Tineke H. Jones

The feline calicivirus as a sample process control for the detection of food and waterborne RNA viruses.

Many food and waterborne outbreaks of infectious disease are caused by viruses. While numerous methods exist and are being developed to test food and water for the presence of enteric viruses, there is no standard control for the comparison of different methods. Potential control viruses should be well characterized, share the physical characteristics of the enterically infecting viruses and not normally be associated with foods. Here, the feline calicivirus (FCV) is proposed as a sample process control for methods aimed at the extraction and detection of RNA viruses in food and water. FCV is shown to be useful as a control for the extraction of hepatitis A virus (HAV) from water using filtration technology and from strawberries using the Pathatrix system. The FCV standard provides a valuable quality control tool when testing potentially contaminated food samples.

Evaluation of the hygienic performances of the processes for cleaning, dressing and cooling pig carcasses at eight packing plants

The hygienic performances of the processes for the production of cooled carcasses at eight pork packing plants were assessed from small sets of microbiological data. At each plant, a single sample was obtained from a randomly selected site on each of 25 randomly selected carcasses at each of three stages of processing, which were after polishing, after washing at the end of the dressing process, and after cooling. The aerobic bacteria, coliforms and Escherichia coli recovered from each sample were enumerated. When bacteria of one type were recovered from > or = 20 of 25 samples, the log mean number of those bacteria on the population of carcasses undergoing processing was estimated on the assumption that the set of counts was normally distributed. The log of the total number recovered from 25 samples was calculated for each set of counts. The log mean numbers of total aerobic bacteria recovered from the polished carcasses at different plants ranged from about 1.9 to 3.8 log cfu cm(-2). At six of the plants, the log mean numbers of total aerobes on the cooled carcasses did not differ substantially from the log mean numbers on the polished carcasses, but the log mean numbers on the cooled carcasses were substantially higher at one plant and substantially lower at another than on the polished carcasses. Coliforms and E. coli were recovered from too few samples in most sets from cooled carcasses for estimation of their log mean numbers. However, the log total numbers of coliforms and E. coli recovered indicated that substantial numbers of those organisms were added to carcasses during the dressing processes at four of the plants, and that the numbers on the carcasses were substantially reduced by the processes for cooling without spraying at two of the plants. At seven of the plants, the total numbers of coliforms and E. coli recovered from cooled carcasses were <3.1 and <2.2 log cfu 2500 cm(-2), respectively. The findings indicate that production processes for pig carcasses can be operated to give cooled carcasses with log mean numbers of total aerobes < 2 cm(-2), and log total numbers of coliforms and E. coli each < 1 2500 cm(-2).

The behaviour of log phase Escherichia coli at temperatures that fluctuate about the minimum for growth

Aims:  To investigate the behaviour of cold‐adapted, log phase Escherichia coli exposed to temperatures that fluctuate below and above the minimum for growth.

The display life of retail packaged pork chops after their storage in master packs under atmospheres of N2, CO2 or O2 + CO2

Commercial, bone-in pork loins were divided into four portions. One portion of each loin was vacuum packaged, then stored at -1.5 °C. The other portions were each divided into three chops, which were retail packaged. The retail packs were master packaged under atmospheres of N(2), CO(2) or O(2) + CO(2) (2:1, v/v), then stored at 2 °C. The pork was assessed after storage for up to 42 days. At each assessment, a vacuum pack and a master pack of each type, each containing product from the same loin, were withdrawn from storage. The vacuum packaged portion was cut into three chops, which were retail packaged. The chops from all packagings were displayed in a retail cabinet which maintained average air temperatures between 3 and 6 °C. The chops were assessed twice daily until they were judged to be of undesirable appearance. After storage for 1 or 2 days, the chops from all master packs appeared less desirable than the freshly cut chops. After all longer storage times, chops from N(2) and CO(2) atmospheres appeared as desirable as freshly cut chops, as did chops from O(2) + CO(2) that were stored for up to 16 days. However, chops stored under O(2) + CO(2) for 21 days appeared undesirable. Chops stored under N(2) or O(2) + CO(2) developed spoilage odours, after storage for 28 or 21 days, respectively. Bacteria were more numerous on the fat than on the muscle tissue. The numbers of bacteria were 10(7) cfu cm(-2) on the fat surfaces of chops stored under vacuum or N(2) for 42 days. The numbers of bacteria were 10(6) cfu cm(-2) on the fat surfaces of chops stored under CO(2) for 42 days or under O(2) + CO(2) for 21 days. At those times, only lactobacilli were isolated from chops stored under CO(2), but small or large fractions of enterobacteria were present in the flora on chops stored under vacuum or N(2), respectively, while the flora on chops stored under O(2) + CO(2) contained large fractions of Brochothrix thermosphacta and Gram negative, strictly aerobic, spoilage bacteria. After all storage times, chops cut from vacuum packaged portions remained of desirable appearance when displayed for 48 hr or longer. Chops stored under N(2) or CO(2) for between 2 and 35 days, or under O(2) + CO(2) for between 4 and 12 days, retained a desirable appearance during display for the same times as the freshly cut chops. Off-odours were apparent in chops after their display following storage under vacuum or CO(2) for 21 days, or under N(2) or O(2) + CO(2) for 12 days. The numbers of bacteria on the fat surfaces of chops spoiled by off-odours were ≥ 10(5) cfu cm(-2). The flora on chops removed from display were generally enriched for B. thermosphacta, enterobacteria and/or Gram negative aerobes as compared with the flora on the chops when they were removed from the storage packs. Those data indicate that the storage life of master packaged, display ready pork will probably be severely limited by the poor hygienic condition of commercial products, to little more than 1 week for product stored under N(2) or O(2) + CO(2) or < 3 weeks for product stored under CO(2).

Coherence among Different Microbial Source Tracking Markers in a Small Agricultural Stream with or without Livestock Exclusion Practices

ABSTRACT Over 1,400 water samples were collected biweekly over 6 years from an intermittent stream protected and unprotected from pasturing cattle. The samples were monitored for host-specific Bacteroidales markers, Cryptosporidium species/genotypes, viruses and coliphages associated with humans or animals, and bacterial zoonotic pathogens. Ruminant Bacteroidales markers did not increase within the restricted cattle access reach of the stream, whereas the ruminant Bacteroidales marker increased significantly in the unrestricted cattle access reach. Human Bacteroidales markers significantly increased downstream of homes where septic issues were documented. Wildlife Bacteroidales markers were detected downstream of the cattle exclusion practice where stream and riparian habitat was protected, but detections decreased after the unrestricted pasture, where the stream and riparian zone was unprotected from livestock. Detection of a large number of human viruses was shown to increase downstream of homes, and similar trends were observed for the human Bacteroidales marker. There was considerable interplay among biomarkers with stream flow, season, and the cattle exclusion practices. There were no to very weak associations with Bacteroidales markers and bacterial, viral, and parasitic pathogens. Overall, discrete sample-by-sample coherence among the different microbial source tracking markers that expressed a similar microbial source was minimal, but spatial trends were physically meaningful in terms of land use (e.g., beneficial management practice) effects on sources of fecal pollution.

Behaviour of log-phase Escherichia coli at temperatures near the minimum for growth

The behaviour of cold-adapted, log-phase Escherichia coli in broth cultures incubated at temperatures between 7 and 15 degrees C was examined by determinations of numbers of colonies recovered on plate count agar (PCA); absorbance at 600 nm (A600); cell lengths from photomicrographs; and cell size distributions by flow cytometry. Cultures incubated between 7 and 10 degrees C were evaluated for 8 days or until A600 values approached 1.0. Cultures incubated at > or =12 degrees C were subcultured to maintain them in the log phase for up to 8 days. Numbers of colonies recovered declined when cultures were incubated at 7 degrees C, but increased when cultures were incubated at higher temperatures. However, A600 values increased during incubation at all temperatures. The mean lengths of cells doubled during incubation at 7 degrees C for 8 days, but remained constant during incubation at 10 degrees C for 1.25 days. Forward angle light scatter (FALS) measurements obtained by flow cytometry indicated that the mean length of cells increased at < or = 8 degrees C, but not at 10 degrees C. A reference value at the 90th percentile of FALS measurements on day 0 was used to determine changes in the distribution of the lengths of cells. About 80% or 17% of the cells were above the reference value after 5 days of incubation at 7 degrees C or 1.25 days of incubation at 10 degrees C, respectively. Cultures that were maintained in the log phase at 12 degrees C became increasingly heterogeneous in cell size after 2 days, but cultures that were maintained at 13 degrees C remained constant in cell size for 8 days. The observations have implications for the prediction of mesophile proliferation at temperatures that approach their minima for growth.

Assessment of the hygienic performances of an air-cooling process for lamb carcasses and a spray-cooling process for pig carcasses.

The air-cooling process for carcasses at a lamb slaughtering plant and the blast-plus-spray-cooling process for carcasses at a pig slaughtering plant were examined. Temperature histories were collected from the deep leg, the aitch bone pocket surface and randomly selected surface sites of carcasses passing through each process. For each process, sets of 25 temperature histories were collected for each type of site, with a single history being collected from each of 75 randomly selected carcasses. A swab sample was obtained from a randomly selected site on each of 25 randomly selected carcasses entering and 25 leaving each process. Total aerobic counts, coliforms and Escherichia coli were enumerated in each sample. Lamb carcasses resided in the chiller for between 17.5 and 66.8 h, and pig carcasses for between 14.8 and 24.5 h. All the lamb carcasses attained deep leg and aitch bone pocket surface temperatures < 7 degrees C as did most pig carcasses. However, those temperatures remained > 13 degrees C in 8% of pig carcasses. Such inadequate cooling of pig carcasses was not apparent in temperature histories from randomly selected surface sites as such sites on both pig and lamb carcasses all attained temperatures < 7 degrees C. Proliferation values for E. coli and psychotrophic pseudomonads calculated for the temperature history from each randomly selected surface site indicated that growth of E. coli on either lamb or pig carcasses would be undetectable, but that increases in the log numbers of total aerobic counts of > 1 and < 1 during the cooling processes could be expected for lamb and pig carcasses respectively. Enumerations of bacteria showed that bacteria on pig carcasses behaved much as would be expected from the temperature histories from randomly selected sites. However, on lamb carcasses the log numbers of bacteria were reduced by about 0.5, 1.5 and 2 for total aerobic counts, coliforms, and E. coli, respectively. The findings indicate that microbiological data are required to properly assess the microbiological effects of carcass cooling processes because, in some, factors other than temperature determine the behavior of the microflora.

Long-term monitoring of waterborne pathogens and microbial source tracking markers in paired agricultural watersheds under controlled and conventional tile drainage management.

ABSTRACT Surface waters from paired agricultural watersheds under controlled tile drainage (CTD) and uncontrolled tile drainage (UCTD) were monitored over 7 years in order to determine if there was an effect of CTD (imposed during the growing season) on occurrences and loadings of bacterial and viral pathogens, coliphages, and microbial source tracking markers. There were significantly lower occurrences of human, ruminant, and livestock (ruminant plus pig) Bacteroidales markers in the CTD watershed in relation to the UCTD watershed. As for pathogens, there were significantly lower occurrences of Salmonella spp. and Arcobacter spp. in the CTD watershed. There were no instances where there were significantly higher quantitative loadings of any microbial target in the CTD watershed, except for F-specific DNA (F-DNA) and F-RNA coliphages, perhaps as a result of fecal inputs from a hobby farm independent of the drainage practice treatments. There was lower loading of the ruminant marker in the CTD watershed in relation to the UCTD system, and results were significant at the level P = 0.06. The odds of Salmonella spp. occurring increased when a ruminant marker was present relative to when the ruminant marker was absent, yet for Arcobacter spp., the odds of this pathogen occurring significantly decreased when a ruminant marker was present relative to when the ruminant marker was absent (but increased when a wildlife marker was present relative to when the wildlife marker was absent). Interestingly, the odds of norovirus GII (associated with human and swine) occurring in water increased significantly when a ruminant marker was present relative to when a ruminant marker was absent. Overall, this study suggests that fecal pollution from tile-drained fields to stream could be reduced by CTD utilization.

Control of the contamination of pig carcasses by Escherichia coli from their mouths

Swab samples from the mouths of 40 pig carcasses all yielded Escherichia coli, at estimated log mean numbers of 2.3 per sample. Single colonies of E. coli O157:H7 were isolated from two, and single colonies of other verotoxigenic strains of E. coli from a further two of those samples. E. coli from the mouth were deposited on the surfaces of carcasses during usual, commercial carcass dressing operations for cutting open the throat and the floor of the mouth, and freeing the tongue. After those operations had been performed, E. coli were recovered from a neck site on 45 of 50 carcasses which were sampled by swabbing that site. When 50 carcasses on which those operations had been performed were pasteurized, the log mean numbers of total aerobic counts recovered from the neck site on carcasses were reduced by two as compared with the unpasteurized carcasses, and only one E. coli was recovered from those carcasses. Pasteurization bleached the small areas of cut muscle exposed during the operations on the mouth and throat, but apparently had no other effect on the appearance of the carcasses. It is suggested that pasteurization of pig carcasses after operating on and regulatory inspection of the mouth and throat may be a commercially viable approach to reducing the contamination of pork with pathogens from the mouths of carcasses.

Filament formation by foodborne bacteria under sublethal stress.

A number of studies have reported that pathogenic and nonpathogenic foodborne bacteria have the ability to form filaments in microbiological growth media and foods after prolonged exposure to sublethal stress or marginal growth conditions. In many cases, nucleoids are evenly spaced throughout the filamentous cells but septa are not visible, indicating that there is a blockage in the early steps of cell division but the mechanism behind filament formation is not clear. The formation of filamentous cells appears to be a reversible stress response. When filamentous cells are exposed to more favorable growth conditions, filaments divide rapidly into a number of individual cells, which may have major health and regulatory implications for the food industry because the potential numbers of viable bacteria will be underestimated and may exceed tolerated levels in foods when filamentous cells that are subjected to sublethal stress conditions are enumerated. Evidence suggests that filament formation under a number of sublethal stresses may be linked to a reduced energy state of bacterial cells. This review focuses on the conditions and extent of filament formation by foodborne bacteria under conditions that are used to control the growth of microorganisms in foods such as suboptimal pH, high pressure, low water activity, low temperature, elevated CO2 and exposure to antimicrobial substances as well as lack a of nutrients in the food environment and explores the impact of the sublethal stresses on the cells inability to divide.