Description of Agathobaculum massiliense sp. nov., a new bacterial species prevalent in the human gut and predicted to produce indole and tryptophan based on genomic analysis.

The novel bacterial strain Marseille-P4005T was isolated from the stool sample of a healthy donor. It is a Gram-stain negative, non-motile, non-spore-forming rod. It grew optimally at 37 °C and at pH 7.0 on 5% sheep blood-enriched Columbia agar after preincubation in a blood-culture bottle supplemented with rumen and blood. This strain does not ferment monosaccharides (except D-tagatose), disaccharides, or polymeric carbohydrates. The major cellular fatty acids were hexadecenoic (24.6%), octadecanoic (22.8%), and tetradecanoic (20.1%) acids. Next-generation sequencing revealed a genome size of 3.2 Mbp with a 56.4 mol% G + C. Phylogenetic analysis based on the 16S rRNA gene highlighted Agathobaculum desmolans strain ATCC 43058T as the closest related strain. The OrthoANI, AAI, and digital DNA-DNA hybridization values were below the critical thresholds of 95%, 95-96%, and 70%, respectively, to define a novel bacterial species. Antibiotic resistance genes APH(3')-IIIa, erm(B), and tet(W) were detected with high identity percentages of 100%, 98.78%, and 97.18% for each gene, respectively. The APH(3')-IIIa gene confers resistance to amikacin, erm(B) gene confers resistance to erythromycin, lincomycin, and clindamycin, while tet(W) gene confers resistance to doxycycline and tetracycline. Based on KEGG BlastKOALA analyses, the annotation results showed that our strain could use glucose to produce L-lactate and pyruvate but not acetate or ethanol. Also, strain Marseille-P4005T was predicted to use phenylalanine to produce indole, a major intercellular signal molecule within the gut microbial ecosystem. Through having a gene coding for tryptophan synthase beta chain (trpB), strain Marseille-P4005T could produce L-tryptophan (an essential amino acid) from indole. Strain Marseille-P4005T showed its highest prevalence in the human gut (34.19%), followed by the reproductive system (17.98%), according to a query carried out on the Integrated Microbial NGS (IMNGS) platform. Based on phylogenetic, phenotypic, and genomic analyses, we classify strain Marseille-P4005T (= CSUR P4005 = CECT 9669), a novel species within the genus Agathobaculum, for which the name of Agathobaculum massiliense sp. nov. is proposed.

Arabiibacter massiliensis gen. nov. sp. nov., New Anaerobic Bacterium Isolated from the Human Gut.

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.

Neglectibacter timonensis gen. nov., sp. nov. and Scatolibacter rhodanostii gen. nov., sp. nov., two anaerobic bacteria isolated from human stool samples.

Strains Marseille-P2265T (=CSUR P2265T =DSM 102082 T) and Marseille-P3890T (=CSUR P3890T =CCUG 72341 T) were isolated from stool samples using the culturomics approach. The 16S rRNA gene sequences of both strains were sequenced and compared by BLASTn to the NCBI database. Strains Marseille-P2265T and Marseille-P3890T were most closely related to Acutalibacter muris with identities of 94.3% and 91.5%, respectively. Between the two strains, the 16S rRNA gene sequence identity was 91.5%. Both strains are anaerobic Gram-positive, oxidase- and catalase-negative. The major fatty acid methyl esters (> 10%) in both strains are C16:0 and anteiso-C15:0. Additionally, strain Marseille-P2265T has iso-C15:0 and C14:0, and strain Marseille-P3890T, iso-C14:0. Strain Marseille-P2265T has a genome size of 3,671,396-bp with a G + C content of 52.8%. As for strain Marseille-P3890T, the genome is 2,702,024-bp-long with a 39.8% G + C content. The genomic comparison of closely related species with strains Marseille-P2265T and Marseille-P3890T showed that all digital DNA-DNA hybridization (dDDH), orthologous average nucleotide identity (OrthoANI) and average amino acid identity (AAI) values were lower than the published species thresholds (70% for dDDH, 95-96% for OrthoANI/AAI). Based on these results, it was concluded that strains Marseille-P2265T and Marseille-P3890T belong to two new genera in the family Oscillospiraceae. For these two genera, the names Neglectibacter gen. nov. and Scatolibacter gen. nov. were proposed, with strains Marseille-P2265T and Marseille-P3890T being the type strains of Neglectibacter timonensis gen. nov., sp. nov. and Scatolibacter rhodanostii gen. nov., sp. nov., respectively.

Gemella massiliensis sp. nov., a new bacterium isolated from the human sputum.

Thanks to its ability to isolate previously uncultured bacterial species, culturomics has dynamized the study of the human microbiota. A new bacterial species, Gemella massiliensis Marseille-P3249T, was isolated from a sputum sample of a healthy French man. Strain Marseille-P3249T is a facultative anaerobe, catalase-negative, Gram positive, coccus, and unable to sporulate. The major fatty acids were C16:0 (34%), C18:1n9 (28%), C18:0 (15%) and C18:2n6 (13%). Its 16S rRNA sequence exhibits a 98.3% sequence similarity with Gemella bergeri strain 617-93T, its phylogenetically closest species with standing in nomenclature. Its digital DNA-DNA hybridization (dDDH) and OrthoANI values with G. bergeri of only 59.7 ± 5.6% and 94.8%, respectively. These values are lower than the thresholds for species delineation (> 70% and > 95%, respectively). This strain grows optimally at 37 °C and its genome is 1.80 Mbp long with a 30.5 mol% G + C content. Based on these results, we propose the creation of the new species Gemella massilienis sp. nov., strain Marseille-P3249T (= CSUR P3249 = DSMZ 103940).

Lagierella massiliensis gen. nov., sp. nov., Isolated from a Stool Sample.

Strain Marseille-P2012T was described to represent a new bacterial genus belonging to the phylum Firmicutes using the taxonogenomics concept. It was isolated from stool samples of a healthy 2-year-old Senegalese boy in a study of the human gut microbiota. This strain is a Gram-positive, anaerobic, non-motile and coccus-shaped bacterium. The 16S rRNA gene sequence of strain Marseille-P2012 exhibited 90.5% similarity with Finegoldia magna strain ATCC 29,328, the phylogenetically closest species with standing in nomenclature. The genome of strain Marseille-P2012T is 1,832,315 bp-long with 32.46 mol% of G + C content. With regard to its phenotypic, biochemical and genomic characteristics, this bacterium was classified as a new bacterial genus and species, Lagierella massiliensis gen. nov., sp. nov., with strain Marseille-P2012T (= CSUR P2012 = DSM100854) as type strain.

Strategies and advancements in human microbiome description and the importance of culturomics.

The human microbiota gained a big interest among the scientific community with numerous studies being performed to better understand its role in health and diseases. Even with all the success achieved in studying the bacterial populations at the different body sites and its interaction among each other and with the host, some links remain missing and might have therapeutic benefits. In this review, we summarize the main means used for bacterial identification, human microbiota description and the role of culturomics in leading the way towards the development of new bacterio-therapeutic approaches.

Dysgonomonas massiliensis sp. nov., a new species isolated from the human gut and its taxonogenomic description.

Culturomics has allowed the isolation of a significant number of new bacterial species from the human gut microbiota and proved to be a valuable complement to culture-independent techniques. Using this culture-based approach, a new bacterial species has been isolated from a stool sample of a 39-year-old healthy Pygmy male and described using the taxonogenomic strategy. Cells of strain Marseille-P4356T are Gram-stain negative cocci. The strain grows optimally at 37 °C and is catalase positive but oxidase negative. Its 16S rRNA gene sequence exhibited 92.96% sequence similarity with Dysgonomonas gadei strain JCM 16698T (NR_113134.1), currently its phylogenetically closest species that has been validly named. The genome of strain Marseille-P4356T is 3,472,011 bp long with 37.3 mol% G+C content. Phenotypic, biochemical, proteomic, genomic and phylogenetic analyses, clearly demonstrate that strain Marseille-P4356T (= CCUG 71356T = CSUR P4356T) represents a new species within the genus Dysgonomonas, for which we propose the name Dysgonomonas massiliensis sp. nov.

Phoenicibacter congonensis gen. nov., sp. nov., a new genus isolated from the human gut and its description using a taxonogenomic approach.

Culturomics has recently allowed the isolation and description of previously uncultured bacteria from the human microbiome at different body sites. As part of a project aiming to describe the human gut microbiota by culturomics, Phoenicibacter congonensis strain Marseille-P3241T was isolated from the gut of a 45 years old Pygmy female. In the present work, we aim to describe this strain via the taxonogenomics approach. The major phenotypic, genomic and biochemical characteristics of this strain were analysed. Strain Marseille-P3241T is an anaerobic, Gram-positive and motile coccobacillus that grows optimally at 37 °C. The genome of strain Marseille-P3241T is 1,447,956 bp long with 43.44% GC content and its 16S rRNA gene sequence exhibited 89% sequence similarity with that of Denitrobacterium detoxificans strain NPOH1T, the phylogenetically closest related species with current standing in nomenclature. After performing a phylogenetic and genomic analysis, we conclude that strain Marseille-P3241T (= CCUG 70681T = CSUR P3241T) represents the type species of a new genus, for which we propose the name Phoenicibacter congonensis gen. nov., sp. nov.

Culturomics, a potential approach paving the way toward bacteriotherapy.

The human microbiota has been extensively studied over the past decade to describe its role in health and diseases. Numerous studies showed the presence of bacterial imbalance in a variety of human health conditions, suggesting great potential for the development of bacteriotherapies. Identifying mechanisms involving the human microbiota has been very challenging due to the complex data generated by molecular approaches and the limited number of organisms isolated by culture and described. This review summarizes the efforts done to describe the human microbiota through culturomics and the advancements in culturing the organisms residing at different body sites.

Peptostreptococcus faecalis sp. nov., new bacterial species isolated from healthy indigenous congolese volunteer.

The Microbial Culturomics Project aiming to discover several bacterial species made it possible to isolate the strain Marseille-P4308T from a stool sample of a healthy indigenous Congolese volunteer. Strain Marseille-P4308T is a Gram-positive coccus shaped bacterium that optimally grows at 37 °C. The 16S rRNA gene sequence of the strain has a 96.2% sequence similarity to Peptostreptococcus anaerobius strain NCTC 11460T (GenBank accession number: NR_042847.1). In addition, the average nucleotide identity of strain Marseille-P4308T with its closest related species was 71.1%, which was far below the recommended threshold (>95-96%). The genome of the strain Marseille-P4308T has a length of 2.14 Mbp with G + C content of 30.4 mol%. Based on phenotypic, biochemical, genomic and phylogenetic analysis, strain Marseille-P4308T (= CSUR P4308 = CECT 9960) clearly appears to be a new species for which the name Peptostreptococcus faecalis sp. nov., is proposed.

Miniphocibacter massiliensis gen. nov., sp. nov., a new species isolated from the human gut and its taxono-genomics description.

With the aim of describing the human microbiota by the means of culture methods, culturomics was developed in order to target previously un-isolated bacterial species and describe it via the taxono-genomics approach. While performing a descriptive study of the human gut microbiota of the pygmy people, strain Marseille-P4678T has been isolated from a stool sample of a healthy 39-year-old pygmy male. Cells of this strain were Gram-positive cocci, spore-forming, non-motile, catalase-positive and oxidase-negative, and grow optimally at 37°C under anaerobic conditions. Its 16S rRNA gene sequence exhibited 89.69% of sequence similarity with Parvimonas micra strain 3119BT (NR 036934.1), its phylogenetically closest species with standing in nomenclature. The genome of strain Marseille-P4678T is 2,083,161 long with 28.26 mol% of G+C content. Based on its phenotypic, biochemical, genotypic and proteomic profile, this bacterium was classified as a new bacterial genus and species Miniphocibacter massiliensis gen. nov., sp. nov. with the type strain Marseille-P4678T .

Taxonogenomics description of Parabacteroides timonensis sp. nov. isolated from a human stool sample.

Intensive efforts have been made to describe the human microbiome and its involvement in health and disease. Culturomics has been recently adapted to target formerly uncultured bacteria and other unclassified bacterial species. This approach enabled us to isolate in the current study a new bacterial species, Parabacteroides timonensis strain Marseille-P3236T , from a stool sample of a healthy 39-year-old pygmy male. This strain, is an anaerobic, gram-negative, nonspore-forming motile rod. Its genome is made up of 6,483,434 bp with 43.41% G+C content, 5046 protein-encoding genes, and 84 RNA genes. We herein provide the full description of Parabacteroides timonensis strain Marseille-P3236T through the taxonogenomic approach.

Noncontiguous finished genome sequence and description of Raoultibacter massiliensis gen. nov., sp. nov. and Raoultibacter timonensis sp. nov, two new bacterial species isolated from the human gut.

As part of the culturomics project aiming at describing the human microbiota, we report in this study the description of the new bacterial genus Raoultibacter gen. nov. that includes two new species, that is, R. massiliensis sp. nov. and R. timonensis sp. nov. The R. massiliensis type strain Marseille-P2849T was isolated from the fecal specimen of a healthy 19-year-old Saudi Bedouin, while R. timonensis type strain Marseille-P3277T was isolated from the feces of an 11-year-old pygmy female living in Congo. Strain Marseille-P2849T exhibited 91.4% 16S rRNA sequence similarity with Gordonibacter urolithinfaciens, its phylogenetic closest neighbor with standing in nomenclature. As well, strain Marseille-P3277T exhibited 97.96% 16S rRNA similarity with strain Marseille-P2849T . Both strains were Gram-positive, motile, nonspore-forming rod and form transparent microcolonies on blood agar in both anaerobic and microaerophilic atmospheres. The genome sizes of strain Marseille-P2849T and strain Marseille-P3277T were 3,657,161 bp and 4,000,215 bp, respectively. Using a taxono-genomic approach combining the phenotypic, biochemical, and genomic characteristics, we propose the genus Raoultibacter gen. nov., which contains strains Marseille-P2849T (= CSUR P2849T , = DSM 103407T ) and Marseille-P3277T (=CCUG 70680T , =CSUR P3277T ) as type strains of the species R. massiliensis sp. nov., and R. timonensis sp. nov., respectively.

The contribution of culturomics to the repertoire of isolated human bacterial and archaeal species.

After a decade of research and metagenomic analyses, our knowledge of the human microbiota appears to have reached a plateau despite promising results. In many studies, culture has proven to be essential in describing new prokaryotic species and filling metagenomic gaps. In 2015, only 2172 different prokaryotic species were reported to have been isolated at least once from the human body as pathogens or commensals. In this review, we update the previous repertoire by reporting the different species isolated from the human body to date, increasing it by 28% to reach a total of 2776 species associated with human beings. They have been classified into 11 different phyla, mostly the Firmicutes, Proteobacteria, Bacteroidetes, and Actinobacteria. Finally, culturomics contributed up to 66.2% towards updating this repertoire by reporting 400 species, of which 288 were novel. This demonstrates the need to continue the culturing work, which seems essential in order to decipher the hidden human microbial content.

Culturing the human microbiota and culturomics.

The gut microbiota has an important role in the maintenance of human health and in disease pathogenesis. This importance was realized through the advent of omics technologies and their application to improve our knowledge of the gut microbial ecosystem. In particular, the use of metagenomics has revealed the diversity of the gut microbiota, but it has also highlighted that the majority of bacteria in the gut remain uncultured. Culturomics was developed to culture and identify unknown bacteria that inhabit the human gut as a part of the rebirth of culture techniques in microbiology. Consisting of multiple culture conditions combined with the rapid identification of bacteria, the culturomic approach has enabled the culture of hundreds of new microorganisms that are associated with humans, providing exciting new perspectives on host-bacteria relationships. In this Review, we discuss why and how culturomics was developed. We describe how culturomics has extended our understanding of bacterial diversity and then explore how culturomics can be applied to the study of the human microbiota and the potential implications for human health.

Genomic and phenotypic description of the newly isolated human species Collinsella bouchesdurhonensis sp. nov.

Using culturomics, a recently developed strategy based on diversified culture conditions for the isolation of previously uncultured bacteria, we isolated strain Marseille-P3296T from a fecal sample of a healthy pygmy female. A multiphasic approach, taxono-genomics, was used to describe the major characteristics of this anaerobic and gram-positive bacillus that is unable to sporulate and is not motile. The genome of this bacterium is 1,878,572 bp-long with a 57.94 mol% G + C content. On the basis of these characteristics and after comparison with its closest phylogenetic neighbors, we are confident that strain Marseille-P3296T (=CCUG 70328 =  CSUR P3296) is the type strain of a novel species for which we propose the name Collinsella bouchesdurhonensis sp. nov.

Eggerthella timonensis sp. nov, a new species isolated from the stool sample of a pygmy female.

Eggerthella timonensis strain Marseille-P3135 is a new bacterial species, isolated from the stool sample of a healthy 8-year-old pygmy female. This strain (LT598568) showed a 16S rRNA sequence similarity of 96.95% with its phylogenetically closest species with standing in nomenclature Eggerthella lenta strain DSM 2243 (AF292375). This bacterium is a nonspore forming, Gram-positive, nonmotile rod with catalase but no oxidase activity. Its genome is 3,916,897 bp long with 65.17 mol% of G + C content. Of the 3,371 predicted genes, 57 were RNAs and 3,314 were protein-coding genes. Here, we report the main phenotypic, biochemical, and genotypic characteristics of E. timonensis strain Marseille-P3135 (=CSUR P3135, =CCUG 70327); ti.mo.nen'sis, N.L. masc. adj., with timonensis referring to La Timone, which is the name of the hospital in Marseille (France) where this work was performed). Strain is a nonmotile Gram-positive rod, unable to sporulate, oxidase negative, and catalase positive. It grows under anaerobic conditions between 25°C and 42°C but optimally at 37°C.

Developmental Cycle and Genome Analysis of Protochlamydia massiliensis sp. nov. a New Species in the Parachlamydiacae Family.

Amoeba-associated microorganisms (AAMs) are frequently isolated from water networks. In this paper, we report the isolation and characterization of Protochlamydia massiliensis, an obligate intracellular Gram-negative bacterium belonging to the Parachlamydiaceae family in the Chlamydiales order, from a cooling water tower. This bacterium was isolated on Vermamoeba vermiformis. It has a multiple range of hosts among amoeba and is characterized by a typical replication cycle of Chlamydiae with a particularity, recently shown in some chlamydia, which is the absence of inclusion vacuoles in the V. vermiformis host, adding by this a new member of Chlamydiae undergoing developmental cycle changes in the newly adapted host V. vermiformis. Draft genome sequencing revealed a chromosome of 2.86 Mb consisting of four contigs and a plasmid of 92 Kb.

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

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 microorganism1. The pure culture of prokaryotes, neglected in recent decades, remains essential to elucidating the role of these organisms2. 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 identification2. 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 studies3-5. 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.