Whole Genome Analyses Accurately Identify Neisseria spp. and Limit Taxonomic Ambiguity.

Genome sequencing facilitates the study of bacterial taxonomy and allows the re-evaluation of the taxonomic relationships between species. Here, we aimed to analyze the draft genomes of four commensal Neisseria clinical isolates from the semen of infertile Lebanese men. To determine the phylogenetic relationships among these strains and other Neisseria spp. and to confirm their identity at the genomic level, we compared the genomes of these four isolates with the complete genome sequences of Neisseria gonorrhoeae and Neisseria meningitidis and the draft genomes of Neisseria flavescens, Neisseria perflava, Neisseria mucosa, and Neisseria macacae that are available in the NCBI Genbank database. Our findings revealed that the WGS analysis accurately identified and corroborated the matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) species identities of the Neisseria isolates. The combination of three well-established genome-based taxonomic tools (in silico DNA-DNA Hybridization, Ortho Average Nucleotide identity, and pangenomic studies) proved to be relatively the best identification approach. Notably, we also discovered that some Neisseria strains that are deposited in databases contain many taxonomical errors. The latter is very important and must be addressed to prevent misdiagnosis and missing emerging etiologies. We also highlight the need for robust cut-offs to delineate the species using genomic tools.

Emergence of a Neisseria flavescens clinical strain with a high level of third-generation cephalosporins resistance in Lebanon.

Third generation cephalosporins (3GC)-resistant Neisseria flavescens could cause difficult-to-treat infections. This study aimed to investigate the genetic determinants of 3GC resistance using whole genome sequence comparisons of three N. flavescens; one of which was 3GC-resistant, while two were susceptible. Mutations in penA, ponA, and porB might be associated with the 3GC-resistance.

Colibacter massiliensis gen. nov. sp. nov., a novel Gram-stain-positive anaerobic diplococcal bacterium, isolated from the human left colon.

The gut microbiota is considered to play a key role in human health. As a consequence, deciphering its microbial diversity is mandatory. A polyphasic taxonogenomic strategy based on the combination of phenotypic and genomic analyses was used to characterize a new bacterium, strain Marseille-P2911. This strain was isolated from a left colon sample of a 60-year old man who underwent a colonoscopy for an etiological investigation of iron-deficiency anemia in Marseille, France. On the basis of 16S rRNA sequence comparison, the closest phylogenetic neighbor was Anaeroglobus geminatus (94.59% 16S rRNA gene sequence similarity) within the family Veillonellaceae. Cells were anaerobic, Gram-stain-positive, non-spore-forming, catalase/oxidase negative cocci grouped in pairs. The bacterium was able to grow at 37 °C after 2 days of incubation. Strain Marseille-P2911 exhibited a genome size of 1,715,864-bp with a 50.2% G + C content, and digital DNA-DNA hybridization (dDDH) and OrthoANI values with A. geminatus of only 19.1 ± 4.5% and 74.42%, respectively. The latter value being lower than the threshold for genus delineation (80.5%), we propose the creation of the new genus Colibacter gen. nov., with strain Marseille-P2911T (=DSM 103304 = CSUR P2911) being the type strain of the new species Colibacter massiliensis gen. nov., sp. nov.

"Mycobacterium mephinesia", a Mycobacterium terrae complex species of clinical interest isolated in French Polynesia.

A 59-year-old tobacco smoker male with chronic bronchitis living in Taravao, French Polynesia, Pacific, presented with a two-year growing nodule in the middle lobe of the right lung. A guided bronchoalveolar lavage inoculated onto Löwenstein-Jensen medium yielded colonies of a rapidly-growing non-chromogenic mycobacterium designed as isolate P7213. The isolate could not be identified using routine matrix-assisted laser desorption ionization-time of flight-mass spectrometry and phenotypic and probe-hybridization techniques and yielded 100% and 97% sequence similarity with the respective 16S rRNA and rpoB gene sequences of Mycobacterium virginiense in the Mycobacterium terrae complex. Electron microscopy showed a 1.15 µm long and 0.38 µm large bacillus which was in vitro susceptible to rifampicin, rifabutin, ethambutol, isoniazid, doxycycline and kanamycin. Its 4,511,948-bp draft genome exhibited a 67.6% G + C content with 4,153 coding-protein genes and 87 predicted RNA genes. Genome sequence-derived DNA-DNA hybridization, OrthoANI and pangenome analysis confirmed isolate P7213 was representative of a new species in the M. terrae complex. We named this species "Mycobacterium mephinesia".

Evaluation of different testing tools for the identification of non-gonococcal Neisseria spp. isolated from Lebanese male semen: a strong and significant association with infertility.

PURPOSE: The aim was to evaluate several microbiological tools for the identification of non-gonococcal Neisseria spp. isolated from semen samples from Lebanese men and to determine the putative link between the presence of Neisseria commensal species and infertility. METHODOLOGY: Within a cross-sectional retrospective study design, the whole population included in this investigation was divided in 2 categories: 173 patients with symptoms of infertility and 139 patients with normal seminograms. Epidemiological and microbiological investigations were performed for 59 strains of Neisseria through several phenotypic and genotypic tools, including seminograms, an analytical profile index of Neisseria and Haemophilus (API-NH), matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), porA PCR, 16S rRNA and rplF gene sequencing, and antimicrobial susceptibility testing. RESULTS: The risk of Neisseria infection was twice as high in infertile patients compared to the control group [odds ratio (OR): 1.95, confidence interval (CI): 1.05-3.65, P =0.03]. Unreliable diagnosis of Neisseria urogenital infection has serious health and social consequences. Our findings showed that API-NH and 16S rRNA sequencing are poor tools to identify Neisseria at the species level. Therefore, reliable diagnosis of cases using MALDI-TOF MS and/or rplF sequencing is needed to provide critical treatment decisions and prevent antimicrobial resistance spreading in the community. CONCLUSION: This work predicted a strong and significant association between the presence of Neisseria spp. in semen and male infertility among the Lebanese population. For a better understanding of this association, it is recommended that more genomic and large-scale epidemiological investigations are undertaken to reach definitive conclusions.