Molecular selection and the taxonomy of rhizobacteria which suppress the growth of Fusarium graminearum on maize (Zea mays L.)

MSc (Biology), North-West University, Mafikeng CampusThe disease-causing agent Fusarium graminearum is considered a global cause of distress to the agricultural economy due to its undesirable effects and its ubiquity in the soil leading to infections in cereal grains. Therefore, healthier and effective inoculant alternatives are necessary to improve the currently applied practices. Plant growth-promoting rhizobacteria (PGPR) are some of the naturally occurring microbes that have a broad spectrum of activities that aid with plant growth and protection. This study investigated the cultural and molecular characteristics of novel Bacillus spp. that are commonly studied as PGPR that are known to attain antagonistic traits effective against maize plant pathogens such as F. graminearum. Forty-six (46) bacteria were successfully isolated from the rhizosphere of healthy maize crops in the North West Province, South Africa. Among these, only 3 strains, LBC4ME, LBC22HL and LBC35MW were found to be highly effective when tested for antifungal activities, particularly against the F. graminearum FG1 strain. These strains were further tested for plant growth-promoting (PGP) traits [Siderophores, indole-acetic-acid (IAA), phosphate solubilization, hydrogen cyanide (HCN), ammonia (NH3) production, nitrogen fixation, hydrolytic enzyme production], antibiotic sensitivity using [Vancomycin (VA), chloramphenicol (C), kanamycin (K), tetracycline (TET), erythromycin (E) and ampicillin (AMP)], and biochemical traits. Molecular approaches (16S rDNA, rpoB, gyrB and tuf) of partial gene sequences were further used for the identification of the isolates and partial gene sequences (ItuD1, Bacc1, AS1, AF2) were used for the detection of lipopeptide biosurfactants biosynthesis. The results showed that LBC4ME, LBC22HL and LBC35MW strains exhibited a substantial fungal inhibition rate above 50% compared to the other isolated bacteria. The taxonomy of the obtained strains was identified as the Firmicutes phyla, genus Bacillus species LBC4ME (Bacillus thuringiensis), LBC22HL (Bacillus velezensis) and LBC35MW (Bacillus thuringiensis). The three (3) strains were also found to produce a broad range of antifungal activities through in vitro and in vivo tests. Antifungal efficiency validated by using pot experiments in the screen-house showed that the treatments containing individual antifungal bacterial cells and the treatments containing combined bacterial cells of the studied strains had a positive effect on disease severity (root length, shoot length, leaf rot, plant wilting) compared to the pathogen only and control treatments. The strains also showed effective antifungal inhibition zones against F. graminearum FG1 under different growth conditions [pH (2-10), temperature (4-45 °C), carbon (Glucose, D-fructose, sucrose, and D-galactose) and nitrogen sources (KNO3, NH4NO3, (NH4)2SO4, and urea)]. Viable production of antifungal lipopeptide biosurfactants was displayed for surfactins, iturins, and fengycins molecular characteristics using chromatographic (Thin-layer chromatography (TLC)), spectroscopic analysis (Fourier transform infrared (FTIR) and the Electrospray ionization-mass spectrometry (ESI-MS)) analysis for the detection of surface-active lipopeptide molecules. The lipopeptide biosurfactants extracted from LBC4ME, LBC22HL and LBC35MW strains showed antifungal properties which demonstrated about 65-66% inhibition rate against F. graminearum FG1. The results obtained from this study have shown that the above-mentioned strains attained stable multipotent broad spectrum of antimicrobial activities and may be targeted and further used as potential biological control agents (BCAs) for F. graminearum, and thus, contribute to improving food safety and security.Master