ABSTRACTBurkholderia gladiolistrain NGJ1 exhibits mycophagous activity on a broad range of fungi, includingRhizoctonia solani, a devastating plant pathogen. Here, we demonstrate that the nicotinic acid (NA) catabolic pathway in NGJ1 is required for mycophagy. NGJ1 is auxotrophic to NA and it potentially sensesR. solanias a NA source. Mutation in thenicCandnicXgenes involved in NA catabolism renders defects in mycophagy and the mutant bacteria are unable to utilizeR. solaniextract as the sole nutrient source. As supplementation of NA, but not FA (fumaric acid, the end product of NA catabolism) restores the mycophagous ability of ΔnicC/ΔnicXmutants, we anticipate that NA is not required as a carbon source for the bacterium during mycophagy. Notably,nicR, a MarR-type of transcriptional regulator that functions as a negative regulator of the NA catabolic pathway is upregulated in ΔnicC/ΔnicXmutant and upon NA supplementation thenicRexpression is reduced to the basal level in both the mutants. The ΔnicRmutant produces excessive biofilm and is completely defective in swimming motility. On the other hand, ΔnicC/ΔnicXmutants are compromised in swimming motility as well as biofilm formation, potentially due to the upregulation ofnicR. Our data suggest that a defect in NA catabolism alters the NA pool in the bacterium and upregulatesnicRwhich in turn suppresses bacterial motility as well as biofilm formation, leading to mycophagy defects.IMPORTANCEMycophagy is an important trait through which certain bacteria forage over fungal mycelia and utilize fungal biomass as a nutrient source to thrive in hostile environments. The present study emphasizes that nicotinic acid (NA) is important for bacterial motility and biofilm formation during mycophagy byBurkholderia gladiolistrain NGJ1. Defects in NA catabolism potentially alter the cellular NA pool, upregulate the expression ofnicR, a negative regulator of biofilm, and therefore suppress bacterial motility as well as biofilm formation, leading to mycophagy defects.