Conservation of virally encoded microRNAs in Kaposi sarcoma--associated herpesvirus in primary effusion lymphoma cell lines and in patients with Kaposi sarcoma or multicentric Castleman disease.

BACKGROUND: MicroRNAs are small noncoding RNAs that posttranscriptionally regulate gene expression. Kaposi sarcoma (KS)-associated herpesvirus (KSHV) encodes 12 distinct microRNA genes, all of which are located within the latency-associated region that is highly expressed in all KSHV-associated malignancies. METHODS: We amplified, cloned, and sequenced a 2.8-kbp-long region containing a cluster of 10 microRNAs plus a 646-bp fragment of K12/T0.7 containing the remaining 2 microRNAs from 5 primary effusion lymphoma-derived cell lines and from 17 patient samples. The patients included 2 with classic KS, 12 with AIDS-KS (8 from the United States, 1 from Europe, 3 from Africa, and 4 from Central/South America), and 2 with multicentric Castleman disease (MCD). Additionally, we analyzed the K1, open reading frame 75, and K15 genes to determine KSHV subtypes, and we performed a phylogenetic analysis. RESULTS: Phylogenetic analysis of the 2.8-kbp microRNA region revealed 2 distinct clusters of sequences: a major (A/C) and a variant (B/Q) cluster. The variant cluster included sequences from 3 patients of African origin and both patients with MCD. Some microRNAs were highly conserved, whereas others had changes that could affect processing and, therefore, biological activity. CONCLUSIONS: These data demonstrate that KSHV microRNA genes are under tight selection in vivo and suggest that they contribute to the biological activity and possibly the pathogenesis of KSHV-associated malignancies.

Hepatitis C virus genotype 4 in Ugandan children and their mothers.

In Kampala, Uganda, in 2001, hepatitis C virus antibodies were found in 27 (4%) of 603 children and in 62 (12%) of 525 of their mothers. However, only approximately 10% of positive results were confirmed by reverse transcription-PCR, which suggests frequent false-positive results or viral clearance. All sequenced types were genotype 4.

Molecular evidence for mother-to-child transmission of Kaposi sarcoma-associated herpesvirus in Uganda and K1 gene evolution within the host.

BACKGROUND: Epidemiological studies of Kaposi sarcoma (KS)-related herpesvirus (KSHV) indicate that having a KSHV-seropositive mother is a risk factor for KSHV infection in children. METHODS: We determined the KSHV K1 sequences in concordantly polymerase chain reaction-positive Ugandan mother-child pairs, to ascertain whether they shared the same viral strain. We also examined sequences amplified from saliva and buffy coat samples from the same subjects, to investigate potential intrasubject sequence differences. RESULTS: We obtained K1 sequences from 6 of 10 mother-child pairs. In 1 pair, the subtypes differed between mother and child. The mother and child in 2 other pairs shared the same subtype, but the sequences differed. The mother and child in 2 pairs shared KSHV strains with exact (100%) nucleotide homology. The last pair showed evidence of viral strain concordance between mother and child but also showed evidence of evolution of the viral sequence within the child. Of 26 study subjects, 19 showed no evidence of intrasubject K1 sequence variability, but, in 7 subjects, all of whom were children, amino acid variation of 1%-4% was observed. CONCLUSIONS: Our findings are consistent with KSHV transmission from maternal and nonmaternal sources in KS-endemic regions. Our results also provide evidence for ongoing evolution of the K1 gene in KSHV-infected children.

Genotypic characterization of Kaposi's sarcoma-associated herpesvirus in asymptomatic infected subjects from isolated populations.

Molecular epidemiological studies of Kaposi's sarcoma-associated herpesvirus (KSHV) have concentrated on characterization of viral strains in tumour biopsy samples from Kaposi's sarcoma (KS) patients, mostly obtained in the United States and Europe. Tumour biopsies are a convenient source of viral DNA, as they have a high viral load compared to peripheral blood. However, sequences obtained from biopsies may not be representative of viral strains in asymptomatic subjects and information on ethnicity is often not available. Here, a population-based approach has been used to study the molecular and seroepidemiology of KSHV in isolated populations in Ecuador and Botswana. Amerindians in Ecuador had a variable prevalence of KSHV and all strains characterized were of subtype E, based on K1 sequencing. All Amerindian strains had predominant (P)-type K15 alleles and had sequences in both T0.7 and ORF 75 that appeared to be characteristic of these strains. The prevalence of KSHV in two ethnic groups in Botswana was extremely high. K1 sequences from both Bantu and San subjects were mostly of subtypes B and A5, which are typical of African KSHV strains, but the sequence from one San subject did not cluster with any known subtype. Considerable heterogeneity was seen in the T0.7 and ORF 75 genes in the San subjects and one had a minor (M)-type K15 allele. The heterogeneity of the KSHV strains found in these subjects from Botswana contrasts with the homogeneity of KSHV strains in Amerindians, reflecting differences in the evolutionary history of these populations.