Novel Cadmium Resistance Determinant in Listeria monocytogenes

Listeria monocytogenes is a foodborne pathogen that can cause severe disease (listeriosis) in susceptible individuals. It is ubiquitous in the environment and often exhibits resistance to heavy metals. One of the determinants that enablesListeria to tolerate exposure to cadmium is thecadAC efflux system, with CadA being a P-type ATPase. Three differentcadA genes (designatedcadA1 tocadA3 ) were previously characterized inL. monocytogenes . A novel putative cadmium resistance gene (cadA4 ) was recently identified through whole-genome sequencing, but experimental confirmation for its involvement in cadmium resistance is lacking. In this study, we characterizedcadA4 inL. monocytogenes strain F8027, a cadmium-resistant strain of serotype 4b. By screening a mariner-based transposon library of this strain, we identified a mutant with reduced tolerance to cadmium and that harbored a single transposon insertion incadA4 . The tolerance to cadmium was restored by genetic complementation with the cadmium resistance cassette (cadA4C ), and enhanced cadmium tolerance was conferred to two unrelated cadmium-sensitive strains via heterologous complementation withcadA4C . Cadmium exposure inducedcadA4 expression, even at noninhibitory levels. Virulence assessments in theGalleria mellonella model suggested that a functionalcadA4 suppressed virulence, potentially promoting commensal colonization of the insect larvae. Biofilm assays suggested thatcadA4 inactivation reduced biofilm formation. These data not only confirmcadA4 as a novel cadmium resistance determinant inL. monocytogenes but also provide evidence for roles in virulence and biofilm formation.IMPORTANCE Listeria monocytogenes is an intracellular foodborne pathogen causing the disease listeriosis, which is responsible for numerous hospitalizations and deaths every year. Among the adaptations that enable the survival ofListeria in the environment are the abilities to persist in biofilms, grow in the cold, and tolerate toxic compounds, such as heavy metals. Here, we characterized a novel determinant that was recently identified on a larger mobile genetic island through whole-genome sequencing. This gene (cadA4 ) was found to be responsible for cadmium detoxification and to be a divergent member of the Cad family of cadmium efflux pumps. Virulence assessments in aGalleria mellonella model suggested thatcadA4 may suppress virulence. Additionally,cadA4 may be involved in the ability ofListeria to form biofilms. Beyond the role in cadmium detoxification, the involvement ofcadA4 in other cellular functions potentially explains its retention and wide distribution inL. monocytogenes .