Diagnosis of cytomegalovirus infection from clinical whole genome sequencing

Nanda Ramchandar, Yan Ding, Lauge Farnaes, David Dimmock, Charlotte Hobbs, Stephen F Kingsmore, Matthew Bainbridge, Nanda Ramchandar, Yan Ding, Lauge Farnaes, David Dimmock, Charlotte Hobbs, Stephen F Kingsmore, Matthew Bainbridge

Abstract

Rapid whole genome sequencing (rWGS) of peripheral blood has been used to detect microbial DNA in acute infections. Cytomegalovirus (CMV) is a herpesvirus capable of causing severe disease in neonates and immunocompromised patients. We identified CMV in patients undergoing diagnostic rWGS by matching reads that did not align to the human reference genome to a database of microbial genomes. rWGS was conducted on peripheral blood obtained from ill pediatric patients (age 1 day to 18 years). Reads not aligning to the human genome were analyzed using an in-house pipeline to identify DNA consistent with CMV infection. Of 669 patients who received rWGS from July 2016 through July 2019, we identified 28 patients (4.2%) with reads that aligned to the CMV reference genome. Six of these patients had clinical findings consistent with symptomatic CMV infection. Positive results were highly correlated (R2 > 0.99, p < 0.001) to a CMV-qPCR assay conducted on DNA isolated from whole blood samples. In acutely ill children receiving rWGS for diagnosis of genetic disease, we propose analysis of patient genetic data to identify CMV, which could impact treatment of up to 4% of children in the intensive care unit.

Conflict of interest statement

All authors have completed the ICMJE uniform disclosure form at www.icmje.org/coi_disclosure.pdf and declare: no financial relationships with any organizations that might have an interest in the submitted work in the previous three years and no other relationships or activities that could appear to have influenced the submitted work.

Figures

Figure 1
Figure 1
Log2 CMV read count from rWGS versus viral load from CMV-quantitative PCR assay for 8 samples where rWGS detected CMV DNA and quantitative PCR could quantify viral load. rWGS read results are highly correlated to infectious unit determination by gold-standard quantitative PCR assay.

References

    1. Farnaes L, et al. Rapid whole-genome sequencing decreases infant morbidity and cost of hospitalization. NPJ Genomic Med. 2018;3:10. doi: 10.1038/s41525-018-0049-4.
    1. Clark MM, et al. Diagnosis of genetic diseases in seriously ill children by rapid whole-genome sequencing and automated phenotyping and interpretation. Sci. Transl. Med. 2019;11:eaat6177. doi: 10.1126/scitranslmed.aat6177.
    1. Blauwkamp TA, et al. Analytical and clinical validation of a microbial cell-free DNA sequencing test for infectious disease. Nat. Microbiol. 2019;4:663. doi: 10.1038/s41564-018-0349-6.
    1. Schlaberg R, et al. Viral pathogen detection by metagenomics and Pan-viral group polymerase chain reaction in children with pneumonia lacking identifiable etiology. J. Infect. Dis. 2017;215:1407–1415. doi: 10.1093/infdis/jix148.
    1. Thoendel MJ, et al. Identification of prosthetic joint infection pathogens using a shotgun metagenomics approach. Clin. Infect. Dis. 2018;67:1333–1338. doi: 10.1093/cid/ciy303.
    1. Leggett RM, et al. Rapid MinION profiling of preterm microbiota and antimicrobial-resistant pathogens. Nat. Microbiol. 2019;5:1–13. doi: 10.1038/s41564-019-0626-z.
    1. Davila S, et al. Viral DNAemia and immune suppression in pediatric sepsis. Pediatr. Crit. Care Med J. Soc. Crit. Care Med. World Fed. Pediatr. Intensive Crit. Care Soc. 2018;19:e14–e22.
    1. Ludwig A, Hengel H. Epidemiological impact and disease burden of congenital cytomegalovirus infection in Europe. Eurosurveillance. 2009;14:19140.
    1. Kimberlin DW, et al. Valganciclovir for symptomatic congenital cytomegalovirus disease. N. Engl. J. Med. 2015;372:933–943. doi: 10.1056/NEJMoa1404599.
    1. Li, H. Aligning sequence reads, clone sequences and assembly contigs with BWA-MEM. Q-Bio, ArXiv (2013).
    1. Uchiyama I. MBGD: microbial genome database for comparative analysis. Nucleic Acids Res. 2003;31:58–62. doi: 10.1093/nar/gkg109.
    1. Tarasov A, Vilella AJ, Cuppen E, Nijman IJ, Prins P. Sambamba: fast processing of NGS alignment formats. Bioinformatics. 2015;31:2032. doi: 10.1093/bioinformatics/btv098.
    1. Bokulich NA, et al. Quality-filtering vastly improves diversity estimates from Illumina amplicon sequencing. Nat. Methods. 2013;10:57–59. doi: 10.1038/nmeth.2276.
    1. Boppana SB, et al. Saliva polymerase-chain-reaction assay for cytomegalovirus screening in newborns. N. Engl. J. Med. 2011;364:2111–2118. doi: 10.1056/NEJMoa1006561.
    1. Ross SA, Novak Z, Pati S, Boppana SB. Diagnosis of cytomegalovirus infections. Infect. Disord. Drug Targets. 2011;11:466–474. doi: 10.2174/187152611797636703.
    1. Rawlinson WD, et al. Congenital cytomegalovirus infection in pregnancy and the neonate: consensus recommendations for prevention, diagnosis, and therapy. Lancet Infect. Dis. 2017;17:e177–e188. doi: 10.1016/S1473-3099(17)30143-3.
    1. Lanzieri TM, et al. Hearing loss in children with asymptomatic congenital cytomegalovirus infection. Pediatrics. 2017;139:e20162610. doi: 10.1542/peds.2016-2610.
    1. Fowler KB, et al. A targeted approach for congenital cytomegalovirus screening within newborn hearing screening. Pediatrics. 2017;139:e20162128. doi: 10.1542/peds.2016-2128.
    1. Park AH, et al. A diagnostic paradigm including cytomegalovirus testing for idiopathic pediatric sensorineural hearing loss. The Laryngoscope. 2014;124:2624–2629. doi: 10.1002/lary.24752.
    1. Sanford E, et al. Concomitant diagnosis of immune deficiency and Pseudomonas sepsis in a 19 month old with ecthyma gangrenosum by host whole-genome sequencing. Mol. Case Stud. 2018;4:a003244. doi: 10.1101/mcs.a003244.
    1. Farnaes L, Coufal NG, Spector SA. Vaccine strain varicella infection in an infant with previously undiagnosed perinatal human immunodeficiency type-1 infection. Pediatr. Infect. Dis. J. 2019;38:413–415. doi: 10.1097/INF.0000000000002183.
    1. Walton AH, et al. Reactivation of multiple viruses in patients with sepsis. PLoS ONE. 2014;9:e98819. doi: 10.1371/journal.pone.0098819.
    1. Ng AP, et al. Cytomegalovirus DNAemia and disease: incidence, natural history and management in settings other than allogeneic stem cell transplantation. Haematologica. 2005;90:1672–1679.
    1. Lisboa LF, et al. The clinical utility of whole blood versus plasma cytomegalovirus viral load assays for monitoring therapeutic response. Transplantation. 2011;91:231–236. doi: 10.1097/TP.0b013e3181ff8719.
    1. Chen XY, Hou PF, Bi J, Ying CM. Detection of human cytomegalovirus DNA in various blood components after liver transplantation. Braz. J. Med. Biol. Res. 2014;47:340–344. doi: 10.1590/1414-431X20133353.
    1. Andersen SC, Fachmann MSR, Kiil K, Møller Nielsen E, Hoorfar J. Gene-based pathogen detection: can we use qPCR to predict the outcome of diagnostic metagenomics? Genes. 2017;8:332. doi: 10.3390/genes8110332.

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