Genome analysis and plant growth promoting activity of Pseudonocardia strain DR1-2 from the root of Dendrobium christyanum Rchb.f

Aims@#The objective of this study was to analyze the genome of endophytic actinomycete associated with orchids and evaluate its plant hormone activities, including phytohormone, siderophore, ammonia production, zinc and phosphate solubilization.@*Methodology and results@#Strain DR1-2 isolated from the roots of the Thai orchid, Dendrobium christyanum Rchb.f., was closely related to Pseudonocardia alni DSM 44104T, P. antarctica DSM 44749T and P. carboxydivorans Y8T (99.93-100% similarity) based 16S rRNA gene sequence. This strain exhibited IAA production (294.10 ± 12.17 μg/mL), phosphate solubilization (2.20 ± 0.08 solubilization Index, SI), positive for siderophore production and ammonia production (36.99 ± 2.24 μg/mL). It showed a maximum IAA of 489.73 ± 8.90 μg/mL, when optimized using 0.5% Ltryptophan, pH 6 and incubated at 30 °C for 7 days. The IAA of strain enhanced the root length, shoot length, number of roots and fresh weight of rice seedlings (Oryza sativa L. cv. RD49). The draft genome of strain DR1-2 was 6,077,423 bp in 23 contigs with G+C content of 74.6%. The average nucleotide identity-Blast (ANIb) and average nucleotide identity-MUMmer (ANIm) values of strain DR1-2 and related type strains were 95.81 to 97.25% and the digital DNA-DNA hybridization (dDDH) values were 72.60 to 74.00%, respectively. Genomic analysis of strain DR1-2 revealed that the gene encodes the enzyme involved in the phytohormones biosynthesis and gene clusters involved in the biosynthesis of bioactive metabolites.@*Conclusion, significance and impact of study@#Endophytic actinomycete, Pseudonocardia strain DR1-2 from Thai orchid, D. christyanum Rchb.f., exhibited significant IAA production and affected the growth of the plant, which was the potential source of plant hormones for agricultural applications.

Identification and characterization of plant growth-promoting properties of bacterial endophytes from selected Zingiberaceae plants

Aims@#This study aims to isolate, characterize and screen the plant growth-promoting bacteria from Zingiberaceae plants. Plant promoting activities such as indole-3-acetic acid (IAA), phosphate solubilization, zinc solubilization and nitrogen-fixing capabilities are determined, and the IAA production of selected isolates are optimized. @*Methodology and results@#Endophytic bacteria were isolated from the plant samples by surface sterilization on nutrient agar (NA) plates and incubated at 30 °C for 2-3 days. The bacteria were identified based on their phenotypic characteristics and 16S rRNA gene sequence analyses. All isolates were identified as genera Bacillus, Lysinibacillus, Kerstersia, Klebsiella and Brucella. The isolates exhibited phosphate solubilization (1.5 ± 0.75-37.5 ± 8.75 Solubilization Index, SI), zinc solubilization (2.5 ± 0-60 ± 1.5 SI) and IAA production (0.1 ± 0.2-115.7 ± 1.6 µg/mL), while 3 isolates possessed nitrogen-fixing capabilities. Five isolates (PHAS-2, PWS-2, PWR-2, PHBS-2 and SCG-2) were selected for IAA optimization. Isolate PWR-2 produced the maximum IAA at 447.7 ± 0 µg/mL when tryptophan concentration was maintained at 1.0%.@*Conclusion, significance and impact of study@#Genera of bacteria included Bacillus, Lysinibacillus, Kerstersia, Klebsiella and Brucella were successfully isolated from Zingiberaceae plants. All the isolates showed the capability to produce IAA, while some isolates exhibited phosphate solubilization and zinc solubilization, and a few possessed nitrogen-fixing capabilities. The potential IAA production isolates could be applied for the enhancement of agricultural production that will be becoming a more widely accepted practice.

Characterization and comparison of phytase production by Bacillus and Paenibacillus strains from Thai soils

Aims: The objective of this research was to isolate, screen and identify phytase-producing bacteria from soils and a potent isolate was selected for its phytase production. Methodology and results: Eight spore-forming bacteria isolated from agricultural soils in Thailand were screened for their phytase production. They were identified as Bacillus and Paenibacillus strains based on their phenotypic characteristics and 16S rRNA gene sequence analyses. The phytase production by Bacillus amyloliquefaciens CH3-1 [Group I(a)] was 20.956 ± 0.099 U/mL, while Bacillus subtilis SR9-3 [Group I(b)] produced 20.588 ± 0.099 U/mL. Five isolates in Group I(c), identified as Bacillus aryabhattai, produced phytase at levels ranging from 2.436 ± 0.116 to 20.910 ± 0.000 U/mL, while Paenibacillus cineris CM5-3 (Group II) produced 1.261 ± 0.111 U/mL. A potent strain, CH3-1, produced the highest phytase when cultivated in Phytate Specific Medium (PSM) supplemented with 1% glucose, at pH 7.0 and incubated at 45 °C. Additionally, wheat bran and sorghum seed (0.5%) substrates were used to induce phytase production by replacing Na-phytate. Conclusion, significance and impact of study: Phytase producing bacteria were isolated from soils in Thailand. Gram-positive spore forming thermotolerant Bacillus strains displayed higher phytase activity than a Paenibacillus strain. A potent strain, CH3-1, could utilize agricultural waste as a substrate, which may be useful for animal feed supplementation.

Identification and lactic acid production of bacteria isolated from soils and tree barks

Aims: The objective of this research was to isolate and identify lactic acid producing bacteria from soils and tree barks in Thailand. Their acid production was also determined. Methodology and results: Eleven bacterial strains isolated from soils and tree barks were screened for their lactic acid production. They were divided into 4 groups based on their phenotypic characteristics and 16S rRNA gene sequence analyses. Group I (3 isolates) identified as Lactococcus produced L-lactic acid ranged from 72.32 ± 0.707-77.47 ± 0.184 g/L, yield of 0.96 ± 0.011-1.06 ± 0.008 g/g, productivity of 1.00 ± 0.010-1.08 ± 0.003 g/L.h and optical purity was 100%. Group II (3 isolates) identified as Enterococcus hirae, produced L-lactic acid ranged from 31.56 ± 0.424-34.86 ± 0.283 g/L, yield of 1.06 ± 0.008-1.23 ± 0.036 g/g, productivity of 0.44 ± 0.006-0.48 ± 0.004 g/L.h and optical purity was 98.6- 100%. Group III (3 isolates) identified as Bacillus coagulans, produced L-lactic acid ranged from 48.48 ± 0.283-93.51 ± 0.552 g/L, yield of 1.00 ± 0.001-1.07 ± 0.005 g/g, productivity of 0.67 ± 0.004-1.30 ± 0.008 g/L.h and high optical purity of the tested isolate was 99.56%. Group IV (2 isolates) identified as Sporoactobacillus. Only NN2 produced D-lactic acid concentration 87.64 ± 0.375 g/L, yield of 0.83 ± 0.000 g/g, productivity of 1.22 ± 0.005 g/L.h and optical purity was 96.16%. All isolates produced lactic acid when 120 g/L of the initial glucose was used as a substrate. Conclusion, significance and impact of study: Lactic acid producing bacteria are isolated from soils and tree barks. The coccal isolates could produce high L-lactic acid concentration with high optical purity while the spore-forming isolates produces L- and D-lactic acid which are useful for food preservation, chemical in pharmaceutical, cosmetic and textile industries.

Characterization and amylolytic activity of yeast and mold strains from Thai sweet rice

Aims: The aim of this study was to isolate and identify the yeast and mold strains from the starter and to investigate their amylolytic activity. Methodology and results: Thirty-two yeasts were isolated from ten samples of look-paeng khao-mak, a traditional starter culture in the production of khao-mak (sweet rice) in several places in Thailand. All isolates were identified based on their morphological and biochemical characteristics including the sequencing of D1/D2 domain of large subunit (26S) ribosomal DNA analyses. They were identified as Saccharomycopsis fibuligera (9 isolates), Candida rugosa (2 isolates), C. tropicalis (1 isolate), Clavispora lusitaniae (1 isolate), Wickerhamomyces anomalus (15 isolates) and Meyerozyma guilliermondii (4 isolates). All isolates of S. fibuligera showed high amylolytic activity. One-hundred isolates of mold were isolated from twenty-one samples of look-paeng khao-mak. They were belonged to Amylomyces sp. (42 isolates), Rhizopus sp. (30 isolates), Mucor sp. (12 isolates) and Penicillium sp. (16 isolates) based on their morphological characteristics. Four isolates, LK4-1, LK8-2, LK12-5 and LK17-1 showed amylase activity ranged from 32.24 to 39.74 unit/mL by dinitrosalicylic acid (DNSA) method. The isolates LK4-1, LK8-2 and LK12-5 were identified as Amylomyces rouxii and LK17-1 was Rhizopus microsporus based on their morphological characteristics and the ribosomal RNAencoding DNA (rDNA) internal transcribed spacer (ITS) sequences. Conclusion, significance and impact study: The characterization and evaluation of yeast and mold species based on their phenotypic and genetic characteristics including their amylase activity will be useful for the diversity and sweet rice production.

Identification and characterization of lipolytic bacteria from Thai fermented foods

Aims: Gram-positive spore forming rod-shaped bacteria from Thai fermented foods were isolated, identified and screened for lipolytic activity. Methodology and results: Bacterial strains were isolated from Thai fermented foods by the standard dilution technique using Tryptic soy agar. Seven isolates which belong to the genus Bacillus and one isolate which belongs to Paenibacillus were characterized based on their phenotypic characteristics and 16S rRNA gene sequence analyses. Isolates NM8-1 (Group 1), PR9-2 (Group 2), PR11-1 (Group 3), KM15-1 and SS49-4 (Group 4), SS48-5 (Group 5) and SS503 (Group 6), were closely related to Bacillus methylotropicus, B. pumilus, B. flexus, B. cereus, B. subtilis and B. anthracis, based on 99.90-100% similarities, respectively. Isolate NM45-3 (Group 7) was closely related to Paenibacillus pasadenensis based on 99.55% similarity. All the isolates were susceptible to amikacin, chloramphenicol, clindamycin, erythromycin, gentamicin, imipenem, kanamycin, levofoxacin, novobiocin, streptomycin, tetracycline and vancomycin. Their lipase production in nutrient broth with Tween 20, Tween 40, Tween 60 or Tween 80 ranged from 0.014±0.129 - 3.231±0.087 U/mL. Bacillus subtilis SS48-5 exhibited highest lipase production when cultured with Tween 80 at pH 7.5 for 24 h. The optimum lipase production of this strain was at 40 °C after incubated for 30 h. Conclusion, significance and impact of study: The characterization and evaluation of the lipolytic activity of Bacillus and Paenibacillus strains isolated from Thai fermented foods will be useful for the species diversity, food fermentation and the lipase production.