The prevalence of porcine cysticercosis in Eastern and Southern provinces of Zambia

The objective of this study was to determine the prevalence and importance of porcine cysticercosis in rural areas of Zambia. The study involved an abattoir survey of 1316 pigs at a slaughter slab in Lusaka and two field surveys in villages in Southern and Eastern provinces. Lingual examination of live pigs and visual inspection of their carcass as well as blood sampling for measuring circulating parasite antigen by enzyme-linked immunosorbent assay (Ag-ELISA) were used as parameters to measure infection. In the field surveys, a questionnaire was administered to every household whose pigs were examined to obtain information on pig husbandry practices and to study risk factors for the infection. Out of the 1316 pigs examined at the slaughter slab, 143 (10.9%) and 271 (20.6%) were positive by lingual examination and meat inspection, respectively. Most of the pigs were very heavily infected with predominantly live cysts. The field surveys revealed that eight (8.2%) out of 98 pigs from Southern province and eight (5.2%) out of 151 pigs from Eastern province were positive for cysticercosis by tongue palpation. Using the Ag-ELISA 20 (20.8%) and 14 (9.3%) pigs were positive in Southern and Eastern provinces, respectively. The questionnaire survey revealed poor pig husbandry practices, absence of meat inspection and control, poor knowledge of the disease and poor sanitation in the surveyed villages. The prevalence of pig cysticercosis found in this study ranks among the highest in the southern African region, in Africa and in the world. The current study suggests the presence of human tapeworm carriers and a high risk of human cysticercosis in the surveyed areas as well as in urban centres where pigs from rural areas are increasingly sold, slaughtered and consumed.     

Risk factors associated with porcine cysticercosis in selected districts of Eastern and Southern provinces of Zambia

To determine the risk factors associated with Taenia solium transmission in humans and pigs in the rural areas of Eastern and Southern provinces of Zambia, a questionnaire was administered in 788 households from 155 villages. Pigs were examined from 800 households. Tongue examination and enzyme-linked immunosorbent assay (Ag-ELISA) for the detection of circulating antigens of T. solium cysticerci were used to measure infection in pigs. A snowballing technique was utilised to select households with pigs. Prevalence of households with pigs infected with T. solium on tongue examination by district ranged from 12.7% to 32.1% with Ag-ELISA having a range of 30.0-51.7%. Of the total number of households visited, 18.8% and 37.6% had at least one pig positive for porcine cysticercosis on tongue examination and Ag-ELISA, respectively. Risk factors associated with T. solium infection were lack of pork inspection at slaughter (96.7%), consumption of pork with cysts (20.1%), selling of pork infected with T. solium cysticerci (18.3%), free-range husbandry system (83.2%) and absence of latrines (58.0). Free-range husbandry system (OR=1.68; 95% CI=1.36-2.07) was a significant risk factor for porcine cysticercosis in the surveyed areas. The result that pigs were mostly kept on free-range and semi-intensive husbandry systems may have permitted them to have access to eating human faeces that could be contaminated with tapeworm eggs. This study has shown that T. solium infection poses a high public health risk in the study areas and urban areas as well. We recommend that a human survey be conducted to verify the human exposure to taeniasis and/or cysticercosis in Zambia.     

Lymphocyte apoptosis in the inflammatory reaction around Taenia solium metacestodes in porcine cysticercosis

In the current research, we report apoptosis of lymphocytes in the inflammatory reaction around metacestodes in muscle tissue from cysticercotic pigs. Two events, high metacestode viability (100%) and high cysteine protease activity were found to be closely related to a high phosphatydilserine expression by inflammatory lymphocytes (56%). Testing the RPMI medium used for washing away inflammatory cells from metacestodes with 100% viability, with the fluorescent substrate Z-Phe-Ala-AFC for measuring cysteine protease activity, significant fluorescent values were found. In contrast, tests performed with RPMI medium used for washing away inflammatory cells from metacestodes with 90% viability or less, showed low fluorescence values. Flow cytometry analyses of inflammatory cells obtained from four naturally cysticercotic pigs, and stained with Annexin-V/PI, showed lymphocytes expressing phosphatidylserine with values of 0, 6, 41 and 56% on their outer surfaces. Electron microscopy studies of inflammatory cells from metacestodes with 100% viability, showed lymphocytes with strangled and fragmented nuclei, and heterochromatin displaced to the nuclear periphery. In addition, DNA from these cells showed fragmentation in electrophoresis assays. Apoptosis of lymphocytes in the inflammatory reaction around Taenia solium metacestodes, might have been induced by the parasite cysteine protease, and may be involved in impairing cell-mediated immune responses in human and porcine cysticercosis.     

Prevalence of Taenia solium cysticercosis in swine from a community-based study in 21 villages of the Eastern Cape Province, South Africa

The pork tapeworm, Taenia solium, causative organism of porcine cysticercosis and human neurocysticercosis is known to occur in areas of South Africa including Eastern Cape Province but, despite increasing reports of its occurrence throughout the subregion, the prevalence is yet to be clearly established. The parasite presents a potentially serious agricultural problem and public health risk in endemic areas. The human populations considered to be at highest risk of infection with this zoonotic helminth are people living in rural areas most of whom earn their livelihood wholly or partially through livestock rearing. Here we report on initial results of a community-based study of pigs owned by resource-poor, emerging pig producers from 21 villages in the Eastern Cape Province. Lingual examination (tongue palpation) in live pigs, two enzyme-linked immunosorbent assays (ELISAs), which detect parasite antigen (B158/B60 Ag-ELISA and HP10 Ag-ELISA) and an enzyme immunotransfer blot (EITB) assay, which detects antiparasite antibody, were used to verify endemicity and estimate apparent prevalence. In the absence of a gold standard true prevalence was obtained, using a Bayesian approach, with a model that uses both available data and prior information. Results indicate that the parasite is indeed present in the study villages and that true prevalence was 64.6%. The apparent prevalences as measured by each of the four tests were: 11.9% for lingual examination, 54.8% for B158/B60 Ag-ELISA, 40.6% for HP10 Ag-ELISA and 33.3% for EITB. This base-line knowledge of the prevalence of T. solium in pigs provides information essential to the design and monitoring of sustainable and appropriate interventions for cysticercosis prevention and control.     

Preliminary evaluation of Community-Led Total Sanitation for the control of Taenia solium cysticercosis in Katete District of Zambia

Taenia solium taeniasis/cysticercosis is a zoonotic disease endemic in sub-Saharan Africa. It is associated with poor sanitary practices, free-range pig husbandry and lack of disease awareness in endemic communities. A comparative research was conducted with pre and post-intervention assessments in nine villages to evaluate Community-Led Total Sanitation (CLTS) as an intervention measure for the control of porcine cysticercosis in Katete District in the Eastern Province of Zambia. Blood samples were collected from pigs for circulating antigen detection and a questionnaire focused on the household was administered to a total of 153 respondents whose pigs were examined (64 pre-intervention, 89 post-intervention), in order to obtain information on general demographic characteristics, pig husbandry practices, sanitation practices and associated knowledge and awareness of T. solium infections. The first sampling was conducted prior to the implementation of the CLTS and second sampling eight months after triggering of CLTS in the selected villages. A total of 379 pig serum samples were examined using the B158/B60 Ag-ELISA to detect T. solium cysticercosis, 104 pre-intervention and 275 post-intervention, of which 14 (13.5%) and 45 (16.4%) were positive, respectively. Wald test p-values were computed to assess significant differences in the variables of interest mentioned above for the pre and post CLTS. The research revealed that CLTS as a control measure did not significantly improve T. solium infections in pigs. The research also revealed that the sanitation practices and awareness of cysticercosis did not change. It is recommended that a longer term evaluation be undertaken when the villages have been declared open defaecation free. In addition, the research recommends that health education, mass drug treatment and pig vaccination be incorporated, as an essential component of prevention and control programmes for T. solium infections.     

猪囊虫DNA疫苗pPCV的生物安全性

以猪囊虫DNA疫苗pPCV为研究对象,其生物安全性问题。将DNA疫苗pPCV以肌注方式免疫仔猪,分别于接种后1d、7d、4周、8周分离各组织,提取组织总DNA,利用PCR技术分析了质粒DNA在各组织的分布,以及与细胞基因组整合的可能性;同时采集环境样品,PCR法分析DNA疫苗上的CMV启动子基因、抗性基因和抗原基因在环境中发生转移和扩散的可能性。结果表明,在疫苗接种后第1天几乎所有组织都能检测到质粒DNA,随着时间的延续,仅在注射部位检测到质粒的存在,并且未发现质粒DNA整合入细胞基因组以及转化环境细菌。实验结果表明肌注DNA疫苗在注射部位以外组织中很快被清除,在注射部位组织及环境细菌中均未发现整合现象,因此认为该DNA疫苗对猪体和环境都是安全的。     

Persistence of passively transferred antibodies in porcine Taenia solium cysticercosis. Cysticercosis Working Group in Peru.

We evaluated the presence and persistence of anticysticercal antibodies in piglets born to Taenia solium infected sows. Infected sows from a disease-endemic area of Peru were transported to a nondisease-endemic area and impregnated. Serum samples were collected from sows and piglets on Day 2 through Week 35 after birth. Using an immunoblot specific for cysticercosis, Ig isotypes to 7 cyst antigens were measured and quantified. Serum samples from the piglets contained detectable antibodies from Week 1 through Week 35 (27 weeks after weaning). The primary Ig isotype present in both sows and piglets was IgG. Antibodies did not appear in piglet serum samples until after suckling, demonstrating that anti-cysticercal antibodies are transferred solely via colostrum. Our data have shown that maternally transferred antibodies to cyst antigens may persist through much of a pig's life. Therefore, the presence of passively transferred antibodies must be considered in studies that examine the prevalence of cysticercosis in pigs. Furthermore, when designing control strategies for cysticercosis, careful evaluation and selection of sentinel pigs becomes a crucial component of sentinel selection.     

PCR test for detecting Taenia solium cysticercosis in pig carcasses

Polymerase chain reaction (PCR) test was employed to detect Taenia solium DNA in muscle lesions for validation of the meat inspection results of slaughtered pigs. Two sets of oligonucleotide primers, one targeted against the large subunit rRNA gene (TBR primers) and the other targeted against cytochrome c oxidase subunit 1 gene (Cox1 primers) of T. solium were used in this study. On reactivity in PCR test, the TBR primers and the Cox1 primers yielded products of 286 and 984 bp, respectively, in cysticercosis positive cases. Both the sets of primers were found to be highly specific, since they did not yield any PCR product in negative controls. A total of 225 pig carcasses were screened for cysticercosis by meat inspection, out of which 25 carcasses with visible cysts (16 viable and 9 degenerated cysts) were also confirmed to be positive for cysticercosis in PCR test. However, out of the 35 carcasses with suspected lesions on meat inspection, only two were found to be positive for cysticercosis in PCR test. The detection limits for both the primer sets were analyzed. The TBR primer set could detect up to 10 pg of cysticercus DNA, whereas the Cox1 primer set could detect only up to 1 ng. It is evident from the study that PCR test is an efficient tool for validation of meat inspection results and also to rule out ambiguity in carcass judgment of suspected cases of porcine cysticercosis.     

Taenia solium cysticercosis: immunity in pigs induced by primary infection

The present study demonstrates that pigs experimentally infected with Taenia solium eggs develop resistance to reinfection that lasts at least five months. Thirteen 2-month-old piglets were infected with eggs of Taenia solium. After 5 months, two pigs were euthanized and five were challenged with eggs from a second tapeworm. Nine months after the first infection, six pigs were challenged with a third tapeworm. All 11 challenged pigs were euthanized 2 months after reinfection. In order to confirm the infectivity of the eggs, several piglets were inoculated with each taenia. Two of the five pigs reinfected after 5 months did not develop metacestodes, two showed few caseous non-infective forms and in the fifth pig, 14% of the metacestodes were vesicular and 86% colloidal and caseous. In the six animals challenged 9 months after the first infection, three were heavily infected with vesicular metacestodes and the other three showed only colloid and caseous forms in muscles. All parasites found in brains were vesicular. We conclude that immunity due to primary infection lasts at least 5 months. At 2 months of infection antigens of 24 and 39-42 kDa were the most frequently recognised. In those pigs with only a few caseous cysts in muscles and/or vesicular ones in brains no antibodies were detected.     

Taenia solium oncosphere antigens induce immunity in pigs against experimental cysticercosis

Immunity to Taenia solium infection was investigated using an experimental intramuscular oncosphere infection assay (IMOA) model in pigs. Three naturally infected pigs with cysticercosis were treated with oxfendazole (OFZ), a drug demonstrated to kill cysts in porcine muscle. These animals were then challenged with oncospheres but did not develop any cysts while three uninfected pigs that were similarly challenged, did develop intramuscular cysts. In another study, two groups of three pigs each were immunized with crude T. solium oncosphere and metacestode antigens, respectively, and tested with the IMOA. Immunization with crude oncosphere antigens (OAs) induced 100% protection, while metacestode antigens provided only partial protection. Immunoblots showed that pigs with complete immune protection to oncosphere intramuscular challenge had antibodies to two OAs at 31.3 and 22.5 kDa, respectively. Antibody to these two antigens was absent in pigs immunized with metacestodes or in uninfected control pigs. This study demonstrated the presence of two antigens that are unique to the oncosphere. Although, antibody to these two antigens is consistently present in pigs that are protected from an oncosphere intramuscular challenge their role in preventing infection by T. solium larval cysts is still hypothetical.     

Performance of the ELISA test for swine cysticercosis using antigens of Taenia solium and Taenia crassiceps cysticerci

Studies were conducted to evaluate antigens of Taenia solium (Tso) and Taenia crassiceps (Tcra) cysticerci in the ELISA test for the diagnosis of swine cysticercosis. The samples analyzed were cysticercosis positive and negative control sera and heterologous sera. Four antigens were assayed: vesicular fluid (VF) and crude (T) Tcra and scolex (S) and crude (T) Tso. All antigens showed good performance, but VF-Tcra was the best followed by T-Tcra. Sensitivity rates of ELISA were respectively, in 2nd and 3rd standard deviation cut-offs, 96.0 and 80.0% for the VF antigen and specificity of 97.5 and 100.0%. Cross-reactivity was verified only for hidatidosis and ascaridiosis. Due to the high performance observed, the ELISA test using Tcra antigens should be recommended for the diagnosis of swine cysticercosis.     

Taenia solium porcine cysticercosis: viability of cysticerci and persistency of antibodies and cysticercal antigens after treatment with oxfendazole

The aim of this study was to assess the effect of treating Taenia solium infected pigs with oxfendazole (OFZ) on viability and clearance of cysticerci and the corresponding persistence of specific antibody isotypes (IgG(total), IgG1, IgG2 and IgA) and circulating cysticercal antigen (CCA). Antibody isotypes and CCA responses were measured by antibody-ELISA (Ab-ELISA) and antigen ELISA (Ag-ELISA), respectively. Correlations were made between antibodies, CCA and the total number of cysticerci enumerated at necropsy. Forty pigs with cysticercosis were randomly allocated into two groups: Treatment group (n=20) was treated with OFZ at 30 mg/kg orally while the treatment control group (n=20) was not treated. Five uninfected pigs served as negative controls. Pigs were killed at 1, 4, 8 and 26 weeks post-treatment (wkpt). Overall, the mean total cyst count in treated pigs was 2904+/-5397 (mean+/-S.D.) while in the controls it was 6235+/-6705. Mean cyst viability was 5+/-11% (mean+/-S.D.) and 97+/-4% in treated and control pigs, respectively. Results showed that OFZ killed muscular cysticerci over a period of 4 weeks but failed to kill cerebral cysticerci. Antibodies, CCA responses and clearance of dead cysts from the meat, depended on the cyst intensity of individual pigs at time of treatment since both antibody and CCA correlated with intensity of cysticerci at necropsy (r=0.441, P=0.005; r=0.654, P<0.001), respectively. IgG1 responses were the best indicator of treatment efficacy because they were predominant in both infected treated and control pigs and disappeared early after treatment. Both Ab/Ag-ELISA failed to detect cysts in the brain. Though dead cysticerci took some time (26 wkpt) to clear from the meat, treatment of porcine cysticercosis with OFZ should, in combination with other intervention measures be considered as an important, cost-effective measure in the control of taeniosis/cysticercosis.     

Human and porcine Taenia solium infections in Mozambique: identifying research priorities

The objective of this paper is to critically review and summarize available scientific and lay literature, and ongoing studies on human and porcine cysticercosis in Mozambique to identify knowledge gaps and direct immediate and long-term research efforts. Data on the spatial distribution and prevalence of the disease in human and swine populations are scarce and fragmented. Human serological studies have shown that 15-21% of apparently healthy adults were positive for cysticercosis antibodies or antigen, while in neuropsychiatric patients seroprevalence was as high as 51%. Slaughterhouse records indicate a countrywide occurrence of porcine cysticercosis, while studies have shown that 10-35% of pigs tested were seropositive for cysticercosis antibodies or antigen. Current research in Mozambique includes studies on the epidemiology, molecular biology, diagnosis and control of the disease. Future research efforts should be directed at better understanding the epidemiology of the disease in Mozambique, particularly risk factors for its occurrence and spread in human and swine populations, documenting the socio-economic impact of the disease, identifying critical control points and evaluating the feasibility and epidemiological impact of control measures and development of local level diagnostic tools for use in humans and swine.     

Impact of naturally acquired Taenia solium cysticercosis on the hormonal levels of free ranging boars

In chronically infected BALBc/AnN male mice, Taenia crassiceps cysticercosis induces changes in the host's sex steroids hormone that lead to their estrogenization and deandrogenization, with possible repercussions on their susceptibility to infections. Here reported are the serum steroid levels in free range cysticercotic male boars. Therefore, the possible effects of Taenia solium cysticerci over the pig steroid levels were evaluated. Herein are described the sex steroids and cortisol levels of non-cysticercotic (n=25) and cysticercotic (n=22) adult boars, as diagnosed by tongue inspection, all free-ranging in a typical village of an endemic rural area in Mexico. A significant reduction of testosterone (P=0.022) and a likely one of 17beta-estradiol (P=0.08) levels were found in the cysticercotic boars in comparison with those non-cysticercotic, whilst no significant differences in the cortisol and DHEA levels were detected. Serum levels of specific antibodies did not correlate with infection nor with the levels of any of the hormones measured. Results suggest that T. solium cysticercosis significantly affects the hormonal status of its porcine host independently of their antibody response.     

Castration and pregnancy of rural pigs significantly increase the prevalence of naturally acquired Taenia solium cysticercosis

Cuentepec is a rural village of central Mexico, where 1300 pigs were bred at the time of the study in conditions that favor Taenia solium transmission. The tongues of 1087 (84%) of these pigs were visually examined and 33% were found to be cysticercotic. Castration of male pigs increased prevalence from 23 to 50% (P < 0.001) and pregnancy in sows also increased their prevalence from 28 to 59% (P < 0.001). Thus, endocrinological conditions characterized by low levels of androgens or high levels of female hormones probably influence the susceptibility of pigs to T. solium cysticercosis as observed in mice infected with Taenia crassiceps. Delaying castration of male pigs and confinement of sows during pregnancy might significantly decrease the prevalence of pig-cysticercosis and help curb transmission without much cost or difficulty.     

Experimental infection model for Taenia solium cysticercosis in swine. Cysticercosis Working Group in Peru

A novel method for infecting pigs with Taenia solium using an intramuscular innoculum of oncospheres was investigated in a series of five experiments in 18 animals. The model is simple to perform, requires a minimal number of oncospheres, permits multiple infections per animal, and decreases the variation inherent in oral infection models. This intramuscular oncosphere assay (IMOA) may provide a valuable tool to evaluate therapeutic agents or potential vaccines for cysticercosis.     

Comparative evaluation of an indirect ELISA test for diagnosis of swine cysticercosis employing antigen from Taenia solium and Taenia crassiceps metacestodes

Taenia solium cysticercosis is still a serious public health problem in several countries where poverty and lack of hygiene favor transmission. Because pigs are the primary intermediate hosts, prevalence of porcine cysticercosis is a reliable indicator of active transmission zones. Serological diagnostic methods are important tools for epidemiological studies since they can be applied to living animals on a large scale. Four antigen preparations (cyst fluid and crude) from T. solium and T. crassiceps metacestodes were compared for swine cysticercosis diagnosis by indirect ELISA (IE). Twenty-eight serum samples from swine naturally and experimentally infected by cysticerci of T. solium and 56 serum samples from swine reared in commercial herds were tested. Best results of overall sensitivity were obtained by the use of cyst fluid and crude antigen of T. crassiceps metacestode (100 and 96.4%, respectively). Using homologous antigen preparations we have observed higher specificity percentage (98.2% for cyst fluid and 96. 4% for crude metacestode T. solium antigen). We concluded that sensitivity is of far more importance than specificity for identification of endemic areas in order to prevent transmission to man. We conclude, therefore, that IE performed with cyst fluid antigen of T. crassiceps metacestode is a better tool for that purpose.     

Isolation of a 14 kDa antigen from Taenia solium cyst fluid by HPLC and its evaluation in enzyme linked immunosorbent assay for diagnosis of porcine cysticercosis

A fraction with a major band of 14kDa was obtained from crude cyst fluid of Taenia solium cysticerci by 2-step chromatography. A first fraction isolated by gel filtration (Sephacryl S-300 high resolution) was purified using an anion exchange column (Mono Q HR 5/5) on high performance liquid chromatography. Evaluation of the analytic sensitivity of this fraction (F3) was carried out in an antibody enzyme linked immunosorbent assay (Ab-ELISA-F3) using serum samples from pigs experimentally infected with different doses of T. solium eggs. The cross-reactivity of F3 was evaluated with serum samples from pigs that were naturally or experimentally infected with Taenia hydatigena, Taenia saginata asiatica, Fasciola hepatica, Trichinella spiralis, Metastrongylus apri, Trypanosoma congolense and Sarcoptes scabiei, and with serum samples of rabbits hyper-immunised with metacestode cyst fluid of T. hydatigena and T. solium. Antibody titres of lightly or heavily infected pigs differed in their kinetics. However, the increase in F3-specific antibodies could not be related to the infection level. Analysis of the specificity of the F3 showed that serum samples of pigs infected with other parasites did not recognise this antigen. Cross-reaction with T. hydatigena occurred in ELISA using cyst fluid as antigen, but the F3 antigen fraction was not recognized by rabbit hyper-immune serum samples to T. hydatigena. Evaluation of the diagnostic sensitivity and specificity of the Ab-ELISA-F3 was done by a non-parametric receiver operating characteristic (ROC) analysis using 66 serum samples from Zambian village pigs. The total number of cysticerci of these pigs was determined by dissection (28 pigs harboured T. solium cysticerci and 38 were negative at dissection). In addition, 58 serum samples from Cameroonian pigs (28 pigs from cysticercosis-free farms and 30 pigs with cysticerci at tongue inspection) were used in a separate ROC analysis. The results from the ROC analysis yielded a low diagnostic value (area under ROC curve=0.48) with the sera from the Zambian pigs while a relatively high diagnostic value was obtained with the sera from Cameroonian pigs (area under ROC curve=0.78). The main factor contributing to a low diagnostic value based on the Zambian serum samples seemed to be the false-positive reactions that were likely caused by the occurrence of transient antibodies in the non-infected animals.     

Fractionation and characterisation of the cysticercus of Taenia solium

Fractionation by chromatography on Sephadex G-200 of a saline extract of Cysticercus cellulosae scolex antigen yielded three distinct fractions associated with distinct peaks. These fractions were analysed by double immunodiffusion (DID) and immunoelectrophoresis (IEP). The three peaks gave five, four and three antigenic determinants, respectively, by DID with homologous hyperimmune rabbit serum. However, the same serum gave nine antigenic determinants of scolex antigen by DID and 11 components by IEP. The IEP demonstrated seven and five antigenic components in the first two peaks. The first peak gave a stronger reaction in indirect haemagglutination than the others. There were common antigenic components in C cellulosae and C tenuicollis antigens.