Low-Frequency Drug Resistance in HIV-Infected Ugandans on Antiretroviral Treatment Is Associated with Regimen Failure

Most patients failing antiretroviral treatment in Uganda continue to fail their treatment regimen even if a dominant drug-resistant HIV-1 genotype is not detected. In a recent retrospective study, we observed that approximately 30% of HIV-infected individuals in the Joint Clinical Research Centre (Kampala, Uganda) experienced virologic failure with a susceptible HIV-1 genotype based on standard Sanger sequencing. Selection of minority drug-resistant HIV-1 variants (not detectable by Sanger sequencing) under antiretroviral therapy pressure can lead to a shift in the viral quasispecies distribution, becoming dominant members of the virus population and eventually causing treatment failure. Here, we used a novel HIV-1 genotyping assay based on deep sequencing (DeepGen) to quantify low-level drug-resistant HIV-1 variants in 33 patients failing a first-line antiretroviral treatment regimen in the absence of drug-resistant mutations, as screened by standard population-based Sanger sequencing. Using this sensitive assay, we observed that 64% (21/33) of these individuals had low-frequency (or minority) drug-resistant variants in the intrapatient HIV-1 population, which correlated with treatment failure. Moreover, the presence of these minority HIV-1 variants was associated with higher intrapatient HIV-1 diversity, suggesting a dynamic selection or fading of drug-resistant HIV-1 variants from the viral quasispecies in the presence or absence of drug pressure, respectively. This study identified low-frequency HIV drug resistance mutations by deep sequencing in Ugandan patients failing antiretroviral treatment but lacking dominant drug resistance mutations as determined by Sanger sequencing methods. We showed that these low-abundance drug-resistant viruses could have significant consequences for clinical outcomes, especially if treatment is not modified based on a susceptible HIV-1 genotype by Sanger sequencing. Therefore, we propose to make clinical decisions using more sensitive methods to detect minority HIV-1 variants.     

A very human being: Sister Marie Simone Roach, 1922–2016

Sister (Sr.) Marie Simone Roach, of the Sisters of St. Martha of Antigonish, Nova Scotia, died at the Motherhouse on 2 July 2016 at the age of 93, leaving behind a rich legacy of theoretical and practical work in the areas of care, caring and nursing ethics. She was a humble soul whose deep and scholarly thinking thrust her onto the global nursing stage where she will forever be tied to a central concept in nursing, caring, through her Six Cs of Caring model. In Canada, she was the lead architect of the Canadian Nurses Association's first code of ethics, and her influence on revisions to it is still profound more than 35 years later. In this paper, four global scholars in nursing and ethics are invited to reflect on Sr. Simone's contribution to nursing and health‐care, and we link her work to nursing and health‐care going forward. 1     

Challenges of mucositis assessment in children: Expert opinion

Mucositis is a challenging treatment-related complication in children receiving therapy for cancer. The conduct of clinical trials that investigate mucositis prevention and treatment requires adequate evaluation of the oral cavity. However, few instruments to measure mucositis in children have been appropriately developed or evaluated. A focus group of nine health care professionals with expertise in mucositis assessment, oral assessment in children and paediatric cancer aimed to determine the challenges and possible solutions to mucositis assessment in children. The results led to the identification of several areas of concern that included: (1) challenges in oral assessment in children related to age and cooperation, (2) the need for proxy responses while recognizing the challenges of reporting pain and function attributed to oral mucositis, (3) the need for an instrument that is simple, quick to complete, and easy to use in almost all children and (4) educational considerations. The results provide a basis from which guidelines for the oral assessment of mucositis in children can begin. This information could be used to aid in the development of a new scale for the assessment of oral mucositis in children.     

Fate and function of hepatitis-C-virus-specific T-cells during peginterferon-alpha2b therapy for acute hepatitis C

BACKGROUND: Strong hepatitis C virus (HCV)-specific T-cell responses are associated with spontaneous clearance of acute hepatitis C. However, recent studies described a decline in HCV-specific CD8+ T-cells during interferon treatment, suggesting that the success of acute HCV therapy might be independent of adaptive immunity. METHODS: T-cell responses of eight human leukocyte antigen (HLA)-A2-positive, acutely infected patients treated with peginterferon-alpha2b were studied by ELISPOT and proliferation assays and flow cytometry analysis using HCV-specific tetramers. RESULTS: HCV-specific T-cells predominately declined during therapy. However, diverse patterns of CD4+ and CD8+ T-cell kinetics were observed. In patients with sustained virological response chemokine receptor 3 (CXCR-3) expression of HCV-specific CD8+ T-cells was upregulated, indicating homing to the liver. Low levels of T-cells remained detectable throughout treatment and follow up. In contrast, T-cells of a relapse patient did not upregulate CXCR-3 but displayed a higher staining for annexin-V, followed by a complete loss of peripheral virus-specific CD8+ T-cells by week 12. CONCLUSIONS: Kinetics of HCV-specific T-cell responses are heterogeneous in interferon-treated patients with acute hepatitis C. The decline of T-cells might be a consequence of both apoptosis and homing. The balance between cell death and regulation of chemokine receptors might lead to different long-term outcomes.     

Correlation of the MIC and dose/MIC ratio of fluconazole to the therapeutic response of patients with mucosal candidiasis and candidemia

We report on the correlation of the outcomes for two cohorts of patients who had been treated for candidemia (126 episodes) or oropharyngeal candidiasis (110 episodes) with various doses of fluconazole and the MIC of fluconazole obtained by using the EUCAST standard for fermentative yeasts. Of 145 episodes caused by an isolate with a fluconazole MIC < or =2 mg/liter, 93.7% (136 of 145) responded to fluconazole treatment. The response for those infected with a strain with a MIC of 4 mg/liter was 66% but reached 100% when the dose was greater than 100 mg/day, whereas the response for those infected with strains with MICs > or =8 mg/liter was only 12%. Hence, a MIC of 2 mg/liter or 4 mg/liter was able to predict successful treatment. A cure rate of 93.9% (140 of 149) was achieved when the dose/MIC ratio was > or =100 but fell to 14.6% (16 of 109) when the ratio was less. The dose/MIC required to achieve a response rate of 50% (the 50% effective concentration) was 43.7 for the cohort of patients with oropharyngeal candidiasis. Classification and regression analysis indicated that a dose/MIC of 35.5 was the threshold for the prediction of cure or failure. However, an increase in exposure above this threshold further increased the probability of cure, and all patients were cured when the dose/MIC exceeded 100. Monte Carlo simulations showed a probability of target attainment of 99% at MICs < or =2 mg/liter and a pharmacodynamic target of a dose/MIC ratio of 100, which was equivalent to an unbound fraction of the fluconazole area under the curve versus the MIC of 79.     

A single unit lymphoma experience: outcome in a Cape Town academic centre.

To document outcome in Hodgkin and other lymphomas from a privately based academic centre the clinical records from 253 consecutive referrals were analysed. Diagnosis was according to World Health Organization criteria, prognosis assigned by the international index and therapy risk-stratified with results subject to appropriate statistical methodology. None of these patients underwent transplantation. For the cohort the median age was 55 years (range 11-94) and 63% were male. Constitutional symptoms were present in 22%; a quarter had previous chemotherapy and a third some form of irradiation prior to referral. Fifty-seven percent were stage I or II and 21% had nodal disease above and below the diaphragm whilst in the remainder cells were present in the circulation and this included the subset of chronic lymphocytic leukaemia -- small lymphocytic lymphoma. Positron emission scanning was not available for these studies. Median survival for the cohort is 3.2 years and reduced to 1.3 years by the presence of unexplained fever, sweating or inappropriate weight loss. Further adverse factors included any prior treatment, intermediate or high-grade histopathology, risk factors defined by the International Prognostic Index as well as late Rai stages. Analysed by disease category Hodgkin lymphoma (n=17) when managed according to the German Study Group protocols and hairy cell leukaemia (n=10) treated with two chlorodeoxyadenosine -- both had a stable plateau in excess of 90%. The corresponding figures for follicular variants (n=31) was 72% in the low risk and 58% in the remainder when treated with cyclophosphamide, vincristine and prednisone. Curves for the aggressive or diffuse large B-cell lymphoma (n=44) fell initially to 48%, but relapse continued in stages III and IV to the current level of 18% when receiving cyclophosphamide, hydroxydaunorubicin, vincristine and prednisone on the 21-day schedule. Chronic lymphocytic leukaemia -- small lymphocytic lymphoma (n=58) were initially given pulsed chlorambucil and sustained response was over 90% with low bulk, but declined to reach 30% as prognostic score rose. The miscellaneous categories (n<5 each) managed variably, but using the same criteria, were pooled and are presently at 62% and 30% for high and low grades. It is concluded that precise diagnosis, accurate staging and therapy on standardised risk-stratified programmes, delivered uniformly by a single multidisciplinary group, creates the all-important centre effect; matching figures are unlikely to apply outside these disciplined circumstances. The expectation from patients and referring physicians alike is that, since lymphomas are potentially curable, such an approach to comprehensive management will be regarded as standard even in an under resourced or Third World country. It follows that late referral and prior therapy will adversely affect performance status and compromise life span: These alternative approaches are inappropriate and strongly discouraged.     

Establishment, value assignment, and characterization of new WHO reference reagents for six molecular forms of human chorionic gonadotropin

BACKGROUND: The International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) established a Working Group to investigate means of improving the comparability of immunoassays for human chorionic gonadotropin (hCG), which was selected as a prototype glycoprotein analyte. The Working Group identified development of unambiguous nomenclature and production of new highly purified International Reference Reagents calibrated in substance concentrations as its primary objectives. METHODS: Preparations of intact hCG, nicked hCG, hCG beta-subunit, nicked hCG beta-subunit, hCG alpha-subunit, and hCG beta-core fragment were purified from a crude urinary hCG preparation, ampouled, lyophilized, and assigned values in substance concentrations (mol/L). Value assignment and accelerated degradation studies were carried out in accordance with WHO protocols for International Reference Reagents. RESULTS: The ampouled standards were assigned final values based on the recovery of immunoreactive material after reconstitution. The degradation studies showed that the standards were highly stable. CONCLUSIONS: The nomenclature of hCG-related molecules and immunoassays has been adopted by the IFCC, and the standards prepared and characterized by the Working Group have been formally adopted by the WHO as the First International Reference Reagents for six hCG-related molecules. These developments will enable better understanding of what assays for hCG measure and should ultimately help to improve the clinical application of these assays.     

Ethyl pyruvate improves systemic and hepatosplanchnic hemodynamics and prevents lipid peroxidation in a porcine model of resuscitated hyperdynamic endotoxemia

OBJECTIVE: To investigate the systemic, pulmonary, and hepatosplanchnic hemodynamic and metabolic effects of delayed treatment with ethyl pyruvate in a long-term porcine model of hyperdynamic endotoxemia. DESIGN: Prospective, randomized, controlled experimental study with repeated measures. SETTING: Investigational animal laboratory. SUBJECTS: Anesthetized, mechanically ventilated, and instrumented pigs. INTERVENTIONS: After 12 hrs of continuous infusion of lipopolysaccharide and hydroxyethyl starch to keep mean arterial pressure >60 mm Hg, swine randomly received placebo (Ringer's solution; control group, n = 11) or ethyl pyruvate in lactated Ringer's solution (n = 8; 0.03 g.kg(-1) loading dose over 10 mins, thereafter 0.03 g.kg(-1)hr(-1) for 12 hrs). MEASUREMENTS AND MAIN RESULTS: Whereas mean arterial pressure significantly decreased in control animals, mean arterial pressure was maintained at the baseline level in pigs treated with ethyl pyruvate. Global oxygen uptake was comparable, so that the trend toward a higher oxygen transport and the significantly higher mixed venous hemoglobin oxygen saturation resulted in a significantly lower oxygen extraction in the ethyl pyruvate group. Ethyl pyruvate reduced intrapulmonary venous admixture and resulted in significantly greater Pa(O2)/F(IO2) ratios. Despite comparable urine production in the two groups during the first 18 hrs of endotoxemia, ethyl pyruvate significantly increased diuresis during the last 6 hrs of the study. Lipopolysaccharide-induced systemic and regional venous metabolic acidosis was significantly ameliorated by ethyl pyruvate. Endotoxemia increased both blood nitrate + nitrite and isoprostane concentrations, and ethyl pyruvate attenuated the response of these markers of nitric oxide production and lipid peroxidation. CONCLUSIONS: Ethyl pyruvate infusion resulted in improved hemodynamic stability and ameliorated acid-base derangements induced by chronic endotoxemia in pigs. Reduced oxidative stress and an decreased nitric oxide release probably contributed to these effects.     

Complications of implantable cardioverter defibrillator therapy in 440 consecutive patients

BACKGROUND: Although more than 150,000 implantable cardioverter defibrillators (ICDs) are implanted yearly worldwide, only few studies systematically examined complications of ICD therapy in large patient cohorts. METHODS: We prospectively analyzed ICD-related complications in 440 consecutive patients who underwent first implantation of an ICD system for primary or secondary prevention of sudden cardiac death within the last 10 years at our institution. All study patients received pectoral nonthoracotomy ICD lead systems with the exception of one patient who had an artificial tricuspid valve. RESULTS: During 46 +/- 37 months follow-up, 136 of 440 patients (31%) experienced at least one complication including implant procedure-related complications in 43 patients (10%), ICD generator-related complications in 28 patients (6%), lead-related complications in 52 patients (12%), and inappropriate shocks in 54 patients (12%). The most serious complications included one perioperative death due to heart failure (0.2%), two ICD system infections necessitating device removal (0.5%) and two perioperative cerebrovascular strokes (0.5%). CONCLUSIONS: We conclude that more than one quarter of ICD patients experience complications during a mean follow-up of almost 4 years, although serious complications such as intraoperative death or ICD system infections are rare in patients with nonthoracotomy ICD systems. Recognition of these complications is the prerequisite for advances in ICD technology and management strategies to avoid their recurrence.