Diversity, astaxanthin production, and genomic analysis of Rhodotorula paludigena SP9-15.

Astaxanthin is a carotenoid known for its powerful antioxidant properties. This study focused on isolating yeast strains capable of producing astaxanthin from flower and fruit samples collected in Thailand. Out of 115 isolates, 11 strains were identified that produced astaxanthin. Molecular identification techniques revealed that these isolates belonged to two species: Rhodotorula paludigena (5 isolates) and Rhodosporidiobolus ruineniae (6 isolates). Whole-genome analysis of one representative strain, R. paludigena SP9-15, identified putative candidate astaxanthin synthesis-associated genes, such as CrtE, CrtYB, CrtI, CrtS, CrtR, CrtW, CrtO, and CrtZ. High-performance liquid chromatography (HPLC) and liquid chromatography-mass spectrometry (LC-MS) confirmed astaxanthin production. Further optimization of astaxanthin production was carried out by investigating the effects of various factors on the growth rate and astaxanthin production. The optimal conditions were 40 g/L glucose as a carbon source, pH 7.5, and cultivation at 25 °C with 200 rpm for 3 days. Under these conditions, R. paludigena SP9-15 synthesized biomass of 11.771 ± 0.003 g/L, resulting in astaxanthin with a content of 0.558 ± 0.018 mg/g DCW (dry cell weight), an astaxanthin yield of 6.565 ± 0.238 mg/L, and astaxanthin productivity of 2.188 ± 0.069 g/L/day. These findings provide insights into astaxanthin production using red yeast strains from Thailand and highlight the potential of R. paludigena SP9-15 for further application.

Sporolactobacillus mangiferae sp. nov., a spore-forming lactic acid bacterium isolated from tree bark in Thailand.

A Gram-stain-positive, facultatively anaerobic and endospore-forming rod-shaped bacterium, designed strain CPB3-1T, was isolated from tree bark. This homofermentative strain produced dl-lactic acid from glucose. It grew at 20-45 °C, pH 4.0-9.5 and in 0-3.0 % (w/v) NaCl. It contained meso-diaminopimelic acid in cell-wall peptidoglycan and had menaquinone with seven isoprene units (MK-7) as the predominant component. The major fatty acid was anteiso-C17 : 0. The polar lipids were phosphatidylglycerol, diphosphatidylglycerol, an unknown phospholipid and an unknown lipid. Based on the results of 16S rRNA gene sequence analysis, strain CPB3-1T belonged to the genus Sporolactobacillus and was closely related to Sporolactobacillus kofuensis DSM 11701T and Sporolactobacillus spathodeae BK117-1T (both 96.7 % similarity), Sporolactobacillus inulinus NRIC 1133T and Sporolactobacillus terrae DSM 11697T (both 96.6 % similarity), and Sporolactobacillus shoreicorticis MK21-7T, Sporolactobacillus laevolacticus DSM 442T, Sporolactobacillus shoreae BK92T and Sporolactobacillus pectinivorans GD201205T (all 95.8-96.5 % similarity). The draft genome of strain CPB3-1T contained 2 930 919 bps with 3117 coding genes. The DNA G+C content was 45.1 mol%. The digital DNA-DNA hybridization values between strain CPB3-1T and closely related type strains were 19.2-24.0 %. The average nucleotide identity (84.0-87.6 %) and average amino acid identity (66.5-76.3 %) values were lower than the cut-off values for species delineation. Strain CPB3-1T was clearly distinguished from related Sporolactobacillus species based on its phenotypic and chemotaxonomic characteristics, 16S rRNA gene sequence similarity and the results of draft genome analysis. Therefore, the strain represents a novel species of the genus Sporolactobacillus, for which the name Sporolactobacillus mangiferae sp. nov. is proposed. The type strain is CPB3-1T (=JCM 35082T=TISTR 10004T).

Genome sequences and functional analysis of Levilactobacillus brevis LSF9-1 and Pediococcus acidilactici LSF1-1 from fermented fish cake (Som-fak) with gamma-aminobutyric acid (GABA) production.

Gamma-aminobutyric acid (GABA) is a crucial inhibitory neurotransmitter in the sympathetic nervous system that exerts regulatory effects on the blood, immune, and nervous systems. GABA production in som-fak, a traditional fermented fish of Thailand, has been attributed to the activity of lactic acid bacteria (LAB). The present study aims to characterize the LAB isolates and compare the genomes and GABA synthesis genes of selected isolates capable of GABA production. Thirteen isolates demonstrating GABA synthesis capability were identified based on their phenotypic and genotypic characteristics. Seven isolates (group I: LSF3-3, LSF8-3, LSF9-1, LSF9-3, LSF9-6, LSF9-7, and LSF10-14) were identified as Levilactobacillus brevis with 99.78-100% similarity. LSF2-1, LSF3-2, LSF5-4, and LSF6-5 (group II) were identified as Lactiplantibacillus pentosus with 99.86-100% similarity. Strain LSF1-1 (group III) was identified as Pediococcus acidilactici (99.47%), and LSF10-4 (group IV) was identified as Pediococcus pentosaceus with 99.93% similarity. The GABA production of isolates ranged from 0.087 to 16.935 g/L. The maximum production of 16.935 g/L from 3% monosodium glutamate was obtained from strain LSF9-1. Gene and genome analysis revealed that L. brevis LSF9-1 has multiple gad genes in the genome, such as gadB1, gadB2, gadC1, and gadC2, making it the potential strain for GABA production. Additionally, the genome analysis of P. acidilactici LSF1-1 consists of gadA, gadB, and gadC, which respond to controlling GABA production and export. Furthermore, strain LSF1-1 was considered safe, containing no virulence factors. Thus, Levilactobacillus brevis LSF9-1 and Pediococcus acidilactici LSF1-1 have the potential for GABA production and probiotic use in future studies.

Genome analysis and optimization of γ-aminobutyric acid (GABA) production by lactic acid bacteria from plant materials.

Gamma-aminobutyric acid (GABA) plays a key role as an inhibitory neurotransmitter in the mammalian sympathetic nervous system and has other health benefits. Molecular characterization, genome analysis, and optimization were investigated to improve GABA production of a selected strain of lactic acid bacteria. Eleven isolates from plant materials were screened for GABA productivity and were identified based on phenotypic and genotypic characteristics. The most potent strain was chosen for genome analysis and GABA production optimization using the response surface methodology (RSM). Each of the two strains was closely related to Lactobacillus plantarum, Lactobacillus brevis, Weissella cibaria, Leuconostoc pseudomesenteroides while each strain was similar to Lactobacillus pentosus, Enterococcus lactis, and Leuconostoc mesenteroides. They produced GABA ranging from 0.036 ± 0.000 to 17.315 ± 0.171 g/L at 72 h-cultivation. Among them, the most potent strain, SL9-6, showed the highest GABA production (17.315 g/L) when cultivated with 10% (v/v) inoculum for 48 h. The draft genome sequence of strain SL9-6 exhibited 96.90% average nucleotide identity value and 74.50% digital DNA-DNA hybridization to Lactobacillus brevis NCTC 13768T. This strain contained a glutamate decarboxylase gene system (gadA, gadB, and gadC). Optimal culture conditions were determined as 40.00 g/L glucose, 49.90 g/L monosodium glutamate, pH 5.94, and 31.10°C by RSM, giving maximum GABA production of 32.48 g/L. Results from RSM also indicated that monosodium glutamate concentration, pH, and temperature were significant variables. GABA production significantly improved here could promise further application of strain SL9-6.

Characterization and comparative genomic analysis of gamma-aminobutyric acid (GABA)-producing lactic acid bacteria from Thai fermented foods.

OBJECTIVES: This study aimed to screen, characterize, and annotate the genome along with the comparison of GABA synthesis genes presented in lactic acid bacteria (LAB). RESULTS: Thirty-five LAB isolates from fermented foods were screened for GABA production using thin-layer chromatography (TLC). Fifteen isolates produced GABA ranging from 0.07 to 22.94 g/L. Based on their GTG5 profiles, phenotypic, and genotypic characteristics, isolates LSI1-1, LSI1-5, LSI2-1, LSI2-2, LSI2-3, LSI2-5, and LSM3-1-4 were identified as Lactobacillus plantarum subsp. plantarum; isolate LSM1-4 was Lactobacillus argentoratensis; isolates CAB1-2, CAB1-5, CAB1-7, and LSI1-4 were Lactobacillus pentosus; and CAB1-1, LSM3-1-1 and LSM3-2-3 were Lactobacillus fermentum. Strains LSI2-1 and CAB1-7 from pickled vegetables were selected for genome analysis. The gadA gene (1410 bp, 470aa) was encountered in GABA production of both strains and no other glutamate decarboxylase (GAD) genes were found in the genomes when compared with other LAB strains. The presence of gadA is evidence for GABA production. Strains LSI2-1 and CAB1-7 produced 22.94 g/L and 11.59 g/L of GABA in GYP broth supplemented with 3% (w/v) MSG at 30 °C for 72 h, respectively. CONCLUSIONS: Our report highlights the characterization of LAB and GABA production of L. plantarum LSI2-1 strain with its GABA synthesis gene. GABA production of strains LSI2-1 and CAB1-7 in GYP broth with 3% (w/v) MSG and comparative GAD genes.

Secundilactobacillus folii sp. nov., isolated from fermented tea leaves in Thailand.

A Gram-stain-positive, catalase-negative, rod-shaped, non-motile, non-spore-forming, and facultatively anaerobic strain CRM56-3T, isolated from fermented tea leaves collected from Chiang Rai province, Thailand, was characterized based on a polyphasic approach. The strain produced dl-lactic acid heterofermentatively from glucose. It grew at 15-42 °C (optimum at 30 °C), pH 3.5-8.0 (optimum pH 6.0) and in 1-4 % (w/v) NaCl. Strain CRM56-3T contained C16:0, C19:0 cyclo ω8c, and C18:1 ω7c, and/or C18:1 ω6c as major cellular fatty acids. Based on 16S rRNA gene sequence analysis, strain CRM56-3T belongs to the genus Secundilactobacillus and was closely related to Secundilactobacillus odoratitofui DSM 19909T (99.2 %), S. collinoides JCM 1123T (98.9 %), and S. paracollinoides DSM 15502T (98.7 %). The draft genome of strain CRM56-3T contained 2681617 bp with 2413 coding sequences and DNA G+C content determined from genome sequence of 44.5 mol%. The digital DNA-DNA hybridization (dDDH) between strain CRM56-3T and S. odoratitofui DSM 19909T, S. collinoides JCM 1123T, and S. paracollinoides DSM 15502T were 19.5, 20.4, and 21.6 %, respectively. The average nucleotide identity (ANIm) and the average amino acid identity (AAI) between strain CRM56-3T and closely related strains were lower than 85.0 and 80.0 %, respectively. The strain CRM56-3T was clearly distinguished from related Secundilactobacillus species by its phenotypic and chemotaxonomic characteristics, 16S rRNA gene sequence similarity, and the draft genome analysis. Therefore, the strain represents a novel species of the genus Secundilactobacillus, for which the name of Secundilactobacillus folii sp. nov. is proposed. The type strain is CRM56-3T (=JCM 34223T=LMG 31663T=TISTR 2851T).

Characterization of a novel Clostridium sp. SP17-B1 and its application for succinic acid production from hevea wood waste hydrolysate.

An anaerobic, gram-positive, rod-shaped bacterium strain SP17-B1, isolated from dog saliva, was taxonomically characterized on the basis of phenotypic, chemotaxonomic, and genotypic characteristics. It was cultured in 4% (w/v) NaCl at a pH range of 5.0-8.0 (optimally at pH 7) and at 30°C-40 °C (optimally at 37 °C). Its major cellular fatty acids are C16:0 (36.3%), C17:0 cyclo (9.7%), C16:1ω9c (13.9%), and C18:1ω9c (10.7%), and its DNA guanine-cytosine content is 40.8 mol%. On the basis of the 16S rRNA gene sequence analysis, it was determined that the strain belonged to the genus Clostridium and was closely related to C. amygdalinum BR-10T (97.8%), C. saccharolyticum WM1T (97.8%), and C. celleracrescens DSM 5628T (97.7%). This strain showed a low level of DNA-DNA relatedness with the closely related strains, suggesting that it is a novel species in the genus Clostridium. Recent studies have demonstrated the production of succinic acid using Clostridium strains. Strain SP17-B1 produced 25.1 ±â€¯1.3 and 15.3 ±â€¯1.5 g/L of succinic acid from 40 g/L of glucose and 30 g/L of hevea wood waste hydrolysate (HH), respectively, after 24 h. When detoxified HH was used as a substrate, the lag phase was reduced and cell growth was enhanced by 7 fold (OD660 0.4-3.0) within 12 h. Detoxification using granular activated carbon may have reduced the levels of furfural and HMF without interfering with the amount of sugars in HH.