HIV-1 subtype is an independent predictor of reverse transcriptase mutation K65R in HIV-1 patients treated with combination antiretroviral therapy including tenofovir.

Subtype-dependent selection of HIV-1 reverse transcriptase resistance mutation K65R was previously observed in cell culture and small clinical investigations. We compared K65R prevalence across subtypes A, B, C, F, G, and CRF02_AG separately in a cohort of 3,076 patients on combination therapy including tenofovir. K65R selection was significantly higher in HIV-1 subtype C. This could not be explained by clinical and demographic factors in multivariate analysis, suggesting subtype sequence-specific K65R pathways.

Decreasing population selection rates of resistance mutation K65R over time in HIV-1 patients receiving combination therapy including tenofovir.

OBJECTIVES: The use of tenofovir is highly associated with the emergence of mutation K65R, which confers broad resistance to nucleoside/nucleotide analogue reverse transcriptase inhibitors (NRTIs), especially when tenofovir is combined with other NRTIs also selecting for K65R. Although recent HIV-1 treatment guidelines discouraging these combinations resulted in reduced K65R selection with tenofovir, updated information on the impact of currently recommended regimens on the population selection rate of K65R is presently lacking. METHODS: In this study, we evaluated changes over time in the selection rate of resistance mutation K65R in a large population of 2736 HIV-1-infected patients failing combination antiretroviral treatment between 2002 and 2010. RESULTS: The K65R resistance mutation was detected in 144 patients, a prevalence of 5.3%. A large majority of observed K65R cases were explained by the use of tenofovir, reflecting its wide use in clinical practice. However, changing patterns over time in NRTIs accompanying tenofovir resulted in a persistent decreasing probability of K65R selection by tenofovir-based therapy. The currently recommended NRTI combination tenofovir/emtricitabine was associated with a low probability of K65R emergence. For any given dual NRTI combination including tenofovir, higher selection rates of K65R were consistently observed with a non-nucleoside reverse transcriptase inhibitor than with a protease inhibitor as the third agent. DISCUSSION: Our finding of a stable time trend of K65R despite elevated use of tenofovir illustrates increased potency of current HIV-1 therapy including tenofovir.

An exploratory survey of professionals on the use of stored tissue samples from minors for genetic research.

he ethical aspects of the use of stored tissue samples collected from minors are of topical interest. However, the views of professionals working in the field of genetics have not been investigated in depth anywhere. We conducted a survey among 194 such professionals in Belgium. This list was composed of the members of the High Council for Anthropogenetics, supplemented with all professionals working in the field of genetics that we found on the websites of the eight Belgian centers of human genetics and of the associated university registries. We achieved a response rate of 35.5%. The vast majority (92%) think that research on stored tissue samples is useful. Most respondents stated that parental consent is valid (82.5%), and 76.5% thought that children should also be given the right to assent when they are able to comprehend the implications of the storage of biological samples and of genetic research. Slightly more than half put the age at which young people can understand storage or research rather high: 16-18 years (51 and 53.1%, respectively). Although there is some consensus in the literature that donors should be allowed to give broad consent for future research on their biological samples, only 47.6% in our survey thought that parents should be allowed to consent to any future research on their children's samples. The aim of our study was to give some basis for future ethical reflections and policies on the subject of stored tissue samples from minors for genetic research. We concluded that a large majority of Belgian researchers and clinicians in the field of genetic research think research on stored tissue samples from minors is useful. They also think that parental consent for such research is valid, but that children should be allowed to assent as they grow older.

Bayesian network analysis of resistance pathways against HIV-1 protease inhibitors.

Interpretation of Human Immunodeficiency Virus 1 (HIV-1) genotypic drug resistance is still a major challenge in the follow-up of antiviral therapy in infected patients. Because of the high degree of HIV-1 natural variation, complex interactions and stochastic behaviour of evolution, the role of resistance mutations is in many cases not well understood. Using Bayesian network learning of HIV-1 sequence data from diverse subtypes (A, B, C, F and G), we could determine the specific role of many resistance mutations against the protease inhibitors (PIs) nelfinavir (NFV), indinavir (IDV), and saquinavir (SQV). Such networks visualize relationships between treatment, selection of resistance mutations and presence of polymorphisms in a graphical way. The analysis identified 30N, 88S, and 90M for nelfinavir, 90M for saquinavir, and 82A/T and 46I/L for indinavir as most probable major resistance mutations. Moreover we found striking similarities for the role of many mutations against all of these drugs. For example, for all three inhibitors, we found that the novel mutation 89I was minor and associated with mutations at positions 90 and 71. Bayesian network learning provides an autonomous method to gain insight in the role of resistance mutations and the influence of HIV-1 natural variation. We successfully applied the method to three protease inhibitors. The analysis shows differences with current knowledge especially concerning resistance development in several non-B subtypes.

Optimization of a genotypic assay applicable to all human immunodeficiency virus type 1 protease and reverse transcriptase subtypes.

Genotypic assays are used often to guide clinicians in decisions concerning the treatment of patients. An optimized sequence-based genotypic assay was used to determine the whole protease and reverse transcriptase (RT) gene, including the gag cleavage site region and RNase H region. Since non-B subtypes are increasing in countries where subtype B was the most prevalent subtype, and treatment becomes more available in developing countries where the epidemic is characterized by a high prevalence of non-B subtypes, it was important that the genotypic test was evaluated using a panel of different subtypes. Amplification was successful for different subtypes: A, B, C, D, F, G, H, J, CRF01_AE, CRF02_AG, CRF11_cpx, CRF13_cpx and an uncharacterized recombinant sample. The detection limit of the PCR was 1000 copies/ml, except for 1 subtype C sample (PL3) and 1 CRF02_AG sample (PL8). The detection limit for these samples was 5000 copies/ml. A sequence could be obtained in both directions for most of the samples.

Evaluation of two commercial kits for the detection of genotypic drug resistance on a panel of HIV type 1 subtypes A through J.

We compared the two commercially available sequencing kits for HIV-1 drug resistance testing, the ViroSeq Genotyping System (Applied Biosystems, Foster City, CA, U.S.A.) and the TRUGENE HIV-1 Genotyping Kit (Visible Genetics, Inc., Toronto, Ontario, Canada), with our in-house genotyping system. Fifteen viral isolates from African patients (6 treated and 9 untreated) covering a panel of HIV-1 subtypes A through J and 7 plasma samples from Belgian and African patients (2 treated and 5 untreated) were tested. All the samples could be amplified and sequenced by the three systems; however, for all systems, alternative amplification/sequencing primers had to be used for some samples belonging to subtype B as well as to other subtypes. The consensus sequence was partially derived from only one strand for the in-house system and for the ViroSeq Genotyping System. The TRUGENE HIV-1 Genotyping Kit scored the highest number of ambiguities, followed by the ViroSeq Genotyping System and the in-house system. For 11 samples, these differences in reporting mixtures affected 14 resistance-related positions, which altered the interpretation toward protease inhibitors for 2 samples when using version 1.2 RetroGram software (Virology Networks, Utrecht, The Netherlands). All three systems were able to sequence diluted samples with a viral load down to 10 3 or 10 4 RNA copies/ml. Our data therefore suggest that the performance of amplification and sequencing primers must be improved to allow fast and reliable resistance testing for all HIV-1 subtypes.

Effect of stenting on coronary flow velocity reserve: comparison of coil and tubular stents.

OBJECTIVE: To determine whether coil stents are as effective as tubular stents in improving coronary flow velocity reserve (CFVR) after stent deployment. METHODS: Distal CFVR was measured with a 0. 014 inch Doppler guide wire before and after stenting in 33 patients. A coil stent was implanted in 16 patients and a tubular stent was used in 17 patients. Coronary flow velocity within the stent was also recorded during a slow pullback. RESULTS: Following placement of the stents, the percentage diameter stenosis was similar for both the tubular and coil stents (mean (SE) 11 (2)% v 13 (2)%, NS). However, distal CFVR was higher after stenting with a tubular stent compared with a coil stent (2.46 (0.13) v 1.96 (0.14), p < 0.05). Furthermore, pullback through the stent detected a major flow velocity increase within coil stents but not in tubular stents (83 (24)% v 5 (5)%, p < 0.05). CONCLUSIONS: In spite of similar angiographic improvement, placement of coil stents was associated with inferior functional results compared with tubular stents. The flow velocity acceleration within the coil stents suggests the presence of a residual narrowing within the stent, which is not appreciated on angiography.

User-friendly and low-cost computer system for immediate review, analysis, and reconstruction of intracoronary ultrasound images.

Rapid review, digital recording, on-line quantification, and three-dimensional reconstruction are all essential in the evaluation of intracoronary ultrasound images during coronary interventions. We describe a low-cost method that offers all these necessary features. The proposed method uses the QuickTime compatible video digitizers of standard multimedia Apple Macintosh or PowerPC desktop computers and the freeware software Object Image 1.60.

An outbreak of Burkholderia cepacia with septicemia on a cardiology ward.

During a 3-day period, eight patients developed septicemia with Burkholderia cepacia. Heparin injection was found to be a risk factor. Heparin was diluted with dextrose solution, which was aspirated from a 1-L bag. B cepacia, genotypically identical to the blood isolates, was isolated from this bag.

Adenosine technetium-99m sestamibi single-photon emission tomography for the assessment of jeopardized myocardium early after acute myocardial infarction. Paradoxical scintigraphic underestimation of jeopardized myocardium in patients with a severe infarct-related stenosis.

This study investigated the value of technetium-99m sestamibi scintigraphy in identifying patients at risk for post-infarct ischaemia (=jeopardized myocardium), especially within the reperfused infarct region. In 51 patients with a recent (<1 month) myocardial infarction, adenosine 99mTc-sestamibi single-photon emission tomography (SPET) and dobutamine stress echocardiography (DSE) were performed and correlated with the presence of significant coronary artery stenosis [% diameter stenosis (DS) >50%] on quantitative coronary angiography. Regional perfusion activity was analysed semi-quantitatively (score 0-4) on a 13-segment left ventricular model. DSE was used for the estimation of the infarct size (low-dose DSE) and for concomitant evaluation of ischaemia (high-dose DSE). A reversible perfusion defect within the infarct region was observed in 20 of the 37 patients with a significant infarct-related lesion (sensitivity of 54%) and only in one patient without a significant infarct-related lesion (specificity of 93%). Further analysis revealed that the scintigraphic assessment of jeopardized myocardium was fairly good in patients with a moderate (DS 51%-64%) infarct-related stenosis but was inadequate in patients with a severe (DS>/=65%) infarct-related stenosis (sensitivity of 80% vs 36%, P<0.01), while the echocardiographic detection of ischaemia was not influenced by stenosis severity (sensitivity of 73% in both subgroups). This scintigraphic underestimation of jeopardized myocardium was mainly related to a severely impaired myocardial perfusion under baseline conditions, as was evidenced by a significantly more severe rest perfusion score in the infarct region in patients with a severe stenosis as compared to those with a moderate stenosis (average score: 1.5+/-0.7 vs 2.1+/-0.6, P<0.01), while infarct size on echocardiography was similar for both subgroups. It may be concluded that early after an acute myocardial infarction, adenosine 99mTc-sestamibi SPET may underestimate reperfused but still jeopardized myocardium, particularly in patients with a severe infarct-related stenosis. In these patients the evaluation of the ischaemic burden on rest-stress scintigraphy is hampered by the presence of a severely impaired myocardial perfusion in resting conditions.

Coronary flow reserve during coronary angioplasty in patients with a recent myocardial infarction: relation to stenosis and myocardial viability.

OBJECTIVES: In the present study, we examined post-stenotic coronary flow before and after percutaneous transluminal coronary angioplasty (PTCA) in patients with and without a recent myocardial infarction (MI) and related it to stenosis severity and residual viability. BACKGROUND: Post-stenotic coronary blood flow velocity reserve (CFVR) has been used with success to estimate functional stenosis severity in patients with stable angina. However, in patients with a recent MI, the impaired coronary vasodilator response of the reperfused myocardium may substantially alter the flow dynamics of the infarct-related artery. METHODS: Distal coronary flow velocities were recorded before and after PTCA in 36 patients at day 13 +/- 7 (mean +/- SD) after acute MI and in 38 patients without MI. The CFVR was assessed by the ratio of distal hyperemic to baseline average peak velocity, using a 0.014-in. Doppler guide wire. Stenosis severity was analyzed by quantitative coronary angiography, and infarct size was assessed scintigraphically. RESULTS: For similar angiographic stenosis severity, pre- and post-PTCA values of CFVR were significantly lower in patients with than without MI: 1.22 +/- 0.26 versus 1.50 +/- 0.45 before PTCA (p < 0.05) and 1.72 +/- 0.43 versus 2.21 +/- 0.74 after PTCA, respectively (p < 0.01). Although CFVR increased significantly (p < 0.0001) after angiographically successful PTCA in both study groups, abnormal CFVR (< or = 2.0) was still observed in 80% of patients with MI and in 44% of those without MI (MI vs. no MI, p = 0.001). Patients with an extensive infarction (relative infarct size > or = 50%) and those with a small infarction (relative infarct size < 50%) had comparable levels of post-PTCA CFVR (1.6 +/- 0.3 vs. 1.8 +/- 0.5, p = NS). Among a variety of factors, angiographic stenosis severity was the most important determinant of CFVR in both study groups. CONCLUSIONS: In patients with a recent MI, CFVR was significantly lower than in those without MI, both before and after PTCA. Besides the presence of this postreperfusion-related impairment of the coronary vasodilating response, CFVR was mainly influenced by stenosis severity and not by residual viability.

Comparative study of rest technetium-99m sestamibi SPET and low-dose dobutamine stress echocardiography for the early assessment of myocardial viability after acute myocardial infarction: importance of the severity of the infarct-related stenosis.

Rest technetium-99m sestamibi single-photon emission tomography (SPET) has been shown to underestimate viability in some patients with chronic ischaemic myocardial dysfunction. The present study was designed to appraise the value of 99mTc-sestamibi as a viability tracer in patients with a recent myocardial infarction and to determine factors that might influence its accuracy in assessing infarct size. Therefore, rest 99mTc-sestamibi SPET, low-dose dobutamines stress echocardiography and quantitative coronary angiography were performed in 51 patients with a recent myocardial infarction. Perfusion activity and regional wall motion were scored semi-quantitatively using the same segmental division of the left ventricle. Assessment of 99mTc-sestamibi uptake as a marker of viability was performed by comparing a binary uptake score (viable=>50% vs necrotic =/=65%-100%) and particularly those with "late" reperfusion therapy (time delay >/=180 min). In patients without a severe infarct-related stenosis, 99mTc-sestamibi was able to accurately distinguish viable from necrotic segments. Thus, rest 99mTc-sestamibi scintigraphy early after acute myocardial infarction may underestimate residual viability within the infarct region, particularly in patients with low flow state coronary anatomy, as a result of a severe infarct-related stenosis and/or late reperfusion therapy.

Aminophylline inhibits adaptation to ischaemia during angioplasty. Role of adenosine in ischaemic preconditioning.

UNLABELLED: The ability of brief periods of ischaemia to protect the heart from subsequent ischaemia has been termed "ischaemic preconditioning'. In order to assess the role of adenosine receptor stimulation in this phenomenon we studied the ischaemic preconditioning effect during angioplasty in 10 control patients and in 10 patients pre-treated with 5 mg.kg-1 aminophylline, an adenosine receptor antagonist. The ischaemic response was assessed by analysis of the intracoronary electrocardiogram every 10 s during three consecutive inflations of 90 s with a reperfusion time of 180 s. The severity of transmural local ischaemia was expressed as the magnitude of the ST segment shift in relation to the time during each inflation. The control patients showed an improved tolerance to myocardial ischaemia: ST segment shift decreased from 1.42 +/- 0.49 mV at the end of the first inflation to 1.03 +/- 0.44 mV at the end of the third inflation (P < 0.001). However, in patients pre-treated with aminophylline, the ischaemic response was not significantly different during three inflations. CONCLUSION: Aminophylline inhibits ischaemic preconditioning, as assessed by analysis of the intracoronary. ST segment changes during angioplasty. This suggests that ischaemic preconditioning is mediated by adenosine receptor stimulation in humans.

Clinical outcome of patients with an uncomplicated myocardial infarction: effect of revascularization.

In 80 patients (pts) with an uncomplicated myocardial infarction (MI) the rate of major cardiac events (MACE) including cardiac death, non-fatal myocardial infarction and recurrent ischemia requiring hospitalization was prospectively assessed over a mean follow-up period of 17 +/- 9 months and related to clinical, angiographic and scintigraphic findings, the latter obtained from adenosine Tc-99m sestamibi SPECT imaging. Decision for revascularization was mainly based on angiographic data and was carried out in a total of 50 patients (angioplasty in 34 pts and cardiac surgery in 16 pts). The overall MACE rate was 24% with a mortality and myocardial infarction rate of 4% and 5%, respectively. Early (< 2 months) revascularization seemed to have a beneficial effect on clinical outcome as was suggested by the following findings: 1) Cardiac events (MACE) were not significantly different in patients with versus without revascularization (MACE 24% versus 23%) although the former constituted a subgroup at higher risk for ischemic events because of a more extensive coronary artery disease state. 2) In the subset of patients with at least one significant coronary artery stenosis the clinical outcome was significantly better in those who were revascularized than in those who underwent no revascularization (MACE 24% vs 47%, p < 0.05. Among a variety of factors, including the scintigraphic and angiographic extent of coronary artery disease and post-MI treatment strategy, multivariate analysis selected hypercholesterolemia (> 240 mg%) as the only independent predictor of MACE with a more than fourfold increase in risk for development of MACE. These data suggest that the natural history, especially the rate of recurrent ischemic events, can be favourably changed by an elective and early revascularization, strategically oriented by the results of the angio-graphic study. Furthermore, our data emphasized the deleterious role of hypercholesterolemia on clinical outcome in patients with a recent MI.

Semi-automatic external defibrillation and implanted cardiac pacemakers: understanding the interactions during resuscitation.

Many emergency medical service (EMS) systems are currently implementing semi-automatic external defibrillation (AED) by emergency medical technicians. Surprisingly little information is available on the possible interactions between AEDs and implanted cardiac pacemakers. Therefore, at present there are no clear guidelines for the use of AEDs on patients having a cardiac pacemaker. During resuscitation, multiple interactions between pacemakers and AEDs are possible. External defibrillation can cause damage to several functions of the pacemaker. On the other hand, the presence of pacemaker spikes during cardiac arrest might prohibit recognition of the ventricular fibrillation by the AED. We report on two resuscitation attempts in which the interaction between the ventricular fibrillation, an implanted dual chamber pacemaker and the AED was decisive for the defibrillation success. A clear understanding of these possible interactions is necessary for the further refining of diagnostic algorithms and clinical strategies of prehospital defibrillation.

Adenosine technetium-99m sestamibi (SPECT) for the early assessment of jeopardized myocardium after acute myocardial infarction.

The purpose of this study was to evaluate the accuracy of adenosine Tc-99m sestamibi single photon emission computed tomography (SPECT) in the detection of jeopardized myocardium early after acute myocardial infarction. Coronary arteriography and myocardial scintigraphy were performed in 50 consecutive patients with an uncomplicated myocardial infarction. Myocardium was considered jeopardized if a significant infarct-related vessel stenosis (> 50% diameter stenosis) supplied an infarct area with residual viable tissue. Perfusion reversibility in the infarct region occurred in 25 patients (50%) and was almost solely observed in the presence of jeopardized myocardium. Non-reversible perfusion defects in the infarct region were found in patients without jeopardized myocardium. This subgroup consisted of either patients without significant vessel stenosis or patients without significant residual viability in the infarct region. Adenosine Tc-99m sestamibi SPECT had an accuracy of 88% for the detection of jeopardized myocardium. Side effects during adenosine infusion were frequently observed but well tolerated. These results suggest that adenosine Tc-99m sestamibi SPECT is an accurate non-invasive method for detecting jeopardized myocardium after acute myocardial infarction and may be a valuable non-invasive test for the early selection of patients at risk for future ischaemic events.

High atrial pacing impedance: efficient or wasteful? A comparison of an active and a passive fixation lead.

We compared the stimulation characteristics of two atrial fixation leads: the CPI model 4269 (n = 45) and Cordis-Telectronics model 327-752 (n = 42). The CPI lead uses an active fixation method, whereas the Cordis-Telectronics lead is fixated passively. Impedance and threshold were measured at implant and during 12 months of follow-up. P wave sensing was good with both types of leads. Follow-up of these 87 leads showed that both the impedance and threshold increased with the active leads, but not with the passive fixation leads. In the active fixation group, regardless of the high atrial pacing impedance, 93% could still be programmed to 2.5 V-0.6 msec (with a 2:1 threshold safety margin). It is concluded that the high chronic pacing impedance of the active fixation leads will be beneficial on current drain if no major increase in pacing threshold occurs simultaneously.

Nosocomial septicaemias in a cardiologic ward. A case-control study.

Nosocomial septicaemias were studied in a cardiologic ward at the University Hospital of Antwerp, Belgium. The incidence of nosocomial infections was found to be 2.7 per 1,000 admissions per year. A case-control study suggested that in contrast with coronarography and pacemaker implantation, percutaneous transluminal coronary angioplasty may be a risk factor for nosocomial septicaemia (OR 4.5, 95% confidence interval 0.7-33.4).

Paradoxic pulmonary vasoconstriction in response to acetylcholine in patients with primary pulmonary hypertension.

Pulmonary vascular reactivity was assessed during diagnostic heart catheterization in two patients with pulmonary hypertension unexplained by pulmonary or cardiac disease and in five patients with atypical chest pain and normal coronary arteriograms. Acetylcholine, an endothelium-dependent vasodilator that also has a direct contracting effect on vascular smooth muscle cells, was infused in the right atrium in a step-wise increasing dose in order to obtain final blood concentrations in the pulmonary circulation ranging from 10(-6) mol/L to 10(-4) mol/L. In the five control patients, acetylcholine induced a dose-related decrease of pulmonary vascular resistance (-52 percent +/- 9 percent). In the patients with primary pulmonary arterial hypertension, however, acetylcholine caused a paradoxic increase of pulmonary arterial pressure and of pulmonary vascular resistance. Thus, it appears that endothelium-dependent vasodilation is impaired in the pulmonary circulation of patients with primary pulmonary arterial hypertension. Endothelial dysfunction in the pulmonary circulation may play a role in the pathophysiology of this disease.