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International audience; Using the culturomics approach, the previously unknown strain Marseille-P8953T, was isolated and classified within the Weizmannia genus. Strain Marseille-P8953T was isolated from the faeces of a healthy subject and consisted of Gram-stain positive, spore-forming, motile rod-shaped cells. A 99.2% similarity was observed between the 16S rRNA gene of strain Marseille-P8953T (accession number LR735539) and that of Weizmannia coagulans strain NBRC 12583T (accession number KX261624), its closest phylogenetic relative, while the genome of strain Marseille-P8953T (3.5 Mpb long, 46.5% GC content) shared the average nucleotide identity by Orthology and digital DNA-DNA Hybridisation values of 95 and 60.4%, respectively. Given the phylogenetic classification and phenotypic characteristics of strain Marseille-P8953T, we propose the creation of a new species within the Weizmannia genus named Weizmannia faecalis (= CSUR P8953T = CECT 9904 T).

Strain Marseille-P8396

A Gram-positive, aerobic, motile, endospore-forming, rod-shaped bacterium was isolated from a stool sample of a child with marasmus. The 16S rRNA gene showed that strain Marseille-P3601

Blood is precious tissue that is normally sterile. With the aim of diagnosing the cause of bacteremia, three bacterial strains were isolated from three different individuals. Strains Marseille-P7157T and Marseille-Q2854T are Gram-stain positive, non-spore-forming rod-shaped bacteria, while strain Marseille-P8049T is a Gram-stain negative, motile, non-spore-forming and rod-shaped bacterium. The major fatty acids found (>30%) were hexadecanoic acid for strain Marseille-P8049T and 12-methyl tetradecanoic acid for both strains Marseille-P7157T and Marseille-Q2854T. The 16S rRNA gene sequence analysis shows that strains Marseille-P8049 and Marseille-Q2854T have sequence similarity of 96.8%, 99.04%, and 98.3% with Acinetobacter ursingii strain LUH3792 (NR_025392.1), Gulosibacter faecalis strain B187 (NR_041812.1), and Schaalia canis strain CCUG 41706 (NR_025366.1), respectively. In addition, strains Marseille-Q2854T, Marseille-P8049T and Marseille-P7157T shared with their closely related species cited above the following DDH values: 19.5%, 24.4%, and 20.2%, respectively. Based on these phenotypic and genomic findings, we consider that strains Marseille-P8049T (= CSUR P8049 = CECT 30350), Marseille-P2854T ( = CSUR Q2854 = CECT 30120) and Marseille-P7157T ( = CSUR P7157 = CECT 30048) are new bacterial species, for which the names Acinetobacter ihumii sp. nov., Microbacterium ihumii sp. nov., and Gulosibacter massiliensis sp. nov., are proposed.

Objectives To compare the demographics, clinical characteristics and severity of patients infected with nine different SARS-CoV-2 variants, during three phases of the COVID-19 epidemic in Marseille. Methods A single centre retrospective cohort study was conducted in 1760 patients infected with SARS-CoV-2 of Nextstrain clades 20A, 20B, and 20C (first phase, February–May 2020), Pangolin lineages B.1.177 (we named Marseille-2) and B.1.160 (Marseille-4) variants (second phase, June–December 2020), and B.1.1.7 (alpha), B.1.351 (beta), P.1 (gamma) and A.27 (Marseille-501) variants (third phase, January 2021-today). Outcomes were the occurrence of clinical failures, including hospitalisation, transfer to the intensive-care unit, and death. Results During each phase, no major differences were observed with regards to age and gender distribution, the prevalence of chronic diseases, and clinical symptoms between variants circulating in a given phase. The B.1.177 and B.1.160 variants were associated with more severe outcomes. Infections occurring during the second phase were associated with a higher rate of death as compared to infections during the first and third phases. Patients in the second phase were more likely to be hospitalised than those in the third phase. Patients infected during the third phase were more frequently obese than others. Conclusion: A large cohort study is recommended to evaluate the transmissibility and to better characterise the clinical severity of emerging variants.

From January 18th to August 13th, 2021, 13,804 unvaccinated and 1,156 patients who had received at least one COVID-19 vaccine dose were tested qPCR-positive for SARS-CoV-2 in our center. Among vaccinated patients, 949, 205 and 2 had received a single, two or three vaccine doses, respectively. Most patients (80.3%) had received the Pfizer-BioNTech vaccine. The SARS-CoV-2 variants infecting vaccinated patients varied over time, reflecting those circulating in the Marseille area, with a predominance of the Marseille-4/20A.EU2 variant from weeks 3 to 6, of the Alpha/20I variant from weeks 7 to 25, and of the Delta/21A variant from week 26. SARS-CoV-2 infection was significantly more likely to occur in the first 13 days post-vaccine injection in those who received a single dose (48.9%) than two doses (27.4%, p< 10–3). Among 161 patients considered as fully vaccinated, i.e., >14 days after the completion of the vaccinal scheme (one dose for Johnson and Johnson and two doses for Pfizer/BioNTech, Moderna and Sputnik vaccines), 10 (6.2%) required hospitalization and four (2.5%) died. Risks of complications increased with age in a nonlinear pattern, with a first breakpoint at 54, 33, and 53 years for death, transfer to ICU, and hospitalization, respectively. Among patients infected by the Delta/21A or Alpha/20I variants, partial or complete vaccination exhibited a protective effect with a risk divided by 3.1 for mortality in patients ≥ 55 years, by 2.8 for ICU transfer in patients ≥ 34 years, and by 1.8 for hospitalization in patients ≥ 54 years. Compared to partial vaccination, complete vaccination provided an even stronger protective effect, confirming effectiveness to prevent severe forms of COVID-19.

Strain Marseille-P3519T isolated from the fecal flora of a 25-year-old healthy French woman was a Gram-positive anaerobic bacterium, non-motile and non-spore forming. The 16S rRNA gene sequence of Marseille-P3519 showed 97.73% of sequence similarity with Limosilactobacillus reuteri DSM 20016, the closest species, phylogenetically. Furthermore, the average nucleotide identity of strain Marseille-3519 with its closest related species was 75.8% that was very below the recommended threshold (>95–96%). Its genome had 2 237 367 bp with 45.42 mol% of G + C content. Major fatty acids were C16:0 (50.8%), C18:1n9 (18.0%), C18:2n6 (9.8%) and C19:1n9 (8.9%). It was catalase negative and fermented glycerol, glucose, fructose, D-maltose, lactose and mannose. These findings support that strain Marseille-P3519 ( = CSURP3519 = CECT 30110) is a new member of the genus Limosilactobacillus for which the name Limosilactobacillus caccae sp. nov., is proposed.

Using microbial culturomics, we were able to isolate strain Marseille-P3078 from a stool sample of a healthy 50-year-old Saudi Arabian woman. To this end, we used taxonogenomics that combines phenotypic, biochemical and genomic analyses, to describe this bacterium. Cells from strain Marseille-P3078 are anaerobic and Gram-negative rods that are motile and unable to sporulate. Its genome size is 3,377,914-bp-long with a 66.33 mol% G + C content. Based on its phenotypic and genomic features, including a 94.6% 16S rRNA similarity with Paraeggerthella hongkongensis strain JCM 14552, its closest phylogenetic neighbor withstanding in nomenclature, we propose that strain Marseille-P3078T (= CSUR P3078 = DSM 104007) is the representative strain of a new genus for which we propose the name Arabiibacter massiliensis gen. nov., sp. nov.

Oceanobacillus timonensis Marseille-P3532(T) (CSUR P3532, CCUG 70981) and Oceanobacillus senegalensis Marseille-P3587(T) (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 P3532(T) and P3587(T) 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-P3532(T) and O. senegalensis Marseille-P3587(T). The genomes are 4,485,335bp 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.

Background Severe acute malnutrition (SAM) is a major public health problem affecting children under the age of five in many low- and middle-income countries, and its resolution would contribute towards achieving the several sustainable development goals. The etiology of SAM is pluri-factorial, including delayed maturation of the gut microbiota, suboptimal feeding practices and dysfunctional breastfeeding. The recent serendipitous detection of Listeria monocytogenes in the breast milk of Malian women, in contrast to French women, suggests a possible association with SAM. Methodology/ Principal findings To investigate the possible association of L. monocytogenes carriage in breast milk and SAM, a case-control study was performed in Senegal, with subjects recruited from two areas. Using 16S amplicon sequencing, a culture independent method, 100% (152/152) of the mothers were positive for L. monocytogenes in their breast milk while qPCR analysis gave lower recovery rates. Interestingly, after enrichment in Fraser broth and seeding on PALCALM agar, all 10 isolated strains were isolated from the milk of 10 mothers who had SAM children which also had a significantly increased relative abundance of L. monocytogenes (0.34 (SD 0.35) vs 0.05 (SD 0.07) in controls, p, Author summary Severe acute malnutrition is a global public health issue which greatly impacts childhood mortality rates. Although still not fully understood, the multi-factorial pathology of severe acute malnutrition has been associated, among other factors, with sub-optimal feeding practices (including dysfunctional breastfeeding) and an altered gut microbiota. The serendipitous detection of Listeria monocytogenes in the breast milk of Malian women has raised the possibility of its involvement in the pathogenesis of severe acute malnutrition. To investigate this possibility, the presence of L. monocytogenes was assessed in a cohort of lactating Senegalese women, both mothers of healthy children as well as those of severely malnourished children using culture-dependent and independent methods. Our study confirms the previously unsuspected presence of L. monocytogenes in the breast milk of Senegalese women, which is increased in the milk of mothers of severely malnourished children. Moreover, breast milk samples from the mothers of healthy children more frequently induced a potent inhibition of L. monocytogenes than those from the mothers of severely malnourished children. An inhibition was also achieved using potential probiotics, Lactobacillus and Streptococcus species, isolated from breast milk. Our study reveals the previously unsuspected carriage of L. monocytogenes in the breast milk of West African women, which is associated with severe acute malnutrition. The inhibitory effect of human selected lactic acid bacterial species against L. monocytogenes might provide new therapeutic and inexpensive options to prevent and treat this neglected public health issue.

Phenotypic, phylogenetic and genomic studies were carried out on three unidentified Gram-stain positive, facultative anaerobic, and cocci-shaped bacteria isolated from the human oral cavity. The 16S rRNA gene of strains Marseille-P5794 T , Marseille-P6264 T and Marseille-P7376 T exhibited a sequence identity of 99,41%, 99.67% and 97.88%, respectively with Streptococcus cristatus, their closest phylogenetic relative with standing in nomenclature. Moreover, the rpoB gene sequence of strains Marseille-P5794 T and Marseille-P6264 T shared a similarity level with 96.1%, and 95.9% with Streptococcus cristatus whereas strain Marseille-P7376 T shared a 93.98% identity with Streptococcus sanguinis. Whole genome comparison of strains Marseille-P5794 T , Marseille-P6264 T and Marseille-P7376 T with their phylogenetic neighbours were under the threshold values set to define new species using digital DNA-DNA hybridization and Orthologous Average Nucleotide Identity. The taxonogenomics analysis thus allowed the classification of these strains as new species within the Streptococcus genus named Streptoccocus resistens sp. nov. Strain Marseille-P5794 T (=CSUR P5794 = CECT9902), Streptococcus buccae sp. nov. Strain Marseille-P6264 T (=CSUR P6264 = CECT9910) and Streptococcus mediterraneus sp. nov. Strain Marseille-P7376 (=CSUR P7376 = CECT30035).

Recent studies have used ethanol stool disinfection as a mean of promoting valuable species’ cultivation in bacteriotherapy trials for Clostridium difficile infections (CDI) treatment with a particular focus on sporulating bacteria. Moreover, the culturomic approach has considerably enriched the repertoire of cultivable organisms in the human gut in recent years. This study aimed to apply this culturomic approach on fecal donor samples treated with ethanol disinfection to evidence potential beneficial microbes that could be used in bacteriotherapy trials for the treatment of CDI. Thereby, a total of 254 bacterial species were identified, 9 of which were novel. Of these, 242 have never been included in clinical trials for the treatment of CDIs, representing potential new candidates for bacteriotherapy trials. While non-sporulating species were nevertheless more affected by the ethanol pretreatment than sporulating species, the ethanol disinfection technique did not specifically select bacteria able to sporulate, as suggested by previous studies. Furthermore, some bacteria previously considered as potential candidates for bacteriotherapy have been lost after ethanol treatment. This study, while enriching the bacterial repertoire of the human intestine, would nevertheless require determining the exact contribution of each of species composing the bacterial consortia intended to be administered for CDI treatment.

Background: Bacterial vaginosis is a very common vaginal disorder, with a gynaeco-obstetrical impact, but remains poorly understood. Current antibiotic treatment most often fails. Our objective was to exhaustively map the bacterial community present in bacterial vaginosis and normal flora, using not only a molecular approach but also a highly efficient culture approach, culturomics. Vaginal bacterial diversity was evaluated using both strategies for 24 Caucasian French women, including 7 cases of bacterial vaginosis, and 10 rural Senegalese women, including 5 cases of bacterial vaginosis. An additional 16 specimens (three cases of bacterial vaginosis and 13 normal flora) obtained during the follow-up visits of five French women with bacterial vaginosis were analysed using metagenomics.Results: The combination of culturomics and metagenomics reveals the richness and diversity of vaginal microbiota. Our data demonstrated that compared to healthy women, those with bacterial vaginosis present a shift in vaginal flora. The microbiota of the bacterial vaginosis group can be easily distinguished from that of healthy group by their increased bacterial diversity, abundance of Bacteroidales and Leptotrichiaeceae, and depletion of Proteobacteria and Lactobacillaceae species. In addition, a complex of 10 genera was associated with bacterial vaginosis: Gardnerella, Atopobium, Snethia, Aerococcus, Prevotella, Gemella, Facklamia, Porphyromonas, Mycoplasma, and Urinacoccus. Overall, the microbial population detected in bacterial vaginosis flora was largely similar to the bacterial communities found in gut microbiota. The two approaches showed a diverse vaginal microbiota that was largely non-overlapping, with only 146 common species. Furthermore, culturomics extends the repertoire of human-associated bacteria with the isolation of 27 new bacterial species.Conclusions: This study provides the most representative topology of the vaginal microbiota of normal flora and bacterial vaginosis and suggests the transplantation of gut microbiota into the vagina during bacterial vaginosis. Several sets of clinically relevant anaerobic bacteria and new species were also isolated from the human vagina, confirming that some previously “uncultivated” species may be cultivated using an appropriate strategy. Exhaustive characterisation of vaginal microbiota will enable a better understanding and management of bacterial vaginosis. Obtaining isolates will allow us to explore the bacterial interactions of the vaginal microbiota and will serve as a basis for the potential development of bacteriotherapy to prevent or treat vaginal dysbiosis.

The increasingly wide knowledge of the microbiota, as well as its continuous relation with several pathologies, favours the interest, discovery and classification of all the bacteria that are part of it. The isolation of strain P3519T is possible thanks to the application of new strategies such as culturomics and taxono-genomics. Strain Marseille-P3519T isolated from the fecal flora of a 25-year-old healthy French woman was a Gram-positive anaerobic bacterium, non-motile and non-spore-forming. The 16S rRNA gene sequence of Marseille-P3519 showed 98.57% of sequence similarity with Lactobacillus caviae MOZM2, the closest species, phylogenetically. Its genome had 2,237,367 bp with 45.42 mol% of G+C content. These findings support that strain Marseille-P3519 is a new member of the genus Lactobacillus for which the name Lactobacillus caccae sp. nov., is proposed.

Freeze-drying technology has been widely considered for decades as a suitable technique to preserve microorganisms. However, protective agents must be added prior to freeze drying to improve the survival and storage stability of the bacteria. The objective of our study was to evaluate the effect of a new protectant medium containing sucrose (10 %), trehalose (10 %), skimmed milk (10 %) and antioxidants on the viability of gut bacteria under different storage conditions. Two strains were tested, Escherichia coli and Akkermansia muciniphila, as examples of facultative aerobic and anaerobic bacteria, respectively. We studied the cell viability and bacterial morphology in 5 fecal samples in the presence and absence of this protectant medium using plating technique, flow cytometry and scanning electron microscopy. The results of bacterial viability assessed by plating method showed that the protectant medium yielded higher survival rates for both strains whatever the storage conditions (85–93 %) compared to normal saline solution (0.36–37.50 %). It also showed its effectiveness on fecal samples, where bacterial viability after freeze-drying was 89.47 ± 7.63 % and 84.01 ± 7.44 %, as evidenced by flow cytometry analysis and plating method. However unprotected samples showed the lowest cell viability at 19.01 ± 12.88 % and 13.23 ± 9.56 %, as measured by flow cytometry and plating method. In addition, bacterial size and shape were conserved in the protectant medium. In contrast, storage without protectant medium severely damaged bacterial morphology. In conclusion, our study is the first to use morphological features as well as culture-dependant and culture-independent tests to evaluate the effectiveness of a new protectant medium.

International audience; A Gram-negative and facultative anaerobic bacterium, designated strain SN4T, was isolated from the stool sample of an obese Amazonian patient. The new isolate was characterized by the taxonogenomics approach. The strain SN4T was beige-colored, circular and not haemolytic. Cells are rod shaped and motile with several flagella. Strain SN4T grows optimally at pH 7 and can survive in the presence of a saline concentration of up to 75 g/l NaCl. The 16S ribosomal RNA gene sequence analysis of the novel strain SN4T showed 95.28% similarity in nucleotide sequence with Gorillibacterium massiliense G5T, the phylogenetically closest neighbor and the type species of this genus. Anteiso-C15:0, iso-C15:0 and C16:0 were found as the major components in the cellular fatty acid analysis of this isolate. The genomic draft of strain SN4T is 5,263,742 bp long with 53.33% of G+C content. The differences in physiological, biochemical characteristics and phylogenetic and genomic data make it possible to clearly distinguish the strain SN4T from G. massiliense G5T. Based on the taxonogenomic description and the phenotypic and biochemical characteristics of this bacterium presented in this article, we propose the SN4T strain (= CSUR P2011 = DSM 100,698) as a new species, Gorillibacterium timonense sp. nov.

International audience; In the context of the current coronavirus disease 2019 (COVID-19) pandemic, we conducted a meta-analysis on the effects of chloroquine derivatives in patients, based on unpublished and published reports available publicly on the internet as of 27 May 2020. The keywords `hydroxychloroquine', `chloroquine', `coronavirus', `COVID-19' and `SARS-Cov-2' were used in the PubMed, Google Scholar and Google search engines without any restrictions as to date or language. Twenty studies were identified involving 105 040 patients (19 270 treated patients) from nine countries (Brazil, China, France, Iran, Saudi Arabia, South Korea, Spain and the USA). Big data observational studies were associated with conflict of interest, lack of treatment dosage and duration, and absence of favourable outcome. Clinical studies were associated with favourable outcomes and details on therapy. Among clinical studies, three of four randomized controlled trials reported a significant favourable effect. Among clinical studies, a significant favourable summary effect was observed for duration of cough (OR 0.19, p 0.00003), duration of fever (OR 0.11, p 0.039), clinical cure (OR 0.21, p 0.0495), death (OR 0.32, p 4.1 x 10(-6)) and viral shedding (OR 0.43, p 0.031). A trend for a favourable effect was noted for the outcome `death and/or intensive care unit transfer' (OR 0.29, p 0.069) with a point estimate remarkably similar to that observed for death (similar to 0.3). In conclusion, a meta-analysis of publicly available clinical reports demonstrates that chloroquine derivatives are effective to improve clinical and virological outcomes, but, more importantly, they reduce mortality by a factor of 3 in patients with COVID-19. Big data are lacking basic treatment definitions and are linked to conflict of interest. The retraction of the only big data study associated with a significantly deleterious effect the day after (June 5, 2020) the acceptance of the present work (June 4, 2020) confirms the relevance of this work. (C) 2020 The Author(s). Published by Elsevier Ltd.

International audience; Tropheryma whipplei is the causative bacterium of Whipple's disease. Its first culture has led to an enlargement of the field of the caused infections. Here, we comprehensively review acute T. whipplei infections. In a cohort study featuring 4000 children, T. whipplei was significantly more common in patients with diarrhea (4%) than in those without (1.7%). A case-controlled study highlighted 58 patients suffering from pneumonia with the detection of T. whipplei in their bronchoalveolar fluids. Finally, a recent study detected T. whipplei in the blood of 36 febrile patients experiencing pulmonary symptoms in a rural area of Senegal. T. whipplei is definitively an agent of acute gastroenteritis, a cause of nonmalarial fever in Africa, and probably a cause of pulmonary infections.

International audience; The strain GD9(T) is the type strain of the newly proposed species Blautia massiliensis sp. nov., belonging to the family Lachnospiraceae. It was isolated from a fresh stool sample collected from a healthy human using the culturomics strategy. Cells are Gram-negative rods, oxygen intolerant, non-motile and non spore forming. The 16S rRNA gene sequencing showed that strain GD9T was closely related to Blautia luti, with a 97.8% sequence similarity. Major fatty acids were C14:0 (19.8%) and C16:0 (53.2%). Strain GD9T exhibits a genome of 3,717,339 bp that contains 3,346 protein-coding genes and 81 RNAs genes including 63 tRNAs. The features of this organism are described here, with its complete genome sequence and annotation. Compared with other Blautia species which are Gram positive, the strain was Gram negative justifying an emended description of the genus Blautia. (C) 2016 Elsevier Ltd. All rights reserved.

Clostridium difficile infections (CDI) are the first cause of healthcare associated diarrhea in both Europe and the USA, causing between 15,000 and 30,000 deaths annually. Over the age of 65, antibiotic treatments are the two main risk factors of developing CDI. Fecal microbiota transplantation has a major role to play in managing these infections. Gut microbiota dysbiosis associated with CDI has been now comprehensively analyzed. Elderly individuals, patients treated with antibiotics or proton pump inhibitors have a dramatically decreased level of gut microbiota diversity as well as undergoing structural changes in taxa composition. In addition to this decreased diversity, patients with CDI present an increase in species belonging to Proteobacteria and a decrease in Clostridiales Incertae Sedis XI, and some commensal bacteria as Ruminococcaceae , Lachnospiraceae or Bifidobacterium longum for patients with CDI, caused by the 027 ribotype. Fecal microbiota transplantation is followed by a reestablishment of diversity, an increase in Firmicutes and Bacteroidetes and a decrease in Proteobacteria, Enterobacteriaceae and Streptoccaceae . Most of the studies are performed using metagenomics and sometimes yield contradictory results. Large studies, including culture dependent techniques and metagenomics using optimized extraction protocols to limit biases should be designed in order to comprehensively highlight the gut microbiota dysbiosis and consider specific microbiome-based therapeutic approaches.

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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.

International audience; 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.

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. Keywords: Arcanobacterium ihumii sp. nov., culturomics, Tessaracoccus timonensis sp. nov., vagina, Varibaculum vaginae sp. nov.

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. Keywords: Arthrobacter senegalensis sp. nov., Corynebacterium senegalense sp. nov., culturomics, human skin, taxonogenomics

International audience; 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.

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.

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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.

International audience; 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 semiauto-mated 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. Breastfeeding has been associated with the health of the child and the mother, regardless of geography and socioeconomic level [1], child's growth and cognitive development [1,2]. Human studies suggest a protective effect against diarrhea, necrotizing enterocolitis, otitis media and respiratory infections in the short term and against leukemia, malocclusion, inflammatory bowel disease, malnutrition including kwashiorkor, obesity and diabetes in the long term [1,3,4]. Up to half of deaths caused by infections in children aged 6-23 months may be associated with the absence of adequate breastfeeding [1]. Breastfeeding has also been associated with a protective effect for mothers against breast cancer and possibly against ovarian cancer and diabetes [1]. In addition to nutrients, bioactive molecules such as human milk oligosaccharides, human maternal cells but also extracellular vesicles [5-8], milk contains many microbes and is therefore not sterile. The importance of the microbial diversity of human milk for the health of the offspring has been largely neglected to date. More recently, an increasing number of studies have reported an unsuspected diversity, including many health-promoting bacteria (probiotics) in the breast, colostrum or milk [9-11] but also bacteria usually considered pathogenic and frequently found in healthy controls [12]. The milk microbiota likely plays a critical role in the colonization of the child's digestive tract and in the development of its immunity [13-16]. Prolonged exclusive breastfeeding is associated with reduced diarrhea-related gut microbiota dysbiosis and microbiota differences that persist for life [8]. The disruption of this ecosystem has also been associated with maternal diseases, such as lactational mastitis and breast cancer [17-20]. Breast bacteria could also play a role in maintaining healthy breast tissue, including stimulating host immunity [17]. Many questions remain unanswered regarding the microbiome, such as the fact that microbes considered pathogenic are frequently found in controls, the mechanism by which the microbiota impacts immune development and how dysbiosis leads to gut inflammation [8]. Researchers suggested that future studies should employ metagenomics, metatranscriptomics and metabolomics approaches to understand the complete taxonomical, func

Strain Vm-5(T) was isolated from the stool specimen of a 10-year-old Amazonian boy. This bacterium is a Gram-positive, strictly aerobic rod, motile by a polar flagellum. Here we describe its phenotypic characteristics and complete genome sequence. The 4 353 177 bp long genome exhibits a G + C content of 36.87% and contains 4394 protein-coding and 125 predicted RNA genes. Phylogenetically and genetically, strain Vm-c is a member of the genus Virgibacillus but is distinct enough to be classified as a new species. We propose the creation of V. massiliensis sp. nov., whose type strain is strain Vm-5(T) (CSUR P971 = DSM 28587).