 amyloid fibrils derived from over 500 (13)C-(13)C, (13)C-(15)N distance and backbone angle structural constraints obtained from high field magic angle spinning NMR spectra. The structure (PDB ID: 5KK3 ) shows that the fibril core consists of a dimer of Aβ42 molecules, each containing four β-strands in a S-shaped amyloid fold, and arranged in a manner that generates two hydrophobic cores that are capped at the end of the chain by a salt bridge. The outer surface of the monomers presents hydrophilic side chains to the solvent. The interface between the monomers of the dimer shows clear contacts between M35 of one molecule and L17 and Q15 of the second. Intermolecular (13)C-(15)N constraints demonstrate that the amyloid fibrils are parallel in register. The RMSD of the backbone structure (Q15-A42) is 0.71 ± 0.12 Å and of all heavy atoms is 1.07 ± 0.08 Å. The structure provides a point of departure for the design of drugs that bind to the fibril surface and therefore interfere with secondary nucleation and for other therapeutic approaches to mitigate Aβ42 aggregation.

  8. Sexually Monomorphic Maps and Dimorphic Responses in Rat Genital Cortex.

    PubMed

    Lenschow, Constanze; Copley, Sean; Gardiner, Jayne M; Talbot, Zoe N; Vitenzon, Ariel; Brecht, Michael

    2016-01-11

    Mammalian external genitals show sexual dimorphism [1, 2] and can change size and shape upon sexual arousal. Genitals feature prominently in the oldest pieces of figural art [3] and phallic depictions of penises informed psychoanalytic thought about sexuality [4, 5]. Despite this longstanding interest, the neural representations of genitals are still poorly understood [6]. In somatosensory cortex specifically, many studies did not detect any cortical representation of genitals [7-9]. Studies in humans debate whether genitals are represented displaced below the foot of the cortical body map [10-12] or whether they are represented somatotopically [13-15]. We wondered what a high-resolution mapping of genital representations might tell us about the sexual differentiation of the mammalian brain. We identified genital responses in rat somatosensory cortex in a region previously assigned as arm/leg cortex. Genital responses were more common in males than in females. Despite such response dimorphism, we observed a stunning anatomical monomorphism of cortical penis and clitoris input maps revealed by cytochrome-oxidase-staining of cortical layer 4. Genital representations were somatotopic and bilaterally symmetric, and their relative size increased markedly during puberty. Size, shape, and erect posture give the cortical penis representation a phallic appearance pointing to a role in sexually aroused states. Cortical genital neurons showed unusual multi-body-part responses and sexually dimorphic receptive fields. Specifically, genital neurons were co-activated by distant body regions, which are touched during mounting in the respective sex. Genital maps indicate a deep homology of penis and clitoris representations in line with a fundamentally bi-sexual layout [16] of the vertebrate brain. PMID:26725197

  9. Diversity of bacteria in ships ballast water as revealed by next generation DNA sequencing.

    PubMed

    Brinkmeyer, Robin

    2016-06-15

    The bacterial diversity in ballast water from five general cargo ships calling at the Port of Houston was determined with ion semiconductor DNA sequencing (Ion Torrent PGM) of PCR amplified 16S rRNA genes. Phylogenetic analysis revealed that the composition of bacteria in ballast water did not resemble that of typical marine habitats or even open ocean waters where BWEs occur. The predominant group of bacteria in ships conducting BWEs was the Roseobacter clade within the Alphaproteobacteria. In contrast, Gammaproteobacteria were predominant in the ship that did not conduct a BWE. All the ships contained human, fish, and terrestrial plant pathogens as well as bacteria indicative of fecal or activated sludge contamination. Most of the 60 pathogens had not been detected in ballast water previously. Among these were the human pathogens Corynebacterium diptheriae and several Legionella species and the fish pathogens Francisella piscicida and Piscirickettsia salmonis. PMID:27076378

  10. Diversity of bacteria in ships ballast water as revealed by next generation DNA sequencing.

    PubMed

    Brinkmeyer, Robin

    2016-06-15

    The bacterial diversity in ballast water from five general cargo ships calling at the Port of Houston was determined with ion semiconductor DNA sequencing (Ion Torrent PGM) of PCR amplified 16S rRNA genes. Phylogenetic analysis revealed that the composition of bacteria in ballast water did not resemble that of typical marine habitats or even open ocean waters where BWEs occur. The predominant group of bacteria in ships conducting BWEs was the Roseobacter clade within the Alphaproteobacteria. In contrast, Gammaproteobacteria were predominant in the ship that did not conduct a BWE. All the ships contained human, fish, and terrestrial plant pathogens as well as bacteria indicative of fecal or activated sludge contamination. Most of the 60 pathogens had not been detected in ballast water previously. Among these were the human pathogens Corynebacterium diptheriae and several Legionella species and the fish pathogens Francisella piscicida and Piscirickettsia salmonis.

  11. Identification of active oxalotrophic bacteria by Bromodeoxyuridine DNA labeling in a microcosm soil experiments.

    PubMed

    Bravo, Daniel; Martin, Gaëtan; David, Maude M; Cailleau, Guillaume; Verrecchia, Eric; Junier, Pilar

    2013-11-01

    The oxalate-carbonate pathway (OCP) leads to a potential carbon sink in terrestrial environments. This process is linked to the activity of oxalotrophic bacteria. Although isolation and molecular characterizations are used to study oxalotrophic bacteria, these approaches do not give information on the active oxalotrophs present in soil undergoing the OCP. The aim of this study was to assess the diversity of active oxalotrophic bacteria in soil microcosms using the Bromodeoxyuridine (BrdU) DNA labeling technique. Soil was collected near an oxalogenic tree (Milicia excelsa). Different concentrations of calcium oxalate (0.5%, 1%, and 4% w/w) were added to the soil microcosms and compared with an untreated control. After 12 days of incubation, a maximal pH of 7.7 was measured for microcosms with oxalate (initial pH 6.4). At this time point, a DGGE profile of the frc gene was performed from BrdU-labeled soil DNA and unlabeled soil DNA. Actinobacteria (Streptomyces- and Kribbella-like sequences), Gammaproteobacteria and Betaproteobacteria were found as the main active oxalotrophic bacterial groups. This study highlights the relevance of Actinobacteria as members of the active bacterial community and the identification of novel uncultured oxalotrophic groups (i.e. Kribbella) active in soils.

  12. Relevance of GC content to the conservation of DNA polymerase III/mismatch repair system in Gram-positive bacteria

    PubMed Central

    Akashi, Motohiro; Yoshikawa, Hirofumi

    2013-01-01

    The mechanism of DNA replication is one of the driving forces of genome evolution. Bacterial DNA polymerase III, the primary complex of DNA replication, consists of PolC and DnaE. PolC is conserved in Gram-positive bacteria, especially in the Firmicutes with low GC content, whereas DnaE is widely conserved in most Gram-negative and Gram-positive bacteria. PolC contains two domains, the 3′-5′exonuclease domain and the polymerase domain, while DnaE only possesses the polymerase domain. Accordingly, DnaE does not have the proofreading function; in Escherichia coli, another enzyme DnaQ performs this function. In most bacteria, the fidelity of DNA replication is maintained by 3′-5′ exonuclease and a mismatch repair (MMR) system. However, we found that most Actinobacteria (a group of Gram-positive bacteria with high GC content) appear to have lost the MMR system and chromosomes may be replicated by DnaE-type DNA polymerase III with DnaQ-like 3′-5′ exonuclease. We tested the mutation bias of Bacillus subtilis, which belongs to the Firmicutes and found that the wild type strain is AT-biased while the mutS-deletant strain is remarkably GC-biased. If we presume that DnaE tends to make mistakes that increase GC content, these results can be explained by the mutS deletion (i.e., deletion of the MMR system). Thus, we propose that GC content is regulated by DNA polymerase and MMR system, and the absence of polC genes, which participate in the MMR system, may be the reason for the increase of GC content in Gram-positive bacteria such as Actinobacteria. PMID:24062730

  13. Characterization of viable bacteria from Siberian permafrost by 16S rDNA sequencing

    NASA Technical Reports Server (NTRS)

    Shi, T.; Reeves, R. H.; Gilichinsky, D. A.; Friedmann, E. I.

    1997-01-01

    Viable bacteria were found in permafrost core samples from the Kolyma-Indigirka lowland of northeast Siberia. The samples were obtained at different depths; the deepest was about 3 million years old. The average temperature of the permafrost is -10 degrees C. Twenty-nine bacterial isolates were characterized by 16S rDNA sequencing and phylogenetic analysis, cell morphology, Gram staining, endospore formation, and growth at 30 degrees C. The majority of the bacterial isolates were rod shaped and grew well at 30 degrees C; but two of them did not grow at or above 28 degrees C, and had optimum growth temperatures around 20 degrees C. Thirty percent of the isolates could form endospores. Phylogenetic analysis revealed that the isolates fell into four categories: high-GC Gram-positive bacteria, beta-proteobacteria, gamma-proteobacteria, and low-GC Gram-positive bacteria. Most high-GC Gram-positive bacteria and beta-proteobacteria, and all gamma-proteobacteria, came from samples with an estimated age of 1.8-3.0 million years (Olyor suite). Most low-GC Gram-positive bacteria came from samples with an estimated age of 5,000-8,000 years (Alas suite).

  14. Cadmium Accumulation and DNA Homology with Metal Resistance Genes in Sulfate-Reducing Bacteria

    PubMed Central

    Naz, Naghma; Young, Hilary K.; Ahmed, Nuzhat; Gadd, Geoffrey M.

    2005-01-01

    Cadmium resistance (0.1 to 1.0 mM) was studied in four pure and one mixed culture of sulfate-reducing bacteria (SRB). The growth of the bacteria was monitored with respect to carbon source (lactate) oxidation and sulfate reduction in the presence of various concentrations of cadmium chloride. Two strains Desulfovibrio desulfuricans DSM 1926 and Desulfococcus multivorans DSM 2059 showed the highest resistance to cadmium (0.5 mM). Transmission electron microscopy of the two strains showed intracellular and periplasmic accumulation of cadmium. Dot blot DNA hybridization using the probes for the smtAB, cadAC, and cadD genes indicated the presence of similar genetic determinants of heavy metal resistance in the SRB tested. DNA sequencing of the amplified DNA showed strong nucleotide homology in all the SRB strains with the known smtAB genes encoding synechococcal metallothioneins. Protein homology with the known heavy metal-translocating ATPases was also detected in the cloned amplified DNA of Desulfomicrobium norvegicum I1 and Desulfovibrio desulfuricans DSM 1926, suggesting the presence of multiple genetic mechanisms of metal resistance in the two strains. PMID:16085855

  15. Biochemical and structural characterization of DNA ligases from bacteria and archaea

    PubMed Central

    Pergolizzi, Giulia; Wagner, Gerd K.; Bowater, Richard P.

    2016-01-01

    DNA ligases are enzymes that seal breaks in the backbones of DNA, leading to them being essential for the survival of all organisms. DNA ligases have been studied from many different types of cells and organisms and shown to have diverse sizes and sequences, with well conserved specific sequences that are required for enzymatic activity. A significant number of DNA ligases have been isolated or prepared in recombinant forms and, here, we review their biochemical and structural characterization. All DNA ligases contain an essential lysine that transfers an adenylate group from a co-factor to the 5′-phosphate of the DNA end that will ultimately be joined to the 3′-hydroxyl of the neighbouring DNA strand. The essential DNA ligases in bacteria use β-nicotinamide adenine dinucleotide (β-NAD+) as their co-factor whereas those that are essential in other cells use adenosine-5′-triphosphate (ATP) as their co-factor. This observation suggests that the essential bacterial enzyme could be targeted by novel antibiotics and the complex molecular structure of β-NAD+ affords multiple opportunities for chemical modification. Several recent studies have synthesized novel derivatives and their biological activity against a range of DNA ligases has been evaluated as inhibitors for drug discovery and/or non-natural substrates for biochemical applications. Here, we review the recent advances that herald new opportunities to alter the biochemical activities of these important enzymes. The recent development of modified derivatives of nucleotides highlights that the continued combination of structural, biochemical and biophysical techniques will be useful in targeting these essential cellular enzymes. PMID:27582505

  16. Optimal DNA isolation method for detection of bacteria in clinical specimens by broad-range PCR.

    PubMed

    Rantakokko-Jalava, Kaisu; Jalava, Jari

    2002-11-01

    Broad-range amplification of bacterial DNA from clinical specimens has proved useful for the diagnosis of various bacterial infections, especially during antimicrobial treatment of the patient. Optimal sample processing protocols for diagnostic broad-range bacterial PCR should release DNA from an array of target organisms with equal efficiencies and wash out inhibitory factors from various sample types without introducing bacterial DNA contamination to the amplification reaction. In the present study, two physical cell wall disintegration methods, bead beating and sonication, for enhanced detection of organisms with difficult-to-lyse cell walls were studied. The analytical sensitivities of several commercially available DNA purification kits, which were used with and without additional cell disintegration steps, were compared by using dilution series of model bacteria. Selected purification methods were used to process routine clinical specimens in parallel with the standard phenol-ether DNA extraction, and the results obtained by bacterial PCR and sequencing with the two template preparations were compared. The method with the DNA isolation kit with the lowest detection limits from the bacterial suspensions (Masterpure) did not prove to be superior to the standard method when the two methods were applied to 69 clinical specimens. For another set of 68 clinical specimens, DNA purified with a glass fiber filter column (High Pure) with an additional sonication step yielded results well in accord with those obtained by the standard method. Furthermore, bacterial DNA was detected in four samples that remained PCR negative by the standard method, and three of these contained DNA from gram-positive pathogens. Three samples were positive by the standard method only, indicating the limitations of applying any single method to all samples.

  17. Algae-bacteria association inferred by 16S rDNA similarity in established microalgae cultures.

    PubMed

    Schwenk, Dagmar; Nohynek, Liisa; Rischer, Heiko

    2014-06-01

    Forty cultivable, visually distinct bacterial cultures were isolated from four Baltic microalgal cultures Chlorella pyrenoidosa, Scenedesmus obliquus, Isochrysis sp., and Nitzschia microcephala, which have been maintained for several years in the laboratory. Bacterial isolates were characterized with respect to morphology, antibiotic susceptibility, and 16S ribosomal DNA sequence. A total of 17 unique bacterial strains, almost all belonging to one of three families, Rhodobacteraceae, Rhizobiaceae, and Erythrobacteraceae, were subsequently isolated. The majority of isolated bacteria belong to Rhodobacteraceae. Literature review revealed that close relatives of the bacteria isolated in this study are not only often found in marine environments associated with algae, but also in lakes, sediments, and soil. Some of them had been shown to interact with organisms in their surroundings. A Basic Local Alignment Search Tool study indicated that especially bacteria isolated from the Isochrysis sp. culture were highly similar to microalgae-associated bacteria. Two of those isolates, I1 and I6, belong to the Cytophaga-Flavobacterium-Bacteroides phylum, members of which are known to occur in close communities with microalgae. An UniFrac analysis revealed that the bacterial community of Isochrysis sp. significantly differs from the other three communities.

  18. DNA methylation in leprosy-associated bacteria: Mycobacterium leprae and Corynebacterium tuberculostearicum.

    PubMed

    Hottat, F; Coene, M; Cocito, C

    1988-01-01

    The DNAs of two kinds of microorganisms from human leprosy lesion, Mycobacterium leprae and Corynebacterium tuberculostearicum (also known as "leprosy-derived corynebacterium" or LDC), have been analysed and compared with the genomes of reference bacteria of the CMN group (genera Corynebacterium, Mycobacterium and Nocardia). The guanine-plus-cytosine content (% GC) of DNA was determined by a double-labelling procedure, which is unaffected by the presence of modified and unusual bases (that alter both buoyant density and mid-melting-point determinations). Accordingly, the DNAs of seven LDC strains had GC values of 54-56 mol %, and that of armadillo-grown M. leprae a value of 54.8 +/- 0.9 mol %. Restriction patterns disclosed no methylated cytosine in the DNA sequences CCGG, GGCC, AGCT and GATC of either LDC or M. leprae DNA. N6-methyl adenine was present in the sequence GATC of all LDC strains, but was missing from the genomes of all others CMN organisms analysed, including M. leprae. By HPLC analysis of LDC-DNA hydrolysates, it was found that N6-methyladenine amounted to 1.8% of total DNA adenine, and was present exclusively within GATC sequences, which appeared all to be methylated. It is concluded that LDC represent a group of corynebacteria endowed with high genetic homogeneity and a unique restriction pattern, whereby their genome is easily distinguished from that of M. leprae, which has a similar base composition.

  19. Deoxynybomycins inhibit mutant DNA gyrase and rescue mice infected with fluoroquinolone-resistant bacteria

    PubMed Central

    Parkinson, Elizabeth I.; Bair, Joseph S.; Nakamura, Bradley A.; Lee, Hyang Y.; Kuttab, Hani I.; Southgate, Emma H.; Lezmi, Stéphane; Lau, Gee W.; Hergenrother, Paul J.

    2015-01-01

    Fluoroquinolones are one of the most commonly prescribed classes of antibiotics, but fluoroquinolone resistance (FQR) is widespread and increasing. Deoxynybomycin (DNM) is a natural-product antibiotic with an unusual mechanism of action, inhibiting the mutant DNA gyrase that confers FQR. Unfortunately, isolation of DNM is difficult and DNM is insoluble in aqueous solutions, making it a poor candidate for development. Here we describe a facile chemical route to produce DNM and its derivatives. These compounds possess excellent activity against FQR methicillin-resistant Staphylococcus aureus and vancomycin-resistant Enterococci clinical isolates and inhibit mutant DNA gyrase in-vitro. Bacteria that develop resistance to DNM are re-sensitized to fluoroquinolones, suggesting that resistance that emerges to DNM would be treatable. Using a DNM derivative, the first in-vivo efficacy of the nybomycin class is demonstrated in a mouse infection model. Overall, the data presented suggest the promise of DNM derivatives for the treatment of FQR infections. PMID:25907309

  20. [Differentiation of bacteria of the genus Aeromonas from other representatives of the Vibrionaceae family on the basis of their DNA].

    PubMed

    Levanova, G F; Lavrovskaia, V M; Shvetsov, Iu P

    1980-08-01

    By comparing the data on the nucleotide composition of DNA, and the phenotypic characteristics, most of the representatives of the genus Aeromonas were clearly differentiated from NAG vibrios isolated in the process of sanitary control of the environment. At the same time some microorganisms with the Aeromonas phenotype, but having a different DNA structure were detected. The use of the method of molecular DNA hybridization indicated that these bacteria were the taxons analogous to the genus Aeromonas.

  1. Classification of Plant Associated Bacteria Using RIF, a Computationally Derived DNA Marker

    PubMed Central

    Schneider, Kevin L.; Marrero, Glorimar; Alvarez, Anne M.; Presting, Gernot G.

    2011-01-01

    A DNA marker that distinguishes plant associated bacteria at the species level and below was derived by comparing six sequenced genomes of Xanthomonas, a genus that contains many important phytopathogens. This DNA marker comprises a portion of the dnaA replication initiation factor (RIF). Unlike the rRNA genes, dnaA is a single copy gene in the vast majority of sequenced bacterial genomes, and amplification of RIF requires genus-specific primers. In silico analysis revealed that RIF has equal or greater ability to differentiate closely related species of Xanthomonas than the widely used ribosomal intergenic spacer region (ITS). Furthermore, in a set of 263 Xanthomonas, Ralstonia and Clavibacter strains, the RIF marker was directly sequenced in both directions with a success rate approximately 16% higher than that for ITS. RIF frameworks for Xanthomonas, Ralstonia and Clavibacter were constructed using 682 reference strains representing different species, subspecies, pathovars, races, hosts and geographic regions, and contain a total of 109 different RIF sequences. RIF sequences showed subspecific groupings but did not place strains of X. campestris or X. axonopodis into currently named pathovars nor R. solanacearum strains into their respective races, confirming previous conclusions that pathovar and race designations do not necessarily reflect genetic relationships. The RIF marker also was sequenced for 24 reference strains from three genera in the Enterobacteriaceae: Pectobacterium, Pantoea and Dickeya. RIF sequences of 70 previously uncharacterized strains of Ralstonia, Clavibacter, Pectobacterium and Dickeya matched, or were similar to, those of known reference strains, illustrating the utility of the frameworks to classify bacteria below the species level and rapidly match unknown isolates to reference strains. The RIF sequence frameworks are available at the online RIF database, RIFdb, and can be queried for diagnostic purposes with RIF sequences obtained

  2. Relationships between 16S-23S rRNA gene internal transcribed spacer DNA and genomic DNA similarities in the taxonomy of phototrophic bacteria

    NASA Astrophysics Data System (ADS)

    Okamura, K.; Hisada, T.; Takata, K.; Hiraishi, A.

    2013-04-01

    Rapid and accurate identification of microbial species is essential task in microbiology and biotechnology. In prokaryotic systematics, genomic DNA-DNA hybridization is the ultimate tool to determine genetic relationships among bacterial strains at the species level. However, a practical problem in this assay is that the experimental procedure is laborious and time-consuming. In recent years, information on the 16S-23S rRNA gene internal transcribed spacer (ITS) region has been used to classify bacterial strains at the species and intraspecies levels. It is unclear how much information on the ITS region can reflect the genome that contain it. In this study, therefore, we evaluate the quantitative relationship between ITS DNA and entire genomic DNA similarities. For this, we determined ITS sequences of several species of anoxygenic phototrophic bacteria belonging to the order Rhizobiales, and compared with DNA-DNA relatedness among these species. There was a high correlation between the two genetic markers. Based on the regression analysis of this relationship, 70% DNA-DNA relatedness corresponded to 92% ITS sequence similarity. This suggests the usefulness of the ITS sequence similarity as a criterion for determining the genospecies of the phototrophic bacteria. To avoid the effects of polymorphism bias of ITS on similarities, PCR products from all loci of ITS were used directly as genetic probes for comparison. The results of ITS DNA-DNA hybridization coincided well with those of genomic DNA-DNA relatedness. These collective data indicate that the whole ITS DNA-DNA similarity can be used as an alternative to genomic DNA-DNA similarity.

  3. Use of DNA probes to study tetracycline resistance determinants in gram-negative bacteria from swine

    SciTech Connect

    Lee, C.Y.

    1989-01-01

    Specific {sup 32}P-labeled DNA probes were prepared and used to evaluate the distribution of tetracycline resistance determinants carried by gram-negative enteric bacteria isolated from pigs in 3 swine herds with different histories of antibiotic exposure. Plasmid DNA, ranging in size from 2.1 to 186 Kb, was observed in over 84% of 114 isolates studied. Two of 78 tetracycline resistant strains did not harbor plasmids. The DNA probes were isolated from plasmids pSL18, pRT29/Tn10, pBR322 and pSL106, respectively, and they represented class A, B, C and D tetracycline resistance determinants. Hybridization conditions using 0.5X SSPE at 65{degrees}C minimize cross-hybridization between the different class of tetracycline resistance genes. Cross-hybridization between class A and class C determinants could be distinguished by simultaneous comparison of the intensity of their hybridization signals. Plasmids from over 44% of the tetracycline resistant isolates did not hybridize to DNA probes for the determinants tested. Class B determinant occurred more frequently than class A or C. None of the isolates hybridized with the class D probe.

  4. The use of genomic DNA fingerprinting in studies of the epidemiology of bacteria in periodontitis.

    PubMed

    Genco, R J; Loos, B G

    1991-07-01

    Recent studies of microbial epidemiology emphasizing the genetic organization and distribution of organisms associated with orofacial infections have led to new insights into the possible origins of pathogenicity. Studies into genetic heterogeneity, acquisition and transmission of these organisms have been markedly advanced by the utilization of the powerful technique of genomic DNA fingerprinting. Characteristic fingerprints for each bacterial isolate can be produced by cleavage of high molecular weight genomic DNA by restriction endonucleases. It is assumed that each DNA fingerprint represents a clonal type. In this report, we review and analyze studies of the epidemiology of bacteria associated with orofacial infections with an emphasis on periodontal disease. Studies of nontypable (NT) Haemophilus influenzae associated with recurrent otitis media illustrate the utility of this technique. DNA fingerprinting clearly demonstrates genetic heterogeneity of NT H. influenzae isolates, and clonality of infection of any individual. Furthermore, DNA fingerprinting has shown that the same clonal type is seen in siblings concurrently suffering from otitis media, suggesting horizontal transmission within the family. Studies of mutans Streptococci also show extensive genetic heterogeneity and show vertical transmission of a predominant clonal type only from mother to infant, but not from father to infant. Studies of Actinobacillus actinomycetemcomitans show considerable genetic heterogeneity among monkey isolates. Thus far, three clonal types have been reported with DNA fingerprinting among isolates from periodontal patients, but additional genetic heterogeneity can be found using specific DNA fragments as probes in hybridization experiments. Intrafamilial transmission of A. actinomycetemcomitans has been demonstrated. Porphyromonas (Bacteroides) gingivalis shows extensive genetic heterogeneity and case reports suggest clonal infection of any one individual. In contrast

  5. A sensitive electrochemical DNA biosensor for specific detection of Enterobacteriaceae bacteria by Exonuclease III-assisted signal amplification.

    PubMed

    Luo, Caihui; Tang, Hua; Cheng, Wei; Yan, Li; Zhang, Decai; Ju, Huangxian; Ding, Shijia

    2013-10-15

    A specific and sensitive methodology was developed successfully for quantitative detection of Enterobacteriaceae bacteria by integrating Exonuclease III-assisted target recycling amplification with a simple electrochemical DNA biosensor. After target DNA hybridizes with capture DNA, Exonuclease III can selectively digest the capture DNA, which releases the target to undergo a new hybridization and cleavage cycle on sensor surface, leading to a successful target recycling. Finally, the left capture DNA is recognized by detection probe to produce the detectable signal, which decreases with the increasing target DNA concentration. Under the optimal conditions, the proposed strategy could detect target DNA down to 8.7 fM with a linear range from 0.01 pM to 1 nM, showing high sensitivity. Meanwhile, the sensing strategy was successfully used for detection of Enterobacteriaceae bacteria down to 40 CFU mL⁻¹ in milk samples. This strategy presented a simple, rapid and sensitive platform for Enterobacteriaceae bacteria detection and would become a versatile and powerful tool for food safety, biothreat detection and environmental monitoring.

  6. Advantages and limitations of ribosomal RNA PCR and DNA sequencing for identification of bacteria in cardiac valves of danish patients.

    PubMed

    Kemp, Michael; Bangsborg, Jette; Kjerulf, Anne; Schmidt, Thomas Andersen; Christensen, John; Irmukhamedov, Akhmadjon 6; Bruun, Niels Eske; Dargis, Rimtas; Andresen, Keld; Christensen, Jens Jørgen

    2013-01-01

    Studies on the value of culture-independent molecular identification of bacteria in cardiac valves are mostly restricted to comparing agreement of identification to what is obtained by culture to the number of identified bacteria in culture-negative cases. However, evaluation of the usefulness of direct molecular identification should also address weaknesses, their relevance in the given setting, and possible improvements. In this study cardiac valves from 56 Danish patients referred for surgery for infective endocarditis were analysed by microscopy and culture as well as by PCR targeting part of the bacterial 16S rRNA gene followed by DNA sequencing of the PCR product. PCR and DNA sequencing identified significant bacteria in 49 samples from 43 patients, including five out of 13 culture-negative cases. No rare, exotic, or intracellular bacteria were identified. There was a general agreement between bacterial identity obtained by ribosomal PCR and DNA sequencing from the valves and bacterial isolates from blood culture. However, DNA sequencing of the 16S rRNA gene did not discriminate well among non-haemolytic streptococci, especially within the Streptococcus mitis group. Ribosomal PCR with subsequent DNA sequencing is an efficient and reliable method of identifying the cause of IE, but exact species identification of some of the most common causes, i.e. non-haemolytic streptococci, may be improved with other molecular methods. PMID:24403979

  7. [Localization of denitrification genes in plasmid DNA of bacteria Azospirillum brasilense].

    PubMed

    Petrova, L P; Varshalomidze, O É; Shelud'ko, A V; Katsy, E I

    2010-07-01

    In 85-Mda plasmid (p85) of plant-associated bacteria Azospirillum brasilense Sp245 model strain, the genes encoding copper-containing nitrite reductase (nirK); heterodimeric NO-reductase (norCB); NorQ and NorD proteins affecting synthesis and (or) activation of NirK and (or) NO-reductase (norQD); catalytic subunit I ofcytochrom c oxidase (CccoN); presumable NO sensor carrying two hemeerythrine domains (orf181); and an enzyme required for synthesis of presumable NO antagonist, homocystein (metC) were identified. In the same region of p85, orf293 encoding transcriptional regulator of LysR type, orf208 whose protein product carries a formylmethanofuran dehydrogenase subunit E domain, and an orf164-encoding conservative secretory protein with unknown function were also found. Localization of a set of denitrification genes in the plasmid DNA A. brasilense Sp245 adjacent to IS elements ISAzba1 and ISAzba2 indicates potential mobility of these genes and high probability of their horizontal transfer among populations of rhizospheric bacteria. A site homologous to p85 nirK-orf208-orf181 genes was detected in the 115 kb plasmid of A. brasilense Sp7 type strain. PMID:20795494

  8. [Localization of denitrification genes in plasmid DNA of bacteria Azospirillum brasilense].

    PubMed

    Petrova, L P; Varshalomidze, O É; Shelud'ko, A V; Katsy, E I

    2010-07-01

    In 85-Mda plasmid (p85) of plant-associated bacteria Azospirillum brasilense Sp245 model strain, the genes encoding copper-containing nitrite reductase (nirK); heterodimeric NO-reductase (norCB); NorQ and NorD proteins affecting synthesis and (or) activation of NirK and (or) NO-reductase (norQD); catalytic subunit I ofcytochrom c oxidase (CccoN); presumable NO sensor carrying two hemeerythrine domains (orf181); and an enzyme required for synthesis of presumable NO antagonist, homocystein (metC) were identified. In the same region of p85, orf293 encoding transcriptional regulator of LysR type, orf208 whose protein product carries a formylmethanofuran dehydrogenase subunit E domain, and an orf164-encoding conservative secretory protein with unknown function were also found. Localization of a set of denitrification genes in the plasmid DNA A. brasilense Sp245 adjacent to IS elements ISAzba1 and ISAzba2 indicates potential mobility of these genes and high probability of their horizontal transfer among populations of rhizospheric bacteria. A site homologous to p85 nirK-orf208-orf181 genes was detected in the 115 kb plasmid of A. brasilense Sp7 type strain.

  9. Role of Protein Phosphorylation in the Regulation of Cell Cycle and DNA-Related Processes in Bacteria

    PubMed Central

    Garcia-Garcia, Transito; Poncet, Sandrine; Derouiche, Abderahmane; Shi, Lei; Mijakovic, Ivan; Noirot-Gros, Marie-Françoise

    2016-01-01

    In all living organisms, the phosphorylation of proteins modulates various aspects of their functionalities. In eukaryotes, protein phosphorylation plays a key role in cell signaling, gene expression, and differentiation. Protein phosphorylation is also involved in the global control of DNA replication during the cell cycle, as well as in the mechanisms that cope with stress-induced replication blocks. Similar to eukaryotes, bacteria use Hanks-type kinases and phosphatases for signal transduction, and protein phosphorylation is involved in numerous cellular processes. However, it remains unclear whether protein phosphorylation in bacteria can also regulate the activity of proteins involved in DNA-mediated processes such as DNA replication or repair. Accumulating evidence supported by functional and biochemical studies suggests that phospho-regulatory mechanisms also take place during the bacterial cell cycle. Recent phosphoproteomics and interactomics studies identified numerous phosphoproteins involved in various aspect of DNA metabolism strongly supporting the existence of such level of regulation in bacteria. Similar to eukaryotes, bacterial scaffolding-like proteins emerged as platforms for kinase activation and signaling. This review reports the current knowledge on the phosphorylation of proteins involved in the maintenance of genome integrity and the regulation of cell cycle in bacteria that reveals surprising similarities to eukaryotes. PMID:26909079

  10. Role of Protein Phosphorylation in the Regulation of Cell Cycle and DNA-Related Processes in Bacteria.

    PubMed

    Garcia-Garcia, Transito; Poncet, Sandrine; Derouiche, Abderahmane; Shi, Lei; Mijakovic, Ivan; Noirot-Gros, Marie-Françoise

    2016-01-01

    In all living organisms, the phosphorylation of proteins modulates various aspects of their functionalities. In eukaryotes, protein phosphorylation plays a key role in cell signaling, gene expression, and differentiation. Protein phosphorylation is also involved in the global control of DNA replication during the cell cycle, as well as in the mechanisms that cope with stress-induced replication blocks. Similar to eukaryotes, bacteria use Hanks-type kinases and phosphatases for signal transduction, and protein phosphorylation is involved in numerous cellular processes. However, it remains unclear whether protein phosphorylation in bacteria can also regulate the activity of proteins involved in DNA-mediated processes such as DNA replication or repair. Accumulating evidence supported by functional and biochemical studies suggests that phospho-regulatory mechanisms also take place during the bacterial cell cycle. Recent phosphoproteomics and interactomics studies identified numerous phosphoproteins involved in various aspect of DNA metabolism strongly supporting the existence of such level of regulation in bacteria. Similar to eukaryotes, bacterial scaffolding-like proteins emerged as platforms for kinase activation and signaling. This review reports the current knowledge on the phosphorylation of proteins involved in the maintenance of genome integrity and the regulation of cell cycle in bacteria that reveals surprising similarities to eukaryotes. PMID:26909079

  11. Ancient bacteria in permafrost soils fact or artefact? Considerations in recovering microbial DNA from geological ancient settings

    NASA Astrophysics Data System (ADS)

    Willerslev, E.

    2003-04-01

    Several recent reports claim that prokaryotic genetic sequences or viable cultures can survive for millions of years in geological settings. If substantiated, these findings could fundamentally alter views about bacterial physiology, ecology and evolution. However, both the culturing of microbes and the amplification of ancient DNA molecules from fossil remains are beset with difficulties. First, theoretical and empirical studies have shown that small DNA fragments (100 200 bp) do not survive in the geosphere for more than 104 years in temperate environments and 105 years in colder ones due to hydrolytic and oxidative damage. Therefore, the revivals of dormant bacteria with no active DNA repair from remains hundreds of thousands to millions of years old is, from a theoretical point, expected to be difficult, if not impossible. Second, the no specificity of the media used to culture micro organisms, as well as the great sensitivity of PCR, makes the risk of contamination with contemporary ubiquitous microbial cells and exogenous DNA molecules extremely high. Contamination poses risks at all stages of sample processing (e.g.) within the samples themselves, in the chemical reagents, on laboratory disposables or through the air. The high risk of contamination strongly suggests the need for standardized procedures within the field such as independent replication of results. This criterion of authenticity has not yet been full field in any of the studies claiming million year old microbial cultures or DNA. In order to tests the long-term survival of ancient bacteria DNA a study on permafrost was conducted using ancient DNA precautions, controls and criteria. Permafrost must be considered among the most promising environments for long term DNA survival due to its constant low temperatures (-10C to 12C Siberian or 20C Antarctica) and high cell numbers (107). We found that bacteria DNA could reproducibly be obtained from samples dated up to 300-400,000 years B.P. but not

  12. The influence of growth patterns on sexual size monomorphism in lemurs.

    PubMed

    Tennenhouse, E M

    2015-09-01

    The lack of sexual size dimorphism among lemurs is puzzling given the high degree of polygyny in this clade. It has been proposed that the unique ecological conditions of Madagascar favour rapid completion of growth, limiting the opportunities for bimaturism and sexual size dimorphism in lemurs. Using recently compiled large data sets on many species across the lemur clade, I examined the prevalence of sexual size monomorphism of body mass among lemurs and tested the hypothesis that limited growth durations constrain sexual size dimorphism. I used segmented regression analyses to accurately model growth in each species. The majority of species analysed exhibited a period of rapid growth followed by a distinct period of slow growth prior to attainment of adult body mass. Whereas the first period of growth was constrained by the need to attain the majority of adult body mass prior to the onset of the infant's first dry season, the subsequent period of slow growth was unconstrained and sufficiently long to promote sexual bimaturism. Sex differences in the duration and rate of growth during this second growth phase appeared to account for the sexual size dimorphism exhibited by three lemur species. Therefore, constraints on growth processes do not limit sexual size dimorphism in lemurs, and other explanations for the prevalence of sexual size monomorphism in this clade should be examined. The importance of considering ontogeny in future investigations of sexual size monomorphism in lemurs is highlighted. PMID:26134876

  13. The use of the terms monomorphic adenoma, basal cell adenoma, and canalicular adenoma as applied to salivary gland tumors.

    PubMed

    Gardner, D G; Daley, T D

    1983-12-01

    There is considerable confusion in the literature concerning the terms monomorphic adenoma, basal cell adenoma, and canalicular adenoma. This article traces the history of these terms as applied to the pathology of salivary gland tumors and attempts to clarify their usage. It is recommended (1) that monomorphic adenoma be used, as it was originally intended, as a nosologic grouping for all benign epithelial salivary gland tumors that are not pleomorphic adenomas, (2) that basal cell adenoma be used to identify a specific entity that is one component of the monomorphic adenoma group and exhibits a number of histologic subtypes, and (3) that canalicular adenoma be used to describe another entity, distinct from basal cell adenomas but also belonging to the monomorphic adenoma group.

  14. Bacteria and bacterial DNA in atherosclerotic plaque and aneurysmal wall biopsies from patients with and without periodontitis

    PubMed Central

    Armingohar, Zahra; Jørgensen, Jørgen J.; Kristoffersen, Anne Karin; Abesha-Belay, Emnet; Olsen, Ingar

    2014-01-01

    Background Several studies have reported an association between chronic periodontitis (CP) and cardiovascular diseases. Detection of periodontopathogens, including red complex bacteria (RCB), in vascular lesions has suggested these bacteria to be involved in the pathogenesis of atherosclerosis and abdominal aortic aneurysms. Objective In this study, we investigate bacteria and their DNA in vascular biopsies from patients with vascular diseases (VD; i.e. abdominal aortic aneurysms, atherosclerotic carotid, and common femoral arteries), with and without CP. Methods DNA was extracted from vascular biopsies selected from 40 VD patients: 30 with CP and 10 without CP. The V3-V5 region of the 16S rDNA (V3-V5) was polymerase chain reaction (PCR)-amplified, and the amplicons were cloned into Escherichia coli, sequenced, and classified (GenBank and the Human Oral Microbiome database). Species-specific primers were used for the detection of Porphyromonas gingivalis. In addition, 10 randomly selected vascular biopsies from the CP group were subjected to scanning electron microscopy (SEM) for visualization of bacteria. Checkerboard DNA–DNA hybridization was performed to assess the presence of RCB in 10 randomly selected subgingival plaque samples from CP patients. Results A higher load and mean diversity of bacteria were detected in vascular biopsies from VD patients with CP compared to those without CP. Enterobacteriaceae were frequently detected in vascular biopsies together with cultivable, commensal oral, and not-yet-cultured bacterial species. While 70% of the subgingival plaque samples from CP patients showed presence of RCB, only P. gingivalis was detected in one vascular biopsy. Bacterial cells were seen in all 10 vascular biopsies examined by SEM. Conclusions A higher bacterial load and more diverse colonization were detected in VD lesions of CP patients as compared to patients without CP. This indicated that a multitude of bacterial species both from the gut and the

  15. Development of a DNA macroarray for simultaneous detection of multiple foodborne pathogenic bacteria in fresh chicken meat.

    PubMed

    Kupradit, Chanida; Rodtong, Sureelak; Ketudat-Cairns, Mariena

    2013-12-01

    A DNA macroarray was developed to provide the ability to detect multiple foodborne pathogens in fresh chicken meat. Probes targeted to the 16S rRNA and genus- and species-specific genes, including fimY, ipaH, prfA, and uspA, were selected for the specific detection of Salmonella spp., Shigella spp., Listeria monocytogenes, and Escherichia coli, respectively. The combination of target gene amplification by PCR and a DNA macroarray in our system was able to distinguish all target bacteria from pure cultures with a detection sensitivity of 10⁵ c.f.u. ml⁻¹. The DNA macroarray was also applied to 10 fresh chicken meat samples. The assay validation demonstrated that by combining the enrichment steps for the target bacteria and the DNA macroarray, all 4 target bacteria could be detected simultaneously from the fresh chicken samples. The sensitivity of L. monocytogenes and Shigella boydii detection in the fresh chicken samples was at least 10 and 3 c.f.u. of the initial contamination in 25 g samples, respectively. The advantages of our developed protocol are high accuracy and time reduction when compared to conventional culture. The macroarray developed in our investigation was cost effective compared to modern oligonucleotide microarray techniques because there was no expensive equipment required for the detection of multiple foodborne pathogens.

  16. Amplification of fluorescently labelled DNA within gram-positive and acid-fast bacteria.

    PubMed

    Vaid, A; Bishop, A H

    1999-10-01

    Representative organisms from a variety of Gram-positive genera were subjected to varying regimes in order to optimise the intracellular amplification of DNA. The bacteria were subjected to treatments with paraformaldehyde, muramidases and mild acid hydrolysis to discover which regime made each organism permeable to the amplification reagents yet allowed retention of the fluorescein-labelled amplified products within the cell. Scanning electron micrographs were used to corroborate the effectiveness of the treatments, as seen by fluorescent photomicrographs, with the damage caused to the bacterial walls. A combination of mutanolysin and lysozyme was found most effective for Bacillus cereus, whereas permeabilisation of Streptomyces coelicolor, Lactococcus lactis and Clostridium sporogenes was most effective when exposed to lysozyme only. Surprisingly, direct amplification with no pre-treatment gave the brightest fluorescence in Mycobacterium phlei. Comparing the techniques of whole cell PCR, primed in situ labelling (PRINS), and cycle PRINS showed that under the conditions used the strongest intensity of fluorescence was obtained with in situ PCR; only L. lactis and M. phlei produced signals with cycle PRINS, fluorescence was not seen for any of the organisms with PRINS.

  17. Multiple DNA Extractions Coupled with Stable-Isotope Probing of Anthracene-Degrading Bacteria in Contaminated Soil▿†

    PubMed Central

    Jones, Maiysha D.; Singleton, David R.; Sun, Wei; Aitken, Michael D.

    2011-01-01

    In many of the DNA-based stable-isotope probing (SIP) studies published to date in which soil communities were investigated, a single DNA extraction was performed on the soil sample, usually using a commercial DNA extraction kit, prior to recovering the 13C-labeled (heavy) DNA by density-gradient ultracentrifugation. Recent evidence suggests, however, that a single extraction of a soil sample may not lead to representative recovery of DNA from all of the organisms in the sample. To determine whether multiple DNA extractions would affect the DNA yield, the eubacterial 16S rRNA gene copy number, or the identification of anthracene-degrading bacteria, we performed seven successive DNA extractions on the same aliquot of contaminated soil either untreated or enriched with [U-13C]anthracene. Multiple extractions were necessary to maximize the DNA yield and 16S rRNA gene copy number from both untreated and anthracene-enriched soil samples. Sequences within the order Sphingomonadales, but unrelated to any previously described genus, dominated the 16S rRNA gene clone libraries derived from 13C-enriched DNA and were designated “anthracene group 1.” Sequences clustering with Variovorax spp., which were also highly represented, and sequences related to the genus Pigmentiphaga were newly associated with anthracene degradation. The bacterial groups collectively identified across all seven extracts were all recovered in the first extract, although quantitative PCR analysis of SIP-identified groups revealed quantitative differences in extraction patterns. These results suggest that performing multiple DNA extractions on soil samples improves the extractable DNA yield and the number of quantifiable eubacterial 16S rRNA gene copies but have little qualitative effect on the identification of the bacterial groups associated with the degradation of a given carbon source by SIP. PMID:21398486

  18. Evaluation of a facile method of template DNA preparation for PCR-based detection and typing of lactic acid bacteria.

    PubMed

    Singh, Atul Kumar; Ramesh, Aiyagari

    2009-08-01

    The objective of our investigation was to develop a convenient and reliable method of generating template DNA for routine PCR-based detection and typing of lactic acid bacteria (LAB). Template DNA extracted from Lactobacillus, Lactococcus, Pediococcus and Leuconostoc using a combination of urea, SDS and NaOH yielded amplicons of expected size in PCR with genus-specific primers. Apart from LAB, the proposed method could also be adopted to generate PCR-compatible template DNA from a number of Gram-positive and Gram-negative bacterial strains. DNA template prepared by the proposed method from various standard strains of Lactobacillus sp. also generated discriminating fingerprints with BOXA1R primer in rep-PCR. A significant finding of the investigation was that a comparable banding profile of LAB strains was obtained in rep-PCR using template DNA prepared by urea-SDS-NaOH method and a commercially available DNA isolation kit. This was further evidenced by high dice coefficient values obtained in the range of 81.8-96.7 when cluster analysis was performed by UPGAMA method. The application potential of this DNA extraction method for PCR-based direct detection of LAB in fermented food samples such as dahi, idli batter and salt-fermented cucumber was validated by detecting specific amplicons of LAB genera in the fermented samples. The applicability of the proposed template DNA extraction method was further substantiated when 29 bacteriocinogenic LAB strains (Bac+) previously detected in salt-fermented cucumber by PCR [Singh, A.K., Ramesh, A., 2008. Succession of dominant and antagonistic lactic acid bacteria in fermented cucumber: Insights from a PCR-based approach. Food. Microbiol. 25, 278-287] generated differentiating fingerprints in BOX element based rep-PCR and formed clusters with reference LAB strains. PMID:19465247

  19. Combined immunofluorescence-DNA-fluorescence staining technique for enumeration of Thiobacillus ferrooxidans in a population of acidophilic bacteria

    SciTech Connect

    Muyzer, G.; De Bruyn, A.; Schmedding, D.J.M.; Bos, P.; Westbroek, P.; Kuenen, G.J.

    1987-04-01

    An antiserum raised against whole cells of Thiobacillus ferroxidans was allowed to react with a variety of acidophilic and nonacidophilic bacteria in an enzyme-linked immunosorbent assay and an indirect immunofluorescence assay. Both experiments demonstrated that the antiserum was specific at the species level. This preparation was used to evaluate the role of T. ferroooxidans in the microbial desulfurization process. Leaching experiments were performed, and the numbers of T. ferrooxidans cells and other bacteria were estimated by using a combined immunofluorescence-DNA-fluorescence staining technique that was adapted for this purpose. Nonsterile coal samples inoculated with T. ferrooxidans yielded high concentrations of soluble iron after 16 days. After this period, however, T. ferrooxidans cells could no longer be detected by the immunofluorescence assay, whereas the DNA-fluorescence staining procedure demonstrated a large number of microorganisms on the coal particles. These results indicate that T. ferrooxidans is removed by competition with different acidophilic microorganisms that were originally present on the coal.

  20. Rapid identification and classification of bacteria by 16S rDNA restriction fragment melting curve analyses (RFMCA).

    PubMed

    Rudi, Knut; Kleiberg, Gro H; Heiberg, Ragnhild; Rosnes, Jan T

    2007-08-01

    The aim of this work was to evaluate restriction fragment melting curve analyses (RFMCA) as a novel approach for rapid classification of bacteria during food production. RFMCA was evaluated for bacteria isolated from sous vide food products, and raw materials used for sous vide production. We identified four major bacterial groups in the material analysed (cluster I-Streptococcus, cluster II-Carnobacterium/Bacillus, cluster III-Staphylococcus and cluster IV-Actinomycetales). The accuracy of RFMCA was evaluated by comparison with 16S rDNA sequencing. The strains satisfying the RFMCA quality filtering criteria (73%, n=57), with both 16S rDNA sequence information and RFMCA data (n=45) gave identical group assignments with the two methods. RFMCA enabled rapid and accurate classification of bacteria that is database compatible. Potential application of RFMCA in the food or pharmaceutical industry will include development of classification models for the bacteria expected in a given product, and then to build an RFMCA database as a part of the product quality control. PMID:17367680

  1. Seasonal Variation in Parental Care Drives Sex-Specific Foraging by a Monomorphic Seabird

    PubMed Central

    Burke, Chantelle M.; Montevecchi, William A.; Regular, Paul M.

    2015-01-01

    Evidence of sex-specific foraging in monomorphic seabirds is increasing though the underlying mechanisms remain poorly understood. We investigate differential parental care as a mechanism for sex-specific foraging in monomorphic Common Murres (Uria aalge), where the male parent alone provisions the chick after colony departure. Using a combination of geolocation-immersion loggers and stable isotopes, we assess two hypotheses: the reproductive role specialization hypothesis and the energetic constraint hypothesis. We compare the foraging behavior of females (n = 15) and males (n = 9) during bi-parental at the colony, post-fledging male-only parental care and winter when parental care is absent. As predicted by the reproductive role specialization hypothesis, we found evidence of sex-specific foraging during post-fledging only, the stage with the greatest divergence in parental care roles. Single-parenting males spent almost twice as much time diving per day and foraged at lower quality prey patches relative to independent females. This implies a potential energetic constraint for males during the estimated 62.8 ± 8.9 days of offspring dependence at sea. Contrary to the predictions of the energetic constraint hypothesis, we found no evidence of sex-specific foraging during biparental care, suggesting that male parents did not forage for their own benefit before colony departure in anticipation of post-fledging energy constraints. We hypothesize that unpredictable prey conditions at Newfoundland colonies in recent years may limit male parental ability to allocate additional time and energy to self-feeding during biparental care, without compromising chick survival. Our findings support differential parental care as a mechanism for sex-specific foraging in monomorphic murres, and highlight the need to consider ecological context in the interpretation of sex-specific foraging behavior. PMID:26575646

  2. Salivary monomorphic adenomas of dermal analogue type: report of two cases.

    PubMed

    Sparrow, S A; Frost, F A

    1993-01-01

    Two cases of monomorphic adenoma, dermal analogue type, are presented illustrating fine-needle aspiration cytology and subsequent histopathology. Aspiration findings were similar in both cases, demonstrating aggregates of mildly disorganised small regular epithelial cells bordered by a thick rim of basement membrane material. In the first case seen, there was a prominent lymphoid component which gave rise to the false suspicion that the lesion was lymphoepithelial in nature. The cytological features of this tumour are characteristic, however, and their recognition is important to enable distinction from malignancy (e.g., adenoid cystic carcinoma), other benign neoplasms (e.g., plemorphic adenoma), and non-neoplastic conditions.

  3. An improved method for extracting bacteria from soil for high molecular weight DNA recovery and BAC library construction.

    PubMed

    Liu, Juan; Li, Jingquan; Feng, Li; Cao, Hui; Cui, Zhongli

    2010-12-01

    Separation of bacterial cells from soil is a key step in the construction of metagenomic BAC libraries with large DNA inserts. Our results showed that when combined with sodium pyro-phosphate and homogenization for soil dispersion, sucrose density gradient centrifugation (SDGC) was more effective at separating bacteria from soil than was low speed centrifugation (LSC). More than 70% of the cells, along with some soil colloids, were recovered with one round of centrifugation. A solution of 0.8% NaCl was used to resuspend these cell and soil pellets for purification with nycodenz density gradient centrifugation (NDGC). After purification, more than 30% of the bacterial cells in the primary soil were extracted. This procedure effectively removed soil contamination and yielded sufficient cells for high molecular weight (HMW) DNA isolation. Ribosomal intergenic spacer analysis (RISA) showed that the microbial community structure of the extracted cells was similar to that of the primary soil, suggesting that this extraction procedure did not significantly change the the soil bacteria community structure. HMW DNA was isolated from bacterial cells extracted from red soil for metagenomic BAC library construction. This library contained DNA inserts of more than 200 Mb with an average size of 75 kb.

  4. Rapid and sensitive identification of marine bacteria by an improved in situ DNA hybridization chain reaction (quickHCR-FISH).

    PubMed

    Yamaguchi, Tsuyoshi; Fuchs, Bernhard Maximilian; Amann, Rudolf; Kawakami, Shuji; Kubota, Kengo; Hatamoto, Masashi; Yamaguchi, Takashi

    2015-09-01

    Catalyzed reporter deposition-fluorescence in situ hybridization (CARD-FISH) with rRNA-targeted oligonucleotide probes has significantly improved the identification of microorganisms in various environmental samples. However, one of the major constraints of CARD-FISH is the low probe penetration due to the high molecular weight of the horseradish peroxidase (HRP) label. Recently, this limitation has been overcome by a novel signal amplification approach termed in situ DNA-hybridization chain reaction (in situ DNA-HCR). In this study, we present an improved and accelerated in situ DNA-HCR protocol (quickHCR-FISH) with increased signal intensity, which was approximately 2 times higher than that of standard in situ DNA-HCR. In addition, the amplification time was only 15 min for the extension of amplifier probes from the initiator probe compared to 2h in the original protocol. The quickHCR-FISH was successfully tested for the quantification of marine bacteria with low rRNA contents in both seawater and sediment samples. It was possible to detect the same number of marine bacteria with quickHCR-FISH compared to CARD-FISH within only 3h. Thus, this newly developed protocol could be an attractive alternative to CARD-FISH for the detection and visualization of microorganisms in their environmental context.

  5. Molecular structure of monomorphic peptide fibrils within a kinetically trapped hydrogel network

    PubMed Central

    Nagy-Smith, Katelyn; Moore, Eric; Schneider, Joel; Tycko, Robert

    2015-01-01

    Most, if not all, peptide- and protein-based hydrogels formed by self-assembly can be characterized as kinetically trapped 3D networks of fibrils. The propensity of disease-associated amyloid-forming peptides and proteins to assemble into polymorphic fibrils suggests that cross-β fibrils comprising hydrogels may also be polymorphic. We use solid-state NMR to determine the molecular and supramolecular structure of MAX1, a de novo designed gel-forming peptide, in its fibrillar state. We find that MAX1 adopts a β-hairpin conformation and self-assembles with high fidelity into a double-layered cross-β structure. Hairpins assemble with an in-register Syn orientation within each β-sheet layer and with an Anti orientation between layers. Surprisingly, although the MAX1 fibril network is kinetically trapped, solid-state NMR data show that fibrils within this network are monomorphic and most likely represent the thermodynamic ground state. Intermolecular interactions not available in alternative structural arrangements apparently dictate this monomorphic behavior. PMID:26216960

  6. Monomorphous Plurihormonal Pituitary Adenoma of Pit-1 Lineage in a Giant Adolescent with Central Hyperthyroidism.

    PubMed

    Pereira, Bernardo Dias; Raimundo, Luísa; Mete, Ozgur; Oliveira, Ana; Portugal, Jorge; Asa, Sylvia L

    2016-03-01

    Thyrotropin (TSH)-secreting pituitary adenomas are exceedingly rare at the pediatric age and no cases of co-secretion with other pituitary hormones in these tumors have been described in this age range. We present a case of a monomorphous plurihormonal pituitary adenoma that co-secreted TSH and GH in a pediatric patient. A 13-year-old male presented with increasing height velocity (17.75 cm/year, 9.55SD), weight loss, and visual impairment. Initial biochemical evaluations revealed secondary hyperthyroidism. A giant pituitary tumor compressing the surrounding structures was detected by magnetic resonance, and a transsphenoidal surgery was initially performed. Pathological examinations revealed an atypical, monomorphous plurihormonal Pit-1 lineage tumor with mixed features of silent subtype 3 adenoma and acidophil stem cell adenoma. In the postoperative period, secondary hyperthyroidism recurred with high levels of both GH and IGF1. In addition, due to tumor re-growth, a multimodality treatment plan was undertaken including surgery, somatostatin analogs, and radiotherapy. We report the first pediatric case of a plurihormonal TSH- and GH-secreting pituitary adenoma, further expanding the clinical manifestations of pediatric pituitary tumors. Comprehensive pathological evaluation and close follow-up surveillance are crucial to the prompt delivery of the best therapeutic options in the context of this particularly aggressive pituitary tumor. PMID:26330191

  7. How long can culturable bacteria and total DNA persist in environmental waters? The role of sunlight and solid particles.

    PubMed

    Gutiérrez-Cacciabue, Dolores; Cid, Alicia G; Rajal, Verónica B

    2016-01-01

    In this work, sunlight inactivation of two indicator bacteria in freshwater, with and without solid particles, was studied and the persistence of culturable cells and total DNA was compared. Environmental water was used to prepare two matrices, with and without solid particles, which were spiked with Escherichia coli and Enterococcus faecalis. These matrices were used to prepare microcosm bags that were placed in two containers: one exposed to sunlight and the other in the dark. During one month, samples were removed from each container and detection was done by membrane filter technique and real-time PCR. Kinetic parameters were calculated to assess sunlight effect. Indicator bacteria without solid particles exposed to sunlight suffered an immediate decay (<4h) compared with the ones which were shielded from them. In addition, the survival of both bacteria with solid particles varied depending on the situation analyzed (T99 from 3 up to 60days), being always culturable E. coli more persistent than E. faecalis. On the other side, E. faecalis DNA persisted much longer than culturable cells (T99>40h in the dark with particles). In this case active cells were more prone to sunlight than total DNA and the protective effect of solid particles was also observed. Results highlight that the effects caused by the parameters which describe the behavior of culturable microorganisms and total DNA in water are different and must be included in simulation models but without forgetting that these parameters will also depend on bacterial properties, sensitizers, composition, type, and uses of the aquatic environment under assessment.

  8. vanA-targeted oligonucleotide DNA probe designed to monitor vancomycin- and teicoplanin-resistant bacteria in surface waters.

    PubMed

    Nakipoglu, Mustafa; Yilmaz, Fadime; Icgen, Bulent

    2016-10-01

    The glycopeptide antibiotics teicoplanin and vancomycin are common to treat severe Gram-positive bacterial infections. The gene vanA confers high-level resistance to these antibiotics, and these phenomena have been shown to be transferable. Release of vancomycin- and teicoplanin-resistant bacteria to surface waters is, therefore, of particular concern since they might proliferate and spread in different environments. Monitoring of the fate of vanA gene in these waters provides information on the exposure and potential threats of those bacteria for the environment and public health. Therefore, this study aimed at preparing a 25-mer-oligonucleotide DNA probe based on the 909 bp BamHI-ClaI fragment from Enterococcus faecium plasmids pVEF1 and pVEF2 through the use of Vector NTI Express Software. The newly designed vanA probe displayed highly specific hybridization with vanA-positive Enterococcus faecalis tested at 46 °C, 55 % formamide, and 0.020 M NaCl stringency conditions. In situ fluorescein hybridizations under the same stringency conditions were also used to monitor river water samples by using fluorescein microscopy. The results showed that the vanA-targeted oligonucleotide DNA probe prepared was not only highly specific but also quantitative tool for monitoring vancomycin- and teicoplanin-resistant bacteria in surface waters. PMID:27640164

  9. Identification of benzo[a]pyrene-metabolizing bacteria in forest soils by using DNA-based stable-isotope probing.

    PubMed

    Song, Mengke; Luo, Chunling; Jiang, Longfei; Zhang, Dayi; Wang, Yujie; Zhang, Gan

    2015-11-01

    DNA-based stable-isotope probing (DNA-SIP) was used in this study to investigate the uncultivated bacteria with benzo[a]pyrene (BaP) metabolism capacities in two Chinese forest soils (Mt. Maoer in Heilongjiang Province and Mt. Baicaowa in Hubei Province). We characterized three different phylotypes with responsibility for BaP degradation, none of which were previously reported as BaP-degrading microorganisms by SIP. In Mt. Maoer soil microcosms, the putative BaP degraders were classified as belonging to the genus Terrimonas (family Chitinophagaceae, order Sphingobacteriales), whereas Burkholderia spp. were the key BaP degraders in Mt. Baicaowa soils. The addition of metabolic salicylate significantly increased BaP degradation efficiency in Mt. Maoer soils, and the BaP-metabolizing bacteria shifted to the microorganisms in the family Oxalobacteraceae (genus unclassified). Meanwhile, salicylate addition did not change either BaP degradation or putative BaP degraders in Mt. Baicaowa. Polycyclic aromatic hydrocarbon ring-hydroxylating dioxygenase (PAH-RHD) genes were amplified, sequenced, and quantified in the DNA-SIP (13)C heavy fraction to further confirm the BaP metabolism. By illuminating the microbial diversity and salicylate additive effects on BaP degradation across different soils, the results increased our understanding of BaP natural attenuation and provided a possible approach to enhance the bioremediation of BaP-contaminated soils. PMID:26253666

  10. High-throughput DNA microarray detection of pathogenic bacteria in shallow well groundwater in the Kathmandu Valley, Nepal.

    PubMed

    Inoue, Daisuke; Hinoura, Takuji; Suzuki, Noriko; Pang, Junqin; Malla, Rabin; Shrestha, Sadhana; Chapagain, Saroj Kumar; Matsuzawa, Hiroaki; Nakamura, Takashi; Tanaka, Yasuhiro; Ike, Michihiko; Nishida, Kei; Sei, Kazunari

    2015-01-01

    Because of heavy dependence on groundwater for drinking water and other domestic use, microbial contamination of groundwater is a serious problem in the Kathmandu Valley, Nepal. This study investigated comprehensively the occurrence of pathogenic bacteria in shallow well groundwater in the Kathmandu Valley by applying DNA microarray analysis targeting 941 pathogenic bacterial species/groups. Water quality measurements found significant coliform (fecal) contamination in 10 of the 11 investigated groundwater samples and significant nitrogen contamination in some samples. The results of DNA microarray analysis revealed the presence of 1-37 pathogen species/groups, including 1-27 biosafety level 2 ones, in 9 of the 11 groundwater samples. While the detected pathogens included several feces- and animal-related ones, those belonging to Legionella and Arthrobacter, which were considered not to be directly associated with feces, were detected prevalently. This study could provide a rough picture of overall pathogenic bacterial contamination in the Kathmandu Valley, and demonstrated the usefulness of DNA microarray analysis as a comprehensive screening tool of a wide variety of pathogenic bacteria.

  11. Identification of Benzo[a]pyrene-Metabolizing Bacteria in Forest Soils by Using DNA-Based Stable-Isotope Probing

    PubMed Central

    Song, Mengke; Jiang, Longfei; Zhang, Dayi; Wang, Yujie; Zhang, Gan

    2015-01-01

    DNA-based stable-isotope probing (DNA-SIP) was used in this study to investigate the uncultivated bacteria with benzo[a]pyrene (BaP) metabolism capacities in two Chinese forest soils (Mt. Maoer in Heilongjiang Province and Mt. Baicaowa in Hubei Province). We characterized three different phylotypes with responsibility for BaP degradation, none of which were previously reported as BaP-degrading microorganisms by SIP. In Mt. Maoer soil microcosms, the putative BaP degraders were classified as belonging to the genus Terrimonas (family Chitinophagaceae, order Sphingobacteriales), whereas Burkholderia spp. were the key BaP degraders in Mt. Baicaowa soils. The addition of metabolic salicylate significantly increased BaP degradation efficiency in Mt. Maoer soils, and the BaP-metabolizing bacteria shifted to the microorganisms in the family Oxalobacteraceae (genus unclassified). Meanwhile, salicylate addition did not change either BaP degradation or putative BaP degraders in Mt. Baicaowa. Polycyclic aromatic hydrocarbon ring-hydroxylating dioxygenase (PAH-RHD) genes were amplified, sequenced, and quantified in the DNA-SIP 13C heavy fraction to further confirm the BaP metabolism. By illuminating the microbial diversity and salicylate additive effects on BaP degradation across different soils, the results increased our understanding of BaP natural attenuation and provided a possible approach to enhance the bioremediation of BaP-contaminated soils. PMID:26253666

  12. Primers and a specific DNA probe for detecting lactic acid bacteria producing 3-hydroxypropionaldehyde from glycerol in spoiled ciders.

    PubMed

    Claisse, O; Lonvaud-Funel, A

    2001-06-01

    Of the 40 strains isolated from several spoiled ciders where glycerol was degraded, 36 were identified as Lactobacillus collinoides, three were Lactobacillus hilgardii, and one was Lactobacillus mali. However, only 30 L. collinoides and two L. hilgardii could degrade glycerol. The glycerol dehydratase activity was shown. The main product of the transformation was 1.3 propanediol. Two DNA primers GD1 and GD2 were chosen in the region encoding one of the subunits of glycerol dehydratase of Citrobacter freundii, Klebsiella pneumoniae, Klebsiella oxytoca, Salmonella Typhimurium, and Clostridium pasteurianum. A 279-bp amplicon in polymerase chain reaction amplification was obtained with the genomic L. collinoides IOEB 9527 DNA as template. The amino acid sequence deduced from the amplicon DNA sequence showed a very high similarity and identity with the gene of gram-negative and C. pasteurianum species. After labeling, the amplicon was used as DNA probe in dot-blot hybridization with the genomic DNA of all the tested strains. Only strains that could degrade glycerol hybridized. Moreover, polymerase chain reactions using GDI and GD2 revealed only glycerol dehydratase genes of positive L. collinoides and L. hilgardii strains. The primers and the amplicon proved to be suitable and reliable tools to detect the lactic acid bacteria involved in the deterioration of cider. PMID:11403134

  13. DNA topoisomerase VIII: a novel subfamily of type IIB topoisomerases encoded by free or integrated plasmids in Archaea and Bacteria.

    PubMed

    Gadelle, Danièle; Krupovic, Mart; Raymann, Kasie; Mayer, Claudine; Forterre, Patrick

    2014-07-01

    Type II DNA topoisomerases are divided into two families, IIA and IIB. Types IIA and IIB enzymes share homologous B subunits encompassing the ATP-binding site, but have non-homologous A subunits catalyzing DNA cleavage. Type IIA topoisomerases are ubiquitous in Bacteria and Eukarya, whereas members of the IIB family are mostly present in Archaea and plants. Here, we report the detection of genes encoding type IIB enzymes in which the A and B subunits are fused into a single polypeptide. These proteins are encoded in several bacterial genomes, two bacterial plasmids and one archaeal plasmid. They form a monophyletic group that is very divergent from archaeal and eukaryotic type IIB enzymes (DNA topoisomerase VI). We propose to classify them into a new subfamily, denoted DNA topoisomerase VIII. Bacterial genes encoding a topoisomerase VIII are present within integrated mobile elements, most likely derived from conjugative plasmids. Purified topoisomerase VIII encoded by the plasmid pPPM1a from Paenibacillus polymyxa M1 had ATP-dependent relaxation and decatenation activities. In contrast, the enzyme encoded by mobile elements integrated into the genome of Ammonifex degensii exhibited DNA cleavage activity producing a full-length linear plasmid and that from Microscilla marina exhibited ATP-independent relaxation activity. Topoisomerases VIII, the smallest known type IIB enzymes, could be new promising models for structural and mechanistic studies.

  14. Differentiation and identification of iron-oxidizing acidophilic bacteria using cultivation techniques and amplified ribosomal DNA restriction enzyme analysis.

    PubMed

    Johnson, D Barrie; Okibe, Naoko; Hallberg, Kevin B

    2005-03-01

    Acidophilic iron-oxidizing microorganisms are important both environmentally and in biotechnological applications. Although, as a group, they are readily detected by their ability to generate ferric iron (resulting in a distinctive color change in liquid media), these microbes highly diverse phylogenetically. Various other characteristics, such as optimum growth temperature, response to organic carbon sources, and cellular morphologies, facilitate, in some cases, identification of isolates to a genus or species level, although this approach has limitations and may give erroneous results. In this study, a combined approach of using physiological traits together with amplified ribosomal DNA restriction enzyme analysis (ARDREA) has been successful in identifying all known acidophilic iron-oxidizing bacteria to the species level. Computer-generated maps were used to identify restriction enzymes that allow the differentiation of the acidophiles, and these were confirmed experimentally using authentic bacterial strains. To test further the validity of this approach, six acidophilic moderately thermophilic iron-oxidizing bacteria isolated from Montserrat (West Indies) were analysed using the ARDREA protocol. Three of the isolates were identified as Sulfobacillus acidophilus-like, and one as Sulfobacillus thermosulfidooxidans-like bacteria. The fifth isolate gave DNA digest patterns that were distinct from all known strains of iron-oxidizing acidophiles. Subsequent sequencing of the 16S rRNA genes of these isolates confirmed the identity of the four Sulfobacillus isolates, and also that the fifth isolate was a novel species. Schematic diagrams showing how ARDREA may be used to rapidly identify all known acidophilic iron-oxidizing bacteria are presented.

  15. Hybridization of cloned Rhodopseudomonas capsulata photosynthesis genes with DNA from other photosynthetic bacteria.

    PubMed Central

    Beatty, J T; Cohen, S N

    1983-01-01

    The homology of Rhodopseudomonas capsulata DNA segments carrying photosynthesis genes with sequences present in total DNA from certain other photosynthetic and non-photosynthetic bacterial species was determined by hybridization. R. capsulata DNA fragments that carry loci for production of peptide components of the photosynthetic reaction center and light-harvesting I antenna complex were found to hybridize to DNA from some photosynthetic species. However, fragments that carry carotenoid or bacteriochlorophyll biosynthesis genes showed either weak or undetectable heterospecific hybridization under the conditions employed. Images PMID:6406432

  16. Gene discovery within the planctomycete division of the domain Bacteria using sequence tags from genomic DNA libraries

    PubMed Central

    Jenkins, Cheryl; Kedar, Vishram; Fuerst, John A

    2002-01-01

    Background The planctomycetes comprise a distinct group of the domain Bacteria, forming a separate division by phylogenetic analysis. The organization of their cells into membrane-defined compartments including membrane-bounded nucleoids, their budding reproduction and complete absence of peptidoglycan distinguish them from most other Bacteria. A random sequencing approach was applied to the genomes of two planctomycete species, Gemmata obscuriglobus and Pirellula marina, to discover genes relevant to their cell biology and physiology. Results Genes with a wide variety of functions were identified in G. obscuriglobus and Pi. marina, including those of metabolism and biosynthesis, transport, regulation, translation and DNA replication, consistent with established phenotypic characters for these species. The genes sequenced were predominantly homologous to those in members of other divisions of the Bacteria, but there were also matches with nuclear genomic genes of the domain Eukarya, genes that may have appeared in the planctomycetes via horizontal gene transfer events. Significant among these matches are those with two genes atypical for Bacteria and with significant cell-biology implications - integrin alpha-V and inter-alpha-trypsin inhibitor protein - with homologs in G. obscuriglobus and Pi. marina respectively. Conclusions The random-sequence-tag approach applied here to G. obscuriglobus and Pi. marina is the first report of gene recovery and analysis from members of the planctomycetes using genome-based methods. Gene homologs identified were predominantly similar to genes of Bacteria, but some significant best matches to genes from Eukarya suggest that lateral gene transfer events between domains may have involved this division at some time during its evolution. PMID:12093378

  17. Monomorphic adenoma of minor salivary glands: a reappraisal and report of nine new cases.

    PubMed

    Levine, J; Krutchkoff, D J; Eisenberg, E

    1981-02-01

    A search of intraoral lesions on which biopsies were performed within a five-year period at the University of Connecticut disclosed 26 benign neoplasms of minor salivary gland origin, of which nine were classified as variants of monomorphic adenoma. Of these, eight were considered to be basal cell, basaloid, or canalicular types. As there was no clear histologic distinction between these histologic subtypes, and as all such lesions (basal cell, basaloid, or canalicular adenomas) displayed a noticeable predilection for the upper lip, it was concluded that the three are variants of one entity and need not be separated. It is recommended that the term basal cell adenoma henceforth be adapted to describe all of such variants.

  18. Phylogenetic Diversity of Lactic Acid Bacteria Associated with Paddy Rice Silage as Determined by 16S Ribosomal DNA Analysis

    PubMed Central

    Ennahar, Saïd; Cai, Yimin; Fujita, Yasuhito

    2003-01-01

    A total of 161 low-G+C-content gram-positive bacteria isolated from whole-crop paddy rice silage were classified and subjected to phenotypic and genetic analyses. Based on morphological and biochemical characters, these presumptive lactic acid bacterium (LAB) isolates were divided into 10 groups that included members of the genera Enterococcus, Lactobacillus, Lactococcus, Leuconostoc, Pediococcus, and Weissella. Analysis of the 16S ribosomal DNA (rDNA) was used to confirm the presence of the predominant groups indicated by phenotypic analysis and to determine the phylogenetic affiliation of representative strains. The virtually complete 16S rRNA gene was PCR amplified and sequenced. The sequences from the various LAB isolates showed high degrees of similarity to those of the GenBank reference strains (between 98.7 and 99.8%). Phylogenetic trees based on the 16S rDNA sequence displayed high consistency, with nodes supported by high bootstrap values. With the exception of one species, the genetic data was in agreement with the phenotypic identification. The prevalent LAB, predominantly homofermentative (66%), consisted of Lactobacillus plantarum (24%), Lactococcus lactis (22%), Leuconostoc pseudomesenteroides (20%), Pediococcus acidilactici (11%), Lactobacillus brevis (11%), Enterococcus faecalis (7%), Weissella kimchii (3%), and Pediococcus pentosaceus (2%). The present study, the first to fully document rice-associated LAB, showed a very diverse community of LAB with a relatively high number of species involved in the fermentation process of paddy rice silage. The comprehensive 16S rDNA-based approach to describing LAB community structure was valuable in revealing the large diversity of bacteria inhabiting paddy rice silage and enabling the future design of appropriate inoculants aimed at improving its fermentation quality. PMID:12514026

  19. Clinical and morphological features of undifferentiated monomorphous GH/TSH-secreting pituitary adenoma.

    PubMed

    Skorić, T; Korsić, M; Zarković, K; Plavsić, V; Besenski, N; Breskovac, L; Giljević, Z; Paladino, J

    1999-06-01

    A 41-year-old male presented with progressive visual defects, acromegaly and hyperthyroidism. After clinical evaluation a giant GH/TSH-secreting pituitary adenoma was diagnosed. Administration of the somatostatin analog octreotide at doses of 150 microg s.c. per day inhibited the secretion of both GH and TSH. A three-week treatment with octreotide prior to surgery led to slight visual improvement and CT scan showed some new necrotic areas within the tumor mass. Transcranial surgery was performed. By immunohistochemical analyses of the adenoma tissue GH, prolactin and beta-chorionic gonadotropin were detected; TSH was negative. Electron microscopy revealed an undifferentiated, monomorphous adenoma with morphological features of an acidophil stem cell adenoma such as the presence of misplaced exocytoses, fibrous bodies and mitochondrial gigantism. However, the tumor cells contained small secretory granules (up to 250 nm) accumulated along the cell membrane characteristic of thyrotrope cells. Furthermore, some adenoma cells were fusiform with long cytoplasmic processes resembling thyrotropes. Two months after the operation CT scan revealed a large residual tumor. Serum GH and TSH levels had increased again and the TSH level was even higher than before the treatment. The patient died suddenly, most probably of lethal arrhythmia. Specimens of the adenoma tissue obtained at autopsy confirmed the previous findings with the exception of positive immunostaining for TSH which was found in less than 1% of the adenoma cells. This undifferentiated, monomorphous GH/TSH-secreting pituitary adenoma represents an entity that is unusual both in its ultrastructural features and clinical manifestations suggesting a cytogenesis from an early, undifferentiated stem cell.

  20. Loss of monomorphic and polymorphic HLA antigens in metastatic breast and colon carcinoma.

    PubMed Central

    Goepel, J. R.; Rees, R. C.; Rogers, K.; Stoddard, C. J.; Thomas, W. E.; Shepherd, L.

    1991-01-01

    MHC class I antigens are intimately involved in intercellular communication, and recognition by cytotoxic T cells. Thus tumour cells that fail to express them may be at a growth or metastatic advantage. A series of ten colorectal and ten breast carcinomas, and their respective lymph node metastases, were examined immunohistologically using monoclonal antibodies (mAb) against both monomorphic and A2 polymorphic determinants, and beta-2-microglobulin (beta 2m). Four colon polypoid adenomas stained positively throughout, but 6/10 primary tumours had partial or complete loss of expression of monomorphic determinants using mAb W6/32: two node and the liver metastasis showed less, four more expression. Similar results were seen for beta 2m. HLA-A2 expression was absent or reduced in 4/4 colon tumours and all their metastases. Among the breast tumours, W6/32 staining was absent or reduced in 2/10, and node deposits showed two with less reactivity than their primary. Beta 2m staining was reduced or absent in 8/10 primaries and all the node metastases; in every case in which beta 2m was detected in the primary tumour their corresponding lymph node metastasis showed a decreased expression. HLA-A2 expression was absent or reduced in 3/4 primary breast carcinomas, and all their metastases. These results show that individual human colon and breast carcinomas often have a reduced HLA class I antigen expression, which apparently confers a metastatic advantage. Images Figure 1 Figure 2 Figure 3 PMID:1718386

  1. Clinical and morphological features of undifferentiated monomorphous GH/TSH-secreting pituitary adenoma.

    PubMed

    Skorić, T; Korsić, M; Zarković, K; Plavsić, V; Besenski, N; Breskovac, L; Giljević, Z; Paladino, J

    1999-06-01

    A 41-year-old male presented with progressive visual defects, acromegaly and hyperthyroidism. After clinical evaluation a giant GH/TSH-secreting pituitary adenoma was diagnosed. Administration of the somatostatin analog octreotide at doses of 150 microg s.c. per day inhibited the secretion of both GH and TSH. A three-week treatment with octreotide prior to surgery led to slight visual improvement and CT scan showed some new necrotic areas within the tumor mass. Transcranial surgery was performed. By immunohistochemical analyses of the adenoma tissue GH, prolactin and beta-chorionic gonadotropin were detected; TSH was negative. Electron microscopy revealed an undifferentiated, monomorphous adenoma with morphological features of an acidophil stem cell adenoma such as the presence of misplaced exocytoses, fibrous bodies and mitochondrial gigantism. However, the tumor cells contained small secretory granules (up to 250 nm) accumulated along the cell membrane characteristic of thyrotrope cells. Furthermore, some adenoma cells were fusiform with long cytoplasmic processes resembling thyrotropes. Two months after the operation CT scan revealed a large residual tumor. Serum GH and TSH levels had increased again and the TSH level was even higher than before the treatment. The patient died suddenly, most probably of lethal arrhythmia. Specimens of the adenoma tissue obtained at autopsy confirmed the previous findings with the exception of positive immunostaining for TSH which was found in less than 1% of the adenoma cells. This undifferentiated, monomorphous GH/TSH-secreting pituitary adenoma represents an entity that is unusual both in its ultrastructural features and clinical manifestations suggesting a cytogenesis from an early, undifferentiated stem cell. PMID:10366409

  2. DNA-binding proteins from marine bacteria expand the known sequence diversity of TALE-like repeats.

    PubMed

    de Lange, Orlando; Wolf, Christina; Thiel, Philipp; Krüger, Jens; Kleusch, Christian; Kohlbacher, Oliver; Lahaye, Thomas

    2015-11-16

    Transcription Activator-Like Effectors (TALEs) of Xanthomonas bacteria are programmable DNA binding proteins with unprecedented target specificity. Comparative studies into TALE repeat structure and function are hindered by the limited sequence variation among TALE repeats. More sequence-diverse TALE-like proteins are known from Ralstonia solanacearum (RipTALs) and Burkholderia rhizoxinica (Bats), but RipTAL and Bat repeats are conserved with those of TALEs around the DNA-binding residue. We study two novel marine-organism TALE-like proteins (MOrTL1 and MOrTL2), the first to date of non-terrestrial origin. We have assessed their DNA-binding properties and modelled repeat structures. We found that repeats from these proteins mediate sequence specific DNA binding conforming to the TALE code, despite low sequence similarity to TALE repeats, and with novel residues around the BSR. However, MOrTL1 repeats show greater sequence discriminating power than MOrTL2 repeats. Sequence alignments show that there are only three residues conserved between repeats of all TALE-like proteins including the two new additions. This conserved motif could prove useful as an identifier for future TALE-likes. Additionally, comparing MOrTL repeats with those of other TALE-likes suggests a common evolutionary origin for the TALEs, RipTALs and Bats.

  3. DNA-binding proteins from marine bacteria expand the known sequence diversity of TALE-like repeats.

    PubMed

    de Lange, Orlando; Wolf, Christina; Thiel, Philipp; Krüger, Jens; Kleusch, Christian; Kohlbacher, Oliver; Lahaye, Thomas

    2015-11-16

    Transcription Activator-Like Effectors (TALEs) of Xanthomonas bacteria are programmable DNA binding proteins with unprecedented target specificity. Comparative studies into TALE repeat structure and function are hindered by the limited sequence variation among TALE repeats. More sequence-diverse TALE-like proteins are known from Ralstonia solanacearum (RipTALs) and Burkholderia rhizoxinica (Bats), but RipTAL and Bat repeats are conserved with those of TALEs around the DNA-binding residue. We study two novel marine-organism TALE-like proteins (MOrTL1 and MOrTL2), the first to date of non-terrestrial origin. We have assessed their DNA-binding properties and modelled repeat structures. We found that repeats from these proteins mediate sequence specific DNA binding conforming to the TALE code, despite low sequence similarity to TALE repeats, and with novel residues around the BSR. However, MOrTL1 repeats show greater sequence discriminating power than MOrTL2 repeats. Sequence alignments show that there are only three residues conserved between repeats of all TALE-like proteins including the two new additions. This conserved motif could prove useful as an identifier for future TALE-likes. Additionally, comparing MOrTL repeats with those of other TALE-likes suggests a common evolutionary origin for the TALEs, RipTALs and Bats. PMID:26481363

  4. Genetic encoding of DNA nanostructures and their self-assembly in living bacteria

    PubMed Central

    Elbaz, Johann; Yin, Peng; Voigt, Christopher A.

    2016-01-01

    The field of DNA nanotechnology has harnessed the programmability of DNA base pairing to direct single-stranded DNAs (ssDNAs) to assemble into desired 3D structures. Here, we show the ability to express ssDNAs in Escherichia coli (32–205 nt), which can form structures in vivo or be purified for in vitro assembly. Each ssDNA is encoded by a gene that is transcribed into non-coding RNA containing a 3′-hairpin (HTBS). HTBS recruits HIV reverse transcriptase, which nucleates DNA synthesis and is aided in elongation by murine leukemia reverse transcriptase. Purified ssDNA that is produced in vivo is used to assemble large 1D wires (300 nm) and 2D sheets (5.8 μm2) in vitro. Intracellular assembly is demonstrated using a four-ssDNA crossover nanostructure that recruits split YFP when properly assembled. Genetically encoding DNA nanostructures provides a route for their production as well as applications in living cells. PMID:27091073

  5. Assessment of methods to recover DNA from bacteria, fungi and archaea in complex environmental samples.

    PubMed

    Guillén-Navarro, Karina; Herrera-López, David; López-Chávez, Mariana Y; Cancino-Gómez, Máximo; Reyes-Reyes, Ana L

    2015-11-01

    DNA extraction from environmental samples is a critical step for metagenomic analysis to study microbial communities, including those considered uncultivable. Nevertheless, obtaining good quality DNA in sufficient quantities for downstream methodologies is not always possible, and it depends on the complexity and stability of each ecosystem, which could be more problematic for samples from tropical regions because those ecosystems are less stable and more complex. Three laboratory methods for the extraction of nucleic acids from samples representing unstable (decaying coffee pulp and mangrove sediments) and relatively stable (compost and soil) environments were tested. The results were compared with those obtained using two commercial DNA extraction kits. The quality of the extracted DNA was evaluated by PCR amplification to verify the recovery of bacterial, archaeal, and fungal genetic material. The laboratory method that gave the best results used a lysis procedure combining physical, chemical, and enzymatic steps.

  6. Assessment of methods to recover DNA from bacteria, fungi and archaea in complex environmental samples.

    PubMed

    Guillén-Navarro, Karina; Herrera-López, David; López-Chávez, Mariana Y; Cancino-Gómez, Máximo; Reyes-Reyes, Ana L

    2015-11-01

    DNA extraction from environmental samples is a critical step for metagenomic analysis to study microbial communities, including those considered uncultivable. Nevertheless, obtaining good quality DNA in sufficient quantities for downstream methodologies is not always possible, and it depends on the complexity and stability of each ecosystem, which could be more problematic for samples from tropical regions because those ecosystems are less stable and more complex. Three laboratory methods for the extraction of nucleic acids from samples representing unstable (decaying coffee pulp and mangrove sediments) and relatively stable (compost and soil) environments were tested. The results were compared with those obtained using two commercial DNA extraction kits. The quality of the extracted DNA was evaluated by PCR amplification to verify the recovery of bacterial, archaeal, and fungal genetic material. The laboratory method that gave the best results used a lysis procedure combining physical, chemical, and enzymatic steps. PMID:26014885

  7. A matter of life or death: modeling DNA damage and repair in bacteria.

    PubMed

    Karschau, Jens; de Almeida, Camila; Richard, Morgiane C; Miller, Samantha; Booth, Ian R; Grebogi, Celso; de Moura, Alessandro P S

    2011-02-16

    DNA damage is a hazard all cells must face, and evolution has created a number of mechanisms to repair damaged bases in the chromosome. Paradoxically, many of these repair mechanisms can create double-strand breaks in the DNA molecule which are fatal to the cell. This indicates that the connection between DNA repair and death is far from straightforward, and suggests that the repair mechanisms can be a double-edged sword. In this report, we formulate a mathematical model of the dynamics of DNA damage and repair, and we obtain analytical expressions for the death rate. We predict a counterintuitive relationship between survival and repair. We can discriminate between two phases: below a critical threshold in the number of repair enzymes, the half-life decreases with the number of repair enzymes, but becomes independent of the number of repair enzymes above the threshold. We are able to predict quantitatively the dependence of the death rate on the damage rate and other relevant parameters. We verify our analytical results by simulating the stochastic dynamics of DNA damage and repair. Finally, we also perform an experiment with Escherichia coli cells to test one of the predictions of our model. PMID:21320424

  8. High-Voltage Electroporation of Bacteria: Genetic Transformation of Campylobacter jejuni with Plasmid DNA

    NASA Astrophysics Data System (ADS)

    Miller, Jeff F.; Dower, William J.; Tompkins, Lucy S.

    1988-02-01

    Electroporation permits the uptake of DNA by mammalian cells and plant protoplasts because it induces transient permeability of the cell membrane. We investigated the utility of high-voltage electroporation as a method for genetic transformation of intact bacterial cells by using the enteric pathogen Campylobacter jejuni as a model system. This report demonstrates that the application of high-voltage discharges to bacterial cells permits genetic transformation. Our method involves exposure of a Campylobacter cell suspension to a high-voltage exponential decay discharge (5-13 kV/cm) for a brief period of time (resistance-capacitance time constant = 2.4-26 msec) in the presence of plasmid DNA. Electrical transformation of C. jejuni results in frequencies as high as 1.2 × 106 transformants per μ g of DNA. We have investigated the effects of pulse amplitude and duration, cell growth conditions, divalent cations, and DNA concentration on the efficiency of transformation. Transformants of C. jejuni obtained by electroporation contained structurally intact plasmid molecules. In addition, evidence is presented that indicates that C. jejuni possesses DNA restriction and modification systems. The use of electroporation as a method for transforming other bacterial species and guidelines for its implementation are also discussed.

  9. Genome-wide study predicts promoter-G4 DNA motifs regulate selective functions in bacteria: radioresistance of D. radiodurans involves G4 DNA-mediated regulation.

    PubMed

    Beaume, Nicolas; Pathak, Rajiv; Yadav, Vinod Kumar; Kota, Swathi; Misra, Hari S; Gautam, Hemant K; Chowdhury, Shantanu

    2013-01-01

    A remarkable number of guanine-rich sequences with potential to adopt non-canonical secondary structures called G-quadruplexes (or G4 DNA) are found within gene promoters. Despite growing interest, regulatory role of quadruplex DNA motifs in intrinsic cellular function remains poorly understood. Herein, we asked whether occurrence of potential G4 (PG4) DNA in promoters is associated with specific function(s) in bacteria. Using a normalized promoter-PG4-content (PG4(P)) index we analysed >60,000 promoters in 19 well-annotated species for (a) function class(es) and (b) gene(s) with enriched PG4(P). Unexpectedly, PG4-associated functional classes were organism specific, suggesting that PG4 motifs may impart specific function to organisms. As a case study, we analysed radioresistance. Interestingly, unsupervised clustering using PG4(P) of 21 genes, crucial for radioresistance, grouped three radioresistant microorganisms including Deinococcus radiodurans. Based on these predictions we tested and found that in presence of nanomolar amounts of the intracellular quadruplex-binding ligand N-methyl mesoporphyrin (NMM), radioresistance of D. radiodurans was attenuated by ~60%. In addition, important components of the RecF recombinational repair pathway recA, recF, recO, recR and recQ genes were found to harbour promoter-PG4 motifs and were also down-regulated in presence of NMM. Together these results provide first evidence that radioresistance may involve G4 DNA-mediated regulation and support the rationale that promoter-PG4s influence selective functions. PMID:23161683

  10. Analysis of several methods for the extraction of high quality DNA from acetic acid bacteria in wine and vinegar for characterization by PCR-based methods.

    PubMed

    Jara, C; Mateo, E; Guillamón, J M; Torija, M J; Mas, A

    2008-12-10

    Acetic acid bacteria (AAB) are fastidious microorganisms with poor recovery in culture. Culture-independent methods are currently under examination. Good DNA extraction is a strict requirement of these methods. We compared five methods for extracting the DNA of AAB directly from wine and vinegar samples. Four matrices (white wine, red wine, superficial vinegar and submerged vinegar) contaminated with two AAB strains belonging to Acetobacter pasteurianus and Gluconacetobacter hansenii were assayed. To improve the yield and quality of the extracted DNA, a sample treatment (washing with polyvinyl pyrrolidone or NaCl) was also tested. DNA quality was measured by amplification of the 16S rRNA gene with conventional PCR. DNA recovery rate was assessed by real-time PCR. DNA amplification was always successful with the Wizard method though DNA recovery was poor. A CTAB-based method and NucleoSpin protocol extracted the highest DNA recoveries from wine and vinegar samples. Both of these methods require treatment to recover suitable DNA for amplification with maximum recovery. Both may therefore be good solutions for DNA extraction in wine and vinegar samples. DNA extraction of Ga hansenii was more effective than that of A. pasteurianus. The fastest and cheapest method we evaluated (the Thermal shock protocol) produced the worst results both for DNA amplification and DNA recovery.

  11. Use of DNA Markers for Investigating Sources of Bacteria in Contaminated Ground Water: Wooster Township, Wayne County, Ohio

    USGS Publications Warehouse

    Dumouchelle, Denise H.

    2006-01-01

    In 2004, a public-health nuisance was declared by the Wayne County Board of Health in the Scenic Heights Drive-Batdorf Road area of Wooster Township, Wayne County, Ohio, because of concerns about the safety of water from local wells. Repeated sampling had detected the presence of fecal-indicator bacteria and elevated nitrate concentrations. In June 2006, the U.S. Geological Survey (USGS), in cooperation with the Ohio Environmental Protection Agency (Ohio EPA), collected and analyzed samples from some of the affected wells to help investigate the possibility of human-origin bacterial contamination. Water samples from 12 wells and 5 home sewage-treatment systems (HSTS) were collected. Bromide concentrations were determined in samples from the 12 wells. Samples from 5 of the 12 wells were analyzed for wastewater compounds. Total coliform, enterococci and Escherichia coli (E. coli) bacteria concentrations were determined for samples from 8 of the 12 wells. In addition, two microbial source-tracking tools that employ DNA markers were used on samples from several wells and a composite sample of water from five septic tanks. The DNA markers from the Enterococcus faecium species and the order Bacteroidales are associated with specific sources, either human or ruminant sources. Bromide concentrations ranged from 0.04 to 0.18 milligrams per liter (mg/L). No wastewater compounds were detected at concentrations above the reporting limits. Samples from the 12 wells also were collected by Ohio EPA and analyzed for chloride and nitrate. Chloride concentrations ranged from 12.6 to 61.6 mg/L and nitrate concentrations ranged from 2.34 to 11.9 mg/L (as N). Total coliforms and enterococci were detected in samples from 8 wells, at concentrations from 2 to 200 colony-forming units per 100 milliliters (CFU/100 mL) and 0.5 to 17 CFU/100 mL, respectively. E. coli were detected in samples from three of the eight wells, at concentrations of 1 or 2 CFU/100 mL. Tests for the human

  12. Development of species-specific hybridization probes for marine luminous bacteria by using in vitro DNA amplification

    SciTech Connect

    Wimpee, C.F.; Nadeau, T.L.; Nealson, K.H. )

    1991-05-01

    By using two highly conserved regions of the luxA gene as primers, polymerase chain reaction amplification methods were used to prepare species-specific probes against the luciferase gene from four major groups of marine luminous bacteria. Laboratory studies with test strains indicated that three of the four probes cross-reacted with themselves and with one or more of the other species at low stringencies but were specific for members of their own species at high stringencies. The fourth probe, generated from Vibrio harveyi DNA, a cross-reacted with DNAs from two closely related species, V. orientalis and V. vulnificus. When nonluminous cultures were tested with the species-specific probes, no false-positive results were observed, even at low stringencies. Two field isolates were correctly identified as Photobacterium phosphoreum by using the species-specific hybridization probes at high stringency. A mixed probe (four different hybridization probes) used at low stringency gave positive results with all of the luminous bacteria tested, including the terrestrial species Xenorhabdus luminescens, and the taxonomically distinct marine bacterial species Shewanella hanedai; minimal cross-hybridization with these species was seen at higher stringencies.

  13. Novel Phenanthrene-Degrading Bacteria Identified by DNA-Stable Isotope Probing

    PubMed Central

    Luo, Chunling; Zhang, Dayi; Zhang, Gan

    2015-01-01

    Microorganisms responsible for the degradation of phenanthrene in a clean forest soil sample were identified by DNA-based stable isotope probing (SIP). The soil was artificially amended with either 12C- or 13C-labeled phenanthrene, and soil DNA was extracted on days 3, 6 and 9. Terminal restriction fragment length polymorphism (TRFLP) results revealed that the fragments of 219- and 241-bp in HaeIII digests were distributed throughout the gradient profile at three different sampling time points, and both fragments were more dominant in the heavy fractions of the samples exposed to the 13C-labeled contaminant. 16S rRNA sequencing of the 13C-enriched fraction suggested that Acidobacterium spp. within the class Acidobacteria, and Collimonas spp. within the class Betaproteobacteria, were directly involved in the uptake and degradation of phenanthrene at different times. To our knowledge, this is the first report that the genus Collimonas has the ability to degrade PAHs. Two PAH-RHDα genes were identified in 13C-labeled DNA. However, isolation of pure cultures indicated that strains of Staphylococcus sp. PHE-3, Pseudomonas sp. PHE-1, and Pseudomonas sp. PHE-2 in the soil had high phenanthrene-degrading ability. This emphasizes the role of a culture-independent method in the functional understanding of microbial communities in situ. PMID:26098417

  14. Bridgehead invasion of a monomorphic plant pathogenic bacterium: Xanthomonas citri pv. citri, an emerging citrus pathogen in Mali and Burkina Faso.

    PubMed

    Leduc, A; Traoré, Y N; Boyer, K; Magne, M; Grygiel, P; Juhasz, C C; Boyer, C; Guerin, F; Wonni, I; Ouedraogo, L; Vernière, C; Ravigné, V; Pruvost, O

    2015-11-01

    Molecular epidemiology studies further our understanding of migrations of phytopathogenic bacteria, the major determining factor in their emergence. Asiatic citrus canker, caused by Xanthomonas citri pv. citri, was recently reported in Mali and Burkina Faso, a region remote from other contaminated areas. To identify the origin and pathways of these emergences, we used two sets of markers, minisatellites and microsatellites, for investigating different evolutionary scales. Minisatellite typing suggested the introduction of two groups of strains in Mali (DAPC 1 and DAPC 2), consistent with microsatellite typing. DAPC 2 was restricted to Bamako district, whereas DAPC 1 strains were found much more invasive. The latter strains formed a major clonal complex based on microsatellite data with the primary and secondary founders detected in commercial citrus nurseries and orchards. This suggests that human activities played a major role in the spread of DAPC 1 strains via the movement of contaminated propagative material, further supported by the frequent lack of differentiation between populations from geographically distant nurseries and orchards. Approximate Bayesian Computation analyses supported the hypothesis that strains from Burkina Faso resulted from a bridgehead invasion from Mali. Multi-locus variable number of tandem repeat analysis and Approximate Bayesian Computation are useful for understanding invasion routes and pathways of monomorphic bacterial pathogens.

  15. Mutual mate choice in a female-dominant and sexually monomorphic primate.

    PubMed

    Gomez, Doris; Huchard, Elise; Henry, Pierre-Yves; Perret, Martine

    2012-03-01

    Sexual dimorphism is common in polygynous species, where intrasexual competition is often thought to drive the evolution of large male body size, and in turn, male behavioral dominance over females. In Madagascar, the entire lemur radiation, which embraces diverse mating systems, lacks sexual dimorphism and exhibits frequent female dominance over males. The evolution of such morphological and behavioral peculiarities, often referred to as "the lemur syndrome," has proven difficult to understand. Among other hypotheses, a potential role of intersexual selection has been repeatedly proposed but hardly ever tested. Here, we investigate whether female choice favors small and compliant males, and whether male choice favors large females in captive gray mouse lemurs (Microcebus murinus). Detailed analysis of a combination of behavioral observations and hormonal data available for both sexes shows that (1) females accept more matings from males with higher fighting abilities, (2) males adjust their investment in intrasexual competition to female fertility, and (3) both male and female strategies are weakly influenced by the body mass of potential partners, in directions contradicting our predictions. These results do not suggest a prominent role of intersexual selection in the evolution and maintenance of the lemur syndrome but rather point to alternative mechanisms relating to male-male competition, specifically highlighting an absence of relationship between male body mass and fighting ability. Finally, our findings add to the growing body of evidence suggesting flexible sex roles, by showing the expression of mutual mate choice in a female-dominant, sexually monomorphic and promiscuous primate.

  16. Monomorphic ants undergo within-colony morphological changes along the metal-pollution gradient.

    PubMed

    Grześ, Irena M; Okrutniak, Mateusz; Woch, Marcin W

    2015-04-01

    In ants, intra and inter-colony variation in body size can be considerable, even in monomorphic species. It has been previously shown that size-related parameters can be environmentally sensitive. The shape of the body size distribution curve is, however, rarely investigated. In this study, we measured head widthes of the black garden ant Lasius niger workers using digital methods. The ants were sampled from 51 colonies originating from 19 sites located along a metal pollution gradient, established in a former mining area in Poland. Total zinc concentrations in random samples of small invertebrates were used as a measure of site pollution levels. We found that the skewness of head size distribution grows significantly in line with the pollution level of the site, ranging from values slightly below zero (about -0.5) in the least polluted site up to a positive value (about 1.5) in the most polluted site. This result indicates that the frequency of small ants grows as pollution levels increase. The coefficient of variation, as well as the measures of central tendency, was not related to the pollution level. Four hypotheses explaining the obtained results are proposed. The bias towards the higher frequency of small workers may result from energy limitation and/or metal toxicity, but may also have an adaptive function.

  17. Presence of species-specific, monomorphic antigens on sheep and goat binucleate cells.

    PubMed

    Gall, J G; MacLaren, L A; Anderson, G B

    1995-07-15

    Immunocytochemical assays for sheep and goat species-specific, monomorphic antigens were developed utilizing polyclonal antisera from sheep and goats immunized by interspecific pregnancy. The assays were applied to cell isolates from sheep and goat fetal cotyledons collected from allogeneic pregnancies at Days 35, 40 and 120 of gestation. The isolates contained 7 to 48% binucleate cells (BNC). Using these assays, the sheep-specific antigen was detected on sheep cotyledonary cell isolates on all days of gestation tested (P < 0.001); the assay also detected the antigen on the BNC subset of the cotyledonary cell isolate population (P < 0.001). The caprine-specific antigen was shown to be present on cotyledonary cell isolates (P < 0.05), although the presence of the antigen could not be demonstrated with statistical confidence on goat BNC due to insufficient numbers of discernible cells. Binucleate cells contribute to the formation of the syncytial layer of the placenta by fusing with maternal epithelial cells and with the syncytium. The species-specific antigen (or antigens) is present on BNC at the appropriate time of gestation at which it (they) could play a role in the humoral immune response to interspecific and hybrid pregnancies observed in ewes and does. PMID:16727726

  18. Monomorphic lymphomas arising in patients with Hodgkin's disease. Correlation of morphologic, immunophenotypic, and molecular genetic findings in 12 cases.

    PubMed Central

    Casey, T. T.; Cousar, J. B.; Mangum, M.; Williams, M. E.; Lee, J. T.; Greer, J. P.; Collins, R. D.

    1990-01-01

    Patients with Hodgkin's Disease (HD) occasionally develop monomorphic lymphomas in which mononuclear cells, usually large in size, grow in sheets, and in which there are few reacting cells or classic Reed-Sternberg (RS) cells. Twelve patients of this type were reviewed to determine the nature of the monomorphic growth. Paraffin-embedded tissue sections from the original diagnostic HD and the monomorphic growths were stained for Leu-M1 (CD15), leukocyte common antigen (LCA, CD45), pan B-cell markers LN1, LN2, and L26, and pan T-cell marker UCHL1 (CD45R) reactive in paraffin-embedded tissues. Cases were included only if the original diagnostic material had the classic histopathologic features of HD, if there was a separate monomorphic growth (in place or time), and if sufficient materials from both phases were available for study. Original diagnoses of HD included nodular sclerosing (NS; 8 cases); lymphocyte predominant (LP; 2 cases); mixed cellularity (MC; 1 case); and lymphocyte depleted (LD: 1 case) types. RS cells in the eight cases of NS HD and one case of MC HD were generally Leu-M1 and LN2 positive, and L26, LN1, UCHL1, and LCA negative. RS cells in one case of NS HD were LCA positive in addition to Leu-M1, LN1, and LN2. Two cases of NS HD showed L26 positive RS cells. Conversely, RS cells and lymphocytic-histiocytic (L and H) variants in the cases of LP HD were Leu-M1 and LN2 negative, and LCA and LN1 positive. The one case of LD HD possessed RS cells that were negative for Leu-M1, but positive for LCA, L26, LN1, and LN2. In seven cases (4 NS, 2 LP, 1 LD) the monomorphic growths possessed a B-cell phenotype (LCA, L26, and LN1 positive; Leu-M1 and UCHL1 negative). In the remaining cases (4 NS, 1 MC), the monomorphic growths were Leu-M1 positive, and displayed phenotypes similar to the RS cells of the original NS HD. Southern blot analysis was performed on the monomorphic components of five cases and showed some form of immunoglobulin gene rearrangement in each

  19. Monomorphic adenomas of the major and minor salivary glands. Report of twenty-one cases and review of the literature.

    PubMed

    Mintz, G A; Abrams, A M; Melrose, R J

    1982-04-01

    Monomorphic adenomas are benign salivary gland tumors that have a predilection for development in the upper lip and parotid gland. Typically, patients are older persons (mean age, 61 years), but a broad age range (32 to 87 years) has been reported in the literature. Adequate treatment consists of superficial or total parotidectomy (depending on extent and location of the tumor) for parotid lesions and excision with a limited border of normal tissue for minor gland tumors. Uniform cellularity, lack of myxoid or chondroid features, and a tendency for multicentric origin are features which separate these tumors from pleomorphic adenomas. Monomorphic adenomas have been mistakenly diagnosed and treated as adenoid cystic carcinomas. Close attention to cytologic detail, histomorphology, and growth pattern at the periphery are important in separating these tumors.

  20. Breakthrough of ultraviolet light from various brands of fluorescent lamps: lethal effects on DNA repair-defective bacteria.

    PubMed

    Hartman, P E; Biggley, W H

    1996-01-01

    In a comparative study of 17 pairs of 15 W fluorescent lamps intended for use in homes and purchased in local stores, we detect over 10-fold differences in UVB + UVC emissions between various lamps. This breakthrough of ultraviolet (UV) light is in part correlated with ability of lamps to kill DNA repair-defective recA-uvrB- Salmonella. Relative proficiency of lamps in eliciting photoreactivation of UV-induced DNA lesions also plays a prominent role in the relative rates of bacterial inactivation by emissions from different lamps. Lamps made in Chile, such as Philips brand lamps and one type of General Electric lamp, produce far less UVB + UVC and fail to kill recA-uvrB- bacteria. In contrast, all tested lamps manufactured in the USA, Hungary, and Japan exhibit readily observed deleterious biological effects. When an E. coli recA-uvrB-phr- (photolyase-negative) triple mutant is used for assay, lethal radiations are detected from all lamps, and single-hit exponential inactivation rates rather closely correlate to amount of directly measured UVB + UVC output of each pair of lamps. Although all lamps tested may meet international and United States standards for radiation safety, optimal practices in lamp manufacture are clearly capable of decreasing human exposure to indoor UV light.

  1. RRNA and dnaK relationships of Bradyrhizobium sp. nodule bacteria from four papilionoid legume trees in Costa Rica.

    PubMed

    Parker, Matthew A

    2004-05-01

    Enzyme electrophoresis and sequencing of rRNA and dnaK genes revealed high genetic diversity among root nodule bacteria from the Costa Rican trees Andira inermis, Dalbergia retusa, Platymiscium pinnatum (Papilionoideae tribe Dalbergieae) and Lonchocarpus atropurpureus (Papilionoideae tribe Millettieae). A total of 21 distinct multilocus genotypes [ETs (electrophoretic types)] was found among the 36 isolates analyzed, and no ETs were shared in common by isolates from different legume hosts. However, three of the ETs from D. retusa were identical to Bradyrhizobium sp. isolates detected in prior studies of several other legume genera in both Costa Rica and Panama. Nearly full-length 16S rRNA sequences and partial 23S rRNA sequences confirmed that two isolates from D. retusa were highly similar or identical to Bradyrhizobium strains isolated from the legumes Erythrina and Clitoria (Papilionoideae tribe Phaseoleae) in Panama. rRNA sequences for five isolates from L. atropurpureus, P. pinnatum and A. inermis were not closely related to any currently known strains from Central America or elsewhere, but had affinities to the reference strains Bradyrhizobium japonicum USDA 110 (three isolates) or to B. elkanii USDA 76 (two isolates). A phylogenetic tree for 21 Bradyrhizobium strains based on 603 bp of the dnaK gene showed several significant conflicts with the rRNA tree, suggesting that genealogical relationships may have been altered by lateral gene transfer events. PMID:15214639

  2. Horizontal DNA Transfer Mechanisms of Bacteria as Weapons of Intragenomic Conflict.

    PubMed

    Croucher, Nicholas J; Mostowy, Rafal; Wymant, Christopher; Turner, Paul; Bentley, Stephen D; Fraser, Christophe

    2016-03-01

    Horizontal DNA transfer (HDT) is a pervasive mechanism of diversification in many microbial species, but its primary evolutionary role remains controversial. Much recent research has emphasised the adaptive benefit of acquiring novel DNA, but here we argue instead that intragenomic conflict provides a coherent framework for understanding the evolutionary origins of HDT. To test this hypothesis, we developed a mathematical model of a clonally descended bacterial population undergoing HDT through transmission of mobile genetic elements (MGEs) and genetic transformation. Including the known bias of transformation toward the acquisition of shorter alleles into the model suggested it could be an effective means of counteracting the spread of MGEs. Both constitutive and transient competence for transformation were found to provide an effective defence against parasitic MGEs; transient competence could also be effective at permitting the selective spread of MGEs conferring a benefit on their host bacterium. The coordination of transient competence with cell-cell killing, observed in multiple species, was found to result in synergistic blocking of MGE transmission through releasing genomic DNA for homologous recombination while simultaneously reducing horizontal MGE spread by lowering the local cell density. To evaluate the feasibility of the functions suggested by the modelling analysis, we analysed genomic data from longitudinal sampling of individuals carrying Streptococcus pneumoniae. This revealed the frequent within-host coexistence of clonally descended cells that differed in their MGE infection status, a necessary condition for the proposed mechanism to operate. Additionally, we found multiple examples of MGEs inhibiting transformation through integrative disruption of genes encoding the competence machinery across many species, providing evidence of an ongoing "arms race." Reduced rates of transformation have also been observed in cells infected by MGEs that

  3. Horizontal DNA Transfer Mechanisms of Bacteria as Weapons of Intragenomic Conflict.

    PubMed

    Croucher, Nicholas J; Mostowy, Rafal; Wymant, Christopher; Turner, Paul; Bentley, Stephen D; Fraser, Christophe

    2016-03-01

    Horizontal DNA transfer (HDT) is a pervasive mechanism of diversification in many microbial species, but its primary evolutionary role remains controversial. Much recent research has emphasised the adaptive benefit of acquiring novel DNA, but here we argue instead that intragenomic conflict provides a coherent framework for understanding the evolutionary origins of HDT. To test this hypothesis, we developed a mathematical model of a clonally descended bacterial population undergoing HDT through transmission of mobile genetic elements (MGEs) and genetic transformation. Including the known bias of transformation toward the acquisition of shorter alleles into the model suggested it could be an effective means of counteracting the spread of MGEs. Both constitutive and transient competence for transformation were found to provide an effective defence against parasitic MGEs; transient competence could also be effective at permitting the selective spread of MGEs conferring a benefit on their host bacterium. The coordination of transient competence with cell-cell killing, observed in multiple species, was found to result in synergistic blocking of MGE transmission through releasing genomic DNA for homologous recombination while simultaneously reducing horizontal MGE spread by lowering the local cell density. To evaluate the feasibility of the functions suggested by the modelling analysis, we analysed genomic data from longitudinal sampling of individuals carrying Streptococcus pneumoniae. This revealed the frequent within-host coexistence of clonally descended cells that differed in their MGE infection status, a necessary condition for the proposed mechanism to operate. Additionally, we found multiple examples of MGEs inhibiting transformation through integrative disruption of genes encoding the competence machinery across many species, providing evidence of an ongoing "arms race." Reduced rates of transformation have also been observed in cells infected by MGEs that

  4. Horizontal DNA Transfer Mechanisms of Bacteria as Weapons of Intragenomic Conflict

    PubMed Central

    Croucher, Nicholas J.; Mostowy, Rafal; Wymant, Christopher; Turner, Paul; Bentley, Stephen D.; Fraser, Christophe

    2016-01-01

    Horizontal DNA transfer (HDT) is a pervasive mechanism of diversification in many microbial species, but its primary evolutionary role remains controversial. Much recent research has emphasised the adaptive benefit of acquiring novel DNA, but here we argue instead that intragenomic conflict provides a coherent framework for understanding the evolutionary origins of HDT. To test this hypothesis, we developed a mathematical model of a clonally descended bacterial population undergoing HDT through transmission of mobile genetic elements (MGEs) and genetic transformation. Including the known bias of transformation toward the acquisition of shorter alleles into the model suggested it could be an effective means of counteracting the spread of MGEs. Both constitutive and transient competence for transformation were found to provide an effective defence against parasitic MGEs; transient competence could also be effective at permitting the selective spread of MGEs conferring a benefit on their host bacterium. The coordination of transient competence with cell–cell killing, observed in multiple species, was found to result in synergistic blocking of MGE transmission through releasing genomic DNA for homologous recombination while simultaneously reducing horizontal MGE spread by lowering the local cell density. To evaluate the feasibility of the functions suggested by the modelling analysis, we analysed genomic data from longitudinal sampling of individuals carrying Streptococcus pneumoniae. This revealed the frequent within-host coexistence of clonally descended cells that differed in their MGE infection status, a necessary condition for the proposed mechanism to operate. Additionally, we found multiple examples of MGEs inhibiting transformation through integrative disruption of genes encoding the competence machinery across many species, providing evidence of an ongoing “arms race.” Reduced rates of transformation have also been observed in cells infected by MGEs that

  5. On-chip concentration of bacteria using a 3D dielectrophoretic chip and subsequent laser-based DNA extraction in the same chip

    NASA Astrophysics Data System (ADS)

    Cho, Yoon-Kyoung; Kim, Tae-hyeong; Lee, Jeong-Gun

    2010-06-01

    We report the on-chip concentration of bacteria using a dielectrophoretic (DEP) chip with 3D electrodes and subsequent laser-based DNA extraction in the same chip. The DEP chip has a set of interdigitated Au post electrodes with 50 µm height to generate a network of non-uniform electric fields for the efficient trapping by DEP. The metal post array was fabricated by photolithography and subsequent Ni and Au electroplating. Three model bacteria samples (Escherichia coli, Staphylococcus epidermidis, Streptococcus mutans) were tested and over 80-fold concentrations were achieved within 2 min. Subsequently, on-chip DNA extraction from the concentrated bacteria in the 3D DEP chip was performed by laser irradiation using the laser-irradiated magnetic bead system (LIMBS) in the same chip. The extracted DNA was analyzed with silicon chip-based real-time polymerase chain reaction (PCR). The total process of on-chip bacteria concentration and the subsequent DNA extraction can be completed within 10 min including the manual operation time.

  6. Gram-positive bacteria with a high DNA G+C content are characterized by a common insertion within their 23S rRNA genes.

    PubMed

    Roller, C; Ludwig, W; Schleifer, K H

    1992-06-01

    An insertion of about 100 bases within the central part of the 23S rRNA genes was found to be a phylogenetic marker for the bacterial line of descent of Gram-positive bacteria with a high DNA G + C content. The insertion was present in 23S rRNA genes of 64 strains representing the major phylogenetic groups of Gram-positive bacteria with a high DNA G+C content, whereas it was not found in 23S rRNA genes of 55 (eu)bacteria representing Gram-positive bacteria with a low DNA G + C content and all other known (eu)bacterial phyla. The presence of the insertion could be easily demonstrated by comparative gel electrophoretic analysis of in vitro-amplified 23S rDNA fragments, which contained the insertion. The nucleotide sequences of the amplified fragments were determined and sequence similarities of at least 44% were found. The overall similarity values are lower than those of 16S and 23S rRNA sequences of the particular organism. Northern hybridization experiments indicated the presence of the insertion within the mature 23S rRNA of Corynebacterium glutamicum.

  7. Coxiella burnetii DNA, but not viable bacteria, in dairy products in France.

    PubMed

    Eldin, Carole; Angelakis, Emmanouil; Renvoisé, Aurélie; Raoult, Didier

    2013-04-01

    Transmission by the oral route of Coxiella burnetii is controversial. Our objective was to evaluate dairy products in the transmission of Q fever. Pasteurized, unpasteurized, and thermized dairy products were tested for C. burnetii by using a quantitative polymerase chain reaction specific for IS1111 and IS30A spacers, culturing in human embryonic lung fibroblasts cells, and inoculation into BALB/c mice. We tested 201 products and C. burnetii was identified in 64%. Cow milk origin products were more frequently positive than goat or ewe products (P = 0.006 and P = 0.0001, respectively), and industrial food was more frequently positive than artisanal food (P < 0.0001). Food made from unpasteurized milk contained higher bacteria concentrations than food made from pasteurized milk (P = 0.02). All cultures were negative and mice did not show signs of illness. Farm animals are highly infected in France but consumption of cheese and yogurt does not seem to pose a public health risk for transmission of Q fever.

  8. Comprehensive Census of Bacteria in Clean Rooms by Using DNA Microarray and Cloning Methods▿ †

    PubMed Central

    La Duc, Myron T.; Osman, Shariff; Vaishampayan, Parag; Piceno, Yvette; Andersen, Gary; Spry, J. A.; Venkateswaran, Kasthuri

    2009-01-01

    A census of clean room surface-associated bacterial populations was derived from the results of both the cloning and sequencing of 16S rRNA genes and DNA microarray (PhyloChip) analyses. Samples from the Lockheed Martin Aeronautics Multiple Testing Facility (LMA-MTF), the Kennedy Space Center Payload Hazard and Servicing Facility (KSC-PHSF), and the Jet Propulsion Laboratory Spacecraft Assembly Facility (JPL-SAF) clean rooms were collected during the various assembly phases of the Phoenix and Mars Science Laboratory (MSL) spacecraft. Clone library-derived analyses detected a larger bacterial diversity prior to the arrival of spacecraft hardware in these clean room facilities. PhyloChip results were in agreement with this trend but also unveiled the presence of anywhere from 9- to 70-fold more bacterial taxa than cloning approaches. Among the facilities sampled, the JPL-SAF (MSL mission) housed a significantly less diverse bacterial population than either the LMA-MTF or KSC-PHSF (Phoenix mission). Bacterial taxa known to thrive in arid conditions were frequently detected in MSL-associated JPL-SAF samples, whereas proteobacterial lineages dominated Phoenix-associated KSC-PHSF samples. Comprehensive bacterial censuses, such as that reported here, will help space-faring nations preemptively identify contaminant biomatter that may compromise extraterrestrial life detection experiments. The robust nature and high sensitivity of DNA microarray technologies should prove beneficial to a wide range of scientific, electronic, homeland security, medical, and pharmaceutical applications and to any other ventures with a vested interest in monitoring and controlling contamination in exceptionally clean environments. PMID:19700540

  9. Comprehensive census of bacteria in clean rooms by using DNA microarray and cloning methods.

    PubMed

    La Duc, Myron T; Osman, Shariff; Vaishampayan, Parag; Piceno, Yvette; Andersen, Gary; Spry, J A; Venkateswaran, Kasthuri

    2009-10-01

    A census of clean room surface-associated bacterial populations was derived from the results of both the cloning and sequencing of 16S rRNA genes and DNA microarray (PhyloChip) analyses. Samples from the Lockheed Martin Aeronautics Multiple Testing Facility (LMA-MTF), the Kennedy Space Center Payload Hazard and Servicing Facility (KSC-PHSF), and the Jet Propulsion Laboratory Spacecraft Assembly Facility (JPL-SAF) clean rooms were collected during the various assembly phases of the Phoenix and Mars Science Laboratory (MSL) spacecraft. Clone library-derived analyses detected a larger bacterial diversity prior to the arrival of spacecraft hardware in these clean room facilities. PhyloChip results were in agreement with this trend but also unveiled the presence of anywhere from 9- to 70-fold more bacterial taxa than cloning approaches. Among the facilities sampled, the JPL-SAF (MSL mission) housed a significantly less diverse bacterial population than either the LMA-MTF or KSC-PHSF (Phoenix mission). Bacterial taxa known to thrive in arid conditions were frequently detected in MSL-associated JPL-SAF samples, whereas proteobacterial lineages dominated Phoenix-associated KSC-PHSF samples. Comprehensive bacterial censuses, such as that reported here, will help space-faring nations preemptively identify contaminant biomatter that may compromise extraterrestrial life detection experiments. The robust nature and high sensitivity of DNA microarray technologies should prove beneficial to a wide range of scientific, electronic, homeland security, medical, and pharmaceutical applications and to any other ventures with a vested interest in monitoring and controlling contamination in exceptionally clean environments. PMID:19700540

  10. Theoretical models for the regulation of DNA replication in fast-growing bacteria

    NASA Astrophysics Data System (ADS)

    Creutziger, Martin; Schmidt, Mischa; Lenz, Peter

    2012-09-01

    Growing in always changing environments, Escherichia coli cells are challenged by the task to coordinate growth and division. In particular, adaption of their growth program to the surrounding medium has to guarantee that the daughter cells obtain fully replicated chromosomes. Replication is therefore to be initiated at the right time, which is particularly challenging in media that support fast growth. Here, the mother cell initiates replication not only for the daughter but also for the granddaughter cells. This is possible only if replication occurs from several replication forks that all need to be correctly initiated. Despite considerable efforts during the last 40 years, regulation of this process is still unknown. Part of the difficulty arises from the fact that many details of the relevant molecular processes are not known. Here, we develop a novel theoretical strategy for dealing with this general problem: instead of analyzing a single model, we introduce a wide variety of 128 different models that make different assumptions about the unknown processes. By comparing the predictions of these models we are able to identify the key quantities that allow the experimental discrimination of the different models. Analysis of these quantities yields that out of the 128 models 94 are not consistent with available experimental data. From the remaining 34 models we are able to conclude that mass growth and DNA replication need either to be truly coupled, by coupling DNA replication initiation to the event of cell division, or to the amount of accumulated mass. Finally, we make suggestions for experiments to further reduce the number of possible regulation scenarios.

  11. Purification of bacterial genomic DNA in less than 20 min using chelex-100 microwave: examples from strains of lactic acid bacteria isolated from soil samples.

    PubMed

    Reyes-Escogido, Lourdes; Balam-Chi, Mario; Rodríguez-Buenfil, Ingrid; Valdés, Jesús; Kameyama, Luis; Martínez-Pérez, Francisco

    2010-11-01

    We established a Chelex 100-Microwave method for the purification of bacterial genomic DNA (gDNA) in less than 20 min with high yield and good quality, useful for multiple purposes. It combines Chelex 100, proteinase K, RNase A and heating in a microwave oven. The resulting gDNA was used directly to identify bacterial species of the Order Lactobacillales by means of PCR amplification of their 16S rDNA gene, isolated from sediments on the Yucatan Peninsula, Mexico. This method produced gDNA free of phenolic and protein residual contaminants from 100 of these isolated bacteria. 16S rDNA amplification and sequencing showed Pediococcus acidilactici to prevail in inland lagoons, and Pediococcus pentosaceus, Lactobacillus plantarum, Lactobacillus sp., and Lactobacillus fermentum to be most abundant in the soils of livestock farms. The combination of Chelex 100, enzymes and microwave heating used in the Chelex 100-Microwave method produced large amounts of highly pure gDNA from Gram-positive and Gram-negative bacteria, in less than 20 min.

  12. Local adaptation and divergence in colour signal conspicuousness between monomorphic and polymorphic lineages in a lizard.

    PubMed

    McLean, C A; Moussalli, A; Stuart-Fox, D

    2014-12-01

    Population differences in visual environment can lead to divergence in multiple components of animal coloration including signalling traits and colour patterns important for camouflage. Divergence may reflect selection imposed by different receivers (conspecifics, predators), which depends in turn on the location of the colour patch. We tested for local adaptation of two genetically and phenotypically divergent lineages of a rock-inhabiting lizard, Ctenophorus decresii, by comparing the visual contrast of colour patches to different receivers in native and non-native environments. The lineages differ most notably in male throat coloration, which is polymorphic in the northern lineage and monomorphic in the southern lineage, but also differ in dorsal and lateral coloration, which is visible to both conspecifics and potential predators. Using models of animal colour vision, we assessed whether lineage-specific throat, dorsal and lateral coloration enhanced conspicuousness to conspecifics, increased crypsis to birds or both, respectively, when viewed against the predominant backgrounds from each lineage. Throat colours were no more conspicuous against native than non-native rock but contrasted more strongly with native lichen, which occurs patchily on rocks inhabited by C. decresii. Conversely, neck coloration (lateral) more closely matched native lichen. Furthermore, although dorsal coloration of southern males was consistently more conspicuous to birds than that of northern males, both lineages had similar absolute conspicuousness against their native backgrounds. Combined, our results are consistent with local adaptation of multiple colour traits in relation to multiple receivers, suggesting that geographic variation in background colour has influenced the evolution of lineage-specific coloration in C. decresii.

  13. Development and application of a new method to extract bacterial DNA from soil based on separation of bacteria from soil with cation-exchange resin.

    PubMed

    Jacobsen, C S; Rasmussen, O F

    1992-08-01

    A new method for the extraction of bacterial DNA from soil has been developed. Soil samples of 50 g were dispersed, and bacteria were released by use of a cation-exchange resin; subsequently, bacteria were separated from soil particles by low-speed centrifugation and lysed with lysozyme and ionic detergent, and the DNA was then purified by CsCl-ethidium bromide equilibrium density centrifugation. The extracted DNA was of high molecular weight and sufficiently pure for restriction enzyme digestion, DNA-DNA hybridization, and amplification by the polymerase chain reaction. The advantages of the new method are that the separation of bacteria from soil is considerably faster than by repeated blending, more samples can be handled, and furthermore no aerosols are formed during separation. Also, we investigated whether the CsCl-ethidium bromide equilibrium density centrifugation could be replaced by purification using Gene-Clean. However, this method produced DNAs which were insufficiently pure for several types of analysis. The new method was used to study survival of a 2,4-dichlorophenoxyacetic acid (2,4-D)-degrading Pseudomonas cepacia DBO1 (pRO101) in unamended soil and in soil amended with 2,4-D. We found that the degrading strain, irrespective of inoculation level, was able to grow to the same high numbers in soil amended with 2,4-D, while the strain in nonamended soil were maintained at the inoculation level. Detection based on DNA extraction and subsequent dot blot DNA-DNA hybridization was in accordance with detection by plating on selective medium. PMID:16348750

  14. Effect of DNA extraction procedure, repeated extraction and ethidium monoazide (EMA)/propidium monoazide (PMA) treatment on overall DNA yield and impact on microbial fingerprints for bacteria, fungi and archaea in a reference soil

    PubMed Central

    Wagner, Andreas O.; Praeg, Nadine; Reitschuler, Christoph; Illmer, Paul

    2015-01-01

    Different DNA extraction protocols were evaluated on a reference soil. A wide difference was found in the total extractable DNA as derived from different extraction protocols. Concerning the DNA yield phenol–chloroform–isomyl alcohol extraction resulted in high DNA yield but also in a remarkable co-extraction of contaminants making PCR from undiluted DNA extracts impossible. By comparison of two different extraction kits, the Macherey&Nagel SoilExtract II kit resulted in the highest DNA yields when buffer SL1 and the enhancer solution were applied. The enhancer solution not only significantly increased the DNA yield but also the amount of co-extracted contaminates, whereas additional disintegration strategies did not. Although a three times repeated DNA extraction increased the total amount of extracted DNA, microbial fingerprints were merely affected. However, with the 5th extraction this changed. A reduction of total DGGE band numbers was observed for archaea and fungi, whereas for bacteria the diversity increased. The application of ethidium monoazide (EMA) or propidium monoazide (PMA) treatment aiming on the selective removal of soil DNA derived from cells lacking cell wall integrity resulted in a significant reduction of total extracted DNA, however, the hypothesized effect on microbial fingerprints failed to appear indicating the need for further investigations. PMID:26339125

  15. A prophage-encoded actin-like protein required for efficient viral DNA replication in bacteria.

    PubMed

    Donovan, Catriona; Heyer, Antonia; Pfeifer, Eugen; Polen, Tino; Wittmann, Anja; Krämer, Reinhard; Frunzke, Julia; Bramkamp, Marc

    2015-05-26

    In host cells, viral replication is localized at specific subcellular sites. Viruses that infect eukaryotic and prokaryotic cells often use host-derived cytoskeletal structures, such as the actin skeleton, for intracellular positioning. Here, we describe that a prophage, CGP3, integrated into the genome of Corynebacterium glutamicum encodes an actin-like protein, AlpC. Biochemical characterization confirms that AlpC is a bona fide actin-like protein and cell biological analysis shows that AlpC forms filamentous structures upon prophage induction. The co-transcribed adaptor protein, AlpA, binds to a consensus sequence in the upstream promoter region of the alpAC operon and also interacts with AlpC, thus connecting circular phage DNA to the actin-like filaments. Transcriptome analysis revealed that alpA and alpC are among the early induced genes upon excision of the CGP3 prophage. Furthermore, qPCR analysis of mutant strains revealed that both AlpA and AlpC are required for efficient phage replication. Altogether, these data emphasize that AlpAC are crucial for the spatio-temporal organization of efficient viral replication. This is remarkably similar to actin-assisted membrane localization of eukaryotic viruses that use the actin cytoskeleton to concentrate virus particles at the egress sites and provides a link of evolutionary conserved interactions between intracellular virus transport and actin.

  16. A prophage-encoded actin-like protein required for efficient viral DNA replication in bacteria.

    PubMed

    Donovan, Catriona; Heyer, Antonia; Pfeifer, Eugen; Polen, Tino; Wittmann, Anja; Krämer, Reinhard; Frunzke, Julia; Bramkamp, Marc

    2015-05-26

    In host cells, viral replication is localized at specific subcellular sites. Viruses that infect eukaryotic and prokaryotic cells often use host-derived cytoskeletal structures, such as the actin skeleton, for intracellular positioning. Here, we describe that a prophage, CGP3, integrated into the genome of Corynebacterium glutamicum encodes an actin-like protein, AlpC. Biochemical characterization confirms that AlpC is a bona fide actin-like protein and cell biological analysis shows that AlpC forms filamentous structures upon prophage induction. The co-transcribed adaptor protein, AlpA, binds to a consensus sequence in the upstream promoter region of the alpAC operon and also interacts with AlpC, thus connecting circular phage DNA to the actin-like filaments. Transcriptome analysis revealed that alpA and alpC are among the early induced genes upon excision of the CGP3 prophage. Furthermore, qPCR analysis of mutant strains revealed that both AlpA and AlpC are required for efficient phage replication. Altogether, these data emphasize that AlpAC are crucial for the spatio-temporal organization of efficient viral replication. This is remarkably similar to actin-assisted membrane localization of eukaryotic viruses that use the actin cytoskeleton to concentrate virus particles at the egress sites and provides a link of evolutionary conserved interactions between intracellular virus transport and actin. PMID:25916847

  17. The 16S rDNA Phylogenetic Composition of Bacteria Implicated in Sulfur Redox Cycles and Associated Sulfur Isotope Fractionation

    NASA Astrophysics Data System (ADS)

    Bicknell, B. T.; Batts, J. E.; Krouse, H. R.

    2006-12-01

    The reduction of sulfate ion to sulfide species by sulfate reducing bacteria (SRB) is accompanied by sulfur isotope fractionation, measured in terms of changes in the δ^{34}S values for sulfate and sulfide ions relative to a defined standard. In open environments, the S-isotope compositions of sulfate and sulfide can be affected by loss from the system of sulfide species as gaseous H2S, insoluble metal sulfides such as FeS2, organic complexes or by re-oxidation. The S-isotope fractionation accompanying bacterial sulfate reduction in nature is often much larger than the maxima obtained in chemical and bacterial sulfate reduction experiments in the laboratory. One mechanism postulated for the large natural S-isotope selectivity depends on repetitive reduction-oxidation cycles. In turn, this would require a level of tolerance to oxygen by SRB in the sedimentary environment, contrary to laboratory experience with SRB strains. Bird Lake (The Coorong, South Australia) is a small calcareous, evaporative lake, where average Δ^{34}S (δ^{34}Ssulfate - δ^{34}Ssulfide) values for groundwater at 16 of the 27 sites sampled periodically since 1974, vary from 15.0 ‰ to 62.3 ‰ within the range -1.8 ‰ to 70.6 ‰. Wide fluctuations in δ34Ssulfide values at individual sites are the significant factor affecting the variability of Δ^{34}S values. Values for δ18Osulfate are elevated over that of the sulfate source to an unusual extent, reflecting re-oxidation of sulfur species and O- isotope exchange between some of these species and water. One aspect of investigations at Bird Lake was the evaluation of bacterial populations in subsurface sediments and their role in sulfur cycling. To achieve this, microcosms were established with subsurface sediment and incubated under a nitrogen atmosphere, for up to 119 days. These were sampled at various times to determine sulfur species concentrations and sulfur isotope fractionation and to generate 16S rDNA clone libraries. Results

  18. Then and now: use of 16S rDNA gene sequencing for bacterial identification and discovery of novel bacteria in clinical microbiology laboratories.

    PubMed

    Woo, P C Y; Lau, S K P; Teng, J L L; Tse, H; Yuen, K-Y

    2008-10-01

    In the last decade, as a result of the widespread use of PCR and DNA sequencing, 16S rDNA sequencing has played a pivotal role in the accurate identification of bacterial isolates and the discovery of novel bacteria in clinical microbiology laboratories. For bacterial identification, 16S rDNA sequencing is particularly important in the case of bacteria with unusual phenotypic profiles, rare bacteria, slow-growing bacteria, uncultivable bacteria and culture-negative infections. Not only has it provided insights into aetiologies of infectious disease, but it also helps clinicians in choosing antibiotics and in determining the duration of treatment and infection control procedures. With the use of 16S rDNA sequencing, 215 novel bacterial species, 29 of which belong to novel genera, have been discovered from human specimens in the past 7 years of the 21st century (2001-2007). One hundred of the 215 novel species, 15 belonging to novel genera, have been found in four or more subjects. The largest number of novel species discovered were of the genera Mycobacterium (n = 12) and Nocardia (n = 6). The oral cavity/dental-related specimens (n = 19) and the gastrointestinal tract (n = 26) were the most important sites for discovery and/or reservoirs of novel species. Among the 100 novel species, Streptococcus sinensis, Laribacter hongkongensis, Clostridium hathewayi and Borrelia spielmanii have been most thoroughly characterized, with the reservoirs and routes of transmission documented, and S. sinensis, L. hongkongensis and C. hathewayi have been found globally. One of the greatest hurdles in putting 16S rDNA sequencing into routine use in clinical microbiology laboratories is automation of the technology. The only step that can be automated at the moment is input of the 16S rDNA sequence of the bacterial isolate for identification into one of the software packages that will generate the result of the identity of the isolate on the basis of its sequence database. However

  19. The art of strain improvement of industrial lactic acid bacteria without the use of recombinant DNA technology

    PubMed Central

    2014-01-01

    The food industry is constantly striving to develop new products to fulfil the ever changing demands of consumers and the strict requirements of regulatory agencies. For foods based on microbial fermentation, this pushes the boundaries of microbial performance and requires the constant development of new starter cultures with novel properties. Since the use of ingredients in the food industry is tightly regulated and under close scrutiny by consumers, the use of recombinant DNA technology to improve microbial performance is currently not an option. As a result, the focus for improving strains for microbial fermentation is on classical strain improvement methods. Here we review the use of these techniques to improve the functionality of lactic acid bacteria starter cultures for application in industrial-scale food production. Methods will be described for improving the bacteriophage resistance of specific strains, improving their texture forming ability, increasing their tolerance to stress and modulating both the amount and identity of acids produced during fermentation. In addition, approaches to eliminating undesirable properties will be described. Techniques include random mutagenesis, directed evolution and dominant selection schemes. PMID:25186244

  20. Rapid and sensitive detection of foodborne pathogenic bacteria (Staphylococcus aureus) using an electrochemical DNA genomic biosensor and its application in fresh beef.

    PubMed

    Abdalhai, Mandour H; Fernandes, António Maximiano; Bashari, Mohand; Ji, Jian; He, Qian; Sun, Xiulan

    2014-12-31

    Rapid early detection of food contamination is the main key in food safety and quality control. Biosensors are emerging as a vibrant area of research, and the use of DNA biosensor recognition detectors is relatively new. In this study a genomic DNA biosensor system with a fixing and capture probe was modified by a sulfhydryl and amino group, respectively, as complementary with target DNA. After immobilization and hybridization, the following sandwich structure fixing DNA-target DNA-capture DNA-PbS NPs was formed to detect pathogenic bacteria (Staphylococuus aureus EF529607.1) by using GCE modified with (multiwalled carbon nanotubes-chitosan-bismuth) to increase the sensitivity of the electrode. The modification procedure was characterized by cyclic voltammetry and electrochemical impedance spectroscopy. The sandwich structure was dissolved in 1 M nitric acid to become accessible to the electrode, and the PbS NPs was measured in solution by differential pulse voltammetry (DPV). The results showed that the detection limit of the DNA sensor was 3.17 × 10(-14) M S. aureus using PbS NPs, whereas the result for beef samples was 1.23 ng/mL. Thus, according to the experimental results presented, the DNA biosensor exhibited high sensitivity and rapid response, and it will be useful for the food matrix.

  1. In Situ Detection of Bacteria within Paraffin-embedded Tissues Using a Digoxin-labeled DNA Probe Targeting 16S rRNA.

    PubMed

    Choi, Yun Sik; Kim, Yong Cheol; Baek, Keum Jin; Choi, Youngnim

    2015-05-21

    The presence of bacteria within the pocket epithelium and underlying connective tissue in gingival biopsies from patients with periodontitis has been reported using various methods, including electron microscopy, immunohistochemistry or immunofluorescence using bacteria-specific antibodies, and fluorescent in situ hybridization (FISH) using a fluorescence-labeled oligonucleotide probe. Nevertheless, these methods are not widely used due to technical limitation or difficulties. Here a method to localize bacteria within paraffin-embedded tissues using DIG-labeled DNA probes has been introduced. The paraffin-embedded tissues are the most common form of biopsy tissues available from pathology banks. Bacteria can be detected either in a species-specific or universal manner. Bacterial signals are detected as either discrete forms (coccus, rod, fusiform, and hairy form) of bacteria or dispersed forms. The technique allows other histological information to be obtained: the epithelia, connective tissue, inflammatory infiltrates, and blood vessels are well distinguished. This method can be used to study the role of bacteria in various diseases, such as periodontitis, cancers, and inflammatory immune diseases.

  2. Ribosomal PCR and DNA sequencing for detection and identification of bacteria: experience from 6 years of routine analyses of patient samples.

    PubMed

    Jensen, Kristine Helander; Dargis, Rimtas; Christensen, Jens Jørgen; Kemp, Michael

    2014-03-01

    The use of broad range PCR and DNA sequencing of bacterial 16S ribosomal RNA genes for routine diagnostics of bacterial infections was evaluated. Here, the results from more than 2600 analyses during a 6-year period (2003-2009) are presented. Almost half of the samples were from joints and bones, and the second most frequent origin of samples was from the central nervous system. Overall, 26% of all samples were positive for bacterial DNA and bacterial identification was obtained in 80% of the PCR-positive samples by subsequent DNA sequencing. Ambiguous species identification was noticed among non-haemolytic streptococci, especially within the mitis group. The data show that ribosomal PCR with subsequent DNA sequencing of the PCR product is a most valuable supplement to culture for identifying bacterial agents of both acute and prolonged infections. However, some bacteria, including non-haemolytic streptococci, may not be precisely identified.

  3. Distribution of human-specific bacteroidales and fecal indicator bacteria in an urban watershed impacted by sewage pollution, determined using RNA- and DNA-based quantitative PCR assays.

    PubMed

    Kapoor, Vikram; Pitkänen, Tarja; Ryu, Hodon; Elk, Michael; Wendell, David; Santo Domingo, Jorge W

    2015-01-01

    The identification of fecal pollution sources is commonly carried out using DNA-based methods. However, there is evidence that DNA can be associated with dead cells or present as "naked DNA" in the environment. Furthermore, it has been shown that rRNA-targeted reverse transcription-quantitative PCR (RT-qPCR) assays can be more sensitive than rRNA gene-based qPCR assays since metabolically active cells usually contain higher numbers of ribosomes than quiescent cells. To this end, we compared the detection frequency of host-specific markers and fecal bacteria using RNA-based RT-qPCR and DNA-based qPCR methods for water samples collected in sites impacted by combined sewer overflows. As a group, fecal bacteria were more frequently detected in most sites using RNA-based methods. Specifically, 8, 87, and 85% of the samples positive for general enterococci, Enterococcus faecalis, and Enterococcus faecium markers, respectively, were detected using RT-qPCR, but not with the qPCR assay counterpart. On average, two human-specific Bacteroidales markers were not detected when using DNA in 12% of the samples, while they were positive for all samples when using RNA (cDNA) as the template. Moreover, signal intensity was up to three orders of magnitude higher in RT-qPCR assays than in qPCR assays. The human-specific Bacteroidales markers exhibited moderate correlation with conventional fecal indicators using RT-qPCR results, suggesting the persistence of nonhuman sources of fecal pollution or the presence of false-positive signals. In general, the results from this study suggest that RNA-based assays can increase the detection sensitivity of fecal bacteria in urban watersheds impacted with human fecal sources.

  4. Oligo-DNA Custom Macroarray for Monitoring Major Pathogenic and Non-Pathogenic Fungi and Bacteria in the Phyllosphere of Apple Trees

    PubMed Central

    He, Ying-Hong; Isono, Sayaka; Shibuya, Makoto; Tsuji, Masaharu; Adkar Purushothama, Charith-Raj; Tanaka, Kazuaki; Sano, Teruo

    2012-01-01

    Background To monitor the richness in microbial inhabitants in the phyllosphere of apple trees cultivated under various cultural and environmental conditions, we developed an oligo-DNA macroarray for major pathogenic and non-pathogenic fungi and bacteria inhabiting the phyllosphere of apple trees. Methods and Findings First, we isolated culturable fungi and bacteria from apple orchards by an agar-plate culture method, and detected 32 fungal and 34 bacterial species. Alternaria, Aureobasidium, Cladosporium, Rhodotorula, Cystofilobasidium, and Epicoccum genera were predominant among the fungi, and Bacillus, Pseudomonas, Sphingomonas, Methylobacterium, and Pantoea genera were predominant among the bacteria. Based on the data, we selected 29 major non-pathogenic and 12 phytopathogenic fungi and bacteria as the targets of macroarray. Forty-one species-specific 40-base pair long oligo-DNA sequences were selected from the nucleotide sequences of rDNA-internal transcribed spacer region for fungi and 16S rDNA for bacteria. The oligo-DNAs were fixed on nylon membrane and hybridized with digoxigenin-labeled cRNA probes prepared for each species. All arrays except those for Alternaria, Bacillus, and their related species, were specifically hybridized. The array was sensitive enough to detect 103 CFU for Aureobasidium pullulans and Bacillus cereus. Nucleotide sequencing of 100 each of independent fungal rDNA-ITS and bacterial 16S-rDNA sequences from apple tree was in agreement with the macroarray data obtained using the same sample. Finally, we analyzed the richness in the microbial inhabitants in the samples collected from apple trees in four orchards. Major apple pathogens that cause scab, Alternaria blotch, and Marssonina blotch were detected along with several non-phytopathogenic fungal and bacterial inhabitants. Conclusions The macroarray technique presented here is a strong tool to monitor the major microbial species and the community structures in the phyllosphere of

  5. Specific oligonucleotide probes for in situ detection of a major group of gram-positive bacteria with low DNA G + C content.

    PubMed

    Meier, H; Amann, R; Ludwig, W; Schleifer, K H

    1999-05-01

    Almost one thousand 16S rRNA sequences of Gram-positive bacteria with a low DNA G + C content from public databases were analyzed using the ARB software package. A signature region was identified between positions 354 and 371 (E. coli numbering) for the Bacillus sub-branch of the Gram-positive bacteria with a low DNA G + C content, the former orders Bacillales and Lactobacillales. Three oligonucleotide probes, namely LGC354A, LGC354B, and LGC354C, were designed to target this diagnostic site. Their fluorescent derivatives were suitable for whole cell detection by fluorescence in situ hybridization (FISH). Hybridization conditions were adjusted for differentiation of target and related non-target reference species. When applying FISH to whole bacterial cells in a sample of activated sludge from a communal wastewater treatment plant, members of the Bacillus sub-branch were detected at levels from 0.01% of cells in samples fixed with paraformaldehyde to over 8 percent in the same samples fixed with ethanol and treated with lysozyme. The problems of quantitative in situ analysis of Gram-positive bacteria with a low DNA G + C content in biofilm flocs are discussed and recommendations made. Members of the Bacillus sub-branch were detected in different abundances in activated sludge samples from different wastewater plants.

  6. [Photoreactivating Activity of Bioluminescence: Repair of UV-damaged DNA of Escherichia coli Occurs with Assistance of lux-Genes of Marine Bacteria].

    PubMed

    Zavilgelsky, G B; Melkina, O E; Kotova, V Yu; Konopleva, M N; Manukhov, I V; Pustovoit, K Ss

    2015-01-01

    The UV resistance of luminescent bacteria Escherichia coli AB1886 uvrA6 (pLeo1) containing the plasmid with luxCDABE genes of marine bacteria Photobacterium leiognathi is approximately two times higher than the UV resistance of non-luminous bacteria E. coli AB1886 uvrA6. Introduction of phr::kan(r) mutations (a defect in the functional activity of photolyase) into the genome of E. coli AB1886 uvrA6 (pLeo1) completely removes the high UV resistance of the cells. Therefore, photoreactivation that involves bacterial photolyase contributes mainly to the bioluminescence-induced DNA repair. It is shown that photoreactivating activity of bioluminescence of P. leiognathi is about 2.5 times lower compared with that one induced by a light source with λ > 385 nm. It is also shown that an increase in the bioluminescence intensity, induced by UV radiation in E. coli bacterial cells with a plasmid containing the luxCD ABE genes under RecA-LexA-regulated promoters, occurs only 25-30 min later after UV irradiation of cells and does not contribute to DNA repair. A quorum sensing regulatory system is not involved in the DNA repair by photolyase.

  7. Hydrocarbon-degrading bacteria enriched by the Deepwater Horizon oil spill identified by cultivation and DNA-SIP

    PubMed Central

    Gutierrez, Tony; Singleton, David R; Berry, David; Yang, Tingting; Aitken, Michael D; Teske, Andreas

    2013-01-01

    The massive influx of crude oil into the Gulf of Mexico during the Deepwater Horizon (DWH) disaster triggered dramatic microbial community shifts in surface oil slick and deep plume waters. Previous work had shown several taxa, notably DWH Oceanospirillales, Cycloclasticus and Colwellia, were found to be enriched in these waters based on their dominance in conventional clone and pyrosequencing libraries and were thought to have had a significant role in the degradation of the oil. However, this type of community analysis data failed to provide direct evidence on the functional properties, such as hydrocarbon degradation of organisms. Using DNA-based stable-isotope probing with uniformly 13C-labelled hydrocarbons, we identified several aliphatic (Alcanivorax, Marinobacter)- and polycyclic aromatic hydrocarbon (Alteromonas, Cycloclasticus, Colwellia)-degrading bacteria. We also isolated several strains (Alcanivorax, Alteromonas, Cycloclasticus, Halomonas, Marinobacter and Pseudoalteromonas) with demonstrable hydrocarbon-degrading qualities from surface slick and plume water samples collected during the active phase of the spill. Some of these organisms accounted for the majority of sequence reads representing their respective taxa in a pyrosequencing data set constructed from the same and additional water column samples. Hitherto, Alcanivorax was not identified in any of the previous water column studies analysing the microbial response to the spill and we discuss its failure to respond to the oil. Collectively, our data provide unequivocal evidence on the hydrocarbon-degrading qualities for some of the dominant taxa enriched in surface and plume waters during the DWH oil spill, and a more complete understanding of their role in the fate of the oil. PMID:23788333

  8. Hydrocarbon-degrading bacteria enriched by the Deepwater Horizon oil spill identified by cultivation and DNA-SIP.

    PubMed

    Gutierrez, Tony; Singleton, David R; Berry, David; Yang, Tingting; Aitken, Michael D; Teske, Andreas

    2013-11-01

    The massive influx of crude oil into the Gulf of Mexico during the Deepwater Horizon (DWH) disaster triggered dramatic microbial community shifts in surface oil slick and deep plume waters. Previous work had shown several taxa, notably DWH Oceanospirillales, Cycloclasticus and Colwellia, were found to be enriched in these waters based on their dominance in conventional clone and pyrosequencing libraries and were thought to have had a significant role in the degradation of the oil. However, this type of community analysis data failed to provide direct evidence on the functional properties, such as hydrocarbon degradation of organisms. Using DNA-based stable-isotope probing with uniformly (13)C-labelled hydrocarbons, we identified several aliphatic (Alcanivorax, Marinobacter)- and polycyclic aromatic hydrocarbon (Alteromonas, Cycloclasticus, Colwellia)-degrading bacteria. We also isolated several strains (Alcanivorax, Alteromonas, Cycloclasticus, Halomonas, Marinobacter and Pseudoalteromonas) with demonstrable hydrocarbon-degrading qualities from surface slick and plume water samples collected during the active phase of the spill. Some of these organisms accounted for the majority of sequence reads representing their respective taxa in a pyrosequencing data set constructed from the same and additional water column samples. Hitherto, Alcanivorax was not identified in any of the previous water column studies analysing the microbial response to the spill and we discuss its failure to respond to the oil. Collectively, our data provide unequivocal evidence on the hydrocarbon-degrading qualities for some of the dominant taxa enriched in surface and plume waters during the DWH oil spill, and a more complete understanding of their role in the fate of the oil.

  9. Distribution of Human-Specific Bacteroidales and Fecal Indicator Bacteria in an Urban Watershed Impacted by Sewage Pollution, Determined Using RNA- and DNA-Based Quantitative PCR Assays

    PubMed Central

    Kapoor, Vikram; Pitkänen, Tarja; Ryu, Hodon; Elk, Michael

    2014-01-01

    The identification of fecal pollution sources is commonly carried out using DNA-based methods. However, there is evidence that DNA can be associated with dead cells or present as “naked DNA” in the environment. Furthermore, it has been shown that rRNA-targeted reverse transcription-quantitative PCR (RT-qPCR) assays can be more sensitive than rRNA gene-based qPCR assays since metabolically active cells usually contain higher numbers of ribosomes than quiescent cells. To this end, we compared the detection frequency of host-specific markers and fecal bacteria using RNA-based RT-qPCR and DNA-based qPCR methods for water samples collected in sites impacted by combined sewer overflows. As a group, fecal bacteria were more frequently detected in most sites using RNA-based methods. Specifically, 8, 87, and 85% of the samples positive for general enterococci, Enterococcus faecalis, and Enterococcus faecium markers, respectively, were detected using RT-qPCR, but not with the qPCR assay counterpart. On average, two human-specific Bacteroidales markers were not detected when using DNA in 12% of the samples, while they were positive for all samples when using RNA (cDNA) as the template. Moreover, signal intensity was up to three orders of magnitude higher in RT-qPCR assays than in qPCR assays. The human-specific Bacteroidales markers exhibited moderate correlation with conventional fecal indicators using RT-qPCR results, suggesting the persistence of nonhuman sources of fecal pollution or the presence of false-positive signals. In general, the results from this study suggest that RNA-based assays can increase the detection sensitivity of fecal bacteria in urban watersheds impacted with human fecal sources. PMID:25326295

  10. Monomorphic adenomas of the salivary glands. A clinico-pathologic study of 12 cases with immunohistochemical observation.

    PubMed

    Cho, K J; Kim, Y I

    1989-06-01

    Twelve cases of monomorphic adenoma of the salivary glands were histologically reclassified and their immunohistochemical reactivity for S-100 and cytokeratin was correlated. All patients underwent a benign clinical course. Individual tumors were well encapsulated and frequently showed a focal cystic change. Histologically, 9 cases were of the epithelial basaloid cell type and 3 cases were of the myoepithelioma variant. About one half of the epithelial type featured a mixture of trabecular and tubular patterns. The immunoreactivity to S-100 and cytokeratin varied. All basaloid cell adenomas were positive for cytokeratin, while S-100 positivity was found mostly in the stroma along with the varied reaction and intensity in the epithelial nests. In myoepitheliomas, cytokeratin was totally negative and S-100 was positive in 2 of 3 cases. The above findings suggest that the degree of participation of myoepithelial cells determines the phenotypic expression of monomorphic adenomas, and supports the hypothesis that the basaloid cell and myoepithelial types may be located on extreme ends of the same tumor spectrum with a wide range of pleomorphic adenoma in-between.

  11. An increase in negative supercoiling in bacteria reveals topology-reacting gene clusters and a homeostatic response mediated by the DNA topoisomerase I gene

    PubMed Central

    Ferrándiz, María-José; Martín-Galiano, Antonio J.; Arnanz, Cristina; Camacho-Soguero, Isabel; Tirado-Vélez, José-Manuel; de la Campa, Adela G.

    2016-01-01

    We studied the transcriptional response to an increase in DNA supercoiling in Streptococcus pneumoniae by using seconeolitsine, a new topoisomerase I inhibitor. A homeostatic response allowing recovery of supercoiling was observed in cells treated with subinhibitory seconeolitsine concentrations. Supercoiling increases of 40.7% (6 μM) and 72.9% (8 μM) were lowered to 8.5% and 44.1%, respectively. Likewise, drug removal facilitated the recovery of cell viability and DNA-supercoiling. Transcription of topoisomerase I depended on the supercoiling level. Also specific binding of topoisomerase I to the gyrase A gene promoter was detected by chromatin-immunoprecipitation. The transcriptomic response to 8 μM seconeolitsine had two stages. An early stage, associated to an increase in supercoiling, affected 10% of the genome. A late stage, manifested by supercoiling recovery, affected 2% of the genome. Nearly 25% of the early responsive genes formed 12 clusters with a coordinated transcription. Clusters were 6.7–31.4 kb in length and included 9–22 responsive genes. These clusters partially overlapped with those observed under DNA relaxation, suggesting that bacteria manage supercoiling stress using pathways with common components. This is the first report of a coordinated global transcriptomic response that is triggered by an increase in DNA supercoiling in bacteria. PMID:27378778

  12. The High Prevalence and Diversity of Chlamydiales DNA within Ixodes ricinus Ticks Suggest a Role for Ticks as Reservoirs and Vectors of Chlamydia-Related Bacteria

    PubMed Central

    Pilloux, Ludovic; Aeby, Sébastien; Gaümann, Rahel; Burri, Caroline; Beuret, Christian

    2015-01-01

    The Chlamydiales order is composed of nine families of strictly intracellular bacteria. Among them, Chlamydia trachomatis, C. pneumoniae, and C. psittaci are established human pathogens, whereas Waddlia chondrophila and Parachlamydia acanthamoebae have emerged as new pathogens in humans. However, despite their medical importance, their biodiversity and ecology remain to be studied. Even if arthropods and, particularly, ticks are well known to be vectors of numerous infectious agents such as viruses and bacteria, virtually nothing is known about ticks and chlamydia. This study investigated the prevalence of Chlamydiae in ticks. Specifically, 62,889 Ixodes ricinus ticks, consolidated into 8,534 pools, were sampled in 172 collection sites throughout Switzerland and were investigated using pan-Chlamydiales quantitative PCR (qPCR) for the presence of Chlamydiales DNA. Among the pools, 543 (6.4%) gave positive results and the estimated prevalence in individual ticks was 0.89%. Among those pools with positive results, we obtained 16S rRNA sequences for 359 samples, allowing classification of Chlamydiales DNA at the family level. A high level of biodiversity was observed, since six of the nine families belonging to the Chlamydiales order were detected. Those most common were Parachlamydiaceae (33.1%) and Rhabdochlamydiaceae (29.2%). “Unclassified Chlamydiales” (31.8%) were also often detected. Thanks to the huge amount of Chlamydiales DNA recovered from ticks, this report opens up new perspectives on further work focusing on whole-genome sequencing to increase our knowledge about Chlamydiales biodiversity. This report of an epidemiological study also demonstrates the presence of Chlamydia-related bacteria within Ixodes ricinus ticks and suggests a role for ticks in the transmission of and as a reservoir for these emerging pathogenic Chlamydia-related bacteria. PMID:26386066

  13. Detection of Sequence Polymorphism in Rubus Occidentalis L. Monomorphic Microsatellite Markers by High Resolution Melting

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Microsatellite, or simple sequence repeat (SSR) markers, are valuable as co-dominant genetic markers with a variety of applications such as DNA fingerprinting, linkage mapping, and population structure analysis. Development of microsatellite primers through the identification of appropriate repeate...

  14. Image findings of monomorphic non-hogdkin lymphoproliferative disorder in a post renal transplant patient diagnosed with fluorine-18 fluorodeoxyglucose-positron emission tomography/computed tomography.

    PubMed

    Kamaleshwaran, Koramadai Karuppusamy; Rajasekar, Thirugnanam; Shibu, Deepu; Radhakrishnan, Edathurthy Kalarikal; Shinto, Ajit Sugunan

    2014-07-01

    Post-transplant lymphoproliferative disorder (PTLD) is a heterogeneous group of lymphoid proliferations caused by immunosuppression after solid organ or bone marrow transplantation. PTLD is categorized by early lesion, polymorphic PTLD and monomorphic PTLD. Fluorine-18 fluorodeoxyglucose-positron emission tomography/computed tomography (F-18 FDG-PET/CT) scans have clinical significance in the evaluation of PTLD following renal transplantation. We report imaging findings of a monomorphic non-Hodgkin lymphoma, post renal transplant seen on FDG PET/CT in a 32-year-old lactating woman. Whole body FDG- ET/CT demonstrated uptake in right external iliac and inguinal lymph nodes.

  15. Image findings of monomorphic non-hogdkin lymphoproliferative disorder in a post renal transplant patient diagnosed with fluorine-18 fluorodeoxyglucose-positron emission tomography/computed tomography

    PubMed Central

    Kamaleshwaran, Koramadai Karuppusamy; Rajasekar, Thirugnanam; Shibu, Deepu; Radhakrishnan, Edathurthy Kalarikal; Shinto, Ajit Sugunan

    2014-01-01

    Post-transplant lymphoproliferative disorder (PTLD) is a heterogeneous group of lymphoid proliferations caused by immunosuppression after solid organ or bone marrow transplantation. PTLD is categorized by early lesion, polymorphic PTLD and monomorphic PTLD. Fluorine-18 fluorodeoxyglucose-positron emission tomography/computed tomography (F-18 FDG-PET/CT) scans have clinical significance in the evaluation of PTLD following renal transplantation. We report imaging findings of a monomorphic non-Hodgkin lymphoma, post renal transplant seen on FDG PET/CT in a 32-year-old lactating woman. Whole body FDG- ET/CT demonstrated uptake in right external iliac and inguinal lymph nodes. PMID:25210292

  16. PprA Protein Is Involved in Chromosome Segregation via Its Physical and Functional Interaction with DNA Gyrase in Irradiated Deinococcus radiodurans Bacteria

    PubMed Central

    Devigne, Alice; Guérin, Philippe; Lisboa, Johnny; Quevillon-Cheruel, Sophie; Armengaud, Jean; Sommer, Suzanne; Bouthier de la Tour, Claire

    2016-01-01

    ABSTRACT PprA, a radiation-induced Deinococcus-specific protein, was previously shown to be required for cell survival and accurate chromosome segregation after exposure to ionizing radiation. Here, we used an in vivo approach to determine, by shotgun proteomics, putative PprA partners coimmunoprecipitating with PprA when cells were exposed to gamma rays. Among them, we found the two subunits of DNA gyrase and, thus, chose to focus our work on characterizing the activities of the deinococcal DNA gyrase in the presence or absence of PprA. Loss of PprA rendered cells hypersensitive to novobiocin, an inhibitor of the B subunit of DNA gyrase. We showed that treatment of bacteria with novobiocin resulted in induction of the radiation desiccation response (RDR) regulon and in defects in chromosome segregation that were aggravated by the absence of PprA. In vitro, the deinococcal DNA gyrase, like other bacterial DNA gyrases, possesses DNA negative supercoiling and decatenation activities. These two activities are inhibited in vitro by novobiocin and nalidixic acid, whereas PprA specifically stimulates the decatenation activity of DNA gyrase. Together, these results suggest that PprA plays a major role in chromosome decatenation via its interaction with the deinococcal DNA gyrase when D. radiodurans cells are recovering from exposure to ionizing radiation. IMPORTANCE D. radiodurans is one of the most radiation-resistant organisms known. This bacterium is able to cope with high levels of DNA lesions generated by exposure to extreme doses of ionizing radiation and to reconstruct a functional genome from hundreds of radiation-induced chromosomal fragments. Here, we identified partners of PprA, a radiation-induced Deinococcus-specific protein, previously shown to be required for radioresistance. Our study leads to three main findings: (i) PprA interacts with DNA gyrase after irradiation, (ii) treatment of cells with novobiocin results in defects in chromosome segregation

  17. Clonetegration Using OSIP Plasmids: One-Step DNA Assembly and Site-Specific Genomic Integration in Bacteria.

    PubMed

    Cui, Lun; Shearwin, Keith E

    2017-01-01

    Clonetegration is a method for site-specific insertion of DNA into prokaryotic chromosomes, based on bacteriophage integrases. The method combines DNA cloning/assembly and chromosomal integration into a single step, providing a simple and rapid strategy for inserting DNA sequences into bacterial chromosomes. PMID:27671938

  18. Characterization of single-stranded DNA-binding proteins from the psychrophilic bacteria Desulfotalea psychrophila, Flavobacterium psychrophilum, Psychrobacter arcticus, Psychrobacter cryohalolentis, Psychromonas ingrahamii, Psychroflexus torquis, and Photobacterium profundum

    PubMed Central

    2014-01-01

    Background Single-stranded DNA-binding proteins (SSBs) play essential roles in DNA replication, recombination and repair in Bacteria, Archaea and Eukarya. In recent years, there has been an increasing interest in SSBs, since they find numerous applications in diverse molecular biology and analytical methods. Results We report the characterization of single-stranded DNA-binding proteins from the psychrophilic bacteria Desulfotalea psychrophila (DpsSSB), Flavobacterium psychrophilum (FpsSSB), Psychrobacter arcticus (ParSSB), Psychrobacter cryohalolentis (PcrSSB), Psychromonas ingrahamii (PinSSB), Photobacterium profundum (PprSSB), and Psychroflexus torquis (PtoSSB). The proteins show a high differential within the molecular mass of their monomers and the length of their amino acid sequences. The high level of identity and similarity in respect to the EcoSSB is related to the OB-fold and some of the last amino acid residues. They are functional as homotetramers, with each monomer encoding one single stranded DNA binding domain (OB-fold). The fluorescence titrations indicated that the length of the ssDNA-binding site size is approximately 30 ± 2 nucleotides for the PinSSB, 31 ± 2 nucleotides for the DpsSSB, and 32 ± 2 nucleotides for the ParSSB, PcrSSB, PprSSB and PtoSSB. They also demonstrated that it is salt independent. However, when the ionic strength was changed from low salt to high, binding-mode transition was observed for the FpsSSB, at 31 ± 2 nucleotides and 45 ± 2 nucleotides, respectively. As expected, the SSB proteins under study cause duplex DNA destabilization. The greatest decrease in duplex DNA melting temperature was observed in the presence of the PtoSSB 17°C. The SSBs in question possess relatively high thermostability for proteins derived from cold-adapted bacteria. Conclusion The results showed that SSB proteins from psychrophilic microorganisms are typical bacterial SSBs and possess relatively high thermostability

  19. Eukaryote-Made Thermostable DNA Polymerase Enables Rapid PCR-Based Detection of Mycoplasma, Ureaplasma and Other Bacteria in the Amniotic Fluid of Preterm Labor Cases

    PubMed Central

    Yoneda, Noriko; Yoneda, Satoshi; Mori, Masashi; Tabata, Homare; Minami, Hiroshi; Saito, Shigeru; Kitajima, Isao

    2015-01-01

    Background Intra-amniotic infection has long been recognized as the leading cause of preterm delivery. Microbial culture is the gold standard for the detection of intra-amniotic infection, but several days are required, and many bacterial species in the amniotic fluid are difficult to cultivate. Methods We developed a novel nested-PCR-based assay for detecting Mycoplasma, Ureaplasma, other bacteria and fungi in amniotic fluid samples within three hours of sample collection. To detect prokaryotes, eukaryote-made thermostable DNA polymerase, which is free from bacterial DNA contamination, is used in combination with bacterial universal primers. In contrast, to detect eukaryotes, conventional bacterially-made thermostable DNA polymerase is used in combination with fungal universal primers. To assess the validity of the PCR assay, we compared the PCR and conventional culture results using 300 amniotic fluid samples. Results Based on the detection level (positive and negative), 93.3% (280/300) of Mycoplasma, 94.3% (283/300) of Ureaplasma, 89.3% (268/300) of other bacteria and 99.7% (299/300) of fungi matched the culture results. Meanwhile, concerning the detection of bacteria other than Mycoplasma and Ureaplasma, 228 samples were negative according to the PCR method, 98.2% (224/228) of which were also negative based on the culture method. Employing the devised primer sets, mixed amniotic fluid infections of Mycoplasma, Ureaplasma and/or other bacteria could be clearly distinguished. In addition, we also attempted to compare the relative abundance in 28 amniotic fluid samples with mixed infection, and judged dominance by comparing the Ct values of quantitative real-time PCR. Conclusions We developed a novel PCR assay for the rapid detection of Mycoplasma, Ureaplasma, other bacteria and fungi in amniotic fluid samples. This assay can also be applied to accurately diagnose the absence of bacteria in samples. We believe that this assay will positively contribute to the

  20. Bacteria capable of degrading anthracene, phenanthrene, and fluoranthene as revealed by DNA based stable-isotope probing in a forest soil.

    PubMed

    Song, Mengke; Jiang, Longfei; Zhang, Dayi; Luo, Chunling; Wang, Yan; Yu, Zhiqiang; Yin, Hua; Zhang, Gan

    2016-05-01

    Information on microorganisms possessing the ability to metabolize different polycyclic aromatic hydrocarbons (PAHs) in complex environments helps in understanding PAHs behavior in natural environment and developing bioremediation strategies. In the present study, stable-isotope probing (SIP) was applied to investigate degraders of PAHs in a forest soil with the addition of individually (13)C-labeled phenanthrene, anthracene, and fluoranthene. Three distinct phylotypes were identified as the active phenanthrene-, anthracene- and fluoranthene-degrading bacteria. The putative phenanthrene degraders were classified as belonging to the genus Sphingomona. For anthracene, bacteria of the genus Rhodanobacter were the putative degraders, and in the microcosm amended with fluoranthene, the putative degraders were identified as belonging to the phylum Acidobacteria. Our results from DNA-SIP are the first to directly link Rhodanobacter- and Acidobacteria-related bacteria with anthracene and fluoranthene degradation, respectively. The results also illustrate the specificity and diversity of three- and four-ring PAHs degraders in forest soil, contributes to our understanding on natural PAHs biodegradation processes, and also proves the feasibility and practicality of DNA-based SIP for linking functions with identity especially uncultured microorganisms in complex microbial biota. PMID:26808242

  1. Disordered nucleiome: Abundance of intrinsic disorder in the DNA- and RNA-binding proteins in 1121 species from Eukaryota, Bacteria and Archaea.

    PubMed

    Wang, Chen; Uversky, Vladimir N; Kurgan, Lukasz

    2016-05-01

    Intrinsically disordered proteins (IDPs) are abundant in various proteomes, where they play numerous important roles and complement biological activities of ordered proteins. Among functions assigned to IDPs are interactions with nucleic acids. However, often, such assignments are made based on the guilty-by-association principle. The validity of the extension of these correlations to all nucleic acid binding proteins has never been analyzed on a large scale across all domains of life. To fill this gap, we perform a comprehensive computational analysis of the abundance of intrinsic disorder and intrinsically disordered domains in nucleiomes (∼548 000 nucleic acid binding proteins) of 1121 species from Archaea, Bacteria and Eukaryota. Nucleiome is a whole complement of proteins involved in interactions with nucleic acids. We show that relative to other proteins in the corresponding proteomes, the DNA-binding proteins have significantly increased disorder content and are significantly enriched in disordered domains in Eukaryotes but not in Archaea and Bacteria. The RNA-binding proteins are significantly enriched in the disordered domains in Bacteria, Archaea and Eukaryota, while the overall abundance of disorder in these proteins is significantly increased in Bacteria, Archaea, animals and fungi. The high abundance of disorder in nucleiomes supports the notion that the nucleic acid binding proteins often require intrinsic disorder for their functions and regulation.

  2. Use of DNA-stable isotope probing and functional gene probes to investigate the diversity of methyl chloride-utilizing bacteria in soil.

    PubMed

    Borodina, Elena; Cox, Michael J; McDonald, Ian R; Murrell, J Colin

    2005-09-01

    Enrichment and isolation of methyl chloride-utilizing bacteria from various terrestrial environments, including woodland and forest soils, resulted in the identification of seven methyl chloride-utilizing strains belonging to the genus Hyphomicrobium, an Aminobacter strain TW23 and strain WG1, which grouped closely with the genus Mesorhizobium. Methyl chloride enrichment cultures were dominated by Hyphomicrobium species, indicating that these bacteria were most suited to growth under the enrichment and isolation conditions used. However, the application of culture-independent techniques such as DNA-stable isotope probing and the use of a functional gene probe targeting cmuA, which encodes the methyltransferase catalysing the first step in bacterial methyl chloride metabolism, indicated a greater diversity of methyl chloride-utilizing bacteria in the terrestrial environment, compared with the diversity of soil isolates obtained via the enrichment and isolation procedure. It also revealed the presence of as yet uncultured and potentially novel methyl chloride-degrading bacteria in soil. PMID:16104855

  3. Disordered nucleiome: Abundance of intrinsic disorder in the DNA- and RNA-binding proteins in 1121 species from Eukaryota, Bacteria and Archaea.

    PubMed

    Wang, Chen; Uversky, Vladimir N; Kurgan, Lukasz

    2016-05-01

    Intrinsically disordered proteins (IDPs) are abundant in various proteomes, where they play numerous important roles and complement biological activities of ordered proteins. Among functions assigned to IDPs are interactions with nucleic acids. However, often, such assignments are made based on the guilty-by-association principle. The validity of the extension of these correlations to all nucleic acid binding proteins has never been analyzed on a large scale across all domains of life. To fill this gap, we perform a comprehensive computational analysis of the abundance of intrinsic disorder and intrinsically disordered domains in nucleiomes (∼548 000 nucleic acid binding proteins) of 1121 species from Archaea, Bacteria and Eukaryota. Nucleiome is a whole complement of proteins involved in interactions with nucleic acids. We show that relative to other proteins in the corresponding proteomes, the DNA-binding proteins have significantly increased disorder content and are significantly enriched in disordered domains in Eukaryotes but not in Archaea and Bacteria. The RNA-binding proteins are significantly enriched in the disordered domains in Bacteria, Archaea and Eukaryota, while the overall abundance of disorder in these proteins is significantly increased in Bacteria, Archaea, animals and fungi. The high abundance of disorder in nucleiomes supports the notion that the nucleic acid binding proteins often require intrinsic disorder for their functions and regulation. PMID:27037624

  4. Competitive binding-based optical DNA mapping for fast identification of bacteria--multi-ligand transfer matrix theory and experimental applications on Escherichia coli.

    PubMed

    Nilsson, Adam N; Emilsson, Gustav; Nyberg, Lena K; Noble, Charleston; Stadler, Liselott Svensson; Fritzsche, Joachim; Moore, Edward R B; Tegenfeldt, Jonas O; Ambjörnsson, Tobias; Westerlund, Fredrik

    2014-09-01

    We demonstrate a single DNA molecule optical mapping assay able to resolve a specific Escherichia coli strain from other strains. The assay is based on competitive binding of the fluorescent dye YOYO-1 and the AT-specific antibiotic netropsin. The optical map is visualized by stretching the DNA molecules in nanofluidic channels. We optimize the experimental conditions to obtain reproducible barcodes containing as much information as possible. We implement a multi-ligand transfer matrix method for calculating theoretical barcodes from known DNA sequences. Our method extends previous theoretical approaches for competitive binding of two types of ligands to many types of ligands and introduces a recursive approach that allows long barcodes to be calculated with standard computer floating point formats. The identification of a specific E. coli strain (CCUG 10979) is based on mapping of 50-160 kilobasepair experimental DNA fragments onto the theoretical genome using the developed theory. Our identification protocol introduces two theoretical constructs: a P-value for a best experiment-theory match and an information score threshold. The developed methods provide a novel optical mapping toolbox for identification of bacterial species and strains. The protocol does not require cultivation of bacteria or DNA amplification, which allows for ultra-fast identification of bacterial pathogens.

  5. Seasonal Sexual Segregation by Monomorphic Sooty Shearwaters Puffinus griseus Reflects Different Reproductive Roles during the Pre-Laying Period

    PubMed Central

    Hedd, April; Montevecchi, William A.; Phillips, Richard A.; Fifield, David A.

    2014-01-01

    Tracking technology has revolutionized knowledge of seabird movements; yet, few studies have examined sex differences in distribution and behavior of small to medium-sized, sexually-monomorphic seabirds. Application of bird-borne geolocation-immersion loggers revealed seasonal segregation in the sexually-monomorphic Sooty Shearwater Puffinus griseus, mainly in the pre-laying period, when there were clear differences in reproductive roles. Shearwaters first returned to the Falkland Islands on 27 Sept±8 d; males, on average, 8 d earlier than females. Prior to egg-laying, distribution at sea, colony attendance and behaviour depended on sex. Males foraged locally over the southern Patagonian Shelf and Burdwood Bank, spending mainly single days at sea and intervening nights in the burrow. Females, who flew for more of the day during this time, foraged in more distant areas of the northern Patagonian Shelf and Argentine Basin that were deeper, warmer and relatively more productive. Attendance of females at the colony was also more variable than that of males and, overall, males were present for significantly more of the pre-laying period (38 vs. 19% of time). Sex differences were reduced following egg-laying, with males and females using similar foraging areas and making trips of similar mean duration in incubation (7.6±2.7 d) and chick-rearing (1.4±1.3 d). Congruence continued into the non-breeding period, with both sexes showing similar patterns of activity and areas of occupancy in the NW Atlantic. Thus, seasonal changes in reproductive roles influenced patterns of sexual segregation; this occurred only early in the season, when male Sooty Shearwaters foraged locally, returning regularly to the colony to defend (or maintain) the burrow or the mate, while females concentrated on building resources for egg development in distant and relatively more productive waters. PMID:24416429

  6. High resolution melting detects sequence polymorphism in rubus occidentalis L. monomorphic microsatellite markers

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Microsatellite, or simple sequence repeat (SSR) markers, are valuable as co-dominant genetic markers with a variety of applications such as DNA fingerprinting, linkage mapping, and population structure analysis. However, primer pairs designed from the regions that flank SSRs often generate fragment...

  7. A novel approach to propagate flavivirus infectious cDNA clones in bacteria by introducing tandem repeat sequences upstream of virus genome.

    PubMed

    Pu, Szu-Yuan; Wu, Ren-Huang; Tsai, Ming-Han; Yang, Chi-Chen; Chang, Chung-Ming; Yueh, Andrew

    2014-07-01

    Despite tremendous efforts to improve the methodology for constructing flavivirus infectious cDNAs, the manipulation of flavivirus cDNAs remains a difficult task in bacteria. Here, we successfully propagated DNA-launched type 2 dengue virus (DENV2) and Japanese encephalitis virus (JEV) infectious cDNAs by introducing seven repeats of the tetracycline-response element (7×TRE) and a minimal cytomegalovirus (CMVmin) promoter upstream of the viral genome. Insertion of the 7×TRE-CMVmin sequence upstream of the DENV2 or JEV genome decreased the cryptic E. coli promoter (ECP) activity of the viral genome in bacteria, as measured using fusion constructs containing DENV2 or JEV segments and the reporter gene Renilla luciferase in an empty vector. The growth kinetics of recombinant viruses derived from DNA-launched DENV2 and JEV infectious cDNAs were similar to those of parental viruses. Similarly, RNA-launched DENV2 infectious cDNAs were generated by inserting 7×TRE-CMVmin, five repeats of the GAL4 upstream activating sequence, or five repeats of BamHI linkers upstream of the DENV2 genome. All three tandem repeat sequences decreased the ECP activity of the DENV2 genome in bacteria. Notably, 7×TRE-CMVmin stabilized RNA-launched JEV infectious cDNAs and reduced the ECP activity of the JEV genome in bacteria. The growth kinetics of recombinant viruses derived from RNA-launched DENV2 and JEV infectious cDNAs displayed patterns similar to those of the parental viruses. These results support a novel methodology for constructing flavivirus infectious cDNAs, which will facilitate research in virology, viral pathogenesis and vaccine development of flaviviruses and other RNA viruses. PMID:24728712

  8. Diagnosis and quantification of glycerol assimilating denitrifying bacteria in an integrated fixed-film activated sludge reactor via 13C DNA stable-isotope probing.

    PubMed

    Lu, Huijie; Chandran, Kartik

    2010-12-01

    Glycerol, a byproduct of biodiesel and oleo-chemical manufacturing operations, represents an attractive alternate to methanol as a carbon and electron donor for enhanced denitrification. However, unlike methanol, little is known about the diversity and activity of glycerol assimilating bacteria in activated sludge. In this study, the microbial ecology of glycerol assimilating denitrifying bacteria in a sequencing batch integrated fixed film activated sludge (SB-IFAS) reactor was investigated using (13)C-DNA stable isotope probing (SIP). During steady state SB-IFAS reactor operation, near complete nitrate removal (92.7 ± 5.8%) was achieved. Based on (13)C DNA clone libraries obtained after 360 days of SB-IFAS reactor operation, bacteria related to Comamonas spp. and Diaphorobacter spp. dominated in the suspended phase communities. (13)C assimilating members in the biofilm community were phylogenetically more diverse and were related to Comamonas spp., Bradyrhizobium spp., and Tessaracoccus spp. Possibly owing to greater substrate availability in the suspended phase, the glycerol-assimilating denitrifying populations (quantified by real-time PCR) were more abundant therein than in the biofilm phase. The biomass in the suspended phase also had a higher specific denitrification rate than the biofilm phase (p = 4.33e-4), and contributed to 69.7 ± 4.5% of the overall N-removal on a mass basis. The kinetics of glycerol based denitrification by suspended phase biomass were approximately 3 times higher than with methanol. Previously identified methanol assimilating denitrifying bacteria were not associated with glycerol assimilation, thereby suggesting limited cross-utilization of these two substrates for denitrification in the system tested.

  9. A trade-off between oxidative stress resistance and DNA repair plays a role in the evolution of elevated mutation rates in bacteria.

    PubMed

    Torres-Barceló, Clara; Cabot, Gabriel; Oliver, Antonio; Buckling, Angus; Maclean, R Craig

    2013-04-22

    The dominant paradigm for the evolution of mutator alleles in bacterial populations is that they spread by indirect selection for linked beneficial mutations when bacteria are poorly adapted. In this paper, we challenge the ubiquity of this paradigm by demonstrating that a clinically important stressor, hydrogen peroxide, generates direct selection for an elevated mutation rate in the pathogenic bacterium Pseudomonas aeruginosa as a consequence of a trade-off between the fidelity of DNA repair and hydrogen peroxide resistance. We demonstrate that the biochemical mechanism underlying this trade-off in the case of mutS is the elevated secretion of catalase by the mutator strain. Our results provide, to our knowledge, the first experimental evidence that direct selection can favour mutator alleles in bacterial populations, and pave the way for future studies to understand how mutation and DNA repair are linked to stress responses and how this affects the evolution of bacterial mutation rates.

  10. Kinetics of killing Listeria monocytogenes by macrophages: correlation of /sup 3/H-DNA release from labeled bacteria and changes in numbers of viable organisms by mathematical model

    SciTech Connect

    Davies, W.A.

    1982-12-01

    Conventional methods of assessing antibacterial activities of macrophages by viable counting are limited by the precision of the statistics and are difficult to interpret quantitatively because of unrestrained extracellular growth of bacteria. An alternative technique based on the release of radioactive DNA from labeled bacteria has been offered as overcoming these drawbacks. To assess it for use with macrophages I have made a correlation with the conventional viable counting method using a mathematical model. Opsonized Listeria monocytogenes labeled with /sup 3/H-thymidine were exposed to rat macrophages for periods up to 4 hr. Numbers of viable bacteria determined after sonication increased exponentially in the absence of live cells and this growth rate was progressively inhibited by increasing numbers of macrophages. After a lag period of 30-60 min soluble /sup 3/H appeared in the supernatant, the amount increasing with time and numbers of macrophages. To correlate these data I developed a mathematical model that considered that changes in numbers of viable organisms were due to the difference between rates of 1) growth of extracellular bacteria and 2) killing within the macrophage. On the basis of this model curves of best fit to the viable counts data were used to predict the release of radioactivity, assuming that death of a bacterium led to the total release of its label. These predictions and the experimental data agreed well, the lag period of 30-60 min between death of the bacterium and release of radioactivity being consistent with intracellular digestion. Release of soluble radioactivity appears to be an accurate reflection of the number of bacteria killed within the macrophage.

  11. Identification of metabolically active bacteria in the gut of the generalist Spodoptera littoralis via DNA stable isotope probing using 13C-glucose.

    PubMed

    Shao, Yongqi; Arias-Cordero, Erika M; Boland, Wilhelm

    2013-01-01

    Guts of most insects are inhabited by complex communities of symbiotic nonpathogenic bacteria. Within such microbial communities it is possible to identify commensal or mutualistic bacteria species. The latter ones, have been observed to serve multiple functions to the insect, i.e. helping in insect reproduction(1), boosting the immune response(2), pheromone production(3), as well as nutrition, including the synthesis of essential amino acids(4,) among others.     Due to the importance of these associations, many efforts have been made to characterize the communities down to the individual members. However, most of these efforts were either based on cultivation methods or relied on the generation of 16S rRNA gene fragments which were sequenced for final identification. Unfortunately, these approaches only identified the bacterial species present in the gut and provided no information on the metabolic activity of the microorganisms. To characterize the metabolically active bacterial species in the gut of an insect, we used stable isotope probing (SIP) in vivo employing (13)C-glucose as a universal substrate. This is a promising culture-free technique that allows the linkage of microbial phylogenies to their particular metabolic activity. This is possible by tracking stable, isotope labeled atoms from substrates into microbial biomarkers, such as DNA and RNA(5). The incorporation of (13)C isotopes into DNA increases the density of the labeled DNA compared to the unlabeled ((12)C) one. In the end, the (13)C-labeled DNA or RNA is separated by density-gradient ultracentrifugation from the (12)C-unlabeled similar one(6). Subsequent molecular analysis of the separated nucleic acid isotopomers provides the connection between metabolic activity and identity of the species. Here, we present the protocol used to characterize the metabolically active bacteria in the gut of a generalist insect (our model system), Spodoptera littoralis (Lepidoptera, Noctuidae). The

  12. Identification of Metabolically Active Bacteria in the Gut of the Generalist Spodoptera littoralis via DNA Stable Isotope Probing Using 13C-Glucose

    PubMed Central

    Boland, Wilhelm

    2013-01-01

    Guts of most insects are inhabited by complex communities of symbiotic nonpathogenic bacteria. Within such microbial communities it is possible to identify commensal or mutualistic bacteria species. The latter ones, have been observed to serve multiple functions to the insect, i.e. helping in insect reproduction1, boosting the immune response2, pheromone production3, as well as nutrition, including the synthesis of essential amino acids4, among others.     Due to the importance of these associations, many efforts have been made to characterize the communities down to the individual members. However, most of these efforts were either based on cultivation methods or relied on the generation of 16S rRNA gene fragments which were sequenced for final identification. Unfortunately, these approaches only identified the bacterial species present in the gut and provided no information on the metabolic activity of the microorganisms. To characterize the metabolically active bacterial species in the gut of an insect, we used stable isotope probing (SIP) in vivo employing 13C-glucose as a universal substrate. This is a promising culture-free technique that allows the linkage of microbial phylogenies to their particular metabolic activity. This is possible by tracking stable, isotope labeled atoms from substrates into microbial biomarkers, such as DNA and RNA5. The incorporation of 13C isotopes into DNA increases the density of the labeled DNA compared to the unlabeled (12C) one. In the end, the 13C-labeled DNA or RNA is separated by density-gradient ultracentrifugation from the 12C-unlabeled similar one6. Subsequent molecular analysis of the separated nucleic acid isotopomers provides the connection between metabolic activity and identity of the species. Here, we present the protocol used to characterize the metabolically active bacteria in the gut of a generalist insect (our model system), Spodoptera littoralis (Lepidoptera, Noctuidae). The phylogenetic analysis of the DNA

  13. Common mechanisms of DNA translocation motors in bacteria and viruses using one-way revolution mechanism without rotation.

    PubMed

    Guo, Peixuan; Zhao, Zhengyi; Haak, Jeannie; Wang, Shaoying; Wu, Dong; Meng, Bing; Weitao, Tao

    2014-01-01

    Biomotors were once described into two categories: linear motor and rotation motor. Recently, a third type of biomotor with revolution mechanism without rotation has been discovered. By analogy, rotation resembles the Earth rotating on its axis in a complete cycle every 24h, while revolution resembles the Earth revolving around the Sun one circle per 365 days (see animations http://nanobio.uky.edu/movie.html). The action of revolution that enables a motor free of coiling and torque has solved many puzzles and debates that have occurred throughout the history of viral DNA packaging motor studies. It also settles the discrepancies concerning the structure, stoichiometry, and functioning of DNA translocation motors. This review uses bacteriophages Phi29, HK97, SPP1, P22, T4, and T7 as well as bacterial DNA translocase FtsK and SpoIIIE or the large eukaryotic dsDNA viruses such as mimivirus and vaccinia virus as examples to elucidate the puzzles. These motors use ATPase, some of which have been confirmed to be a hexamer, to revolve around the dsDNA sequentially. ATP binding induces conformational change and possibly an entropy alteration in ATPase to a high affinity toward dsDNA; but ATP hydrolysis triggers another entropic and conformational change in ATPase to a low affinity for DNA, by which dsDNA is pushed toward an adjacent ATPase subunit. The rotation and revolution mechanisms can be distinguished by the size of channel: the channels of rotation motors are equal to or smaller than 2 nm, that is the size of dsDNA, whereas channels of revolution motors are larger than 3 nm. Rotation motors use parallel threads to operate with a right-handed channel, while revolution motors use a left-handed channel to drive the right-handed DNA in an anti-chiral arrangement. Coordination of several vector factors in the same direction makes viral DNA-packaging motors unusually powerful and effective. Revolution mechanism that avoids DNA coiling in translocating the lengthy genomic

  14. Common mechanisms of DNA translocation motors in bacteria and viruses using one-way revolution mechanism without rotation.

    PubMed

    Guo, Peixuan; Zhao, Zhengyi; Haak, Jeannie; Wang, Shaoying; Wu, Dong; Meng, Bing; Weitao, Tao

    2014-01-01

    Biomotors were once described into two categories: linear motor and rotation motor. Recently, a third type of biomotor with revolution mechanism without rotation has been discovered. By analogy, rotation resembles the Earth rotating on its axis in a complete cycle every 24h, while revolution resembles the Earth revolving around the Sun one circle per 365 days (see animations http://nanobio.uky.edu/movie.html). The action of revolution that enables a motor free of coiling and torque has solved many puzzles and debates that have occurred throughout the history of viral DNA packaging motor studies. It also settles the discrepancies concerning the structure, stoichiometry, and functioning of DNA translocation motors. This review uses bacteriophages Phi29, HK97, SPP1, P22, T4, and T7 as well as bacterial DNA translocase FtsK and SpoIIIE or the large eukaryotic dsDNA viruses such as mimivirus and vaccinia virus as examples to elucidate the puzzles. These motors use ATPase, some of which have been confirmed to be a hexamer, to revolve around the dsDNA sequentially. ATP binding induces conformational change and possibly an entropy alteration in ATPase to a high affinity toward dsDNA; but ATP hydrolysis triggers another entropic and conformational change in ATPase to a low affinity for DNA, by which dsDNA is pushed toward an adjacent ATPase subunit. The rotation and revolution mechanisms can be distinguished by the size of channel: the channels of rotation motors are equal to or smaller than 2 nm, that is the size of dsDNA, whereas channels of revolution motors are larger than 3 nm. Rotation motors use parallel threads to operate with a right-handed channel, while revolution motors use a left-handed channel to drive the right-handed DNA in an anti-chiral arrangement. Coordination of several vector factors in the same direction makes viral DNA-packaging motors unusually powerful and effective. Revolution mechanism that avoids DNA coiling in translocating the lengthy genomic

  15. Spatial Dependence of DNA Damage in Bacteria due to Low-Temperature Plasma Application as Assessed at the Single Cell Level

    PubMed Central

    Privat-Maldonado, Angela; O’Connell, Deborah; Welch, Emma; Vann, Roddy; van der Woude, Marjan W.

    2016-01-01

    Low temperature plasmas (LTPs) generate a cocktail of reactive nitrogen and oxygen species (RNOS) with bactericidal activity. The RNOS however are spatially unevenly distributed in the plasma. Here we test the hypothesis that this distribution will affect the mechanisms underpinning plasma bactericidal activity focussing on the level of DNA damage in situ. For the first time, a quantitative, single cell approach was applied to assess the level of DNA damage in bacteria as a function of the radial distance from the centre of the plasma jet. Salmonella enterica on a solid, dry surface was treated with two types of LTP: an atmospheric-pressure dielectric barrier discharge plasma jet (charged and neutral species) and a radio-frequency atmospheric-pressure plasma jet (neutral species). In both cases, there was an inverse correlation between the degree of DNA damage and the radial distance from the centre of the plasma, with the highest DNA damage occurring directly under the plasma. This trend was also observed with Staphylococcus aureus. LTP-generated UV radiation was eliminated as a contributing factor. Thus valuable mechanistic information can be obtained from assays on biological material, which can inform the development of LTP as a complementary or alternative therapy for (topical) bacterial infections. PMID:27759098

  16. Lytic enzyme, labiase for a broad range of Gram-positive bacteria and its application to analyze functional DNA/RNA.

    PubMed

    Niwa, Takashi; Kawamura, Yoshiaki; Katagiri, Yoshihiro; Ezaki, Takayuki

    2005-05-01

    The lytic activity of labiase and achromopeptidase for bacterial DNA/RNA extraction were compared. Rapid lysis of many bacterial strains was observed with labiase followed by SDS treatment. Both labiase and achromopeptidase showed high lytic activity against bacterial strains with the A1alpha chemotype (e.g., Aerococcus viridans) and the A3alpha chemotype (e.g., Staphylococcus epidermidis) for cell wall peptidoglycan structures. The lytic activity of labiase was higher than that of achromopeptidase against strains with the A1gamma chemotype (e.g., Bacillus subtilis). The activity of labiase was not detrimentally affected with increasing NaCl concentration. Labiase lysates were successfully used for rapid extraction of DNA and RNA, whereas achromopeptidase lysates degraded RNA. The DNA and RNA obtained were successfully used for 16S rRNA amplification and real-time RT-PCR detection. It is concluded that labiase is useful for rapid lysis of a wide variety of Gram-positive bacteria and can be used for DNA/RNA isolation protocols.

  17. Lytic enzyme, labiase for a broad range of Gram-positive bacteria and its application to analyze functional DNA/RNA.

    PubMed

    Niwa, Takashi; Kawamura, Yoshiaki; Katagiri, Yoshihiro; Ezaki, Takayuki

    2005-05-01

    The lytic activity of labiase and achromopeptidase for bacterial DNA/RNA extraction were compared. Rapid lysis of many bacterial strains was observed with labiase followed by SDS treatment. Both labiase and achromopeptidase showed high lytic activity against bacterial strains with the A1alpha chemotype (e.g., Aerococcus viridans) and the A3alpha chemotype (e.g., Staphylococcus epidermidis) for cell wall peptidoglycan structures. The lytic activity of labiase was higher than that of achromopeptidase against strains with the A1gamma chemotype (e.g., Bacillus subtilis). The activity of labiase was not detrimentally affected with increasing NaCl concentration. Labiase lysates were successfully used for rapid extraction of DNA and RNA, whereas achromopeptidase lysates degraded RNA. The DNA and RNA obtained were successfully used for 16S rRNA amplification and real-time RT-PCR detection. It is concluded that labiase is useful for rapid lysis of a wide variety of Gram-positive bacteria and can be used for DNA/RNA isolation protocols. PMID:15722152

  18. Common Mechanisms of DNA translocation motors in Bacteria and Viruses Using One-way Revolution Mechanism without Rotation

    PubMed Central

    Guo, Peixuan; Zhao, Zhengyi; Haak, Jeannie; Wang, Shaoying; Weitao, Tao

    2014-01-01

    Biomotors were once classified into two categories: linear motor and rotation motor. For decades, the viral DNA-packaging motor has been popularly believed to be a five-fold rotation motor. Recently, a third type of biomotor with revolution mechanism without rotation has been discovered. By analogy, rotation resembles the Earth rotating on its axis in a complete cycle every 24 hours, while revolution resembles the Earth revolving around the Sun one circle per 365 days (see animations http://nanobio.uky.edu/movie.html). The action of revolution that enables a motor free of coiling and torque has solved many puzzles and debates that have occurred throughout the history of viral DNA packaging motor studies. It also settles the discrepancies concerning the structure, stoichiometry, and functioning of DNA translocation motors. This review uses bacteriophages Phi29, HK97, SPP1, P22, T4, T7 as well as bacterial DNA translocase FtsK and SpoIIIE as examples to elucidate the puzzles. These motors use a ATPase, some of which have been confirmed to be a hexamer, to revolve around the dsDNA sequentially. ATP binding induces conformational change and possibly an entropy alteration in ATPase to a high affinity toward dsDNA; but ATP hydrolysis triggers another entropic and conformational change in ATPase to a low affinity for DNA, by which dsDNA is pushed toward an adjacent ATPase subunit. The rotation and revolution mechanisms can be distinguished by the size of channel: the channels of rotation motors are equal to or smaller than 2 nm, whereas channels of revolution motors are larger than 3 nm. Rotation motors use parallel threads to operate with a right-handed channel, while revolution motors use a left-handed channel to drive the right-handed DNA in an anti-parallel arrangement. Coordination of several vector factors in the same direction makes viral DNA-packaging motors unusually powerful and effective. Revolution mechanism avoids DNA coiling in translocating the lengthy

  19. Co-evolution of segregation guide DNA motifs and the FtsK translocase in bacteria: identification of the atypical Lactococcus lactis KOPS motif

    PubMed Central

    Nolivos, Sophie; Touzain, Fabrice; Pages, Carine; Coddeville, Michele; Rousseau, Philippe; El Karoui, Meriem; Le Bourgeois, Pascal; Cornet, François

    2012-01-01

    Bacteria use the global bipolarization of their chromosomes into replichores to control the dynamics and segregation of their genome during the cell cycle. This involves the control of protein activities by recognition of specific short DNA motifs whose orientation along the chromosome is highly skewed. The KOPS motifs act in chromosome segregation by orienting the activity of the FtsK DNA translocase towards the terminal replichore junction. KOPS motifs have been identified in γ-Proteobacteria and in Bacillus subtilis as closely related G-rich octamers. We have identified the KOPS motif of Lactococcus lactis, a model bacteria of the Streptococcaceae family harbouring a compact and low GC% genome. This motif, 5′-GAAGAAG-3, was predicted in silico using the occurrence and skew characteristics of known KOPS motifs. We show that it is specifically recognized by L. lactis FtsK in vitro and controls its activity in vivo. L. lactis KOPS is thus an A-rich heptamer motif. Our results show that KOPS-controlled chromosome segregation is conserved in Streptococcaceae but that KOPS may show important variation in sequence and length between bacterial families. This suggests that FtsK adapts to its host genome by selecting motifs with convenient occurrence frequencies and orientation skews to orient its activity. PMID:22373923

  20. Similarities between the DNA replication initiators of Gram-negative bacteria plasmids (RepA) and eukaryotes (Orc4p)/archaea (Cdc6p).

    PubMed

    Giraldo, R; Diaz-Orejas, R

    2001-04-24

    The proteins responsible for the initiation of DNA replication are thought to be essentially unrelated in bacteria and archaea/eukaryotes. Here we show that RepA, the initiator from the Pseudomonas plasmid pPS10, and the C-terminal domain of ScOrc4p, a subunit of Saccharomyces cerevisiae (Sc) origin recognition complex (ORC), share sequence similarities. Based on biochemical and spectroscopic evidence, these similarities include common structural elements, such as a winged-helix domain and a leucine-zipper dimerization motif. We have also found that ScOrc4p, as previously described for RepA-type initiators, interacts with chaperones of the Hsp70 family both in vitro and in vivo, most probably to regulate the assembly of active ORC. In evolutionary terms, our results are compatible with the recruitment of the same protein module for initiation of DNA replication by the ancestors of present-day Gram-negative bacteria plasmids, archaea, and eukaryotes. PMID:11296251

  1. 40 CFR 798.5500 - Differential growth inhibition of repair proficient and repair deficient bacteria: “Bacterial DNA...

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... increase of turbidity of the paired cultures. (3) Number of cultures. When using a plate diffusion.... Bacteria should be exposed to the test substance both in the presence and absence of an appropriate... substance for tests including a metabolic activation system should be selected on the basis of......

  2. USE OF COMPETITIVE DNA HYBRIDIZATION TO IDENTIFY DIFFERENCES IN THE GENOMES OF TWO CLOSELY RELATED FECAL INDICATOR BACTERIA

    EPA Science Inventory

    Although recent technological advances in DNA sequencing and computational biology now allow scientists to compare entire microbial genomes, comparisons of closely related bacterial species and individual isolates by whole-genome sequencing approaches remains prohibitively expens...

  3. Seasonal Succession Leads to Habitat-Dependent Differentiation in Ribosomal RNA:DNA Ratios among Freshwater Lake Bacteria

    PubMed Central

    Denef, Vincent J.; Fujimoto, Masanori; Berry, Michelle A.; Schmidt, Marian L.

    2016-01-01

    Relative abundance profiles of bacterial populations measured by sequencing DNA or RNA of marker genes can widely differ. These differences, made apparent when calculating ribosomal RNA:DNA ratios, have been interpreted as variable activities of bacterial populations. However, inconsistent correlations between ribosomal RNA:DNA ratios and metabolic activity or growth rates have led to a more conservative interpretation of this metric as the cellular protein synthesis potential (PSP). Little is known, particularly in freshwater systems, about how PSP varies for specific taxa across temporal and spatial environmental gradients and how conserved PSP is across bacterial phylogeny. Here, we generated 16S rRNA gene sequencing data using simultaneously extracted DNA and RNA from fractionated (free-living and particulate) water samples taken seasonally along a eutrophic freshwater estuary to oligotrophic pelagic transect in Lake Michigan. In contrast to previous reports, we observed frequent clustering of DNA and RNA data from the same sample. Analysis of the overlap in taxa detected at the RNA and DNA level indicated that microbial dormancy may be more common in the estuary, the particulate fraction, and during the stratified period. Across spatiotemporal gradients, PSP was often conserved at the phylum and class levels. PSPs for specific taxa were more similar across habitats in spring than in summer and fall. This was most notable for PSPs of the same taxa when located in the free-living or particulate fractions, but also when contrasting surface to deep, and estuary to Lake Michigan communities. Our results show that community composition assessed by RNA and DNA measurements are more similar than previously assumed in freshwater systems. However, the similarity between RNA and DNA measurements and taxa-specific PSPs that drive community-level similarities are conditional on spatiotemporal factors. PMID:27199936

  4. Seasonal succession leads to habitat-dependent differentiation in ribosomal RNA:DNA ratios among freshwater lake bacteria

    DOE PAGES

    Denef, Vincent J.; Fujimoto, Masanori; Berry, Michelle A.; Schmidt, Marian L.

    2016-04-29

    Relative abundance profiles of bacterial populations measured by sequencing DNA or RNA of marker genes can widely differ. These differences, made apparent when calculating ribosomal RNA:DNA ratios, have been interpreted as variable activities of bacterial populations. However, inconsistent correlations between ribosomal RNA:DNA ratios and metabolic activity or growth rates have led to a more conservative interpretation of this metric as the cellular protein synthesis potential (PSP). Little is known, particularly in freshwater systems, about how PSP varies for specific taxa across temporal and spatial environmental gradients and how conserved PSP is across bacterial phylogeny. Here, we generated 16S rRNA genemore » sequencing data using simultaneously extracted DNA and RNA from fractionated (free-living and particulate) water samples taken seasonally along a eutrophic freshwater estuary to oligotrophic pelagic transect in Lake Michigan. In contrast to previous reports, we observed frequent clustering of DNA and RNA data from the same sample. Analysis of the overlap in taxa detected at the RNA and DNA level indicated that microbial dormancy may be more common in the estuary, the particulate fraction, and during the stratified period. Across spatiotemporal gradients, PSP was often conserved at the phylum and class levels. PSPs for specific taxa were more similar across habitats in spring than in summer and fall. This was most notable for PSPs of the same taxa when located in the free-living or particulate fractions, but also when contrasting surface to deep, and estuary to Lake Michigan communities. Our results show that community composition assessed by RNA and DNA measurements are more similar than previously assumed in freshwater systems. Furthermore, the similarity between RNA and DNA measurements and taxa-specific PSPs that drive community-level similarities are conditional on spatiotemporal factors.« less

  5. Seasonal Succession Leads to Habitat-Dependent Differentiation in Ribosomal RNA:DNA Ratios among Freshwater Lake Bacteria.

    PubMed

    Denef, Vincent J; Fujimoto, Masanori; Berry, Michelle A; Schmidt, Marian L

    2016-01-01

    Relative abundance profiles of bacterial populations measured by sequencing DNA or RNA of marker genes can widely differ. These differences, made apparent when calculating ribosomal RNA:DNA ratios, have been interpreted as variable activities of bacterial populations. However, inconsistent correlations between ribosomal RNA:DNA ratios and metabolic activity or growth rates have led to a more conservative interpretation of this metric as the cellular protein synthesis potential (PSP). Little is known, particularly in freshwater systems, about how PSP varies for specific taxa across temporal and spatial environmental gradients and how conserved PSP is across bacterial phylogeny. Here, we generated 16S rRNA gene sequencing data using simultaneously extracted DNA and RNA from fractionated (free-living and particulate) water samples taken seasonally along a eutrophic freshwater estuary to oligotrophic pelagic transect in Lake Michigan. In contrast to previous reports, we observed frequent clustering of DNA and RNA data from the same sample. Analysis of the overlap in taxa detected at the RNA and DNA level indicated that microbial dormancy may be more common in the estuary, the particulate fraction, and during the stratified period. Across spatiotemporal gradients, PSP was often conserved at the phylum and class levels. PSPs for specific taxa were more similar across habitats in spring than in summer and fall. This was most notable for PSPs of the same taxa when located in the free-living or particulate fractions, but also when contrasting surface to deep, and estuary to Lake Michigan communities. Our results show that community composition assessed by RNA and DNA measurements are more similar than previously assumed in freshwater systems. However, the similarity between RNA and DNA measurements and taxa-specific PSPs that drive community-level similarities are conditional on spatiotemporal factors. PMID:27199936

  6. Mechanisms of recombination and function of DNA in bacteria: Final report, April 1969 through November 30, 1985

    SciTech Connect

    Guild, W.R.

    1987-09-01

    A brief overview of the authors research from 1969 to 1985 is presented. In the course of the work, we have developed a description of how the extracellular DNA binds to and enters the cell and how the cutting events on the surface and the stripping down to only one strand affects the biological end result. We also used these insights to develop a picture of the fate of the DNA after entry, both for transforming the chromosome or for creating a new replicon from either phage DNA or from plasmid DNA. In the course of doing this, we had first to develop a usable phage system, show transfection with its DNA, and find a novel ''pseudotransduction'' phenomenon. Later work, stimulated by the appearance of drug resistance in clinical isolates, led to the study of plasmids in pneumococcus and to showing that they transform the cells by processes similar to those used by phage DNA in transfection. These are two versions of the same process - creating a new replicon in the cell - and a full understanding of it has been helpful for others. A comparable understanding of the process in other species - Hemophilus in particular, remains to be established. 37 refs.

  7. A novel universal DNA labeling and amplification system for rapid microarray-based detection of 117 antibiotic resistance genes in Gram-positive bacteria.

    PubMed

    Strauss, Christian; Endimiani, Andrea; Perreten, Vincent

    2015-01-01

    A rapid and simple DNA labeling system has been developed for disposable microarrays and has been validated for the detection of 117 antibiotic resistance genes abundant in Gram-positive bacteria. The DNA was fragmented and amplified using phi-29 polymerase and random primers with linkers. Labeling and further amplification were then performed by classic PCR amplification using biotinylated primers specific for the linkers. The microarray developed by Perreten et al. (Perreten, V., Vorlet-Fawer, L., Slickers, P., Ehricht, R., Kuhnert, P., Frey, J., 2005. Microarray-based detection of 90 antibiotic resistance genes of gram-positive bacteria. J.Clin.Microbiol. 43, 2291-2302.) was improved by additional oligonucleotides. A total of 244 oligonucleotides (26 to 37 nucleotide length and with similar melting temperatures) were spotted on the microarray, including genes conferring resistance to clinically important antibiotic classes like β-lactams, macrolides, aminoglycosides, glycopeptides and tetracyclines. Each antibiotic resistance gene is represented by at least 2 oligonucleotides designed from consensus sequences of gene families. The specificity of the oligonucleotides and the quality of the amplification and labeling were verified by analysis of a collection of 65 strains belonging to 24 species. Association between genotype and phenotype was verified for 6 antibiotics using 77 Staphylococcus strains belonging to different species and revealed 95% test specificity and a 93% predictive value of a positive test. The DNA labeling and amplification is independent of the species and of the target genes and could be used for different types of microarrays. This system has also the advantage to detect several genes within one bacterium at once, like in Staphylococcus aureus strain BM3318, in which up to 15 genes were detected. This new microarray-based detection system offers a large potential for applications in clinical diagnostic, basic research, food safety and

  8. Design and Performance Testing of a DNA Extraction Assay for Sensitive and Reliable Quantification of Acetic Acid Bacteria Directly in Red Wine Using Real Time PCR

    PubMed Central

    Longin, Cédric; Guilloux-Benatier, Michèle; Alexandre, Hervé

    2016-01-01

    Although strategies exist to prevent AAB contamination, the increased interest for wines with low sulfite addition leads to greater AAB spoilage. Hence, there is a real need for a rapid, specific, sensitive, and reliable method for detecting these spoilage bacteria. All these requirements are met by real time Polymerase Chain Reaction (or quantitative PCR; qPCR). Here, we compare existing methods of isolating DNA and their adaptation to a red wine matrix. Two different protocols for isolating DNA and three PCR mix compositions were tested to select the best method. The addition of insoluble polyvinylpolypyrrolidone (PVPP) at 1% (v/v) during DNA extraction using a protocol succeeded in eliminating PCR inhibitors from red wine. We developed a bacterial internal control which was efficient in avoiding false negative results due to decreases in the efficiency of DNA isolation and/or amplification. The specificity, linearity, repeatability, and reproducibility of the method were evaluated. A standard curve was established for the enumeration of AAB inoculated into red wines. The limit of quantification in red wine was 3.7 log AAB/mL and about 2.8 log AAB/mL when the volume of the samples was increased from 1 to 10 mL. Thus, the DNA extraction method developed in this paper allows sensitive and reliable AAB quantification without underestimation thanks to the presence of an internal control. Moreover, monitoring of both the AAB population and the amount of acetic acid in ethanol medium and red wine highlighted that a minimum about 6.0 log cells/mL of AAB is needed to significantly increase the production of acetic acid leading to spoilage. PMID:27313572

  9. DNA.

    ERIC Educational Resources Information Center

    Felsenfeld, Gary

    1985-01-01

    Structural form, bonding scheme, and chromatin structure of and gene-modification experiments with deoxyribonucleic acid (DNA) are described. Indicates that DNA's double helix is variable and also flexible as it interacts with regulatory and other molecules to transfer hereditary messages. (DH)

  10. Design and testing of a functional group-specific DNA probe for the study of natural populations of acetogenic bacteria.

    PubMed Central

    Lovell, C R; Hui, Y

    1991-01-01

    The acetogens, although phylogenetically diverse, can be characterized by their possession of the acetyl coenzyme A (acetyl-CoA) pathway for autotrophic CO2 fixation. The gene encoding formyltetrahydrofolate synthetase, a key enzyme of the acetyl-CoA pathway, was previously cloned from the thermophilic acetogen Clostridium thermoaceticum and has now been tested as a group-specific probe for acetogens. Stable hybrids were formed between the probe and single DNA fragments from eight known acetogens representing six genera. A hybrid was also formed between the probe and a DNA fragment from one sulfate reducer known to be capable of both autotrophic CO2 fixation and acetate catabolism. No such hybrid was formed between the probe and DNA from a homoacetate fermenter not known to use the acetyl-CoA pathway, with two known formyltetrahydrofolate synthetase-producing purine fermenters, or with DNA from 27 other species representing 16 genera of organisms that do not use the acetyl-CoA pathway. DNA purified from cells extracted from horse manure was also screened with the acetogen probe. Six hybrids, indicating at least six detectable acetogen "strains," were observed. Images PMID:1768134

  11. Identification and characterization of single-stranded DNA-binding protein from the facultative psychrophilic bacteria Pseudoalteromonas haloplanktis.

    PubMed

    Olszewski, Marcin; Nowak, Marta; Cyranka-Czaja, Anna; Kur, Józef

    2014-01-01

    Single-stranded DNA-binding protein (SSB) plays an important role in DNA metabolism such as DNA replication, repair, and recombination, and is essential for cell survival. This study reports on the ssb-like gene cloning, gene expression and characterization of a single-stranded DNA-binding protein of Pseudoalteromonas haloplanktis (PhaSSB) and is the first report of such a protein from psychrophilic microorganism. PhaSSB possesses a high sequence similarity to Escherichia coli SSB (48% identity and 57% similarity) and has the longest amino acid sequence (244 amino acid residues) of all the known bacterial SSBs with one OB-fold per monomer. An analysis of purified PhaSSB by means of chemical cross-linking experiments, sedimentation analysis and size exclusion chromatography revealed a stable tetramer in solution. Using EMSA, we characterized the stoichiometry of PhaSSB complexed with a series of ssDNA homopolymers, and the size of the binding site was determined as being approximately 35 nucleotides long. In fluorescence titrations, the occluded site size of PhaSSB on poly(dT) is 34 nucleotides per tetramer under low-salt conditions (2mM NaCl), but increases to 54-64 nucleotides at higher-salt conditions (100-300mM NaCl). This suggests that PhaSSB undergoes a transition between ssDNA binding modes, which is observed for EcoSSB. The binding properties of PhaSSB investigated using SPR technology revealed that the affinity of PhaSSB to ssDNA is typical of SSB proteins. The only difference in the binding mode of PhaSSB to ssDNA is a faster association phase, when compared to EcoSSB, though compensated by faster dissociation rate. When analyzed by differential scanning calorimetry (DSC), the melting temperature (Tm) was determined as 63 °C, which is only a few degrees lower than for EcoSSB.

  12. The Structure and Stability of the Monomorphic HLA-G Are Influenced by the Nature of the Bound Peptide

    SciTech Connect

    Walpole, Nicholas G.; Kjer-Nielsen, Lars; Kostenko, Lyudmila; McCluskey, James; Brooks, Andrew G.; Rossjohn, Jamie; Clements, Craig S.

    2010-03-26

    The highly polymorphic major histocompatibility complex class Ia (MHC-Ia) molecules present a broad array of peptides to the clonotypically diverse {alpha}{beta} T-cell receptors. In contrast, MHC-Ib molecules exhibit limited polymorphism and bind a more restricted peptide repertoire, in keeping with their major role in innate immunity. Nevertheless, some MHC-Ib molecules do play a role in adaptive immunity. While human leukocyte antigen E (HLA-E), the MHC-Ib molecule, binds a very restricted repertoire of peptides, the peptide binding preferences of HLA-G, the class Ib molecule, are less stringent, although the basis by which HLA-G can bind various peptides is unclear. To investigate how HLA-G can accommodate different peptides, we compared the structure of HLA-G bound to three naturally abundant self-peptides (RIIPRHLQL, KGPPAALTL and KLPQAFYIL) and their thermal stabilities. The conformation of HLA-G{sup KGPPAALTL} was very similar to that of the HLA-G{sup RIIPRHLQL} structure. However, the structure of HLA-G{sup KLPQAFYIL} not only differed in the conformation of the bound peptide but also caused a small shift in the {alpha}2 helix of HLA-G. Furthermore, the relative stability of HLA-G was observed to be dependent on the nature of the bound peptide. These peptide-dependent effects on the substructure of the monomorphic HLA-G are likely to impact on its recognition by receptors of both innate and adaptive immune systems.

  13. The costs of risky male behaviour: sex differences in seasonal survival in a small sexually monomorphic primate

    PubMed Central

    Kraus, Cornelia; Eberle, Manfred; Kappeler, Peter M

    2008-01-01

    Male excess mortality is widespread among mammals and frequently interpreted as a cost of sexually selected traits that enhance male reproductive success. Sex differences in the propensity to engage in risky behaviours are often invoked to explain the sex gap in survival. Here, we aim to isolate and quantify the survival consequences of two potentially risky male behavioural strategies in a small sexually monomorphic primate, the grey mouse lemur Microcebus murinus: (i) most females hibernate during a large part of the austral winter, whereas most males remain active and (ii) during the brief annual mating season males roam widely in search of receptive females. Using a 10-year capture–mark–recapture dataset from a population of M. murinus in Kirindy Forest, western Madagascar, we statistically modelled sex-specific seasonal survival probabilities. Surprisingly, we did not find any evidence for direct survival benefits of hibernation—winter survival did not differ between males and females. By contrast, during the breeding season males survived less well than females (sex gap: 16%). Consistent with the ‘risky male behaviour’ hypothesis, the period for lowered male survival was restricted to the short mating season. Thus, sex differences in survival in a promiscuous mammal can be substantial even in the absence of sexual dimorphism. PMID:18426751

  14. Small and ugly? Phylogenetic analyses of the "selfing syndrome" reveal complex evolutionary fates of monomorphic primrose flowers.

    PubMed

    de Vos, Jurriaan M; Wüest, Rafael O; Conti, Elena

    2014-04-01

    One of the most common trends in plant evolution, loss of self-incompatibility and ensuing increases in selfing, is generally assumed to be associated with a suite of phenotypic changes, notably a reduction of floral size, termed the selfing syndrome. We investigate whether floral morphological traits indeed decrease in a deterministic fashion after losses of self-incompatibility, as traditionally expected, using a phylogeny of 124 primrose species containing nine independent transitions from heterostyly (heteromorphic incompatibility) to homostyly (monomorphic self-compatibility), a classic system for evolution of selfing. We find similar overall variability of homostylous and heterostylous species, except for diminished herkogamy in homostyles. Bayesian mixed models demonstrate differences between homostylous and heterostylous species in all traits, but net effects across species are small (except herkogamy) and directionality differs among traits. Strongly drift-like evolutionary trajectories of corolla tube length and corolla diameter inferred by Ornstein-Uhlenbeck models contrast with expected deterministic trajectories toward small floral size. Lineage-specific population genetic effects associated with evolution of selfing may explain that reductions of floral size represent one of several possible outcomes of floral evolution after loss of heterostyly in primroses. Contrary to the traditional paradigm, selfing syndromes may, but do not necessarily evolve in response to increased selfing.

  15. Using a DNA Microarray To Investigate the Distribution of Insect Virulence Factors in Strains of Photorhabdus Bacteria

    PubMed Central

    Marokhazi, Judit; Waterfield, Nicholas; LeGoff, Gaelle; Feil, Edward; Stabler, Richard; Hinds, Jason; Fodor, Andras; ffrench-Constant, Richard H.

    2003-01-01

    Photorhabdus is an insect-pathogenic bacterium in which oral toxicity to insects is found in two distinct taxonomic groups. Using a DNA microarray and comparative genomics, we show that oral toxicity is associated with toxin complex genes tcaABC and that this locus can be mobilized or deleted within different strains.   PMID:12867479

  16. Behavior of variable V3 region from 16S rDNA of lactic acid bacteria in denaturing gradient gel electrophoresis.

    PubMed

    Ercolini, D; Moschetti, G; Blaiotta, G; Coppola, S

    2001-03-01

    Separation of amplified V3 region from 16S rDNA by denaturing gradient gel electrophoresis (DGGE) was tested as a tool for differentiation of lactic acid bacteria commonly isolated from food. Variable V3 regions of 21 reference strains and 34 wild strains referred to species belonging to the genera Pediococcus, Enterococcus, Lactococcus, Lactobacillus, Leuconostoc, Weissella, and Streptococcus were analyzed. DGGE profiles obtained were species-specific for most of the cultures tested. Moreover, it was possible to group the remaining LAB reference strains according to the migration of their 16S V3 region in the denaturing gel. The results are discussed with reference to their potential in the analysis of LAB communities in food, besides shedding light on taxonomic aspects.

  17. Molecular characterization by amplified ribosomal DNA restriction analysis and antimicrobial potential of endophytic fungi isolated from Luehea divaricata (Malvaceae) against plant pathogenic fungi and pathogenic bacteria.

    PubMed

    Bernardi-Wenzel, J; Garcia, A; Azevedo, J L; Pamphile, J A

    2013-01-01

    Luehea divaricata is an important plant in popular medicine; it is used for its depurative, anti-inflammatory, and other therapeutic activities. We evaluated the antimicrobial activity of endophytic fungi isolated from leaves of L. divaricata against phytopathogens and pathogenic bacteria, and characterized the isolates based on amplified ribosomal DNA restriction analysis (ARDRA). The in vitro antagonistic activity of these endophytes against the phytopathogen Alternaria alternata was assayed by dual culture technique. Based on this evaluation of antimicrobial activity, we extracted secondary metabolites from nine endophytic fungi by partitioning in ethyl acetate and methanol. These were tested against the phytopathogens A. alternata, Colletotrichum sp and Moniliophthora perniciosa, and against the human pathogenic bacteria Escherichia coli and Staphylococcus aureus. Molecular characterization by ARDRA technique was used for phylogenetic analysis, based on comparison with sequences in GenBank. The endophytes had varied effects on A. alternata. One isolate produced an inhibition halo against M. perniciosa and against E. coli. This antibiosis activity indicates a role in the protection of the plant against microbial pathogens in nature, with potential for pharmaceutical and agricultural applications. Based on ARDRA, the 13 isolates were grouped. We found three different haplotypes of Phomopsis sp, showing interspecific variability. It appears that examination of the microbial community associated with medicinal plants of tropical regions has potential as a useful strategy to look for species with biotechnological applications. PMID:24301768

  18. Culturable bacteria present in the fluid of the hooded-pitcher plant Sarracenia minor based on 16S rDNA gene sequence data.

    PubMed

    Siragusa, Alex J; Swenson, Janice E; Casamatta, Dale A

    2007-08-01

    The culturable microbial community within the pitcher fluid of 93 Sarracenia minor carnivorous plants was examined over a 2-year study. Many aspects of the plant/bacterial/insect interaction within the pitcher fluid are minimally understood because the bacterial taxa present in these pitchers have not been identified. Thirteen isolates were characterized by 16S rDNA sequencing and subsequent phylogenetic analysis. The Proteobacteria were the most abundant taxa and included representatives from Serratia, Achromobacter, and Pantoea. The Actinobacteria Micrococcus was also abundant while Bacillus, Lactococcus, Chryseobacterium, and Rhodococcus were infrequently encountered. Several isolates conformed to species identifiers (>98% rDNA gene sequence similarity) including Serratia marcescens (isolates found in 27.5% of pitchers), Achromobacter xylosoxidans (37.6%), Micrococcus luteus (40.9%), Bacillus cereus (isolates found in 10.2%), Bacillus thuringiensis (5.4%), Lactococcus lactis (17.2%), and Rhodococcus equi (2.2%). Species-area curves suggest that sampling efforts were sufficient to recover a representative culturable bacterial community. The bacteria present represent a diverse community probably as a result of introduction by insect vectors, but the ecological significance remains under explored. PMID:17380356

  19. In situ probing of gram-positive bacteria with high DNA G + C content using 23S rRNA-targeted oligonucleotides.

    PubMed

    Roller, C; Wagner, M; Amann, R; Ludwig, W; Schleifer, K H

    1994-10-01

    23S-rRNA-targeted oligonucleotide probes were designed for the phylogenetic group 'Gram-positive bacteria with high G + C content of DNA' (GPBHGC). A sequence idiosyncrasy in two adjacent base pairs in the stem of helix 69 in domain IV of the 23S rRNA is present in all hitherto analysed strains of GPBHGC. An oligonucleotide probe targeted to this region hybridized only with strains of GPBHGC and was successfully used for in situ monitoring of these cells in activated sludge. Another unique feature of the 23S rRNA molecules of GPBHGC is a large insertion in domain III. Fluorescent oligonucleotides targeted to the highly variable regions of the rRNA within the insertions of Corynebacterium glutamicum DSM 20300, Aureobacterium testaceum DSM 20166 and Brevibacterium sp. DSM 20165 hybridized specifically to their target strains, whereas probing with oligonucleotides complementary to the rRNA-coding strand of the 23S rDNA and to the spacer between 16S and 23S rRNA of C. glutamicum did not result in detectable fluorescence. This confirmed that the large 23S insertions are indeed present in 23S rRNAs of GPBHGC and provide potential target sites for highly specific nucleic acid probes.

  20. Culturable bacteria present in the fluid of the hooded-pitcher plant Sarracenia minor based on 16S rDNA gene sequence data.

    PubMed

    Siragusa, Alex J; Swenson, Janice E; Casamatta, Dale A

    2007-08-01

    The culturable microbial community within the pitcher fluid of 93 Sarracenia minor carnivorous plants was examined over a 2-year study. Many aspects of the plant/bacterial/insect interaction within the pitcher fluid are minimally understood because the bacterial taxa present in these pitchers have not been identified. Thirteen isolates were characterized by 16S rDNA sequencing and subsequent phylogenetic analysis. The Proteobacteria were the most abundant taxa and included representatives from Serratia, Achromobacter, and Pantoea. The Actinobacteria Micrococcus was also abundant while Bacillus, Lactococcus, Chryseobacterium, and Rhodococcus were infrequently encountered. Several isolates conformed to species identifiers (>98% rDNA gene sequence similarity) including Serratia marcescens (isolates found in 27.5% of pitchers), Achromobacter xylosoxidans (37.6%), Micrococcus luteus (40.9%), Bacillus cereus (isolates found in 10.2%), Bacillus thuringiensis (5.4%), Lactococcus lactis (17.2%), and Rhodococcus equi (2.2%). Species-area curves suggest that sampling efforts were sufficient to recover a representative culturable bacterial community. The bacteria present represent a diverse community probably as a result of introduction by insect vectors, but the ecological significance remains under explored.

  1. Mechanisms of recombination and function of DNA in bacteria. Progress report, January 15, 1983-January 17, 1984

    SciTech Connect

    Guild, W.R.

    1984-01-01

    Studies on gene transfer in pneumococcus are described. Specifically transformation by chromosomal and plasmid DNAs, transfection by phage and plasmid DNA, and the novel kind of conjugative transfer of drug resistance elements among the chromosomes of streptococci are examined. Some of these elements appear to be transposons. There are numerous questions about their structure, functions, and relation to plasmids and other better characterized genetic elements. The experiments involve extensive use of transformation, cloning in both pneumococcal and E. coli systems, and analysis of DNAs by gel electrophoresis. For restriction mapping, we have concentrated on the insertion carrying genes for resistance to chloramphenicol and tetracycline.

  2. A Monomorphic Haplotype of Chromosome Ia Is Associated with Widespread Success in Clonal and Nonclonal Populations of Toxoplasma gondii

    PubMed Central

    Khan, Asis; Miller, Natalie; Roos, David S.; Dubey, J. P.; Ajzenberg, Daniel; Dardé, Marie Laure; Ajioka, James W.; Rosenthal, Benjamin; Sibley, L. David

    2011-01-01

    ABSTRACT Toxoplasma gondii is a common parasite of animals that also causes a zoonotic infection in humans. Previous studies have revealed a strongly clonal population structure that is shared between North America and Europe, while South American strains show greater genetic diversity and evidence of sexual recombination. The common inheritance of a monomorphic version of chromosome Ia (referred to as ChrIa*) among three clonal lineages from North America and Europe suggests that inheritance of this chromosome might underlie their recent clonal expansion. To further examine the diversity and distribution of ChrIa, we have analyzed additional strains with greater geographic diversity. Our findings reveal that the same haplotype of ChrIa* is found in the clonal lineages from North America and Europe and in older lineages in South America, where sexual recombination is more common. Although lineages from all three continents harbor the same conserved ChrIa* haplotype, strains from North America and Europe are genetically separate from those in South America, and these respective geographic regions show limited evidence of recent mixing. Genome-wide, array-based profiling of polymorphisms provided evidence for an ancestral flow from particular older southern lineages that gave rise to the clonal lineages now dominant in the north. Collectively, these data indicate that ChrIa* is widespread among nonclonal strains in South America and has more recently been associated with clonal expansion of specific lineages in North America and Europe. These findings have significant implications for the spread of genetic loci influencing transmission and virulence in pathogen populations. PMID:22068979

  3. DNA

    ERIC Educational Resources Information Center

    Stent, Gunther S.

    1970-01-01

    This history for molecular genetics and its explanation of DNA begins with an analysis of the Golden Jubilee essay papers, 1955. The paper ends stating that the higher nervous system is the one major frontier of biological inquiry which still offers some romance of research. (Author/VW)

  4. High-throughput DNA sequencing of the ruminal bacteria from moose (Alces alces) in Vermont, Alaska, and Norway.

    PubMed

    Ishaq, Suzanne L; Wright, André-Denis

    2014-08-01

    In the present study, the rumen bacteria of moose (Alces alces) from three distinct geographic locations were investigated. Moose are large, browsing ruminants in the deer family, which subsist on fibrous, woody browse, and aquatic plants. Subspecies exist which are distinguished by differing body and antler size, and these are somewhat geographically isolated. Seventeen rumen samples were collected from moose in Vermont, Alaska, and Norway, and bacterial 16S ribosomal RNA genes were sequenced using Roche 454 pyrosequencing with titanium chemistry. Overall, 109,643 sequences were generated from the 17 individual samples, revealing 33,622 unique sequences. Members of the phylum Bacteroidetes were dominant in samples from Alaska and Norway, but representatives of the phylum Firmicutes were dominant in samples from Vermont. Within the phylum Bacteroidetes, Prevotellaceae was the dominant family in all three sample locations, most of which belonged to the genus Prevotella. Within the phylum Firmicutes, the family Lachnospiraceae was the most prevalent in all three sample locations. The data set supporting the results of this article is available in the Sequence Read Archive (SRA), available through NCBI [study accession number SRP022590]. Samples clustered by geographic location and by weight and were heterogenous based on gender, location, and weight class (p < 0.05). Location was a stronger factor in determining the core microbiome than either age or weight, but gender did not appear to be a strong factor. There were no shared operational taxonomic units across all 17 samples, which indicates that these moose may have been isolated long enough to preclude a core microbiome among moose. Other potential factors discussed include differences in climate, food quality and availability, gender, and life cycle.

  5. Protective role of probiotic lactic acid bacteria against dietary fumonisin B1-induced toxicity and DNA-fragmentation in sprague-dawley rats.

    PubMed

    Khalil, Ashraf A; Abou-Gabal, Ashgan E; Abdellatef, Amira A; Khalid, Ahmed E

    2015-08-18

    The genus Fusarium, especially F. verticillioides and F. proliferatum, has been found in several agricultural products worldwide, especially in maize. Regardless the occurrence of symptoms, the presence of Fusarium in maize constitutes an imminent risk due to its ability to produce fumonisins, mycotoxins with proven carcinogenic effect on rats, swine, and equines and already classified as possible carcinogens to humans. The toxicity of incremental levels of fumonisin B1 (FB1), that is, 50, 100, and 200 mg FB1/kg diet, and the role of Lactobacillus delbrueckii subsp. lactis DSM 20076 (LL) and Pediococcus acidilactici NNRL B-5627 (PA) supplementation in counteracting the FB1 effects in intoxicated rats were monitored over a period of 4 weeks. Effects on the feed intake and body weight gain were noticed. A significant (p ≤ 0.05) increase in the level of liver and kidney functions markers and DNA fragmentation was also noticed in rat groups T100 and T200. The lactic acid bacteria (LAB) supplementation could bring back the normal serum biochemical parameters in rats fed on fumonisin B1-contaminated diets (T50 and T100) compared to FB1-treated groups. In rats of high-dosage dietary groups supplemented with LAB (T200-LL and T200-PA), the supplementation reduced the serum activity levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), and creatinine by 11.3, 11.9, 32, and 20%, respectively. DNA fragmentations were observed in the rat group treated with 200 mg FB1 after 3 weeks, while fragmentation was noticed in treated groups with 100 and 200 mg FB1 after 4 weeks. No DNA fragmentation was apparent in FB1-treated rats co-administered the LL or PA strain. These results suggest that in male rats consuming diets containing FB1, there is a time- and dose-dependent increase in serum enzyme activities and DNA lesions. Moreover, Lb. delbrueckii subsp. lactis (LL) and P. acidilactici (PA) strains have a protective effect

  6. In situ DNA hybridized chain reaction (FISH-HCR) as a better method for quantification of bacteria and archaea within marine sediment

    NASA Astrophysics Data System (ADS)

    Buongiorno, J.; Lloyd, K. G.; Shumaker, A.; Schippers, A.; Webster, G.; Weightman, A.; Turner, S.

    2015-12-01

    Nearly 75% of the Earth's surface is covered by marine sediment that is home to an estimated 2.9 x 1029 microbial cells. A substantial impediment to understanding the abundance and distribution of cells within marine sediment is the lack of a consistent and reliable method for their taxon-specific quantification. Catalyzed reporter fluorescent in situ hybridization (CARD-FISH) provides taxon-specific enumeration, but this process requires passing a large enzyme through cell membranes, decreasing its precision relative to general cell counts using a small DNA stain. In 2015, Yamaguchi et al. developed FISH hybridization chain reaction (FISH-HCR) as an in situ whole cell detection method for environmental microorganisms. FISH-HCR amplifies the fluorescent signal, as does CARD-FISH, but it allows for milder cell permeation methods that might prevent yield loss. To compare FISH-HCR to CARD-FISH, we examined bacteria and archaea cell counts within two sediment cores, Lille Belt (~78 meters deep) and Landsort Deep (90 meters deep), which were retrieved from the Baltic Sea Basin during IODP Expedition 347. Preliminary analysis shows that CARD-FISH counts are below the quantification limit for most depths across both cores. By contrast, quantification of cells was possible with FISH-HCR in all examined depths. When quantification with CARD-FISH was above the limit of detection, counts with FISH-HCR were up to 11 fold higher for Bacteria and 3 fold higher for Archaea from the same sediment sample. Further, FISH-HCR counts follow the trends of on board counts nicely, indicating that FISH-HCR may better reflect the cellular abundance within marine sediment than other quantification methods, including qPCR. Using FISH-HCR, we found that archaeal cell counts were on average greater than bacterial cell counts, but within the same order of magnitude.

  7. Geographic Separation of Domestic and Wild Strains of Toxoplasma gondii in French Guiana Correlates with a Monomorphic Version of Chromosome1a

    PubMed Central

    Khan, Asis; Ajzenberg, Daniel; Mercier, Aurélien; Demar, Magalie; Simon, Stéphane; Dardé, Marie Laure; Wang, Qiuling; Verma, Shiv Kumar; Rosenthal, Benjamin M.; Dubey, Jitender P.; Sibley, L. David

    2014-01-01

    Background Previous studies have stressed the genetic divergence and high pathogenicity of strains of T. gondii from French Guiana. Although strains from coastal, human adapted environments (so called anthropized) resemble those found in other regions of the Caribbean, strains collected from inland jungle environment are genetically quite diverse. To better understand the composition of these distinct strain types, we undertook a more in depth analysis of T. gondii strains from French Guiana including profiling of chromosome 1a (Chr1a), which is often shared as a single monomorphic haplotype among lineages that are otherwise genetically distinct. Methodology/Principal Findings Comparison of intron sequences from selectively neutral genes indicated that anthropized strains were most closely related to clonal type III strains from North America, although wider RFLP analysis revealed that they are natural hybrids. In contrast, strains isolated from the jungle were genetically very diverse. Remarkably, nearly all anthropized strains contained the monomorphic version of Chr1a while wild stains were extremely divergent. The presence of the monomorphic Chr1a strongly correlated with greater transmission in domestic cats, although there were several exceptions, indicating that other factors also contribute. Anthropized strains also varied in their virulence in laboratory mice, and this pattern could not be explained by the simple combination of previously identified virulence factors, indicating that other genetic determinants influence pathogenicity. Conclusions/Significance Our studies underscore the marked genetic separation of anthropized and wild strains of T. gondii in French Guiana and provide additional evidence that the presence of Chr1a is associated with successful expansion of widely different lineages within diverse geographic areas. The predominance of Chr1a among strains in the anthropized environment suggests that it may confer an advantage for transmission

  8. Sensitive Visual Detection of AHPND Bacteria Using Loop-Mediated Isothermal Amplification Combined with DNA-Functionalized Gold Nanoparticles as Probes.

    PubMed

    Arunrut, Narong; Kampeera, Jantana; Sirithammajak, Sarawut; Sanguanrut, Piyachat; Proespraiwong, Porranee; Suebsing, Rungkarn; Kiatpathomchai, Wansika

    2016-01-01

    Acute hepatopancreatic necrosis disease (AHPND) is a component cause of early mortality syndrome (EMS) of shrimp. In 2013, the causative agent was found to be unique isolates of Vibrio parahaemolyticus (VPAHPND) that contained a 69 kbp plasmid (pAP1) carrying binary Pir-like toxin genes PirvpA and PirvpB. In Thailand, AHPND was first recognized in 2012, prior to knowledge of the causative agent, and it subsequently led to a precipitous drop in shrimp production. After VPAHPND was characterized, a major focus of the AHPND control strategy was to monitor broodstock shrimp and post larvae for freedom from VPAHPND by nucleic acid amplification methods, most of which required use of expensive and sophisticated equipment not readily available in a shrimp farm setting. Here, we describe a simpler but equally sensitive approach for detection of VPAHPND based on loop-mediated isothermal amplification (LAMP) combined with unaided visual reading of positive amplification products using a DNA-functionalized, ssDNA-labled nanogold probe (AuNP). The target for the special set of six LAMP primers used was the VPAHPND PirvpA gene. The LAMP reaction was carried out at 65°C for 45 min followed by addition of the red AuNP solution and further incubation at 65°C for 5 min, allowing any PirvpA gene amplicons present to hybridize with the probe. Hybridization protected the AuNP against aggregation, so that the solution color remained red upon subsequent salt addition (positive test result) while unprotected AuNP aggregated and underwent a color change from red to blue and eventually precipitated (negative result). The total assay time was approximately 50 min. The detection limit (100 CFU) was comparable to that of other commonly-used methods for nested PCR detection of VPAHPND and 100-times more sensitive than 1-step PCR detection methods (104 CFU) that used amplicon detection by electrophoresis or spectrophotometry. There was no cross reaction with DNA templates derived from non

  9. Sensitive Visual Detection of AHPND Bacteria Using Loop-Mediated Isothermal Amplification Combined with DNA-Functionalized Gold Nanoparticles as Probes

    PubMed Central

    Arunrut, Narong; Kampeera, Jantana; Sirithammajak, Sarawut; Sanguanrut, Piyachat; Proespraiwong, Porranee; Suebsing, Rungkarn; Kiatpathomchai, Wansika

    2016-01-01

    Acute hepatopancreatic necrosis disease (AHPND) is a component cause of early mortality syndrome (EMS) of shrimp. In 2013, the causative agent was found to be unique isolates of Vibrio parahaemolyticus (VPAHPND) that contained a 69 kbp plasmid (pAP1) carrying binary Pir-like toxin genes PirvpA and PirvpB. In Thailand, AHPND was first recognized in 2012, prior to knowledge of the causative agent, and it subsequently led to a precipitous drop in shrimp production. After VPAHPND was characterized, a major focus of the AHPND control strategy was to monitor broodstock shrimp and post larvae for freedom from VPAHPND by nucleic acid amplification methods, most of which required use of expensive and sophisticated equipment not readily available in a shrimp farm setting. Here, we describe a simpler but equally sensitive approach for detection of VPAHPND based on loop-mediated isothermal amplification (LAMP) combined with unaided visual reading of positive amplification products using a DNA-functionalized, ssDNA-labled nanogold probe (AuNP). The target for the special set of six LAMP primers used was the VPAHPND PirvpA gene. The LAMP reaction was carried out at 65°C for 45 min followed by addition of the red AuNP solution and further incubation at 65°C for 5 min, allowing any PirvpA gene amplicons present to hybridize with the probe. Hybridization protected the AuNP against aggregation, so that the solution color remained red upon subsequent salt addition (positive test result) while unprotected AuNP aggregated and underwent a color change from red to blue and eventually precipitated (negative result). The total assay time was approximately 50 min. The detection limit (100 CFU) was comparable to that of other commonly-used methods for nested PCR detection of VPAHPND and 100-times more sensitive than 1-step PCR detection methods (104 CFU) that used amplicon detection by electrophoresis or spectrophotometry. There was no cross reaction with DNA templates derived from non

  10. Sensitive Visual Detection of AHPND Bacteria Using Loop-Mediated Isothermal Amplification Combined with DNA-Functionalized Gold Nanoparticles as Probes.

    PubMed

    Arunrut, Narong; Kampeera, Jantana; Sirithammajak, Sarawut; Sanguanrut, Piyachat; Proespraiwong, Porranee; Suebsing, Rungkarn; Kiatpathomchai, Wansika

    2016-01-01

    Acute hepatopancreatic necrosis disease (AHPND) is a component cause of early mortality syndrome (EMS) of shrimp. In 2013, the causative agent was found to be unique isolates of Vibrio parahaemolyticus (VPAHPND) that contained a 69 kbp plasmid (pAP1) carrying binary Pir-like toxin genes PirvpA and PirvpB. In Thailand, AHPND was first recognized in 2012, prior to knowledge of the causative agent, and it subsequently led to a precipitous drop in shrimp production. After VPAHPND was characterized, a major focus of the AHPND control strategy was to monitor broodstock shrimp and post larvae for freedom from VPAHPND by nucleic acid amplification methods, most of which required use of expensive and sophisticated equipment not readily available in a shrimp farm setting. Here, we describe a simpler but equally sensitive approach for detection of VPAHPND based on loop-mediated isothermal amplification (LAMP) combined with unaided visual reading of positive amplification products using a DNA-functionalized, ssDNA-labled nanogold probe (AuNP). The target for the special set of six LAMP primers used was the VPAHPND PirvpA gene. The LAMP reaction was carried out at 65°C for 45 min followed by addition of the red AuNP solution and further incubation at 65°C for 5 min, allowing any PirvpA gene amplicons present to hybridize with the probe. Hybridization protected the AuNP against aggregation, so that the solution color remained red upon subsequent salt addition (positive test result) while unprotected AuNP aggregated and underwent a color change from red to blue and eventually precipitated (negative result). The total assay time was approximately 50 min. The detection limit (100 CFU) was comparable to that of other commonly-used methods for nested PCR detection of VPAHPND and 100-times more sensitive than 1-step PCR detection methods (104 CFU) that used amplicon detection by electrophoresis or spectrophotometry. There was no cross reaction with DNA templates derived from non

  11. Anaerobic bacteria

    MedlinePlus

    Anaerobic bacteria are bacteria that do not live or grow when oxygen is present. In humans, these ... Goldstein EJ. Diseases caused by non-spore forming anaerobic bacteria. In: Goldman L, Schafer AI, eds. Goldman's ...

  12. A comparison of four new time-domain techniques for discriminating monomorphic ventricular tachycardia from sinus rhythm using ventricular waveform morphology.

    PubMed

    Throne, R D; Jenkins, J M; DiCarlo, L A

    1991-06-01

    Electrical management of intractable tachycardia via implantable antitachycardia devices has become a major form of therapy. Newly advanced methods of ventricular tachycardia detection propose examination of changes in intraventricular electrogram morphology in addition to or in combination with earlier rate-based detection algorithms. Unfortunately, most of the proposed morphology analysis techniques have computational demands beyond the capabilities of present devices or may be adversely affected by amplitude and baseline fluctuations of the intraventricular electrogram. We have designed four new computationally efficient time-domain algorithms for distinguishing ventricular electrograms during monomorphic ventricular tachycardia (VT) from those during sinus rhythm using direct analysis of the ventricular electrogram morphology. All four techniques are independent of amplitude fluctuations and three of the four are independent of baseline changes. These new techniques were compared to correlation waveform analysis, a previously proposed method for distinction of VT from sinus rhythm. Evaluation of these four new algorithms was performed on data from 19 consecutive patients with 31 distinct monomorphic ventricular tachycardia morphologies. Three of the algorithms performed as well or better than correlation waveform analysis but with one-tenth to one-half the computational demands.

  13. Development of a real-time PCR method for the detection of fossil 16S rDNA fragments of phototrophic sulfur bacteria in the sediments of Lake Cadagno.

    PubMed

    Ravasi, D F; Peduzzi, S; Guidi, V; Peduzzi, R; Wirth, S B; Gilli, A; Tonolla, M

    2012-05-01

    Lake Cadagno is a crenogenic meromictic lake situated in the southern range of the Swiss Alps characterized by a compact chemocline that has been the object of many ecological studies. The population dynamics of phototrophic sulfur bacteria in the chemocline has been monitored since 1994 with molecular methods such as 16S rRNA gene clone library analysis. To reconstruct paleo-microbial community dynamics, we developed a quantitative real-time PCR methodology for specific detection of 16S rRNA gene sequences of purple and green sulfur bacteria populations from sediment samples. We detected fossil 16S rDNA of nine populations of phototrophic sulfur bacteria down to 9-m sediment depth, corresponding to about 9500 years of the lake's biogeological history. These results provide the first evidence for the presence of 16S rDNA of anoxygenic phototrophic bacteria in Holocene sediments of an alpine meromictic lake and indicate that the water column stratification and the bacterial plume were already present in Lake Cadagno thousands of years ago. The finding of Chlorobium clathratiforme remains in all the samples analyzed shows that this population, identified in the water column only in 2001, was already a part of the lake's biota in the past.

  14. DNase I and Proteinase K eliminate DNA from injured or dead bacteria but not from living bacteria in microbial reference systems and natural drinking water biofilms for subsequent molecular biology analyses.

    PubMed

    Villarreal, Jessica Varela; Jungfer, Christina; Obst, Ursula; Schwartz, Thomas

    2013-09-01

    Molecular techniques, such as polymerase chain reaction (PCR) and quantitative PCR (qPCR), are very sensitive, but may detect total DNA present in a sample, including extracellular DNA (eDNA) and DNA coming from live and dead cells. DNase I is an endonuclease that non-specifically cleaves single- and double-stranded DNA. This enzyme was tested in this study to analyze its capacity of digesting DNA coming from dead cells with damaged cell membranes, leaving DNA from living cells with intact cell membranes available for DNA-based methods. For this purpose, an optimized DNase I/Proteinase K (DNase/PK) protocol was developed. Intact Staphylococcus aureus cells, heat-killed Pseudomonas aeruginosa cells, free genomic DNA of Salmonella enterica, and a mixture of these targets were treated according to the developed DNase/PK protocol. In parallel, these samples were treated with propidium monoazide (PMA) as an already described assay for live-dead discrimination. Quantitative PCR and PCR-DGGE of the eubacterial 16S rDNA fragment were used to test the ability of the DNase/PK and PMA treatments to distinguish DNA coming from cells with intact cell membranes in the presence of DNA from dead cells and free genomic DNA. The methods were applied to three months old autochthonous drinking water biofilms from a pilot facility built at a German waterworks. Shifts in the DNA patterns observed after DGGE analysis demonstrated the applicability of DNase/PK as well as of the PMA treatment for natural biofilm investigation. However, the DNase/PK treatment demonstrated some practical advantages in comparison with the PMA treatment for live/dead discrimination of bacterial targets in drinking water systems.

  15. Detection of fecal bacteria and source tracking identifiers in environmental waters using rRNA-based RT-qPCR and rDNA-based qPCR assays.

    PubMed

    Pitkänen, Tarja; Ryu, Hodon; Elk, Michael; Hokajärvi, Anna-Maria; Siponen, Sallamaari; Vepsäläinen, Asko; Räsänen, Pia; Santo Domingo, Jorge W

    2013-01-01

    In this study, we evaluated the use of RT-qPCR assays targeting rRNA gene sequences for the detection of fecal bacteria in water samples. We challenged the RT-qPCR assays against RNA extracted from sewage effluent (n = 14), surface water (n = 30), and treated source water (n = 15) samples. Additionally, we applied the same assays using DNA as the qPCR template. The targeted fecal bacteria were present in most of the samples tested, although in several cases, the detection frequency increased when RNA was used as the template. For example, the majority of samples that tested positive for E. coli and Campylobacter spp. in surface waters, and for human-specific Bacteroidales, E. coli, and Enterococcus spp. in treated source waters were only detected when rRNA was used as the original template. The difference in detection frequency using rRNA or rDNA (rRNA gene) was sample- and assay-dependent, suggesting that the abundance of active and nonactive populations differed between samples. Statistical analyses for each population exhibiting multiple quantifiable results showed that the rRNA copy numbers were significantly higher than the rDNA counterparts (p < 0.05). Moreover, the detection frequency of rRNA-based assays were in better agreement with the culture-based results of E. coli, intestinal enterococci, and thermotolerant Campylobacter spp. in surface waters than that of rDNA-based assays, suggesting that rRNA signals were associated to active bacterial populations. Our data show that using rRNA-based approaches significantly increases detection sensitivity for common fecal bacteria in environmental waters. These findings have important implications for microbial water quality monitoring and public health risk assessments.

  16. Simple & Safe Genomic DNA Isolation.

    ERIC Educational Resources Information Center

    Moss, Robert; Solomon, Sondra

    1991-01-01

    A procedure for purifying DNA using either bacteria or rat liver is presented. Directions for doing a qualitative DNA assay using diphenylamine and a quantitative DNA assay using spectroscopy are included. (KR)

  17. EBV-negative monomorphic B-cell post-transplant lymphoproliferative disorders are pathologically distinct from EBV-positive cases and frequently contain TP53 mutations.

    PubMed

    Courville, Elizabeth L; Yohe, Sophia; Chou, David; Nardi, Valentina; Lazaryan, Aleksandr; Thakral, Beenu; Nelson, Andrew C; Ferry, Judith A; Sohani, Aliyah R

    2016-10-01

    Monomorphic post-transplant lymphoproliferative disorder commonly resembles diffuse large B-cell lymphoma or Burkitt lymphoma, and most are Epstein-Barr virus (EBV) positive. We retrospectively identified 32 cases of monomorphic post-transplant lymphoproliferative disorder from two institutions and evaluated EBV in situ hybridization; TP53 mutation status; p53, CD30, myc, and BCL2 expression by immunohistochemistry; proliferation index by Ki67; and germinal center vs non-germinal center immunophenotype by Hans criteria. Post-transplant lymphoproliferative disorder arose after hematopoietic stem cell transplant in five and solid organ transplant in 27 patients, a median of 4 and 96 months after transplant, respectively (overall median latency 71 months, range 2-295). The most common morphology was diffuse large B-cell lymphoma (28 cases), with three cases of Burkitt lymphoma, and one case of plasmablastic lymphoma. Ten cases (31%) were EBV negative. Of those with the morphology of diffuse large B-cell lymphoma, the EBV-negative cases were more frequently TP53-mutated (P<0.001), p53 positive by immunohistochemistry (P<0.001), CD30 negative (P<0.01), and of germinal center immunophenotype (P=0.01) compared with EBV-positive cases. No statistically significant difference in overall survival was identified based on EBV, TP53 mutation status, germinal center vs non-germinal center immunophenotype, or other immunohistochemical parameters evaluated. Patients who died of post-transplant lymphoproliferative disorder were older with a longer latency from time of transplant to diagnosis (P<0.05). Our study demonstrates that diffuse large B-cell lymphoma-related immunohistochemical prognostic markers have limited relevance in the post-transplant setting and underscores differences between EBV-positive and EBV-negative post-transplant lymphoproliferative disorder in terms of immunophenotype and TP53 mutation frequency, supporting an alternative pathogenesis for EBV-negative post

  18. Isolation and Identification of Concrete Environment Bacteria

    NASA Astrophysics Data System (ADS)

    Irwan, J. M.; Anneza, L. H.; Othman, N.; Husnul, T.; Alshalif, A. F.

    2016-07-01

    This paper presents the isolation and molecular method for bacteria identification through PCR and DNA sequencing. Identification of the bacteria species is required in order to fully utilize the bacterium capability for precipitation of calcium carbonate in concrete. This process is to enable the addition of suitable catalyst according to the bacterium enzymatic pathway that is known through the bacteria species used. The objective of this study is to isolate, enriched and identify the bacteria species. The bacteria in this study was isolated from fresh urine and acid mine drainage water, Kota Tinggi, Johor. Enrichment of the isolated bacteria was conducted to ensure the bacteria survivability in concrete. The identification of bacteria species was done through polymerase chain reaction (PCR) and rRDNA sequencing. The isolation and enrichment of the bacteria was done successfully. Whereas, the results for bacteria identification showed that the isolated bacteria strains are Bacillus sp and Enterococus faecalis.

  19. Analyzing arthropods for the presence of bacteria.

    PubMed

    Andrews, Elizabeth S

    2013-02-01

    Bacteria within arthropods can be identified using culture-independent methods. This unit describes protocols for surface sterilization of arthropods, DNA extraction of whole bodies and tissues, touchdown PCR amplification using 16S rDNA general bacteria primers, and profiling the bacterial community using denaturing gradient gel electrophoresis.

  20. Genetic and functional characterization of a yet-unclassified rhizobial Dtr (DNA-transfer-and-replication) region from a ubiquitous plasmid conjugal system present in Sinorhizobium meliloti, in Sinorhizobium medicae, and in other nonrhizobial Gram-negative bacteria.

    PubMed

    Giusti, María de los Ángeles; Pistorio, Mariano; Lozano, Mauricio J; Tejerizo, Gonzalo A Torres; Salas, María Eugenia; Martini, María Carla; López, José Luis; Draghi, Walter O; Del Papa, María Florencia; Pérez-Mendoza, Daniel; Sanjuán, Juan; Lagares, Antonio

    2012-05-01

    Rhizobia are Gram-negative bacteria that live in soils and associate with leguminous plants to establish nitrogen-fixing symbioses. The ability of these bacteria to undergo horizontal gene transfer (HGT) is thought to be one of the main features to explain both the origin of their symbiotic life-style and the plasticity and dynamics of their genomes. In our laboratory we have previously characterized at the species level the non-pSym plasmid mobilome in Sinorhizobium meliloti, the symbiont of Medicago spp., and have found a high incidence of conjugal activity in many plasmids (Pistorio et al., 2008). In this work we characterized the Dtr (DNA-transfer-and-replication) region of one of those plasmids, pSmeLPU88b. This mobilization region was found to represent a previously unclassified Dtr type in rhizobia (hereafter type-IV), highly ubiquitous in S. meliloti and found in other genera of Gram-negative bacteria as well; including Agrobacterium, Ochrobactrum, and Chelativorans. The oriT of the type-IV Dtr described here could be located by function within a DNA fragment of 278 bp, between the divergent genes parA and mobC. The phylogenetic analysis of the cognate relaxase MobZ indicated that this protein groups close to the previously defined MOB(P3) and MOB(P4) type of enzymes, but is located in a separate and novel cluster that we have designated MOB(P0). Noteworthy, MOB(P0) and MOB(P4) relaxases were frequently associated with plasmids present in rhizospheric soil bacteria. A comparison of the nod-gene locations with the phylogenetic topology of the rhizobial relaxases revealed that the symbiotic genes are found on diverse plasmids bearing any of the four Dtr types, thus indicating that pSym plasmids are not specifically associated with any particular mobilization system. Finally, we demonstrated that the type-IV Dtr promoted the mobilization of plasmids from S. meliloti to Sinorhizobium medicae as well as from these rhizobia to other bacteria by means of their own

  1. Cost-effective pooling of DNA from nasopharyngeal swab samples for large-scale detection of bacteria by real-time PCR.

    PubMed

    Edouard, Sophie; Prudent, Elsa; Gautret, Philippe; Memish, Ziad A; Raoult, Didier

    2015-03-01

    We investigated the potential of pooling DNA from nasopharyngeal specimens to reduce the cost of real-time PCR (RT-PCR) for bacterial detection. Lyophilization is required to reconcentrate DNA. This strategy yields a high specificity (86%) and a high sensitivity (96%). We estimate that compared to individual testing, 37% fewer RT-PCR tests are needed.

  2. The flexibility of UV-inducible mutation in Deinococcus ficus as evidenced by the existence of the imuB-dnaE2 gene cassette and generation of superior feather degrading bacteria.

    PubMed

    Zeng, You-Hong; Shen, Fo-Ting; Tan, Chen-Chung; Huang, Chieh-Chen; Young, Chiu-Chung

    2011-12-20

    The lexA-imuB-dnaE2 gene cassette contributing to the TLS (translesion synthesis) polymerase activity and can easily cause mutation after DNA damage in many bacteria. But it was previously thought that TLS polymerase activity was unlikely to exist in the radio-resistant genus Deinococcus. In our preliminary studies, the lexA-imuB-dnaE2 gene cassette was found in a newly isolated feather-degrading Deinococcus ficus. Here we have attempted to determine the imuB gene sequence from another Deinococcus species namely D. grandis, by using the newly designed primers. The destroying of either imuB or dnaE2 gene in D. ficus leads to the increase in UV sensitivity and decrease in UV-induced mutations, which demonstrated the existence of TLS polymerase activity in D. ficus. In the presence of lexA-imuB-dnaE2, it is possible to obtain mutants with various keratinolytic activities after UV exposure. The keratinolytic activity of mutant strain CC-ZG207 increased by approximately twofold during growth in liquid feather medium. In contrast, the mutant strain CC-ZG227 showed only half of the keratinolytic activity compared with the wild type strain. By utilizing SDS-PAGE and zymogram profile analysis, the change in the protease activity was observed. We have proposed that the superior mutants of D. ficus can be created under UV stress, which is mediated by the lexA-imuB-dnaE2 gene cassette.

  3. Magnetic Bacteria.

    ERIC Educational Resources Information Center

    Nelson, Jane Bray; Nelson, Jim

    1992-01-01

    Describes the history of Richard Blakemore's discovery of magnetotaxic organisms. Discusses possible reasons why the magnetic response in bacteria developed. Proposes research experiments integrating biology and physics in which students investigate problems using cultures of magnetotaxic organisms. (MDH)

  4. Leading ladies: leadership of group movements in a pair-living, co-dominant, monomorphic primate across reproductive stages and fruit availability seasons.

    PubMed

    Tecot, Stacey R; Romine, Natalie K

    2012-07-01

    For gregarious species, individuals must maintain cohesion while minimizing the costs of coordinated travel. Leaders of group movements potentially influence energy expenditure, energy intake, and predation risk for individuals in the group, which can have important fitness consequences. Models of pair-living species predict that energetic asymmetries lead to an emergent leader, with those in greater need leading. We investigated sex differences in leadership in pairs of red-bellied lemurs, Eulemur rubriventer, a monomorphic species with bisexual dispersal and no discernible hierarchy, to determine whether higher energetic requirements by adult females lead to female leadership. We collected leadership data in Ranomafana National Park, Madagascar on six groups of habituated E. rubriventer for 13 consecutive months between 2004-2005. To determine whether females led group movements more than males, we examined the difference in leadership frequencies of progressions in adult males and adult females within each group (n = 1,346 progressions). We further investigated the behavioral context (i.e. travel followed by feeding or not) and seasonal contexts (fruit availability, reproduction) of leadership. Group leadership was distributed, with different individuals leading the group at different times. However, females led significantly more than males, a pattern which was consistent in both feeding and non-feeding contexts and throughout all fruiting seasons and reproductive stages. While disparities in energetic status among the sexes may impact leadership in this species, leadership did not differ with changes in food availability or reproductive stage, and thus we were unable to determine whether female leadership might be related to changes in energetic status. Females may have higher energetic needs than males at all times, not merely seasonally, or female leadership may be unrelated to immediate energetic need. Rather, female leadership may be a legacy of female

  5. Basal cell (monomorphic) and minimally pleomorphic adenomas of the salivary glands. Distinction from the solid (anaplastic) type of adenoid cystic carcinoma in fine-needle aspiration.

    PubMed

    Stanley, M W; Horwitz, C A; Rollins, S D; Powers, C N; Bardales, R H; Korourain, S; Stern, S J

    1996-07-01

    Cytologic features of the cell-stroma interface are useful in distinguishing between monomorphic adenomas of the basal cell type and adenoid cystic carcinoma. In basal cell adenomas, the collagenous stroma interdigitates with adjacent cells, whereas in adenoid cystic carcinoma, the two are separated by a sharp smooth border. Furthermore, the stroma of basal cell adenomas can contain rare spindle cells or capillaries, but the cylinders of adenoid cystic carcinoma are acellular. The authors review their experience with five cases of basal cell adenoma, and three cases that were designated "minimally pleomorphic adenomas." The latter group showed the small blue cell pattern of basal cell adenoma at the time of fine-needle aspiration, and histology revealed only small foci of typical pleomorphic adenoma. With the exception of one cystic case, the cell-stroma interface of basal cell adenoma was observed in all eight cases. These cases are contrasted with three adenoid cystic carcinomas with extensive solid (anaplastic) areas. All showed the small blue cell pattern and cell-stroma interface features of basal cell adenoma. Neither showed the smooth-bordered cylinders of adenoid cystic carcinoma. Two of these three were incorrectly interpreted as benign at the time of fine-needle aspiration. The authors suggest that the stroma aspirated from solid adenoid cystic carcinoma represents desmoplastic tumor stroma that mimics the pattern of basal cell adenoma in smear material. Distinction between basal cell adenoma and the solid type of adenoid cystic carcinoma at the time of fine-needle aspiration remains a very difficult problem.

  6. Studies on DNA binding behaviour of biologically active transition metal complexes of new tetradentate N2O2 donor Schiff bases: Inhibitory activity against bacteria

    NASA Astrophysics Data System (ADS)

    Sobha, S.; Mahalakshmi, R.; Raman, N.

    A series of Cu(II), Ni(II) and Zn(II) complexes of the type ML have been synthesized with Schiff bases derived from o-acetoacetotoluidide, 2-hydroxybenzaldehyde and o-phenylenediamine/1,4-diaminobutane. The complexes are insoluble in common organic solvents but soluble in DMF and DMSO. The measured molar conductance values in DMSO indicate that the complexes are non-electrolytic in nature. All the six metal complexes have been fully characterized with the help of elemental analyses, molecular weights, molar conductance values, magnetic moments and spectroscopic data. The analytical data helped to elucidate the structure of the metal complexes. The Schiff bases are found to act as tetradentate ligands using N2O2 donor set of atoms leading to a square-planar geometry for the complexes around all the metal ions. The binding properties of metal complexes with DNA were investigated by absorption spectra, viscosity measurements and cyclic voltammetry. Detailed analysis reveals that the metal complexes intercalate into the DNA base stack as intercalators. All the metal complexes cleave the pUC19 DNA in presence of H2O2. The Schiff bases and their complexes have been screened for their antibacterial activity against five bacterial strains (Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli, Staphylococcus epidermidis, Klebsiella pneumoniae) by disk diffusion method. All the metal complexes have potent biocidal activity than the free ligands.

  7. Methanotrophic bacteria.

    PubMed Central

    Hanson, R S; Hanson, T E

    1996-01-01

    Methane-utilizing bacteria (methanotrophs) are a diverse group of gram-negative bacteria that are related to other members of the Proteobacteria. These bacteria are classified into three groups based on the pathways used for assimilation of formaldehyde, the major source of cell carbon, and other physiological and morphological features. The type I and type X methanotrophs are found within the gamma subdivision of the Proteobacteria and employ the ribulose monophosphate pathway for formaldehyde assimilation, whereas type II methanotrophs, which employ the serine pathway for formaldehyde assimilation, form a coherent cluster within the beta subdivision of the Proteobacteria. Methanotrophic bacteria are ubiquitous. The growth of type II bacteria appears to be favored in environments that contain relatively high levels of methane, low levels of dissolved oxygen, and limiting concentrations of combined nitrogen and/or copper. Type I methanotrophs appear to be dominant in environments in which methane is limiting and combined nitrogen and copper levels are relatively high. These bacteria serve as biofilters for the oxidation of methane produced in anaerobic environments, and when oxygen is present in soils, atmospheric methane is oxidized. Their activities in nature are greatly influenced by agricultural practices and other human activities. Recent evidence indicates that naturally occurring, uncultured methanotrophs represent new genera. Methanotrophs that are capable of oxidizing methane at atmospheric levels exhibit methane oxidation kinetics different from those of methanotrophs available in pure cultures. A limited number of methanotrophs have the genetic capacity to synthesize a soluble methane monooxygenase which catalyzes the rapid oxidation of environmental pollutants including trichloroethylene. PMID:8801441

  8. Studies of the repair of O/sup 6/-alkylguanine and O/sup 4/-alkylthymine in DNA by alkyltransferases from mammalian cells and bacteria

    SciTech Connect

    Pegg, A.E.; Dolan, M.E.; Acchitano, D.; Morimoto, K.

    1985-10-01

    O/sup 6/-Methylguanine in DNA is repaired by the action of a protein termed O/sup 6/-alkylguanine-DNA alkyltransferase (AT) which transfers the methyl group to a cysteine residue in its own sequence. Since the cysteine which is methylated is not regenerated rapidly, if at all, the capacity for repair of O/sup 6/-methylguanine is limited by the number of molecules of the AT available within the cell. The level and inducibility of the AT differed greatly in different mammalian cell types and species with the highest levels in human tissues and in liver and the lowest levels in brain. In E. coli such exposure increased the activity more than 100-fold. The protein isolated from E. coli removed methyl groups much more rapidly than the larger alkyl groups but the mammalian AT isolated from rat liver showed much less difference in rate with adducts of different size. Ethyl and n-propyl groups were removed by the rat liver AT only three to four times more slowly than methyl groups. Another important difference between the bacterial and mammalian ATs is that the bacterial protein was also able to remove methyl groups from the O/sup 4/-position of thymine in methylated DNA or poly(dT) but the AT from rat liver or human fibroblasts did not repair O/sup 4/-methylthymidine. These results indicate that the results obtained with the E. coli system may not be a suitable model for extrapolation to predictions of the effects of alkylating agents in initiating tumors or mutations in mammalian cells.

  9. Stress responses of bacteria.

    PubMed

    Marles-Wright, Jon; Lewis, Richard J

    2007-12-01

    Bacteria, irrespective of natural habitat, are exposed to constant fluctuations in their growth conditions. Consequently they have developed sophisticated responses, modulated by the re-modelling of protein complexes and by phosphorylation-dependent signal transduction systems, to adapt to and to survive a variety of insults. Ultimately these signalling systems affect transcriptional regulons either by activating an alternative sigma factor subunit of RNA polymerase, for example, sigma E (sigma(E)) of Escherichia coli and sigma B (sigma(B)) and sigma F (sigma(F)) in Bacillus subtilis or by activating DNA-binding two-component response regulators. Recent structure determinations, and systems biology analysis of key regulators in well-characterised stress-responsive pathways, illustrate conserved and novel mechanisms in these representative model bacteria.

  10. Evaluation of direct 16S rDNA sequencing as a metagenomics-based approach to screening bacteria in bottled water.

    PubMed

    Hansen, Trine; Skånseng, Beate; Hoorfar, Jeffrey; Löfström, Charlotta

    2013-09-01

    Deliberate or accidental contamination of food, feed, and water supplies poses a threat to human health worldwide. A rapid and sensitive detection technique that could replace the current labor-intensive and time-consuming culture-based methods is highly desirable. In addition to species-specific assays, such as PCR, there is a need for generic methods to screen for unknown pathogenic microorganisms in samples. This work presents a metagenomics-based direct-sequencing approach for detecting unknown microorganisms, using Bacillus cereus (as a model organism for B. anthracis) in bottled water as an example. Total DNA extraction and 16S rDNA gene sequencing were used in combination with principle component analysis and multicurve resolution to study detection level and possibility for identification. Results showed a detection level of 10(5) to 10(6) CFU/L. Using this method, it was possible to separate 2 B. cereus strains by the principal component plot, despite the close sequence resemblance. A linear correlation between the artificial contamination level and the relative amount of the Bacillus artificial contaminant in the metagenome was observed, and a relative amount value above 0.5 confirmed the presence of Bacillus. The analysis also revealed that background flora in the bottled water varied between the different water types that were included in the study. This method has the potential to be adapted to other biological matrices and bacterial pathogens for fast screening of unknown bacterial threats in outbreak situations.

  11. Evaluation of direct 16S rDNA sequencing as a metagenomics-based approach to screening bacteria in bottled water.

    PubMed

    Hansen, Trine; Skånseng, Beate; Hoorfar, Jeffrey; Löfström, Charlotta

    2013-09-01

    Deliberate or accidental contamination of food, feed, and water supplies poses a threat to human health worldwide. A rapid and sensitive detection technique that could replace the current labor-intensive and time-consuming culture-based methods is highly desirable. In addition to species-specific assays, such as PCR, there is a need for generic methods to screen for unknown pathogenic microorganisms in samples. This work presents a metagenomics-based direct-sequencing approach for detecting unknown microorganisms, using Bacillus cereus (as a model organism for B. anthracis) in bottled water as an example. Total DNA extraction and 16S rDNA gene sequencing were used in combination with principle component analysis and multicurve resolution to study detection level and possibility for identification. Results showed a detection level of 10(5) to 10(6) CFU/L. Using this method, it was possible to separate 2 B. cereus strains by the principal component plot, despite the close sequence resemblance. A linear correlation between the artificial contamination level and the relative amount of the Bacillus artificial contaminant in the metagenome was observed, and a relative amount value above 0.5 confirmed the presence of Bacillus. The analysis also revealed that background flora in the bottled water varied between the different water types that were included in the study. This method has the potential to be adapted to other biological matrices and bacterial pathogens for fast screening of unknown bacterial threats in outbreak situations. PMID:23971801

  12. Transformation of gram positive bacteria by sonoporation

    DOEpatents

    Yang, Yunfeng; Li, Yongchao

    2014-03-11

    The present invention provides a sonoporation-based method that can be universally applied for delivery of compounds into Gram positive bacteria. Gram positive bacteria which can be transformed by sonoporation include, for example, Bacillus, Streptococcus, Acetobacterium, and Clostridium. Compounds which can be delivered into Gram positive bacteria via sonoporation include nucleic acids (DNA or RNA), proteins, lipids, carbohydrates, viruses, small organic and inorganic molecules, and nano-particles.

  13. A Novel HURRAH Protocol Reveals High Numbers of Monomorphic MHC Class II Loci and Two Asymmetric Multi-Locus Haplotypes in the Père David's Deer

    PubMed Central

    Wan, Qiu-Hong; Zhang, Pei; Ni, Xiao-Wei; Wu, Hai-Long; Chen, Yi-Yan; Kuang, Ye-Ye; Ge, Yun-Fa; Fang, Sheng-Guo

    2011-01-01

    The Père David's deer is a highly inbred, but recovered, species, making it interesting to consider their adaptive molecular evolution from an immunological perspective. Prior to this study, genomic sequencing was the only method for isolating all functional MHC genes within a certain species. Here, we report a novel protocol for isolating MHC class II loci from a species, and its use to investigate the adaptive evolution of this endangered deer at the level of multi-locus haplotypes. This protocol was designated “HURRAH” based on its various steps and used to estimate the total number of MHC class II loci. We confirmed the validity of this novel protocol in the giant panda and then used it to examine the Père David's deer. Our results revealed that the Père David's deer possesses nine MHC class II loci and therefore has more functional MHC class II loci than the eight genome-sequenced mammals for which full MHC data are currently available. This could potentially account at least in part for the strong survival ability of this species in the face of severe bottlenecking. The results from the HURRAH protocol also revealed that: (1) All of the identified MHC class II loci were monomorphic at their antigen-binding regions, although DRA was dimorphic at its cytoplasmic tail; and (2) these genes constituted two asymmetric functional MHC class II multi-locus haplotypes: DRA1*01 ∼ DRB1 ∼ DRB3 ∼ DQA1 ∼ DQB2 (H1) and DRA1*02 ∼ DRB2 ∼ DRB4 ∼ DQA2 ∼ DQB1 (H2). The latter finding indicates that the current members of the deer species have lost the powerful ancestral MHC class II haplotypes of nine or more loci, and have instead fixed two relatively weak haplotypes containing five genes. As a result, the Père David's deer are currently at risk for increased susceptibility to infectious pathogens. PMID:21267075

  14. Identification of Nitrogen-Incorporating Bacteria in Petroleum-Contaminated Arctic Soils by Using [15N]DNA-Based Stable Isotope Probing and Pyrosequencing ▿ †

    PubMed Central

    Bell, Terrence H.; Yergeau, Etienne; Martineau, Christine; Juck, David; Whyte, Lyle G.; Greer, Charles W.

    2011-01-01

    Arctic soils are increasingly susceptible to petroleum hydrocarbon contamination, as exploration and exploitation of the Arctic increase. Bioremediation in these soils is challenging due to logistical constraints and because soil temperatures only rise above 0°C for ∼2 months each year. Nitrogen is often added to contaminated soil in situ to stimulate the existing microbial community, but little is known about how the added nutrients are used by these microorganisms. Microbes vary widely in their ability to metabolize petroleum hydrocarbons, so the question becomes: which hydrocarbon-degrading microorganisms most effectively use this added nitrogen for growth? Using [15N]DNA-based stable isotope probing, we determined which taxonomic groups most readily incorporated nitrogen from the monoammonium phosphate added to contaminated and uncontaminated soil in Canadian Forces Station-Alert, Nunavut, Canada. Fractions from each sample were amplified with bacterial 16S rRNA and alkane monooxygenase B (alkB) gene-specific primers and then sequenced using lage-scale parallel-pyrosequencing. Sequence data was combined with 16S rRNA and alkB gene C quantitative PCR data to measure the presence of various phylogenetic groups in fractions at different buoyant densities. Several families of Proteobacteria and Actinobacteria that are directly involved in petroleum degradation incorporated the added nitrogen in contaminated soils, but it was the DNA of Sphingomonadaceae that was most enriched in 15N. Bacterial growth in uncontaminated soils was not stimulated by nutrient amendment. Our results suggest that nitrogen uptake efficiency differs between bacterial groups in contaminated soils. A better understanding of how groups of hydrocarbon-degraders contribute to the catabolism of petroleum will facilitate the design of more targeted bioremediation treatments. PMID:21498745

  15. Identification of nitrogen-incorporating bacteria in petroleum-contaminated arctic soils by using [15N]DNA-based stable isotope probing and pyrosequencing.

    PubMed

    Bell, Terrence H; Yergeau, Etienne; Martineau, Christine; Juck, David; Whyte, Lyle G; Greer, Charles W

    2011-06-01

    Arctic soils are increasingly susceptible to petroleum hydrocarbon contamination, as exploration and exploitation of the Arctic increase. Bioremediation in these soils is challenging due to logistical constraints and because soil temperatures only rise above 0°C for ∼2 months each year. Nitrogen is often added to contaminated soil in situ to stimulate the existing microbial community, but little is known about how the added nutrients are used by these microorganisms. Microbes vary widely in their ability to metabolize petroleum hydrocarbons, so the question becomes: which hydrocarbon-degrading microorganisms most effectively use this added nitrogen for growth? Using [(15)N]DNA-based stable isotope probing, we determined which taxonomic groups most readily incorporated nitrogen from the monoammonium phosphate added to contaminated and uncontaminated soil in Canadian Forces Station-Alert, Nunavut, Canada. Fractions from each sample were amplified with bacterial 16S rRNA and alkane monooxygenase B (alkB) gene-specific primers and then sequenced using large-scale parallel-pyrosequencing. Sequence data was combined with 16S rRNA and alkB gene C quantitative PCR data to measure the presence of various phylogenetic groups in fractions at different buoyant densities. Several families of Proteobacteria and Actinobacteria that are directly involved in petroleum degradation incorporated the added nitrogen in contaminated soils, but it was the DNA of Sphingomonadaceae that was most enriched in (15)N. Bacterial growth in uncontaminated soils was not stimulated by nutrient amendment. Our results suggest that nitrogen uptake efficiency differs between bacterial groups in contaminated soils. A better understanding of how groups of hydrocarbon-degraders contribute to the catabolism of petroleum will facilitate the design of more targeted bioremediation treatments.

  16. Bacteria Counter

    NASA Technical Reports Server (NTRS)

    1981-01-01

    Science Applications, Inc.'s ATP Photometer makes a rapid and accurate count of the bacteria in a body fluid sample. Instrument provides information on the presence and quantity of bacteria by measuring the amount of light emitted by the reaction between two substances. Substances are ATP adenosine triphosphate and luciferase. The reactants are applied to a human body sample and the ATP Photometer observes the intensity of the light emitted displaying its findings in a numerical output. Total time lapse is usually less than 10 minutes, which represents a significant time savings in comparison of other techniques. Other applications are measuring organisms in fresh and ocean waters, determining bacterial contamination of foodstuffs, biological process control in the beverage industry, and in assay of activated sewage sludge.

  17. Identification of biomass utilizing bacteria in a carbon-depleted glacier forefield soil by the use of 13C DNA stable isotope probing.

    PubMed

    Zumsteg, Anita; Schmutz, Stefan; Frey, Beat

    2013-06-01

    As Alpine glaciers are retreating rapidly, bare soils with low organic C and N contents are becoming exposed. Carbon availability is a key factor regulating microbial diversity and ecosystem functioning in these soils. The aim of this study was to investigate how bacterial activity, community structure and composition are influenced by organic carbon availability. Bare soils were supplied with (13)C-labelled fungal (Penicillium sp.) and green algal (Chlorella sp.) biomass and the CO2 evolution and its δ(13)C signature were monitored up to 60 days. These organisms have previously been isolated near the glacier terminus. DNA stable isotope probing followed by T-RFLP profiling and sequencing of 16S rRNA genes was employed to identify consumers able to assimilate carbon from these biomass amendments. Higher respiration and higher bacterial activity indicated a more efficient utilization of algal cells than fungal cells. Flavobacterium sp. predominantly incorporated fungal-derived C, whereas the algal-derived C was mainly incorporated by Acidobacteria and Proteobacteria. This study emphasizes the important role of both fungal and algal biomass in increasing the carbon pool in recently deglaciated bare soils, as only 20% of the added C was respired as CO2, and the rest, we presume, remained in the soil.

  18. DNA Methylation

    PubMed Central

    Marinus, M.G.; Løbner-Olesen, A.

    2014-01-01

    The DNA of E. coli contains 19,120 6-methyladenines and 12,045 5-methylcytosines in addition to the four regular bases and these are formed by the postreplicative action of three DNA methyltransferases. The majority of the methylated bases are formed by the Dam and Dcm methyltransferases encoded by the dam (DNA adenine methyltransferase) and dcm (DNA cytosine methyltransferase) genes. Although not essential, Dam methylation is important for strand discrimination during repair of replication errors, controlling the frequency of initiation of chromosome replication at oriC, and regulation of transcription initiation at promoters containing GATC sequences. In contrast, there is no known function for Dcm methylation although Dcm recognition sites constitute sequence motifs for Very Short Patch repair of T/G base mismatches. In certain bacteria (e.g., Vibrio cholerae, Caulobacter crescentus) adenine methylation is essential and in C. crescentus, it is important for temporal gene expression which, in turn, is required for coordinating chromosome initiation, replication and division. In practical terms, Dam and Dcm methylation can inhibit restriction enzyme cleavage; decrease transformation frequency in certain bacteria; decrease the stability of short direct repeats; are necessary for site-directed mutagenesis; and to probe eukaryotic structure and function. PMID:26442938

  19. Deterioration to extinction of wastewater bacteria by non-thermal atmospheric pressure air plasma as assessed by 16S rDNA-DGGE fingerprinting

    PubMed Central

    El-Sayed, Wael S.; Ouf, Salama A.; Mohamed, Abdel-Aleam H.

    2015-01-01

    The use of cold plasma jets for inactivation of a variety of microorganisms has recently been evaluated via culture-based methods. Accordingly, elucidation of the role of cold plasma in decontamination would be inaccurate because most microbial populations within a system remain unexplored owing to the high amount of yet uncultured bacteria. The impact of cold atmospheric plasma on the bacterial community structure of wastewater from two different industries was investigated by metagenomic-based polymerase chain reaction-denaturing gradient gel electrophoresis (DGGE) utilizing 16S rRNA genes. Three doses of atmospheric pressure dielectric barrier discharge plasma were applied to wastewater samples on different time scales. DGGE revealed that the bacterial community gradually changed and overall abundance decreased to extinction upon plasma treatment. The bacterial community in food processing wastewater contained 11 key operational taxonomic units that remained almost completely unchanged when exposed to plasma irradiation at 75.5 mA for 30 or 60 s. However, when exposure time was extended to 90 s, only Escherichia coli, Coliforms, Aeromonas sp., Vibrio sp., and Pseudomonas putida survived. Only E. coli, Aeromonas sp., Vibrio sp., and P. putida survived treatment at 81.94 mA for 90 s. Conversely, all bacterial groups were completely eliminated by treatment at 85.34 mA for either 60 or 90 s. Dominant bacterial groups in leather processing wastewater also changed greatly upon exposure to plasma at 75.5 mA for 30 or 60 s, with Enterobacter aerogenes, Klebsiella sp., Pseudomonas stutzeri, and Acidithiobacillus ferrooxidans being sensitive to and eliminated from the community. At 90 s of exposure, all groups were affected except for Pseudomonas sp. and Citrobacter freundii. The same trend was observed for treatment at 81.94 mA. The variability in bacterial community response to different plasma treatment protocols revealed that plasma had a selective impact on bacterial

  20. Microsatellite DNA markers in Populus tremuloides.

    PubMed

    Rahman, M H; Dayanandan, S; Rajora, O P

    2000-04-01

    Markers for eight new microsatellite DNA or simple sequence repeat (SSR) loci were developed and characterized in trembling aspen (Populus tremuloides) from a partial genomic library. Informativeness of these microsatellite DNA markers was examined by determining polymorphisms in 38 P. tremuloides individuals. Inheritance of selected markers was tested in progenies of controlled crosses. Six characterized SSR loci were of dinucleotide repeats (two perfect and four imperfect), and one each of trinucleotide and tetranucleotide repeats. The monomorphic SSR locus (PTR15) was of a compound imperfect dinucleotide repeat. The primers of one highly polymorphic SSR locus (PTR7) amplified two loci, and alleles could not be assigned to a specific locus. At the other six polymorphic loci, 25 alleles were detected in 38 P. tremuloides individuals; the number of alleles ranged from 2 to 7, with an average of 4.2 alleles per locus, and the observed heterozygosity ranged from 0.05 to 0.61, with an average of 0.36 per locus. The two perfect dinucleotide and one trinucleotide microsatellite DNA loci were the most informative. Microsatellite DNA variants of four SSR loci characterized previously followed a single-locus Mendelian inheritance pattern, whereas those of PTR7 from the present study showed a two-locus Mendelian inheritance pattern in controlled crosses. The microsatellite DNA markers developed and reported here could be used for assisting various genetic, breeding, biotechnology, genome mapping, conservation, and sustainable forest management programs in poplars. PMID:10791817

  1. [Genetic resources of nodule bacteria].

    PubMed

    Rumiantseva, M L

    2009-09-01

    Nodule bacteria (rhizobia) form highly specific symbiosis with leguminous plants. The efficiency of accumulation of biological nitrogen depends on molecular-genetic interaction between the host plant and rhizobia. Genetic characteristics of microsymbiotic strains are crucial in developing highly productive and stress-resistant symbiotic pairs: rhizobium strain-host plant cultivar (species). The present review considers the issue of studying genetic resources of nodule bacteria to identify genes and their blocks, responsible for the ability of rhizobia to form highly effective symbiosis in various agroecological conditions. The main approaches to investigation of intraspecific and interspecific genetic and genomic diversity of nodule bacteria are considered, from MLEE analysis to the recent methods of genomic DNA analysis using biochips. The data are presented showing that gene centers of host plants are centers of genetic diversification of nodule bacteria, because the intraspecific polymorphism of genetic markers of the core and the accessory rhizobial genomes is extremely high in them. Genotypic features of trapped and nodule subpopulations of alfalfa nodule bacteria are discussed. A survey of literature showed that the genomes of natural strains in alfalfa gene centers exhibit significant differences in genes involved in control of metabolism, replication, recombination, and the formation of defense response (hsd genes). Natural populations of rhizobia are regarded as a huge gene pool serving as a source of evolutionary innovations.

  2. Magnetic bacteria against MIC

    SciTech Connect

    Javaherdashti, R.

    1997-12-01

    In this article, it is suggested to use the sensitivity of magnetotactic bacteria to changes of magnetic field direction and the natural ability of this bacteria in rapid growth during relatively short time intervals against corrosion-enhancing bacteria and especially sulfate-reducing bacteria. If colonies of sulfate-reducing bacteria could be packed among magnetotactic bacteria, then, by applying sufficiently powerful magnetic field (about 0.5 gauss), all of these bacteria (magnetic and non-magnetic) will be oriented towards an Anti-bacteria agent (oxygen or biocide). So, Microbiologically-Influenced Corrosion in the system would be controlled to a large extent.

  3. Back To Bacteria.

    ERIC Educational Resources Information Center

    Flannery, Maura C.

    1997-01-01

    Explores new research about bacteria. Discusses bacterial genomes, archaea, unusual environments, evolution, pathogens, bacterial movement, biofilms, bacteria in the body, and a bacterial obsession. Contains 29 references. (JRH)

  4. Recombinant DNA means and method

    SciTech Connect

    Alford, B.L.; Mao, J.I.; Moir, D.T.; Taunton-Rigby, A.; Vovis, G.F.

    1987-05-19

    This patent describes a transformed living cell selected from the group consisting of fungi, yeast and bacteria, and containing genetic material derived from recombinant DNA material and coding for bovine rennin.

  5. Functional Encyclopedia of Bacteria and Archaea

    SciTech Connect

    Blow, M. J.; Deutschbauer, A. M.; Hoover, C. A.; Lamson, J.; Lamson, J.; Price, M. N.; Waters, J.; Wetmore, K. M.; Bristow, J.; Arkin, A. P.

    2013-03-20

    Bacteria and Archaea exhibit a huge diversity of metabolic capabilities with fundamental importance in the environment, and potential applications in biotechnology. However, the genetic bases of these capabilities remain unclear due largely to an absence of technologies that link DNA sequence to molecular function. To address this challenge, we are developing a pipeline for high throughput annotation of gene function using mutagenesis, growth assays and DNA sequencing. By applying this pipeline to annotate gene function in 50 diverse microbes we hope to discover thousands of new gene functions and produce a proof of principle `Functional Encyclopedia of Bacteria and Archaea?.

  6. Comparison of mitochondrial DNA control region sequence and microsatellite DNA analyses in estimating population structure and gene flow rates in Atlantic sturgeon Acipenser oxyrinchus

    USGS Publications Warehouse

    Wirgin, I.; Waldman, J.; Stabile, J.; Lubinski, B.; King, T.

    2002-01-01

    Atlantic sturgeon Acipenser oxyrinchus is large, long-lived, and anadromous with subspecies distributed along the Atlantic (A. oxyrinchus oxyrinchus) and Gulf of Mexico (A. o. desotoi) coasts of North America. Although it is not certain if extirpation of some population units has occurred, because of anthropogenic influences abundances of all populations are low compared with historical levels. Informed management of A. oxyrinchus demands a detailed knowledge of its population structure, levels of genetic diversity, and likelihood to home to natal rivers. We compared the use of mitochondrial DNA (mtDNA) control region sequence and microsatellite nuclear DNA (nDNA) analyses in identifying the stock structure and homing fidelity of Atlantic and Gulf coast populations of A. oxyrinchus. The approaches were concordant in that they revealed moderate to high levels of genetic diversity and suggested that populations of Atlantic sturgeon are highly structured. At least six genetically distinct management units were detected using the two approaches among the rivers surveyed. Mitochondrial DNA sequences revealed a significant cline in haplotype diversity along the Atlantic coast with monomorphism observed in Canadian populations. High levels of nDNA diversity were also observed among populations along the Atlantic coast, including the two Canadian populations, probably resulting from the more rapid rate of mutational and evolutionary change at microsatellite loci. Estimates of gene flow among populations were similar between both approaches with the exception that because of mtDNA monomorphism in Canadian populations, gene flow estimates between them were unobtainable. Analyses of both genomes provided high resolution and confidence in characterizing the population structure of Atlantic sturgeon. Microsatellite analysis was particularly informative in delineating population structure in rivers that were recently glaciated and may prove diagnostic in rivers that are

  7. Mechanism of Transcriptional Bursting in Bacteria

    PubMed Central

    Chong, Shasha; Chen, Chongyi; Ge, Hao; Xie, X. Sunney

    2014-01-01

    SUMMARY Transcription of highly expressed genes has been shown to occur in stochastic bursts. But the origin of such ubiquitous phenomenon has not been understood. Here we present the mechanism in bacteria. We developed a high-throughput in vitro single-molecule assay to follow transcription on individual DNA templates in real time. We showed that positive supercoiling buildup on a DNA segment by transcription slows down transcription elongation and eventually stops transcription initiation. Transcription can be resumed upon gyrase binding to the DNA segment. Furthermore, using single-cell mRNA counting fluorescence in situ hybridization (FISH), we found the extent of transcriptional bursting depends on the intracellular gyrase concentration. Together, these findings prove that transcriptional bursting of highly expressed genes in bacteria is primarily caused by reversible gyrase dissociation from and rebinding to a DNA segment, changing the supercoiling level of the segment. PMID:25036631

  8. High efficiency recombineering in lactic acid bacteria

    PubMed Central

    van Pijkeren, Jan-Peter; Britton, Robert A.

    2012-01-01

    The ability to efficiently generate targeted point mutations in the chromosome without the need for antibiotics, or other means of selection, is a powerful strategy for genome engineering. Although oligonucleotide-mediated recombineering (ssDNA recombineering) has been utilized in Escherichia coli for over a decade, the successful adaptation of ssDNA recombineering to Gram-positive bacteria has not been reported. Here we describe the development and application of ssDNA recombineering in lactic acid bacteria. Mutations were incorporated in the chromosome of Lactobacillus reuteri and Lactococcus lactis without selection at frequencies ranging between 0.4% and 19%. Whole genome sequence analysis showed that ssDNA recombineering is specific and not hypermutagenic. To highlight the utility of ssDNA recombineering we reduced the intrinsic vancomymycin resistance of L. reuteri >100-fold. By creating a single amino acid change in the d-Ala-d-Ala ligase enzyme we reduced the minimum inhibitory concentration for vancomycin from >256 to 1.5 µg/ml, well below the clinically relevant minimum inhibitory concentration. Recombineering thus allows high efficiency mutagenesis in lactobacilli and lactococci, and may be used to further enhance beneficial properties and safety of strains used in medicine and industry. We expect that this work will serve as a blueprint for the adaptation of ssDNA recombineering to other Gram-positive bacteria. PMID:22328729

  9. Ejecting Phage DNA against Cellular Turgor Pressure

    PubMed Central

    Marion, Sanjin; Šiber, Antonio

    2014-01-01

    We examine in vivo ejection of noncondensed DNA from tailed bacteriophages into bacteria. The ejection is dominantly governed by the physical conditions in the bacteria. The confinement of the DNA in the virus capsid only slightly helps the ejection, becoming completely irrelevant during its last stages. A simple calculation based on the premise of condensed DNA in the cell enables us to estimate the maximal bacterial turgor pressure against which the ejection can still be fully realized. The calculated pressure (∼5 atm) shows that the ejection of DNA into Gram-negative bacteria could proceed spontaneously, i.e., without the need to invoke active mechanisms. PMID:25418173

  10. Ejecting phage DNA against cellular turgor pressure.

    PubMed

    Marion, Sanjin; Siber, Antonio

    2014-10-21

    We examine in vivo ejection of noncondensed DNA from tailed bacteriophages into bacteria. The ejection is dominantly governed by the physical conditions in the bacteria. The confinement of the DNA in the virus capsid only slightly helps the ejection, becoming completely irrelevant during its last stages. A simple calculation based on the premise of condensed DNA in the cell enables us to estimate the maximal bacterial turgor pressure against which the ejection can still be fully realized. The calculated pressure (~5 atm) shows that the ejection of DNA into Gram-negative bacteria could proceed spontaneously, i.e., without the need to invoke active mechanisms.

  11. Programmed Death in Bacteria

    PubMed Central

    Lewis, Kim

    2000-01-01

    Programmed cell death (PCD) in bacteria plays an important role in developmental processes, such as lysis of the mother cell during sporulation of Bacillus subtilis and lysis of vegetative cells in fruiting body formation of Myxococcus xanthus. The signal transduction pathway leading to autolysis of the mother cell includes the terminal sporulation sigma factor EςK, which induces the synthesis of autolysins CwlC and CwlH. An activator of autolysin in this and other PCD processes is yet to be identified. Autolysis plays a role in genetic exchange in Streptococcus pneumoniae, and the gene for the major autolysin, lytA, is located in the same operon with recA. DNA from lysed cells is picked up by their neighbors and recombined into the chromosome by RecA. LytA requires an unknown activator controlled by a sensory kinase, VncS. Deletion of vncS inhibits autolysis and also decreases killing by unrelated antibiotics. This observation suggests that PCD in bacteria serves to eliminate damaged cells, similar to apoptosis of defective cells in metazoa. The presence of genes affecting survival without changing growth sensitivity to antibiotics (vncS, lytA, hipAB, sulA, and mar) indicates that bacteria are able to control their fate. Elimination of defective cells could limit the spread of a viral infection and donate nutrients to healthy kin cells. An altruistic suicide would be challenged by the appearance of asocial mutants without PCD and by the possibility of maladaptive total suicide in response to a uniformly present lethal factor or nutrient depletion. It is proposed that a low rate of mutation serves to decrease the probability that asocial mutants without PCD will take over the population. It is suggested that PCD is disabled in persistors, rare cells that are resistant to killing, to ensure population survival. It is suggested that lack of nutrients leads to the stringent response that suppresses PCD, producing a state of tolerance to antibiotics, allowing cells to

  12. Susceptibility-weighted imaging and diffusion-weighted imaging findings in central nervous system monomorphic B cell post-transplant lymphoproliferative disorder before and after treatment and comparison with primary B cell central nervous system lymphoma.

    PubMed

    Ginat, Daniel Thomas; Purakal, Alixandra; Pytel, Peter

    2015-11-01

    The purpose of this article is to review the MRI features of monomorphic central nervous system post-transplant lymphoproliferative disorder (CNS PTLD), including diffusion-weighted and susceptibility-weighted sequences before and after treatment and to compare the imaging findings with those of primary central nervous system B cell lymphoma (PCNS BCL). Retrospective review of the brain MRI characteristics in patients with pathology proven monomorphic CNS PTLD and PCNS BCL was performed. In particular, the enhancement, diffusion-weighted, susceptibility-weighted MRI characteristics of the lesions were evaluated. In addition, the diffusion-weighted, susceptibility-weighted MRI features after treatment for CNS PTLD were evaluated. A total of 12 lesions in six patients with CNS PTLD and 12 lesions in nine patients with PCNS BCL were identified on MRI. Among the CNS PTLD lesions with post-contrast images, 80 % demonstrated peripheral enhancement. All of the CNS PTLD lesions contained foci of intratumoral susceptibility signal (ITSS) and the average mean ADC values and ratios were 0.892 × 10(-3) mm(2)/s (standard deviation: 0.082 × 10(-3) mm(2)/s) and 1.19 (standard deviation: 0.15), respectively. On the other hand, 75 % of the PCNS BCL displayed diffuse enhancement, two cases (16.7 %) contained ITSS, and the mean ADC values and ratios were 0.721 × 10(-3) mm(2)/s (standard deviation: 0.093 × 10(-3) mm(2)/s), and 0.99 (standard deviation: 0.17), respectively. Thus, the presence of heterogeneous lesions with ITSS that do not necessarily have as extensive restricted diffusion as PCNS BCL is suggestive of CNS PTLD in the appropriate clinical setting. The preliminary data in this series suggests that diffusion-weighted imaging may serve as a useful biomarker for monitoring treatment response, in which successful treatment of CNS PTLD may result in increased ADC values. In addition, foci of susceptibility effect in CNS PTLD tend to persist or increase over the course of

  13. Bacteria isolated from amoebae/bacteria consortium

    DOEpatents

    Tyndall, R.L.

    1995-05-30

    New protozoan derived microbial consortia and method for their isolation are provided. Consortia and bacteria isolated therefrom are useful for treating wastes such as trichloroethylene and trinitrotoluene. Consortia, bacteria isolated therefrom, and dispersants isolated therefrom are useful for dispersing hydrocarbons such as oil, creosote, wax, and grease.

  14. Bacteria isolated from amoebae/bacteria consortium

    DOEpatents

    Tyndall, Richard L.

    1995-01-01

    New protozoan derived microbial consortia and method for their isolation are provided. Consortia and bacteria isolated therefrom are useful for treating wastes such as trichloroethylene and trinitrotoluene. Consortia, bacteria isolated therefrom, and dispersants isolated therefrom are useful for dispersing hydrocarbons such as oil, creosote, wax, and grease.

  15. Metabolic engineering of bacteria.

    PubMed

    Kumar, Ravi R; Prasad, Satish

    2011-07-01

    Yield and productivity are critical for the economics and viability of a bioprocess. In metabolic engineering the main objective is the increase of a target metabolite production through genetic engineering. Metabolic engineering is the practice of optimizing genetic and regulatory processes within cells to increase the production of a certain substance. In the last years, the development of recombinant DNA technology and other related technologies has provided new tools for approaching yields improvement by means of genetic manipulation of biosynthetic pathway. Industrial microorganisms like Escherichia coli, Actinomycetes, etc. have been developed as biocatalysts to provide new or to optimize existing processes for the biotechnological production of chemicals from renewable plant biomass. The factors like oxygenation, temperature and pH have been traditionally controlled and optimized in industrial fermentation in order to enhance metabolite production. Metabolic engineering of bacteria shows a great scope in industrial application as well as such technique may also have good potential to solve certain metabolic disease and environmental problems in near future. PMID:22754024

  16. Detection of fecal bacteria and source tracking identifiers in environmental waters using rRNA-based RT-qPCR and rDNA-based qPCR assays

    EPA Science Inventory

    The identification of fecal pollution sources is commonly performed using DNA-based methods. However, there is evidence that DNA can be associated with dead cells or present as “naked DNA” in the environment. To this end, we compared the detection frequency of host specific marke...

  17. Intrarenal solitary fibrous tumor of the kidney report of a case with emphasis on the differential diagnosis in the wide spectrum of monomorphous spindle cell tumors of the kidney.

    PubMed

    Magro, Gaetano; Cavallaro, Vincenzo; Torrisi, Antonietta; Lopes, Maria; Dell'Albani, Marcello; Lanzafame, Salvatore

    2002-01-01

    Solitary fibrous tumor (SFT) is a neoplasm that can occur in the urogenital tract, and is also reported occurring in the spermatic cord, seminal vesicles, urinary bladder, prostate, and kidney. Furthermore, it is most important to consider its existence in the kidney, because it is usually diagnosed as renal cell carcinoma pre-operatively. To our knowledge, only 10 cases of SFT have been reported in the kidney to date. We report the clinico-pathological features of an intrarenal SFT occurring in a 31-year-old woman. The tumor, measuring 8.6 cm in its greatest diameter, completely replaced the cortex and the medulla of the middle region of the right kidney, compressing the pelvis. Radiological imaging was consistent with a renal cell carcinoma. Histologically, the tumor was composed of a proliferation of bland-looking vimentin+, CD34+, bcl2+ and CD99+ spindle cells exhibiting a haphazard to storiform growth pattern, pushing borders, and a low mitotic rate (2 mitoses x 10 HPF). We placed emphasis on the differential diagnostic problems, i.e., its differentiation from other primary monomorphous benign and malignant spindle cell tumors of the kidney, such as fibroma, benign fibrous histiocytoma, hemangiopericytoma, inflammatory myofibroblastic (pseudo-)tumor, leiomyoma, angiomyolipoma with predominant spindle cell smooth muscle component, benign peripheral nerve sheath tumors, renal mixed epithelial/stromal tumors, adult type mesoblastic nephroma, fibrous type monophasic synovial sarcoma, malignant peripheral nerve sheath tumors, fibrosarcoma, and low-grade fibromyxoid sarcoma.

  18. The use of dimorphic Alu insertions in human DNA fingerprinting

    SciTech Connect

    Novick, G.E.; Gonzalez, T.; Garrison, J.; Novick, C.C.; Herrera, R.J.; Batzer, M.A.; Deininger, P.L.

    1992-12-04

    We have characterized certain Human Specific Alu Insertions as either dimorphic (TPA25, PV92, APO), sightly dimorphic (C2N4 and C4N4) or monomorphic (C3N1, C4N6, C4N2, C4N5, C4N8), based on studies of Caucasian, Asian, American Black and African Black populations. Our approach is based upon: (1) PCR amplification using primers directed to the sequences that flank the site of insertion of the different Alu elements studied; (2) gel electrophoresis and scoring according to the presence or absence of an Alu insertion in one or both homologous chromosomes; (3) allelic frequencies calculated and compared according to Hardy-Weinberg equilibrium. Our DNA fingerprinting procedure using PCR amplification of dimorphic Human Specific Alu insertions, is stable enough to be used not only as a tool for genetic mapping but also to characterize populations, study migrational patterns and track the inheritance of human genetic disorders.

  19. Intracellular Bacteria in Protozoa

    NASA Astrophysics Data System (ADS)

    Görtz, Hans-Dieter; Brigge, Theo

    Intracellular bacteria in humans are typically detrimental, and such infections are regarded by the patients as accidental and abnormal. In protozoa it seems obvious that many bacteria have coevolved with their hosts and are well adapted to the intracellular way of life. Manifold interactions between hosts and intracellular bacteria are found, and examples of antibacterial resistance of unknown mechanisms are observed. The wide diversity of intracellular bacteria in protozoa has become particularly obvious since they have begun to be classified by molecular techniques. Some of the bacteria are closely related to pathogens; others are responsible for the production of toxins.

  20. Bacteriophage biosensors for antibiotic-resistant bacteria.

    PubMed

    Sorokulova, Irina; Olsen, Eric; Vodyanoy, Vitaly

    2014-03-01

    An increasing number of disease-causing bacteria are resistant to one or more anti-bacterial drugs utilized for therapy. Early and speedy detection of these pathogens is therefore very important. Traditional pathogen detection techniques, that include microbiological and biochemical assays are long and labor-intensive, while antibody or DNA-based methods require substantial sample preparation and purification. Biosensors based on bacteriophages have demonstrated remarkable potential to surmount these restrictions and to offer rapid, efficient and sensitive detection technique for antibiotic-resistant bacteria.

  1. Identification of bacteria in scuba divers' rinse tanks.

    PubMed

    Washburn, Brian K; Levin, Andrew E; Hennessy, Kristen; Miller, Michael R

    2010-01-01

    Scuba divers typically rinse equipment in communal tanks. Studies show these tanks are contaminated with bacteria, but the types of bacteria have not been studied. We sought to identify bacteria in rinse tanks at a dive facility at San Pedro, Belize, to determine the origin of the bacteria and determine whether the bacteria represented potential threats to human health. The identity of bacteria was investigated using reverse line blot (RLB) assays based on 28 different rDNA probes designed to detect known pathogens of sepsis, as well as by sequencing 23S rDNA from isolates and performing VITEK identification of several isolates. Based on the identities of bacteria in divers' rinse tanks, many likely originate from the ocean, and others likely originate from the divers themselves. None of the bacteria identified would be considered overt human pathogens. However, some of the bacteria found in the tanks are known to be associated with unsanitary conditions and can cause opportunistic infections, which may pose health problems to some individuals. Rinsing scuba equipment in communal tanks has the potential to transmit disease among some divers. Equipment, especially regulators and masks, should be rinsed/cleaned individually and not be placed in communal tanks.

  2. Genomics of Probiotic Bacteria

    NASA Astrophysics Data System (ADS)

    O'Flaherty, Sarah; Goh, Yong Jun; Klaenhammer, Todd R.

    Probiotic bacteria from the Lactobacillus and Bifidobacterium species belong to the Firmicutes and the Actinobacteria phylum, respectively. Lactobacilli are members of the lactic acid bacteria (LAB) group, a broadly defined family of microorganisms that ferment various hexoses into primarily lactic acid. Lactobacilli are typically low G + C gram-positive species which are phylogenetically diverse, with over 100 species documented to date. Bifidobacteria are heterofermentative, high G + C content bacteria with about 30 species of bifidobacteria described to date.

  3. Structural investigation into physiological DNA phosphorothioate modification

    PubMed Central

    Lan, Wenxian; Hu, Zhongpei; Shen, Jie; Wang, Chunxi; Jiang, Feng; Liu, Huili; Long, Dewu; Liu, Maili; Cao, Chunyang

    2016-01-01

    DNA phosphorothioate (PT) modification, with sulfur replacing a nonbridging phosphate oxygen in a sequence and stereo specific manner, is a novel physiological variation in bacteria. But what effects on DNA properties PT modification has is still unclear. To address this, we prepared three double-stranded (ds) DNA decamers, d(CGPXGCCGCCGA) with its complementary strand d(TCGGCGPXGCCG) (where X = O or S, i.e., PT-free dsDNA, [Sp, Sp]-PT dsDNA or [Rp, Rp]-PT dsDNA) located in gene of Streptomyces lividans. Their melting temperature (Tm) measurement indicates that [Rp, Rp]-PT dsDNA is most unstable. Their electron transfer potential detection presents an order of anti-oxidation properties: Sp-PT DNA > Rp-PT DNA > PT-free DNA. Their NMR structures demonstrate that PT modification doesn’t change their B-form conformation. The sulfur in [Rp, Rp]-PT dsDNA locates in the major groove, with steric effects on protons in the sugar close to modification sites, resulting in its unstability, and facilitating its selectively interactions with ScoMcrA. We thought that PT modification was dialectical to the bacteria. It protects the hosting bacteria by working as antioxidant against H2O2, and acts as a marker, directing restriction enzyme observed in other hosts, like ScoMcrA, to correctly cleave the PT modified DNA, so that bacteria cannot spread and survive. PMID:27169778

  4. In vitro antibacterial activity of AZD0914, a new spiropyrimidinetrione DNA gyrase/topoisomerase inhibitor with potent activity against Gram-positive, fastidious Gram-Negative, and atypical bacteria.

    PubMed

    Huband, Michael D; Bradford, Patricia A; Otterson, Linda G; Basarab, Gregory S; Kutschke, Amy C; Giacobbe, Robert A; Patey, Sara A; Alm, Richard A; Johnstone, Michele R; Potter, Marie E; Miller, Paul F; Mueller, John P

    2015-01-01

    AZD0914 is a new spiropyrimidinetrione bacterial DNA gyrase/topoisomerase inhibitor with potent in vitro antibacterial activity against key Gram-positive (Staphylococcus aureus, Staphylococcus epidermidis, Streptococcus pneumoniae, Streptococcus pyogenes, and Streptococcus agalactiae), fastidious Gram-negative (Haemophilus influenzae and Neisseria gonorrhoeae), atypical (Legionella pneumophila), and anaerobic (Clostridium difficile) bacterial species, including isolates with known resistance to fluoroquinolones. AZD0914 works via inhibition of DNA biosynthesis and accumulation of double-strand cleavages; this mechanism of inhibition differs from those of other marketed antibacterial compounds. AZD0914 stabilizes and arrests the cleaved covalent complex of gyrase with double-strand broken DNA under permissive conditions and thus blocks religation of the double-strand cleaved DNA to form fused circular DNA. Whereas this mechanism is similar to that seen with fluoroquinolones, it is mechanistically distinct. AZD0914 exhibited low frequencies of spontaneous resistance in S. aureus, and if mutants were obtained, the mutations mapped to gyrB. Additionally, no cross-resistance was observed for AZD0914 against recent bacterial clinical isolates demonstrating resistance to fluoroquinolones or other drug classes, including macrolides, β-lactams, glycopeptides, and oxazolidinones. AZD0914 was bactericidal in both minimum bactericidal concentration and in vitro time-kill studies. In in vitro checkerboard/synergy testing with 17 comparator antibacterials, only additivity/indifference was observed. The potent in vitro antibacterial activity (including activity against fluoroquinolone-resistant isolates), low frequency of resistance, lack of cross-resistance, and bactericidal activity of AZD0914 support its continued development. PMID:25385112

  5. Bleach vs. Bacteria

    MedlinePlus

    ... Inside Life Science > Bleach vs. Bacteria Inside Life Science View All Articles | Inside Life Science Home Page Bleach vs. Bacteria By Sharon Reynolds ... For Proteins, Form Shapes Function This Inside Life Science article also appears on LiveScience . Learn about related ...

  6. Bacteria turn tiny gears

    SciTech Connect

    2009-01-01

    Swarms of bacteria turn two 380-micron long gears, opening the possibility of building hybrid biological machines at the microscopic scale. Read more at Wired: http://www.wired.com/wiredscience/2009/12/bacterial-micro-machine/#more-15684 or Scientific American: http://www.scientificamerican.com/article.cfm?id=brownian-motion-bacteria

  7. Some Bacteria Are Beneficial!

    USGS Publications Warehouse

    McMahon, Peter B.

    1995-01-01

    Most people would agree that bacteria usually spell trouble where the quality of drinking water is con cerned. However, recent studies conducted by the U.S. Geological Survey (USGS) under the National Water-Quality Assessment (NAWQA) program have shown that some bacteria can improve the quality of water.

  8. [Genetic virulence markers of opportunistic bacteria].

    PubMed

    Bondarenko, V M

    2011-01-01

    The analysis of opportunistic bacteria phenotypic and genetic virulence markers indicates that pathogenicity formation is based on a structural modification of bacterial DNA which is linked with migration of interbacterial pathogenicity "islands" genetic determinants. Structural organization features of these mobile genetic elements determine high expression probability, and PCR detection of pathogenicity "islands" determinants that control adhesins, invasins, cytotoxic and cytolitic toxines synthesis may indicate etiopathogenetic significance of clinical isolates.

  9. Precision genome engineering in lactic acid bacteria

    PubMed Central

    2014-01-01

    Innovative new genome engineering technologies for manipulating chromosomes have appeared in the last decade. One of these technologies, recombination mediated genetic engineering (recombineering) allows for precision DNA engineering of chromosomes and plasmids in Escherichia coli. Single-stranded DNA recombineering (SSDR) allows for the generation of subtle mutations without the need for selection and without leaving behind any foreign DNA. In this review we discuss the application of SSDR technology in lactic acid bacteria, with an emphasis on key factors that were critical to move this technology from E. coli into Lactobacillus reuteri and Lactococcus lactis. We also provide a blueprint for how to proceed if one is attempting to establish SSDR technology in a lactic acid bacterium. The emergence of CRISPR-Cas technology in genome engineering and its potential application to enhancing SSDR in lactic acid bacteria is discussed. The ability to perform precision genome engineering in medically and industrially important lactic acid bacteria will allow for the genetic improvement of strains without compromising safety. PMID:25185700

  10. Isolation of isoprene degrading bacteria from soils, development of isoA gene probes and identification of the active isoprene-degrading soil community using DNA-stable isotope probing.

    PubMed

    El Khawand, Myriam; Crombie, Andrew T; Johnston, Antonia; Vavlline, Dmitrii V; McAuliffe, Joseph C; Latone, Jacob A; Primak, Yuliya A; Lee, Sang-Kyu; Whited, Gregg M; McGenity, Terry J; Murrell, J Colin

    2016-09-01

    Emissions of biogenic volatile organic compounds (bVOCs), are an important element in the global carbon cycle, accounting for a significant proportion of fixed carbon. They contribute directly and indirectly to global warming and climate change and have a major effect on atmospheric chemistry. Plants emit isoprene to the atmosphere in similar quantities to emissions of methane from all sources and each accounts for approximately one third of total VOCs. Although methanotrophs, capable of growth on methane, have been intensively studied, we know little of isoprene biodegradation. Here, we report the isolation of two isoprene-degrading strains from the terrestrial environment and describe the design and testing of polymerase chain reaction (PCR) primers targeting isoA, the gene encoding the active-site component of the conserved isoprene monooxygenase, which are capable of retrieving isoA sequences from isoprene-enriched environmental samples. Stable isotope probing experiments, using biosynthesized (13) C-labelled isoprene, identified the active isoprene-degrading bacteria in soil. This study identifies novel isoprene-degrading strains using both culture-dependent and, for the first time, culture-independent methods and provides the tools and foundations for continued investigation of the biogeography and molecular ecology of isoprene-degrading bacteria.

  11. Genetic manipulation of acidophilic bacteria

    SciTech Connect

    Ward, T.E.; Rowland, M.L.; Glenn, A.W.; Watkins, C.S.; Bruhn, D.F.; Bulmer, D.; Roberto, F.F.

    1989-01-01

    Thiobacillus ferrooxidans is important in leaching of metals from mineral ores and in the removal of pyritic sulfur from coal. It is also intimately involved in production of acid mine drainage. Other acidophilic bacteria, including members of the genus Acidiphilium, are usually present in the same environments as T. ferrooxidans, and there is evidence to suggest that these acidophilic heterotrophs may increase the rate of T. ferrooxidans' attack on inorganic sulfides. Our laboratory is studying the genetic characteristics of these acidophilic bacteria and developing techniques for introducing desirable genes into them. Several endogenous plasmids from Acidiphilium strains have been cloned into E. coli vectors. Some of the resulting plasmids are able to confer antibiotic resistance to Acidiphilium after transformation by electroporation. In addition, a broad-host range plasmid conferring resistance to tetracycline has been introduced into Acidiphilium strains by electroporation. This same plasmid, has also been transferred to Acidiphilium from E. coli directly by conjugation. A temperate bacteriophage which infects a number of Acidiphilium isolates has been discovered and partially characterized. It has a lambdoid morphology and a genome of approximately 97 kb, comprised of double-stranded DNA which is probably modified. 16 refs., 2 figs., 4 tabs.

  12. Visualization of DNA double-strand break repair in live bacteria reveals dynamic recruitment of Bacillus subtilis RecF, RecO and RecN proteins to distinct sites on the nucleoids.

    PubMed

    Kidane, Dawit; Sanchez, Humberto; Alonso, Juan C; Graumann, Peter L

    2004-06-01

    We have found that SMC-like RecN protein, RecF and RecO proteins that are involved in DNA recombination play an important role in DNA double-strand break (DSB) repair in Bacillus subtilis. Upon induction of DNA DSBs, RecN, RecO and RecF localized as a discrete focus on the nucleoids in a majority of cells, whereas two or three foci were rarely observed. RecN, RecO and RecF co-localized to the induced foci, with RecN localizing first, while RecO localized later, followed by RecF. Thus, three repair proteins were differentially recruited to distinct sites on the nucleoids, potentially constituting active DSB repair centres (RCs). RecF did not form regular foci in the absence of RecN and failed to form any foci in recO cells, demonstrating a central role for RecN and RecO in initializing the formation of RCs. RecN/O/F foci were detected in recA, recG or recU mutant cells, indicating that the proteins act upstream of proteins involved in synapsis or post-synapsis. In the absence of exogenous DNA damage, RCs were rare, but they accumulated in recA and recU cells, suggesting that DSBs occur frequently in the absence of RecA or RecU. The results suggest a model in which RecN that forms multimers in solution and high-molecular-weight complexes in cells containing DSBs initiates the formation of RCs that mediate DSB repair with the homologous sister chromosome, which presents a novel concept for DSB repair in prokaryotes. PMID:15186413

  13. Antibiotics from predatory bacteria

    PubMed Central

    Korp, Juliane; Vela Gurovic, María S

    2016-01-01

    Summary Bacteria, which prey on other microorganisms, are commonly found in the environment. While some of these organisms act as solitary hunters, others band together in large consortia before they attack their prey. Anecdotal reports suggest that bacteria practicing such a wolfpack strategy utilize antibiotics as predatory weapons. Consistent with this hypothesis, genome sequencing revealed that these micropredators possess impressive capacities for natural product biosynthesis. Here, we will present the results from recent chemical investigations of this bacterial group, compare the biosynthetic potential with that of non-predatory bacteria and discuss the link between predation and secondary metabolism. PMID:27340451

  14. Antibiotics from predatory bacteria.

    PubMed

    Korp, Juliane; Vela Gurovic, María S; Nett, Markus

    2016-01-01

    Bacteria, which prey on other microorganisms, are commonly found in the environment. While some of these organisms act as solitary hunters, others band together in large consortia before they attack their prey. Anecdotal reports suggest that bacteria practicing such a wolfpack strategy utilize antibiotics as predatory weapons. Consistent with this hypothesis, genome sequencing revealed that these micropredators possess impressive capacities for natural product biosynthesis. Here, we will present the results from recent chemical investigations of this bacterial group, compare the biosynthetic potential with that of non-predatory bacteria and discuss the link between predation and secondary metabolism. PMID:27340451

  15. Compartmentalization of prokaryotic DNA replication.

    PubMed

    Bravo, Alicia; Serrano-Heras, Gemma; Salas, Margarita

    2005-01-01

    It becomes now apparent that prokaryotic DNA replication takes place at specific intracellular locations. Early studies indicated that chromosomal DNA replication, as well as plasmid and viral DNA replication, occurs in close association with the bacterial membrane. Moreover, over the last several years, it has been shown that some replication proteins and specific DNA sequences are localized to particular subcellular regions in bacteria, supporting the existence of replication compartments. Although the mechanisms underlying compartmentalization of prokaryotic DNA replication are largely unknown, the docking of replication factors to large organizing structures may be important for the assembly of active replication complexes. In this article, we review the current state of this subject in two bacterial species, Escherichia coli and Bacillus subtilis, focusing our attention in both chromosomal and extrachromosomal DNA replication. A comparison with eukaryotic systems is also presented.

  16. Immunoelectronmicroscopic study of the nucleoid structure of hydrogen bacteria.

    PubMed

    Mogilnaya, O A; Kiselyova, E V; Medvedeva, S E; Puzir, A P; Guseynov, O A; Kulyba, N N; Kozlov, A V

    1992-01-01

    Electron microscopical studies of the nucleoid structure of hydrogen bacteria using ultrahin sections and spread DNA from bacterial cell lysates revealed a different DNA packaging in the cell. A compact state of the major part of DNA at all growth stages and stability of nucleosome-like structures were shown. The use of antibodies to HU protein of E. coli labelled by protein A-colloidal gold demonstrated the immunological relationship between HU protein of E. coli and histone-like proteins of Alcaligenes eutrophus and their possible role in the nucleosome-like DNA packaging in procariotic genome.

  17. Evaluation of six commercial DNA extraction kits for recovery of Burkholderia pseudomallei DNA.

    PubMed

    Marques, Maria Angela de Mello; Zimmermann, Pia; Messelhäußer, Ute; Sing, Andreas

    2012-12-01

    Six commercially available DNA extraction kits, as well as thermal lysis and proteinase K DNA extraction were evaluated regarding bacterial inactivation, DNA yield and purity, and their use in a Burkholderia pseudomallei real-time PCR. While all methods successfully inactivated the bacteria, by measuring DNA purity and the level of detection by real-time PCR, the proteinase K method was the most sensitive.

  18. [Darwin and bacteria].

    PubMed

    Ledermann D, Walter

    2009-02-01

    As in 2009 the scientific world celebrates two hundreds years from the birthday of Charles Darwin and one hundred and fifty from the publication of The Origin of Species, an analysis of his complete work is performed, looking for any mention of bacteria. But it seems that the great naturahst never took knowledge about its existence, something rather improbable in a time when the discovery of bacteria shook the medical world, or he deliberately ignored them, not finding a place for such microscopic beings into his theory of evolution. But the bacteria badly affected his familiar life, killing scarlet fever one of his children and worsening to death the evolution of tuberculosis of his favourite Annie. Darwin himself could suffer the sickness of Chagas, whose etiological agent has a similar level to bacteria in the scale of evolution. PMID:19350162

  19. Bacteria subsisting on antibiotics.

    PubMed

    Dantas, Gautam; Sommer, Morten O A; Oluwasegun, Rantimi D; Church, George M

    2008-04-01

    Antibiotics are a crucial line of defense against bacterial infections. Nevertheless, several antibiotics are natural products of microorganisms that have as yet poorly appreciated ecological roles in the wider environment. We isolated hundreds of soil bacteria with the capacity to grow on antibiotics as a sole carbon source. Of 18 antibiotics tested, representing eight major classes of natural and synthetic origin, 13 to 17 supported the growth of clonal bacteria from each of 11 diverse soils. Bacteria subsisting on antibiotics are surprisingly phylogenetically diverse, and many are closely related to human pathogens. Furthermore, each antibiotic-consuming isolate was resistant to multiple antibiotics at clinically relevant concentrations. This phenomenon suggests that this unappreciated reservoir of antibiotic-resistance determinants can contribute to the increasing levels of multiple antibiotic resistance in pathogenic bacteria. PMID:18388292

  20. Indicator For Pseudomonas Bacteria

    NASA Technical Reports Server (NTRS)

    Margalit, Ruth

    1990-01-01

    Characteristic protein extracted and detected. Natural protein marker found in Pseudomonas bacteria. Azurin, protein containing copper readily extracted, purified, and used to prepare antibodies. Possible to develop simple, fast, and accurate test for marker carried out in doctor's office.

  1. [Darwin and bacteria].

    PubMed

    Ledermann D, Walter

    2009-02-01

    As in 2009 the scientific world celebrates two hundreds years from the birthday of Charles Darwin and one hundred and fifty from the publication of The Origin of Species, an analysis of his complete work is performed, looking for any mention of bacteria. But it seems that the great naturahst never took knowledge about its existence, something rather improbable in a time when the discovery of bacteria shook the medical world, or he deliberately ignored them, not finding a place for such microscopic beings into his theory of evolution. But the bacteria badly affected his familiar life, killing scarlet fever one of his children and worsening to death the evolution of tuberculosis of his favourite Annie. Darwin himself could suffer the sickness of Chagas, whose etiological agent has a similar level to bacteria in the scale of evolution.

  2. Gut bacteria and cancer

    PubMed Central

    Erdman, Susan E.; Poutahidis, Theofilos

    2015-01-01

    Microbiota on the mucosal surfaces of the gastrointestinal (GI) tract greatly outnumber the cells in the human body. Effects of antibiotics indicate that GI tract bacteria may be determining the fate of distal cancers. Recent data implicate dysregulated host responses to enteric bacteria leading to cancers in extra-intestinal sites. Together these findings point to novel anti-cancer strategies aimed at promoting GI tract homeostasis. PMID:26050963

  3. Natural soil reservoirs for human pathogenic and fecal indicator bacteria

    USGS Publications Warehouse

    Boschiroli, Maria L; Falkinham, Joseph; Favre-Bonte, Sabine; Nazaret, Sylvie; Piveteau, Pascal; Sadowsky, Michael J.; Byappanahalli, Muruleedhara; Delaquis, Pascal; Hartmann, Alain

    2016-01-01

    Soils receive inputs of human pathogenic and indicator bacteria through land application of animal manures or sewage sludge, and inputs by wildlife. Soil is an extremely heterogeneous substrate and contains meso- and macrofauna that may be reservoirs for bacteria of human health concern. The ability to detect and quantify bacteria of human health concern is important in risk assessments and in evaluating the efficacy of agricultural soil management practices that are protective of crop quality and protective of adjacent water resources. The present chapter describes the distribution of selected Gram-positive and Gram-negative bacteria in soils. Methods for detecting and quantifying soilborne bacteria including extraction, enrichment using immunomagnetic capture, culturing, molecular detection and deep sequencing of metagenomic DNA to detect pathogens are overviewed. Methods for strain phenotypic and genotypic characterization are presented, as well as how comparison with clinical isolates can inform the potential for human health risk.

  4. Thermal control of virulence factors in bacteria: A hot topic

    PubMed Central

    Lam, Oliver; Wheeler, Jun; Tang, Christoph M

    2014-01-01

    Pathogenic bacteria sense environmental cues, including the local temperature, to control the production of key virulence factors. Thermal regulation can be achieved at the level of DNA, RNA or protein and although many virulence factors are subject to thermal regulation, the exact mechanisms of control are yet to be elucidated in many instances. Understanding how virulence factors are regulated by temperature presents a significant challenge, as gene expression and protein production are often influenced by complex regulatory networks involving multiple transcription factors in bacteria. Here we highlight some recent insights into thermal regulation of virulence in pathogenic bacteria. We focus on bacteria which cause disease in mammalian hosts, which are at a significantly higher temperature than the outside environment. We outline the mechanisms of thermal regulation and how understanding this fundamental aspect of the biology of bacteria has implications for pathogenesis and human health. PMID:25494856

  5. The fecal bacteria

    USGS Publications Warehouse

    Sadowsky, Michael J.; Whitman, Richard L.

    2011-01-01

    The Fecal Bacteria offers a balanced, integrated discussion of fecal bacteria and their presence and ecology in the intestinal tract of mammals, in the environment, and in the food supply. This volume covers their use in examining and assessing water quality in order to offer protection from illnesses related to swimming in or ingesting contaminated water, in addition to discussing their use in engineering considerations of water quality, modeling, monitoring, and regulations. Fecal bacteria are additionally used as indicators of contamination of ready-to-eat foods and fresh produce. The intestinal environment, the microbial community structure of the gut microbiota, and the physiology and genomics of this broad group of microorganisms are explored in the book. With contributions from an internationally recognized group of experts, the book integrates medicine, public health, environmental, and microbiological topics in order to provide a unique, holistic understanding of fecal bacteria. Moreover, it shows how the latest basic science and applied research findings are helping to solve problems and develop effective management strategies. For example, readers will discover how the latest tools and molecular approaches have led to our current understanding of fecal bacteria and enabled us to improve human health and water quality. The Fecal Bacteria is recommended for microbiologists, clinicians, animal scientists, engineers, environmental scientists, food safety experts, water quality managers, and students. It will help them better understand fecal bacteria and use their knowledge to protect human and environmental health. They can also apply many of the techniques and molecular tools discussed in this book to the study of a broad range of microorganisms in a variety of habitats.

  6. Stress-Induced Mutagenesis in Bacteria

    PubMed Central

    Foster, Patricia L.

    2009-01-01

    Bacteria spend their lives buffeted by changing environmental conditions. To adapt to and survive these stresses, bacteria have global response systems that result in sweeping changes in gene expression and cellular metabolism. These responses are controlled by master regulators, which include: alternative sigma factors, such as RpoS and RpoH; small molecule effectors, such as ppGpp; gene repressors such as LexA; and, inorganic molecules, such as polyphosphate. The response pathways extensively overlap and are induced to various extents by the same environmental stresses. These stresses include nutritional deprivation, DNA damage, temperature shift, and exposure to antibiotics. All of these global stress responses include functions that can increase genetic variability. In particular, up-regulation and activation of error-prone DNA polymerases, down-regulation of error-correcting enzymes, and movement of mobile genetic elements are common features of several stress responses. The result is that under a variety of stressful conditions, bacteria are induced for genetic change. This transient mutator state may be important for adaptive evolution. PMID:17917873

  7. DNA Charge Transport within the Cell

    PubMed Central

    Grodick, Michael A.; Muren, Natalie B.; Barton, Jacqueline K.

    2015-01-01

    The unique characteristics of DNA charge transport (CT) have prompted an examination of roles for this chemistry within a biological context. Not only can DNA CT facilitate long range oxidative damage of DNA, but redox-active proteins can couple to the DNA base stack and participate in long range redox reactions using DNA CT. DNA transcription factors with redox-active moieties such as SoxR and p53 can use DNA CT as a form of redox sensing. DNA CT chemistry also provides a means to monitor the integrity of the DNA, given the sensitivity of DNA CT to perturbations in base stacking as arise with mismatches and lesions. Enzymes that utilize this chemistry include an interesting and ever-growing class of DNA-processing enzymes involved in DNA repair, replication, and transcription that have been found to contain 4Fe-4S clusters. DNA repair enzymes containing 4Fe-4S clusters, that include Endonuclease III (EndoIII), MutY, and DinG from bacteria, as well as XPD from archaea, have been shown to be redox-active when bound to DNA, share a DNA-bound redox potential, and can be reduced and oxidized at long range via DNA CT. Interactions between DNA and these proteins in solution, in addition to genetics experiments within E. coli, suggest that DNA-mediated CT can be used as a means of cooperative signaling among DNA repair proteins that contain 4Fe-4S clusters as a first step in finding DNA damage, even within cells. Based on these data, we can consider also how DNA-mediated CT may be used as a means of signaling to coordinate DNA processing across the genome. PMID:25606780

  8. DNA charge transport within the cell.

    PubMed

    Grodick, Michael A; Muren, Natalie B; Barton, Jacqueline K

    2015-02-01

    The unique characteristics of DNA charge transport (CT) have prompted an examination of roles for this chemistry within a biological context. Not only can DNA CT facilitate long-range oxidative damage of DNA, but redox-active proteins can couple to the DNA base stack and participate in long-range redox reactions using DNA CT. DNA transcription factors with redox-active moieties such as SoxR and p53 can use DNA CT as a form of redox sensing. DNA CT chemistry also provides a means to monitor the integrity of the DNA, given the sensitivity of DNA CT to perturbations in base stacking as arise with mismatches and lesions. Enzymes that utilize this chemistry include an interesting and ever-growing class of DNA-processing enzymes involved in DNA repair, replication, and transcription that have been found to contain 4Fe-4S clusters. DNA repair enzymes containing 4Fe-4S clusters, that include endonuclease III (EndoIII), MutY, and DinG from bacteria, as well as XPD from archaea, have been shown to be redox-active when bound to DNA, share a DNA-bound redox potential, and can be reduced and oxidized at long-range via DNA CT. Interactions between DNA and these proteins in solution, in addition to genetics experiments within Escherichia coli, suggest that DNA-mediated CT can be used as a means of cooperative signaling among DNA repair proteins that contain 4Fe-4S clusters as a first step in finding DNA damage, even within cells. On the basis of these data, we can consider also how DNA-mediated CT may be used as a means of signaling to coordinate DNA processing across the genome.

  9. Antibiotic preparations contain DNA: a source of drug resistance genes?

    PubMed Central

    Webb, V; Davies, J

    1993-01-01

    Fluorescence measurements and polymerase chain reaction amplification of streptomycete 16S ribosomal DNA sequences were used to show that a number of antibiotic preparations employed for human and animal use are contaminated with chromosomal DNA of the antibiotic-producing organism. The DNA contains identifiable antibiotic resistance gene sequences; the uptake of this DNA by bacteria and its functional incorporation into bacterial replicons would lead to the generation of antibiotic resistance determinants. We propose that the presence of DNA encoding drug resistance in antibiotic preparations has been a factor in the rapid development of multiple antibiotic resistance in bacteria. Images PMID:8285621

  10. Anaerobic bacteria in otitis media.

    PubMed

    Fulghum, R S; Daniel, H J; Yarborough, J G

    1977-01-01

    Anaerobic bacteria, Peptostrepotococcus intermedius and Propionibacterium acnes, were found in mixed culture specimens from four to ten tested cases of chronic secretory otitis media. These anaerobic bacteria were in a mixed infection flora with aerobic bacteria most often Staphylococcus epidermidis and Cornybacterium sp. which do not fit any established species. The findings of anaerobic bacteria in otitis media is consistent with the sporadic report of the involvement of anaerobic bacteria in otitis media in the literature since 1898.

  11. Conjugative plasmid transfer in gram-positive bacteria.

    PubMed

    Grohmann, Elisabeth; Muth, Günther; Espinosa, Manuel

    2003-06-01

    Conjugative transfer of bacterial plasmids is the most efficient way of horizontal gene spread, and it is therefore considered one of the major reasons for the increase in the number of bacteria exhibiting multiple-antibiotic resistance. Thus, conjugation and spread of antibiotic resistance represents a severe problem in antibiotic treatment, especially of immunosuppressed patients and in intensive care units. While conjugation in gram-negative bacteria has been studied in great detail over the last decades, the transfer mechanisms of antibiotic resistance plasmids in gram-positive bacteria remained obscure. In the last few years, the entire nucleotide sequences of several large conjugative plasmids from gram-positive bacteria have been determined. Sequence analyses and data bank comparisons of their putative transfer (tra) regions have revealed significant similarities to tra regions of plasmids from gram-negative bacteria with regard to the respective DNA relaxases and their targets, the origins of transfer (oriT), and putative nucleoside triphosphatases NTP-ases with homologies to type IV secretion systems. In contrast, a single gene encoding a septal DNA translocator protein is involved in plasmid transfer between micelle-forming streptomycetes. Based on these clues, we propose the existence of two fundamentally different plasmid-mediated conjugative mechanisms in gram-positive microorganisms, namely, the mechanism taking place in unicellular gram-positive bacteria, which is functionally similar to that in gram-negative bacteria, and a second type that occurs in multicellular gram-positive bacteria, which seems to be characterized by double-stranded DNA transfer. PMID:12794193

  12. [Innovative treatments for multidrug-resistant bacteria].

    PubMed

    Pierre, Tattevin; Aurélien, Lorleac'h; Matthieu, Revest

    2014-03-01

    The spread of multidrug-resistant bacteria has accelerated sharply in the last decade. According to the World Health Organization they are responsible for an estimated 25 000 deaths in Europe each year. In addition, few new antibiotics are under development, raising the spectrum of a return to the "pre-antibiotic era". Non antibiotic antibacterial agents have recently attracted renewed interest. The most promising candidates are: i) phages (bacteria-infecting viruses) have been widely used in Eastern European countries since the 1930s but come up against logistic and regulatory obstacles due to the evolutionary nature of these biologic agents, while convincing clinical data are lacking; ii) bacteriocines are smallantibacterialpeptidesproducedby numerous bacteria; some have a rapid bactericidal effect, good tolerability, and a limited impact on the commensal flora; however, clinical use of bacteriocines is complicated by their fragility, poor penetration, and substantial risk of resistance selection ; iii) antisense oligonucleo tides act by inactivating genes through specific interaction with a complementary DNA or RNA fragment, potentially allowing specific inhibition of selected bacterial virulence factors. However, this therapeutic class may be more suitable for viral or genetic diseases than for multidrug-resistant bacterial infections, owing to the difficulty of delivering them inside bacteria. PMID:26427289

  13. Have sex or not? Lessons from bacteria.

    PubMed

    Lodé, T

    2012-01-01

    Sex is one of the greatest puzzles in evolutionary biology. A true meiotic process occurs only in eukaryotes, while in bacteria, gene transcription is fragmentary, so asexual reproduction in this case really means clonal reproduction. Sex could stem from a signal that leads to increased reproductive output of all interacting individuals and could be understood as a secondary consequence of primitive metabolic reactions. Meiotic sex evolved in proto-eukaryotes to solve a problem that bacteria did not have, namely a large amount of DNA material, occurring in an archaic step of proto-cell formation and genetic exchanges. Rather than providing selective advantages through reproduction, sex could be thought of as a series of separate events which combines step-by-step some very weak benefits of recombination, meiosis, gametogenesis and syngamy.

  14. Bacterial identification and subtyping using DNA microarray and DNA sequencing.

    PubMed

    Al-Khaldi, Sufian F; Mossoba, Magdi M; Allard, Marc M; Lienau, E Kurt; Brown, Eric D

    2012-01-01

    The era of fast and accurate discovery of biological sequence motifs in prokaryotic and eukaryotic cells is here. The co-evolution of direct genome sequencing and DNA microarray strategies not only will identify, isotype, and serotype pathogenic bacteria, but also it will aid in the discovery of new gene functions by detecting gene expressions in different diseases and environmental conditions. Microarray bacterial identification has made great advances in working with pure and mixed bacterial samples. The technological advances have moved beyond bacterial gene expression to include bacterial identification and isotyping. Application of new tools such as mid-infrared chemical imaging improves detection of hybridization in DNA microarrays. The research in this field is promising and future work will reveal the potential of infrared technology in bacterial identification. On the other hand, DNA sequencing by using 454 pyrosequencing is so cost effective that the promise of $1,000 per bacterial genome sequence is becoming a reality. Pyrosequencing technology is a simple to use technique that can produce accurate and quantitative analysis of DNA sequences with a great speed. The deposition of massive amounts of bacterial genomic information in databanks is creating fingerprint phylogenetic analysis that will ultimately replace several technologies such as Pulsed Field Gel Electrophoresis. In this chapter, we will review (1) the use of DNA microarray using fluorescence and infrared imaging detection for identification of pathogenic bacteria, and (2) use of pyrosequencing in DNA cluster analysis to fingerprint bacterial phylogenetic trees.

  15. Bacteria-surface interactions

    PubMed Central

    Tuson, Hannah H.; Weibel, Douglas B.

    2013-01-01

    The interaction of bacteria with surfaces has important implications in a range of areas, including bioenergy, biofouling, biofilm formation, and the infection of plants and animals. Many of the interactions of bacteria with surfaces produce changes in the expression of genes that influence cell morphology and behavior, including genes essential for motility and surface attachment. Despite the attention that these phenotypes have garnered, the bacterial systems used for sensing and responding to surfaces are still not well understood. An understanding of these mechanisms will guide the development of new classes of materials that inhibit and promote cell growth, and complement studies of the physiology of bacteria in contact with surfaces. Recent studies from a range of fields in science and engineering are poised to guide future investigations in this area. This review summarizes recent studies on bacteria-surface interactions, discusses mechanisms of surface sensing and consequences of cell attachment, provides an overview of surfaces that have been used in bacterial studies, and highlights unanswered questions in this field. PMID:23930134

  16. Communication among Oral Bacteria

    PubMed Central

    Kolenbrander, Paul E.; Andersen, Roxanna N.; Blehert, David S.; Egland, Paul G.; Foster, Jamie S.; Palmer, Robert J.

    2002-01-01

    Human oral bacteria interact with their environment by attaching to surfaces and establishing mixed-species communities. As each bacterial cell attaches, it forms a new surface to which other cells can adhere. Adherence and community development are spatiotemporal; such order requires communication. The discovery of soluble signals, such as autoinducer-2, that may be exchanged within multispecies communities to convey information between organisms has emerged as a new research direction. Direct-contact signals, such as adhesins and receptors, that elicit changes in gene expression after cell-cell contact and biofilm growth are also an active research area. Considering that the majority of oral bacteria are organized in dense three-dimensional biofilms on teeth, confocal microscopy and fluorescently labeled probes provide valuable approaches for investigating the architecture of these organized communities in situ. Oral biofilms are readily accessible to microbiologists and are excellent model systems for studies of microbial communication. One attractive model system is a saliva-coated flowcell with oral bacterial biofilms growing on saliva as the sole nutrient source; an intergeneric mutualism is discussed. Several oral bacterial species are amenable to genetic manipulation for molecular characterization of communication both among bacteria and between bacteria and the host. A successful search for genes critical for mixed-species community organization will be accomplished only when it is conducted with mixed-species communities. PMID:12209001

  17. PATHOGENICITY OF BIOFILM BACTERIA

    EPA Science Inventory

    There is a paucity of information concerning any link between the microorganisms commonly found in biofilms of drinking water systems and their impacts on human health. For bacteria, culture-based techniques detect only a limited number of the total microorganisms associated wit...

  18. Antibiotic-Resistant Bacteria.

    ERIC Educational Resources Information Center

    Longenecker, Nevin E.; Oppenheimer, Dan

    1982-01-01

    A study conducted by high school advanced bacteriology students appears to confirm the hypothesis that the incremental administration of antibiotics on several species of bacteria (Escherichia coli, Staphylococcus epidermis, Bacillus sublitus, Bacillus megaterium) will allow for the development of antibiotic-resistant strains. (PEB)

  19. R-body-producing bacteria.

    PubMed

    Pond, F R; Gibson, I; Lalucat, J; Quackenbush, R L

    1989-03-01

    Until 10 years ago, R bodies were known only as diagnostic features by which endosymbionts of paramecia were identified as kappa particles. They were thought to be limited to the cytoplasm of two species in the Paramecium aurelia species complex. Now, R bodies have been found in free-living bacteria and other Paramecium species. The organisms now known to form R bodies include the cytoplasmic kappa endosymbionts of P. biaurelia and P. tetraurelia, the macronuclear kappa endosymbionts of P. caudatum, Pseudomonas avenae (a free-living plant pathogen), Pseudomonas taeniospiralis (a hydrogen-oxidizing soil microorganism), Rhodospirillum centenum (a photosynthetic bacterium), and a soil bacterium, EPS-5028, which is probably a pseudomonad. R bodies themselves fall into five distinct groups, distinguished by size, the morphology of the R-body ribbons, and the unrolling behavior of wound R bodies. In recent years, the inherent difficulties in studying the organization and assembly of R bodies by the obligate endosymbiont kappa, have been alleviated by cloning and expressing genetic determinants for these R bodies (type 51) in Escherichia coli. Type 51 R-body synthesis requires three low-molecular-mass polypeptides. One of these is modified posttranslationally, giving rise to 12 polypeptide species, which are the major structural subunits of the R body. R bodies are encoded in kappa species by extrachromosomal elements. Type 51 R bodies, produced in Caedibacter taeniospiralis, are encoded by a plasmid, whereas bacteriophage genomes probably control R-body synthesis in other kappa species. However, there is no evidence that either bacteriophages or plasmids are present in P. avenae or P. taeniospiralis. No sequence homology was detected between type 51 R-body-encoding DNA and DNA from any R-body-producing species, except C. varicaedens 1038. The evolutionary relatedness of different types of R bodies remains unknown. PMID:2651865

  20. Ultrasensitive Detection of Bacteria by Targeting Abundant Transcripts.

    PubMed

    Wang, Xinhui; Li, Xinran; Liu, Shiwei; Ren, Hang; Yang, Mingjuan; Ke, Yuehua; Huang, Liuyu; Liu, Chao; Liu, Bo; Chen, Zeliang

    2016-01-01

    Molecular detection assays are increasingly becoming routine diagnostic techniques for bacterial infection; however, their sensitivities are restricted by the low concentrations of bacteria in clinical samples. Here, we report a new paradigm for ultrasensitive detection of bacteria. The principle of this approach is that by choosing highly transcribed genes as signature sequences and detecting both DNA and its RNA transcripts, assay sensitivity can be greatly improved. First, signature genes with abundant transcripts were screened by RNA-Seq. We confirmed that RT-PCR efficiently amplifies both DNA and RNA, while PCR amplifies only DNA. Unexpectedly, we found that the RNA extraction efficiency is relatively low, while simplified denaturation was more appropriate for transcript detection. For highly transcribed genes, RT-PCR consistently generated lower cycle threshold (Ct) values than those of PCR. The sensitivity of RT-PCR targeting abundant transcripts could detect quantities as low as one bacterium, which was not possible using PCR. Amplification of different genes among several other common bacteria also confirmed that transcript detection by RT-PCR is more sensitive than is DNA detection by PCR. Therefore, abundant transcript detection represents a universal strategy for ultrasensitive detection of bacteria.

  1. Plasmid incidence in bacteria from deep subsurface sediments.

    PubMed

    Fredrickson, J K; Hicks, R J; Li, S W; Brockman, F J

    1988-12-01

    Bacteria were isolated from deep terrestrial subsurface sediments underlying the coastal plain of South Carolina. A total of 163 isolates from deep sediments, surface soil, and return drill muds were examined for plasmid DNA content and resistance to the antibiotics penicillin, ampicillin, carbenicillin, streptomycin, kanamycin, and tetracycline. MICs of Cu, Cr, and Hg for each isolate were also determined. The overall frequency of plasmid occurrence in the subsurface bacteria was 33%. Resistance was most frequent to penicillin (70% of all isolates), ampicillin (49%), and carbenicillin (32%) and was concluded to be related to the concentrations of the individual antibiotics in the disks used for assaying resistance and to the production of low levels of beta-lactamase. The frequencies of resistance to penicillin and ampicillin were significantly greater for isolates bearing plasmids than for plasmidless isolates; however, resistance was not transferable to penicillin-sensitive Escherichia coli. Hybridization of subsurface bacterial plasmids and chromosomal DNA with a whole-TOL-plasmid (pWWO) probe revealed some homology of subsurface bacterial plasmid and chromosomal DNAs, indicating a potential for those bacteria to harbor catabolic genes on plasmids or chromosomes. The incidences of antibiotic resistance and MICs of metals for subsurface bacteria were significantly different from those for drill mud bacteria, ruling out the possibility that bacteria from sediments were derived from drill muds. PMID:16347789

  2. Caenorhabditis elegans responses to bacteria from its natural habitats.

    PubMed

    Samuel, Buck S; Rowedder, Holli; Braendle, Christian; Félix, Marie-Anne; Ruvkun, Gary

    2016-07-01

    Most Caenorhabditis elegans studies have used laboratory Escherichia coli as diet and microbial environment. Here we characterize bacteria of C. elegans' natural habitats of rotting fruits and vegetation to provide greater context for its physiological responses. By the use of 16S ribosomal DNA (rDNA)-based sequencing, we identified a large variety of bacteria in C. elegans habitats, with phyla Proteobacteria, Bacteroidetes, Firmicutes, and Actinobacteria being most abundant. From laboratory assays using isolated natural bacteria, C. elegans is able to forage on most bacteria (robust growth on ∼80% of >550 isolates), although ∼20% also impaired growth and arrested and/or stressed animals. Bacterial community composition can predict wild C. elegans population states in both rotting apples and reconstructed microbiomes: alpha-Proteobacteria-rich communities promote proliferation, whereas Bacteroidetes or pathogens correlate with nonproliferating dauers. Combinatorial mixtures of detrimental and beneficial bacteria indicate that bacterial influence is not simply nutritional. Together, these studies provide a foundation for interrogating how bacteria naturally influence C. elegans physiology. PMID:27317746

  3. Caenorhabditis elegans responses to bacteria from its natural habitats

    PubMed Central

    Rowedder, Holli; Braendle, Christian; Félix, Marie-Anne; Ruvkun, Gary

    2016-01-01

    Most Caenorhabditis elegans studies have used laboratory Escherichia coli as diet and microbial environment. Here we characterize bacteria of C. elegans' natural habitats of rotting fruits and vegetation to provide greater context for its physiological responses. By the use of 16S ribosomal DNA (rDNA)-based sequencing, we identified a large variety of bacteria in C. elegans habitats, with phyla Proteobacteria, Bacteroidetes, Firmicutes, and Actinobacteria being most abundant. From laboratory assays using isolated natural bacteria, C. elegans is able to forage on most bacteria (robust growth on ∼80% of >550 isolates), although ∼20% also impaired growth and arrested and/or stressed animals. Bacterial community composition can predict wild C. elegans population states in both rotting apples and reconstructed microbiomes: alpha-Proteobacteria-rich communities promote proliferation, whereas Bacteroidetes or pathogens correlate with nonproliferating dauers. Combinatorial mixtures of detrimental and beneficial bacteria indicate that bacterial influence is not simply nutritional. Together, these studies provide a foundation for interrogating how bacteria naturally influence C. elegans physiology. PMID:27317746

  4. REVIEW ARTICLE: DNA protein interactions and bacterial chromosome architecture

    NASA Astrophysics Data System (ADS)

    Stavans, Joel; Oppenheim, Amos

    2006-12-01

    Bacteria, like eukaryotic organisms, must compact the DNA molecule comprising their genome and form a functional chromosome. Yet, bacteria do it differently. A number of factors contribute to genome compaction and organization in bacteria, including entropic effects, supercoiling and DNA-protein interactions. A gamut of new experimental techniques have allowed new advances in the investigation of these factors, and spurred much interest in the dynamic response of the chromosome to environmental cues, segregation, and architecture, during both exponential and stationary phases. We review these recent developments with emphasis on the multifaceted roles that DNA-protein interactions play.

  5. How-to-Do-It: Recombinant DNA Made Easy II. Gene, Gene, Who's Got the Gene?

    ERIC Educational Resources Information Center

    Thomson, Robert G.

    1989-01-01

    Described is an activity in which students are able to determine that DNA can be transferred between bacteria and should be able to predict the type of DNA transferred. Methods, materials, and results are discussed. (CW)

  6. DNA phosphorothioate modifications influence the global transcriptional response and protect DNA from double-stranded breaks

    PubMed Central

    Gan, Rui; Wu, Xiaolin; He, Wei; Liu, Zhenhua; Wu, Shuangju; Chen, Chao; Chen, Si; Xiang, Qianrong; Deng, Zixin; Liang, Dequan; Chen, Shi; Wang, Lianrong

    2014-01-01

    The modification of DNA by phosphorothioate (PT) occurs when the non-bridging oxygen in the sugar-phosphate backbone of DNA is replaced with sulfur. This DNA backbone modification was recently discovered and is governed by the dndABCDE genes in a diverse group of bacteria and archaea. However, the biological function of DNA PT modifications is poorly understood. In this study, we employed the RNA-seq analysis to characterize the global transcriptional changes in response to PT modifications. Our results show that DNA without PT protection is susceptible to DNA damage caused by the dndFGHI gene products. The DNA double-stranded breaks then trigger the SOS response, cell filamentation and prophage induction. Heterologous expression of dndBCDE conferring DNA PT modifications at GPSA and GPST prevented the damage in Salmonella enterica. Our data provide insights into the physiological role of the DNA PT system. PMID:25319634

  7. Spatial regulation and organization of DNA replication within the nucleus.

    PubMed

    Natsume, Toyoaki; Tanaka, Tomoyuki U

    2010-01-01

    Duplication of chromosomal DNA is a temporally and spatially regulated process. The timing of DNA replication initiation at various origins is highly coordinated; some origins fire early and others late during S phase. Moreover, inside the nuclei, the bulk of DNA replication is physically organized in replication factories, consisting of DNA polymerases and other replication proteins. In this review article, we discuss how DNA replication is organized and regulated spatially within the nucleus and how this spatial organization is linked to temporal regulation. We focus on DNA replication in budding yeast and fission yeast and, where applicable, compare yeast DNA replication with that in bacteria and metazoans.

  8. Mitochondrial DNA.

    ERIC Educational Resources Information Center

    Wright, Russell G.; Bottino, Paul J.

    1986-01-01

    Provides background information for teachers on mitochondrial DNA, pointing out that it may have once been a free-living organism. Includes a ready-to-duplicate exercise titled "Using Microchondrial DNA to Measure Evolutionary Distance." (JN)

  9. Radiobiological effects of heavy ions and protons. [on cells of mammals, bacteria and viruses

    NASA Technical Reports Server (NTRS)

    Ryzhov, N. I.; Vorozhtsova, S. V.; Krasavin, Y. A.; Mashinskaya, T. Y.; Savchenko, N. Y.; Fedorov, B. S.; Khlaponina, V. F.; Shelegedin, V. N.; Gut, L.; Sabo, L.

    1974-01-01

    Radiobiological effects of heavy ions and protons are studied on cells of mammals, bacteria, viruses and DNA of bacteria. Results show that the dose effect dependence bears an exponential character; the reduction of RBE as LET of particle increases reflects the different character of microdistribution of absorbed energy in biological objects with different levels of biological organization.

  10. DNA Banking

    SciTech Connect

    Reilly, P.R. )

    1992-11-01

    The author is involved in the ethical, legal, and social issues of banking of DNA and data from DNA analysis. In his attempt to determine the extent of DNA banking in the U.S., the author surveyed some commercial companies performing DNA banking services. This article summarizes the results of that survey, with special emphasis on the procedures the companies use to protect the privacy of individuals. 4 refs.

  11. Structural and Thermodynamic Signatures of DNA Recognition by Mycobacterium tuberculosis DnaA

    SciTech Connect

    Tsodikov, Oleg V.; Biswas, Tapan

    2011-09-06

    An essential protein, DnaA, binds to 9-bp DNA sites within the origin of replication oriC. These binding events are prerequisite to forming an enigmatic nucleoprotein scaffold that initiates replication. The number, sequences, positions, and orientations of these short DNA sites, or DnaA boxes, within the oriCs of different bacteria vary considerably. To investigate features of DnaA boxes that are important for binding Mycobacterium tuberculosis DnaA (MtDnaA), we have determined the crystal structures of the DNA binding domain (DBD) of MtDnaA bound to a cognate MtDnaA-box (at 2.0 {angstrom} resolution) and to a consensus Escherichia coli DnaA-box (at 2.3 {angstrom}). These structures, complemented by calorimetric equilibrium binding studies of MtDnaA DBD in a series of DnaA-box variants, reveal the main determinants of DNA recognition and establish the [T/C][T/A][G/A]TCCACA sequence as a high-affinity MtDnaA-box. Bioinformatic and calorimetric analyses indicate that DnaA-box sequences in mycobacterial oriCs generally differ from the optimal binding sequence. This sequence variation occurs commonly at the first 2 bp, making an in vivo mycobacterial DnaA-box effectively a 7-mer and not a 9-mer. We demonstrate that the decrease in the affinity of these MtDnaA-box variants for MtDnaA DBD relative to that of the highest-affinity box TTGTCCACA is less than 10-fold. The understanding of DnaA-box recognition by MtDnaA and E. coli DnaA enables one to map DnaA-box sequences in the genomes of M. tuberculosis and other eubacteria.

  12. Recombinant bacteria for mosquito control.

    PubMed

    Federici, B A; Park, H-W; Bideshi, D K; Wirth, M C; Johnson, J J

    2003-11-01

    Bacterial insecticides have been used for the control of nuisance and vector mosquitoes for more than two decades. Nevertheless, due primarily to their high cost and often only moderate efficacy, these insecticides remain of limited use in tropical countries where mosquito-borne diseases are prevalent. Recently, however, recombinant DNA techniques have been used to improve bacterial insecticide efficacy by markedly increasing the synthesis of mosquitocidal proteins and by enabling new endotoxin combinations from different bacteria to be produced within single strains. These new strains combine mosquitocidal Cry and Cyt proteins of Bacillus thuringiensis with the binary toxin of Bacillus sphaericus, improving efficacy against Culex species by 10-fold and greatly reducing the potential for resistance through the presence of Cyt1A. Moreover, although intensive use of B. sphaericus against Culex populations in the field can result in high levels of resistance, most of this can be suppressed by combining this bacterial species with Cyt1A; the latter enables the binary toxin of this species to enter midgut epithelial cells via the microvillar membrane in the absence of a midgut receptor. The availability of these novel strains and newly discovered mosquitocidal proteins, such as the Mtx toxins of B. sphaericus, offers the potential for constructing a range of recombinant bacterial insecticides for more effective control of the mosquito vectors of filariasis, Dengue fever and malaria. PMID:14506223

  13. Isolation of methanotrophic bacteria from termite gut.

    PubMed

    Reuss, Julia; Rachel, Reinhard; Kämpfer, Peter; Rabenstein, Andreas; Küver, Jan; Dröge, Stefan; König, Helmut

    2015-10-01

    The guts of termites feature suitable conditions for methane oxidizing bacteria (MOB) with their permanent production of CH4 and constant supply of O2 via tracheae. In this study, we have isolated MOB from the gut contents of the termites Incisitermes marginipennis, Mastotermes darwiniensis, and Neotermes castaneus for the first time. The existence of MOB was indicated by detecting pmoA, the gene for the particulate methane monooxygenase, in the DNA of gut contents. Fluorescence in situ hybridization and quantitative real-time polymerase chain reaction supported those findings. The MOB cell titer was determined to be 10(2)-10(3) per gut. Analyses of the 16S rDNA from isolates indicated close similarity to the genus Methylocystis. After various physiological tests and fingerprinting methods, no exact match to a known species was obtained, indicating the isolation of new MOB species. However, MALDI-TOF MS analyses revealed a close relationship to Methylocystis bryophila and Methylocystis parvus.

  14. Recovering glycoside hydrolase genes from active tundra cellulolytic bacteria.

    PubMed

    Pinnell, Lee J; Dunford, Eric; Ronan, Patrick; Hausner, Martina; Neufeld, Josh D

    2014-07-01

    Bacteria responsible for cellulose hydrolysis in situ are poorly understood, largely because of the relatively recent development of cultivation-independent methods for their detection and characterization. This study combined DNA stable-isotope probing (DNA-SIP) and metagenomics for identifying active bacterial communities that assimilated carbon from glucose and cellulose in Arctic tundra microcosms. Following DNA-SIP, bacterial fingerprint analysis of gradient fractions confirmed isotopic enrichment. Sequenced fingerprint bands and clone library analysis of 16S rRNA genes identified active bacterial taxa associated with cellulose-associated labelled DNA, including Bacteroidetes (Sphingobacteriales), Betaproteobacteria (Burkholderiales), Alphaproteobacteria (Caulobacteraceae), and Chloroflexi (Anaerolineaceae). We also compared glycoside hydrolase metagenomic profiles from bulk soil and heavy DNA recovered from DNA-SIP incubations. Active populations consuming [(13)C]glucose and [(13)C]cellulose were distinct, based on ordinations of light and heavy DNA. Metagenomic analysis demonstrated a ∼3-fold increase in the relative abundance of glycoside hydrolases in DNA-SIP libraries over bulk-soil libraries. The data also indicate that multiple displacement amplification introduced bias into the resulting metagenomic analysis. This research identified DNA-SIP incubation conditions for glucose and cellulose that were suitable for Arctic tundra soil and confirmed that DNA-SIP enrichment can increase target gene frequencies in metagenomic libraries.

  15. Flowthrough Bacteria-Detection System

    NASA Technical Reports Server (NTRS)

    Grana, D. C.; Wilkins, J. R.

    1983-01-01

    Online system allows repetitive cycling of sample intake, bacteria counting and sterilization. System measures bacteria count by using sample/incubate/ measure cycle. Steps in cycle are on/off operations to cycle automated easily.

  16. Phylogenetic analysis on the soil bacteria distributed in karst forest

    PubMed Central

    Zhou, JunPei; Huang, Ying; Mo, MingHe

    2009-01-01

    Phylogenetic composition of bacterial community in soil of a karst forest was analyzed by culture-independent molecular approach. The bacterial 16S rRNA gene was amplified directly from soil DNA and cloned to generate a library. After screening the clone library by RFLP, 16S rRNA genes of representative clones were sequenced and the bacterial community was analyzed phylogenetically. The 16S rRNA gene inserts of 190 clones randomly selected were analyzed by RFLP and generated 126 different RFLP types. After sequencing, 126 non-chimeric sequences were obtained, generating 113 phylotypes. Phylogenetic analysis revealed that the bacteria distributed in soil of the karst forest included the members assigning into Proteobacteria, Acidobacteria, Planctomycetes, Chloroflexi (Green nonsulfur bacteria), Bacteroidetes, Verrucomicrobia, Nitrospirae, Actinobacteria (High G+C Gram-positive bacteria), Firmicutes (Low G+C Gram-positive bacteria) and candidate divisions (including the SPAM and GN08). PMID:24031430

  17. Small Talk: Cell-to-Cell Communication in Bacteria

    ScienceCinema

    Bassler, Bonnie [Princeton University, Princeton, New Jersey, United States

    2016-07-12

    Cell-cell communication in bacteria involves the production, release, and subsequent detection of chemical signaling molecules called autoinducers. This process, called quorum sensing, allows bacteria to regulate gene expression on a population-wide scale. Processes controlled by quorum sensing are usually ones that are unproductive when undertaken by an individual bacterium but become effective when undertaken by the group. For example, quorum sensing controls bioluminescence, secretion of virulence factors, biofilm formation, sporulation, and the exchange of DNA. Thus, quorum sensing is a mechanism that allows bacteria to function as multi-cellular organisms. Bacteria make, detect, and integrate information from multiple autoinducers, some of which are used exclusively for intra-species communication while others enable communication between species. Research is now focused on the development of therapies that interfere with quorum sensing to control bacterial virulence.

  18. Small Talk: Cell-to-Cell Communication in Bacteria

    SciTech Connect

    Bassler, Bonnie

    2008-05-14

    Cell-cell communication in bacteria involves the production, release, and subsequent detection of chemical signaling molecules called autoinducers. This process, called quorum sensing, allows bacteria to regulate gene expression on a population-wide scale. Processes controlled by quorum sensing are usually ones that are unproductive when undertaken by an individual bacterium but become effective when undertaken by the group. For example, quorum sensing controls bioluminescence, secretion of virulence factors, biofilm formation, sporulation, and the exchange of DNA. Thus, quorum sensing is a mechanism that allows bacteria to function as multi-cellular organisms. Bacteria make, detect, and integrate information from multiple autoinducers, some of which are used exclusively for intra-species communication while others enable communication between species. Research is now focused on the development of therapies that interfere with quorum sensing to control bacterial virulence.

  19. Small Talk: Cell-to-Cell Communication in Bacteria

    SciTech Connect

    Bassler, Bonnie

    2008-12-03

    Cell-cell communication in bacteria involves the production, release, and subsequent detection of chemical signaling molecules called autoinducers. This process, called quorum sensing, allows bacteria to regulate gene expression on a population-wide scale. Processes controlled by quorum sensing are usually ones that are unproductive when undertaken by an individual bacterium but become effective when undertaken by the group. For example, quorum sensing controls bioluminescence, secretion of virulence factors, biofilm formation, sporulation, and the exchange of DNA. Thus, quorum sensing is a mechanism that allows bacteria to function as multi-cellular organisms. Bacteria make, detect, and integrate information from multiple autoinducers, some of which are used exclusively for intra-species communication while others enable communication between species. Research is now focused on the development of therapies that interfere with quorum sensing to control bacterial virulence.

  20. Dna Sequencing

    DOEpatents

    Tabor, Stanley; Richardson, Charles C.

    1995-04-25

    A method for sequencing a strand of DNA, including the steps off: providing the strand of DNA; annealing the strand with a primer able to hybridize to the strand to give an annealed mixture; incubating the mixture with four deoxyribonucleoside triphosphates, a DNA polymerase, and at least three deoxyribonucleoside triphosphates in different amounts, under conditions in favoring primer extension to form nucleic acid fragments complementory to the DNA to be sequenced; labelling the nucleic and fragments; separating them and determining the position of the deoxyribonucleoside triphosphates by differences in the intensity of the labels, thereby to determine the DNA sequence.

  1. Lipoprotein sorting in bacteria.

    PubMed

    Okuda, Suguru; Tokuda, Hajime

    2011-01-01

    Bacterial lipoproteins are synthesized as precursors in the cytoplasm and processed into mature forms on the cytoplasmic membrane. A lipid moiety attached to the N terminus anchors these proteins to the membrane surface. Many bacteria are predicted to express more than 100 lipoproteins, which play diverse functions on the cell surface. The Lol system, composed of five proteins, catalyzes the localization of Escherichia coli lipoproteins to the outer membrane. Some lipoproteins play vital roles in the sorting of other lipoproteins, lipopolysaccharides, and β-barrel proteins to the outer membrane. On the basis of results from biochemical, genetic, and structural studies, we discuss the biogenesis of lipoproteins in bacteria, their importance in cellular functions, and the molecular mechanisms underlying efficient sorting of hydrophobic lipoproteins to the outer membrane through the hydrophilic periplasm. PMID:21663440

  2. Manufacture of Probiotic Bacteria

    NASA Astrophysics Data System (ADS)

    Muller, J. A.; Ross, R. P.; Fitzgerald, G. F.; Stanton, C.

    Lactic acid bacteria (LAB) have been used for many years as natural biopreservatives in fermented foods. A small group of LAB are also believed to have beneficial health effects on the host, so called probiotic bacteria. Probiotics have emerged from the niche industry from Asia into European and American markets. Functional foods are one of the fastest growing markets today, with estimated growth to 20 billion dollars worldwide by 2010 (GIA, 2008). The increasing demand for probiotics and the new food markets where probiotics are introduced, challenges the industry to produce high quantities of probiotic cultures in a viable and stable form. Dried concentrated probiotic cultures are the most convenient form for incorporation into functional foods, given the ease of storage, handling and transport, especially for shelf-stable functional products. This chapter will discuss various aspects of the challenges associated with the manufacturing of probiotic cultures.

  3. Bacteria, food, and cancer

    PubMed Central

    Rooks, Michelle G.

    2011-01-01

    Gut microbes are essential components of the human organism—helping us metabolize food into energy, produce micronutrients, and shape our immune systems. Having a particular pattern of gut microbes is also increasingly being linked to medical conditions including obesity, inflammatory bowel disease, and diabetes. Recent studies now indicate that our resident intestinal bacteria may also play a critical role in determining one's risk of developing cancer, ranging from protection against cancer to promoting its initiation and progression. Gut bacteria are greatly influenced by diet and in this review we explore evidence that they may be the missing piece that explains how dietary intake influences cancer risk, and discuss possible prevention and treatment strategies. PMID:21876723

  4. Reanimation of Ancient Bacteria

    SciTech Connect

    Vreeland, Russell H.

    2009-01-09

    Recent highly publicized experiments conducted on salt crystals taken from the Permian Salado Formation in Southeastern New Mexico have shown that some ancient crystals contain viable microorganisms trapped within fluid inclusions. Stringent geological and microbiological selection criteria were used to select crystals and conduct all sampling. This talk will focus on how each of these lines of data support the conclusion that such isolated bacteria are as old as the rock in which they are trapped. In this case, the isolated microbes are salt tolerant bacilli that grow best in media containing 8% NaCl, and respond to concentrated brines by forming spores. One of the organisms is phylogenetically related to several bacilli, but does have several unique characteristics. This talk will trace the interdisciplinary data and procedures supporting these discoveries, and describe the various isolated bacteria.

  5. Bacteria in Confined Spaces

    NASA Astrophysics Data System (ADS)

    Wilking, Connie; Weitz, David

    2010-03-01

    Bacterial cells can display differentiation between several developmental pathways, from planktonic to matrix-producing, depending upon the colony conditions. We study the confinement of bacteria in hydrogels as well as in liquid-liquid double emulsion droplets and observe the growth and morphology of these colonies as a function of time and environment. Our results can give insight into the behavior of bacterial colonies in confined spaces that can have applications in the areas of food science, cosmetics, and medicine.

  6. Characterization of lactic acid bacteria and other gut bacteria in pigs by a macroarraying method.

    PubMed

    Thanantong, Narut; Edwards, Sandra; Sparagano, Olivier A E

    2006-10-01

    Lactic acid bacteria (LAB) consist of many genera, Gram-positive, and nonspore-forming micro-organisms; some members being used as probiotics while some others have negative effects on pig health. Bacterial species in the gastrointestinal tract can produce antibacterial substances, reduce serum cholesterol in their host, or can be responsible for growth reduction, diarrhea, and intestinal epithelial damage. It is therefore important for the pig industry to evaluate the impact of food and farm management on the presence of "good" or "bad" bacteria and the risk for consumers. This articles focuses on the molecular identification of gut microflora species following different diets given to pigs in UK and correlating the data on growth, health, and welfare. First of all, pig feces were individually collected from sows before and after farrowing and also from piglets before and after weaning over several months. Bacteria colonies were grown on MRS agar plates from feces and DNA was extracted (QIAamp DNA stool kit) and amplified using 16S rDNA (27f and 519r) primers. DNA sequencing and sequence alignment allowed us to identify species-specific zones, which were used as probes in a macroarray system also known as reverse line blot hybridization. Some probes were found to be species specific for the following species: Lactobacillus acidophilus, L. animalis, L. gallinarum, L. kitasanotis, L salivarius, Streptococcus alactolyticus, S. hyointestinalis, and Sarcina ventriculi. Actual studies are now focusing on the impact of diets of the microflora in different gut parts and at different stages of the animal's life.

  7. Growing Unculturable Bacteria

    PubMed Central

    2012-01-01

    The bacteria that can be grown in the laboratory are only a small fraction of the total diversity that exists in nature. At all levels of bacterial phylogeny, uncultured clades that do not grow on standard media are playing critical roles in cycling carbon, nitrogen, and other elements, synthesizing novel natural products, and impacting the surrounding organisms and environment. While molecular techniques, such as metagenomic sequencing, can provide some information independent of our ability to culture these organisms, it is essentially impossible to learn new gene and pathway functions from pure sequence data. A true understanding of the physiology of these bacteria and their roles in ecology, host health, and natural product production requires their cultivation in the laboratory. Recent advances in growing these species include coculture with other bacteria, recreating the environment in the laboratory, and combining these approaches with microcultivation technology to increase throughput and access rare species. These studies are unraveling the molecular mechanisms of unculturability and are identifying growth factors that promote the growth of previously unculturable organisms. This minireview summarizes the recent discoveries in this area and discusses the potential future of the field. PMID:22661685

  8. Denitrification by extremely halophilic bacteria

    NASA Technical Reports Server (NTRS)

    Hochstein, L. I.; Tomlinson, G. A.

    1985-01-01

    Extremely halophilic bacteria were isolated from widely separated sites by anaerobic enrichment in the presence of nitrate. The anaerobic growth of several of these isolates was accompanied by the production of nitrite, nitrous oxide, and dinitrogen. These results are a direct confirmation of the existence of extremely halophilic denitrifying bacteria, and suggest that such bacteria may be common inhabitants of hypersaline environments.

  9. Real-time PCR quantification of nitrifying bacteria in a municipal wastewater treatment plant.

    PubMed

    Harms, Gerda; Layton, Alice C; Dionisi, Hebe M; Gregory, Igrid R; Garrett, Victoria M; Hawkins, Shawn A; Robinson, Kevin G; Sayler, Gary S

    2003-01-15

    Real-time PCR assays using TaqMan or Molecular Beacon probes were developed and optimized for the quantification of total bacteria, the nitrite-oxidizing bacteria Nitrospira, and Nitrosomonas oligotropha-like ammonia oxidizing bacteria (AOB) in mixed liquor suspended solids (MLSS) from a municipal wastewater treatment plant (WWTP) using a single-sludge nitrification process. The targets for the real-time PCR assays were the 16S rRNA genes (16S rDNA) for bacteria and Nitrospira spp. and the amoA gene for N. oligotropha. A previously reported assay for AOB 16S rDNA was also tested for its application to activated sludge. The Nitrospira 16S rDNA, AOB 16S rDNA, and N. oligotropha-like amoA assays were log-linear over 6 orders of magnitude and the bacterial 16S rDNA real-time PCR assay was log-linear over 4 orders of magnitude with DNA standards. When these real-time PCR assays were applied to DNA extracted from MLSS, dilution of the DNA extracts was necessary to prevent PCR inhibition. The optimal DNA dilution range was broad for the bacterial 16S rDNA (1000-fold) and Nitrospira 16S rDNA assays (2500-fold) but narrow for the AOB 16S rDNA assay (10-fold) and N. oligotropha-like amoA real-time PCR assay (5-fold). In twelve MLSS samples collected over one year, mean cell per L values were 4.3 +/- 2.0 x 10(11) for bacteria, 3.7 +/- 3.2 x 10(10) for Nitrospira, 1.2 +/- 0.9 x 10(10) for all AOB, and 7.5 +/- 6.0 x 10(9) for N. oligotropha-like AOB. The percent of the nitrifying population was 1.7% N. oligotropha-like AOB based on the N. oligotropha amoA assay, 2.9% total AOB based on the AOB 16S rDNA assay, and 8.6% nitrite-oxidizing bacteria based on the Nitrospira 16S rDNA assay. Ammonia-oxidizing bacteria in the wastewater treatment plant were estimated to oxidize 7.7 +/- 6.8 fmol/hr/cell based on the AOB 16S rDNA assay and 12.4 +/- 7.3 fmol/hr/cell based on the N. oligotropha amoA assay.

  10. Molecular analysis of polyphosphate accumulation in bacteria.

    PubMed

    Kuroda, A; Ohtake, H

    2000-03-01

    The dynamic behavior of inorganic polyphosphate (polyP), its accumulation and disappearance, is the most striking aspect of polyP metabolism in bacteria. Imbalance between polyP synthesis and degradation results in fluctuations of polyP by 100- to 1000-fold. We here review recent results with respect to this polyP metabolism in bacteria. PolyP accumulation in response to amino acid starvation, accompanied by increased levels of stringent factors, has been observed in Escherichia coli. Inhibition by stringent factors of polyphosphatase interrupts the dynamic balance between the synthesis and degradation of polyP, accounting for polyP accumulation. Polyphosphate kinase is required for activation of intracellular protein degradation, which is required for adaptation at the onset of amino acid starvation. The adaptation to amino acid starvation is mediated by the network of stringent response and polyP metabolism. PolyP accumulation independent of stringent response has also been observed. Novobiocin, an inhibitor for DNA gyrase, stimulated accumulation of polyP but not that of stringent factors. However, a temperature-sensitive DNA gyrase mutant did not exhibit polyP accumulation at the non-permissive temperature. Antagonistic relationship of polyP to nucleic acid synthesis, explored by Harold, appears to be more complicated. We discuss relationship of Pi regulation to polyP accumulation in E. coli and Klebsiella aerogenes. A function of polyP as an in vivo phosphagen affecting polyP accumulation is also discussed.

  11. The Origin And Spread Of Airborne Bacteria

    NASA Astrophysics Data System (ADS)

    Henderson-Begg, S. K.; Moffett, B. F.

    2009-12-01

    The presence of bacteria in clouds may affect their radiation and precipitation properties as some species are able to catalyse the freezing of water at high temperatures (-2C to -10C). Where cloud-borne bacteria originate and the distances they are able to travel in the air remains a mystery. In this study we have attempted to address these issues by comparing metagenomic DNA sequences from air samples with those from other environmental sources. Air samples were collected on 1 July 2009 from a hill top at Thursley Nature Reserve in Surrey, United Kingdom, a rural site, 31 miles from the nearest stretch of coastline, and on 6 July 2009 from the top of a six storey building in Stratford on the East end of London, 38 miles from the nearest coastal area. Samples were collected using the Karcher DS5500 vacuum into a liquid filled collection vessel at an air flow rate of 3.3 m3 min-1 over a 4 hour period. Samples were then concentrated and the bacterial content was investigated by PCR, cloning and sequencing of 16S rRNA genes. During the collection period on 1 July the Royston Weather Station in the South East of England recorded wind speed of 1.9 miles/hour in an Easterly direction, with no cloud cover, relative humidity of 74% and atmospheric pressure of 1021.6 mB. On 6 July wind speed was 9.8 miles/hour in a South Westerly direction, there was light cloud cover, relative humidity was 73.8% and atmospheric pressure was 1002.8 mB. Twenty cloned 16S PCR products from each air sample were sequenced. The species identification of each clone is shown in Table 1. The diversity of bacteria found at both sites was similar, with Stenotrophomona and Pedobacteria species dominating both samples. When the DNA sequences were blasted against the environmental samples database, all sequences were found to display greatest homology to metagenomic DNA from marine sources. This may suggest that the most numerous bacteria in air samples originate in the oceans. Taking account of the

  12. Predominant bacteria diversity in Chinese traditional sourdough.

    PubMed

    Zhang, Guohua; He, Guoqing

    2013-08-01

    The purpose of this study was to identify the major bacteria in Chinese traditional sourdough (CTS). Five CTS samples (Hn-87, Sx-91, Gs-107, Hf-112, and Hr-122) were collected from different Chinese steamed breads shops or private households. The total bacterial DNA was extracted from sourdough samples and sequenced using Illumina Hiseq 2000 system. Illumina tags were assigned to BLASTN server based on 16S rRNA libraries to reveal a genetic profile. Phylogenetic analysis revealed that the bacteria in traditional sourdough samples were dominated by the genera Leuconostoc and Lactobacillus. Beta diversity analysis, principal component analysis, and cluster analysis compared the bacterial differences in traditional sourdough samples. The results showed that Leuconostoc, Lactobacillus, and Weissella were the predominant genera among the 5 samples. This differentiated the sourdoughs into 3 typologies, namely, 1) Gs-107 and Sx-91, 2) Hr-122 and Hn-87, and 3) Hf-112. This study identified 3 unique major bacteria genus in CTS bread ecosystems.

  13. Multitasking SecB chaperones in bacteria.

    PubMed

    Sala, Ambre; Bordes, Patricia; Genevaux, Pierre

    2014-01-01

    Protein export in bacteria is facilitated by the canonical SecB chaperone, which binds to unfolded precursor proteins, maintains them in a translocation competent state and specifically cooperates with the translocase motor SecA to ensure their proper targeting to the Sec translocon at the cytoplasmic membrane. Besides its key contribution to the Sec pathway, SecB chaperone tasking is critical for the secretion of the Sec-independent heme-binding protein HasA and actively contributes to the cellular network of chaperones that control general proteostasis in Escherichia coli, as judged by the significant interplay found between SecB and the trigger factor, DnaK and GroEL chaperones. Although SecB is mainly a proteobacterial chaperone associated with the presence of an outer membrane and outer membrane proteins, secB-like genes are also found in Gram-positive bacteria as well as in certain phages and plasmids, thus suggesting alternative functions. In addition, a SecB-like protein is also present in the major human pathogen Mycobacterium tuberculosis where it specifically controls a stress-responsive toxin-antitoxin system. This review focuses on such very diverse chaperone functions of SecB, both in E. coli and in other unrelated bacteria.

  14. Bacteria counting method based on polyaniline/bacteria thin film.

    PubMed

    Zhihua, Li; Xuetao, Hu; Jiyong, Shi; Xiaobo, Zou; Xiaowei, Huang; Xucheng, Zhou; Tahir, Haroon Elrasheid; Holmes, Mel; Povey, Malcolm

    2016-07-15

    A simple and rapid bacteria counting method based on polyaniline (PANI)/bacteria thin film was proposed. Since the negative effects of immobilized bacteria on the deposition of PANI on glass carbon electrode (GCE), PANI/bacteria thin films containing decreased amount of PANI would be obtained when increasing the bacteria concentration. The prepared PANI/bacteria film was characterized with cyclic voltammetry (CV) technique to provide quantitative index for the determination of the bacteria count, and electrochemical impedance spectroscopy (EIS) was also performed to further investigate the difference in the PANI/bacteria films. Good linear relationship of the peak currents of the CVs and the log total count of bacteria (Bacillus subtilis) could be established using the equation Y=-30.413X+272.560 (R(2)=0.982) over the range of 5.3×10(4) to 5.3×10(8)CFUmL(-1), which also showed acceptable stability, reproducibility and switchable ability. The proposed method was feasible for simple and rapid counting of bacteria. PMID:26921555

  15. Plasmid incidence in bacteria from deep subsurface sediments

    SciTech Connect

    Fredrickson, J.K.; Hicks, R.J.; Li, S.W.; Brockman, F.J. )

    1988-12-01

    Bacteria were isolated from deep terrestrial subsurface sediments underlying the coastal plain of South Carolina. A total of 163 isolates from deep sediments, surface soil, and return drill muds were examined for plasmid DNA content and resistance to the antibiotics penicillin, ampicillin, carbenicillin, streptomycin, kanamycin, and tetracycline. MICs of Cu{sup 2+}, Cr{sup 3+}, and Hg{sup 2+} for each isolate were also determined. The overall frequency of plasmid occurrence in the subsurface bacteria was 33%. Resistance was most frequent to penicillin (70% of all isolates), ampicillin (49%), and carbenicillin (32%) and was concluded to be related to the concentrations of the individual antibiotics in the disks used for assaying resistance and to the production of low levels of {beta}-lactamase. The frequencies of resistance to penicillin and ampicillin were significantly greater for isolates bearing plasmids than for plasmidless isolates; however, resistance was not transferable to penicillin-sensitive Escherichia coli. Hybridization of subsurface bacterial plasmids and chromosomal DNA with a whole-TOL-plasmid (pWWO) probe revealed some homology of subsurface bacterial plasmid and chromosomal DNAs, indicating a potential for those bacterial to harbor catabolic genes on plasmids or chromosomes. The incidences of antibiotic resistance and MICs of metals for subsurface bacteria were significantly different from those drill mud bacteria, ruling out the possibility that bacteria from sediments were derived from drill muds.

  16. Bioprobes Based on Aptamer and Silica Fluorescent Nanoparticles for Bacteria Salmonella typhimurium Detection.

    PubMed

    Wang, Qiu-Yue; Kang, Yan-Jun

    2016-12-01

    In this study, we have developed an efficient method based on single-stranded DNA (ssDNA) aptamers along with silica fluorescence nanoparticles for bacteria Salmonella typhimurium detection. Carboxyl-modified Tris(2,2'-bipyridyl)dichlororuthenium(II) hexahydrate (RuBPY)-doped silica nanoparticles (COOH-FSiNPs) were prepared using reverse microemulsion method, and the streptavidin was conjugated to the surface of the prepared COOH-FSiNPs. The bacteria S. typhimurium was incubated with a specific ssDNA biotin-labeled aptamer, and then the aptamer-bacteria conjugates were treated with the synthetic streptavidin-conjugated silica fluorescence nanoprobes (SA-FSiNPs). The results under fluorescence microscopy show that SA-FSiNPs can be applied effectively for the labeling of bacteria S. typhimurium with great photostable property. To further verify the specificity of SA-FSiNPs out of multiple bacterial conditions, variant concentrations of bacteria mixtures composed of bacteria S. typhimurium, Escherichia coli, and Bacillus subtilis were treated with SA-FSiNPs.In addition, the feasibility of SA-FSiNPs for bacteria S. typhimurium detection in chicken samples was assessed. All the results display that the established aptamer-based nanoprobes exhibit the superiority for bacteria S. typhimurium detection, which is referentially significant for wider application prospects in pathogen detection.

  17. Visualization of yeast chromosomal DNA

    NASA Technical Reports Server (NTRS)

    Lubega, Seth

    1990-01-01

    The DNA molecule is the most significant life molecule since it codes the blue print for other structural and functional molecules of all living organisms. Agarose gel electrophoresis is now being widely used to separate DNA of virus, bacteria, and lower eukaryotes. The task was undertaken of reviewing the existing methods of DNA fractionation and microscopic visualization of individual chromosonal DNA molecules by gel electrophoresis as a basis for a proposed study to investigate the feasibility of separating DNA molecules in free fluids as an alternative to gel electrophoresis. Various techniques were studied. On the molecular level, agarose gel electrophoresis is being widely used to separate chromosomal DNA according to molecular weight. Carl and Olson separate and characterized the entire karyotype of a lab strain of Saccharomyces cerevisiae. Smith et al. and Schwartz and Koval independently reported the visualization of individual DNA molecules migrating through agarose gel matrix during electrophoresis. The techniques used by these researchers are being reviewed in the lab as a basis for the proposed studies.

  18. Evolution of Eukaryotic DNA Polymerases via Interaction Between Cells and Large DNA Viruses.

    PubMed

    Takemura, Masaharu; Yokobori, Shin-ichi; Ogata, Hiroyuki

    2015-08-01

    B-family DNA-directed DNA polymerases are DNA replication enzymes found in Eukaryota, Archaea, large DNA viruses, and in some, but not all, bacteria. Several polymerase domains are conserved among the B-family DNA polymerases from these organisms, suggesting that the B-family DNA polymerases evolved from a common ancestor. Eukaryotes retain at least three replicative B-family DNA polymerases, DNA polymerase α, δ, and ε, and one translesion B-family DNA polymerase, DNA polymerase ζ. Here, we present molecular evolutionary evidence that suggests DNA polymerase genes evolved through horizontal gene transfer between the viral and archaeal-eukaryotic lineages. Molecular phylogenetic analyses of the B-family DNA polymerases from nucleo-cytoplasmic large DNA viruses (NCLDVs), eukaryotes, and archaea suggest that different NCLDV lineages such as Poxviridae and Mimiviridae were involved in the evolution of different DNA polymerases (pol-α-, δ-, ε-, and ζ-like genes) in archaeal-eukaryotic cell lineages, putatively through horizontal gene transfer. These results support existing theories that link the evolution of NCLDVs and the origin of the eukaryotic nucleus.

  19. Beneficial bacteria inhibit cachexia

    PubMed Central

    Varian, Bernard J.; Goureshetti, Sravya; Poutahidis, Theofilos; Lakritz, Jessica R.; Levkovich, Tatiana; Kwok, Caitlin; Teliousis, Konstantinos; Ibrahim, Yassin M.; Mirabal, Sheyla; Erdman, Susan E.

    2016-01-01

    Muscle wasting, known as cachexia, is a debilitating condition associated with chronic inflammation such as during cancer. Beneficial microbes have been shown to optimize systemic inflammatory tone during good health; however, interactions between microbes and host immunity in the context of cachexia are incompletely understood. Here we use mouse models to test roles for bacteria in muscle wasting syndromes. We find that feeding of a human commensal microbe, Lactobacillus reuteri, to mice is sufficient to lower systemic indices of inflammation and inhibit cachexia. Further, the microbial muscle-building phenomenon extends to normal aging as wild type animals exhibited increased growth hormone levels and up-regulation of transcription factor Forkhead Box N1 [FoxN1] associated with thymus gland retention and longevity. Interestingly, mice with a defective FoxN1 gene (athymic nude) fail to inhibit sarcopenia after L. reuteri therapy, indicating a FoxN1-mediated mechanism. In conclusion, symbiotic bacteria may serve to stimulate FoxN1 and thymic functions that regulate inflammation, offering possible alternatives for cachexia prevention and novel insights into roles for microbiota in mammalian ontogeny and phylogeny. PMID:26933816

  20. Nitrogen control in bacteria.

    PubMed

    Merrick, M J; Edwards, R A

    1995-12-01

    Nitrogen metabolism in prokaryotes involves the coordinated expression of a large number of enzymes concerned with both utilization of extracellular nitrogen sources and intracellular biosynthesis of nitrogen-containing compounds. The control of this expression is determined by the availability of fixed nitrogen to the cell and is effected by complex regulatory networks involving regulation at both the transcriptional and posttranslational levels. While the most detailed studies to date have been carried out with enteric bacteria, there is a considerable body of evidence to show that the nitrogen regulation (ntr) systems described in the enterics extend to many other genera. Furthermore, as the range of bacteria in which the phenomenon of nitrogen control is examined is being extended, new regulatory mechanisms are also being discovered. In this review, we have attempted to summarize recent research in prokaryotic nitrogen control; to show the ubiquity of the ntr system, at least in gram-negative organisms; and to identify those areas and groups of organisms about which there is much still to learn. PMID:8531888

  1. Beneficial bacteria inhibit cachexia.

    PubMed

    Varian, Bernard J; Goureshetti, Sravya; Poutahidis, Theofilos; Lakritz, Jessica R; Levkovich, Tatiana; Kwok, Caitlin; Teliousis, Konstantinos; Ibrahim, Yassin M; Mirabal, Sheyla; Erdman, Susan E

    2016-03-15

    Muscle wasting, known as cachexia, is a debilitating condition associated with chronic inflammation such as during cancer. Beneficial microbes have been shown to optimize systemic inflammatory tone during good health; however, interactions between microbes and host immunity in the context of cachexia are incompletely understood. Here we use mouse models to test roles for bacteria in muscle wasting syndromes. We find that feeding of a human commensal microbe, Lactobacillus reuteri, to mice is sufficient to lower systemic indices of inflammation and inhibit cachexia. Further, the microbial muscle-building phenomenon extends to normal aging as wild type animals exhibited increased growth hormone levels and up-regulation of transcription factor Forkhead Box N1 [FoxN1] associated with thymus gland retention and longevity. Interestingly, mice with a defective FoxN1 gene (athymic nude) fail to inhibit sarcopenia after L. reuteri therapy, indicating a FoxN1-mediated mechanism. In conclusion, symbiotic bacteria may serve to stimulate FoxN1 and thymic functions that regulate inflammation, offering possible alternatives for cachexia prevention and novel insights into roles for microbiota in mammalian ontogeny and phylogeny. PMID:26933816

  2. Chemical communication in bacteria

    NASA Astrophysics Data System (ADS)

    Suravajhala, Srinivasa Sandeep; Saini, Deepak; Nott, Prabhu

    Luminescence in Vibrio fischeri is a model for quorum-sensing-gene-regulation in bacteria. We study luminescence response of V. fischeri to both internal and external cues at the single cell and population level. Experiments with ES114, a wild-type strain, and ainS mutant show that luminescence induction in cultures is not always proportional to cell-density and there is always a basal level of luminescence. At any given concentration of the exogenously added signals, C6-HSL and C8-HSL, luminescence per cell reaches a maximum during the exponential phase and decreases thereafter. We hypothesize that (1) C6-HSL production and LuxR activity are not proportional to cell-density, and (2) there is a shift in equilibrium from C6-HSL to C8-HSL during the later stages of growth of the culture. RT-PCR analysis of luxI and luxR shows that the expression of these genes is maximum corresponding to the highest level of luminescence. The shift in equilibrium is shown by studying competitive binding of C6-HSL and C8-HSL to LuxR. We argue that luminescence is a unicellular behaviour, and an intensive property like per cell luminescence is more important than gross luminescence of the population in understanding response of bacteria to chemical signalling. Funding from the Department of Science and Technology, India is acknowledged.

  3. Nitrogen control in bacteria.

    PubMed Central

    Merrick, M J; Edwards, R A

    1995-01-01

    Nitrogen metabolism in prokaryotes involves the coordinated expression of a large number of enzymes concerned with both utilization of extracellular nitrogen sources and intracellular biosynthesis of nitrogen-containing compounds. The control of this expression is determined by the availability of fixed nitrogen to the cell and is effected by complex regulatory networks involving regulation at both the transcriptional and posttranslational levels. While the most detailed studies to date have been carried out with enteric bacteria, there is a considerable body of evidence to show that the nitrogen regulation (ntr) systems described in the enterics extend to many other genera. Furthermore, as the range of bacteria in which the phenomenon of nitrogen control is examined is being extended, new regulatory mechanisms are also being discovered. In this review, we have attempted to summarize recent research in prokaryotic nitrogen control; to show the ubiquity of the ntr system, at least in gram-negative organisms; and to identify those areas and groups of organisms about which there is much still to learn. PMID:8531888

  4. DNA nanomachines.

    PubMed

    Bath, Jonathan; Turberfield, Andrew J

    2007-05-01

    We are learning to build synthetic molecular machinery from DNA. This research is inspired by biological systems in which individual molecules act, singly and in concert, as specialized machines: our ambition is to create new technologies to perform tasks that are currently beyond our reach. DNA nanomachines are made by self-assembly, using techniques that rely on the sequence-specific interactions that bind complementary oligonucleotides together in a double helix. They can be activated by interactions with specific signalling molecules or by changes in their environment. Devices that change state in response to an external trigger might be used for molecular sensing, intelligent drug delivery or programmable chemical synthesis. Biological molecular motors that carry cargoes within cells have inspired the construction of rudimentary DNA walkers that run along self-assembled tracks. It has even proved possible to create DNA motors that move autonomously, obtaining energy by catalysing the reaction of DNA or RNA fuels.

  5. Trail following by gliding bacteria.

    PubMed Central

    Burchard, R P

    1982-01-01

    Slime trails, which are deposited on surfaces by gliding bacteria and which serve as preferential pathways for gliding motility, were tested for the species specificity of their support of movement. Among the pairs of bacteria tested, a variety of gliding bacteria and a flagellated bacterium moved along trails of unrelated species. Thus, the trails did not serve as pheromones. Rather, they may have guided gliding elasticotactically. Some biological implications of this finding are considered. Images PMID:6811562

  6. The bacterial DnaA-trio replication origin element specifies single-stranded DNA initiator binding.

    PubMed

    Richardson, Tomas T; Harran, Omar; Murray, Heath

    2016-06-16

    DNA replication is tightly controlled to ensure accurate inheritance of genetic information. In all organisms, initiator proteins possessing AAA+ (ATPases associated with various cellular activities) domains bind replication origins to license new rounds of DNA synthesis. In bacteria the master initiator protein, DnaA, is highly conserved and has two crucial DNA binding activities. DnaA monomers recognize the replication origin (oriC) by binding double-stranded DNA sequences (DnaA-boxes); subsequently, DnaA filaments assemble and promote duplex unwinding by engaging and stretching a single DNA strand. While the specificity for duplex DnaA-boxes by DnaA has been appreciated for over 30 years, the sequence specificity for single-strand DNA binding has remained unknown. Here we identify a new indispensable bacterial replication origin element composed of a repeating trinucleotide motif that we term the DnaA-trio. We show that the function of the DnaA-trio is to stabilize DnaA filaments on a single DNA strand, thus providing essential precision to this binding mechanism. Bioinformatic analysis detects DnaA-trios in replication origins throughout the bacterial kingdom, indicating that this element is part of the core oriC structure. The discovery and characterization of the novel DnaA-trio extends our fundamental understanding of bacterial DNA replication initiation, and because of the conserved structure of AAA+ initiator proteins these findings raise the possibility of specific recognition motifs within replication origins of higher organisms. PMID:27281207

  7. Bacteria responsive antibacterial surfaces for indwelling device infections.

    PubMed

    Traba, Christian; Liang, Jun F

    2015-01-28

    Indwelling device infections now represent life-threatening circumstances as a result of the biofilms' tolerance to antibiotic treatments. Current antibiotic impregnation approaches through sustained antibiotic release have some unsolved problems which include short life-span, narrowed antibacterial spectrum, ineffectiveness towards resistant mutants, and the potential to hasten the antibiotic resistance process. In this study, bacteria responsive anti-biofilm surfaces were developed using bioactive peptides with proved activity to antibiotic resistant bacteria and biofilms. Resulting surfaces were stable under physiological conditions and in the presence of high concentrations of salts (0.5M NaCl) and biomacromolcules (1.0% DNA and 2.0% alginate), and thus showed good biocompatibility to various tissue cells. However, lytic peptide immobilized surfaces could sense bacteria adhesion and kill attached bacteria effectively and specifically, so biofilms were unable to develop on the lytic peptide immobilized surfaces. Bacteria responsive catheters remained biofilm free for up to a week. Therefore, the bacteria responsive antibacterial surfaces developed in this study represent new opportunities for indwelling device infections.

  8. Toxic effects of gold nanoparticles on Salmonella typhimurium bacteria

    PubMed Central

    Wang, Shuguang; Lawson, Rasheeda; Ray, Paresh C; Yu, Hongtao

    2013-01-01

    Nanometer-sized gold, due to its beautiful and bountiful color and unique optical properties, is a versatile material for many industrial and societal applications. We have studied the effect of gold nanoparticles on Salmonella typhimurium strain TA 102. The gold nanoparticles in solution prepared using the citrate reduction method is found not to be toxic or mutagenic but photomutagenic to the bacteria; however, careful control experiments indicate that the photomutagenicity is due to the co-existing citrate and Au3+ ions, not due to the gold nanoparticle itself. Au3+ is also found to be photomutagenic to the bacteria at concentrations lower than 1 µM, but toxic at higher concentrations. The toxicity of Au3+ is enhanced by light irradiation. The photomutagenicity of both citrate and Au3+ is likely due to the formation of free radicals, as a result of light-induced citrate decarboxylation or Au3+ oxidation of co-existing molecules. Both processes can generate free radicals that may cause DNA damage and mutation. Studies of the interaction of gold nanoparticles with the bacteria indicate that gold nanoparticles can be absorbed onto the bacteria surface but not able to penetrate the bacteria wall to enter the bacteria. PMID:21415096

  9. Bacteria associated with the bleached and cave coral Oculina patagonica.

    PubMed

    Koren, Omry; Rosenberg, Eugene

    2008-04-01

    The relative abundance of bacteria in the mucus and tissues of Oculina patagonica taken from bleached and cave (azooxanthellae) corals was determined by analyses of the 16S rRNA genes from cloned libraries of extracted DNA and from isolated colonies. The results were compared to previously published data on healthy O. patagonica. The bacterial community of bleached, cave, and healthy corals were completely different from each other. A tight cluster (>99.5% identity) of bacteria, showing 100% identity to Acinetobacter species, dominated bleached corals, comprising 25% of the 316 clones sequenced. The dominant bacterial cluster found in cave corals, representing 29% of the 97 clones sequenced, showed 98% identity to an uncultured bacterium from the Great Barrier Reef. Vibrio splendidus was the most dominant species in healthy O. patagonica. The culturable bacteria represented 0.1-1.0% of the total bacteria (SYBR Gold staining) of the corals. The most abundant culturable bacteria in bleached, cave, and healthy corals were clusters that most closely matched Microbulbifer sp., an alpha-proteobacterium previously isolated from healthy corals and an alpha-protobacterium (AB026194), respectively. Three generalizations emerge from this study on O. patagonica: (1) More bacteria are associated with coral tissue than mucus; (2) tissue and mucus populations are different; (3) bacterial populations associated with corals change dramatically when corals lack their symbiotic zooxanthellae, either as a result of the bleaching disease or when growing in the absence of light.

  10. Bacteria Responsive Antibacterial Surfaces for Indwelling Device Infections

    PubMed Central

    Traba, Christian; Liang, Jun F.

    2014-01-01

    Indwelling device infections now represents life-threatening circumstances as a result of the biofilms’ tolerance to antibiotic treatments. Current antibiotic impregnation approaches through sustained antibiotic release have some unsolved problems which include short life-span, narrowed antibacterial spectrum, ineffectiveness towards resistant mutants, and the potential to hasten the antibiotic resistance process. In this study, bacteria responsive anti-biofilm surfaces were developed using bioactive peptides with proved activity to antibiotic resistant bacteria and biofilms. Resulting surfaces were stable under physiological conditions and in the presence of high concentrations of salts (0.5 M NaCl) and biomacromolcules (1.0% DNA and 2.0% alginate), and thus showed good biocompatibility to various tissue cells. However, lytic peptide immobilized surfaces could sense bacteria adhesion and kill attached bacteria effectively and specifically, so biofilms were unable to develop on the lytic peptide immobilized surfaces. Bacteria responsive catheters remained biofilm free for up to a week. Therefore, the bacteria responsive antibacterial surfaces developed in this study represent new opportunities for indwelling device infections. PMID:25481445

  11. Bacteria responsive antibacterial surfaces for indwelling device infections.

    PubMed

    Traba, Christian; Liang, Jun F

    2015-01-28

    Indwelling device infections now represent life-threatening circumstances as a result of the biofilms' tolerance to antibiotic treatments. Current antibiotic impregnation approaches through sustained antibiotic release have some unsolved problems which include short life-span, narrowed antibacterial spectrum, ineffectiveness towards resistant mutants, and the potential to hasten the antibiotic resistance process. In this study, bacteria responsive anti-biofilm surfaces were developed using bioactive peptides with proved activity to antibiotic resistant bacteria and biofilms. Resulting surfaces were stable under physiological conditions and in the presence of high concentrations of salts (0.5M NaCl) and biomacromolcules (1.0% DNA and 2.0% alginate), and thus showed good biocompatibility to various tissue cells. However, lytic peptide immobilized surfaces could sense bacteria adhesion and kill attached bacteria effectively and specifically, so biofilms were unable to develop on the lytic peptide immobilized surfaces. Bacteria responsive catheters remained biofilm free for up to a week. Therefore, the bacteria responsive antibacterial surfaces developed in this study represent new opportunities for indwelling device infections. PMID:25481445

  12. Phage-bacteria interaction network in human oral microbiome.

    PubMed

    Wang, Jinfeng; Gao, Yuan; Zhao, Fangqing

    2016-07-01

    Although increasing knowledge suggests that bacteriophages play important roles in regulating microbial ecosystems, phage-bacteria interaction in human oral cavities remains less understood. Here we performed a metagenomic analysis to explore the composition and variation of oral dsDNA phage populations and potential phage-bacteria interaction. A total of 1,711 contigs assembled with more than 100 Gb shotgun sequencing data were annotated to 104 phages based on their best BLAST matches against the NR database. Bray-Curtis dissimilarities demonstrated that both phage and bacterial composition are highly diverse between periodontally healthy samples but show a trend towards homogenization in diseased gingivae samples. Significantly, according to the CRISPR arrays that record infection relationship between bacteria and phage, we found certain oral phages were able to invade other bacteria besides their putative bacterial hosts. These cross-infective phages were positively correlated with commensal bacteria while were negatively correlated with major periodontal pathogens, suggesting possible connection between these phages and microbial community structure in oral cavities. By characterizing phage-bacteria interaction as networks rather than exclusively pairwise predator-prey relationships, our study provides the first insight into the participation of cross-infective phages in forming human oral microbiota.

  13. Bacteriophages of methanotrophic bacteria

    SciTech Connect

    Tyutikow, F.M.; Bespalova, I.A.; Rebentish, B.A.; Aleksandrushkina, N.N.; Krivisky, A.S.

    1980-10-01

    Bacteriophages of methanotrophic bacteria have been found in 16 out of 88 studied samples (underground waters, pond water, soil, gas and oil installation waters, fermentor cultural fluids, bacterial paste, and rumen of cattle) taken in different geographic zones of the Soviet Union. Altogether, 23 phage strains were isolated. By fine structure, the phages were divided into two types (with very short or long noncontractile tails); by host range and serological properties, they fell into three types. All phages had guanine- and cytosine-rich double-stranded deoxyribonucleic acid consisting of common nitrogen bases. By all of the above-mentioned properties, all phages within each of the groups were completely identical to one another, but differed from phages of other groups.

  14. Kin Recognition in Bacteria.

    PubMed

    Wall, Daniel

    2016-09-01

    The ability of bacteria to recognize kin provides a means to form social groups. In turn these groups can lead to cooperative behaviors that surpass the ability of the individual. Kin recognition involves specific biochemical interactions between a receptor(s) and an identification molecule(s). Recognition specificity, ensuring that nonkin are excluded and kin are included, is critical and depends on the number of loci and polymorphisms involved. After recognition and biochemical perception, the common ensuing cooperative behaviors include biofilm formation, quorum responses, development, and swarming motility. Although kin recognition is a fundamental mechanism through which cells might interact, microbiologists are only beginning to explore the topic. This review considers both molecular and theoretical aspects of bacterial kin recognition. Consideration is also given to bacterial diversity, genetic relatedness, kin selection theory, and mechanisms of recognition. PMID:27359217

  15. [DNA computing].

    PubMed

    Błasiak, Janusz; Krasiński, Tadeusz; Popławski, Tomasz; Sakowski, Sebastian

    2011-01-01

    Biocomputers can be an alternative for traditional "silicon-based" computers, which continuous development may be limited due to further miniaturization (imposed by the Heisenberg Uncertainty Principle) and increasing the amount of information between the central processing unit and the main memory (von Neuman bottleneck). The idea of DNA computing came true for the first time in 1994, when Adleman solved the Hamiltonian Path Problem using short DNA oligomers and DNA ligase. In the early 2000s a series of biocomputer models was presented with a seminal work of Shapiro and his colleguas who presented molecular 2 state finite automaton, in which the restriction enzyme, FokI, constituted hardware and short DNA oligomers were software as well as input/output signals. DNA molecules provided also energy for this machine. DNA computing can be exploited in many applications, from study on the gene expression pattern to diagnosis and therapy of cancer. The idea of DNA computing is still in progress in research both in vitro and in vivo and at least promising results of these research allow to have a hope for a breakthrough in the computer science. PMID:21735816

  16. Scarce evidence of yogurt lactic acid bacteria in human feces after daily yogurt consumption by healthy volunteers.

    PubMed

    del Campo, Rosa; Bravo, Daniel; Cantón, Rafael; Ruiz-Garbajosa, Patricia; García-Albiach, Raimundo; Montesi-Libois, Alejandra; Yuste, Francisco-Javier; Abraira, Victor; Baquero, Fernando

    2005-01-01

    In a double-blind prospective study including 114 healthy young volunteers, the presence in human feces of the yogurt organisms Lactobacillus delbrueckii and Streptococcus thermophilus after repeated yogurt consumption (15 days) was analyzed by culture, specific PCR, and DNA hybridization of total fecal DNA. Detection of yogurt lactic acid bacteria in total fecal DNA by bacterial culture and PCR assay was consistently negative. DNA compatible with yogurt bacteria was found by hybridization experiments in only 10 (10.52%) of 96 individuals after consumption of fresh yogurt and in 2 (2.10%) of 96 individuals after consumption of pasteurized yogurt (P = 0.01).

  17. Isolation of marine bacteria by in situ culture on media-supplemented polyurethane foam.

    PubMed

    Yasumoto-Hirose, Mina; Nishijima, Miyuki; Ngirchechol, Metiek Kimie; Kanoh, Kaneo; Shizuri, Yoshikazu; Miki, Wataru

    2006-01-01

    Polyurethane foam (PUF) supplemented with various agar media was used in situ to trap marine bacteria and it consequently provided a substrate on which they could be cultivated while exposed to natural seawater in the coral reef area. The bacterial population on the PUF blocks was analyzed by denaturing gradient gel electrophoresis (DGGE) of polymerase chain reaction (PCR)-amplified 16S rDNA fragments. Changing the composition of the cultivation medium in the PUF blocks and selecting different sampling sites resulted in different bacteria being detected on the PUF blocks. For example, iron-utilizing (IU) bacteria, siderophore-producing (SP) bacteria, and petroleum-degrading (PD) bacteria were isolated from PUF blocks and it was discovered that IU and SP contained iron and PD contained hydrocarbon. This method opens up the possibility for isolating novel and useful marine bacteria.

  18. Development of Mucosal Vaccines Based on Lactic Acid Bacteria

    NASA Astrophysics Data System (ADS)

    Bermúdez-Humarán, Luis G.; Innocentin, Silvia; Lefèvre, Francois; Chatel, Jean-Marc; Langella, Philippe

    Today, sufficient data are available to support the use of lactic acid bacteria (LAB), notably lactococci and lactobacilli, as delivery vehicles for the development of new mucosal vaccines. These non-pathogenic Gram-positive bacteria have been safely consumed by humans for centuries in fermented foods. They thus constitute an attractive alternative to the attenuated pathogens (most popular live vectors actually studied) which could recover their pathogenic potential and are thus not totally safe for use in humans. This chapter reviews the current research and advances in the use of LAB as live delivery vectors of proteins of interest for the development of new safe mucosal vaccines. The use of LAB as DNA vaccine vehicles to deliver DNA directly to antigen-presenting cells of the immune system is also discussed.

  19. cpDNA microsatellite markers for Lemna minor (Araceae): Phylogeographic implications1

    PubMed Central

    Wani, Gowher A.; Shah, Manzoor A.; Reshi, Zafar A.; Atangana, Alain R.; Khasa, Damase P.

    2014-01-01

    • Premise of the study: A lack of genetic markers impedes our understanding of the population biology of Lemna minor. Thus, the development of appropriate genetic markers for L. minor promises to be highly useful for population genetic studies and for addressing other life history questions regarding the species. • Methods and Results: For the first time, we characterized nine polymorphic and 24 monomorphic chloroplast microsatellite markers in L. minor using DNA samples of 26 individuals sampled from five populations in Kashmir and of 17 individuals from three populations in Quebec. Initially, we designed 33 primer pairs, which were tested on genomic DNA from natural populations. Nine loci provided markers with two alleles. Based on genotyping of the chloroplast DNA fragments from 43 sampled individuals, we identified one haplotype in Quebec and 11 haplotypes in Kashmir, of which one occurs in 56% of the genotypes, one in 8%, and nine in 4%, respectively. There was a maximum of two alleles per locus. • Conclusions: These new chloroplast microsatellite markers for L. minor and haplotype distribution patterns indicate a complex phylogeographic history that merits further investigation. PMID:25202636

  20.  Acetobacter bacteria are found in Zhenjiang vinegar grains.

    PubMed

    Wang, C Y; Zhang, J; Gui, Z Z

    2015-01-01

    Zhenjiang vinegar, the grains of which contain a unique microbial flora, is one of the four famous traditional Chinese vinegars. We investigated the components of Zhenjiang vinegar grains. Unique acetic acid bacteria were randomly isolated from Zhenjiang vinegar grains, and the obtained strains were qualitatively analyzed to compare their capacities for acetate decomposition and acid production. Acetic acid bacteria with a high acid-producing rate were identified by 16S rDNA sequencing, and further confirmation was performed using the Basic Local Alignment Search Tool comparison method. Six significant strains of acetic acid bacteria were isolated. Qualitative analysis showed that these strains produced no brown precipitate and had a capacity for acetate decomposition. Based on physiological and biochemical evaluation, the two strains with the highest acid yield were sequenced, and the results identified strain W1 as Acetobacter aceti and strain W6 as A. pasteurianus.

  1. Dancing DNA.

    ERIC Educational Resources Information Center

    Pennisi, Elizabeth

    1991-01-01

    An imaging technique that uses fluorescent dyes and allows scientists to track DNA as it moves through gels or in solution is described. The importance, opportunities, and implications of this technique are discussed. (KR)

  2. DNA Dynamics.

    ERIC Educational Resources Information Center

    Warren, Michael D.

    1997-01-01

    Explains a method to enable students to understand DNA and protein synthesis using model-building and role-playing. Acquaints students with the triplet code and transcription. Includes copies of the charts used in this technique. (DDR)

  3. Swimming bacteria in liquid crystal

    NASA Astrophysics Data System (ADS)

    Sokolov, Andrey; Zhou, Shuang; Aranson, Igor; Lavrentovich, Oleg

    2014-03-01

    Dynamics of swimming bacteria can be very complex due to the interaction between the bacteria and the fluid, especially when the suspending fluid is non-Newtonian. Placement of swimming bacteria in lyotropic liquid crystal produces a new class of active materials by combining features of two seemingly incompatible constituents: self-propelled live bacteria and ordered liquid crystals. Here we present fundamentally new phenomena caused by the coupling between direction of bacterial swimming, bacteria-triggered flows and director orientations. Locomotion of bacteria may locally reduce the degree of order in liquid crystal or even trigger nematic-isotropic phase transition. Microscopic flows generated by bacterial flagella disturb director orientation. Emerged birefringence patterns allow direct optical observation and quantitative characterization of flagella dynamics. At high concentration of bacteria we observed the emergence of self-organized periodic texture caused by bacteria swimming. Our work sheds new light on self-organization in hybrid bio-mechanical systems and can lead to valuable biomedical applications. Was supported by the US DOE, Office of Basic Energy Sciences, Division of Materials Science and Engineering, under the Contract No. DE AC02-06CH11357.

  4. Multilocus sequence typing of bacteria.

    PubMed

    Maiden, Martin C J

    2006-01-01

    Multilocus sequence typing (MLST) was proposed in 1998 as a portable, universal, and definitive method for characterizing bacteria, using the human pathogen Neisseria meningitidis as an example. In addition to providing a standardized approach to data collection, by examining the nucleotide sequences of multiple loci encoding housekeeping genes, or fragments of them, MLST data are made freely available over the Internet to ensure that a uniform nomenclature is readily available to all those interested in categorizing bacteria. At the time of writing, over thirty MLST schemes have been published and made available on the Internet, mostly for pathogenic bacteria, although there are schemes for pathogenic fungi and some nonpathogenic bacteria. MLST data have been employed in epidemiological investigations of various scales and in studies of the population biology, pathogenicity, and evolution of bacteria. The increasing speed and reduced cost of nucleotide sequence determination, together with improved web-based databases and analysis tools, present the prospect of increasingly wide application of MLST.

  5. DNA Adductomics

    PubMed Central

    2015-01-01

    Systems toxicology is a broad-based approach to describe many of the toxicological features that occur within a living system under stress or subjected to exogenous or endogenous exposures. The ultimate goal is to capture an overview of all exposures and the ensuing biological responses of the body. The term exposome has been employed to refer to the totality of all exposures, and systems toxicology investigates how the exposome influences health effects and consequences of exposures over a lifetime. The tools to advance systems toxicology include high-throughput transcriptomics, proteomics, metabolomics, and adductomics, which is still in its infancy. A well-established methodology for the comprehensive measurement of DNA damage resulting from every day exposures is not fully developed. During the past several decades, the 32P-postlabeling technique has been employed to screen the damage to DNA induced by multiple classes of genotoxicants; however, more robust, specific, and quantitative methods have been sought to identify and quantify DNA adducts. Although triple quadrupole and ion trap mass spectrometry, particularly when using multistage scanning (LC–MSn), have shown promise in the field of DNA adductomics, it is anticipated that high-resolution and accurate-mass LC–MSn instrumentation will play a major role in assessing global DNA damage. Targeted adductomics should also benefit greatly from improved triple quadrupole technology. Once the analytical MS methods are fully mature, DNA adductomics along with other -omics tools will contribute greatly to the field of systems toxicology. PMID:24437709

  6. The awakening of DNA repair at Yale.

    PubMed

    Hanawalt, Philip C

    2013-12-13

    As a graduate student with Professor Richard Setlow at Yale in the late 1950s, I studied the effects of ultraviolet and visible light on the syntheses of DNA, RNA, and protein in bacteria. I reflect upon my research in the Yale Biophysics Department, my subsequent postdoctoral experiences, and the eventual analyses in the laboratories of Setlow, Paul Howard-Flanders, and myself that constituted the discovery of the ubiquitous pathway of DNA excision repair in the early 1960s. I then offer a brief perspective on a few more recent developments in the burgeoning DNA repair field and their relationships to human disease.

  7. The Awakening of DNA Repair at Yale

    PubMed Central

    Hanawalt, Philip C.

    2013-01-01

    As a graduate student with Professor Richard Setlow at Yale in the late 1950s, I studied the effects of ultraviolet and visible light on the syntheses of DNA, RNA, and protein in bacteria. I reflect upon my research in the Yale Biophysics Department, my subsequent postdoctoral experiences, and the eventual analyses in the laboratories of Setlow, Paul Howard-Flanders, and myself that constituted the discovery of the ubiquitous pathway of DNA excision repair in the early 1960s. I then offer a brief perspective on a few more recent developments in the burgeoning DNA repair field and their relationships to human disease. PMID:24348216

  8. Endosymbiotic bacteria in the parasitic ciliate Ichthyophthirius multifiliis.

    PubMed

    Sun, H Y; Noe, J; Barber, J; Coyne, R S; Cassidy-Hanley, D; Clark, T G; Findly, R C; Dickerson, H W

    2009-12-01

    Endosymbiotic bacteria were identified in the parasitic ciliate Ichthyophthirius multifiliis, a common pathogen of freshwater fish. PCR amplification of DNA prepared from two isolates of I. multifiliis, using primers that bind conserved sequences in bacterial 16S rRNA genes, generated an approximately 1,460-bp DNA product, which was cloned and sequenced. Sequence analysis demonstrated that 16S rRNA gene sequences from three classes of bacteria were present in the PCR product. These included Alphaproteobacteria (Rickettsiales), Sphingobacteria, and Flavobacterium columnare. DAPI (4',6-diamidino-2-phenylindole) staining showed endosymbionts dispersed throughout the cytoplasm of trophonts and, in most, but not all theronts. Endosymbionts were observed by transmission electron microscopy in the cytoplasm, surrounded by a prominent, electron-translucent halo characteristic of Rickettsia. Fluorescence in situ hybridization demonstrated that bacteria from the Rickettsiales and Sphingobacteriales classes are endosymbionts of I. multifiliis, found in the cytoplasm, but not in the macronucleus or micronucleus. In contrast, F. columnare was not detected by fluorescence in situ hybridization. It likely adheres to I. multifiliis through association with cilia. The role that endosymbiotic bacteria play in the life history of I. multifiliis is not known.

  9. Recombinant DNA products: Insulin, interferon and growth hormone

    SciTech Connect

    Bollon, A.P.

    1984-01-01

    This book provides the discussion of products of biotechnology of recombinant DNA. The contents include: Recombinant DNA techniques; isolation, cloning, and expression of genes; from somatostatin to human insulin; yeast; an alternative organism for foreign protein production; background in human interferon; preclinical assessment of biological properties of recombinant DNA derived human interferons; human clinical trials of bacteria-derived human ..cap alpha.. interferon.f large scale production of human alpha interferon from bacteria; direct expression of human growth hormone in escherichia coli with the lipoprotein promoter; biological actions in humans of recombinant DNA synthesized human growth hormone; NIH guidelines for research involving recombinant DNA molecules; appendix; viral vectors and the NHY guidelines; FDA's role in approval and regulation of recombinant DNA drugs; and index.

  10. DNA: Polymer and molecular code

    NASA Astrophysics Data System (ADS)

    Shivashankar, G. V.

    1999-10-01

    gene expression a prime example of a biological code. We developed a novel method of making DNA micro- arrays, the so-called DNA chip. Using the optical tweezer concept, we were able to pattern biomolecules on a solid substrate, developing a new type of sub-micron laser lithography. A laser beam is focused onto a thin gold film on a glass substrate. Laser ablation of gold results in local aggregation of nanometer scale beads conjugated with small DNA oligonucleotides, with sub-micron resolution. This leads to specific detection of cDNA and RNA molecules. We built a simple micro-array fabrication and detection in the laboratory, based on this method, to probe addressable pools (genes, proteins or antibodies). We have lately used molecular beacons (single stranded DNA with a stem-loop structure containing a fluorophore and quencher), for the direct detection of unlabelled mRNA. As a first step towards a study of the dynamics of the biological code, we have begun to examine the patterns of gene expression during virus (T7 phage) infection of E-coli bacteria.

  11. Physical mode of bacteria and virus coevolution

    NASA Astrophysics Data System (ADS)

    Han, Pu; Niestemski, Liang; Deem, Michael

    2013-03-01

    Single-cell hosts such as bacteria or archaea possess an adaptive, heritable immune system that protects them from viral invasion. This system, known as the CRISPR-Cas system, allows the host to recognize and incorporate short foreign DNA or RNA sequences from viruses or plasmids. The sequences form what are called ``spacers'' in the CRISPR. Spacers in the CRISPR loci provide a record of the host and predator coevolution history. We develop a physical model to study the dynamics of this coevolution due to immune pressure. Hosts and viruses reproduce, die, and evolve due to viral infection pressure, host immune pressure, and mutation. We will discuss the differing effects of point mutation and recombination on CRISPR evolution. We will also discuss the effect of different spacer deletion mechanisms. We will describe population structure of hosts and viruses, how spacer diversity depends on position within CRISPR, and match of the CRISPR spacers to the virus population.

  12. SOS, the formidable strategy of bacteria against aggressions.

    PubMed

    Baharoglu, Zeynep; Mazel, Didier

    2014-11-01

    The presence of an abnormal amount of single-stranded DNA in the bacterial cell constitutes a genotoxic alarm signal that induces the SOS response, a broad regulatory network found in most bacterial species to address DNA damage. The aim of this review was to point out that beyond being a repair process, SOS induction leads to a very strong but transient response to genotoxic stress, during which bacteria can rearrange and mutate their genome, induce several phenotypic changes through differential regulation of genes, and sometimes acquire characteristics that potentiate bacterial survival and adaptation to changing environments. We review here the causes and consequences of SOS induction, but also how this response can be modulated under various circumstances and how it is connected to the network of other important stress responses. In the first section, we review articles describing the induction of the SOS response at the molecular level. The second section discusses consequences of this induction in terms of DNA repair, changes in the genome and gene expression, and sharing of genomic information, with their effects on the bacteria's life and evolution. The third section is about the fine tuning of this response to fit with the bacteria's 'needs'. Finally, we discuss recent findings linking the SOS response to other stress responses. Under these perspectives, SOS can be perceived as a powerful bacterial strategy against aggressions.

  13. Sociomicrobiology and pathogenic bacteria

    PubMed Central

    Xavier, Joao B.

    2015-01-01

    The study of microbial pathogenesis has been primarily a reductionist science since Koch's principles. Reductionist approaches are essential to identify the causal agents of infectious disease, their molecular mechanisms of action and potential drug targets, and much of medicine's success in the treatment of infectious disease comes from this approach. But many bacterial caused diseases cannot be explained by focusing on a single bacterium. Many aspects of bacterial pathogenesis will benefit from a more holistic approach that takes into account social interaction within bacteria of the same species and between different species in consortia such as the human microbiome. I discuss recent advances in the emerging discipline of sociomicrobiology and how it provides a framework to dissect microbial interactions in single and multispecies communities without compromising mechanistic detail. The study of bacterial pathogenesis can benefit greatly from incorporating concepts from other disciplines such as social evolution theory and microbial ecology where communities, their interactions with hosts and with the environment play key roles. PMID:27337482

  14. Cultivation-independent identification of candidate dehalorespiring bacteria in tetrachloroethylene degradation.

    PubMed

    Yamasaki, Shouhei; Nomura, Nobuhiko; Nakajima, Toshiaki; Uchiyama, Hiroo

    2012-07-17

    Tetrachloroethylene (PCE) is one of the major pollutants and is degraded by dissimilation by dehalorespiring bacteria. The dehalorespiring bacteria are anaerobic, and most cannot be cultured by conventional agar plating methods. Therefore, to identify the dehalorespiring bacteria that dissimilatively degrade PCE, a cultivation-independent method is required. To achieve accurate and detailed analysis of the bacteria, we developed a novel stable isotope probing (SIP) method. This technique involves 2 steps, namely, a labeling step, in which a labeled carbon source is incorporated into the sample's DNA, and an analysis step, in which the DNA is isolated, fractionated, and analyzed by polymerase chain reaction denaturing gradient gel electrophoresis (PCR-DGGE). Subsequently, 16S rRNA sequencing and phylogenetic analysis were performed to identify the bacteria. Initially, we examined the effectiveness of this method by using Dehalococcoides ethenogenes 195 consortium as a defined model system. The result indicated the method was able to correctly identify the dehalorespiring bacteria D. ethenogenes 195 from the consortium. Moreover, in an artificially contaminated microcosm experiment, we confirmed that the method was able to identify the indigenous dehalorespiring bacteria Dehalobacter sp. Thus, we concluded that this novel method was a feasible tool to identify dehalorespiring bacteria in natural environments.

  15. What Is Mitochondrial DNA?

    MedlinePlus

    ... DNA What is mitochondrial DNA? What is mitochondrial DNA? Although most DNA is packaged in chromosomes within ... proteins. For more information about mitochondria and mitochondrial DNA: Molecular Expressions, a web site from the Florida ...

  16. A direct pre-screen for marine bacteria producing compounds inhibiting quorum sensing reveals diverse planktonic bacteria that are bioactive.

    PubMed

    Linthorne, Jamie S; Chang, Barbara J; Flematti, Gavin R; Ghisalberti, Emilio L; Sutton, David C

    2015-02-01

    A promising new strategy in antibacterial research is inhibition of the bacterial communication system termed quorum sensing. In this study, a novel and rapid pre-screening method was developed to detect the production of chemical inhibitors of this system (quorum-quenching compounds) by bacteria isolated from marine and estuarine waters. This method involves direct screening of mixed populations on an agar plate, facilitating specific isolation of bioactive colonies. The assay showed that between 4 and 46 % of culturable bacteria from various samples were bioactive, and of the 95 selectively isolated bacteria, 93.7 % inhibited Vibrio harveyi bioluminescence without inhibiting growth, indicating potential production of quorum-quenching compounds. Of the active isolates, 21 % showed further activity against quorum-sensing-regulated pigment production by Serratia marcescens. The majority of bioactive isolates were identified by 16S ribosomal DNA (rDNA) amplification and sequencing as belonging to the genera Vibrio and Pseudoalteromonas. Extracts of two strongly bioactive Pseudoalteromonas isolates (K1 and B2) were quantitatively assessed for inhibition of growth and quorum-sensing-regulated processes in V. harveyi, S. marcescens and Chromobacterium violaceum. Extracts of the isolates reduced V. harveyi bioluminescence by as much as 98 % and C. violaceum pigment production by 36 % at concentrations which had no adverse effect on growth. The activity found in the extracts indicated that the isolates may produce quorum-quenching compounds. This study further supports the suggestion that quorum quenching may be a common attribute among culturable planktonic marine and estuarine bacteria.

  17. Freeing Water from Viruses and Bacteria

    NASA Technical Reports Server (NTRS)

    2004-01-01

    Four years ago, Argonide Corporation, a company focused on the research, production, and marketing of specialty nano materials, was seeking to develop applications for its NanoCeram[R] fibers. Only 2 nanometers in diameter, these nano aluminum oxide fibers possessed unusual bio-adhesive properties. When formulated into a filter material, the electropositive fibers attracted and retained electro-negative particles such as bacteria and viruses in water-based solutions. This technology caught the interest of NASA as a possible solution for improved water filtration in space cabins. NASA's Johnson Space Center awarded Sanford, Florida-based Argonide a Phase I Small Business Innovation Research (SBIR) contract to determine the feasibility of using the company's filter for purifying recycled space cabin water. Since viruses and bacteria can be carried aboard space cabins by space crews, the ability to detect and remove these harmful substances is a concern for NASA. The Space Agency also desired an improved filter to polish the effluent from condensed and waste water, producing potable drinking water. During its Phase I partnership with NASA, Argonide developed a laboratory-size filter capable of removing greater than 99.9999 percent of bacteria and viruses from water at flow rates more than 200 times faster than virus-rated membranes that remove particles by sieving. Since the new filter s pore size is rather large compared to other membranes, it is also less susceptible to clogging by small particles. In September 2002, Argonide began a Phase II SBIR project with Johnson to develop a full-size cartridge capable of serving a full space crew. This effort, which is still ongoing, enabled the company to demonstrate that its filter media is an efficient absorbent for DNA and RNA.

  18. Bacteria-mediated disease therapy.

    PubMed

    Yu, Hao

    2011-12-01

    Laboratory bacteria have been model systems for studying gene function or developing molecular biotechnologies; industrial bacteria have been used to produce drugs, foods, fuel etc.; while pathogenic bacteria have been the subject of infectious diseases studies with aim of controlling them. Recently, with our increasing knowledge in molecular biology, bacterial genetics, and immunology, bacteria have gained increased interest in therapeutic applications. This review will summarize recent advances toward applying bacteria in treatment of diseases, such as inflammatory disease, cancer, and virus infection. Probiotics have long been appreciated for their immunomodulatory effect, which are discussed in the anti-inflammatory section. Anaerobic bacteria have been applied for destructing tumor cells or used as a magic bullet to specifically deliver therapeutic agents to tumor tissue. Examples and strategies of applying those tumor-targeting bacteria are discussed in the antitumor section. Finally, the pioneer study of applying Salmonella in delivering RNase P that was specifically engineered to target essential mRNAs of virus is summarized.

  19. Interactions between Diatoms and Bacteria

    PubMed Central

    Amin, Shady A.; Parker, Micaela S.

    2012-01-01

    Summary: Diatoms and bacteria have cooccurred in common habitats for hundreds of millions of years, thus fostering specific associations and interactions with global biogeochemical consequences. Diatoms are responsible for one-fifth of the photosynthesis on Earth, while bacteria remineralize a large portion of this fixed carbon in the oceans. Through their coexistence, diatoms and bacteria cycle nutrients between oxidized and reduced states, impacting bioavailability and ultimately feeding higher trophic levels. Here we present an overview of how diatoms and bacteria interact and the implications of these interactions. We emphasize that heterotrophic bacteria in the oceans that are consistently associated with diatoms are confined to two phyla. These consistent bacterial associations result from encounter mechanisms that occur within a microscale environment surrounding a diatom cell. We review signaling mechanisms that occur in this microenvironment to pave the way for specific interactions. Finally, we discuss known interactions between diatoms and bacteria and exciting new directions and research opportunities in this field. Throughout the review, we emphasize new technological advances that will help in the discovery of new interactions. Deciphering the languages of diatoms and bacteria and how they interact will inform our understanding of the role these organisms have in shaping the ocean and how these interactions may change in future oceans. PMID:22933565

  20. Elimination of bioweapons agents from forensic samples during extraction of human DNA.

    PubMed

    Timbers, Jason; Wilkinson, Della; Hause, Christine C; Smith, Myron L; Zaidi, Mohsin A; Laframboise, Denis; Wright, Kathryn E

    2014-11-01

    Collection of DNA for genetic profiling is a powerful means for the identification of individuals responsible for crimes and terrorist acts. Biologic hazards, such as bacteria, endospores, toxins, and viruses, could contaminate sites of terrorist activities and thus could be present in samples collected for profiling. The fate of these hazards during DNA isolation has not been thoroughly examined. Our goals were to determine whether the DNA extraction process used by the Royal Canadian Mounted Police eliminates or neutralizes these agents and if not, to establish methods that render samples safe without compromising the human DNA. Our results show that bacteria, viruses, and toxins were reduced to undetectable levels during DNA extraction, but endospores remained viable. Filtration of samples after DNA isolation eliminated viable spores from the samples but left DNA intact. We also demonstrated that contamination of samples with some bacteria, endospores, and toxins for longer than 1 h compromised the ability to complete genetic profiling. PMID:25069670

  1. Ancient DNA.

    PubMed

    Willerslev, Eske; Cooper, Alan

    2005-01-01

    In the past two decades, ancient DNA research has progressed from the retrieval of small fragments of mitochondrial DNA from a few late Holocene specimens, to large-scale studies of ancient populations, phenotypically important nuclear loci, and even whole mitochondrial genome sequences of extinct species. However, the field is still regularly marred by erroneous reports, which underestimate the extent of contamination within laboratories and samples themselves. An improved understanding of these processes and the effects of damage on ancient DNA templates has started to provide a more robust basis for research. Recent methodological advances have included the characterization of Pleistocene mammal populations and discoveries of DNA preserved in ancient sediments. Increasingly, ancient genetic information is providing a unique means to test assumptions used in evolutionary and population genetics studies to reconstruct the past. Initial results have revealed surprisingly complex population histories, and indicate that modern phylogeographic studies may give misleading impressions about even the recent evolutionary past. With the advent and uptake of appropriate methodologies, ancient DNA is now positioned to become a powerful tool in biological research and is also evolving new and unexpected uses, such as in the search for extinct or extant life in the deep biosphere and on other planets.

  2. Ancient DNA

    PubMed Central

    Willerslev, Eske; Cooper, Alan

    2004-01-01

    In the past two decades, ancient DNA research has progressed from the retrieval of small fragments of mitochondrial DNA from a few late Holocene specimens, to large-scale studies of ancient populations, phenotypically important nuclear loci, and even whole mitochondrial genome sequences of extinct species. However, the field is still regularly marred by erroneous reports, which underestimate the extent of contamination within laboratories and samples themselves. An improved understanding of these processes and the effects of damage on ancient DNA templates has started to provide a more robust basis for research. Recent methodological advances have included the characterization of Pleistocene mammal populations and discoveries of DNA preserved in ancient sediments. Increasingly, ancient genetic information is providing a unique means to test assumptions used in evolutionary and population genetics studies to reconstruct the past. Initial results have revealed surprisingly complex population histories, and indicate that modern phylogeographic studies may give misleading impressions about even the recent evolutionary past. With the advent and uptake of appropriate methodologies, ancient DNA is now positioned to become a powerful tool in biological research and is also evolving new and unexpected uses, such as in the search for extinct or extant life in the deep biosphere and on other planets. PMID:15875564

  3. DNA vaccines

    NASA Astrophysics Data System (ADS)

    Gregersen, Jens-Peter

    2001-12-01

    Immunization by genes encoding immunogens, rather than with the immunogen itself, has opened up new possibilities for vaccine research and development and offers chances for new applications and indications for future vaccines. The underlying mechanisms of antigen processing, immune presentation and regulation of immune responses raise high expectations for new and more effective prophylactic or therapeutic vaccines, particularly for vaccines against chronic or persistent infectious diseases and tumors. Our current knowledge and experience of DNA vaccination is summarized and critically reviewed with particular attention to basic immunological mechanisms, the construction of plasmids, screening for protective immunogens to be encoded by these plasmids, modes of application, pharmacokinetics, safety and immunotoxicological aspects. DNA vaccines have the potential to accelerate the research phase of new vaccines and to improve the chances of success, since finding new immunogens with the desired properties is at least technically less demanding than for conventional vaccines. However, on the way to innovative vaccine products, several hurdles have to be overcome. The efficacy of DNA vaccines in humans appears to be much less than indicated by early studies in mice. Open questions remain concerning the persistence and distribution of inoculated plasmid DNA in vivo, its potential to express antigens inappropriately, or the potentially deleterious ability to insert genes into the host cell's genome. Furthermore, the possibility of inducing immunotolerance or autoimmune diseases also needs to be investigated more thoroughly, in order to arrive at a well-founded consensus, which justifies the widespread application of DNA vaccines in a healthy population.

  4. Sampling bacteria with a laser

    NASA Astrophysics Data System (ADS)

    Schwarzwälder, Kordula; Rutschmann, Peter

    2014-05-01

    Water quality is a topic of high interest and it's getting more and more important due to climate change and the implementation of European Water Framework Directive (WFD). One point of interest here is the inflow of bacteria into a river caused by combined sewer overflows which lead untreated wastewater including bacteria directly into a river. These bacteria remain in the river for a certain time, they settle down and can be remobilised again. In our study we want to investigate these processes of sedimentation and resuspension and use the results for the development of a software module coupled with the software Flow3D. Thereby we should be able to simulate and therefore predict the water quality influenced by combined sewer overflows. Hence we need to get information about the bacteria transport and fate. We need to know about the size of the bacteria or of the bacteria clumps and the size of the particles the bacteria are attached to. The agglomerates lead to different characteristics and velocities of settlement. The timespan during this bacteria can be detected in the bulk phase depends on many factors like the intensity of UV light, turbidity of the water, the temperature of the water, if there are grazers and a lot more. The size, density and composition of the agglomerates is just a part of all these influencing factors, but it is extremely difficult to differ between the other effects if we have no information about the simple sedimentation in default of these basic information. However we have a big problem getting the data. The chaining between bacteria or bacteria and particles is not too strong, so filtering the water to get a sieving curve may destroy these connections. We did some experiments similar to PIV (particle image velocimetry) measurements and evaluated the pictures with a macro written for the software ImageJ. Doing so we were able to get the concentration of bacteria in the water and collect information about the size of the bacteria. We

  5. DNA Repair and Genome Maintenance in Bacillus subtilis

    PubMed Central

    Lenhart, Justin S.; Schroeder, Jeremy W.; Walsh, Brian W.

    2012-01-01

    Summary: From microbes to multicellular eukaryotic organisms, all cells contain pathways responsible for genome maintenance. DNA replication allows for the faithful duplication of the genome, whereas DNA repair pathways preserve DNA integrity in response to damage originating from endogenous and exogenous sources. The basic pathways important for DNA replication and repair are often conserved throughout biology. In bacteria, high-fidelity repair is balanced with low-fidelity repair and mutagenesis. Such a balance is important for maintaining viability while providing an opportunity for the advantageous selection of mutations when faced with a changing environment. Over the last decade, studies of DNA repair pathways in bacteria have demonstrated considerable differences between Gram-positive and Gram-negative organisms. Here we review and discuss the DNA repair, genome maintenance, and DNA damage checkpoint pathways of the Gram-positive bacterium Bacillus subtilis. We present their molecular mechanisms and compare the functions and regulation of several pathways with known information on other organisms. We also discuss DNA repair during different growth phases and the developmental program of sporulation. In summary, we present a review of the function, regulation, and molecular mechanisms of DNA repair and mutagenesis in Gram-positive bacteria, with a strong emphasis on B. subtilis. PMID:22933559

  6. DNA topoisomerases.

    PubMed

    Wang, J C

    1996-01-01

    The various problems of disentangling DNA strands or duplexes in a cell are all rooted in the double-helical structure of DNA. Three distinct subfamilies of enzymes, known as the DNA topoisomerases, have evolved to solve these problems. This review focuses on work in the past decade on the mechanisms and cellular functions of these enzymes. Newly discovered members and recent biochemical and structural results are reviewed, and mechanistic implications of these results are summarized. The primary cellular functions of these enzymes, including their roles in replication, transcription, chromosome condensation, and the maintenance of genome stability, are then discussed. The review ends with a summary of the regulation of the cellular levels of these enzymes and a discussion of their association with other cellular proteins.

  7. Distribution and Diversity of Symbiotic Thermophiles, Symbiobacterium thermophilum and Related Bacteria, in Natural Environments

    PubMed Central

    Ueda, Kenji; Ohno, Michiyo; Yamamoto, Kaori; Nara, Hanae; Mori, Yujiro; Shimada, Masafumi; Hayashi, Masahiko; Oida, Hanako; Terashima, Yuko; Nagata, Mitsuyo; Beppu, Teruhiko

    2001-01-01

    Symbiobacterium thermophilum is a tryptophanase-positive thermophile which shows normal growth only in coculture with its supporting bacteria. Analysis of the 16S rRNA gene (rDNA) indicated that the bacterium belongs to a novel phylogenetic branch at the outermost position of the gram-positive bacterial group without clustering to any other known genus. Here we describe the distribution and diversity of S. thermophilum and related bacteria in the environment. Thermostable tryptophanase activity and amplification of the specific 16S rDNA fragment were effectively employed to detect the presence of Symbiobacterium. Enrichment with kanamycin raised detection sensitivity. Mixed cultures of thermophiles containing Symbiobacterium species were frequently obtained from compost, soil, animal feces, and contents in the intestinal tracts, as well as feeds. Phylogenetic analysis and denaturing gradient gel electrophoresis of the specific 16S rDNA amplicons revealed a diversity of this group of bacteria in the environment. PMID:11525967

  8. Functions of single-strand DNA-binding proteins in DNA replication, recombination, and repair.

    PubMed

    Marceau, Aimee H

    2012-01-01

    Double-stranded (ds) DNA contains all of the necessary genetic information, although practical use of this information requires unwinding of the duplex DNA. DNA unwinding creates single-stranded (ss) DNA intermediates that serve as templates for myriad cellular functions. Exposure of ssDNA presents several problems to the cell. First, ssDNA is thermodynamically less stable than dsDNA, which leads to spontaneous formation of duplex secondary structures that impede genome maintenance processes. Second, relative to dsDNA, ssDNA is hypersensitive to chemical and nucleolytic attacks that can cause damage to the genome. Cells deal with these potential problems by encoding specialized ssDNA-binding proteins (SSBs) that bind to and stabilize ssDNA structures required for essential genomic processes. SSBs are essential proteins found in all domains of life. SSBs bind ssDNA with high affinity and in a sequence-independent manner and, in doing so, SSBs help to form the central nucleoprotein complex substrate for DNA replication, recombination, and repair processes. While SSBs are found in every organism, the proteins themselves share surprisingly little sequence similarity, subunit composition, and oligomerization states. All SSB proteins contain at least one DNA-binding oligonucleotide/oligosaccharide binding (OB) fold, which consists minimally of a five stranded beta-sheet arranged as a beta barrel capped by a single alpha helix. The OB fold is responsible for both ssDNA binding and oligomerization (for SSBs that operate as oligomers). The overall organization of OB folds varies between bacteria, eukaryotes, and archaea. As part of SSB/ssDNA cellular structures, SSBs play direct roles in the DNA replication, recombination, and repair. In many cases, SSBs have been found to form specific complexes with diverse genome maintenance proteins, often helping to recruit SSB/ssDNA-processing enzymes to the proper cellular sites of action. This clustering of genome maintenance

  9. An advanced PCR method for the specific detection of viable total coliform bacteria in pasteurized milk.

    PubMed

    Soejima, Takashi; Minami, Jun-ichi; Yaeshima, Tomoko; Iwatsuki, Keiji

    2012-07-01

    Pasteurized milk is a complex food that contains various inhibitors of polymerase chain reaction (PCR) and may contain a large number of dead bacteria, depending on the milking conditions and environment. Ethidium monoazide bromide (EMA)-PCR is occasionally used to distinguish between viable and dead bacteria in foods other than pasteurized milk. EMA is a DNA-intercalating dye that selectively permeates the compromised cell membranes of dead bacteria and cleaves DNA. Usually, EMA-PCR techniques reduce the detection of dead bacteria by up to 3.5 logs compared with techniques that do not use EMA. However, this difference may still be insufficient to suppress the amplification of DNA from dead Gram-negative bacteria (e.g., total coliform bacteria) if they are present in pasteurized milk in large numbers. Thus, false positives may result. We developed a new method that uses real-time PCR targeting of a long DNA template (16S-23S rRNA gene, principally 2,451 bp) following EMA treatment to completely suppress the amplification of DNA of up to 7 logs (10(7) cells) of dead total coliforms. Furthermore, we found that a low dose of proteinase K (25 U/ml) removed PCR inhibitors and simultaneously increased the signal from viable coliform bacteria. In conclusion, our simple protocol specifically detects viable total coliforms in pasteurized milk at an initial count of ≥1 colony forming unit (CFU)/2.22 ml within 7.5 h of total testing time. This detection limit for viable cells complies with the requirements for the analysis of total coliforms in pasteurized milk set by the Japanese Sanitation Act (which specifies <1 CFU/2.22 ml). PMID:22644523

  10. Gene transfer in bacteria: speciation without species?

    PubMed

    Lawrence, Jeffrey G

    2002-06-01

    Although Bacteria and Archaea reproduce by binary fission, exchange of genes among lineages has shaped the diversity of their populations and the diversification of their lineages. Gene exchange can occur by two distinct routes, each differentially impacting the recipient genome. First, homologous recombination mediates the exchange of DNA between closely related individuals (those whose sequences are sufficient similarly to allow efficient integration). As a result, homologous recombination mediates the dispersal of advantageous alleles that may rise to high frequency among genetically related individuals via periodic selection events. Second, lateral gene transfer can introduce novel DNA into a genome from completely unrelated lineages via illegitimate recombination. Gene exchange by this route serves to distribute genes throughout distantly related clades and therefore may confer complex abilities--not otherwise found among closely related lineages--onto the recipient organisms. These two mechanisms of gene exchange play complementary roles in the diversification of microbial populations into independent, ecologically distinct lineages. Although the delineation of microbial "species" then becomes difficult--if not impossible--to achieve, a cogent process of speciation can be predicted.

  11. Isolation of methanotrophic bacteria from termite gut.

    PubMed

    Reuss, Julia; Rachel, Reinhard; Kämpfer, Peter; Rabenstein, Andreas; Küver, Jan; Dröge, Stefan; König, Helmut

    2015-10-01

    The guts of termites feature suitable conditions for methane oxidizing bacteria (MOB) with their permanent production of CH4 and constant supply of O2 via tracheae. In this study, we have isolated MOB from the gut contents of the termites Incisitermes marginipennis, Mastotermes darwiniensis, and Neotermes castaneus for the first time. The existence of MOB was indicated by detecting pmoA, the gene for the particulate methane monooxygenase, in the DNA of gut contents. Fluorescence in situ hybridization and quantitative real-time polymerase chain reaction supported those findings. The MOB cell titer was determined to be 10(2)-10(3) per gut. Analyses of the 16S rDNA from isolates indicated close similarity to the genus Methylocystis. After various physiological tests and fingerprinting methods, no exact match to a known species was obtained, indicating the isolation of new MOB species. However, MALDI-TOF MS analyses revealed a close relationship to Methylocystis bryophila and Methylocystis parvus. PMID:26411892

  12. Further studies of infectious DNA extracted from mycobacteriophages.

    PubMed

    Sellers, M I; Tokunaga, T

    1966-02-01

    Results of the previous investigation in which it was found that DNA extracted from D29 mycobacteriophage was infectious for Mycobacterium smegmatis 607, have been extended. DNA extracted from mycobacteriophage D4 and D32 produced plaques when plated on their respective hosts; D28 DNA, extracted in the same manner and tested under similar conditions, failed to show infectivity. Species barriers were not crossed by mycobacteriophage DNA; bacteria resistant to intact phage were not infected with the phage DNA. The efficiency of plating of the DNA is very much lower than that of intact phage; infection of a given host was not accomplished by DNA when titration for plaque formation by the intact phage was less than 10(9) PFU. The base composition of DNA extracted from the four mycobacteriophages and the three propagating hosts was very similar. The bases were paired, adenine with thymine and guanine with cytosine. A relatively higher per cent of guanine-cytosine than of adenine-thymine, was found. The buoyant density of each DNA in CsCl was linearly related to its guanine-cytosine content whereas with the exception of D28 DNA, thermal denaturation temperatures failed to show this relationship. However, the thermal transition profiles were characteristic of double stranded DNA. Additional evidence that D29 DNA forms complexes with basic proteins was obtained. Binding between calf thymus histone and between RNAase and D29 DNA readily occurs with a resultant loss in DNA infectivity. Trypsin and D29 DNA are only weakly reactive.

  13. Environmental sources of fecal bacteria

    USGS Publications Warehouse

    Byappanahalli, Muruleedhara N.; Ishii, Satoshi; Sadowsky, Michael J.; Whitman, Richard L.

    2011-01-01

    This chapter provides a review of the research on environmental occurrences of faecal indicator bacteria in a variety of terrestrial and aquatic habitats under different geographic and climatic conditions, and discusses how these external sources may affect surface water quality.

  14. Magnetosome biogenesis in magnetotactic bacteria.

    PubMed

    Uebe, René; Schüler, Dirk

    2016-09-13

    Magnetotactic bacteria derive their magnetic orientation from magnetosomes, which are unique organelles that contain nanometre-sized crystals of magnetic iron minerals. Although these organelles have evident potential for exciting biotechnological applications, a lack of genetically tractable magnetotactic bacteria had hampered the development of such tools; however, in the past decade, genetic studies using two model Magnetospirillum species have revealed much about the mechanisms of magnetosome biogenesis. In this Review, we highlight these new insights and place the molecular mechanisms of magnetosome biogenesis in the context of the complex cell biology of Magnetospirillum spp. Furthermore, we discuss the diverse properties of magnetosome biogenesis in other species of magnetotactic bacteria and consider the value of genetically 'magnetizing' non-magnetotactic bacteria. Finally, we discuss future prospects for this highly interdisciplinary and rapidly advancing field. PMID:27620945

  15. Biopreservation by lactic acid bacteria.

    PubMed

    Stiles, M E

    1996-10-01

    Biopreservation refers to extended storage life and enhanced safety of foods using the natural microflora and (or) their antibacterial products. Lactic acid bacteria have a major potential for use in biopreservation because they are safe to consume and during storage they naturally dominate the microflora of many foods. In milk, brined vegetables, many cereal products and meats with added carbohydrate, the growth of lactic acid bacteria produces a new food product. In raw meats and fish that are chill stored under vacuum or in an environment with elevated carbon dioxide concentration, the lactic acid bacteria become the dominant population and preserve the meat with a "hidden' fermentation. The same applies to processed meats provided that the lactic acid bacteria survive the heat treatment or they are inoculated onto the product after heat treatment. This paper reviews the current status and potential for controlled biopreservation of foods. PMID:8879414

  16. Clinical microbiology of coryneform bacteria.

    PubMed Central

    Funke, G; von Graevenitz, A; Clarridge, J E; Bernard, K A

    1997-01-01

    Coryneform bacteria are aerobically growing, asporogenous, non-partially-acid-fast, gram-positive rods of irregular morphology. Within the last few years, there has been a massive increase in the number of publications related to all aspects of their clinical microbiology. Clinical microbiologists are often confronted with making identifications within this heterogeneous group as well as with considerations of the clinical significance of such isolates. This review provides comprehensive information on the identification of coryneform bacteria and outlines recent changes in taxonomy. The following genera are covered: Corynebacterium, Turicella, Arthrobacter, Brevibacterium, Dermabacter. Propionibacterium, Rothia, Exiguobacterium, Oerskovia, Cellulomonas, Sanguibacter, Microbacterium, Aureobacterium, "Corynebacterium aquaticum," Arcanobacterium, and Actinomyces. Case reports claiming disease associations of coryneform bacteria are critically reviewed. Minimal microbiological requirements for publications on disease associations of coryneform bacteria are proposed. PMID:8993861

  17. Biopreservation by lactic acid bacteria.

    PubMed

    Stiles, M E

    1996-10-01

    Biopreservation refers to extended storage life and enhanced safety of foods using the natural microflora and (or) their antibacterial products. Lactic acid bacteria have a major potential for use in biopreservation because they are safe to consume and during storage they naturally dominate the microflora of many foods. In milk, brined vegetables, many cereal products and meats with added carbohydrate, the growth of lactic acid bacteria produces a new food product. In raw meats and fish that are chill stored under vacuum or in an environment with elevated carbon dioxide concentration, the lactic acid bacteria become the dominant population and preserve the meat with a "hidden' fermentation. The same applies to processed meats provided that the lactic acid bacteria survive the heat treatment or they are inoculated onto the product after heat treatment. This paper reviews the current status and potential for controlled biopreservation of foods.

  18. The Mechanical World of Bacteria

    PubMed Central

    Persat, Alexandre; Nadell, Carey D.; Kim, Minyoung Kevin; Ingremeau, Francois; Siryaporn, Albert; Drescher, Knut; Wingreen, Ned S.; Bassler, Bonnie L.; Gitai, Zemer; Stone, Howard A.

    2015-01-01

    Summary In the wild, bacteria are predominantly associated with surfaces as opposed to existing as free-swimming, isolated organisms. They are thus subject to surface-specific mechanics including hydrodynamic forces, adhesive forces, the rheology of their surroundings and transport rules that define their encounters with nutrients and signaling molecules. Here, we highlight the effects of mechanics on bacterial behaviors on surfaces at multiple length scales, from single bacteria to the development of multicellular bacterial communities such as biofilms. PMID:26000479

  19. Bacteriophage Infection of Model Metal Reducing Bacteria

    NASA Astrophysics Data System (ADS)

    Weber, K. A.; Bender, K. S.; Gandhi, K.; Coates, J. D.

    2008-12-01

    Microbially-mediated metal reduction plays a significant role controlling contaminant mobility in aqueous, soil, and sedimentary environments. From among environmentally relevant microorganisms mediating metal reduction, Geobacter spp. have been identified as predominant metal-reducing bacteria under acetate- oxidizing conditions. Due to the significance of these bacteria in environmental systems, it is necessary to understand factors influencing their metabolic physiology. Examination of the annotated finished genome sequence of G. sulfurreducens PCA, G. uraniumreducens Rf4, G. metallireduceans GS-15 as well as a draft genome sequence of Geobacter sp. FRC-32 have identified gene sequences of putative bacteriophage origin. Presence of these sequences indicates that these bacteria are susceptible to phage infection. Polymerase chain reaction (PCR) primer sets designed tested for the presence of 12 of 25 annotated phage-like sequences in G. sulfurreducens PCA and 9 of 17 phage-like sequences in FRC- 32. The following genes were successfully amplified in G. sulfurreducens PCA: prophage type transcription regulator, phage-induced endonuclease, phage tail sheath, 2 phage tail proteins, phage protein D, phage base plate protein, phage-related DNA polymerase, integrase, phage transcriptional regulator, and Cro-like transcription regulator. Nine of the following sequences were present in FRC-32: 4 separate phage- related proteins, phage-related tail component, viron core protein, phage Mu protein, phage base plate, and phage tail sheath. In addition to the bioinformatics evidence, incubation of G. sulfurreducens PCA with 1 μg mL-1 mytomycin C (mutagen stimulating prophage induction) during mid-log phase resulted in significant cell lysis relative to cultures that remained unamended. Cell lysis was concurrent with an increase in viral like particles enumerated using epifluorescent microscopy. In addition, samples collected following this lytic event (~44hours) were

  20. Bacterial natural transformation by highly fragmented and damaged DNA.

    PubMed

    Overballe-Petersen, Søren; Harms, Klaus; Orlando, Ludovic A A; Mayar, J Victor Moreno; Rasmussen, Simon; Dahl, Tais W; Rosing, Minik T; Poole, Anthony M; Sicheritz-Ponten, Thomas; Brunak, Søren; Inselmann, Sabrina; de Vries, Johann; Wackernagel, Wilfried; Pybus, Oliver G; Nielsen, Rasmus; Johnsen, Pål Jarle; Nielsen, Kaare Magne; Willerslev, Eske

    2013-12-01

    DNA molecules are continuously released through decomposition of organic matter and are ubiquitous in most environments. Such DNA becomes fragmented and damaged (often <100 bp) and may persist in the environment for more than half a million years. Fragmented DNA is recognized as nutrient source for microbes, but not as potential substrate for bacterial evolution. Here, we show that fragmented DNA molecules (≥ 20 bp) that additionally may contain abasic sites, cross-links, or miscoding lesions are acquired by the environmental bacterium Acinetobacter baylyi through natural transformation. With uptake of DNA from a 43,000-y-old woolly mammoth bone, we further demonstrate that such natural transformation events include ancient DNA molecules. We find that the DNA recombination is RecA recombinase independent and is directly linked to DNA replication. We show that the adjacent nucleotide variations generated by uptake of short DNA fragments escape mismatch repair. Moreover, double-nucleotide polymorphisms appear more common among genomes of transformable than nontransformable bacteria. Our findings reveal that short and damaged, including truly ancient, DNA molecules, which are present in large quantities in the environment, can be acquired by bacteria through natural transformation. Our findings open for the possibility that natural genetic exchange can occur with DNA up to several hundreds of thousands years old.

  1. Bacterial natural transformation by highly fragmented and damaged DNA.

    PubMed

    Overballe-Petersen, Søren; Harms, Klaus; Orlando, Ludovic A A; Mayar, J Victor Moreno; Rasmussen, Simon; Dahl, Tais W; Rosing, Minik T; Poole, Anthony M; Sicheritz-Ponten, Thomas; Brunak, Søren; Inselmann, Sabrina; de Vries, Johann; Wackernagel, Wilfried; Pybus, Oliver G; Nielsen, Rasmus; Johnsen, Pål Jarle; Nielsen, Kaare Magne; Willerslev, Eske

    2013-12-01

    DNA molecules are continuously released through decomposition of organic matter and are ubiquitous in most environments. Such DNA becomes fragmented and damaged (often <100 bp) and may persist in the environment for more than half a million years. Fragmented DNA is recognized as nutrient source for microbes, but not as potential substrate for bacterial evolution. Here, we show that fragmented DNA molecules (≥ 20 bp) that additionally may contain abasic sites, cross-links, or miscoding lesions are acquired by the environmental bacterium Acinetobacter baylyi through natural transformation. With uptake of DNA from a 43,000-y-old woolly mammoth bone, we further demonstrate that such natural transformation events include ancient DNA molecules. We find that the DNA recombination is RecA recombinase independent and is directly linked to DNA replication. We show that the adjacent nucleotide variations generated by uptake of short DNA fragments escape mismatch repair. Moreover, double-nucleotide polymorphisms appear more common among genomes of transformable than nontransformable bacteria. Our findings reveal that short and damaged, including truly ancient, DNA molecules, which are present in large quantities in the environment, can be acquired by bacteria through natural transformation. Our findings open for the possibility that natural genetic exchange can occur with DNA up to several hundreds of thousands years old. PMID:24248361

  2. Bacterial natural transformation by highly fragmented and damaged DNA

    PubMed Central

    Overballe-Petersen, Søren; Harms, Klaus; Orlando, Ludovic A. A.; Mayar, J. Victor Moreno; Rasmussen, Simon; Dahl, Tais W.; Rosing, Minik T.; Poole, Anthony M.; Sicheritz-Ponten, Thomas; Brunak, Søren; Inselmann, Sabrina; de Vries, Johann; Wackernagel, Wilfried; Pybus, Oliver G.; Nielsen, Rasmus; Johnsen, Pål Jarle; Nielsen, Kaare Magne; Willerslev, Eske

    2013-01-01

    DNA molecules are continuously released through decomposition of organic matter and are ubiquitous in most environments. Such DNA becomes fragmented and damaged (often <100 bp) and may persist in the environment for more than half a million years. Fragmented DNA is recognized as nutrient source for microbes, but not as potential substrate for bacterial evolution. Here, we show that fragmented DNA molecules (≥20 bp) that additionally may contain abasic sites, cross-links, or miscoding lesions are acquired by the environmental bacterium Acinetobacter baylyi through natural transformation. With uptake of DNA from a 43,000-y-old woolly mammoth bone, we further demonstrate that such natural transformation events include ancient DNA molecules. We find that the DNA recombination is RecA recombinase independent and is directly linked to DNA replication. We show that the adjacent nucleotide variations generated by uptake of short DNA fragments escape mismatch repair. Moreover, double-nucleotide polymorphisms appear more common among genomes of transformable than nontransformable bacteria. Our findings reveal that short and damaged, including truly ancient, DNA molecules, which are present in large quantities in the environment, can be acquired by bacteria through natural transformation. Our findings open for the possibility that natural genetic exchange can occur with DNA up to several hundreds of thousands years old. PMID:24248361

  3. Cable Bacteria in Freshwater Sediments

    PubMed Central

    Kristiansen, Michael; Frederiksen, Rasmus B.; Dittmer, Anders Lindequist; Bjerg, Jesper Tataru; Trojan, Daniela; Schreiber, Lars; Damgaard, Lars Riis; Schramm, Andreas; Nielsen, Lars Peter

    2015-01-01

    In marine sediments cathodic oxygen reduction at the sediment surface can be coupled to anodic sulfide oxidation in deeper anoxic layers through electrical currents mediated by filamentous, multicellular bacteria of the Desulfobulbaceae family, the so-called cable bacteria. Until now, cable bacteria have only been reported from marine environments. In this study, we demonstrate that cable bacteria also occur in freshwater sediments. In a first step, homogenized sediment collected from the freshwater stream Giber Å, Denmark, was incubated in the laboratory. After 2 weeks, pH signatures and electric fields indicated electron transfer between vertically separated anodic and cathodic half-reactions. Fluorescence in situ hybridization revealed the presence of Desulfobulbaceae filaments. In addition, in situ measurements of oxygen, pH, and electric potential distributions in the waterlogged banks of Giber Å demonstrated the presence of distant electric redox coupling in naturally occurring freshwater sediment. At the same site, filamentous Desulfobulbaceae with cable bacterium morphology were found to be present. Their 16S rRNA gene sequence placed them as a distinct sister group to the known marine cable bacteria, with the genus Desulfobulbus as the closest cultured lineage. The results of the present study indicate that electric currents mediated by cable bacteria could be important for the biogeochemistry in many more environments than anticipated thus far and suggest a common evolutionary origin of the cable phenotype within Desulfobulbaceae with subsequent diversification into a freshwater and a marine lineage. PMID:26116678

  4. Cable Bacteria in Freshwater Sediments.

    PubMed

    Risgaard-Petersen, Nils; Kristiansen, Michael; Frederiksen, Rasmus B; Dittmer, Anders Lindequist; Bjerg, Jesper Tataru; Trojan, Daniela; Schreiber, Lars; Damgaard, Lars Riis; Schramm, Andreas; Nielsen, Lars Peter

    2015-09-01

    In marine sediments cathodic oxygen reduction at the sediment surface can be coupled to anodic sulfide oxidation in deeper anoxic layers through electrical currents mediated by filamentous, multicellular bacteria of the Desulfobulbaceae family, the so-called cable bacteria. Until now, cable bacteria have only been reported from marine environments. In this study, we demonstrate that cable bacteria also occur in freshwater sediments. In a first step, homogenized sediment collected from the freshwater stream Giber Å, Denmark, was incubated in the laboratory. After 2 weeks, pH signatures and electric fields indicated electron transfer between vertically separated anodic and cathodic half-reactions. Fluorescence in situ hybridization revealed the presence of Desulfobulbaceae filaments. In addition, in situ measurements of oxygen, pH, and electric potential distributions in the waterlogged banks of Giber Å demonstrated the presence of distant electric redox coupling in naturally occurring freshwater sediment. At the same site, filamentous Desulfobulbaceae with cable bacterium morphology were found to be present. Their 16S rRNA gene sequence placed them as a distinct sister group to the known marine cable bacteria, with the genus Desulfobulbus as the closest cultured lineage. The results of the present study indicate that electric currents mediated by cable bacteria could be important for the biogeochemistry in many more environments than anticipated thus far and suggest a common evolutionary origin of the cable phenotype within Desulfobulbaceae with subsequent diversification into a freshwater and a marine lineage.

  5. Bioreporter bacteria for landmine detection

    SciTech Connect

    Burlage, R.S.; Youngblood, T.; Lamothe, D.

    1998-04-01

    Landmines (and other UXO) gradually leak explosive chemicals into the soil at significant concentrations. Bacteria, which have adapted to scavenge low concentrations of nutrients, can detect these explosive chemicals. Uptake of these chemicals results in the triggering of specific bacterial genes. The authors have created genetically recombinant bioreporter bacteria that detect small concentrations of energetic chemicals. These bacteria are genetically engineered to produce a bioluminescent signal when they contact specific explosives. A gene for a brightly fluorescent compound can be substituted for increased sensitivity. By finding the fluorescent bacteria, you find the landmine. Detection might be accomplished using stand-off illumination of the minefield and GPS technology, which would result in greatly reduced risk to the deminers. Bioreporter technology has been proven at the laboratory scale, and will be tested under field conditions in the near future. They have created a bacterial strain that detects sub-micromolar concentrations of o- and p-nitrotoluene. Related bacterial strains were produced using standard laboratory protocols, and bioreporters of dinitrotoluene and trinitrotoluene were produced, screening for activity with the explosive compounds. Response time is dependent on the growth rate of the bacteria. Although frill signal production may require several hours, the bacteria can be applied over vast areas and scanned quickly, producing an equivalent detection speed that is very fast. This technology may be applicable to other needs, such as locating buried explosives at military and ordnance/explosive manufacturing facilities.

  6. Filtrating forms of soil bacteria

    NASA Astrophysics Data System (ADS)

    Van'kova, A. A.; Ivanov, P. I.; Emtsev, V. T.

    2013-03-01

    Filtrating (ultramicroscopic) forms (FF) of bacteria were studied in a soddy-podzolic soil and the root zone of alfalfa plants as part of populations of the most widespread physiological groups of soil bacteria. FF were obtained by filtering soil solutions through membrane filters with a pore diameter of 0.22 μm. It was established that the greater part of the bacteria in the soil and in the root zone of the plants has an ultramicroscopic size: the average diameter of the cells is 0.3 μm, and their length is 0.6 μm, which is significantly less than the cell size of banal bacteria. The number of FF varies within a wide range depending on the physicochemical conditions of the habitat. The FF number's dynamics in the soil is of a seasonal nature; i.e., the number of bacteria found increases in the summer and fall and decreases in the winter-spring period. In the rhizosphere of the alfalfa, over the vegetation period, the number of FF and their fraction in the total mass of the bacteria increase. A reverse tendency is observed in the rhizoplane. The morphological particularities (identified by an electron microscopy) and the nature of the FF indicate their physiological activity.

  7. Anaerobic carboxydotrophic bacteria in geothermal springs identified using stable isotope probing.

    PubMed

    Brady, Allyson L; Sharp, Christine E; Grasby, Stephen E; Dunfield, Peter F

    2015-01-01

    Carbon monoxide (CO) is a potential energy and carbon source for thermophilic bacteria in geothermal environments. Geothermal sites ranging in temperature from 45 to 65°C were investigated for the presence and activity of anaerobic CO-oxidizing bacteria. Anaerobic CO oxidation potentials were measured at up to 48.9 μmoles CO g(-1) (wet weight) day(-1) within five selected sites. Active anaerobic carboxydotrophic bacteria were identified using (13)CO DNA stable isotope probing (SIP) combined with pyrosequencing of 16S rRNA genes amplified from labeled DNA. Bacterial communities identified in heavy DNA fractions were predominated by Firmicutes, which comprised up to 95% of all sequences in (13)CO incubations. The predominant bacteria that assimilated (13)C derived from CO were closely related (>98% 16S rRNA gene sequence identity) to genera of known carboxydotrophs including Thermincola, Desulfotomaculum, Thermolithobacter, and Carboxydocella, although a few species with lower similarity to known bacteria were also found that may represent previously unconfirmed CO-oxidizers. While the distribution was variable, many of the same OTUs were identified across sample sites from different temperature regimes. These results show that bacteria capable of using CO as a carbon source are common in geothermal springs, and that thermophilic carboxydotrophs are probably already quite well known from cultivation studies.

  8. Anaerobic carboxydotrophic bacteria in geothermal springs identified using stable isotope probing

    PubMed Central

    Brady, Allyson L.; Sharp, Christine E.; Grasby, Stephen E.; Dunfield, Peter F.

    2015-01-01

    Carbon monoxide (CO) is a potential energy and carbon source for thermophilic bacteria in geothermal environments. Geothermal sites ranging in temperature from 45 to 65°C were investigated for the presence and activity of anaerobic CO-oxidizing bacteria. Anaerobic CO oxidation potentials were measured at up to 48.9 μmoles CO g−1 (wet weight) day−1 within five selected sites. Active anaerobic carboxydotrophic bacteria were identified using 13CO DNA stable isotope probing (SIP) combined with pyrosequencing of 16S rRNA genes amplified from labeled DNA. Bacterial communities identified in heavy DNA fractions were predominated by Firmicutes, which comprised up to 95% of all sequences in 13CO incubations. The predominant bacteria that assimilated 13C derived from CO were closely related (>98% 16S rRNA gene sequence identity) to genera of known carboxydotrophs including Thermincola, Desulfotomaculum, Thermolithobacter, and Carboxydocella, although a few species with lower similarity to known bacteria were also found that may represent previously unconfirmed CO-oxidizers. While the distribution was variable, many of the same OTUs were identified across sample sites from different temperature regimes. These results show that bacteria capable of using CO as a carbon source are common in geothermal springs, and that thermophilic carboxydotrophs are probably already quite well known from cultivation studies. PMID:26388850

  9. Bioprobes Based on Aptamer and Silica Fluorescent Nanoparticles for Bacteria Salmonella typhimurium Detection

    NASA Astrophysics Data System (ADS)

    Wang, Qiu-Yue; Kang, Yan-Jun

    2016-03-01

    In this study, we have developed an efficient method based on single-stranded DNA (ssDNA) aptamers along with silica fluorescence nanoparticles for bacteria Salmonella typhimurium detection. Carboxyl-modified Tris(2,2'-bipyridyl)dichlororuthenium(II) hexahydrate (RuBPY)-doped silica nanoparticles (COOH-FSiNPs) were prepared using reverse microemulsion method, and the streptavidin was conjugated to the surface of the prepared COOH-FSiNPs. The bacteria S. typhimurium was incubated with a specific ssDNA biotin-labeled aptamer, and then the aptamer-bacteria conjugates were treated with the synthetic streptavidin-conjugated silica fluorescence nanoprobes (SA-FSiNPs). The results under fluorescence microscopy show that SA-FSiNPs can be applied effectively for the labeling of bacteria S. typhimurium with great photostable property. To further verify the specificity of SA-FSiNPs out of multiple bacterial conditions, variant concentrations of bacteria mixtures composed of bacteria S. typhimurium, Escherichia coli, and Bacillus subtilis were treated with SA-FSiNPs.

  10. Anaerobic carboxydotrophic bacteria in geothermal springs identified using stable isotope probing.

    PubMed

    Brady, Allyson L; Sharp, Christine E; Grasby, Stephen E; Dunfield, Peter F

    2015-01-01

    Carbon monoxide (CO) is a potential energy and carbon source for thermophilic bacteria in geothermal environments. Geothermal sites ranging in temperature from 45 to 65°C were investigated for the presence and activity of anaerobic CO-oxidizing bacteria. Anaerobic CO oxidation potentials were measured at up to 48.9 μmoles CO g(-1) (wet weight) day(-1) within five selected sites. Active anaerobic carboxydotrophic bacteria were identified using (13)CO DNA stable isotope probing (SIP) combined with pyrosequencing of 16S rRNA genes amplified from labeled DNA. Bacterial communities identified in heavy DNA fractions were predominated by Firmicutes, which comprised up to 95% of all sequences in (13)CO incubations. The predominant bacteria that assimilated (13)C derived from CO were closely related (>98% 16S rRNA gene sequence identity) to genera of known carboxydotrophs including Thermincola, Desulfotomaculum, Thermolithobacter, and Carboxydocella, although a few species with lower similarity to known bacteria were also found that may represent previously unconfirmed CO-oxidizers. While the distribution was variable, many of the same OTUs were identified across sample sites from different temperature regimes. These results show that bacteria capable of using CO as a carbon source are common in geothermal springs, and that thermophilic carboxydotrophs are probably already quite well known from cultivation studies. PMID:26388850

  11. DNA Investigations.

    ERIC Educational Resources Information Center

    Mayo, Ellen S.; Bertino, Anthony J.

    1991-01-01

    Presents a simulation activity that allow students to work through the exercise of DNA profiling and to grapple with some analytical and ethical questions involving a couple arranging with a surrogate mother to have a baby. Can be used to teach the principles of restriction enzyme digestion, gel electrophoresis, and probe hybridization. (MDH)

  12. Molecular Evidence for Metabolically Active Bacteria in the Atmosphere.

    PubMed

    Klein, Ann M; Bohannan, Brendan J M; Jaffe, Daniel A; Levin, David A; Green, Jessica L

    2016-01-01

    Bacterial metabolisms are responsible for critical chemical transformations in nearly all environments, including oceans, freshwater, and soil. Despite the ubiquity of bacteria in the atmosphere, little is known about the metabolic functioning of atmospheric bacterial communities. To gain a better understanding of the metabolism of bacterial communities in the atmosphere, we used a combined empirical and model-based approach to investigate the structure and composition of potentially active bacterial communities in air sampled at a high elevation research station. We found that the composition of the putatively active bacterial community (assayed via rRNA) differed significantly from the total bacterial community (assayed via rDNA). Rare taxa in the total (rDNA) community were disproportionately active relative to abundant taxa, and members of the order Rhodospirillales had the highest potential for activity. We developed theory to explore the effects of random sampling from the rRNA and rDNA communities on observed differences between the communities. We found that random sampling, particularly in cases where active taxa are rare in the rDNA community, will give rise to observed differences in community composition including the occurrence of "phantom taxa", taxa which are detected in the rRNA community but not the rDNA community. We show that the use of comparative rRNA/rDNA techniques can reveal the structure and composition of the metabolically active portion of bacterial communities. Our observations suggest that metabolically active bacteria exist in the atmosphere and that these communities may be involved in the cycling of organic compounds in the atmosphere.

  13. Molecular Evidence for Metabolically Active Bacteria in the Atmosphere

    PubMed Central

    Klein, Ann M.; Bohannan, Brendan J. M.; Jaffe, Daniel A.; Levin, David A.; Green, Jessica L.

    2016-01-01

    Bacterial metabolisms are responsible for critical chemical transformations in nearly all environments, including oceans, freshwater, and soil. Despite the ubiquity of bacteria in the atmosphere, little is known about the metabolic functioning of atmospheric bacterial communities. To gain a better understanding of the metabolism of bacterial communities in the atmosphere, we used a combined empirical and model-based approach to investigate the structure and composition of potentially active bacterial communities in air sampled at a high elevation research station. We found that the composition of the putatively active bacterial community (assayed via rRNA) differed significantly from the total bacterial community (assayed via rDNA). Rare taxa in the total (rDNA) community were disproportionately active relative to abundant taxa, and members of the order Rhodospirillales had the highest potential for activity. We developed theory to explore the effects of random sampling from the rRNA and rDNA communities on observed differences between the communities. We found that random sampling, particularly in cases where active taxa are rare in the rDNA community, will give rise to observed differences in community composition including the occurrence of “phantom taxa”, taxa which are detected in the rRNA community but not the rDNA community. We show that the use of comparative rRNA/rDNA techniques can reveal the structure and composition of the metabolically active portion of bacterial communities. Our observations suggest that metabolically active bacteria exist in the atmosphere and that these communities may be involved in the cycling of organic compounds in the atmosphere. PMID:27252689

  14. Browning of freeze-dried probiotic bacteria cultures in relation to loss of viability during storage.

    PubMed

    Kurtmann, Lone; Skibsted, Leif H; Carlsen, Charlotte U

    2009-08-12

    Freeze-dried cultures of Lactobacillus acidophilus (La-5) showed visible brown discoloration even after a short storage at relatively mild conditions (a(w) = 0.22 and 30 degrees C), and the browning processes were found to coincide with bacteria inactivation. It was demonstrated, by using high-pressure treatment for obtaining bacteria samples with different ratios of live/dead bacteria, that death of bacteria is not a prerequisite for the browning processes. Furthermore, it was shown that hydroxymethylfurfural (HMF) (or condensation products of HMF) introduces accelerated viability loss when HMF is added to the freeze-drying medium. Discoloration of bacteria cultures containing only sucrose/maltodextrin or lactose/maltodextrin in the freeze-drying matrices is suggested to be related to various types of nonenzymatic browning reactions, including carbonyl-protein (or carbonyl-DNA) interactions and carbohydrate condensation/polymerization (without involvement of proteins), the latter proceeding at low a(w) following hydrolysis of the peptidoglycan layer in the bacteria cell wall. More than one single type of browning reaction is accordingly concluded to be related to bacteria death, and the loss of viability in freeze-dried bacteria seems to be influenced by oxidation reactions, browning reactions, and the physical instability of the bacteria membrane/cell wall. PMID:19591471

  15. Methods for Baiting and Enriching Fungus-Feeding (Mycophagous) Rhizosphere Bacteria

    PubMed Central

    Ballhausen, Max-Bernhard; van Veen, Johannes A.; Hundscheid, Maria P. J.; de Boer, Wietse

    2015-01-01

    Mycophagous soil bacteria are able to obtain nutrients from living fungal hyphae. However, with exception of the soil bacterial genus Collimonas, occurrence of this feeding strategy has not been well examined. Evaluation of the importance of mycophagy in soil bacterial communities requires targeted isolation methods. In this study, we compared two different approaches to obtain mycophagous bacteria from rhizospheric soil. A short-term method based on baiting for bacteria that can rapidly adhere to fungal hyphae and a long-term method based on the enrichment of bacteria on fungal hyphae via repeated transfer. Hyphae-adhering bacteria were isolated, identified by 16S rDNA sequencing and tested for antifungal activity and the ability to feed on fungi as the sole source of carbon. Both methods yielded a range of potentially mycophagous bacterial isolates with little phylogenetic overlap. We also found indications for feeding preferences among the potentially mycophagous bacteria. Our results indicate that mycophagy could be an important growth strategy for rhizosphere bacteria. To our surprise, we found several potential plant pathogenic bacteria among the mycophagous isolates. We discuss the possible benefits that these bacteria might gain from colonizing fungal hyphae. PMID:26733962

  16. DNA repair in Mycoplasma gallisepticum

    PubMed Central

    2013-01-01

    Background DNA repair is essential for the maintenance of genome stability in all living beings. Genome size as well as the repertoire and abundance of DNA repair components may vary among prokaryotic species. The bacteria of the Mollicutes class feature a small genome size, absence of a cell wall, and a parasitic lifestyle. A small number of genes make Mollicutes a good model for a “minimal cell” concept. Results In this work we studied the DNA repair system of Mycoplasma gallisepticum on genomic, transcriptional, and proteomic levels. We detected 18 out of 22 members of the DNA repair system on a protein level. We found that abundance of the respective mRNAs is less than one per cell. We studied transcriptional response of DNA repair genes of M. gallisepticum at stress conditions including heat, osmotic, peroxide stresses, tetracycline and ciprofloxacin treatment, stationary phase and heat stress in stationary phase. Conclusions Based on comparative genomic study, we determined that the DNA repair system M. gallisepticum includes a sufficient set of proteins to provide a cell with functional nucleotide and base excision repair and mismatch repair. We identified SOS-response in M. gallisepticum on ciprofloxacin, which is a known SOS-inducer, tetracycline and heat stress in the absence of established regulators. Heat stress was found to be the strongest SOS-inducer. We found that upon transition to stationary phase of culture growth transcription of DNA repair genes decreases dramatically. Heat stress does not induce SOS-response in a stationary phase. PMID:24148612

  17. Commensal Bacteria Aid Mate-selection in the Fruit Fly, Bactrocera dorsalis.

    PubMed

    Damodaram, Kamala Jayanthi Pagadala; Ayyasamy, Arthikirubha; Kempraj, Vivek

    2016-10-01

    Commensal bacteria influence many aspects of an organism's behaviour. However, studies on the influence of commensal bacteria in insect mate-selection are scarce. Here, we present empirical evidence that commensal bacteria mediate mate-selection in the Oriental fruit fly, Bactrocera dorsalis. Male flies were attracted to female flies, but this attraction was abolished when female flies were fed with antibiotics, suggesting the role of the fly's microbiota in mediating mate-selection. We show that male flies were attracted to and ejaculated more sperm into females harbouring the microbiota. Using culturing and 16S rDNA sequencing, we isolated and identified different commensal bacteria, with Klebsiella oxytoca being the most abundant bacterial species. This preliminary study will enhance our understanding of the influence of commensal bacteria on mate-selection behaviour of B. dorsalis and may find use in devising control operations against this devastating pest. PMID:27423980

  18. Commensal Bacteria Aid Mate-selection in the Fruit Fly, Bactrocera dorsalis.

    PubMed

    Damodaram, Kamala Jayanthi Pagadala; Ayyasamy, Arthikirubha; Kempraj, Vivek

    2016-10-01

    Commensal bacteria influence many aspects of an organism's behaviour. However, studies on the influence of commensal bacteria in insect mate-selection are scarce. Here, we present empirical evidence that commensal bacteria mediate mate-selection in the Oriental fruit fly, Bactrocera dorsalis. Male flies were attracted to female flies, but this attraction was abolished when female flies were fed with antibiotics, suggesting the role of the fly's microbiota in mediating mate-selection. We show that male flies were attracted to and ejaculated more sperm into females harbouring the microbiota. Using culturing and 16S rDNA sequencing, we isolated and identified different commensal bacteria, with Klebsiella oxytoca being the most abundant bacterial species. This preliminary study will enhance our understanding of the influence of commensal bacteria on mate-selection behaviour of B. dorsalis and may find use in devising control operations against this devastating pest.

  19. Genetic Diversity and Association Characters of Bacteria Isolated from Arbuscular Mycorrhizal Fungal Spore Walls.

    PubMed

    Selvakumar, Gopal; Krishnamoorthy, Ramasamy; Kim, Kiyoon; Sa, Tong-Min

    2016-01-01

    Association between arbuscular mycorrhizal fungi (AMF) and bacteria has long been studied. However, the factors influencing their association in the natural environment is still unknown. This study aimed to isolate bacteria associated with spore walls of AMF and identify their potential characters for association. Spores collected from coastal reclamation land were differentiated based on their morphology and identified by 18S rDNA sequencing as Funneliformis caledonium, Racocetra alborosea and Funneliformis mosseae. Bacteria associated with AMF spore walls were isolated after treating them with disinfection solution at different time intervals. After 0, 10 and 20 min of spore disinfection, 86, 24 and 10 spore associated bacteria (SAB) were isolated, respectively. BOX-PCR fingerprinting analysis showed that diverse bacterial communities were associated to AMF spores. Bacteria belonging to the same genera could associate with different AMF spores. Gram positive bacteria were more closely associated with AMF spores. Isolated SAB were characterized and tested for spore association characters such as chitinase, protease, cellulase enzymes and exopolysaccharide production (EPS). Among the 120 SAB, 113 SAB were able to show one or more characters for association and seven SAB did not show any association characters. The 16S rDNA sequence of SAB revealed that bacteria belonging to the phyla Firmicutes, Proteobacteria, Actinobacteria and Bactereiodes were associated with AMF spore walls. PMID:27479250

  20. Genetic Diversity and Association Characters of Bacteria Isolated from Arbuscular Mycorrhizal Fungal Spore Walls

    PubMed Central

    Selvakumar, Gopal; Krishnamoorthy, Ramasamy; Kim, Kiyoon; Sa, Tong-Min

    2016-01-01

    Association between arbuscular mycorrhizal fungi (AMF) and bacteria has long been studied. However, the factors influencing their association in the natural environment is still unknown. This study aimed to isolate bacteria associated with spore walls of AMF and identify their potential characters for association. Spores collected from coastal reclamation land were differentiated based on their morphology and identified by 18S rDNA sequencing as Funneliformis caledonium, Racocetra alborosea and Funneliformis mosseae. Bacteria associated with AMF spore walls were isolated after treating them with disinfection solution at different time intervals. After 0, 10 and 20 min of spore disinfection, 86, 24 and 10 spore associated bacteria (SAB) were isolated, respectively. BOX-PCR fingerprinting analysis showed that diverse bacterial communities were associated to AMF spores. Bacteria belonging to the same genera could associate with different AMF spores. Gram positive bacteria were more closely associated with AMF spores. Isolated SAB were characterized and tested for spore association characters such as chitinase, protease, cellulase enzymes and exopolysaccharide production (EPS). Among the 120 SAB, 113 SAB were able to show one or more characters for association and seven SAB did not show any association characters. The 16S rDNA sequence of SAB revealed that bacteria belonging to the phyla Firmicutes, Proteobacteria, Actinobacteria and Bactereiodes were associated with AMF spore walls. PMID:27479250

  1. Phylogenetic Diversity of Bacteria Associated with the Marine Sponge Rhopaloeides odorabile†

    PubMed Central

    Webster, Nicole S.; Wilson, Kate J.; Blackall, Linda L.; Hill, Russell T.

    2001-01-01

    Molecular techniques were employed to document the microbial diversity associated with the marine sponge Rhopaloeides odorabile. The phylogenetic affiliation of sponge-associated bacteria was assessed by 16S rRNA sequencing of cloned DNA fragments. Fluorescence in situ hybridization (FISH) was used to confirm the presence of the predominant groups indicated by 16S rDNA analysis. The community structure was extremely diverse with representatives of the Actinobacteria, low-G+C gram-positive bacteria, the β- and γ-subdivisions of the Proteobacteria, Cytophaga/Flavobacterium, green sulfur bacteria, green nonsulfur bacteria, planctomycetes, and other sequence types with no known close relatives. FISH probes revealed the spatial location of these bacteria within the sponge tissue, in some cases suggesting possible symbiotic functions. The high proportion of 16S rRNA sequences derived from novel actinomycetes is good evidence for the presence of an indigenous marine actinomycete assemblage in R. odorabile. High microbial diversity was inferred from low duplication of clones in a library with 70 representatives. Determining the phylogenetic affiliation of sponge-associated microorganisms by 16S rRNA analysis facilitated the rational selection of culture media and isolation conditions to target specific groups of well-represented bacteria for laboratory culture. Novel media incorporating sponge extracts were used to isolate bacteria not previously recovered from this sponge. PMID:11133476

  2. Genetic diversity of bacteria associated with the hindgut of the terrestrial crustacean Porcellio scaber (Crustacea: Isopoda).

    PubMed

    Kostanjsek, Rok; Strus, Jasna; Avgustin, Gorazd

    2002-06-01

    Molecular approaches were used to examine the genetic diversity of bacteria associated with the gut wall of the terrestrial isopod Porcellio scaber and to determine whether an autochthonous microflora exists in the P. scaber hindgut. 16S ribosomal genes were amplified from the total DNA isolated from thoroughly washed papillate regions of the hindgut, where the highest concentrations of bacteria are commonly found. The amplified genes were cloned, sequenced and phylogenetically analysed. The results implied an unexpectedly large diversity of microflora associated with the cuticle of the hindgut. Almost half of the retrieved sequences were found to be less than 80% homologous with any of the known sequences available at DNA data banks. Most of these sequences were clustered in one of three groups, and were clearly distant from the sequences of other bacterial taxa, indicating that they could represent novel bacterial species or even genera. More than two thirds of the sequences were found to be phylogenetically related to sequences from bacteria typically isolated from human and animal intestines, e.g. streptococci, enterococci, and members of the genus Bacteroides. The majority of the remaining sequences were most closely related to typical soil bacteria, e.g. bacilli and pseudomonads. The facts that a large proportion of the retrieved sequences was related to the sequences of bacteria, which are autochthonous to intestinal ecosystems, and that bacteria, specifically attached to the cuticular spines, were observed, indicate that truly autochthonous bacteria may well be present in the hindgut of P. scaber.

  3. Enumeration of total bacteria and bacteria with genes for proteolytic activity in pure cultures and in environmental samples by quantitative PCR mediated amplification.

    PubMed

    Bach, H-J; Tomanova, J; Schloter, M; Munch, J C

    2002-05-01

    Real-time quantitative PCR assays were developed for the absolute quantification of different groups of bacteria in pure cultures and in environmental samples. 16S rRNA genes were used as markers for eubacteria, and genes for extracellular peptidases were used as markers for potentially proteolytic bacteria. For the designed 16S rDNA TaqMan assay, specificity of the designed primer-probe combination for eubacteria, a high amplification efficiency over a wide range of starting copy numbers and a high reproducibility is demonstrated. Cell concentrations of Bacillus cereus, B. subtilis and Pseudomonas fluorescens in liquid culture were monitored by TaqMan-PCR using the 16S rDNA target sequence of Escherichia coli as external standard for quantification. Results agree with plate counts and microscopic counts of DAPI stained cells. The significance of 16S rRNA operon multiplicity to the quantification of bacteria is discussed.Furthermore, three sets of primer pair together with probe previously designed for targeting different classes of bacterial extracellular peptidases were tested for their suitability for TaqMan-PCR based quantification of proteolytic bacteria. Since high degeneracy of the probes did not allow accurate quantification, SybrGreen was used instead of molecular probes to visualize and quantify PCR products during PCR. The correlation between fluorescence and starting copy number was of the same high quality as for the 16S rDNA TaqMan assay for all the three peptidase gene classes. The detected amount of genes for neutral metallopeptidase of B. cereus, for subtilisin of B. subtilis and for alkaline metallopeptidase of P. fluorescens corresponded exactly to the numbers of bacteria investigated by the 16S rDNA targeting assay. The developed assays were applied for the quantification of bacteria in soil samples.

  4. quenched-smFISH: Counting small RNA in Pathogenic Bacteria

    NASA Astrophysics Data System (ADS)

    Shepherd, Douglas; Li, Nan; Micheva-Viteva, Sofiya; Munsky, Brian; Hong-Geller, Elizabeth; Werner, James

    2014-03-01

    Here, we present a modification to single-molecule fluorescence in situ hybridization, quenched smFISH (q-smFISH), that enables quantitative detection and analysis of small RNA (sRNA) expressed in bacteria. We show that short nucleic acid targets can be detected when the background of unbound singly dye-labeled DNA oligomers is reduced through hybridization with a set of complementary DNA oligomers labeled with a fluorescence quencher. Exploiting an automated, multi-color wide-field microscope and GPU-accelerated data analysis package, we analyzed the statistics of sRNA expression in thousands of individual Yersinia pseudotuberculosis and Yersinia pestis bacteria before and during a simulated infection. Before infection, we find only a small fraction of either bacteria express the small RNAs YSR35 or YSP8. The copy numbers of these RNA are increased during simulated infection, suggesting a role in pathogenesis. The ability to directly quantify expression level changes of sRNA in single cells as a function of external stimuli provides key information on the role of sRNA in bacterial regulatory networks.

  5. Aptamer-based viability impedimetric sensor for bacteria.

    PubMed

    Labib, Mahmoud; Zamay, Anna S; Kolovskaya, Olga S; Reshetneva, Irina T; Zamay, Galina S; Kibbee, Richard J; Sattar, Syed A; Zamay, Tatiana N; Berezovski, Maxim V

    2012-11-01

    The development of an aptamer-based viability impedimetric sensor for bacteria (AptaVISens-B) is presented. Highly specific DNA aptamers to live Salmonella typhimurium were selected via the cell-systematic evolution of ligands by exponential enrichment (SELEX) technique. Twelve rounds of selection were performed; each comprises a positive selection step against viable S. typhimurium and a negative selection step against heat killed S. typhimurium and a mixture of related pathogens, including Salmonella enteritidis, Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa, and Citrobacter freundii to ensure the species specificity of the selected aptamers. The DNA sequence showing the highest binding affinity to the bacteria was further integrated into an impedimetric sensor via self-assembly onto a gold nanoparticle-modified screen-printed carbon electrode (GNP-SPCE). Remarkably, this aptasensor is highly selective and can successfully detect S. typhimurium down to 600 CFU mL(-1) (equivalent to 18 live cells in 30 μL of assay volume) and distinguish it from other Salmonella species, including S. enteritidis and S. choleraesuis. This report is envisaged to open a new venue for the aptamer-based viability sensing of a variety of microorganisms, particularly viable but nonculturable (VBNC) bacteria, using a rapid, economic, and label-free electrochemical platform.

  6. Antibacterial activity of caffeine against plant pathogenic bacteria.

    PubMed

    Sledz, Wojciech; Los, Emilia; Paczek, Agnieszka; Rischka, Jacek; Motyka, Agata; Zoledowska, Sabina; Piosik, Jacek; Lojkowska, Ewa

    2015-01-01

    The objective of the present study was to evaluate the antibacterial properties of a plant secondary metabolite - caffeine. Caffeine is present in over 100 plant species. Antibacterial activity of caffeine was examined against the following plant-pathogenic bacteria: Ralstonia solanacearum (Rsol), Clavibacter michiganesis subsp. sepedonicus (Cms), Dickeya solani (Dsol), Pectobacterium atrosepticum (Pba), Pectobacterium carotovorum subsp. carotovorum (Pcc), Pseudomonas syringae pv. tomato (Pst), and Xanthomonas campestris subsp. campestris (Xcc). MIC and MBC values ranged from 5 to 20 mM and from 43 to 100 mM, respectively. Caffeine increased the bacterial generation time of all tested species and caused changes in cell morphology. The influence of caffeine on the synthesis of DNA, RNA and proteins was investigated in cultures of plant pathogenic bacteria with labelled precursors: [(3)H]thymidine, [(3)H]uridine or (14)C leucine, respectively. RNA biosynthesis was more affected than DNA or protein biosynthesis in bacterial cells treated with caffeine. Treatment of Pba with caffeine for 336 h did not induce resistance to this compound. Caffeine application reduced disease symptoms caused by Dsol on chicory leaves, potato slices, and whole potato tubers. The data presented indicate caffeine as a potential tool for the control of diseases caused by plant-pathogenic bacteria, especially under storage conditions. PMID:26307771

  7. Genetics of bacteria that utilize one-carbon compounds

    SciTech Connect

    Hanson, R.S.

    1989-11-01

    Methylotrophic bacteria that grow on the one-carbon compounds; methane, methanol, methylamines and dichloromethane are a morphologically and physiologically diverse group of eubacteria. The 16S rRNA molecules of several gram-negative methylotrophs have been sequenced. Two phylogenetically related groups containing type I and type II methylotrophs have been identified. Each group contains two subgroups of bacteria. DNA probes homologous to 16S rRNA's of each group of methanotrophs have been synthesized and have been shown to hybridize only to the 16S rRNA's from target bacteria. We have mapped the positions of 15 genes controlling the synthesis of methanol dehydrogenase, cytochrome C{sub L} and other functions required for the oxidation of methanol to formaldehyde in three species of type II methanotrophs. We have isolated a DNA-binding protein that binds to a cloned 172 bp sequence that is located upstream from the MDH structural gene. The function of this protein in the regulation of MDH synthesis will be investigated. The gene encoding the methane monooxygenase B component of {ital Methylosinus trichosporium} OB3b has been cloned and expressed in {ital Escherichia coli}. We intend to clone and map all five genes required for the expression of soluble MMO activity in {ital M. trichosporium} OB3b and to study the regulation of their synthesis. 5 refs.

  8. Antibacterial activity of caffeine against plant pathogenic bacteria.

    PubMed

    Sledz, Wojciech; Los, Emilia; Paczek, Agnieszka; Rischka, Jacek; Motyka, Agata; Zoledowska, Sabina; Piosik, Jacek; Lojkowska, Ewa

    2015-01-01

    The objective of the present study was to evaluate the antibacterial properties of a plant secondary metabolite - caffeine. Caffeine is present in over 100 plant species. Antibacterial activity of caffeine was examined against the following plant-pathogenic bacteria: Ralstonia solanacearum (Rsol), Clavibacter michiganesis subsp. sepedonicus (Cms), Dickeya solani (Dsol), Pectobacterium atrosepticum (Pba), Pectobacterium carotovorum subsp. carotovorum (Pcc), Pseudomonas syringae pv. tomato (Pst), and Xanthomonas campestris subsp. campestris (Xcc). MIC and MBC values ranged from 5 to 20 mM and from 43 to 100 mM, respectively. Caffeine increased the bacterial generation time of all tested species and caused changes in cell morphology. The influence of caffeine on the synthesis of DNA, RNA and proteins was investigated in cultures of plant pathogenic bacteria with labelled precursors: [(3)H]thymidine, [(3)H]uridine or (14)C leucine, respectively. RNA biosynthesis was more affected than DNA or protein biosynthesis in bacterial cells treated with caffeine. Treatment of Pba with caffeine for 336 h did not induce resistance to this compound. Caffeine application reduced disease symptoms caused by Dsol on chicory leaves, potato slices, and whole potato tubers. The data presented indicate caffeine as a potential tool for the control of diseases caused by plant-pathogenic bacteria, especially under storage conditions.

  9. Natural Transformation of Azotobacter vinelandii by Adsorbed Chromosomal DNA: Role of Adsorbed DNA Conformation

    NASA Astrophysics Data System (ADS)

    Lv, N.; Zilles, J.; Nguyen, H.

    2008-12-01

    Recent increases in antibiotic resistance among pathogenic microorganisms and the accompanying public health concerns result both from the widespread use of antibiotics and from the transfer of antibiotic resistance genes among microorganisms. To understand the transfer of antibiotic resistance genes and identify efficient measures to minimize these transfers, an interdisciplinary approach was used to identify physical and chemical factors that control the fate and biological availability of extracellular DNA. Quartz crystal microbalance with dissipation (QCM-D) was used to study extracellular DNA adsorption and the conformation of the adsorbed DNA on silica and natural organic matter (NOM) surfaces. Solution chemistry was varied systematically to investigate the role of adsorbed DNA conformation on transformation. Gene transfer was assessed under the same conditions using natural transformation of chromosomal DNA into the soil bacteria Azotobacter vinelandii. DNA adsorbed to both silica and NOM surfaces has a more compact and rigid conformation in the presence of Ca2+ compared to Na+. Extracellular DNA adsorbed on silica and NOM surfaces transformed A. vinelandii. The transformation efficiency of adsorbed DNA was up to 4 orders of magnitude lower than that of dissolved DNA. Preliminary results suggest that the presence of Ca2+ in groundwater (e.g. hardness) reduces the availability of adsorbed DNA for transformation.

  10. Unexpected photoreactivation of Vibrio harveyi bacteria living in ionization environment

    SciTech Connect

    Alifano, P.; Tala, A.; Tredici, S. M.; Nassisi, V.; Siciliano, M. V.

    2011-05-15

    Bacteria undergoing environmental effects is extremely interesting for structural, mechanistic, and evolutionary implications. Luminescent bacteria that have evolved in a specific ambient have developed particular responses and their behavior can give us new suggestions on the task and production of luciferina proteins. To analyze the UV interaction under controlled laboratory conditions, we used photoluminescent bacterial strains belonging to a new species evolutionarily close to Vibrio harveyi sampled from a coastal cave with a high radon content that generates ionizing radiation. The survival of the bacterial strains was analyzed, in the light and in the dark, following a variety of genotoxic treatments including UV radiation exposure. The strains were irradiated by a germicide lamp. The results demonstrated that most of the strains exhibited a low rate of survival after the UV exposure. After irradiation by visible light following the UV exposure, all strains showed a high capability of photoreactivation when grown. This capability was quite unexpected because these bacteria were sampled from a dark ambient without UV radiation. This leads us to hypothesize that the photoreactivation in these bacteria might have been evolved to repair DNA lesions also induced by different radiation sources other than UV (e.g., x-ray) and that the luminescent bacteria might use their own light emission to carry out the photoreactivation. The high capability of photoreactivation of these bacteria was also justified by the results of deconvolution. The deconvolution was applied to the emission spectra and it was able to show evidence of different light peaks. The presence of the visible peak could control the photolysis enzyme.

  11. Methylotrophic bacteria in sustainable agriculture.

    PubMed

    Kumar, Manish; Tomar, Rajesh Singh; Lade, Harshad; Paul, Diby

    2016-07-01

    Excessive use of chemical fertilizers to increase production from available land has resulted in deterioration of soil quality. To prevent further soil deterioration, the use of methylotrophic bacteria that have the ability to colonize different habitats, including soil, sediment, water, and both epiphytes and endophytes as host plants, has been suggested for sustainable agriculture. Methylotrophic bacteria are known to play a significant role in the biogeochemical cycle in soil ecosystems, ultimately fortifying plants and sustaining agriculture. Methylotrophs also improve air quality by using volatile organic compounds such as dichloromethane, formaldehyde, methanol, and formic acid. Additionally, methylotrophs are involved in phosphorous, nitrogen, and carbon cycling and can help reduce global warming. In this review, different aspects of the interaction between methylotrophs and host plants are discussed, including the role of methylotrophs in phosphorus acquisition, nitrogen fixation, phytohormone production, iron chelation, and plant growth promotion, and co-inoculation of these bacteria as biofertilizers for viable agriculture practices.

  12. Genetic transfer in acidophilic bacteria

    SciTech Connect

    Roberto, F.F.; Glenn, A.W.; Bulmer, D.; Ward, T.E.

    1990-01-01

    There is increasing interest in the use of microorganisms to recover metals from ores, as well as to remove sulfur from coal. These so-called bioleaching processes are mediated by a number of bacteria. The best-studied of these organisms are acidophiles including Thiobacillus and Acidiphilium species. Our laboratory has focused on developing genetic strategies to allow the manipulation of acidophilic bacteria to improve and augment their utility in large scale operations. We have recently been successful in employing conjugation for interbacterial transfer of genetic information, as well as in directly transforming Acidiphilium by use of electroporation. We are now testing the properties of IncPl, IncW and IncQ plasmid vectors in Acidiphilium to determine their relative usefulness in routine manipulation of acidophiles and transfer between organisms. This study also allows us to determine the natural ability of these bacteria to transfer genetic material amongst themselves in their particular environment. 21 refs., 3 figs., 2 tabs.

  13. Chemical signature of magnetotactic bacteria.

    PubMed

    Amor, Matthieu; Busigny, Vincent; Durand-Dubief, Mickaël; Tharaud, Mickaël; Ona-Nguema, Georges; Gélabert, Alexandre; Alphandéry, Edouard; Menguy, Nicolas; Benedetti, Marc F; Chebbi, Imène; Guyot, François

    2015-02-10

    There are longstanding and ongoing controversies about the abiotic or biological origin of nanocrystals of magnetite. On Earth, magnetotactic bacteria perform biomineralization of intracellular magnetite nanoparticles under a controlled pathway. These bacteria are ubiquitous in modern natural environments. However, their identification in ancient geological material remains challenging. Together with physical and mineralogical properties, the chemical composition of magnetite was proposed as a promising tracer for bacterial magnetofossil identification, but this had never been explored quantitatively and systematically for many trace elements. Here, we determine the incorporation of 34 trace elements in magnetite in both cases of abiotic aqueous precipitation and of production by the magnetotactic bacterium Magnetospirillum magneticum strain AMB-1. We show that, in biomagnetite, most elements are at least 100 times less concentrated than in abiotic magnetite and we provide a quantitative pattern of this depletion. Furthermore, we propose a previously unidentified method based on strontium and calcium incorporation to identify magnetite produced by magnetotactic bacteria in the geological record.

  14. Radioresistant Bacteria Came From Mars?

    NASA Astrophysics Data System (ADS)

    Pavlov, A.; Kalinin, V.; Konstantinov, A.; Shelegedin, V.

    We propose that the radioresistant bacteria (i.e. Deinococcus radiodurans) has been originated on Mars. This bacteria possesses an ability which should have been ab- solutely "unnecessary" in the Earth environment. It can survive very high doses of the ionizing radiation. Our experiments demonstrate that different kinds of non- radioresistant bacteria are able to develop very high radioresistance ability also. To develop radioresistance we exposed different bacterial cultures to several dozens of cycles of high irradiation. Therefore, radioresistance is not a result of some early spontaneous bacterial mutation but rather a consequence of the very specific plane- tary environment. Polar regions of Mars are the most probable (if not the only) place in the Solar System for such a periodical high-dosage irradiation process. We pro- pose a plausible scenario of where and when such an adaptation process could have taken place and also discuss indirect arguments of the Martian origin of Deinococcus Radiodurans based on their specific genetic structure.

  15. Methylotrophic bacteria in sustainable agriculture.

    PubMed

    Kumar, Manish; Tomar, Rajesh Singh; Lade, Harshad; Paul, Diby

    2016-07-01

    Excessive use of chemical fertilizers to increase production from available land has resulted in deterioration of soil quality. To prevent further soil deterioration, the use of methylotrophic bacteria that have the ability to colonize different habitats, including soil, sediment, water, and both epiphytes and endophytes as host plants, has been suggested for sustainable agriculture. Methylotrophic bacteria are known to play a significant role in the biogeochemical cycle in soil ecosystems, ultimately fortifying plants and sustaining agriculture. Methylotrophs also improve air quality by using volatile organic compounds such as dichloromethane, formaldehyde, methanol, and formic acid. Additionally, methylotrophs are involved in phosphorous, nitrogen, and carbon cycling and can help reduce global warming. In this review, different aspects of the interaction between methylotrophs and host plants are discussed, including the role of methylotrophs in phosphorus acquisition, nitrogen fixation, phytohormone production, iron chelation, and plant growth promotion, and co-inoculation of these bacteria as biofertilizers for viable agriculture practices. PMID:27263015

  16. Intracellular endosymbiotic bacteria of Camponotus species (carpenter ants): systematics, evolution and ultrastructural characterization.

    PubMed

    Schröder, D; Deppisch, H; Obermayer, M; Krohne, G; Stackebrandt, E; Hôlldobler, B; Goebel, W; Gross, R

    1996-08-01

    Intracellular endosymbiotic bacteria inherent to ants of the genus Camponotus were characterized. The bacteria were localized in bacteriocytes, which are specialized cells of both workers and queen ants; these cells are intercalated between epithelial cells of the midgut. The bacteriocytes show a different morphology from the normal epithelial cells and carry a large number of the rod-shaped Gram-negative bacteria free in the cytoplasm. The bacteria were never observed in the neighbouring epithelial cells, but they were found intracellularly in oocytes, strongly indicating a maternal transmission of the bacteria. The 16S DNA encoding rrs loci of the endosymbionts of four species of the genus Camponotus derived either from Germany (C. herculeanus and C. ligniperdus), North America (C. floridanus) or South America (C. rufipes) were cloned after polymerase chain reaction (PCR) amplification using oligonucleotides complementary to all so far known eubacterial rrs sequences. The DNA sequences of the rrs loci of the four endosymbionts were determined, and, using various genus- and species-specific oligonucleotides derived from variable regions in the rrs sequences, the identity of the bacteria present in the bacteriocytes and the ovarian cells was confirmed by PCR and in situ hybridization techniques. Comparison of the 16S DNA sequences with the available database showed the endosymbiotic bacteria to be members of the gamma-subclass of Proteobacteria. They formed a distinct taxonomic group, a sister taxon of the taxons defined by the tsetse fly and aphid endosymbionts. Within the gamma-subclass, the cluster of the ant, tsetse fly and aphid endosymbionts are placed adjacent to the family of Enterobacteriaceae. The evolutionary tree of the ant endosymbionts reflects the systematic classification and geographical distribution of their host insects, indicating an early co-evolution of the symbiotic partners and a vertical transmission of the bacteria.

  17. Intracellular endosymbiotic bacteria of Camponotus species (carpenter ants): systematics, evolution and ultrastructural characterization.

    PubMed

    Schröder, D; Deppisch, H; Obermayer, M; Krohne, G; Stackebrandt, E; Hôlldobler, B; Goebel, W; Gross, R

    1996-08-01

    Intracellular endosymbiotic bacteria inherent to ants of the genus Camponotus were characterized. The bacteria were localized in bacteriocytes, which are specialized cells of both workers and queen ants; these cells are intercalated between epithelial cells of the midgut. The bacteriocytes show a different morphology from the normal epithelial cells and carry a large number of the rod-shaped Gram-negative bacteria free in the cytoplasm. The bacteria were never observed in the neighbouring epithelial cells, but they were found intracellularly in oocytes, strongly indicating a maternal transmission of the bacteria. The 16S DNA encoding rrs loci of the endosymbionts of four species of the genus Camponotus derived either from Germany (C. herculeanus and C. ligniperdus), North America (C. floridanus) or South America (C. rufipes) were cloned after polymerase chain reaction (PCR) amplification using oligonucleotides complementary to all so far known eubacterial rrs sequences. The DNA sequences of the rrs loci of the four endosymbionts were determined, and, using various genus- and species-specific oligonucleotides derived from variable regions in the rrs sequences, the identity of the bacteria present in the bacteriocytes and the ovarian cells was confirmed by PCR and in situ hybridization techniques. Comparison of the 16S DNA sequences with the available database showed the endosymbiotic bacteria to be members of the gamma-subclass of Proteobacteria. They formed a distinct taxonomic group, a sister taxon of the taxons defined by the tsetse fly and aphid endosymbionts. Within the gamma-subclass, the cluster of the ant, tsetse fly and aphid endosymbionts are placed adjacent to the family of Enterobacteriaceae. The evolutionary tree of the ant endosymbionts reflects the systematic classification and geographical distribution of their host insects, indicating an early co-evolution of the symbiotic partners and a vertical transmission of the bacteria. PMID:8866472

  18. Unique and Universal Features of Epsilonproteobacterial Origins of Chromosome Replication and DnaA-DnaA Box Interactions

    PubMed Central

    Jaworski, Pawel; Donczew, Rafal; Mielke, Thorsten; Thiel, Marcel; Oldziej, Stanislaw; Weigel, Christoph; Zawilak-Pawlik, Anna

    2016-01-01

    In bacteria, chromosome replication is initiated by the interaction of the initiator protein DnaA with a defined region of a chromosome at which DNA replication starts (oriC). While DnaA proteins share significant homology regardless of phylogeny, oriC regions exhibit more variable structures. The general architecture of oriCs is universal, i.e., they are composed of a cluster of DnaA binding sites, a DNA-unwinding element, and sequences that bind regulatory proteins. However, detailed structures of oriCs are shared by related species while being significantly different in unrelated bacteria. In this work, we characterized Epsilonproteobacterial oriC regions. Helicobacter pylori was the only species of the class for which oriC was characterized. A few unique features were found such as bipartite oriC structure, not encountered in any other Gram-negative species, and topology-sensitive DnaA-DNA interactions, which have not been found in any other bacterium. These unusual H. pylori oriC features raised questions of whether oriC structure and DnaA-DNA interactions are unique to this bacterium or whether they are common to related species. By in silico and in vitro analyses we identified putative oriCs in three Epsilonproteobacterial species: pathogenic Arcobacter butzleri, symbiotic Wolinella succinogenes, and free-living Sulfurimonas denitrificans. We propose that oriCs typically co-localize with ruvC-dnaA-dnaN in Epsilonproteobacteria, with the exception of Helicobacteriaceae species. The clusters of DnaA boxes localize upstream (oriC1) and downstream (oriC2) of dnaA, and they likely constitute bipartite origins. In all cases, DNA unwinding was shown to occur in oriC2. Unlike the DnaA box pattern, which is not conserved in Epsilonproteobacterial oriCs, the consensus DnaA box sequences and the mode of DnaA-DnaA box interactions are common to the class. We propose that the typical Epsilonproteobacterial DnaA box consists of the core nucleotide sequence 5′-TTCAC-3

  19. Adaptation, Bacteria and Maxwell's Demons

    NASA Astrophysics Data System (ADS)

    Galajda, Peter; Keymer, Juan E.; Austin, Robert H.

    2007-03-01

    We propose a method to study the adaptation of bacterial populations with an asymmetric wall of Maxwell Demon openings. A Maxwell Demon opening is a funnel which is easier to enter than to leave. The interaction of swimming cells with such a Maxwell Demon Wall results in a population density separation, in apparent (but not real) violation of the Second Law of Thermodynamics, as we will show. Bacteria can be exposed to spatial challenges in order to move to e. g. higher food levels. The question we address in these experiments is: do the bacteria adapt and overcome the Maxwell Demon Wall?

  20. Isolation of naphthalene-degrading bacteria from tropical marine sediments.

    PubMed

    Zhuang, W Q; Tay, J H; Maszenan, A M; Tay, S T L

    2003-01-01

    Oil pollution is a major environmental concern in many countries, and this has led to a concerted effort in studying the feasibility of using oil-degrading bacteria for bioremediation. Although many oil-degrading bacteria have been isolated from different environments, environmental conditions can impose a selection pressure on the types of bacteria that can reside in a particular environment. This study reports the successful isolation of two indigenous naphthalene-degrading bacteria from oil-contaminated tropical marine sediments by enrichment culture. Strains MN-005 and MN-006 were characterized using an extensive range of biochemical tests. The 16S ribosomal deoxyribonucleic acid (rDNA) sequence analysis was also performed for the two strains. Their naphthalene degradation capabilities were determined using gas chromatography and DAPI counting of bacterial cells. Strains MN-005 and MN-006 are phenotypically and phylogenetically different from each other, and belong to the genera Staphylococcus and Micrococcus, respectively. Strains MN-005 and MN-006 had maximal specific growth rates (micro(max)) of 0.082 +/- 0.008 and 0.30 +/- 0.02 per hour, respectively, and half-saturation constants (K(s)) of 0.79 +/- 0.10 and 2.52 +/- 0.32 mg per litre, respectively. These physiological and growth studies are useful in assessing the potential of these indigenous isolates for in situ or ex situ naphthalene pollutant bioremediation in tropical marine environments.

  1. 'Rare biosphere' bacteria as key phenanthrene degraders in coastal seawaters.

    PubMed

    Sauret, Caroline; Séverin, Tatiana; Vétion, Gilles; Guigue, Catherine; Goutx, Madeleine; Pujo-Pay, Mireille; Conan, Pascal; Fagervold, Sonja K; Ghiglione, Jean-François

    2014-11-01

    By coupling DNA-SIP and pyrosequencing approaches, we identified Cycloclasticus sp. as a keystone degrader of polycyclic aromatic hydrocarbons (PAH) despite being a member of the 'rare biosphere' in NW Mediterranean seawaters. We discovered novel PAH-degrading bacteria (Oceanibaculum sp., Sneathiella sp.) and we identified other groups already known to possess this function (Alteromonas sp., Paracoccus sp.). Together with Cycloclasticus sp., these groups contributed to potential in situ phenanthrene degradation at a rate >0.5 mg l(-1) day(-1), sufficient to account for a considerable part of PAH degradation. Further, we characterized the PAH-tolerant bacterial communities, which were much more diverse in the polluted site by comparison to unpolluted marine references. PAH-tolerant bacteria were also members of the rare biosphere, such as Glaciecola sp. Collectively, these data show the complex interactions between PAH-degraders and PAH-tolerant bacteria and provide new insights for the understanding of the functional ecology of marine bacteria in polluted waters.

  2. Antibiotic-producing bacteria from stag beetle mycangia.

    PubMed

    Miyashita, Atsushi; Hirai, Yuuki; Sekimizu, Kazuhisa; Kaito, Chikara

    2015-02-01

    The search for new antibiotics or antifungal agents is crucial for the chemotherapies of infectious diseases. The limited resource of soil bacteria makes it difficult to discover such new drug candidate. We, therefore, focused on another bacterial resource than soil bacteria, the microbial flora of insect species. In the present study, we isolated 40 strains of bacteria and fungi from the mycangia of three species of stag beetle, Dorcus hopei binodulosus, Dorcus rectus, and Dorcus titanus pilifer. We identified those species with their ribosomal DNA sequences, and revealed that Klebsiella spp. are the most frequent symbiont in the stag beetle mycangia. We examined whether these microorganisms produce antibiotics against a Gram-negative bacterium, Escherichia coli, a Gram-positive bacterium, Staphylococcus aureus, or a fungus, Cryptococcus neoformans. Culture supernatants from 33, 29, or 18 strains showed antimicrobial activity against E. coli, S. aureus, or C. neoformans, respectively. These findings suggest that bacteria present in the mycangia of stag beetles are useful resources for screening novel antibiotics. PMID:25639488

  3. The DNA-Uptake Process of Naturally Competent Vibrio cholerae.

    PubMed

    Matthey, Noémie; Blokesch, Melanie

    2016-02-01

    The sophisticated DNA-uptake machinery used during natural transformation is still poorly characterized, especially in Gram-negative bacteria where the transforming DNA has to cross two membranes as well as the peptidoglycan layer before entering the cytoplasm. The DNA-uptake machinery was hypothesized to take the form of a pseudopilus, which, upon repeated cycles of extension and retraction, would pull external DNA towards the cell surface or into the periplasmic space, followed by translocation across the cytoplasmic membrane. In this review, we summarize recent advances on the DNA-uptake machinery of V. cholerae, highlighting the presence of an extended competence-induced pilus and the contribution of a conserved DNA-binding protein that acts as a ratchet and reels DNA into the periplasm. PMID:26614677

  4. Charting the Structure and Energetics of Packaged DNA in Bacteriophages

    NASA Astrophysics Data System (ADS)

    Qiu, Xiangyun; Rau, Donald C.; Parsegian, V. Adrian; Fang, Li Tai; Knobler, Charles M.; Gelbart, William M.

    2009-03-01

    Many bacterial viruses resort to pressure in order to infect bacteria, e.g., lambda phage stores its dsDNA genome at surprisingly high pressure and then uses this pressure to drive delivery of the genome. We report on a biophysical interrogation of the DNA configuration and pressure in lambda phage by combining structural and thermodynamic measurements with theoretical modeling. Changes in DNA organization in the capsid are monitored using solution small angle x-ray scattering (SAXS). We vary the DNA-DNA repulsion and DNA bending contributions to the capsid pressure by changing salt concentrations and packaged length, and augment SAXS data with osmotic stress measurements to elicit the evolving structure and energetics of the packaged DNA.

  5. Methods for Engineering Sulfate Reducing Bacteria of the Genus Desulfovibrio

    SciTech Connect

    Chhabra, Swapnil R; Keller, Kimberly L.; Wall, Judy D.

    2011-03-15

    Sulfate reducing bacteria are physiologically important given their nearly ubiquitous presence and have important applications in the areas of bioremediation and bioenergy. This chapter provides details on the steps used for homologous-recombination mediated chromosomal manipulation of Desulfovibrio vulgaris Hildenborough, a well-studied sulfate reducer. More specifically, we focus on the implementation of a 'parts' based approach for suicide vector assembly, important aspects of anaerobic culturing, choices for antibiotic selection, electroporation-based DNA transformation, as well as tools for screening and verifying genetically modified constructs. These methods, which in principle may be extended to other sulfate-reducing bacteria, are applicable for functional genomics investigations, as well as metabolic engineering manipulations.

  6. Isolation and identification of bacteria from spent nuclear fuel pools.

    PubMed

    Chicote, Eduardo; García, Ana M; Moreno, Diego A; Sarró, M Isabel; Lorenzo, Petra I; Montero, Felipe

    2005-04-01

    The aim of the present research was to isolate and identify bacteria from spent nuclear fuel pools of a Spanish nuclear power plant. Water samples were collected and inoculated onto different culture media to isolate the highest number of species. 16S rDNA fragments from colonies growing on solid media were amplified and analysed by denaturing gradient gel electrophoresis. Sequencing revealed the presence of 21 different bacteria belonging to several phylogenetic groups (alpha, beta, and gamma-Proteobacteria, Actinomycetales, Flavobacterium, and the Bacillus/Staphylococcus group). The isolation of these microorganisms in this particular environment (oligotrophic and radioactive) is highly interesting because of the possibility of their being used for the bioremediation of radionuclide-contaminated waters.

  7. Comparative Genomics via Wavelet Analysis for Closely Related Bacteria

    NASA Astrophysics Data System (ADS)

    Song, Jiuzhou; Ware, Tony; Liu, Shu-Lin; Surette, M.

    2004-12-01

    Comparative genomics has been a valuable method for extracting and extrapolating genome information among closely related bacteria. The efficiency of the traditional methods is extremely influenced by the software method used. To overcome the problem here, we propose using wavelet analysis to perform comparative genomics. First, global comparison using wavelet analysis gives the difference at a quantitative level. Then local comparison using keto-excess or purine-excess plots shows precise positions of inversions, translocations, and horizontally transferred DNA fragments. We firstly found that the level of energy spectra difference is related to the similarity of bacteria strains; it could be a quantitative index to describe the similarities of genomes. The strategy is described in detail by comparisons of closely related strains: S.typhi CT18, S.typhi Ty2, S.typhimurium LT2, H.pylori 26695, and H.pylori J99.

  8. DNA Microarrays

    NASA Astrophysics Data System (ADS)

    Nguyen, C.; Gidrol, X.

    Genomics has revolutionised biological and biomedical research. This revolution was predictable on the basis of its two driving forces: the ever increasing availability of genome sequences and the development of new technology able to exploit them. Up until now, technical limitations meant that molecular biology could only analyse one or two parameters per experiment, providing relatively little information compared with the great complexity of the systems under investigation. This gene by gene approach is inadequate to understand biological systems containing several thousand genes. It is essential to have an overall view of the DNA, RNA, and relevant proteins. A simple inventory of the genome is not sufficient to understand the functions of the genes, or indeed the way that cells and organisms work. For this purpose, functional studies based on whole genomes are needed. Among these new large-scale methods of molecular analysis, DNA microarrays provide a way of studying the genome and the transcriptome. The idea of integrating a large amount of data derived from a support with very small area has led biologists to call these chips, borrowing the term from the microelectronics industry. At the beginning of the 1990s, the development of DNA chips on nylon membranes [1, 2], then on glass [3] and silicon [4] supports, made it possible for the first time to carry out simultaneous measurements of the equilibrium concentration of all the messenger RNA (mRNA) or transcribed RNA in a cell. These microarrays offer a wide range of applications, in both fundamental and clinical research, providing a method for genome-wide characterisation of changes occurring within a cell or tissue, as for example in polymorphism studies, detection of mutations, and quantitative assays of gene copies. With regard to the transcriptome, it provides a way of characterising differentially expressed genes, profiling given biological states, and identifying regulatory channels.

  9. Transport of microspheres and indigenous bacteria through a sandy aquifer: Results of natural- and forced-gradient tracer experiments

    USGS Publications Warehouse

    Harvey, R.W.; George, L.H.; Smith, R.L.; LeBlanc, D.R.

    1989-01-01

    Transport of indigenous bacteria through sandy aquifer sediments was investigated in forced- and natural-gradient tracer teste. A diverse population of bacteria was collected and concentrated from groundwater at the site, stained with a DNA-specific fluorochrome, and injected back into the aquifer. Included with the injectate were a conservative tracer (Br- or Cl-) and bacteria-sized (0.2-1.3-??m) microspheres having carboxylated, carbonyl, or neutral surfaces. Transport of stained bacteria and all types and size classes of microspheres was evident. In the natural-gradient test, both surface characteristics and size of microspheres affected attenuation. Surface characteristics had the greatest effect upon retardation. Peak break-through of DAPI-stained bacteria (forced-gradient experiment) occurred well in advance of bromide at the more distal sampler. Transport behavior of bacteria was substantially different from that of carboxylated microspheres of comparable size. ?? 1988 American Chemical Society.

  10. Influence of Chicken Manure Fertilization on Antibiotic-Resistant Bacteria in Soil and the Endophytic Bacteria of Pakchoi

    PubMed Central

    Yang, Qingxiang; Zhang, Hao; Guo, Yuhui; Tian, Tiantian

    2016-01-01

    Animal manure is commonly used as fertilizer for agricultural crops worldwide, even though it is believed to contribute to the spread of antibiotic resistance from animal intestines to the soil environment. However, it is unclear whether and how there is any impact of manure fertilization on populations and community structure of antibiotic-resistant endophytic bacteria (AREB) in plant tissues. To investigate the effect of manure and organic fertilizer on endophytic bacterial communities, pot experiments were performed with pakchoi grown with the following treatments: (1) non-treated; (2) chicken manure-treated and (3) organic fertilizer-treated. Manure or organic fertilizer significantly increased the abundances of total cultivable endophytic bacteria (TCEB) and AREB in pakchoi, and the effect of chicken manure was greater than that of organic fertilizer. Further, 16S rDNA sequencing and the phylogenetic analysis indicated that chicken manure or organic fertilizer application increased the populations of multiple antibiotic-resistant bacteria (MARB) in soil and multiple antibiotic-resistant endophytic bacteria (MAREB) in pakchoi. The identical multiple antibiotic-resistant bacterial populations detected in chicken manure, manure- or organic fertilizer-amended soil and the vegetable endophytic system were Brevundimonas diminuta, Brachybacterium sp. and Bordetella sp., suggesting that MARB from manure could enter and colonize the vegetable tissues through manure fertilization. The fact that some human pathogens with multiple antibiotic resistance were detected in harvested vegetables after growing in manure-amended soil demonstrated a potential threat to human health. PMID:27376311

  11. Influence of Chicken Manure Fertilization on Antibiotic-Resistant Bacteria in Soil and the Endophytic Bacteria of Pakchoi.

    PubMed

    Yang, Qingxiang; Zhang, Hao; Guo, Yuhui; Tian, Tiantian

    2016-01-01

    Animal manure is commonly used as fertilizer for agricultural crops worldwide, even though it is believed to contribute to the spread of antibiotic resistance from animal intestines to the soil environment. However, it is unclear whether and how there is any impact of manure fertilization on populations and community structure of antibiotic-resistant endophytic bacteria (AREB) in plant tissues. To investigate the effect of manure and organic fertilizer on endophytic bacterial communities, pot experiments were performed with pakchoi grown with the following treatments: (1) non-treated; (2) chicken manure-treated and (3) organic fertilizer-treated. Manure or organic fertilizer significantly increased the abundances of total cultivable endophytic bacteria (TCEB) and AREB in pakchoi, and the effect of chicken manure was greater than that of organic fertilizer. Further, 16S rDNA sequencing and the phylogenetic analysis indicated that chicken manure or organic fertilizer application increased the populations of multiple antibiotic-resistant bacteria (MARB) in soil and multiple antibiotic-resistant endophytic bacteria (MAREB) in pakchoi. The identical multiple antibiotic-resistant bacterial populations detected in chicken manure, manure- or organic fertilizer-amended soil and the vegetable endophytic system were Brevundimonas diminuta, Brachybacterium sp. and Bordetella sp., suggesting that MARB from manure could enter and colonize the vegetable tissues through manure fertilization. The fact that some human pathogens with multiple antibiotic resistance were detected in harvested vegetables after growing in manure-amended soil demonstrated a potential threat to human health.

  12. Influence of Chicken Manure Fertilization on Antibiotic-Resistant Bacteria in Soil and the Endophytic Bacteria of Pakchoi.

    PubMed

    Yang, Qingxiang; Zhang, Hao; Guo, Yuhui; Tian, Tiantian

    2016-01-01

    Animal manure is commonly used as fertilizer for agricultural crops worldwide, even though it is believed to contribute to the spread of antibiotic resistance from animal intestines to the soil environment. However, it is unclear whether and how there is any impact of manure fertilization on populations and community structure of antibiotic-resistant endophytic bacteria (AREB) in plant tissues. To investigate the effect of manure and organic fertilizer on endophytic bacterial communities, pot experiments were performed with pakchoi grown with the following treatments: (1) non-treated; (2) chicken manure-treated and (3) organic fertilizer-treated. Manure or organic fertilizer significantly increased the abundances of total cultivable endophytic bacteria (TCEB) and AREB in pakchoi, and the effect of chicken manure was greater than that of organic fertilizer. Further, 16S rDNA sequencing and the phylogenetic analysis indicated that chicken manure or organic fertilizer application increased the populations of multiple antibiotic-resistant bacteria (MARB) in soil and multiple antibiotic-resistant endophytic bacteria (MAREB) in pakchoi. The identical multiple antibiotic-resistant bacterial populations detected in chicken manure, manure- or organic fertilizer-amended soil and the vegetable endophytic system were Brevundimonas diminuta, Brachybacterium sp. and Bordetella sp., suggesting that MARB from manure could enter and colonize the vegetable tissues through manure fertilization. The fact that some human pathogens with multiple antibiotic resistance were detected in harvested vegetables after growing in manure-amended soil demonstrated a potential threat to human health. PMID:27376311

  13. Application of immobilized synthetic anti-lipopolysaccharide peptides for the isolation and detection of bacteria.

    PubMed

    Sandetskaya, N; Engelmann, B; Brandenburg, K; Kuhlmeier, D

    2015-08-01

    The molecular detection of microorganisms in liquid samples generally requires their enrichment or isolation. The aim of our study was to evaluate the capture and pre-concentration of bacteria by immobilized particular cationic antimicrobial peptides, called synthetic anti-lipopolysaccharide peptides (SALP). For the proof-of-concept and screening of different SALP, the peptides were covalently immobilized on glass slides, and the binding of bacteria was confirmed by microscopic examination of the slides or their scanning, in case of fluorescent bacterial cells. The most efficient SALP was further tethered to magnetic beads. SALP beads were used for the magnetic capture of Escherichia coli in liquid samples. The efficiency of this strategy was evaluated using polymerase chain reaction (PCR). Covalently immobilized SALP were capable of capturing bacteria in liquid samples. However, PCR was hampered by the unspecific binding of DNA to the positively charged peptide. We developed a method for DNA recovery by the enzymatic digestion of the peptide, which allowed for a successful PCR, though the method had its own adverse impact on the detection and, thus, did not allow for the reliable quantitative analysis of the pathogen enrichment. Immobilized SALP can be used as capture molecules for bacteria in liquid samples and can be recommended for the design of the assays or decontamination of the fluids. For the accurate subsequent detection of bacteria, DNA-independent methods should be used.

  14. Occurrence of sulfonamide and tetracycline-resistant bacteria and resistance genes in aquaculture environment.

    PubMed

    Gao, Panpan; Mao, Daqing; Luo, Yi; Wang, Limei; Xu, Bingjie; Xu, Lin

    2012-05-01

    The occurrence of sulfonamide and tetracycline resistance and their pollution profile in the aquaculture environment of Tianjin, northern China, were investigated. The presence of antibiotic-resistant bacteria was identified and the corresponding antibiotic resistance genes (ARGs) were quantified at 6 aquaculture farms in Tianjin. Sulfonamide-resistance genes were prevalent and their concentrations were the highest detected (3.0 × 10(-5) to 3.3 × 10(-4) for sul1/16S rDNA, 2.0 × 10(-4) to 1.8 × 10(-3) for sul2/16S rDNA) among the various ARGs, most likely because the use of sulfonamides is more prevalent than tetracyclines in this area. Bacillus was the most dominant bacterial genus in both sulfamethoxazole resistant bacteria (63.27% of the total resistant bacteria) and tetracycline-resistant bacteria (57.14% of the total resistant bacteria). At least two of those genes (tetM, tetO, tetT, tetW, sul1 and sul2) were detected in the isolates of Bacillus cereus, Bacillus subtilis, Bacillus megaterium and Acinetobacter lwofii, and all of the above genes were detected in B. cereus, suggesting the occurrence of multi-resistance in the studied area. The genetic transfer of sul1 between intestinal bacteria (e.g., Enterococcus spp.) and indigenous bacteria (e.g., Bacillus spp.) was implied by phylogenetic analysis. Several strains of resistant opportunistic pathogens (e.g., Acinetobacter spp.) were found in indigenous bacteria, which increase the risk of ARGs to public health. Overall, this is the first study to comprehensively investigate the antibiotic resistance profile by analyzing the species of antibiotic-resistant bacteria and adopting qualitative and quantitative methods to investigate ARGs at a typical aquaculture area in northern China.

  15. Manipulating Genetic Material in Bacteria

    NASA Technical Reports Server (NTRS)

    1998-01-01

    Lisa Crawford, a graduate research assistant from the University of Toledo, works with Laurel Karr of Marshall Space Flight Center (MSFC) in the molecular biology laboratory. They are donducting genetic manipulation of bacteria and yeast for the production of large amount of desired protein. Photo credit: NASA/Marshall Space Flight Center (MSFC)

  16. Role of Bacteria in Oncogenesis

    PubMed Central

    Chang, Alicia H.; Parsonnet, Julie

    2010-01-01

    Summary: Although scientific knowledge in viral oncology has exploded in the 20th century, the role of bacteria as mediators of oncogenesis has been less well elucidated. Understanding bacterial carcinogenesis has become increasingly important as a possible means of cancer prevention. This review summarizes clinical, epidemiological, and experimental evidence as well as possible mechanisms of bacterial induction of or protection from malignancy. PMID:20930075

  17. Hydrocarbon degradation by antarctic bacteria

    SciTech Connect

    Cavanagh, J.A.E.; Nichols, P.D.; McMeekin, T.A.; Franzmann, P.D.

    1996-12-31

    Bacterial cultures obtained from sediment samples collected during a trial oil spill experiment conducted at Airport beach, Eastern Antarctica were selectively enriched for n-alkane-degrading and phenanthrenedegrading bacteria. Samples were collected from a control site and sites treated with different hydrocarbon mixtures - Special Antarctic blend (SAB), BP-Visco and orange roughy oils. One set of replicate sites was also treated with water from Organic Lake which had previously been shown to contain hydrocarbon-degrading bacteria. No viable bacteria were obtained from samples collected from sites treated with orange roughy oil. Extensive degradation of n-alkanes by enrichment cultures obtained from sites treated with SAB and BP-Visco occurred at both 25{degrees}C and 10{degrees}C. Extensive degradation of phenanthrene also occurred in enrichment cultures from these sites grown at 25{degrees}C. Concurrent increases of polar lipid in these cultures were also observed. The presence of 1,4-naphthaquinone and 1-naphthol during the growth of the cultures on phenanthrene is unusual and warrants further investigation of the mechanism of phenanthrene-degradation by these Antarctic bacteria.

  18. Heteropolysaccharides from lactic acid bacteria.

    PubMed

    De Vuyst, L; Degeest, B

    1999-04-01

    Microbial exopolysaccharides are biothickeners that can be added to a wide variety of food products, where they serve as viscosifying, stabilizing, emulsifying or gelling agents. Numerous exopolysaccharides with different composition, size and structure are synthesized by lactic acid bacteria. The heteropolysaccharides from both mesophilic and thermophilic lactic acid bacteria have received renewed interest recently. Structural analysis combined with rheological studies revealed that there is considerable variation among the different exopolysaccharides; some of them exhibit remarkable thickening and shear-thinning properties and display high intrinsic viscosities. Hence, several slime-producing lactic acid bacterium strains and their biopolymers have interesting functional and technological properties, which may be exploited towards different products, in particular, natural fermented milks. However, information on the biosynthesis, molecular organization and fermentation conditions is rather scarce, and the kinetics of exopolysaccharide formation are poorly described. Moreover, the production of exopolysaccharides is low and often unstable, and their downstream processing is difficult. This review particularly deals with microbiological, biochemical and technological aspects of heteropolysaccharides from, and their production by, lactic acid bacteria. The chemical composition and structure, the biosynthesis, genetics and molecular organization, the nutritional and physiological aspects, the process technology, and both food additive and in situ applications (in particular in yogurt) of heterotype exopolysaccharides from lactic acid bacteria are described. Where appropriate, suggestions are made for strain improvement, enhanced productivities and advanced modification and production processes (involving enzyme and/or fermentation technology) that may contribute to the economic soundness of applications with this promising group of biomolecules.

  19. Antibacterial susceptibility of plaque bacteria.

    PubMed

    Newman, M G; Hulem, C; Colgate, J; Anselmo, C

    1979-07-01

    Selected anaerobic, capnophilic and facultative bacteria isolated from patients with various forms of periodontal health and disease were tested for their susceptibility to antibiotics and antimicrobial agents. Specific bactericidal and minimum inhibitory concentrations were compared to disc zone diameters, thereby generating new standards for the potential selection of antimicrobial agents. PMID:286720

  20. Photosynthetic reaction centers in bacteria

    SciTech Connect

    Norris, J.R. Univ. of Chicago, IL ); Schiffer, M. )

    1990-07-30

    The photochemistry of photosynthesis begins in complexes called reaction centers. These have become model systems to study the fundamental process by which plants and bacteria convert and store solar energy as chemical free energy. In green plants, photosynthesis occurs in two systems, each of which contains a different reaction center, working in series. In one, known as photosystem 1, oxidized nicotinamide adenine dinucleotide phosphate (NADP[sup +]) is reduced to NADPH for use in a series of dark reactions called the Calvin cycle, named for Nobel Laureate Melvin Calvin, by which carbon dioxide is converted into useful fuels such as carbohydrates and sugars. In the other half of the photosynthetic machinery of green plants, called photosystem 2, water is oxidized to produce molecular oxygen. A different form of photosynthesis occurs in photosynthetic bacteria, which typically live at the bottom of ponds and feed on organic debris. Two main types of photosynthetic bacteria exist: purple and green. Neither type liberates oxygen from water. Instead, the bacteria feed on organic media or inorganic materials, such as sulfides, which are easier to reduce or oxidize than carbon dioxide or water. Perhaps in consequence, their photosynthetic machinery is simpler than that of green, oxygen-evolving plants and their primary photochemistry is better understood.

  1. Antibacterial susceptibility of plaque bacteria.

    PubMed

    Newman, M G; Hulem, C; Colgate, J; Anselmo, C

    1979-07-01

    Selected anaerobic, capnophilic and facultative bacteria isolated from patients with various forms of periodontal health and disease were tested for their susceptibility to antibiotics and antimicrobial agents. Specific bactericidal and minimum inhibitory concentrations were compared to disc zone diameters, thereby generating new standards for the potential selection of antimicrobial agents.

  2. Marine bacteria which produce tetrodotoxin.

    PubMed Central

    Simidu, U; Noguchi, T; Hwang, D F; Shida, Y; Hashimoto, K

    1987-01-01

    A number of type strains of marine bacteria, including members of the family Vibrionaceae, were cultured and examined for tetrodotoxin productivity by high-performance liquid chromatography and gas chromatography-mass spectrometry. Most of the Vibrionaceae strains produced tetrodotoxin, anhydrotetrodotoxin, or both. PMID:3310884

  3. Endobiotic bacteria and their pathogenic potential in cnidarian tentacles

    NASA Astrophysics Data System (ADS)

    Schuett, Christian; Doepke, Hilke

    2010-09-01

    Endobiotic bacteria colonize the tentacles of cnidaria. This paper provides first insight into the bacterial spectrum and its potential of pathogenic activities inside four cnidarian species. Sample material originating from Scottish waters comprises the jellyfish species Cyanea capillata and C. lamarckii, hydrozoa Tubularia indivisa and sea anemone Sagartia elegans. Mixed cultures of endobiotic bacteria, pure cultures selected on basis of haemolysis, but also lyophilized samples were prepared from tentacles and used for DGGE-profiling with subsequent phylogenetic analysis of 16S rDNA fragments. Bacteria were detected in each of the cnidarian species tested. Twenty-one bacterial species including four groups of closely related organisms were found in culture material. The species within these groups could not be differentiated from each other (one group of Pseudoalteromonas spp., two groups of Shewanella spp., one group of Vibrio spp.). Each of the hosts exhibits a specific endobacterial spectrum. Solely Cyanea lamarckii harboured Moritella viscosa. Only in Cyanea capillata, members of the Shewanella group #2 and the species Pseudoalteromonas arctica, Shewanella violacea, Sulfitobacter pontiacus and Arcobacter butzleri were detected. Hydrozoa Tubularia indivisa provided an amazingly wide spectrum of nine bacterial species. Exclusively, in the sea anemone Sagartia elegans, the bacterial species P. aliena was found. Overall eleven bacterial species detected were described recently as novel species. Four 16S rDNA fragments generated from lyophilized material displayed extremely low relationship to their next neighbours. These organisms are regarded as members of the endobiotic “terra incognita”. Since the origin of cnidarian toxins is unclear, the possible pathogenic activity of endobiotic bacteria has to be taken into account. Literature data show that their next neighbours display an interesting diversity of haemolytic, septicaemic and necrotic actions including

  4. INTERNAL AMPLIFICATION CONTROL FOR USE IN QUANTITATIVE POLYMERASE CHAIN REACTION FECAL INDICATOR BACTERIA ASSAYS

    EPA Science Inventory

    Quantitative polymerase chain reaction (QPCR) can be used as a rapid method for detecting fecal indicator bacteria. Because false negative results can be caused by PCR inhibitors that co-extract with the DNA samples, an internal amplification control (IAC) should be run with eac...

  5. Comparison of phenotypic and molecular tests to identify lactic acid bacteria.

    PubMed

    Moraes, Paula Mendonça; Perin, Luana Martins; Júnior, Abelardo Silva; Nero, Luís Augusto

    2013-01-01

    Twenty-nine lactic acid bacteria (LAB) isolates were submitted for identification using Biolog, API50CHL, 16S rDNA sequencing, and species-specific PCR reactions. The identification results were compared, and it was concluded that a polyphasic approach is necessary for proper LAB identification, being the molecular analyzes the most reliable. PMID:24159291

  6. Detection and mapping of homologous, repeated and amplified DNA sequences by DNA renaturation in agarose gels.

    PubMed Central

    Roninson, I B

    1983-01-01

    A new molecular hybridization approach to the analysis of complex genomes has been developed. Tracer and driver DNAs were digested with the same restriction enzyme(s), and tracer DNA was labeled with 32P using T4 DNA polymerase. Tracer DNA was mixed with an excess amount of driver, and the mixture was electrophoresed in an agarose gel. Following electrophoresis, DNA was alkali-denatured in situ and allowed to reanneal in the gel, so that tracer DNA fragments could hybridize to the driver only when homologous driver DNA sequences were present at the same place in the gel, i.e. within a restriction fragment of the same size. After reannealing, unhybridized single-stranded DNA was digested in situ with S1 nuclease. The hybridized tracer DNA was detected by autoradiography. The general applicability of this technique was demonstrated in the following experiments. The common EcoRI restriction fragments were identified in the genomes of E. coli and four other species of bacteria. Two of these fragments are conserved in all Enterobacteriaceae. In other experiments, repeated EcoRI fragments of eukaryotic DNA were visualized as bands of various intensity after reassociation of a total genomic restriction digest in the gel. The situation of gene amplification was modeled by the addition of varying amounts of lambda phage DNA to eukaryotic DNA prior to restriction enzyme digestion. Restriction fragments of lambda DNA were detectable at a ratio of 15 copies per chicken genome and 30 copies per human genome. This approach was used to detect amplified DNA fragments in methotrexate (MTX)-resistant mouse cells and to identify commonly amplified fragments in two independently derived MTX-resistant lines. Images PMID:6310499

  7. Optical DNA

    NASA Astrophysics Data System (ADS)

    Vijaywargi, Deepak; Lewis, Dave; Kirovski, Darko

    A certificate of authenticity (COA) is an inexpensive physical object with a random and unique structure S which is hard to near-exactly replicate. An inexpensive device should be able to scan object’s physical “fingerprint,” a set of features that represents S. In this paper, we explore one set of requirements that optical media such as DVDs should satisfy, to be considered as COAs. As manufacturing of such media produces inevitable errors, we use the locations and count of these errors as a “fingerprint” for each optical disc: its optical DNA. The “fingerprint” is signed using publisher’s private-key and the resulting signature is stored onto the optical medium using a post-production process. Standard DVD players with altered firmware that includes publisher’s public-key, should be able to verify the authenticity of DVDs protected with optical DNA. Our key finding is that for the proposed protocol, only DVDs with exceptional wear-and-tear characteristics would result in an inexpensive and viable anti-counterfeiting technology.

  8. Influence of water chlorination on the counting of bacteria with DAPI (4',6-diamidino-2-phenylindole).

    PubMed Central

    Saby, S; Sibille, I; Mathieu, L; Paquin, J L; Block, J C

    1997-01-01

    Counting bacteria in drinking water samples by the epifluorescence technique after 4',6-diamidino-2-phenylindole (DAPI) staining is complicated by the fact that bacterial fluorescence varies with exposure of the cells to sodium hypochlorite. An Escherichia coli laboratory-grown suspension treated with sodium hypochlorite (5 to 15 mg of chlorine liter-1) for 90 min was highly fluorescent after DAPI staining probably due to cell membrane permeation and better and DAPI diffusion. At chlorine concentrations greater than 25 mg liter-1, DAPI-stained bacteria had only a low fluorescence. Stronger chlorine doses altered the DNA structure, preventing the DAPI from complexing with the DNA. When calf thymus DNA was exposed to sodium hypochlorite (from 15 to 50 mg of chlorine liter-1 for 90 min), the DNA lost the ability to complex with DAPI. Exposure to monochloramine did not have a similar effect. Treatment of drinking water with sodium hypochlorite (about 0.5 mg of chlorine liter-1) caused a significant increase in the percentage of poorly fluorescent bacteria, from 5% in unchlorinated waters (40 samples), to 35 to 39% in chlorinated waters (40 samples). The presence of the poorly fluorescent bacteria could explain the underestimation of the real number of bacteria after DAPI staining. Microscopic counting of both poorly and highly fluorescent bacteria is essential under these conditions to obtain the total number of bacteria. A similar effect of chlorination on acridine orange-stained bacteria was observed in treated drinking waters. The presence of the poorly fluorescent bacteria after DAPI staining could be interpreted as a sign of dead cells. PMID:9097452

  9. Diversity and Detection of Nitrate Assimilation Genes in Marine Bacteria

    PubMed Central

    Allen, Andrew E.; Booth, Melissa G.; Frischer, Marc E.; Verity, Peter G.; Zehr, Jonathan P.; Zani, Sabino

    2001-01-01

    A PCR approach was used to construct a database of nasA genes (called narB genes in cyanobacteria) and to detect the genetic potential for heterotrophic bacterial nitrate utilization in marine environments. A nasA-specific PCR primer set that could be used to selectively amplify the nasA gene from heterotrophic bacteria was designed. Using seawater DNA extracts obtained from microbial communities in the South Atlantic Bight, the Barents Sea, and the North Pacific Gyre, we PCR amplified and sequenced nasA genes. Our results indicate that several groups of heterotrophic bacterial nasA genes are common and widely distributed in oceanic environments. PMID:11679368

  10. DNA maintenance in plastids and mitochondria of plants

    PubMed Central

    Oldenburg, Delene J.; Bendich, Arnold J.

    2015-01-01

    The DNA molecules in plastids and mitochondria of plants have been studied for over 40 years. Here, we review the data on the circular or linear form, replication, repair, and persistence of the organellar DNA (orgDNA) in plants. The bacterial origin of orgDNA appears to have profoundly influenced ideas about the properties of chromosomal DNA molecules in these organelles to the point of dismissing data inconsistent with ideas from the 1970s. When found at all, circular genome-sized molecules comprise a few percent of orgDNA. In cells active in orgDNA replication, most orgDNA is found as linear and branched-linear forms larger than the size of the genome, likely a consequence of a virus-like DNA replication mechanism. In contrast to the stable chromosomal DNA molecules in bacteria and the plant nucleus, the molecular integrity of orgDNA declines during leaf development at a rate that varies among plant species. This decline is attributed to degradation of damaged-but-not-repaired molecules, with a proposed repair cost-saving benefit most evident in grasses. All orgDNA maintenance activities are proposed to occur on the nucleoid tethered to organellar membranes by developmentally-regulated proteins. PMID:26579143

  11. Activity and Regulation of Archaeal DNA Alkyltransferase

    PubMed Central

    Perugino, Giuseppe; Vettone, Antonella; Illiano, Giuseppina; Valenti, Anna; Ferrara, Maria C.; Rossi, Mosè; Ciaramella, Maria

    2012-01-01

    Agents that form methylation adducts in DNA are highly mutagenic and carcinogenic, and organisms have evolved specialized cellular pathways devoted to their repair, including DNA alkyltransferases. These are proteins conserved in eucarya, bacteria and archaea, acting by a unique reaction mechanism, which leads to direct repair of DNA alkylation damage and irreversible protein alkylation. The alkylated form of DNA alkyltransferases is inactive, and in eukaryotes, it is rapidly directed to degradation. We report here in vitro and in vivo studies on the DNA alkyltransferase from the thermophilic archaeon Sulfolobus solfataricus (SsOGT). The development of a novel, simple, and sensitive fluorescence-based assay allowed a careful characterization of the SsOGT biochemical and DNA binding activities. In addition, transcriptional and post-translational regulation of SsOGT by DNA damage was studied. We show that although the gene transcription is induced by alkylating agent treatment, the protein is degraded in vivo by an alkylation-dependent mechanism. These experiments suggest a striking conservation, from archaea to humans, of this important pathway safeguarding genome stability. PMID:22167184

  12. Lactic acid bacteria production from whey.

    PubMed

    Mondragón-Parada, María Elena; Nájera-Martínez, Minerva; Juárez-Ramírez, Cleotilde; Galíndez-Mayer, Juvencio; Ruiz-Ordaz, Nora; Cristiani-Urbina, Eliseo

    2006-09-01

    The main purpose of this work was to isolate and characterize lactic acid bacteria (LAB) strains to be used for biomass production using a whey-based medium supplemented with an ammonium salt and with very low levels of yeast extract (0.25 g/L). Five strains of LAB were isolated from naturally soured milk after enrichment in whey-based medium. One bacterial isolate, designated MNM2, exhibited a remarkable capability to utilize whey lactose and give a high biomass yield on lactose. This strain was identified as Lactobacillus casei by its 16S rDNA sequence. A kinetic study of cell growth, lactose consumption, and titratable acidity production of this bacterial strain was performed in a bioreactor. The biomass yield on lactose, the percentage of lactose consumption, and the maximum increase in cell mass obtained in the bioreactor were 0.165 g of biomass/g of lactose, 100%, and 2.0 g/L, respectively, which were 1.44, 1.11, and 2.35 times higher than those found in flask cultures. The results suggest that it is possible to produce LAB biomass from a whey-based medium supplemented with minimal amounts of yeast extract.

  13. Pneumococcus: the sugar-coated bacteria.

    PubMed

    López, Rubens

    2006-09-01

    The study of Streptococcus pneumoniae (the pneumococcus) had been a central issue in medicine for many decades until the use of antibiotics became generalized. Many fundamental contributions to the history of microbiology should credit this bacterium: the capsular precipitin reaction, the major role this reaction plays in the development of immunology through the identification of polysaccharides as antigens, and, mainly, the demonstration, by genetic transformation, that genes are composed of DNA-the finding from the study of bacteria that has had the greatest impact on biology. Currently, pneumococcus is the most common etiologic agent in acute otitis media, sinusitis, and pneumonia requiring the hospitalization of adults. Moreover, meningitis is the leading cause of death among children in developing countries. Here I discuss the contributions that led to the explosion of knowledge about pneumococcus and also report some of the contributions of our group to the understanding of the molecular basis of three important virulence factors: lytic enzymes, pneumococcal phages, and the genes coding for capsular polysaccharides.

  14. Hexameric ring structure of the N-terminal domain of Mycobacterium tuberculosis DnaB helicase

    SciTech Connect

    Biswas, Tapan; Tsodikov, Oleg V.

    2009-01-15

    Hexameric DnaB helicase unwinds the DNA double helix during replication of genetic material in bacteria. DnaB is an essential bacterial protein; therefore, it is an important potential target for antibacterial drug discovery. We report a crystal structure of the N-terminal region of DnaB from the pathogen Mycobacterium tuberculosis (MtDnaBn), determined at 2.0 {angstrom} resolution. This structure provides atomic resolution details of formation of the hexameric ring of DnaB by two distinct interfaces. An extensive hydrophobic interface stabilizes a dimer of MtDnaBn by forming a four-helix bundle. The other, less extensive, interface is formed between the dimers, connecting three of them into a hexameric ring. On the basis of crystal packing interactions between MtDnaBn rings, we suggest a model of a helicase-primase complex that explains previously observed effects of DnaB mutations on DNA priming.

  15. Hexameric ring structure of the N-terminal domain of Mycobacterium tuberculosis DnaB helicase.

    PubMed

    Biswas, Tapan; Tsodikov, Oleg V

    2008-06-01

    Hexameric DnaB helicase unwinds the DNA double helix during replication of genetic material in bacteria. DnaB is an essential bacterial protein; therefore, it is an important potential target for antibacterial drug discovery. We report a crystal structure of the N-terminal region of DnaB from the pathogen Mycobacterium tuberculosis (MtDnaBn), determined at 2.0 A resolution. This structure provides atomic resolution details of formation of the hexameric ring of DnaB by two distinct interfaces. An extensive hydrophobic interface stabilizes a dimer of MtDnaBn by forming a four-helix bundle. The other, less extensive, interface is formed between the dimers, connecting three of them into a hexameric ring. On the basis of crystal packing interactions between MtDnaBn rings, we suggest a model of a helicase-primase complex that explains previously observed effects of DnaB mutations on DNA priming.

  16. Environment or kin: whence do bees obtain acidophilic bacteria?

    PubMed

    McFrederick, Quinn S; Wcislo, William T; Taylor, Douglas R; Ishak, Heather D; Dowd, Scot E; Mueller, Ulrich G

    2012-04-01

    As honey bee populations decline, interest in pathogenic and mutualistic relationships between bees and microorganisms has increased. Honey bees and bumble bees appear to have a simple intestinal bacterial fauna that includes acidophilic bacteria. Here, we explore the hypothesis that sweat bees can acquire acidophilic bacteria from the environment. To quantify bacterial communities associated with two species of North American and one species of Neotropical sweat bees, we conducted 16S rDNA amplicon 454 pyrosequencing of bacteria associated with the bees, their brood cells and their nests. Lactobacillus spp. were the most abundant bacteria in many, but not all, of the samples. To determine whether bee-associated lactobacilli can also be found in the environment, we reconstructed the phylogenetic relationships of the genus Lactobacillus. Previously described groups that associate with Bombus and Apis appeared relatively specific to these genera. Close relatives of several bacteria that have been isolated from flowers, however, were isolated from bees. Additionally, all three sweat bee species associated with lactobacilli related to flower-associated lactobacilli. These data suggest that there may be at least two different means by which bees acquire putative probiotics. Some lactobacilli appear specific to corbiculate apids, possibly because they are largely maternally inherited (vertically transmitted). Other lactobacilli, however, may be regularly acquired from environmental sources such as flowers. Sweat bee-associated lactobacilli were found to be abundant in the pollen and frass inside the nests of halictids, suggesting that they could play a role in suppressing the growth of moulds and other spoilage organisms.

  17. Characterization of unusual alkaliphilic gram-positive bacteria isolated from degraded brown alga thalluses.

    PubMed

    Ivanova, E P; Wright, J P; Lysenko, A M; Zhukova, N V; Alexeeva, Y V; Buljan, V; Kalinovskaya, N I; Nicolau, D V; Christen, R; Mikhailov, V V

    2006-01-01

    Two orange-pigmented Gram-positive, aerobic bacteria were isolated from enrichment culture during degradation of brown alga Fucus evanescens thalluses. In this work, atomic force microscopy (AFM) has been used to study the cell morphology. The non-contact mode imaging revealed unusual irregular coccoid shape of cells, possessing a single flagellum. Bacteria produced carotenoid pigments, were chemo-organotrophic, alkaliphilic and halo-tolerant growing well on nutrient media containing up to 15% NaCl. Growth temperature ranged from 5 to 45 degrees C. The DNA base compositions were 48 mol% G + C and the level of DNA similarity of two strains was conspecific (98%). A comparative phylogenetic analysis of 16S rRNA gene sequences revealed that the strain KMM 3738 tightly clustered with recently described Planococcus maritimus (99.9% 16S rRNA gene sequence similarity). DNA-DNA hybridisation experiments revealed that DNA from the KMM 3738 showed 12-15% and 16-35% of genetic relatedness with the DNA of type strains of the genera Planomicrobium and Planococcus, respectively, and 87% with DNA from Planococcus maritimus, indicating that new isolates belong to the later species. PMID:17100323

  18. DNA mimicry by proteins.

    PubMed

    Dryden, D T F; Tock, M R

    2006-04-01

    It has been discovered recently, via structural and biophysical analyses, that proteins can mimic DNA structures in order to inhibit proteins that would normally bind to DNA. Mimicry of the phosphate backbone of DNA, the hydrogen-bonding properties of the nucleotide bases and the bending and twisting of the DNA double helix are all present in the mimics discovered to date. These mimics target a range of proteins and enzymes such as DNA restriction enzymes, DNA repair enzymes, DNA gyrase and nucleosomal and nucleoid-associated proteins. The unusual properties of these protein DNA mimics may provide a foundation for the design of targeted inhibitors of DNA-binding proteins. PMID:16545103

  19. DNA Damage Responses in Prokaryotes: Regulating Gene Expression, Modulating Growth Patterns, and Manipulating Replication Forks

    PubMed Central

    Kreuzer, Kenneth N.

    2013-01-01

    Recent advances in the area of bacterial DNA damage responses are reviewed here. The SOS pathway is still the major paradigm of bacterial DNA damage response, and recent studies have clarified the mechanisms of SOS induction and key physiological roles of SOS including a very major role in genetic exchange and variation. When considering diverse bacteria, it is clear that SOS is not a uniform pathway with one purpose, but rather a platform that has evolved for differing functions in different bacteria. Relating in part to the SOS response, the field has uncovered multiple apparent cell-cycle checkpoints that assist cell survival after DNA damage and remarkable pathways that induce programmed cell death in bacteria. Bacterial DNA damage responses are also much broader than SOS, and several important examples of LexA-independent regulation will be reviewed. Finally, some recent advances that relate to the replication and repair of damaged DNA will be summarized. PMID:24097899

  20. Mutagenic DNA repair in enterobacteria.

    PubMed Central

    Sedgwick, S G; Ho, C; Woodgate, R

    1991-01-01

    Sixteen species of enterobacteria have been screened for mutagenic DNA repair activity. In Escherichia coli, mutagenic DNA repair is encoded by the umuDC operon. Synthesis of UmuD and UmuC proteins is induced as part of the SOS response to DNA damage, and after induction, the UmuD protein undergoes an autocatalytic cleavage to produce the carboxy-terminal UmuD' fragment needed for induced mutagenesis. The presence of a similar system in other species was examined by using a combined approach of inducible-mutagenesis assays, cross-reactivity to E. coli UmuD and UmuD' antibodies to test for induction and cleavage of UmuD-like proteins, and hybridization with E. coli and Salmonella typhimurium umu DNA probes to map umu-like genes. The results indicate a more widespread distribution of mutagenic DNA repair in other species than was previously thought. They also show that umu loci can be more complex in other species than in E. coli. Differences in UV-induced mutability of more than 200-fold were seen between different species of enteric bacteria and even between multiple natural isolates of E. coli, and yet some of the species which display a poorly mutable phenotype still have umu-like genes and proteins. It is suggested that umDC genes can be curtailed in their mutagenic activities but that they may still participate in some other, unknown process which provides the continued stimulus for their retention. Images PMID:1885540

  1. Killer Pigments in Bacteria: An Ecological Nightmare.

    ERIC Educational Resources Information Center

    Benathen, Isaiah A.; Saccardi, Marion

    2000-01-01

    Describes an alternative to teaching ecology by using bacteria to test competitor survival. Students observe a time-dependent selective killing of other unrelated bacteria by Pseudomonas aeruginosa. (SAH)

  2. Gene therapy for cancer: bacteria-mediated anti-angiogenesis therapy.

    PubMed

    Gardlik, R; Behuliak, M; Palffy, R; Celec, P; Li, C J

    2011-05-01

    Several bacterial species have inherent ability to colonize solid tumors in vivo. However, their natural anti-tumor activity can be enhanced by genetic engineering that enables these bacteria express or transfer therapeutic molecules into target cells. In this review, we summarize latest research on cancer therapy using genetically modified bacteria with particular emphasis on blocking tumor angiogenesis. Despite recent progress, only a few recent studies on bacterial tumor therapy have focused on anti-angiogenesis. Bacteria-mediated anti-angiogenesis therapy for cancer, however, is an attractive approach given that solid tumors are often characterized by increased vascularization. Here, we discuss four different approaches for using modified bacteria as anti-cancer therapeutics--bactofection, DNA vaccination, alternative gene therapy and transkingdom RNA interference--with a specific focus on angiogenesis suppression. Critical areas and future directions for this field are also outlined.

  3. Effect of different nitroheterocyclic compounds on aerobic, microaerophilic, and anaerobic bacteria.

    PubMed Central

    Hof, H; Ströder, J; Buisson, J P; Royer, R

    1986-01-01

    The antibacterial activities of different nitroheterocyclic compounds were assessed by an agar dilution method against aerobic, microaerophilic, and anaerobic bacteria. Nitronaphthofurans inhibited the multiplication of aerobic bacteria at low concentrations (MIC for 50% of strains tested [MIC50], 1 mg/liter). Under anaerobic growth conditions the MICs were found to be even lower. The rough, DNA repair-deficient mutants of Salmonella typhimurium were more susceptible, whereas nitroreductase-deficient strains were resistant. Microaerophilic campylobacter isolates could be divided into two groups, one of which was as susceptible as aerobic bacteria (MIC50, 1 mg/liter) and the other of which was more highly susceptible (MIC50, 0.015 mg/liter). All anaerobic bacteria tested were susceptible to nitronaphthofurans (MIC50, 0.125 mg/liter). Nitrothiazole exerted antibacterial activities similar to those of the nitronaphthofurans. Metronidazole, a nitroimidazole derivative, and nitrofurans were definitely less active. Nitrobenzofurans showed relatively high MICs. PMID:3800344

  4. Further evidence for the absence of bacteria in horsehair worms (Nematomorpha: Gordiidae).

    PubMed

    Hudson, Andrew J; Floate, Kevin D

    2009-12-01

    We used molecular techniques to characterize bacteria associated with the nematomorph Gordius robustus (Leidy). This worm is a parasite of the fall field cricket, Gryllus pennsylvanicus (Burmeister), which is infected with the symbiotic bacteria, Wolbachia. Because of this close association, our a priori expectation was that G. robustus may be similarly infected. However, results of denaturing gradient gel electrophoresis and sequencing of amplified 16S rDNA failed to detect any bacteria (symbiotic or non-symbiotic) in G. robustus. These unexpected findings suggest that G. robustus has no internal bacterial community and indicate that close association with a Wolbachia-infected host is insufficient for the transmission of bacteria from insect to nematomorph.

  5. Heterologous surface display on lactic acid bacteria: non-GMO alternative?

    PubMed Central

    Zadravec, Petra; Štrukelj, Borut; Berlec, Aleš

    2015-01-01

    Lactic acid bacteria (LAB) are food-grade hosts for surface display with potential applications in food and therapy. Alternative approaches to surface display on LAB would avoid the use of recombinant DNA technology and genetically-modified organism (GMO)-related regulatory requirements. Non-covalent surface display of proteins can be achieved by fusing them to various cell-wall binding domains, of which the Lysine motif domain (LysM) is particularly well studied. Fusion proteins have been isolated from recombinant bacteria or from their growth medium and displayed on unmodified bacteria, enabling heterologous surface display. This was demonstrated on non-viable cells devoid of protein content, termed bacteria-like particles, and on various species of genus Lactobacillus. Of the latter, Lactobacillus salivarius ATCC 11741 was recently shown to be particularly amenable for LysM-mediated display. Possible regulatory implications of heterologous surface display are discussed, particularly those relevant for the European Union. PMID:25880164

  6. Bactericidal Activity and Mechanism of Photoirradiated Polyphenols against Gram-Positive and -Negative Bacteria.

    PubMed

    Nakamura, Keisuke; Ishiyama, Kirika; Sheng, Hong; Ikai, Hiroyo; Kanno, Taro; Niwano, Yoshimi

    2015-09-01

    The bactericidal effect of various types of photoirradiated polyphenols against Gram-positive and -negative bacteria was evaluated in relation to the mode of action. Gram-positive bacteria (Enterococcus faecalis, Staphylococcus aureus, and Streptococcus mutans) and Gram-negative bacteria (Aggregatibacter actinomycetemcomitans, Escherichia coli, and Pseudomonas aeruginosa) suspended in a 1 mg/mL polyphenol aqueous solution (caffeic acid, gallic acid, chlorogenic acid, epigallocatechin, epigallocatechin gallate, and proanthocyanidin) were exposed to LED light (wavelength, 400 nm; irradiance, 260 mW/cm(2)) for 5 or 10 min. Caffeic acid and chlorogenic acid exerted the highest bactericidal activity followed by gallic acid and proanthocyanidin against both Gram-positive and -negative bacteria. It was also demonstrated that the disinfection treatment induced oxidative damage of bacterial DNA, which suggests that polyphenols are incorporated into bacterial cells. The present study suggests that blue light irradiation of polyphenols could be a novel disinfection treatment.

  7. Heterologous surface display on lactic acid bacteria: non-GMO alternative?

    PubMed

    Zadravec, Petra; Štrukelj, Borut; Berlec, Aleš

    2015-01-01

    Lactic acid bacteria (LAB) are food-grade hosts for surface display with potential applications in food and therapy. Alternative approaches to surface display on LAB would avoid the use of recombinant DNA technology and genetically-modified organism (GMO)-related regulatory requirements. Non-covalent surface display of proteins can be achieved by fusing them to various cell-wall binding domains, of which the Lysine motif domain (LysM) is particularly well studied. Fusion proteins have been isolated from recombinant bacteria or from their growth medium and displayed on unmodified bacteria, enabling heterologous surface display. This was demonstrated on non-viable cells devoid of protein content, termed bacteria-like particles, and on various species of genus Lactobacillus. Of the latter, Lactobacillus salivarius ATCC 11741 was recently shown to be particularly amenable for LysM-mediated display. Possible regulatory implications of heterologous surface display are discussed, particularly those relevant for the European Union.

  8. Survival of bacteria in carcasses.

    PubMed

    Gill, C O; Penney, N

    1979-04-01

    Bacteria injected into the bloodstream of guinea pigs shortly before death decreased in number in carcass tissues for about 1 h after death. If initial bacterial numbers were sufficiently low, all bacteria were eliminated, and carcass tissues were sterile 24 h after death. Carcass tissue sterility was maintained with an initial density of Clostridium perfringens or Salmonella typhimurium of 20 cells per g or with an initial density of the other species examined of several hundred cells per gram. With larger numbers of strict and facultative anaerobes, growth commenced after 3 h in carcasses incubated at 30 degrees C. Spores of C. perfringens were killed over the same period as vegetative cells, but growth did not commence until 8 h after death. Bactericidal activity in carcass tissues must therefore be taken into account in evaluating the significance of reports of deep-tissue contamination of carcasses from meat animals.

  9. Genetics of Lactic Acid Bacteria

    NASA Astrophysics Data System (ADS)

    Zagorec, Monique; Anba-Mondoloni, Jamila; Coq, Anne-Marie Crutz-Le; Champomier-Vergès, Marie-Christine

    Many meat (or fish) products, obtained by the fermentation of meat originating from various animals by the flora that naturally contaminates it, are part of the human diet since millenaries. Historically, the use of bacteria as starters for the fermentation of meat, to produce dry sausages, was thus performed empirically through the endogenous micro-biota, then, by a volunteer addition of starters, often performed by back-slopping, without knowing precisely the microbial species involved. It is only since about 50 years that well defined bacterial cultures have been used as starters for the fermentation of dry sausages. Nowadays, the indigenous micro-biota of fermented meat products is well identified, and the literature is rich of reports on the identification of lactic acid bacteria (LAB) present in many traditional fermented products from various geographical origin, obtained without the addition of commercial starters (See Talon, Leroy, & Lebert, 2007, and references therein).

  10. Viability of bacteria in peatlands

    NASA Astrophysics Data System (ADS)

    Bogdanova, O. Yu.; Golovchenko, A. V.; Lysak, L. V.; Glukhova, T. V.; Zvyagintsev, D. G.

    2014-04-01

    The viability of bacteria in oligotrofic bogs and fens was determined by the luminescent microscopy method with the help of a two-component fluorescent dye (L7012 LIVE/DEAD). Living bacterial cells were found in the entire peat profiles. Their portion was maximal (up to 60%) in the upper layers and did not exceed 25% in the lower layers. The portion of dead bacterial cells varied from 3 to 19%, and dormant cells constituted 25 to 95% of the total number of bacterial cells. The numbers of dormant cells increased down the profiles irrespectively of the peat type. The portion of nanoforms did not exceed 5% of the total. The cells of the nanoforms, unlike the bacteria of typical sizes, were characterized by their high viability (93-98%).

  11. Swimming bacteria power microscopic gears

    SciTech Connect

    Sokolov, Andrey; Apodaca, Mario M.; Grzybowski, Bartosz A.; Aranson, Igor S.

    2010-01-19

    Whereas the laws of thermodynamics prohibit extraction of useful work from the Brownian motion of particles in equilibrium, these motions can be “rectified” under nonequilibrium conditions, for example, in the presence of asymmetric geometrical obstacles. Here, we describe a class of systems in which aerobic bacteria Bacillus subtilis moving randomly in a fluid film power submillimeter gears and primitive systems of gears decorated with asymmetric teeth. The directional rotation is observed only in the regime of collective bacterial swimming and the gears’ angular velocities depend on and can be controlled by the amount of oxygen available to the bacteria. The ability to harness and control the power of collective motions appears an important requirement for further development of mechanical systems driven by microorganisms.

  12. Swimming bacteria power microscopic gears.

    SciTech Connect

    Sokolov, A.; Apodaca, M. M.; Grzybowski, B. A.; Aranson, I. S.; Materials Science Division; Princeton Univ.; Northwestern Univ.

    2010-01-19

    Whereas the laws of thermodynamics prohibit extraction of useful work from the Brownian motion of particles in equilibrium, these motions can be 'rectified' under nonequilibrium conditions, for example, in the presence of asymmetric geometrical obstacles. Here, we describe a class of systems in which aerobic bacteria Bacillus subtilis moving randomly in a fluid film power submillimeter gears and primitive systems of gears decorated with asymmetric teeth. The directional rotation is observed only in the regime of collective bacterial swimming and the gears angular velocities depend on and can be controlled by the amount of oxygen available to the bacteria. The ability to harness and control the power of collective motions appears an important requirement for further development of mechanical systems driven by microorganisms.

  13. Re-engineering bacteria for ethanol production

    SciTech Connect

    Yomano, Lorraine P; York, Sean W; Zhou, Shengde; Shanmugam, Keelnatham; Ingram, Lonnie O

    2014-05-06

    The invention provides recombinant bacteria, which comprise a full complement of heterologous ethanol production genes. Expression of the full complement of heterologous ethanol production genes causes the recombinant bacteria to produce ethanol as the primary fermentation product when grown in mineral salts medium, without the addition of complex nutrients. Methods for producing the recombinant bacteria and methods for producing ethanol using the recombinant bacteria are also disclosed.

  14. Sand Beach Bacteria: Enumeration and Characterization

    PubMed Central

    Khiyama, H. M.; Makemson, J. C.

    1973-01-01

    Bacteria in the water-saturated sand of a relatively unpolluted sand beach were enumerated by direct microscope and viable counting. The number of interstitial bacteria was estimated to be a significant fraction of the total number of bacteria present. Three hundred sixty-two strains were isolated and submitted to cultural and biochemical tests. Fermentational abilities and the production of indole suggested that a significant number of these bacteria were symbiotically associated with resident metazoans. PMID:4356458

  15. Bacteria turn a tiny gear

    SciTech Connect

    2009-01-01

    Thousands of tiny Bacillus subtillis bacteria turn a single gear, just 380 microns across. (A human hair is about 100 microns across.) The method could be used to create micro-machines. Argonne National Laboratory scientist Igor Aronson pioneered this technique. Read more at the New York Times: http://ow.ly/ODfI or at Argonne: http://ow.ly/ODfa Video courtesy Igor Aronson.

  16. Endocytosis of Viruses and Bacteria

    PubMed Central

    Cossart, Pascale; Helenius, Ari

    2014-01-01

    Of the many pathogens that infect humans and animals, a large number use cells of the host organism as protected sites for replication. To reach the relevant intracellular compartments, they take advantage of the endocytosis machinery and exploit the network of endocytic organelles for penetration into the cytosol or as sites of replication. In this review, we discuss the endocytic entry processes used by viruses and bacteria and compare the strategies used by these dissimilar classes of pathogens. PMID:25085912

  17. Enteric bacteria: friend or foe?

    PubMed

    Batt, R M; Rutgers, H C; Sancak, A A

    1996-06-01

    The normal gastrointestinal tract contains an enormous number of aerobic and anaerobic bacteria which normally enjoy a symbiotic relationship with the host but can have adverse effects with local and systemic consequences. The small intestine constitutes a zone of transition between the sparsely populated stomach and the luxuriant bacterial flora of the colon. Regulation of the intestinal flora depends on complex interactions between many factors including secretion of gastric acid, intestinal motility, biliary and pancreatic secretions, local immunity, the surface glycocalyx and mucus layer, and diet. Microbial interactions are also important, and can involve alterations in redox potential, substrate depletion and production of substances such as bacteriocins that inhibit bacterial growth. The beneficial effect of the normal enteric flora include the competitive exclusion of potentially pathogenic organisms, and the production of nutrients such as short-chain fatty acids (which represent an important energy source for the colonic mucosa) and vitamins. Detrimental effects of the enteric flora include competition for calories and essential nutrients, particularly by bacteria located in the small intestine, and a capacity to damage the mucosa, in some circumstances causing or contributing to inflammatory bowel disease. These problems can be accentuated by interference with the physiological regulation of intraluminal bacteria allowing overgrowth by a normal resident, or colonisation by transient pathogens. The pathophysiological consequences may involve direct damage to the intestinal mucosa, and bacterial metabolism of intraluminal constituents, for example forming deconjugated bile acids and hydroxylated fatty acids which stimulate fluid secretion. Additional problems arise if there is interference with the mucosal barrier since this can result in increased passage of bacteria and bacterial products stimulating mucosal inflammation, while bacterial translocation

  18. Compartmentalization of bacteria in microcapsules.

    PubMed

    van Wijk, Judith; Heunis, Tiaan; Harmzen, Elrika; Dicks, Leon M T; Meuldijk, Jan; Klumperman, Bert

    2014-12-18

    Lactobacillus plantarum strain 423 was encapsulated in hollow poly(organosiloxane) microcapsules by templating water-in-oil Pickering emulsion droplets via the interfacial reaction of alkylchlorosilanes. The bacteria were suspended in growth medium or buffer to protect the cells against pH changes during the interfacial reactions with alkylchlorosilanes. The results of this work open up novel avenues for the encapsulation of microbial cells.

  19. DNA ligase I, the replicative DNA ligase

    PubMed Central

    Howes, Timothy R.L.; Tomkinson, Alan E.

    2013-01-01

    Multiple DNA ligation events are required to join the Okazaki fragments generated during lagging strand DNA synthesis. In eukaryotes, this is primarily carried out by members of the DNA ligase I family. The C-terminal catalytic region of these enzymes is composed of three domains: a DNA binding domain, an adenylation domain and an OB-fold domain. In the absence of DNA, these domains adopt an extended structure but transition into a compact ring structure when they engage a DNA nick, with each of the domains contacting the DNA. The non-catalytic N-terminal region of eukaryotic DNA ligase I is responsible for the specific participation of these enzymes in DNA replication. This proline-rich unstructured region contains the nuclear localization signal and a PCNA interaction motif that is critical for localization to replication foci and efficient joining of Okazaki fragments. DNA ligase I initially engages the PCNA trimer via this interaction motif which is located at the extreme N-terminus of this flexible region. It is likely that this facilitates an additional interaction between the DNA binding domain and the PCNA ring. The similar size and shape of the rings formed by the PCNA trimer and the DNA ligase I catalytic region when it engages a DNA nick suggest that these proteins interact to form a double-ring structure during the joining of Okazaki fragments. DNA ligase I also interacts with replication factor C, the factor that loads the PCNA trimeric ring onto DNA. This interaction, which is regulated by phosphorylation of the non-catalytic N-terminus of DNA ligase I, also appears to be critical for DNA replication. PMID:22918593

  20. Chemical signature of magnetotactic bacteria

    PubMed Central

    Amor, Matthieu; Busigny, Vincent; Durand-Dubief, Mickaël; Tharaud, Mickaël; Ona-Nguema, Georges; Gélabert, Alexandre; Alphandéry, Edouard; Menguy, Nicolas; Benedetti, Marc F.; Chebbi, Imène; Guyot, François

    2015-01-01

    There are longstanding and ongoing controversies about the abiotic or biological origin of nanocrystals of magnetite. On Earth, magnetotactic bacteria perform biomineralization of intracellular magnetite nanoparticles under a controlled pathway. These bacteria are ubiquitous in modern natural environments. However, their identification in ancient geological material remains challenging. Together with physical and mineralogical properties, the chemical composition of magnetite was proposed as a promising tracer for bacterial magnetofossil identification, but this had never been explored quantitatively and systematically for many trace elements. Here, we determine the incorporation of 34 trace elements in magnetite in both cases of abiotic aqueous precipitation and of production by the magnetotactic bacterium Magnetospirillum magneticum strain AMB-1. We show that, in biomagnetite, most elements are at least 100 times less concentrated than in abiotic magnetite and we provide a quantitative pattern of this depletion. Furthermore, we propose a previously unidentified method based on strontium and calcium incorporation to identify magnetite produced by magnetotactic bacteria in the geological record. PMID:25624469