Publications by authors named "Dipankar Bachar"

23 Publications

  • Page 1 of 1

Impact of smoking cessation, coffee and bread consumption on the intestinal microbial composition among Saudis: A cross-sectional study.

PLoS One 2020 29;15(4):e0230895. Epub 2020 Apr 29.

Special Infectious Agents Unit, King Fahd Medical Research Center, King Abdulaziz University, Jeddah, Saudi Arabia.

The gut microbiota is often affected by the dietary and lifestyle habits of the host, resulting in a better efficacy that favors energy harvesting from the consumed food. Our objective was to characterize the composition of gut microbiota in adult Saudis and investigate possible association with lifestyle and dietary practices. Feces from 104 Saudi volunteers (48% males) were tested for microbiota by sequencing the V3-V4 region of bacterial 16S ribosomal RNA (rRNA). For all participants, data were collected related to their lifestyle habits and dietary practices. The relative abundance (RA) of Fusobacteria was significantly higher in normal weight Saudis (P = 0.005, false discovery rate-FDR = 0.014). Individuals who consumed more coffee presented marginally significant more RA of Fusobacteria (P = 0.02, FDR = 0.20) in their gut microbiota compared to those reporting low or no coffee intake, but the RA of Fusobacteria was significantly higher in smokers compared to non-smokers (P = 0.009, FDR = 0.027). The RA of Fusobacteria was also significantly higher in those reporting daily consumption of bread (P = 0.005, FDR = 0.015). At the species level, the gut microbiota of people who consumed coffee was dominated by Bacteroides thetaiotaomicron followed by Phascolarctobacterium faecium and Eubacterium rectale. Similarly, the gut microbiota of smokers was also enriched by B. thetaiotaomicron and Lactobacillus amylovorus. Smoking cessation, bread and coffee consumption induce changes in the intestinal microbial composition of Saudis. This indicates the significance of diet and lifestyle practices in the determination of the composition of the gut microbiota, which could possibly lead later to changes in metabolic profile and weight.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0230895PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7190147PMC
July 2020

Metagenomic and culturomic analysis of gut microbiota dysbiosis during Clostridium difficile infection.

Sci Rep 2019 09 5;9(1):12807. Epub 2019 Sep 5.

Aix Marseille Univ, IRD, MEPHI, IHU-Méditerranée Infection, Marseille, France.

Recently, cocktail of bacteria were proposed in order to treat Clostridium difficile infection (CDI), but these bacteriotherapies were selected more by chance than experimentation. We propose to comprehensively explore the gut microbiota of patients with CDI compared to healthy donors in order to propose a consortium of bacteria for treating C. difficile. We compared stool samples composition from 11 CDI patients and 8 healthy donors using two techniques: metagenomics, 16S V3-V4 region amplification and sequencing and culturomics, high throughout culture using six culture conditions and MALDI-TOF identification. By culturomics, we detected 170 different species in the CDI group and 275 in the control group. Bacteroidetes were significantly underrepresented in the CDI group (p = 0.007). By metagenomics, 452 different operational taxonomic units assigned to the species level were detected in the CDI group compared to 522 in the control group. By these two techniques, we selected 37 bacteria only found in control group in more than 75% of the samples and/or with high relative abundance, 10 of which have already been tested in published bacteriotherapies against CDI, and 3 of which (Bifidobacterium adolescentis, Bifidobacterium longum and Bacteroides ovatus) have been detected by these two techniques. This controlled number of bacteria could be administrated orally in a non-invasive way in order to treat CDI.
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http://dx.doi.org/10.1038/s41598-019-49189-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6728329PMC
September 2019

Gut Microbiota Alteration is Characterized by a Proteobacteria and Fusobacteria Bloom in Kwashiorkor and a Bacteroidetes Paucity in Marasmus.

Sci Rep 2019 06 24;9(1):9084. Epub 2019 Jun 24.

Aix-Marseille Univ, IRD, APHM, MEPHI, IHU-Méditerranée Infection, Marseille, France.

Kwashiorkor and marasmus are considered to be two different clinical diseases resulting from severe malnutrition, but this distinction has been questioned. In a previous study comparing children with kwashiorkor and healthy children from Niger and Senegal, we found a dramatic gut microbiota alteration with a predominant depletion of anaerobes and enrichment in Proteobacteria and Fusobacteria in kwashiorkor. However, it remained unknown whether this association was related to malnutrition or was a specific feature of kwashiorkor. In this continuation study, we added 7 new marasmus subjects and 71,162 new colonies from the same countries. Our results showed that, compared to marasmus, the kwashiorkor gut microbiota was characterized by an increased proportion of Proteobacteria (culturomics, Marasmus 5.0%, Kwashiorkor 16.7%, p < 0.0001; metagenomics, Marasmus 14.7%, Kwashiorkor 22.0%, p = 0.001), but there was a decreased proportion of Bacteroidetes in marasmus (culturomics, Marasmus 0.8%, Kwashiorkor 6.5%, p = 0.001; metagenomics, Marasmus 5.4%, Kwashiorkor 7.0%, p = 0.03). Fusobacterium was more frequently cultured from kwashiorkor. All detected potential pathogenic species were enriched in the kwashiorkor gut microbiota. These results provide a biological basis to support the usage of an antibiotic therapy more effective in suppressing the overgrowth of bacterial communities resistant to penicillin, combined with antioxidants and probiotics for nutritional recovery therapies, particularly for kwashiorkor.
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http://dx.doi.org/10.1038/s41598-019-45611-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6591176PMC
June 2019

Repertoire of the gut microbiota from stomach to colon using culturomics and next-generation sequencing.

BMC Microbiol 2018 10 24;18(1):157. Epub 2018 Oct 24.

Aix Marseille Univ, IRD, MEPHI, IHU - Méditerranée Infection, 19-21 Boulevard Jean Moulin, 13005, Marseille, France.

Background: Most studies on the human microbiota have analyzed stool samples, although a large proportion of the absorption of nutrients takes place in upper gut tract. We collected samples from different locations along the entire gastrointestinal tract from six patients who had simultaneously undergone upper endoscopy and colonoscopy, to perform a comprehensive analysis using culturomics with matrix assisted laser desorption ionisation - time of flight (MALDI-TOF) identification and by metagenomics targeting the 16S ribosomal ribonucleic acid (rRNA) gene.

Results: Using culturomics, we isolated 368 different bacterial species, including 37 new species. Fewer species were isolated in the upper gut: 110 in the stomach and 106 in the duodenum, while 235 were isolated from the left colon (p < 0.02). We isolated fewer aero-intolerant species in the upper gut: 37 from the stomach and 150 from the left colon (p < 0.004). Using metagenomics, 1,021 species were identified. The upper gut microbiota was revealed to be less rich than the lower gut microbiota, with 37,622 reads from the stomach, 28,390 from the duodenum, and 79,047 from the left colon (p < 0.009). There were fewer reads for aero-intolerant species in the upper gut (8,656 in the stomach, 5,188 in the duodenum and 72,262 in the left colon, p < 0.02). Patients taking proton pump inhibitors (PPI) were then revealed to have a higher stomach pH and a greater diversity of species in the upper digestive tract than patients not receiving treatment (p < 0.001).

Conclusion: Significant modifications in bacterial composition and diversity exist throughout the gastrointestinal tract. We suggest that the upper gut may be key to understanding the relationship between the gut microbiota and health.
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http://dx.doi.org/10.1186/s12866-018-1304-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6201554PMC
October 2018

16S Metagenomic Comparison of -Infected and Noninfected and Microbiota from Senegal.

Am J Trop Med Hyg 2018 12 18;99(6):1489-1498. Epub 2018 Oct 18.

Aix Marseille Université, IRD, AP-HM, Microbes, Evolution, Phylogeny and Infection (MEPHI), Marseille, France.

In the context of the pre-elimination of malaria, biological control may provide an alternative or additional tool to current malaria control strategies. During their various stages of development, mosquitoes undergo subsequent changes in their associated microbiota, depending on their environment and nutritional status. Although s.l. and are the two major malaria vectors in Senegal, the composition of their microbiota is not yet well known. In this study, we explored the microbiota of mosquitoes naturally infected or not by () using the 16S ribosomal RNA gene-based bacterial metagenomic approach. In both vector species, the microbiota was more diverse in infected samples than in the noninfected ones, although the total number of reads appeared to be higher in noninfected mosquitoes. Overall, the microbiota was different between the two vector species. Noteworthy, the bacterial microbiota was significantly different between -positive and -negative groups whatever the species, but was similar between individuals of the same infection status within a species. Overall, the phylum of was the most predominant in both species, with bacteria of the genus outweighing the others in noninfected vectors. The presence of some specific bacterial species such as , , , and was also observed in -free samples only. These preliminary observations pave the way for further characterization of the mosquito microbiota to select promising bacterial candidates for potential use in an innovative approach to controlling malaria and overcoming the challenges to achieving a malaria-free world.
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http://dx.doi.org/10.4269/ajtmh.18-0263DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6283497PMC
December 2018

The nasopharyngeal microbiota in patients with viral respiratory tract infections is enriched in bacterial pathogens.

Eur J Clin Microbiol Infect Dis 2018 Sep 22;37(9):1725-1733. Epub 2018 Jul 22.

Aix Marseille Univ, IRD, APHM, MEPHI, IHU-Méditerranée Infection, 19-21 Bd Jean Moulin, 13005, Marseille, France.

The nasopharynx is the primary site of colonization by respiratory pathogen that constitutes the port of entrance in the respiratory tract. The role of mucosal respiratory microbiota in infection has been recently emphasized; therefore, we aimed to assess if a specific respiratory microbiota profile was associated with symptomatic infection and/or with presence of respiratory viruses. We performed a case-control study to characterize the healthy respiratory microbiota and its alteration during acute viral infections. Next-generation sequencing of the 16S rRNA gene was applied to 225 nasopharyngeal samples from 177 patients with viral respiratory infection and 48 matched healthy controls. We evidenced an important decrease of bacterial alpha-diversity in patients with symptomatic respiratory infection and a loss of the healthy core microbiota, specifically anaerobes and Prevotella spp. Moreover, eight respiratory pathogens were enriched in these patients, including Staphylococcus aureus, Haemophilus influenzae, Streptococcus pneumoniae, Moraxella catarrhalis, Dol osigranulum pigrum and Corynebacterium propinquum/pseudodiphtheriticum, whose role in respiratory infection is unclear. The asymptomatic carrier of influenza harbors a microbiota similar to healthy subjects, suggesting a critical role of microbiota in the clinical expression of viruses. These data suggest that the commensal microbiota plays a significant role in susceptibility to viral infection. The frequent co-detection of virus and bacteria raises the question of a strategy to prevent bacterial disease, focusing on the prevention of nasopharyngeal colonization through effective antibiotic treatment. In addition to antibiotics, further studies should test preventive or therapeutic interventions for maintaining or restoring a healthy nasopharyngeal microbiota.
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http://dx.doi.org/10.1007/s10096-018-3305-8DOI Listing
September 2018

Clostridium scindens Is Present in the Gut Microbiota during Clostridium difficile Infection: a Metagenomic and Culturomic Analysis.

J Clin Microbiol 2018 05 25;56(5). Epub 2018 Apr 25.

Aix-Marseille Université, IRD, MEPHI, APHM, IHU Méditerranée Infection, Marseille, France

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http://dx.doi.org/10.1128/JCM.01663-17DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5925712PMC
May 2018

Gut Bacteria Missing in Severe Acute Malnutrition, Can We Identify Potential Probiotics by Culturomics?

Front Microbiol 2017 23;8:899. Epub 2017 May 23.

URMITE, Aix Marseille Université, UM63, Centre National de la Recherche Scientifique 7278, IRD 198, Institut National de la Santé Et de la Recherche Médicale 1095, IHU-Méditerranée InfectionMarseille, France.

Severe acute malnutrition is the world-leading cause of children under-five's death. Recent metagenomics studies have established a link between gut microbiota and severe acute malnutrition, describing an immaturity with a striking depletion in oxygen-sensitive prokaryotes. Amoxicillin and therapeutic diet cure most of the children with severe acute malnutrition but an irreversible disruption of the gut microbiota is suspected in the refractory and most severe cases. In these cases, therapeutic diet may be unable to reverse the microbiota alteration leading to persistent impaired development or death. In addition, as enteric sepsis is a major cause of death in this context, identification of missing gut microbes to be tested as probiotics (live bacteria that confer a benefit to the host) to restore rapidly the healthy gut microbiota and prevent the gut pathogenic invasion is of foremost importance. In this study, stool samples of malnourished patients with kwashiorkor and healthy children were collected from Niger and Senegal and analyzed by culturomics and metagenomics. We found a globally decreased diversity, a decrease in the hitherto unknown diversity (new species isolation), a depletion in oxygen-sensitive prokaryotes including and an enrichment in potentially pathogenic and . A complex of 12 species identified only in healthy children using culturomics and metagenomics were identified as probiotics candidates, providing a possible, defined, reproducible, safe, and convenient alternative to fecal transplantation to restore a healthy gut microbiota in malnourished children. Microbiotherapy based on selected strains has the potential to improve the current treatment of severe acute malnutrition and prevent relapse and death by reestablishing a healthy gut microbiota.
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http://dx.doi.org/10.3389/fmicb.2017.00899DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5440526PMC
May 2017

Culture of previously uncultured members of the human gut microbiota by culturomics.

Nat Microbiol 2016 Nov 7;1:16203. Epub 2016 Nov 7.

Institut Louis Malardé, Papeete, Tahiti, Polynésie Française.

Metagenomics revolutionized the understanding of the relations among the human microbiome, health and diseases, but generated a countless number of sequences that have not been assigned to a known microorganism. The pure culture of prokaryotes, neglected in recent decades, remains essential to elucidating the role of these organisms. We recently introduced microbial culturomics, a culturing approach that uses multiple culture conditions and matrix-assisted laser desorption/ionization-time of flight and 16S rRNA for identification. Here, we have selected the best culture conditions to increase the number of studied samples and have applied new protocols (fresh-sample inoculation; detection of microcolonies and specific cultures of Proteobacteria and microaerophilic and halophilic prokaryotes) to address the weaknesses of the previous studies. We identified 1,057 prokaryotic species, thereby adding 531 species to the human gut repertoire: 146 bacteria known in humans but not in the gut, 187 bacteria and 1 archaea not previously isolated in humans, and 197 potentially new species. Genome sequencing was performed on the new species. By comparing the results of the metagenomic and culturomic analyses, we show that the use of culturomics allows the culture of organisms corresponding to sequences previously not assigned. Altogether, culturomics doubles the number of species isolated at least once from the human gut.
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http://dx.doi.org/10.1038/nmicrobiol.2016.203DOI Listing
November 2016

Gut microbiome and dietary patterns in different Saudi populations and monkeys.

Sci Rep 2016 08 31;6:32191. Epub 2016 Aug 31.

Unité de Recherche sur les Maladies Infectieuses et Tropicales Emergentes: URMITE CNRS-IRD 198 UMR 6236, Aix Marseille Université, Faculté de Médecine, 27 Bd Jean Moulin, 13385 Marseille, France.

Host genetics, environment, lifestyle and proximity between hosts strongly influence the composition of the gut microbiome. To investigate the association of dietary variables with the gut microbiota, we used 16S rDNA sequencing to test the fecal microbiome of Bedouins and urban Saudis and we compared it to the gut microbiome of baboons living in close contact with Bedouins and eating their leftovers. We also analyzed fermented dairy products commonly consumed by Bedouins in order to investigate their impact on the gut microbiome of this population. We found that the gut microbiomes of westernized urban Saudis had significantly lower richness and biodiversity than the traditional Bedouin population. The gut microbiomes of baboons were more similar to that of Bedouins compared to urban Saudis, probably due the dietary overlap between baboons and Bedouins. Moreover, we found clusters that were compositionally similar to clusters identified in humans and baboons, characterized by differences in Acinetobacter, Turicibacter and Collinsella. The fermented food presented significantly more bacteria genera common to the gut microbiome of Bedouins compared to urban Saudis. These results support the hypothesis that dietary habits influence the composition of the gut microbiome.
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http://dx.doi.org/10.1038/srep32191DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5006041PMC
August 2016

Gut microbiota associated with HIV infection is significantly enriched in bacteria tolerant to oxygen.

BMJ Open Gastroenterol 2016 28;3(1):e000080. Epub 2016 Jul 28.

Faculté de Médecine, URMITE, UMR CNRS 6236-IRD 198, Aix-Marseille Université, Marseille, France; Pôle des Maladies Infectieuses et Tropicales Clinique et Biologique, Fédération de Bactériologie-Hygiène-Virologie, University Hospital Centre Timone, Institut Hospitalo-Universitaire (IHU) Méditerranée Infection, Assistance Publique-Hôpitaux de Marseille, Marseille, France; Special Infectious Agents Unit, King Fahd Medical Research Center, King Abdulaziz University, Jeddah, Saudi Arabia.

Objectives: Gut microbiota modifications occurring during HIV infection have recently been associated with inflammation and microbial translocation. However, discrepancies between studies justified a comprehensive analysis performed on a large sample size.

Design And Methods: In a case-control study, next-generation sequencing of the 16S rRNA gene was applied to the faecal microbiota of 31 HIV-infected patients, of whom 18 were treated with antiretroviral treatment (ART), compared with 27 healthy controls. 21 sera samples from HIV-infected patients and 7 sera samples from control participants were used to test the presence of 25 markers of inflammation and/or immune activation.

Results: Diversity was significantly reduced in HIV individuals when compared with controls and was not restored in the ART group. The relative abundance of several members of Ruminococcaceae such as Faecalibacterium prausnitzii was critically less abundant in the HIV-infected group and inversely correlated with inflammation/immune activation markers. Members of Enterobacteriaceae and Enterococcaceae were found to be enriched and positively correlated with these markers. There were significantly more aerotolerant species enriched in HIV samples (42/52 species, 80.8%) when compared with the control group (14/87 species, 16.1%; χ(2) test, p<10(-5), conditional maximum-likelihood estimate (CMLE) OR=21.9).

Conclusions: Imbalance between aerobic and anaerobic flora observed in HIV faecal microbiota could be a consequence of the gut impairment classically observed in HIV infection via the production of oxygen. Overgrowth of proinflammatory aerobic species during HIV infection raises the question of antioxidant supplementation, such as vitamin C, E or N-acetylcysteine.
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http://dx.doi.org/10.1136/bmjgast-2016-000080DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4985784PMC
August 2016

Glycans affect DNA extraction and induce substantial differences in gut metagenomic studies.

Sci Rep 2016 05 18;6:26276. Epub 2016 May 18.

URMITE, UM63, CNRS 7278, IRD 198, Inserm 1095, Aix Marseille Université,13005 Marseille, France.

Exopolysaccharides produced by bacterial species and present in feces are extremely inhibitory to DNA restriction and can cause discrepancies in metagenomic studies. We determined the effects of different DNA extraction methods on the apparent composition of the gut microbiota using Illumina MiSeq deep sequencing technology. DNA was extracted from the stool from an obese female using 10 different methods and the choice of DNA extraction method affected the proportional abundance at the phylum level, species richness (Chao index, 227 to 2,714) and diversity (non parametric Shannon, 1.37 to 4.4). Moreover DNA was extracted from stools obtained from 83 different individuals by the fastest extraction assay and by an extraction assay that degradated exopolysaccharides. The fastest extraction method was able to detect 68% to 100% genera and 42% to 95% species whereas the glycan degradation extraction method was able to detect 56% to 93% genera and 25% to 87% species. To allow a good liberation of DNA from exopolysaccharides commonly presented in stools, we recommend the mechanical lysis of stools plus glycan degradation, used here for the first time. Caution must be taken in the interpretation of current metagenomic studies, as the efficiency of DNA extraction varies widely among stool samples.
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http://dx.doi.org/10.1038/srep26276DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4870698PMC
May 2016

Increased Gut Redox and Depletion of Anaerobic and Methanogenic Prokaryotes in Severe Acute Malnutrition.

Sci Rep 2016 05 17;6:26051. Epub 2016 May 17.

Aix-Marseille Université, Unité de Recherche sur les Maladies Infectieuses et Tropicales Emergentes, UM63, CNRS 7278, IRD 198, INSERM 1095, Marseille, France.

Severe acute malnutrition (SAM) is associated with inadequate diet, low levels of plasma antioxidants and gut microbiota alterations. The link between gut redox and microbial alterations, however, remains unexplored. By sequencing the gut microbiomes of 79 children of varying nutritional status from three centers in Senegal and Niger, we found a dramatic depletion of obligate anaerobes in malnutrition. This was confirmed in an individual patient data meta-analysis including 107 cases and 77 controls from 5 different African and Asian countries. Specifically, several species of the Bacteroidaceae, Eubacteriaceae, Lachnospiraceae and Ruminococceae families were consistently depleted while Enterococcus faecalis, Escherichia coli and Staphylococcus aureus were consistently enriched. Further analyses on our samples revealed increased fecal redox potential, decreased total bacterial number and dramatic Methanobrevibacter smithii depletion. Indeed, M. smithii was detected in more than half of the controls but in none of the cases. No causality was demonstrated but, based on our results, we propose a unifying theory linking microbiota specificity, lacking anaerobes and archaea, to low antioxidant nutrients, and lower food conversion.
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http://dx.doi.org/10.1038/srep26051DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4869025PMC
May 2016

A Metagenomic Investigation of the Duodenal Microbiota Reveals Links with Obesity.

PLoS One 2015 10;10(9):e0137784. Epub 2015 Sep 10.

URMITE CNRS-IRD 198 UMR 6236, Aix Marseille Université, Faculté de Médecine, 27 Bd Jean Moulin, 13385, Marseille, France.

Background: Few studies have tested the small intestine microbiota in humans, where most nutrient digestion and absorption occur. Here, our objective was to examine the duodenal microbiota between obese and normal volunteers using metagenomic techniques.

Methodology/principal Findings: We tested duodenal samples from five obese and five normal volunteers using 16S rDNA V6 pyrosequencing and Illumina MiSeq deep sequencing. The predominant phyla of the duodenal microbiota were Firmicutes and Actinobacteria, whereas Bacteroidetes were absent. Obese individuals had a significant increase in anaerobic genera (p < 0.001) and a higher abundance of genes encoding Acyl-CoA dehydrogenase (p = 0.0018) compared to the control group. Obese individuals also had a reduced abundance of genes encoding sucrose phosphorylase (p = 0.015) and 1,4-alpha-glucan branching enzyme (p = 0.05). Normal weight people had significantly increased FabK (p = 0.027), and the glycerophospholipid metabolism pathway revealed the presence of phospholipase A1 only in the control group (p = 0.05).

Conclusions/significance: The duodenal microbiota of obese individuals exhibit alterations in the fatty acid and sucrose breakdown pathways, probably induced by diet imbalance.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0137784PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4565581PMC
May 2016

Exploration of Deinococcus-Thermus molecular diversity by novel group-specific PCR primers.

Microbiologyopen 2013 Oct 29;2(5):862-72. Epub 2013 Aug 29.

CEA, DSV, IBEB, SBVME, LIPM, F-13108, Saint-Paul-lez-Durance, France; CNRS, UMR 7265, F-13108, Saint-Paul-lez-Durance, France; Université d'Aix-Marseille, F-13108, Saint-Paul-lez-Durance, France; IRSN, PRP-ENV, SERIS, L2BT, F-13115, Saint Paul-lez-Durance, France.

The deeply branching Deinococcus-Thermus lineage is recognized as one of the most extremophilic phylum of bacteria. In previous studies, the presence of Deinococcus-related bacteria in the hot arid Tunisian desert of Tataouine was demonstrated through combined molecular and culture-based approaches. Similarly, Thermus-related bacteria have been detected in Tunisian geothermal springs. The present work was conducted to explore the molecular diversity within the Deinococcus-Thermus phylum in these extreme environments. A set of specific primers was designed in silico on the basis of 16S rRNA gene sequences, validated for the specific detection of reference strains, and used for the polymerase chain reaction (PCR) amplification of metagenomic DNA retrieved from the Tataouine desert sand and Tunisian hot spring water samples. These analyses have revealed the presence of previously undescribed Deinococcus-Thermus bacterial sequences within these extreme environments. The primers designed in this study thus represent a powerful tool for the rapid detection of Deinococcus-Thermus in environmental samples and could also be applicable to clarify the biogeography of the Deinococcus-Thermus phylum.
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http://dx.doi.org/10.1002/mbo3.119DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3831646PMC
October 2013

Microbacterium lemovicicum sp. nov., a bacterium isolated from a natural uranium-rich soil.

Int J Syst Evol Microbiol 2013 Jul 21;63(Pt 7):2600-2606. Epub 2012 Dec 21.

Université d'Aix-Marseille, 13108 Saint-Paul-lez-Durance, France.

An actinobacterial strain, designated ViU22(T), was isolated from a natural uranium-rich soil and was studied using a polyphasic approach. Cells formed orange-pigmented colonies, were rod-shaped, Gram-positive (non-staining method), non-motile and non-spore-forming. This organism grew in 0-4.5 % (w/v) NaCl and at 15-37 °C, with optimal growth occurring in 0.5 % (w/v) NaCl and at 30 °C. Comparative 16S rRNA gene sequence analysis revealed that the strain ViU22(T) belonged to the genus Microbacterium. It exhibited highest 16S rRNA gene sequence similarity with the type strains of Microbacterium testaceum (98.14 %) and Microbacterium binotii (98.02 %). The DNA-DNA relatedness of strains ViU22(T) with the most closely related type strains Microbacterium testaceum and Microbacterium binotii DSM 19164(T) was 20.10 % (± 0.70) and 28.05 % (± 0.35), respectively. Strain ViU22(T) possessed a type B2β peptidoglycan with partial substitution of glutamic acid by 3-hydroxy glutamic acid. The major menaquinones were MK-11 and MK-12. Major polar lipids detected in the strain ViU22(T) were diphosphatidylglycerol, phosphatidylglycerol, an unknown phospholipid and unknown glycolipids. The predominant fatty acids were anteiso-C15 : 0, anteiso-C17 : 0 and iso-C16 : 0, a pattern reported for other Microbacterium species. The major cell-wall sugars were galactose, xylose and mannose and the DNA G+C content was 71 mol%. Together, the DNA-DNA hybridization results and the differentiating phenotypic characteristics, showed that strain ViU22(T) should be classified as the type strain of a novel species within the genus Microbacterium, for which the name Microbacterium lemovicicum sp. nov. is proposed. The type strain is ViU22(T) ( = ATCC BAA-2396(T) = CCUG 62198(T) = DSM 25044(T)).
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http://dx.doi.org/10.1099/ijs.0.048454-0DOI Listing
July 2013

The Protist Ribosomal Reference database (PR2): a catalog of unicellular eukaryote small sub-unit rRNA sequences with curated taxonomy.

Nucleic Acids Res 2013 Jan 27;41(Database issue):D597-604. Epub 2012 Nov 27.

CNRS, UMR 7144, Adaptation et Diversité en Milieu Marin, 29682 Roscoff, France.

The interrogation of genetic markers in environmental meta-barcoding studies is currently seriously hindered by the lack of taxonomically curated reference data sets for the targeted genes. The Protist Ribosomal Reference database (PR(2), http://ssu-rrna.org/) provides a unique access to eukaryotic small sub-unit (SSU) ribosomal RNA and DNA sequences, with curated taxonomy. The database mainly consists of nuclear-encoded protistan sequences. However, metazoans, land plants, macrosporic fungi and eukaryotic organelles (mitochondrion, plastid and others) are also included because they are useful for the analysis of high-troughput sequencing data sets. Introns and putative chimeric sequences have been also carefully checked. Taxonomic assignation of sequences consists of eight unique taxonomic fields. In total, 136 866 sequences are nuclear encoded, 45 708 (36 501 mitochondrial and 9657 chloroplastic) are from organelles, the remaining being putative chimeric sequences. The website allows the users to download sequences from the entire and partial databases (including representative sequences after clustering at a given level of similarity). Different web tools also allow searches by sequence similarity. The presence of both rRNA and rDNA sequences, taking into account introns (crucial for eukaryotic sequences), a normalized eight terms ranked-taxonomy and updates of new GenBank releases were made possible by a long-term collaboration between experts in taxonomy and computer scientists.
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http://dx.doi.org/10.1093/nar/gks1160DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3531120PMC
January 2013

Significant and persistent impact of timber harvesting on soil microbial communities in Northern coniferous forests.

ISME J 2012 Dec 2;6(12):2199-218. Epub 2012 Aug 2.

Department of Microbiology and Immunology, Life Sciences Institute, University of British Columbia, Vancouver, British Columbia V6T 1Z3, Canada.

Forest ecosystems have integral roles in climate stability, biodiversity and economic development. Soil stewardship is essential for sustainable forest management. Organic matter (OM) removal and soil compaction are key disturbances associated with forest harvesting, but their impacts on forest ecosystems are not well understood. Because microbiological processes regulate soil ecology and biogeochemistry, microbial community structure might serve as indicator of forest ecosystem status, revealing changes in nutrient and energy flow patterns before they have irreversible effects on long-term soil productivity. We applied massively parallel pyrosequencing of over 4.6 million ribosomal marker sequences to assess the impact of OM removal and soil compaction on bacterial and fungal communities in a field experiment replicated at six forest sites in British Columbia, Canada. More than a decade after harvesting, diversity and structure of soil bacterial and fungal communities remained significantly altered by harvesting disturbances, with individual taxonomic groups responding differentially to varied levels of the disturbances. Plant symbionts, like ectomycorrhizal fungi, and saprobic taxa, such as ascomycetes and actinomycetes, were among the most sensitive to harvesting disturbances. Given their significant ecological roles in forest development, the fate of these taxa might be critical for sustainability of forest ecosystems. Although abundant bacterial populations were ubiquitous, abundant fungal populations often revealed a patchy distribution, consistent with their higher sensitivity to the examined soil disturbances. These results establish a comprehensive inventory of bacterial and fungal community composition in northern coniferous forests and demonstrate the long-term response of their structure to key disturbances associated with forest harvesting.
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http://dx.doi.org/10.1038/ismej.2012.84DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3504969PMC
December 2012

Midgut microbiota of the malaria mosquito vector Anopheles gambiae and interactions with Plasmodium falciparum infection.

PLoS Pathog 2012 31;8(5):e1002742. Epub 2012 May 31.

UMR MIVEGEC (IRD 224- CNRS 5290- UM1- UM2), Montpellier, France.

The susceptibility of Anopheles mosquitoes to Plasmodium infections relies on complex interactions between the insect vector and the malaria parasite. A number of studies have shown that the mosquito innate immune responses play an important role in controlling the malaria infection and that the strength of parasite clearance is under genetic control, but little is known about the influence of environmental factors on the transmission success. We present here evidence that the composition of the vector gut microbiota is one of the major components that determine the outcome of mosquito infections. A. gambiae mosquitoes collected in natural breeding sites from Cameroon were experimentally challenged with a wild P. falciparum isolate, and their gut bacterial content was submitted for pyrosequencing analysis. The meta-taxogenomic approach revealed a broader richness of the midgut bacterial flora than previously described. Unexpectedly, the majority of bacterial species were found in only a small proportion of mosquitoes, and only 20 genera were shared by 80% of individuals. We show that observed differences in gut bacterial flora of adult mosquitoes is a result of breeding in distinct sites, suggesting that the native aquatic source where larvae were grown determines the composition of the midgut microbiota. Importantly, the abundance of Enterobacteriaceae in the mosquito midgut correlates significantly with the Plasmodium infection status. This striking relationship highlights the role of natural gut environment in parasite transmission. Deciphering microbe-pathogen interactions offers new perspectives to control disease transmission.
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http://dx.doi.org/10.1371/journal.ppat.1002742DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3364955PMC
October 2012

Patho-Genes.org: a website dedicated to gene sequences of potential bioterror bacteria and PCR primers used to amplify them.

Microb Biotechnol 2012 Sep 10;5(5):594-8. Epub 2012 Jun 10.

CNRS UMR 7138 Systématique Adaptation et Evolution, Université de Nice-Sophia Antipolis, Parc Valrose BP71, F06108, Nice cedex, 02, France.

Pathogenic agents can be very hard to detect, and usually they do not cause illness for several hours or days. To improve the speed and the accuracy of detection tests and satisfy the needs of early diagnosis, molecular biology methods such as PCR are now used. However, selecting a proper target gene and designing good primers is often not easy. We present a dedicated website, http://patho-genes.org, where we provide every sequence, functional annotation, published primer and relevant article for every annotated gene of major pathogenic bacterial species listed as key agents to be used for a bioterrorism attack. Each published primer was analysed to determine its melting temperature, its specificity and its coverage (i.e. its sensitivity against every allele of its target gene). Data generated have been organized in the form of data sheet for each gene, which are available through multiple browser panels and query systems.
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http://dx.doi.org/10.1111/j.1751-7915.2012.00353.xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3815871PMC
September 2012

Molecular biomass and MetaTaxogenomic assessment of soil microbial communities as influenced by soil DNA extraction procedure.

Microb Biotechnol 2012 Jan 12;5(1):135-41. Epub 2011 Oct 12.

INRA-Université de Bourgogne, UMR Microbiologie du Sol et de l'Environnement, CMSE, 17, rue Sully, B.V. 86510, 21065 Dijon Cedex, France.

Three soil DNA extraction procedures (homemade protocols and commercial kit) varying in their practicability were applied to contrasting soils to evaluate their efficiency in recovering: (i) soil DNA and (ii) bacterial diversity estimated by 16S rDNA pyrosequencing. Significant differences in DNA yield were systematically observed between tested procedures. For certain soils, 10 times more DNA was recovered with one protocol than with the others. About 15,000 sequences of 16S rDNA were obtained for each sample which were clustered to draw rarefaction curves. These curves, as well as the PCA ordination of community composition based on OTU clustering, did not reveal any significant difference between procedures. Nevertheless, significant differences between procedures were highlighted by the taxonomic identification of sequences obtained at the phylum to genus levels. Depending on the soil, differences in the number of genera detected ranged from 1% to 26% between the most and least efficient procedures, mainly due to a poorer capacity to recover populations belonging to Actinobacteria, Firmicutes or Crenarchaeota. This study enabled us to rank the relative efficiencies of protocols for their recovery of soil molecular microbial biomass and bacterial diversity and to help choosing an appropriate soil DNA extraction procedure adapted to novel sequencing technologies.
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http://dx.doi.org/10.1111/j.1751-7915.2011.00307.xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3815280PMC
January 2012

Ultra-deep sequencing of foraminiferal microbarcodes unveils hidden richness of early monothalamous lineages in deep-sea sediments.

Proc Natl Acad Sci U S A 2011 Aug 25;108(32):13177-82. Epub 2011 Jul 25.

Department of Genetics and Evolution, University of Geneva, CH-1211 Geneva 4, Switzerland.

Deep-sea floors represent one of the largest and most complex ecosystems on Earth but remain essentially unexplored. The vastness and remoteness of this ecosystem make deep-sea sampling difficult, hampering traditional taxonomic observations and diversity assessment. This problem is particularly true in the case of the deep-sea meiofauna, which largely comprises small-sized, fragile, and difficult-to-identify metazoans and protists. Here, we introduce an ultra-deep sequencing-based metagenetic approach to examine the richness of benthic foraminifera, a principal component of deep-sea meiofauna. We used Illumina sequencing technology to assess foraminiferal richness in 31 unsieved deep-sea sediment samples from five distinct oceanic regions. We sequenced an extremely short fragment (36 bases) of the small subunit ribosomal DNA hypervariable region 37f, which has been shown to accurately distinguish foraminiferal species. In total, we obtained 495,978 unique sequences that were grouped into 1,643 operational taxonomic units, of which about half (841) could be reliably assigned to foraminifera. The vast majority of the operational taxonomic units (nearly 90%) were either assigned to early (ancient) lineages of soft-walled, single-chambered (monothalamous) foraminifera or remained undetermined and yet possibly belong to unknown early lineages. Contrasting with the classical view of multichambered taxa dominating foraminiferal assemblages, our work reflects an unexpected diversity of monothalamous lineages that are as yet unknown using conventional micropaleontological observations. Although we can only speculate about their morphology, the immense richness of deep-sea phylotypes revealed by this study suggests that ultra-deep sequencing can improve understanding of deep-sea benthic diversity considered until now as unknowable based on a traditional taxonomic approach.
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http://dx.doi.org/10.1073/pnas.1018426108DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3156150PMC
August 2011

Eukaryotic richness in the abyss: insights from pyrotag sequencing.

PLoS One 2011 Apr 4;6(4):e18169. Epub 2011 Apr 4.

Department of Genetics and Evolution, University of Geneva, Geneva, Switzerland.

Background: The deep sea floor is considered one of the most diverse ecosystems on Earth. Recent environmental DNA surveys based on clone libraries of rRNA genes confirm this observation and reveal a high diversity of eukaryotes present in deep-sea sediment samples. However, environmental clone-library surveys yield only a modest number of sequences with which to evaluate the diversity of abyssal eukaryotes.

Methodology/principal Findings: Here, we examined the richness of eukaryotic DNA in deep Arctic and Southern Ocean samples using massively parallel sequencing of the 18S ribosomal RNA (rRNA) V9 hypervariable region. In very small volumes of sediments, ranging from 0.35 to 0.7 g, we recovered up to 7,499 unique sequences per sample. By clustering sequences having up to 3 differences, we observed from 942 to 1756 Operational Taxonomic Units (OTUs) per sample. Taxonomic analyses of these OTUs showed that DNA of all major groups of eukaryotes is represented at the deep-sea floor. The dinoflagellates, cercozoans, ciliates, and euglenozoans predominate, contributing to 17%, 16%, 10%, and 8% of all assigned OTUs, respectively. Interestingly, many sequences represent photosynthetic taxa or are similar to those reported from the environmental surveys of surface waters. Moreover, each sample contained from 31 to 71 different metazoan OTUs despite the small sample volume collected. This indicates that a significant faction of the eukaryotic DNA sequences likely do not belong to living organisms, but represent either free, extracellular DNA or remains and resting stages of planktonic species.

Conclusions/significance: In view of our study, the deep-sea floor appears as a global DNA repository, which preserves genetic information about organisms living in the sediment, as well as in the water column above it. This information can be used for future monitoring of past and present environmental changes.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0018169PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3070721PMC
April 2011
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