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Background The aim of this large multicenter study was to determine variations in cerebrospinal fluid (CSF) HIV-RNA in different phases of untreated human immunodeficiency virus type 1 (HIV-1) infection and its associations with plasma HIV-RNA and other biomarkers. Methods Treatment naive adults with available CSF HIV-RNA quantification were included and divided into groups representing significant disease phases. Plasma HIV-RNA, CSF white blood cell count (WBC), neopterin, and albumin ratio were included when available. Results In total, 1018 patients were included. CSF HIV-RNA was in median (interquartile range [IQR]) 1.03 log10 (0.37–1.86) copies/mL lower than in plasma, and correlated with plasma HIV-RNA (r = 0.44, P plasma discordance was found in 126 of 972 individuals (13%) and varied between groups, from 1% in primary HIV, 11% in neuroasymptomatic groups, up to 30% of patients with HAD. Conclusions Our study confirms previous smaller observations of variations in CSF HIV-RNA in different stages of HIV disease. Overall, CSF HIV-RNA was approximately 1 log10 copies/mL lower in CSF than in plasma, but CSF discordance was found in a substantial minority of subjects, most commonly in patients with HAD, indicating increasing CNS compartmentalization paralleling disease progression.

The risk of venous thromboembolism (VTE) and of recurrent VTE remain elevated in people living with HIV compared to controls still with contemporary antiretroviral therapy (ART). The pathophysiology of VTE in HIV is multi factorial and includes an interplay among traditional risk factors, HIV-specific factors, behavioral factors, exposure to ART and other therapies, coinfections, and co-morbidities.

Summary Objective HIV-infection increases the risk to progress to active-tuberculosis (TB). Detection of latent TB infection (LTBI) is needed to eventually propose preventive-therapy and reduce TB reservoir. QuantiFERON-TB Plus (QFT-Plus)-test identifies LTBI. Currently, only two studies on QFT-Plus accuracy in HIV-infected-population are available in high TB-endemic-countries. Therefore we aimed to evaluate the effect of HIV-infection on QFT-Plus accuracy to detect LTBI in a low TB-endemic-country. Methods We enrolled 465 participants, among the 167 HIV-infected-persons: 32 with active-TB (HIV-TB), 45 remote-LTBI (HIV-LTBI) and 90 at low M. tuberculosis (Mtb)-infection risk. Among the 298 HIV-uninfected-persons: 170 with active-TB, 76 recent-LTBI, 34 remote-LTBI and 18 with low Mtb-infection risk. Results QFT-Plus sensitivity was similar in TB regardless of HIV-status. CD4-count did not influence the distribution of IFN-γ values in HIV-TB and HIV-LTBI. Moreover HIV-LTBI and HIV-uninfected remote LTBI had a similar proportion of results in the uncertain range (IFNγ ≥0.2 ≤ 0.7 IU/ml) differently from those LTBI-persons reporting recent-exposure (p = 0.016). Cytometry results demonstrated that CD8-response was similar in HIV-infected- and -uninfected-persons whereas CD4-response was impaired in HIV-infected-persons (p = 0.011). Conclusions HIV-infection does not affect QFT-Plus response in active-TB, whereas the time of exposure influences the proportion of uncertain-results in LTBI.

ObjectivesOur aim was to investigate the durability of different initial regimens in patients starting ART with CD4+ counts 5 log10 copies/mL.MethodsThis was a retrospective study of HIV-infected patients prospectively followed in the ICONA cohort. Those who started ART with boosted protease inhibitors (bPIs), NNRTIs or integrase strand transfer inhibitors (InSTIs), with CD4+ 5 log10 copies/mL, were included. The primary endpoint was treatment failure (TF), a composite endpoint defined as virological failure (VF, first of two consecutive HIV-RNA >50 copies/mL after 6 months of treatment), discontinuation of class of the anchor drug or death. Independent associations were investigated by Poisson regression analysis in a model including age, gender, mode of HIV transmission, CDC stage, HCV and HBV co-infection, pre-treatment HIV-RNA, CD4+ count and CD4+/CD8+ ratio, ongoing opportunistic disease, fibrosis FIB-4 index, estimated glomerular filtration rate, haemoglobin, platelets, neutrophils, calendar year of ART initiation, anchor drug class (treatment group) and nucleos(t)ide backbone.ResultsA total of 1195 patients fulfilled the inclusion criteria: 696 started ART with a bPI, 315 with an InSTI and 184 with an NNRTI. During 2759 person-years of follow up, 642 patients experienced TF. Starting ART with bPIs [adjusted incidence rate ratio (aIRR) (95% CI) 1.62 (1.29–2.03) versus starting with NNRTIs; P ConclusionsIn patients starting ART with 5 log10 HIV-RNA copies/mL, the durability of regimens based on InSTIs was longer than that of NNRTI- and bPI-based regimens.

Introduction: The use of steroid therapy in patients within the context of SARS-CoV-2 infection is still a matter of debate. This study aimed to evaluate if potential steroid benefits could be predicted by the ratio of arterial oxygen partial pressure (PaO2 in mmHg) to fractional inspired oxygen (FiO2) (P/F) in COVID-19 patients at admission. Materials and Methods: Medical records were retrospectively collected from all adult patients admitted because of COVID-19 from 29 January to 31 July 2020. The association of steroid therapy with 28-day all-cause mortality outcome was analysed in a multivariable logistic regression model adjusted for confounding factors. Results: Overall, 511 patients were analysed, of which 39.1% underwent steroid therapy. Steroid treated patients were mostly male, older, and more frequently treated with antiviral drugs and aminoquinolines, the most common comorbidities were hypertension, followed by cardiovascular disease. Overall, 51 patients died within 28-days, and overall 28-days mortality was 19.5% in the cohort of patients exposed to steroids versus 3.9% mortality in unexposed patients (p &lt, 0.001). Steroid therapy on patients with P/F ratio of 235 mmHg or higher at admission can be considered as detrimental, with an 8% increased probability of death. Conclusions: Steroid therapy is associated with increased 28-day mortality in COVID-19 in patients with mild or no ARDS.

Objectives: To investigate the association between sex hormones and the severity of coronavirus disease 2019 (COVID-19). Furthermore, associations between sex hormones and systemic inflammation markers, viral shedding and length of hospital stay were studied. Design and methods: This case–control study included a total of 48 male patients with COVID-19 admitted to an Italian reference hospital. The 24 cases were patients with PaO2/FiO2 300 mmHg at all times and who may have required low-flow oxygen supplementation during hospitalization (mild COVID-19). For each group, sex hormones were evaluated on hospital admission. Results: Patients with severe COVID-19 (cases) had a significantly lower testosterone level compared with patients with mild COVID-19 (controls). Median total testosterone (TT) was 1.4 ng/mL in cases and 3.5 ng/mL in controls (P = 0.005); median bioavailable testosterone (BioT) was 0.49 and 1.21 in cases and controls, respectively (P = 0.008); and median calculated free testosterone (cFT) was 0.029 ng/mL and 0.058 ng/mL in cases and controls, respectively (P = 0.015). Low TT, low cFT and low BioT were correlated with hyperinflammatory syndrome (P = 0.018, P = 0.048 and P = 0.020, respectively) and associated with longer length of hospital stay (P = 0.052, P = 0.041 and P = 0.023, respectively). No association was found between sex hormone level and duration of viral shedding, or between sex hormone level and mortality rate. Conclusions: A low level of testosterone was found to be a marker of clinical severity of COVID-19.

Background Few data about predictors and outcomes associated with prolonged SARS-CoV-2 RNA shedding (VS) are available. Methods Retrospective study including all patients admitted with COVID-19 in an Italian reference hospital for infectious diseases between March 1 and July 1, 2020. Predictors of viral clearance (VC) and prolonged VS from upper respiratory tract were assessed by Poisson regression and logistic regression analyses. The causal relation between duration of VS and probability of clinical outcomes was evaluated through inverse probability weighted Cox model. Results 536 subjects were included. Median duration of VS from symptoms onset was 18 days (IQR 12-26). The estimated 30-day probability of VC was 70.2% (95%CI:65-75). At multivariable analysis, patients with comorbidities (aIRR = 0.88, p = 0.004), lymphopenia at hospital admission (aIRR = 0.75, p = 0.032) and with moderate/severe respiratory disease (aIRR = 0.42, p 1000 ng/mL at admission (aOR = 1.76, p = 0.035) independently predicted prolonged VS. The achievement of VC doubled the chance of clinical recovery (aHR = 2.17, p

Molecular investigation of primary HIV infections (PHI) is crucial to describe current dynamics of HIV transmission. Aim of the study was to investigate HIV transmission clusters (TC) in PHI referred during the years 2013&ndash, 2020 to the National Institute for Infectious Diseases in Rome (INMI), that is the Lazio regional AIDS reference centre, and factors possibly associated with inclusion in TC. These were identified by phylogenetic analysis, based on population sequencing of pol, a more in depth analysis was performed on TC of B subtype, using ultra-deep sequencing (UDS) of env. Of 270 patients diagnosed with PHI during the study period, 229 were enrolled (median follow-up 168 (IQR 96&ndash, 232) weeks). Median age: 39 (IQR 32&ndash, 48) years, 94.8% males, 86.5% Italians, 83.4% MSM, 56.8% carrying HIV-1 subtype B. Of them, 92.6% started early treatment within a median of 4 (IQR 2&ndash, 7) days after diagnosis, median time to sustained suppression was 20 (IQR 8&ndash, 32) weeks. Twenty TC (median size 3, range 2&ndash, 9 individuals), including 68 patients, were identified. A diagnosis prior to 2015 was the unique factor associated with inclusion in a TC. Added value of UDS was the identification of shared quasispecies components in transmission pairs within TC.

Background Little evidence on COVID‐19 in people living with HIV (PLWH) is currently available. Material and Methods We reported clinical and viro‐immunological data of all HIV‐positive patients admitted to our centre with COVID‐19 from March 1 to May 12,2020. Results Overall, five patients were included: all were virologically‐suppressed on antiretroviral therapy and CD4+ count was >350 cell/mm3 in all but two patients. Although all patients had evidence of pneumonia on admission, only one developed respiratory failure. SARS‐CoV‐2‐RNA was never detected from nasopharyngeal swabs in two patients, whereas, in the others, viral clearance occurred within a maximum of 43 days. IgG production was elicited in all patients and neutralizing antibodies in all but one patient. Specific‐T‐cell response developed in all patients but was stronger in those with more severe presentation. Similarly, the highest level of pro‐inflammatory cytokines was found in the only patient experiencing respiratory failure. Despite a mild presentation, patients with more pronounced immunosuppression showed high degrees of both cytokines production and immune‐activation. Conclusions Our study did not find an increased risk and severity of COVID‐19 in PLWH. Adaptative cellular immune response to SARS‐CoV‐2 appeared to correlate to disease severity. The mild clinical picture showed in advanced HIV patients, despite a significant T‐cell activation and inflammatory profile, suggests a potential role of HIV‐driven immunological dysregulation in avoiding immune‐pathogenetic processes. However, other possible explanations, as a protective role of certain antiretroviral drugs, should be considered. Further larger studies are needed to better clarify the impact of HIV infection on COVID‐19. This article is protected by copyright. All rights reserved.

Background Human Immunodeficiency Virus (HIV) infection is a risk factor for tuberculosis (TB). Antiretroviral therapy (ART) changed HIV clinical management but it is still unclear how pre-existing HIV/Mycobacterium tuberculosis (Mtb)-specific CD4+ and CD8+ T-cells are restored. Aim to evaluate the impact of ART and TB therapies on the functional and phenotypic profile of Mtb-specific antigen-response of CD4+ and CD8+ T-cells in prospectively enrolled HIV-TB co-infected patients. Methods ART-naive HIV-infected patients, with or without active TB or latent TB infection (LTBI), were enrolled before and after starting ART and TB therapies. Peripheral blood mononuclear cells (PBMC) were stimulated overnight with Mtb and HIV antigens (GAG). Cytokine expression and phenotype profile were evaluated by flow cytometry. Cytomegalovirus (CMV) and staphylococcal enterotoxin B (SEB) were also used. Results The median of absolute number of CD4+ T-cells increased after ART and TB therapies in all groups analyzed, while the median of absolute number of CD8+ T-cells decreases in HIV and HIV-LTBI groups. Treatments significantly increased the frequency of Mtb-specific CD4+ T-cells in the HIV-LTBI (p = 0.015) with a rise of the central memory compartment. The magnitude of the CD4+ T-cell response to HIV-GAG significantly increased in active TB (p = 0.03), whereas the magnitude of CMV-specific CD4+ T-cell response decreased in all the groups. Similarly, the treatments increased the number of Mtb-specific CD8+ responders in both HIV-LTBI and HIV-TB groups, whereas the phenotype distribution was dependent on the antigens used and on the stage of infection/disease. Conclusions After therapies the median of absolute number and the proportion of CD4+ T-cells increased in all groups whereas the median of absolute count and proportion of CD8+ T-cells decreased in the HIV and HIV-LTBI subjects. Interestingly, an increased frequency of CD4+ T-cell response to RD1 proteins in HIV-LTBI subjects was found. These results contribute to a better understanding of the effect of ART and TB therapies on the modulation of Mtb-specific CD4+ and CD8+ T-cells subsets.

Abacavir is a widely used nucleotide reverse transcriptase inhibitor, for which cerebrospinal fluid (CSF) exposure has been previously assessed in twice-daily recipients. We studied abacavir CSF concentrations in 61 and nine HIV-positive patients taking abacavir once daily and twice daily, respectively. Patients on once-daily abacavir had higher plasma and CSF concentrations (96 vs. 22 ng ml-1 , P = 0.038 and 123 vs. 49 ng ml-1 , P = 0.038) but similar CSF-to-plasma ratios (0.8 vs. 0.5, P = 0.500). CSF abacavir concentrations were adequate in patients receiving once-daily treatment.

The impact of early antiretroviral therapy (ART) during Primary HIV Infection (PHI) on the hematopoietic progenitor cells (HPCs) homeostasis is not available. This study aimed to characterize HPCs and their relationship with cytokines regulating progenitors function in ART-treated patients with PHI. We enrolled HIV infected patients treated with ART during PHI. Circulating HPCs, Lymphoid-HPCs (L-HPCs) frequency and plasmatic concentrations of IL-7, IL-18 and Stem Cell Factor (SCF) were analysed at baseline and after 6 months of therapy. ART introduction during PHI restored the decline of L-HPCs, induced a decrease in the level of pro-inflammatory IL-18 cytokine and a parallel increase of SCF. Moreover, L-HPCs frequency positively correlated with IL-18 at baseline, and with SCF after 6 months of therapy, suggesting that different signals impact L-HPCs expansion and maintenance before and after treatment. Finally, the SCF receptor expression on HPCs decreased after early ART initiation. These insights may open new perspectives for the evaluation of cytokine-driven L-HPCs expansion and their impact on the homeostasis of hematopoietic compartment during HIV infection.

Background: Hepatitis C virus (HCV) chronic infection has been associated with increased risk of non-Hodgkin lymphoma (NHL) in people living with human immunodeficiency virus (HIV) as well as with a trend of inferior overall survival (OS) in HIV-associated NHL in the modern antiretroviral therapy (ART) era (Besson 2020). The recent introduction of interferon (IFN)-free direct-acting antivirals (DAAs) led to the achievement of sustained virologic response (SVR) in nearly all treated patients (pts) with negligible toxicity in all settings, including HIV/HCV coinfected pts, in which, however, careful attention to interactions with ART is required. We recently showed that DAAs' administration after immuno-chemotherapy (I-CT) may improve long-term outcome in HIV-negative HCV-associated diffuse large B-cell lymphomas (DLBCL) pts (Merli 2019), however, only scant data have been reported so far about the use of DAAs in HIV/HCV coinfected NHL pts. METHODS: We retrospectively collected clinical and virological features, treatments and outcome data of all consecutive pts with NHL and HIV/HCV co-infection, diagnosed and treated at 13 Italian centers between 2005 and 2021, with a special focus on pts affected by DLBCL and treated with DAAs. Only pts who received ART were included. The primary endpoints were SVR rate after DAAs and 2-year OS in HIV/HCV-positive NHL pts. RESULTS: Overall, we collected data of 74 HIV/HCV coinfected pts with NHL (69 males, 93%), including 52 DLBCL, 13 Burkitt lymphoma (BL), 5 plasmablastic lymphoma (PL), 1 anaplastic large-cell ALK negative, 1 T lymphoblastic, 1 gastric MALT and 1 lymphoplasmacytic lymphoma (Table 1). Median age was 51 years (22-57). Previous AIDS defining event was recorded in 16 cases (22%). The main HIV transmission group was represented by intravenous drug users (69%). Stage was III-IV in 63 pts (85%) and aaIPI was ≥2 in 57 pts (77%). At NHL diagnosis 38% of pts had CD4+ DAAs use was associated with improved OS in all pts (p=0.01) and in DLBCL (p=0.04) and with better PFS (p=0.01) in all pts. Similarly, the achievement of SVR after either DAAs or IFN predicted a better OS (Fig.2) and PFS in all pts (p=0.005 and p=0.008, respectively) and in DLBCL (p=0.018 and p=0.047, respectively). The impact of DAAs and SVR on OS remained significant also if considering only pts who achieved CR or PR after I-CT (p At univariate analysis, age >60 years (p=0.02), ARL-IPI (p=0.013), PS ECOG ≥2 (p=0.018) were associated with inferior OS. By applying multivariate Cox regression analysis, age >60 years (HR 67.9, 95% CI 7.2- 643.3, p CONCLUSIONS: In this very high risk series of HIV/HCV coinfected pts with NHL, mainly represented by DLBCL, the administration of DAAs after I-CT resulted feasible and effective (SVR 95%), and displayed an independent favourable influence on OS. These results strongly support DAAs' use in this hard to treat population. Figure 1 Figure 1. Disclosures Tisi: Incyte: Membership on an entity's Board of Directors or advisory committees; BWS: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; Gilead: Honoraria, Membership on an entity's Board of Directors or advisory committees. Zilioli: Roche, Italfarmaco: Consultancy, Honoraria; MSD, Janssen: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations; Takeda: Other: travel expenses, accommodation; Gentili, Takeda, Gilead, Servier: Consultancy, Speakers Bureau. Passamonti: AbbVie: Speakers Bureau; Celgene: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; BMS: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Janssen: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Novartis: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Arcaini: Celgene, Roche, Janssen-Cilag, Gilead: Other: Travel expenses; Bayer, Celgene, Gilead Sciences, Roche, Sandoz, Janssen-Cilag, VERASTEM: Consultancy; Gilead Sciences: Research Funding; Celgene: Speakers Bureau.

Background: Immunological nonresponse represents the Achilles heel in the combination antiretroviral therapy (cART) effectiveness, and increases risk of clinical events and death. CD8 T cells play a crucial role in controlling HIV replication, and polyfunctional HIV-specific CD8 T cells have been associated with nonprogressive HIV infection. However, the possible role of polyfunctional CD8 T cells in predicting posttreatment immune reconstitution has not yet been explored. The aim of this study was to identify functional markers predictive of immunological response to cART in chronic HIV-infected patients. Methods: A cohort of chronic HIV-infected individuals naive to cART were enrolled in the ALPHA study. CD4/CD8 T-cell subsets, their differentiation/activation, as well as susceptibility to apoptosis were analyzed before and after 12 months of cART. Moreover, CD8 T cells polyfunctional response after HIV antigenic stimulation was also assessed. Results: Results showed a significant correlation between worse CD4 T-cell restoration and low frequency of naive CD4 T cells, high frequency of effector memory CD4 T cells, and high susceptibility to apoptosis of CD4 T cells all before cART. Moreover, CD8 functional subsets expressing total C-C motif chemokine ligand 4 (CCL-4) or in combination with CD107a and interferon gamma (IFNγ) were negatively associated with immune reconstitution. Conclusions: In conclusion, our study shows that a more differentiated phenotype of CD4 T cells and CCL-4-producing CD8 T cells could represent valuable predictors of worse immune reconstitution. These parameters may be used as tools for identifying patients at risk of immunological failure during cART and eventually represent the basis for innovative therapeutic strategies.

Background: Few studies have evaluated in pregnant women with HIV the prevalence of smoking and its associations with maternal and neonatal outcomes. Objectives: to assess the prevalence of smoking among women with HIV in early pregnancy and the association between smoking and pregnancy outcomes in this particular population. Methods: We used data from a multicenter observational study to define the prevalence of smoking in women with HIV in early pregnancy, and the role of smoking status and intensity as risk factors for adverse maternal and neonatal outcomes. Main outcome measures were fetal growth restriction [FGR], preterm delivery [PD] and low birthweight [LB], evaluated in univariate and multivariate analyses. Results: The overall (2001-2018) prevalence of reported smoking (at least one cigarette/day) was 25.6% (792/3097), with a significant decrease in recent years (19.0% in 2013-2018). Women who smoked were less commonly African, had lower body mass index, older age, a longer history of HIV infection and higher CD4 counts. In univariate analyses, smokers were significantly more likely to have PD, LB, FGR and detectable HIV viral load at third trimester. Multivariable analyses confirmed for smokers a significantly higher risk of LB (adjusted odds ratio [AOR]: 1.69, 95%CI 1.22-2.34) and FGR (AOR 1.88, 95%CI 1.27-2.80), while the associations with detectable HIV and PD were not maintained. Conclusions: The common prevalence of smoking among pregnant women with HIV and its association with adverse outcomes indicates that smoking cessation programs in this population may have a significant impact on neonatal and maternal health.

ObjectivesTo evaluate the prevalence and therapeutic relevance of drug resistance among isolates from ART-experienced HIV-1-infected patients over the past two decades in Italy.MethodsDynamics of resistance to one, two and three or more antiretroviral classes were evaluated from 1999–2018. Virological success (VS) after the latest therapy switch was evaluated according to cumulative class resistance and cumulative genotypic susceptibility score (Stanford HIV_DB algorithm).ResultsAmong 13 663 isolates (from 6739 patients), resistance to at least one drug class decreased sharply from 1999 to 2010 (≤2001, 84.6%; 2010, 43.6%; P ConclusionsA dramatic drop of HIV-1 drug resistance at failure has been achieved over the last two decades in Italy; resistance to three or more classes is low but present among currently failing patients. Its management still requires a rational and careful diagnostic and therapeutic approach.

The aim of this study was to evaluate the factors that can influence an incomplete viral response (IVR) after acute and early HIV infection (AEHI).This was a retrospective, observational study including patients with AEHI (Fiebig stages I-V) diagnosed between January 2008 and December 2014 at 20 Italian centres. IVR was defined by: (1) viral blip (51-1000 HIV-1 RNA copies/mL after achievement of 50 HIV-1 RNA copies/mL); (2) virologic failure [ 1000 copies/mL after achievement of 200 copies/mL, or ≥ 200 copies/mL after 24 weeks on an antiretroviral therapy (ART)]; (3) suboptimal viral response ( 50 copies/mL after 48 weeks on ART or two consecutive HIV-1 RNA levels with ascending trend during ART). Cox regression analysis was used to calculate the hazard ratios (HRs) and 95% confidence intervals (95% CIs) for IVR.In all, 263 patients were studied, 227 (86%) males, with a median [interquartile range (IQR)] age of 38 (30-46) years. During a median follow-up of 13.0 (5.7-31.1) months, 38 (14.4%) had IVR. The presence of central nervous system (CNS) symptoms was linked to a higher risk of IVR (HR = 4.70, 95% CI: 1.56-14.17), while a higher CD4/CD8 cell count ratio (HR = 0.13, 95% CI: 0.03-0.51 for each point increase) and first-line ART with three-drug regimens recommended by current guidelines (HR = 0.40, 95% CI: 0.18-0.91 compared with other regimens including four or five drugs, older drugs or non-standard backbones) were protective against IVR.Patients with lower CD4/CD8 ratio and CNS symptoms could be at a higher risk of IVR after AEHI. The use of recommended ART may be relevant for improving short-term viral efficacy in this group of patients.

Highlights • Impact of coinfection with SARS-CoV-2 on HIV replication in CNS is unknown. • SARS-CoV-2 could alter the BBB permeability and cause a CNS HIV escape. • Few symptoms during COVID-19 suggest a contained inflammatory response in CNS/plasma. • Poor immunological recovery might contribute to avoid immune-pathogenetic processes., We report two cases of HIV positive patients with COVID-19 infection and a recent diagnosis of opportunistic infections of central nervous system (CNS). We investigated the potential impact of coinfection with SARS-CoV-2 on HIV replication in CNS.

Antiretroviral treatment (ART) of Primary HIV Infection (PHI) has demonstrated virological and immunological benefits. The effect of early ART during PHI on the level of growth factors and chemokines modulating immune cell functions remains to be established. The aim of our work was to analyze the dynamics of 27 cytokines, chemokines and growth/regulation factors in plasma of HIV infected patients treated during PHI. Patients with PHI (n = 43) were enrolled before, 24 and 48 weeks after therapy initiation. Quantification of soluble immune mediators was performed in plasma from HIV infected patients and healthy donors (HD, n = 7) by Luminex technology. The cytokines profile was strongly perturbed in primary HIV infected patients when compared to healthy donors (HD). After 48 weeks of ART, some of these factors were restored to HD level (IL-2, IL-5, IL-7, IL-9, IL12p70, TNFα) while others persisted higher than HD (IL-6, IL-10, IL-13). Interestingly, a subset of chemokines, such as IL-8, MCP-1, RANTES and CCL27, and growth factors such as HGF, SCF and GM-CSF, increased during ART, reaching values significantly higher than HD after 48 weeks. Moreover, the G-CSF and MIP-1β soluble mediators were persistently altered and showed an inverse correlation with the CD4/CD8 T cell ratio. The increase of chemokines with antiviral activity and of growth factors with hematopoietic and immunomodulatory properties may have beneficial effects. Other studies are mandatory to evaluate the effects of long lasting levels of these factors to clarify their possible role in the context of protection/pathogenesis.

Myeloid-derived suppressor cells (MDSC) are expanded during HIV-1 infection and correlated with disease progression. MDSC expand in the early phase of primary infection depending on TRAIL level. In this study we evaluated the effect of ART on the frequency of MDSC in patients with primary HIV infection (PHI), and their impact on CD4 T cell reconstitution. MDSC frequency was evaluated by flow-cytometry in 60 PHI patients at 12, 24 and 48 weeks after ART initiation. Cytokine plasma levels were evaluated by Luminex technology at the same time points. The capacity of MDSC to modulate hematopoietic early progenitor cells' expansion was evaluated using the OP9/Dl1 in vitro system. As previously described, polymorphonuclear-MDSC (PMN-MDSC) frequency was higher in PHI compared to healthy donors. Interestingly, 48 weeks of successful ART failed to normalize the PMN-MDSC frequency. Moreover, PMN-MDSC frequency was not correlated with residual viral load, suggesting that the persistence of PMN-MDSC was not due to residual viral replication. Interestingly, patients with low PMN-MDSC frequency (6%). We also found an inverse correlation between PMN-MDSC frequency and CD4-T cell count at 48 weeks post-ART, which was confirmed by multivariate analysis adjusting for age and CD4 T cell number at baseline. These data suggest that the persistence of PMN-MDSC may impact CD4 T cell recovery. Indeed, in vitro PMN-MDSC impaired the expansion of CD34+CD38- hematopoietic early progenitors. Further, a balance between TRAIL and GM-CSF may be necessary to maintain a low MDSC level. In conclusion, early ART initiation was not able to normalize PMN-MDSC frequency that might impact the CD4 T cell recovery. These data open new questions regarding the clinical impact of MDSC persistence in HIV+ patients, in particular on non-AIDS related diseases.

Primary and chronic human immunodeficiency virus (HIV) infection alters γδ T-cell features. However, there is no evidence about early combined antiretroviral therapy (cART) and γδ T-cell dynamics. In the present study, HIV-positive individuals were divided into those with early primary infection (EPI) and those with late primary infection (LPI). The analysis of γδ T cells was performed by flow cytometry before and after therapy. Polyfunctional profile was assessed after in vitro peripheral blood mononuclear cell (PBMC) exposure to specific antigens. The results show that primary infection induced an expansion of Vδ1 T cells in LPI. Before treatment, a massive activation of γδ T-cell subsets was observed in both groups of patients, that correlated with disease progression and was significantly reduced after cART introduction. Despite this, CD107A-expressing Vδ1 T cells in both groups were significantly fewer than in healthy donors, but were restored by therapy introduction. Polyfunctional analysis of Vδ1 T cells from HIV-positive individuals revealed a lower frequency of CD107A+CCL-4+ Vδ1 T-cell subsets than healthy donors that persists after therapy. Functional profile of Vδ2 was similar to that in healthy donors before therapy but, at 6months, a lower frequency of CD107A, interferon-γ- or tumor necrosis factor-α-producing Vδ2 T cells was observed in the EPI group. Finally, individuals with LPI showed a lower frequency of quadruple-functional Vδ2 T-cell subset. In conclusion, during primary HIV infection, the baseline Vδ1 T-cell activation is correlated with immune reconstitution potential. Moreover, an altered γδ polyfunctional profile occurred, persisting after cART. Further studies are needed to understand whether a longer treatment of primary infection may increase γδ T-cell functionality.

New methods of HIV-1 RNA quantification based on dual-target detection are increasingly used in HIV viral load monitoring, but clinical implications and impact of dual-target detection on HIV-1 infection management are not established. Aptima HIV-1 Quant Dx assay is a last generation HIV viral load method, that uses pol and LTR as simultaneous target, providing quantitative results based mainly on pol target, while LTR target is used to report the results when pol signal is absent. In our laboratory, about 6% of results of all HIV-1 viral load tests performed with this platform in one year period resulted from LTR signal. Interestingly, LTR-based viremia (sometimes exceeding 1,000 copies/mL) was observed in a small proportion (up to 1%) of patients under ART, considered for long time virologically suppressed on the basis of a single target (pol-based) assay. Male gender, >700 vs 200 copies/mL, longitudinal assessments showed parallel kinetics between plasma viremia and proviral DNA. Sequencing of pol region for drug resistance assessment in patients with LTR-based viremia failed on plasma HIV-1 RNA, while it was successful on proviral DNA. The detection/quantification of HIV-1 viremia based only on LTR signal with a dual target assay in samples resulting undetectable with the more conventional target pol needs accurate evaluation; unravelling the biological basis of this phenomenon, here described for the first time, is mandatory to establish relevance and implication by both pathogenetic (i.e. infectivity of LTR-detected viruses, reservoir turnover, immune activation, etc.) and clinical standpoint.

The impact of baseline HIV-1 reverse transcriptase (RT) polymorphisms on response to first-line modern HAART containing tenofovir disoproxil fumarate (TDF) and emtricitabine (FTC) was evaluated. The impact of each RT polymorphism on virological success (VS) was evaluated in 604 HIV-1 subtype B-infected patients starting TDF+FTC-containing HAART. TDF and FTC antiviral activity was also tested in PBMCs infected by mutagenised HIV. Structural analysis based on docking simulations was performed. A98S was the only mutation significantly correlated with an increased proportion of patients achieving VS at 24 weeks (94.0% vs. 84.3%; P=0.03). Multivariate regression and Cox model analyses confirmed this result. At concentrations close to the minimal concentration achieved in patient plasma, TDF and FTC exhibited higher potency in the presence of A98S-mutated virus compared with wild-type (IC90,TDF, 8.6±1.1 vs. 19.3±3.5nM; and IC90,FTC, 12.4±7.7 vs. 16.8±9.8nM, respectively). The efficacy of FTC, abrogated by M184V, was partially restored by A98S (IC90,FTC, 5169±5931nM for A98S+M184V vs. 18477±12478nM for M184V alone). Docking analysis showed the higher potency of TDF and FTC in the presence of A98S-mutated virus was mainly due to higher binding affinity between drugs and mutated RT compared with wild-type. In the presence of FTC, A98S also partially restored the RT binding affinity impaired by M184V alone. A98S polymorphism improves virological response to TDF+FTC-containing HAART. This may help clinicians in the choice of the optimal NRTI backbone aimed at achieving maximal virological inhibition.

Objectives We evaluated the virological response in patients starting a regimen based on darunavir/ritonavir (DRV/r), which is currently the most widely used ritonavir-boosted protease inhibitor. Methods Data from 206 drug-naive and 327 PI-experienced patients starting DRV/r 600/100 mg twice daily (DRV600) or 800/100 mg once daily (DRV800) were examined. The probabilities of virological success (VS) and virological rebound (VR) were evaluated in survival analyses. Baseline DRV/r resistance and its evolution at failure were also examined. Results DRV600 was preferentially administered in patients with complex requirements (older age, higher viraemia, lower CD4 cell count and DRV/PI resistance) compared with DRV800. By 12 months, the probability of achieving VS was 93.2% and 84.3% in drug-naive and PI-experienced patients, respectively. The higher the baseline viraemia, the longer was the time required to achieve VS, both in drug-naive patients [>500 000 HIV-1 RNA copies/mL: median [interquartile range (IQR)] 6.1 (5.1–10.3) months; 100 000–500 000 copies/mL: median (IQR) 4.9 (3.8–6.1) months

Background: Low vitamin D levels are associated with higher odds of cognitive dysfunction in the older population, and in subjects with mental disorders or with chronic neurologic diseases. With combination antiretroviral therapy (cART), incidence of HIV-associated dementia has reduced, while the prevalence of milder forms of neurocognitive impairment (NCI) persisted stable over time. Hypovitaminosis D is often found in HIV infection but its association with NCI has not been investigated yet. Aim was to explore this association in a clinic-based HIV-positive population. Methods: A retrospective, cross-sectional analysis of an existing monocentre dataset obtained from patients undergoing neuropsychological assessment in routine clinical care between January, 2011 and December, 2016 was carried out. NCI was assessed through a standardized battery of 13 tests on 5 different cognitive domains and HIV-associated neurocognitive deficit (HAND) was classified according to Frascati's criteria. Vitamin D deficiency was defined by 25 hydroxy-vitamin D 25(OH)D levels

ObjectiveWe evaluated the characteristics of HIV-1 molecular transmission clusters (MTCs) in 1890 newly diagnosed individuals infected with non-B subtypes between 2005 and 2017 in Italy.MethodsPhylogenetic analyses were performed on pol sequences to characterise subtypes/circulating recombinant forms and identify MTCs. MTCs were divided into small (SMTCs, 2–3 sequences), medium (MMTCs, 4–9 sequences) and large (LMTCs, ≥10 sequences). Factors associated with MTCs were evaluated using logistic regression analysis.Results145 MTCs were identified and involved 666 individuals (35.2%); 319 of them (16.9%) were included in 13 LMTCs, 111 (5.9%) in 20 MMTCs and 236 (12.5%) in 112 SMTCs. Compared with individuals out of MTCs, individuals involved in MTCs were prevalently Italian (72.7% vs 30.9%, p9/L: 0.4 (0.265–0.587) vs 0.246 (0.082–0.417), p10 copies/mL: 4.8 (4.2–5.5) vs 5.0 (4.3–5.5), p=0.87). Logistic regression confirmed that certain factors such as being MSM, of Italian origin, younger age and higher CD4 cell count were significantly associated with MTCs.ConclusionsOur findings show that HIV-1 newly diagnosed individuals infected with non-B subtypes are involved in several MTCs in Italy. These MTCs include mainly Italians and MSM and highlight the complex phenomenon characterising the HIV-1 spread. This is important especially in view of monitoring the HIV epidemic and guiding the public health response.

OBJECTIVE Antiretrovirals with long half-lives, such as tenofovir disoproxil fumarate/emtricitabine (TDF/FTC) and efavirenz (EFV), are suitable for reduced frequency dosing, with potential for improved adherence and reduced toxicity and costs. The objective of this study was to investigate the noninferiority of the TDF/FTC/EFV fixed-dose combination on alternate-days versus standard regimen in virologically suppressed patients. DESIGN A randomized-controlled open-label noninferiority trial enrolling HIV-1-infected patients treated for at least 6 months with TDF/FTC/EFV fixed-dose combination, virologically suppressed (

No consensus has been reached on how to define cerebrospinal fluid HIV-1 escape (CSF-E). We describe its prevalence in 1095 paired CSF-plasma HIV-RNA measurements from antiretroviral-treated patients according to several definitions and neurological affections. CSF-E prevalence varied substantially (9.0-38.9%) and was higher in patients with cerebrovascular disorders, HIV-associated dementia and white matter abnormalities. Considering the variability in HIV-RNA quantification assays, the biological relevance of viral escape at different thresholds needs to be accurately assessed.

The possibility of performing genotypic tropism testing (GTT) with proviral DNA (pvDNA) even during suppressed viremia would facilitate the use of CCR5 inhibitors as part of switching, simplification, or intensification strategies. Thus, we aimed to evaluate the tropism concordance between plasma RNA and pvDNA samples and to assess which factors could affect possible discrepancies between the two compartments. GTT was performed using both plasma RNA and pvDNA from 55 sample pairs from drug-experienced patients. Potential differences between the two compartments were evaluated by analyzing coreceptor usage and genetic variability. Paired samples were also stratified in three levels of viremia (500 copies/ml). Overall, Geno2Pheno comparisons of false-positive rates in the two compartments showed good correlation ( r = 0.72). A high level of concordance in tropism predictions for the two compartments was found (46/55 sample pairs [83.6%]). Among the 9 sample pairs with discordant tropisms, a larger proportion of pvDNA samples harboring CXCR4/dual-mixed-tropic viruses was found, in comparison with plasma RNA samples (88.9% versus 11.1%; P = 0.0034). Discordant samples were characterized by greater genetic variability than were concordant samples. With stratification of the paired samples according to viremia levels, the prevalence of discordant samples decreased with increasing viremia (500 copies/ml, 6.7%; P = 0.2). Our findings confirm that prediction of viral tropism using pvDNA is feasible even in low-level viremia and provides useful information for therapy optimization for patients with low or suppressed viremia.

Herpes simplex virus encephalitis (HSE) is a rare but often fatal disease if left untreated. MRI typically shows the characteristic findings of medial temporal lobe and insular involvement, while diagnosis in confirmed by CSF PCR. In immunocompromised state, HSE may have atypical clinical and radiological features. We report a MS patient under natalizumab treatment with HSE, who presented with MRI lesions exclusively in the right parietal lobe. The patient was timely started on acyclovir resulting in marked improvement. A high index of suspicion for HSE should be maintained when a patient presents with fever and extratemporal lesions, even more in immunocompromised subjects.

Background Genotypic resistance test (GRT) performed in peripheral blood mononuclear cells (PBMC) represents a chance to evaluate resistance in virologically suppressed HIV infected patients. Objectives To evaluate the impact of baseline resistance detected through PBMC GRT on virological rebound after switching treatment. Study design Baseline genotypic susceptibility scores (GSS) from PBMC GRT (DNA-GSS) and from previous cumulative plasma GRTs (when available, pRNA-GSS) were evaluated. Survival analysis was used to assess the probability and predictors of virological rebound (VR). Results 227 virologically suppressed patients were analysed. Twenty-four months after switching therapy, the probability of VR was 15.3%. Patients showing an intermediate or full resistant DNA-GSS had a higher probability of experiencing VR compared to those carrying a fully susceptible DNA-GSS (27.2% vs. 13.7%, p = 0.001). By multivariable Cox regression, patients with an intermediate/full resistant DNA-GSS, with a nadir CD4 count In a sub-group of 114 patients with previous plasma GRTs available, patients with an intermediate or fully resistance showed by both GSSs (from plasma and PBMCs) had the highest probability of experiencing VR. Conclusions Resistance detected in proviral DNA, together with a low nadir CD4 count and a short previous virological control, predicts VR after therapy switching in virologically suppressed patients. PBMC GRT can be a useful tool for tailoring treatment switch, especially if paired with information about previous cumulative resistance and previous viro-immunological history.

Background Virological success (VS) and immunological reconstitution (IR) of antiretroviral-naive HIV-1-infected patients with pre-therapy viral load (VL) >500,000 copies/ml was assessed after 12 months of treatment according to initial drug-class regimens. Methods An observational multicentre retrospective study was performed. VS was defined as the first VL + T-lymphocytes from treatment start. Survival analysis was used to estimate the probability and predictors of VS and IR by 12 months of therapy. Results 428 HIV-1-infected patients were analysed. Patients were grouped according to the different first-line drug-classes used: a non-nucleoside reverse transcriptase inhibitor (NNRTI) plus two nucleoside reverse transcriptase inhibitors (NRTIs; NNRTI-group; n=105 [24.5%]); a protease inhibitor (PI) plus two NRTIs (PI-group; n=260 [60.8%]); a four-drug regimen containing a PI-regimen plus an integrase inhibitor (PI+INI-group; n=63 [14.7%]). Patients in the PI-group showed the lowest probability of VS (PI-group: 72.4%; NNRTI-group: 75.5%; PI+INI-group: 81.0%; PConclusions In this multicentre retrospective study, patients with viraemia >500,000 copies/ml who start a first-line regimen containing PI+INI or NNRTI yield a better VS compared to those receiving a PI-based regimen.

Introduction: It would be important to identify subjects at risk to develop tuberculosis (TB) disease. RD1-based Interferon-γ Release Assays (IGRAs) cannot distinguish latent from active TB disease. Conversely, a positive response to HBHA-based IGRAs, among TB-infected subjects, correlates with Mtb containment and low risk of TB progression. Aims: to characterize HBHA-immune responses in HIV-infected and uninfected subjects with active TB or latent TB infection (LTBI). Methods: 49 subjects were prospectively enrolled, 22 HIV-uninfected (13 TB, 9 LTBI) and 27 HIV-infected (12 HIV-TB, 15 HIV-LTBI). Whole blood and PBMC were stimulated with HBHA and RD1 antigens. IFNγ release was evaluated by ELISA whereas cytokine profile (IFNγ, TNFα, IL2) and phenotype (CD45RA, CD27) by FACS. Results: Among LTBI individuals, HBHA stimulation induced IFNγ release in all the HIV-uninfected, while only 4/15 HIV-infected responded. Within the active TB, only 6/13 HIV-uninfected and 1/12 HIV-TB patients responded. Differently than what observed for RD1, the cytokine profile of HBHA-specific T cells evaluated by FACS showed that the CD4 T cells were mostly monofunctional. Conversely, CD8-specific T cells were mostly monofunctional for both HBHA and RD1 stimulations. The phenotype of HBHA-specific T cells showed a predominantly central memory (CM) and effector memory (EM) phenotype in CD4 T cells without differences among the groups. Differently, HBHA-specific CD8 T cells, showed mainly a CM and naive phenotype in LTBI group while TB, HIV-LTBI and HIV-TB groups were characterized by EM or TEMRA phenotypes. Conclusions: These results may help to better define immunological correlates of protection from TB disease.

Background: There is limited information on pregnancy loss in women with HIV, and it is still debate whether HIV-related markers may play a role. Objectives: To explore potential risk factors for pregnancy loss in women with HIV, with particular reference to modifiable risk factors and markers of HIV disease. Methods: Multicenter observational study of HIV-positive pregnant women. The main outcome measure was pregnancy loss, including both miscarriage (

Background. We evaluated reliabilityand clinical usefulness of genotypic resistance testing (GRT) in patients for whom combination antiretroviral therapy (cART) was unsuccessful with viremia levels 50–1000 copies/mL, for whom GRT is generally not recommended by current guidelines. Methods. The genotyping success rate was evaluated in 12 828 human immunodeficiency virus type 1 (HIV-1) plasma samples with viremia >50 copies/mL, tested using the commercial ViroSeq HIV-1 Genotyping System or a homemade system. Phylogenetic analysis was performed to test the reliability and reproducibility of the GRT at lowlevel viremia (LLV). Drug resistance was evaluated in 3895 samples from 2200 patients for whom treatment was unsuccessful (viremia >50 copies/mL) by considering the resistance mutations paneled in the 2013 International Antiviral Society list. Results. Overall, the success rate of amplification/sequencing was 96.4%. Viremia levels of 50–200 and 201–500 copies/mL afforded success rates of 67.2% and 88.1%, respectively, reaching 93.2% at 501–1000 copies/mL and ≥97.3% above 1000 copies/mL. A high homology among sequences belonging to the same subject for 96.4% of patients analyzed was found. The overall resistance prevalence was 74%. Drug resistance was commonly found also at LLV. In particular, by stratifying for different viremia ranges, detection of resistance was as follows: 50–200 copies/ mL = 52.8%; 201–500 = 70%; 501–1000 = 74%; 1001–10 000 = 86.1%; 10 001–100 000 = 76.7%; and >100 000 = 63% (P< .001). Similar bell-shaped results were found when the GRT analysis was restricted to 2008–2012, although at a slightly lower prevalence. Conclusions. In patients failing cART with LLV, HIV-1 genotyping provides reliable and reproducible results that are informative about emerging drug resistance.