RESULTS:

A Gram-stain-negative endospore-forming rod-shaped indole-producing bacterial strain designated YZC6T was isolated from fermented cabbage. Strain YZC6T grew at 10–37  °C pH 5.5–8.5 and with up to 2  % (w/v) NaCl. The major cellular fatty acids were C16 : 0 and C18 : 1 cis 11 dimethyl acetal. Phylogenetic analysis of the 16S rRNA gene revealed that strain YZC6T belonged to the genus Lacrimispora and was closely related to Lacrimispora aerotolerans DSM 5434T (98.3  % sequence similarity) Lacrimispora saccharolytica WM1T (98.1  %) and Lacrimispora algidixylanolytica SPL73T (98.1  %). The average nucleotide identity based on blast (below 87.8  %) and digital DNA–DNA hybridization (below 36.1 %) values between the novel isolate and its corresponding relatives showed that strain YZC6T could be readily distinguished from its closely related species. Based on genotypic phenotypic and chemotaxonomic data a novel Lacrimispora species Lacrimispora brassicae sp. nov. was proposed with YZC6T as the type strain (=MAFF 212518T=JCM 32810T=DSM 112100T). This study also proposed Clostridium indicum Gundawar et al. 2019 as a later heterotypic synonym of Lacrimispora amygdalina (Parshina et al. 2003) Haas and Blanchard 2020 and Clostridium methoxybenzovorans Mechichi et al. 1999 as a later heterotypic synonym of Lacrimispora indolis (McClung and McCpy 1957) Haas and Blanchard 2020.

A neutrophilic iron-oxidizing and -reducing bacterium strain MIZ03T was previously isolated from a wetland in Ibaraki Japan. Here we report the detailed characteristics of this strain. It was motile with a single polar flagellum and Gram-stain-negative. It could grow not only chemolithoautotrophically but also chemoorganotrophically by aerobic respiration and fermentation. Major cellular fatty acids were C16 : 1  ω7c/C16 : 1  ω6c and C16 : 0. Phylogenetic analyses indicated that strain MIZ03T belonged to the genus Rhodoferax. This strain was closely related to Rhodoferax ferrireducens with 98.5 % of 16S rRNA gene sequence similarity. Based on its phenotypic and genomic based characteristics we conclude that strain MIZ03T represents a new species in the genus Rhodoferax. We propose the name Rhodoferax lithotrophicus sp. nov. to accommodate this strain. The type strain is MIZ03T (=JCM 34246T=DSM 113266T). We also propose the name Rhodoferax koreensis sp. nov. of which the type strain is DCY110T (=KCTC 52288T=JCM 31441T) for the effectively but not yet validly published name ‘Rhodoferax koreense’.

Obligately anaerobic Gram-stain-positive bacilli strains 12BBH14T 9CFEGH4 and 10CPCBH12 were isolated from faecal samples of healthy Japanese people. Strain 12BBH14T showed the highest 16S rRNA gene sequence similarity to Sellimonas monacensis Cla-CZ-80T (97.5 %) and ‘Lachnoclostridium phocaeense’ Marseille-P3177T (97.2 %). Strain 12BBH14T was also closely related to Eubacterium sp. c-25 with 99.7 % 16S rRNA gene sequence similarity. The 16S rRNA gene sequence analysis showed that strains 12BBH14T 9CFEGH4 and 10CPCBH12 formed a monophyletic cluster with Eubacterium sp. c-25. Near this monophyletic cluster S. monacensis Cla-CZ-80T and ‘L. phocaeense’ Marseille-P3177T formed a cluster and did not form a cluster with other Sellimonas species. The digital DNA–DNA hybridization (dDDH) and average nucleotide identity (ANI) values between strains 12BBH14T 9CFEGH4 10CPCBH12 and Eubacterium sp. c-25 were higher than the cut-off values of species demarcation (>88 % dDDH and >98 % ANI) indicating that these four strains are the same species. On the other hand the dDDH and ANI values of these strains were lower than the cut-off values of species demarcation against other strains (<29 % dDDH and <76 % ANI). Moreover the average amino acid identity values among these strains were higher than the genus boundary. These results indicate that the isolates should be considered to belong to a new genus of the family Lachnospiraceae . Based on the collected data strains 12BBH14T 9CFEGH4 and 10CPCBH12 represent a novel species of a novel genus for which the name Claveliimonas bilis gen. nov. sp. nov. is proposed. The type strain of C. bilis is 12BBH14T (=JCM 35899T=DSM 115701T). Eubacterium sp. c-25 belongs to C. bilis. In addition S. monacensis is transferred to the genus Claveliimonas as Claveliimonas monacensis comb. nov.

Nucleotide sequence similarity including k-mer plasmid composition has been used for prediction of plasmid evolutionary host range representing the hosts in which a plasmid has replicated at some point during its evolutionary history. However the relationships between the bacterial taxa of experimentally identified transconjugants and the predicted evolutionary host ranges are poorly understood. Here four different PromA group plasmids showing different k-mer compositions were used as model plasmids. Filter mating assays were performed with a donor harbouring plasmids and recipients of bacterial communities extracted from environmental samples. A broad range of transconjugants was obtained with different bacterial taxa. A calculation of the dissimilarities in k-mer compositions as Mahalanobis distance between the plasmid and its sequenced transconjugant chromosomes revealed that each plasmid and transconjugant were significantly more similar than the plasmid and other non-transconjugant chromosomes. These results indicate that plasmids with different k-mer compositions clearly have different host ranges to which the plasmid will be transferred and replicated. The similarity of the nucleotide compositions could be used for predicting not only the plasmid evolutionary host range but also future host ranges.

Obligately anaerobic Gram-stain-positive small-chain coccobacilli strains 12EGH17T and 18CBH55 were isolated from faecal samples of healthy Japanese humans. Strain 12EGH17T showed the highest 16S rRNA gene sequence similarity to Sellimonas intestinalis BR72T (95.5 %) Coprococcus comes ATCC 27758T (94.4 %) and Clostridium nexile DSM 1787T (93.7 %). The percentage of conserved proteins values between the genome of strain 12EGH17T and that of the members of the genus Sellimonas were >54 % suggesting that strain 12EGH17T belongs to the genus Sellimonas . The digital DNA–DNA hybridization (dDDH) and average nucleotide identity (ANI) values between strains 12EGH17T and 18CBH55 were higher than the cut-off values of species demarcation (90 % dDDH and 99 % ANI) indicating these two strains are the same species. However the dDDH and ANI values of these strains were lower than the cut-off values of species demarcation against other strains (<30 % dDDH and <79 % ANI). These results indicate that the isolates should be considered to represent a new species of the genus Sellimonas . The isolates were differentiated from the type species S. intestinalis by the ability of aesculin hydrolysis. Based on the collected data strains 12EGH17T and 18CBH55 represent a novel species in the genus Sellimonas for which the name Sellimonas catena sp. nov. is proposed. The type strain of S. catena is 12EGH17T (=JCM 35622T=DSM 114916T).

A co-culture of a novel thermoacidophilic obligate symbiotic archaeon designated as strain MJ1T with its specific host archaeon Metallosphaera sedula strain MJ1HA was obtained from a terrestrial hot spring in Japan. Strain MJ1T grew in the co-culture under aerobic conditions. Coccoid cells of strain MJ1T were 200–500 nm in diameter and attached to the MJ1HA cells in the co-culture. The ranges and optima of the growth temperature and pH of strain MJ1T in the co-culture were 60–75 °C (optimum 65–70 °C) and pH 1.0–4.0 (optimum pH 2.5) respectively. Core lipids of dialkyl glycerol tetraethers (GDGT)−3 and GDGT-4 were highly abundant in MJ1T cells concentrated from the co-culture. Strain MJ1T has a small genome (0.67 Mbp) lacking genes for biosynthesis of essential biomolecules such as nucleotides lipids and ATP. The genomic DNA G+C content was 24.9 mol%. The 16S rRNA gene sequence of strain MJ1T was most closely related to that of the cultivated species ‘Nanopusillus acidilobi’ strain N7A (85.8 % similarity). Based on phylogenetic and physiological characteristics we propose the name Nanobdella aerobiophila gen. nov. sp. nov. to accommodate the strain MJ1T (=JCM 33616T=DSM 111728T). In addition we propose the names Nanobdellaceae fam. nov. Nanobdellales ord. nov. and Nanobdellia class. nov. to accommodate the novel genus.

Faecalibacterium prausnitzii is one of the most important butyrate-producing bacteria in the human gut. Previous studies have suggested the presence of several phylogenetic groups with differences at the species level in the species and a taxonomic re-evaluation is thus essential for further understanding of ecology of the important human symbiont. Here we examine the phenotypic physiological chemotaxonomic and phylogenomic characteristics of six F. prausnitzii strains (BCRC 81047T=ATCC 27768T A2-165T=JCM 31915T APC918/95b=JCM 39207 APC942/30−2=JCM 39208 APC924/119=JCM 39209 and APC922/41−1T=JCM 39210T) deposited in public culture collections with two reference strains of Faecalibacterium butyricigenerans JCM 39212T and Faecalibacterium longum JCM 39211T. Faecalibacterium sp. JCM 17207T isolated from caecum of broiler chicken was also included. Three strains of F. prausnitzii (BCRC 81047T JCM 39207 and JCM 39209) shared more than 96.6 % average nucleotide identity (ANI) and 69.6 % digital DNA–DNA hybridization (dDDH) values indicating that the three strains are members of the same species. On the other hand the remaining three strains of F. prausnitzii (JCM 31915T JCM 39208 and JCM 39210T) were clearly separated from the above three strains based on the ANI and dDDH values. Rather JCM 39208 showed ANI and dDDH values over the cut-off values of species discrimination (>70 % dDDH and >95–96 % ANI) with F. longum JCM 39211T whereas JCM 31915T JCM 39210T and JCM 17207T did not share dDDH and ANI values over the currently accepted cut-off values with any of the tested strains including among them. Furthermore the cellular fatty acid patterns of these strains were slightly different from other F. prausnitzii strains. Based on the collected data F. prausnitzii JCM 31915T F. prausnitzii JCM 39210T and Faecalibacterium sp. JCM 17207T represent three novel species of the genus Faecalibacterium for which the names Faecalibacterium duncaniae sp. nov. (type strain JCM 31915T=DSM 17677T=A2-165T) Faecalibacterium hattorii sp. nov. (type strain JCM 39210T=DSM 107841T=APC922/41-1T) and Faecalibacterium gallinarum sp. nov. (type strain JCM 17207T=DSM 23680T=ic1379T) are proposed.

In Japan during a screening of lactic acid bacteria in spent mushroom substrates an unknown bacterium was isolated and could not be assigned to any known species. Strain YK48GT is Gram-stain-positive rod-shaped non-motile non-spore-forming and catalase-negative. The isolate grew in 0–4 % (w/v) NaCl at 15–37 °C (optimum 30 °C) and at pH 4.0–8.0 (optimum pH 6.0). The genomic DNA G+C content of strain YK48GT was 42.5 mol%. Based on its 16S rRNA gene sequence strain YK48GT represented a member of the genus Lentilactobacillus and showed the highest pairwise similarity to Lentilactobacillus rapi DSM 19907T (97.86 %). Phylogenetic analyses based on amino acid sequences of 466 shared protein-encoding genes also revealed that the strain was phylogenetically positioned in the genus Lentilactobacillus but did not suggest an affiliation with previously described species. The average nucleotide identity and digital DNA–DNA hybridization values between strain YK48GT and the type strains of phylogenetically related species were 72.2–76.6% and 19.0–21.2 % respectively indicating that strain YK48GT represents a novel species within the genus Lentilactobacillus . Phenotypic data further confirmed the differentiation of strain YK48GT from other members of the genus Lentilactobacillus . According to the results of the polyphasic characterization presented in this study strain YK48GT represents a novel species of the genus Lentilactobacillus for which the name Lentilactobacillus fungorum sp. nov. is proposed. The type strain is YK48GT (=JCM 32598T=DSM 107968T).

An actinomycete strain LCR2-06T isolated from a lichen sample on rock collected from Chiang Rai Province (Pong Phra Bat Waterfall) Thailand was characterized using a polyphasic approach. The strain grew at 25–45 °C pH 6–11 and on International Streptomyces Project 2 agar plate with 5 % (w/v) NaCl. It contained meso-diaminopimelic acid as the diamino acid in whole-cell hydrolysates. Rhamnose ribose xylose madurose glucose and galactose were detected as whole-cell sugar hydrolysates. Mycolic acids were absent. The N-acyl type of muramic acid was acetyl. The strain contained C16 : 0 TBSA 10-methyl C18 : 0 and 2-hydroxy C16 : 0 as the predominant fatty acids and MK-9(H6) MK-9(H4) and MK-9(H8) as the major menaquinones. The major polar lipids were diphosphatidylglycerol phosphatidylinositol and unidentified phospholipid. The draft genome of strain LCR2-06T was closely related to Actinomadura barringtoniae TBRC 7225T (99.2 %) Actinomadura nitritigenes NBRC 15918T (98.8 %) Actinomadura montaniterrae TISTR 2400T (98.5 %) and Actinomadura physcomitrii JCM 33455T (97.9 %). The draft genome of LCR2-06T was 11.1 Mb with 10 588 coding sequences with an average G+C content of 72.7 mol%. Results of genomic analysis revealed that the ANIb and ANIm values between strain LCR2-06T and A. montaniterrae TISTR 2400T were 90.0 and 92.0 % respectively. The digital DNA–DNA hybridization value was 43.9 % in comparison with the draft genome of A. montaniterrae TISTR 2400T. The strain produced an antibacterial compound active against Bacillus subtilis ATCC 6633 and Kocuria rhizophila ATCC 9341. The results of taxonomic analysis suggested that strain LCR2-06T represented a novel species of the genus Actinomadura for which the name Actinomadura violacea sp. nov. is proposed. The type strain is LCR2-06T (=JCM 33065T=KCTC 49547T=NBRC 114810T=LMG 32136T=TISTR 2935T).

A novel nitrogen-fixing fermentative bacterium designated as YA01T was isolated from Nakabusa hot springs in Japan. The short-rod cells of strain YA01T were Gram-positive and non-sporulating. Phylogenetic trees of the 16S rRNA gene sequence and concatenated sequences of 40 single-copy ribosomal genes revealed that strain YA01T belonged to the genus Caldicellulosiruptor and was closely related to Caldicellulosiruptor hydrothermalis 108T Caldicellulosiruptor bescii DSM 6725T and Caldicellulosiruptor kronotskyensis 2002T. The 16S rRNA gene sequence of strain YA01T shares less than 98.1 % identity to the known Caldicellulosiruptor species. The G+C content of the genomic DNA was 34.8 mol%. Strain YA01T shares low genome-wide average nucleotide identity (90.31–91.10 %) average amino acid identity (91.45–92.10 %) and <70 % digital DNA–DNA hybridization value (41.8–44.2 %) with the three related species of the genus Caldicellulosiruptor . Strain YA01T grew at 50–78 °C (optimum 70 °C) and at pH 5.0–9.5 (optimum pH 6.5). Strain YA01T mainly produced acetate by consuming d(+)-glucose as a carbon source. The main cellular fatty acids were iso-C17 : 0 (35.7 %) C16 : 0 (33.3 %) DMA16 : 0 (6.6 %) and iso-C15 : 0 (5.9 %). Based on its distinct phylogenetic position biochemical and physiological characteristics and the major cellular fatty acids strain YA01T is considered to represent a novel species of the genus Caldicellulosiruptor for which the name Caldicellulosiruptor diazotrophicus sp. nov. is proposed (type strain YA01T=DSM 112098T=JCM 34253T).