Caroline T. Tiemessen

Genetic diversity and molecular epidemiology of respiratory syncytial virus over four consecutive seasons in South Africa: identification of new subgroup A and B genotypes.

The molecular epidemiology of respiratory syncytial virus (RSV) was studied over four consecutive seasons (1997-2000) in a single tertiary hospital in South Africa: 225 isolates were subgrouped by RT-PCR and the resulting products sequenced. Subgroup A predominated in two seasons, while A and B co-circulated approximately equally in the other seasons. The nucleotide sequences of the C-terminal of the G-protein were compared to sequences representative of previously defined RSV genotypes. South African subgroup A and subgroup B isolates clustered into four and five genotypes respectively. One new subgroup A and three new subgroup B genotypes were identified. Different genotypes co-circulated in every season. Different circulation patterns were identified for group A and B isolates. Subgroup A revealed more variability and displacement of genotypes while subgroup B remained more consistent.

Human Immunodeficiency Virus (HIV)–Specific Cellular Immune Responses in Newborns Exposed to HIV In Utero

Significant immunological changes are associated with intrauterine human immunodeficiency virus (HIV) encounter among uninfected infants of HIV-infected mothers. Peripheral blood cells of more than one-third of these exposed-uninfected infants proliferate and produce IL-2 after stimulation with HIV, and HIV-specific CD4+ T helper cell responses can be quantified in nearly all when sensitive intracellular cytokine assays are used. HIV-specific CD8+ cytotoxic T lymphocyte responses can be elicited in some, although less frequently. It is difficult to demonstrate that these responses are components of protective immunity and not simply epiphenomena of exposure. However, HIV-specific responses are associated with lack of infection, even with prolonged reexposure through breast-feeding. Elevations in nonspecific markers of immune activation provide further corroboration, as do similar findings in adults, consistent across all known routes of HIV transmission. Many questions remain, but much can be learned from this special population that may be informative for development of effective immunity in response to HIV vaccines.

African infants’ CCL3 gene copies influence perinatal HIV transmission in the absence of maternal nevirapine

Background:Individuals with more copies of CCL3L1 (CCR5 ligand) than their population median have been found to be less susceptible to HIV infection. We investigated whether maternal or infant CCL3L1 gene copy numbers are associated with perinatal HIV transmission when single-dose nevirapine is given for prevention. Method:A nested case–control study was undertaken combining data from four cohorts including 849 HIV-infected mothers and their infants followed prospectively in Johannesburg, South Africa. Access to antiretroviral drugs for the prevention of perinatal transmission differed across the cohorts. Maternal and infant CCL3L1 gene copy numbers per diploid genome (pdg) were determined by real-time polymerase chain reaction for 79 out of 83 transmitting pairs (∼10% transmission rate) and 235 randomly selected non-transmitting pairs. Results:Higher numbers of infant, but not maternal, CCL3L1 gene copies were associated with reduced HIV transmission (P = 0.004) overall, but the association was attenuated if mothers took single-dose nevirapine or if the maternal viral load was low. Maternal nevirapine was also associated with reduced spontaneously released CCL3 (P = 0.007) and phytohemagglutinin-stimulated CCL3 (P = 0.005) production in cord blood mononuclear cells from uninfected infants. Conclusion:We observed a strong association between higher infant CCL3L1 gene copies and reduced susceptibility to HIV in the absence of maternal nevirapine. We also observed a reduction in newborn CCL3 production with nevirapine exposure. Taken together, we hypothesize that nevirapine may have direct or indirect effects that partly modify the role of the CCR5 ligand CCL3 in HIV transmission, obscuring the relationship between this genetic marker and perinatal HIV transmission.

Cutting Edge: Unusual NK Cell Responses to HIV-1 Peptides Are Associated with Protection against Maternal-Infant Transmission of HIV-1

Most infants exposed to HIV-1 in utero and at delivery do not acquire infection. We show that mothers and infants who have CD3-negative cells that respond to HIV-1 peptides are substantially less likely to transmit and acquire infection, respectively. The CD3-negative cells, shown to be NK cells, respond with remarkable specificity and high magnitude to HIV-1 peptides from Env (envelope) and Reg (regulatory) protein regions, as measured by a whole blood intracellular cytokine assay only in the context of HIV-1 infection or exposure. These findings identify an important new measure of protective immunity to HIV-1 that highlights the importance of innate immunity in preventing the establishment of HIV-1 infection.

Evolution and diversity of HIV-1 in Africa--a review.

The HIV/AIDS pandemic represents a major development crisis for the African continent, which is the worst affected region in the world. Currently, almost 30 of the 42 million people infected with HIV worldwide live in Africa. AIDS in humans is caused by two lentiviruses, HIV-1 and HIV-2, which entered the human population by zoonotic transmissions from at least two different African primate species. Extensive phylogenetic analyses of partial and full-length genome sequences have helped to gain insights into the evolutionary biology and population dynamics of HIV. One of the major characteristics of HIV is its rapid evolution, which has resulted in substantial genetic diversity amongst different isolates, the majority of which are represented in Africa. Genetic variability of HIV and any consequent phenotypic variation poses a significant challenge to disease control and surveillance in different geographic regions of Africa. This review focuses on the origins and evolution of HIV, current classification and diversity of HIV isolates in Africa and provides an extensive account of the geographic distribution of HIV types, groups, and subtypes in each of the 49 African countries. Numerous epidemiological studies have provided a picture of HIV distribution patterns in most countries in Africa, and these show increasing evidence of the importance of HIV-1 recombinants. In particular, this review highlights that our current understanding of HIV distribution in Africa is incomplete and inadequately represents the diversity of the virus, and underscores the need for ongoing surveillance.

Reduced Expression of Interleukin-8 Receptors A and B on Polymorphonuclear Neutrophils from Persons with Human Immunodeficiency Virus Type 1 Disease and Pulmonary Tuberculosis

The expression of the two human interleukin (IL)-8 receptors, designated IL-8RA (CXCR-1) and IL-8RB (CXCR-2), on the surface of whole blood polymorphonuclear leukocytes (PMNL) was determined by use of receptor-specific monoclonal antibodies and flow cytometry. Sixteen subjects each were included in 4 study groups: healthy blood donors (ND), patients with pulmonary tuberculosis (TB), human immunodeficiency virus type 1-seropositive patients (HIV), and HIV-1-seropositive subjects with pulmonary tuberculosis (HIV/TB). A significant reduction in the percentage of PMNL expressing IL-8RA and IL-8RB and in their respective fluorescence intensities was found in TB, HIV, and HIV/TB groups compared with that obtained for the ND group. The greatest down-regulation of both receptors occurred in the HIV/TB group. Furthermore, associated with this reduced expression of IL-8 receptors was impairment of both intracellular calcium flux and migration of PMNL in response to IL-8 in a group of HIV/TB patients compared with that in healthy persons.

FOXP3 Expression Is Upregulated in CD4+T Cells in Progressive HIV-1 Infection and Is a Marker of Disease Severity

Background Understanding the role of different classes of T cells during HIV infection is critical to determining which responses correlate with protective immunity. To date, it is unclear whether alterations in regulatory T cell (Treg) function are contributory to progression of HIV infection. Methodology FOXP3 expression was measured by both qRT-PCR and by flow cytometry in HIV-infected individuals and uninfected controls together with expression of CD25, GITR and CTLA-4. Cultured peripheral blood mononuclear cells were stimulated with anti-CD3 and cell proliferation was assessed by CFSE dilution. Principal Findings HIV infected individuals had significantly higher frequencies of CD4+FOXP3+ T cells (median of 8.11%; range 1.33%–26.27%) than healthy controls (median 3.72%; range 1.3–7.5%; P = 0.002), despite having lower absolute counts of CD4+FOXP3+ T cells. There was a significant positive correlation between the frequency of CD4+FOXP3+ T cells and viral load (rho = 0.593 P = 0.003) and a significant negative correlation with CD4 count (rho = −0.423 P = 0.044). 48% of our patients had CD4 counts below 200 cells/µl and these patients showed a marked elevation of FOXP3 percentage (median 10% range 4.07%–26.27%). Assessing the mechanism of increased FOXP3 frequency, we found that the high FOXP3 levels noted in HIV infected individuals dropped rapidly in unstimulated culture conditions but could be restimulated by T cell receptor stimulation. This suggests that the high FOXP3 expression in HIV infected patients is likely due to FOXP3 upregulation by individual CD4+ T cells following antigenic or other stimulation. Conclusions/Significance FOXP3 expression in the CD4+ T cell population is a marker of severity of HIV infection and a potential prognostic marker of disease progression.

The subgroup F adenoviruses.

Background. Introduction. Human subgroup F adenoviruses (Ad) differ from all other adenoviruses in having been discovered by electron microscopy rather than culture (Flewett et al., 1974). Ironically, the first isolate, designated strain Tak, was noted in 1973 as an unidentifiable adenovirus giving CPE in HeLa cells, but remained uncharacterized for several years (de Jong et al., 1983). After more than 20 years, and despite appreciable advances in their genomic characterization, the fastidiousness of subgroup F adenoviruses in culture remains largely unexplained. This has baffled experts in adenovirus diagnosis and strain characterization and continues to intrigue others adopting a more molecular approach to their recalcitrant nature. Subgroup F adenoviruses present a quite different phenotype to that of the prototype human adenovirus (Ad2), which, by its ability to replicate efficiently in human cell cultures, allowed elucidation of eukaryotic splicing mechanisms.

Respiratory Syncytial Virus Nucleoprotein-Specific Cytotoxic T-Cell Epitopes in a South African Population of Diverse HLA Types Are Conserved in Circulating Field Strains

ABSTRACT This study identifies memory cytotoxic T lymphocyte (CTL) epitopes to respiratory syncytial virus (RSV) in healthy South African adults and demonstrates the conservation of those epitopes in circulating field strains of RSV in South Africa. Thirty-seven healthy adults from a population with diverse HLA backgrounds were screened by gamma interferon (IFN-γ) enzyme-linked immunospot for memory CTL activity in response to overlapping peptides representing the complete nucleoprotein (N) of RSV. Responses of more than 40 spot-forming cells/million cells were detectable in 21 individuals. The significant responses were further characterized, and 14-mer peptides were identified that induced cytolytic activity. Fine mapping of peptides with the highest cytolytic activity identified an HLA-B*08-restricted RSV-specific CTL epitope. The extended 14-mer peptide containing this epitope also induced lysis in the context of A*02-restricted target cells in some individuals. These HLA types are common in the target population; thus, the epitope is useful for studies of CTL responses to RSV in humans. The epitope was detected in healthy adults, reflecting the response generated in the course of previous natural RSV infection. We obtained a large panel of naturally occurring isolates of RSV to determine whether there was evidence of escape from CTL activity in circulating strains. We found that this epitope and a previously identified B*07-restricted N protein epitope were conserved in RSV field strains representing the diversity of circulating genotypes. This work suggests that escape from CTL activity is not common for this acute respiratory infection.

KIR-HLA and Maternal-Infant HIV-1 Transmission in Sub-Saharan Africa

Numerous studies have suggested a role for natural killer (NK) cells in attenuation of HIV-1 disease progression via recognition by killer-cell immunoglobulin-like receptors (KIRs) of specific HLA class I molecules. The role of KIR and HLA class I has not been addressed in the context of maternal-infant HIV-1 transmission. KIR and HLA class I B and C genes from 224 HIV-1-infected mothers and 222 infants (72 infected and 150 uninfected) from South Africa were characterized. Although a number of significant associations were determined in both the total group and in the nevirapine (NVP) exposed group, the most significant findings involved KIR2DL2 and KIR2DL3 and HLA-C. KIR2DL2/KIR2DL3 was underrepresented in intrapartum (IP)-transmitting mothers compared to non-transmitting (NT) mothers (P = 0.008) and remained significant (P = 0.036) after correction for maternal viral load (MVL). Homozygosity for KIR2DL3 alone and in combination with HLA-C allotype heterozygosity (C1C2) was elevated in IP-transmitting mothers compared to NT mothers (P = 0.034 and P = 0.01 respectively), and after MVL correction (P = 0.033 and P = 0.027, respectively). In infants, KIR2DL3 in combination with its HLA-C1 ligand (C1) as well as homozygosity for KIR2DL3 with C1C2, were both found to be underrepresented in infected infants compared to exposed uninfected infants in the total group (P = 0.06 and P = 0.038, respectively) and in the sub-group of infants whose mothers received NVP (P = 0.007 and P = 0.03, respectively). These associations were stronger post MVL adjustment (total group: P = 0.02 and P = 0.009, respectively; NVP group: P = 0.004 and P = 0.02, respectively). Upon stratification according to low and high MVL, all significant associations fell within the low MVL group, suggesting that with low viral load, the effects of genotype can be more easily detected. In conclusion this study has identified a number of significant associations that suggest an important role for NK cells in maternal-to-infant HIV-1 transmission.