Isolation and identification of antimicrobial compounds produced by Bacillus nakamurai CBAS-959 and their activity against phytopathogenic bacteria

IF 5.2 2区 农林科学 Q1 AGRICULTURE, MULTIDISCIPLINARY
Carmen Julia Pedroza-Padilla, Sergio Orduz, Luiz Alberto Beraldo Moraes
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Abstract

Background

The application of beneficial microorganisms is an effective solution for the control of crop diseases caused by phytopathogenic bacteria. Bacillus sp. plays a significant role as a biofactory of antimicrobial compounds, and bioformulations containing species of the Bacillus subtilis complex are commercialized. However, there is limited information on the chemical structural diversity of antimicrobial compounds produced by Bacillus nakamurai and its own potential use in agriculture as a suppressor agent of phytopathogenic bacteria.

Results

In this study, the bacterium strain CBAS-959, which produces diffusible black pigmentation in the growth medium, was identified as Bacillus nakamurai through biochemical characterization and the analysis of the 16S rRNA and gyrA genes. Co-culture assays confirmed a strong antagonistic effect against Ralstonia solanacearum and Pectobacterium carotovorum, producing inhibition halos of 23.9 ± 1.8 mm and 23.2 ± 1.8 mm, respectively. The crude extract and pure fractions analyzed by MS/MS mass spectrometry and evaluated by disk diffusion assays demonstrated that B. nakamurai CBAS-959 produces iturin A, bacillibactin, bacillaene, bacillaene B, and dihydrobacillaene, and five isoforms with different surfactin homologs. Extracts obtained from PDB and M1 media showed zones of inhibition of phytopathogenic bacteria between 9.5 ± 0.5 and 23.5 ± 0.5 mm, while fractions of purified compounds between 13.2 ± 0.8 and 21.0 ± 2.6 mm. Bacillibactin showed a MIC of 31.25 µg/mL against R. solanacearum and P. carotovorum but did not inhibit X. citri. PDB-2 L extract was the most active against X. citri, with an MIC of 62.5 µg/mL.

Conclusions

This is the first study that purified and demonstrated that bacillibactin has a direct antibacterial effect against R. solanacearum and P. carotovorum. In addition, this work reports that B. nakamurai CBAS-959 produces bacillaene B, five isoforms of surfactins, inhibits the growth of R. solanacearum on co-culture, and the extracts obtained in PDB and M1 inhibit the growth of X. citri. Therefore, this research provides new information on the metabolic diversity and antimicrobial activity of B. nakamurai CBAS-959 and its potential use for future studies to explore new alternatives for the control of phytopathogens.

Graphical Abstract

中村芽孢杆菌CBAS-959抗菌化合物的分离鉴定及其抗植物病原菌活性研究
背景有益微生物的应用是防治植物病原菌引起的作物病害的有效途径。芽孢杆菌作为抗菌化合物的生物工厂发挥着重要作用,含有枯草芽孢杆菌复合物的生物制剂已被商业化。然而,关于中村芽孢杆菌产生的抗菌化合物的化学结构多样性及其在农业中作为植物致病菌抑制剂的潜在用途的信息有限。结果通过生化鉴定和16S rRNA和gyrA基因分析,本研究鉴定的菌株CBAS-959为中村芽孢杆菌(Bacillus nakamurai)。共培养实验证实,该菌对茄枯菌(Ralstonia solanacearum)和胡萝卜乳杆菌(Pectobacterium carotovorum)具有较强的拮抗作用,分别产生23.9±1.8 mm和23.2±1.8 mm的抑制晕。采用MS/MS质谱法和光盘扩散法对粗提物和纯组分进行了分析,结果表明,B. nakamurai cba -959可产生iturin A、bacillibactin、bacillaene、bacillaene B和dihydrobacillaene,以及5种具有不同表面素同源物的异构体。PDB和M1培养基提取物对植物致病菌的抑制区在9.5±0.5 ~ 23.5±0.5 mm之间,纯化后的化合物提取物对植物致病菌的抑制区在13.2±0.8 ~ 21.0±2.6 mm之间。Bacillibactin对番茄红霉(R. solanacearum)和胡萝卜红霉(P. carotovorum)的MIC为31.25µg/mL,对柑橘红霉(X. citri)没有抑制作用。PDB-2 L提取物对柑桔的抑制作用最强,MIC为62.5µg/mL。结论本研究首次纯化并证实了杆菌素对茄青霉和胡萝卜霉具有直接抑菌作用。此外,本工作报道了B. nakamurai c巴斯-959在共培养中产生5种表面素异构体bacillaene B,抑制茄青霉的生长,PDB和M1中获得的提取物抑制X. citri的生长。因此,本研究为B. nakamurai CBAS-959的代谢多样性和抗菌活性提供了新的信息,并为未来的研究探索控制植物病原体的新选择提供了潜在的应用价值。图形抽象
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来源期刊
Chemical and Biological Technologies in Agriculture
Chemical and Biological Technologies in Agriculture Biochemistry, Genetics and Molecular Biology-Biotechnology
CiteScore
6.80
自引率
3.00%
发文量
83
审稿时长
15 weeks
期刊介绍: Chemical and Biological Technologies in Agriculture is an international, interdisciplinary, peer-reviewed forum for the advancement and application to all fields of agriculture of modern chemical, biochemical and molecular technologies. The scope of this journal includes chemical and biochemical processes aimed to increase sustainable agricultural and food production, the evaluation of quality and origin of raw primary products and their transformation into foods and chemicals, as well as environmental monitoring and remediation. Of special interest are the effects of chemical and biochemical technologies, also at the nano and supramolecular scale, on the relationships between soil, plants, microorganisms and their environment, with the help of modern bioinformatics. Another special focus is the use of modern bioorganic and biological chemistry to develop new technologies for plant nutrition and bio-stimulation, advancement of biorefineries from biomasses, safe and traceable food products, carbon storage in soil and plants and restoration of contaminated soils to agriculture. This journal presents the first opportunity to bring together researchers from a wide number of disciplines within the agricultural chemical and biological sciences, from both industry and academia. The principle aim of Chemical and Biological Technologies in Agriculture is to allow the exchange of the most advanced chemical and biochemical knowledge to develop technologies which address one of the most pressing challenges of our times - sustaining a growing world population. Chemical and Biological Technologies in Agriculture publishes original research articles, short letters and invited reviews. Articles from scientists in industry, academia as well as private research institutes, non-governmental and environmental organizations are encouraged.
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