Identification of genes responsible for the potent antifungal activity of Burkholderia ambifaria T16

FELDMAN N; ALVAREZ F; SIMONETTI E; RUIZ JA; VINACOUR M

San Luis

Congreso; . XIII Congreso Argentino de Microbiología General SAMIGE. San Luis. Argentina. 8 al 10 de agosto de 2018; 2018

SAMIGE

Burkholderia ambifaria T16 is a bacterial strain isolated from the rhizosphere of barley which showed theability to inhibit mycelial growth and conidial germination of several phytopathogenic fungi, includingdifferent Fusarium spp. With the aim to identify the genes responsible for this potent antifungal activity, aninsertional mini-Tn5 library was constructed in B. ambifaria T16 by triparental mating. A total of 8,950mutants were screened for the ability to inhibit growth of F. oxysporum by using an overlay assay withhomogenized mycelium. The insertion sites of the mini-Tn5 were mapped in 20 mutants, which showedreduced or null antifungal activity, by an arbitrary primed polymerase chain reaction (AP-PCR) method. In halfof these mutants, the mini-Tn5 was inserted in a modular non ribosomal peptide synthetase (NRPS) genecluster, encoding proteins involved in the biosynthesis of cyclic antifungal lipopeptides (CLPs) known asoccidiofungins/burkholdines (bks). In these mutants, the mini-Tn5 insertion abolished completely the abilityof Burkholderia ambifaria T16 to inhibit mycelial growth of F. oxysporum. When these mutants were testedagainst other important pathogenic fungi, such as F. graminearum, Macrophomina phaseolina and Candidaalbicans, the antifungal activity was significantly reduced compared to the wild type, indicating thatburkholdines were responsible for most of the antifungal activity. The identification and characterization ofNRPS gene clusters responsible of strong antifungal activity against phytopathogenic fungi would provide keyinformation for engineered biosynthesis of innovative antifungal compounds.