Prevalence and clinical impact of minority resistant variants in patients failing an integrase inhibitor-based regimen by ultra-deep sequencing.

Background: Integrase strand transfer inhibitors (INSTIs) are recommended by international guidelines as first-line therapy in antiretroviral-naive and -experienced HIV-1-infected patients. Objectives: This study aimed at evaluating the prevalence at failure of INSTI-resistant variants and the impact of baseline minority resistant variants (MiRVs) on the virological response to an INSTI-based regimen. Methods: Samples at failure of 134 patients failing a raltegravir-containing (n = 65), an elvitegravir-containing (n = 20) or a dolutegravir-containing (n = 49) regimen were sequenced by Sanger sequencing and ultra-deep sequencing (UDS). Baseline samples of patients with virological failure (VF) (n = 34) and of those with virological success (VS) (n = 31) under INSTI treatment were sequenced by UDS. Data were analysed using the SmartGene platform, and resistance was interpreted according to the ANRS algorithm version 27. Results: At failure, the prevalence of at least one INSTI-resistant variant was 39.6% by Sanger sequencing and 57.5% by UDS, changing the interpretation of resistance in 17/134 (13%) patients. Among 53 patients harbouring at least one resistance mutation detected by both techniques, the most dominant INSTI resistance mutations were N155H (45%), Q148H/K/R (23%), T97A (19%) and Y143C (11%). There was no difference in prevalence of baseline MiRVs between patients with VF and those with VS. MiRVs found at baseline in patients with VF were not detected at failure either in majority or minority mutations. Conclusions: UDS is more sensitive than Sanger sequencing at detecting INSTI MiRVs at treatment failure. The presence of MiRVs at failure could be important to the decision to switch to other INSTIs. However, there was no association between the presence of baseline MiRVs and the response to INSTI-based therapies in our study.

Emerging resistance mutations in PI-naive patients failing an atazanavir-based regimen (ANRS multicentre observational study).

Background: Atazanavir is a PI widely used as a third agent in combination ART. We aimed to determine the prevalence and the patterns of resistance in PI-naive patients failing on an atazanavir-based regimen. Methods: We analysed patients failing on an atazanavir-containing regimen used as a first line of PI therapy. We compared the sequences of reverse transcriptase and protease before the introduction of atazanavir and at failure [two consecutive viral loads (VLs) >50 copies/mL]. Resistance was defined according to the 2014 Agence Nationale de Recherche sur le SIDA et les Hépatites Virales (ANRS) algorithm. Results: Among the 113 patients, atazanavir was used in the first regimen in 71 (62.8%) patients and in the first line of a PI-based regimen in 42 (37.2%). Atazanavir was boosted with ritonavir in 95 (84.1%) patients and combined with tenofovir/emtricitabine or lamivudine (n = 81) and abacavir/lamivudine or emtricitabine (n = 22). At failure, median VL was 3.05 log10 copies/mL and the median CD4+ T cell count was 436 cells/mm3. The median time on atazanavir was 21.2 months. At failure, viruses were considered resistant to atazanavir in four patients (3.5%) with the selection of the following major atazanavir-associated mutations: I50L (n = 1), I84V (n = 2) and N88S (n = 1). Other emergent PI mutations were L10V, G16E, K20I/R, L33F, M36I/L, M46I/L, G48V, F53L, I54L, D60E, I62V, A71T/V, V82I/T, L90M and I93L/M. Emergent NRTI substitutions were detected in 21 patients: M41L (n = 2), D67N (n = 3), K70R (n = 1), L74I/V (n = 3), M184V/I (n = 16), L210W (n = 1), T215Y/F (n = 3) and K219Q/E (n = 2). Conclusions: Resistance to atazanavir is rare in patients failing the first line of an atazanavir-based regimen according to the ANRS. Emergent NRTI resistance-associated mutations were reported in 18% of patients.

Disparities in HIV-1 transmitted drug resistance detected by ultradeep sequencing between men who have sex with men and heterosexual populations.

OBJECTIVES: Transmitted drug resistance (TDR) can impair the response to first-line antiretroviral therapy. In treatment-naïve patients chronically infected with HIV type 1 (HIV-1), it was previously shown through Sanger sequencing that TDR was more common in men who have sex with men (MSM) than in other transmission risk groups. We aimed to compare two HIV-1 transmission groups in terms of the presence of TDR mutations. METHODS: We investigated, through Sanger sequencing and ultradeep sequencing (UDS), the presence of resistance mutations, both in majority (> 20%) and in minority (1-20%) proportions, in 70 treatment-naïve MSM and 70 treatment-naïve heterosexual patients who recently screened positive for HIV-1. RESULTS: The global prevalence of TDR was not significantly different between the two groups, either by Sanger or by UDS. Nevertheless, a higher frequency of nucleoside reverse transcriptase inhibitor TDR was observed among heterosexual patients (P = 0.04). There was also a trend for a higher frequency of TDR among MSM infected with HIV-1 subtype B compared with MSM infected with HIV-1 non-B subtypes (P = 0.06). CONCLUSIONS: Ultradeep sequencing UDS allowed sensitive monitoring of TDR, and highlighted some disparities between transmission groups.

Prevalence of HIV-1 drug resistance in treated patients with viral load >50 copies/mL: a 2014 French nationwide study.

Background: Surveillance of HIV-1 resistance in treated patients with a detectable viral load (VL) is important to monitor, in order to assess the risk of spread of resistant viruses and to determine the proportion of patients who need new antiretroviral drugs with minimal cross-resistance. Methods: The HIV-1 protease and reverse transcriptase (RT) and integrase genes were sequenced in plasma samples from 782 consecutive patients on failing antiretroviral regimens, seen in 37 specialized centres in 2014. The genotyping results were interpreted using the ANRS v24 algorithm. Prevalence rates were compared with those obtained during a similar survey conducted in 2009. Results: The protease and RT sequences were obtained in 566 patients, and the integrase sequence in 382 patients. Sequencing was successful in 60%, 78%, 78% and 87% of patients with VLs of 51-200, 201-500, 501-1000 and >1000 copies/mL, respectively. Resistance to at least one antiretroviral drug was detected in 56.3% of samples. Respectively, 3.9%, 8.7%, 1.5% and 3.4% of patients harboured viruses that were resistant to any NRTI, NNRTI, PI and integrase inhibitor (INI). Resistance rates were lower in 2014 than in 2009. Resistance was detected in 48.5% of samples from patients with a VL between 51 and 200 copies/mL. Conclusion: In France in 2014, 90.0% of patients in AIDS care centres were receiving antiretroviral drugs and 12.0% of them had VLs >50 copies/mL. Therefore, this study suggests that 6.7% of treated patients in France might transmit resistant strains. Resistance testing may be warranted in all treated patients with VL > 50 copies/mL.

Usefulness of an HIV DNA resistance genotypic test in patients who are candidates for a switch to the rilpivirine/emtricitabine/tenofovir disoproxil fumarate combination.

OBJECTIVES: In the context of a rilpivirine/emtricitabine/tenofovir disoproxil fumarate switch in HIV-1-infected patients with at least 1 year of virological success, we determined whether proviral DNA is an alternative to plasma HIV RNA for resistance genotyping. METHODS: Resistance-associated mutations (RAMs) in DNA after at least 1 year of virological success [viral load (VL) <50 copies/mL] were compared with those identified in the last plasma RNA genotype available. Rilpivirine/emtricitabine/tenofovir disoproxil fumarate RAMs studied were K65R, L100I, K101E/P, E138A/G/K/R/Q, V179L, Y181C/I/V, M184V/I, Y188L, H221Y, F227C and M230I/L in the RT. We studied patients without virological failure (VF) and with at least 1 VF (two consecutive VLs >50 copies/mL). Kappa's coefficient was used to measure agreement between the DNA and RNA genotypes. RESULTS: In patients without VF (n = 130) and with VF (n = 114), RNA and DNA showed resistance to at least one drug of the rilpivirine/emtricitabine/tenofovir disoproxil fumarate combination in 8% and 9% and in 60% and 45%, respectively. For rilpivirine RAMs, correlation between RNA and DNA was higher in patients without VF than in patients with VF (kappa = 0.60 versus 0.19, P = 0.026). Overall, the prevalence of RAMs was lower in DNA than in RNA. CONCLUSIONS: Incomplete information provided by the DNA genotypic test is more notable in patients with VF, suggesting that all resistance mutations associated with prior VF have not been archived in the proviral DNA or decreased to a level below the threshold of detection. In the case where no historical plasma genotypic test is available, DNA testing might be useful to rule out switching to rilpivirine/emtricitabine/tenofovir disoproxil fumarate.

Assessment of coronary arterial stents by multislice-CT angiography.

PURPOSE: To assess patency and lumen visibility of coronary artery stents by multislice-CT angiography (MSCTA) in comparison with conventional coronary angiography as the standard of reference. MATERIAL AND METHODS: 47 stents of 13 different types were evaluated in 29 patients. MSCTA was performed on a 4-slice scanner with a standard coronary protocol (detector collimation 4 x 1 mm; table feed 1.5 mm/rotation, 400 mAs, 120 kV). Image evaluation was performed by two readers who were blinded to the reports from the catheter angiography. MIP reconstructions were evaluated for image quality on a 4-point scale (1 = poor, 4 = excellent) and stent patency (contrast distal to the stent as an indirect patency sign). Axial images and multiplanar reformations through the stents were used for assessment of stent lumen visibility (measurement of the visible stent lumen diameter) and detection of relevant in-stent stenosis (> or =50%). RESULTS: Image quality was fair to good on average (score 2.64 +/- 1.0) and depended on the heart rate (heart rate 45-60: average score 3.2, heart rate 61-70: average score 2.8, heart rate >71: average score 1.4). Thirty-seven stents were correctly classified as patent, 1 was correctly classified as occluded and 9 stents were not assessible due to insufficient image quality because of triggering artifacts. Parts of the stent lumen could be visualized in 30 cases. On average, 20-40% of the stent lumen diameter was visible. Twenty-five stents were correctly classified as having no stenosis, 1 was falsely classified as stenosed, 1 was correctly classified as occluded. In 20 stents lumen visibility was not sufficient for stenosis evaluation. CONCLUSION: Although the stent lumen may be partly visualized in most stents, a reliable evaluation of in-stent stenoses does not seem practical by 4-slice MSCT. Nevertheless, for stent patency evaluation, MS-CTA might provide valuable clinical information. With submillimeter MSCT (e.g., 16-slice scanners) and more sophisticated reconstruction algorithms, further improvements may be expected.

[Aneurysm of the subclavian artery after operatively corrected coarctation of the aorta: 20 years of follow-up and review of the current literature]. / Aneurysma der Arteria subclavia nach Korrektur einer Aortenisthmusstenose. 20-jähriger Langzeitverlauf und Literaturübersicht.

We report of long-term follow-up of a combined fusiform aneurysm of the right subclavian artery extending to the thyreocervical trunk (3.2 x 2.8 x 2.2 cm (width x height x depth)) in a 33-year old patient. As a newborn, the clinical diagnosis of an aortic isthmus stenosis was made without need for intervention at this stage. Further development of the child remained unremarkable until the age of eleven years when he experienced dizziness after sporting activities. Due to clinically proven progress, cardiac catheterization was performed and confirmed the initial diagnosis of a juxtaductale stenosis of the aortic isthmus, which was operated thereafter with an end-to-end anastomosis. Furthermore, an aneurysm of the right subclavian artery was revealed. Since then, non-invasive routine follow-up showed no significant worsening of this aneurysm, which extends to the thyreocervical trunk. The patient has been event free and completely asymptomatic. This case report illustrates the more than twenty years of follow-up of an asymptomatic combined fusiform aneurysm of the subclavian artery and thyreocervical trunk and provides a review of the literature on this topic.

Contrast transcranial doppler ultrasound in the detection of right-to-left shunts : time window and threshold in microbubble numbers.

BACKGROUND AND PURPOSE: Cardiac right-to-left shunts can be identified by transesophageal echocardiography (TEE) and by transcranial Doppler ultrasound (TCD) with the use of contrast agents and a Valsalva maneuver (VM) as provocation procedure. Currently, data on the appropriate timing of the VM, the use of a diagnostic time window, and a threshold in contrast agent microbubbles detected are insufficient. METHODS: Fifty-eight patients were investigated by both TEE and bilateral TCD of the middle cerebral artery. The following protocol with injections of 10 mL of the commercial galactose-based contrast agent Echovist was applied in a randomized way: (1) no VM, (2) VM for 5 seconds starting 2 seconds after the beginning of contrast injection, (3) VM for 5 seconds starting 5 seconds after the beginning of contrast injection, (4) VM for 5 seconds starting 8 seconds after the beginning of contrast injection, and (5) repetitive short VMs in between 2 and 13 seconds after the beginning of contrast injection. In addition to the single tests, we also tested the sensitivity and specificity of combined results of the tests with VM. RESULTS: In 21 patients, a right-to-left shunt was demonstrated by TEE and contrast TCD (shunt positive). Twenty-one patients were negative in both investigations, no patient was positive on TEE and negative on TCD, and 16 patients were only positive on at least 1 TCD investigation but negative during TEE. Test 3 was the most appropriate test when combined with the results of 1 of the other tests with VM. The highest sensitivities were achieved with a diagnostic time window of 40 seconds and when the presence of a single microbubble was sufficient for the diagnosis of a shunt. CONCLUSIONS: TCD performed twice with 2 provocation maneuvers with Echovist is a sensitive method to identify TEE-proven cardiac right-to-left shunts. The VM should be performed for 5 seconds starting at 5 seconds after the beginning of contrast injection.

Anaphylactoid reactions during hemodialysis in sheep are ACE inhibitor dose-dependent and mediated by bradykinin.

Anaphylactoid reactions (AR) have been attributed to the generation of bradykinin (BK) when AN69 membranes are used together with angiotensin converting enzyme (ACE) inhibitors during hemodialysis. However, conclusive evidence for the involvement of the BK as the mediator of these AR is still lacking. This study examined the degree of contact activation in an animal model caused by three PAN membranes--AN69, PAN DX, and SPAN- and the effects of different doses of the ACE inhibitor enalapril (ENA) and the BK B2-receptor antagonist icatibant on AR during hemodialysis. Six sheep were dialyzed for one hour with or without ENA pre-treatment using the different membranes in random order. Severe AR were observed only during hemodialysis with AN69 dialyzers together with ENA pre-treatment; the severity of AR increased with the ENA dose. Mild hypotension was noted during hemodialysis with AN69 without ACE inhibition and with PAN DX and 20 mg ENA. Compared to pre-dialysis values, maximum generation of BK after blood passage through the dialyzer was found at five minutes: 73-fold (AN69 without ENA), 161-fold (AN69 with 10 mg ENA), 97-fold (AN69 with 20 mg ENA), 108-fold (AN69 with 30 mg ENA), 154-fold (AN69 with 30 mg ENA and 0.1 mg/kg icatibant), 18-fold (PAN DX without ENA), and 42-fold (PAN DX with 20 mg ENA). Elevated BK levels in arterial blood were detected during hemodialysis with AN69 membranes even without ACE inhibition (2.5-fold); pre-treatment with 20 mg ENA further increased arterial BK concentrations (4-fold). Furthermore, a marked decline of prekallikrein and high molecular weight kininogen concentrations was noted for both AN69 and PAN DX membranes. Anaphylactoid reactions during hemodialysis were completely prevented by icatibant even after pre-treatment with ENA and in the presence of high BK concentrations. Concentrations of prekallikrein, high molecular weight kininogen, and BK remained unchanged and no AR were observed during hemodialysis with SPAN and pre-treatment with 20 mg ENA. Our findings confirm that AR during hemodialysis with the negatively charged AN69 membrane are mediated by BK, since they can be prevented by the BK B2-receptor antagonist icatibant.