Insight into the symbiotic lifestyle of DPANN archaea revealed by cultivation and genome analyses.

Decades of culture-independent analyses have resulted in proposals of many tentative archaeal phyla with no cultivable representative. Members of DPANN (an acronym of the names of the first included phyla Diapherotrites, Parvarchaeota, Aenigmarchaeota, Nanohaloarchaeota, and Nanoarchaeota), an archaeal superphylum composed of at least 10 of these tentative phyla, are generally considered obligate symbionts dependent on other microorganisms. While many draft/complete genome sequences of DPANN archaea are available and their biological functions have been considerably predicted, only a few examples of their successful laboratory cultivation have been reported, limiting our knowledge of their symbiotic lifestyles. Here, we investigated physiology, morphology, and host specificity of an archaeon of the phylum "Candidatus Micrarchaeota" (ARM-1) belonging to the DPANN superphylum by cultivation. We constructed a stable coculture system composed of ARM-1 and its original host Metallosphaera sp. AS-7 belonging to the order Sulfolobales Further host-switching experiments confirmed that ARM-1 grew on five different archaeal species from three genera-Metallosphaera, Acidianus, and Saccharolobus-originating from geologically distinct hot, acidic environments. The results suggested the existence of DPANN archaea that can grow by relying on a range of hosts. Genomic analyses showed inferred metabolic capabilities, common/unique genetic contents of ARM-1 among cultivated micrarchaeal representatives, and the possibility of horizontal gene transfer between ARM-1 and members of the order Sulfolobales Our report sheds light on the symbiotic lifestyles of DPANN archaea and will contribute to the elucidation of their biological/ecological functions.

Hitherto-Unnoticed Self-Transmissible Plasmids Widely Distributed among Different Environments in Japan.

Various conjugative plasmids were obtained by exogenous plasmid capture, biparental mating, and/or triparental mating methods from different environmental samples in Japan. Based on phylogenetic analyses of their whole-nucleotide sequences, new IncP/P-1 plasmids that could be classified into novel subgroups were obtained. Mini-replicons of the plasmids were constructed, and each of them was incompatible with at least one of the IncP/P-1 plasmids, although they showed diverse iteron sequences in their oriV regions. There were two large clades of IncP/P-1 plasmids, clade I and II. Plasmids in clade I and II included antibiotic resistance genes. Notably, nucleotide compositions of newly found plasmids exhibited different tendencies compared with those of the previously well-studied IncP/P-1 plasmids. Indeed, the host range of plasmids of clade II was different from that of clade I. Although few PromA plasmids have been reported, the number of plasmids belonging to PromAß, and -γ subgroups detected in this study was close to that of IncP/P-1 plasmids. The host ranges of PromAγ and PromAδ plasmids were broad and transferred to different and distinct classes of Proteobacteria. Interestingly, PromA plasmids and many IncP/P-1 plasmids do not carry any accessory genes. These findings indicate the presence of "hitherto-unnoticed" conjugative plasmids, including IncP/P-1 or PromA derivative ones in nature. These plasmids would have important roles in the exchange of various genes, including antibiotic resistance genes, among different bacteria in nature. IMPORTANCE Plasmids are known to spread among different bacteria. However, which plasmids spread among environmental samples and in which environments they are present is still poorly understood. This study showed that unidentified conjugative plasmids were present in various environments. Different novel IncP/P-1 plasmids were found, whose host ranges were different from those of known plasmids, showing wide diversity of IncP/P-1 plasmids. PromA plasmids, exhibiting a broad host range, were diversified into several subgroups and widely distributed in varied environments. These findings are important for understanding how bacteria naturally exchange their genes, including antibiotic resistance genes, a growing threat to human health worldwide.

Nanobdella aerobiophila gen. nov., sp. nov., a thermoacidophilic, obligate ectosymbiotic archaeon, and proposal of Nanobdellaceae fam. nov., Nanobdellales ord. nov. and Nanobdellia class. nov.

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.

Nocardia terrae sp. nov., an actinomycete isolated from soil in Thailand.

A novel actinomycete strain ET3-3T isolated from soil collected from Chachoengsao province, Thailand was taxonomic evaluated using a polyphasic approach. Chemotaxonomic analysis indicated that meso-diaminopimelic acid was the diagnostic diamino acid in the cell-wall peptidoglycan. Mycolic acids were present. Ribose, arabinose, and galactose were detected in its whole-cell hydrolysates. The strain comprised C16:0 and TBSA 10-methyl C18:0 as the main fatty acids and MK-8(H4ω-cycl) as the predominant menaquinone. Major polar lipids were phosphatidylinositol, phosphatidylinositol mannoside, phosphatidylethanolamine, and diphosphatidylglycerol. The results of 16S rRNA gene sequence analysis showed that strain ET3-3T was closely related to Nocardia seriolae JCM 30082T (99.2%), Nocardia yunnanensis JCM 30082T (98.4%), and Nocardia concava JCM 12351T (98.2%). The draft genome of ET3-3T was 9.31 Mb with 8826 coding sequences with an average G + C content of 68.0%. The comparison of the draft genome of strain ET3-3T and N. seriolae NBRC 15557T showed the ANIb, ANIm and the digital DNA-DNA hybridization values of 86.3%, 88.5% and 32.9%, respectively. The results of the taxonomic analysis suggested that strain ET3-3T represented a novel species of the genus Nocardia for which the name Nocardia terrae sp. nov. is proposed. The type strain is ET3-3T (= JCM 33776T = TISTR 2837T).

Actinoplanes lichenicola sp. nov. and Actinoplanes ovalisporus sp. nov., isolated from lichen in Thailand.

Two actinobacteria, designated as strain LDG1-01T and LDG1-06T, were isolated from lichen samples collected in Thailand. Results of morphological characterization, chemotaxonomic studies and 16S rRNA gene analysis indicated that both strains were members of the genus Actinoplanes. MK-9(H4) was found as the major menaquinone. The major fatty acids were anteiso-C15 : 0, iso-C15 : 0, iso-C16 : 0 and anteiso-C17 : 0. Phosphatidylglycerol, diphosphatidylglycerol, phosphatidylethanolamine and phosphatidylinositol were observed as the polar lipids, but differences in minor unidentified polar lipids were found between the strains. Results of comparative genome analysis based on average nucleotide identity and digital DNA-DNA hybridization calculations revealed that both strains showed values below 95 and 70 %, respectively, from each other and closely related Actinoplanes type strains. Based on data from this polyphasic study, strains LDG1-01T and LDG1-06T represent novel species of the genus Actinoplanes. The names proposed are Actinoplanes lichenicola sp. nov. (type strain, LDG1-01T (=JCM 33066T=TISTR 2982T) and Actinoplanes ovalisporus sp. nov. (type strain, LDG1-06T=JCM 33067T=TISTR 2983T).

Actinomadura violacea sp. nov., a madurastatin A1-producing strain isolated from lichen in Thailand.

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).

Adlercreutzia hattorii sp. nov., an equol non-producing bacterium isolated from human faeces.

Two obligately anaerobic, Gram-stain-positive, rod-shaped bacteria were isolated from faecal samples of healthy humans in Japan. 16S rRNA gene sequence analysis indicated that these two strains (8CFCBH1T and 9CBH6) belonged to the genus Adlercreutzia, which is known as an equol-producing bacterium. The closest neighbours of strain 8CFCBH1T were Adlercreutzia equolifaciens subsp. equolifaciens DSM 19450T (98.6%), Adlercreutzia equolifaciens subsp. celatus do03T (98.4%), Adlercreutzia muris WCA-131-CoC-2T (96.6%), Parvibacter caecicola NR06T (96.4%), Adlercreutzia caecimuris B7T (95.3%) and Adlercreutzia mucosicola Mt1B8T (95.3%). The closest relatives to strain 9CBH6 were A. equolifaciens subsp. equolifaciens DSM 19450T (99.8%), A. equolifaciens subsp. celatus do03T (99.6%) and A. muris WCA-131-CoC-2T (96.8%). Strain 8CFCBH1T showed 22.3-53.5% digital DNA-DNA hybridization (dDDH) values with its related species. In addition, the average nucleotide identity (ANI) values between strain 8CFCBH1T and its related species ranged from 75.4 to 93.3%. On the other hand, strain 9CBH6 was considered as A. equolifaciens based on the dDDH and ANI values (>70% dDDH and >95-96% ANI). Strain 9CBH6 showed daidzein-converting activity, as expected from the result of genome analysis. The genome of strain 8CFCBH1T lacked four genes involved in equol production. Growing cells of strain 8CFCBH1T were not capable of converting daidzein. Based on the collected data, strain 8CFCBH1T represents a novel species in the genus Adlercreutzia, for which the name Adlercreutzia hattorii sp. nov. is proposed. The type strain of A. hattorii is 8CFCBH1T (=JCM 34083T=DSM 112284T).

Conexivisphaera calida gen. nov., sp. nov., a thermophilic sulfur- and iron-reducing archaeon, and proposal of Conexivisphaeraceae fam. nov., Conexivisphaerales ord. nov., and Conexivisphaeria class. nov. in the phylum Thaumarchaeota.

A thermoacidophilic, anaerobic, and iron- and sulfur-reducing archaeon, strain NAS-02T, was isolated from a terrestrial hot spring in Japan, as previously reported. This organism is the first non-ammonia-oxidizing isolate in the phylum Thaumarchaeota. Here, we propose Conexivisphaera calida gen. nov., sp. nov. to accommodate this strain. The type strain of the type species is NAS-02T (=JCM 31663T=DSM 105898T). The values of 16S rRNA gene similarity and average amino acid identity between NAS-02T and its closest relatives are <86 and <42 %, respectively. Based on the phylogeny and physiology, we propose the family Conexivisphaeraceae fam. nov., the order Conexivisphaerales ord. nov. and the class Conexivisphaeria class. nov. to accommodate the novel genus.

Oxygen concentration affects frequency and range of transconjugants for the incompatibility (Inc) P-1 and P-7 plasmids pBP136 and pCAR1.

The frequency of transconjugants were compared for the incompatibility (Inc) P-1 and P-7 plasmids pBP136 and pCAR1 under aerobic and anaerobic conditions. Filter mating assays were performed with one donor strain and one recipient strain using different donors of Pseudomonas and recipient strains, including Pseudomonas, Pantoea, and Buttiauxella. Under anaerobic condition, frequencies of transconjugants for both plasmids were 101-103-fold lower than those under aerobic condition regardless of whether aerobically or anaerobically grown donors and recipients were used. To compare the transconjugant ranges under aerobic and anaerobic conditions, conjugation was performed between the donor of pBP136 and recipient bacteria extracted from environmental samples. Several transconjugants were uniquely obtained from each aerobic or anaerobic condition. Our findings indicate that a plasmid can differently spread among bacteria depending on the oxygen concentrations of the environment.

Nonomuraea phyllanthi sp. nov., an endophytic actinomycete isolated from the leaf of Phyllanthus amarus.

A novel actinomycete, strain PA1-10T, isolated from the leaf of Phyllanthus amarus collected from Bangkok, Thailand, was characterized taxonomically using a polyphasic approach. This strain contained the characteristics consistent with those of members of the genus Nonomuraea. It formed short rugose spore chain on aerial mycelium. The diamino acid in cell wall peptidoglycan was meso-diaminopimelic acid. Galactose, glucose, madurose, mannose, and ribose were found in whole-cell hydrolysates. Predominant menaquinones were MK-9 (H2), MK-9 (H4), and MK-9 (H6). Major cellular fatty acids were iso-C16:0 and C17:0 10-methyl. Phospholipid profiles were composed of phosphatidylinositol mannoside (PIM), lyso-phosphatidylethanolamine (lyso-PE), phosphatidylethanolamine (PE), methylphosphatidylethanolamine (PME), diphosphatidylglycerol (DPG), and phosphatidylglycerol (PG). The G + C content of DNA was 71.2 mol%. Strain PA1-10T showed the highest 16S rRNA gene sequence similarity with Nonomuraea candida JCM 15928T (98.35%) and shared the same node with Nonomuraea maritima JCM 18321T in the phylogenetic tree analysis. Based on the phenotypic characteristics, DNA-DNA relatedness, and average nucleotide identity (ANI), the strain is considered to represent a novel species of the genus Nonomuraea, for which the name Nonomuraea phyllanthi is proposed. The type strain is PA1-10T (= JCM 33073T = NBRC 112774T = TISTR 2497T).

Thermaurantimonas aggregans gen. nov., sp. nov., a moderately thermophilic heterotrophic aggregating bacterium isolated from microbial mats at a terrestrial hot spring.

A moderately thermophilic and heterotrophic bacterial strain, LAT, was isolated from microbial mats sampled at a hot spring in Nakabusa, Nagano, Japan. The cells of strain LAT were aerobic, Gram-stain-negative, non-sporulating, non-motile and rod-shaped (2.0-4.1 µm long). The formation of dense cell aggregates in liquid medium was a unique characteristic of the strain. Strain LAT grew optimally at 50°C and at pH 7.5. Phylogenetic analysis based on the 16S rRNA gene sequence revealed that the nearest neighbour of strain LAT was Schleiferia thermophila JCM 30197T with 94.1 % sequence similarity. The draft genome sequence of strain LAT (2 671 880 bp) showed 18.0 % digital DNA-DNA hybridization, 70.9 % average nucleotide identity and 72.1 % average amino acid identity (AAI) values in comparison with the genome sequence of S. thermophila JCM 30197T (2 606 763 bp); the 16S rRNA gene sequence similarity and AAI values are lower than the cutoffs used for assignment to a separate genus. On the basis of phenotypic features, major cellular fatty acid composition, genome sequencing and phylogenetic position, a novel genus and species are proposed for strain LAT, to be named Thermaurantimonas aggregans (= JCM 31826T=DSM 106522T).

Nocardia aurantiaca sp. nov., isolated from soil in Thailand.

A novel actinomycete strain, CT2-14T, belonging to the genus Nocardia, was isolated from a soil sample collected from Phichit Province, Thailand. The taxonomic position of the strain was characterized using a polyphasic approach. The strain grew at 15-40 °C (optimum, 28-37 °C), pH 6-11 (optimum, pH 6-8) and on an International Streptomyces Project 2 with 4 % (w/v) NaCl agar plate. Meso-diaminopimelic acid was detected in the cell-wall peptidoglycan. Ribose, arabinose and galactose were detected in its whole-cell hydrolysates. Mycolic acids were present. The strain contained C16 : 0, summed feature 3, C17 : 0 10-methyl and C18 : 1 ω9c as the major fatty acids and MK-8(H4ω-cycl) as the major menaquinone. The major polar lipids were phosphatidylethanolamine, diphosphatidylglycerol and phosphatidylinositol mannosides. Strain CT2-14T showed the highest 16S rRNA gene similarity to Nocardia veterana JCM 11307T (98.4 %), Nocardia africana JCM 11438T (98.2 %) and Nocardia kruczakiae JCM 13032T (98.0 %). The draft genome of strain CT2-14T was 7.37 Mb with 6685 coding sequences with an average G+C content of 67.9 mol %. Based on the phylogenomic tree analysis, the strain was closely related to Nocardia niigatensis NBRC 100131T. On the basis of polyphasic and genome analyses, strain CT2-14T represented a novel species of the genus Nocardia for which the name Nocardia aurantiaca sp. nov. is proposed. The type strain is CT2-14T (=JCM 33775T=TISTR 2838T).

Streptomyces mimosae sp. nov., an endophytic actinomycete isolated from the root of Mimosa pudica in Thailand.

An endophytic actinomycete, strain 3MP-10T, isolated from the root of Mimosa pudica was taxonomically studied based upon polyphasic approaches. This strain formed spiral spore chains on aerial mycelia. ll-Diaminopimelic acid, glucose and ribose were found in the whole-cell hydrolysates. It belonged to the genus Streptomyces and was closely related to Streptomyces zhaozhouensis DSM 42101T (98.9 %) and Streptomyces sedi JCM 16909T (98.6 %) based on 16S rRNA gene sequence analysis results. The major menaquinones were MK-10(H8), MK-10(H6) and MK-9(H8). The predominant cellular fatty acids were iso-C16 : 0, anteiso-C15 : 0 and anteiso-C17 : 0. The detected phospholipids were diphosphatidylglycerol, phosphatidylinositol mannoside, phosphatidylinositol, phosphatidylethanolamine and phosphatidylglycerol. Strain 3MP-10T had a genome size of 7.2 Mb with a genome G+C content of 73.4 mol%. Results of in silico genome-based similarity analysis revealed ANIb values of 84.94 and 84.77 %, ANIm values of 88.01 and 87.92 %, and dDDH values of 29.9 and 29.6 % when compared with S. zhaozhouensis DSM 42101T and S. sedi JCM 16909T, respectively. Based on the polyphasic approach, digital DNA-DNA relatedness and average nucleotide identity, we propose that the novel actinomycete represents a novel species, Streptomyces mimosae, with type strain 3MP-10T (=JCM 33328T=TISTR 2646T).

Streptomyces bauhiniae sp. nov., isolated from tree bark of Bauhinia variegata Linn. in Thailand.

A novel actinomycete strain, Bv016T, belonging to the genus Streptomyces, was isolated from the bark of tree, Bauhinia variegata Linn., collected in Thailand. The taxonomic position of the strain was characterized by using a polyphasic approach. ll-Diaminopimelic acid, glucose, mannose, ribose and galactose were detected in its whole-cell hydrolysates. The N-acyl type of muramic acid was acetyl. The strain contained anteiso-C15  :  0, iso-C16  :  0, iso-C15  :  0 and iso-C14  :  0 as the major fatty acids and MK-9(H8), MK-9(H6) and MK-9(H4) as the major menaquinones. The major polar lipids were diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, phosphatidylinositol and phosphatidylinositol mannoside. The strain was closely related to Streptomyces griseoluteus JCM 4041T (98.4 %), Streptomyces seoulensis JCM 10116T (98.4 %) and Streptomyces recifensis JCM 4408T (98.2 %). The draft genome of Bv016T was 6.74 Mb with 5949 coding sequences with an average G+C content of 71.7 mol%. The ANIb and ANIm values of strain Bv016T were 94.1 and 95.2 %, respectively, and the digital DNA-DNA hybridization value was 60.6 % in comparison with the draft genome of S. griseoluteus JCM 4765T. The results of the taxonomic analysis suggested that strain Bv016T represented a novel species of the genus Streptomyces for which the name Streptomyces bauhiniae sp. nov. is proposed. The type strain is Bv016T (=JCM 33208T=TISTR 2645T).

Microbispora catharanthi sp. nov., a novel endophytic actinomycete isolated from the root of Catharanthus roseus.

Strain CR1-09T, an actinomycete isolated from the root of Catharanthus roseus, was taxonomically studied based upon polyphasic approaches. The isolate formed a pair of ovular to circular, smooth-surfaced spores on short sporophores alternately branched from aerial mycelia. It contained meso-diaminopimelic acid in cell wall peptidoglycans. The major menaquinones were MK-9 (H4) and MK-9 (H2). The predominant cellular fatty acids were iso-C16 : 0 and C16 : 0. The polar lipids profile consisted of diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, hydroxyl phosphatidylethanolamine, phosphatidylinositol, phosphatidylinositol mannosides, and unidentified ninhydrin positive phosphoglycolipids. Strain CR1-09T showed the highest 16S rRNA gene sequence similarity with Microbispora tritici DSM 104650T (99.5 %). Based on the polyphasic approach, DNA-DNA relatedness and average nucleotide identity (ANI), the strain is considered to represent a novel species of the genus Microbispora, for which the name Microbispora catharanthi is proposed. The type strain is strain CR1-09T (=JCM 30045T=TISTR 2273T).

Cutibacterium modestum sp. nov., isolated from meibum of human meibomian glands, and emended descriptions of Cutibacterium granulosum and Cutibacterium namnetense.

An anaerobic and aerotolerant bacterium, strain M12T, was isolated from the meibum of inflamed human meibomian glands. Cells of the strain was Gram-stain-positive, non-spore-forming and non-motile rods. Growth on trypticase soy agar plates supplemented with 5 % sheep blood was fastest at 30-37 °C under anaerobic conditions. The 16S rRNA gene sequence of the strain revealed that it belongs to the genus Cutibacterium with a 98.0 % similarity value to the closest species, Cutibacterium acnes. Genome analysis of the strain with type strains of the other Cutibacterium species resulted in digital DNA-DNA hybridization values of 32.3-22.3% and average nucleotide identity (OrthoANI) values of 86.7-73.6 %. Biochemical and physiological analyses using API rapid ID 32A and API Coryne kits revealed relatively low reactivity of the strain compared with C. acnes and Cutibacterium namnetense. The most abundant major cellular fatty acid was iso-C15 : 0. Fermentation end-products from glucose were propionate, lactate, succinate and acetate. The diagnostic diamino acid of the peptidoglycan was meso-diaminopimelic acid. Major menaquinones were MK-9(H4), MK-9(H2) and MK-9. The major peaks of the MALDI-TOF mass spectrometry spectrum were at 3493, 3712, 6986 and 7424 Da. The DNA G+C content was 59.9 mol%. Based on these findings, we propose a novel species, Cutibacterium modestum. The type strain of C. modestum is M12T (=JCM 33380T=DSM 109769T). On the basis of further genomic analysis, we also provide emended descriptions of Cutibacterium granulosum (Prévot 1938) Scholz and Kilian 2016 and Cutibacterium namnetense (Aubin et al. 2016) Nouioui et al. 2018.

Lactococcus insecticola sp. nov. and Lactococcus hodotermopsidis sp. nov., isolated from the gut of the wood-feeding lower termite Hodotermopsis sjostedti.

Two strains of lactic acid bacteria, designated Hs20B0-1T and Hs30E4-3T, were isolated from the gut of the damp-wood termite Hodotermopsis sjostedti. These strains were characterized genetically and phenotypically. Strain Hs20B0-1T was related to Lactococcus piscium DSM 6634T showing 96.3 and 84.2 % sequence similarity in 16S rRNA gene and rpoB gene sequences, respectively. Strain Hs30E4-3T was related to Lactococcus plantarum DSM 20686T showing 94.8 and 82.2 % sequence similarity in 16S rRNA gene and rpoB gene sequences, respectively. The 16S rRNA gene sequence similarity between strains Hs20B0-1T and Hs30E4-3T was 95.7 %. Furthermore, genomic comparisons using pairwise average nucleotide identity (ANI) and digital DNA-DNA hybridization (DDH) analyses between strain Hs20B0-1T and L. piscium DSM 6634T resulted in values of 73.5 and 20.1 %, respectively. Strain Hs30E4-3T had 72.8 % ANI similarity and 21.3 % DDH similarity to L. plantarum DSM 20686T. Strains Hs20B0-1T and Hs30E4-3T had 75.4 % ANI similarity and 21.1 % DDH similarity to each other. The cell-wall peptidoglycan types of strains Hs20B0-1T and Hs30E4-3T were A4α, Lys-Asp and A3α, Lys-Thr-Ala, respectively. The two strains, Hs20B0-1T and Hs30E4-3T, are distinguishable from each other and other established Lactococcus species phylogenetically and phenotypically. In conclusion, two novel species of the genus Lactococcus are proposed, namely Lactococcus insecticola Hs20B0-1T (=JCM 33485T=DSM 110147T) and Lactococcus hodotermopsidis Hs30E4-3T (=JCM 33486T=DSM 110148T), respectively.

Comparative genomics of Parolsenella catena and Libanicoccus massiliensis: Reclassification of Libanicoccus massiliensis as Parolsenella massiliensis comb. nov.

Previous analyses based on 16S rRNA and hsp60 genes indicated that Parolsenella catena and Libanicoccus massiliensis were closely related to each other and formed a monophyletic cluster independent of the related Olsenella species. To clarify the relationship of these two species, we determined the genome sequence of P. catena JCM 31932T and compared it with that already sequenced for L. massiliensis Marseille-P3237T. Phylogenetic trees based on the concatenated 37 single-copy ribosomal proteins or RpoB robustly supported the relationship observed in the previous studies. Digital DNA-DNA hybridization and average nucleotide identity (ANI) values between P. catena JCM 31932T and L. massiliensis Marseille-P3237T were 32.6 and 87.8 %, respectively, indicating that P. catena JCM 31932T and L. massiliensis Marseille-P3237T are independent species. Alignment fraction and ANI values between the two genomes were 0.75 and 88.84 %, respectively, thus indicating that the two species should be classified into the same genus. The number of putative orthologous genes shared between the two genomes was 1321, which was significantly larger than those (482-928) reported between L. massiliensis Marseille-P3237T and other closely related species. In addition, the genome of P. catena JCM 31932T had a high degree of synteny conservation with that of L. massiliensis Marseille-P3237T. On the basis of these findings, we propose that L. massiliensis should be reclassified as Parolsenella massiliensis comb. nov.; the type strain is Marseille-P3237T (=JCM 33000T=CCUG 71182T).

Micromonospora radicis sp. nov., isolated from roots of Azadirachta indica var. siamensis Valenton, and reclassification of Jishengella zingiberis as Micromonospora zingiberis comb. nov.

A novel endophytic actinomycete strain AZ1-13T was isolated from roots of Azadirachta indica, and its taxonomic position was investigated using a polyphasic approach. Pairwise 16S rRNA gene sequence similarities of strain AZ1-13T and its closest species, Jishegella zingiberis PLAI1-1T and Micromonospora endophytica 202201T, were 99.7 and 99.2 %, respectively. Phylogenetic analyses of the family Micromonosporaceae based on 16S rRNA gene sequences indicated strains AZ1-13T and J. zingiberis PLAI1-1Tare located within the genus Micromonospora. The approximate genome size of the strain was 5.96 Mb with 71.9 mol% of G+C content. The strain AZ1-13T exhibited ANIb values of 87.4 % with J. zingiberis PLAI1-1T and 85.1 % with M. endophytica 202201T. Chemotaxonomic characteristics of strain AZ1-13T were consistent within the genus Micromonospora: cell-wall peptidoglycan of the strain contained meso-diaminopimelic acid; glucose, mannose, ribose and xylose are presented as the whole-cell sugars; the predominant menaquinones were MK-9(H4) and MK-9(H6); major cellular fatty acids were iso-C15 : 0, 10-methyl C17 : 0, C17 : 0, anteiso-C17 : 0 and iso-C17 : 1ω8c; diphosphatidylglycerol, phosphatidylethanolamine and phosphatidylinositol were detected as distinguished phospholipids. Based on phenotypic properties, phylogeny and genomic data, the strain AZ1-13T could be distinguished from its closest neighbours, representing a novel species of the genus Micromonospora, for which the name Micromonospora radicis sp. nov. is proposed. The type strain is AZ1-13T (=KCTC 39786T=NBRC 112324T=JCM 32147T = TISTR 2404T). This study also proposed that J. zingiberisis transferred to the genus Micromonospora as Micromonospora zingiberis comb. nov. (type strain PLAI1-1T=TBRC 7644T=NBRC 113144T=JCM 32592T).

Proteolytic analysis of Trichoderma reesei in celluase-inducing condition reveals a role for trichodermapepsin (TrAsP) in cellulase production.

Filamentous fungi produce a variety of proteases with significant biotechnological potential and show diverse substrate specificities. Proteolytic analysis of the industrial enzyme producer Trichoderma reesei has been sparse. Therefore, we determined the substrate specificity of T. reesei secretome and its main protease Trichodermapepsin (TrAsP) up to P1 position using FRETS-25Xaa-libraries. The role of TrAsP was analyzed using T. reesei QM9414 and the deletant QM∆trasp in Avicel. We observed higher activities of CMCase, Avicelase, and Xylanase in QM∆trasp compared to that of QM9414. Saccharification rate of cellulosic biomass also increased when using secretome of QM∆trasp but the effect was not significant due to the absence of difference in BGL activity compared to QM9414. Higher TrAsP was produced when monosaccharides were used as a carbon source compared to cellulase inducers such as Avicel and α-sophorose. These results elucidate the relationship between TrAsP and cellulase production in T. reesei and suggest a physiological role for TrAsP.