Diagnosis of porcine and bovine enteric coronavirus infections using cloned cDNA probes

Molecular clones representing the first 2,000 bases from the 3' end of the porcine transmissible gastroenteritis coronavirus genome and the first 2,160 bases from the 3' end of the bovine enteric coronavirus genome were used in dot blot hybridization assays to detect viral RNA from cell culture and from fecal specimens. In each case, the cloned DNA represents approximately 10% of the genome. The cloned sequence for each virus encompasses the 3' noncoding region, the nucleocapsid protein gene, and a large portion of the matrix protein gene. 32P-labeled cDNA probes prepared from these clones detected as little as 25 pg of RNA from the parental virus but did not detect RNA from the nonparental virus even when amounts of up to 10 ng per dot were used. This specificity reflects the antigenic diversity between these two coronaviruses. The hybridization assay could also detect coronaviruses antigenically closely related to the parental virus but not coronaviruses belonging to an antigenically unrelated subgroup. Dot blot hybridization for transmissible gastroenteritis coronavirus diagnosis was compared with the routine procedures of virus isolation and electron microscopy as a diagnostic test.</jats:p

Detection and Titration of Human Herpesvirus-8–Specific Antibodies in Sera From Blood Donors, Acquired Immunodeficiency Syndrome Patients, and Kaposi's Sarcoma Patients Using a Whole Virus Enzyme-Linked Immunosorbent Assay

A human herpesvirus-8 (HHV-8) enzyme-linked immunosorbent assay (ELISA) with a whole virus lysate as antigen was developed and used to measure the seroprevalence rate and levels of IgG antibodies to HHV-8 in sera/plasma of various patient groups and blood donors. The virus antigen was prepared from the KS-1 cell line, which produces lytic virus, and therefore contains a broad array of viral proteins. Seroprevalence studies using this ELISA showed the following: 10 of 91 blood donors (11%) had an average HHV-8 antibody titer of 118; 67 of 72 (93%) classic Kaposi's sarcoma (KS) patients were positive with an average titer of 14,111; and 57 of 62 (92%) KS/human immunodeficiency virus (HIV) patients were positive with an average titer of 4,000. A study on a very limited number of serial serum samples from patients before and after diagnosis with KS showed highly elevated antibody titers to HHV-8 virus after KS lesions developed. Preliminary data show that 50% of the sera from HIV-1+ homosexual patients contain IgG antibodies to HHV-8 suggesting that this population is at high risk for developing KS. Antibody results correlated well with the confirmatory immunofluorescent assays (IFA) using KS-1 cells as the substrate. This HHV-8 IgG antibody detection ELISA is sensitive and specific and does not cross-react with Epstein-Barr virus (EBV) or other human herpesviruses. The results of this HHV-8 antibody survey suggest that this rapid ELISA assay can be used to screen large numbers of sera to find those at risk for developing KS.</jats:p