Molluscum contagiosum virus expresses late genes in primary human fibroblasts but does not produce infectious progeny.

Molluscum contagiosum virus (MCV), a member of the family Poxviridae, can be isolated from skin-lesion material of patients with molluscum contagiosum infection. MCV replicates efficiently in human keratinocytes in vivo but viral replication has not been observed in vitro in cell or tissue culture systems. We investigated a variety of established cell lines for productive MCV infection and found that: (i) MCV induces a typical cytopathogenic effect (CPE) only in human primary fibroblast cells (MRC5 ATCC-CCL 171 and HEPM ATCC-CRL 1486) but not in permanent eucaryotic cell lines of human or simian origin; (ii) UV irradiated MCV virions and heat inactivated virions do not induce a CPE; (iii) decreasing amounts of MCV viral DNA are detectable in infected human embryonic fibroblasts for at least 14 days post infection (p.i.); (iv) MCV early mRNAs are detectable by RT-PCR between one and two hours p.i. and remain detectable upon passaging of the infected cells; (iv) transcripts of viral late genes (mc095L and mc106L) are detectable by RT-PCR from day 5 p.i.; (v) MCV viral antigens can be detected on the surface of infected cells using human and rabbit polyclonal antisera against MCV from 24 h p.i.; (vi) a CPE can not be observed if cell free supernatants or homogenizates of MCV infected cells are used to try passage of the virus onto uninfected human embryonic fibroblasts, indicating that infectious viral progeny is not produced. This is the first report demonstrating long time persistence of MCV viral DNA and expression of late proteins in an in vitro cell culture system.

Mapping of mRNA transcripts in the genome of molluscum contagiosum virus: transcriptional analysis of the viral slam gene family.

Molluscum contagiosum virus (MCV) is a member of the poxvirus family and causes benign skin tumors in children and immunocompromised individuals. The primary structure and coding capacity of MCV was previously determined by DNA nucleotide sequencing (Senkevich et al., Science 273, 813-816, 1996). Hypothetical genes were predicted based on (i) amino acid homologies with known genes, (ii) presence or absence of conserved transcription regulation signals, and (iii) algorithms based on learning sets of coding sequences. These methods provide a rational basis for the prediction of MCV coding sequences. However, the existence and exact size of MCV open reading frames and the precise position of transcription regulation signals can only be determined by MCV mRNA transcript mapping experiments. We developed methods for the characterization of the mRNA transcripts of MCV genes in infected skin tissue and abortively infected human fibroblast cell cultures. Using these methods the properties of the mRNA transcripts of the MCV SLAM (signaling lymphocytic activating molecule) gene family (mc002L, mc161R, and mc162R) were analyzed. The mRNA start site found for the mc161R transcript suggests that a second start codon is used leading to a mc161R open reading frame that is nine amino acid residues shorter than predicted.