Berberine at non-lethal concentrations attenuates virulence of Chromobacterium violaceum

Chromobacterium violaceum is a gram-negative bacteria, intrinsically resistant to multiple antibiotics, considered as a pathogen of emerging importance. C. violaceum pre-exposed to a phytochemical, berberine (0.5-20 ppm), when allowed to attack the model host Caenorhabditis elegans, could kill lesser worms than its berberine-non-exposed control counterpart. While all the test concentrations between 5-20 ppm were able to attenuate this pathogen’s virulence, 10 ppm had the maximum (60±10%; p<0.001) virulence-attenuating effect. Interestingly, concentrations higher than 10 ppm had lesser anti-virulence effect. Berberine’s anti-virulence effect between 5-20 ppm can be said to follow an inverted-U shaped pattern of dose-response relation. Berberine at none of these concentrations had any effect on bacterial growth or quorum sensing-regulated violacein production. Whole transcriptome of berberine-treated C. violaceum revealed 9 genes to be differently expressed (FDR<0.01; log FC>2). Among the down-regulated genes were those (nirK: copper containing nitrite reductase; and norB: nitric oxide reductase large subunit) pertaining to nitrogen metabolism. Co-occurrence analysis shows both the down-regulated target genes to be absent from humans. Further investigation on targetability of these genes with respect to development of novel anti-virulence agents, and correlation of nitrogen metabolism with virulence of C. violaceum is warranted. Repeated exposure of C. violaceum to berberine (ten subculturings in 10-20 ppm berberine containing media) did not induce resistance against this phytochemical in the bacteria, as this berberine-habituated C. violaceum still could kill lesser worms than the berberine non-exposed control.