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Phenotypic diversity of cytomegalovirus DNA polymerase gene variants observed after antiviral therapy.

ABSTRACT: Cytomegalovirus UL54 DNA polymerase mutations observed in clinical specimens are of diagnostic significance if confirmed to affect antiviral drug susceptibility.Validate an updated recombinant phenotyping method to determine the degree of drug resistance conferred by previously uncharacterized UL54 sequence variants, in comparison with known resistance-related mutations.Bacterial artificial chromosome clones of viral DNA were mutagenized by recombination, transfected to produce live virus and phenotyped by standardized reporter-based yield reduction assays.Sixteen recombinant viruses were constructed, representing baseline sequences, known resistance-related mutations and amino acid changes of unproven significance from clinical specimens. Phenotypes of baseline strains and known mutants were comparable to results from prior methods and helped to resolve some published inconsistencies. Mutations F412L, F412S, L545W were newly confirmed to confer ganciclovir and cidofovir resistance, while Q578H conferred ganciclovir and foscarnet resistance with borderline cidofovir resistance. Some foscarnet-resistant mutants were appreciably growth-retarded.Results add to known exonuclease domain mutations that confer ganciclovir-cidofovir cross-resistance, polymerase domain mutations that confer foscarnet resistance with variably decreased ganciclovir/cidofovir susceptibility, and increase the list of sequence variants with no measurable impact on drug susceptibility. The phenotypic diversity of similar UL54 genotypic variants complicates the interpretation of genotypic resistance testing. Technical improvements are facilitating the phenotyping of remaining unknown sequence variants.

SUBMITTER: Chou S

PROVIDER: S-EPMC3059355 | biostudies-literature | 2011 Apr

REPOSITORIES: biostudies-literature

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Phenotypic diversity of cytomegalovirus DNA polymerase gene variants observed after antiviral therapy.

Chou Sunwen S

Journal of clinical virology : the official publication of the Pan American Society for Clinical Virology 20110203 4

<h4>Background</h4>Cytomegalovirus UL54 DNA polymerase mutations observed in clinical specimens are of diagnostic significance if confirmed to affect antiviral drug susceptibility.<h4>Objectives</h4>Validate an updated recombinant phenotyping method to determine the degree of drug resistance conferred by previously uncharacterized UL54 sequence variants, in comparison with known resistance-related mutations.<h4>Study design</h4>Bacterial artificial chromosome clones of viral DNA were mutagenized ...[more]

PMID: 21295516

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Project description:Mitochondria are arising as critical modulators of antiviral tolerance by releasing mtDNA/mtRNA fragments to cytoplasm upon infection, activating virus sensors and type-I interferon response. The relevance of these mechanisms for mitochondrial diseases remains understudied. Here, we investigated mitochondrial recessive ataxia syndrome (MIRAS), caused by a common European founder mutation in DNA polymerase gamma (POLG1). The patients homozygous for the MIRAS variant p.W748S show exceptionally variable ages-of-onset and symptoms, indicating unknown modifying factors contributing to disease manifestation. We report that the mitochondrial DNA (mtDNA) replicase POLG1 has a role in antiviral defence mechanisms to double-stranded DNA and positive-strand RNA virus infections (HSV-1, TBEV, SARS-CoV-2) and its p.W748S variant dampens innate immune responses. Our patient and knock-in mouse data show that p.W748S compromises mtDNA replisome stability, causing mtDNA depletion, aggravated by virus infection. Low mtDNA and mtRNA release to cytoplasm and slow interferon response in MIRAS allow an early replicative advantage for viruses, leading to augmented pro-inflammatory response, subacute loss of GABAergic neurons, liver inflammation and necrosis. A population databank of ~300,000 Finns demonstrates enrichment of immunodeficient traits in p.W748S carriers. Our evidence suggests that POLG1 defects compromise antiviral tolerance, triggering epilepsy and liver disease. The finding has important implications to mitochondrial disease spectrum, including epilepsy, ataxia, and parkinsonism. In this metabolomic dataset: To investigate the in vivo response of MIRAS to viral infection, we generated a MIRAS-knock-in mouse model and subjected the mice to TBEV infection. These animals carry a homozygous knock-in mutation and the accompanying polymorphic variant homologous to the human ancestral MIRAS allele (p.W726S+E1121G in mice and p.W748S+E1143G in humans). We performed metabolomic analyses on the mouse brain (cerebral cortex) isolated from the control (wildtype) and MIRAS mice at 4 days post TBEV infection (n=5 mice per condition).

Project description:BackgroundWe aimed to identify the risk factors associated with colonic cytomegalovirus (CMV) infection in ulcerative colitis (UC) and to compare the clinical course between antiviral therapy-treated and -untreated groups in mucosal CMV-polymerase chain reaction (PCR) -positive cases.MethodsWe retrospectively selected 46 UC patients (>15 years old) in active phase who underwent colonoscopy with biopsy and were analyzed for CMV infection by mucosal PCR between October 2011 and December 2015 at our institution. Colonic CMV in inflamed mucosa was detected using quantitative real-time PCR. The clinical course was evaluated, including need for drug therapy/surgery or drug therapy intensification. In addition, we evaluated the clinical course between CMV-DNA- cases and CMV-DNA+ cases with low viral load.ResultsAt baseline, CMV-DNA+ patients were significantly older, had higher endoscopic scores, and required higher corticosteroid doses during the past 4 weeks than CMV-DNA- patients (p< 0.05). No significant differences were observed in disease duration, disease distribution, laboratory data, or use of other medication between CMV-DNA+ and CMV-DNA- patients. In the anti-CMV-treated group with a median (range) DNA load of 16,000 (9,000-36,400), 3patients achieved remission without additional UC therapy, 2 required additional UC therapy, and 1 required colectomy despite azathioprine and infliximab therapy. In the CMV-untreated group with a median (range) DNA load of 919 (157-5,480), all patients achieved remission with UC therapy alone. No significant difference was observed in the clinical course between CMV-DNA- cases and CMV-DNA+ cases with low viral loads.ConclusionsAging, endoscopic UC activity, and corticosteroid dose predispose to colonic CMV infection, as determined by mucosal PCR, in UC. UC treatment without anti-CMV therapy may be warranted, particularly in patients with low-load CMV-DNA. Anti-CMV therapy alone does not always achieve clinical response in UC even in cases with high-load PCR.

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Phenotypic diversity of cytomegalovirus DNA polymerase gene variants observed after antiviral therapy.

Publications

Phenotypic diversity of cytomegalovirus DNA polymerase gene variants observed after antiviral therapy.

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