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Introduction: Few data on the diagnostic performance of serological tests for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection are currently available. We evaluated sensitivity and specificity of five different widely used commercial serological assays for the detection of SARS-CoV-2–specific IgG, IgM, and IgA antibodies using reverse transcriptase-PCR assay in nasopharyngeal swab as reference standard test.Methods: A total of 337 plasma samples collected in the period April–June 2020 from SARS-CoV-2 RT-PCR positive (n = 207) and negative (n = 130) subjects were investigated by one point-of-care lateral flow immunochromatographic assay (LFIA IgG and IgM, Technogenetics) and four fully automated assays: two chemiluminescence immunoassays (CLIA-iFlash IgG and IgM, Shenzhen YHLO Biotech and CLIA-LIAISON® XL IgG, DiaSorin), one electrochemiluminescence immunoassay (ECLIA-Elecsys® total predominant IgG, Roche), and one enzyme-linked immunosorbent assay (ELISA IgA, Euroimmune).Results: The overall sensitivity of all IgG serological assays was >80% and the specificity was >97%. The sensitivity of IgG assays was lower within 2 weeks from the onset of symptoms ranging from 70.8 to 80%. The LFIA and CLIA-iFlash IgM showed an overall low sensitivity of 47.6 and 54.6%, while the specificity was 98.5 and 96.2%, respectively. The ELISA IgA yielded a sensitivity of 84.3% and specificity of 81.7%. However, the ELISA IgA result was indeterminate in 11.7% of cases.Conclusions: IgG serological assays seem to be a reliable tool for the retrospective diagnosis of SARS-CoV-2 infection. IgM assays seem to have a low sensitivity and IgA assay is limited by a substantial rate of indeterminate results.

Rationale Targeted cystic fibrosis (CF) therapy with lumacaftor/ivacaftor partly restores chloride channel function and improves epithelial fluid transport in the airways. Consequently, changes may occur in the microbiome, which is adapted to CF lungs. Objectives To investigate the effects of lumacaftor/ivacaftor on respiratory microbial composition and microbial metabolic activity by repeatedly sampling the lower respiratory tract. Methods This was a single-centre longitudinal observational cohort study in adult CF patients with a homozygous Phe508del mutation. Lung function measurements and microbial cultures of sputum were performed as part of routine care. An oral and nasal wash, and a breath sample, were collected before and every 3 months after starting therapy, for up to 12 months. Results Twenty patients were included in this study. Amplicon 16S RNA and metagenomics sequencing revealed that Pseudomonas aeruginosa was most abundant in sputum and seemed to decrease after 6 months of treatment, although this did not reach statistical significance after correction for multiple testing. Two types of untargeted metabolomics analyses in sputum showed a change in metabolic composition between 3 and 9 months that almost returned to baseline levels after 12 months of treatment. The volatile metabolic composition of breath was significantly different after 3 months and remained different from baseline until 12 months follow-up. Conclusions After starting CF transmembrane conductance regulator (CFTR) modulating treatment in CF patients with a homozygous Phe508del mutation, a temporary and moderate change in the lung microbiome is observed, which is mainly characterised by a reduction in the relative abundance of Pseudomonas aeruginosa., Lumacaftor/ivacaftor in adult cystic fibrosis patients with homozygous Phe508del results in temporal and moderate changes in lung microbiome and metabolome, that are mainly characterised by a reduction in the relative abundance of Pseudomonas aeruginosa https://bit.ly/3pcPUfX

[This corrects the article DOI: 10.3389/fmed.2020.00415.].

Microbicides are considered a promising strategy for preventing human immunodeficiency virus (HIV-1) transmission and disease. In this report, we first analyzed the antiviral activity of the miniCD4 M48U1 peptide formulated in hydroxyethylcellulose (HEC) hydrogel in activated peripheral blood mononuclear cells (PBMCs) infected with R5- and X4-tropic HIV-1 strains. The results demonstrate that M48U1 prevented infection by several HIV-1 strains including laboratory strains, and HIV-1 subtype B and C strains isolated from the activated PBMCs of patients. M48U1 also inhibited infection by two HIV-1 transmitted/founder infectious molecular clones (pREJO.c/2864 and pTHRO.c/2626). In addition, M48U1 was administered in association with tenofovir, and these two antiretroviral drugs synergistically inhibited HIV-1 infection. In the next series of experiments, we tested M48U1 alone or in combination with tenofovir in HEC hydrogel with an organ-like structure mimicking human cervicovaginal tissue. We demonstrated a strong antiviral effect in absence of significant tissue toxicity. Together, these results indicate that co-treatment with M48U1 plus tenofovir is an effective antiviral strategy that may be used as a new topical microbicide to prevent HIV-1 transmission.

Antiretroviral therapy has effectively tackled HIV replication and prevented the development of AIDS-related complications in the majority of HIV-positive patients. This pharmacological approach has dramatically increased the life expectancy of HIV-positive subjects transforming HIV infection into a chronic disease. Notwithstanding this major improvement in HIV disease management, several HIV-positive patients show an earlier and significant onset of aging related chronic conditions such as cardiovascular disease, osteoporosis, diabetes and neoplasias with respect to uninfected individuals. In particular, cardiovascular diseases are associated with both HIV infection and antiretroviral treatment, and represent major clinical complications in HIV-positive patients. Here, we discuss the interaction between antiretroviral therapy and cardiovascular system in HIV-positive patients focusing on the antiretroviral-related mechanisms involved in cardiovascular alterations.

The routine determination of drug resistance in newly HIV-1 infected individuals records a potential increase in transmissions of drug-resistant variants. Plasma samples from 38 individuals classified as newly infected (seroconversion time

The possible relationships between the intensity of humoral response to full length Tat protein, the amount of proviral DNA reservoir in peripheral blood mononuclear cells and RNA viral load were analyzed in plasma samples obtained from a group of HIV-1 seropositive subjects, who never received any antiretroviral therapy. All HIV-1 patients showed detectable levels of serum IgG to full-length Tat by immunoenzymatic assay. We found a higher percentage of HIV-1 seropositive subjects with low levels of antibody in the presence of barely detectable proviral DNA copies (or =10 copies/1.5x10(5) PBMCs) and a high anti-Tat antibody response accompanied by variable (from10(1) toor =10(3) copies/1.5x10(5) PBMCs) levels of DNA load (p=0.011). Moreover, an inverse relationship between anti-Tat antibody titers and HIV-1 RNA viral load was demonstrated HIV-1 seropositive patients. In HIV-1-infected patients, a strong humoral immune response against HIV-1 transactivating Tat protein, able to down-modulate viral replication in peripheral blood, does not seem to inhibit the number of proviral DNA molecules in peripheral blood mononuclear cells. Even though our data strongly confirm the "positive" role of anti-Tat antibody on viral replication, the persistence of significant amount of DNA viral load in peripheral blood mononuclear cells, despite high level of anti Tat antibody, suggests a more cautious approach to HIV-1 Tat-containing vaccines, able to stimulate an immune specific response to transactivating Tat protein sufficient in inhibiting circulating virus, but not completely efficient in decreasing proviral DNA integration.

Background The routine determination of drug resistance in newly HIV-1 infected individuals documents a potential increase in the transmission of drug-resistant variants. Plasma samples from twenty seven therapy naive HIV-1 infected Italian patients were analyzed by the line probe assay (LIPA) and the TruGene HIV-1 assay for the detection of mutations conferring resistance to HIV-1. Results Both tests disclosed amino-acid substitutions associated with resistance in a variable number of patients. In particular, two mutations (K70R and V118I), detectable by LIPA and by sequencing analysis respectively, revealed resistance to NRTIs in two plasma samples. At least three mutations conferring resistance to NNRTIs, not detectable by commercial LIPA, able to reveal mutations associated only with nucleoside reverse transcriptase analogues, were disclosed by viral sequence analysis. Moreover, most samples showed mutations correlated with resistance to protease inhibitors. Remarkably, a key mutation, like V82A (found as a mixture), and some "indeterminate" results (9 samples), due the absence of signal on the lines corresponding to a specific probe, was revealed only by LIPA, while a variable number of secondary mutations was detectable only by TruGene HIV-1 assay. Conclusion Even if further studies are necessary to establish the impact of different tests on the evaluation of drug-resistant strains transmission, LIPA might be useful in a wide population analysis, where bulk results are needed in a short time, while sequencing analysis, able to detect mutations conferring resistance to both NRTIs and NNRTIs, might be considered a more complete assay, albeit more expensive and more technically complex.

Standard serological tests (both EIA and Immunoblotting) have reached high levels of sensitivity and reproducibility, but do not indicate whether infection is recent or longstanding. Since many patients with HIV-1 infection are not usually diagnosed until symptom presentation, the possibility to distinguish between acute and chronic infection has become increasingly important for the purposes of therapeutic decision-making, partner notification and epidemiological surveillance. We evaluated a guanidine-based-antibody-avidity assay in a selected group of recent (within six months from seroconversion) and chronic (more than forty eight months) HIV-1 infections in an attempt to shed more light on the significance of the avidity index in establishing the time of infection. Sera from newly infected individuals showed a low mean avidity index (ranging from 0.35 to 0.60 with a standard deviation 0.09) at baseline and a clear increasing value at the following times of observation. Our data showed that an avidity index

Anemia is the most common hematological abnormality in human immunodeficiency virus (HIV)-infected patients. Besides chronic disease, opportunistic infections, nutritional deficiencies and antiretroviral drug toxicity, the direct role of HIV in the development of anemia has not yet been fully investigated. To explore the HIV-related mechanisms involved in the genesis of anemia, we used two experimental designs. In the first, HPCs purified from cord blood were challenged with HIV-1IIIb or recombinant gp120 (rgp120) and then committed to erythrocyte differentiation (EPO-post-treated HPCs). In the second, HPCs were first committed to differentiate towards the erythroid lineage and only afterwards challenged with HIV-1IIIb or rgp120 (EPO-pre-treated HPCs). Our results showed that HPCs and EPO-induced HPCs were not susceptible to HIV-1 infection. In addition, the two experimental designs (EPO post or pre-treated HPCs) independently showed that HIV-1IIIb or rgp120 were able to induce the impairment of survival, proliferation, and differentiation albeit differing in kinetics and extent. Interestingly, the gp120 interaction with CD4 and CXCR4 played a pivotal role in the impairment of erythrocyte differentiation by inducing TGF-b1 expression. These observations reveal an important additional mechanism involved in the genesis of anemia suggesting a complex competition between EPO-positive regulation and HIV-negative priming regarding erythrocyte survival, proliferation and maturation.

Standard serological tests have reached high levels of sensitivity and reproducibility but do not indicate whether infection is recent or long-standing. Among the 59960 sera analyzed for HIV positivity at the Retrovirus Laboratory, Operative Unit of Microbiology, Bologna, Italy, from January 2010 to July 2011, 134 samples showed an initial positive result. Application of the avidity test, able to distinguish between recent or long-standing HIV infection, classified 59 subjects as recently infected and 75 as chronically infected. Besides all the public health implications, the distinction between acute and chronic infection might serve to establish the time of infection and therefore reach any potential partners who might have been infected in a specific period of time. Although our results are limited to subjects referred to our laboratory and hence represent only a limited part of the problem, the routine application of methods able to distinguish recent from long-lasting infection could help monitor disease incidence, identify high-risk groups, and enhance epidemiological conclusions.

Genotypic resistance analysis on viral DNA and plasma was performed in 56 therapy naive patients with recent and chronic infection to assess the prevalence of mutations associated with drug resistance and compare cell-free and cell-associated strains. Direct sequencing of DNA provirus disclosed key mutations to RT inhibitors more frequently than in plasma RNA. In addition, major mutations associated with drug resistance in the PR region were only found in PBMCs. Although our data are limited to a small cohort, they show a different resistance profile between plasma and PBMC compartments and may yield additional information for first-line antiretroviral regimens.

It is crucial to establish the timing of infection and distinguish between early and long-lasting HIV-1 infections not only for partner notification and epidemiological surveillance, but also to offer early drug treatment and contain the spread of infection. This study analyzed serum and/or plasma samples with a first positive HIV antibody/antigen result coming from different Medical Centers in the Emilia Romagna Region, North East Italy, using the avidity assay, Western Blotting, RNA viral load, CD4 cell counts and genotyping assay. From May 2013 to May 2016, we certified 845 new HIV-1 infections, 18.7% of which were classified on the basis of avidity index as recent infections and 81.3% as long-lasting infections, with an estimated conversion time exceeding six months at the time of study. Western Blotting showed reactivity to only one or two HIV-1 proteins in recently infected patients (RIPs), while a complete pattern to gag, env and pol proteins was observed in most long-lasting infected patients (LLIPs). The median age, gender, nationality and risk transmission factors were comparable in RIPs and LLIPs. Phylogenetic analysis performed in available plasma disclosed B strains, non-B subtypes and circulating recombinant forms (CRFs) in both groups of patients, with a major presence of CRFs in non-Italian HIV subjects. The large number of patients unaware of their HIV status makes it crucial to discover hidden epidemics and implement appropriate targeted public health interventions.

Since the discovery of 3'-azido-3'deoxthymidine (zidovudine) as an effective antiretroviral agent against human immunodeficiency virus type 1 (HIV-1), drug therapy has been widely used in the treatment of AIDS. To date, new combination therapies have significantly altered the longterm prognosis for HIV-infected patients showing a reduction of plasma viral load, associated with clinical and immunological recovery. Nevertheless, in various circumstances treatment can fail for several reasons, such as patient noncompliance with the therapeutic regimen, suboptimal antiviral drug concentrations, drug pharmacokinetics, and virus resistance to one or more drugs. Virus drug resistance is the most important factor contributing to the failure of antiretroviral therapy. Since some evidence indicates that viral resistance and treatment failure are closely linked, this brief review explores the routine determination of drug resistance and its importance to shed more light on the meaning of mutations correlated to drug resistance.

We describe the case of a young woman with an acute HIV infection characterized at onset by neurological features. The patient spontaneously controlled her HIV infection and recovered in a short period of time. The patient's clinical and virological history showed a peculiar evolution of HIV infection, with an MDR HIV-1 in CSF and a wild HIV strain in PBMCs. The patient's PBMC showed a rapid shift from a wild type to an MDR strain in few days.

In this study, we showed the existence of a positive correlation between the amount of human immunodeficiency virus-type 1 (HIV-1) RNA in HIV-1 seropositive subjects and the plasma levels of TRAIL. Since it has been previously demonstrated that HIV-1 Tat protein up-regulates the expression of TRAIL in monocytic cells whereas tat-expressing lymphoid cells are more resistant to TRAIL cytotoxicity, we next investigated the effect of Tat on the expression/activity of both apical caspase-8 and -10, which play a key role in mediating the initial phases of apoptosis by TRAIL, and c-FLIP. Jurkat lymphoblastoid human T cell lines stably transfected with a plasmid expressing wild-type (HIV-1) tat gene showed normal levels of caspase-8 but significantly decreased levels of caspase-10 at both mRNA and protein levels with respect to Jurkat transfected with the control plasmid or with a mutated (cys22) non-functional tat cDNA. A significant decrease of caspase-10 expression/activity was also observed in transient transfection experiments with plasmid carrying tat cDNA. Moreover, c-FLIP(L) and c-FLIP(S) isoforms were up-regulated in tat-expressing cells at both mRNA and protein level in comparison with control cells. Taken together, these results provide a molecular basis to explain the resistance of tat-expressing Jurkat cells to apoptosis induced by TRAIL and, possibly, to other death-inducing ligands. Copyright 2004 Wiley-Liss, Inc.

HIV-infected patients have a significantly higher risk of developing cardiovascular events during the progression of HIV disease. Atherosclerosis, myocardial infarction, cerebrovascular injury, pulmonary hypertension and thrombosis are consistently described in both combined antiretroviral therapy (cART)-treated and naive HIV-positive patients as major clinical complications. Recent studies indicate that the pathogenesis of cardiovascular lesions in HIV-positive patients is related to direct and indirect effects of HIV infection on vessel structures, independently of traditional risk factors. HIV infection strongly interferes with the biology of several cellular targets such as macrophage and endothelial cells. Moreover, HIV induces a profound derangement of lipid metabolism and inflammatory cytokine networks that are directly involved in atherogenesis and progressive impairment of the cardiovascular system.In this review, we discuss these major HIV-related mechanisms able to promote atherosclerosis and cardiovascular diseases in HIV-positive patients.