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Recent molecular identification of Paramphistomum epiclitum in Pakistan raises concerns about its epidemiology and pathologies in infected tissues of ruminants. The present study aimed to find the seroepidemiological and histopathological record of Paramphistomum epiclitum from cattle and buffaloes. Indirect ELISA on animal sera and histology of infected rumen with hematological and biochemical analyses were performed. The overall prevalence of P. epiclitum was noted as 15.3% in the abattoir survey and 37.6% in the serological examination. The sensitivity and specificity of the diagnostic test were 100% and 83.3% respectively. The paramphistomosis was significantly (p= 0.001) higher during August (6.4%) followed by September (5.4%), whereas the lowest prevalence was recorded during April (0.4%). The hematological and biochemical variations showed significant increase in total leukocyte count (p= 0.002), alanine aminotransferase (p= 0.05), glucose (p= 0.01) and cholesterol (p = 0.024) levels. However, significant decrease in the level of erythrocyte (p= 0.019), hemoglobin (p= 0.001), mean corpuscular haemoglobin concentration (p= 0.05), mean corpuscular volume (p= 0.038) and platelets count (p= 0.048) was observed. The histopathology of rumen tissue showed haemorrhages, atrophy of ruminal papillae, sloughed mucosa, cellular vacuolation, and infiltration of lymphocytes inflammatory cells. The present study provides the prevalence and histopathological record of P. epiclitum in Pakistan for the first time in order to take control measures in the country.

Fasciolosis, caused by liver fluke species of the genus Fasciola, are well recognized because of its high veterinary impact. Stool examination for Fasciola eggs is not a sensitive method, and limited efforts to find a reliable and cheaper means of detection are available. The present study aimed to develop rapid diagnostic ELISA test against fasciolosis. The excretory/secretory (ES) and somatic (SA) products of Fasciola helminths were analysed using polyacrylamide gel electrophoresis (PAGE). Immunogenicity was evaluated by immunoblotting using hyperimmune sera raised in rabbits and seroprevalence was determined by indirect ELISA.  The results of SA antigen of Fasciola species showed polypeptide bands ranged from 10kDa-100kDa, while ES antigen of Fasciola showed bands of 15kDa-55kDa. The immunoblotting results showed the most prominent bands against ES antibodies were 25, 35, 55-70, 100 and 250 kDa and SA antigens showed 10, 15-25, 35, 70, 100 and 250 kDa polypeptide bands. The sensitivity and specificity of developed indirect ELISA for SA antigens was 95.45% and 87.1%, while for ES antigens was 100% and 77.42% respectively. The overall seroprevalence recorded for fascioliasis based on SA antigen was 39.8% and 29.8% for ES antigen. The fasciolosis did not show significant association with host type, sex and age groups of examined animals, however significantly higher infection was found in months of September and October. The result provides sensitive in house immunodetection assay for diagnosis of fasciolosis alternative to commercial kits with high import cost.

Fascioliasis causes huge economic losses in livestock industry by reducing the growth, fertility rate, meat and milk yield. In the present study, somatic antigens isolated from Fasciola gigantica were assessed for the early detection of infection by developing antibody detection enzyme immunoassay. The somatic antigens extracted from adult F. gigantica worms was separated on SDS-PAGE and immunogenicity was determined by Western blot (WB). The results showed polypeptides bands between 15 to 95 kDa, and most prominent bands were 15, 28, 36, 38, 55, 72 and 95 kDa polypeptides. The WB revealed a single polypeptide of size between 36 to 55 kDa as antigenic. The sensitivity and specificity of ELISA test established for 36 to 55 kDa somatic antigens was 95.45% (95% CI: 77.16% to 99.88%) and 87.1% (95% CI: 70.17% to 96.37%), respectively. Kappa value revealed that the strength of agreement is considered to be very ‘good’. In house established ELISA was implemented in the field and 12.1% (31/256) large ruminants were found positive, while 4.68% (12/256) with postmortem examination. The fasciolosis did not show significant (P>0.05) association with host type, breed and age groups, while significant (P<0.01) association with host sexes. The result confirmed that in-house established ELISA test had good value for serodiagnosis of fasciolosis in cattle and buffaloes for large scale epidemiological studies.

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