Dario Alberto Dilernia

HLA-driven convergence of HIV-1 viral subtypes B and F toward the adaptation to immune responses in human populations.

Background Cytotoxic T-Lymphocyte (CTL) response drives the evolution of HIV-1 at a host-level by selecting HLA-restricted escape mutations. Dissecting the dynamics of these escape mutations at a population-level would help to understand how HLA-mediated selection drives the evolution of HIV-1. Methodology/Principal Findings We undertook a study of the dynamics of HIV-1 CTL-escape mutations by analyzing through statistical approaches and phylogenetic methods the viral gene gag sequenced in plasma samples collected between the years 1987 and 2006 from 302 drug-naïve HIV-positive patients. By applying logistic regression models and after performing correction for multiple test, we identified 22 potential CTL-escape mutations (p-value<0.05; q-value<0.2); 10 of these associations were confirmed in samples biologically independent by a Bayesian Markov Chain Monte-Carlo method. Analyzing their prevalence back in time we found that escape mutations that are the consensus residue in samples collected after 2003 have actually significantly increased in time in one of either B or F subtype until becoming the most frequent residue, while dominating the other viral subtype. Their estimated prevalence in the viral subtype they did not dominate was lower than 30% for the majority of samples collected at the end of the 80s. In addition, when screening the entire viral region, we found that the 75% of positions significantly changing in time (p<0.05) were located within known CTL epitopes. Conclusions Across HIV Gag protein, the rise of polymorphisms from independent origin during the last twenty years of epidemic in our setting was related to an association with an HLA allele. The fact that these mutations accumulated in one of either B or F subtypes have also dominated the other subtype shows how this selection might be causing a convergence of viral subtypes to variants which are more likely to evade the immune response of the population where they circulate.

Lack of viral selection in human immunodeficiency virus type 1 mother-to-child transmission with primary infection during late pregnancy and/or breastfeeding

Mother-to-child transmission (MTCT) of human immunodeficiency virus type 1 (HIV-1) as described for women with an established infection is, in most cases, associated with the transmission of few maternal variants. This study analysed virus variability in four cases of maternal primary infection occurring during pregnancy and/or breastfeeding. Estimated time of seroconversion was at 4 months of pregnancy for one woman (early seroconversion) and during the last months of pregnancy and/or breastfeeding for the remaining three (late seroconversion). The C2V3 envelope region was analysed in samples of mother-child pairs by molecular cloning and sequencing. Comparisons of nucleotide and amino acid sequences as well as phylogenetic analysis were performed. The results showed low variability in the virus population of both mother and child. Maximum-likelihood analysis showed that, in the early pregnancy seroconversion case, a minor viral variant with further evolution in the child was transmitted, which could indicate a selection event in MTCT or a stochastic event, whereas in the late seroconversion cases, the mothers and childs sequences were intermingled, which is compatible with the transmission of multiple viral variants from the mothers major population. These results could be explained by the less pronounced selective pressure exerted by the immune system in the early stages of the mothers infection, which could play a role in MTCT of HIV-1.

Host genetic factors associated with symptomatic primary HIV infection and disease progression among Argentinean seroconverters.

Background Variants in HIV-coreceptor C-C chemokine receptor type 5 (CCR5) and Human leukocyte antigen (HLA) genes are the most important host genetic factors associated with HIV infection and disease progression. Our aim was to analyze the association of these genetic factors in the presence of clinical symptoms during Primary HIV Infection (PHI) and disease progression within the first year. Methods Seventy subjects diagnosed during PHI were studied (55 symptomatic and 15 asymptomatic). Viral load (VL) and CD4 T-cell count were evaluated. HIV progression was defined by presence of B or C events and/or CD4 T-cell counts <350 cell/mm3. CCR5 haplotypes were characterized by polymerase chain reaction and SDM-PCR-RFLP. HLA-I characterization was performed by Sequencing. Results Symptoms during PHI were significantly associated with lower frequency of CCR5-CF1 (1.8% vs. 26.7%, p = 0.006). Rapid progression was significantly associated with higher frequency of CCR5-CF2 (16.7% vs. 0%, p = 0.024) and HLA-A*11 (16.7% vs. 1.2%, p = 0.003) and lower frequency of HLA-C*3 (2.8% vs. 17.5%, p = 0.035). Higher baseline VL was significantly associated with presence of HLA-A*11, HLA-A*24, and absence of HLA-A*31 and HLA-B*57. Higher 6-month VL was significantly associated with presence of CCR5-HHE, HLA-A*24, HLA-B*53, and absence of HLA-A*31 and CCR5-CF1. Lower baseline CD4 T-cell count was significantly associated with presence of HLA-A*24/*33, HLA-B*53, CCR5-CF2 and absence of HLA-A*01/*23 and CCR5-HHA. Lower 6-month CD4 T-cell count was associated with presence of HLA-A*24 and HLA-B*53, and absence of HLA-A*01 and HLA-B*07/*39. Moreover, lower 12-month CD4 T-cell count was significantly associated with presence of HLA-A*33, HLA-B*14, HLA-C*08, CCR5-CF2, and absence of HLA-B*07 and HLA-C*07. Conclusion Several host factors were significantly associated with disease progression in PHI subjects. Most results agree with previous studies performed in other groups. However, some genetic factor associations are being described for the first time, highlighting the importance of genetic studies at a local level.

Estimation of HIV-Testing Rates to Maximize Early Diagnosis-Derived Benefits at the Individual and Population Level

Background In HIV infection, initiation of treatment is associated with improved clinical outcom and reduced rate of sexual transmission. However, difficulty in detecting infection in early stages impairs those benefits. We determined the minimum testing rate that maximizes benefits derived from early diagnosis. Methods We developed a mathematical model of HIV infection, diagnosis and treatment that allows studying both diagnosed and undiagnosed populations, as well as determining the impact of modifying time to diagnosis and testing rates. The model’s external consistency was assessed by estimating time to AIDS and death in absence of treatment as well as by estimating age-dependent mortality rates during treatment, and comparing them with data previously reported from CASCADE and DHCS cohorts. Results In our model, life expectancy of patients diagnosed before 8 years post infection is the same as HIV-negative population. After this time point, age at death is significantly dependent on diagnosis delay but initiation of treatment increases life expectancy to similar levels as HIV-negative population. Early mortality during HAART is dependent on treatment CD4 threshold until 6 years post infection and becomes dependent on diagnosis delay after 6 years post infection. By modifying testing rates, we estimate that an annual testing rate of 20% leads to diagnosis of 90% of infected individuals within the first 8.2 years of infection and that current testing rate in middle-high income settings stands close to 10%. In addition, many differences between low-income and middle-high incomes can be predicted by solely modifying the diagnosis delay. Conclusions To increase testing rate of undiagnosed HIV population by two-fold in middle-high income settings will minimize early mortality during initiation of treatment and global mortality rate as well as maximize life expectancy. Our results highlight the impact of achieving early diagnosis and the importance of strongly work on improving HIV testing rates.

Centrifugation improves the detection of HIV-1 p24 antigen in plasma from children born to mothers infected with HIV-1.

Detection of HIV proteins and/or nucleic acids is necessary for the diagnosis of perinatal HIV infection. Despite its low sensitivity, detection of p24 antigen in plasma is a simple and economic method for the diagnosis of HIV in exposed children. The aim of this study was to improve the sensitivity of detection of p24 using centrifugation of plasma. Forty-seven selected stored samples from 37 children (23 infected, 14 uninfected, median age of 137 days) were examined. Plasma samples (volume 0.3-1.5 ml) were defrosted, centrifuged at 23,500 x g at 4 degrees C for 60 min and determination of p24 was carried out in the resuspended pellet (0.12 ml). In 32 plasma samples from infected children, p24 was found originally in 6 (18.7%) and resulted positive in 24 (75%) pellets. When only one sample per child was considered, sensitivity was significantly higher in pellets, 3/23 uncentrifuged plasma samples and 15/23 pellets (McNemar Test, p<0.001). Specificity was 100%. The absorbance/cut-off ratio was always higher in the pellets from positive children (p=0.028). Plasma samples with volumes of 1 ml or more achieved a higher sensitivity (91.7% vs. 36.4%, p=0.009). Centrifugation of plasma samples prior to determination of p24 in pediatric patients resulted in a significant increase in sensitivity.

Stable human T-cell lymphotropic virus type 1 (HTLV-1) subtype a/subgroup a endemicity in Amerindians from Northwest Argentina: a health problem to be resolved.

Jujuy province, in Northwest Argentina, is known to be endemic for HTLV‐1 infection. Moreover, foci of HTLV‐1 associated pathologies have also been described in this region. To gain an insight into the current situation of HTLV‐1/2 in this endemic area, a seroprevalence and phylogenetic study was performed among a Kolla community from Abra Pampa city and surroundings. Out of 112 individuals, 11 (9.8%) were confirmed as HTLV‐1 positive and no HTLV‐2 infection was detected. The phylogenetic analysis of the LTR region showed that all the HTLV‐1 sequences belonged to the Cosmopolitan subtype a/transcontinental subgroup A, and were closely related to reference sequences from Peru, Argentina, and the South of Brazil (P = 0.82). Considering the cultural and historical features of this community and in spite of the mandatory detection of anti‐HTLV‐1/2 antibodies in blood banks since 2005, it would be important to implement new public health measures focused on decreasing HTLV‐1 transmission in this endemic area. J. Med. Virol. 82:2116–2122, 2010.

Low selection rate of HLA-anchor escape mutations in HIV after transmission of subtype B and recombinants BF strains patients from Argentina

Background The immune response of HIV-infected individuals shapes the evolution of the virus by selecting escape mutation. After transmission to a new host, the HLA-mediated immune pressure changes. The objective of our study was to characterize the dynamics of HLA-anchor escape mutations after transmission. Methods We studied 6 transmission events between members of serodiscordant couples collecting blood samples from the donor and the previously seronegative-partner at the moment of seroconversion, and a second sample at least 6 months post-infection. HLA-I typing was performed by the SSOP-PCR method. The viral gene gag was amplified by RT-PCR and cloned into the pGEM-T vector for viral quasiespecies analysis. Transmitted strain was identified by phylogenetic analysis. Escape mutation was defined as viral polimorphisms located in HLA-anchor position that eliminate an epitope predicted by the NetMHC (CBS Prediction Server). Significant variations in the number

Differential HLA-dependent HIV evolution among subtypes

Population HLA-dependent evolution has been documented over the principal HIV proteins. Effect of subtype variability is uncertain. In Argentina, B subtype and recombinant BF variants circulate in equal proportions.