Pangenomic analysis of Coxiella burnetii unveils new traits in genome architecture.

Coxiella burnetii is the etiological agent of Q fever, a worldwide zoonosis able to cause large outbreaks. The disease is polymorphic. Symptomatic primary infection is named acute Q fever and is associated with hepatitis, pneumonia, fever, and auto-immune complications while persistent focalized infections, mainly endocarditis, and vascular infections, occur in a minority of patients but are potentially lethal. In order to evaluate the genomic features, genetic diversity, evolution, as well as genetic determinants of antibiotic resistance, pathogenicity, and ability to cause outbreaks of Q fever, we performed a pangenomic analysis and genomic comparison of 75 C. burnetii strains including 63 newly sequenced genomes. Our analysis demonstrated that C. burnetii has an open pangenome, unique genes being found in many strains. In addition, pathogenicity islands were detected in all genomes. In consequence C. burnetii has a high genomic plasticity, higher than that of other intracellular bacteria. The core- and pan-genomes are made of 1,211 and 4,501 genes, respectively (ratio 0.27). The core gene-based phylogenetic analysis matched that obtained from multi-spacer typing and the distribution of plasmid types. Genomic characteristics were associated to clinical and epidemiological features. Some genotypes were associated to specific clinical forms and countries. MST1 genotype strains were associated to acute Q fever. A significant association was also found between clinical forms and plasmids. Strains harboring the QpRS plasmid were never found in acute Q fever and were only associated to persistent focalized infections. The QpDV and QpH1 plasmids were associated to acute Q fever. In addition, the Guyanese strain CB175, the most virulent strain to date, exhibited a unique MST genotype, a distinct COG profile and an important variation in gene number that may explain its unique pathogenesis. Therefore, strain-specific factors play an important role in determining the epidemiological and clinical manifestations of Q fever alongside with host-specific factors (valvular and vascular defects notably).

Buttiauxella massiliensis sp. nov., Isolated from a Human Bone Infection.

Strain Marseille-P9829 was isolated from a bone sample collected from an open right fibula fracture from a 46-years old patient. Strain Marseille-P9829 (= CSUR P9829 = DSM 110695) was a Gram-negative, non-spore-forming and non-motile bacterium. This strain had a positive catalase activity but was oxidase-negative. The major fatty acids methyl esters were hexadecanoic acid (45.6%) and 9-hexadecenoic acid (28.4%). Matrix Assisted Laser Desorption/Ionization-Time of Flight Mass Spectrometry analysis suggested that this strain belongs to the species Buttiauxella gaviniae. Since there were few reports of clinical infections with this species in humans, whole genome sequencing was performed and a polyphasic taxono-genomic approach was followed in order to verify the classification of strain Marseille-P9829. The 16S rRNA gene sequence BLAST against the NCBI database yielded the highest similarity of 99.8% with Buttiauxella agrestis, suggesting that strain Marseille-P9829 belongs to this species. However, genomic comparison by digital DNA-DNA hybridization showed that values between strain Marseille-P9829 and other validly published Buttiauxella species were all lower than 70%. Furthermore, all average nucleotide identities were lower than 95-96%. Therefore, these results confirmed that strain Marseille-P9829 belonged to a new Buttiauxella species for which we propose the name Buttiauxella massiliensis sp. nov., with strain Marseille-P9829 as type strain.

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.

Genomic Characterization of the Novel Bartonella refiksaydamii sp. Isolated from the Blood of a Crocidura suaveolens (Pallas, 1811).

Bartonella species are reemerging infectious agents that are transmitted by arthropod vectors among animals and/or humans. At least 13 of the 35 currently recognized Bartonella species are pathogenic for humans. Most of the pathogenic species, except Bartonella quintana and Bartonella bacilliformis, are zoonotic agents with animal reservoirs, including cats, dogs, coyotes, foxes, cattle, and rodents. In this study, a novel Bartonella species was isolated from the blood of a Crocidura suaveolens (Pallas, 1811) Lesser shrew that was captured in the Bartin region of Northwestern Turkey. The strain, RSKK 19006, was characterized using whole-genome sequencing and comparison, multilocus sequence typing (gltA, rpoB, ssrA, nuoG, and 16S rRNA) and internal transcribed spacer sequencing, electron microscopy scanning, biochemical tests, and MALDI-TOF MS (matrix assisted laser desorption ionization-time of flight mass spectrometry). This novel Bartonella is a Gram-negative, rod-shaped, microaerophilic bacterium and has neither flagella nor pilus. As a consequence, we propose to name this new species Bartonella refiksaydamii sp. nov. in Bartonella genus. The zoonotic potential of this novel Bartonella species is as yet unknown.

New human-associated species of the family Atopobiaceae and proposal to reclassify members of the genus Olsenella.

Five novel bacterial strains, Marseille-P1476T (=CSURP1476T=DSM 100642T), Marseille-P3256T (=CSURP3256T=CECT 9977T), Marseille-P2936T (=CSURP2936T=DSM 103159T), Marseille-P2912T (=CSURP2912T=DSM 103345T) and Marseille-P3197T (=CSURP3197T=CCUG 71847T), were isolated from various human specimens. These five strains were not identified at the species level by matrix-assisted laser desorption/ionization time of flight mass spectrometry. Following 16S rRNA gene sequence comparisons with the GenBank database, the highest nucleotide sequence similarities of all studied strains were obtained to members of the paraphyletic genus Olsenella. A polyphasic taxono-genomic strategy (16S rRNA gene-based and core genome-based phylogeny, genomic comparison, phenotypic and biochemical characteristics) enabled us to better classify these strains and reclassify Olsenella species. Among the studied strains, Marseille-P1476T, Marseille-P2936T and Marseille-P3197T belonged to new species of the genus Olsenella for which we propose the names Olsenella massiliensis sp. nov., Olsenella phocaeensis sp. nov. and Olsenella urininfantis sp. nov., respectively. Strains Marseille-P2912T and Marseille-P3256T belonged to a new genus for which the names Thermophilibacter provencensis gen. nov., sp. nov. and Thermophilibacter mediterraneus gen. nov., sp. nov. are proposed, respectively. We also propose the creation of the genera Parafannyhessea gen. nov., Tractidigestivibacter gen. nov. and Paratractidigestivibacter gen. nov. and the reclassification of Olsenella umbonata as Parafannyhessea umbonata comb. nov., Olsenella scatoligenes as Tractidigestivibacter scatoligenes comb. nov., and Olsenella faecalis as Paratractidigestivibacter faecalis comb. nov.

Running after ghosts: are dead bacteria the dark matter of the human gut microbiota?

The human gut microbiota has been explored by a wide range of culture-dependent and culture-independent methods, revealing that many microbes remain uncharacterized and uncultured. In this work, we aimed to confirm the hypothesis that some of the species present in the human gut microbiota remain uncultured not because of culture limitations, but because all members of such species are dead before reaching the end of the gastro-intestinal tract.We evaluate this phenomenon by studying the microbial viability and culturability of the human gut microbiota from the fresh fecal materials of eight healthy adults. For the first time, we applied fluorescence-activated cell sorting (FACS) combined with 16S metagenomics analysis and microbial culturomics.We identified a total of 1,020 bacterial OTUs and 495 bacterial isolates through metagenomics and culturomics, respectively. Among the FACS metagenomics results, only 735 bacterial OTUs were alive, comprising on average 42% of known species and 87% of relative abundance per individual. The remaining uncultured bacteria were rare, dead, or injured.Our strategy allowed us to shed light on the dark matter of the human gut microbiota and revealed that both metagenomics and culturomics approaches are needed for greater insight into the diversity and richness of bacteria in the human gut microbiota. Further work on culture is needed to enhance the repertoire of cultured gut bacteria by targeting low abundance bacteria and optimizing anaerobic sample conditioning and processing to preserve the viability of bacteria.

Draft Genome Sequence of Vogesella oryzae L3B39T, Isolated from the Rhizosphere of Saline-Tolerant Pokkali Rice.

Here, we report the draft genome sequence of Vogesella oryzae L3B39T (CSUR Q2602T = DSM 28780), which is a Vogesella species isolated from the rhizosphere of saline-tolerant pokkali rice. The genome sequence was assembled into 58 contigs for a total size of 3,415,129 bp, with a G+C content of 62.3%.

Interest of bacterial pangenome analyses in clinical microbiology.

Thanks to the progress and decreasing costs in genome sequencing technologies, more than 250,000 bacterial genomes are currently available in public databases, covering most, if not all, of the major human-associated phylogenetic groups of these microorganisms, pathogenic or not. In addition, for many of them, sequences from several strains of a given species are available, thus enabling to evaluate their genetic diversity and study their evolution. In addition, the significant cost reduction of bacterial whole genome sequencing as well as the rapid increase in the number of available bacterial genomes have prompted the development of pangenomic software tools. The study of bacterial pangenome has many applications in clinical microbiology. It can unveil the pathogenic potential and ability of bacteria to resist antimicrobials as well identify specific sequences and predict antigenic epitopes that allow molecular or serologic assays and vaccines to be designed. Bacterial pangenome constitutes a powerful method for understanding the history of human bacteria and relating these findings to diagnosis in clinical microbiology laboratories in order to optimize patient management.

Draft Genome Sequence of Salirhabdus euzebyi Strain Q1438.

In 2007, Salirhabdus euzebyi was first described as a bacterial isolate from a sea salt evaporation pond. As no genome sequence was previously available for this species, we performed whole-genome sequencing. The chromosome of strain Q1438 was 3,784,443 bp long with 36% G+C content, 3,830 protein-coding genes, and 74 RNA genes.

Chitinophaga vietnamensis sp. nov., a multi-drug resistant bacterium infecting humans.

We describe a new multidrug resistant Chitinophaga species that was isolated from patients with type 2 diabetes in Vietnam. Strain BD 01T was cultivated in 2017 from a blood sample of a patient suffering from bacteremia. Strain VP 7442 was isolated in 2018 from a pleural fluid sample of a patient who had presented with lung abscess and pleural effusion. Both strains are aerobic, Gram-negative, non-motile and non-spore-forming. The 16S rRNA gene sequences of both strains are 100 % similar and share a highest 16S sequence identity with Chitinophaga polysaccharea MRP-15T of 97.42 %. Their predominant fatty acid is iso-C15 : 0 (73.8 % for strain BD 01T and 79.8 % for strain VP 7442). The draft genome sizes of strains BD 01T and VP 7442 are 6 308 408 and 6 308 579 bp, respectively. They are resistant to beta-lactams, aminoglycosides, fluoroquinolones, metronidazole, fosfomycin, vancomycin and macrolides, and exhibit 20 and 18 antimicrobial resistance-related genes, respectively. Using the multiphasic taxonogenomic approach, we propose that strains BD 01T (=CSUR P9622=VTCC 70981) and VP 7442 (=CSUR P9623=VTCC 70982) represent a new species, for which we propose the name Chitinophaga vietnamensis sp. nov. Strain BD 01T was chosen as type strain of C. vietnamensis sp. nov.

Case Report: Vibrio cholerae Biliary Tract Infections in Two North Africans in France.

The origin of a cholera outbreak may be unclear, as recently in Algeria. In two patients from North Africa, Vibrio cholerae was isolated in the context of hepatobiliary tract infections without any known outbreak. Gallbladder and asymptomatic long-term carriers might play a role in the emergence of cholera.

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.

Whole-Genome Sequence of French Clinical Peptoniphilus catoniae Strain P8546.

In 2016, Peptoniphilus catoniae was reported as a bacterial species isolated from a healthy Peruvian male. In 2018, a clinical strain from the same species was isolated from the stool of a French patient with kidney cancer. The genome of this strain, P8546, was 1,725,465 bp long, with 33.4% G+C content.

Whole-Genome Sequence of Haloimpatiens lingqiaonensis Strain P8956.

In 2016, Haloimpatiens lingqiaonensis was described as a bacterial isolate from paper mill wastewater. Previously, no whole-genome sequence was available for this microorganism. Whole-genome sequencing of strain P8956 yielded a 3,295,388-bp genome with a 30.7% G+C content, 2,917 protein-coding genes, and 95 predicted RNA genes.

Whole-Genome Sequence of French Clinical Olivibacter jilunii Strain P8502.

In 2013, Olivibacter jilunii was reported as a bacterial species isolated from contaminated soil. In 2018, a clinical strain from the same species was isolated from the rectal swab of a Hajj pilgrim. Genome sequencing yielded 6,704,032 bp, with 41.2% G+C content, 5,406 protein-coding genes, and 54 predicted RNA genes in strain P8502.

Draft genome and description of Chryseobacterium phocaeense sp. nov.: a new bacterial species isolated from the sputum of a cystic fibrosis patient.

Strain 6021061333T was isolated from the sputum of 16-year-old girl with cystic fibrosis following a pulmonary exacerbation. This bacterial strain could not be identified by our systematic MALDI-TOF mass spectrometry screening on a MicroFlex. This led to the sequencing of the 16S rRNA gene, which shows 97.83% sequence identity with Chryseobacterium kwangjuense strain KJ1R5T, the phylogenetic closely related type strain of a species with standing in nomenclature, which putatively classifies it as a new species. Colonies are yellow, circular and 0.5-1 mm in diameter after cultivation at 28 °C for 24 h on 5% sheep blood-enriched Colombia agar. Growth occurs at temperatures in the range of 28-37 °C (optimally at 28 °C). Strain 6021061333T is Gram-negative, non-motile and strictly aerobic bacillus. It is catalase and oxidase positive. The 4,864,678 bp-long genome, composed of five contigs, has a G+C content of 38.86%. Out of the 4427 predicted genes, 4342 were protein-coding genes and 85 were RNAs. The major fatty acids are branched (13-methyl-tetradecanoic acid and 15-methyl-hexadecenoic acid). Digital DNA-DNA hybridization (dDDH) estimation and average nucleotide identity (ANI) of the strain 6021061333T against genomes of the type strains of related species ranged between 23.60 and 50.40% and between 79.31 and 93.06%, respectively. According to our taxonogenomics results, we propose the creation of Chryseobacterium phocaeense sp. nov. that contains the type strain 6021061333T (= CSUR P2660, = CECT 9670).

Draft genome and description of Merdibacter massiliensis gen.nov., sp. nov., a new bacterium genus isolated from the human ileum.

We used phenotypic, genomic and phylogenetic information following the taxono-genomics approach to demonstrate that strain Marseille-P3254, isolated from an ileal sample of a 76-year old woman who underwent upper and lower digestive tract endoscopy for esophagitis and colonic polyp, is representative of a novel bacterial genus within the family Erysipelotrichaceae in the phylum Firmicutes. It is an anaerobic Gram-negative bacterium without catalase and oxidase activities. The genome of strain Marseille-P3254 is 2,468,496-bp long with a 40.1% G + C content. This new bacterium is most closely related to Eubacterium dolichum, with which it shares 90.7% 16S rRNA sequence similarity. In addition, genomic comparison using the digital DNA-DNA hybridization and OrthoANI analyses between the novel organism and the E. dolichum type strain revealed identities of 25.2 and 68.91%, respectively. The major fatty acids were C16: 0, C18: 1n9 and C18: 0. Based on these data, we propose the creation of the new genus Merdibacter gen. nov., with strain Marseille-P3254T (=CSUR P3254 = DSM 103534) being the type strain of the new species Merdibacter massiliensis gen. nov., sp. nov.

Description of Janibacter massiliensis sp. nov., cultured from the vaginal discharge of a patient with bacterial vaginosis.

Strain Marseille-P4121T was isolated from a vaginal sample of a 45-year-old French woman with bacterial vaginosis. It is a Gram-positive, asporogenous, non-motile and aerobic bacterium. Strain Marseille-P4121T exhibits 98.2% 16S rRNA sequence similarity with Janibacter alkaliphilus strain SCSIO 10480T, a phylogenetically closely related species with standing in nomenclature. Its major fatty acids were identified as C18:1ω9 (34.4%), C16:0 (30.1%), and C18:0 (19%). The draft genome size of strain Marseille-P4121T is 2,452,608 bp long with a 72.5% G+C content and contains 2351 protein-coding genes and 49 RNA genes including 3 rRNA genes. We propose that strain Marseille-P4121T (= CECT 9671T = CSUR P4121T) is the type strain of the new species Janibacter massiliensis sp. nov.

Dihydroxyacetone as a definitive treatment for aluminium phosphide poisoning in rats.

Aluminium phosphide (AlP), a very toxic pesticide also known as the rice tablet, releases phosphine gas upon contact with water, moisture, or gastric acid. Its mortality rate in humans is 70-100 % due to cardiogenic shock and refractory hypotension. Dihydroxyacetone (DHA) is a simple ketonic carbohydrate, mainly used for sunless skin tanning. It also plays a beneficial role in the treatment of hypotension and cardiogenic shock by restoring blood volume and cellular respiration. The aim of this study was to investigate the its effect on the haemodynamics and electrocardiogram (ECG) in male rats poisoned with AlP. The animals were divided into the following groups: control (received 1 mL corn oil, orally), AlP (received 15 mg kg-1 AlP solved in corn oil, orally), AlP plus DHA (treated with 50 mg kg-1 of DHA 30 min after receiving AlP), and AlP plus N-acetyl cysteine (NAC) (treated with 200 mg kg-1 of NAC 30 min after receiving AlP). The animals were then anaesthetised and ECG, blood pressure, and heart rate were recorded for 120 min. Treatment with AlP alone and in combination with NAC was associated with progressive hypotension, tachycardia, and ECG disturbances in rats, resulting in 100 % mortality 3 h after poisoning. However, DHA achieved 100 % survival in the poisoned rats and prevented AlP-induced ECG and haemodynamic abnormalities. The main mechanism of DHA in the treatment of AlP poisoning is unclear, but the findings suggest the promising therapeutic potential of DHA against AlP poisoning.