Antibody Banding Patterns on the Enzyme-Linked Immunoelectrotransfer Blot (EITB) Assay Clearly Discriminate Viable Cysticercosis in Naturally Infected Pigs.

Enzyme-linked immunoelectrotransfer blot (EITB) detects antibodies against seven Taenia solium larvae antigens in three protein families (GP50, T24/42, and 8-kDa) with different structures and functions. EITB banding patterns against these antigens in pigs provide information about the course of infection and may discriminate viable cysticercosis. We analyzed the banding patterns and infection outcomes (presence of viable cysts, degenerated cysts, and any cysts) of 512 rural pigs. Banding patterns were grouped into homogenous classes using latent class analysis, and relationships with infection outcomes were assessed. Four classes were identified: 1 (n = 308, EITB-negative or positive for the GP50 family), 2 (n = 127, positive for GP50 (GP50 family), GP42-39 and GP24 (T24/42 family), but negative for 8-kDa antigens), 3 (n = 22, positive for GP50 and T24/42 antigens (GP42-39 and GP24), as well as to 8-kDa bands GP13, GP14, and GP18, but negative for GP21), and 4 (n = 55, positive for GP50 and T24/42 antigens, as well as to 8-kDa antigens GP21 and GP18 in combination). Pigs in classes 3 and 4 were more likely to have viable cysts (72.6% and 96.4%, respectively) than pigs in classes 1 and 2 (0.7% and 27.6%, respectively; p < 0.001). The number of infections with any cysts was higher in classes 3 and 4 (77.3% and 98.2%, respectively) and lower in classes 2 and 1 (34.7% and 4.9%, respectively; p < 0.001). Pigs with viable cysts represented >90% of pigs with any cysts in classes 3 and 4 (94.1% and 98.2%, respectively), while degenerated cysts were frequent among pigs with any cysts in classes 1, 3, and 2 (86.7%, 47.1%, and 43.2%, respectively; p < 0.001). EITB banding patterns strongly correlate with cysticercosis infection status in rural pigs, with classes 3 and 4 being more predictive of viable infections.

Validity of the Enzyme-linked Immunoelectrotransfer Blot (EITB) for naturally acquired porcine cysticercosis.

The Enzyme-linked Immunoelectrotransfer Blot (EITB) has been used widely as a screening test for Taenia solium cysticercosis in swine. However, the relation between seropositivity and infection in pig populations from endemic areas has not been well defined. The aim of this study is to relate EITB seropositivity with infection and infection burden, analyse the trade-off between sensitivity and specificity with various cut-off points for the EITB assay, and finally describe the serology changes in a cohort of rural pigs raised under natural conditions. A group of 107 pigs that were used as controls during a vaccination field trial in Peru was our study population. The prevalence of porcine cysticercosis determined by necropsy examination was 16.82% (18/107) in these animals. Using EITB reactivity to ≥ 1 band as a cut-off point for the assay, the sensitivity was 88.89% (65.29-98.62, 95% CI) and the specificity was 48.31% (37.59-59.16, 95% CI). Comparing other cut-off points, involving up to as many as 7 reactive bands, a reactivity of ≥ 3 bands provided the best trade-offs in sensitivity and specificity. Using this cut-off point for the assay, the sensitivity was 77.77% (52.36-93.59, 95% CI) and the specificity was 76.40% (66.22-84.76, 95% CI). A significant association was found between cyst counts over 100 cysts and reactivity to ≥ 3 bands in the EITB assay (Fisher's exact test, p<0.05). The results of this study suggest that the use of the EITB assay to study porcine cysticercosis may require setting different cut-offs under field and experimental conditions, and depending upon the objective of the screening process.

Characterisation of antibody responses in pigs induced by recombinant oncosphere antigens from Taenia solium.

Recombinant antigens cloned from the oncosphere life cycle stage of the cestode parasite Taenia solium (T. solium) have been proven to be effective as vaccines for protecting pigs against infections with T. solium. Previous studies have defined three different host protective oncosphere antigens, TSOL18, TSOL16 and TSOL45. In this study, we evaluated the potential for combining the antigens TSOL16 and TSOL18 as a practical vaccine. Firstly, in a laboratory trial, we compared the immunogenicity of the combined antigens (TSOL16/18) versus the immunogenicity of the antigens separately. Secondly, in a field trial, we tested the ability of the TSOL16/18 vaccine to induce detectable antibody responses in animals living under environmental stress and traditionally reared in areas where T. solium cysticercosis is endemic; and finally, we characterised the immune response of the study population. Pigs of 8-16 weeks of age were vaccinated with 200 µg each of TSOL16 and TSOL18, plus 5mg of Quil-A. Specific total IgG, IgG(1) and IgG(2) antibody responses induced by TSOL16 and TSOL18 were determined with ELISA. The immunogenicity of both antigens was retained in the combined TSOL16/18 vaccine. The combined vaccine TSOL16/18 induced detectable specific anti-TSOL18 antibody responses in 100% (113/113) and specific anti-TSOL16 in 99% (112/113) of the vaccinated animals measured at 2 weeks following the booster vaccination. From the two IgG antibody subtypes analysed we found there was stronger response to IgG(2).

Seroprevalence and risk factors for Taenia solium cysticercosis in rural pigs of northern Peru.

Taenia solium is a cestode parasite that causes cysticercosis in both humans and pigs. A serological survey was undertaken to assess the seroprevalence and risk factors associated with porcine cysticercosis in the rural district of Morropon, Peru. Pigs aged between 2 and 60 months were assessed by the Enzyme-linked Immunoelectrotransfer blot (EITB) assay to determine their serological status against porcine cysticercosis in a cross-sectional study. A total of 1,153 pigs were sampled. Porcine seroprevalence was 45.19% (42.31-48.06). The information about the animals and households was analyzed and risk factors associated with seroprevalence were determined by a multivariate logistic regression analysis. In the porcine population, the risk of being seropositive increased by 7% with every month of age (OR 1.07, 95% CI 1.05-1.09), and by 148% for pigs living in East Morropon (OR 2.48, 95% CI 1.82-3.37). Whereas, the presence of latrines in a household decreased the risk of being seropositive by 49% (OR 0.51; 95% CI 0.39-0.67). Sex and rearing system did not represent either risk or protective factors associated with the seroprevalence of porcine cysticercosis. The findings of this study could be used for further development of control programs that might focus on similar population groups within rural communities of developing countries where cysticercosis is endemic.

Successful immunization of naturally reared pigs against porcine cysticercosis with a recombinant oncosphere antigen vaccine.

Taenia solium causes cysticercosis in pigs and taeniasis and neurocysticercosis in humans. Oncosphere antigens have proven to be effective as vaccines to protect pigs against an experimental infection with T. solium. A pair-matched vaccination trial field, using a combination of two recombinant antigens, TSOL16 and TSOL18, was undertaken in rural villages of Peru to evaluate the efficacy of this vaccine under natural conditions. Pairs of pigs (n=137) comprising one vaccinated and one control animal, were allocated to local villagers. Animals received two vaccinations with 200 µg of each of TSOL16 and TSOL18, plus 5mg Quil-A. Necropsies were performed 7 months after the animals were distributed to the farmers. Vaccination reduced 99.7% and 99.9% (p<0.01) the total number of cysts and the number of viable cysts, respectively. Immunization with the TSOL16-TSOL18 vaccines has the potential to control T. solium transmission in areas where the disease is endemic, reducing the source for tapeworm infections in humans.

Protection of pigs against Taenia solium cysticercosis by immunization with novel recombinant antigens.

Recombinant antigens from the oncosphere stage of the parasite Taenia solium were expressed in Escherichia coli. The TSOL16, TSOL45-1A and TSOL45-1B recombinant antigens, each consisting of fibronectin type III (FnIII) domain S, were produced as fusion proteins with glutathione S-transferase (GST) and maltose binding protein (MBP). Groups of pigs were immunized twice with the GST fusions of the antigens and boosted a third time with the MBP fusions prior to receiving a challenge infection with T. solium eggs. The TSOL16 antigen was found to be capable of inducing high levels of immunity in pigs against a challenge infection with T. solium. Immunological investigations identified differences in immune responses in the pigs vaccinated with the various antigens. The results demonstrate that the TSOL16 antigen could be a valuable adjunct to current porcine vaccination approaches and may allow the further development of new vaccination strategies against T. solium cysticercosis.