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Background Disparities in mortality in human immunodeficiency virus (HIV)–associated Kaposi sarcoma have been described, particularly in Black men in the southern United States. It is unclear if there are racial/ethnic differences in the seroprevalence of Kaposi sarcoma–associated herpesvirus (KSHV) that may be contributing. Methods This is a cross-sectional study of men who have sex with men (MSM) and transgender women with HIV. Participants were recruited from an outpatient HIV clinic in Dallas, Texas, for a 1-time study visit and were excluded from analysis if they had any history of KSHV disease. Plasma was tested for antibodies to KSHV K8.1 or ORF73 antigens, and KSHV DNA was measured in oral fluids and blood by polymerase chain reaction. KSHV seroprevalence and viral shedding in blood and oral fluids were calculated. Additionally, independent risk factors for KSHV seropositivity were assessed by multivariable logistic regression analysis. Results Two hundred five participants were included in our analysis. Overall, KSHV seroprevalence was high (68%) with no significant difference between racial/ethnic groups. Among seropositive participants, KSHV DNA was detected in 28.6% of oral fluids and 10.9% of peripheral blood specimens, respectively. The factors most strongly associated with KSHV seropositivity were oral–anal sex (odds ratio [OR], 3.02), oral–penile sex (OR, 4.63), and methamphetamine use (OR, 4.67). Conclusions High local seroprevalence of KSHV is likely a key driver of the high burden of KSHV-associated diseases regionally, though it does not explain the observed disparities in KSHV-associated disease prevalence among racial/ethnic groups. Our findings support that KSHV is primarily transmitted via exchange of oral fluids.

We previously found that age, sex, and malaria were associated with KSHV viral load in individuals from Uganda. In this study, we have evaluated factors associated with presence of EBV DNA in blood and oral fluids among the same individuals, using the same biological samples. Overall, 74% of oral fluids samples and 46% of PBMCs had detectable EBV, compared to 24% and 11% for KSHV respectively Individuals with EBV in PBMCs were more likely to have KSHV in PBMCs (P=0.016). The peak age for detection of EBV in oral fluids was 3-5 years while that of KSHV was 6-12 years. In PBMCs, the peak age for detection of EBV was 66+ years and KSHV was 3-5 years. Individuals with malaria had higher levels of EBV in PBMCs compared to malaria-negative individuals (P=0.002). In summary, our results show that younger age and malaria are associated with higher levels of EBV and KSHV in PBMCs suggesting malaria impacts immunity to EBV and KSHV.

Infection by Kaposi sarcoma-associated herpesvirus (KSHV) can cause severe consequences, such as cancers and lymphoproliferative diseases. Whole inactivated viruses (WIV) with chemically destroyed genetic materials have been used as antigens in several licensed vaccines. During KSHV productive replication, virus-like vesicles (VLVs) that lack capsids and viral genomes are generated along with virions. Here, we investigated the immunogenicity of KSHV VLVs produced from a viral mutant that was defective in capsid formation and DNA packaging. Mice immunized with adjuvanted VLVs generated KSHV-specific T cell and antibody responses. Neutralization of KSHV infection by the VLV immune serum was low but was markedly enhanced in the presence of the complement system. Complement-enhanced neutralization and complement deposition on KSHV-infected cells was dependent on antibodies targeting viral open reading frame 4 (ORF4). However, limited complement-mediated enhancement was detected in the sera of a small cohort of KSHV-infected humans which contained few neutralizing antibodies. Therefore, vaccination that induces antibody effector functions can potentially improve infection-induced humoral immunity. Overall, our study highlights a potential benefit of engaging complement-mediated antibody functions in future KSHV vaccine development. IMPORTANCE KSHV is a virus that can lead to cancer after infection. A vaccine that prevents KSHV infection or transmission would be helpful in preventing the development of these cancers. We investigated KSHV VLV as an immunogen for vaccination. We determined that antibodies targeting the viral protein ORF4 induced by VLV immunization could engage the complement system and neutralize viral infection. However, ORF4-specific antibodies were seldom detected in the sera of KSHV-infected humans. Moreover, these human sera did not potently trigger complement-mediated neutralization, indicating an improvement that immunization can confer. Our study suggests a new antibody-mediated mechanism to control KSHV infection and underscores the benefit of activating the complement system in a future KSHV vaccine.

Liver transplantation (LT) from donors-with-HIV to recipients-with-HIV (HIV D+/R+) is permitted under the HOPE Act. There are only three international single-case reports of HIV D+/R+ LT, each with limited follow-up. We performed a prospective multicenter pilot study comparing HIV D+/R+ to donors-without-HIV to recipients-with-HIV (HIV D-/R+) LT. We quantified patient survival, graft survival, rejection, serious adverse events (SAEs), human immunodeficiency virus (HIV) breakthrough, infections, and malignancies, using Cox and negative binomial regression with inverse probability of treatment weighting. Between March 2016-July 2019, there were 45 LTs (8 simultaneous liver-kidney) at 9 centers: 24 HIV D+/R+, 21 HIV D-/R+ (10 D- were false-positive). The median follow-up time was 23 months. Median recipient CD4 was 287 cells/µL with 100% on antiretroviral therapy; 56% were hepatitis C virus (HCV)-seropositive, 13% HCV-viremic. Weighted 1-year survival was 83.3% versus 100.0% in D+ versus D- groups (p = .04). There were no differences in one-year graft survival (96.0% vs. 100.0%), rejection (10.8% vs. 18.2%), HIV breakthrough (8% vs. 10%), or SAEs (all p .05). HIV D+/R+ had more opportunistic infections, infectious hospitalizations, and cancer. In this multicenter pilot study of HIV D+/R+ LT, patient and graft survival were better than historical cohorts, however, a potential increase in infections and cancer merits further investigation.

Background Kaposi sarcoma-associated herpesvirus (KSHV) seroprevalence in sub-Saharan African children can range up to 50% by age 2 years but factors affecting early age of KSHV infection are not well understood. Malaria during pregnancy has been associated with hindered transplacental transfer of antibodies to several pathogens but whether it affects transplacental transfer of KSHV antibodies is unknown. We aimed to determine if in utero malaria exposure reduced the transfer of KSHV antibodies across the placenta. Methods A cohort study in Kisumu, Kenya enrolled pregnant women at their first antenatal clinic (ANC) visit and followed them through delivery. We included 70 KSHV-positive, HIV-negative mothers and their children. KSHV antibody levels were measured by ELISA (K8.1, ORF73) and multiplex assay (K8.1, ORF73, K10.5, ORF38, ORF50). Transplacental transfer of antibodies was measured by the cord to maternal blood ratio (CMR) of KSHV antibodies. Malaria during pregnancy was defined as detection of Plasmodium falciparum (Pf) DNA at any ANC visit or delivery. Among women with malaria during pregnancy, we examined time of last malaria infection prior to delivery ( Results KSHV seroprevalence (positive for K8.1 or ORF73 by ELISA) among pregnant women was 88%. Neither malaria during pregnancy, malaria infection timing, nor MIR were associated with maternal delivery KSHV antibody blood levels. Maternal delivery and cord blood KSHV antibody levels were highly correlated but these correlations did not differ by malaria during pregnancy. KSHV transplacental antibody transfer was not associated with malaria during pregnancy, malaria infection timing, nor MIR. Conclusions Malaria during pregnancy does not appear to affect transfer of KSHV antibodies across the placenta.

OBJECTIVE To assess the cytokine and viral profiles of effusions and peripheral blood among patients diagnosed with HIV and Kaposi sarcoma herpesvirus [KSHV, also known as human herpesvirus 8 (HHV-8)]-associated conditions. DESIGN Retrospective comparative study evaluating clinicopathologic findings in patients with HIV and KSHV-associated conditions presenting with an effusion between 2010 and 2018. METHODS Paired peripheral blood and effusion samples collected at the time of pathological diagnosis of KSHV-associated conditions [Kaposi sarcoma, KSHV-associated multicentric Castleman disease (KSHV-MCD), primary effusion lymphoma (PEL), or KSHV-associated inflammatory cytokine syndrome (KICS)] were evaluated for disease-specific and compartment-specific (effusion vs. blood) characteristics. We assessed 12 cytokines, KSHV viral DNA (KSHV-VL), and Epstein--Barr virus (EBV) viral DNA (EBV-VL). RESULTS Nine patients had PEL, five patients had KSHV-MCD, and eight patients met criteria for KICS; all but one patient had concurrent Kaposi sarcoma in addition to these conditions. PEL effusions had substantially higher levels of IL-13 (median 16.9 pg/ml; interquartile range 9.7--26.9 pg/ml) compared with KSHV-MCD (median

Kaposi's sarcoma-associated herpesvirus (KSHV) and Epstein-Barr virus (EBV) are prevalent in sub-Saharan Africa, together with HIV; the consequent burden of disease is grave. The cofactors driving transmission of the two viruses and pathogenesis of associated malignancies are not well understood. We measured KSHV and EBV DNA in whole blood and saliva as well as serum antibodies levels in 175 Cameroonians with Kaposi's sarcoma and 1,002 age- and sex-matched controls with and without HIV. KSHV seroprevalence was very high (81%) in controls, while EBV seroprevalence was 100% overall. KSHV DNA was detectable in the blood of 36-46% of cases and 6-12% of controls; EBV DNA was detected in most participants (72-89%). In saliva, more cases (50-58%) than controls (25-28%) shed KSHV, regardless of HIV infection. EBV shedding was common (75-100%); more HIV+ than HIV- controls shed EBV. Cases had higher KSHV and EBV VL in blood and saliva then controls, only among HIV+ participants. KSHV and EBV VL were also higher in HIV+ than in HIV- controls. Cases (but not controls) were more likely to have detectable KSHV in blood if they also had EBV, whereas shedding of each virus in saliva was independent. While EBV VL in saliva and blood were modestly correlated, no correlation existed for KSHV. Numerous factors, several related to parasitic coinfections, were associated with detection of either virus or with VL. These findings may help better understand the interplay between the two gammaherpesviruses and generally among copathogens contributing to cancer burden in sub-Saharan Africa.

Background Detectable Kaposi’s sarcoma–associated herpesvirus (KSHV) DNA in blood and increased antibody titres may indicate KSHV reactivation, while the transmission of KSHV occurs via viral shedding in saliva. Methods We investigated the risk factors for KSHV DNA detection by real-time polymerase chain reaction in blood and by viral shedding in saliva, in 878 people aged 3 to 89 years of both sexes in a rural Ugandan population cohort. Helminths were detected using microscopy and the presence of malaria parasitaemia was identified using rapid diagnostic tests. Regression modelling was used for a statistical analysis. Results The KSHV viral load in blood did not correlate with the viral load in saliva, suggesting separate immunological controls within each compartment. The proportions of individuals with a detectable virus in blood were 23% among children aged 3–5 years and 22% among those 6–12 years, thereafter reducing with increasing age. The proportions of individuals with a detectable virus in saliva increased from 30% in children aged 3–5 years to 45% in those aged 6–12 years, and decreased subsequently with increasing age. Overall, 29% of males shed in saliva, compared to 19% of females (P = .008). Conclusions Together, these data suggest that young males may be responsible for much of the onward transmission of KSHV. Individuals with a current malaria infection had higher levels of viral DNA in their blood (P = .031), compared to uninfected individuals. This suggests that malaria may lead to KSHV reactivation, thereby increasing the transmission and pathogenicity of the virus.

Background We aimed to determine whether Plasmodium falciparum infection affects age of Kaposi sarcoma-associated herpesvirus (KSHV) seroconversion in Kenyan children. Methods Kenyan children (n = 144) enrolled at age 1 month, from 2 sites with different levels of malaria transmission (stable/high vs unstable/low) were followed to age 24 months. Plasma was tested for KSHV antibodies using enzyme-linked immunosorbent assay (ELISA; K8.1 and LANA) and a multiplex bead-based assay (K8.1, K10.5, ORF38, ORF50, and LANA) and whole blood tested for P. falciparum DNA using quantitative PCR. Cox proportional hazards models were used to assess associations between P. falciparum DNA detection, malaria annualized rate (P. falciparum detections/person-years), and enrollment site (malaria-high vs malaria-low) with time to KSHV seroconversion. Results KSHV seroprevalence was 63% by age 2 years when assessed by multiplex assay. Children with P. falciparum were at increased hazards of earlier KSHV seroconversion and, among children with malaria, the hazard of becoming KSHV seropositive increased significantly with increasing malaria annualized rate. Children from the malaria-high transmission region had no significant difference in hazards of KSHV seroconversion at 12 months but were more likely to become KSHV seropositive by age 24 months. Discussion Malaria exposure increases the risk for KSHV seroconversion early in life.

Kaposi’s sarcoma-associated herpesvirus (KSHV) and Epstein-Barr Virus (EBV) establish life-long infections and are associated with malignancies. Striking geographic variation in incidence and the fact that virus alone is insufficient to cause disease, suggests other co-factors are involved. Here we present epidemiological analysis and genome-wide association study (GWAS) in 4365 individuals from an African population cohort, to assess the influence of host genetic and non-genetic factors on virus antibody responses. EBV/KSHV co-infection (OR = 5.71(1.58–7.12)), HIV positivity (OR = 2.22(1.32–3.73)) and living in a more rural area (OR = 1.38(1.01–1.89)) are strongly associated with immunogenicity. GWAS reveals associations with KSHV antibody response in the HLA-B/C region (p = 6.64 × 10−09). For EBV, associations are identified for VCA (rs71542439, p = 1.15 × 10−12). Human leucocyte antigen (HLA) and trans-ancestry fine-mapping substantiate that distinct variants in HLA-DQA1 (p = 5.24 × 10−44) are driving associations for EBNA-1 in Africa. This study highlights complex interactions between KSHV and EBV, in addition to distinct genetic architectures resulting in important differences in pathogenesis and transmission., Disease prognosis after infection with Kaposi’s sarcoma-associated herpesvirus and Epstein-Barr Virus is highly variable. Here the authors carry out epidemiological and genetic analysis of a Ugandan cohort and suggest complex interactions may influence pathogenesis and transmission.

Kaposi’s sarcoma-associated herpesvirus (KSHV) transmission within endemic areas may vary. KSHV seroprevalence has been studied by different groups of researchers using different methods, making it difficult to make direct comparisons. Here we show results on KSHV seroprevalence using the same laboratory method from four different but geographically proximate populations in Uganda.Blood samples from the urban Entebbe Mother and Baby Study (EMaBS), the rural General Population Cohort (GPC), the fishing community Lake Victoria Island Intervention Study on Worms and Allergy related Diseases (LaVIISWA) and the high-risk sexual behaviour Good Health for Women Project (GHWP), were tested for IgG antibody levels to K8.1 and ORF73 recombinant proteins using ELISA.All adult participants of the EMaBS study and the GHWP were women, while the GPC (54% female) and LaVIISWA (52% female) studies had both males and females. EMaBS children were all 5 years of age while their mothers were 14 to 47 years of age. GHWP women were 15 to 45 years old, LaVIISWA participants were 1 to 72 years old while GPC participants were 1 to 103 years old. KSHV seropositivity varied in the different populations. In children aged 5 years, EMaBS had the lowest prevalence of 15% followed by GPC at 35% and LaVIISWA at 54%. In adult women, seropositivity varied from 69% (EMaBS) to 80% (LaVIISWA) to 87% (GPC) to 90% (GHWP).The reasons for the variation in prevalence are unclear but may reflect differences in the prevalence of cofactors between these four geographically proximate populations.

We investigated the impact of helminths and malaria infection on Kaposi’s sarcoma associated herpesvirus (KSHV) seropositivity, using samples and data collected from a cluster-randomised trial of intensive versus standard anthelminthic treatment. The trial was carried out in 2012 to 2016 among fishing communities on Lake Victoria islands in Uganda. Plasma samples from 2881 participants from two household surveys, the baseline (1310 participants) and the final (1571 participants) surveys were tested for KSHV IgG antibody responses to K8.1 and ORF73 recombinant proteins using ELISA. The baseline survey was carried out before the trial intervention while the final survey was carried out after three years of the trial intervention. Additionally, a subset sample of 372 participants from the final survey was tested for IgE, IgG and IgG4 antibody concentrations to S. mansoni adults worm antigen (SWA) and S. mansoni egg antigen (SEA) using ELISA. Infection by helminths (S. mansoni, N. americanus, T. trichiura and S. stercoralis) was diagnosed using real-time PCR, urine circulating cathodic antigen (CCA) and stool microscopy (Kato-Katz method) while malaria infection was diagnosed using microscopy. We analysed the relationship between helminth and malaria infections and KSHV seropositivity using regression modelling, allowing for survey design. At baseline, 56% of the participants were male while 48% of the participants were male in the final survey. The most prevalent helminth infection was S. mansoni (at baseline 52% and 34% in the final survey by microscopy, 86% by CCA and 50% by PCR in the final survey). KSHV seropositivity was 66% (baseline) and 56% (final survey) among those 1–12 years and >80% in those 13+ years in both surveys; malaria parasitaemia prevalence was 7% (baseline) and 4% (final survey). At baseline, individuals infected with S. mansoni (detected by microscopy) were more likely to be KSHV seropositive (aOR = 1.86 (1.16, 2.99) p = 0.012) and had higher anti-K8.1 antibody levels (acoefficient = 0.03 (0.01, 0.06) p = 0.02). In the final survey, S. mansoni (by microscopy, adjusted Odds Ratio (aOR = 1.43 (1.04–1.95), p = 0.028) and malaria parasitaemia (aOR = 3.49 (1.08–11.28), p = 0.038) were positively associated with KSHV seropositivity. Additionally, KSHV seropositive participants had higher S. mansoni-specific IgE and IgG antibody concentrations in plasma. Furthermore, HIV infected individuals on cART were less likely to be KSHV seropositive compared to HIV negative individuals (aOR = 0.46 (0.30, 0.71) p = 0.002). Schistosoma species skew the immune response towards Th2 and regulatory responses, which could impact on KSHV reactivation if co-infected with both organisms., Author summary Kaposi’s sarcoma associated herpesvirus (KSHV), the causative agent of Kaposi’s sarcoma cancer, varies geographically. KSHV infections are highest in sub-Saharan Africa, with Uganda having the highest prevalence reported to date. Infection with KSHV is lifelong with an intermittent revival of the virus, leading to viral spread. In this study, we show that infection with Schistosoma mansoni and malaria parasites is associated with being infected or exposed to KSHV. These parasite infections interfere with the proper functioning of the immune system to control viral infections. Although not shown in the current study, these parasite infections might lead to reactivation of KSHV in infected people increasing the likelihood of having detectable KSHV antibodies. Consequently, this viral reactivation may increase the spread of KSHV in sub-Saharan Africa.

Several studies published to date report associations between human leukocyte antigen (HLA) alleles and different types of Kaposi’s Sarcoma (KS). However, there is little concordance between the HLA alleles identified and the populations studied. To test whether HLA alleles associate with KS in a Cameroonian case–control study, we performed high-resolution HLA typing in KSHV seropositive individuals. Among HIV-positive individuals, carriers of HLA-B*14:01 were at a significantly higher risk of AIDS-KS (p = 0.033). For HIV-negative patients, a gene-wise comparison of allele frequencies identified the HLA-B (p = 0.008) and -DQA1 (p = 0.002) loci as possible risk factors for endemic KS. Our study provides additional understanding of genetic determinants of KS and their implications in disease pathogenesis. Further validation of these findings is needed to define the functional relevance of these associations.

IgA antibodies targeting Epstein-Barr virus (EBV) have been proposed for screening for nasopharyngeal carcinoma (NPC). However, methods differ, and the antigens used in these assays differ considerably between laboratories. To enable formal comparisons across a range of established EBV serology assays, we created a panel of 66 pooled serum samples and 66 pooled plasma samples generated from individuals with a broad range of IgA antibody levels. Aliquots from these panels were distributed to six laboratories and were tested by 26 assays measuring antibodies against VCA, EBNA1, EA-EBNA1, Zta, or EAd antigens. We estimated the correlation between assay pairs using Spearman coefficients (continuous measures) and percentages of agreement (positive versus negative, using predefined positivity cutoffs by each assay developer/manufacturer). While strong correlations were observed between some assays, considerable differences were also noted, even for assays that targeted the same protein. For VCA-IgA assays in serum, two distinct clusters were identified, with a median Spearman coefficient of 0.41 (range, 0.20 to 0.66) across these two clusters. EBNA1-IgA assays in serum grouped into a single cluster with a median Spearman coefficient of 0.79 (range, 0.71 to 0.89). Percentages of agreement differed broadly for both VCA-IgA (12% to 98%) and EBNA1-IgA (29% to 95%) assays in serum. Moderate-to-strong correlations were observed across assays in serum that targeted other proteins (correlations ranged from 0.44 to 0.76). Similar results were noted for plasma. We conclude that standardization of EBV serology assays is needed to allow for comparability of results obtained in different translational research studies across laboratories and populations.

Primary effusion lymphoma (PEL) is an aggressive HIV-associated lymphoma with a relatively poor prognosis in the era of effective HIV therapy. Kaposi sarcoma herpesvirus (KSHV) is the etiologic agent, and ∼80% of tumors are coinfected with Epstein-Barr virus (EBV). A better understanding of how KSHV-related immune dysregulation contributes to the natural history of PEL will improve outcomes. Twenty patients with PEL diagnosed between 2000 and 2013, including 19 treated with modified infusional etoposide, vincristine, and doxorubicin with cyclophosphamide and prednisone (EPOCH), were identified. We compared their clinical, virologic, and immunologic features vs 20 patients with HIV-associated diffuse large B-cell lymphoma and 19 patients with symptomatic interleukin (IL)-6 related KSHV-associated multicentric Castleman disease. Survival analyses of treated patients with PEL were then performed to identify prognostic factors and cancer-specific mortality. Compared with HIV-associated diffuse large B-cell lymphoma, PEL was associated with significant hypoalbuminemia (P < .0027), thrombocytopenia (P = .0045), and elevated IL-10 levels (P < .0001). There were no significant differences in these parameters between PEL and KSHV-associated multicentric Castleman disease. Median overall survival in treated patients with PEL was 22 months, with a plateau in survival noted after 2 years. Three-year cancer-specific survival was 47%. EBV-positive tumor status was associated with improved survival (hazard ratio, 0.27; P = .038), and elevated IL-6 level was associated with inferior survival (hazard ratio, 6.1; P = .024). Our analysis shows that IL-6 and IL-10 levels contribute to the natural history of PEL. Inflammatory cytokines and tumor EBV status are the strongest prognostic factors. Pathogenesis-directed first-line regimens are needed to improve overall survival in PEL.

We recently showed that the interaction between Kaposi's sarcoma-associated herpesvirus (KSHV) tegument proteins ORF33 and ORF45 is crucial for progeny virion production, but the exact functions of KSHV ORF33 during lytic replication were unknown (J. Gillen, W. Li, Q. Liang, D. Avey, J. Wu, F. Wu, J. Myoung, and F. Zhu, J Virol 89: 4918–4931, 2015, http://dx.doi.org/10.1128/JVI.02925-14 ). Therefore, here we investigated the relationship between ORF33 and ORF38, whose counterparts in both alpha- and betaherpesviruses interact with each other. Using specific monoclonal antibodies, we found that both proteins are expressed during the late lytic cycle with similar kinetics and that both are present in mature virions as components of the tegument. Furthermore, we confirmed that ORF33 interacts with ORF38. Interestingly, we observed that ORF33 tightly associates with the capsid, whereas ORF38 associates with the envelope. We generated ORF33-null, ORF38-null, and double-null mutants and found that these mutants apparently have identical phenotypes: the mutations caused no apparent effect on viral gene expression but reduced the yield of progeny virion by about 10-fold. The progeny virions also lack certain virion component proteins, including ORF45. During viral lytic replication, the virions associate with cytoplasmic vesicles. We also observed that ORF38 associates with the membranes of vesicles and colocalizes with the Golgi membrane or early endosome membrane. Further analyses of ORF33/ORF38 mutants revealed the reduced production of virion-containing vesicles, suggesting that ORF33 and ORF38 are involved in the transport of newly assembled viral particles into cytoplasmic vesicles, a process important for viral maturation and egress. IMPORTANCE Herpesvirus assembly is an essential step in virus propagation that leads to the generation of progeny virions. It is a complicated process that depends on the delicate regulation of interactions among virion proteins. We previously revealed an essential role of ORF45-ORF33 binding for virus assembly. Here, we report that ORF33 and its binding partner, ORF38, are required for infectious virus production due to their important role in the tegumentation process. Moreover, we found that both ORF33 and ORF38 are involved in the transportation of virions through vesicles during maturation and egress. Our results provide new insights into the important roles of ORF33 and ORF38 during viral assembly, a process critical for virus propagation that is intimately linked to KSHV pathobiology.

Background Epstein-Barr virus (EBV) and Kaposi sarcoma-associated herpesvirus (KSHV) are transmitted via saliva, but factors associated with salivary shedding are unknown. Methods We measured the DNA load of both viruses in saliva specimens collected from approximately 500 Ugandan mothers and their 6-year-old children, testing all participants for EBV and KSHV-seropositive individuals for KSHV. Results EBV and KSHV were shed by 72% and 22% of mothers, respectively, and by 85% and 40% of children, respectively; boys were more likely than girls to shed KSHV (48% vs 30%) but were equally likely to shed EBV. Children shed more KSHV and EBV than mothers, but salivary loads of EBV and KSHV were similar. KSHV shedding increased with increasing anti-KSHV (K8.1) antibodies in mothers and with decreasing antimalarial antibodies both in mothers and children. Among mothers, 40% of KSHV shedders also shed EBV, compared with 75% of KSHV nonshedders; among children, EBV was shed by 65% and 83%, respectively. Conclusions In summary, in this population, individuals were more likely to shed EBV than KSHV in saliva. We identified several factors, including child's sex, that influence KSHV shedding, and we detected an inverse relationship between EBV and KSHV shedding, suggesting a direct or indirect interaction between the two viruses.

We examined anemia and malaria as risk factors for Kaposi sarcoma-associated herpesvirus (KSHV) seropositivity and antibody levels in a long-standing rural Ugandan cohort, in which KSHV is prevalent. Samples from 4134 children, aged 1-17 years, with a sex ratio of 1:1, and 3149 adults aged 18-103 years, 41% of whom were males, were analyzed. Among children, malaria infection was associated with higher KSHV prevalence (61% vs 41% prevalence among malaria infected and uninfected, respectively); malaria was not assessed in adults. Additionally, lower hemoglobin level was associated with an increased prevalence of KSHV seropositivity, both in children and in adults.

Kaposi's sarcoma-associated herpesvirus (KSHV) is the causative agent of several human cancers such as Kaposi's sarcoma (KS), which represents the most common AIDS-associated malignancy that lacks effective treatment options. Despite its clear role in AIDS malignancies, the fact that only a small set of KSHV-infected patients will eventually develop these tumors implies that additional co-factors are required for the development of KSHV-related cancers. In the current study, we demonstrate for the first time that KSHV de novo infection or viral latent proteins are able to transactivate human endogenous retrovirus K (HERV-K) through a variety of cellular signaling pathways and transcriptional factors. Moreover, we found that HERV-K transactivation, particularly activation of its encoded oncogenic NP9 protein, plays an important role in KSHV pathogenesis and tumorigenesis in vitro and in vivo. Our data provide innovative insights into the mechanisms of HERV-K transactivation contributing to viral oncogenesis, which may represent a promising target for KS treatment.

SummaryInvading viral DNA can be recognized by the host cytosolic DNA sensor, cyclic GMP-AMP (cGAMP) synthase (cGAS), resulting in production of the second messenger cGAMP, which directs the adaptor protein STING to stimulate production of type I interferons (IFNs). Although several DNA viruses are sensed by cGAS, viral strategies targeting cGAS are virtually unknown. We report here that Kaposi’s sarcoma-associated herpesvirus (KSHV) ORF52, an abundant gammaherpesvirus-specific tegument protein, subverts cytosolic DNA sensing by directly inhibiting cGAS enzymatic activity through a mechanism involving both cGAS binding and DNA binding. Moreover, ORF52 homologs in other gammaherpesviruses also inhibit cGAS activity and similarly bind cGAS and DNA, suggesting conserved inhibitory mechanisms. Furthermore, KSHV infection evokes cGAS-dependent responses that can limit the infection, and an ORF52 null mutant exhibits increased cGAS signaling. Our findings reveal a mechanism through which gammaherpesviruses antagonize host cGAS DNA sensing.

OBJECTIVE: To determine the effect of the introduction of combination antiretroviral treatment (cART) in the HIV-1 infected US population on the epidemiology of Kaposi’s sarcoma herpesvirus (KSHV). DESIGN, SETTING AND PARTICIPANTS: We investigated the epidemiology of KSHV in 5022 HIV-1-infected, antiretroviral naïve US persons participating in six AIDS Clinical Trials Group (ACTG) randomized clinical trials, and followed in a long-term cohort study. We tested the first and last available sera of each participant for antibodies to KSHV K8.1 and ORF73. MAIN OUTCOME MEASURES: We studied prevalence and incidence of KSHV infection, incidence of Kaposi’s sarcoma (KS), and overall survival. RESULTS: KSHV prevalence was 38.1% (95% CI 36.8-39.5%). Male gender, Caucasian race, age between 30 and 49, residence in North-eastern or Western US, and enrolment after 2001 were independently associated with prevalent infection. KSHV incidence was 4.07/100 person/years (95%CI 3.70-4.47). Male gender, Caucasian race, age below 30, and enrolment after 2001 were associated with incident infection. CD4 count increase following cART was associated with lower risk. KS incidence was 104.05/100,000 person/years (95% CI 71.17-146.89). Higher baseline CD4 count, but not increase in CD4 count after cART, was associated with lower hazard of KS. Randomized assignment of protease inhibitors was not associated with better KSHV outcomes. CONCLUSIONS: HIV-1 infected individuals, in particular Caucasian men, remain at significant risk for KSHV co- infection and KS. Thus, optimal management of HIV-1 infection should continue to include vigilance for manifestations of KSHV co-infection, including KS. Video abstract at http://ncifrederick.cancer.gov/services/spgm/filedownload/pdsDownload.aspx?id=c472803f-1d66–4391-b18b-24e367b6b190

The known human tumor viruses include the DNA viruses Epstein-Barr virus (EBV), Kaposi sarcoma herpesvirus (KSHV), Merkel cell polyomavirus (MCPyV), human papillomavirus (HPV), and hepatitis B virus (BV). RNA tumor viruses include human T-cell lymphotrophic virus type 1 (HTLV-1) and hepatitis C virus (HCV). The serological identification of antigens/antibodies in serum is a rapidly progressing field with utility for both scientists and clinicians. Serology is useful for conducting seroepidemiology studies and to inform on the pathogenesis and host immune response to a particular viral agent. Clinically, serology is useful for diagnosing current or past infection and for aiding in clinical management decisions. Serology is useful for screening blood donations for infectious agents and for monitoring the outcome of vaccination against these viruses. Serodiagnosis of human tumor viruses has improved in recent years with increased specificity and sensitivity of the assays, as well as reductions in cost and the ability to assess multiple antibody/antigens in single assays. Serodiagnosis of tumor viruses plays an important role in our understanding of the prevalence and transmission of these viruses and ultimately in the ability to develop treatments/preventions for these globally important diseases.

Prior studies of T-cell responses to KSHV have included relatively few participants and focused on relatively few KSHV antigens. To provide a more comprehensive analysis, we investigated T-cell responses to the whole KSHV proteome using IFN-γ ELISpot. Using ∼7,500 overlapping 15mer peptides we generated one to three peptide pools for each of the 82 KSHV ORFs. IFN-γ ELISpot analysis of PBMCs from 19 patients with a history of KSHV-associated disease and 24 healthy donors (11 KSHV seropositive) detected widely varied responses. Fifty six of the 82 ORFs were recognized by at least one individual but there was little overlap between participants. Responses to at least one ORF pool were observed in all 19 patients and in 7 seropositive donors. Four seropositive donors and 10 seronegative donors had no detectable responses while 3 seronegative donors had weak responses to one ORF. Patients recognised more ORFs than the donors (p=0.04) but the response intensity (spot forming units: SFU per million cells) was similar in the two groups. In four of the responding donors, individual peptides eliciting the predominant responses were identified: three donors responded to only one peptide per ORF, while one recognized five. Using intracellular cytokine staining in four participant samples, we detected peptide-induced IFN-γ, MIP1-β, and TNF-α as well as CD107a degranulation, consistent with multifunctional effector responses in CD8+ and CD4+ T cells. Sequence analysis of TCRs present in peptide specific T-cell clones generated from two participants showed both mono- and multi-clonotypic responses. Finally, we molecularly cloned the KSHV specific TCRs and incorporated the sequences into retroviral vectors to transfer the specificities to fresh donor cells for additional studies. This study suggests that KSHV infected individuals respond to diverse KSHV antigens, consistent with a lack of shared immunodominance and establishes useful tools to facilitate KSHV immunology studies.

Objective Unlike other herpes viruses, Kaposi's sarcoma-associated herpes virus (KSHV) is not ubiquitous worldwide and is most prevalent in sub-Saharan Africa. The reasons for this are unclear. As part of a wider investigation of factors that facilitate transmission in Uganda, a high prevalence country, we examined the association between antimalaria antibodies and seropositivity against KSHV. Methods Antibodies against P. falciparum merozoite surface protein (PfMSP)-1, P. falciparum apical membrane antigen (PfAMA)-1 and KSHV antigens (ORF73 and K8.1) were measured in samples from 1164 mothers and 1227 children. Results Kaposi's sarcoma-associated herpes virus seroprevalence was 69% among mothers and 15% children. Among mothers, KSHV seroprevalence increased with malaria antibody titres: from 60% to 82% and from 54% to 77%, comparing those with the lowest and highest titres for PfMSP-1 and PfAMA-1, respectively (P < 0.0001). Among children, only antibodies to PfAMA-1 were significantly associated with KSHV seropositivity, (P < 0.0001). In both mothers and children, anti-ORF73 antibodies were more strongly associated with malaria antibodies than anti-K8.1 antibodies. Conclusion The association between malaria exposure and KSHV seropositivity suggests that malaria is a cofactor for KSHV infection or reactivation.

HIV-associated Kaposi's sarcoma (KS) is a public health challenge in sub-Saharan Africa since both the causative agent, Kaposi's sarcoma associated-herpesvirus (KSHV), and the major risk factor, HIV, are prevalent. In a nested case-control study within a long-standing clinical cohort in rural Uganda, we used stored sera to examine the evolution of antibody titres against the KSHV antigens K8.1 and latency-associated nuclear antigen (LANA) among 30 HIV-infected subjects who subsequently developed HIV-related KS (cases) and among 108 matched HIV/KSHV coinfected controls who did not develop KS. Throughout the 6 years prior to diagnosis, antibody titres to K8.1 and LANA were significantly higher among cases than controls (p < 0.0001), and titres increased prior to diagnosis in the cases. K8.1 titres differed more between KS cases and controls, compared to LANA titres. These differences in titre between cases and controls suggest a role for lytic viral replication in the pathogenesis of HIV-related KS in this setting.

Background Individuals co-infected with Kaposi's sarcoma herpesvirus (KSHV) and Human Immunodeficiency Virus (HIV) are at greatly increased risk of developing Kaposi's sarcoma (KS). The objective of the current analysis is to identify risk cofactors for KS among HIV-positive individuals. Methods We conducted a case-control study of KS in Cameroon on 161 HIV-positive and 14 HIV-negative cases and 680 HIV-positive and 322 HIV-negative controls. Participants answered a physician-administered questionnaire and provided blood and saliva specimens. Antibodies against KSHV lytic, K8.1, and latent, ORF73, antigens were measured by ELISA to determine KSHV serostatus. Conditional logistic regression was performed to determine multivariate odds ratios (OR) and 95% confidence intervals (CI) for risk factors associated with KS among HIV-positive cases and controls. Results Overall, 98% (158) of HIV-positive cases, 100% (14) of HIV-negative cases, 81% (550) of HIV-positive controls, and 80% (257) of HIV-negative controls were KSHV seropositive. Risk factors for KS among HIV-positive individuals included KSHV seropositivity (OR = 9.6; 95% CI 2.9, 31.5), non-use of a mosquito bed net (OR 1.9; 95% CI 1.2, 2.9), minority ethnicity (OR = 3.1; 95% CI 1.1, 9.3), treatment from a traditional healer (OR = 2.3; 95% CI 1.5, 3.7), history of transfusion (OR = 2.4; 95% CI 1.5, 3.9), and family history of cancer (OR = 1.9; 95% CI 1.1, 3.1). Conclusion KSHV seroprevalence of ≥80% indicates a high prevalence in the general population in Cameroon. Among HIV-positive individuals, the strong association of KS with non-use of mosquito nets and treatment from traditional healers are compelling findings, consistent with recently reported data from East Africa.

Background: Determinants of Kaposi sarcoma–associated herpesvirus (KSHV) seropositivity among children living in sub-Saharan African populations where infection is endemic are not well understood. Local environmental factors, including other infectious agents, may be key. Methods: Within the context of a well-characterized birth cohort, we examined associations between various factors and antibodies against KSHV, measured in stored plasma samples from 1823 mother–child pairs in Entebbe, Uganda. Results: Seroprevalence increased with increasing age of the child (P = 0.0003) and was higher among those with KSHV seropositive mothers than in those without (12% vs 9%; odds ratio: 1.4, 95% confidence interval: 1.1 to 2.0). It was also higher among children with HIV infection (29% vs 10%; odds ratio: 3.1, 95% confidence interval: 1.2 to 8.3) or malaria parasitemia (30% vs 10%; odds ratio: 4.1, 95% confidence interval: 2.4 to 7.0) than in children without. These associations were not explained by socioeconomic status. Conclusions: The finding that KSHV serostatus is associated with malaria parasitemia in children is novel. In a country endemic for KSHV, malaria may be a cofactor for KSHV infection or reactivation among children.

Background:\ud Immune modulation by parasites may influence susceptibility to bacteria and viruses. We examined the association between current parasite infections, HIV and syphilis (measured in blood or stool samples using standard methods) and antibodies against Kaposi's sarcoma herpesvirus (KSHV), measured by ELISA, in 1915 stored plasma samples from pregnant women in Entebbe, Uganda.\ud \ud Results:\ud Seroprevalence of KSHV was higher in women with malaria parasitaemia (73% vs 60% p = 0.01), hookworm (67% vs 56% p = 0.001) and Mansonella perstans (69% vs 59% p = 0.05); seroprevalence increased with increasing intensity of hookworm infection (p < 0.001[trend]). No associations were found for HIV, five other parasites or active syphilis. These effects were not explained by socioeconomic status or education.\ud \ud Conclusions:\ud Specific parasite infections are associated with presence of antibodies against KSHV, perhaps mediated via their effect on immune function.

Objective. Human T-cell lymphotropic virus type I (HTLV-I) infection in childhood is believed to play an important role in risk for adult T-cell leukemia/lymphoma. Although HTLV-I is known to be associated with infective dermatitis in childhood, other HTLV-I-associated morbidity in children has not been well studied. We sought to determine the HTLV-I-associated health effects in Jamaican children. Methods. We compared incidence rates of several health outcomes in 28 HTLV-I-infected and 280 uninfected children clinically followed from age 6 weeks to a maximum of 10 years. Cox proportional hazards regression analysis was used to analyze these prospectively collected data, adjusting for confounding effects of other variables as necessary. Results. HTLV-I-infected children had significantly higher incidence rates of seborrheic dermatitis (rate ratio [RR] = 4.8, 95% confidence interval [CI] = 1.9–12.5), eczema (RR = 3.1, CI = 1.2–7.9) and persistent hyperreflexia (RR = 3.7, CI = 1.6–8.2). Additionally, HTLV-I infected children had increased rates of severe anemia (RR = 2.5, CI = 0.8–7.9) and abnormal lymphocytes (RR = 2.4, CI = 0.8–7.6) that were of borderline statistical significance. Conclusions. Our study suggests that HTLV-I-associated skin diseases of childhood may include seborrheic dermatitis and eczema. Additionally, these data suggest that persistent hyperreflexia of the lower limbs may be an early sign of HTLV-I-associated neurologic involvement in children. Expansion and continued clinical observation of this cohort would be valuable.

Background: Hepatitis C virus (HCV) infection is a significant global health issue because it is widespread and persistent and can cause serious liver diseases. Objectives: The aim of this study is to estimate HCV prevalence in women from the general population in different geographical areas worldwide and to assess the potential role of sexual behaviour in the virus transmission. Study design: Each participating centre recruited a random sample of women from the general population aged from less than 20 to more than 75 years. The study included 8130 women from 8 countries with information on sociodemographic factors, reproductive and sexual behaviour, smoking habit and HPV DNA through individual interviews. A blood sample was also collected to perform serological tests. We estimated the prevalence ratios associated to HCV to evaluate the effect of sexual behaviour in viral transmission. Results: Women were reactive to a minimum of two HCV antigens, including at least one non structural protein were considered as positive (33% of the samples were classified as positive, 40% as negative, and 27% as indeterminate (N = 402), that were considered as not positive). The age-adjusted HCV seroprevalence varied significantly by regions (0.3% in Argentina to 21.1% in Nigeria). We found no association between HCV prevalence and age, educational level, smoking habit and any of the available variables for sexual behaviour and reproductive history. Conclusions: This large study showed heterogeneous distribution of HCV seroprevalence in female and provides evidence of the null impact of sexual behaviour in HCV transmission. (C) 2015 Elsevier B.V. All rights reserved.