Molecular identification of zoonotic hookworm species in dog faeces from Tanzania.

The presence and distribution of various species of canine hookworms in Africa are poorly known. The main objective of this study, therefore, was to identify the hookworm species present in canine faecal samples from Morogoro, Tanzania, using molecular techniques. Faecal samples from 160 local dogs were collected and hookworm positive samples processed to recover larvae for further molecular characterization. DNA was extracted from pools of larvae from individual samples (n = 66), which were analysed subsequently using two different molecular approaches, polymerase chain reaction-linked restriction fragment length polymorphism (PCR-RFLP) and species-specific PCR coupled with Sanger sequencing. The PCR-RFLP technique detected only the presence of the ubiquitous Ancylostoma caninum in the 66 samples. However, by species-specific PCR coupled with Sanger sequencing we identified ten samples with A. braziliense, two with Uncinaria stenocephala and five with A. ceylanicum. Thus, all four known species of canine hookworms were identified in Morogoro, Tanzania. To our knowledge this is the first report of the detection of the presence of U. stenocephala and A. ceylanicum in Africa using molecular techniques. In addition to their veterinary importance, canine hookworms have zoonotic potential and are of public health concern.

Piloting the effectiveness of pig health education in combination with oxfendazole treatment on prevention and/or control of porcine cysticercosis, gastrointestinal parasites, African swine fever and ectoparasites in Angónia District, Mozambique.

A community-based intervention combining health education (HE) and treatment of pigs for control of porcine cysticercosis (PC), gastrointestinal (GI) helminths, African swine fever (ASF) and external parasites was tested involving six villages of resource-poor smallholder pig farmers. Farmers and pigs of six rural villages were randomly allocated into group 1 (HE), which served as controls, and group 2 (HE + OFZ) pigs received a single oral dose of 30 mg/kg OFZ. Farmers were trained in pig health, housing and feeding. The proportion of farmers with confined pigs, the adoption rate of the introduced pig pen, the sero-prevalence of PC and ASF, the prevalence and intensity of GI nematodes and the prevalence of ectoparasites were measured at 9, 15 and 24 months after initiation and compared to a baseline survey to seek the effectiveness of the interventions. There was no clear effect of the intervention on the sero-prevalence of PC, but analysis of the rate of change in prevalence between the two groups showed significant effect with the rate of change to lower prevalence in the HE + OFZ group compared to the HE group. Although HE managed to improve the farmer's knowledge in the control and prevention of ASF and ectoparasites, there was no significant reduction in the sero-prevalence of ASF and the prevalence of ectoparasites throughout the two-year period. The reported ineffectiveness of the intervention in this study suggested that more research is needed to develop more effective methods for controlling PC, ASF and pig parasites.

Distribution and histopathological changes induced by cysts of Taenia solium in the brain of pigs from Tanzania.

Neurocysticercosis (NCC) caused by Taenia solium cysts is a frequent but neglected parasitic disease of the central nervous system (CNS) worldwide. The aim of this study was to describe anatomical locations of cysts in the CNS and the corresponding inflammation. A total of 17 naturally infected pigs were used to evaluate the distribution of cysts and, of these, seven were used to evaluate the corresponding inflammation further, through histopathology. Clinical signs in the pigs included dullness, sluggishness, somnolence, apathy and loss of consciousness. Cysts were distributed in all cerebral lobes, i.e. 39.7% in the frontal lobe, 20.3% in the parietal lobe, 20.0% in the occipital lobe and 19.7% in the temporal lobe, and only 0.4% in the cerebellum. No cysts were found in the spinal cord. Cysts were localized as follows: 47.9% in the dorsal subarachnoid, 46.9% in the parenchyma, 4.4% in the subarachnoid base and 0.9% in the ventricles. The results of the histopathology revealed lesions in an early inflammatory stage, i.e. stage I, in all anatomical locations except for two, which showed more of an inflammatory reaction, stage III, in one pig. It was concluded that clinical signs in pigs were neither pathognomonic nor consistent. These signs, therefore, cannot be used as a reliable indicator of porcine NCC. Furthermore, T. solium cysts were found to be in abundance in all cerebral lobes, and only a few were found in the cerebellum. Regarding the inflammatory response, no significant differences were found in the location and total number of cysts. Thus, further studies are needed to explain the determinants of cyst distribution in the CNS and assess in detail clinical signs associated with porcine NCC.

Distribution and histopathological changes induced by cysts of Taenia solium in the brain of pigs from Tanzania.

Neurocysticercosis (NCC) caused by Taenia solium cysts is a frequent but neglected parasitic disease of the central nervous system (CNS) worldwide. The aim of this study was to describe anatomical locations of cysts in the CNS and the corresponding inflammation. A total of 17 naturally infected pigs were used to evaluate the distribution of cysts and, of these, seven were used to evaluate the corresponding inflammation further, through histopathology. Clinical signs in the pigs included dullness, sluggishness, somnolence, apathy and loss of consciousness. Cysts were distributed in all cerebral lobes, i.e. 39.7% in the frontal lobe, 20.3% in the parietal lobe, 20.0% in the occipital lobe and 19.7% in the temporal lobe, and only 0.4% in the cerebellum. No cysts were found in the spinal cord. Cysts were localized as follows: 47.9% in the dorsal subarachnoid, 46.9% in the parenchyma, 4.4% in the subarachnoid base and 0.9% in the ventricles. The results of the histopathology revealed lesions in an early inflammatory stage, i.e. stage I, in all anatomical locations except for two, which showed more of an inflammatory reaction, stage III, in one pig. It was concluded that clinical signs in pigs were neither pathognomonic nor consistent. These signs, therefore, cannot be used as a reliable indicator of porcine NCC. Furthermore, T. solium cysts were found to be in abundance in all cerebral lobes, and only a few were found in the cerebellum. Regarding the inflammatory response, no significant differences were found in the location and total number of cysts. Thus, further studies are needed to explain the determinants of cyst distribution in the CNS and assess in detail clinical signs associated with porcine NCC.

The immune response in Taenia solium neurocysticercosis in pigs is associated with astrogliosis, axonal degeneration and altered blood-brain barrier permeability.

Immunohistochemistry was used to examine the immuno-pathological changes and the extent of neuronal damage caused by either viable or dead Taenia solium cysticerci during porcine neurocysticercosis. Thirty pig brains with cerebral cysticercosis and 5 brains from T. solium free pigs were used in this study. Results revealed extensive astrogliosis, neuronal and mostly axonal damage in both early (grade I) and late (grades III and V) lesions as evidenced by an increased expression of glial fibrillary acidic protein (GFAP) and neurofilament protein (NFP). In many late lesions, astrocyte end-feet formed glial scars that surrounded the dead parasite. Rapid angiogenesis resulted in blood vessels lacking astrocyte end-feet suggesting loss of blood-brain barrier (BBB) hence allowing an influx of peripheral blood immune cells such as eosinophils, macrophages, CD3+ T cells, B lymphocytes and plasma cells into lesions. This study showed that porcine NCC was associated with severe nervous tissue damage, the host response of which is a collaborative effort between the local and peripheral immune responses comparable to that observed in human NCC. Results further implied that porcine NCC could be a useful model for understanding the course of NCC in human as well as provide useful information for therapeutic and/or immune strategies.

Persistent immune responses in late infection and after treatment in experimental Schistosoma bovis infections in goats.

This study explored host immune responses and their possible relationship to the anti-fecundity phenomenon in Schistosoma bovis-infected goats. The design comprised a primary infection with or without treatment at week (wk) 13, and with or without challenge at wk 36. Necropsy was performed at 36 or 52wk. Serum levels of anti-egg IgG, and anti-worm IgG and IgM, were measured by ELISA. In chronic infection, anti-worm antibodies stayed high, reflecting persisting worm burdens, whereas anti-egg IgG remained high despite minimized egg excretion. After treatment, anti-worm IgM and anti-egg IgG were minimized, but anti-worm IgG remained above the values of the uninfected controls. Histopathology showed lowered numbers of perioval granulomas in chronic infection and resolution of liver fibrosis with time, but intestinal lymphoplasmacytic perivasculitis and hepatic eosinophilic infiltrates were maintained at wk 52. Significant splenic plasmacytosis persisted after treatment. The results indicated that persistent immune responses, in chronically infected and in treated goats, may explain sustained worm fecundity depression at challenge infection.

Real-time PCR for detection of low intensity Schistosoma japonicum infections in a pig model.

Decades of successful Schistosoma japonicum control have increased the interest in how to diagnose low intensity infections. A real-time PCR assay targeting the mitochondrial NADH dehydrogenase I gene in S. japonicum was evaluated in infected pigs with very low egg output. Six out of 12 S. japonicum infected pigs were treated with praziquantel 8 weeks after infection and all pigs were followed for 16 weeks post-infection. One commercial and one non-commercial extraction method were evaluated in combination with PCR on faecal samples. PCR with either extraction method were equally sensitive as the DBL-filtration/sedimentation technique in the acute, productive stage. PCR recovered slightly more positive samples in the chronic stage, but most faecal samples were negative for both PCR and microscopy from week 9 post-infection irrespective of treatment. IgG antibody titers against soluble egg antigen IgG remained high throughout the study in both the treated and non-treated group. PCR was consistently negative in serum and urine samples and negative in most of the caecal biopsies. We conclude that the S. japonicum faecal PCR is a highly sensitive test. However, in clinical samples when faecal egg output almost reaches nil in the chronic stage despite persistent worm burdens, both the faecal PCR and microscopy results were negative. Real-time PCR is less labour intensive than most microscopy methods, but has a higher material cost per sample.

Host-cell apoptosis in Taenia solium-induced brain granulomas in naturally infected pigs.

To assess whether apoptosis occurs in pig brain granulomas due to Taenia solium cysticerci, brain tissues from 30 pigs naturally infected with T. solium cysticercosis were evaluated by terminal deoxynucleotidyl transferase-end labelling (TUNEL) staining. In addition, tissues were stained with CD3 marker to identify T lymphocytes. Examination of TUNEL-stained tissues showed apoptotic cells in early lesions that contained viable cysticerci. Apoptotic cells were primarily found interspersed with normal cell types, and were mostly located in the inflammatory infiltrate. Late or advanced granulomas with disintegrated scolices did not show TUNEL-positive cells. CD3+ cells were found in both early and advanced lesions and apoptosis mainly co-localized with CD3+ T lymphocytes. This suggests that these cells are constantly undergoing apoptosis and thus die as soon as they arrive at the site of infection. Apoptosis indeed may be one way by which T. solium cysticerci down-regulate the host's cellular immune response in early cysticercosis. Therefore, further research is needed to establish if other cells besides T-lymphocytes are also a target for destruction by cysticerci in early cysticercosis as well as studies to assess if cysteine protease is expressed by viable cysticerci in situ.

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.

Field testing of the WHO dose pole for administration of praziquantel in the treatment of opisthorchiasis in Lao PDR.

In 2001, the WHO developed a dose pole that employs height measurements for estimation of the dose of praziquantel. In the present study, conducted in December 2005 during a mass treatment campaign for the control of opisthorchiasis in 232 individuals in Nala village, Keo Udom district, Lao PDR, performance of the dose pole in estimating dosages of praziquantel was compared with a bathroom scale; a digital scale was used as the gold standard. Results showed that the bathroom scale performed significantly better than the dose pole in delivering dosages of 40-50 mg/kg for opisthorchiasis treatment (70.7% vs. 44.8%). Furthermore, the dose pole performed significantly better for children than adults. The reason for the poor performance of the dose pole among adults is likely to be due to the high percentage (19.4%) of overweight individuals in the adult population of the village. It was concluded that the WHO dose pole is not recommended for distribution of praziquantel for the treatment of opisthorchiasis in populations where being overweight is common.

Hepatic changes in congenital Schistosoma japonicum infections in pigs.

The inflammatory response in liver tissue from piglets congenitally infected with Schistosoma japonicum was examined at two different timepoints after infection. The piglets, which were the offspring of three sows infected with 9000 S. japonicum cercariae in the 10th week of gestation, were allocated into two groups (n=9 and 17) killed 5 or 11 weeks after birth, respectively. All piglets developed a low level infection,with no significant difference between the groups. Inflammatory lesions in the liver consisted mainly of granulomas in portal areas, often obliterating the portal veins, and frequently with central eggs or egg remnants. The granulomatous reaction consisted of epithelioid cells and occasional giant cells surrounded by layers of lymphocytes, eosinophils, plasma cells, and various amounts of collagen and fibroblasts. Mild to moderate infiltration of portal and septal connective tissue with eosinophils and lymphocytes was common, but the connective tissue was generally not increased. At the two timepoints, slight differences were observed in the numbers of eosinophils and lymphocytes in the granulomas and in the size of the granulomatous reaction. The same pattern of immunohistochemical labelling was seen in both groups. CD79alpha(+) B cells were scarce except in granuloma-associated lymphoid follicles;the majority of lymphocytes in granulomas and at other sites were CD3epsilon(+) T cells. The granulomatous reaction in the livers of piglets to schistosoma eggs from prenatal S. japonicum infection was similar to that seen in postnatal infection. Signs of immunomodulation of granulomas between the two timepoints of infection were not demonstrable.

Control of Taenia solium taeniasis/cysticercosis: The best way forward for sub-Saharan Africa?

Taenia solium taeniasis/cysticercosis is a neglected parasitic zoonosis with significant economic and public health impacts. Control measures can be broadly grouped into community health education, improvements in hygiene and sanitary conditions, proper meat handling at household and community level, improved standards of meat inspection, pig management, treatment of individual patients and possibly human populations, and treatment and/or vaccination of porcine populations. This manuscript looks critically into currently existing control options and provides suggestions on which (combination of) tools would be most effective in the control of T. solium taeniasis/cysticercosis in sub-Saharan Africa. Field data and disease transmission simulations suggest that implementation of a single intervention control strategy will not lead to a satisfactory reduction of disease morbidity or transmission. A feasible strategy to combat T. solium taeniasis/cysticercosis would include a combination of approaches focussing on both human (health education and treatment) and animal host (management, treatment and vaccination), which can vary for different communities and different geographical locations. Selection of the specific strategy depends on cost-effectiveness analyses based on solid field data, currently unavailable, though urgently needed; as well as on health priorities and resources of the country. A One Health approach involving medical, veterinary, environmental and social sectors is essential for T. solium to be controlled and eventually eliminated. Finally the success of any intervention is largely dependent on the level of societal and political acceptance, commitment and engagement.

Does multiple hosts mean multiple parasites? Population genetic structure of Schistosoma japonicum between definitive host species.

Multi-host parasites, those capable of infecting more than one species of host, are responsible for the majority of all zoonotic, emerging or persistent human and animal diseases and are considered one of the major challenges for the biomedical sciences in the 21st century. We characterized the population structure of the multi-host parasite Schistosoma japonicum in relation to its definitive host species by genotyping miracidia collected from humans and domestic animals across five villages around the Yangtze River in Anhui Province, mainland China, using microsatellite markers. High levels of polymorphisms were observed and two main genetic clusters were identified which separated water buffalo, cattle and humans from goats, pigs, dogs and cats. We thereby believe that we present the first evidence of definitive host-based genetic variation in Schistosoma japonicum which has important epidemiological, evolutionary, medical and veterinary implications.

Evaluation of PCR based coprodiagnosis of human opisthorchiasis.

In this study, a recently developed PCR test for the detection of Opisthorchis viverrini in human faecal samples was evaluated using two parasitological methods as references. During a survey of foodborne trematodes (FBT) in the Vientiane Province, Lao PDR, 85 samples were collected and evaluated for FBT eggs by the Kato Katz (KK) technique, the formalin ethyl acetate concentration technique (FECT) and a PCR analysis for the distinction between O. viverrini and other FBT. The two parasitological methods did not differ in the ability of detecting FBT eggs, and a single KK reading was characterized by a sensitivity of 85% when compared to two FECT readings. The PCR tested positive only in cases where eggs had been demonstrated by parasitological examination. However, the PCR tested negative in some samples with very high egg counts. Demonstrating a PCR sensitivity of approximately 50% in samples with faecal egg counts>1000, the previously reported PCR sensitivity based on in vitro studies was not supported. It is believed that technical problems rather than diagnostic reference related issues were responsible for the relatively low PCR performance. Further studies should aim at optimizing DNA extraction and amplification, and future PCR evaluation should include specificity control such as the scanning electron microscopy of eggs in test samples or the expulsion of adult trematodes from PCR tested individuals.

World association for the advancement of veterinary parasitology (WAAVP): second edition of guidelines for evaluating the efficacy of anthelmintics in swine.

Guidelines are provided for evaluating the efficacy of anthelmintics in swine which, in conjunction with other sets of guidance such as those of the International Cooperation on Harmonization of Technical Requirements for Registration of Veterinary Medicinal Products (VICH GL7 and VICH GL16), should encourage the adoption of uniform registration requirements globally. Testing of efficacy should be carried out according to the principles of "Good Clinical Practice" (VICH GL9, 2000). Data obtained according to these guidelines should be internationally acceptable for the registration of anthelmintics for swine. Further, the use of the guidelines should expedite development, government review, and approval of anthelmintics for swine, as well as contribute towards reducing costs and the number of experimental animals used for drug testing.

Novel real-time PCr for detection of Schistosoma japonicum in stool.

Chemotherapy has been used on a large scale in countries where the blood fluke Schistosoma japonicum is endemic. This has led to a lower intensity of infections and consequently lower diagnostic values of commonly used diagnostic tests like serology and Kato-Katz stool smear. We designed a novel real-time PCR method for detection of S. japonicum in stool samples. Further, we evaluated different versions of an inexpensive, non-commercial extraction method, ROSE, as well as the commercial QIAamp DNA Stool Mini Kit. PCR primer sequences were designed targeting the mitochondrial NADH dehydrogenase I gene. Bovine serum albumin was added to the DNA extracts and SYBR Green was used for detection. The PCR method was evaluated with non-infected stool samples spiked with S. japonicum eggs. It demonstrated high sensitivity, even in samples containing a single egg. The two extraction methods were equally effective. The PCR was specific for S. japonicum when tested against other Schistosoma species, Trichuris trichiura, hookworm and Taenia sp. We conclude that this novel real-time PCR, in combination with either ROSE or QIAamp DNA Stool Mini Kit extraction, is a sensitive and specific tool for diagnosing S. japonicum in human stool samples.

Estimating sensitivity and specificity of a faecal examination method for Schistosoma japonicum infection in cats, dogs, water buffaloes, pigs, and rats in Western Samar and Sorsogon Provinces, The Philippines.

Schistosoma japonicum causes a chronic parasitic disease, which persists as a major public health concern in The Philippines, the People's Republic of China and Indonesia. This infection is unique among helminthic zoonoses because it can infect humans and more than 40 other mammals. The objective of this study was to estimate the sensitivity and specificity of the Danish Bilharziasis Laboratory technique in cats, dogs, pigs, water buffaloes and rats in the Philippines. Faecal samples from each animal were collected on up to five occasions on five consecutive days in four villages of Sorsogon and Western Samar Provinces between January and July 2003. The faecal samples were analysed with the filtration and sedimentation Danish Bilharziasis Laboratory technique. Sensitivity and specificity of one, two, three, four, and five faecal samples were estimated using a Bayesian latent class approach. A total of 59, 43, 74, and 80% of the censored cats, dogs, pigs, and water buffaloes in the four villages were sampled, respectively. For all species, the sensitivity estimates when using the results of only 1 day of sampling were less than 80%. However, the sensitivity improved to at least 96% in all species when three or more faecal samples were collected on three separate days. The specificity was estimated to be above 92% across all species, even if just a single sample is used. The prevalences and 95% credible intervals of S. japonicum, adjusted for imperfect sensitivity and specificity, in cats, dogs, pigs, rats, and water buffaloes were 11.9% (6.8-18.3%), 19.9% (15.1-25.2%), 2.9% (1.1-5.2%), 31.3% (18.3-45.6%) and 6.3% (2.1-12.6%), respectively. Our results suggest that the Danish Bilharziasis Laboratory technique is valid for the detection of infection with S. japonicum in animals, and that sensitivity estimates are excellent when faecal samples are collected on at least three different days. Monitoring S. japonicum infection in animal reservoirs with a valid test could contribute to more effective public health control programmes.

Detection of African swine fever virus DNA in blood samples stored on FTA cards from asymptomatic pigs in Mbeya region, Tanzania.

The aim of the study was to assess whether blood samples collected onto FTA(®) cards could be used in combination with real-time PCR for the detection of African swine fever virus (ASFV) DNA in samples from resource-poor settings under the assumption that asymptomatically (sub-clinically) infected pigs may be present. Blood samples were collected from clinically healthy pigs from Mbeya Region, Tanzania. The blood samples were stored on FTA(®) cards and analysed by real-time PCR assays in duplicate; three pigs had high levels of viral DNA (Ct values of 27-29), and three pigs had a low level of viral DNA (Ct 36-45). Four pigs were positive in one of the duplicate samples only, but clear products of the expected size were obtained when the reactions were analysed by gel electrophoresis. For comparison, blood samples from pigs experimentally infected with either a pathogenic (OURT T88/1) or a non-pathogenic (OURT T88/3) isolate of ASFV were collected, stored on FTA(®) cards and analysed in the same way. The blood from pigs infected with the OURT T88/1 isolate showed high levels of viral DNA (Ct 22-33), whereas infection with non-pathogenic OURT T88/3 isolate resulted in only low levels of viral DNA (Ct 39) in samples collected at 10-14 days after inoculation.

Human migration and pig/pork import in the European Union: What are the implications for Taenia solium infections?

Taenia solium taeniasis/cysticercosis is a neglected zoonotic disease complex occurring primarily in developing countries. Though claimed eradicated from the European Union (EU), an increasing number of human neurocysticercosis cases is being detected. Risk factors such as human migration and movement of pigs/pork, as well as the increasing trend in pig rearing with outside access are discussed in this review. The entry of a tapeworm carrier into the EU seems a lot more plausible than the import of infected pork. The establishment of local transmission in the EU is presently very unlikely. However, considering the potential changes in risk factors, such as the increasing trend in pig farming with outdoor access, the increasing human migration from endemic areas into the EU, this situation might change, warranting the establishment of an early warning system, which should include disease notification of taeniasis/cysticercosis both in human and animal hosts. As currently human-to-human transmission is the highest risk, prevention strategies should focus on the early detection and treatment of tapeworm carriers, and should be designed in a concerted way, across the EU and across the different sectors.

Schistosoma japonicum: interactions of successive infections in pigs and mice using polymerase chain reaction-based identification of individual worms.

The study reported here investigated acquired resistance of mice and pigs to challenge-infections with Schistosoma japonicum. Two morphologically indistinguishable isolates of the parasite (from the Anhui and Zhejiang provinces of China), which could be typed by polymerase chain reaction-linked restriction fragment length polymorphism analysis (PCR-RFLP), were used for the infections. In two parallel infection studies, 60 female outbred NMRI mice and 29 Danish Landrace/Yorkshire/Duroc crossbred pigs were used. Two of the groups received a primary infection with either the Anhui or the Zhejiang isolate, respectively. The remaining groups received a primary infection with the Zhejiang isolate and challenge-infections with the Anhui isolate at either week 2, 3, 4 or 6 post primary infection. The results of the study indicated that both mice and pigs are partially resistant to challenge-infection from week 4 post primary infection. Resistance appeared to decrease in pigs 6 weeks after primary infection, while it remained effective in mice. These results suggest that the mechanism responsible for acquired resistance in mice and pigs may not be the same and support the theory that worm burdens in pigs receiving repeated infection are in a balance between acquisition and loss of worms.