Salmonella contamination during production of cantaloupe: a binational study.

Six cantaloupe farms and packing plants in South Texas (950 cantaloupe, 140 water, and 45 environmental samples), including the Rio Grande Valley area, and three farms in Colima State, Mexico (300 cantaloupe, 45 water, and 15 environmental samples), were sampled to evaluate cantaloupe contamination with Salmonella and Escherichia coli during production and processing. Samples collected from external surfaces of cantaloupes, water, and the environments of packing sheds on cantaloupe farms were examined for the presence of Salmonella and E. coli. Of a total of 1,735 samples collected, 31 (1.8%) tested positive for Salmonella. Fifteen Salmonella serotypes were isolated from samples collected in Texas, and nine from samples collected in Colima. Two serotypes (Poona and Oranienburg) that have been associated with three large Salmonella outbreaks in the United States and Canada linked to the consumption of contaminated cantaloupe were found in water samples collected at four farms (three from the United States). Susceptibility of Salmonella isolates to 10 antimicrobials was evaluated by disk diffusion. Eighty-eight percent of the isolates from the United States and Mexico were pansusceptible to the antimicrobials tested; eight isolates from the United States demonstrated an intermediate susceptibility to streptomycin and only two isolates were resistant to the same antimicrobial. From Mexico, four isolates showed an intermediate susceptibility to streptomycin and one isolate was resistant to nalidixic acid and streptomycin. Repetitive sequence-based PCR analysis of Salmonella isolates helped to trace potential sources of Salmonella contamination in source water and in subsequent water samples obtained after the filtration systems of U.S. and Mexican cantaloupe farms. No differences could be seen between the levels of Salmonella contamination in melons from both countries.

Isolation of Arcobacter spp. from retail meats and cytotoxic effects of isolates against vero cells.

A survey of Arcobacter spp. was conducted over a 12-month period in Guadalajara, Mexico. A total of 135 samples (45 lean ground beef samples, 45 lean ground pork samples, and 45 chicken samples, including drumsticks, gizzards, and ground or chopped breast) were collected from local butcheries. The samples were enriched in Johnson-Murano enrichment medium and then streaked onto Johnson-Murano agar plates. Typical colonies were subjected to microscopic and biochemical identification followed by polymerase chain reaction confirmation of the genus Arcobacter. All isolates confirmed to be Arcobacter isolates were then inoculated into Eagle's minimum essential medium to determine their cytotoxicity against Vero cells. Arcobacter spp. were detected in 28.8, 51.1, and 40.0% of beef, pork, and chicken samples, respectively. From these samples, 101 isolates were confirmed to be Arcobacter spp. by polymerase chain reaction. Overall, the species most frequently identified was A. butzleri, followed by A. skirrowii. A. cryaerophilus was isolated only from pork meat. Ninety-five (95%) of the Arcobacter isolates produced a virulence mechanism against Vero cells, and 38 of them induced cell elongation, indicating enterotoxin production. Eighteen isolates produced the formation of vacuoles, and 39 produced both vacuolization and elongation. The vacuolization effect may be related to a vacuolizing toxin. The production of a vacuolizing toxin by Arcobacter spp. has not previously been reported. Results obtained in this study indicate that Arcobacter spp. may show cytotoxic effects other than the recognized enterotoxin production.

Comparison of three protocols for the isolation of Arcobacter from poultry.

The microaerophilic bacterium Arcobacter has received increasing attention in recent years regarding its presence in food products. There exist a limited number of methods for the detection of this microorganism, with currently available methods being cumbersome to perform, time consuming, and limited in specificity. The objective of this study was to develop a selective enrichment broth to isolate accurately three Arcobacter spp. from concentrated chicken microflora by comparing the efficacy of various selective and growth-promoting additives in order. This newly developed enrichment broth was incorporated into an isolation protocol using a previously developed plating medium, and this new protocol was compared with two existing methods for the isolation of Arcobacter from poultry. Method 1 consisted of enrichment in Ellinghausen-McCullough-Johnson-Harris Polysorbate 80 broth followed by plating on cefoperazone-vancomycin-amphotericin B medium. Method 2 consisted of enrichment in Arcobacter selective broth and plating onto Arcobacter selective medium. Method 3 (the JM method), used a newly developed enrichment broth followed by plating on a previously described JM agar. The JM method isolated Arcobacter strains in 42 out of 50 broiler chicken samples, while methods 1 and 2 detected the organism in only 24 and 15 out of 50 samples, respectively.

Development of a new medium for the isolation of Arcobacter spp.

Arcobacter, the newly reclassified Campylobacter species, has been shown to cause diarrhea in both humans and animals. Few studies have been conducted regarding its occurrence in foods because of the lack of effective isolation and identification methods. The purpose of this study was to develop a plating medium that would be selective for the three most commonly found Arcobacter species. The effect of common components used in media intended for the isolation of Campylobacter, Helicobacter, and other gram-negative rods was examined. These components were divided into five distinct groups: (1) basic growth nutrients, (2) reducing and growth-promoting agents, (3) detoxifying agents, (4) antibiotics, and (5) color-enhancing compounds. Components from each of these groups were tested for their ability to recover Arcobacter on a solid medium when incubated aerobically at 30 degrees C for up to 72 h. Growth was evaluated by the ecometric technique, colony size, and differential colony morphology after incubation. After initial evaluations, five formulas showing the best results were selected and tested in detail and compared with brucella agar. A medium containing a basal nutrient mix along with 0.05% thioglycolic acid, 0.05% sodium pyruvate, and 5% sheep's blood (pH 6.9+/-0.2) was found to be the most effective for the growth of A. butzleri, A. cryaerophilus, and A. nitrofigilis. In addition to superior growth characteristics, a deep red color around the colonies also was observed with this formulation.

Application of high hydrostatic pressure to eliminate Listeria monocytogenes from fresh pork sausage.

Ground pork patties were inoculated separately with 10(9) CFU/g each of three strains of Listeria monocytogenes obtained from the National Animal Disease Center (NADC). Inoculated patties were packaged under vacuum and treated at 414 megapascals (60,000 lb/in2) for up to 60 min by high hydrostatic pressure (HHP). Survivors were determined by surface plating onto modified Oxford agar and trypticase soy agar with yeast extract, as well as by the most probable number method using Listeria enrichment broth. Average D values ranged from 1.89 to 4.17 min, depending on the strain, with the most virulent strain (reported by the NADC) having the highest D value. We tested the usefulness of applying a mild heat treatment at 50 degrees C, simultaneously with HHP, to lower these values. Average D values ranged from 0.37 to 0.63 min, depending on the strain. Thus, a 10-log10 reduction could be achieved even in the most pressure-resistant strain of L. monocytogenes by a 6-min application of heat and HHP. Shelf life studies were also conducted, with spoilage levels reached after 5 days of storage at 4 degrees C for controls versus 28 days for treated samples. Sensory evaluation of uninoculated grilled patties showed that panelists could not distinguish between those treated by heat and HHP and untreated controls (P<0.05). Thus, treatment by HHP in combination with mild heating can be used successfully to produce safer, longer-lasting fresh pork without affecting quality.

Influence of various commercial packaging conditions on survival of Escherichia coli O157:H7 to irradiation by electron beam versus gamma rays.

Irradiation of ground beef patties inoculated with the organism Escherichia coli O157:H7 was performed either by gamma rays from a cobalt 60 source or by electron beam generated by a linear accelerator. Patties were packaged in one of the following materials: nylon/polyethylene bags, Saran/polyester/polyethylene bags (PM2), or Saran overwrap with a Styrofoam tray inside. Bags were sealed in air or under vacuum and were irradiated at either 5 or -15 degrees C. Average D10 values (dose required to inactivate 90% of a microbial population) ranged from 0.27 to 0.63 kGy, depending on the conditions. Overall, higher D10 values (P<0.0001) were obtained upon irradiation at -15 degrees C as compared with 5 degrees C. Cells inoculated in samples packaged in PM2 had the highest D10 values, but only if irradiated by electron beam at -15 degrees C (P<0.001). Since PM2 had the lowest oxygen permeability rate and since the temperature was too low for radicals to migrate easily, these conditions may have minimized the effect of oxygen- and water-derived radicals on microbial survival. Irradiation by gamma rays resulted in higher D10 values (P<0.047) than irradiation by electron beam, with the highest values being observed at -15 degrees C. Differences may be attributed to dose rate (1.0 kGy/h for gamma, 17 kGy/min for electron beam) since it is possible that, at low dose rates, microbial enzymes may have more time to repair damage to the cell due to irradiation, resulting in higher D10 values.

Survival of heat-shocked Yersinia enterocolitica after irradiation in ground pork.

Earlier studies conducted in our laboratory showed that heat-shocked Yersinia enterocolitica (45 degrees C for 60 min) are more resistant to a subsequent beat treatment of 55 or 60 degrees C in ground pork than cells not previously heat-shocked. The increased thermotolerance was partly attributed to the production of stress proteins. The present study was performed to determine if the stress proteins produced by heating could also afford protection to the cells to irradiation. As part of the study, the effect of air versus vacuum packaging on survival of Yersinia to irradiation was also examined. Irradiating the inoculated pork at 1.0 kGy was sufficient to completely eliminate this pathogen. The irradiation D value for both heat-shocked and non-heat-shocked cells was statistically the same (0.15 kGy). Neither heat-shocking Yersinia nor packaging under vacuum resulted in increased resistance of this organism to irradiation. In addition, no effect was seen in virulence of the cells after these treatments, when compared with controls.

Food preservation using ionizing radiation.

Irradiation processing has been researched extensively and is now in use worldwide for many food commodities. Irradiation has been successfully used to reduce pathogenic bacteria, eliminate parasites, decrease postharvest sprouting, and extend the shelf life of fresh perishable foods. Although food irradiation is widely accepted in world food markets, U.S. markets have been slower to accept the idea of irradiated food products. For fruits and vegetables, irradiation is not a cure for shelf life problems; cost and quality problems damage preclude its general use. It appears that the most likely use of irradiation in fruits and vegetables is as an insect control in those commodities for which there is no effective alternative method. For grains such as rice and wheat, irradiation has been used primarily to control insect infestation when insects have been shown to develop resistance to the traditional fumigation methods. Treatment of spices with irradiation doses of 10 kGy has proved to extend shelf life without causing significant changes in sensory or chemical quality. Higher doses that effectively sterilize spices, however, may cause undesirable chemical and sensorial changes. For meat, especially red meat, irradiation is considered a viable alternative in the effort to improve the safety of meat products. With time, the authors believe that economic realities and the technical superiority of irradiation for specific poultry products will lead to public acceptance of the process. Irradiation of seafood products is still being considered for approval by the USFDA, although it is currently used in Asian and European markets, especially for shrimp. It is our belief that scientifically based research in food irradiation and the positive results thereof will also prove economical in the twenty-first century. As we move to a more peaceful world with reduced threat of nuclear holocaust, these valid opinions will prevail and will overshadow the distortions and misinformation generated by the opponents of irradiation.

Shelf life extension, safety, and quality of fresh pork loin treated with high hydrostatic pressure.

The optimal conditions of pressure, time, and processing temperature required to eliminate Listeria monocytogenes Scott A and Salmonella typhimurium ATCC 13311 in fresh pork loin and the effect of these optimal conditions on quality and shelf life were determined. Twenty-five grams of fresh pork loin were inoculated with either of the two organisms and were subjected to pressures between 414 and 827 MPa at either 2 or 25 degrees C for 30 min. The D414MPa(25 degrees C) was determined to be 2.17 min for L. monocytogenes and the D414 MPa (2 degrees C) was determined to be 1.48 min for S. typhimurium. Samples subjected to a 6D process were evaluated by sensory and objective tests as well as for shelf life. These samples were found to be different (P < 0.05) from controls when evaluated after cooking by a triangle test of difference, but only when the pressure was applied at 2 degrees C and not at 25 degrees C. The descriptive analysis test showed that cooked samples treated at 25 degrees C were not different (P > 0.05) from controls in flavor, juiciness, and firmness. Color, peak load, water-holding capacity, and moisture were not found to be different (P > 0.05) between samples treated at 25 degrees C and controls when both were cooked. However, in the raw state, differences were found in the values for color parameters L and b. The level of psychrotrophs was 5.7 log CFU/g for samples treated at 25 degrees C after 33 days of storage at 4 degrees C, as compared with 7.0 log CFU/g for controls. The color and peak load (texture) did not change over the storage period (P > 0.05) in any of the samples. All samples spoiled in 5 days when stored at 25 degrees C.

D values of Salmonella enteritidis isolates and quality attributes of shell eggs and liquid whole eggs treated with irradiation.

Irradiation sensitivity of five Salmonella enteritidis isolates inoculated either on the surface or inside of whole shell eggs were determined. The shell eggs were irradiated at doses of 0, 0.5, 1.0, and 1.5 kGy. A minimal dose of 0.5 kGy was sufficient to eliminate all the isolates from the surface of whole eggs; however, the same isolates were more resistant to irradiation when present inside the eggs. The ATCC 13076 isolate was significantly more sensitive to irradiation, with a D value of 0.32 kGy, than the other four isolates from animal origin. Irradiation D values of the latter ranged from 0.39 to 0.41 kGy. Liquid whole eggs were also inoculated (2.4 x 10(6) cells per milliliter) with two S. enteritidis isolates and were heat-treated at 50 C for 0, 20, 40, or 60 min followed by irradiation at 0, 0.25, 0.5, 0.75, or 1.0 kGy. The results indicate that mild heating prior to irradiation was ineffective in reducing the irradiation D values. However, on the basis of the D values obtained, an irradiation dose of 1.5 kGy should be sufficient to reduce Salmonella counts by approximately 4 log10 in both whole shell and liquid eggs. Results also indicate that color and thermal characteristics of the whole or liquid eggs were unaffected by a 1.5-kGy dose of irradiation.

Detection of Arcobacter spp. in Ground Pork by Modified Plating Methods.

A modified cefsulodin-irgasan-novobiocin (CIN) medium was developed for the recovery of Arcobacter spp. from meats. Modified CIN was compared to brain heart infusion agar supplemented with 10% bovine blood and cephalothin, vancomycin, and amphotericin B (CVA) as well as brain heart infusion agar supplemented with 10% bovine blood and no antibiotics. The three media were used to recover Arcobacter spp. in a survey of pork-processing plants. Examination of ground pork (149 samples) from one Iowa slaughter facility (Plant #1) revealed that 89 percent of the samples were positive for Arcobacter spp. In a second survey conducted 9 months later involving that same plant and four others, only 5% of the samples from the four plants were found to be positive for Arcobacter spp. Again, 90% of the samples were positive from Plant #1. It was not determined whether the sanitary practices during slaughter or the rearing of pigs on the source farms contributed to the prevalence of Arcobacter spp. in one plant versus another.

Role of respiratory-burst products from polymorphonuclear leukocytes in the antitumor activity of Propionibacterium acnes vaccine.

Tumor cells injected into Balb/c mice together with heat-killed 48-h P. acnes cells were rendered nontumorigenic as early as 12 h after injection, as determined by the inability of the tumor cells to give rise to tumors when transferred to a new host. Determination of tumor cell antigen levels by ELISA indicated that the tumour antigens had virtually disappeared by 24 h after injection of tumor cells and P. acnes. In contrast, in control animals injected with tumor cells only, there was an initial drop in tumor antigen levels at 12 h, after which the level rose steadily and tumors developed in 7-10 days. Since the cellular exudate at 12 h was almost entirely composed of polymorphonuclear leukocytes (PMN), we tested the ability of PMN, stimulated by phagocytosis of 48-h P. acnes cells, to produce substances toxic to tumor cells. Results indicated that the supernatant fluid from a phagocytosis mixture of PMN and P. acnes contained material toxic to tumor cells and also to Chinese hamster ovary cells. Tests with scavengers and inhibitors of oxygen-derived radicals suggested that the toxic material is either hydrogen peroxide (H2O2) or hydroxyl radicals (OH). Suspensions of 12-h P. acnes, P. acnes cells walls, P. freudenreichii, or latex beads were ineffective in preventing tumor growth, and induced little toxicity when phagocytosed. We conclude that in this test system 48-h P. acnes prevents tumor growth by stimulating the production of toxic oxygen metabolites during phagocytosis by PMN.