Rapid Competitive Enzyme-Linked Immunosorbent Assay for Detection of Antibodies to Peste des Petits Ruminants Virus

Abstract

Peste des petits ruminants (PPR) is a contagious viral disease of small ruminants that is of economic importance in Africa, the Middle East, and Asia. We developed a rapid competitive enzyme-linked immunosorbent assay (rapid c-ELISA) for the diagnosis and surveillance of PPR. This assay detects PPR virus (PPRV) antibodies in serum samples by quantifying the amount of monoclonal antibody (MAb) P-3H12 after 30 min of incubation of a serum-MAb conjugate mixture on plates coated with a PPRV recombinant nucleocapsid protein (rPPRV-N). We tested 249 PPRV-positive serum samples and 733 PPRV-negative serum samples from field ruminants. The threshold of percent inhibition (PI) was determined to be <50 on the basis of the mean PI plus 3 standard deviations for sera from PPRV-negative ruminants. The relative specificity and sensitivity of the rapid c-ELISA were 98.5% (722 of 733 serum samples) and 93.4% (234 of 249 serum samples), respectively. The rapid c-ELISA sensitively detected PPRV antibodies in hyperimmune sera (virus neutralization test [VNT] titer, >512), even at dilutions >512 in normal goat serum, and as early as 6 to 13 days postinfection from 12 goats, each of which was infected with one of the four PPRV lineages. Hyperimmune sera from animals experimentally vaccinated with rinderpest virus gave positive results by the rapid c-ELISA when the rinderpest virus VNT titers were >512, although the rapid c-ELISA titers were very low (2 to 16). However, the rapid c-ELISA was negative when the rinderpest virus VNT titer was <128. The rapid c-ELISA developed in the present work provides a short turnaround time and could be a useful tool for the diagnosis of PPR and screening for PPRV in the field.Peste des petits ruminants (PPR) is an acute and highly contagious viral disease of small ruminants, such as sheep and goats, with high rates of morbidity and sometimes high rates of mortality. Economically, it has been the most important disease of these species in sub-Saharan Africa, the Middle East, and southwest Asia since it was first described in West Africa in 1942 (20). It also results in subclinical infection in large ruminants, which act as carriers of infection to small ruminants (1, 14). The PPR virus (PPRV) belongs to the genus Morbillivirus in the family Paramyxoviridae and is closely related to rinderpest virus (RPV) (2, 16). At least four distinct genetic lineages (lineages I, II, III, and IV) of PPRV with different geographical distributions circulate among small ruminants in regions of endemicity (6,20).A vigorous program for PPR eradication has been carried out in these regions, including vaccination, seromonitoring, serosurveillance, and the destruction of infected animals and those that have been in contact with them. Rapid detection of infected animals is very important for PPR controls to be effective. Severe cases in which animals show clinical signs in the field can easily be detected through clinical surveillance and the detection of antigen in clinical samples, while the diagnosis of PPRV infection ...

Global distribution of peste des petits ruminants virus and prospects for improved diagnosis and control

“…Currently, the OIE recommend the use of the competitive PPRVspecific anti-H monoclonal based ELISA (cH-ELISA) (Anderson & McKay, 1994) and virus neutralization tests (FAO, 1996). However, several alternatives exist (Choi et al, 2005;Libeau et al, 1995) including the indirect N ELISA (Ismail et al, 1995), immunofiltration (Dhinakar Raj et al, 2008), a novel sandwich ELISA (Saravanan et al, 2008), haemagglutination tests (Dhinakar Raj et al, 2000;Ezeibe et al, 2008) and latex agglutination tests (Keerti et al, 2009).…”

Diagnosis and control of peste des petits ruminants: a comprehensive review

“…They used recombinant N protein of Nigeria 75/1 strain of PPRV expressed in baculovirus as antigen and reported that the c-ELISA is as sensitive and specific as VNT, with the added advantage that it uses an antigen that is safe and can be produced in large quantities. Choi et al [31] developed a rapid c-ELISA for diagnosis and surveillance of PPR. This assay detects PPRV antibodies in serum samples by quantifying the amount of MAb after 30 min of incubation of a serum-MAb conjugate mixture on plates coated with a PPRV recombinant N protein (rPPRV-N) expressed in baculovirus system.…”

Prevalence of Peste Des Petits Ruminants Virus PPRV in Mardan, Hangu and Kohat District of Pakistan; Comparative Analysis of PPRV Suspected serum samples using Competitive ELISA cELISA and Agar Gel Immunodiffusion AGID

“…The results of their study suggest a need for continuous serological and clinical surveillance of PPR in wild ruminants in order to determine the prevalence of PPR, its effects on wildlife conservation and the possible role of these species in the transmission cycle of PPRV (Ogunsanmi et al., 2003). Choi et al, 2005 reported that the procedure of c-ELISA consists of four reaction steps: adsorption of the antigen onto a solid phase, competition between the serum and MAb, detection of the MAbbound to the antigen, and a substrate reaction. The presence of antibodies in serum samples will block reactivity of monoclonal antibody resulting in reduction of expected coloration enzyme labeled anti mouse conjugate and substrate-chromogen solution.…”

“…The same procedure was used in this study for PPR detection. The capability of the c-ELISA to deal with a large number of samples at a time and its short turnaround time may better serve the needs of surveillance and control programs (Choi et al, 2005).…”