STRATEGY FOR IDENTIFYING THE GENE ENCODING THE DNA-POLYMERASE OF MOLLUSCUM CONTAGIOSUM VIRUS TYPE-1

Molluscum contagiosum virus (MCV) is a member of the family Poxviridae and pathogenic to humans. MCV causes benign epidermal tumors mainly i n children and young adults and is a common pathogen in immunecompromi sed individuals. The viral DNA polymerase is the essential enzyme invo lved in the replication of the genome of DNA viruses. The identificati on and characterization of the gene encoding the DNA polymerase of mol luscum contagiosum virus type 1 (MCV-1) was carried out by PCR technol ogy and nucleotide sequence analysis. Computer-aided analysis of known amino acid sequences of DNA polymerases from two members of the poxvi rus family revealed a high amino acid sequence homology of about 49.7% as detected between the DNA polymerases of vaccinia virus (genus Orth opoxvirus) and fowlpoxvirus (genus Avipoxvirus). Specific oligonucleot ide primers were designed and synthesized according to the distinct co nserved regions of amino acid sequences of the DNA polymerases in whic h the codon usage of the MCV-I genome was considered. Using this techn ology a 228 bp DNA fragment was amplified and used as hybridization pr obe for identifying the corresponding gene of the MCV-1 genome. It was found that the PCR product was able to hybridize to the BamHI MCV-1 D NA fragment G (9.2 kbp, 0.284 to 0.332 map units). The nucleotide sequ ence of this particular region of the MCV-1 genome (7267 bp) between m ap coordinates 0.284 and 0.315 was determined. The analysis of the DNA sequences revealed the presence of 22 open reading frames (ORFs-1 to -22). ORF-13 (3012 bp; nucleotide positions 6624 to 3612) codes for a putative protein of a predicted size of 115 kDa (1004 aa) which shows 40.1% identity and 35% similarity to the amino acid sequences of the D NA polymerases of vaccinia, variola, and fowlpoxvirus. In addition sig nificant homologies (30% to 55%) were found between the amino acid seq uences of the ORFs 3, -5, -9, and -14 and the amino acid sequences of the E6R, E8R, E10R, and a 7.3 kDa protein of vaccinia and variola viru s, respectively. Comparative analysis of the genomic positions of the loci of the detected viral genes including the DNA polymerases of MCV- 1, vaccinia, and variola virus revealed a similar gene organization an d arrangement.