ORIGINAL RESEARCH Comparative efficacy of immunological, molecular and culture assays for detection of group A rotavirus from faecal samples of buffalo (Bubalus bubalis) calves Balvinder Kumar Manuja & Minakshi Prasad & Baldev R. Gulati & Anju Manuja & Gaya Prasad Accepted: 1 May 2009 / Published online: 6 July 2010 # Springer Science+Business Media B.V. 2010 Abstract Group A rotaviruses play an important role in causing gastroenteritis and mortality in buffalo (Bubalus bubalis) calves. A number of assays like RNA-polyacrylamide gel electro- phoresis (RNA-PAGE), enzyme-linked immunosorbent assay (ELISA), reverse transcription polymerase chain reaction (RT-PCR) and virus isolation have been employed for rotavirus diagnosis. We evaluated the comparative efficacy of different assays for detection of group A rotavirus in buffalo calves. A total of 455 faecal samples collected from five organized farms in northern India were screened by monoclonal antibody based ELISA, 33 (7.25%) samples were positive for group A rotavirus. The percent positivity ranged from 3.22% to 28% in different organized farms. The same samples were also tested by RNA-PAGE, which revealed classical 11 segments with 4:2:3:2 migration patterns in 14 faecal samples showing 3.08% positivity. Virus isolation was successfully done from 21 (4.61%) samples. However, only 15 (3.3%) samples yielded a specific product of 864 and 1,011 bp for VP4 and VP7 genes, respectively, by RT-PCR. The sensitivity and specificity of ELISA, RNA-PAGE and RT-PCR was 100%, 66.67% and 71.43% and 97%, 100% and 100%, respectively, considering virus isolation as standard test. ELISA being simple, fast and sensitive assay can be used as routine laboratory test for the diagnosis of group A rotavirus and field epidemiological studies. Keywords Buffalo . Group A rotavirus . RT-PCR . RNA-PAGE . Virus isolation Introduction Rotavirus, the prototype virus of the genus Rotavirus in the family Reoviridae, causes severe diarrheal disease in neonates of man and animals throughout the world. Seven distinct groups (A to G) of viruses have been described. Group A, B and C rotaviruses have been found in both humans and animals; group D to G rotaviruses have been found only in animals (Saif and Jiang 1994). The presence of rotavirus in buffalo calves and its association with diarrhoea has been reported in various countries like Sri Lanka (Chandra and Mahalingam 1995) and Italy (Martella et al. 1999, 2005; Pisanelli et al. 2005), including India (Mittal et al. 1986; Singh and Pandey 1988; Gulati et al. 1995; Grover et al. 1998; Jindal et al. 2000; Manuja et al. 2008; Nataraju et al. 2009). Buffalo (Bubalus bubalis) contributes significantly to the economy of dairy farmers in India. The country has over 98 million buffaloes which constitute 56.4% of the world buffalo population and contribute 66.3% of total buffalo milk in the world (http://dahd.nic.in/stat_files/BAHS2006). Calf mortality due to gastroenteritis is major concern for farmers and animal scientists in India (Rathore 1998). Incidence of rotavirus-associated diarrhoea in buffalo calves has been reported between 3% (Gulati et al. 1995) and 64.28% (Mittal et al. 1986). The mortality due to B. K. Manuja : M. Prasad : G. Prasad Department of Animal Biotechnology, CCS Haryana Agricultural University, Hisar, Haryana 125 004, India B. R. Gulati : A. Manuja National Research Centre on Equines, Hisar, Haryana 125 001, India Present Address: B. K. Manuja (*) National Research Centre on Equines, Hisar, Haryana 125 001, India e-mail: bmanuja@lycos.com Trop Anim Health Prod (2010) 42:1817–1820 DOI 10.1007/s11250-010-9642-y