Multiple Homologies of Oligonucleotide Size Exist between Nucleic Acids of Picornaviruses

Abstract

SUMMARYA semi-quantitative analysis of hybrid formation between restriction enzymegenerated subgenomic fragments of cloned cDNA prepared from RNA of foot-andmouth disease virus (FMDV) strain O1K and radiolabelled RNA from bovine enterovirus, bovine rhinovirus or Mengo virus indicated that the hybrids were of oligonucleotide size. They were located in those parts of the FMDV O1K genome that code for the two capsid proteins VP3 and VP1 and the precursor protein P52 as well as at the Tend. No hybridization was observed with poliovirus type 1 RNA.Picornaviruses (for review, see Cooper et al., 1978) consist of a single-stranded RNA molecule 7000 to 8000 bases long of (+) polarity which is encapsidated by 60 copies of each of the four capsid proteins. The primary product of protein synthesis is a precursor polyprotein from which the mature proteins are released after extensive processing. The aphthoviruses represent one of the four genera grouped in this virus family. They can be distinguished from the others (enteroviruses, rhinoviruses and cardioviruses) by their different buoyant densities in caesium chloride, stabilities in the pH range 3 to 7 or ribonucleotide compositions. The Y-proximal RNA sequences of aphthoviruses are highly homologous, an observation providing evidence for the close relationship of the serotypes. The Y-proximal sequences coding for the capsid proteins VP1 and VP3 are less homologous (Marquardt, 1982); this may explain the antigenic variation between the serotypes noticed some time ago (Galloway et al., 1948;Wild et al., 1969).The specific distribution of sequence homology among aphthoviruses made us consider whether sequence homologies exist between distinct genome areas of aphthoviruses and those of other picornaviruses. The availability of bacterial clones which contain foot-and-mouth disease virus (FMDV) O~ K-specific complementary DNA (K/ipper et al., 1981) facilitated such studies. The experimental strategy is based on the correlation of the cloned cDNA to the map of FMDVinduced polypeptides (Fig. 1). It takes advantage of restriction enzymes which cleave FMDVspecific double-stranded DNA into defined subgenomic fragments. These fragments are analysed individually as to whether or not they hybridize with radiolabelled RNA by use of the technique of Southern (1975). Since RNA that did not hybridize is removed by RNase A digestion, the remaining RNA/DNA hybrids consist of true double-strands. They require a minimum size between more than 6 and about 30 base pairs (bp) to be detectable.Such a technique which detects homologies of oligonucleotide size is required to reveal common picornavirus sequences, because a study comparing the RNase T1 digestion products of the RNAs of representative members of three genera (Frisby et al., 1976) did not find large common oligonucleotides. However, a pentanucleotide resistant to the RNases T1 and A was shown to be shared by at least four RNAs. This could form the core of a common oligonucleotide consisting of more than six bases. The only homology thu...

Use of in Situ Hybridization for the Detection of Foot-and-Mouth Disease Virus in Cell Culture

“…In addition to being highly conserved among serotypes, the polymerase gene is also specific for FMDV with no hybridization of this fragment occurring with other picornaviruses, including bovine enterovirus (BEV), bovine rhinovirus, or poliovirus. 9 It was the aim of this study to investigate the use of biotinylated probes derived from the polymerase gene either as a DNA probe or as an RNA probe for detection of FMDV in cell cultures. Preparation of complementary DNA (cDNA) probe.…”