Four methods were compared for the diagnosis of human taeniasis caused by Taenia solium. Fecal samples from persons living in a T. solium endemic region of Madagascar were examined for taeniid eggs by the Kato–Katz method. Subsequently, samples positive (n = 16) and negative (n = 200) for T. solium eggs were examined by (i) amplification of the fragment of small subunit of the mitochondrial ribosomal RNA (rrnS) gene using conventional polymerase chain reaction (PCR) and (ii) a nested PCR of a fragment of the T. solium Tso31 gene. Additionally, 12 egg-positive and all egg-negative samples were tested for coproantigen detection. A further 9 egg-positive fecal samples were examined using both PCRs. Of the 12 egg-positive samples tested by PCRs and coproantigen methods, 9 (75%) were positive by rrnS PCR, 3 (25%) using Tso31-nested PCR and 9 (75%) by coproantigen testing. None of the 200 egg-negative fecal samples was positive in either rrnS or Tso31-nested PCR. Twenty of the 25 egg-positive samples (80%) were positive in rrnS PCR, and DNA sequencing of PCR amplicons was obtained from 18 samples, all confirmed to be T. solium. Twelve of the 25 egg-positive samples (48%) were positive in the Tso31-nested PCR, all of which were also positive by rrnS PCR. It is suggested that species-specific diagnosis of T. solium taeniasis may be achieved by either coprological examination to detect eggs or coproantigen testing, followed by rrnS PCR and DNA sequencing to confirm the tapeworm species in egg-positive or coproantigen-positive samples.

Risk factors related to herd and farmer status, farm and pasture management, and environmental factors derived by satellite data were examined for their association with the prevalence of F. hepatica in sheep and goat farms in Thessaly, Greece. Twelve farms (16·2%) and 58 farms (78·4%) of 74 had evidence of infection using coproantigen and serology respectively. The average normalized difference vegetation index (NDVI) of farm location for 12 months before sampling was the most significant environmental risk factor for F. hepatica infection based on high seropositivity. The risk of infection increased by 1% when the value of NDVI increased by 0·01 degree. A geospatial map was constructed to show the relative risk (RR) of Fasciola infection in sheep and goat farms in Thessaly. In addition, geospatial maps of the model-based predicted RR for the presence of Fasciola infection in farms in Thessaly and the entire area of Greece were constructed from the developed model based on NDVI. In conclusion, this study demonstrated that Thessaly should be regarded as an endemic region for Fasciola infection and it represents the first prediction model of Fasciola infection in small ruminants in the Mediterranean basin.

Juvenile hedgehogs having insufficient body weight are often brought for overwintering to hedgehog rehabilitation centres. Faecal samples of juvenile hedgehogs and overwintering hedgehogs (n=188) collected prior to releasing them back into the wilderness were examined for the presence of Cryptosporidium coproantigen and oocysts. Altogether 56 (29·8%) submitted samples were positive for coproantigen. Forty-five (39·5%, n=114) of the positive samples originated from newly rescued hedgehogs, while 11 (14·8%, n=74) positive samples were from animals that spent several months at the station. Fifteen samples subjected to PCR-RFLP analysis on the partial 18S rRNA locus suggested the presence of C. parvum. Multilocus sequence typing on partial 60 kDa glycoprotein gene, 18S rRNA, actin gene, 70 kDa heat shock protein gene sequences revealed 3 different subtype families: IIa, IIc and a new, proposed as VIIa subtype family. Cryptosporidium sp. genotype belonging to VIIa subtype family is closely related to C. parvum but is genetically distinct being probably a hedgehog-specific Cryptosporidium sp. genotype with unknown zoonotical potential. Hedgehogs excreting Cryptosporidium oocysts represent a potential source for human infections, but also an anthroponotic nature of the IIc subtype family should be reviewed.

Three ELISA assays, based on hyperimmune rabbit serum raised against adult cestode somatic antigen, were applied in this study for the detection of Taenia- and Echinococcus-specific antigens in host faeces. The first assay, using an antiserum against Taenia pisiformis antigen extract, was used in a time-course of T. pisiformis experimental infection in dogs. The assay was shown to be considerably more sensitive than microscopical detection of eggs in faeces. Antigen was present in faeces before patency and antigen levels were independent of T. pisiformis egg output. The second assay, involving a test for human taeniasis based on antibodies against T. solium, was applied in two field studies carried out in China and Guatemala. The test was highly specific, no false positive reactions occurred with human faecal samples and the test was capable of diagnosing individuals who would not have been detected by coproscopy or treatment to recover the tapeworm. A third assay was designed for E. granulosus and demonstrated 87·5% sensitivity and 96·5% specificity with samples from naturally and experimentally infected dogs with Echinococcus or Taenia infections. In both the human Taenia and canine Echinococcus studies antigen could be detected in faecal samples from infected hosts stored in 5% formalin for 6 months. Further refinements to these tests for field application are discussed.

A dipstick dot ELISA for detection of Taenia-specific coproantigens was developed. The test was based on a sandwich ELISA using antibodies raised against adult Taenia solium. Antibodies were adsorbed to nitrocellulose paper previously adhered to acetate plastic to form dipsticks. Once blocked with 5% skimmed milk and dried the antibody-coated dipsticks were stable for several weeks at room temperature. Both micro and dot ELISA formats were genus specific although the dot ELISA was less sensitive than the micro ELISA based on the same antiserum. During field studies, in which the majority of samples were tested in rural villages soon after collection, 3728 samples were tested. All samples were also examined by microscopy using formol ether concentration and individuals questioned to determine whether they were aware of being infected. After the initial diagnostic work individuals were treated with taeniacidal drugs for worm recovery. Use of the coproantigen test significantly increased the number of cases diagnosed. Of the 41 cases diagnosed by the three diagnostic techniques combined 31 were detected by the dipstick assay making it the most sensitive technique employed. The specificity of the dipstick assay was 99·9% with a positive predictive value of 88·%. The combined diagnostic approach did not, however, diagnose all cases. The coproantigen test was fast and easy to use. Further improvements may make the dipstick test suitable for wide-scale use in field studies and diagnostic laboratories.

Immunodiagnostic tests for Taenia-specific faecal antigen based on polyclonal rabbit antisera against Taenia saginata or Taenia solium proglottid extracts in capture-type ELISA assays have been developed. Taenia-specific antigen was detected in detergent-solubilized faecal extracts from T. solium- and T. saginata-infected hosts. Coproantigen from T. solium-infected hamsters did not cross-react with faeces from rodents infected with Hymenolepis diminuta, H. citelli, H. micro-stoma, Necator americanus, Strongyloides ratti or Nematospiroides dubius and faeces from uninfected animals. When the T. saginata-capture ELISA was tested with faecal samples positive for T. solium antigen, no cross-reactions were obtained. However, faecal samples from humans infected with T. solium or T. saginata, including some with extremely low egg counts, were cross-reactive by either test. Nevertheless, considerably higher O.D. values were obtained with stool samples from Taenia patients compared to Hymenolepis nana-infected or uninfected individuals. Two individuals, infected with Taenia sp. and positive for coproantigens by ELISA, became antigen-negative 6 days after treatment with Niclosamide. The possibility of developing species-specific immunodiagnostic tests for human taeniasis through coproantigen detection is discussed.

Dogs infected with adult tapeworms of Echinococcus granulosus release antigens (coproantigens) in faeces which can be detected by a capture ELISA. Supernatants prepared from E. granulosus-infected dog faecal samples were fractionated by size-exclusion fast protein liquid chromatography (FPLC) on a Superose-6 column. Coproantigen ELISA and Western blotting were used to demonstrate the immunoreactivity of eluted fractions. Two main FPLC peaks of antigenic activity were detected and designated as fraction F1 and fraction F2 with approximate relative molecular weights >670 kDa, and in the range of 146 to 440 kDa respectively. These two antigenic fractions (F1 and F2) fractionated from infected dog faeces were heat stable and largely protease-insensitive, but were highly sensitive to sodium periodate treatment, which strongly suggested the involvement of carbohydrates. Capture IgG antibodies against E. granulosus proglottis somatic extracts, detected a molecule with an approximate molecular weight of 155 kDa in fraction F2 after immunoblotting. The 155 kDa antigen could be completely ablated by sodium periodate treatment, but not after protease or lipase treatment. A surface tegument preparation of adult E. granulosus tapeworms contained large amounts of antigen that corresponded in size range and antigenicity to that observed in the FPLC fraction F2. There was also a peak of antigenic activity at >670 kDa corresponding to fraction F1 from a culture derived excretory–secretory (E–S) adult tapeworm preparation. The involvement of carbohydrate moieties in coproantigen activity present in the FPLC fractions F1 and F2 from faecal supernatants of E. granulosus-infected dogs was confirmed by lectin-binding assays and exoglycosidase treatment, which showed that α-D-mannose and/or α-D-glucose, β-galactose and N-acetyl-β-glucosamine residues were the most important carbohydrate components in putative coproantigens present in both fractions. N-acetyl-β-glucosamine and sialic acid residues were also contained in coproantigen molecules present in fraction F2. These results suggested that coproantigens detected in faeces of E. granulosus-infected dogs are large molecular weight molecules that may be derived from the carbohydrate-rich surface glycocalyx of adult worms, and are shed, released or secreted during the life-span of the tapeworm.

Two novel approaches for diagnosis of intestinal Echinococcus multilocularis infection, the detection of E. multilocularis-specific coproantigens in ELISA and of copro-DNA by PCR, have been successfully implemented. These methods have proven their value for the post mortem and the intra vitam diagnosis of E. multilocularis in definitive hosts. They have also made novel approaches possible to study the transmission biology of the parasite as they allow detection of the infection in faecal samples collected in the environment. Coproantigen detection is the diagnostic method of choice as it is sensitive, fast and cheap. Studies on faecal samples collected in the field revealed that coproantigen detection did reflect the different prevalences in fox populations as assessed from foxes at necropsy and also the effect of deworming efforts in foxes as achieved by long-term distribution of praziquantel-containing baits. The use of PCR for routine diagnostic or large-scale purposes is hampered by the fact that DNA extraction from faecal material is a very laborious task. Therefore, PCR is rationally used for confirmatory purposes of copro-antigen-positive samples. As taeniid eggs cannot further be differentiated morphologically, PCR is the method of choice to identify E. multilocularis infections in faecal or environmental samples containing taeniid eggs. In intermediate rodent hosts, PCR is routinely used in epidemiological studies for identifying E. multilocularis from liver lesions which are often very small, atypical or calcified.

The study was conducted to evaluate a mAb-based coproantigen detection assay for monitoring fox prevalence of Echinococcus multilocularis infections in the Shiretoko National Park in Hokkaido, Japan. Eight fox families, each consisting of 2–4 adults and their pups, were selected because their territories have been determined. Faeces of each family (total n=537) were collected monthly from April to October, 1994. Detection of coproantigen and taeniid eggs was recognized in 58 and 27 faeces, respectively, but mostly restricted in 1 family, in which coproantigen ELISA OD values had dual peaks, one in June and the other in October, whereas taeniid eggs were detected only from April to July. Fox pup faeces (n=51) collected around the dens used by the positive family were all coproantigen and taeniid egg negative except for 1 faeces. These results suggest that the prevalence and infection pressure in the study area were low and the infected family terminated the infection in the summer and acquired a new one in the early autumn, 1994. This monitoring method for fox infection will be a practical management tool for E. multilocularis infection, especially at the tourist spots in Hokkaido where close contact of fox with humans is frequently observed.