Methods Used for Detection and Recovery of Escherichia coli O157:H7 Associated with a Food-Borne Disease Outbreak †.

The current Food Safety and Inspection Service method for detection and recovery of Escherichia coli O157:H7, (including modified EC broth with novobiocin (mEC+n) and a direct blot ELISA). was used to analyze beef and environmental samples during an investigation of a food-borne disease outbreak attributed to consumption of undercooked hamburger patties. Double-modified trypticase soy broth (dmTSB) and a commercially available dipstick immunoassay were also used to improve detection/recovery of E. coli O157:H7. A total of 1,115 beef and environmental samples was screened with the direct blot ELISA and the dipstick immunoassay; 178 presumptive-positive samples (by either or both of the screening methods) were subjected to recovery/isolation procedures. Toxigenic E. coli O157:H7 was recovered from 45 samples: 40 hamburger-patty samples produced on the epidemiologically identified date, 3 hamburger-patty samples produced on another date, and 2 beef briskets. The organism was not recovered from environmental samples. Limited quantitative analyses indicated that contaminated hamburger patties contained fewer than 4.3 CFU of E. coli O157:H7 per g. Atypical, toxigenic ornithine decarboxylase-negative E. coli O157:H7 and nontoxigenic sorbitol-positive E. coli O157:H29 were also recovered. Both enrichment broths gave strong positive reactions with the two immunoassay screening methods, but E. coli O157:H7 was recovered more often from mEC+n broth than from dmTSB. Both screening methods gave positive results for 44 of the 45 beef samples found to contain E. coli O157:H7. False-positive results were frequently observed with both screening methods.

Presence of Yersinia enterocolitica Serotype O:5,27 in Slaughter Pigs.

The tonsils, tongue, mesenteric lymph nodes, cecal contents, and feces from 50 slaughter pigs were evaluated for the presence of pathogenic Yersinia enterocolitica . The cecal contents and feces of two pigs were positive (4%) with the pathogenic serotype O:5,27, biotype 3, whereas all other pigs were negative for pathogenic serotypes of Y. enterocolitica . The pathogenic serotypes were isolated from the cecal contents and feces using the stomacher sampling method. The cold enrichment in phosphate buffered saline and plating in Cefsulodin-irgasan-novobiocin medium. No pathogenic serotypes were recovered using the swab sampling technique, enrichment in Irgasan-ticarcillin-chlorate, and plating in modified Salmonella-Shigella deoxycholate-calcium chloride medium. The isolation of only 4% positive pathogenic Y. enterocolitica in our sampling of pigs in one production unit provides some encouragement that detection and control procedures might be effectively implemented to reduce or eliminate serotype O:5,27 from that herd.

Effect of Freezing on Infectivity of Toxoplasma Gondii Tissue Cysts in Pork.

To study the effect of low temperatures on infectivity of Toxoplasma gondii tissue cysts, pork from infected pigs was mixed with infected mouse brains and homogenized thoroughly. Twenty-gram samples of infected homogenized meat which were sealed in plastic pouches, pressed to a uniform thickness resulting in samples having the dimensions of ~.2 × 16 × 18 cm, were subjected to temperatures of -1 to -171.1 °C for 1 s to 67.2 d. Treated samples were digested in HCl-pepsin solution and infectivity assayed in mice. A regression model from these data is described by the least squares linear regression: Square root of time for the inactivation of T. gondii (h) = 26.72 + 2.16 temperature (°C) with r = 0.77. T. gondii tissue cysts remained viable usually up to 22.4 d at -1 and -3.9°C and 11.2 d at -6.7°C but were usually rendered nonviable by freezing at -12.37°C. These data demonstrate that T. gondii tissue cysts are inactivated by freezing more readily than encysted Trichinella spiralis larvae.

Effect of high temperature on infectivity of Toxoplasma gondii tissue cysts in pork.

To study the effect of high temperature on infectivity of Toxoplasma gondii tissue cysts, pork from infected pigs was mixed with infected mouse brains and homogenized thoroughly. Twenty-gram samples of infected homogenized meat were sealed in plastic pouches, pressed to a uniform thickness of 2 mm, and subjected to water-bath temperatures of 49, 52, 55, 58, 61, 64, and 67 C for 0.01, 3, 6, 9, 12, 24, 48, and 96 min. Treated samples were digested in HCl-pepsin solution and bioassayed in mice. Toxoplasma gondii tissue cysts remained viable at 52 C for 9.5 min but not for 9.5 min at 58 C; tissue cysts were generally rendered nonviable by heating to 61 C or higher temperature for 3.6 min. Tissue cysts survived once at 64 C for 3 min. These data demonstrate that T. gondii tissue cysts are less heat resistant than encysted Trichinella spiralis larvae.

Infectivity of Trichinella spiralis from Frozen Pork.

This research was carried out to determine the time/temperature exposure of Trichinella spiralis to freezing conditions necessary to destroy the infectivity of the trichinae. Experimentally infected pork was subjected to temperatures of -1 to -193°C for one sec to 182 d and the treated pork samples, which contained about 1000 larvae per gram of tissue, were subjected to rat bioassay to determine infectivity of the larvae. A linear regression equation, log10t = 5.98 + 0.40T where t = required inactivation time in hours and T = temperature in degrees Celsius, described the exposure necessary to destroy the trichinae. The correlation for that relationship was r = 0.942. The predicted thermal death times (+7 min) at -20, -15, and -10°C were 8 min, 64 min, and 4.0 d, respectively. The predicted upper confidence limits (99%) for the thermal death times (+7 min) for exposure at -20, -15, and - 10°C, were 48 min, 63 h, and 266 d, respectively. These data provide a continuum of definitive times and temperatures necessary to destroy T. spiralis by freezing and are of value to the meat industry and the regulatory agencies.