Zika virus infection during pregnancy and vertical transmission: case reports and peptide-specific cell-mediated immune responses.

Zika virus (ZIKV) infection in pregnant women is associated with birth defects, which are more prevalent and severe the earlier in pregnancy the infection occurs. Pregnant women at risk of possible ZIKV exposure (n = 154) were screened using ELISA for ZIKV IgM and IgG. Nine of 154 (5.84%) pregnant women who underwent screening exhibited positive ZIKV serology. Of these, two maternal infections were confirmed by real-time RT-PCR and five were considered probable, but only three of those were retained for further analysis based on strict diagnostic criteria. Plaque reduction neutralization tests (PRNT) confirmed ZIKV infection in nine cases (5.84%). Two cases of vertical ZIKV transmission were confirmed by PCR. One infant showed no signs of congenital ZIKV syndrome and had a normal developmental profile despite first-trimester maternal infection. In the second case, pregnancy was terminated. Production of interferon γ (IFN-γ) by peripheral blood mononuclear cells obtained from pregnant women and umbilical cord blood was measured using enzyme-linked immunospot assay (ELISpot) after stimulation with panels of synthetic peptides derived from the sequence of ZIKV proteins. This analysis revealed that, among all peptide pools tested, those derived from the ZIKV envelope protein generated the strongest IFN-γ response.

HIV-1 group O integrase displays lower enzymatic efficiency and higher susceptibility to raltegravir than HIV-1 group M subtype B integrase.

HIV-1 group O (HIV-O) is a rare HIV-1 variant characterized by a high number of polymorphisms, especially in the integrase coding region. As HIV-O integrase enzymes have not previously been studied, our aim was to assess the impact of HIV-O integrase polymorphisms on enzyme function and susceptibility to integrase inhibitors. Accordingly, we cloned and purified integrase proteins from each of HIV-1 group O clades A and B, an HIV-O divergent strain, and HIV-1 group M (HIV-M, subtype B), used as a reference. To assess enzymatic function of HIV-O integrase, we carried out strand transfer and 3' processing assays with various concentrations of substrate (DNA target and long terminal repeats [LTR], respectively) and characterized these enzymes for susceptibility to integrase strand transfer inhibitors (INSTIs) in cell-free assays and in tissue culture, in the absence or presence of various concentrations of several INSTIs. The inhibition constant (Ki) and 50% effective concentration (EC50) values were calculated for HIV-O integrases and HIV-O viruses, respectively, and compared with those of HIV-M. The results showed that HIV-O integrase displayed lower activity in strand transfer assays than did HIV-M enzyme, whereas 3' processing activities were similar to those of HIV-M. HIV-O integrases were more susceptible to raltegravir (RAL) in competitive inhibition assays and in tissue culture than were HIV-M enzymes and viruses, respectively. Molecular modeling suggests that two key polymorphic residues that are close to the integrase catalytic site, 74I and 153A, may play a role in these differences.

Baseline genotypic and phenotypic susceptibilities of HIV-1 group O to enfuvirtide.

We assessed the natural genotypic and phenotypic susceptibilities to enfuvirtide of 171 HIV group O (HIV-O) samples and 29 strains, respectively. The N42D resistance-associated mutation in the gp41 region was detected in 98% of cases. The phenotypic assay showed a wide range of baseline susceptibilities, with 50% inhibitory concentrations (IC(50)s) from 4 to 5,000 nM, a range similar to that reported for HIV-1 group M. Thus, despite the natural genotypic resistance conferred by the N42D signature mutation, HIV-O variants appear to be phenotypically susceptible. Enfuvirtide could therefore potentially be used in antiretroviral treatments for HIV-O-infected patients.

HIV-1 Group O Resistance Against Integrase Inhibitors.

BACKGROUND: HIV-1 group O (HIV-O) is a rare variant that is characterized by a high number of natural polymorphisms in the integrase coding region that may impact on susceptibility to integrase strand transfer inhibitors (INSTIs) and on the emergence of resistance substitutions. We previously reported that HIV-O is more susceptible to RAL than HIV-1 group M (HIV-M). METHODS: The aim of this study was to assess pathways of resistance to INSTIs in group 0 variants. Accordingly, we selected for resistance to each of raltegravir (RAL), elvitegravir (EVG), and dolutegravir (DTG) in cord blood mononuclear cells using HIV group O subtypes A and B, an HIV-O divergent isolate, and HIV-1 group M (subtype B, which served as a reference). Site-directed mutagenesis was performed on the pCOM2.5 HIV group 0 infectious clone to ascertain the impact of INSTI resistance substitutions at positions Q148R, N155H, and R263K within integrase on susceptibility to INSTIs and infectiousness. RESULTS: Cell culture selections of group O variants yielded similar patterns of resistance to RAL, EVG, and DTG as observed for subtype B. In the DTG selections, subtype B yielded S153Y, whereas a natural S153A polymorphism sometimes led to A153V in group O. The pCMO2.5/Q148R and pCMO2.5/N155H variants displayed far higher levels of resistance to DTG (>1000 FC) than was seen for group M viruses. CONCLUSIONS: HIV-O harboring Q148R and N155H shows higher resistance to DTG compared with HIV-M subtype B.

HIV-1 group O integrase displays lower susceptibility to raltegravir and has a different mutational pathway for resistance than HIV-1 group M.

INTRODUCTION: HIV-1 group O (HIV-O) is a rare HIV-1 variant characterized by a high number of polymorphisms, especially in the integrase gene, e.g. positions L74I, S153A, G163Q and T206S. As HIV-O integrase enzymes have not previously been studied, our aim was to assess the impact of HIV-O integrase polymorphisms on susceptibility to integrase inhibitors and emergence of resistance associated mutations. Viruses and Methods: We cloned and purified integrase proteins from each of HIV-1 Group O clades A (HIV-O/A) and B (HIV-O/B), a HIV-O divergent strain (HIV-O/Div), and HIV-1 group M (subtype B, HIV-M/B) and characterized these enzymes for susceptibility to integrase strand transfer inhibitors (INSTIs) in cell-free assays and in tissue culture, in the absence or presence of varying concentrations of several INSTIs. The inhibition constant (Ki) and IC50 were calculated and compared for HIV-M and HIV-O integrases. Selections for resistance-related mutations were performed using cord blood mononuclear cells and increasing concentration of INSTIs. RESULTS: HIV-O integrase and viruses were more susceptible to raltegravir (RAL) in competitive inhibition assays and in tissue culture than were HIV-M enzymes and viruses, respectively. During selection, we observed different pathways of resistance depending on the drug and clade. Mutations selected in HIV-O can be classified as follows: (1) mutations described for HIV-M such as T97A, Q148R, V151A/I (RAL), T66I, E92Q, E157Q (EVG) and M50I, R263K (DTG) and (2) signature mutations for HIV-O (i.e. not described in HIV-M) F121C (HIV-O/B for RAL), V75I (HIV-O/A for RAL) and S153V (HIV-O/A for DTG). Only the HIV-O/Div selected the Q148R mutation for RAL and R263K+M50I for DTG, as previously described for HIV-M. None of the HIV-O viruses selected either N155H or Y143C. The selection of the specific S153V mutation could be explained at the nucleotide level: HIV-O at this position contains an alanine and substitution of alanine to valine (153AGGC→153VGTC) is easier than substitution of alanine to tyrosine (153AGGC→153YTAC), with only a transversion needed instead of one transition plus one transversion. CONCLUSIONS: This is the first report of susceptibility and resistance in vitro to INSTIs for HIV-O. Our study confirmed the impact of HIV-O polymorphism, on susceptibility to INSTIs and the emergence of resistance mutations.

Impact of HIV-1 group O genetic diversity on genotypic resistance interpretation by algorithms designed for HIV-1 group M.

BACKGROUND: HIV-1 group O (HIV-O) is characterized by a high genetic divergence from HIV-1 group M viruses. Little is known about the therapeutic impact of this diversity. The aim of this study was to assess in a large series of samples (1) the genotypic impact of natural polymorphism of the HIV-O reverse transcriptase and protease genes; and (2) the predictive value of resistance interpretation algorithms developed for HIV-1 group M when used for highly mutated HIV-O viruses. METHODS: Sixty-eight antiretroviral-naive and 9 highly antiretroviral-experienced HIV-O-infected patients were included. The viruses were sequenced and resistance-associated mutations were identified using 3 different algorithms (Agence Nationale de Recherches sur le SIDA et les hépatites virales, Rega, Stanford). RESULTS: All HIV-O samples naturally exhibited the A98G and V179E resistance mutations in the reverse transcriptase region; 54% of samples presented the Y181C mutation, conferring resistance to nonnucleoside reverse transcriptase inhibitors. Twelve minor resistance mutations, present in more than 75% of the protease sequences, led to the different algorithms giving discrepant results for nelfinavir and saquinavir susceptibility. A marked virological response was observed in 8 of the 9 antiretroviral-experienced patients, despite the prediction of limited activity of the combination for 5 to 8 patients according to the algorithm used. CONCLUSIONS: The high level of natural polymorphism in HIV-O genes, and the important discrepancies between genotypic resistance interpretation and the virological response, emphasize the need for resistance algorithm rules better adapted to HIV-O.

Diagnosis and monitoring of HIV-1 group O-infected patients in Cameroun.

OBJECTIVE: To define a routine algorithm for the specific diagnosis and complete follow-up of HIV-1 group O (HIV-O) infections in Cameroun. METHODS: During 18 months, samples referred to Centre Pasteur du Cameroun for HIV testing or viral monitoring were screened for HIV-O infection with an in-house serotyping assay. HIV-O viral load was quantified by real-time polymerase chain reaction in the LTR gene and resistance genotyping was performed on pol and env sequences. RESULTS: Of the 7030 samples tested, 78 HIV-O infections (1.1%) were identified, including 7 M and O dually seroreactive samples (9%). All treatment-naive patients and 59% of the patients receiving HAART had detectable viral loads. Analysis of pol sequences from 15 treatment-naive patients revealed a high number of polymorphisms in the protease region, with natural residues implicated in genotypic resistance to tipranavir and saquinavir for HIV-1 group M according to the Agence Nationale de Recherches sur le Sida et les Hépatites virales algorithm. Six patients (40%) harbored the 181C mutation conferring natural resistance to nonnucleoside reverse transcriptase inhibitors. Among antiretroviral-treated patients, major resistance mutations described for HIV-1 group M were found. CONCLUSIONS: HIV-O prevalence remains relatively low in Cameroun. The cocirculation of groups M and O in this country leads to replicative dual infections. HIV-O-infected patients in this region can now benefit from effective and specific tools for a complete monitoring of infection. However, further studies are needed to understand long-term response to antiretrovirals of these complex variants.