Abstract. The spike (S) glycoprotein of the Miller strain of transmissible gastroenteritis virus (TGEV) was recently cloned and expressed in baculovirus. The recombinant S protein was used as the coating antigen in a competition (blocking) enzyme-linked immunosorbent assay (ELISA) in combination with monoclonal antibodies to the S protein epitope A (conserved on TGEV and porcine respiratory coronavirus [PRCV]) or epitope D (present on TGEV only) to differentiate PRCV-from TGEV-induced antibodies. One set (set A) of 125 serum samples were collected at different times after inoculation of caesarean-derived, colostrum-deprived (n ϭ 52) and conventional young pigs (n ϭ 73) with 1 of the 2 porcine coronaviruses or uninoculated negative controls (TGEV/PRCV/negative ϭ 75/30/20). A second set (set B) of 63 serum samples originated from adult sows inoculated with PRCV and the recombinant TGEV S protein or with mock-protein control and then exposed to virulent TGEV after challenge of their litters. Sera from set A were used to assess the accuracy indicators (sensitivity, specificity, accuracy) of the fixed-cell blocking ELISA, which uses swine testicular cells infected with the M6 strain of TGEV as the antigen source (ELISA 1) and the newly developed ELISA based on the recombinant S protein as antigen (ELISA 2). The sera from set B (adults) were tested for comparison. The plaque reduction virus neutralization test was used as a confirmatory test for the presence of antibodies to TGEV/PRCV in the test sera. The accuracy indicators for both ELISAs suggest that differential diagnosis can be of practical use at least 3 weeks after inoculation by testing the dual (acute/convalescent) samples from each individual in conjunction with another confirmatory (virus neutralization) antibody assay to provide valid and complete differentiation information. Moreover, whereas ELISA 1 had 10-20% false positive results to epitope D for PRCV-infected pigs (set A samples), no false-positive results to epitope D occurred using ELISA 2, indicating its greater specificity. The progression of seroresponses to the TGEV S protein epitopes A or D, as measured by the 2 ELISAs, was similar for both sets (A and B) of samples. Differentiation between TGEV and PRCV antibodies (based on seroresponses to epitope D) was consistently measured after the third week of inoculation.Transmissible gastroenteritis virus (TGEV), a prominent cause of neonatal diarrhea, causes death in 90-100% of seronegative pigs under 2 weeks of age and costs the US swine industry nearly $200 million/ year. 15,19 Surveys of TGEV antibodies in swine sera collected prior to the detection of porcine respiratory coronavirus (PRCV) indicated that 19-54% of swine herds in North America have antibodies, 17 but few differential serosurveys have been done since the detection of PRCV. 32 A variant of TGEV, referred to as PRCV, was isolated from pigs in Europe in 1986 and identified in the USA in 1989. 15,18,19 As reported from Iowa swine herds, a recent increase observed in TGEV/PRCV seropre...

Real-time RT-PCR assay, based on light upon extension (LUX) fluorogenic primer and LightCycle technology, was developed for rapid detection of transmissible gastroenteritis virus (TGEV). Viral RNA from different TGEV isolates and clinical specimens was detected. To evaluate the sensitivity of the assay, a gel-based RT-PCR method targeted at the same 101 bp sequence was also developed. Serial 10-fold dilutions of TGEV RNA were detected by the two methods. Although the real time method used only 2 microl RNA for each reaction, a 10-fold increase of sensitivity over that of the gel-based method, which used 10 microl RNA was demonstrated. The study indicates that the LUX assay reported below is rapid, reliable and sensitive and it has the potential for use as an alternative molecular method for TGEV diagnosis.

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