Low salt exposure results in inactivation of Toxoplasma gondii bradyzoites during formulation of dry cured ready-to-eat pork sausage.

The production of safe and healthy food products represents one of the main objectives of the food industry. The presence of microorganisms in meat and products containing meat can result in a range of human health problems, as well as economic losses to producers of these products. However, contaminated meat products continue to initiate serious and large-scale outbreaks of disease in consumers. In addition to outbreaks of diseases caused by bacteria and viruses, parasitic organisms, such as Toxoplasma gondii, are responsible for foodborne infections worldwide, and in the case of T. gondii, is considered the 2nd leading cause of death from foodborne illness in the U.S. Transmission of Toxoplasma gondii has historically been linked to the consumption of raw or undercooked meat products, including pork. Specific concerns with respect to pork products are ready-to-eat (RTE) pork meals. These are pork or products containing pork that are prepared by curing or drying, and are not intended to be cooked before being consumed. Previous studies have demonstrated that T. gondii is inactivated during dry cured sausage preparation, apparently in the batter during fermentation. In this study, we have analyzed timing of inactivation of T. gondii in freshly prepared pepperoni batter to confirm our previous findings, to determine how quickly inactivation occurs during fermentation, and to confirm what parameters of the sausage preparation are involved in inactivation of the parasite. Results from the current and previous study indicate that rapid inactivation of T. gondii bradyzoites occurs in low salt batter for dry cured sausage within 4 h of initiation of fermentation.

Rapid inactivation of Toxoplasma gondii bradyzoites during formulation of dry cured ready-to-eat pork sausage.

Curing processes for pork meat in the U.S. currently require individual validation of methods to demonstrate inactivation of Trichinella spiralis, a nematode parasite historically associated with pork. However, for protozoan parasites, no such strictures exist. It has been assumed, with little evidence, that curing processes required to inactivate Trichinella also inactivate Toxoplasma gondii. Currently no model of meat chemistry exists that can be correlated with inactivation of T. gondii. Given the possibility of the presence of T. gondii in pork meat, and the frequent use of pork for ready-to-eat (RTE) products not intended to be cooked, curing methods which inactivate T. gondii early in the curing process would be of great value to producers. In this study, we tested the effect of five variables - salt/brine concentration, water activity (aw), pH, temperature, and time on inactivation of T. gondii bradyzoites in tissue cysts using low and high endpoints for common curing treatments during preparation of dry cured pork sausage. Survival of T. gondii bradyzoites at each stage of preparation was assessed using a mouse bioassay. Results indicated that encysted T. gondii bradyzoites do not survive the early stages of the dry curing process within the endpoint parameters tested here, even at levels of NaCl that are lower than typically used for dry curing (1.3%). Exposure of T. gondii encysted bradyzoites to curing components in the formulated batter resulted in rapid inactivation of bradyzoites. These data suggest that the use of dry curing components may be effective for controlling T. gondii potentially transmitted through RTE meats, rendering them safe from risk with respect to T. gondii transmission to human consumers.

Curing conditions to inactivate Trichinella spiralis muscle larvae in ready-to-eat pork sausage.

Curing processes are one method by which pork products, which are considered ready to eat (RTE) and have not been otherwise tested or treated, can be rendered safe from risk for exposure to Trichinella muscle larvae (ML). Curing processes in the U.S. currently require individual validation of methods to demonstrate inactivation of Trichinella. This is a major undertaking for each process; currently no model of meat chemistry exists that can be correlated with inactivation of Trichinella. Given the potential for new RTE products (e.g., lower salt), the availability of a wider range of tested methods for inactivation of Trichinella in pork would be of substantial value to the industry. In this study, five variables were tested - salt/brine concentration, water activity (aw), pH, temperature, and time, using low and high endpoints for common curing treatments for dry cured pork sausage. The data demonstrated that NaCl concentrations above 1.3%, in combination with fermentation to pH 5.2 or below, resulted in inactivation of > 96% of Trichinella ML in stuffed sausages within 24-28 h. All ML were inactivated by 7-10 days post-stuffing. These curing processes reliably predict inactivation of Trichinella spiralis, and can be used within the defined upper and lower endpoint parameters to reduce or eliminate the need for individual product validation.

Cessation of Trichinella spiralis transmission among scavenging mammals after the removal of infected pigs from a poorly managed farm: implications for trichinae transmission in the US.

Pigs infected with the zoonotic parasite Trichinella spiralis were detected on a farm in Maryland during an animal welfare investigation. Sera and/or tissues were collected from 49 pigs and three pig carcasses (7 weeks of age to adult, mixed sex). The tissues were tested for the presence of T. spiralis muscle larvae (ML) by tissue digestion, and the sera were tested for the presence of anti-Trichinella antibodies by ELISA. Seventeen of 50 (34%) pigs were infected with T. spiralis based on tissue digestion. Of these 17 pigs, sera were collected from 16; nine were serologically positive, three sera had OD values that were very close to the positive cut-off (0.30), but were still negative, and four were negative (suggesting that they had become infected within a few weeks of testing). All pigs that tested negative by tissue digestion for ML were also ELISA negative. The farm was subsequently depopulated of pigs. Six months later, testing of trapped scavenging mammals in the farm environment demonstrated that 41% were infected with T. spiralis. After 12 months, 10% of trapped animals were T. spiralis positive, and after 18 months, T. spiralis could not be detected in the scavenging mammal population surrounding the farm. Results of the study suggest that T. spiralis, typically transmitted in the peridomestic rat-pig-human cycle in the US, was not maintained in scavenging mammals in the absence of infected pigs.

Effects of time and temperature on the viability of Toxoplasma gondii tissue cysts in enhanced pork loin.

Enhanced or pumped pork products represent a significant proportion (40 to 50%) of the commercially available pork cuts available to consumers at the retail level. In a previous study, pork loins containing viable Toxoplasma gondii tissue cysts were pumped with solutions containing 2% sodium chloride or 1.4% or higher potassium or sodium lactate and stored at 4 degrees C for 7 days. This treatment prevented transmission of T. gondii to cats. In the present study, enhanced pork loins were stored for 0, 8, 16, 24, 32, or 40 h at 4 degrees C and then fed to T. gondii-seronegative cats to determine how quickly the loss of tissue cyst viability occurred. In a second experiment, pork loins collected from pigs experimentally infected with T. gondii were stored at temperatures found in retail meat cases and then fed to T. gondii-seronegative cats to determine the effect of typical meat case storage temperatures on tissue cyst viability. In both experiments, cat feces were examined for 14 days after the infected meat meal to assess oocyst shedding. The results indicate that solutions containing 2% sodium chloride or 1.4% potassium or sodium lactate are effective within 8 h of injection for killing T. gondii tissue cysts in pork loins and that storage at meat case temperatures at or below 0 degrees C (32 degrees F) for 7 days also killed T. gondii tissue cysts in pork loins.

Detection of Hammondia heydorni-like organisms and their differentiation from Neospora caninum using random-amplified polymorphic DNA-polymerase chain reaction.

Neospora caninum and Hammondia heydorni are morphologically and phylogenetically related coccidians that are found in dogs. New diagnostic genetic loci, based on random-amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR), were developed to aid in the detection of H. heydorni-like parasites and to discriminate them from N. caninum and other related coccidians of dogs. On the basis of the data obtained from 5 random decamers, H. heydorni (Manhattan-1) and N. caninum (NC1) were characterized by distinct banding patterns (similarity index = 0.068). High-stringency PCR assays were developed from the sequences of 2 cloned bands (GenBank BZ592549 and BZ592593), uniquely amplified from H. heydorni. Interestingly, using these primers, PCR amplification was achieved only from 2 of the 5 isolates presumed to represent H. heydorni. The same result was obtained from these 5 isolates using a recently described PCR assay directed to the H. heydorni internal transcribed spacer-1. It is concluded that H. heydorni and N. caninum are genetically distinct and that such tools may be useful for more detailed characterization of the diversity of related parasites occurring in dogs.