Detection of Low Frequency Multi-Drug Resistance and Novel Putative Maribavir Resistance in Immunocompromised Pediatric Patients with Cytomegalovirus.
Human cytomegalovirus (HCMV) is a significant pathogen in immunocompromised individuals, with the potential to cause fatal pneumonitis and colitis, as well as increasing the risk of organ rejection in transplant patients. With the advent of new anti-HCMV drugs there is therefore considerable interest in using virus sequence data to monitor emerging resistance to antiviral drugs in HCMV viraemia and disease, including the identification of putative new mutations. We used target-enrichment to deep sequence HCMV DNA from 11 immunosuppressed pediatric patients receiving single or combination anti-HCMV treatment, serially sampled over 1-27 weeks. Changes in consensus sequence and resistance mutations were analyzed for three ORFs targeted by anti-HCMV drugs and the frequencies of drug resistance mutations monitored. Targeted-enriched sequencing of clinical material detected mutations occurring at frequencies of 2%. Seven patients showed no evidence of drug resistance mutations. Four patients developed drug resistance mutations a mean of 16 weeks after starting treatment. In two patients, multiple resistance mutations accumulated at frequencies of 20% or less, including putative maribavir and ganciclovir resistance mutations P522Q (UL54) and C480F (UL97). In one patient, resistance was detected 14 days earlier than by PCR. Phylogenetic analysis suggested recombination or superinfection in one patient. Deep sequencing of HCMV enriched from clinical samples excluded resistance in 7 of 11 subjects and identified resistance mutations earlier than conventional PCR-based resistance testing in 2 patients. Detection of multiple low level resistance mutations was associated with poor outcome.
Infection, Immunity, Inflammation and Physiological Medicine, Institute of Child Health, University College LondonLondon, UK; Division of Infection and Immunity, University College LondonLondon, UK.
Charlotte J
CJ
Houldcroft
Division of Infection and Immunity, University College London London, UK.
Josephine M
JM
Bryant
Division of Infection and Immunity, University College London London, UK.
Daniel P
DP
Depledge
Infection, Immunity, Inflammation and Physiological Medicine, Institute of Child Health, University College LondonLondon, UK; Centre for Mathematics and Physics in the Life Sciences and Experimental Biology (CoMPLEX), University College LondonLondon, UK.
Ben K
BK
Margetts
Great Ormond Street Hospital for Children NHS Foundation Trust London, UK.
Jacob
J
Simmonds
Division of Infection and Immunity, University College London London, UK.
Stephanos
S
Nicolaou
Division of Infection and Immunity, University College London London, UK.
Helena J
HJ
Tutill
Division of Infection and Immunity, University College London London, UK.
Rachel
R
Williams
Great Ormond Street Hospital for Children NHS Foundation Trust London, UK.
Austen J J
AJ
Worth
Infection, Immunity, Inflammation and Physiological Medicine, Institute of Child Health, University College LondonLondon, UK; Great Ormond Street Hospital for Children NHS Foundation TrustLondon, UK.
Stephen D
SD
Marks
Great Ormond Street Hospital for Children NHS Foundation Trust London, UK.
Paul
P
Veys
Great Ormond Street Hospital for Children NHS Foundation Trust London, UK.
Elizabeth
E
Whittaker
Infection, Immunity, Inflammation and Physiological Medicine, Institute of Child Health, University College LondonLondon, UK; Division of Infection and Immunity, University College LondonLondon, UK; Great Ormond Street Hospital for Children NHS Foundation TrustLondon, UK.
Judith
J
Breuer
antivirals; herpesviruses; immune deficiency; immune suppression; next-generation sequencing
Curr Protoc Bioinformatics. 2002 Aug;Chapter 2:Unit 2.3
18792934
Antimicrob Agents Chemother. 2009 Jan;53(1):81-5
18981262
Comput Struct Biotechnol J. 2015 Feb 10;13:153-9
25750703
J Virol. 2010 Jul;84(14):7195-203
20463084
Antimicrob Agents Chemother. 2014;58(1):274-8
24145545
Antimicrob Agents Chemother. 2011 Sep;55(9):4019-27
21709106
Antimicrob Agents Chemother. 2007 Nov;51(11):4160-2
17709468
Am J Transplant. 2012 Jan;12(1):202-9
21967659
Mol Biol Evol. 2013 Dec;30(12):2725-9
24132122
Antiviral Res. 2011 Nov;92(2):313-8
21906628
Am J Transplant. 2007 Sep;7(9):2106-13
17640310
Virol J. 2008 Mar 27;5:47
18371229
Bone Marrow Transplant. 2013 Jun;48(6):803-8
23178547
Rev Med Virol. 2008 Jul-Aug;18(4):233-46
18383425
J Infect Dis. 2002 Jan 15;185(2):162-9
11807689
Blood. 2009 Jun 4;113(23):5711-9
19299333
Curr Opin Infect Dis. 2015 Aug;28(4):293-9
26098499
J Virol. 2015 May 13;:null
25972543
Antimicrob Agents Chemother. 2014 Aug;58(8):4697-702
24890586
J Clin Virol. 2016 Jul;80:74-81
27214758
J Clin Microbiol. 2013 Nov;51(11):3700-10
23985916
Antivir Ther. 2012;17(1):45-51
22267468
J Infect Dis. 2014 Feb 15;209(4):557-61
23983215
Antimicrob Agents Chemother. 2015 Oct;59(10):6588-93
26259791
Epidemiol Infect. 2014 Sep;142(9):1990-5
24160893
J Virol. 2005 Aug;79(15):9492-502
16014912
PLoS One. 2011;6(11):e27805
22125625
J Clin Virol. 2011 Apr;50(4):287-91
21295516
J Antimicrob Chemother. 2010 Dec;65(12):2628-40
20961907