Physiological and Transcriptional Characterization of Persistent and Nonpersistent Listeria monocytogenes Isolates

Abstract

This study aimed to characterize physiological differences between persistent and presumed nonpersistent Listeria monocytogenes strains isolated at processing facilities and to investigate the molecular basis for this by transcriptomic sequencing. Full metabolic profiles of two strains, one persistent and one nonpersistent, were initially screened using Biolog's Phenotype MicroArray (PM) technology. Based on these results, in which major differences from selected antimicrobial agents were detected, another persistent strain and two nonpersistent strains were characterized using two antimicrobial PMs. Resistance to quaternary ammonium compounds (QACs) was shown to be higher among persistent strains. Growth of persistent and nonpersistent strains in various concentrations of the QACs benzethonium chloride (BZT) and cetylpyridinium chloride (CPC) was determined. Transcriptomic sequencing of a persistent and a presumed nonpersistent strain was performed to compare gene expression among these strains in the presence and absence of BZT. Two strains, designated "frequent persisters" because they were the most frequently isolated at the processing facility, showed overall higher resistance to QACs. Transcriptome analysis showed that BZT induced a complex peptidoglycan (PG) biosynthesis response, which may play a key role in BZT resistance. Comparison of persistent and nonpersistent strains indicated that transcription of many genes was upregulated among persistent strains. This included three gene operons: pdu, cob-cbi, and eut. These genes may play a role in the persistence of L. monocytogenes outside the human host.Listeria monocytogenes is a human food-borne pathogen that causes listeriosis in humans. Because infection is primarily food borne, control of the organism in food-processing environments, including dairy food production facilities, is important. There are many studies detailing the occurrence of L. monocytogenes in food-processing facilities (18,22,25,28). In order to better understand the ecology and transmission of L. monocytogenes strains through the processing facility, however, techniques such as molecular strain typing are necessary. Such studies not only play a crucial role in determining possible transmission vectors, they provide insights into the persistence of certain strains at the facility itself. This is of critical importance to the manufacturer, as strains that can successfully persist at the production facility have an increased chance of cross-contamination of the product and thus lead to an increased likelihood of product recall or human infection. A study published in 1995 by Lawrence and Gilmour (20) characterized strains isolated over a 12-month period at a poultryprocessing plant located in Ireland. This study identified 18 different randomly amplified polymorphic DNA (RAPD) profiles among the L. monocytogenes strains, and interestingly, one of these RAPD types was seen throughout the 12 months of the study. Similar evidence of persistent strains has subsequently been documented in o...

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

“…The tendency of L. monocytogenes to persist in food processing plants after disinfection can lead to the selection and dissemination of clones with decreased susceptibility to disinfectants. However, persistent strains of L. monocytogenes showing reduced susceptibility to disinfectants have been characterized only in a few cases (6)(7)(8)(9)(10). Therefore, it is necessary to elucidate further the ecological and genetic characteristics of persistent strains of L. monocytogenes with reduced susceptibility to disinfectants.…”

“…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)(7)(8)(9)(10), and stress resistance mechanisms (11,12).…”

Emerging RNA-Seq Applications in Food Science

“…With regard to the investigation on foodborne pathogens, RNA-Seq has been highly valuable in providing global transcriptomic profiles of persistent and nonpersistent Listeria monocytogenes isolates [55]. This foodborne pathogen is the causative bacterium of serious invasive disease in animals and in humans.…”

Listeria monocytogenes in Milk Products

“…Various factors have been studied for their role in the persistence of L. monocytogenes strains, including disinfectant and desiccation resistance (Aase et al 2000;Holah et al 2002;Kastbjerg and Gram 2009;Vogel et al 2010), differences in gene expression (Fox et al 2011b) and biofilm formation (Norwood and Gilmour 1999;Lunden et al 2000;Djordjevic et al 2002).…”