Advances in Molecular Technologies and Platforms for the Diagnosis of Infectious Diseases

Abstract:

nfectious microbial pathogens constitute the largest cause of morbidity and mortality worldwide. Early diagnosis and rapid infection control measures can lead to improved outcomes, earlier discharges and reduced nosocomial infections. Conventional diagnostic methods for infectious diseases such as microscopy, culture, and immunological methods, in most cases, are not universally applicable, less sensitive and could take from days to months to complete depending on the pathogen. Molecular assays based on nucleic acids such as polymerase chain reaction (PCR) have improved the sensitivity, specificity and turn-around time in diagnostic microbiology laboratories. These tests are particularly important to detect very low levels of pathogens in clinical samples, and for organisms that have long half-lives or are non-culturable. However, individual molecular tests are available for only a limited number of the more common infectious agents. Moreover, infectious disease events arising from novel pathogens or genetic variants have significantly increased, recently, for which, routine diagnostic methods are not yet available. Therefore, molecular methods and technologies capable of detecting multiple pathogens in a single test have become available over the last few years. Although, these methods are based on the conventional nucleic acid amplification and hybridization chemistry, enhanced multiplexing capability has been achieved through innovations in nucleic acid labeling techniques, and post-amplification analytic methods and instrumentation. The availability of these test kits brought a new level of convenience to the physicians ordering practices, and to the laboratory personnel, as they require very little hands on time. However, these tests are yet unaffordable to many laboratories, and in many cases, the sensitivity is poor compared to that of single-target, real-time PCR assays. Looking into the future, the revolutionary, next generation sequencing (NGS) technology is now being considered as a potential method for rapid identification of hundreds of pathogens, in an unbiased manner, with a single test that could significantly benefit patients who are critically ill with undiagnosed disease.

• Periodical: Advanced Materials Research (Volume 810)
• Pages: 77-125
• DOI: https://doi.org/10.4028/www.scientific.net/AMR.810.77
• Online since: September 2013
• Authors: M. Rubayet Hasan
• Keywords: Diagnosis, DNA Sequencing, Emerging Technologies, Infectious Disease, Mass-Spectrometry, Microarray, Molecular Diagnostics, Multiplex PCR, Next-Generation Sequencing, Nucleic Acid Based Tests (NAT), Nucleic Acid Hybridization, Pathogen , Polymerase Chain Reaction (PCR), Real-Time PCR

References

[1] D. A. Relman, S. Falkow, A Molecular Perspective of Microbial Pathogenicity, in: G.L. Mandell, J.E. Bennett, R. Dolin (Eds.), Principles and practice of infectious diseases, Elsevier, Philadelphia, 2010, pp.3-13.

DOI: 10.1016/b978-0-443-06839-3.00001-1

[2] Information on http://www.who.int/infectious-disease-report/, Accessed June 15, 2013.

[3] W. Levinson, Review of Medical Microbiology and Immunology, 12th ed., McGraw-Hill, New York 2012.

[4] A. Zeimet, D.R. McBride, R. Basilan, W.E. Roland, D. McCrary, H. Koo, Infectious Diseases, in: R.E. Rakel, D.P. Rakel (Eds.), Textbook of Family Medicine, Elsevier Inc., Philadelphia, PA, 2011, pp.207-247.

DOI: 10.1016/b978-1-4377-1160-8.10016-8

[5] A.C. Yu, G. Vatcher, X. Yue, Y. Dong, M.H. Li, P.H. Tam, P.Y. Tsang, A.K. Wong, M. H. Hui, B. Yang, H. Tang, and L.T. Lau, Nucleic acid-based diagnostics for infectious diseases in public health affairs, Front Med. 6 (2012) 173-186.

DOI: 10.1007/s11684-012-0195-5

[6] L.S. Garcia, Clinical Microbiology Procedures Handbook, 3rd ed., ASM Press, Washington, DC 2010.

[7] S.A. Bustin, Kessler, H.H., Amplification and Detection Methods, in: H.H. Kessler (Ed.), Molecular Diagnostics of Infectious Diseases, Walter de Gruyter, Berlin, DEU, 2010, pp.65-78.

DOI: 10.1515/9783110214864.53

[8] J. Versalovic, K.C. Carroll, G. Funke, J.H. Jorgensen, M.L. Landry, D.W. Warnock, Manual of Clinical Microbiology, 10th ed., ASM Press, Washington, DC 2013.

DOI: 10.1128/9781555816728

[9] Information on http://textbookofbacteriology.net/themicrobialworld/medical.html, Accessed June 15, 2013.

[10] G.L. Mandell, J.E. Bennett, R. Dolin, Principles and Practice of Infectious Diseases, 7th ed., Elsevier, Philadelphia 2010.

DOI: 10.1086/655696

[11] J.C. O'Horo, D. Thompson, N. Safdar, Is the gram stain useful in the microbiologic diagnosis of VAP? A meta-analysis, Clin. Infect. Dis. 55 (2012) 551-561.

DOI: 10.1093/cid/cis512

[12] A.A. Faraj, O.D. Omonbude, P. Godwin, Gram staining in the diagnosis of acute septic arthritis, Acta Orthop. Belg. 68 (2002) 388-391.

[13] R.C. Reyes, L. Stoakes, S. Milburn, G. Lennox, J. Daniel, S. N. Silver, E. Wijnker, M.A. John, Z. Hussain, Evaluation of a new chromogenic medium for the detection of methicillin-resistant Staphylococcus aureus carriage on nasal and perianal specimens, Diagn. Microbiol. Infect. Dis. 60 (2008) 225-227.

DOI: 10.1016/j.diagmicrobio.2007.09.011

[14] M.R. Hasan, J.D. Brunstein, G. Al-Rawahi, R. Tan, E. Thomas, P. Tilley, Optimal use of MRSASelect and PCR to maximize sensitivity and specificity of MRSA detection, Curr Microbiol. 66 (2012) 61-63.

DOI: 10.1007/s00284-012-0241-1

[15] C.H. Cho, B. Chulten, C.K. Lee, M.H. Nam, S.Y. Yoon, C.S. Lim, Y. Cho, Y.K. Kim, Evaluation of a novel real-time RT-PCR using TOCE technology compared with culture and Seeplex RV15 for simultaneous detection of respiratory viruses, J. Clin. Virol. 57 (2013) 338-342.

DOI: 10.1016/j.jcv.2013.04.014

[16] J. Heritage, N. Ransome, P.A. Chambers, M. H. Wilcox, A comparison of culture and PCR to determine the prevalence of ampicillin-resistant bacteria in the faecal flora of general practice patients, J. Antimicrob. Chemother 48 (2001) 287-289.

DOI: 10.1093/jac/48.2.287

[17] H. Frickmann, D. Dekker, K. Boahen, S. Acquah, N. Sarpong, Y. Adu-Sarkodie, N.G. Schwarz, J. May, F. Marks, S. Poppert, D.F. Wiemer, R.M. Hagen, Increased detection of invasive enteropathogenic bacteria in pre-incubated blood culture materials by real-time PCR in comparison with automated incubation in Sub-Saharan Africa, Scand. J. Infect. Dis. (2013).

DOI: 10.3109/00365548.2013.777777

[18] R.C. Tilton, Immunologic Methods for Detection of Microbial Antigens, in: W.J.H.J.A. Balows, M. Ohashi, A. Turano, E. H. Lennete (Eds.), Laboratory Diagnosis of Infectious Diseases, Springer, New York, 1988, pp.14-21.

DOI: 10.1007/978-1-4612-3898-0_3

[19] A.M. Schotthoefer, J.K. Meece, L.C. Ivacic, P.D. Bertz, K. Zhang, T. Weiler, T.S. Uphoff, T. R. Fritsche, Comparison of a Real-time PCR Method with Serology and Blood Smear Analysis for Diagnosis of Human Anaplasmosis: Importance of Infection Time Course for Optimal Test Utilization, J. Clin. Microbiol. 51 (2013) 2147-2153.

DOI: 10.1128/jcm.00347-13

[20] K. E. Templeton, S.A. Scheltinga, A.W. Graffelman, J.M. Van Schie, J.W. Crielaard, P. Sillekens, P.J. Van Den Broek, H. Goossens, M.F. Beersma, E. C. Claas, Comparison and evaluation of real-time PCR, real-time nucleic acid sequence-based amplification, conventional PCR, and serology for diagnosis of Mycoplasma pneumoniae, J. Clin. Microbiol. 41 (2003) 4366-4371.

DOI: 10.1128/jcm.41.9.4366-4371.2003

[21] F. Gharabaghi, R. Tellier, R. Cheung, C. Collins, G. Broukhanski, S.J. Drews, S.E. Richardson, Comparison of a commercial qualitative real-time RT-PCR kit with direct immunofluorescence assay (DFA) and cell culture for detection of influenza A and B in children, J. Clin. Virol. 42 (2008) 190-193.

DOI: 10.1016/j.jcv.2008.01.013

[22] K.A. Enan, H. Rennert, A.M. El-Eragi, A.R. El Hussein, I. M. Elkhidir, Comparison of Real-time PCR to ELISA for the detection of human cytomegalovirus infection in renal transplant patients in the Sudan, Virol. J. 8 (2011) 1-4.

DOI: 10.1186/1743-422x-8-222

[23] M.R. Hasan, R. Tan, G.N. Al-Rawahi, E. Thomas, P. Tilley, Short-term stability of pathogen-specific nucleic acid targets in clinical samples, J. Clin. Microbiol. 50 (2012) 4147-4150.

DOI: 10.1128/jcm.02659-12

[24] S. Mattoo, J.D. Cherry, Molecular pathogenesis, epidemiology, and clinical manifestations of respiratory infections due to Bordetella pertussis and other Bordetella subspecies, Clin. Microbiol. Rev. 18 (2005) 326-382.

DOI: 10.1128/cmr.18.2.326-382.2005

[25] D. Raoult, P.E. Fournier, M. Drancourt, What does the future hold for clinical microbiology?, Nat. Rev. Microbiol. 2 (2004) 151-159.

DOI: 10.1038/nrmicro820

[26] CLSI, Molecular Diagnostic Methods for Infectious Diseases; Approved Guideline-Second Edition. MM3-A2., ed., Clinical and Laboratory Standards Institute, Wayne, PA 2006.

[27] S.P. Naber, Molecular pathology--diagnosis of infectious disease, N Engl J Med 331 (1994) 1212-1215.

DOI: 10.1056/nejm199411033311808

[28] K.L. Muldrew, Molecular diagnostics of infectious diseases, Curr. Opin. Pediatr. 21 (2009) 102-111.

[29] K.B. Mullis, The unusual origin of the polymerase chain reaction, Sci. Am. 262 (1990) 56-65.

DOI: 10.1038/scientificamerican0490-56

[30] R.P. Viscidi, R.G. Yolken, Molecular diagnosis of infectious diseases by nucleic acid hybridization, Mol. Cell Probes. 1 (1987) 3-14.

DOI: 10.1016/0890-8508(87)90003-x

[31] R.P. Peters, P.H. Savelkoul, A.M. Simoons-Smit, S.A. Danner, C.M. Vandenbroucke-Grauls, M.A. van Agtmael, Faster identification of pathogens in positive blood cultures by fluorescence in situ hybridization in routine practice, J. Clin. Microbiol. 44 (2006) 119-123.

DOI: 10.1128/jcm.44.1.119-123.2006

[32] J.J. Carrino, H.H. Lee, Nucleic acid amplification methods, J. Microbiol. Methods 23 (1995) 3-20.

[33] L. Garibyan, N. Avashia, Polymerase chain reaction, J. Invest. Dermatol. 133 (2013) e6.

[34] S. Wilhelm, U. Truyen, Real-time reverse transcription polymerase chain reaction assay to detect a broad range of feline calicivirus isolates, J. Virol. Methods 133 (2006) 105-108.

DOI: 10.1016/j.jviromet.2005.10.011

[35] H.M. Welch, The polymerase chain reaction, Methods Mol. Biol. 878 (2012) 71-88.

[36] K.M. Ririe, R.P. Rasmussen, C.T. Wittwer, Product differentiation by analysis of DNA melting curves during the polymerase chain reaction, Anal. Biochem. 245 (1997) 154-160.

DOI: 10.1006/abio.1996.9916

[37] J. Wilhelm, A. Pingoud, Real-time polymerase chain reaction, Chembiochem 4 (2003) 1120-1128.

DOI: 10.1002/cbic.200300662

[38] S.A. Marras, S. Tyagi, F.R. Kramer, Real-time assays with molecular beacons and other fluorescent nucleic acid hybridization probes, Clin. Chim. Acta 363 (2006) 48-60.

DOI: 10.1016/j.cccn.2005.04.037

[39] P. Craw, W. Balachandran, Isothermal nucleic acid amplification technologies for point-of-care diagnostics: a critical review, Lab Chip 12 (2012) 2469-2486.

DOI: 10.1039/c2lc40100b

[40] P. Gill, A. Ghaemi, Nucleic acid isothermal amplification technologies: a review, Nucleosides Nucleotides Nucleic Acids 27 (2008) 224-243.

DOI: 10.1080/15257770701845204

[41] A.E. Men, P. Wilson, K. Siemering, S. Forrest, Sanger DNA Sequencing, in: M. Janitz (Ed.), Next-Generation Genome Sequencing: Towards Personalized Medicine., WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim, 2008.

DOI: 10.1002/9783527625130.ch1

[42] J.M. Janda, S.L. Abbott, 16S rRNA gene sequencing for bacterial identification in the diagnostic laboratory: pluses, perils, and pitfalls, J. Clin. Microbiol. 45 (2007) 2761-2764.

DOI: 10.1128/jcm.01228-07

[43] S. Mignard, J.P. Flandrois, 16S rRNA sequencing in routine bacterial identification: a 30-month experiment, J. Microbiol. Methods 67 (2006) 574-581.

DOI: 10.1016/j.mimet.2006.05.009

[44] S. Persson, J.N. Jensen, K.E. Olsen, Multiplex PCR method for detection of Clostridium difficile tcdA, tcdB, cdtA, and cdtB and internal in-frame deletion of tcdC, J. Clin. Microbiol. 49 (2011) 4299-4300.

DOI: 10.1128/jcm.05161-11

[45] C.G. Fedele, M. Ciardi, S. Delia, J.M. Echevarria, A. Tenorio, Multiplex polymerase chain reaction for the simultaneous detection and typing of polyomavirus JC, BK and SV40 DNA in clinical samples, J. Virol. Methods 82 (1999) 137-144.

DOI: 10.1016/s0166-0934(99)00095-6

[46] J.E. Echevarria, D.D. Erdman, E.M. Swierkosz, B.P. Holloway, L.J. Anderson, Simultaneous detection and identification of human parainfluenza viruses 1, 2, and 3 from clinical samples by multiplex PCR, J. Clin. Microbiol. 36 (1998) 1388-1391.

DOI: 10.1128/jcm.36.5.1388-1391.1998

[47] S.A. Marras, Selection of fluorophore and quencher pairs for fluorescent nucleic acid hybridization probes, Methods Mol. Biol. 335 (2006) 3-16.

DOI: 10.1385/1-59745-069-3:3

[48] Information on http://www.invitrogen.com/site/us/en/home/Products-and-Services/Applications/PCR/real-time-pcr/real-time-pcr-instruments.html, Accessed June 15, 2013.

[49] Information on http://www.roche-applied-science.com/webapp/wcs/stores/servlet/CategoryDisplay?catalogId=10001&tab=&identifier=Real-Time+PCR+Overview&langId=-1&storeId=15016., Accessed June 15, 2013.

[50] Information on http://www.qiagen.com/Products/Catalog/Automated-Solutions/Detection-and-Analysis/Rotor-Gene-Q#orderinginformation, Accessed June 15, 2013.

[51] Information on http://www.bio-rad.com/evportal/en/CA/LSR/Category/059db09c-88a4-44ad-99f8-78635d8d54db/Real-Time-PCR-Detection-Systems, Accessed June 15, 2013.

[52] J.L. Silva, G.G. Leite, G.M. Bastos, B.C. Lucas, D.K. Shinohara, J.S. Takinami, M. Miyata, C.M. Fajardo, A.D. Luchessi, C.Q. Leite, R.F. Cardoso, R.D. Hirata, M.H. Hirata, Plasmid-based controls to detect rpoB mutations in Mycobacterium tuberculosis by quantitative polymerase chain reaction-high-resolution melting, Mem. Inst. Oswaldo Cruz. 108 (2013) 106-109.

DOI: 10.1590/s0074-02762013000100017

[53] Information on http://find.lifetechnologies.com/Global/FileLib/qPCR/RealTimePCR_Handbook_Update_FLR.pdf, Accessed June 15, 2013.

[54] I.M. Mackay, Real-time PCR in the microbiology laboratory, Clin Microbiol Infect 10 (2004) 190-212.

[55] M.L. Choudhary, S.P. Anand, M. Heydari, G. Rane, V.A. Potdar, M.S. Chadha, A.C. Mishra, Development of a multiplex one step RT-PCR that detects eighteen respiratory viruses in clinical specimens and comparison with real time RT-PCR, J Virol Methods 189 (2013) 15-19.

DOI: 10.1016/j.jviromet.2012.12.017

[56] M.J. Loeffelholz, D.L. Pong, R.B. Pyles, Y. Xiong, A.L. Miller, K.K. Bufton, T. Chonmaitree, Comparison of the FilmArray Respiratory Panel and Prodesse real-time PCR assays for detection of respiratory pathogens, J. Clin. Microbiol. 49 (2011) 4083-4088.

DOI: 10.1128/jcm.05010-11

[57] K.A. Thurman, A.K. Warner, K.C. Cowart, A.J. Benitez, J.M. Winchell, Detection of Mycoplasma pneumoniae, Chlamydia pneumoniae, and Legionella spp. in clinical specimens using a single-tube multiplex real-time PCR assay, Diagn. Microbiol. Infect. Dis. 70 (2011) 1-9.

DOI: 10.1016/j.diagmicrobio.2010.11.014

[58] M. Hindiyeh, D.R. Hillyard, K.C. Carroll, Evaluation of the Prodesse Hexaplex multiplex PCR assay for direct detection of seven respiratory viruses in clinical specimens, Am. J. Clin. Pathol. 116 (2001) 218-224.

DOI: 10.1309/f1r7-xd6t-rn09-1u6l

[59] F. Barletta, E.H. Mercado, A. Lluque, J. Ruiz, T.G. Cleary, T.J. Ochoa, Multiplex real-time polymerase chain reaction for the diagnosis of Campylobacter, Salmonella, and Shigella, J. Clin. Microbiol. (2011) Published ahead of print.

DOI: 10.1128/JCM.01397-13

[60] R.R. Higgins, M. Beniprashad, M. Cardona, S. Masney, D.E. Low, J.B. Gubbay, Evaluation and verification of the Seeplex Diarrhea-V ACE assay for simultaneous detection of adenovirus, rotavirus, and norovirus genogroups I and II in clinical stool specimens, J. Clin. Microbiol. 49 (2011) 3154-3162.

DOI: 10.1128/jcm.00599-11

[61] M. Pavlovic, I. Huber, H. Skala, R. Konrad, H. Schmidt, A. Sing, U. Busch, Development of a multiplex real-time polymerase chain reaction for simultaneous detection of enterohemorrhagic Escherichia coli and enteropathogenic Escherichia coli strains, Foodborne Pathog. Dis. 7 (2010) 801-808.

DOI: 10.1089/fpd.2009.0457

[62] Introduction to Flow Cytometry: A Learning Guide, ed., Becton, Dickinson and Company, San Jose, CA 2000.

[63] A.M. Caliendo, Multiplex PCR and emerging technologies for the detection of respiratory pathogens, Clin. Infect. Dis. 52(4) (2011) S326-330.

DOI: 10.1093/cid/cir047

[64] Information on http://www.luminexcorp.com/TechnologiesScience/xMAPTechnology/, Accessed June 15, 2013.

[65] Information on http://www.luminexcorp.com/TechnologiesScience/xTAGTechnology/, Accessed June 15, 2013.

[66] K. Loens, A.M. van Loon, F. Coenjaerts, Y. van Aarle, H. Goossens, P. Wallace, E.J. Claas, M. Ieven, Performance of different mono- and multiplex nucleic acid amplification tests on a multipathogen external quality assessment panel, J. Clin. Microbiol. 50 (2012) 977-987.

DOI: 10.1128/jcm.00200-11

[67] K. Pabbaraju, K.L. Tokaryk, S. Wong, J.D. Fox, Comparison of the Luminex xTAG respiratory viral panel with in-house nucleic acid amplification tests for diagnosis of respiratory virus infections, J. Clin. Microbiol. 46 (2008) 3056-3062.

DOI: 10.1128/jcm.00878-08

[68] J.M. Balada-Llasat, H. LaRue, C. Kelly, L. Rigali, P. Pancholi, Evaluation of commercial ResPlex II v2.0, MultiCode-PLx, and xTAG respiratory viral panels for the diagnosis of respiratory viral infections in adults, J. Clin. Virol. 50 (2011) 42-45.

DOI: 10.1016/j.jcv.2010.09.022

[69] K. Pabbaraju, S. Wong, K.L. Tokaryk, K. Fonseca, and S.J. Drews, Comparison of the Luminex xTAG respiratory viral panel with xTAG respiratory viral panel fast for diagnosis of respiratory virus infections, J. Clin. Microbiol. 49 (2011) 1738-1744.

DOI: 10.1128/jcm.02090-10

[70] E. Claas, C.A. Burnham, T. Mazulli, K. Templeton, and F. Topin, Performance of the xTAG(R) Gastrointestinal Pathogen Panel (GPP), a multiplex molecular assay for simultaneous detection of bacterial, viral and parasitic causes of infectious gastroenteritis, J. Microbiol. Biotechnol. (2013).

DOI: 10.4014/jmb.1212.12042

[71] M. Raymaekers, B. de Rijke, I. Pauli, A.M. Van den Abeele, R. Cartuyvels, Timely diagnosis of respiratory tract infections: evaluation of the performance of the Respifinder assay compared to the xTAG respiratory viral panel assay, J. Clin. Virol. 52 (2011) 314-316.

DOI: 10.1016/j.jcv.2011.08.017

[72] P.F. Wolffs, C. Vink, J. Keijdener, B. Habek, M. Reijans, G. Simons, C.A. Bruggeman, A. J. van den Brule, Evaluation of MeningoFinder, a novel multiplex ligation-dependent probe amplification assay for simultaneous detection of six virus species causing central nervous system infections, J. Clin. Microbiol. 47 (2009) 2620-2622.

DOI: 10.1128/jcm.02436-08

[73] L. Hlousek, S. Voronov, V. Diankov, A.B. Leblang, P.J. Wells, D.M. Ford, J. Nolling, K.W. Hart, P.A. Espinoza, M.R. Bristol, G.J. Tsongalis, B. Yen-Lieberman, V.I. Slepnev, L.I. Kong, M.C. Lee, Automated high multiplex qPCR platform for simultaneous detection and quantification of multiple nucleic acid targets, Biotechniques 52 (2012) 316-324.

DOI: 10.2144/0000113852

[74] Information on http://www.primeradx.com/technology.php, Accessed June 15, 2013.

[75] E.B. Popowitch, S.S. O'Neill, M.B. Miller, Comparison of the Biofire FilmArray RP, Genmark eSensor RVP, Luminex xTAG RVPv1, and Luminex xTAG RVP fast multiplex assays for detection of respiratory viruses, J. Clin. Microbiol. 51 (2013) 1528-1533.

DOI: 10.1128/jcm.03368-12

[76] V.M. Pierce, M. Elkan, M. Leet, K.L. McGowan, R.L. Hodinka, Comparison of the Idaho Technology FilmArray system to real-time PCR for detection of respiratory pathogens in children, J. Clin. Microbiol. 50 (2012) 364-371.

DOI: 10.1128/jcm.05996-11

[77] M. Wilm, Principles of electrospray ionization, Mol Cell Proteomics 10 M111 009407.

[78] S. Sauer, Typing of single nucleotide polymorphisms by MALDI mass spectrometry: principles and diagnostic applications, Clin. Chim. Acta 363 (2006) 95-105.

DOI: 10.1016/j.cccn.2005.05.040

[79] H. Oberacher, On the use of different mass spectrometric techniques for characterization of sequence variability in genomic DNA, Anal. Bioanal. Chem. 391 (2008) 135-149.

DOI: 10.1007/s00216-008-1929-8

[80] D.M. Wolk, E.J. Kaleta, V.H. Wysocki, PCR-electrospray ionization mass spectrometry: the potential to change infectious disease diagnostics in clinical and public health laboratories, J. Mol. Diagn. 14 (2012) 295-304.

DOI: 10.1016/j.jmoldx.2012.02.005

[81] L. Murillo, J. Hardick, K. Jeng, C.A. Gaydos, Evaluation of the Pan Influenza detection kit utilizing the PLEX-ID and influenza samples from the 2011 respiratory season, J. Virol. Methods (2013).

DOI: 10.1016/j.jviromet.2013.06.006

[82] Y.W. Tang, K.S. Lowery, A. Valsamakis, V.C. Schaefer, J.D. Chappell, J. White-Abell, C.D. Quinn, H. Li, C.A. Washington, J. Cromwell, C.M. Giamanco, M. Forman, J. Holden, R.E. Rothman, M.L. Parker, E.V. Ortenberg, L. Zhang, Y.L. Lin, C.A. Gaydos, Clinical accuracy of a PLEX-ID flu device for simultaneous detection and identification of influenza viruses A and B, J. Clin. Microbiol. 51 (2012) 40-45.

DOI: 10.1128/jcm.01978-12

[83] S. Cordey, Y. Thomas, P. Suter, L. Kaiser, Pilot Evaluation of RT-PCR/Electrospray Ionization Mass Spectrometry (PLEX-ID/Flu assay) on Influenza-Positive Specimens, Open Virol J 6 (2012) 64-67.

DOI: 10.2174/1874357901206010064

[84] D. Jacob, U. Sauer, R. Housley, C. Washington, K. Sannes-Lowery, D.J. Ecker, R. Sampath, R. Grunow, Rapid and high-throughput detection of highly pathogenic bacteria by Ibis PLEX-ID technology, PLoS One 7 (2012) e39928.

DOI: 10.1371/journal.pone.0039928

[85] P.J. Simner, J.R. Uhl, L. Hall, M.M. Weber, R.C. Walchak, S. Buckwalter, N.L. Wengenack, Broad-Range Direct Detection and Identification of Fungi by Use of the PLEX-ID PCR-Electrospray Ionization Mass Spectrometry (ESI-MS) System, J. Clin. Microbiol. 51 (2013) 1699-1706.

DOI: 10.1128/jcm.03282-12

[86] S.M. Yoo, J.H. Choi, S.Y. Lee, and N.C. Yoo, Applications of DNA microarray in disease diagnostics, J. Microbiol. Biotechnol. 19 (2009) 635-646.

[87] M.B. Miller, Y.W. Tang, Basic concepts of microarrays and potential applications in clinical microbiology, Clin. Microbiol. Rev. 22 (2009) 611-633.

DOI: 10.1128/cmr.00019-09

[88] M.L. Metzker, Sequencing technologies - the next generation, Nat. Rev. Genet. 11 (2010) 31-46.

[89] L. Liu, Y. Li, S. Li, N. Hu, Y. He, R. Pong, D. Lin, L. Lu, M. Law, Comparison of next-generation sequencing systems, J. Biomed. Biotechnol. 2012 (2012).

DOI: 10.1155/2012/251364

[90] T. Tucker, M. Marra, J.M. Friedman, Massively parallel sequencing: the next big thing in genetic medicine, Am. J. Hum. Genet. 85 (2009) 142-154.

DOI: 10.1016/j.ajhg.2009.06.022

[91] J. Shendure, E. Lieberman Aiden, The expanding scope of DNA sequencing, Nat. Biotechnol. 30 (2012) 1084-1094.

DOI: 10.1038/nbt.2421

[92] J. Shendure, H. Ji, Next-generation DNA sequencing, Nat Biotechnol 26 (2008) 1135-1145.

DOI: 10.1038/nbt1486

[93] P. Tang, C. Chiu, Metagenomics for the discovery of novel human viruses, Future Microbiol 5 177-189.

[94] D. Bogaert, B. Keijser, S. Huse, J. Rossen, R. Veenhoven, E. van Gils, J. Bruin, R. Montijn, M. Bonten, E. Sanders, Variability and diversity of nasopharyngeal microbiota in children: a metagenomic analysis, PLoS One 6 (2011) e17035.

DOI: 10.1371/journal.pone.0017035

[95] L. Barzon, E. Lavezzo, V. Militello, S. Toppo, G. Palu, Applications of next-generation sequencing technologies to diagnostic virology, Int. J. Mol. Sci. 12 (2011) 7861-7884.

DOI: 10.3390/ijms12117861

[96] J. Yang, F. Yang, L. Ren, Z. Xiong, Z. Wu, J. Dong, L. Sun, T. Zhang, Y. Hu, J. Du, J. Wang, Q. Jin, Unbiased parallel detection of viral pathogens in clinical samples by use of a metagenomic approach, J. Clin. Microbiol. 49 (2011) 3463-3469.

DOI: 10.1128/jcm.00273-11

[97] S. Nakamura, C.S. Yang, N. Sakon, M. Ueda, T. Tougan, A. Yamashita, N. Goto, K. Takahashi, T. Yasunaga, K. Ikuta, T. Mizutani, Y. Okamoto, M. Tagami, R. Morita, N. Maeda, J. Kawai, Y. Hayashizaki, Y. Nagai, T. Horii, T. Iida, T. Nakaya, Direct metagenomic detection of viral pathogens in nasal and fecal specimens using an unbiased high-throughput sequencing approach, PLoS One 4 (2009) e4219.

DOI: 10.1371/journal.pone.0004219

[98] J.G. Caporaso, C.L. Lauber, W.A. Walters, D. Berg-Lyons, C.A. Lozupone, P.J. Turnbaugh, N. Fierer, R. Knight, Global patterns of 16S rRNA diversity at a depth of millions of sequences per sample, Proc. Natl. Acad. Sci. USA, 108 (2010) 4516-4522.

DOI: 10.1073/pnas.1000080107

[99] S.J. Salipante, D.J. Sengupta, C. Rosenthal, G. Costa, J. Spangler, E.H. Sims, M.A. Jacobs, S.I. Miller, D.R. Hoogestraat, B.T. Cookson, C. McCoy, F.A. Matsen, J. Shendure, C.C. Lee, T.T. Harkins, N.G. Hoffman, Rapid 16S rRNA Next-Generation Sequencing of Polymicrobial Clinical Samples for Diagnosis of Complex Bacterial Infections, PLoS One 8 (2013) e65226.

DOI: 10.1371/journal.pone.0065226

[100] S.D. Boyd, Diagnostic applications of high-throughput DNA sequencing, Annu. Rev. Pathol. 8 (2012) 381-410.

[101] E.M. Burd, Validation of laboratory-developed molecular assays for infectious diseases, Clin. Microbiol. Rev. 23 (2010) 550-576.

DOI: 10.1128/cmr.00074-09