Bacterial Quorum-Quenching Lactonase Hydrolyzes Fungal Mycotoxin and Reduces Pathogenicity of Penicillium expansum—Suggesting a Mechanism of Bacterial Antagonism

Abstract

<i>Penicillium expansum</i> is a necrotrophic wound fungal pathogen that secrets virulence factors to kill host cells including cell wall degrading enzymes (CWDEs), proteases, and mycotoxins such as patulin. During the interaction between <i>P. expansum</i> and its fruit host, these virulence factors are strictly modulated by intrinsic regulators and extrinsic environmental factors. In recent years, there has been a rapid increase in research on the molecular mechanisms of pathogenicity in <i>P. expansum</i>; however, less is known regarding the bacteria-fungal communication in the fruit environment that may affect pathogenicity. Many bacterial species use quorum-sensing (QS), a population density-dependent regulatory mechanism, to modulate the secretion of quorum-sensing signaling molecules (QSMs) as a method to control pathogenicity. N-acyl homoserine lactones (AHLs) are Gram-negative QSMs. Therefore, QS is considered an antivirulence target, and enzymes degrading these QSMs, named quorum-quenching enzymes, have potential antimicrobial properties. Here, we demonstrate that a bacterial AHL lactonase can also efficiently degrade a fungal mycotoxin. The mycotoxin is a lactone, patulin secreted by fungi such as <i>P. expansum</i>. The bacterial lactonase hydrolyzed patulin at high catalytic efficiency, with a <i>k_cat</i> value of 0.724 ± 0.077 s^-1 and K_M value of 116 ± 33.98 μM. The calculated specific activity (<i>k_cat</i>/K_M) showed a value of 6.21 × 10³ s^-1M^-1. While the incubation of <i>P. expansum</i> spores with the purified lactonase did not inhibit spore germination, it inhibited colonization by the pathogen in apples. Furthermore, adding the purified enzyme to <i>P. expansum</i> culture before infecting apples resulted in reduced expression of genes involved in patulin biosynthesis and fungal cell wall biosynthesis. Some AHL-secreting bacteria also express AHL lactonase. Here, phylogenetic and structural analysis was used to identify putative lactonase in <i>P. expansum</i>. Furthermore, following recombinant expression and purification of the newly identified fungal enzyme, its activity with patulin was verified. These results indicate a possible role for patulin and lactonases in inter-kingdom communication between fungi and bacteria involved in fungal colonization and antagonism and suggest that QQ lactonases can be used as potential antifungal post-harvest treatment.