Analysis of transmitted HIV-1 drug resistance using 454 ultra-deep-sequencing and the DeepChek(®)-HIV system.

Introduction Next-generation sequencing (NGS) is capable of detecting resistance-associated mutations (RAMs) present at frequencies of 1% or below. Several studies have found that baseline low-frequency RAMs are associated with failure to first-line HAART. One major limitation to the expansion of this technology in routine diagnostics is the complexity and laboriousness integral to bioinformatics analysis. DeepChek (ABL, TherapyEdge) is a CE-marked software that allows automated analysis and resistance interpretation of NGS data. Objective To evaluate the use of 454 ultra-deep-sequencing (Roche® 454, Life Sciences; 454-UDS) and DeepChek for routine baseline resistance testing in a clinical diagnostic laboratory. Methods 107 newly diagnosed HIV-1-infected patients (subtypes: A, n=9; B, n=52; C, n=21; D, n=2; F, n=3; G, n=1; CRF01, n=7; CRF02, n=7; CRF06, n=1; CRF07, n=1; CRF10, n=1 and unassigned complex, n=2) with a median plasma viral load of 88,727 copies/mL (range: 1380–2,143,543) were tested by 454-UDS and Sanger sequencing for the detection of protease and reverse transcriptase RAMs. In addition, integrase RAMs were investigated in 57 of them. Sequence analysis and resistance interpretation were performed using DeepChek applying 1% and 20% thresholds for variant detections; filters applied were comparison between Sanger and 454-UDS, and Stanford and IAS list for resistance interpretation. Results The time elapsed from generation of raw 454 data (between 2,000–5,000 sequences/sample) to elaboration of a resistance report was approximately 10 minutes per sample, equivalent to the time required for the same process using Sanger sequencing. Four patients (3.7%) showed baseline resistance by Sanger and 454-UDS at frequencies above 20%, which affected both NRTIs (n=2) and NNRTIs (n=2). In addition, 12 patients (11.2%) showed transmitted drug resistance (TDR) by 454-UDS at frequencies below 20% affecting NRTIs (n=9), NNRTIs (n=7) and PIs (n=2). Integrase resistance was not detected at baseline by 454-UDS or Sanger sequencing. Conclusions DeepChek allowed easy and rapid analysis and interpretation of NGS data, thus facilitating the incorporation of this technology in routine diagnostics. The use of NGS considerably increased the detection rates of TDR to NRTI, NNRTIs and PIs. No transmitted resistance to integrase inhibitors was found in our population by Sanger sequencing or UDS

HIV and Other Sexually Transmitted Infections among Men Who Have Sex with Men Recruited by RDS in Buenos Aires, Argentina: High HIV and HPV Infection

The aim of this study was to estimate the prevalence of HIV and other STIs, among MSM from Buenos Aires (2007-2009).Responding Driven Sampling was used for recruitment of MSM. Participants completed a structured web-based survey and provided biological samples.A total of 496 MSM were studied for HIV, HBV, HCV, and T. pallidum infections. Chlamydia and HPV diagnoses were only performed in 98 and 109 participants, respectively. Prevalence of HIV was 17.3%, HBV 22.9%, HCV 7.5%, T. pallidum 20.5%, HPV 83.5%, and C. trachomatis 1.7%. In the year prior to the evaluation, 71% of the participants had had sex with men and/or trans and women (MMW) while 29% had not had sex with women (MM). Comparing MM to MMW, prevalence of HIV (30.7% vs. 11.9%, p<0.001), HBV (36.4% vs. 17.8%, p<0.001), T. pallidum (32.1% vs. 15.7%, p<0.001), and HPV (88.3% vs. 70.4%, p = 0.039) were significantly higher among MM, whereas no significant differences were found for HCV and C. trachomatis. The MM group had also significantly higher HIV incidence (5.60 vs. 4.28 per 100 persons-year, p = 0.032). HPV genotypes 16, 6, and 11 were the most frequently found; 40.7% of the MSM had more than one genotype and one high risk genotype was detected in 43.6% of participants.Both MM and MMW are at high risk of infection for HIV and other STIs. Rates of HIV, HBV, T. pallidum and HPV infections are higher in the MM group

Effect of angiotensin-converting enzyme inhibitor and angiotensin receptor blocker initiation on organ support-free days in patients hospitalized with COVID-19

IMPORTANCE Overactivation of the renin-angiotensin system (RAS) may contribute to poor clinical outcomes in patients with COVID-19. Objective To determine whether angiotensin-converting enzyme (ACE) inhibitor or angiotensin receptor blocker (ARB) initiation improves outcomes in patients hospitalized for COVID-19. DESIGN, SETTING, AND PARTICIPANTS In an ongoing, adaptive platform randomized clinical trial, 721 critically ill and 58 non–critically ill hospitalized adults were randomized to receive an RAS inhibitor or control between March 16, 2021, and February 25, 2022, at 69 sites in 7 countries (final follow-up on June 1, 2022). INTERVENTIONS Patients were randomized to receive open-label initiation of an ACE inhibitor (n = 257), ARB (n = 248), ARB in combination with DMX-200 (a chemokine receptor-2 inhibitor; n = 10), or no RAS inhibitor (control; n = 264) for up to 10 days. MAIN OUTCOMES AND MEASURES The primary outcome was organ support–free days, a composite of hospital survival and days alive without cardiovascular or respiratory organ support through 21 days. The primary analysis was a bayesian cumulative logistic model. Odds ratios (ORs) greater than 1 represent improved outcomes. RESULTS On February 25, 2022, enrollment was discontinued due to safety concerns. Among 679 critically ill patients with available primary outcome data, the median age was 56 years and 239 participants (35.2%) were women. Median (IQR) organ support–free days among critically ill patients was 10 (–1 to 16) in the ACE inhibitor group (n = 231), 8 (–1 to 17) in the ARB group (n = 217), and 12 (0 to 17) in the control group (n = 231) (median adjusted odds ratios of 0.77 [95% bayesian credible interval, 0.58-1.06] for improvement for ACE inhibitor and 0.76 [95% credible interval, 0.56-1.05] for ARB compared with control). The posterior probabilities that ACE inhibitors and ARBs worsened organ support–free days compared with control were 94.9% and 95.4%, respectively. Hospital survival occurred in 166 of 231 critically ill participants (71.9%) in the ACE inhibitor group, 152 of 217 (70.0%) in the ARB group, and 182 of 231 (78.8%) in the control group (posterior probabilities that ACE inhibitor and ARB worsened hospital survival compared with control were 95.3% and 98.1%, respectively). CONCLUSIONS AND RELEVANCE In this trial, among critically ill adults with COVID-19, initiation of an ACE inhibitor or ARB did not improve, and likely worsened, clinical outcomes. TRIAL REGISTRATION ClinicalTrials.gov Identifier: NCT0273570

Characterization of avian Chlamydia psittaci strains using omp1 restriction mapping and serovar-specific monoclonal antibodies

In the present study, 60 avian Chlamydia psittaci isolates were characterized using restriction fragment length polymorphism as well as serovar-specific monoclonal antibodies, enabling a comparison between the two characterization methods. Sixty avian C. psittaci isolates were characterized by AluI restriction mapping of the major outer membrane protein gene omp1 obtained after amplification by the polymerase chain reaction. The 60 avian C. psittaci strains were also characterized using serovar-specific monoclonal antibodies in a microimmunofluorescence test. Digestion of 60 avian C. psittaci omp1 amplicons by AluI generated 5 of the 6 known distinct restriction patterns (A, B, D, E and F). Restriction pattern C was not observed. Serotyping revealed 4 avian C. psittaci serovars (A, B, C and D). None of the 60 isolates was typed as serovar E. AluI restriction patterns A, B, D and E corresponded in 98% of the cases to serovars A, B, C and D, respectively. One isolate, classified as serovar A, generated restriction pattern F instead of A. Genotyping enabled a more precise differentiation of avian C. psittaci serovar A strains. Serovar A strains were divided into two groups according to their AluI restriction pattern (A or F). For epidemiological studies, genotyping can thus be a highly valuable alternative to serotyping, especially when applied directly to the clinical samples