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Access type: UnknownAuthor: search.filters.author.Nuwong, Warinya

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    Correlation of Antifungal Activities and Nonribosomal Peptide Synthetase Gene Expression of Bacillus siamensis 1021
    (2022-03-01)
    Nuwong, Warinya
    ;
    Kittiwongwattana, Chokchai
    Members of the genus Bacillus were biocontrol agents against various phytopathogenic fungi. Several factors affected the regulation and production of nonribosomal peptides (NRPs) in Bacillus species. The aim of this study was to examine how culture media influenced the antagonistic activities and gene expression of Bacillus siamensis 1021. The bacterial supernatant from potato dextrose broth (PDB) exhibited higher inhibitory effects against mycelia of Pyricularia oryzae and Colletotrichum fructicola, when compared with those from nutrient broth (NB) and minimal salt medium (MSM). However, the biocontrol activities of these supernatants were not significantly different when they were tested against P. oryzae and C. fructicola conidia. Additionally, the bacterial supernatant from PDB significantly reduced the disease severity caused by P. oryzae in rice seedlings when compared with the water-treated control group. The genome of strain 1021 was sequenced, and biosynthetic gene clusters of nonribosomal peptides surfactin, fengycin and bacillibactin were identified. Effects of PDB, NB and MSM media on expression levels of core biosynthetic genes of surfactin (srfAA), fengycin (fenC) and bacillibactin (dhbE) gene clusters were determined by reverse-transcription quantitative PCR. fenC expression was significantly increased in PDB and corresponded with the antagonistic activities against fungal mycelia. Conversely, expression of the regulatory genes comA and codY in PDB were highly reduced in PDB, indicating their negative relation with fenC expression. This expression analysis indirectly suggested that fengycin was potentially the bioactive compound of B. siamensis 1021 against P. oryzae and C. fructicola.
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    Validation of reference genes for the normalization of RT-qPCR gene expression in Bacillus siamensis 1021 grown in different culture media
    (2022-01-01)
    Nuwong, Warinya
    ;
    Kittiwongwattana, Chokchai
    Background and Objectives: House-keeping genes are generally selected as reference genes in gene expression analysis. However, some genes may not be stably expressed across all experimental conditions. Thus, this study aimed to validate seven house-keeping genes for gene expression analysis in Bacillus siamensis 1021. Materials and Methods: Strain 1021 was grown in potato dextrose broth, nutrient broth and mineral salt medium. Re-verse-transcription quantitative PCR was used to determine Cq values of seven reference genes including gyrA, gyrB, ssb and dnaB, rpsU, gat_Yqey and udp in these media. Expression stability of these genes was analyzed, using geNorm and Normfinder applications. The target gene ftsZ was used for assessment of the best candidate genes. Results: Based on geNorm and Normfinder, ssb was the most-stably expressed gene, while udp was the least-stably expressed gene. Pairwise variation indicated the combination of ssb, gyrA, gyrB and gatB_Yqey was suitable for the normalization of ftsZ expression. ftsZ expression in potato dextrose broth and mineral salt medium was higher than that in nutrient broth. In contrast, the normalization against udp resulted in an under-and overestimation of ftsZ expression in potato dextrose broth and mineral salt medium, respectively. Conclusion: The combination of ssb, gyrA, gyrB and gatB_Yqey was the best candidate for normalization of target gene expression in B. siamensis 1021 in these media. This study emphasized the significance of reference gene validation for gene expression analysis and provided a guideline for future gene expression studies in B. siamensis.