Modified Activated Carbon: A Supporting Material for Improving Clostridium beijerinckii TISTR1461 Immobilized Fermentation.

This study aimed to investigate the effect of activated carbon (AC) as an immobilization material in acetone-butanol-ethanol fermentation. The AC surface was modified with different physical (orbital shaking and refluxing) and chemical (nitric acid, sodium hydroxide and, (3-aminopropyl)triethoxysilane (APTES)) treatments to enhance the biobutanol production by Clostridium beijerinckii TISTR1461. The effect of surface modification on AC was evaluated using Fourier-transform infrared spectroscopy, field emission scanning electron microscopy, surface area analyses, and X-ray photoelectron spectroscopy, while the fermented broth was examined by high-performance liquid chromatography. The chemical functionalization significantly modified the physicochemical properties of the different treated ACs and further enhanced the butanol production. The AC treated with APTES under refluxing provided the best fermentation results at 10.93 g/L of butanol, 0.23 g/g of yield, and 0.15 g/L/h of productivity, which were 1.8-, 1.5-, and 3.0-fold higher, respectively, than that in the free-cell fermentation. The obtained dried cell biomass also revealed that the treatment improved the AC surface for cell immobilization. This study demonstrated and emphasized the importance of surface properties to cell immobilization.

Novosphingobium kaempferiae sp. nov., a phosphate-solubilizing bacterium isolated from stem of Kaempferia marginata Carey.

A Gram-stain-negative, aerobic, rod-shaped, non-spore-forming and yellow-pigment-producing bacterium, designated as Sx8-5T, was isolated from stem tissue of Kaempferia marginata Carey in Kanchanaburi Province, Thailand. The strain exhibited tricalcium phosphate solubilizing activity. Its taxonomic position was investigated using a polyphasic approach. Sx8-5T grew at 25-37 °C (optimum 30 °C), pH 6-9 (optimum 7) and with 0 and 1% NaCl (optimum 0 %). According to the 16S rRNA gene phylogeny, Sx8-5T represents a member of genus Novosphingobium and shared the highest sequence similarities to Novosphingobium barchaimii LL02T of 99.4 % and shared sequence similarities with other species of the genus Novosphingobium of less than 99.4 %. The whole-genome size was 5.7 Mb, comprised of one contig, with a DNA G+C content of 66 %. The average nucleotide identity using BLASTn (ANIb) or MUMMER (ANIm) values for whole genome comparisons between Sx8-5T and Novosphingobium barchaimii LL02T and six closely related type strains were 72.33-82.14 % and 83.82-87.38 %, respectively, and the digital DNA-DNA hybridization (dDDH) values ranged from 21.0 to 28.6% when compared with the type strains of the members of the genus Novosphingobium. Major fatty acids were summed feature 8 (C18 : 1 ω7c and/or C18 : 1 ω6c), C16 : 0 and summed feature 3 (C16 : 1 ω7c and/or C16 : 1 ω6c), respectively. Polar lipids were diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylmonomethylethanolamine, phosphatidylglycerol, unidentified phospholipids and unidentified polar lipids. The major isoprenoid quinone was Q-10. According to results obtained using a polyphasic approach, Sx8-5T represents a novel species of the genus Novosphingobium, the name Novosphingobium kaempferiae sp. nov. is proposed. The type strain is Sx8-5T (=JCM 35076T =TBRC 15600T).

Metahyphopichia suwanaadthiae sp. nov., an anamorphic yeast species in the order Saccharomycetales and reassignment of Candida silvanorum to the genus Metahyphopichia.

Seven yeast strains, representing a single novel anamorphic species, were isolated in Thailand. They consisted of five strains (DMKU-MRY16T, DMKU-SK18, DMKU-SK25, DMKU-SK30 and DMKU-SK32) obtained from five different mushrooms, and two strains (ST-224 and 11-14.2) derived from insect frass and soil, respectively. The pairwise sequence analysis indicated that all seven strains had identical sequences in the D1/D2 domains of the large subunit (LSU) rRNA gene and the internal transcribed spacer (ITS) region. Metahyphopichia silvanorum was the most closely related species, but with 11.9-12.4% nucleotide substitutions in the D1/D2 domains of the LSU rRNA gene and 13.1-13.3% nucleotide substitutions in the ITS region. The phylogenetic analyses based on the concatenated sequences of the ITS region and the D1/D2 domains of the LSU rRNA gene showed that the seven strains form a well-separated subclade in a clade containing M. silvanorum and Metahyphopichia laotica with high bootstrap support. A phylogenetic analysis of a multilocus dataset including the small subunit (SSU) rRNA gene, the ITS region, the D1/D2 domains of the LSU rRNA gene, translation elongation factor 1-alpha gene, actin gene and the RNA polymerase II subunit 2 gene, confirmed the presence of the monophyletic clade that also includes M. silvanorum and M. laotica, and strongly supported the phylogenetic isolation of the seven strains from its neighbouring species. Therefore, the seven strains were assigned as a single novel species of the genus Metahyphopichia, according to their phylogenetic relationships. The name Metahyphopichia suwanaadthiae sp. nov. is proposed to accommodate the seven strains. The holotype is DMKU-MRY16T (TBRC 11775T=NBRC 114386T=PYCC 8655T). The MycoBank number of the novel species is MB 841280. In addition, Candida silvanorum is reassigned to the genus Metahyphopichia. The MycoBank number of M. silvanorum comb. nov. is MB 841279.

Allobacillus salarius sp. nov., and Allobacillus saliphilus sp. nov., isolated from shrimp paste (ka-pi) in Thailand.

Two strains of moderately halophilic, Gram-stain-positive and spore-forming rods, designated as SKP4-8T and SKP8-2T isolated from shrimp paste (Ka-pi), were taxonomic studied based on polyphasic approach. Strain SKP4-8T grew at pH 6.0-9.0 (optimum 7.0), at 25-45 °C (optimum 37 °C) and in 1-16% (w/v) NaCl (optimum 5-10%). Strain SKP8-2T grew at pH 6.0-9.0 (optimum 8.0), at 25-45 °C (optimum 37 °C) and in 0-20% (w/v) NaCl (optimum 3-10%). The strains contained meso-diaminopimelic acid in cell-wall peptidoglycan and the major menaquinone was MK-7. Strain SKP4-8T contained iso-C15:0, anteiso-C15:0 and iso-C17:0; and strain SKP8-2T contained anteiso-C15:0, iso-C15:0, iso-C16:0 and antesio-C17:0 as major cellular fatty acids. Phosphatidylglycerol, diphosphatidylglycerol, unknown phospholipids and an unknown glycolipid were detected as major polar lipids. On the basis of 16S rRNA gene sequence analysis, strains SKP4-8T and SKP8-2T belonged to the genus Allobacillus and were closely related to Allobacillus halotolerans LMG 24826T with 98.8% and 99.3% similarity, respectively. The comparative genome analysis based on average nucleotide identity (ANI) and digital DNA-DNA hybridization revealed that both strains showed the values below 95 and 70%, from each other and from Allobacillus halotolerans LMG 24826T, respectively. Based on the data from this polyphasic study, strains SKP4-8T and SKP8-2T represent novel species of the genus Allobacillus and the name Allobacillus salarius sp. nov. was proposed for SKP4-8T (= KCTC 33905T = LMG 30016T = TISTR 2499T); and Allobacillus saliphilus sp. nov. for SKP8-2T (= KCTC 33906T = LMG 29682T = TISTR 2558T).

Identification and lipolytic activity of yeasts isolated from foods and wastes.

Thirty-three yeasts were isolated from palm oil industrial wastes and traditional fermented foods in Thailand. Based on the analysis of the sequences of the D1/D2 region of the large subunit ribosomal RNA gene (LSU rDNA) and their phenotypic characteristics, they were identified as Pichia kudriavzevii (11 isolates), Candida ethanolica (1 isolate), Clavispora lusitaniae (2 isolates), Ogataea polymorpha (1 isolate), Hanseniaspora opuntiae (1 isolate), Lodderomyces elongisporus (1 isolate), Saccharomyces cerevisiae (2 isolates), C. tropicalis (5 isolates), Yarrowia lipolytica (2 isolates), Magnusiomyces ingens (1 isolate), and Magnusiomyces capitatus (3 isolates), Trichosporon insectorum (1 isolate), and Trichosporon asteroides (2 isolates). Seven strains, T. insectorum 4E-1D, M. capitatus 5E-1T and 5E-2D, T. asteroides 8E-1T and 8E-1D, and Y. lipolytica Fy-12 and Fy-13, showed high lipolytic activity ranged from 5.21 ± 0.09 to 45.68 ± 2.37 U/mL. Moreover, these seven strains exhibited good lipolytic activity after culturing in the medium containing palm oil (11.79 ± 0.67 to 28.19 ± 4.84 U/mL) and soy oil (9.14 ± 1.08 to 22.97 ± 0.69 U/mL) as lipase inducers. The result of this study suggests that the palm oil industrial wastes and Thai fermented foods could be promised as the invaluable sources of lipolytic yeasts.

Heterocephalacria mucosa sp. nov., a new basidiomycetous yeast species isolated from a mangrove forest in Thailand.

Strains of yeast were isolated under a nitrogen-depleted culture condition from decaying tree bark (strain N-12.1) and from mangrove forest water (strain 1-7W.1) sampled at different locations within a mangrove forest site in Ranong province, Thailand. They were found to be genetically and phenotypically different from any currently recognised yeast species. Phylogenetic analysis of nucleotide sequence of three genes, the internal transcribed spacer region 1 and 2 plus 5.8S ribosomal RNA gene (ITS), D1/D2 domain of the large subunit ribosomal RNA gene (LSU D1/D2) and the small subunit of the ribosomal RNA gene (SSU), revealed that these two strains were related to but distinguished from Heterocephalacriaarrabidensis. Several distinct physiological characteristics of these two strains were detected, namely inability to assimilate glycerol, dl-lactic acid, succinic acid, citric acid, d-gluconic acid, and ability to grow well at 25 °C, which were different from those of H. arrabidensis. Accordingly, the name Heterocephalacria mucosa sp. nov., is proposed to accommodate this novel species. The MycoBank number is MB 828624.

Occurrence of oleaginous yeast from mangrove forest in Thailand.

A total of 191 yeasts were isolated from 197 samples collected from eight estuarine mangrove forests along four different coastlines of Thailand (Andaman Sea and the East, North and West coasts of the Gulf of Thailand). Of these, 178 isolates were identified as 32 species in 16 genera of Ascomycota, 12 species in nine genera of Basidiomycota, and 13 isolates as potential new species, respectively. Mangroves located along the Andaman Sea coastline had a higher yeast diversity at the species and genera levels than those along the Gulf of Thailand. Kluyveromyces siamensis was the most frequently isolated species, whilst Candida tropicalis was the only species isolated at all eight sites. Screening isolated yeast strains belonging to genera previously reported as oleaginous yeast plus the 13 potential new species, revealed two oleaginous strains, Rhodotorula sphaerocarpa 11-14.4 and Saitozyma podzolica 11-11.3.1. Both of these strains were isolated from the same mangrove forest on the Andaman Sea coastline. They could accumulate lipid when suspended in glucose solution without any supplementation, while the fatty acid composition and oil profile of Rh. sphaerocarpa 11-14.4 and Sait. podzolica 11-11.3.1 were similar to vegetable oil and cocoa butter, respectively.

Streptomyces lichenis sp. nov., isolated from lichen.

A new actinobacterial strain, LCR6-01T, was isolated from a lichen sample collected from Chiang Rai Province, Thailand. Its taxonomic position was determined using a polyphasic approach. The strain had morphological characteristics and chemotaxonomic properties identical to those of members of the genus Streptomyces. The predominant menaquinones were MK-9(H6) and MK-9(H8). The major fatty acids were anteiso-C15 : 0, iso-C15 : 0, iso-C16 : 0, iso-C14 : 0 and C16 : 0. The polar lipids were diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, an unidentified phosphoglycolipid, an unidentified phospholipid and an unidentified lipid. The genomic DNA G+C content was 74.5 mol%. Phylogenetic analysis based on 16S rRNA gene sequences suggested that the strain belonged to the genus Streptomyces. The strain showed highest 16S rRNA gene sequence similarity to Streptomyces palmae TBRC 1999T (98.6 %), Streptomyces misionensis JCM 4497T (98.6 %), Streptomyces matensis JCM 4268T (98.5 %), Streptomyces althioticus KCTC 9752T (98.5 %) and Streptomyces wuyuanensis KCTC 29112T (98.4 %). DNA-DNA relatedness values among the strain and closely related Streptomyces species were below 70 %. On the basis of the phenotypic characteristics and genotypic distinctiveness, strain LCR6-01T is considered to represent a novel species of the genus Streptomyces, for which the name Streptomyces lichenis sp. nov. is proposed. The type strain is LCR6-01T (=KCTC 39908T=TISTR 2500T).

Diversity and characterization of cultivable oleaginous yeasts isolated from mangrove forests.

A total of 198 yeasts were isolated from 140 samples collected from 7 mangrove forests in 4 provinces of Thailand, and were found to belong to 30 genera, 45 described species and at least 12 undescribed species based on their 26S rRNA (D1/D2 domain) gene sequence. The most prevalent species was Candida tropicalis, followed by Candida pseudolambica and Rhodosporidium paludigena. Lipid accumulation, as determined by Nile red staining, of the isolated yeasts revealed that 69 and 18 strains were positive and strongly positive, respectively, while quantitative analysis of the intracellular lipid accumulated in the latter indicated that 10 of these strains, Pseudozyma tsukubaensis (YWT7-2 and YWT7-3), Rhodotorula sphaerocarpa (YWW6-1 and SFL14-1SF), Saitozyma podzolica (YWT1-1, NS3-3 and NS10-2), Prototheca zopfii var. hydrocarbonea OMS6-1 and Prototheca sp. (YMTW3-1 and YMTS5-2), were oleaginous. In this study we found that under nitrogen depletion condition (155 C/N ratio) Pseudozyma tsukubaensis YWT7-2 accumulated the highest level of intracellular lipid at 32.4% (w/w, dry cell weight), with a broadly similar fatty acid composition to that in palm oil.