Methanomethylophilus alvi gen. nov., sp. nov., a Novel Hydrogenotrophic Methyl-Reducing Methanogenic Archaea of the Order Methanomassiliicoccales Isolated from the Human Gut and Proposal of the Novel Family Methanomethylophilaceae fam. nov.

The methanogenic strain Mx-05T was isolated from the human fecal microbiome. A phylogenetic analysis based on the 16S rRNA gene and protein marker genes indicated that the strain is affiliated with the order Methanomassiliicoccales. It shares 86.9% 16S rRNA gene sequence identity with Methanomassiliicoccus luminyensis, the only member of this order previously isolated. The cells of Mx-05T were non-motile cocci, with a diameter range of 0.4-0.7 µm. They grew anaerobically and reduced methanol, monomethylamine, dimethylamine, and trimethylamine into methane, using H2 as an electron donor. H2/CO2, formate, ethanol, and acetate were not used as energy sources. The growth of Mx-05T required an unknown medium factor(s) provided by Eggerthella lenta and present in rumen fluid. Mx-05T grew between 30 °C and 40 °C (optimum 37 °C), over a pH range of 6.9-8.3 (optimum pH 7.5), and between 0.02 and 0.34 mol.L-1 NaCl (optimum 0.12 mol.L-1 NaCl). The genome is 1.67 Mbp with a G+C content of 55.5 mol%. Genome sequence annotation confirmed the absence of the methyl branch of the H4MPT Wood-Ljungdahl pathway, as described for other Methanomassiliicoccales members. Based on an average nucleotide identity analysis, we propose strain Mx-05T as being a novel representative of the order Methanomassiliicoccales, within the novel family Methanomethylophilaceae, for which the name Methanomethylophilus alvi gen. nov, sp. nov. is proposed. The type strain is Mx-05T (JCM 31474T).

Physicochemical and Microbiological Water Quality Assessment of a Northwestern Algerian Dam: Detection of Ichtyopathogenic Bacteria.

Freshwater fish are often exposed to threats from anthropogenic or natural origins, such as pathogenic or opportunistic microorganisms responsible for a broad range of severe infections. In this study, we aimed to assess this microbiological threat to fish in an Algerian northwestern dam Sekkak (Tlemcen) by evaluating the diversity of ichtyopathogenic bacteria. In order to determine the water quality, physicochemical analyses of the dam water were carried out in situ. Ichtyopathogenic bacteria were isolated on selective media and identified by API galleries and molecular techniques (PCR and sequencing of the 16S rRNA gene). Besides, the antibiograms were constructed for all the isolates. The physicochemical and bacteriological analyses allowed us to classify the dam water as moderately polluted to polluted. Furthermore, an important diversity of ichtyopathogenic bacterial species was observed as Aeromonas hydrophila, Providencia rettgeri, and Pseudomonas aeruginosa were retrieved. The antibiogram test revealed notable resistance. The antibiotic family for which most resistances were found was the ß-lactam family, followed by aminoglycosides and macrolides. These results indicate that aquatic environments can shelter multidrug-resistant pathogenic bacteria representing a threat to the endemic fauna. Therefore, it is important to closely monitor these waters in order to improve the fish's living environment and ensure healthier production.

Aminithiophilus ramosus gen. nov., sp. nov., a sulphur-reducing bacterium isolated from a pyrite-forming enrichment culture, and taxonomic revision of the family Synergistaceae.

A novel sulphur-reducing bacterium was isolated from a pyrite-forming enrichment culture inoculated with sewage sludge from a wastewater treatment plant. Based on phylogenetic data, strain J.5.4.2-T.3.5.2T could be affiliated with the phylum Synergistota. Among type strains of species with validly published names, the highest 16S rRNA gene sequence identity value was found with Aminiphilus circumscriptus ILE-2T (89.2 %). Cells of the new isolate were Gram-negative, non-spore-forming, straight to slightly curved rods with tapered ends. Motility was conferred by lateral flagella. True branching of cells was frequently observed. The strain had a strictly anaerobic, asaccharolytic, fermentative metabolism with peptides and amino acids as preferred substrates. Sulphur was required as an external electron acceptor during fermentative growth and was reduced to sulphide, whereas it was dispensable during syntrophic growth with a Methanospirillum species. Major fermentation products were acetate and propionate. The cellular fatty acid composition was dominated by unsaturated and branched fatty acids, especially iso-C15 : 0. Its major polar lipids were phosphatidylglycerol, phosphatidylethanolamine and distinct unidentified polar lipids. Respiratory lipoquinones were not detected. Based on the obtained data we propose the novel species and genus Aminithiophilus ramosus, represented by the type strain J.5.4.2-T.3.5.2T (=DSM 107166T=NBRC 114655T) and the novel family Aminithiophilaceae fam. nov. to accommodate the genus Aminithiophilus. In addition, we suggest reclassifying certain members of the Synergistaceae into new families to comply with current standards for the classification of higher taxa. Based on phylogenomic data, the novel families Acetomicrobiaceae fam. nov., Aminiphilaceae fam. nov., Aminobacteriaceae fam. nov., Dethiosulfovibrionaceae fam. nov. and Thermovirgaceae fam. nov. are proposed.

Anaerohalosphaera lusitana gen. nov., sp. nov., and Limihaloglobus sulfuriphilus gen. nov., sp. nov., isolated from solar saltern sediments, and proposal of Anaerohalosphaeraceae fam. nov. within the order Sedimentisphaerales.

Two strains of anaerobic, coccoid, saccharolytic, Gram-stain-negative bacteria were isolated from samples of anoxic hypersaline sediments of evaporation ponds in Tavira (Portugal) and Mallorca (Spain). Both isolates were moderately halophilic, neutrophilic and had a temperature optimum at 37 °C. The highest 16S rRNA gene sequence identity values were found with members of the genus Sedimentisphaera (84.9-88.2 %) within the order Sedimentisphaerales, class Phycisphaerae. The strain SM-Chi-D1T could be assigned to the family Sedimentisphaeraceae, while phylogenetic analyses based on 16S rRNA gene sequences and genomic data indicate that strain ST-NAGAB-D1T is both a member of a novel genus and a novel family. SM-Chi-D1T could be distinguished from other cultured members of the Sedimentisphaeraceae mainly by the stimulatory effect of sulfur on growth, lack of ethanol production during fermentation and several differences in the cellular fatty acids and polar lipids patterns. Main differential characteristics of ST-NAGAB-D1T were a polytrichous flagellation, the absence of branched chain fatty acids and presence of large proportions of the unsaturated cellular fatty acids C16 : 1 c9 and C18 : 1 c11. On the basis of genomic, chemotaxonomic, biochemical and physiological data, we propose the novel species and genera Anaerohalosphaera lusitana gen. nov., sp. nov., and Limihaloglobus sulfuriphilus gen. nov., sp. nov., represented by the type strains ST-NAGAB-D1T (=DSM 103484T=JCM 31926T=KCTC 15600T) and SM-Chi-D1T (=DSM 100118T=JCM 31927T=KCTC 15601T), respectively. In addition, we propose the novel family Anaerohalosphaeraceae fam. nov. to accommodate the genus Anaerohalosphaera.

Multidrug-resistant Opportunistic and Pathogenic Bacteria Contaminate Algerian Banknotes Currency.

Currency is one of the most exchanged items in human communities as it is used daily in exchange for goods and services. It is handled by persons with different hygiene standards and can transit in different environments. Hence, money can constitute a reservoir for different types of human pathogens. This study aimed to evaluate the potential of Algerian banknotes to shelter opportunistic pathogenic and multiresistant bacteria. To that end, 200 circulating notes of four different denominations were collected from various places and analyzed for their bacterial loads and contents. Besides, predominant strains were identified and characterized by biochemical and molecular methods, and their resistance profiles against 34 antibiotics were determined. Our results indicated that 100% of the studied banknotes were contaminated with bacteria. The total bacterial concentrations were relatively high, and different bacterial groups were grown, showing important diversity. In total, 48 predominant strains were identified as belonging to 17 genera. Staphylococcus and Micrococcus were the most prevalent genera, followed by Bacillus, Pseudomonas, and Acinetobacter. Antibiotic susceptibility testing showed that all the isolates harbored resistance to at least two molecules, and worrying resistance levels were observed. These findings prove that Algerian currency harbors opportunistic multiresistant bacteria and could potentially act as a vehicle for the spread of bacterial diseases and as a reservoir for antibiotic resistance genes among the community. Therefore, no cash payment systems should be developed and generalized to minimize cash handling and subsequent potential health risks.

Purification and biochemical characterization of a novel acido-halotolerant and thermostable endochitinase from Melghiribacillus thermohalophilus strain Nari2AT.

An extracellular acido-thermostable endochitinase (called ChiA-Mt45) from thermohalophilic Melghiribacillus thermohalophilus strain Nari2AT gen. nov. sp. nov., was purified and biochemically characterized. The maximum chitinase activity recorded after 48-h of incubation at 55 °C was 9000 U/mL. Pure enzyme was obtained after heat treatment (20 min at 90 °C) followed by sequential column chromatographies on fast performance liquid chromatography (FPLC) and high performance liquid chromatography (HPLC). Based on MALDI-TOF/MS analysis, the purified enzyme is a monomer with a molecular mass of 45201.10 Da. The 27 residue NH2-terminal sequence of the enzyme showed high homology with Bacillus GH-18 chitinases family. The optimum pH and temperature values for chitinase activity were pH 3.5 and 90 °C, respectively. In addition, the enzyme was halotolerant and can be classified as an extremozyme. The pure enzyme was completely inhibited by p-chloromercuribenzoic acid (p-CMB) and N-ethylmaleimide (NEM). Its Km and kcat values were 0.253 mg colloidal chitin/mL and 47000 s-1, respectively. Interestingly, its catalytic efficiency was higher than those of chitinases ChiA-Hh59 from Hydrogenophilus hirchii KB-DZ44 and chitodextrinase from Streptomyces griseus, and N-acetyl-ß-glucosaminidase from Trichoderma viride. The studied chitinase exhibited high activity towards colloidal chitin, chitin azure, glycol chitin, while it did not hydrolyse chitibiose and amylose. Additionally, thin-layer chromatography (TLC) analysis from chitin-oligosaccharides showed that ChiA-Mt45 acted as an endosplitting enzyme. Overall, the chitinase ChiA-Mt45 may have great potential for the enzymatic degradation of chitin.

Bacterial diversity obtained by culturable approaches in the gut of Glossina pallidipes population from a non sleeping sickness focus in Tanzania: preliminary results.

BACKGROUND: Glossina pallidipes is a haematophagous insect that serves as a cyclic transmitter of trypanosomes causing African Trypanosomiasis (AT). To fully assess the role of G. pallidipes in the epidemiology of AT, especially the human form of the disease (HAT), it is essential to know the microbial diversity inhabiting the gut of natural fly populations. This study aimed to examine the diversity of G. pallidipes fly gut bacteria by culture-dependent approaches. RESULTS: 113 bacterial isolates were obtained from aerobic and anaerobic microorganisms originating from the gut of G. pallidipes. 16S rDNA of each isolate was PCR amplified and sequenced. The overall majority of identified bacteria belonged in descending order to the Firmicutes (86.6%), Actinobacteria (7.6%), Proteobacteria (5.5%)and Bacteroidetes (0.3%). Diversity of Firmicutes was found higher when enrichments and isolation were performed under anaerobic conditions than aerobic ones. Experiments conducted in the absence of oxygen (anaerobiosis) led to the isolation of bacteria pertaining to four phyla (83% Firmicutes, 15% Actinobacteria, 1% Proteobacteria and 0.5% Bacteroidetes, whereas those conducted in the presence of oxygen (aerobiosis) led to the isolation of bacteria affiliated to two phyla only (90% Firmicutes and 10% Proteobacteria). Phylogenetic analyses placed these isolates into 11 genera namely Bacillus, Acinetobacter, Mesorhizobium, Paracoccus, Microbacterium, Micrococcus, Arthrobacter, Corynobacterium, Curtobacterium, Vagococcus and Dietzia spp.which are known to be either facultative anaerobes, aerobes, or even microaerobes. CONCLUSION: This study shows that G. pallidipes fly gut is an environmental reservoir for a vast number of bacterial species, which are likely to be important for ecological microbial well being of the fly and possibly on differing vectorial competence and refractoriness against AT epidemiology.

Towards a congruent reclassification and nomenclature of the thermophilic species of the genus Pseudothermotoga within the order Thermotogales.

The phylum Thermotogae gathers thermophilic, hyperthermophic, mesophilic, and thermo-acidophilic anaerobic bacteria that are mostly originated from geothermally heated environments. The metabolic and phenotypic properties harbored by the Thermotogae species questions the evolutionary events driving the emergence of this early branch of the universal tree of life. Recent reshaping of the Thermotogae taxonomy has led to the description of a new genus, Pseudothermotoga, a sister group of the genus Thermotoga within the order Thermotogales. Comparative genomics of both Pseudothermotoga and Thermotoga spp., including 16S-rRNA-based phylogenetic, pan-genomic analysis as well as signature indel conservation, provided evidence that Thermotoga caldifontis and Thermotoga profunda species should be reclassified within the genus Pseudothermotoga and renamed as Pseudothermotoga caldifontis comb. nov. (type strain=AZM44c09T) and Pseudothermotoga profunda comb. nov. (type strain=AZM34c06T), respectively. In addition, based upon whole-genome relatedness indices and DNA-DNA Hybridization results, the reclassification of Pseudothermotoga lettingae and Pseudothermotoga subterranea as latter heterotypic synonyms of Pseudothermotoga elfii is proposed. Finally, potential genetic elements resulting from the distinct evolutionary story of the Thermotoga and Pseudothermotoga clades are discussed.

Obligate sugar oxidation in Mesotoga spp., phylum Thermotogae, in the presence of either elemental sulfur or hydrogenotrophic sulfate-reducers as electron acceptor.

Mesotoga prima strain PhosAc3 is a mesophilic representative of the phylum Thermotogae comprising only fermentative bacteria so far. We show that while unable to ferment glucose, this bacterium is able to couple its oxidation to reduction of elemental sulfur. We demonstrate furthermore that M. prima strain PhosAc3 as well as M. prima strain MesG1 and Mesotoga infera are able to grow in syntrophic association with sulfate-reducing bacteria (SRB) acting as hydrogen scavengers through interspecies hydrogen transfer. Hydrogen production was higher in M. prima strain PhosAc3 cells co-cultured with SRB than in cells cultured alone in the presence of elemental sulfur. We propose that the efficient sugar-oxidizing metabolism by M. prima strain PhosAc3 in syntrophic association with a hydrogenotrophic sulfate-reducing bacterium can be extrapolated to all members of the Mesotoga genus. Genome comparison of Thermotogae members suggests that the metabolic difference between Mesotoga and Thermotoga species (sugar oxidation versus fermentation) is mainly due to the absence of the bifurcating [FeFe]-hydrogenase in the former. Such an obligate oxidative process for using sugars, unusual within prokaryotes, is the first reported within the Thermotogae. It is hypothesized to be of primary ecological importance for growth of Mesotoga spp. in the environments that they inhabit.

Desulfonatronum parangueonense sp. nov., a sulfate-reducing bacterium isolated from sediment of an alkaline crater lake.

Novel Gram-stain-negative, non-spore-forming, vibrio-shaped, anaerobic, alkaliphilic, sulfate-reducing bacteria, designated strains PAR180T and PAR190, were isolated from sediments collected at an alkaline crater lake in Guanajuato (Mexico). Strain PAR180T grew at temperatures between 15 and 40 °C (optimum 35 °C), and at pH between 8.3 and 10.4 (optimum 9). It was halotolerant, growing with up to 8 % (w/v) NaCl. Lactate, formate, pyruvate and ethanol were used as electron donors in the presence of sulfate and were incompletely oxidized to acetate and CO2. The isolate was able to grow with hydrogen and with CO2 as a carbon source. Beside sulfate, sulfite and thiosulfate were used as terminal electron acceptors. The isolate was able to grow by disproportionation of sulfite and thiosulfate, but not elemental sulfur, using acetate as a carbon source. The predominant fatty acids were C16 : 0, C16 : 1ω7c and summed feature 10 (C18 : 1ω7c and/or C18 : 1ω9t and/or C18 : 1ω12t). The DNA G+C content was 56.1 mol%. Phylogenetic analysis based on 16S rRNA gene sequences showed that it belongs to the genus Desulfonatronum, class Deltaproteobacteria. Its closest relative is Desulfonatronum thiosulfatophilum (98.7 % 16S rRNA gene sequence similarity). The DNA-DNA relatedness value between strain PAR180T and the type strain of D. thiosulfatophilum was 37.1±2.5 %. On the basis of phylogenetic, phenotypic and chemotaxonomic characteristics, the isolates is considered to represent a novel species of the genus Desulfonatronum, for which the name Desulfonatronum parangueonense sp. nov. is proposed. The type strain is PAR180T (=DSM 103602T=JCM 31598T).