RESULTS:

A polyphasic approach was used to describe strain RB6PN24T a novel actinobacterium isolated from peat swamp forest soil collected from Rayong province Thailand. Phylogenetic analysis based on 16S rRNA gene sequences indicated that the strain belonged to the genus Kitasatospora and showed the highest sequence similarities to Kitasatospora kifunensis IFO 15206T (98.7 %) and Kitasatospora acidiphila MMS16-CNU292T (98.5 %). Strain RB6PN24T contained major amounts of meso-diaminopimelic acid galactose mannose and ribose in the whole-cell hydrolysates. MK-9(H6) and MK-9(H8) were the predominant menaquinones of the micro-organism. The polar lipids consisted of diphosphatidylglycerol phosphatidylethanolamine phosphatidylglycerol phosphatidylinositol phosphatidylinositol mannosides an unidentified lipid four unidentified aminolipids and six unidentified phospholipids. Mycolic acids were not present. The major fatty acids were iso-C15 : 0 iso-C16 : 0 anteiso-C15 : 0 iso-C17:0 anteiso-C17 : 0 and C16 : 0. The draft genome size of strain RB6PN24T was 8.09 Mbp with 72.1 mol% G+C content and predicted to contain at least 44 biosynthetic gene clusters encoding diverse secondary metabolites. Furthermore the strain exhibited low average nucleotide identity and digital DNA–DNA hybridization values with K. acidiphila MMS16-CNU292T (89.1 % 42.4 %) and K. kifunensis DSM 41654T (79.5 % 25.5 %). The results of phenotypic chemotaxonomic genotypic and phylogenetic analyses revealed that strain RB6PN24T represents a novel species of the genus Kitasatospora for which the name Kitasatospora humi sp. nov. is proposed. The type strain is RB6PN24T (=TBRC 14818T=NBRC 115116T). In addition the comparison of the whole genome sequences and phenotypic features suggested that Kitasatospora aureofaciens and Kitasatospora psammotica belong to the same species. Therefore it is proposed that K. psammotica is reclassified as a later heterotypic synonym of K. aureofaciens .

The taxonomic position of a novel actinomycete designated strain RS10V-4T was determined using a polyphasic approach. Strain RS10V-4T was isolated from paddy rhizosphere soil of rice plant (Oryzae sativa L.). The morphological physiological and chemotaxonomic properties were consistent with its classification in the genus Streptomyces . On the basis of 16S rRNA gene sequence analysis strain RS10V-4T belongs to the genus Streptomyces and had the highest sequence similarity to Streptomyces noursei NBRC 15452T (98.3 %). The G+C content of the genomic DNA was 73.8 %. Digital DNA–DNA hybridization and average nucleotide identity values between the genome sequences of strain RS10V-4T and S.noursei ATCC 11455T were lower than the recommendation threshold values for the recognition of species within the same genus. The whole-cell hydrolysates of strain RS10V-4 T contained ll -diaminopimelic acid as the diagnostic diamino acid and the whole-cell sugars were glucose and ribose. The predominant menaquinones were MK-9(H6) and MK-9(H8). The predominant cellular fatty acids (>10 %) were iso-C16 : 0 anteiso-C15 : 0 iso-C14 : 0 and iso-C15 : 0. The polar lipids of strain RS10V-4T contained diphosphatidylglycerol hydroxyphosphatidylethanolamine phosphatidylethanolamine phosphatidylinositol an unidentified aminolipid two unidentified lipids and an unidentified phospholipid. On the basis of these phenotypic and genotypic characteristics it is supported that strain RS10V-4T represents a novel species of the genus Streptomyces for which the name Streptomyces rhizoryzae sp. nov. is proposed. The type strain is RS10V-4T (=TBRC 15167T=NBRC 115345T). In addition the comparison of the whole genome sequences and phenotypic features suggested that S. noursei and S. albulus belong to the same species. Therefore it is proposed that S. albulus is reclassified as a later heterotypic synonym of S. noursei .

A novel actinobacterium designated strain SF1T was isolated from the rhizosphere soil of a star fruit plant (Averrhoa carambola L.) collected from Bangkok Thailand and its taxonomic position was evaluated. The strain showed morphological chemotaxonomic and phylogenetic characteristics consistent with its classification in the genus Actinomycetospora . Strain SF1T was an aerobic Gram-stain-positive and non-motile actinobacterium. Growth occurred at 15–35 °C at pH 4.0–12.0 and in the presence of 0–10 % (w/v) NaCl. The 16S rRNA gene sequence of strain SF1T showed the highest similarity to Actinomycetospora chiangmaiensis YIM 0006T (99.5 %) Actinomycetospora corticicola 014-5T (98.8 %) and Actinomycetospora endophytica A-T 8314T (98.8 %). The genome sequencing revealed a genome size of 6.52 Mbp and a DNA G+C content of 74.0 %. In addition the average nucleotide identity values between strain SF1T and reference strains A. chiangmaiensis DSM 45062T A. corticicola DSM 45772T and A. endophytica TBRC 5722T were found to be 86.1 86.5 and 79.7 % respectively and the level of digital DNA–DNA hybridization between them were 32.4 32.4 and 23.3 % respectively. The whole-cell hydrolysates of strain SF1T contained meso-diaminopimelic acid as the diagnostic diamino acid with arabinose galactose glucose and ribose as whole-cell sugars. The predominant menaquinone was MK-8(H4). Major cellular fatty acids were iso-C16 : 0 and a summed feature consisting of C16 : 1 ω6c/C16 : 1 ω7c. Characterization based on chemotaxonomic phylogenetic phenotypic and genomic evidence demonstrated that strain SF1T represents a novel species of the genus Actinomycetospora for which the name Actinomycetospora soli sp. nov. is proposed. The type strain is strain SF1T (=TBRC 15166T= NBRC 115295T).

The genus Exophiala is polymorphic able to transition between yeast hyphal and pseudohyphal forms. Species of the genus Exophiala are ubiquitous fungi that are distributed in various environments around the world. During a survey of fungal diversity in the gut of amphipods (Floresorchestia amphawaensis and undescribed Dogielinotid amphipods) from the Amphawa estuary Samut Songkhram province Thailand five black yeast strains (DMKU-MG01 DMKU-MG07 DMKU-MG08 DMKU-HG10 and DMKU-FG04) were identified as representing a novel taxon on the basis of a combination of morphological and molecular phylogenetic features. The five strains did not produce filamentous hyphae or pseudohyphae. Only budding yeast cells were observed. On the basis of the phenotypic characteristics and the results of molecular analyses of the D1/D2 region of the large subunit (LSU) rRNA gene and the internal transcribed spacer (ITS) region the five strains were identified as representing a novel species via applied nucleotide pairwise analysis. They differed from the most closely related species Exophiala alcalophiala by 3.54 % nucleotide substitutions (20 nucleotide substitutions in 572 bp) in the D1/D2 domains of the LSU rRNA gene. Moreover the sequences of the ITS region of the five strains differed from those of the most closely related species E. alcalophiala by 7.44–9.62 % nucleotide substitutions and Exophiala halophiala by 7.2–7.53 % nucleotide substitutions. The results of phylogenetic analyses based on the concatenated sequences of the ITS regions and the D1/D2 domains of the LSU rRNA gene confirmed that the five black yeast strains represented a single novel species of the genus Exophiala. In this study Exophiala amphawaensis sp. nov. is proposed to accommodate these strains. The holotype is TBRC 15626T and the isotype is PYCC9020. The MycoBank accession number of the novel species is MB 851477.

During a study on the diversity of culturable actinobacteria from coastal halophytes in Thailand strain LSe6-5T was isolated from leaves of sea purslane (Sesuvium portulacastrum L.) and a polyphasic approach was employed to determine its taxonomic position. The 16S rRNA gene sequences analysis indicated that the strain was most closely related to Klenkia brasiliensis Tu 6233T (99.2 %) Klenkia marina YIM M13156T (99.1 %) and Klenkia terrae PB261T (98.7 %). The genome of strain LSe6-5T was estimated to be 4.33 Mbp in size with DNA G+C contents of 74.3%. A phylogenomic tree based on whole-genome sequences revealed that strain LSe6-5T formed a clade with Klenkia marina DSM 45722T indicating their close relationship. However the average nucleotide identity (ANI)-blast ANI-MUMmer and dDDH values between strain LSe6-5T with K. marina DSM 45722T (87.1 88.9 and 33.0 %) were below the thresholds of 95–96 % ANI and 70 % dDDH for identifying a novel species. Furthermore strain LSe6-5T showed morphological and chemotaxonomic characteristics of the genus Klenkia. Cells were motile rod-shaped and Gram-stain-positive. Optimal growth of strain LSe6-5T occurred at 28 °C pH 7.0 and 0–3 % NaCl. The whole-cell hydrolysates contained meso-diaminopimelic acid as the diagnostic diamino acid with galactose glucose mannose and ribose as whole-cell sugars. The predominant menaquinones were MK-9(H4) and MK-9(H0). The polar lipid profile was composed of diphosphatidylglycerol hydroxyphosphatidylethanolamine phosphatidylinositol glycophosphatidylinositol an unidentified phospholipid and an unidentified lipid. Major cellular fatty acids were iso-C15 : 0 iso-C16 : 0 and iso-C17 : 0. From the distinct phylogenetic position and combination of genotypic and phenotypic characteristics it is supported that strain LSe6-5T represents a novel species of the genus Klenkia for which the name Klenkia sesuvii sp. nov. is proposed. The type strain is strain LSe6-5T (=TBRC 16417T= NBRC 115929T).

Two strains of genus Kodamaea representing a novel anamorphic yeast species were isolated from two samples of Marasmiellus sp. collected in Thailand. Analysis of the sequences of the internal transcribed spacer (ITS) regions and the D1/D2 domains of the large subunit (LSU) rRNA gene showed that the two strains differed by 27–42 nucleotide substitutions in the D1/D2 domains of the LSU rRNA gene and 7–34 nucleotide substitutions in the ITS region of a group of related species Kodamaea smagusa CBS 11430T Kodamaea fungicola JCM 10142T Kodamaea plutei ATCC MYA-4329T Kodamaea lidongshanica SD5S01T and Kodamaea jinghongensis NYNU 167162T. Phylogenetic analysis based on the concatenated sequences of the ITS and the D1/D2 domains of the LSU rRNA gene showed that the two strains were placed in the Kodamaea clade and clearly separated from other recognized species of the genus. Therefore the two strains were assigned as a novel species of the genus Kodamaea for which we propose the name Kodamaea samutsakhonensis f.a. sp. nov. The holotype is TBRC 16043T (=DMKU-BP19T) and the isotype is PYCC 9354. The MycoBank number of the novel species is MB 846490.

Two novel actinobacterial strains designated RB6PN23T and K1PA1T were isolated from peat swamp soil samples in Thailand and characterized using a polyphasic taxonomic approach. The strains were filamentous Gram-stain-positive bacteria containing ll -diaminopimelic acid in their whole-cell hydrolysates. Phylogenetic analysis of their 16S rRNA gene sequences revealed that strain RB6PN23T was most closely related to Streptomyces rubrisoli (99.1 % sequence similarity) and Streptomyces ferralitis (98.5%) while strain K1PA1T showed 98.8 and 98.7% sequence similarities to Streptomyces coacervatus and Streptomyces griseoruber respectively. However the average nucleotide identity (ANI) and digital DNA–DNA hybridization (dDDH) values were below the species-level thresholds (95–96 % ANI and 70 % dDDH). The genomes of strains RB6PN23T and K1PA1T were estimated to be 7.88 Mbp and 7.39 Mbp in size respectively with DNA G+C contents of 70.2 and 73.2 mol%. Moreover strains RB6PN23T and K1PA1T encode 37 and 24 putative biosynthetic gene clusters respectively and in silico analysis revealed that these new species have a high potential to produce unique natural products. Genotypic and phenotypic characteristics confirmed that strains RB6PN23T and K1PA1T represented two novel species in the genus Streptomyces . The names proposed for these strains are Streptomyces silvisoli sp. nov. (type strain RB6PN23T=TBRC 17040T=NBRC 116113T) and Streptomyces tropicalis sp. nov. (type strain K1PA1T=TBRC 17041T=NBRC 116114T). Additionally a giant linear polyene compound neotetrafibricin A exhibiting antifungal activity in strain RB6PN23T was identified through HPLC and quadrupole time-of-flight MS analysis. The crude extract from the culture broth of strain RB6PN23T exhibited strong antifungal activity against Fusarium verticillioides Fusarium fujikuroi and Bipolaris zeicola. This finding suggests that strain RB6PN23T could be a promising candidate for biological control of fungal diseases.

Four strains (NYNU 178247 NYNU 178251 DMKU-PAL160 and DMKU-PAL137) representing a novel yeast species were isolated from the external surfaces of rice and pineapple leaves collected in China and Thailand. Phylogenetic analysis based on the concatenated sequences of the internal transcribed spacer (ITS) regions and the D1/D2 domains of the large subunit rRNA gene revealed that the novel species belonged to the genus Spencerozyma. The D1/D2 sequence of the novel species differed from its closest relative Spencerozyma acididurans SYSU-17T by 3.2 % sequence divergence. The species also differed from Spencerozyma crocea CBS 2029T and Spencerozyma siamensis DMKU13-2T by 3.0–6.9 % sequence divergence in the D1/D2 sequences out of 592 bp. In the ITS regions the novel species displayed 19.8–29.2% sequence divergence from S. acididurans SYSU-17T S. crocea CBS 2029T and S. siamensis DMKU13-2T out of 655 bp. Furthermore the novel species could also be differentiated from the closely related species by some physiological characteristics. The species name of Spencerozyma pingqiaoensis sp. nov. (Holotype CBS 15238 Mycobank MB 844734) is proposed to accommodate these four strains.

The taxonomic position of two novel Actinoallomurus strains isolated from rhizosphere soil of wild rice (Oryza rufipogon Griff.) was established using a polyphasic approach. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strains WRP6H-15T and WRP9H-5T were closely related to Actinoallomurus spadix JCM 3146T and Actinoallomurus purpureus TTN02-30T. Chemotaxonomic and morphological characteristics of both strains were consistent with members of the genus Actinoallomurus while phenotypic properties genome-based comparisons and phylogenomic analyses distinguished strains WRP6H-15T and WRP9H-5T from their closest phylogenetic relatives. The two strains showed nearly identical 16S rRNA gene sequences (99.9 %). Strain WRP6H-15T showed 68.7 % digital DNA–DNA hybridization 95.9 % average nucleotide identity (ANI) based on blast and 96.4 % ANI based on MUMmer to strain WRP9H-5T. A phylogenomic tree based on draft genome sequences of the strains and representative of the genus Actinoallomurus confirmed the phylogenetic relationships. The genomes sizes of strains WRP6H-15T and WRP9H-5T were 9.42 Mb and 9.68 Mb with DNA G+C contents of 71.5 and 71.3 mol% respectively. In silico analysis predicted that the strains contain biosynthetic gene clusters encoding for specialized metabolites. Characterization based on chemotaxonomic phylogenetic phenotypic and genomic evidence demonstrated that strains WRP6H-15T and WRP9H-5T represent two novel species of the genus Actinoallomurus for which the names Actinoallomurus soli sp. nov. (type strain WRP6H-15T=TBRC 15726T=NBRC 115556T) and Actinoallomurus rhizosphaericola sp. nov. (type strain WRP9H-5T=TBRC 15727T=NBRC 115557T) are proposed.

Six strains representing two novel ascomycetous yeast species were isolated from mushroom fruiting bodies and cocoa leaves collected in Thailand. Analysis of the internal transcribed spacer (ITS) regions and the D1/D2 domains of the large subunit rRNA gene sequences showed that the six strains were divided into two groups. The first group consisted of four strains (DMKU-SSK46 DMKU-SK1 SCCL3-5 and SCCL19-3) that were closely related to the type strains of Candida conglobata Candida insectorum Yamadazyma dushanensis Yamadazyma mexicana and Yamadazyma riverae but with 12–14 (2.5–2.9 %) and 28–50 (5.4–8.8 %) nucleotide substitutions in the D1/D2 domains and the ITS regions respectively. However two strains (DMKU-KMY40 and DMKU-KO18) of the second group differed from a group of described species Candida diddensiae Candida dendronema Candida germanica Candida kanchanaburiensis Candida naeodendra Candida vaughaniae and Yamadazyma siamensis by 8–15 (1.5–2.8 %) and 45–53 (8.2–9.6 %) nucleotide substitutions in the D1/D2 domains and the ITS regions respectively. Phylogenetic analysis based on the concatenated sequences of the ITS regions and D1/D2 domains showed that these strains represented two species of the Yamadazyma clade that were distinct from the other related species. Based on the phylogenetic analysis and phenotypic characteristics these six strains were assigned to two novel species of the genus Yamadazyma although formation of ascospores was not observed. Yamadazyma sisaketensis f.a. sp. nov. is proposed for the first group (four strains). The holotype is TBRC 17139T (ex-type culture: PYCC 9797). The MycoBank number is MB 849637. Yamadazyma koratensis f.a. sp. nov. is proposed for the second group (two strains). The holotype is TBRC 14868T (ex-type culture: PYCC 8907). The MycoBank number is MB 849638. In addition it is proposed that Candida andamanensis Candida jaroonii and Candida songkhlaensis are reassigned to the genus Yamadazyma as Yamadazyma andamanensis comb. nov. Y. jaroonii comb. nov. and Y. songkhlaensis comb. nov. respectively.