Doctoral Dissertations

Date of Award

12-1991

Degree Type

Dissertation

Degree Name

Doctor of Philosophy

Major

Comparative and Experimental Medicine

Major Professor

Michael A. Breider

Committee Members

Leon N. D. Potgieter, J. Erby Wilkinson, Barry T. Rouse

Abstract

Classification of respiratory syncytial viruses into subgroups is based primarily on genetic and antigenic divergence of an important immunogenic envelope glycoprotein that may be important in disease immunopathogenesis and immunoprophylaxis. Currently, there are two human respiratory syncytial virus (HRSV) subgroups, but relatively little is known about bovine, caprine, and ovine respiratory syncytial virus isolates. Laboratory work was conducted in 1989-1991 to gather data on subgroup divergence and G glycoprotein heterogeneity among respiratory syncytial viruses. Seven bovine (BRSV 375, 391-2, FS1, NMK7, 1143, 1144, and 16186), two human (HRSV A2 and B8/60), one caprine (CRSV), and one ovine (ORSY) respiratory syncytial virus strains from various geographic sources were analyzed using current biomolecular techniques including Western blot analysis, polymerase chain reaction, molecular cloning, dideoxy nucleotide DNA sequencing, and RNase mismatch cleavage. Western blot analysis was performed using bovine anti-BRSV strain 375 and human anti-HRSV sera. The bovine antiserum cross-reacted with the G glycoproteins from all BRSV strains tested, whereas the human antiserum reacted with the G glycoprotein from HRSV A2. Neither antiserum cross-reacted with the G glycoproteins from HRSV strain B8/60, CRSV, or ORSV. Oligonucleotide primers based on the nucleotide sequence of the BRSV strain 391-2 G glycoprotein gene were used in the polymerase chain reaction to amplify virus-specific G glycoprotein gene cDNAs from genomic RNA of CRSV and BRSV strains 391-2 and 375. Virus-specific G glycoprotein gene cDNA could not be amplified from ORSV genomic RNA. Nucleotide sequence of glycoprotein G gene cDNA from BRSV strain 375 was compared to a similar sequence from BRSV strain 391-2 indicating 95.0% and 89.9% nucleotide and amino acid identities, respectively. A radiolabelled riboprobe was made from a cDNA copy of the BRSV strain 375 G glycoprotein gene and all virus strains were compared by RNase mismatch cleavage analysis. The data indicated a close genetic relationship among CRSV and all BRSV strains suggesting that these strains be included in the same subgroup. In agreement with previously published data, the HRSV strains tested belong to additional RSV subgroups. Additionally, ORSV likely should be classified in a separate RSV subgroup.

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