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•Occidiofungins (Ocfs) are essential for the antifungal activity of B. ambifaria T16.•T16 produces four Ocfs variants with or without xylose addition.•T16 is able to synthesize a ...membrane-associated antifungal lipopeptide (AFC-BC11)•Ocfs biosynthesis involves a multifunctional enzyme, containing a FAAL domain.•The afc-BC11 cluster includes a FAAL domain containing enzyme and a type III PKS.
Burkholderia ambifaria T16 is a bacterial strain isolated from the rhizosphere of barley plants, which showed a potent antifungal activity against several Fusarium species. In this work, the construction of a mini-Tn5 insertional library in this strain allowed us the identification of several genes involved in the antifungal activity. In particular, several mutants with mini-Tn5 insertions located in a biosynthetic gene cluster (BGC) involved in the production of cyclic glycolipopeptides known as occidiofungins (Ocfs) were completely unable to inhibit the in vitro growth of F. oxysporum. Tandem mass spectrometry analysis of lipopeptide extracts obtained from culture supernatants of the wild-type strain allowed the identification of eight Ocfs structural variants, that were not detected in extracts of the mutants with insertions in the ocf gene cluster. These results indicate that the antifungal activity of B. ambifaria T16 mainly relies on Ocfs production. Moreover, an insertion in a BGC predicted to be involved in the biosynthesis of a membrane-associated lipopeptide designated AFC-BC11 reduced the antifungal activity of B. ambifaria T16 by approximately 50%. Accordingly, membrane extracts from the insertional mutant in the afc-BC11 cluster showed a marked reduction in the antifungal activity against several fungi. In contrast, the antifungal activity of the membrane fraction of mutants unable to produce Ocfs was not affected. Furthermore, a detailed bioinformatic analysis of the proteins encoded by the ocf and afc-BC11 gene clusters revealed that the biosynthesis of these lipopeptides would be initiated by fatty acyl-AMP ligase (FAAL) domain containing enzymes and continued by a hybrid non-ribosomal peptide synthetase (NRPS)/polyketide synthase (PKS) or a type III PKS, respectively. These results provide new insights into lipopeptide biosynthesis, which may be valuable in the field of novel antifungal compound design.
Burkholderia ambifaria
T16 is a bacterium isolated from the rhizosphere of barley plants that showed a remarkable antifungal activity. This strain was also able to degrade fusaric acid ...(5-Butylpyridine-2-carboxylic acid) and detoxify this mycotoxin in inoculated barley seedlings. Genes and enzymes responsible for fusaric acid degradation have an important biotechnological potential in the control of fungal diseases caused by fusaric acid producers, or in the biodegradation/bio catalysis processes of pyridine derivatives. In this study, the complete genome of
B. ambifaria
T16 was sequenced and analyzed to identify genes involved in survival and competition in the rhizosphere, plant growth promotion, fungal growth inhibition, and degradation of aromatic compounds. The genomic analysis revealed the presence of several operons for the biosynthesis of antimicrobial compounds, such as pyrrolnitrin, ornibactin, occidiofungin and the membrane-associated AFC-BC11. These compounds were also detected in bacterial culture supernatants by mass spectrometry analysis. In addition, this strain has multiple genes contributing to its plant growth-promoting profile, including those for acetoin, 2,3-butanediol and indole-3-acetic acid production, siderophores biosynthesis, and solubilisation of organic and inorganic phosphate. A pan-genomic analysis demonstrated that the genome of strain T16 possesses large gene clusters that are absent in the genomes of
B. ambifaria
reference strains. According to predictions, most of these clusters would be involved in aromatic compounds degradation. One genomic region, encoding flavin-dependent monooxygenases of unknown function, is proposed as a candidate responsible for fusaric acid degradation.
Fusaric acid (FA) is an important virulence factor produced by several
species. These fungi are responsible for wilt and rot diseases in a diverse range of crops. FA is toxic for animals, humans and ...soil-borne microorganisms. This mycotoxin reduces the survival and competition abilities of bacterial species able to antagonize
spp., due to its negative effects on viability and the production of antibiotics effective against these fungi. FA biodegradation is not a common characteristic among bacteria, and the determinants of FA catabolism have not been identified so far in any microorganism. In this study, we identified genes, enzymes, and metabolic pathways involved in the degradation of FA in the soil bacterium
T16. Our results provide insights into the catabolism of a pyridine-derivative involved in plant pathogenesis by a rhizosphere bacterium.
