The effect of genetic mutations on biofilm production in Listeria monocytogenes

Abstract

The foodborne pathogen Listeria monocytogenes is responsible for listeriosis, a disease that endangers immuno-compromised individuals, the elderly, and pregnant women. The pathogen is capable of surviving harsh conditions, allowing it to inhabit food-processing facilities for long amounts of time. This persistence is due to the ability of Listeria spp. to form biofilms that protect against disinfectants. Understanding of the influence genes have in biofilm formation of L. monocytogenes is limited. To examine the genetic role in this process, a transposon library of 4300 Tn917 mutants of L. monocytogenes 568 serotype 1/2a (Lm568) was screened for biofilm variation. The screening process consisted of a crystal violet staining assay, in which the mutants were cultured in 96 well micotitre plates in modified Welshimer's broth. After 72 h, 0.3% crystal violet was added, and then ethanol was used to extract the dye from the stained cells of the biofilm. The stained ethanol was then measured spectrophotometrically for quantitative analysis. Mutants exhibiting variability in biofilm formation were isolated but were unable to be sequenced. Assays were also conducted on previously sequenced mutants with known transposon insertions to see how specific genes alter biofilm production when treated with a preservative or cleaning solution ingredients (NaCl, H2O2 and ethanol). Some tests showed significant variability in biofilm production. Future studies into specific genes that affect biofilm production may aid in preventing the pathogen from inhabiting food-processing facilities.