HIV Infection of Naturally Occurring and Genetically Reprogrammed Human Regulatory T-cells

A T-cell subset, defined as CD4(+)CD25(hi) (regulatory T-cells [Treg cells]), was recently shown to suppress T-cell activation. We demonstrate that human Treg cells isolated from healthy donors express the HIV-coreceptor CCR5 and are highly susceptible to HIV infection and replication. Because Treg cells are present in very few numbers and are difficult to expand in vitro, we genetically modified conventional human T-cells to generate Treg cells in vitro by ectopic expression of FoxP3, a transcription factor associated with reprogramming T-cells into a Treg subset. Overexpression of FoxP3 in naïve human CD4(+) T-cells recapitulated the hyporesponsiveness and suppressive function of naturally occurring Treg cells. However, FoxP3 was less efficient in reprogramming memory T-cell subset into regulatory cells. In addition, FoxP3-transduced T-cells also became more susceptible to HIV infection. Remarkably, a portion of HIV-positive individuals with a low percentage of CD4(+) and higher levels of activated T-cells have greatly reduced levels of FoxP3(+)CD4(+)CD25(hi) T-cells, suggesting disruption of the Treg cells during HIV infection. Targeting and disruption of the T-cell regulatory system by HIV may contribute to hyperactivation of conventional T-cells, a characteristic of HIV disease progression. Moreover, the ability to reprogram human T-cells into Treg cells in vitro will greatly aid in decoding their mechanism of suppression, their enhanced susceptibility to HIV infection, and the unique markers expressed by this subset

Evidence for Histidine-Rich Protein 2 Immune Complex Formation in Symptomatic Patients in Southern Zambia

Background: Rapid diagnostic tests based on histidine-rich protein 2 (HRP2) detection are the primary tools used to detect Plasmodium falciparum malaria infections. Recent conflicting reports call into question whether α-HRP2 antibodies are present in human host circulation and if resulting immune complexes could interfere with HRP2 detection on malaria RDTs. This study sought to determine the prevalence of immune-complexed HRP2 in a low-transmission region of Southern Zambia. Methods: An ELISA was used to quantify HRP2 in patient sample DBS extracts before and after heat-based immune complex dissociation. A pull-down assay reliant on proteins A, G, and L was developed and applied for IgG and IgM capture and subsequent immunoprecipitation of any HRP2 present in immune complexed form. A total of 104 patient samples were evaluated using both methods. Results: Immune-complexed HRP2 was detectable in 17% (18/104) of all samples evaluated and 70% (16/23) of HRP2-positive samples. A majority of the patients with samples containing immune-complexed HRP2 had P. falciparum infections (11/18) and were also positive for free HRP2 (16/18). For 72% (13/18) of patients with immune-complexed HRP2, less than 10% of the total HRP2 present was in immune-complexed form. For the remaining samples, a large proportion (≥ 20%) of total HRP2 was complexed with α-HRP2 antibodies. Conclusions: Endogenous α-HRP2 antibodies form immune complexes with HRP2 in the symptomatic patient population of a low-transmission area in rural Southern Zambia. For the majority of patients, the percentage of HRP2 in immune complexes is low and does not affect HRP2-based malaria diagnosis. However, for some patients, a significant portion of the total HRP2 was in immune-complexed form. Future studies investigating the prevalence and proportion of immune-complexed HRP2 in asymptomatic individuals with low HRP2 levels will be required to assess whether α-HRP2 antibodies affect HRP2 detection for this portion of the transmission reservoir

Identification of a CCR5-Expressing T Cell Subset That Is Resistant to R5-Tropic HIV Infection

Infection with HIV-1 perturbs homeostasis of human T cell subsets, leading to accelerated immunologic deterioration. While studying changes in CD4(+) memory and naïve T cells during HIV-1 infection, we found that a subset of CD4(+) effector memory T cells that are CCR7(−)CD45RO(−)CD45RA(+) (referred to as T(EMRA) cells), was significantly increased in some HIV-infected individuals. This T cell subset displayed a differentiated phenotype and skewed Th1-type cytokine production. Despite expressing high levels of CCR5, T(EMRA) cells were strikingly resistant to infection with CCR5 (R5)–tropic HIV-1, but remained highly susceptible to CXCR4 (X4)–tropic HIV-1. The resistance of T(EMRA) cells to R5-tropic viruses was determined to be post-entry of the virus and prior to early viral reverse transcription, suggesting a block at the uncoating stage. Remarkably, in a subset of the HIV-infected individuals, the relatively high proportion of T(EMRA) cells within effector T cells strongly correlated with higher CD4(+) T cell numbers. These data provide compelling evidence for selection of an HIV-1–resistant CD4(+) T cell population during the course of HIV-1 infection. Determining the host factors within T(EMRA) cells that restrict R5-tropic viruses and endow HIV-1–specific CD4(+) T cells with this ability may result in novel therapeutic strategies against HIV-1 infection

Multi-center Evaluation of the Cepheid Xpert® Xpress SARS-CoV-2/Flu/RSV Test.

With the approach of respiratory virus season in the Northern Hemisphere, clinical microbiology and public health laboratories will need rapid diagnostic assays to distinguish SARS-CoV-2 from influenza and respiratory syncytial virus (RSV) infections for diagnosis and surveillance. In this study, the clinical performance of the Xpert® Xpress SARS-CoV-2/Flu/RSV test (Cepheid, Sunnyvale, CA, USA) for nasopharyngeal swab specimens was evaluated in four centers: Johns Hopkins Medical Microbiology Laboratory, Northwell Health Laboratories, NYC Public Health Laboratory, and Los Angeles County/University of Southern California (LAC+USC) Medical Center. A total of 319 nasopharyngeal swab specimens, positive for SARS-CoV-2 (n = 75), influenza A (n = 65), influenza B (n = 50), RSV (n = 38), or negative (n = 91) by the standard of care nucleic acid amplification tests at each site were tested using the Cepheid panel test. The overall positive percent agreement for the SARS-CoV-2 target was 98.7% (n= 74/75) and the negative agreement was 100% (n= 91) with all other analytes showing 100% total agreement (n= 153). Standard of care tests to which the Cepheid panel was compared included the Cepheid Xpert Xpress SARS-CoV-2, Cepheid Xpert Xpress Flu/RSV, the GenMark ePlex respiratory panel, the BioFire Respiratory panels 2.1 and v1.7, the DiaSorin Simplexa COVID-19 Direct, and the Hologic Panther Fusion SARS-CoV-2 assays. The Xpert Xpress SARS-CoV-2/Flu/RSV test showed high sensitivity and accuracy for all analytes included in the test. This test will provide a valuable clinical diagnostic and public health solution for detecting and differentiating SARS-CoV-2, influenza A and B, and RSV infections during the current respiratory virus season

Cytokine Production by FoxP3-Transduced T-cells

<p>CD4⁺ naïve T-cells isolated from CB (CB-naïve) and AB (AB-naïve) and memory T-cells from AB (AB-memory) were transduced with HDV or HDV.FoxP3 as described in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0020198#pbio-0020198-g005" target="_blank">Figure 5</a>. Transduced T-cells were purified through magnetic sorting of mCD24⁺ cells and activated using plate-bound anti-CD3 and soluble anti-CD28 antibodies. Supernatants were collected at 18–24 h postactivation and analyzed for (A) IL-2 production or (B) IFNγ, IL-4, and IL-5 production from HDV or HDV.FoxP3-tranduced naïve T-cells, using CBA assay. The results represent five separate experiments from different donors with similar relative differences in cytokine production.</p