Enzyme immunofiltration assay for measurement of antibodies to transmissible gastroenteritis virus of swine: comparison with enzyme immunosorbence assay, serum neutralization, and indirect immunofluorescent antibody techniques.

Abstract

There are 3 recognized coronaviruses of swine: transmissible gastroenteritis virus (TGEV), l,2 porcine epidemic diarrhea virus (PEDV), and porcine respiratory coronavirus (PRCV). The TGEV and PRCV cross-react serologically and are closely related. The PEDV is not serologically related to TGEV or PRCV. Transmissible gastroenteritis virus causes a highly contagious disease of swine characterized by vomiting, diarrhea, and dehydration. 1,2 High mortality occurs in pigs that are ≤ 10 days of age when exposed to TGEV. Pigs recovering from the disease may become chronic carriers of TGEV and a source of infection for susceptible animals. The carrier animal, especially older swine, are symptomless; therefore, breeding stock should be tested for TGEV antibodies before introduction into a susceptible herd. The serum neutralization (SN) test is the standard TGEV serologic test. The SN test is a useful and valid test but may lack sensitivity. The enzyme-linked immunosorbent assay (ELISA) and the indirect fluorescent antibody test (IFAT) are possibly more sensitive. 4-6 The purpose of this study was to compare a newer approach, the enzyme immunofiltration assay (ELIFA) to the other 3 tests. The ELIFA is a more rapid assay and could decrease the time required for TGEV serology. A total of 229 samples were taken from swine sera submitted to the diagnostic laboratory of the Department of Veterinary Diagnosis, College of Veterinary Medicine, Kansas State University, Manhattan, Kansas. The sera were inactivated at 56 C for 30 minutes prior to being used in the different tests. The swine testicle (ST) cell line was obtained from the National Veterinary Service Laboratory (NVSL) at Ames, Iowa. These cells were used to produce viral antigen needed in the different tests. Cell cultures were maintained in minimum essential medium (MEM) with 5% irridated fetal calf serum, 5% iron-supplemented calf serum, 0.5% lactalbumin hydrolysate, and 5 μg gentamycin per ml. The Purdue strain of TGEV was obtained from NVSL and propagated in ST cells. The stock virus was treated with trypsin at a final concentration of 5 μg/ml at for 30 minutes before inoculation onto confluent monolayers of ST cells (in 75-cm flask) at a multiplicity of infection of 10 tissue culture infective doses 50% (TCID50) per cell. After adsorption at 37 C for 1 hour, the inoculum was removed, and MEM was added, supplemented with 5 μg trypsin per ml. The infected cultures were incubated at 37 C for 24 hours. When the cytopathic effect was approximately 95%, the virus was harvested and cultures were frozen.