Thermal and nonthermal factors affecting survival of Salmonella and Listeria monocytogenes in animal manure-based compost mixtures.

Reduction of enteric pathogens in animal manures before field application is essential for mitigating the risk of foodborne illness associated with produce. Aerobic composting of manures has been advocated as an effective treatment for reducing pathogen populations, and heat is a major factor contributing to pathogen inactivation. This study was initiated to determine the potential contribution of both thermal and nonthermal (pH, volatile acids, and ammonia) factors to pathogen inactivation during aerobic composting in bioreactors for mixtures containing manure from various sources (dairy, chicken, and swine). The test mixtures were formulated with an initial moisture content of 60% and a C:N ratio of 20:1, using straw and cottonseed meal as amendments. Mixtures were then inoculated with Salmonella and Listeria monocytogenes labeled with green fluorescent protein at initial populations of ca. 10(7) CFU/g. Three replicate trials of each treatment were conducted. Temperatures within the bioreactors were recorded at 30-min intervals, and duplicate samples were withdrawn daily from two sampling locations within the bioreactor. Significant regression models were derived relating decreases in pathogen populations to the degree of heat generated in the mixture (cumulative heat) and the pH of the mixture on the day before the pathogen losses were calculated (P < 0.0002). Although pathogens in swine manure compost mixtures were inactivated by the third day of composting, very little heat was generated in these mixtures, which were characterized by significantly higher levels of volatile acids compared with the other two compost mixtures. Therefore, volatile acids could help achieve pathogen inactivation when temperatures are too low such as when heat is lost too quickly at the surface of static compost piles or during winter composting.

Examination of factors for use as potential predictors of human enteric pathogen survival in soil.

AIMS: Three soils that varied in their physicochemical characteristics and microbial diversity were inoculated with Escherichia coli O157:H7 and Salmonella to determine the relative impact of abiotic and biotic factors on the pathogens' survival when the soil was held at 25°C. METHODS AND RESULTS: Three soils that were classified as having low, medium and high microbial diversity were divided into two batches for adjustment to 20% of water-holding capacity and to 40% of water-holding capacity. Soils were inoculated with both green fluorescent-labelled E. coli O157:H7 and red fluorescent-labelled Salmonella (5 log CFU g(-1) dry weight) and held at 25°C. Pathogens inoculated into an acidic soil died off within 9 weeks, whereas they were still detected in the other two soils by enrichment culture after 18 weeks. Moisture did not affect inactivation of E. coli O157:H7, but did affect Salmonella inactivation in soil having the greatest organic load and microbial diversity. Using multiple linear regression analysis, 98.7% of the variability in the inactivation rate for E. coli O157:H7 was explained by a model that included the variables of initial pH and electrical conductivity. Salmonella's inactivation rate was predicted by a model that included pH and initial cell numbers of copiotrophic and oligotrophic bacteria. CONCLUSION: This study provided evidence of specific properties that impact inactivation of E. coli O157:H7 and Salmonella in soils at 25°C. SIGNIFICANCE AND IMPACT OF THE STUDY: Identification of factors influential in the die-off of enteric pathogens will assist in developing guidelines for safe intervals between field contamination events and planting or harvesting of fresh-cut produce crops.

Evidence for horizontal and vertical transmission in Campylobacter passage from hen to her progeny.

Campylobacter is an important human pathogen, and consumption of undercooked poultry has been linked to significant human illnesses. To reduce human illness, intervention strategies targeting Campylobacter reduction in poultry are in development. For more than a decade, there has been an ongoing national and international controversy about whether Campylobacter can pass from one generation of poultry to the next via the fertile egg. We recognize that there are numerous sources of Campylobacter entry into flocks of commercial poultry (including egg transmission), yet the environment is often cited as the only source. There has been an abundance of published research globally that refutes this contention, and this article lists and discusses many of them, along with other studies that support environment as the sole or primary source. One must remember that egg passage can mean more than vertical, transovarian transmission. Fecal bacteria, including Campylobacter, can contaminate the shell, shell membranes, and albumen of freshly laid fertile eggs. This contamination is drawn through the shell by temperature differential, aided by the presence of moisture (the "sweating" of the egg); then, when the chick emerges from the egg, it can ingest bacteria such as Campylobacter, become colonized, and spread this contamination to flock mates in the grow house. Improvements in cultural laboratory methods continue to advance our knowledge of the ecology of Campylobacter, and in the not-so-distant future, egg passage will not be a subject continuously debated but will be embraced, thus allowing the development and implementation of more effective intervention strategies.

Prevalence of Salmonella on retail broiler chicken meat carcasses in Colombia.

A cross-sectional study was performed to estimate the prevalence of Salmonella on retail market chicken carcasses in Colombia. A total of 1,003 broiler chicken carcasses from 23 departments (one city per department) were collected via a stratified sampling method. Carcass rinses were tested for the presence of Salmonella by conventional culture methods. Salmonella strains were isolated from 27 % of the carcasses sampled. Logistic regression analysis was used to determine potential risk factors for Salmonella contamination associated with the chicken production system (conventional versus free-range), storage condition (chilled versus frozen), retail store type (supermarket, independent, and wet market), poultry company (integrated company versus nonintegrated company), and socioeconomic stratum. Chickens from a nonintegrated poultry company were associated with a significantly (P < 0.05) greater risk of Salmonella contamination (odds ratio, 2.0) than were chickens from an integrated company. Chilled chickens had a significantly (P < 0.05) higher risk of Salmonella contamination (odds ratio, 4.3) than did frozen chicken carcasses.

Prevalence of Salmonella on raw poultry at retail markets in China.

Data regarding Salmonella on raw poultry are very limited in China. The objective of this study was to determine the prevalence of Salmonella on raw poultry at the retail level in six provinces and two national cities in China. Whole chicken carcasses (n = 1,152) were collected from three types of retail markets (large, small, and wet). All samples were analyzed for the presence of Salmonella by using the U.S. Department of Agriculture, Food Safety Inspection Service method. Of 1,152 chicken samples, overall Salmonella prevalence was 52.2%. The highest prevalence was observed in Guangxi Province (65.3%), next in Guangdong Province (64.6%), and then in Beijing (63.9%), Shaanxi Province (50.7%), Henan Province (47.9%), Shanghai (44.4%), and Fujian Province (42.4%), and lowest prevalence was observed in Sichuan Province (38.9%). Salmonella prevalence was significantly different among the six provinces and two national cities. Salmonella prevalence was highest in the wet markets (54.4%) compared with the large markets (50.3%) and the small markets (52.1%), but differences were not significant (P > 0.05). Good manufacturing practices, good agricultural practices, and hazard analysis critical control point systems for Salmonella control in poultry production at the farm, processing, and retail level should be implemented.

Use of a mixture of bacteriophages for biological control of Salmonella enterica strains in compost.

Bacteriophages specific to Salmonella strains were isolated from sewage effluent and characterized. A five-strain bacteriophage mixture was applied to dairy manure compost inoculated with Salmonella enterica serotype Typhimurium. Bacteriophage treatment resulted in a greater than 2-log-unit reduction of Salmonella within 4 h at all moisture levels compared to the controls.

Comparison of neck skin excision and whole carcass rinse sampling methods for microbiological evaluation of broiler carcasses before and after immersion chilling.

Sampling protocols for detecting Salmonella on poultry differ among various countries. In the United States, the U.S. Department of Agriculture Food Safety and Inspection Service dictates that whole broiler carcasses should be rinsed with 400 ml of 1% buffered peptone water, whereas in the European Union 25-g samples composed of neck skin from three carcasses are evaluated. The purpose of this study was to evaluate a whole carcass rinse (WCR) and a neck skin excision (NS) procedure for Salmonella and Escherichia coli isolation from the same broiler carcass. Carcasses were obtained from three broiler processing plants. The skin around the neck area was aseptically removed and bagged separately from the carcass, and microbiological analysis was performed. The corresponding carcass was bagged and a WCR sample was evaluated. No significant difference (alpha

Microbiological analysis of composts produced on South Carolina poultry farms.

AIMS: The purpose of this study was to determine whether the methods used in compost operations of small and medium-sized poultry farms resulted in the production of an amendment free of foodborne pathogens. METHODS AND RESULTS: Nine compost heaps on five South Carolina poultry farms were surveyed at different stages of the composting process. Compost samples were analysed for coliforms and enriched for Escherichia coli O157:H7, Salmonella spp. and Listeria monocytogenes. The waste materials and composting practices differed among the surveyed farms. On two farms, new materials were added to heaps that had previously completed the active composting phase. Five compost heaps did not reach an internal temperature of 55 degrees C, and c. 62% of all internal samples in the first composting phase contained moisture contents <40%. Escherichia coli was detected in 63% of the surface samples (n = 38) and 9.8% of the internal samples (n = 82) from the first composting phase, as compared with 16.7% of the surface samples (n = 12) and 0% internal samples (n = 24) from the second composting phase. Salmonella was detected in 26 and 6.1% of all surface and internal samples collected from heaps in the first composting phase, respectively, but was absent in all compost samples undergoing a second composting phase. The predominant Salmonella serotypes were Thompson, Montevideo and Anatum. Neither E. coli O157:H7 nor L. monocytogenes was detected in any of the samples. CONCLUSIONS: Our results indicate that the conditions at the compost surface are suitable for pathogen survival, and the complete composting process can result in the elimination of pathogens in poultry wastes. SIGNIFICANCE AND IMPACT OF THE STUDY: This research provides information regarding the effectiveness of the composting practices and microbiological quality of poultry compost produced by small- and medium-sized farms. Ensuring the safety of compost that may be applied to soils should be an integral part of preharvest food safety programme.

Evolving public health approaches to the global challenge of foodborne infections.

The landscape of foodborne infections is in flux. New pathogens emerge, established pathogens may acquire new characteristics and appear in unexpected food vehicles, while many existing problems remain unsolved. Consumers want more fresh foods year round, populations age and migrate, and the technologies and trade practices that produce foods change. Protecting the public health and minimizing the burden of foodborne illness mean expecting the unexpected, and being prepared to understand it when it occurs, so that prevention can be improved. Public health surveillance is also constantly evolving, as new diseases emerge and are judged worthy of notification, as new diagnostic tests change the ease and specificity of routine diagnosis and as social interest in particular issues waxes and wanes. Accurate health information, including reliable estimates of the burden of foodborne disease, can improve foodborne disease prevention, foster global health security, promote economic growth and development and strengthen evidence-based policy making.

Summer meeting 2007 - the problems with fresh produce: an overview.

In Fall 2006, four separate outbreaks of foodborne illness associated with the consumption of fresh produce occurred in the United States. In follow-up investigations, spinach, lettuce, and tomatoes were identified as the vehicles of illness. Epidemiologic investigations subsequently focused on finding the specific growing regions using traceback records. While the areas most likely involved in the outbreaks have been identified, the specific mode of contamination remains unconfirmed. Suspected risk factors in these cases include: proximity of irrigation wells and surface waterways exposed to faeces from cattle and wildlife; exposure in fields to wild animals and their waste materials; and improperly composted animal manure used as fertilizer. Difficulty in deciphering these and other on-farm routes of contamination is due to the sporadic nature of these events. Hence, evidence to support these contamination modes is based largely on experimental studies in the laboratory and field. Still at issue is the relevance of internalization of pathogens, whether this occurs through the roots and plant vascular tissues of vegetables and fruits or through plant surfaces into cracks and crevices. Potential for these events, conditions under which the events occur, and pathogen survival following these events, are questions that still need to be answered. Answers to these questions will ultimately affect the type of interventions needed for application postharvest. Currently, many chemical and biological interventions can reduce surface pathogens and minimize cross-contamination, however, they are largely ineffective on internalized pathogens. In the event internalization is a significant route of contamination in the field, physical interventions (irradiation and high pressure) may be needed to minimize risk. Ultimately, risk assessment studies will be useful tools in developing risk management strategies for the produce industry.

Reducing the carriage of foodborne pathogens in livestock and poultry.

Several foodborne pathogens, including Salmonella species and campylobacters, are common contaminants in poultry and livestock. Typically, these pathogens are carried in the animal's intestinal tract asymptomatically; however, they can be shed in feces in large populations and be transmitted by other vectors from feces to animals, produce, or humans. A wide array of interventions has been developed to reduce the carriage of foodborne pathogens in poultry and livestock, including genetic selection of animals resistant to colonization, treatments to prevent vertical transmission of enteric pathogens, sanitation practices to prevent contamination on the farm and during transportation, elimination of pathogens from feed and water, feed and water additives that create an adverse environment for colonization by the pathogen, and biological treatments that directly or indirectly inactivate the pathogen within the host. To successfully reduce the carriage of foodborne pathogens, it is likely that a combination of intervention strategies will be required.

Role of colanic acid exopolysaccharide in the survival of enterohaemorrhagic Escherichia coli O157:H7 in simulated gastrointestinal fluids.

AIM: This study evaluated the production of colanic acid (CA) exopolysaccharide (EPS) by Escherichia coli O157:H7 in relation to the pathogen's ability to survive under acidic conditions simulating the environment in the human gastrointestinal tract. METHODS AND RESULTS: Escherichia coli O157:H7 W6-13 and its CA-deficient mutant M4020 were examined for their resistance to bile salts, and their ability to survive in simulated gastric fluid containing pepsin (pH 2.0) and simulated intestinal fluid containing pancreatin (pH 8.0). The effect of acid adaptation at pH 5.5 on the survival of E. coli O157:H7 in simulated gastric fluid was also determined. The results indicated that the survivability of M4020, under conditions simulating the environment in the human gastrointestinal tract, reduced more drastically than the viability of W6-13. The presence of bile salts had a slight effect on both types of E. coli O157:H7 cells. The loss of CA did not change the ability of M4020 to respond to acid adaptation. CONCLUSION: The EPS CA may serve as a protective barrier to E. coli O157:H7 for its survival in the human gastrointestinal tract. SIGNIFICANCE AND IMPACT OF THE STUDY: The study contributes to a better understanding of the EPS affecting the ability of E. coli O157:H7 to combat acid stress.

Effect of fat content on infection by Listeria monocytogenes in a mouse model.

An estimated 2,500 cases of listeriosis occur annually in the United States. Listeriosis is particularly severe among pregnant women and immunocompromised individuals. Little is known regarding the effect of the food matrix on the ability of L. monocytogenes to survive in the gastrointestinal tract and cause systemic infection. Mice were inoculated with various doses of L. monocytogenes in skim milk, Half & Half, or whipping cream to determine whether differences in milk fat content influence the ability of L. monocytogenes to survive passage through the gut and infect the liver or spleen. The number of fecal samples positive for L. monocytogenes increased with increasing doses of L. monocytogenes for all three vehicles. The number of L. monocytogenes cells isolated from liver or spleen of mice dosed with L. monocytogenes was not significantly different among treatment vehicles. Dose-response models revealed that as the dosage of L. monocytogenes was increased in different milk vehicles, the number of L. monocytogenes cells in liver or spleen also increased. Although fat content of food had no dose-dependent effect on L. monocytogenes infection in the murine gastrointestinal tract, we cannot discount the possibility that it may be a factor in L. monocytogenes infections of humans because of differences in the physiology of gastrointestinal tracts of mice and humans.

Emerging microbiological food safety issues related to meat.

Avian influenza viruses and antibiotic-resistant pathogens have become topics of current public health interest. This paper will focus on the significance of these pathogens to the meat industry as well as other emerging microbiological food safety topics likely to impact the meat industry. These include surveillance of foodborne pathogens, microbial source tracking, risk assessment, and human populations at increased risk of infection by foodborne microbes. These emerging issues will likely lead to even greater challenges to producing microbiologically safe meat products than the industry has ever experienced. However, accompanying such challenges will be innovative solutions that provide even greater public health protection to meat-containing foods.

Ethanol-mediated variations in cellular fatty acid composition and protein profiles of two genotypically different strains of Escherichia coli O157:H7.

Two strains of Escherichia coli O157:H7 were grown in tryptic soy broth (TSB, pH 7.1) supplemented with 0, 2.5, 5.0, 7.5, and 10% ethanol at 30 degrees C for up to 54 h. Growth rates in TSB supplemented with 0, 2.5, and 5.0% ethanol decreased with an increase in ethanol concentration. Growth was not observed in TSB supplemented with 7.5 or 10% ethanol. The pH of TSB containing 5.0% ethanol decreased to 5.8 within 12 h and then increased to 7.0 at 54 h. The ethanol content in TSB supplemented with 2.5 or 5.0% ethanol did not change substantially during the first 36 h of incubation but decreased slightly thereafter, indicating utilization or degradation of ethanol by both strains. Glucose was depleted in TSB supplemented with 0, 2.5, or 5.0% ethanol within 12 h. Cells grown under ethanol stress contained a higher amount of fatty acids. With the exceptions of cis-oleic acid and nonadecanoic acid, larger amounts of fatty acid were present in stationary-phase cells of the two strains grown in TSB supplemented with 5.0% ethanol for 30 h than in cells grown in TSB without ethanol for 22 h. The trans-oleic acid content was 10-fold higher in the cells grown in TSB with 5.0% ethanol than those grown in TSB without ethanol. In contrast, cis-oleic acid was not detected in ethanol-stressed cells but was present at concentrations of 0.32 and 0.36 mg/g of cells of the two strains grown in TSB without ethanol. Protein content was higher in ethanol-stressed cells than in nonstressed cells. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis protein profiles varied qualitatively as affected by the strain and the presence of ethanol in TSB. An ethanol-mediated protein (28 kDa) was observed in the ethanol-stressed cells but not in control cells. It is concluded that the two test strains of E. coli O157:H7 underwent phenotypic modifications in cellular fatty acid composition and protein profiles in response to ethanol stress. The potential for cross protection against subsequent stresses applied in food preservation technologies as a result of these changes is under investigation.

Consumer acceptance of fresh-cut iceberg lettuce treated with 2% hydrogen peroxide and mild heat.

An antibacterial treatment consisting of 2% hydrogen peroxide at 50 degrees C for 60 s was evaluated for its effects on the sensory quality of fresh-cut iceberg lettuce. Treated lettuce was packaged in polyethylene film by a protocol used in the fresh-cut produce industry and then stored along with untreated controls for 3, 10, and 15 days at 5 degrees C. Gas chromatographic analysis confirmed that the desired initial volume (approximately 6 liters) and oxygen gas content (approximately 10%) were maintained during storage. Consumers from the local community who were the primary shoppers for their households and who purchased and ate lettuce regularly evaluated the appearance, color, aroma, flavor, and texture of the lettuce and their overall liking of the lettuce. Forty consumers were recruited to evaluate processing replication 1 on 26 April, and another 40 were chosen to evaluate processing replication 2 on 3 May. Because replication differences were observed in instrumental color measurements and in mean sensory ratings, replications were not pooled for the two test dates. Lettuce purchased for processing replication 2 was considerably greener than that used in replication 1. Overall, the antibacterial treatment was more effective than the control treatment in maintaining sensory quality over 15 days of storage, provided that the lettuce was initially intensely green. Three-fourths of the participants indicated that they would be willing to buy precut packaged lettuce that had already been treated at the packinghouse or processing plant with an antibacterial solution, and of these participants, 62.5% indicated that they would be willing to pay 5 to 10 cents more per bag.

Fate of Escherichia coli O157:H7 in coleslaw during storage.

An outbreak of Escherichia coli O157:H7 infection associated with the consumption of coleslaw in several units of a restaurant chain prompted a study to determine the fate of the pathogen in two commercial coleslaw preparations (pH 4.3 and 4.5) held at 4, 11, and 21 degrees C for 3 days. At an initial population of 5.3 log10 CFU/g of coleslaw, E. coli O157:H7 did not grow in either coleslaw stored at the three temperatures. Rather, the population of E. coli O157:H7 decreased by 0.1 to 0.5 log10 CFU/g within 3 days. The greatest reduction (0.4 and 0.5 log10 CFU/g) in population occurred at 21 degrees C, whereas only slight decreases (0.1 to 0.2 log10 CFU/g) occurred at 4 and 11 degrees C. A pH of 4.3 to 4.5 of coleslaw had little effect on reducing E. coli O157:H7 populations. Results suggest that the tolerance of E. coli O157:H7 to acid pH, not temperature abuse, is a major factor influencing the pathogen's fate in restaurant-prepared coleslaw.

Consumer acceptance of raw apples treated with an antibacterial solution designed for home use.

An antibacterial treatment consisting of 1.5% lactic acid plus 1.5% hydrogen peroxide at 40 degrees C for 15 min was effective in reducing foodborne bacterial pathogens on raw apples. However, the effects of this treatment on an apple's sensory characteristics and the extent of consumers' willingness to use the treatment at home were not known. This study was undertaken to determine the sensory acceptability and chemical characteristics (pH, soluble solids, and total acidity) of apples subjected to the sanitizing treatment and to obtain information on consumers' purchase behavior, apple handling and consumption practices, and willingness to use an antibacterial treatment. Untrained consumers (n = 80) evaluated the appearance, color, aroma, flavor, texture, and overall appeal of untreated (control) and treated Red Delicious apples that had been stored at 5 degrees C for 0, 6, and 10 days. Panelists used a nine-point hedonic scale (1 = "dislike extremely"; 5 = "neither like nor dislike"; 9 = "like extremely") to evaluate sensory acceptability. Treatment and storage had no significant effect on the appearance, color, or aroma of the samples. Flavor ratings ranged from 6.2 ("like slightly") to 7.0 ("like moderately"). There was no significant difference among any of the control and treated apples stored for 0 days or among those stored for 6 days. Although apples stored for 10 days received the lowest ratings (6.2 to 6.3), they still had an acceptable flavor (6, "like slightly"), and panelists could not perceive differences between the control samples and the treated samples on day 10. The same trends were noted in texture ratings and in overall liking ratings. Treatment and storage had a minimal effect on pH (range, 3.96 to 4.02), soluble solids (range, 11.8 to 12.9 degrees Brix), and total acidity (range, 0.20 to 0.23% malic acid), which are important for apple flavor. Many consumers (87%) were concerned about fruit safety, and 53.2% were willing to try an antibacterial treatment at home. However, 74% would not be willing to use it if a 15-min heating-and-soaking step were required. Implementation of the treatment may be more feasible in the packinghouse than in the home.

Evaluation of universal preenrichment broth for growth of heat-injured pathogens.

Universal preenrichment broth (UPB) was developed to enable enrichment of injured foodborne pathogens of different genera simultaneously in lieu of having to undergo separate simultaneous enrichment cultures for subsequent detection or isolation of each pathogen. Enrichment conditions in UPB for growth of injured pathogens to populations that will enable pathogen detection by rapid immuno-based or polymerase chain reaction (PCR)-based assays have not been defined. Hence, studies were done to determine recovery and growth rates of heat-injured Escherichia coli O157:H7, Salmonella enterica ser. Typhimurium, Salmonella enterica ser. Enteritidis. and Listeria monocytogenes in UPB. Bacterial cells were heat injured in tryptic phosphate broth at 57.2 degrees C and inoculated at populations of ca. 0.17 to 63 injured cells per ml with raw ground beef, fresh chicken, lettuce, and environmental sponge samples. Enrichment cultures were sampled at 1, 2, 3, 4, 5, 6, and 24 h at 37 degrees C postinoculation, and pathogens were enumerated on appropriate selective media. Results revealed that recovery and growth of pathogens during the first 6 h of enrichment were not sufficient to ensure adequate numbers of bacteria (> 10(3) CFU/ ml) for detection by most immunoassays or PCR assays. Cells often required 3 to 4 h for recovery before growth was initiated. Salmonella Typhimurium, Salmonella Enteritidis, E. coli O157:H7, or L. monocytogenes cell populations in enrichment cultures with ground beef or lettuce at 6 h were 0.5 to 2.9 log10 CFU/ml. At 24 h of incubation, cell counts of enrichment samples for the three pathogens from all food and environmental sponge samples ranged from 4.0 to 8.3 log10 CFU/ml. Enrichment in UPB at 37 degrees C of foods or environmental sponge samples containing heat-injured cells of Salmonella Typhimurium, Salmonella Enteritidis, E. coli O157:H7, or L. monocytogenes reliably provides at 24 h of incubation-but not at 6 h-sufficient cell populations for detection by rapid immunoassay or PCR assay procedures that can detect at least 4 log10 CFU/ml. These results raise questions regarding the sensitivity of rapid detection methods that employ an abbreviated enrichment protocol of 6 h or less.

Highly stereoselective syntheses of five- and seven-membered ring heterocycles from ylides generated by catalytic reactions of styryldiazoacetates with aldehydes and imines.

[reaction--see text] Styryldiazoacetates are effective reactants for ylide formation that results in the formation of dihydropyrroles and dihydroazepines with high stereocontrol and in high yields.