Lesley L. Duffy

Growth of Listeria monocytogenes on vacuum-packed cooked meats : effects of pH, aw, nitrite and ascorbate

Slices of cooked meats were inoculated with Listeria monocytogenes strain Murray B, vacuum-packed and stored at either 0 or 5 degrees C. Decreases in pH (6.9-5.9) and aw (0.993-0.960; adjusted with sodium chloride) of the cooked meats increased the lag time and reduced the growth rate at 5 degrees C. The type of meat (beef, pork, chicken or turkey) had no effect on the growth of the organism after allowance was made for pH. Sodium tripolyphosphate (0.3%) increased growth by increasing the pH of the cooked meat. Sodium nitrite reduced the growth rate and increased the lag time. Three microM of residual undissociated nitrite doubled the time taken for a 3 log increase in numbers of L. monocytogenes. The effectiveness of nitrite was significantly increased by sodium ascorbate (0.042%). In the absence of nitrite, ascorbate had no detectable effect on growth. The extent of growth at 0 degree C was similarly influenced by the interaction of pH, aw, nitrite and ascorbate, and was considerably less than at 5 degrees C. Quadratic equations were developed to describe some of the combined effects of pH, aw and residual nitrite on lag, growth rate and time for a 3 log increase in numbers of L. monocytogenes.

Distribution, prevalence and persistence of Cronobacter (Enterobacter sakazakii) in the nonprocessing and processing environments of five milk powder factories

Aims:  To characterise the occurrence of Cronobacter in milk powder factories.

Acid resistance of enterohaemorrhagic and generic Escherichia coli associated with foodborne disease and meat

As part of the Australia New Zealand Food Authorities (ANZFA) food standards code, salami manufacturers are required to demonstrate that their process is capable of achieving a 3-log reduction in Escherichia coli. Non-pathogenic E. coli strains with similar or greater acid resistance to enterohaemorrhagic E. coli (EHEC) are needed if industry is to conduct challenge studies to demonstrate compliance with the standard. In the present study, E. coli isolates from sheep and beef carcasses and meat were shown to have wide-ranging acid resistance in broth when preadapted to growth in acidic conditions. Times required for a 3-log reduction in E. coli ranged from less than I day to more than 28 days. Variable acid resistance was observed in both EHEC strains associated with foodborne outbreaks and generic E. coli strains. Generic E. coli strains with the greatest acid resistance were assessed for pathogenicity markers and their survival in fermented meat compared with EHEC strains. It was demonstrated that generic E. coli strains could be used for challenge studies to determine compliance with or validate performance standards designed for the control of EHEC.

The Effects of Transport and Lairage on Counts of Escherichia coli O157 in the Feces and on the Hides of Individual Cattle

OBJECTIVES The main objective of this study was to determine the impact of transport and lairage on the isolation rate and the number of Escherichia coli O157 on cattle. MATERIALS Ninety animals, divided into three groups (A, B, and C) of 30 animals each, were used in this study. Individual animals were tagged, and samples were collected from the hides and feces of each at a feedlot and again after slaughter. The carcass of each animal was also sampled. Samples were also collected from the feedlot pens, the sides and floors of the transport trucks, and abattoir holding pens. The isolation rate and the number of E. coli O157 were estimated using a combination of immunomagnetic separation and the Most Probable Number technique. RESULTS Cattle hides were more likely to be contaminated with E. coli O157 at the feedlot (31%) than at the abattoir (4%). E. coli O157 was detected in 18% and 12% of cattle feces collected at the feedlot and after slaughter, respectively. E. coli O157 was isolated from truck floors (26%), truck sides (11%), abattoir pen rails (47%), and pen floors (42%). The mean count of E. coli O157 in positive feces was log(10) 1.17 and 2.37 MPN/g at the feedlot and slaughter, respectively. A 3 log(10) increase in the number of E. coli O157 was observed between the feedlot (2.66 MPN/g) and slaughter (5.66 MPN/g) in the feces of one animal in group B. E. coli O157 was isolated from the hide and carcass of this animal. CONCLUSIONS Transport and lairage did not lead to an increase in the number or isolation rate of E. coli O157 from cattle. APPLICATIONS Intervention strategies for reducing E. coli O157 contamination of cattle carcasses should target mechanisms that limit the impact of animals shedding a high number throughout production and processing.

Quantitative effects of in-line operations on Campylobacter and Escherichia coli through two Australian broiler processing plants

Campylobacter is an important food borne pathogen, mainly associated with poultry. A lack of through-chain quantitative Campylobacter data has been highlighted within quantitative risk assessments. The aim of this study was to quantitatively and qualitatively measure Campylobacter and Escherichia coli concentration on chicken carcasses through poultry slaughter. Chickens (n=240) were sampled from each of four flocks along the processing chain, before scald, after scald, before chill, after chill, after packaging and from individual caeca. The overall prevalence of Campylobacter after packaging was 83% with a median concentration of 0.8log10CFU/mL. The processing points of scalding and chilling had significant mean reductions of both Campylobacter (1.8 and 2.9log10CFU/carcase) and E. coli (1.3 and 2.5log10CFU/carcase). The concentration of E. coli and Campylobacter was significantly correlated throughout processing indicating that E. coli may be a useful indicator organism for reductions in Campylobacter concentration. The carriage of species varied between flocks, with two flocks dominated by Campylobacter coli and two flocks dominated by Campylobacter jejuni. Current processing practices can lead to significant reductions in the concentration of Campylobacter on carcasses. Further understanding of the variable effect of processing on Campylobacter and the survival of specific genotypes may enable more targeted interventions to reduce the concentration of this poultry associated pathogen.

Prevalence and Antimicrobial Resistance of Salmonella and Escherichia coli from Australian Cattle Populations at Slaughter.

Antimicrobial agents are used in cattle production systems for the prevention and control of bacteria associated with diseases. Australia is the worlds third largest exporter of beef; however, this country does not have an ongoing surveillance system for antimicrobial resistance (AMR) in cattle or in foods derived from these animals. In this study, 910 beef cattle, 290 dairy cattle, and 300 veal calf fecal samples collected at slaughter were examined for the presence of Escherichia coli and Salmonella, and the phenotypic AMR of 800 E. coli and 217 Salmonella isolates was determined. E. coli was readily isolated from all types of samples (92.3% of total samples), whereas Salmonella was recovered from only 14.4% of samples and was more likely to be isolated from dairy cattle samples than from beef cattle or veal calf samples. The results of AMR testing corroborate previous Australian animal and retail food surveys, which have indicated a low level of AMR. Multidrug resistance in Salmonella isolates from beef cattle was detected infrequently; however, the resistance was to antimicrobials of low importance in human medicine. Although some differences in AMR between isolates from the different types of animals were observed, there is minimal evidence that specific production practices are responsible for disproportionate contributions to AMR development. In general, resistance to antimicrobials of critical and high importance in human medicine was low regardless of the isolate source. The low level of AMR in bacteria from Australian cattle is likely a result of strict regulation of antimicrobials in food animals in Australia and animal management systems that do not favor bacterial disease.

The ability of Campylobacter jejuni cells to attach to stainless steel does not change as they become nonculturable.

The ability of Campylobacter jejuni strains to attach to stainless steel as they became nonculturable during storage in distilled water at 4 degrees C for 30 days was investigated. From an initial count of approximately 7 log colony-forming units/mL all strains completely lost culturability by day 20, but the numbers of cells attaching to stainless steel remained constant at approximately 3.5 log cells/cm(2). These findings suggest that viable but nonculturable Campylobacter in a liquid matrix can still attach to stainless steel.

Hanging drop: an in vitro air toxic exposure model using human lung cells in 2D and 3D structures.

Using benzene as a candidate air toxicant and A549 cells as an in vitro cell model, we have developed and validated a hanging drop (HD) air exposure system that mimics an air liquid interface exposure to the lung for periods of 1h to over 20 days. Dose response curves were highly reproducible for 2D cultures but more variable for 3D cultures. By comparing the HD exposure method with other classically used air exposure systems, we found that the HD exposure method is more sensitive, more reliable and cheaper to run than medium diffusion methods and the CULTEX(®) system. The concentration causing 50% of reduction of cell viability (EC50) for benzene, toluene, p-xylene, m-xylene and o-xylene to A549 cells for 1h exposure in the HD system were similar to previous in vitro static air exposure. Not only cell viability could be assessed but also sub lethal biological endpoints such as DNA damage and interleukin expressions. An advantage of the HD exposure system is that bioavailability and cell concentrations can be derived from published physicochemical properties using a four compartment mass balance model. The modelled cellular effect concentrations EC50cell for 1h exposure were very similar for benzene, toluene and three xylenes and ranged from 5 to 15 mmol/kgdry weight, which corresponds to the intracellular concentration of narcotic chemicals in many aquatic species, confirming the high sensitivity of this exposure method.

Mixture Effects of Benzene, Toluene, Ethylbenzene, and Xylenes (BTEX) on Lung Carcinoma Cells via a Hanging Drop Air Exposure System

A recently developed hanging drop air exposure system for toxicity studies of volatile chemicals was applied to evaluate the cell viability of lung carcinoma A549 cells after 1 and 24 h of exposure to benzene, toluene, ethylbenzene, and xylenes (BTEX) as individual compounds and as mixtures of four or six components. The cellular chemical concentrations causing 50% reduction of cell viability (EC50) were calculated using a mass balance model and came to 17, 12, 11, 9, 4, and 4 mmol/kg cell dry weight for benzene, toluene, ethylbenzene, m-xylene, o-xylene, and p-xylene, respectively, after 1 h of exposure. The EC50 decreased by a factor of 4 after 24 h of exposure. All mixture effects were best described by the mixture toxicity model of concentration addition, which is valid for chemicals with the same mode of action. Good agreement with the model predictions was found for benzene, toluene, ethylbenzene, and m-xylene at four different representative fixed concentration ratios after 1 h of exposure, but lower agreement with mixture prediction was obtained after 24 h of exposure. A recreated car exhaust mixture, which involved the contribution of the more toxic p-xylene and o-xylene, yielded an acceptable, but lower quality, prediction as well.

Prevalence and Concentration of Arcobacter spp. on Australian Beef Carcasses

The International Commission on Microbiological Specifications for Foods (ICMSF) classified Arcobacter spp. as emerging pathogens in 2002. Arcobacter spp. have been isolated from numerous food products at retail and from animal carcasses and feces at slaughter. A survey was conducted to determine both the prevalence and concentration of Arcobacter spp. on pre-chill beef carcasses. Surface swab samples were collected from 130 beef carcasses at the end of processing, prior to chilling. The concentration of Arcobacter spp. was determined by a most-probable-number per square centimeter (3 by 3) method with a limit of detection of 0.12 CFU/cm(2). Of the 100 carcasses examined from export abattoirs, 20 (20.0%) were contaminated with Arcobacter spp., and 5 of these had quantifiable levels of contamination ranging from 0.12 to 0.31 CFU/cm(2). Of the 30 carcasses examined at a pet food abattoir, 25 (83.3%) were contaminated with Arcobacter spp., and 10 of these had quantifiable levels of contamination ranging from 0.12 to 0.95 CFU/cm(2). Three species of Arcobacter, A. butzleri, A. cryaerophilus, and A. skirowii, were identified by PCR. Each of the species was present in an approximately equal ratio from export abattoirs. This study demonstrates that slaughter practices at export abattoirs are sufficient to maintain both low prevalence and low levels of contamination of beef carcasses with Arcobacter spp.