Sensitivity of Next-Generation Sequencing Metagenomic Analysis for Detection of RNA and DNA Viruses in Cerebrospinal Fluid: The Confounding Effect of Background Contamination

  • Iwona Bukowska-Ośko
  • Karol Perlejewski
  • Shota Nakamura
  • Daisuke Motooka
  • Tomasz Stokowy
  • Joanna Kosińska
  • Marta Popiel
  • Rafał Płoski
  • Andrzej Horban
  • Dariusz Lipowski
  • Kamila Caraballo Cortés
  • Agnieszka Pawełczyk
  • Urszula Demkow
  • Adam Stępień
  • Marek Radkowski
  • Tomasz Laskus
Chapter

Abstract

Next-generation sequencing (NGS) followed by metagenomic analysis enables the detection and identification of known as well as novel pathogens. It could be potentially useful in the diagnosis of encephalitis, caused by a variety of microorganisms. The aim of the present study was to evaluate the sensitivity of isothermal RNA amplification (Ribo-SPIA) followed by NGS metagenomic analysis in the detection of human immunodeficiency virus (HIV) and herpes simplex virus (HSV) in cerebrospinal fluid (CSF). Moreover, we analyzed the contamination background. We detected 102 HIV copies and 103 HSV copies. The analysis of control samples (two water samples and one CSF sample from an uninfected patient) revealed the presence of human DNA in the CSF sample (91 % of all reads), while the dominating sequences in water were qualified as ‘other’, related to plants, plant viruses, and synthetic constructs, and constituted 31 % and 60 % of all reads. Bacterial sequences represented 5.9 % and 21.4 % of all reads in water samples and 2.3 % in the control CSF sample. The bacterial sequences corresponded mainly to Psychrobacter, Acinetobacter, and Corynebacterium genera. In conclusion, Ribo-SPIA amplification followed by NGS metagenomic analysis is sensitive for detection of RNA and DNA viruses. Contamination seems common and thus the results should be confirmed by other independent methods such as RT-PCR and PCR. Despite these reservations, NGS seems to be a promising method for the diagnosis of viral infections.

Keywords

Bacteria Cerebrospinal fluid DNA Encephalitis Next-generation sequencing Metagenomics analysis Pathogens Viruses 

References

  1. Barzon L, Lavezzo E, Costanzi G, Franchin E, Toppo S, Palu G (2013) Next-generation sequencing technologies in diagnostic virology. J Clin Virol 58(2):346–350CrossRefPubMedGoogle Scholar
  2. Capobianchi MR, Giombini E, Rozera G (2013) Next-generation sequencing technology in clinical virology. Clin Microbiol Infect 19(1):15–22CrossRefPubMedGoogle Scholar
  3. Chaudhuri A, Kennedy PG (2002) Diagnosis and treatment of viral encephalitis. Postgrad Med J 78(924):575–583CrossRefPubMedPubMedCentralGoogle Scholar
  4. Cheval J, Sauvage V, Frangeul L, Dacheux L, Guigon G, Dumey N, Pariente K, Rousseaux C, Dorange F, Berthet N, Brisse S, Moszer I, Bourhy H, Manuguerra CJ, Lecuit M, Burguiere A, Caro V, Eloit M (2011) Evaluation of high-throughput sequencing for identifying known and unknown viruses in biological samples. J Clin Microbiol 49(9):3268–3275CrossRefPubMedPubMedCentralGoogle Scholar
  5. Chomczynski P, Sacchi N (1987) Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem 162(1):156–159CrossRefPubMedGoogle Scholar
  6. Clement-Ziza M, Gentien D, Lyonnet S, Thiery JP, Besmond C, Decraene C (2009) Evaluation of methods for amplification of picogram amounts of total RNA for whole genome expression profiling. BMC Genomics 10:246CrossRefPubMedPubMedCentralGoogle Scholar
  7. Dickins B, Rebolledo-Jaramillo B, Su MS, Paul IM, Blankenberg D, Stoler N, Makova KD, Nekrutenko A (2014) Controlling for contamination in re-sequencing studies with a reproducible web-based phylogenetic approach. Biotechniques 56(3):134–136, 138–141CrossRefPubMedPubMedCentralGoogle Scholar
  8. Dunne WM, Westblade LF, Ford B (2012) Next-generation and whole-genome sequencing in the diagnostic clinical microbiology laboratory. Eur J Clin Microbiol Infect Dis 31(8):1719–1726CrossRefPubMedGoogle Scholar
  9. Feng H, Shuda M, Chang Y, Moore PS (2008) Clonal integration of a polyomavirus in human Merkel cell carcinoma. Science 319(5866):1096–1100CrossRefPubMedPubMedCentralGoogle Scholar
  10. Frey KG, Herrera-Galeano JE, Redden CL, Luu TV, Servetas SL, Mateczun AJ, Mokashi VP, Bishop-Lilly KA (2014) Comparison of three next-generation sequencing platforms for metagenomic sequencing and identification of pathogens in blood. BMC Genomics 15:96CrossRefPubMedPubMedCentralGoogle Scholar
  11. Glaser CA, Gilliam S, Schnurr D, Forghani B, Honarmand S, Khetsuriani N, Fischer M, Cossen CK, Anderson LJ, California Encephalitis (2003) In search of encephalitis etiologies: diagnostic challenges in the California Encephalitis Project, 1998–2000. Clin Infect Dis 36(6):731–742CrossRefPubMedGoogle Scholar
  12. Glaser CA, Honarmand S, Anderson LJ, Schnurr DP, Forghani B, Cossen CK, Schuster FL, Christie LJ, Tureen JH (2006) Beyond viruses: clinical profiles and etiologies associated with encephalitis. Clin Infect Dis 43(12):1565–1577CrossRefPubMedGoogle Scholar
  13. Granerod J, Crowcroft NS (2007) The epidemiology of acute encephalitis. Neuropsychol Rehabil 17(4–5):406–428CrossRefPubMedGoogle Scholar
  14. Granerod J, Cunningham R, Zuckerman M, Mutton K, Davies NW, Walsh AL, Ward KN, Hilton DA, Ambrose HE, Clewley JP, Morgan D, Lunn MP, Solomon T, Brown DW, Crowcroft NS (2010a) Causality in acute encephalitis: defining aetiologies. Epidemiol Infect 138(6):783–800CrossRefPubMedGoogle Scholar
  15. Granerod J, Tam CC, Crowcroft NS, Davies NW, Borchert M, Thomas SL (2010b) Challenge of the unknown. A systematic review of acute encephalitis in non-outbreak situations. Neurology 75(10):924–932CrossRefPubMedGoogle Scholar
  16. Kircher M, Sawyer S, Meyer M (2012) Double indexing overcomes inaccuracies in multiplex sequencing on the Illumina platform. Nucleic Acids Res 40(1):e3CrossRefPubMedGoogle Scholar
  17. Kurn N, Chen P, Heath JD, Kopf-Sill A, Stephens KM, Wang S (2005) Novel isothermal, linear nucleic acid amplification systems for highly multiplexed applications. Clin Chem 51(10):1973–1981CrossRefPubMedGoogle Scholar
  18. Laurence M, Hatzis C, Brash DE (2014) Common contaminants in next-generation sequencing that hinder discovery of low-abundance microbes. PLoS One 9(5):e97876CrossRefPubMedPubMedCentralGoogle Scholar
  19. Lecuit M, Eloit M (2014) The diagnosis of infectious diseases by whole genome next generation sequencing: a new era is opening. Front Cell Infect Microbiol 4:25CrossRefPubMedPubMedCentralGoogle Scholar
  20. Leek JT, Scharpf RB, Bravo HC, Simcha D, Langmead B, Johnson WE, Geman D, Baggerly K, Irizarry RA (2010) Tackling the widespread and critical impact of batch effects in high-throughput data. Nat Rev Genet 11(10):733–739CrossRefPubMedGoogle Scholar
  21. Lusk RW (2014) Diverse and widespread contamination evident in the unmapped depths of high throughput sequencing data. PLoS One 9(10):e110808CrossRefPubMedPubMedCentralGoogle Scholar
  22. Malboeuf CM, Yang X, Charlebois P, Qu J, Berlin AM, Casali M, Pesko KN, Boutwell CL, DeVincenzo JP, Ebel GD, Allen TM, Zody MC, Henn MR, Levin JZ (2013) Complete viral RNA genome sequencing of ultra-low copy samples by sequence-independent amplification. Nucleic Acids Res 41(1):e13CrossRefPubMedGoogle Scholar
  23. Miller RR, Montoya V, Gardy JL, Patrick DM, Tang P (2013) Metagenomics for pathogen detection in public health. Genome Med 5(9):81CrossRefPubMedPubMedCentralGoogle Scholar
  24. Miyamoto M, Motooka D, Gotoh K, Imai T, Yoshitake K, Goto N, Iida T, Yasunaga T, Horii T, Arakawa K, Kasahara M, Nakamura S (2014) Performance comparison of second- and third-generation sequencers using a bacterial genome with two chromosomes. BMC Genomics 15:699CrossRefPubMedPubMedCentralGoogle Scholar
  25. Naccache SN, Federman S, Veeraraghavan N, Zaharia M, Lee D, Samayoa E, Bouquet J, Greninger AL, Luk KC, Enge B, Wadford DA, Messenger SL, Genrich GL, Pellegrino K, Grard G, Leroy E, Schneider BS, Fair JN, Martinez MA, Isa P, Crump JA, DeRisi JL, Sittler T, Hackett J, Miller S, Chiu CY (2014) A cloud-compatible bioinformatics pipeline for ultrarapid pathogen identification from next-generation sequencing of clinical samples. Genome Res 24(7):1180–1192CrossRefPubMedPubMedCentralGoogle Scholar
  26. Nakamura S, Yang CS, Sakon N, Ueda M, Tougan T, Yamashita A, Goto N, Takahashi K, Yasunaga T, Ikuta K, Mizutani T, Okamoto Y, Tagami M, Morita R, Maeda N, Kawai J, Hayashizaki Y, Nagai Y, Horii T, Iida T, Nakaya T (2009) Direct metagenomic detection of viral pathogens in nasal and fecal specimens using an unbiased high-throughput sequencing approach. PLoS One 4(1), e4219CrossRefPubMedPubMedCentralGoogle Scholar
  27. Newsome T, Li BJ, Zou N, Lo SC (2004) Presence of bacterial phage-like DNA sequences in commercial Taq DNA polymerase reagents. J Clin Microbiol 42(5):2264–2267CrossRefPubMedPubMedCentralGoogle Scholar
  28. Palacios G, Druce J, Du L, Tran T, Birch C, Briese T, Conlan S, Quan PL, Hui J, Marshall J, Simons JF, Egholm M, Paddock CD, Shieh WJ, Goldsmith CS, Zaki SR, Catton M, Lipkin WI (2008) A new arenavirus in a cluster of fatal transplant-associated diseases. N Engl J Med 358(10):991–998Google Scholar
  29. Paradowska-Stankiewicz I, Piotrowska A (2014) Meningitis and encephalitis in Poland in 2012. Przegl Epidemiol 68(2):21–218, 333–216Google Scholar
  30. Perlejewski K, Popiel M, Laskus T, Nakamura S, Motooka D, Stokowy T, Lipowski D, Pollak A, Lechowicz U, Caraballo Cortes K, Stepien A, Radkowski M, Bukowska-Osko I (2015) Next-generation sequencing (NGS) in the identification of encephalitis-causing viruses: unexpected detection of human herpesvirus 1 while searching for RNA pathogens. J Virol Methods 226:1–6CrossRefPubMedGoogle Scholar
  31. Quinones-Mateu ME, Avila S, Reyes-Teran G, Martinez MA (2014) Deep sequencing: becoming a critical tool in clinical virology. J Clin Virol 61(1):9–19CrossRefPubMedPubMedCentralGoogle Scholar
  32. Rasool V, Rasool S, Mushtaq S (2014) Viral encephalitis and its management through advanced molecular diagnostic methods: a review. Clin Pediatr (Phila) 53(2):118–120CrossRefGoogle Scholar
  33. Salter SJ, Cox MJ, Turek EM, Calus ST, Cookson WO, Moffatt MF, Turner P, Parkhill J, Loman NJ, Walker AW (2014) Reagent and laboratory contamination can critically impact sequence-based microbiome analyses. BMC Biol 12:87CrossRefPubMedPubMedCentralGoogle Scholar
  34. Shen H, Rogelj S, Kieft TL (2006) Sensitive, real-time PCR detects low-levels of contamination by Legionella pneumophila in commercial reagents. Mol Cell Probes 20(3–4):147–153CrossRefPubMedGoogle Scholar
  35. Silva MT (2013) Viral encephalitis. Arq Neuropsiquiatr 71(9B):703–709CrossRefPubMedGoogle Scholar
  36. Solomon T, Hart IJ, Beeching NJ (2007) Viral encephalitis: a clinician’s guide. Pract Neurol 7(5):288–305CrossRefPubMedGoogle Scholar
  37. Steiner I, Budka H, Chaudhuri A, Koskiniemi M, Sainio K, Salonen O, Kennedy PG (2005) Viral encephalitis: a review of diagnostic methods and guidelines for management. Eur J Neurol 12(5):331–343CrossRefPubMedGoogle Scholar
  38. Strong MJ, Xu G, Morici L, Splinter Bon-Durant S, Baddoo M, Lin Z, Fewell C, Taylor CM, Flemington EK (2014) Microbial contamination in next generation sequencing: implications for sequence-based analysis of clinical samples. PLoS Pathog 10(11):e1004437CrossRefPubMedPubMedCentralGoogle Scholar
  39. Tan le V, van Doorn HR, Nghia HD, Chau TT, le TP T, de Vries M, Canuti M, Deijs M, Jebbink MF, Baker S, Bryant JE, Tham NT NTBK, Boni MF, Loi TQ, Phuong le T, Verhoeven JT, Crusat M, Jeeninga RE, Schultsz C, Chau NV, Hien TT, van der Hoek L, Farrar J, de Jong MD (2013) Identification of a new cyclovirus in cerebrospinal fluid of patients with acute central nervous system infections. MBio 4(3):e00231–e00313PubMedPubMedCentralGoogle Scholar
  40. Towner JS, Sealy TK, Khristova ML, Albarino CG, Conlan S, Reeder SA, Quan PL, Lipkin WI, Downing R, Tappero JW, Okware S, Lutwama J, Bakamutumaho B, Kayiwa J, Comer JA, Rollin PE, Ksiazek TG, Nichol ST (2008) Newly discovered ebola virus associated with hemorrhagic fever outbreak in Uganda. PLoS Pathog 4(11):e1000212CrossRefPubMedPubMedCentralGoogle Scholar
  41. Virgin HW, Todd JA (2011) Metagenomics and personalized medicine. Cell 147(1):44–56CrossRefPubMedPubMedCentralGoogle Scholar
  42. Wilson MR, Naccache SN, Samayoa E, Biagtan M, Bashir H, Yu G, Salamat SM, Somasekar S, Federman S, Miller S, Sokolic R, Garabedian E, Candotti F, Buckley RH, Reed KD, Meyer TL, Seroogy CM, Galloway R, Henderson SL, Gern JE, DeRisi JL, Chiu CY (2014) Actionable diagnosis of neuroleptospirosis by next-generation sequencing. N Engl J Med 370(25):2408–2417CrossRefPubMedPubMedCentralGoogle Scholar
  43. Yozwiak NL, Skewes-Cox P, Stenglein MD, Balmaseda A, Harris E, DeRisi JL (2012) Virus identification in unknown tropical febrile illness cases using deep sequencing. PLoS Negl Trop Dis 6(2):e1485CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer International Publishing Switzerland 2016

Authors and Affiliations

  • Iwona Bukowska-Ośko
    • 1
  • Karol Perlejewski
    • 1
  • Shota Nakamura
    • 2
  • Daisuke Motooka
    • 2
  • Tomasz Stokowy
    • 3
  • Joanna Kosińska
    • 4
  • Marta Popiel
    • 1
  • Rafał Płoski
    • 4
  • Andrzej Horban
    • 5
  • Dariusz Lipowski
    • 5
  • Kamila Caraballo Cortés
    • 1
  • Agnieszka Pawełczyk
    • 1
  • Urszula Demkow
    • 6
  • Adam Stępień
    • 7
  • Marek Radkowski
    • 1
  • Tomasz Laskus
    • 1
  1. 1.Department of Immunopathology of Infectious and Parasitic DiseasesWarsaw Medical UniversityWarsawPoland
  2. 2.Department of Infection Metagenomics, Genome Information Research Center, Research Institute for Microbial DiseasesOsaka UniversitySuitaJapan
  3. 3.Department of Clinical ScienceBergen UniversityBergenNorway
  4. 4.Department of the Medical GeneticsWarsaw Medical UniversityWarsawPoland
  5. 5.Municipal Hospital for Infectious DiseasesWarsawPoland
  6. 6.Department of Laboratory Medicine and Clinical Immunology of Developmental AgeWarsaw Medical UniversityWarsawPoland
  7. 7.Department of NeurologyMilitary Institute of MedicineWarsawPoland

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