Prevalence of baseline polymorphisms for potential resistance to NS5A inhibitors in drug-naive individuals infected with hepatitis C genotypes 1–4

Background: The non-structural 5A (NS5A) protein of HCV is a multifunctional phosphoprotein involved in regulation of viral replication and virion assembly. NS5A inhibitors targeting domain I of NS5A protein have demonstrated high potency and pan-genotypic antiviral activity, however they possess a low genetic barrier to resistance. At present, only genotype 1, the most prevalent HCV genotype has been studied in detail for resistant variants. Methods: Utilising a panel of genotypic-specific resistance assays, population sequencing was performed on plasma derived viral RNA isolated from 138 patients infected with HCV genotypes 1-4 and not treated with directly acting anti-viral agents (DAAs). Amino acid changes in HCV NS5A domain I at codon positions 28, 30, 31, 32 and 93, reported to confer reduced susceptibility to certain NS5A inhibitors were examined. Additionally, genotypic outcome based on NS5A sequences were compared with LiPA and Abbott® real time. Results: Amino acid substitutions associated with moderate to high level resistance to NS5A inhibitors were detected in 2/42 (4.76%) HCV-1a, 3/23 (13.04%) HCV-1b, 4/26 ( 15.38% ) HCV-2, 1/24 (4.17%) HCV-3 and 1/23 (4.35%) HCV-4 infected patients who had not been treated with NS5A inhibitors. Genotype prediction based on NS5A sequences were concordant with LiPA and/or Abbott® real-time for 97.10% of cases. Conclusion: Primary resistance mutations associated with resistance to first generation NS5A inhibitors such as Daclatasvir (DCV) were observed in all genotypes, albeit at low frequencies. An excellent correlation based on NS5A genotyping and LiPA or Abbott® real-time was achieved

The Cholecystectomy As A Day Case (CAAD) Score: A Validated Score of Preoperative Predictors of Successful Day-Case Cholecystectomy Using the CholeS Data Set

Background Day-case surgery is associated with significant patient and cost benefits. However, only 43% of cholecystectomy patients are discharged home the same day. One hypothesis is day-case cholecystectomy rates, defined as patients discharged the same day as their operation, may be improved by better assessment of patients using standard preoperative variables. Methods Data were extracted from a prospectively collected data set of cholecystectomy patients from 166 UK and Irish hospitals (CholeS). Cholecystectomies performed as elective procedures were divided into main (75%) and validation (25%) data sets. Preoperative predictors were identified, and a risk score of failed day case was devised using multivariate logistic regression. Receiver operating curve analysis was used to validate the score in the validation data set. Results Of the 7426 elective cholecystectomies performed, 49% of these were discharged home the same day. Same-day discharge following cholecystectomy was less likely with older patients (OR 0.18, 95% CI 0.15–0.23), higher ASA scores (OR 0.19, 95% CI 0.15–0.23), complicated cholelithiasis (OR 0.38, 95% CI 0.31 to 0.48), male gender (OR 0.66, 95% CI 0.58–0.74), previous acute gallstone-related admissions (OR 0.54, 95% CI 0.48–0.60) and preoperative endoscopic intervention (OR 0.40, 95% CI 0.34–0.47). The CAAD score was developed using these variables. When applied to the validation subgroup, a CAAD score of ≤5 was associated with 80.8% successful day-case cholecystectomy compared with 19.2% associated with a CAAD score >5 (p < 0.001). Conclusions The CAAD score which utilises data readily available from clinic letters and electronic sources can predict same-day discharges following cholecystectomy

Denominations Routines of identification in Northern Irish politics

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Heart rate variability in older workers during work under the Threshold Limit Values for heat exposure

Background: The Threshold Limit Values (TLV) of the American Conference of Governmental and Industrial Hygienists indicate the levels of heat stress that all workers may be repeatedly exposed to without adverse health effects. In this study, we evaluated heart rate variability (HRV) during moderate-to-heavy work performed continuously or according to different TLV work-rest (WR) allocations in healthy physically active older workers. Methods: Nine healthy older (58 ± 5 years) males performed three different 120-minute conditions in accordance with TLV guidelines for moderate-to-heavy intensity work (360 W fixed rate of heat production) in different wet-bulb globe temperatures (WBGT): continuous cycling at 28°C WBGT (CON), as well as intermitted work performed at WR of 3:1 in 29°C WBGT (WR3:1), and at WR of 1:1 at 30°C (WR1:1). Rectal temperature and HRV (3-lead electrocardiogram [ECG]) were assessed throughout. Results: Coefficient of Variation, Poincaré SD2, and Shannon Entropy were decreased during the CON compared with the WR3:1 when core temperature exceeded 38°C and after 1 hour of continuous work (P <.05). Also, 4 of the 12 HRV indices studied were reduced at CON compared with WR1:1 after 2 hours of accumulated work time (P <.05). Participants worked longer before core temperature reached 38°C during the WR1:1 and the WR3:1, compared with CON (P <.05). Conclusions: Incorporating breaks during moderate-to-heavy work in the heat for older adults can reduce autonomic stress and prolong the work performed at safe core temperature levels. The TLV WR1:1 provides increased cardiac protection for older workers, as compared with the CON and the WR3:1. © 2020 Wiley Periodicals LL

Evaluation of sequencing of HCV core/E1, NS5A and NS5B as a genotype predictive tool in comparison with commercial assays targeting 5′UTR

Background Hepatitis C virus (HCV) genotyping is required for tailoring the dose and duration of antiviral therapy, predicting virological response rates, and selecting future treatment options. Objective To establish whether baseline genotypes, performed by INNO-LiPA Version 1.0 (v1.0), before 2008, were valid for making treatment decisions now or whether genotypic determination should be repeated. Furthermore, to evaluate concordance between Abbott RealTime genotype II assay (RT) and genotyping by sequencing HCV C/E1, NS5A, NS5B. Study design Genotyping by RT and sequencing was performed on paired historic and current specimens from 50 patients previously baseline genotyped using INNO-LiPA. Results Of 100 samples from 50 patients, ≥2 of HCV genomic target regions yielded a sequence that was suitable for genotyping, with 100% concordance, providing no evidence of recombination events. Genotype and subtype prediction based on RT and sequencing agreed in 62.8% historic and 72.7% current specimens, with a kappa coefficient score of 0.48 and 0.76, respectively. LiPA could not subtype 46% of HCV gt1 infections, and LiPA subgenotype was only in agreement with RT and sequencing in 28.6% cases, where matched baseline and historic specimens were available. Three patients were indeterminate by RT, and five patients with HCV gt1 infections could not be subtyped by RT. However, RT revealed mixed infections in five patients where sequencing detected only single HCV infection at 20% threshold. Conclusion Genotyping by sequencing, exhibited excellent concordance, with moderate to good agreement with RT, and could resolve RT indeterminates and subtype HCV-gt1 infections not possible by LiPA

Evaluation of the Roche prototype 454 HIV-1 ultradeep sequencingdrug resistance assay in a routine diagnostic laboratory

Background - Studies have shown that low-frequency resistance mutations can influence treatment outcome. However, the lack of a standardized high-throughput assay has precluded their detection in clinical settings. Objective - To evaluate the performance of the Roche prototype 454 UDS HIV-1 drug resistance assay (UDS assay) in a routine diagnostic laboratory. Study design - 50 plasma samples, previously characterized by population sequencing and that had shown ≥1 resistance associated mutation (RAM), were retrospectively tested by the UDS assay, including 18 B and 32 non-B subtypes; viral loads between 114–1,806,407 cp/ml; drug-naive (n = 27) and drug-experienced (n = 23) individuals. Results - The UDS assay was successful for 37/50 (74%) samples. It detected all RAMs found by population sequencing at frequencies above 20%. In addition, 39 low-frequency RAMs were exclusively detected by the UDS assay at frequencies below 20% in both drug-naïve (19/26, 73%) and drug-experienced (9/18, 50%) individuals. UDS results would lead to changes from susceptible to resistant to efavirenz (EFV) in one drug-naive individual with suboptimal response to an EFV-containing regimen and from susceptible to resistance to lamivudine (3TC) in one drug naïve subject who subsequently failed a 3TC-containing regimen and in a treatment experienced subject who had failed a 3TC-containing regimen. Conclusions - The UDS assay performed well across a wide range of subtypes and viral loads; it showed perfect agreement with population sequencing for all RAMs analyzed. In addition, the UDS assay detected additional mutations at frequencies below 20% which correlate with patients’ treatment history and had in some cases important prognostic implications

Age-related reductions in heart rate variability do not worsen during exposure to humid compared to dry heat: A secondary analysis

We conducted a secondary analysis to investigate whether age-related attenuations in heart rate variability (HRV) worsen during exposure to moderate, dry (36.5°C, 20% RH) or humid (36.5°C, 60% RH) heat conditions that resulted in greater body heat storage among older compared to young participants, and during humid compared to dry heat, regardless of age. Six HRV indices [heart rate (HR), coefficient of variation (CoV), detrended fluctuation analysis: α1, low frequency power, high frequency power, and low/high frequency ratio] were assessed in 10 young (21 ± 3 y) and 9 older (65 ± 5 y) adults for 15-min prior to (baseline), and at the end of a 120-min exposure to dry and humid heat while seated at rest. Our results demonstrated a condition (dry and humid) x time (baseline and end) interaction effect on HR (p = 0.047) such that HR gradually increased during humid heat exposure yet remained similar during dry heat exposure across groups. We also found an age-related attenuation in CoV at baseline for both the dry (young: 0.097 ± 0.023%; older: 0.054 ± 0.016%) and humid (young: 0.093 ± 0.034%; older: 0.056 ± 0.014%) heat conditions (p 0.05). While older adults stored more heat during a brief 120-min exposure to dry heat compared to their young counterparts, this was not paralleled by further age-related impairments in HRV even when body heat storage and cardiovascular strain were exacerbated by exposure to humid heat. © 2019, © 2019 Informa UK Limited, trading as Taylor & Francis Group

The utility of genotypic tropism testing in clinical practice

A review of a large number of HIV-1 tropism test requests (n = 1148) performed at a London tertiary referral centre was carried out. The aim was to establish whether these were being performed in line with recommendations from published guidelines and whether this represented the most cost-effective use of these tests in informing prescribing decisions of the CCR5 antagonist drug, Maraviroc. The cost of these assays within the UK was covered by commercial funding until April 2013 which has subsequently been withdrawn. Furthermore, all healthcare settings are under increasing cost constraints and hence establishing the real utility and appropriate use of these tests is of vital importance

The utility of different bioinformatics algorithms for genotypic HIV-1 tropism testing in a large clinical cohort with multiple subtypes

Objectives: HIV-1 tropism needs to be determined before the use of CCR5 antagonist drugs such as maraviroc (MVC), which are ineffective against CXCR4-using HIV-1. This study assessed how different computational methods for predicting tropism from HIV sequence data performed in a large clinical cohort. The value of adding clinical data to these algorithms was also investigated. Design and methods: PCR amplification and sequence analysis of the HIV-1 gp120 V3 loop region was performed on triple replicates of plasma viral RNA or proviral DNA extracted from peripheral blood monocytes (PBMCs) in 242 patients. Coreceptor usage was predicted from V3 sequences using seven bioinformatics interpretation algorithms, combined with clinical data where appropriate. An intention-to-treat approach was employed for exploring outcomes and performance for different viral subtypes was examined. Results: The frequency of R5 predictions varied by 22.6%, with all seven algorithms agreeing for only 75.3% of tests. The identification of individuals likely to fail was poor for all algorithms. The addition of clinical data improved this, but at the expense of their ability to predict success. The clinical algorithms varied across subtypes, whereas other algorithms were more consistent. Furthermore, individuals with discordant clonal and clinical predictions were more likely to fail MVC treatment. Conclusion: Eligibility for MVC varied depending on the algorithm method used. The addition of clinical parameters alongside sequence data may help predict X4 emergence during treatment. It could be that V3 loop analysis in isolation may not be the best method for selecting individuals for MVC