Maria A. Papathanasopoulos

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.

Emergence of X4 usage among HIV-1 subtype C: evidence for an evolving epidemic in South Africa.

This study investigated the genotype and phenotype of HIV-1 isolates of 20 South African AIDS patients. We found the highest percentage of CXCR4 usage among primary isolates, in which 30% efficiently utilized CXCR4 and exhibited the syncytium-inducing phenotype. Phylogenetic analysis of env confirmed that 19 of the 20 were subtype C, and syncytium-inducing viruses had genetic changes in the V3 loop, characteristic of CXCR4 usage. Results imply that the frequency of CXCR4-utilizing subtype C is increasing with time.

Multiple bacteriocin production by Leuconostoc mesenteroides TA33a and other Leuconostoc/Weissella strains.

Abstract.Leuconostoc (Lc.) mesenteroides TA33a produced three bacteriocins with different inhibitory activity spectra. Bacteriocins were purified by adsorption/desorption from producer cells and reverse phase high-performance liquid chromatography. Leucocin C-TA33a, a novel bacteriocin with a predicted molecular mass of 4598 Da, inhibited Listeria and other lactic acid bacteria (LAB). Leucocin B-TA33a has a predicted molecular mass of 3466 Da, with activity against Leuconostoc/Weissella (W.) strains, and appears similar to mesenterocin 52B and dextranicin 24, while leucocin A-TA33a, which also inhibited Listeria and other LAB strains, is identical to leucocin A-UAL 187. A survey of other known bacteriocin-producing Leuconostoc/Weissella strains for the presence of the three different bacteriocins revealed that production of leucocin A-, B- and C-type bacteriocins was widespread. Lc. carnosum LA54a, W. paramesenteroides LA7a, and Lc. gelidum UAL 187-22 produced all three bacteriocins, whereas W. paramesenteroides OX and Lc. carnosum TA11a produced only leucocin A- and B-type bacteriocins.

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.

Full-Length Genome Analysis of HIV-1 Subtype C Utilizing CXCR4 and Intersubtype Recombinants Isolated in South Africa

The HIV-1 epidemic in South Africa is largely due to subtype C viruses, which preferentially use CCR5 as a coreceptor for infection. We describe full-length genome sequences of two CXCR4-utilizing HIV-1 subtype C viruses and two intersubtype recombinants from South Africa. Three of the viruses (99ZACM4, 99ZACM9, and 99ZASW7) were isolated in 1999 from AIDS patients in Johannesburg, and a fourth virus (98ZADu178) was isolated in Durban in 1998 from an asymptomatic female sex worker. Isolates 99ZASW7 and 99ZACM9 from Johannesburg were subtype C throughout the genome, 99ZASW7 used the CXCR4 coreceptor, and 99ZACM9 used both CCR5 and CXCR4. Isolate 98ZADu178 from Durban was a novel recombinant between subsubtype A2 and subtype C. The third isolate from Johannesburg, 99ZACM4, was a complex, novel recombinant with multiple breakpoints and contained segments of subtypes A, C, D, G, and K. These results establish the presence of intersubtype recombinants in South Africa, indicating that ongoing surveillance for other subtypes and recombinants is necessary.

Characterization of leucocin B-Ta11a: a bacteriocin fromLeuconostoc carnosum Ta11a isolated from meat.

Leuconostoc (Lc.)carnosum Ta11a, isolated from vacuum-packaged processed meats, produced a bacteriocin designated leucocin B-Ta11a. The crude bacteriocin was heat stable and sensitive to proteolytic enzymes, but not to catalase, lysozyme, or chloroform. It was active againstListeria monocytogenes and several lactic acid bacteria. Leucocin B-Ta11a was optimally produced at 25°C in MRS broth at an initial pH of 6.0 or 6.5 An 8.9-MDa plasmid inLeuconostoc carnosum Ta11a hybridized to a 36-mer oligonucleotide probe (JF-1) that was homologous to leucocin A-UAL187. A 4.9-kbSau3A fragment from a partial digest of the 8.9-MDa plasmid was cloned into pUC118. The 8.1-kb recombinant plasmid (pJF8.1) was used for sequencing and revealed the presence of two open reading frames (ORFs). ORF1 codes for a protein of 61 amino acids comprising a 37-amino-acid bacteriocin that was determined to be the leucocin B-Ta11a structural gene by virtue of its homology to leucocin A-UAL 187 (Hastings et al. 1991. J. Bacteriol 173: 7491–7500). The 24-amino-acid N-terminal extension, however, differs from that of leucocin A-UAL187 by seven residues. The predicted protein of the ORF2 has 113 amino acids and is identical with the amino acid sequence of the cognate ORF of the leucocin A-UAL 187 operon.

Sequence and structural relationships of leucocins A-, B- and C-TA33a from Leuconostoc mesenteroides TA33a

Amino acid sequences of two of the three bacteriocins from Leuconostoc mesenteroides TA33a were determined and their sequence-structure relationships investigated. Leucocin B-TA33a consists of 31 amino acid residues, with a molecular mass of 3466 Da. Leucocin B-TA33a does not belong to the pediocin family of bacteriocins, but shares 62% homology with mesenterocin 52B. A partial sequence of 36 amino acids of leucocin C-TA33a (4598 Da) was determined. Leucocin C-TA33a belongs to the class II bacteriocins having the consensus YGNGV motif. The third bacteriocin, leucocin A-TA33a, is identical to leucocin A-UAL 187. Circular dichroism spectra of the leucocins in aqueous solution and micellar SDS indicated that they undergo a structural transition when in a membrane-mimicking environment. Theoretical predictions from circular dichroism data suggest that leucocins A-, B- and C-TA33a adopt a beta-structure (48%) in membrane-mimicking environments. Sequence alignments and secondary structure predictions for the N-terminus of leucocins A- and C-TA33a predicted that Cys-9 and Cys-14 are connected by a disulfide bridge and form two beta-strands.

Sensitivity of HIV Type 1 Subtype C Isolates to the Entry Inhibitor T-20

T-20 is the first in a new class of antiretroviral drugs targeting the entry stage of the virus life cycle. It is a 36 amino acid peptide that binds to the HR1 region of gp41 preventing gp41-mediated fusion with the host cell membrane. T-20 was designed based on the HR2 sequence of HIV-1 subtype B gp41, a region that shows significant genetic variation with HIV-1 subtype C sequences. In order to assess the efficacy of T-20 to inhibit subtype C isolates, a total of 23 isolates were tested for their ability to replicate in the presence of T-20. This included 15 isolates that used CCR5, five that used both CCR5 and CXCR4, and three that used CXCR4. Five of these were from patients failing other antiretroviral therapies. Sequence analysis of the HR2 region indicated that there were 10-16 amino acid changes in the region corresponding to T-20. However, all isolates were effectively inhibited by T-20 at 1 microg/ml. There were no significant differences between viruses that used CCR5 or CXCR4 to enter cells. All isolates, except one, had GIV at positions 36-38 in the HR1 region. One isolate had a GVV motif but this did not affect its sensitivity to T-20. Therefore, T-20 inhibited subtype C viruses despite significant genetic differences in the HR2 region and there was no evidence for baseline resistance to T-20. These data suggest that T-20 would be highly effective in patients with HIV-1 subtype C infection, including those failing existing antiretroviral drug regimens.

Natural Killer Cells That Respond to Human Immunodeficiency Virus Type 1 (HIV-1) Peptides Are Associated with Control of HIV-1 Infection

Human immunodeficiency virus (HIV)-specific natural killer (CD3- cells), CD4, and CD8 T cellular responses were determined in 79 HIV‐1-infected women in response to HIV‐1 peptide pools (Gag, Pol, Nef, Reg, and Env) with use of a whole‐blood intracellular cytokine staining assay that measures interferon-γ and/or interleukin-2. HIV‐specific CD3- cell responses to any region (Env and Reg predominantly targeted) were associated with lower viral load (P = .031) and higher CD4 T cell count (P = .015). Env‐specific CD3- cell responses were stronger in women who had both Gag CD4 and CD8 T cell responses and, in turn, was associated with lower viral load (P = .005). CD3- cell responders had significantly higher representation of CD4 T cell responses to Env and Reg (P = .012 and P = .015, respectively) and higher magnitudes of CD4 T cell responses (P = .017 and P = .037, respectively) than did nonresponders. Peptide‐specific natural killer cells are associated with markers of less severe disease progression among HIV‐1-infected women (lower viral load and higher CD4 T cell count) and with stronger HIV‐specific T cell responses.

High prevalence of the K65R mutation in HIV-1 subtype C infected patients failing tenofovir-based first-line regimens in South Africa.

Background Tenofovir (TDF) has replaced stavudine (d4T) as the preferred nucleoside reverse transcriptase inhibitor (NRTI) in first-line regimens in South Africa, but limited information is available on the resistance patterns that develop after the introduction of TDF. This study investigated the antiretroviral drug resistance patterns in South African HIV-1 subtype C-infected patients failing stavudine- (d4T) and tenofovir- (TDF) based first-line regimens and assess the suitability of TDF as the preferred first-line nucleotide reverse transcriptase inhibitor (NRTI). Methods Resistance patterns of HIV-1 from 160 adult patients virologically failing TDF- (n = 80) and d4T- (n = 80) based first-line regimens were retrospectively analyzed. The pol gene was sequenced using an in-house protocol and mutations were analysed using the IAS-USA 2014 Drug Resistance Mutation list. Results Compared to d4T-exposed patients (n = 7), patients failing on a TDF-containing regimen (n = 43) were almost 5 times more likely to present with a K65R mutation (aRR 4.86 95% CI 2.29 – 10.34). Y115F was absent in the d4T group, and detected in 13.8% (n = 11) of TDF-exposed patients, p = 0.0007. Virus from 9 of the 11 patients (82.0%) who developed the Y115F mutation also developed K65R. Intermediate or high-level resistance to most NRTIs was common in the TDF-treatment group, but these patients twice more likely to remain susceptible to AZT as compared to those exposed to d4T (aRR 2.09 95% CI 1.13 – 3.90). Conclusion The frequency of the TDF induced K65R mutation was higher in our setting compared to non-subtype C dominated countries. However, despite the higher frequency of cross-resistance to NRTIs, most patients remained susceptible to AZT, which is reflected in the South African treatment guidelines that recommend AZT as an essential component of second-line regimens.