Heterogeneity and evolution of thymidine kinase and DNA polymerase mutants of herpes simplex virus type 1 (HSV-1): implications for antiviral therapy.
ABSTRACT Background. Infections caused by acyclovir resistant isolates of herpes simplex virus (HSV) are an emerging concern after hematopoietic stem cell transplantation (HSCT). Understanding evolutionary aspects of HSV infection is crucial to the design of effective therapeutic and control strategies.Methods. Eight sequential HSV-1 isolates were recovered from a HSCT patient who suffered from recurrent herpetic gingivostomatitis and was treated alternatively with acyclovir, ganciclovir, and foscavir. The diverse spectra and temporal changes of HSV drug resistance were determined phenotypically (drug-resistance profiling) and genotypically (sequencing of the viral thymidine kinase and DNA polymerase genes).Results. Analysis of sixty clones recovered from the different isolates demonstrated that most of these isolates were heterogeneous mixtures of variants indicating the simultaneous infection with different drug-resistant viruses. The phenotype/genotype of several clones associated with resistance to acyclovir and/or foscavir were identified. Two novel mutations (E798 K and I922T) in the viral DNA polymerase could be linked to drug-resistance.Conclusions. The heterogeneity within the viral populations and the temporal changes of drug-resistant viruses found in this HSCT recipient were remarkable, showing a rapid evolution of HSV-1. Drug-resistance surveillance is highly recommended among immunocompromised patients to manage the clinical syndrome and to avoid the emergence of multi-drug resistant isolates.
Article: Antiviral resistance testing[Show abstract] [Hide abstract]
ABSTRACT: Purpose of reviewCurrent genotypic resistance tests fail to amplify drug-resistant minority variants when they are present below 20% of the total virus population. Next-generation sequencing (NGS) addresses this issue and is being introduced into diagnostic laboratories. This review gives an overview of the resistance tests currently used and explores the opportunities and challenges that NGS genotypic resistance tests will bring.Recent findingsThe technical challenges of NGS, such as PCR and sequence-related errors, are being addressed and various assays are currently undergoing technical validation for clinical use. Although not conclusive, the data seem to suggest that NGS will be valuable for low genetic barrier drugs and certain types of tests such as the HIV-1 tropism test. Clinical validation of the reporting and interpretation of minority variant results are essential when laboratories start reporting these results.SummaryThe first wave of NGS technology is being rolled out in diagnostic laboratories. Antiviral test benefits include increased sensitivity and eventually cheaper antiviral resistance tests. There is a risk that low percentage minority variants may be over interpreted. This could result in antiviral drugs, which may have been effective, being possibly denied to patients if proper clinical validation studies are not performed.Current Opinion in Infectious Diseases 10/2014; 27(6). DOI:10.1097/QCO.0000000000000108 · 5.03 Impact Factor
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ABSTRACT: Real-time PCR is the traditional face of nucleic acid detection in the diagnostic microbiology laboratory and is now generally regarded as robust enough to be widely adopted. Methods based on nucleic acid detection of this type are bringing increased accuracy to diagnosis in areas where culture is difficult and/or expensive, and these methods are often effective partners to other rapid molecular diagnostic tools such as matrix-assisted laser desorption ionisation-time of flight mass spectrometry (MALDI-TOF MS). This change in practice has particularly affected the recognition of viruses and fastidious or antibiotic-exposed bacteria, but has been also shown to be effective in the recognition of troublesome or specialised phenotypes such as antiviral resistance and transmissible antibiotic resistance in the Enterobacteriaceae. Quantitation and high-intensity sequencing (of multiple whole genomes) has brought new opportunities as well as new challenges to the microbiology community. Diagnostic microbiologists currently training might be expected to deal less with the culture-based techniques of the last half-century than with the high-volume data and complex analyses of the next.Pathology 02/2015; DOI:10.1097/PAT.0000000000000241 · 2.62 Impact Factor
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ABSTRACT: The seminal discovery of acyclovir 40 years ago heralded the modern era of truly selective antiviral therapies and this drug remains the therapy of choice for herpes simplex virus infections. Yet by modern standards, its antiviral activity is modest and new drugs against novel molecular targets such as the helicase-primase have the potential to improve clinical outcome, particularly in high-risk patients. A brief synopsis of current therapies for these infections and clinical need is provided to help provide an initial perspective. The function of the helicase-primase complex is then summarized and the development of new inhibitors of the helicase-primase complex, such as pritelivir and amenamevir, is discussed. We review their mechanism of action, propensity for drug resistance, and pharmacokinetic characteristics and discuss their potential to advance current therapeutic options. Strategies that include combinations of these inhibitors with acyclovir are also considered, as they will likely maximize clinical efficacy.Clinical Pharmacology & Therapeutics 11/2014; 97(1). DOI:10.1002/cpt.3 · 7.39 Impact Factor