Hepatitis E Virus Quasispecies in Cerebrospinal Fluid with Neurological Manifestations.

Hepatitis E virus (HEV) infection can lead to a variety of neurological disorders. While HEV RNA is known to be present in the central nervous system, HEV quasispecies in serum and cerebrospinal fluid (CSF) have rarely been explored. We studied the virus' quasispecies in the blood and the CSF of five patients at the onset of their neurological symptoms.

Th22 cells are efficiently recruited in the gut by CCL28 as an alternative to CCL20 but do not compensate for the loss of Th17 cells in treated HIV-1-infected individuals.

Gut CD4 T cells are incompletely restored in most HIV-1-infected individuals on antiretroviral therapy, notably Th17 cells, a key subset in mucosal homeostasis. By contrast, gut Th22 cells are usually restored at normal frequencies. Th22 cells display a CCR6CCR10 phenotype and could thus respond to CCL20- and CCL28-mediated chemotaxis, while Th17 cells, which express CCR6 but not CCR10, depend on CCL20.

Platelets from HIV-infected individuals on antiretroviral drug therapy with poor CD4 T cell recovery can harbor replication-competent HIV despite viral suppression.

In addition to hemostasis, human platelets have several immune functions and interact with infectious pathogens including HIV in vitro. Here, we report that platelets from HIV-infected individuals on combined antiretroviral drug therapy (ART) with low blood CD4 T cell counts (<350 cells/μl) contained replication-competent HIV despite viral suppression. In vitro, human platelets harboring HIV propagated the virus to macrophages, a process that could be prevented with the biologic abciximab, an anti-integrin αIIb/β3 Fab.

THETA: a new genotypic approach for predicting HIV-1 CRF02-AG coreceptor usage.

Motivation: The circulating recombinant form of HIV-1 CRF02-AG is the most frequent non-B subtype in Europe. Anti-HIV therapy and pathophysiological studies on the impact of HIV-1 tropism require genotypic determination of HIV-1 tropism for non-B subtypes. But genotypic approaches based on analysis of the V3 envelope region perform poorly when used to determine the tropism of CRF02-AG.

Long-term evolution of transmitted CXCR4-using HIV-1 under effective antiretroviral therapy.

Objective: To study the long-term evolution of the transmitted CXCR4-using viruses. CCR5-using viruses (R5 viruses) predominate during primary HIV-1 infections (PHI) while CXCR4-using viruses are isolated in less than 10% of PHI.

Design: Six patients infected with an R5X4 virus, detected by a sensitive phenotypic assay during PHI, were matched with six patients infected with a pure R5 virus for sex, Fiebig stage, time of antiretroviral initiation and duration of follow-up.

Increased CXCR3+ T Cells Impairs Recruitment of T-Helper Type 17 Cells via Interferon γ and Interleukin 18 in the Small Intestine Mucosa During Treated HIV-1 Infection.

The restoration of CD4+ T cells, especially T-helper type 17 (Th17) cells, remains incomplete in the gut mucosa of most human immunodeficiency virus type 1 (HIV-1)-infected individuals despite sustained antiretroviral therapy (ART). Herein, we report an increase in the absolute number of CXCR3+ T cells in the duodenal mucosa during ART. The frequencies of Th1 and CXCR3+ CD8+ T cells were increased and negatively correlated with CCL20 and CCL25 expression in the mucosa.

Detection of Zika, dengue and chikungunya viruses using single-reaction multiplex real-time RT-PCR.

Zika (ZIKV), Dengue (DENV) and Chikungunya viruses (CHIKV) co-circulate in the same geographical areas during the same seasonal period through the same biting arthropods. Therefore a rapid sensitive and specific molecular assay for these viruses would be a considerable help in the disease management and the epidemiological survey. We developed a one-step multiplex real-time PCR for the simultaneous detection of these viruses.

Performance comparison of next-generation sequencing platforms for determining HIV-1 coreceptor use.

The coreceptor used by HIV-1 must be determined before a CCR5 antagonist, part of the arsenal of antiretroviral drugs, is prescribed because viruses that enter cells using the CXCR4 coreceptor are responsible for treatment failure. HIV-1 tropism is also correlated with disease progression and so must be determined for virological studies. Tropism can be determined by next-generation sequencing (NGS), but not all of these new technologies have been fully validated for use in clinical practice.

Quantification of HEV RNA by Droplet Digital PCR.

The sensitivity of real-time PCR for hepatitis E virus (HEV) RNA quantification differs greatly among techniques. Standardized tools that measure the real quantity of virus are needed. We assessed the performance of a reverse transcription droplet digital PCR (RT-ddPCR) assay that gives absolute quantities of HEV RNA.

Brief Report: HIV-1 Tropism During Primary Infections in France: 1996-2014.

HIV-1 was mainly CCR5 tropic in recent seroconverters. We analyzed the coreceptor use in 239 primary HIV-1 infections (PHIs) between 1996 and 2014 using a validated recombinant virus phenotypic entry assay. CXCR4-using viruses were detected in 8.

CCR6(-) regulatory T cells blunt the restoration of gut Th17 cells along the CCR6-CCL20 axis in treated HIV-1-infected individuals.

The gut CD4(+) T cells, particularly the T helper type 17 (Th17) subset, are not completely restored in most HIV-1-infected individuals despite combined antiretroviral therapy, when initiated at the chronic phase of infection. We show here that the CCR6-CCL20 chemotactic axis is altered, with reduced CCL20 production by small intestine epithelial cells in treated HIV-1-infected individuals. This leads to impaired CCR6(+)CD4(+) T-cell homing, particularly Th17 cells, to the small intestine mucosa.

No selection of CXCR4-using variants in cell reservoirs of dual-mixed HIV-infected patients on suppressive maraviroc therapy.

We used ultradeep sequencing to investigate the evolution of the frequency of CXCR4-using viruses in the peripheral blood mononuclear cells of 22 patients infected with both CCR5 and CXCR4-using viruses treated with the CCR5 antagonist maraviroc for 24 weeks and a stable antiviral therapy. The mean CXCR4-using virus frequency in peripheral blood mononuclear cells was 59% before maraviroc intensification and 52% after 24 weeks of effective treatment, indicating no selection by maraviroc.

Position-specific automated processing of V3 env ultra-deep pyrosequencing data for predicting HIV-1 tropism.

HIV-1 coreceptor usage must be accurately determined before starting CCR5 antagonist-based treatment as the presence of undetected minor CXCR4-using variants can cause subsequent virological failure. Ultra-deep pyrosequencing of HIV-1 V3 env allows to detect low levels of CXCR4-using variants that current genotypic approaches miss. However, the computation of the mass of sequence data and the need to identify true minor variants while excluding artifactual sequences generated during amplification and ultra-deep pyrosequencing is rate-limiting.

Evolution of HIV-1 quasispecies and coreceptor use in cell reservoirs of patients on suppressive antiretroviral therapy.

Objectives: To track changes in the V3 env region of HIV-1 quasispecies and determine virus coreceptor use in cell reservoirs of patients on long-term suppressive antiretroviral therapy (ART).

Patients And Methods: Ten patients whose plasma viraemia had been suppressed for a median of 5.5 years were followed for 5 years.

Primary resistance of CCR5-tropic HIV-1 to maraviroc cannot be predicted by the V3 sequence.

Objectives: Resistance of HIV-1 to CCR5 antagonists can occur without coreceptor switching by mutations in envelope glycoproteins that enable virus entry using the inhibitor-bound form of CCR5. We investigated whether mutations in the V3 region of HIV-1 from subjects naive to maraviroc could be associated with primary resistance to this drug.

Methods: The frequency of CCR5-tropic HIV-1 subtype B isolates harbouring putative V3 maraviroc resistance mutations was assessed among the HIV tropism database of Toulouse University Hospital, France.

Altered CD4+ T cell homing to the gut impairs mucosal immune reconstitution in treated HIV-infected individuals.

Depletion of CD4+ T cells from the gut occurs rapidly during acute HIV-1 infection. This has been linked to systemic inflammation and disease progression as a result of translocation of microbial products from the gut lumen into the bloodstream. Combined antiretroviral therapy (cART) substantially restores CD4+ T cell numbers in peripheral blood, but the gut compartment remains largely depleted of such cells for poorly understood reasons.

Genotypic prediction of HIV-1 subtype D tropism.

Background: HIV-1 subtype D infections have been associated with rapid disease progression and phenotypic assays have shown that CXCR4-using viruses are very prevalent. Recent studies indicate that the genotypic algorithms used routinely to assess HIV-1 tropism may lack accuracy for non-B subtypes. Little is known about the genotypic determinants of HIV-1 subtype D tropism.

Frequency of CXCR4-using viruses in primary HIV-1 infections using ultra-deep pyrosequencing.

We used ultra-deep pyrosequencing and the Toulouse Tropism Test phenotypic assay to determine the prevalence of CXCR4-using viruses in 21 patients with primary HIV-1 infections. We found X4-containing virus populations in 9% of patients by ultra-deep pyrosequencing using position-specific scoring matrices (PSSM(X4/R5)) or geno2pheno(5.75) and in 14% using the combined 11/25 and net charge rule.

CXCR4-using viruses in plasma and peripheral blood mononuclear cells during primary HIV-1 infection and impact on disease progression.

Objective: Cysteine-cysteine receptor 5 (CCR5)-using viruses classically predominate during HIV-1 primary infection but the frequency of cysteine-X-cysteine receptor 4 (CXCR4)-using viruses varies between studies and could be different between plasma and peripheral blood mononuclear cells (PBMCs). Thus, we determined HIV-1 tropism in both these compartments during primary infection and evaluated the impact of CXCR4-using viruses on disease progression.

Design: One hundred and thirty-three patients with primary HIV-1 infection were screened for HIV-1 coreceptor usage in plasma and PBMCs using both genotypic and phenotypic methods.

Development and performance of a new recombinant virus phenotypic entry assay to determine HIV-1 coreceptor usage.

Background: Clinical trials of CCR5 antagonists have relied on the phenotypic determination of HIV-1 coreceptor usage. Few phenotypic assays are available, with few data on their concordance, and none has been designed to determine tropism from cell-associated HIV-1 DNA.

Objectives: To assess the performance of the new Toulouse Tropism Test (TTT) phenotypic assay to characterize HIV-1 tropism using blood plasma and peripheral blood mononuclear cells (PBMC).