Direct Repeat 6 from Human Herpesvirus-6B Encodes a Nuclear Protein that Forms a Complex with the Viral DNA Processivity Factor p41

The SalI-L fragment from human herpesvirus 6A (HHV-6A) encodes a protein DR7 that has been reported to produce fibrosarcomas when injected into nude mice, to transform NIH3T3 cells, and to interact with and inhibit the function of p53. The homologous gene in HHV-6B is dr6. Since p53 is deregulated in both HHV-6A and -6B, we characterized the expression of dr6 mRNA and the localization of the translated protein during HHV-6B infection of HCT116 cells. Expression of mRNA from dr6 was inhibited by cycloheximide and partly by phosphonoacetic acid, a known characteristic of herpesvirus early/late genes. DR6 could be detected as a nuclear protein at 24 hpi and accumulated to high levels at 48 and 72 hpi. DR6 located in dots resembling viral replication compartments. Furthermore, a novel interaction between DR6 and the viral DNA processivity factor, p41, could be detected by confocal microscopy and by co-immunoprecipitation analysis. In contrast, DR6 and p53 were found at distinct subcellular locations. Together, our data imply a novel function of DR6 during HHV-6B replication

Cytomegalovirus Replicon-Based Regulation of Gene Expression In Vitro and In Vivo

There is increasing evidence for a connection between DNA replication and the expression of adjacent genes. Therefore, this study addressed the question of whether a herpesvirus origin of replication can be used to activate or increase the expression of adjacent genes. Cell lines carrying an episomal vector, in which reporter genes are linked to the murine cytomegalovirus (MCMV) origin of lytic replication (oriLyt), were constructed. Reporter gene expression was silenced by a histone-deacetylase-dependent mechanism, but was resolved upon lytic infection with MCMV. Replication of the episome was observed subsequent to infection, leading to the induction of gene expression by more than 1000-fold. oriLyt-based regulation thus provided a unique opportunity for virus-induced conditional gene expression without the need for an additional induction mechanism. This principle was exploited to show effective late trans-complementation of the toxic viral protein M50 and the glycoprotein gO of MCMV. Moreover, the application of this principle for intracellular immunization against herpesvirus infection was demonstrated. The results of the present study show that viral infection specifically activated the expression of a dominant-negative transgene, which inhibited viral growth. This conditional system was operative in explant cultures of transgenic mice, but not in vivo. Several applications are discussed

Micronutrient Fortification to Improve Growth and Health of Maternally HIV-Unexposed and Exposed Zambian Infants: A Randomised Controlled Trial

Background: The period of complementary feeding, starting around 6 months of age, is a time of high risk for growth faltering and morbidity. Low micronutrient density of locally available foods is a common problem in low income countries. Children of HIV-infected women are especially vulnerable. Although antiretroviral prophylaxis can reduce breast milk HIV transmission in early infancy, there are no clear feeding guidelines for after 6 months. There is a need for acceptable, feasible, affordable, sustainable and safe (AFASS by WHO terminology) foods for both HIV-exposed and unexposed children after 6 months of age.Methods and Findings: We conducted in Lusaka, Zambia, a randomised double-blind trial of two locally made infant foods: porridges made of flour composed of maize, beans, bambaranuts and groundnuts. One flour contained a basal and the other a rich level of micronutrient fortification. Infants (n = 743) aged 6 months were randomised to receive either regime for 12 months. The primary outcome was stunting (length-for-age Z < -2) at age 18 months. No significant differences were seen between trial arms overall in proportion stunted at 18 months (adjusted odds ratio 0.87; 95% CI 0.50, 1.53; P = 0.63), mean length-for-age Z score, or rate of hospital referral or death. Among children of HIV-infected mothers who breastfed <6 months (53% of HIV-infected mothers), the richly-fortified porridge increased length-for-age and reduced stunting (adjusted odds ratio 0.17; 95% CI 0.04, 0.84; P = 0.03). Rich fortification improved iron status at 18 months as measured by hemoglobin, ferritin and serum transferrin receptors.Conclusions: In the whole study population, the rich micronutrient fortification did not reduce stunting or hospital referral but did improve iron status and reduce anemia. Importantly, in the infants of HIV-infected mothers who stopped breastfeeding before 6 months, the rich fortification improved linear growth. Provision of such fortified foods may benefit health of these high risk infants

Complete Genome Sequence of the Human Herpesvirus 6A Strain AJ from Africa Resembles Strain GS from North America.

The genome sequence of human herpesvirus 6A (HHV-6A) strain AJ was determined in a comparison of target enrichment and long-range PCR using next-generation sequencing methodologies. The analyses show 85 predicted open reading frames (ORFs), conservation with sequenced HHV-6A reference strain U1102, and closest identity to the recently determined GS strain, despite different geographic origins (United States and Gambia)

Analyses of germline, chromosomally integrated human herpesvirus 6A and B genomes indicate emergent infection and new inflammatory mediators.

Human herpesvirus-6A (HHV-6A) is rarer than HHV-6B in many infant populations. However, they are similarly prevalent as germline, chromosomally integrated genomes (ciHHV-6A/B). This integrated form affects 0.1-1 % of the human population, where potentially virus gene expression could be in every cell, although virus relationships and health effects are not clear. In a Czech/German patient cohort ciHHV-6A was more common and diverse than ciHHV-6B. Quantitative PCR, nucleotide sequencing and telomeric integration site amplification characterized ciHHV-6 in 44 German myocarditis/cardiomyopathy and Czech malignancy/inflammatory disease (MI) patients plus donors. Comparisons were made to sequences from global virus reference strains, and blood DNA from childhood-infections from Zambia (HHV-6A mainly) and Japan (HHV-6B). The MI cohort were 86 % (18/21) ciHHV-6A, the cardiac cohort 65 % (13/20) ciHHV-6B, suggesting different disease links. Reactivation was supported by findings of 1) recombination between ciHHV-6A and HHV-6B genes in 20 % (4/21) of the MI cohort; 2) expression in a patient subset, of early/late transcripts from the inflammatory mediator genes chemokine receptor U51 and chemokine U83, both identical to ciHHV-6A DNA sequences; and 3) superinfection shown by deep sequencing identifying minor virus-variants only in ciHHV-6A, which expressed transcripts, indicating virus infection reactivates latent ciHHV-6A. Half the MI cohort had more than two copies per cell, median 5.2, indicative of reactivation. Remarkably, the integrated genomes encoded the secreted-active form of virus chemokines, rare in virus from childhood-infections. This shows integrated virus genomes can contribute new human genes with links to inflammatory pathology and supports ciHHV-6A reactivation as a source for emergent infection