Evaluation of the PrioCHECK™ Trichinella AAD kit to detect Trichinella spiralis, T. britovi, and T. pseudospiralis larvae in pork using the automated digestion method Trichomatic-35.

Trichinellosis is a potentially deadly parasitic zoonosis that is contracted by consuming undercooked infected meat. Reliable detection of infectious Trichinella spp. larvae in meat is therefore pivotal to ensure consumer's safety. The recently authorised PrioCHECK™ Trichinella Alternative Artificial Digestion (AAD) test kit appears promising when used with the standard magnetic stirrer method, but evaluation with other apparatus types is lacking. In this study, the performance of the AAD kit in an adapted Trichomatic-35 (TM35) instrument was evaluated, first, at the Swiss National Reference Laboratory for trichinellosis (NRL); second, in a ring trial involving four Swiss official laboratories. Proficiency pork samples spiked with larvae of Trichinella spiralis, T. britovi, or T. pseudospiralis were tested with the AAD kit and with the reference pepsin-HCl digestion method in TM35 instruments. At the NRL, both methods yielded identical qualitative and similar quantitative results independently of the Trichinella species. In the ring trial, satisfactory results were obtained for 47/50 (94.0%) (AAD) and 62/67 (92.5%) (reference method) of the analysed samples. Technical problems impairing analysis were more frequently observed with the AAD kit (n = 22) than with the reference method (n = 5) and were mainly (16/22) reported by one of the external labs. When no technical issues were recorded, the performance of both methods was comparable, in agreement with the observations at the NRL; however, these results suggest a need for further training with the kit and standardisation of the adapted TM35 instruments.

Detection of Sarcocystis spp. and Toxoplasma gondii in swine and detection of DNA of these protozoa in tissues and sausages / Soroprevalência de Sarcocystis spp. e Toxoplasma gondii em suínos e detecção do DNA desses protozoários em tecidos e embutidos

Abstract The seroprevalence of Sarcocystis spp. and Toxoplasma gondii was researched in swine raised in Santa Maria, RS, Brazil. Serum samples from 84 pigs from 31 farms were tested using indirect immunofluorescence assay (IFA) for both agents. Additionally, 53 samples of pork sausages and tissues destined for human consumption, including: salami, sausage, black pudding, heart, tongue, brain, and rib muscle, were submitted to PCR to detect DNA for each agent. The frequency of anti-Sarcocystis spp. antibodies was 36.9% (31/84), with titers ranging from 32 to 1024, and 25% (21/84) for anti-T. gondii antibodies, with titers ranging from 64 to 2048. Sarcocystis spp. and T. gondii DNA were detected in 67.9% (36/53) and 13.2% (7/53) of samples, respectively. The presence of antibodies and the detection of DNA from Sarcocystis spp., and T. gondii suggests that the pigs were infected and may serve as an important reservoir for both parasites. The infection by these protozoa in the swine population is relevant to public health due to their zoonotic potential.

Resumo A soroprevalência de Sarcocystis spp. e Toxoplasma gondii foi pesquisada em suínos criados em Santa Maria, RS, Brasil. Amostras de soro de 84 suínos de 31 fazendas foram testadas pela reação deimunofluorescência indireta (IFA) para ambos os agentes. Adicionalmente, 53 amostras de embutidos suínos e tecidos cárneos destinados ao consumo humano, incluindo: salame, linguiça, morcela, coração, língua, cérebro e músculo da costela foram submetidas à PCR para detecção de DNA para cada agente. A frequência de anticorpos anti-Sarcocystis spp. foi de 36,9% (31/84), com títulos variando de 32 a 1.024; e 25% (21/84) para anticorpos anti-T. gondii, com títulos variando de 64 a 2048. A presença de DNA de Sarcocystis spp. e T. gondii foi detectada em 67,9% (36/53) e 13,2% (7/53) das amostras avaliadas, respectivamente. A detecção de anticorpos e DNA de Sarcocystis spp. e T. gondii sugere que os suínos foram infectados e podem servir como um importante reservatório de ambos os parasitas. A circulação desses agentes na população suína é relevante para a saúde pública devido ao seu potencial zoonótico.

Prevalence of Alaria alata mesocercariae in wild boars from Brandenburg, Germany.

Since 2002, Alaria (A.) alata mesocercariae (AM) have been found during routine Trichinella inspection of wild boars in many European countries. To date, human infection with AM through consumption of undercooked or raw AM infested wild boar meat cannot be excluded. In Germany, data on the parasite's prevalence in wild boars are scarce. To better understand temporal and spatial fluctuations of this parasite, this study investigated the prevalence of AM in wild boars in the German federal state of Brandenburg during three hunting seasons from 2017 to 2020. In total, 28.3% (100/354, 95% CI: 23.3-33.3%) of all wild boars sampled in eight counties of Brandenburg were tested positive for AM by Alaria alata mesocercariae migration technique (AMT). AM were detected in wild boars from seven different counties. Samples from one county (Havelland) tested completely negative for AM (0/16). Prevalences of the seven AM positive counties of Brandenburg ranged from 11.5 (3/26, 95% CI: 2.5-30.1%) in Märkisch-Oderland to 64.1% (25/39, 95% CI: 47.2-78.8%) in Uckermark. An association between sex and A. alata positivity could not be determined. A statistically significant increase in frequency of older AM positive wild boars was observed (p = 0.001). For a nationwide assessment of the prevalence of A. alata in wild boars and the risk for consumers of ingesting viable AM by consumption of raw or undercooked AM infested wild boar meat, further long-term studies in different regions of Germany are needed.

The wild boar (Sus scrofa, Linnaeus, 1758) as an important reservoir host for Alaria alata in the Baltic region and potential risk of infection in humans.

Alaria alata trematode is widely distributed throughout Europe and has a complex life cycle in which wild boar serve as a reservoir host. The primary aims of the present study are to establish the occurrence of A. alata mesocercariae in naturally infected wild boar in Latvia and to assess the risk for humans to acquire A. alata infection via consumption of wild boar meat. By summarizing long-term data using the Trichinella inspection method from 2014 to 2019, the overall A. alata prevalence was 8.3%, of which significantly higher A. alata prevalence was observed during the summer seasons. Additionally, 43.9% (n = 485) of wild boar were found to be infected with A. alata using Alaria mesocercariae migration technique. The present study indicates that the probability for humans to acquire A. alata mesocercariae is possible, yet improbable and varies from 0.2% to 2.2%. Most likely, it depends on both frequency of A. alata presence in wild boar population and of a method of preparing wild boar meat for consumption that will allow for the parasite inactivation.

All about Toxoplasma gondii infections in pigs: 2009-2020.

Toxoplasma gondii infections are common in humans and animals worldwide. Toxoplasma gondii infection in pigs continues to be of public health concern. Pigs are important for the economy of many countries, particularly, USA, China, and European countries. Among the many food animals, pigs are considered the most important for T. gondii transmission in USA and China because viable parasites have rarely been isolated from beef or indoor raised chickens. Besides public health issues, T. gondii causes outbreaks of clinical toxoplasmosis in pigs in China, associated with a unique genotype of T. gondii (ToxoDB genotype #9 or Chinese 1), rarely found in other countries. The safety of ready to eat pork products with respect to T. gondii infection is a matter of recent debate. Here, we review in detail seroprevalence, prevalence of viable and nonviable T. gondii, epidemiology, risk assessment, diagnosis, and curing of pork products containing T. gondii for the past decade. This review will be of interest to biologists, parasitologists, veterinarians, and public health workers.

The effect of temperature and time on the viability of Taenia solium metacestodes in pork.

OBJECTIVE: Taenia solium taeniosis is a growing health problem in large parts of the world including Sub-Saharan Africa. Humans are infected by eating undercooked pork with T. solium metacestodes, which cause taeniosis. The aim of the present study was to assess the effect of increasing temperatures on T. solium metacestode viability in pork. METHODS: Heavily T. solium infected pieces of pork were cooked in a water-bath at five different temperatures (40 °C, 50 °C, 60 °C, 70 °C and 80 °C) for 10-60 min. At each temperature/time point, five 5x5x5 cm pieces of pork fitted with thermometers at the core were placed in the water-bath. Controls were kept at 5 °C throughout exposure. After exposure, approximately 100 intact metacestodes were harvested and after a maximum of 6 h incubation at 37 °C in a culture media consisting of 50% porcine bile and 50% saline, the metacestodes were evaluated for viability. RESULTS: The metacestodes were fully viable after cooking at a core temperature of 40 °C for up to an hour. The metacestodes were non-viable after cooking for >40 min at a core temperature over 50 °C. All metacestodes were dead after cooking for 30 min at a core temperature of 60 °C; at 70 °C, non-viability was found after 20 min and all metacestodes were dead after 10 min cooking at 80 °C. CONCLUSION: Findings showed that pork pieces cooked at >80 °C for >10 min proved safe for human consumption. This means that dishes consisting of pork pieces in sizes not greater than a 5 cm cube which are immersed in continuously boiling water for at least 10 min would be safe to eat. However, pork deep-fried in oil may entail a risk due to generally shorter cooking time. More research on the cooking practises is needed in order to produce safe guidelines for risk-free pork consumption.

Use of meat juice and blood serum with a miniaturised protein microarray assay to develop a multi-parameter IgG screening test with high sample throughput potential for slaughtering pigs.

BACKGROUND: Serological screening of pig herds at the abattoir is considered a potential tool to improve meat inspection procedures and herd health management. Therefore, we previously reported the feasibility of a miniaturised protein microarray as a new serological IgG screening test for zoonotic agents and production diseases in pigs. The present study investigates whether the protein microarray-based assay is applicable for high sample throughput using either blood serum or meat juice. MATERIAL AND METHODS: Microarrays with 12 different antigens were produced by Abbott (formerly Alere Technologies GmbH) Jena, Germany in a previously offered 'ArrayTube' platform and in an 'ArrayStrip' platform for large-scale use. A test protocol for the use of meat juice on both microarray platforms was developed. Agreement between serum and meat juice was analysed with 88 paired samples from three German abattoirs. Serum was diluted 1:50 and meat juice 1:2. ELISA results for all tested antigens from a preceding study were used as reference test to perform Receiver Operating Characteristic analysis for both test specimens on both microarray platforms. RESULTS: High area under curve values (AUC > 0.7) were calculated for the analysis of T. gondii (0.87), Y. enterocolitica (0.97), Mycoplasma hyopneumoniae (0.84) and Actinobacillus pleuropneumoniae (0.71) with serum as the test specimen and for T. gondii (0.99), Y. enterocolitica (0.94), PRRSV (0.88), A. pleuropneumoniae (0.78) and Salmonella spp. (0.72) with meat juice as the test specimen on the ArrayStrip platform. Cohens kappa values of 0.92 for T. gondii and 0.82 for Y. enterocolitica were obtained for the comparison between serum and meat juice. When applying the new method in two further laboratories, kappa values between 0.63 and 0.94 were achieved between the laboratories for these two pathogens. CONCLUSION: Further development of a miniaturised pig-specific IgG protein microarray assay showed that meat juice can be used on microarray platforms. Two out of twelve tested antigens (T. gondii, Y. enterocolitica) showed high test accuracy on the ArrayTube and the ArrayStrip platform with both sample materials.

Consumer knowledge and practices to pork safety in two Taenia solium cysticercosis endemic districts in Eastern Cape Province of South Africa.

BACKGROUND: Globally, Taenia solium can cause cysticercosis in humans (including neurocysticercosis) and in pigs through ingestion of eggs and taeniasis in humans through ingestion of raw/undercooked pork contaminated with mature cysts. It is now recognised globally as one of the most prevalent food-borne parasitic diseases. The majority of cases have been reported in developing countries where consumption of food produced under unhygienic conditions is prevalent, exacerbated by lack of food safety education. The aim of this study was to determine the knowledge and practices of consumers towards pork safety in two districts of the Eastern Cape Province of South Africa, where T. solium cysticercosis is endemic in pigs and humans. METHODS: Three-hundred-and-sixty-one (361) participants were conveniently interviewed on consumer knowledge (harmfulness of T. solium cysticercosis, ability to identify cysts, trustworthiness of registered butcheries and legal requirements) and practices (storage of pork and method(s) of cooking pork safely) through a structured questionnaire. Chi-square for association of variables was used to compare differences in the districts. RESULTS: Overall, 73.1% of the study group from both districts agreed that pork forms an important part of their diet. Consumers (54.2%: 189/349) agreed that pork infected with T. solium cysts could be harmful, and 57.3% (188/328) indicated their inability to identify T. solium cysts in pork when slaughtered at home. Although 69.5% (234/352) trusted pork bought from butcheries, only 52.2% (187/358) were aware that butcheries must present a registration certificate in order to operate. This coincides with the fact that very few (< 10%) were aware of the legal requirements in terms of disease control, slaughter and food preparation. Most consumers (88.7%: 268/302) kept pork in the fridge and only 11.3% (34/302) kept it in a freezer (p = 0.02). Although not significantly different between the districts (p = 0.15), consumers in Alfred Nzo (71.4%: 152/213) and OR Tambo (61.2%: 74/12) mostly cooked pork as a stew, followed by braai/barbeque and frying or baking. This was in line with the fact that consumers in Alfred Nzo (79%: 147/186) and OR Tambo (80.8%: 120) preferred well-cooked pork; the main reason for this was the belief that cooking kills germs (43.6%: 121/277) followed by rendering the meat tasty (26.4%: 73/277). CONCLUSIONS: Consumers surveyed in the two districts were somewhat aware that T. solium cysticercosis could be harmful, although some were not able to identify T. solium cysts in pork. They also lacked sufficient knowledge regarding butchery certification and other legal requirements related to disease control, slaughter and food preparation. Practices related to cooking have the potential to promote the transmission of human taeniasis and the fact that most respondents preferred stewed pork could be a positive sign, as the cysts are destroyed during the cooking process. Results from this study are useful for the development of a control and prevention strategy targeted towards consumers, and the creation of awareness of food safety, with special emphasis on T, solium cysticercosis.

High anti-Ascaris seroprevalence in fattening pigs in Sichuan, China, calls for improved management strategies.

BACKGROUND: Ascariasis, caused by Ascaris suum, is an important soil-transmitted parasitic disease of pigs worldwide. It leads to significant economic losses in the pork industry, as a consequence of low feed conversion efficiency in pigs and liver condemnation at slaughter. Despite ascariasis still being widespread on pig farms in many developing and the industrialised countries, there are surprisingly limited data on porcine ascariasis in China, where nearly half of the world's total pork is produced. METHODS: In the present study, using the recently developed A. suum-haemoglobin (As-Hb) antigen-based serological test, we screened 512 individual serum samples from fattening pigs from 13 farms across seven distinct locations of Sichuan Province in China for anti-Ascaris antibody. RESULTS: The prevalence of anti-Ascaris antibody ranged from 0% to 100% on the distinct farms, with the mean (overall) seroprevalence being > 60%. There was no significant difference in seroprevalence between the intensive and extensive farms. CONCLUSIONS: To our knowledge, this is the first study to measure anti-Ascaris seroprevalence in China. The results of this 'snapshot' investigation indicate that Ascaris infection in commercial pig farms in Sichuan Province is seriously underestimated, encouraging future, large-scale serological studies to assess the distribution and extent of Ascaris exposure and infection in various regions of China and the world.

Trichinella spiralis Infecting Wild Boars in Southern Chile: Evidence of an Underrated Risk.

Trichinellosis in Chilean wild animals has scarcely been documented. The introduction of wild boars into the wild environment represents a viable new host with a potential risk of infection for human health. The aim of this study was to determine the presence and prevalence of Trichinella in wild boars. Two hundred seventy eight wild boars from of the Southern Chile were examined by compression and artificial digestion techniques. The larvae in the positive samples were collected for taxonomic analysis through polymerase chain reaction-inter-simple sequence repeats and to calculate the parasitic burden. A prevalence of 1.8% (5/278) of infected animals and an average parasitic burden of 6.8 ± 2.1 larvae per gram were estimated. The only species identified by molecular techniques was Trichinella spiralis. Prevalence of T. spiralis in wild boars was similar to those described around the world. T. spiralis infection rate and parasite burden detected in Chilean wild boars represent a certain food-borne risk for human population.