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Molecular Identification and Quantification of Human Rhinoviruses in Respiratory Samples

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Rhinoviruses

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1221))

Abstract

PCR-based molecular assays have become standard diagnostic procedures for the identification and quantification of human rhinoviruses (HRVs) and other respiratory pathogens in most, if not all, clinical microbiology laboratories. Molecular assays are significantly more sensitive than traditional culture-based and serological methods. This advantage has led to the recognition that HRV infections are common causes for not only upper airway symptoms but also more severe lower respiratory illnesses. In addition, molecular assays improve turnaround time, can be performed by technicians with ordinary skills, and can easily be automated. This chapter describes two highly sensitive and specific PCR-based methods for identifying and quantifying HRVs. The first is a two-step PCR method for the detection and typing of HRV. The second is a pan-HRV real-time quantitative (q) PCR method for measuring viral loads in respiratory samples.

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References

  1. Turner RB, Couch RB (2007) Rhinoviruses. In: Knipe DM, Howley PM (eds) Fields virology. Lippincott Williams and Wilkins, Philadelphia, PA, pp 895–909

    Google Scholar 

  2. Turner RB, Lee WM (2009) Rhinovirus. In: Richman DD, Whitley RJ, Hayden FG (eds) Clinical virology. ASM, Washington, DC, pp 1063–1082

    Google Scholar 

  3. Rueckert R (1996) Picornaviridae: the viruses and their replication. In: Fields BN, Knipe DM, Howley PM et al (eds) Fields virology, 3rd edn. Lippincott-Raven, Philadelphia, PA, pp 609–654

    Google Scholar 

  4. Busse WW, Lemanske RF Jr, Gern JE (2010) Role of viral respiratory infections in asthma and asthma exacerbations. Lancet 376:826–834

    Article  PubMed  PubMed Central  Google Scholar 

  5. Gern JE (2010) The ABCs of rhinoviruses, wheezing, and asthma. J Virol 84:7418–7426

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  6. Ruuskanen O, Waris M, Ramilo O (2013) New aspects on human rhinovirus infections. Pediatr Infect Dis J 32:553–555

    Article  PubMed  Google Scholar 

  7. Jartti L, Langen H, Soderlund-Venermo M, Vuorinen T, Ruuskanen O, Jartti T (2011) New respiratory viruses and the elderly. Open Respir Med J 5:61–69

    Article  PubMed  PubMed Central  Google Scholar 

  8. Kaiser L, Aubert JD, Pache JC, Deffernez C, Rochat T, Garbino J, Wunderli W, Meylan P, Yerly S, Perrin L et al (2006) Chronic rhinoviral infection in lung transplant recipients. Am J Respir Crit Care Med 174:1392–1399

    Article  PubMed  Google Scholar 

  9. Miller EK, Lu X, Erdman DD, Poehling KA, Zhu Y, Griffin MR, Hartert TV, Anderson LJ, Weinberg GA, Hall CB et al (2007) Rhinovirus-associated hospitalizations in young children. J Infect Dis 195:773–781

    Article  PubMed  Google Scholar 

  10. Peltola V, Waris M, Osterback R, Susi P, Ruuskanen O, Hyypia T (2008) Rhinovirus transmission within families with children: incidence of symptomatic and asymptomatic infections. J Infect Dis 197:382–389

    Article  PubMed  Google Scholar 

  11. Wright PF, Deatly AM, Karron RA, Belshe RB, Shi JR, Gruber WC, Zhu Y, Randolph VB (2007) Comparison of results of detection of rhinovirus by PCR and viral culture in human nasal wash specimens from subjects with and without clinical symptoms of respiratory illness. J Clin Microbiol 45:2126–2129

    Article  PubMed  PubMed Central  Google Scholar 

  12. Lee WM, Lemanske RF Jr, Evans MD, Vang F, Pappas T, Gangnon R, Jackson DJ, Gern JE (2012) Human rhinovirus species and season of infection determine illness severity. Am J Respir Crit Care Med 186:886–891

    Article  PubMed  PubMed Central  Google Scholar 

  13. Horsnell C, Gama RE, Hughes PJ, Stanway G (1995) Molecular relationships between 21 human rhinovirus serotypes. J Gen Virol 76(Pt 10):2549–2555

    Article  PubMed  CAS  Google Scholar 

  14. Kistler AL, Webster DR, Rouskin S, Magrini V, Credle JJ, Schnurr DP, Boushey HA, Mardis ER, Li H, DeRisi JL (2007) Genome-wide diversity and selective pressure in the human rhinovirus. Virol J 4:40

    Article  PubMed  PubMed Central  Google Scholar 

  15. Ledford RM, Patel NR, Demenczuk TM, Watanyar A, Herbertz T, Collett MS, Pevear DC (2004) VP1 sequencing of all human rhinovirus serotypes: insights into genus phylogeny and susceptibility to antiviral capsid-binding compounds. J Virol 78:3663–3674

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  16. Palmenberg AC, Spiro D, Kuzmickas R, Wang S, Djikeng A, Rathe JA, Fraser-Liggett CM, Liggett SB (2009) Sequencing and analyses of all known human rhinovirus genomes reveal structure and evolution. Science 324:55–59

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  17. Savolainen C, Blomqvist S, Mulders MN, Hovi T (2002) Genetic clustering of all 102 human rhinovirus prototype strains: serotype 87 is close to human enterovirus 70. J Gen Virol 83:333–340

    PubMed  CAS  Google Scholar 

  18. Tapparel C, Junier T, Gerlach D, Cordey S, Van Belle S, Perrin L, Zdobnov EM, Kaiser L (2007) New complete genome sequences of human rhinoviruses shed light on their phylogeny and genomic features. BMC Genomics 8:224

    Article  PubMed  PubMed Central  Google Scholar 

  19. Price WH (1956) The isolation of a new virus associated with respiratory clinical disease in humans. Proc Natl Acad Sci USA 42:892–896

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  20. Hamparian VV, Colonno RJ, Cooney MK, Dick EC, Gwaltney JM Jr, Hughes JH, Jordan WS Jr, Kapikian AZ, Mogabgab WJ, Monto A et al (1987) A collaborative report: rhinoviruses–extension of the numbering system from 89 to 100. Virology 159:191–192

    Article  PubMed  CAS  Google Scholar 

  21. McErlean P, Shackelton LA, Lambert SB, Nissen MD, Sloots TP, Mackay IM (2007) Characterisation of a newly identified human rhinovirus, HRV-QPM, discovered in infants with bronchiolitis. J Clin Virol 39:67–75

    Article  PubMed  CAS  Google Scholar 

  22. Lee WM, Kiesner C, Pappas T, Lee I, Grindle K, Jartti T, Jakiela B, Lemanske RF Jr, Shult PA, Gern JE (2007) A diverse group of previously unrecognized human rhinoviruses are common causes of respiratory illnesses in infants. PLoS One 2:e966

    Article  PubMed  PubMed Central  Google Scholar 

  23. Lau SK, Yip CC, Tsoi HW, Lee RA, So LY, Lau YL, Chan KH, Woo PC, Yuen KY (2007) Clinical features and complete genome characterization of a distinct human rhinovirus (HRV) genetic cluster, probably representing a previously undetected HRV species, HRV-C, associated with acute respiratory illness in children. J Clin Microbiol 45:3655–3664

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  24. Kistler A, Avila PC, Rouskin S, Wang D, Ward T, Yagi S, Schnurr D, Ganem D, Derisi JL, Boushey HA (2007) Pan-viral screening of respiratory tract infections in adults with and without asthma reveals unexpected human coronavirus and human rhinovirus diversity. J Infect Dis 196:817–825

    Article  PubMed  CAS  Google Scholar 

  25. Bochkov YA, Palmenberg AC, Lee WM, Rathe JA, Amineva SP, Sun X, Pasic TR, Jarjour NN, Liggett SB, Gern JE (2011) Molecular modeling, organ culture and reverse genetics for a newly identified human rhinovirus C. Nat Med 17:627–632

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  26. Conant RM, Hamparian VV (1968) Rhinoviruses: basis for a numbering system. II. Serologic characterization of prototype strains. J Immunol 100:114–119

    PubMed  CAS  Google Scholar 

  27. Kapikian AZ, Conant RM, Hamparian VV et al (1967) Rhinoviruses: a numbering system. Nature 213:761–762

    Article  Google Scholar 

  28. Lee WM, Grindle K, Pappas T, Marshall D, Moser M, Beaty E, Shult P, Prudent J, Gern J (2007) High-throughput, sensitive, and accurate multiplex PCR-microsphere flow cytometry system for large-scale comprehensive detection of respiratory viruses. J Clin Microbiol 45:2626–2634

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  29. Lemanske RF Jr (2002) The childhood origins of asthma (COAST) study. Pediatr Allergy Immunol 13(Suppl 15):38–43

    Article  PubMed  Google Scholar 

  30. Gern JE (2010) The urban environment and childhood asthma study. J Allergy Clin Immunol 125:545–549

    Article  PubMed  PubMed Central  Google Scholar 

  31. Kloepfer KM, Olenec JP, Lee WM, Liu G, Vrtis RF, Roberg KA, Evans MD, Gangnon RE, Lemanske RF Jr, Gern JE (2012) Increased H1N1 infection rate in children with asthma. Am J Respir Crit Care Med 185:1275–1279

    Article  PubMed  PubMed Central  Google Scholar 

  32. Denlinger LC, Sorkness RL, Lee WM, Evans MD, Wolff MJ, Mathur SK, Crisafi GM, Gaworski KL, Pappas TE, Vrtis RF et al (2011) Lower airway rhinovirus burden and the seasonal risk of asthma exacerbation. Am J Respir Crit Care Med 184:1007–1014

    Article  PubMed  PubMed Central  Google Scholar 

  33. Ireland DC, Kent J, Nicholson KG (1993) Improved detection of rhinoviruses in nasal and throat swabs by seminested RT-PCR. J Med Virol 40:96–101

    Article  PubMed  CAS  Google Scholar 

  34. Don RH, Cox PT, Wainwright BJ, Baker K, Mattick JS (1991) ‘Touchdown’ PCR to circumvent spurious priming during gene amplification. Nucleic Acids Res 19:4008

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  35. Mathur SK, Fichtinger PS, Kelly JT, Lee WM, Gern JE, Jarjour NN (2013) Interaction between allergy and innate immunity: model for eosinophil regulation of epithelial cell interferon expression. Ann Allergy Asthma Immunol 111(25–31):e21

    Google Scholar 

  36. Mifflin TE (2003) Setting up a PCR laboratory. In: Dieffenbach GS, Dveksler GS (eds) PCR primer: a laboratory manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, pp 5–14

    Google Scholar 

  37. Chenna R, Sugawara H, Koike T, Lopez R, Gibson TJ, Higgins DG, Thompson JD (2003) Multiple sequence alignment with the Clustal series of programs. Nucleic Acids Res 31:3497–3500

    Article  PubMed  CAS  PubMed Central  Google Scholar 

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Author information

Authors and Affiliations

  1. Biological Mimetics Inc., 124 Byte Drive, Frederick, MD, 21702, USA

    Wai-Ming Lee

  2. Department of Pediatrics, School of Medicine and Public Health, University of Wisconsin, Madison, WI, USA

    Kris Grindle, Rose Vrtis, Tressa Pappas, Fue Vang & Iris Lee

  3. Department of Pediatrics and Medicine, University of Wisconsin School of Medicine and Public Health, Madison, WI, USA

    James E. Gern

Authors
  1. Wai-Ming Lee
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  2. Kris Grindle
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  3. Rose Vrtis
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  4. Tressa Pappas
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  5. Fue Vang
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  6. Iris Lee
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  7. James E. Gern
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Corresponding author

Correspondence to Wai-Ming Lee .

Editor information

Editors and Affiliations

  1. Department of Biochemistry and Molecular Biology, Faculty of Health, Nursing and Medical Sciences,, Monash University, Melbourne, Victoria, Australia

    David A. Jans

  2. Centre for Research in Therapeutic Solutions, Faculty of Education, Science, Technology and Mathematics, University of Canberra, Canberra, Aust Capital Terr, Australia

    Reena Ghildyal

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Lee, WM. et al. (2015). Molecular Identification and Quantification of Human Rhinoviruses in Respiratory Samples. In: Jans, D., Ghildyal, R. (eds) Rhinoviruses. Methods in Molecular Biology, vol 1221. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1571-2_3

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  • DOI: https://doi.org/10.1007/978-1-4939-1571-2_3

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-1570-5

  • Online ISBN: 978-1-4939-1571-2

  • eBook Packages: Springer Protocols

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