Survival of methicillin-resistant Staphylococcus aureus during thermal processing of frankfurters, summer sausage, and ham.

Infections from antibiotic-resistant bacteria are a major concern for human health professionals around the world. Methicillin-resistant Staphylococcus aureus (MRSA) is just one of the resistant organisms of concern. MRSA prevalence has also been recently reported in retail meat products at rates higher than originally thought. Although the risk of contracting an infection from handling contaminated meat products is thought to be low, very little is known about this organism from a food safety perspective. The objective of this study was to determine the survival of MRSA during thermal processing of frankfurters, summer sausage, and boneless ham. Frankfurters, summer sausage, and boneless ham were manufactured using formulations and processing procedures developed at the Iowa State University meat laboratory. Thermal processing resulted in a significant log reduction (p<0.05) for boneless ham, summer sausage, and frankfurters when compared to uncooked, positive controls for each of the three processed meat products. All products were thermally processed to an internal temperature of 70°C and promptly cooled to 7.2°C. Boneless ham showed the highest log reduction (7.28 logs) from cooking, followed by summer sausage (6.75 logs) and frankfurters (5.53 logs). The results of this study indicate that thermal processing of ham, summer sausage, and frankfurters to 70°C is sufficient to reduce the risk of MRSA as a potential food safety hazard.

Inactivation of feline calicivirus as a surrogate for norovirus on lettuce by electron beam irradiation.

Caliciviridae, including norovirus, are considered important sources of human gastroenteritis. As leafy green vegetables are commonly consumed without additional processing, it is important to evaluate interventions to reduce the presence of human pathogens in these products. Feline calicivirus was used as a model for small round structured viruses on lettuce. The lettuce was inoculated by immersion to simulate contamination from irrigation or wash water. The inoculated lettuce was then exposed to electron beam irradiation at various dose levels to determine survival. The D10-value of the calicivirus on lettuce was determined to be 2.95 kGy. Irradiation to reduce bacterial pathogens on cut lettuce could also reduce the risk associated with small round structured viruses on lettuce.

Impact of irradiation on the safety and quality of poultry and meat products: a review.

For more than 100 years research on food irradiation has demonstrated that radiation will make food safer and improve the shelf life of irradiated foods. Using the current food safety technology, we may have reached the point of diminishing returns even though recent figures from the CDC show a significant drop in the number of foodborne illnesses. However, too many people continue to get sick and die from eating contaminated food. New and under utilized technologies such as food irradiation need to be re-examined to achieve new levels of safety for the food supply. Effects of irradiation on the safety and quality of meat and poultry are discussed. Irradiation control of the principle microbial pathogens including viruses, the differences among at-risk sub-populations, factors affecting the diminished rate of improvement in food safety and published D values for irradiating raw meat and poultry are presented. Currently permitted levels of irradiation are probably not sufficient to control pathogenic viruses. Typical gram-negative spoilage organisms are very sensitive to irradiation. Their destruction leads to a significant increase in the acceptable shelf life. In addition, the destruction of these normal spoilage organisms did not provide a competitive growth advantage for irradiation injured food pathogens. Another of the main focuses of this review is a detailed compilation of the effects of most of the food additives that have been proposed to minimize the negative quality effect of irradiation. Most of the antimicrobials and antioxidants used singly or in combination produced an increased lethality of irradiation and a decrease in oxidation by-products. Combinations of dosage, temperature, dietary and direct additives, storage temperature and packaging atmosphere can produce meats that the average consumer will find indistinguishable from non-irradiated meats. A discussion of the production of unique radiological by-products is also included.

Isolation and Purification of Colicin ECL 12, a Bacteriocin That Inhibits Strains of Escherichia coli O157:H7 †.

A swine fecal isolate, identified as Escherichia coli ECL12, was found to produce an antimicrobial substance designated as colicin ECL12. Colicin ECL12 was inhibitory against 20 strains of E. coli O157:H7 previously isolated from both human and bovine feces. Identification of the producer strain was determined phenotypically by biochemical and morphological tests. Colicin ECL12 was sensitive to several proteolytic enzymes. Adsorption of colicin ECL12 to sensitive cells of E. coli O157:H7 was bactericidal, resulting in a 2 log reduction in viable cell counts. Colicin ECL12 was purified from strain ECL12 by cell extraction and ion-exchange chromatography. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of colicin ECL12 resolved a single protein with a molecular weight of approximately 65,000.

Irradiation and Packaging of Fresh Meat and Poultry.

Extensive research on the irradiation of fresh meat and poultry has been carried out during the past 40 years; yet there is a need for consideration of combined use with other technologies such as modified atmosphere (MA) packaging. Some of the past work has focused on spoilage microorganisms and pathogens, whereas other reports emphasize the sensory quality of fresh meat and poultry. Reports published indicate that the effects of irradiation in conjunction with packaging vary depending upon the kind of meat and poultry and the atmosphere composition in the package. Irradiation may result in off-flavor and/or odor and discoloration of fresh meat and poultry in packages containing air (oxygen). One concern is that pathogens may grow and/or produce toxins in irradiated meat or poultry packaged using modified atmospheres because of a lack of competing organisms. This is of even greater concern if spoilage is suppressed and does not provide the usual warning signals. On the other hand, even though there is some evidence of the growth of pathogens in temperature-abuse conditions, most reports have indicated that spoilage preceded toxin production. Therefore, considering the sensory quality and concerns for safety, the effects of irradiation in combination with vacuum or MA packaging of fresh meat and poultry should be studied further. More complete information is needed to ensure the appropriate use of vacuum or MA packaging in combination with irradiation for the safety of fresh meat and poultry.

Survival and Injury of Listeria monocytogenes , Listeria innocua and Listeria ivanovii in Ground Pork Following Electron Beam Irradiation †.

The sensitivity of five strains of Listeria to electron beam irradiation in ground pork as well as the extent of sublethal radiation injury exhibited by each were investigated. Ground pork was inoculated with one of five strains of Listeria and irradiated with from 0 to 1.25 kGy at 0.25 kGy intervals. Listeria innocua NADC 2841 was more radiation-resistant (D10 = 0.638 kGy) than L. monocytogenes NADC 2045 Scott A (D10 = 0.447 kGy), L. monocytogenes NADC 2783 (a hamburger isolate) (D10 = 0.424 kGy), L. monocytogenes ATCC 15313 (D10 = 0.445 kGy), and L. ivanovii NADC 3518 (D10 = 0.372 kGy), when recovered on tryptic soy agar supplemented with 0.6% yeast extract. D10 values for L. innocua , L. ivanovii , and L. monocytogenes ATCC 15313 were lower when cells were recovered on modified Oxford medium. These three strains were susceptible to radiation-induced sublethal injury, with the numbers of injured organisms increasing with irradiation dose. The two pathogenic strains of L. monocytogenes were not injured significantly at the dose levels used. The results show that the dose range currently being considered by the Food and Drug Administration for the irradiation of beef and pork (1.5 to 4.5 kGy) is adequate for the elimination of L. monocytogenes from pork.

Use of High Hydrostatic Pressure and Irradiation To Eliminate Clostridium sporogenes Spores in Chicken Breast.

High pressure has been studied for its usefulness in reducing microbial contaminants in foods. We sought to determine whether this technology could be used in combination with irradiation to develop shelf-stable products. We first determined the optimal pressure, temperature, and time conditions that would result in maximum reduction of Clostridium sporogenes spores in fresh chicken. At ambient temperature, a pressure of 6,800 atm for up to 60 min resulted in a 5-log-unit reduction. Heating the samples during pressurization at 80°C for 20 min resulted in the lowest number of survivors compared to samples that were heated and pressurized for only 1 and 10 min. Further, irradiation at a medium dose (3.0 kGy) before and after pressurization at 6,800 atm and 80°C for 1, 10, and 20 min revealed no significant differences in spore counts between samples that were pressurized and then irradiated or vice-versa. We then examined the effect of high pressure in lowering the irradiation dose necessary to eliminate all spores. The irradiation D value of C. sporogenes spores was calculated to be 4.1 kGy. Samples were then irradiated at various doses followed by pressurization at 6,800 atm at 80°C for 20 min. The irradiation D value was lowered to approximately 2 kGy, indicating that a combination of high hydrostatic pressure and irradiation can be used to produce chicken with an extended shelf life without the use of high irradiation doses.

Isolation and Purification of Enterocin EL1, a Bacteriocin Produced by a Strain of Enterococcus faecium †.

A meat isolate, identified as Enterococcus faecium L1, was found to produce a bacteriocin designated enterocin EL1 Enterocin EL1 was active against a narrow spectrum of microorganisms, inhibiting all tested strains of Listeria . Identification of the producer strain was determined phenotypically by biochemical and morphological tests. Enterocin EL1 was heat stable, sensitive to several proteolytic enzymes, and stable from pH 2 to 11. Adsorption of the bacteriocin to producer cells was dependent on ionic interaction of the bacteriocin and the cell surface at various pHs. By changing the pH of the extraction buffer, enterocin EL1 was extracted from E. faecium L1 cells in a concentrated form. Enterocin EL1 isolated by cell extraction was resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis as a protein with an approximate molecular weight of 2,300. Partially purified enterocin EL1 added to sensitive cells of Listeria ivanovii was bactericidal; however, the bacteriocin did not inhibit the producer strain L1.

Elimination of Pathogens of Significance in Food by Low-dose Irradiation: A Review.

Food irradiation is a processing technology that has been shown to be a wholesome process by many scientific studies conducted worldwide during the past 40 years. The research has been supported by the World Health Organization, the Food and Agricultural Organization, and govemmental agencies in many different countries. Industrial support also has been substantial. Some of the benefits ascribed to this technology include improved shelf life, reduced use of Chemicals as preservatives, and reduced levels of pathogens in foods. Pathogens such as Listeria monocytogenes , Yersinia enterocolitica , and Aeromonas hydrophila are capable of growing at temperatures as low as 0°C and are considered to pose a threat to the safety of refrigerated products. The number of cases of foodborne illness caused by contamination by Salmonella and Campylobacter spp. continues to increase. Researchers have been investigating ways in which food safety can be improved without sacrificing product quality and wholesomeness. The sensitivity of these pathogens to low-dose irradiation has been studied in several food products. Survival curves have been elucidated, and some studies on the effects of storage atmosphere, storage temperature, heating, and various treatments in combination with irradiation have been conducted. This review presents background information on this technology, with an emphasis on the radiation sensitivity of some pathogens of importance. Suggestions for future work in this area are also discussed.

Metal Ions Reverse the Inhibitory Effects of Selected Food-Grade Phosphates in Staphylococcus aureus.

Previous studies showed that levels as low as 0.1% sodium ultraphosphate (UP), 0.1% sodium polyphosphate glassy (SPG) and 0.5% tetrasodium pyrophosphate (TSPP) were bactericidal and bacteriolytic to early-exponential phase cells of Staphylococcus aureus ISP40 8325. In the present study, Ca2+ (0.01 M) or Mg2+ (0.01 M) reversed the bacteriolytic effects of UP (0.1%) and SPG (0.1%) to S. aureus . In addition, Ca2+ (0.01 M) or Mg2+ (0.01 M), when added to the culture medium before inoculation, protected cells from growth inhibition by UP and SPG. Moreover, the bactericidal effects of UP or SPG were reversed by Ca2+ or Mg2+ in metal-rescue experiments in which the metals were added to polyphosphate-containing medium after 1 h of incubation. No additive effect existed between Mg2+ and Ca2+. Growth inhibition of TSPP was not reversed by Mg2+ or Ca2+, but it was reversed by Fe3+ when Fe3+ was added to protect cells 1 h before the addition of TSPP. These studies show that the antibacterial effects of phosphates can be altered substantially by the metal-ion content of the environment.

Antibacterial Mechanism of Long-Chain Polyphosphates in Staphylococcus aureus.

The results of previous studies indicated that the antibacterial effects of long-chain polyphosphates (sodium polyphosphate glassy [SPG] and sodium ultraphosphate [UP]) to Staphylococcus aureus ISP40 8325 could be attributed to damage to the cell envelope (cell wall or cell membrane). Also, Ca2+ (0.01 M) or Mg2+ (0.01 M) reversed the bactericidal and bacteriolytic effects of polyphosphates in S. aureus . In the present study, 0.4 M sodium chloride (NaCl) protected the cells from leakage caused by SPG and 0.6 M NaCl protected the cells from leakage by UP. Polymyxin, a peptide antibiotic that causes cell membrane damage, induced leakage even in the presence of 0.6 M NaCl. In the presence of 0.4 M NaCl, bacterial leakage was significantly reduced by disodium ethylenediamine tetraacetate (EDTA), a metal chelator that causes cell wall damage. Bacterial leakage by polyphosphates was significantly greater at pH 8 than at pH 6, which suggested that metal-ion chelation was involved in the antibacterial mechanism. A dialysis membrane (MWCO 100) was used to separate free metal and polyphosphate-bound metal. Levels of free Ca2+ and Mg2+ in polyphosphate-treated cells were significantly lower than those of the cells without polyphosphate. This free-metal dialysis study provided Chemical evidence to show that long-chain polyphosphates interacted with S. aureus cell walls by a metal-ion chelation mechanism. In addition, long-chain polyphosphates were shown to bind to the cell wall, chelate metals, and remain bound without releasing metal ions from the cell wall into the suspending medium. A hypothesis is proposed in which the antibacterial mechanism of long-chain polyphosphates is caused by binding of long-chain polyphosphates to the cell wall of early-exponential phase cells of S. aureus ISP40 8325. The polyphosphates chelate structurally essential metals (Ca2+ and Mg2+) of the cell wall, resulting in bactericidal and bacteriolytic effects. The structurally essential metals probably form cross bridges between the teichoic acid chains in the cell walls of gram-positive bacteria.

Bactericidal and Bacteriolytic Effects of Selected Food-Grade Phosphates, Using Staphylococcus aureus as a Model System.

Phosphates have been approved for use in meat products primarily to protect flavor and increase yields. It also is known that phosphates have antimicrobial properties. The objective of this study was to compare the effects of different phosphates in a model system. Minimum inhibitory concentrations (MICs) of selected food-grade phosphates added to early-exponential-phase cells of Staphylococcus aureus ISP40 8325 in a synthetic medium were determined to be 0.1% for sodium ultraphosphate and sodium polyphosphate glassy and 0.5% for sodium acid pyrophosphate, sodium tripolyphosphate and tetrasodium pyrophosphate. Thus, the MIC values for the very long chain-length phosphates were lower than the MIC values for shorter chain-length phosphates. Leakage of intracellular nucleotides was observed both spectrophotometrically (release of A260-absorbing material) and microscopically (appearance of gelatinous cellular aggregates). Treatment of the gelatinous cellular aggregates with DNase, RNase and proteinase indicated that the aggregates contained DNA, RNA and protein, thus indicating cellular lysis in the presence of phosphates.

DEVELOPMENT OF AN ENZYME-LINKED IMMUNOSORBENT ASSAY (ELISA) FOR ANALYSIS OF LISTERIOLYSIN O PRODUCED BY LISTERIA MONOCYTOGENES.

Listeriolysin O (LLO) is a heat-labile hemolysin produced by Listeria mono-cytogenes. Its hemolytic activity has been evaluated qualitatively by sodium dodecyl sulfate (SDS) electrophoresis and immunoblotting. In this experiment, an enzyme-linked immunosorbent assay (ELISA) was developed for quantitative analysis of LLO by using Streptolysin O (SLO) and antistreptolysin O (ASO) as the reagents. The selected coating and blocking buffers were 0.05 M Tris buffer (pH 8.5) and 0.25% casein solution with phosphate-buffered saline solution + 0.05% Tween 20 (PBS-T), respectively. A relationship between ASO and antibody was achieved with 5 mg/ml ASO and a 1:1,000 dilution of conjugate. The heat stability of LLO at 48, 62, 72, and 80C was examined by using this method and compared with a traditional hemolysis assay. Although the LLO is inactivated easily at those temperatures, the protein structure was not affected at temperatures lower than 80C for 3 min, pointing to a need for both hemolysis and ELISA to be conducted in determining both the activity and presence of LLO in foods.