W.C. Hazeleger

Survival of foodborne pathogens on stainless steel surfaces and cross-contamination to foods.

The retention of bacteria on food contact surfaces increases the risk of cross-contamination of these microorganisms to food. The risk has been considered to be lowered when the surfaces are dry, partly because bacterial growth and survival would be reduced. However, some non-spore-forming bacteria might be able to withstand dry conditions on surfaces for an extensive period of time. In this study the survival of Salmonella enteritidis, Staphylococcus aureus and Campylobacter jejuni on stainless steel surfaces at different initial levels was determined at room temperature. The transfer rates of these pathogens from kitchen sponges to stainless steel surfaces and from these surfaces to foods were also investigated. Staph. aureus was recovered from the surfaces for at least 4 days when the contamination level was high (10(5) CFU/cm2) or moderate (10(3) CFU/cm2). At low levels (10 CFU/cm2), the surviving numbers decreased below the detection limit (4 CFU/100 cm2) within 2 days. S. enteritidis was recovered from surfaces for at least 4 days at high contamination levels, but at moderate level, the numbers decreased to the detection limit within 24 h and at low level within 1 h. C. jejuni was the most susceptible to slow-air-drying on surfaces; at high contamination levels, the numbers decreased below the detection limit within 4 h. The test microorganisms were readily transmitted from the wet sponges to the stainless steel surfaces and from these surfaces to the cucumber and chicken fillet slices, with the transfer rates varied from 20% to 100%. This study has highlighted the fact that pathogens remain viable on dry stainless steel surfaces and present a contamination hazard for considerable periods of time, dependent on the contamination levels and type of pathogen. Systematic studies on the risks of pathogen transfer associated with surface cleaning using contaminated sponges provide quantitative data from which a model of risks assessment in domestic setting could lead.

Darkling Beetles (Alphitobius diaperinus) and Their Larvae as Potential Vectors for the Transfer of Campylobacter jejuni and Salmonella enterica Serovar Paratyphi B Variant Java between Successive Broiler Flocks

ABSTRACT Broiler flocks often become infected with Campylobacter and Salmonella, and the exact contamination routes are still not fully understood. Insects like darkling beetles and their larvae may play a role in transfer of the pathogens between consecutive cycles. In this study, several groups of beetles and their larvae were artificially contaminated with a mixture of Salmonella enterica serovar Paratyphi B Variant Java and three C. jejuni strains and kept for different time intervals before they were fed to individually housed chicks. Most inoculated insects were positive for Salmonella and Campylobacter just before they were fed to the chicks. However, Campylobacter could not be isolated from insects that were kept for 1 week before they were used to mimic an empty week between rearing cycles. All broilers fed insects that were inoculated with pathogens on the day of feeding showed colonization with Campylobacter and Salmonella at levels of 50 to 100%. Transfer of both pathogens by groups of insects that were kept for 1 week before feeding to the chicks was also observed, but at lower levels. Naturally contaminated insects that were collected at a commercial broiler farm colonized broilers at low levels as well. In conclusion, the fact that Salmonella and Campylobacter can be transmitted via beetles and their larvae to flocks in successive rearing cycles indicates that there should be intensive control programs for exclusion of these insects from broiler houses.

Residual Viral and Bacterial Contamination of Surfaces after Cleaning and Disinfection

ABSTRACT Environmental surfaces contaminated with pathogens can be sources of indirect transmission, and cleaning and disinfection are common interventions focused on reducing contamination levels. We determined the efficacy of cleaning and disinfection procedures for reducing contamination by noroviruses, rotavirus, poliovirus, parechovirus, adenovirus, influenza virus, Staphylococcus aureus, and Salmonella enterica from artificially contaminated stainless steel surfaces. After a single wipe with water, liquid soap, or 250-ppm free chlorine solution, the numbers of infective viruses and bacteria were reduced by 1 log10 for poliovirus and close to 4 log10 for influenza virus. There was no significant difference in residual contamination levels after wiping with water, liquid soap, or 250-ppm chlorine solution. When a single wipe with liquid soap was followed by a second wipe using 250- or 1,000-ppm chlorine, an extra 1- to 3-log10 reduction was achieved, and except for rotavirus and norovirus genogroup I, no significant additional effect of 1,000 ppm compared to 250 ppm was found. A reduced correlation between reduction in PCR units (PCRU) and reduction in infectious particles suggests that at least part of the reduction achieved in the second step is due to inactivation instead of removal alone. We used data on infectious doses and transfer efficiencies to estimate a target level to which the residual contamination should be reduced and found that a single wipe with liquid soap followed by a wipe with 250-ppm free chlorine solution was sufficient to reduce the residual contamination to below the target level for most of the pathogens tested.

Survival, Elongation, and Elevated Tolerance of Salmonella enterica Serovar Enteritidis at Reduced Water Activity

Growing microorganisms on dry surfaces, which results in exposure to low water activity (a(w)), may change their normal morphology and physiological activity. In this study, the morphological changes and cell viability of Salmonella enterica serovar Enteritidis challenged to low a(w) were analyzed. The results indicated that exposure to reduced a(w) induced filamentation of the cells. The amount of filamentous cells at a(w) 0.94 was up to 90% of the total number of cells. Surviving filamentous cells maintained their membrane integrity after exposure to low a(w) for 21 days. Furthermore, cells prechallenged to low a(w), obtained with an ionic humectant, demonstrated higher resistance to sodium hypochlorite than control cells. These resistant cells are able to survive disinfection more efficiently and can therefore cause contamination of foods coming in contact with surfaces. This points to the need for increased attention to cleaning of surfaces in household environments and disinfection procedures in processing plants.

Listeria monocytogenes: diagnostic problems

The first isolation methods for the detection of Listeria spp. were generally based on the direct culture of samples on simple agar media, but isolation of the pathogenic Listeria monocytogenes was difficult. In time, new techniques were developed, based on a variety of selective and elective agents in isolation and enrichment media, which gained better and quicker results. Current reference methods allow the recovery of L. monocytogenes from a variety of foods with relative ease. However, more comparative studies are needed to select one horizontal method. It is suggested that the procedure of the International Organization for Standardization is a good base for such comparisons.

Tolerance of Salmonella Enteritidis and Staphylococcus aureus to Surface Cleaning and Household Bleach

Effective cleaning and sanitizing of food preparation sites is important because pathogens are readily spread to food contact surfaces after preparation of contaminated raw products. Tolerance of Salmonella Enteritidis and Staphylococcus aureus to surface cleaning by wiping with regular, microfiber, and antibacterial-treated cloths was investigated. Wiping with cleaning cloths resulted in a considerable reduction of microorganisms from surfaces, despite the greater difficulty in removing S. aureus than Salmonella Enteritidis. Depending on the cloth type, S. aureus were reduced on surfaces from initial numbers of approximately 10(5) CFU/100 cm2 to numbers from less than 4 CFU/100 cm2 (below the detection limit) to 100 CFU/100 cm2. Directly after the cloths were used to clean the contaminated surfaces, they contained high numbers of bacteria (10(4) to 10(5) CFU/100 cm2), except for the disposable antibacterial-treated cloths, in which no bacteria could be detected. The tolerance of these pathogens to sodium hypochlorite was studied in the suspension test and in cloths. S. aureus showed a better tolerance for sodium hypochlorite than Salmonella Enteritidis. Inactivation of microorganisms in cloths required a higher concentration of sodium hypochlorite than was needed in the suspension test. Repeated exposure to sodium hypochlorite, however, resulted in an increase in susceptibility to this compound. This study provides essential information about the transfer of bacteria when wiping surfaces and highlights the need for a hygiene procedure with cleaning cloths that sufficiently avoids cross-contamination in the household environment.

Quantifying strain variability in modeling growth of Listeria monocytogenes.

Prediction of microbial growth kinetics can differ from the actual behavior of the target microorganisms. In the present study, the impact of strain variability on maximum specific growth rate (μmax) (h(-1)) was quantified using twenty Listeria monocytogenes strains. The μmax was determined as function of four different variables, namely pH, water activity (aw)/NaCl concentration [NaCl], undissociated lactic acid concentration ([HA]), and temperature (T). The strain variability was compared to biological and experimental variabilities to determine their importance. The experiment was done in duplicate at the same time to quantify experimental variability and reproduced at least twice on different experimental days to quantify biological (reproduction) variability. For all variables, experimental variability was clearly lower than biological variability and strain variability; and remarkably, biological variability was similar to strain variability. Strain variability in cardinal growth parameters, namely pHmin, [NaCl]max, [HA]max, and Tmin was further investigated by fitting secondary growth models to the μmax data, including a modified secondary pH model. The fitting results showed that L. monocytogenes had an average pHmin of 4.5 (5-95% prediction interval (PI) 4.4-4.7), [NaCl]max of 2.0mM (PI 1.8-2.1), [HA]max of 5.1mM (PI 4.2-5.9), and Tmin of -2.2°C (PI (-3.3)-(-1.1)). The strain variability in cardinal growth parameters was benchmarked to available literature data, showing that the effect of strain variability explained around 1/3 or less of the variability found in literature. The cardinal growth parameters and their prediction intervals were used as input to illustrate the effect of strain variability on the growth of L. monocytogenes in food products with various characteristics, resulting in 2-4 logCFU/ml(g) difference in growth prediction between the most and least robust strains, depending on the type of food product. This underlined the importance to obtain quantitative knowledge on variability factors to realistically predict the microbial growth kinetics.

Survival of Listeria monocytogenes on a conveyor belt material with or without antimicrobial additives

Survival of Listeria monocytogenes on a conveyor belt material with or without antimicrobial additives, in the absence or presence of food debris from meat, fish and vegetables and at temperatures of 10, 25 and 37 degrees C was investigated. The pathogen survived best at 10 degrees C, and better at 25 degrees C than at 37 degrees C on both conveyor belt materials. The reduction in the numbers of the pathogen on belt material with antimicrobial additives in the first 6h at 10 degrees C was 0.6 log unit, which was significantly higher (P<0.05) than the reduction of 0.2 log unit on belt material without additives. Reductions were significantly less (P<0.05) in the presence of food residue. At 37 degrees C and 20% relative humidity, large decreases in the numbers of the pathogen on both conveyor belt materials during the first 6h were observed. Under these conditions, there was no obvious effect of the antimicrobial substances. However, at 25 degrees C and 10 degrees C and high humidity (60-75% rh), a rapid decrease in bacterial numbers on the belt material with antimicrobial substances was observed. Apparently the reduction in numbers of L. monocytogenes on belt material with antimicrobial additives was greater than on belt material without additives only when the surfaces were wet. Moreover, the presence of food debris neutralized the effect of the antimicrobials. The results suggest that the antimicrobial additives in conveyor belt material could help to reduce numbers of microorganisms on belts at low temperatures when food residues are absent and belts are not rapidly dried.

Quantifying variability on thermal resistance of Listeria monocytogenes

Knowledge of the impact of strain variability and growth history on thermal resistance is needed to provide a realistic prediction and an adequate design of thermal treatments. In the present study, apart from quantifying strain variability on thermal resistance of Listeria monocytogenes, also biological variability and experimental variability were determined to prioritize their importance. Experimental variability was defined as the repeatability of parallel experimental replicates and biological variability was defined as the reproducibility of biologically independent reproductions. Furthermore, the effect of growth history was quantified. The thermal inactivation curves of 20 L. monocytogenes strains were fitted using the modified Weibull model, resulting in total 360 D-value estimates. The D-value ranged from 9 to 30 min at 55 °C; from 0.6 to 4 min at 60 °C; and from 0.08 to 0.6 min at 65 °C. The estimated z-values of all strains ranged from 4.4 to 5.7 °C. The strain variability was ten times higher than the experimental variability and four times higher than the biological variability. Furthermore, the effect of growth history on thermal resistance variability was not significantly different from that of strain variability and was mainly determined by the growth phase.

Transfer of noroviruses between fingers and fomites and food products

Human norovirus (NoV) contaminated hands are important routes for transmission. Quantitative data on transfer during contact with surfaces and food are scarce but necessary for a quantitative risk assessment. Therefore, transfer of MNV1 and human NoVs GI.4 and GII.4 was studied by artificially contaminating human finger pads, followed by pressing on stainless steel and Trespa® surfaces and also on whole tomatoes and cucumber slices. In addition, clean finger pads were pressed on artificially contaminated stainless steel and Trespa® surfaces. The transfers were performed at a pressure of 0.8-1.9 kg/cm(2) for approximately 2s up to 7 sequential transfers either to carriers or to food products. MNV1 infectivity transfer from finger pads to stainless steel ranged from 13 ± 16% on the first to 0.003 ± 0.009% on the sixth transfer on immediate transfer. After 10 min of drying, transfer was reduced to 0.1 ± 0.2% on the first transfer to 0.013 ± 0.023% on the fifth transfer. MNV1 infectivity transfer from stainless steel and Trespa® to finger pads after 40 min of drying was 2.0 ± 2.0% and 4.0 ± 5.0% respectively. MNV1 infectivity was transferred 7 ± 8% to cucumber slices and 0.3 ± 0.5% to tomatoes after 10 min of drying, where the higher transfer to cucumber was probably due to the higher moisture content of the cucumber slices. Similar results were found for NoVs GI.4 and GII.4 transfers measured in PCR units. The results indicate that transfer of the virus is possible even after the virus is dried on the surface of hands or carriers. Furthermore, the role of fingers in transmission of NoVs was quantified and these data can be useful in risk assessment models and to establish target levels for efficacy of transmission intervention methods.