Publications by authors named "Lee J Kerkhof"

39 Publications

Lactobacillus rhamnosus GG modifies the metabolome of pathobionts in gnotobiotic mice.

BMC Microbiol 2021 Jun 3;21(1):165. Epub 2021 Jun 3.

Department of Pharmacology, Physiology and Neurosciences, Medical Science Building, New Jersey Medical School, Rutgers University, Newark, NJ, 07103, USA.

Background: Lactobacillus rhamnosus GG (LGG) is the most widely used probiotic, but the mechanisms underlying its beneficial effects remain unresolved. Previous studies typically inoculated LGG in hosts with established gut microbiota, limiting the understanding of specific impacts of LGG on host due to numerous interactions among LGG, commensal microbes, and the host. There has been a scarcity of studies that used gnotobiotic animals to elucidate LGG-host interaction, in particular for gaining specific insights about how it modifies the metabolome. To evaluate whether LGG affects the metabolite output of pathobionts, we inoculated with LGG gnotobiotic mice containing Propionibacterium acnes, Turicibacter sanguinis, and Staphylococcus aureus (PTS).

Results: 16S rRNA sequencing of fecal samples by Ion Torrent and MinION platforms showed colonization of germ-free mice by PTS or by PTS plus LGG (LTS). Although the body weights and feeding rates of mice remained similar between PTS and LTS groups, co-associating LGG with PTS led to a pronounced reduction in abundance of P. acnes in the gut. Addition of LGG or its secretome inhibited P. acnes growth in culture. After optimizing procedures for fecal metabolite extraction and metabolomic liquid chromatography-mass spectrometry analysis, unsupervised and supervised multivariate analyses revealed a distinct separation among fecal metabolites of PTS, LTS, and germ-free groups. Variables-important-in-projection scores showed that LGG colonization robustly diminished guanine, ornitihine, and sorbitol while significantly elevating acetylated amino acids, ribitol, indolelactic acid, and histamine. In addition, carnitine, betaine, and glutamate increased while thymidine, quinic acid and biotin were reduced in both PTS and LTS groups. Furthermore, LGG association reduced intestinal mucosal expression levels of inflammatory cytokines, such as IL-1α, IL-1β and TNF-α.

Conclusions: LGG co-association had a negative impact on colonization of P. acnes, and markedly altered the metabolic output and inflammatory response elicited by pathobionts.
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http://dx.doi.org/10.1186/s12866-021-02178-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8176599PMC
June 2021

Is Oxford Nanopore sequencing ready for analyzing complex microbiomes?

Authors:
Lee J Kerkhof

FEMS Microbiol Ecol 2021 03;97(3)

Department of Marine and Coastal Sciences, Rutgers, the State University of New Jersey, New Brunswick, NJ 08901, USA.

This minireview will discuss the improvements in Oxford Nanopore (Oxford; sequencing technology that make the MinION a viable platform for microbial ecology studies. Specific issues being addressed are the increase in sequence accuracy from 65 to 96.5% during the last 5 years, the ability to obtain a quantifiable/predictive signal from the MinION with respect to target molecule abundance, simple-to-use GUI-based pathways for data analysis and the modest additional equipment needs for sequencing in the field. Coupling these recent improvements with the low capital costs for equipment and the reasonable per sample cost makes MinION sequencing an attractive option for virtually any laboratory.
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http://dx.doi.org/10.1093/femsec/fiab001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8068755PMC
March 2021

Species-level evaluation of the human respiratory microbiome.

Gigascience 2020 04;9(4)

Department of Marine and Coastal Sciences, Rutgers-the State University of New Jersey, 71 Dudley Road, New Brunswick, NJ USA 08901.

Background: Changes to human respiratory tract microbiome may contribute significantly to the progression of respiratory diseases. However, there are few studies examining the relative abundance of microbial communities at the species level along the human respiratory tract.

Findings: Bronchoalveolar lavage, throat swab, mouth rinse, and nasal swab samples were collected from 5 participants. Bacterial ribosomal operons were sequenced using the Oxford Nanopore MinION to determine the relative abundance of bacterial species in 4 compartments along the respiratory tract. More than 1.8 million raw operon reads were obtained from the participants with ∼600,000 rRNA reads passing quality assurance/quality control (70-95% identify; >1,200 bp alignment) by Discontiguous MegaBLAST against the EZ BioCloud 16S rRNA gene database. Nearly 3,600 bacterial species were detected overall (>750 bacterial species within the 5 dominant phyla: Firmicutes, Proteobacteria, Actinobacteria, Bacteroidetes, and Fusobacteria. The relative abundance of bacterial species along the respiratory tract indicated that most microbes (95%) were being passively transported from outside into the lung. However, a small percentage (<5%) of bacterial species were at higher abundance within the lavage samples. The most abundant lung-enriched bacterial species were Veillonella dispar and Veillonella atypica while the most abundant mouth-associated bacterial species were Streptococcus infantis and Streptococcus mitis.

Conclusions: Most bacteria detected in lower respiratory samples do not seem to colonize the lung. However, >100 bacterial species were found to be enriched in bronchoalveolar lavage samples (compared to mouth/nose) and may play a substantial role in lung health.
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http://dx.doi.org/10.1093/gigascience/giaa038DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7162353PMC
April 2020

Arctic tundra soil bacterial communities active at subzero temperatures detected by stable isotope probing.

FEMS Microbiol Ecol 2020 02;96(2)

School of Environmental and Biological Sciences, Rutgers, The State University of New Jersey, New Brunswick NJ 08901, USA.

Arctic soils store vast amounts of carbon and are subject to intense climate change. While the effects of thaw on the composition and activities of Arctic tundra microorganisms has been examined extensively, little is known about the consequences of temperature fluctuations within the subzero range in seasonally frozen or permafrost soils. This study identified tundra soil bacteria active at subzero temperatures using stable isotope probing (SIP). Soils from Kilpisjärvi, Finland, were amended with 13C-cellobiose and incubated at 0, -4 and -16°C for up to 40 weeks. 16S rRNA gene sequence analysis of 13C-labelled DNA revealed distinct subzero-active bacterial taxa. The SIP experiments demonstrated that diverse bacteria, including members of Candidatus Saccharibacteria, Melioribacteraceae, Verrucomicrobiaceae, Burkholderiaceae, Acetobacteraceae, Armatimonadaceae and Planctomycetaceae, were capable of synthesising 13C-DNA at subzero temperatures. Differences in subzero temperature optima were observed, for example, with members of Oxalobacteraceae and Rhizobiaceae found to be more active at 0°C than at -4°C or -16°C, whereas Melioribacteriaceae were active at all subzero temperatures tested. Phylogeny of 13C-labelled 16S rRNA genes from the Melioribacteriaceae, Verrucomicrobiaceae and Candidatus Saccharibacteria suggested that these taxa formed subzero-active clusters closely related to members from other cryo-environments. This study demonstrates that subzero temperatures impact active bacterial community composition and activity, which may influence biogeochemical cycles.
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http://dx.doi.org/10.1093/femsec/fiz192DOI Listing
February 2020

Identification of a Chlorodibenzo--dioxin Dechlorinating by Stable Isotope Probing.

Environ Sci Technol 2019 12 5;53(24):14409-14419. Epub 2019 Dec 5.

Department of Biochemistry and Microbiology, Rutgers , The State University of New Jersey , New Brunswick , New Jersey 08901 , United States.

Polychlorinated dibenzo--dioxins (PCDDs) are released into the environment from a variety of both anthropogenic and natural sources. While highly chlorinated dibenzo--dioxins are persistent under oxic conditions, in anoxic environments, these organohalogens can be reductively dechlorinated to less chlorinated compounds that are then more amenable to subsequent aerobic degradation. Identifying the microorganisms responsible for dechlorination is an important step in developing bioremediation approaches. In this study, we demonstrated the use of a DNA-stable isotope probing (SIP) approach to identify the bacteria active in dechlorination of PCDDs in river sediments, with 1,2,3,4-tetrachlorodibenzo--dioxin (1,2,3,4-TeCDD) as a model. In addition, pyrosequencing of reverse transcribed 16S rRNA of TeCDD dechlorinating enrichment cultures was used to reveal active members of the bacterial community. A set of operational taxonomic units (OTUs) responded positively to the addition of 1,2,3,4-TeCDD in SIP microcosms assimilating C-acetate as the carbon source. Analysis of bacterial community profiles of the C labeled heavy DNA fraction revealed that an OTU corresponding to accounted for a significantly greater abundance in cultures amended with 1,2,3,4-TeCDD than in cultures without 1,2,3,4-TeCDD. This implies the involvement of this strain in the reductive dechlorination of 1,2,3,4-TeCDD and suggests the applicability of SIP for a robust assessment of the bioremediation potential of organohalogen contaminated sites.
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http://dx.doi.org/10.1021/acs.est.9b05395DOI Listing
December 2019

Offline Next Generation Metagenomics Sequence Analysis Using MinION Detection Software (MINDS).

Genes (Basel) 2019 07 30;10(8). Epub 2019 Jul 30.

US Army, CCDC-Chemical Biological Center, Aberdeen Proving Ground, MD 21010, USA.

Field laboratories interested in using the MinION often need the internet to perform sample analysis. Thus, the lack of internet connectivity in resource-limited or remote locations renders downstream analysis problematic, resulting in a lack of sample identification in the field. Due to this dependency, field samples are generally transported back to the lab for analysis where internet availability for downstream analysis is available. These logistics problems and the time lost in sample characterization and identification, pose a significant problem for field scientists. To address this limitation, we have developed a stand-alone data analysis packet using open source tools developed by the Nanopore community that does not depend on internet availability. Like Oxford Nanopore Technologies' (ONT) cloud-based What's In My Pot (WIMP) software, we developed the offline MinION Detection Software (MINDS) based on the Centrifuge classification engine for rapid species identification. Several online bioinformatics applications have been developed surrounding ONT's framework for analysis of long reads. We have developed and evaluated an offline real time classification application pipeline using open source tools developed by the Nanopore community that does not depend on internet availability. Our application has been tested on ATCC's 20 strain even mix whole cell (ATCC MSA-2002) sample. Using the Rapid Sequencing Kit (SQK-RAD004), we were able to identify all 20 organisms at species level. The analysis was performed in 15 min using a Dell Precision 7720 laptop. Our offline downstream bioinformatics application provides a cost-effective option as well as quick turn-around time when analyzing samples in the field, thus enabling researchers to fully utilize ONT's MinION portability, ease-of-use, and identification capability in remote locations.
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http://dx.doi.org/10.3390/genes10080578DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6723491PMC
July 2019

Sulfate-Reducing Naphthalene Degraders Are Picky Eaters.

Microorganisms 2018 06 25;6(3). Epub 2018 Jun 25.

Department of Environmental Sciences, Rutgers University, New Brunswick, NJ 08901, USA.

Polycyclic aromatic hydrocarbons (PAHs) are common organic contaminants found in anoxic environments. The capacity for PAH biodegradation in unimpacted environments, however, has been understudied. Here we investigate the enrichment, selection, and sustainability of a microbial community from a pristine environment on naphthalene as the only amended carbon source. Pristine coastal sediments were obtained from the Jacques Cousteau National Estuarine Research Reserve in Tuckerton, New Jersey, an ecological reserve which has no direct input or source of hydrocarbons. After an initial exposure to naphthalene, primary anaerobic transfer cultures completely degraded 500 µM naphthalene within 139 days. Subsequent transfer cultures mineralized naphthalene within 21 days with stoichiometric sulfate loss. Enriched cultures efficiently utilized only naphthalene and 2-methylnaphthalene from the hydrocarbon mixtures in crude oil. To determine the microorganisms responsible for naphthalene degradation, stable isotope probing was utilized on cultures amended with fully labeled C-naphthalene as substrate. Three organisms were found to unambiguously synthesize C-DNA from C-naphthalene within 7 days. Phylogenetic analysis revealed that 16S rRNA genes from two of these organisms are closely related to the known naphthalene degrading isolates NaphS2 and NaphS3 from PAH-contaminated sites. A third 16S rRNA gene was only distantly related to its closest relative and may represent a novel naphthalene degrading microbe from this environment.
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http://dx.doi.org/10.3390/microorganisms6030059DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6163709PMC
June 2018

Discerning autotrophy, mixotrophy and heterotrophy in marine TACK archaea from the North Atlantic.

FEMS Microbiol Ecol 2018 03;94(3)

Department of Marine and Coastal Sciences, Rutgers University, New Brunswick, NJ 08901-8521, USA.

DNA stable isotope probing (SIP) was used to track the uptake of organic and inorganic carbon sources for TACK archaea (Thaumarchaeota/Aigarchaeota/Crenarchaeota/Korarchaeota) on a cruise of opportunity in the North Atlantic. Due to water limitations, duplicate samples from the deep photic (60-115 m), the mesopelagic zones (local oxygen minimum; 215-835 m) and the bathypelagic zone (2085-2835 m) were amended with various combinations of 12C- or 13C-acetate/urea/bicarbonate to assess cellular carbon acquisition. The SIP results indicated the majority of TACK archaeal operational taxonomic units (OTUs) incorporated 13C from acetate and/or urea into newly synthesized DNA within 48 h. A small fraction (16%) of the OTUs, often representing the most dominant members of the archaeal community, were able to incorporate bicarbonate in addition to organic substrates. Only two TACK archaeal OTUs were found to incorporate bicarbonate but not urea or acetate. These results further demonstrate the utility of SIP to elucidate the metabolic capability of mesothermal archaea in distinct oceanic settings and suggest that TACK archaea play a role in organic carbon recycling in the mid-depth to deep ocean.
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http://dx.doi.org/10.1093/femsec/fiy014DOI Listing
March 2018

Profiling bacterial communities by MinION sequencing of ribosomal operons.

Microbiome 2017 09 15;5(1):116. Epub 2017 Sep 15.

Department of Marine and Coastal Sciences, Rutgers University, 71 Dudley Rd, New Brunswick, NJ, 08901-8521, USA.

Background: An approach utilizing the long-read capability of the Oxford Nanopore MinION to rapidly sequence bacterial ribosomal operons of complex natural communities was developed. Microbial fingerprinting employs domain-specific forward primers (16S rRNA subunit), reverse primers (23S rRNA subunit), and a high-fidelity Taq polymerase with proofreading capabilities. Amplicons contained both ribosomal subunits for broad-based phylogenetic assignment (~ 3900 bp of sequence), plus the intergenic spacer (ITS) region (~ 300 bp) for potential strain-specific identification.

Results: To test the approach, bacterial rRNA operons (~ 4200 bp) were amplified from six DNA samples employing a mixture of farm soil and bioreactor DNA in known concentrations. Each DNA sample mixture was barcoded, sequenced in quadruplicate (n = 24), on two separate 6-h runs using the MinION system (R7.3 flow cell; MAP005 and 006 chemistry). From nearly 90,000 MinION reads, roughly 33,000 forward and reverse sequences were obtained. This yielded over 10,000 2D sequences which were analyzed using a simplified data analysis pipeline based on NCBI Blast and assembly with Geneious software. The method could detect over 1000 operational taxonomic units in the sample sets in a quantitative manner. Global sequence coverage for the various rRNA operons ranged from 1 to 1951x. An iterative assembly scheme was developed to reconstruct those rRNA operons with > 35x coverage from a set of 30 operational taxonomic units (OTUs) among the Proteobacteria, Actinobacteria, Acidobacteria, Firmicutes, and Gemmatimonadetes. Phylogenetic analysis of the 16S rRNA and 23S rRNA genes from each operon demonstrated similar tree topologies with species/strain-level resolution.

Conclusions: This sequencing method represents a cost-effective way to profile microbial communities. Because the MinION is small, portable, and runs on a laptop, the possibility of microbiota characterization in the field or on robotic platforms becomes realistic.
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http://dx.doi.org/10.1186/s40168-017-0336-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5599880PMC
September 2017

Forensic Analysis of Polychlorinated Dibenzo-p-Dioxin and Furan Fingerprints to Elucidate Dechlorination Pathways.

Environ Sci Technol 2017 Sep 30;51(18):10485-10493. Epub 2017 Aug 30.

Department of Environmental Sciences, Rutgers University , 14 College Farm Road, New Brunswick, New Jersey 08901, United States.

Polychlorinated dibenzo-p-dioxins and -furans (PCDD/Fs) are persistent organic pollutants whose main removal process in the environment is due to biodegradation, and particularly anaerobic reductive dechlorination. Since PCDD/F congeners that are substituted in the lateral 2, 3, 7, and 8 positions are the most toxic, removal of these chlorines is advantageous, but previous studies have only demonstrated their removal under laboratory conditions. We evaluated a concentration data set of PCDD/F congeners with four or more chlorines along with all 209 polychlorinated biphenyl (PCB) congeners in surface water, treated and untreated wastewater, landfill leachate, and biosolids (NY CARP data set) to determine whether peri and peri/lateral dechlorination of PCDD/Fs occurs in these environments. Positive Matrix Factorization (PMF) applied to the data set revealed a factor indicative of the microbial dechlorination of PCBs, and this factor also contained a variety of non-2,3,7,8 substituted PCDD/F congeners. These results suggest that dechlorination of PCDD/Fs at the lateral positions is facile if not preferred in these environments. The relative lack of tetra- and penta-chlorinated PCDD/Fs suggested that dechlorination proceeds to PCDD/F congeners with less than four chlorines. The PMF results were confirmed by examining three samples that contained >90% PCB dechlorination products from the Fresh Kills Landfill and the Hudson River. Even without factor analysis, these samples demonstrated almost identical PCDD/F congener patterns. This study suggests that PCDD/Fs are reductively dechlorinated to nontoxic non-2,3,7,8 PCDD/F congeners in sewers and landfills as well as in the sediment of the Upper Hudson River.
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http://dx.doi.org/10.1021/acs.est.7b02705DOI Listing
September 2017

Novel reductive dehalogenases from the marine sponge associated bacterium Desulfoluna spongiiphila.

Environ Microbiol Rep 2017 10 21;9(5):537-549. Epub 2017 Jul 21.

Departments of Biochemistry and Microbiology, School of Environmental and Biological Sciences, Rutgers, The State University of New Jersey, New Brunswick, NJ 08901, USA.

Desulfoluna spongiiphila strain AA1 is an organohalide respiring bacterium, isolated from the marine sponge Aplysina aerophoba, that can use brominated and iodinated phenols, in addition to sulfate and thiosulfate as terminal electron acceptors. The genome of Desulfoluna spongiiphila strain AA1 is approximately 6.5 Mb. Three putative reductive dehalogenase (rdhA) genes involved in respiratory metabolism of organohalides were identified within the sequence. Conserved motifs found in respiratory reductive dehalogenases (a twin arginine translocation signal sequence and two iron-sulfur clusters) were present in all three putative AA1 rdhA genes. Transcription of one of the three rdhA genes was significantly upregulated during respiration of 2,6-dibromophenol and sponge extracts. Strain AA1 appears to have the ability to synthesize cobalamin, the key cofactor of most characterized reductive dehalogenase enzymes. The genome contains genes involved in cobalamin synthesis and uptake and can grow without cobalamin supplementation. Identification of this target gene associated with debromination lays the foundation for understanding how dehalogenating bacteria control the fate of organohalide compounds in sponges and their role in a symbiotic organobromine cycle. In the sponge environment, D. spongiiphila strain AA1 may thus take advantage of both brominated compounds and sulfate as electron acceptors for respiration.
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http://dx.doi.org/10.1111/1758-2229.12556DOI Listing
October 2017

Comparison of a modified phenol/chloroform and commercial-kit methods for extracting DNA from horse fecal material.

J Microbiol Methods 2016 10 25;129:14-19. Epub 2016 Jul 25.

Microbial Biology Graduate Program, Rutgers-The State University of New Jersey, New Brunswick, NJ 08901, USA; Department of Animal Science, Rutgers-The State University of New Jersey, New Brunswick, NJ 08901, USA. Electronic address:

There are many choices for methods of extracting bacterial DNA for Next Generation Sequencing (NGS) from fecal samples. Here, we compare our modifications of a phenol/chloroform extraction method plus an inhibitor removal solution (C3) (ph/Chl+C3) to the PowerFecal® DNA Isolation Kit (MoBio-K). DNA quality and quantity coupled to NGS results were used to assess differences in relative abundance, Shannon diversity index, unique species, and principle coordinate analysis (PCoA) between biological replicates. Six replicate samples, taken from a single ball of horse feces manually collected from the rectum, were subjected to each extraction method. The Ph/Chl+C3 method produced 100× higher DNA yields with less shearing than the MoBio-K method. To assess the methods, the two method samples were sent for sequencing of the bacterial V3-V4 region of 16S rRNA gene using the Illumina MiSeq platform. The relative abundance of Bacteroidetes was greater and there were more unique species assigned to this group in MoBio-K than in Ph/Chl+C3 (P<0.05). In contrast, Firmicutes had greater relative abundance and more unique species in Ph/Chl+C3 extracts than in MoBio-K (P<0.05). The other major bacterial phyla were equally abundant in samples using both extraction methods. Alpha diversity and Shannon Weaver indices showed greater evenness of bacterial distribution in Ph/Chl+C3 compared with MoBio-K (P<0.05), but there was no difference in the OTU richness. Principle coordinate analysis (PCoA) indicated a distinct separation between the two methods (P<0.05) and tighter clustering (less variability) in Ph/Chl+C3 than in MoBio-K. These results suggest that the Ph/Chl+C3 may be preferred for research to identify specific Firmicutes taxa such as Clostridium, and Bacillus. However; MoBio-K may be a better choice for projects focusing on Bacteroidetes abundance. The Ph/Chl+C3 method required less time, but has some safety concerns associated with exposure and disposal of phenol and chloroform. While the MoBio-K may be better choice for researchers with less access to safety equipment like a fume hood.
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http://dx.doi.org/10.1016/j.mimet.2016.07.019DOI Listing
October 2016

The subzero microbiome: microbial activity in frozen and thawing soils.

FEMS Microbiol Ecol 2016 Jun 21;92(6):fiw081. Epub 2016 Apr 21.

Department of Biochemistry and Microbiology

Most of the Earth's biosphere is characterized by low temperatures (<5°C) and cold-adapted microorganisms are widespread. These psychrophiles have evolved a complex range of adaptations of all cellular constituents to counteract the potentially deleterious effects of low kinetic energy environments and the freezing of water. Microbial life continues into the subzero temperature range, and this activity contributes to carbon and nitrogen flux in and out of ecosystems, ultimately affecting global processes. Microbial responses to climate warming and, in particular, thawing of frozen soils are not yet well understood, although the threat of microbial contribution to positive feedback of carbon flux is substantial. To date, several studies have examined microbial community dynamics in frozen soils and permafrost due to changing environmental conditions, and some have undertaken the complicated task of characterizing microbial functional groups and how their activity changes with changing conditions, either in situ or by isolating and characterizing macromolecules. With increasing temperature and wetter conditions microbial activity of key microbes and subsequent efflux of greenhouse gases also increase. In this review, we aim to provide an overview of microbial activity in seasonally frozen soils and permafrost. With a more detailed understanding of the microbiological activities in these vulnerable soil ecosystems, we can begin to predict and model future expectations for carbon release and climate change.
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http://dx.doi.org/10.1093/femsec/fiw081DOI Listing
June 2016

The Effect of Diet and Exercise on Intestinal Integrity and Microbial Diversity in Mice.

PLoS One 2016 8;11(3):e0150502. Epub 2016 Mar 8.

Department of Marine and Coastal Sciences, Rutgers University, New Brunswick, NJ 08901, United States of America.

Background: The gut microbiota is now known to play an important role contributing to inflammatory-based chronic diseases. This study examined intestinal integrity/inflammation and the gut microbial communities in sedentary and exercising mice presented with a normal or high-fat diet.

Methods: Thirty-six, 6-week old C57BL/6NTac male mice were fed a normal or high-fat diet for 12-weeks and randomly assigned to exercise or sedentary groups. After 12 weeks animals were sacrificed and duodenum/ileum tissues were fixed for immunohistochemistry for occludin, E-cadherin, and cyclooxygenase-2 (COX-2). The bacterial communities were assayed in fecal samples using terminal restriction fragment length polymorphism (TRFLP) analysis and pyrosequencing of 16S rRNA gene amplicons.

Results: Lean sedentary (LS) mice presented normal histologic villi while obese sedentary (OS) mice had similar villi height with more than twice the width of the LS animals. Both lean (LX) and obese exercise (OX) mice duodenum and ileum were histologically normal. COX-2 expression was the greatest in the OS group, followed by LS, LX and OX. The TRFLP and pyrosequencing indicated that members of the Clostridiales order were predominant in all diet groups. Specific phylotypes were observed with exercise, including Faecalibacterium prausnitzi, Clostridium spp., and Allobaculum spp.

Conclusion: These data suggest that exercise has a strong influence on gut integrity and host microbiome which points to the necessity for more mechanistic studies of the interactions between specific bacteria in the gut and its host.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0150502PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4783017PMC
August 2016

Identification of a Ruminococcaceae Species as the Methyl tert-Butyl Ether (MTBE) Degrading Bacterium in a Methanogenic Consortium.

Environ Sci Technol 2016 Feb 15;50(3):1455-64. Epub 2016 Jan 15.

Department of Biochemistry and Microbiology, Rutgers University , New Brunswick, New Jersey 08901, United States.

The widespread use of methyl tert-butyl ether (MTBE) has caused major contamination of groundwater sources and is a concern due to its taste and odor problems, as well as its toxicity. MTBE can be degraded anaerobically which makes bioremediation of contaminated aquifers a potential solution. Nevertheless, the organisms and mechanisms that are responsible for anaerobic MTBE degradation are still unknown. The aim of our research was to identify the organisms actively degrading MTBE. For this purpose we characterized an anaerobic methanogenic culture enriched with MTBE as the sole carbon source from the New Jersey Arthur Kill intertidal strait sediment. The cultures were analyzed using stable isotope probing (SIP) combined with terminal restriction fragment length polymorphism (T-RFLP), high-throughput sequencing and clone library analysis of bacterial 16S rRNA genes. The sequence data indicated that phylotypes belonging to the Ruminococcaceae in the Firmicutes were predominant in the methanogenic cultures. SIP experiments also showed sequential incorporation of the (13)C labeled MTBE by the bacterial community with a bacterium most closely related to Saccharofermentans acetigenes identified as the bacterium active in O-demethylation of MTBE. Identification of the microorganisms responsible for the activity will help us better understand anaerobic MTBE degradation processes in the field and determine biomarkers for monitoring natural attenuation.
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http://dx.doi.org/10.1021/acs.est.5b04731DOI Listing
February 2016

BioDry: An Inexpensive, Low-Power Method to Preserve Aquatic Microbial Biomass at Room Temperature.

PLoS One 2015 28;10(12):e0144686. Epub 2015 Dec 28.

Department of Marine and Coastal Science, Rutgers University, New Brunswick, New Jersey, United States of America.

This report describes BioDry (patent pending), a method for reliably preserving the biomolecules associated with aquatic microbial biomass samples, without the need of hazardous materials (e.g. liquid nitrogen, preservatives, etc.), freezing, or bulky storage/sampling equipment. Gel electrophoresis analysis of nucleic acid extracts from samples treated in the lab with the BioDry method indicated that molecular integrity was protected in samples stored at room temperature for up to 30 days. Analysis of 16S/18S rRNA genes for presence/absence and relative abundance of microorganisms using both 454-pyrosequencing and TRFLP profiling revealed statistically indistinguishable communities from control samples that were frozen in liquid nitrogen immediately after collection. Seawater and river water biomass samples collected with a portable BioDry "field unit", constructed from off-the-shelf materials and a battery-operated pumping system, also displayed high levels of community rRNA preservation, despite a slight decrease in nucleic acid recovery over the course of storage for 30 days. Functional mRNA and protein pools from the field samples were also effectively conserved with BioDry, as assessed by respective RT-PCR amplification and western blot of ribulose-1-5-bisphosphate carboxylase/oxygenase. Collectively, these results demonstrate that BioDry can adequately preserve a suite of biomolecules from aquatic biomass at ambient temperatures for up to a month, giving it great potential for high resolution sampling in remote locations or on autonomous platforms where space and power are limited.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0144686PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4692454PMC
August 2016

Identification of Bacteria Synthesizing Ribosomal RNA in Response to Uranium Addition During Biostimulation at the Rifle, CO Integrated Field Research Site.

PLoS One 2015 18;10(9):e0137270. Epub 2015 Sep 18.

Department of Marine and Coastal Science, Rutgers University, New Brunswick, NJ, United States of America.

Understanding which organisms are capable of reducing uranium at historically contaminated sites provides crucial information needed to evaluate treatment options and outcomes. One approach is determination of the bacteria which directly respond to uranium addition. In this study, uranium amendments were made to groundwater samples from a site of ongoing biostimulation with acetate. The active microbes in the planktonic phase were deduced by monitoring ribosomes production via RT-PCR. The results indicated several microorganisms were synthesizing ribosomes in proportion with uranium amendment up to 2 μM. Concentrations of U (VI) >2 μM were generally found to inhibit ribosome synthesis. Two active bacteria responding to uranium addition in the field were close relatives of Desulfobacter postgateii and Geobacter bemidjiensis. Since RNA content often increases with growth rate, our findings suggest it is possible to rapidly elucidate active bacteria responding to the addition of uranium in field samples and provides a more targeted approach to stimulate specific populations to enhance radionuclide reduction in contaminated sites.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0137270PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4575074PMC
May 2016

Identification of Anaerobic Aniline-Degrading Bacteria at a Contaminated Industrial Site.

Environ Sci Technol 2015 Sep 26;49(18):11079-88. Epub 2015 Aug 26.

Department of Environmental Sciences, Rutgers University , 14 College Farm Road, New Brunswick, New Jersey 08901, United States.

Anaerobic aniline biodegradation was investigated under different electron-accepting conditions using contaminated canal and groundwater aquifer sediments from an industrial site. Aniline loss was observed in nitrate- and sulfate-amended microcosms and in microcosms established to promote methanogenic conditions. Lag times of 37 days (sulfate amended) to more than 100 days (methanogenic) were observed prior to activity. Time-series DNA-stable isotope probing (SIP) was used to identify bacteria that incorporated (13)C-labeled aniline in the microcosms established to promote methanogenic conditions. In microcosms from heavily contaminated aquifer sediments, a phylotype with 92.7% sequence similarity to Ignavibacterium album was identified as a dominant aniline degrader as indicated by incorporation of (13)C-aniline into its DNA. In microcosms from contaminated canal sediments, a bacterial phylotype within the family Anaerolineaceae, but without a match to any known genus, demonstrated the assimilation of (13)C-aniline. Acidovorax spp. were also identified as putative aniline degraders in both of these two treatments, indicating that these species were present and active in both the canal and aquifer sediments. There were multiple bacterial phylotypes associated with anaerobic degradation of aniline at this complex industrial site, which suggests that anaerobic transformation of aniline is an important process at the site. Furthermore, the aniline degrading phylotypes identified in the current study are not related to any known aniline-degrading bacteria. The identification of novel putative aniline degraders expands current knowledge regarding the potential fate of aniline under anaerobic conditions.
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http://dx.doi.org/10.1021/acs.est.5b02166DOI Listing
September 2015

Spatial distribution of an uranium-respiring betaproteobacterium at the Rifle, CO field research site.

PLoS One 2015 13;10(4):e0123378. Epub 2015 Apr 13.

Inst. of Marine and Coastal Science, Rutgers University, New Brunswick, New Jersey, United States of America.

The Department of Energy's Integrated Field-Scale Subsurface Research Challenge Site (IFRC) at Rifle, Colorado was created to address the gaps in knowledge on the mechanisms and rates of U(VI) bioreduction in alluvial sediments. Previous studies at the Rifle IFRC have linked microbial processes to uranium immobilization during acetate amendment. Several key bacteria believed to be involved in radionuclide containment have been described; however, most of the evidence implicating uranium reduction with specific microbiota has been indirect. Here, we report on the cultivation of a microorganism from the Rifle IFRC that reduces uranium and appears to utilize it as a terminal electron acceptor for respiration with acetate as electron donor. Furthermore, this bacterium constitutes a significant proportion of the subsurface sediment community prior to biostimulation based on TRFLP profiling of 16S rRNA genes. 16S rRNA gene sequence analysis indicates that the microorganism is a betaproteobacterium with a high similarity to Burkholderia fungorum. This is, to our knowledge, the first report of a betaproteobacterium capable of uranium respiration. Our results indicate that this microorganism occurs commonly in alluvial sediments located between 3-6 m below ground surface at Rifle and may play a role in the initial reduction of uranium at the site.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0123378PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4395306PMC
January 2016

Crenarchaeal heterotrophy in salt marsh sediments.

ISME J 2014 Jul 20;8(7):1534-43. Epub 2014 Feb 20.

Institute of Marine and Coastal Science, Rutgers University, New Brunswick, NJ, USA.

Mesophilic Crenarchaeota (also known as Thaumarchaeota) are ubiquitous and abundant in marine habitats. However, very little is known about their metabolic function in situ. In this study, salt marsh sediments from New Jersey were screened via stable isotope probing (SIP) for heterotrophy by amending with a single (13)C-labeled compound (acetate, glycine or urea) or a complex (13)C-biopolymer (lipids, proteins or growth medium (ISOGRO)). SIP incubations were done at two substrate concentrations (30-150 μM; 2-10 mg ml(-1)), and (13)C-labeled DNA was analyzed by terminal restriction fragment length polymorphism (TRFLP) analysis of 16S rRNA genes. To test for autotrophy, an amendment with (13)C-bicarbonate was also performed. Our SIP analyses indicate salt marsh crenarchaea are heterotrophic, double within 2-3 days and often compete with heterotrophic bacteria for the same organic substrates. A clone library of (13)C-amplicons was screened to find matches to the (13)C-TRFLP peaks, with seven members of the Miscellaneous Crenarchaeal Group and seven members from the Marine Group 1.a Crenarchaeota being discerned. Some of these crenarchaea displayed a preference for particular carbon sources, whereas others incorporated nearly every (13)C-substrate provided. The data suggest salt marshes may be an excellent model system for studying crenarchaeal metabolic capabilities and can provide information on the competition between crenarchaea and other microbial groups to improve our understanding of microbial ecology.
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http://dx.doi.org/10.1038/ismej.2014.15DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4069397PMC
July 2014

Bacterial genome replication at subzero temperatures in permafrost.

ISME J 2014 Jan 29;8(1):139-49. Epub 2013 Aug 29.

Institute of Marine and Coastal Science, School of Environmental and Biological Sciences, Rutgers University, New Brunswick, NJ, USA.

Microbial metabolic activity occurs at subzero temperatures in permafrost, an environment representing ∼25% of the global soil organic matter. Although much of the observed subzero microbial activity may be due to basal metabolism or macromolecular repair, there is also ample evidence for cellular growth. Unfortunately, most metabolic measurements or culture-based laboratory experiments cannot elucidate the specific microorganisms responsible for metabolic activities in native permafrost, nor, can bulk approaches determine whether different members of the microbial community modulate their responses as a function of changing subzero temperatures. Here, we report on the use of stable isotope probing with (13)C-acetate to demonstrate bacterial genome replication in Alaskan permafrost at temperatures of 0 to -20 °C. We found that the majority (80%) of operational taxonomic units detected in permafrost microcosms were active and could synthesize (13)C-labeled DNA when supplemented with (13)C-acetate at temperatures of 0 to -20 °C during a 6-month incubation. The data indicated that some members of the bacterial community were active across all of the experimental temperatures, whereas many others only synthesized DNA within a narrow subzero temperature range. Phylogenetic analysis of (13)C-labeled 16S rRNA genes revealed that the subzero active bacteria were members of the Acidobacteria, Actinobacteria, Chloroflexi, Gemmatimonadetes and Proteobacteria phyla and were distantly related to currently cultivated psychrophiles. These results imply that small subzero temperature changes may lead to changes in the active microbial community, which could have consequences for biogeochemical cycling in permanently frozen systems.
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http://dx.doi.org/10.1038/ismej.2013.140DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3869017PMC
January 2014

Impact of mercury on denitrification and denitrifying microbial communities in nitrate enrichments of subsurface sediments.

Biodegradation 2013 Feb 8;24(1):33-46. Epub 2012 Jun 8.

Department of Biochemistry and Microbiology, Rutgers University, 223C Lipman Hall, 76 Lipman Dr., New Brunswick, NJ 08901, USA.

The contamination of groundwater with mercury (Hg) is an increasing problem worldwide. Yet, little is known about the interactions of Hg with microorganisms and their processes in subsurface environments. We tested the impact of Hg on denitrification in nitrate reducing enrichment cultures derived from subsurface sediments from the Oak Ridge Integrated Field Research Challenge site, where nitrate is a major contaminant and where bioremediation efforts are in progress. We observed an inverse relationship between Hg concentrations and onset and rates of denitrification in nitrate enrichment cultures containing between 53 and 1.1 μM of inorganic Hg; higher Hg concentrations increasingly extended the time to onset of denitrification and inhibited denitrification rates. Microbial community complexity, as indicated by terminal restriction fragment length polymorphism (tRFLP) analysis of the 16S rRNA genes, declined with increasing Hg concentrations; at the 312 nM Hg treatment, a single tRFLP peak was detected representing a culture of Bradyrhizobium sp. that possessed the merA gene indicating a potential for Hg reduction. A culture identified as Bradyrhizobium sp. strain FRC01 with an identical 16S rRNA sequence to that of the enriched peak in the tRFLP patterns, reduced Hg(II) to Hg(0) and carried merA whose amino acid sequence has 97 % identity to merA from the Proteobacteria and Firmicutes. This study demonstrates that in subsurface sediment incubations, Hg may inhibit denitrification and that inhibition may be alleviated when Hg resistant denitrifying Bradyrhizobium spp. detoxify Hg by its reduction to the volatile elemental form.
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http://dx.doi.org/10.1007/s10532-012-9555-8DOI Listing
February 2013

Phase preference by active, acetate-utilizing bacteria at the rifle, CO integrated field research challenge site.

Environ Sci Technol 2011 Feb 12;45(4):1250-6. Epub 2011 Jan 12.

Institute of Marine and Coastal Science, Rutgers University, New Brunswick, NJ 08901-8521, USA.

Previous experiments at the Rifle, Colorado Integrated Field Research Challenge (IFRC) site demonstrated that field-scale addition of acetate to groundwater reduced the ambient soluble uranium concentration. In this report, sediment samples collected before and after acetate field addition were used to assess the active microbes via (13)C acetate stable isotope probing on 3 phases [coarse sand, fines (8-approximately 150 μm), groundwater (0.2-8 μm)] over a 24-day time frame. TRFLP results generally indicated a stronger signal in (13)C-DNA in the "fines" fraction compared to the sand and groundwater. Before the field-scale acetate addition, a Geobacter-like group primarily synthesized (13)C-DNA in the groundwater phase, an alpha Proteobacterium primarily grew on the fines/sands, and an Acinetobacter sp. and Decholoromonas-like OTU utilized much of the (13)C acetate in both groundwater and particle-associated phases. At the termination of the field-scale acetate addition, the Geobacter-like species was active on the solid phases rather than the groundwater, while the other bacterial groups had very reduced newly synthesized DNA signal. These findings will help to delineate the acetate utilization patterns of bacteria in the field and can lead to improved methods for stimulating distinct microbial populations in situ.
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http://dx.doi.org/10.1021/es102893rDOI Listing
February 2011

The effect of co-substrate activation on indigenous and bioaugmented PCB dechlorinating bacterial communities in sediment microcosms.

Appl Microbiol Biotechnol 2011 Mar 3;89(6):2005-17. Epub 2010 Nov 3.

Department of Biochemistry and Microbiology, Rutgers, The State University of New Jersey, New Brunswick, NJ 08901, USA.

Microbial reductive dechlorination by members of the phylum Chloroflexi, including the genus Dehalococcoides, may play an important role in natural detoxification of highly chlorinated environmental pollutants, such as polychlorinated biphenyls (PCBs). Previously, we showed the increase of an indigenous bacterial population belonging to the Pinellas subgroup of Dehalococcoides spp. in Anacostia River sediment (Washington DC, USA) microcosms treated with halogenated co-substrates ("haloprimers"), tetrachlorobenzene (TeCB), or pentachloronitrobenzene (PCNB). The PCNB-amended microcosms exhibited enhanced dechlorination of weathered PCBs, while TeCB-amended microcosms did not. We therefore developed and used different phylogenetic approaches to discriminate the effect of the two different haloprimers. We also developed complementary approaches to monitor the effects of haloprimer treatments on 12 putative reductive dehalogenase (rdh) genes common to Dehalococcoides ethenogenes strain 195 and Dehalococcoides sp. strain CBDB1. Our results indicate that 16S rRNA gene-based phylogenetic analyses have a limit in their ability to distinguish the effects of two haloprimer treatments and that two of rdh genes were present in high abundance when microcosms were amended with PCNB, but not TeCB. rdh gene-based phylogenetic analysis supports that these two rdh genes originated from the Pinellas subgroup of Dehalococcoides spp., which corresponds to the 16S rRNA gene-based phylogenetic analysis.
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http://dx.doi.org/10.1007/s00253-010-2958-8DOI Listing
March 2011

Community characterization of anaerobic methyl tert-butyl ether (MTBE)-degrading enrichment cultures.

FEMS Microbiol Ecol 2010 May 22;72(2):279-88. Epub 2010 Jan 22.

Department of Biochemistry and Microbiology, Rutgers, The State University of New Jersey, New Brunswick, NJ 08901-8525, USA.

Use of the fuel oxygenate methyl tert-butyl ether (MTBE) has led to widespread environmental contamination. Anaerobic biodegradation of MTBE observed under different redox conditions is a potential means for remediation of contaminated aquifers; however, no responsible microorganisms have been identified as yet. We analyzed the bacterial communities in anaerobic-enriched cultures originating from three different contaminated sediments that have retained MTBE-degrading activity for over a decade. MTBE was transformed to tert-butyl alcohol and the methyl group used as a carbon and energy source. Terminal restriction fragment length polymorphism (T-RFLP) analysis of bacterial 16S rRNA genes showed that the MTBE-utilizing microcosms established from different sediment sources had substantially different community profiles, suggesting that multiple species are capable of MTBE biodegradation. The 16S rRNA genes from one enrichment culture were cloned and sequenced. Phylogenetic analysis showed a diverse community, with phylotypes belonging to the Proteobacteria, Bacteroidetes, Firmicutes, Chloroflexi and Thermotogae. Continued enrichment on MTBE further reduced the community to three predominant phylotypes, as evidenced by T-RFLP analysis, which were most closely related to the Deltaproteobacteria, Firmicutes and Chloroflexi. These three common operational taxonomic units were detectable in the enrichments from Atlantic and Pacific coastal samples. Identification of the microorganisms important in mediating anaerobic MTBE transformation will provide the foundation for developing tools for site assessment and bioremediation monitoring.
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http://dx.doi.org/10.1111/j.1574-6941.2010.00841.xDOI Listing
May 2010

Detection of 2,4,6-trinitrotoluene-utilizing anaerobic bacteria by 15N and 13C incorporation.

Appl Environ Microbiol 2010 Mar 15;76(5):1695-8. Epub 2010 Jan 15.

Department of Environmental Sciences, the State University of New Jersey, New Brunswick, New Jersey 08901, USA.

2,4,6-Trinitrotoluene ((15)N or (13)C labeled) was added to Norfolk Harbor sediments to test whether anaerobic bacteria use TNT for growth. Stable-isotope probing (SIP)-terminal restriction fragment length polymorphism (TRFLP) detected peaks in the [(15)N]TNT cultures (60, 163, and 168 bp). The 60-bp peak was also present in the [(13)C]TNT cultures and was related to Lysobacter taiwanensis.
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http://dx.doi.org/10.1128/AEM.02274-09DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2832382PMC
March 2010

PCB dechlorination enhancement in Anacostia River sediment microcosms.

Water Res 2009 Oct 6;43(18):4549-58. Epub 2009 Aug 6.

Department of Environmental Sciences, Rutgers-The State University of New Jersey, 14 College Farm Road, New Brunswick, NJ 08901, USA.

In situ treatment of PCB contaminated sediments via microbial dechlorination is a promising alternative to dredging, which may be reserved for only the most contaminated areas. Reductive dechlorination of low levels of weathered PCB mixtures typical of urban environments may occur at slow rates. Here, we report that biostimulation and bioaugmentation enhanced dechlorination of low concentration (2.1 mg PCBs/kg dry weight) historical PCBs in microcosms prepared with Anacostia River, Washington, DC, sediment. Treatments included electron donors butyrate, lactate, propionate and acetate (1 mM each); alternate halogenated electron acceptors (haloprimers) tetrachlorobenzene (TeCB, 25 microM), pentachloronitrobenzene (PCNB, 25 microM), or 2,3,4,5,6-PCB (PCB116, 2.0 microM); and/or bioaugmentation with a culture containing Dehalococcoides ethenogenes strain 195 (3 x 10(6)cells/mL). Dechlorination rates were enhanced in microcosms receiving bioaugmentation, PCNB and PCNB plus bioaugmentation, compared to other treatments. Microcosm subcultures generated after 415 days and spiked with PCB116 showed sustained capacity for dechlorination of PCB116 in PCNB, PCNB plus bioaugmentation, and TeCB treatments, relative to other treatments. Analysis of Chloroflexi 16S rRNA genes showed that TeCB and PCNB increased native Dehalococcoides spp. from the Pinellas subgroup; however this increase was correlated to enhanced dechlorination of low concentration weathered PCBs only in PCNB-amended microcosms. D. ethenogenes strain 195 was detected only in bioaugmented microcosms and decreased over 281 days. Bioaugmentation with D. ethenogenes strain 195 increased PCB dechlorination rates initially, but enhanced capacity for dechlorination of a model congener, PCB116, after 415 days occurred only in microcosms with enhanced native Dehalococcoides spp.
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http://dx.doi.org/10.1016/j.watres.2009.08.003DOI Listing
October 2009

Desulfoluna spongiiphila sp. nov., a dehalogenating bacterium in the Desulfobacteraceae from the marine sponge Aplysina aerophoba.

Int J Syst Evol Microbiol 2009 Sep 15;59(Pt 9):2133-9. Epub 2009 Jul 15.

Department of Biochemistry and Microbiology and Biotechnology Center for Agriculture and the Environment, The State University of New Jersey, New Brunswick, NJ, USA.

A reductively dehalogenating, strictly anaerobic, sulfate-reducing bacterium, designated strain AA1T, was isolated from the marine sponge Aplysina aerophoba collected in the Mediterranean Sea and was characterized phenotypically and phylogenetically. Cells of strain AA1T were Gram-negative, short, curved rods. Growth of strain AA1T was observed between 20 and 37 degrees C (optimally at 28 degrees C) at pH 7-8. NaCl was required for growth; optimum growth occurred in the presence of 25 g NaCl l(-1). Growth occurred with lactate, propionate, pyruvate, succinate, benzoate, glucose and sodium citrate as electron donors and carbon sources and either sulfate or 2-bromophenol as electron acceptors, but not with acetate or butyrate. Strain AA1T was able to dehalogenate several different bromophenols, and 2- and 3-iodophenol, but not monochlorinated or fluorinated phenols. Lactate, pyruvate, fumarate and malate were not utilized without an electron acceptor. The G+C content of the genomic DNA was 58.5 mol%. The predominant cellular fatty acids were C14:0, iso-C14:0, C14:0 3-OH, anteiso-C15:0, C16:0, C16:1omega7c and C18:1omega7c. Phylogenetic analysis based on 16S rRNA gene sequence comparisons placed the novel strain within the class Deltaproteobacteria. Strain AA1T was related most closely to the type strains of Desulfoluna butyratoxydans (96% 16S rRNA gene sequence similarity), Desulfofrigus oceanense (95%) and Desulfofrigus fragile (95%). Based on its phenotypic, physiological and phylogenetic characteristics, strain AA1T is considered to represent a novel species of the genus Desulfoluna, for which the name Desulfoluna spongiiphila sp. nov. is proposed. The type strain is AA1T (=DSM 17682T=ATCC BAA-1256T).
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http://dx.doi.org/10.1099/ijs.0.005884-0DOI Listing
September 2009

Quantifying nonspecific TEM beta-lactamase (blaTEM) genes in a wastewater stream.

Appl Environ Microbiol 2009 Jan 7;75(1):203-11. Epub 2008 Nov 7.

The Biological Laboratories, Harvard University, 16 Divinity Ave., Cambridge, MA 02138, USA.

To control the antibiotic resistance epidemic, it is necessary to understand the distribution of genetic material encoding antibiotic resistance in the environment and how anthropogenic inputs, such as wastewater, affect this distribution. Approximately two-thirds of antibiotics administered to humans are beta-lactams, for which the predominant bacterial resistance mechanism is hydrolysis by beta-lactamases. Of the beta-lactamases, the TEM family is of overriding significance with regard to diversity, prevalence, and distribution. This paper describes the design of DNA probes universal for all known TEM beta-lactamase genes and the application of a quantitative PCR assay (also known as Taqman) to quantify these genes in environmental samples. The primer set was used to study whether sewage, both treated and untreated, contributes to the spread of these genes in receiving waters. It was found that while modern sewage treatment technologies reduce the concentrations of these antibiotic resistance genes, the ratio of bla(TEM) genes to 16S rRNA genes increases with treatment, suggesting that bacteria harboring bla(TEM) are more likely to survive the treatment process. Thus, beta-lactamase genes are being introduced into the environment in significantly higher concentrations than occur naturally, creating reservoirs of increased resistance potential.
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http://dx.doi.org/10.1128/AEM.01254-08DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2612200PMC
January 2009

Temporal patterns of microbial community structure in the Mid-Atlantic Bight.

FEMS Microbiol Ecol 2008 Sep 25;65(3):484-93. Epub 2008 Jul 25.

Pt Pleasant High School, Pt Pleasant, NJ, USA.

Although open ocean time-series sites have been areas of microbial research for years, relatively little is known about the population dynamics of bacterioplankton communities in the coastal ocean on kilometer spatial and seasonal temporal scales. To gain a better understanding of microbial community variability, monthly samples of bacterial biomass were collected in 1995-1996 along a 34-km transect near the Long-Term Ecosystem Observatory (LEO-15) off the New Jersey coast. Surface and bottom sampling was performed at seven stations along a transect line with depths ranging from 1 to 35 m (n=178). Microbial populations were fingerprinted using ribosomal 16S rRNA genes and terminal restriction fragment length polymorphism analysis. Results from cluster analysis revealed distinct temporal patterns among the bacterioplankton communities in the Mid-Atlantic Bight rather than grouping by sample location or depth. Principal components analysis models supported the temporal patterns. In addition, partial least squares regression modeling could not discern a significant correlation from traditional oceanographic physical and phytoplankton nutrient parameters on overall bacterial community variability patterns at LEO-15. These results suggest factors not traditionally measured during oceanographic studies are structuring coastal microbial communities.
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http://dx.doi.org/10.1111/j.1574-6941.2008.00553.xDOI Listing
September 2008