Savitreea siamensis sp. nov., an ascomycetous yeast species in the family Saccharomycetaceae discovered in Thailand.

Four yeast strains, representing a novel anamorphic species, were isolated in Thailand. The two strains (ST-3660T and ST-3647) were obtained from two different estuarine water samples in a mangrove forest. Strain DMKU-FW1-37 was derived from a grease sample, and another strain (TSU57) was isolated from a fruiting body of Phallus sp. Pairwise sequence analysis showed that the four strains had identical or differed by only one nucleotide substitution in the D1/D2 domains of the large subunit (LSU) rRNA gene, and differed by one to three nucleotide substitutions in the internal transcribed spacer (ITS) regions. Savitreea pentosicarens is the most closely related species to the four strains, but with 9-10 (1.57-1.72 %) nucleotide substitutions in the D1/D2 domains of the LSU rRNA gene and 29-31 (4.22-4.45 %) nucleotide substitutions in the ITS regions. Phylogenetic analyses based on the concatenated sequences of the ITS regions and the D1/D2 domains of the LSU rRNA gene showed that the four strains form a well-separated lineage from S. pentosicarens with high bootstrap support, confirming that they represent a distinct species. Therefore, the four strains are assigned as representives of a novel species of the genus Savitreea, for which the name Savitreea siamensis sp. nov. is proposed. The holotype is TBRC 4481T and the ex-type is PYCC 9794T (=ST-3660T). The MycoBank number of the novel species is MB 851951.

Microbiome sequencing revealed the abundance of uncultured bacteria in the Phatthalung sago palm-growing soil.

Environmental variations have been observed to influence bacterial community composition, thereby impacting biological activities in the soil. Together, the information on bacterial functional groups in Phatthalung sago palm-growing soils remains limited. In this work, the core soil bacterial community in the Phatthalung sago palm-growing areas during both the summer and rainy seasons was examined using V3-V4 amplicon sequencing. Our findings demonstrated that the seasons had no significant effects on the alpha diversity, but the beta diversity of the community was influenced by seasonal variations. The bacteria in the phyla Acidobacteriota, Actinobacteriota, Chloroflexi, Methylomirabilota, Planctomycetota, and Proteobacteria were predominantly identified across the soil samples. Among these, 26 genera were classified as a core microbiome, mostly belonging to uncultured bacteria. Gene functions related to photorespiration and methanogenesis were enriched in both seasons. Genes related to aerobic chemoheterotrophy metabolisms and nitrogen fixation were more abundant in the rainy season soils, while, human pathogen pneumonia-related genes were overrepresented in the summer season. The investigation not only provides into the bacterial composition inherent to the sago palm-cultivated soil but also the gene functions during the shift in seasons.

Yamadazyma sisaketensis f.a., sp. nov. and Yamadazyma koratensis f.a., sp. nov., two novel ascomycetous yeast species from mushrooms and cocoa leaves in Thailand, and reassignment of Candida andamanensis, Candida jaroonii and Candida songkhlaensis to the genus Yamadazyma.

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.

Vegetative insecticidal protein (Vip3A) production by Bacillus thuringiensis Bt294 and its efficacy against Lepidopteran pests (Spodoptera exigua).

A vegetative insecticidal protein, Vip3A, is highly active against lepidopteran pests, which are the most important pests in most tropical countries. An important aspect of the successful commercial production of this bacterial insecticide is the development of bacterial culture media that maximize the titres of this protein and cost reduction. This study aimed to investigate and optimize Vip3A production by Bacillus thuringiensis Bt294 using statistical methods and 3-step sequential approaches. The experimental design showed that the production of Vip3A was maximized to 300 mg/L when the bacterium was cultivated in medium composed of 5.05 g/L glycerol, 49.17 g/L soytone, 30.05 g/L casein hydrolysate, 1.99 g/L CaCl2.2H2O, 7.5 mg/L CuSO4, 15 mg/L MnSO4.H2O, 9.4 g/L K2HPO4, 2.2 g/L KH2PO4, 0.2 g/L MgSO4.7H2O, 5 g/L yeast extract, 2.5 mg/L NiCl2.6H2O and 3 mL/L vitamin solution. B. thuringiensis Bt294 Vip3A toxin was highly toxic to Spodoptera exigua with LC50 values of 187.1 ng/cm2 at 7 days. This result demonstrated that a high titre of Vip3A produced by B. thuringiensis Bt294 will be useful as a biological control agent. This optimization will allow production to be scaled up for commercial production in the future.

Spencerozyma pingqiaoensis sp. nov., a yeast species isolated from the external surface of rice leaves in China.

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.

A Novel Yeast Genus and Two Novel Species Isolated from Pineapple Leaves in Thailand: Savitreella phatthalungensis gen. nov., sp. nov. and Goffeauzyma siamensis sp. nov.

Four yeast strains, representing one genus and two novel anamorphic yeast species, were isolated from pineapple leaves collected in Thailand. Analysis of the sequences of the D1/D2 domains of the large subunit (LSU) rRNA gene and the internal transcribed spacer (ITS) regions showed the two strains (DMKU-PAL186 and DMKU-PAL178) were closely related to the type strains of the Protomyces and Taphrina species, but with high nucleotide divergence. Two strains (DMKU-PAL39 and DMKU-PAL18) were found to be closely related to the type strains of Goffeauzyma iberica, but with eight nucleotide substitutions in the D1/D2 domains and 26 nucleotide substitutions in the ITS regions. In phylogenetic analyses, the strains DMKU-PAL186 and DMKU-PAL178 formed a well-separated lineage from Protomyces and Taphrina genera, confirming that they represented a distinct genus, while the strains DMKU-PAL39 and DMKU-PAL18 represented a species in the genus Goffeauzyma, which was phylogenetically distinct from other recognized species of the genus. Based on molecular analyses and phenotypic characteristics, the names Savitreella gen. nov. (Taphrinomycetes, Ascomycota) and Savitreella phatthalungensis sp. nov. are proposed to accommodate the strains DMKU-PAL186 and DMKU-PAL178, and the name Goffeauzyma siamensis sp. nov. (Tremellomycetes, Basidiomycota) is proposed to accommodate the strains DMKU-PAL39 and DMKU-PAL18.

Production of Indole-3-Acetic Acid by Enterobacter sp. DMKU-RP206 Using Sweet Whey as a Low-Cost Feed Stock.

In this study, we investigated Indole-3-acetic acid (IAA) production by a rice phylloplane bacteria, Enterobacter sp. DMKU-RP206, using sweet whey as a feed stock instead of lactose. We succeeded in using sweet whey for Enterobacter sp. DMKU-RP206 to produce 3,963.0 mg IAA/l with the optimal medium containing 1.48% sweet whey, 1.42% yeast extract and 0.88% L-tryptophan. The medium pH was adjusted to 6 and the culture conditions were shaking at 200 rpm on an orbital shaker at 30°C for 3 days. We also evaluated the effect of IAA in culture filtrates of Enterobacter sp. DMKU-RP206 on the promotion of jasmine rice growth in a pot experiment. Compared with the negative control (without IAA), the result showed that biosynthetic IAA produced by Enterobacter sp. DMKU-RP206 significantly increased the growth of jasmine rice (Oryza sativa L. cv. KDML105) in terms of length and dry weight of shoot. This work thus reveals the impact of IAA produced by Enterobacter sp. on the promotion of jasmine rice growth.

High-yield production of indole-3-acetic acid by Enterobacter sp. DMKU-RP206, a rice phyllosphere bacterium that possesses plant growth-promoting traits.

Enterobacter sp. DMKU-RP206 was isolated from rice leaves in Thailand and identified by the 16S rRNA gene and multilocus sequence (gyrB, rpoB, atpD, and infB genes) analysis. The bacterium was assessed on plant growth-promoting traits including indole-3-acetic acid (IAA) production. Phosphate solubilization, ammonia production, and antagonism to fungal plant pathogens, as well as siderophore production, were shown by this bacterium. However, only IAA production was focused on. The production of IAA by Enterobacter sp. DMKU-RP206 was optimized by statistical methods. A Box-Behnken design was used for the investigation of interactions among the basic influencing factors and for the optimization of IAA production. The results showed that l-tryptophan had a significant importance in terms of IAA production. Enterobacter sp. DMKU-RP206 produced a higher amount of IAA than previously reported for the genus Enterobacter. 0.85% of lactose as a carbon source, 1.3% of yeast extract as a nitrogen source, 1.1% of l-tryptophan as a precursor, 0.4% of NaCl, an initial pH of 5.8, an incubation temperature at 30 °C, and a shaking speed of 200 rpm were found to be the optimum conditions for IAA production. In addition, IAA production was performed to scale up IAA production, and the highest amount, 5561.7 mg l-1, was obtained. This study reported a 13.4-fold improvement in IAA production by Enterobacter sp. DMKU-RP206.

Indole-3-acetic acid biosynthetic pathways in the basidiomycetous yeast Rhodosporidium paludigenum.

Microorganisms produce plant growth regulators, such as auxins, cytokinins and gibberellins, to promote plant growth. Auxins are a group of compounds with an indole ring that have a positive effect on plant growth. Indole-3-acetic acid (IAA) is a plant growth hormone classified as an indole derivative of the auxin family. IAA biosynthesis pathways have been reported and widely studied in several groups of bacteria. Only a few studies on IAA biosynthesis pathways have been conducted in yeast. This study aimed to investigate IAA biosynthesis pathways in a basidiomycetous yeast (Rhodosporidium paludigenum DMKU-RP301). Investigations were performed both with and without a tryptophan supplement. Indole compound intermediates were detected by gas chromatography-mass spectrometry. Indole-3-lactic acid and indole-3-ethanol were found as a result of the enzymatic reduction of indole-3-pyruvic acid and indole-3-acetaldehyde, in IAA biosynthesis via an indole-3-pyruvic acid pathway. In addition, we also found indole-3-pyruvic acid in culture supernatants determined by high-performance liquid chromatography. Identification of tryptophan aminotransferase activity supports indole-3-pyruvic acid-routed IAA biosynthesis in R. paludigenum DMKU-RP301. We hence concluded that R. paludigenum DMKU-RP301 produces IAA through an indole-3-pyruvic acid pathway.

Roseomonas elaeocarpi sp. nov., isolated from olive (Elaeocarpus hygrophilus Kurz.) phyllosphere.

An aerobic, Gram-stain-negative, coccobacillus-shaped, non-endospore-forming, pink-pigmented bacterium, designated PN2T, was isolated from an olive leaf. The strain grew at 15-35 °C with an optimum temperature for growth at 30 °C, and at pH 5.0-7.5 with an optimum pH for growth at 6.0. Growth was observed in the presence of up to 1.02 % (w/v) NaCl. The major fatty acids were C19 : 0 cyclo ω8c, C16 : 0 and C18 : 1ω7c. The polar lipids were diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylcholine, phosphatidylglycerol, unknown aminolipids, an unknown phospholipid and an unknown lipid. The respiratory quinone was ubiquinone-10. The DNA G+C content of strain PN2T was 70.4 mol%. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain PN2T was closely related to members of the genus Roseomonas and shared highest similarity with Roseomonas mucosa ATCC BAA-692T (96.5 %), Roseomonas gilardii subsp. gilardii ATCC 49956T (96.2 %) and Roseomonas gilardii subsp. rosea ATCC BAA-691T (96.2 %). Furthermore, the DNA-DNA relatedness value between strain PN2T and the closest related species R. mucosa ATCC BAA-692T was 27 %. These data allowed the phenotypic and genotypic differentiation of strain PN2T from its closest phylogenetic neighbour (R. mucosa ATCC BAA-692T). Based on phenotypic and genotypic characteristics, strain PN2T is classified as representing a novel species of the genus Roseomonas for which the name Roseomonas elaeocarpi sp. nov. is proposed. The type strain is PN2T ( = BCC 44864T = NBRC 107871T).

Indole-3-acetic acid production by newly isolated red yeast Rhodosporidium paludigenum.

Indole 3-acetic acid (IAA) is the principal hormone which regulates various developmental and physiological processes in plants. IAA production is considered as a key trait for supporting plant growth. Hence, in this study, production of indole-3-acetic acid (IAA) by a basidiomycetous red yeast Rhodosporidium paludigenum DMKU-RP301 (AB920314) was investigated and improved by the optimization of the culture medium and culture conditions using one factor at a time (OFAT) and response surface methodology (RSM). The study considered the effects of incubation time, carbon and nitrogen sources, growth factor, tryptophan, temperature, shaking speed, NaCl and pH, on the production of IAA. The results showed that all the factors studied, except NaCl, affected IAA production by R. paludigenum DMKU-RP301. Maximum IAA production of 1,623.9 mg/l was obtained as a result of the studies using RSM. The optimal medium and growth conditions observed in this study resulted in an increase of IAA production by a factor of up to 5.0 compared to the unoptimized condition, i.e. when yeast extract peptone dextrose (YPD) broth supplemented with 0.1% l-tryptophan was used as the production medium. The production of IAA was then scaled up in a 2-l stirred tank fermenter, and the maximum IAA of 1,627.1 mg/l was obtained. This experiment indicated that the obtained optimal medium and condition (pH and temperature) from shaking flask production can be used for the production of IAA in a larger size production. In addition, the present research is the first to report on the optimization of IAA production by the yeast Rhodosporidium.

Plant growth-promoting traits of epiphytic and endophytic yeasts isolated from rice and sugar cane leaves in Thailand.

A total of 1035 yeast isolates, obtained from rice and sugar cane leaves, were screened primarily for indole-3-acetic acid (IAA) production. Thirteen isolates were selected, due to their IAA production ranging from 1.2 to 29.3 mg g(-)(1) DCW. These isolates were investigated for their capabilities of calcium phosphate and ZnO(3) solubilisation, and also for production of NH(3), polyamine, and siderophore. Their 1-aminocyclopropane-1-carboxylate (ACC) deaminase, catalase and fungal cell wall-degrading enzyme activities were assessed. Their antagonism against rice fungal pathogens was also evaluated. Strain identification, based on molecular taxonomy, of the thirteen yeast isolates revealed that four yeast species - i.e. Hannaella sinensis (DMKU-RP45), Cryptococcus flavus (DMKU-RE12, DMKU-RE19, DMKU-RE67, and DMKU-RP128), Rhodosporidium paludigenum (DMKU-RP301) and Torulaspora globosa (DMKU-RP31) - were capable of high IAA production. Catalase activity was detected in all yeast strains tested. The yeast R. paludigenum DMKU-RP301 was the best IAA producer, yielding 29.3 mg g(-)(1) DCW, and showed the ability to produce NH3 and siderophore. Different levels of IAA production (7.2-9.7 mg g(-)(1) DCW) were found in four strains of C. flavus DMKU-RE12, DMKU-RE19, and DMKU-RE67, which are rice leaf endophytes, and strain DMKU-RP128, which is a rice leaf epiphyte. NH(3) production and carboxymethyl cellulase (CMCase) activity was also detected in these four strains. Antagonism to fungal plant pathogens and production of antifungal volatile compounds were exhibited in T. globosa DMKU-RP31, as well as a moderate level of IAA production (4.9 mg g(-)(1) DCW). The overall results indicated that T. globosa DMKU-RP31 might be used in two ways: enhancing plant growth and acting as a biocontrol agent. In addition, four C. flavus were also found to be strains of interest for optimal IAA production.

Roseomonas musae sp. nov., a new bacterium isolated from a banana phyllosphere.

A Gram-negative, coccobacilli, non-spore forming and non-motile bacterium, designated PN1(T), was isolated from a banana leaf collected in Mattra island, Thailand. This isolate was observed to grow optimally at 30 °C and pH 7.0, and to grow with 0-3 % NaCl. Comparative 16S rRNA gene sequence analysis showed that strain PN1(T) is closely related to members of the genus Roseomonas, exhibiting the highest 16S rRNA gene sequence similarity to Roseomonas aestuarii JC17(T) (96.5 %). The DNA G + C content of strain PN1(T) was determined to be 69.7 mol %. Based on physiological and biochemical tests, and genotypic differences between strain PN1(T) and the validly named species of the genus Roseomonas, it is proposed that the strain be classified as a new species of Roseomonas for which the name Roseomonas musae sp. nov. is proposed. The type strain is PN1(T) (= BCC 44863(T) = NBRC 107870(T)).