Thesis (Ph. D.)--University of Virginia, 1998. / Includes bibliographical references (p. 157-170). Also available online through Digital Dissertations.

No description available.

Herpes Simplex Virus (HSV) requires the host cell secretory apparatus for the maturation and egress of newly synthesized viral particles. Not only do viral glycoproteins rely on the host ER and Golgi compartments for their proper processing, it is believed that enveloped particles are transported through these same organelles for their export out of the cells. Brefeldin A (BFA) is a compound that induces retrograde movement of material from the Golgi apparatus to the ER and causes the disassembly of the Golgi complex. In this study, the effects of BFA on the propagation of HSV-1 in infected cells were examined. Release of viral particles from infected cells was inhibited by as little as 1 µg/ml BFA. Further analysis revealed that BFA did not affect the normal assembly of viral nucleocapsids, but did block the movement of newly-enveloped particles from the nucleus into the cytoplasm. Naked nucleocapsids were found in the cytoplasm of infected cells treated with BFA, however, these particles were neither infectious, nor were they released from the cells. Although BFA altered the distribution of viral glycoproteins in infected cells, this alteration was reversed within 2 hours after the removal of BFA. In contrast, the BFA-induced blockage to viral release was not fully reversed after BFA was removed and cells were allowed to recover in fresh medium for 3 hours. These findings indicate that the BFA-induced retrograde movement of material from the Golgi complex to the ER early in infection arrests the ability of the host cell to support the maturation and egress of enveloped viral particles. Furthermore, exposure of infected cells to BFA during the exponential release phase of the viral life cycle can cause irreversible damage to the egressing particles. This suggests that productive growth of HSV-1 in infected cells relies on a series of events that, once disrupted by agents such as BFA, cannot be easily reconstituted. / Science, Faculty of / Microbiology and Immunology, Department of / Graduate

The aim of this work was to purify type-specific glycoproteins from herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) for diagnostic use. The most likely candidate for a type-specific glycoprotein of HSV-1 is glycoprotein C (gC), although it has recently been shown to contain some type-common antigenic determinants. HSV-1 and HSV-2 were produced in BHK-21 cells and labelled with either (³H)-glucosamine ((³H)-gln) or a mixture of (¹⁴C)-amino acids ((¹⁴C)-aa). Analysis of the radiolabelled products by analytical sodium dodecyl sulphate polyacrylamide gel electrophoresis (SOS-PAGE) and autoradiography revealed that in the HSV-1 infected cells the radiolabelled components were incorporated into viral specific proteins only, whereas in the HSV-2 infected cells they were incorporated into host cell proteins as well as viral proteins. Preparative polyacrylamide gel electrophoresis (Prep-PAGE) was used as an initial step in separating HSV-1 infected cell proteins labelled with (³H)-gln. Two cycles of Prep-PAGE were sufficient to produce solutions containing either glycoprotein B ( gB) or glyco- protein C (gC), which were free of other HSV-1 glycoproteins. However, these solutions still contained a number of non-glycosylated proteins. Two different techniques were utilized to remove the non-glycosylated proteins from the glycoprotein solutions. Hydroxylapatite (HAUltrogel) chromatography in the presence of sodium dodecyl sulphate (SDS-HTP) did not separate the different HSV-1 glycoproteins and was not satisfactory for removing the non-glycosylated proteins. Gel-bound lectin affinity chromatography using wheat germ lectin and Helix pomatia lectin was not successful in purifying the glycoproteins because the glycoproteins which bound to the lectins could not be eluted under normal conditions. Difficulties encountered in eluting the HSV-1 glycoproteins from the lectins may have been due to the sodium dodecyl sulphate (SOS) in which the proteins were solubilized. For this reason, the gelbound lectin affinity chromatography was repeated using HSV-1 membrane proteins solubilized in a non-ionic detergent, Triton X-100. Using material prepared in this way, several HSV-1 glycoproteins were bound by wheat germ lectin and eluted under normal conditions to yield glycoproteins which were purified with respect to nonglycosylated proteins.

Thesis (Ph.D. (Public Health))--University of the Witwatersrand, Faculty of Health Sciences, 2012 / Factors associated with the transmission of Kaposi’s sarcoma-associated herpesvirus (KSHV) are inconclusive. In countries where KS and KSHV are confined to men who have sex with other men (MSM), KSHV is associated with sexual risk factors. In countries where KSHV is endemic, it affects adults and children of all ages and irrespective of sexual orientation, suggesting the existence of non-sexual risk factors for KSHV infection. In this thesis, three distinct cross sectional studies aiming to define the seroprevalence of KSHV in South African populations and to identify plausible risk factors for KSHV infection were undertaken. The studies measured KSHV seropositivity in relation to sociodemographic factors and HIV status. In children, factors associated with horizontal mother to child transmission were also explored. In adults KSHV seropositivity was also measured in relation to sexually transmitted infections and/or measures of sexual behaviour. Calculated risk factors were expressed as odds ratios (95% confidence interval) for KSHV. Methods Mother to Child KSHV seroepidemiology Study: KSHV seroprevalence (reactive to either lytic K8.1 or latent Orf73) was measured in 1287 children and their 1179 biological mothers. Association between KSHV seropositivity in children was measured against KSHV seropositivity and HIV status of their mothers. KSHV seroepidemiology in women attending antenatal clinics: Antibodies to KSHV lytic K8.1 and latent Orf73 antigens were tested in 1740 pregnant women attending antenatal clinics in South Africa in 2001. Information on HIV and syphilis serology, age, education, residential area, gravidity, and parity was anonymously linked to evaluate risk factors for KSHV seropositivity. Clinics were grouped by municipal regions and their proximity to the two main river catchments defined. Carletonville Community KSHV seroepidemiology Study: Sera from 2103 South African individuals (862 miners, 95 sex workers, 731 female and 415 male township residents) were tested for antibodies to KSHV lytic K8.1 and latent Orf73, HIV gonococcus, herpes simplex virus type 2 (HSV-2), syphilis and chlamydia. Information on social, demographic and highrisk sexual behaviour was linked to laboratory data. Results Mother to Child KSHV seroepidemiology Study: KSHV seroprevalence (reactive to either lytic K8.1 or latent Orf73) was 15.9% (204 of 1287 subjects) in children and 29.7% (350 of 1179 subjects) in mothers. The risk of KSHV seropositivity was significantly higher in children of KSHV seropositive mothers compared with those of KSHV-seronegative mothers. The HIV status of mothers was marginally associated with an increased risk of KSHV seropositivity in their children (AOR = 1.6, 95% CI: 1.0 to 2.6; P = 0.07). KSHV seroprevalence was significantly higher in HIV-infected subjects (P = 0.0005), and HIV-infected subjects had significantly higher lytic and latent KSHV antibody levels than HIV-negative subjects. KSHV seroepidemiology in women attending antenatal clinics: KSHV seroprevalence was nearly twice that of HIV (44.6% vs. 23.1%). HIV and syphilis seropositivity was 12.7% and 14.9% respectively in women without KSHV, and 36.1% and 19.9% respectively in those with KSHV. Women who were KSHV seropositive were 4 times more likely to be HIV positive than those who were KSHV seronegative (AOR 4.1 95%CI: 3.4 - 5.7). Although, women with HIV infection were more likely to be syphilis seropositive (AOR 1.8 95%CI: 1.3 - 2.4), no association between KSHV and syphilis seropositivity was observed. Those with higher levels of education had lower levels of KSHV seropositivity compared to those with lower education levels. KSHV seropositivity showed a heterogeneous pattern of prevalence in some localities. Carletonville Community KSHV seroepidemiology Study: Overall KSHV and HIV prevalences were 47.5 and 40%, respectively (P<0.43). The risk of HIV infection was highest in sex workers followed by female residents and miners, compared with male residents (P<0.001). HSV-2 infection was highly prevalent (66%) and lower, but still substantial, prevalence (6–8%) was observed for other sexually transmitted infections (STI). No significant difference in KSHV infection was observed among the residential groups (P>0.05). KSHV was not associated with any of the STI or any measures of sexual behaviour. Conclusion The findings of these three studies contribute substantially to global KSHV seroepidemiology and show that in Southern African settings KSHV is associated with non-sexual mode of transmission. Firstly KSHV is common in very young children up to ten years of age and increases with age until adulthood. The high prevalence of KSHV in the South African populations remained evident in all populations. In children, the risk of acquisition of KSHV was higher among children of KSHV-seropositive mothers than if the mother was KSHV negative. The association between KSHV and HIV was also noted in the study of pregnant women attending antenatal clinics and in the mother to child study. However this association was not evident in the Carletonville population where both KSHV and HIV were highly prevalent. In both the adult studies the lack of association between KSHV and syphilis was evident. KSHV infection was also not associated with other sexually transmitted infections and measures of sexual behaviour. As expected, the pattern of HIV and STI in sex workers suggests high rates of high-risk sexual behaviour in this population; however KSHV seropositivity was the same amongst sexworkers and all the other community groups. This pattern of the lack of association with high-risk sexual behaviour, particularly in sex workers and with any markers of STI strongly suggests that the sexual mode does not play a significant role in KSHV transmission in this South African population. This may also suggest that KSHV transmission may involve geographical and cultural factors other than sexual transmission.

Thesis (doctoral)--University of Göteborg, 1993. / Added t.p. with thesis statement inserted. Includes bibliographical references.

Thesis (doctoral)--University of Göteborg, 1993. / Added t.p. with thesis statement inserted. Includes bibliographical references.

Thesis (Ph.D. (Microbiology))--Mahidol University, 1978.

Human Herpes Simplex viruses and Simian Herpes Simplex viruses share a high degree of genome homology, but despite this, important differences arise when the viruses are compared at the level of gene expression and virulence in non-host primates. In Human Herpes viruses (HSV-1 and HSV-2); 5 genes (RL02, US01, RS01, UL54 and US12) are expressed with an immediate early kinetics, i.e. their transcriptional activation does not require de novo synthesis of host or viral factors. The five immediate early (IE) genes regulate the cascade of expression of the other early and late HSV genes. Literature indicates that in HSV-1 infections, ICP4, ICP27 and to a lesser extent, ICP0, are mandatory for the full expression of the early and late gene classes. In contrast, our data on the Simian simplexviruses SA8, HVP-2 and B virus indicate that ICP0 (RL2) is the only gene with true IE kinetics. It is possible that in Simian Herpes viruses, ICP0 is necessary for the expression of all other viral genes, and to test this hypothesis I have cloned and expressed in Vero cells the ICP0 protein for the simian simplexvirus SA8 and studied its effect on the SA8 genes that are homologous to the immediate early genes in HSV. Results demonstrate that ICP0 does not appear to be sufficient to activate the transcription of the other IE genes but it is likely that ICP0 functionality is a necessary component in the activation process.

Human Herpes Simplex viruses and Simian Herpes Simplex viruses share a high degree of genome homology, but despite this, important differences arise when the viruses are compared at the level of gene expression and virulence in non-host primates. In Human Herpes viruses (HSV-1 and HSV-2); 5 genes (RL02, US01, RS01, UL54 and US12) are expressed with an immediate early kinetics, i.e. their transcriptional activation does not require de novo synthesis of host or viral factors. The five immediate early (IE) genes regulate the cascade of expression of the other early and late HSV genes. Literature indicates that in HSV-1 infections, ICP4, ICP27 and to a lesser extent, ICP0, are mandatory for the full expression of the early and late gene classes. In contrast, our data on the Simian simplexviruses SA8, HVP-2 and B virus indicate that ICP0 (RL2) is the only gene with true IE kinetics. It is possible that in Simian Herpes viruses, ICP0 is necessary for the expression of all other viral genes, and to test this hypothesis I have cloned and expressed in Vero cells the ICP0 protein for the simian simplexvirus SA8 and studied its effect on the SA8 genes that are homologous to the immediate early genes in HSV. Results demonstrate that ICP0 does not appear to be sufficient to activate the transcription of the other IE genes but it is likely that ICP0 functionality is a necessary component in the activation process.