Development of an antibody-ELISA for seroprevalence of Trypanosoma evansi in equids of North and North-western regions of India
Introduction
Trypanosomosis (surra) caused by T. evansi is one of the important diseases of equines and transmitted mechanically by biting of blood sucking flies including tabanids (Tabanus, Haematopota, Haematobia). The rainy and post rainy seasons are most favourable for larval development of tabanid flies resulting in to propagation of the disease in these seasons. However, the cases of surra are come across throughout the year. Though T. evansi has wide distribution, there is no up-to-date information available on the prevalence and economic importance of equine trypanosomosis in India. Presently, T. evansi has been emerging in non-endemic areas and infecting new hosts including occasional cases in humans raising concern for zoonotic potential of this infection (Joshi et al., 2005, Powar et al., 2006).
Under field conditions, the disease is diagnosed on basis of clinical signs which are not sufficiently pathognomonic and generally confused with other chronic wasting diseases, notably helminthosis and malnutrition. Attention has recently been focused on the development of more sensitive and specific serological and DNA based tests and a number of tests have been described for the diagnosis of surra in domestic animals including ELISA (Rae and Luckins, 1992, Singh et al., 1995, Reid and Copeman, 2003, Singh et al., 2004, Desquesnes et al., 2009). During chronic and sub-clinical stages which is the most common form of the disease when host has no detectable parasite, the standard parasitological methods (examination of wet blood film, stained blood smear, haematocrit centrifugation) generally miss most of T. evansi infected cases due to poor sensitivity. Therefore, sensitive, specific diagnostic tests are required for quantifying disease prevalence in areas of concern, socio-economic impact assessment, vector studies, and for modelling of disease transmission and control strategies. The present study reports development of an antibody-ELISA and its application in assessing seroprevalence of T. evansi in equids of North and North-western regions of India.
Section snippets
The parasite
Trypanosoma evansi isolate was collected from a naturally infected camel and cryopreserved in liquid nitrogen after propagation in Swiss albino mice procured from Disease Free Small Animal House, Lala Lajpat Rai University of Animal and Veterinary Sciences, Hisar, India. The cryopreserved T. evansi stock (Camel/1/Bikaner/Rajasthan) was maintained in the Parasitology Laboratory of National Research Centre on Equines, Hisar, India for further use. For preparation of whole cell lysate antigen the
Confirmation of T. evansi infection in experimentally infected animals
All six experimentally infected donkeys became positive for T. evansi infection in microscopic examination of wet blood film by 7th dpi. Thereafter, parasites were observed intermittently in experimental donkeys by wet blood film examination at various intervals during the course of experiment. The infected donkeys exhibited clinical signs as intermittent fever, weakness, emaciation, anaemia, lacrimation and oedema in brisket and vulvar regions in initial stage of infection. These signs
Discussion
In the present study we have developed an antibody-ELISA using WCL antigen of T. evansi prepared from purified trypanosomes derived from infected mice blood at first parasitaemic peak, considering presence of immunodominant antigens in this peak as reported by earlier workers (Holland et al., 2002, Verloo et al., 2000, Verloo et al., 2001). Laha and Sasmal (2008) characterised WCL antigens of T. evansi prepared from buffalo, horse and cattle origins of eastern region of India by western
Conflict of interest statement
All authors disclose that they have no financial and personal relationships with other people or organization that could inappropriately influence (bias) their work, including employment, consultancies, stock ownership, honoraria, paid expert testimony, patent applications/registration, and grants or other funding.
Ethics
All authors certified that the animal experimentations carried out in the present work after approval of Institute Animal Ethic Committee and as per guidelines set by the Committee for the Purpose of Control and Supervision of Experiments on Animals (CPCSEA), Animal Welfare Division, Government of India.
Acknowledgements
The authors wish to acknowledge their gratitude to the Director, National Research Centre on Equines, Hisar, India for providing research facilities to carry out the present study. Authors are thankful to Prof. M. B. Chhabra for critically reading the manuscript and giving valuable feedback. They would also like to thank Mr. R. K. Dayal and Mr. Neeraj Kumar Yadav for technical assistance provided during the investigation.
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