The Connection between Persistent, Disinfectant-Resistant Listeria monocytogenes Strains from Two Geographically Separate Iberian Pork Processing Plants: Evidence from Comparative Genome Analysis

Abstract

The aim of this study was to investigate the basis of the putative persistence of Listeria monocytogenes in a new industrial facility dedicated to the processing of ready-to-eat (RTE) Iberian pork products. Quaternary ammonium compounds, which included benzalkonium chloride (BAC), were repeatedly used as surface disinfectants in the processing plant. Clean and disinfected surfaces were sampled to evaluate if resistance to disinfectants was associated with persistence. Of the 14 isolates obtained from product contact and non-product contact surfaces, only five different pulsed-field gel electrophoresis (PFGE) types were identified during the 27-month study period. Two of these PFGE types (S1 and S10-1) were previously identified to be persistent and BAC-resistant (BAC r ) strains in a geographically separate slaughterhouse belonging to the same company. The remaining three PFGE types, which were first identified in this study, were also BAC r . Whole-genome sequencing and in silico multilocus sequence typing (MLST) analysis of five BAC r isolates of the different PFGE types identified in this study showed that the isolate of the S1 PFGE type belonged to MLST sequence type 31 (ST31), a low-virulence type characterized by mutations in the inlA and prfA genes. The isolates of the remaining four PFGE types were found to belong to MLST ST121, a persistent type that has been isolated in several countries. The ST121 strains contained the BAC resistance transposon Tn6188. The disinfection-resistant L. monocytogenes population in this RTE pork product plant comprised two distinct genotypes with different multidrug resistance phenotypes. This work offers insight into the L. monocytogenes subtypes associated with persistence in food processing environments. Listeria monocytogenes is a Gram-positive bacterium of the phylum Firmicutes. Often found in raw foods, L. monocytogenes bacteria can cause the life-threatening disease listeriosis. Most cases of human listeriosis appear to be caused by ready-to-eat (RTE) foods, and the risk of illness increases with the number of cells ingested and with RTE foods that support the growth of L. monocytogenes (1). Processed foods can be contaminated by contact with equipment, by the handling of raw products by staff, or from postprocessing environmental niches in which L. monocytogenes can survive despite the routine use of thorough disinfection procedures (2, 3). Mechanisms that facilitate the survival of L. monocytogenes in food processing environments include biofilm formation (4, 5), acquisition of antimicrobial resistance (6-10), and stress resistance mechanisms (11,12).L. monocytogenes is capable of colonizing food production plants with certain subtypes that are found only in specific sections of the plants (13, 14). Furthermore, some of these subtypes may persist in food processing environments for years (15-17). Persistent strains have been identified to be major postprocessing contaminants of RTE foods, and in many cases, listeriosis outbreaks have been associated with cases of p...

Human pathogens in plant biofilms: Formation, physiology, and detection

“…Mechanisms that facilitate the survival of L. monocytogenes in food processing environments include biofilm formation (Nakamura et al, ; Wang et al, ), acquisition of antimicrobial resistance (Fox et al, ; Ortiz et al, , ), and stress resistance mechanisms (Cabrita et al, ; Lundén et al, ; Ortiz et al, ). L. monocytogenes is a saprophytic soil organism and intracellular pathogen that causes infections via the gastrointestinal route, and is able to survive and grow under a wide range of temperatures and stressful environmental conditions including acid and osmotic stress (Gahan and Hill, ; Milillo et al, ).…”

Design Elements of Listeria Environmental Monitoring Programs in Food Processing Facilities: A Scoping Review of Research and Guidance Materials

“…For example, nine outbreak investigations from years 1989 to 2016 reported focused and detailed sampling investigations of food processing facilities, taking between 19 and 282 samples when production was stopped in order to find the source of Lm associated with cases of human listeriosis (Acciari et al., ; Awofisayo‐Okuyelu et al., ; Dalton et al., ; Gaul et al., ; Jacks et al., ; McLauchlin, Greenwood, & Pini, ; Salvat, Toquin, Michel, & Colin, ; Wenger et al., ; Winter et al., ). On the other hand, studies evaluating the efficacy of interventions conducted sampling immediately before and after the intervention, but may also have followed up after months or years of production to determine the long‐term effect of plant‐specific interventions, such as employee training, equipment disassembly during sanitation, or a new disinfectant compound or procedure (Eglezos, Thygesen, Huang, & Dykes, ; Lappi et al., ; Lappi et al., ; Ortiz, Lopez‐Alonso, Rodriguez, & Martinez‐Suarez, ).…”

Comparison of selected disinfectants efficiency against Listeria monocytogenes biofilm formed on various surfaces

“…Poimenidou et al (2016) found that quaternary ammonium compounds were more effective against the biofilm formed on polystyrene as compared to the one formed on stainless steel. In contrast, Ortiz et al (2016) reported the resistance of bacilli to quaternary ammonium compounds caused by the long-term use of those particular disinfectants. In the present study, it was found that sodium hydroxide was the least efficient in eliminating L. monocytoges strains.…”