Sanna Hellström

Adaptive and cross-adaptive responses of persistent and non-persistent Listeria monocytogenes strains to disinfectants

Persistent and non-persistent Listeria monocytogenes strains were tested for initial resistance and adaptive and cross-adaptive responses towards two quaternary ammonium compounds, alkyl-benzyl-dimethyl ammonium chloride and n-alkyldimethyl ethylbenzyl ammonium chloride, one tertiary alkylamine, 1,3-propanediamine-N-(3-aminopropyl)N-dodecyl, sodium hypochlorite and potassium persulphate. The initial resistance of two persistent and two non-persistent L. monocytogenes strains was observed to differ. Both types of strains adapted after a 2-h sublethal exposure to the quaternary ammonium compounds and the tertiary alkylamine, the highest increase in the minimum inhibitory concentration (MIC) being 3-fold. Progressively increasing disinfecting concentrations at 10 and 37 degrees C resulted in adaptation of L. monocytogenes to all disinfectants except potassium sulphate. The highest observed increase in MIC was over 15-fold, from 0.63 to 10 microg/ml of n-alkyldimethyl ethylbenzyl ammonium chloride. All strains reached approximately similar MICs. Stability of the increased resistance was tested by measuring MICs every seventh day for 28 days. The increased resistance to sodium hypochlorite disappeared in 1 week, but the quaternary ammonium compounds and the tertiary alkylamine showed increased resistance for 28 days. These results suggest that cellular changes due to adaptive responses continue to have an effect on the resistance some time after the exposure. All disinfectants were shown to cause cross-adaptation of L. monocytogenes, the highest increase in MIC being almost 8-fold. The only agent that L. monocytogenes could not be shown to cross-adapt to was potassium persulphate which did, however, cause cross-adaptation to the other disinfectants. The mechanism behind these adaptive responses seemed to be non-specific as cross-adaptation was observed not only between related but also unrelated disinfectants. These findings suggest that sustaining high disinfectant effectiveness may be unsuccessful by rotation, even when using agents with different mechanisms of action.

Efficacy of disinfectants to reduce Listeria monocytogenes on precut iceberg lettuce.

The efficacy of water, chlorinated water (100 ppm), peracetic acid solution (0.05%), and commercial citric acid-based produce wash (0.25%) to reduce the population of Listeria monocytogenes on precut lettuce was tested. Samples were inoculated with a mixture of equal amounts of five L. monocytogenes strains at a level of 4.7 log CFU/g, and analyzed on the day of washing and after 3 and 6 days of storage at 6 degrees C. Sanitizer reduced the number of L. monocytogenes at maximum 1.7 log CFU/g and number of L. monocytogenes reached the inoculation level during 6 days of storage. Thus, disinfectants do not eliminate L. monocytogenes on precut lettuce and cannot be solely relied on in producing precut lettuce safely. The inoculated L. monocytogenes strains were recovered at different rates after 6 days of storage; one of these strains was not recovered at all. Thus, strain-specific differences exist in the ability of L. monocytogenes to survive the washing treatments of the lettuce.

Listeria monocytogenes contamination in pork can originate from farms.

The presence of Listeria monocytogenes in the pork production chain was followed from farm to slaughterhouse by examining the farm and slaughterhouse levels in the same 364 pigs, and finally by analyzing the cut meats from the same pig lots. Both organic and conventional farms were included in the study. Altogether, 1,962 samples were collected, and the 424 L. monocytogenes isolates were analyzed by pulsed-field gel electrophoresis. The results from microbial analyses were combined with data from an on-farm observation and a questionnaire to clarify the associations between farm factors and prevalence of L. monocytogenes. The prevalence of L. monocytogenes was 11, 1, 1, 24, 5, 1, and 4% in feed and litter, rectal swabs, intestinal contents, tonsils, pluck sets (including lungs, heart, liver, and kidney), carcasses, and meat cuts, respectively. The prevalence was significantly higher in organic than in conventional pig production at the farm and slaughterhouse level, but not in meat cuts. Similar L. monocytogenes genotypes were recovered in different steps of the production chain in pigs originating from the same farm. Specific farm management factors, i.e., large group size, contact with pet and pest animals, manure treatment, use of coarse feed, access to outdoor area, hygiene practices, and drinking from the trough, influenced the presence of L. monocytogenes in pigs. L. monocytogenes was present in the production chain, and transmission of the pathogen was possible throughout the chain, from the farm to pork. Good farm-level practices can therefore be utilized to reduce the prevalence of this pathogen.

Listeria monocytogenes is common in wild birds in Helsinki region and genotypes are frequently similar with those found along the food chain

Aims:  To evaluate the prevalence and genetic diversity of Listeria monocytogenes in wild birds and to compare the genotypes with isolates previously collected from foods and food processing environments.

Prevalence and Genetic Diversity of Listeria monocytogenes in the Tonsils of Pigs

This study was set up to establish the prevalence of Listeria monocytogenes in the tonsils of sows and fattening pigs from five Finnish slaughterhouses and to evaluate the genetic similarity of L. monocytogenes strains isolated from the tonsils. A total of 271 pig tonsils (132 tonsils from fattening pigs and 139 from sows) from five different slaughterhouses in various parts of Finland were studied from June 1999 to March 2000. Overall, 14 and 4% of pig tonsils harbored L. monocytogenes and Listeria innocua, respectively. The prevalence of L. monocytogenes in tonsils of fattening pigs (22%) was significantly higher than in sows (6%). The isolates (n = 38) recovered from tonsils showed a wide genetic diversity by means of 24 different pulsed-field gel electrophoresis (PFGE) types presented by the strains. Moreover, in numerical analyses of restriction patterns, no association was found between the clustering of strains and the slaughterhouses, and strains showing a similar PFGE type were recovered from pigs of different slaughterhouses. The high prevalence of L. monocytogenes showing various PFGE types in the tonsils of pigs could indicate a potential source of contamination of pluck sets, carcasses, and the slaughterhouse environment and of subsequent processing steps.

Contamination patterns of Listeria monocytogenes in cold-smoked pork processing.

Contamination patterns of Listeria monocytogenes were studied in a cold-smoked pork processing plant to identify the sources and possible reasons for the contamination. Environmental sampling combined with pulsed-field gel electrophoresis (PFGE) subtyping and serotyping were applied to investigate the genetic diversity of L. monocytogenes in the plant environment and ready-to-eat (RTE) cold-smoked pork products. A total of 183 samples were collected for contamination analyses, including samples of the product at different stages during manufacture (n = 136) and environmental samples (n = 47) in 2009. L. monocytogenes isolates, previously recovered from 73 RTE cold-smoked pork samples and collected from the same meat processing plant in 2004, were included in this study. The brining machine and personnel working with brining procedures were the most contaminated places with L. monocytogenes. The overall prevalence of L. monocytogenes in raw pork (18%) increased to 60% after the brining injections. The brining machine harbored six different PFGE types belonging to serotypes 1/2a, 1/2c, 4b, and 4d, which were found on the feeding teeth, smooth surfaces, and spaces of the machine, thus potentially facilitating dissemination of L. monocytogenes contamination. Two PFGE types (2 and 8) belonging to serotypes 1/2a and 1/2c were recovered from RTE cold-smoked pork collected in 2004, and from surfaces of the brining machine sampled in 2009, and may indicate the presence of persistent L. monocytogenes strains in the plant. Due to poor hygiene design, removal of the brining machine from the production of cold-smoked meat products should be considered to reduce L. monocytogenes contamination in the finished products.

Causal hidden variable model of pathogenic contamination from pig to pork

Risk assessments relating to food safety over more than one step along a production chain are frequently hampered by lack of detailed quantitative data. This study set out to develop a Bayesian hidden variable model to integrate available limited data of the combined occurrence of three bacterial pathogens, Listeria monocytogenes, Yersinia enterocolitica and Yersinia pseudotuberculosis, with causal assumptions along three steps of pork production chain. The pathogen occurrence data were animal specific both on conventional and organic pig farms and at the abattoir, but merely farm specific at meat cutting plants. The model was able to incorporate all data concerning different types of testing at different steps of the chain, and missing data values were dealt with in a straightforward manner. It provides a tool for quantitative risk assessments and for estimating the causal risk mitigation effects by combining external data with the specific follow-up data. Intervention effects are provided with Bayesian credible intervals indicating the uncertainty due to all information sources included in the model. Combined prevalence in Finnish pork was estimated to be 1–11% and it could be reduced to 0–2% if head was removed intact and rectum sealed off.

Survival of Listeria monocytogenes Strains in a Dry Sausage Model

The survival of five inoculated Listeria monocytogenes strains (DCS 31, DCS 184, AT3E, HT4E, and HR5E) was studied in dry fermented sausages prepared using two different starter cultures (starter A and B) with or without a protective Lactobacillus plantarum DDEN 2205 strain. L. monocytogenes was detected throughout ripening in every sausage sample in which the L. plantarum DDEN 2205 strain had not been used. The use of either starter A, with a high concentration of protective culture, or starter B, with a low concentration of protective culture, resulted in L. monocytogenes-negative sausages after 17 days of ripening. Differential survival was noted among the L. monocytogenes strains during fermentation. Strains AT3E and DCS 31 survived in sausages with protective cultures more often than did the other strains, whereas HT4E and HR5E were inhibited during ripening by all starter and protective cultures used. Protective cultures such as L. plantarum may be used as part of a hurdle strategy in dry sausage processing, but variations in susceptibility of different L. monocytogenes strains can create problems if other hurdles are not included.