Showing papers by "Jean-Christophe Lagier published in 2019"

Repertoire of human breast and milk microbiota: a systematic review.

Abstract: Breastfeeding is a major determinant of human health. Breast milk is not sterile and ecological large-scale sequencing methods have revealed an unsuspected microbial diversity that plays an important role. However, microbiological analysis at the species level has been neglected while it is a prerequisite before understanding which microbe is associated with symbiosis or dysbiosis, and health or disease. We review the currently known bacterial repertoire from the human breast and milk microbiota using a semiautomated strategy. Total 242 articles from 38 countries, 11,124 women and 15,489 samples were included. Total 820 species were identified mainly composed of Proteobacteria and Firmicutes. We report variations according to the analytical method (culture or molecular method), the anatomical site (breast, colostrum or milk) and the infectious status (healthy control, mastitis, breast abscess, neonatal infection). In addition, we compared it with the other human repertoires. Finally, we discuss its putative origin and role in health and disease.

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

Abstract: 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.

Human Bacterial Repertoire of the Urinary Tract: a Potential Paradigm Shift.

Abstract: The aim of this article is to review the human repertoire of bacteria in urine already described by culture and metagenomic techniques and published in the literature. Our study led us to compare this repertoire with other available human repertoires. We followed automatic and manual bibliographical methods and found 562 bacterial species reported in the literature as part of the human urinary microbiota. Of the 562 species, 322 were described only by culture, 101 by both culture and metagenomics, and 139 only by metagenomics. A total of 352 species (62.6%) have been associated with at least one case report of human infection, of which 225 (40.0%) have been described as causative agents of urinary tract infection. The urinary tract bacterial repertoire contains 21.4% of the known prokaryotic diversity associated with human beings (464 species in common), and it shares 23.6% species with the human gut microbiota (350 species in common, 62.3% of the urine species). The urinary repertoire shares a significant difference in aerointolerant species compared with those of the gut microbiota (100/562 [17.8%] and 505/1,484 [34.0%], respectively; P < 0.001; odds ratio [OR] = 9.0 [7.0 to 11.4]). Studies using high-throughput sequencing show a higher proportion of aerointolerant bacteria in urine (74/240 [30.8%]) than studies using culture techniques (40/423 [9.5%]). Most pathogenic bacteria are part of the commensal human urinary tract bacteria, and their pathogenicity may occur following any imbalance of this microbiota. The restoration of urinary tract health can occur following a fecal transplantation. The potential gut origin of the human bacterial microbiota has to be explored.

Among Live and Dead Bacteria, the Optimization of Sample Collection and Processing Remains Essential in Recovering Gut Microbiota Components.

Abstract: The human gut microbiota plays an important role in human health. Accessing and culturing the gut microbes remains critical in microbiology. The culturomics approach, combined with sequencing strategies, provides new insights in the study of gut microbiota components. However, we are still far from having described all the microbes. Many factors are involved in recovering as much bacteria as possible. One of the most important factors is sample preparation and conservation. The aim of the present work was to evaluate three different types of stool sample conditioning by mainly studying the effect of atmospheric oxygen on bacterial viability and culturability. Quantitative analysis of fecal samples from eight different healthy individuals was carried out using a culture-independent method (flow cytometry) and a culture-dependent method (plating technique). We found that the cultured bacteria reached a 50% yield when the samples were exposed to oxygen for 120 min without any protectant medium, while the percentage of culturability increased to 67% in the presence of antioxidants. More importantly, when samples were exposed to oxygen for less than 2 min, combined with the work under the anaerobic chamber, no discordance was found between the two counting techniques and the culturability increased to 87%. Our study confirmed the importance of sample conditioning to preserve the bacterial viability in samples, especially for oxygen-sensitive intestinal bacteria.

Extensive culturomics of 8 healthy samples enhances metagenomics efficiency.

Abstract: Molecular approaches have long led to the assumption that the human gut microbiota is dominated by uncultivable bacteria. The recent advent of large-scale culturing methods, and in particular that of culturomics have demonstrated that these prokaryotes can in fact be cultured. This is increasing in a dramatic manner the repertoire of commensal microbes inhabiting the human gut. Following eight years of culturomics approach applied on more than 900 samples, we propose herein a remake of the pioneering study applying a dual approach including culturomics and metagenomics on a cohort of 8 healthy specimen. Here we show that culturomics enable a 20% higher richness when compared to molecular approaches by culturing 1 archaeal species and 494 bacterial species of which 19 were new taxa. Species discovered as a part of previous culturomics studies represent 30% of the cultivated isolates, while sequences derived from these new taxa enabled to increase by 22% the bacterial richness retrieved by metagenomics. Overall, 67% of the total reads generated were covered by cultured isolates, significantly reducing the hidden content of sequencing methods compared to the pioneering study. By redefining culture conditions to recover microbes previously considered fastidious, there are greater opportunities than ever to eradicate metagenomics dark matter.

Successful Fecal Microbiota Transplantation in a Patient Suffering From Irritable Bowel Syndrome and Recurrent Urinary Tract Infections.

Abstract: Background Irritable bowel syndrome (IBS) is a chronic and debilitating functional gastrointestinal disorder affecting 9%-23% of the population across the world. The relative efficacy of fecal microbiota transplantation (FMT) on IBS symptoms was demonstrated in a double-blind, randomized study. Methods We describe the case of a 73-year-old woman suffering from IBS (abdominal pain, bloating, and abundant and disabling diarrhea, with 10-15 stools a day) and repetitive urinary tract infection (UTI; 5 episodes in 6 months, including 3 the last 2 months) for several years, generating an impaired quality of life. She received an FMT with 400 mL of fecal infusion from a healthy donor via a nasogastric tube after bowel lavage. Her digestive microbiota was analyzed using culturomic and metagenomic targeting 16S rRNA sequencing methods. Results Eight months after transplantation, we observed a significant reduction in frequency and improvement in stool consistency (3-4 molded stools a day against 10-15 before the transplant) and no recurrence of urinary infection (as previously reported). Using culturomics, we found 12 bacteria present in the fecal infusion and post-transplant stool; these were absent pretransplant. Three of them (Intestinimonas massiliensis, Oscillibacter massiliensis, and Provencibacter massiliensis) were previously discovered and cultivated in our laboratory using culturomics. Using metagenomics, we also observed 12 bacteria, different from those observed during culture, that could have been transferred to the patient by FMT. Conclusions In this case report, IBS symptoms and UTI frequency decreased after FMT UTI. Further studies involving more patients would be relevant to confirm this work and develop bacteriotherapy.

Passive Filtration, Rapid Scanning Electron Microscopy, and Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry for Treponema Culture and Identification from the Oral Cavity

Abstract: We present here a new passive-filtration-based culture device combined with rapid identification with a new electron microscope (Hitachi TM4000) for the detection and culture of Treponema species from the human oral cavity. Of the 44 oral samples cultivated, 15 (34%) were found to be positive for Treponema using electron microscopy and were also culture positive. All were subcultured on agar plates; based on genome sequencing and analyses, 10 were strains of Treponema pectinovorum and 5 were strains of Treponema denticola The 29 samples that were negative for Treponema remained culture negative. In addition, 14 Treponema species ordered from the DSMZ collection were cultured in the T-Raoult culture medium optimized here. Finally, matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) was used and 30 novel spectra were added to the MALDI-TOF MS database. We have successfully developed a new and effective method for treponemal detection, culture, and identification.

Feasibility, Acceptability, and Accuracy of Vaginal Self-Sampling for Screening Human Papillomavirus Types in Women from Rural Areas in Senegal.

Abstract: Vaginal self-sampling and human papillomavirus (HPV) DNA testing can be useful tools for women with limited access to health care living in sub-Saharan Africa. To assess the feasibility and acceptability of vaginal self-sampling and high-risk HPV prevalence in two villages of central Senegal, women were asked to self-sample vaginal swabs for HPV detection in May, 2016. Vaginal swabs were collected from 133 women and were tested for HPV genotyping. The acceptability rate of vaginal self-sampling was 98.5%, and 99.2% of the women (133/134) used the device correctly. The quality of self-sampling was satisfactory in 100% of the samples; 10.5% of the samples were positive for HPV, including 6% with high-risk HPV types and 4% with low-risk HPV types. This preliminary study indicates that vaginal self-sampling is a valuable strategy for high-risk HPV detection and cervical cancer screening in a population of women not attending gynecologic screening in rural areas of Senegal.

Noncontiguous finished genome sequence and description of Intestinimonas massiliensis sp. nov strain GD2T , the second Intestinimonas species cultured from the human gut.

Abstract: Intestinimonas massiliensis sp. nov strain GD2T is a new species of the genus Intestinimonas (the second, following Intestinimonas butyriciproducens gen. nov., sp. nov). First isolated from the gut microbiota of a healthy subject of French origin using a culturomics approach combined with taxono-genomics, it is strictly anaerobic, nonspore-forming, rod-shaped, with catalase- and oxidase-negative reactions. Its growth was observed after preincubation in an anaerobic blood culture enriched with sheep blood (5%) and rumen fluid (5%), incubated at 37°C. Its phenotypic and genotypic descriptions are presented in this paper with a full annotation of its genome sequence. This genome consists of 3,104,261 bp in length and contains 3,074 predicted genes, including 3,012 protein-coding genes and 62 RNA-coding genes. Strain GD2T significantly produces butyrate and is frequently found among available 16S rRNA gene amplicon datasets, which leads consideration of Intestinimonas massiliensis as an important human gut commensal.

Arcanobacterium ihumii sp. nov., Varibaculum vaginae sp. nov. and Tessaracoccus timonensis sp. nov., isolated from vaginal swabs from healthy Senegalese women.

Abstract: Culturomics studies the microbial variety of the human microbiome by combining diversified culture conditions, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and 16S rRNA gene identification. This study identifies three putative new bacterial species: Arcanobacterium ihumii sp. nov. strain Marseille-P5647T, Varibaculum vaginae sp. nov. strain Marseille-P5644T and Tessaracoccus timonensis sp. nov. strain Marseille-P5995T, which we describe according to the concept of taxonogenomics.

Description and genomic characterization of Massiliimalia massiliensis gen. nov., sp. nov., and Massiliimalia timonensis gen. nov., sp. nov., two new members of the family Ruminococcaceae isolated from the human gut.

Abstract: Using the culturomics approach, we isolated two strains, Marseille-P2963 and Marseille-P3753, from the intestinal microbiota of a 19-year-old healthy Saudi Arabian Bedouin male and from a 32-year-old healthy Senegalese male faecal transplant donor. Here, we studied their phenotypic, phylogenetic and genomic characteristics. Both strains were phylogenetically related, but different from Ruminococcus species. Bacterial cells were anaerobic, rod-shaped, non-spore-forming and not motile, with neither catalase nor oxidase activities. Their growth temperatures ranged from 28 to 45 °C, with an optimal growth at 37 °C. The genomes are 2,842,720 bp- and 2,707,061 bp-long respectively. The G + C contents are 47.18% and 46.90%, respectively. Based on these characteristics, we propose the creation of a new genus within the family Ruminococcaceae named Massiliimalia gen. nov., that contains the new species Massiliimalia massiliensis gen. nov., sp. nov., and Massiliimalia timonensis gen. nov., sp. nov. Strains Marseille-P2963T (= CSUR P2963 = DSM 106837) and Marseille-P3753T (= CSUR P3753 = CCUG 71632) are their type strains, respectively.

Limbic encephalitis as a relapse of Whipple’s disease with digestive involvement and spondylodiscitis

Abstract: Many clinical manifestations can be related to Tropheryma whipplei infection. We report a Tropheryma whipplei limbic encephalitis developed as a relapse of classical Whipple’s disease. This case is to the best of our knowledge the first proof of the effective brain–blood barrier crossing of both doxycycline and hydroxychloroquine as demonstrated by direct concentration monitoring on brain biopsy.

Corynebacterium pacaense sp. nov., Alistipes megaguti sp. nov., Alistipes provencensis sp. nov., 3 new bacteria isolated from fresh human stool specimens.

Abstract: Here we describe the basic characteristics of Corynebacterium pacaense strain Marseille-P2417T (= CSUR P2417), Alistipes megaguti strain Marseille-P5997T (= CSUR P5997) and Alistipes provencensis strain Marseille-P2431T (= CSUR P2431 = DSM 102308). The phenotypic criteria, the 16S ribosomal RNA sequencing and MALDI-TOF MS spectra analysis were used to identify and characterize these new bacteria species, which were isolated from fresh human stool specimens.

Oceanobacillus timonensis sp. nov. and Oceanobacillus senegalensis sp. nov., two new moderately halophilic, Gram-stain positive bacteria isolated from stools sample of healthy young Senegalese

Abstract: Oceanobacillus timonensis Marseille-P3532T (CSUR P3532, CCUG 70981) and Oceanobacillus senegalensis Marseille-P3587T (CSUR P3587, CCUG 70613), are the type strains of O. timonensis sp. nov. and O. senegalensis sp. nov., respectively. They are moderately halophilic, aerobic, motile and Gram-stain positive bacteria. The strains P3532T and P3587T were isolated from stools with 3.8% and 2.1% sodium chloride (NaCl) of healthy 10 year old female and male 7-year-old children, respectively and living respectively at Dielmo and N’diop two villages in Senegal (West Africa). This study aimed to describe the genome and phenotypic characteristics of O. timonensis Marseille-P3532T and O. senegalensis Marseille-P3587T. The genomes are 4,485,335 bp long for O. timonensis and 4,300,331 bp for O. senegalensis with 38.78% and 36.92% G+C content, respectively. They contain 4306 and 3979 protein-coding and 87 and 273 RNAs genes, respectively.

Routine Culture-Resistant Mycobacterium tuberculosis Rescue and Shell-Vial Assay, France.

Abstract: We used shell-vial assay with a medium that buffered rifampin to isolate routine culture-resistant Mycobacterium tuberculosis bacteria from cerebrospinal fluid and rifampin-containing intervertebral disc and vertebral corpus of a patient in treatment for Pott's disease and disseminated tuberculosis. Whole-genome sequencing confirmed M. tuberculosis lineage 4 (Euro-American) strain.

Corynebacterium senegalense sp. nov. and Arthrobacter senegalensis sp. nov., two new Actinobacteria isolated from skin swab from the palm of hand.

Abstract: Corynebacterium senegalense strain Marseille-P4329T (= CSURP4329) and Arthrobacter senegalensis strain Marseille-P4329T (= CSURP4198) are new species first isolated from human skin. A culturomics approach and taxonogenomics methods were used for these new bacterial species.

Neoactinobaculum massilliense gen. nov., a new genesis and Pseudopropionibacterium massiliense sp. nov., a new bacterium isolated from the human oral microbiota.

Abstract: Neoactinobaculum massilliense gen. nov., strain Marseille-P6182T (= CSUR P6182) and Pseudopropionibacterium massiliense sp. nov., strain Marseille-P6184T (= CSUR P6184) are a new bacterial genus and new bacterial species belonging to the Actinobacteria phylum that have been isolated from the human oral microbiota.