1215. Longitudinal Assessment of Genotypic Resistance by Deep Sequencing of Cytomegalovirus DNA in Congenitally Infected Infants Treated with Valganciclovir
Session: Poster Abstract Session: Pediatric CMV and Other Herpes Infections
Saturday, October 22, 2011
Room: Poster Hall B1
Background: Genotypic resistance analysis of cytomegalovirus (CMV) DNA from congenitally infected infants who participated in a phase I/II clinical trial of valganciclovir (VGCV) has previously been reported.  Deep sequencing has been shown to have greater sensitivity in identifying subpopulations not typically detectable by Sanger sequencing.  In order to assess for the possible emergence of minor subpopulations of resistant strains, deep sequencing analysis of sequential specimens obtained before, during and after VGCV treatment was performed on our study population.  

Methods: CMV DNA samples collected over multiple study days from 3 subjects with symptomatic congenital CMV disease who received 6 weeks of antiviral therapy were obtained from the Collaborative Antiviral Study Group Central Lab.  UL97 open reading frames were amplified using a double nested polymerase chain reaction procedure and each resulting amplicon underwent deep sequencing analysis with the Illumina GAIIx Genome Analyzer on a paired end, multiplexed sequencing run.  A lower limit of detection of 5% was used.    

Results: In 1 subject, deep sequencing analysis revealed a subpopulation of ganciclovir-resistant virus containing the A594V mutation known to confer ganciclovir resistance.  This resistant strain comprised 12% of the total CMV population by study day 42 and was associated with a 0.8 log increase in viral load in that study subject in the weeks preceding its appearance.  A K355M mutation, which alters the invariant lysine of the UL97 substrate-binding region and results in a growth restricted kinase-null mutant, was also noted to occur in 1 study subject.  No other populations of known resistance-conferring UL97 mutations were detected, although numerous polymorphisms were noted to arise in each of the 3 study subjects throughout their treatment course. 

Conclusion: The increased sensitivity of deep sequencing genotypic analysis allows for the detection of low-level subpopulations of resistant CMV strains that may arise during the treatment of congenitally infected infants.  Appearance of a resistant subpopulation was associated with a rise in viral load in 1 subject.  Correlation of the presence of resistant CMV subpopulations and clinical outcome is uncertain at this time.

Subject Category: P. Pediatric and perinatal infections

Scott H. James, MD1, David W. Kimberlin, MD1, Caroll Hartline, BS1, Elizabeth Driebe, MS2, James A. Schupp, BS MBA2, David M. Engelthaler, MS2, Paul S. Keim, PhD2,3, Richard Whitley, MD, FIDSA1 and Mark N. Prichard, PhD1, (1)University of Alabama at Birmingham, Birmingham, AL, (2)Translational Genomics Research Institute, Flagstaff, AZ, (3)Northern Arizona University, Flagstaff, AZ


S. H. James, None

D. W. Kimberlin, None

C. Hartline, None

E. Driebe, None

J. A. Schupp, None

D. M. Engelthaler, None

P. S. Keim, None

R. Whitley, None

M. N. Prichard, None

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