Identification of Pathogens by Nonculturing Molecular Techniques

Reference work entry

Abstract

This chapter will focus on the use of non-culture-based methods to identify prokaryotic human pathogens. The methods discussed rely primarily on nucleic acid detection and analysis for pathogens commonly encountered in diagnostic and clinical research settings. Diagnostic capabilities in clinical microbiology have exponentially grown due to the impact of molecular methods and tools, like (but not limited to) polymerase chain reaction (PCR) and DNA sequencing, for the detection of human pathogens. Increasingly, technologies such as mass spectrometry and next-generation sequencing that have been incorporated into routine research laboratory use for sometime will be implemented for routine use in a clinical setting along with PCR-based assays. In addition, the pace and continued development of molecular methods for clinical applications of pathogen detection may ultimately transform diagnostic microbiology into a “culture less” diagnostic science, in the future.

Keywords

Polymerase Chain Reaction Assay Multiplex Polymerase Chain Reaction Peptide Nucleic Acid Peptide Nucleic Acid Probe Chlamydophila Pneumoniae 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

References

  1. Alanio A, Beretti JL et al (2010) Matrix-assisted laser desorption ionization time-of-flight mass spectrometry for fast and accurate identification of clinically relevant Aspergillus species. Clin Microbiol Infect 17(5):750–755Google Scholar
  2. Anhalt JP, Fenselau C (1975) Identification of bacteria using mass spectrometry. Anal Chem 47:219–225Google Scholar
  3. Babady NE, Stiles J et al (2010) Evaluation of the Cepheid Xpert Clostridium difficile Epi assay for diagnosis of Clostridium difficile infection and typing of the NAP1 strain at a cancer hospital. J Clin Microbiol 48(12):4519–4524PubMedGoogle Scholar
  4. Baldwin CD, Howe GB et al (2009) Usefulness of multilocus polymerase chain reaction followed by electrospray ionization mass spectrometry to identify a diverse panel of bacterial isolates. Diagn Microbiol Infect Dis 63(4):403–408PubMedGoogle Scholar
  5. Battaglia A, Schweighardt AJ et al (2011) Pathogen detection using a liquid array technology. J Forensic Sci 56(3):760–765PubMedGoogle Scholar
  6. Benson R, Tondella ML et al (2008) Development and evaluation of a novel multiplex PCR technology for molecular differential detection of bacterial respiratory disease pathogens. J Clin Microbiol 46(6):2074–2077PubMedGoogle Scholar
  7. Blanc DS, Basset P et al (2011) High proportion of wrongly identified methicillin-resistant Staphylococcus aureus carriers by use of a rapid commercial PCR assay due to presence of staphylococcal cassette chromosome element lacking the mecA gene. J Clin Microbiol 49(2):722–724PubMedGoogle Scholar
  8. Bourdon N, Berenger R et al (2010) Rapid detection of vancomycin-resistant enterococci from rectal swabs by the Cepheid Xpert vanA/vanB assay. Diagn Microbiol Infect Dis 67(3):291–293PubMedGoogle Scholar
  9. Bravo LT, Procop GW (2009) Recent advances in diagnostic microbiology. Semin Hematol 46(3):248–258PubMedGoogle Scholar
  10. Brenwald NP, Baker N et al (2010) Feasibility study of a real-time PCR test for methicillin-resistant Staphylococcus aureus in a point of care setting. J Hosp Infect 74(3):245–249PubMedGoogle Scholar
  11. Brunstein JD, Cline CL et al (2008) Evidence from multiplex molecular assays for complex multipathogen interactions in acute respiratory infections. J Clin Microbiol 46(1):97–102PubMedGoogle Scholar
  12. Chapin K, Musgnug M (2003) Evaluation of three rapid methods for the direct identification of Staphylococcus aureus from positive blood cultures. J Clin Microbiol 41(9):4324–4327PubMedGoogle Scholar
  13. Cherkaoui A, Hibbs J et al (2010) Comparison of two matrix-assisted laser desorption ionization-time of flight mass spectrometry methods with conventional phenotypic identification for routine identification of bacteria to the species level. J Clin Microbiol 48(4):1169–1175PubMedGoogle Scholar
  14. Ciardo DE, Burger S et al (2010) GeneXpert captures unstable methicillin-resistant Staphylococcus aureus prone to rapidly losing the mecA gene. J Clin Microbiol 48(8):3030–3032PubMedGoogle Scholar
  15. Claydon MA, Davey SN et al (1996) The rapid identification of intact microorganisms using mass spectrometry. Nat Biotechnol 14(11):1584–1586PubMedGoogle Scholar
  16. Crowder CD, Matthews HE et al (2010) Genotypic variation and mixtures of Lyme Borrelia in Ixodes ticks from North America and Europe. PLoS One 5(5):e10650PubMedGoogle Scholar
  17. Dekeyser S, Beclin E et al (2011) Implementation of vanA and vanB genes by PCR technique research interest in system (Xpert vanA/vanB CepheidR) closed in a laboratory of microbiology in managing an outbreak to Enterococcus faecium resistant glycopeptide (EfRG). Pathol Biol (Paris) 59(2):73–78Google Scholar
  18. Demirev PA, Fenselau C (2008) Mass spectrometry in biodefense. J Mass Spectrom 43:1441–1457PubMedGoogle Scholar
  19. Dunbar SA (2006) Applications of Luminex xMAP technology for rapid, high-throughput multiplexed nucleic acid detection. Clin Chim Acta 363(1–2):71–82PubMedGoogle Scholar
  20. Dunbar SA, Jacobson JW (2007a) Parallel processing in microbiology: detection of infectious pathogens by Luminex xMAP multiplexed suspension array technology. Clin Microbiol Newslett 29(11):79–86Google Scholar
  21. Dunbar SA, Jacobson JW (2007b) Quantitative, multiplexed detection of Salmonella and other pathogens by Luminex xMAP suspension array. Methods Mol Biol 394:1–19PubMedGoogle Scholar
  22. Ecker DJ, Sampath R et al (2008) Ibis T5000: a universal biosensor approach for microbiology. Nat Rev Microbiol 6(7):553–558PubMedGoogle Scholar
  23. Ecker DJ, Massire C et al (2009) Molecular genotyping of microbes by multilocus PCR and mass spectrometry: a new tool for hospital infection control and public health surveillance. Methods Mol Biol 551:71–87PubMedGoogle Scholar
  24. Emmadi R, Boonyaratanakornkit JB et al (2011) Molecular methods and platforms for infectious diseases testing a review of FDA-approved and cleared assays. J Mol Diagn 13(6):583–604PubMedGoogle Scholar
  25. Emonet S, Shah HN et al (2010) Application and use of various mass spectrometry methods in clinical microbiology. Clin Microbiol Infect 16(11):1604–1613PubMedGoogle Scholar
  26. Endimiani A, Hujer KM et al (2010) Rapid identification of bla KPC-possessing Enterobacteriaceae by PCR/electrospray ionization-mass spectrometry. J Antimicrob Chemother 65(8):1833–1834PubMedGoogle Scholar
  27. Endimiani A, Hujer KM et al (2011) Are we ready for novel detection methods to treat respiratory pathogens in hospital-acquired pneumonia? Clin Infect Dis 52(Suppl 4):S373–S383PubMedGoogle Scholar
  28. Eshoo MW, Crowder CD et al (2010) Detection and identification of Ehrlichia species in blood by use of PCR and electrospray ionization mass spectrometry. J Clin Microbiol 48(2):472–478PubMedGoogle Scholar
  29. Espy MJ, Uhl JR et al (2002) Detection of vaccinia virus, herpes simplex virus, varicella-zoster virus, and Bacillus anthracis DNA by LightCycler polymerase chain reaction after autoclaving: implications for biosafety of bioterrorism agents. Mayo Clin Proc 77(7):624–628PubMedGoogle Scholar
  30. Espy MJ, Uhl JR et al (2006) Real-time PCR in clinical microbiology: applications for routine laboratory testing. Clin Microbiol Rev 19(1):165–256PubMedGoogle Scholar
  31. Evans CA (2011) GeneXpert–a game-changer for tuberculosis control? PLoS Med 8(7):e1001064PubMedGoogle Scholar
  32. Fenselau C, Demirev PA (2001) Characterization of intact microorganisms by MALDI mass spectrometry. Mass Spectrom Rev 20(4):157–171PubMedGoogle Scholar
  33. Ferreira L, Vega CS et al (2010) Identification of Brucella by MALDI-TOF mass spectrometry fast and reliable identification from agar plates and blood cultures. PLoS One 5(12):e14235PubMedGoogle Scholar
  34. Forrest GN (2007) PNA FISH: present and future impact on patient management. Expert Rev Mol Diagn 7(3):231–236PubMedGoogle Scholar
  35. Forrest GN, Roghmann MC et al (2008) Peptide nucleic acid fluorescent in situ hybridization for hospital-acquired enterococcal bacteremia: delivering earlier effective antimicrobial therapy. Antimicrob Agents Chemother 52(10):3558–3563PubMedGoogle Scholar
  36. Gavino M, Wang E (2007) A comparison of a new rapid real-time polymerase chain reaction system to traditional culture in determining group B streptococcus colonization. Am J Obstet Gynecol 197(4):388–384PubMedGoogle Scholar
  37. Gazin M, Lammens C et al (2011) Evaluation of GeneOhm VanR and Xpert vanA/vanB molecular assays for the rapid detection of vancomycin-resistant enterococci. Eur J Clin Microbiol Infect Dis 31(3):273–276PubMedGoogle Scholar
  38. Graber JH, O'Donnell MJ et al (1998) Advances in DNA diagnostics. Curr Opin Biotechnol 9(1):14–18PubMedGoogle Scholar
  39. Hall TA, Sannes-Lowery KA et al (2009) Base composition profiling of human mitochondrial DNA using polymerase chain reaction and direct automated electrospray ionization mass spectrometry. Anal Chem 81(18):7515–7526PubMedGoogle Scholar
  40. Hanif SN, Eldeen HS et al (2011) GeneXpert(R) MTB/RIF for rapid detection of Mycobacterium tuberculosis in pulmonary and extra-pulmonary samples. Int J Tuberc Lung Dis 15(9):1274–1275PubMedGoogle Scholar
  41. Hannis JC, Manalili SM et al (2008) High-resolution genotyping of Campylobacter species by use of PCR and high-throughput mass spectrometry. J Clin Microbiol 46(4):1220–1225PubMedGoogle Scholar
  42. Hayden RT, Kolbert CP et al (2001a) Characterization of culture-derived spiral bacteria by 16 S ribosomal RNA gene sequence analysis. Diagn Microbiol Infect Dis 39(1):55–59PubMedGoogle Scholar
  43. Hayden RT, Uhl JR et al (2001b) Direct detection of Legionella species from bronchoalveolar lavage and open lung biopsy specimens: comparison of LightCycler PCR, in situ hybridization, direct fluorescence antigen detection, and culture. J Clin Microbiol 39(7):2618–2626PubMedGoogle Scholar
  44. Hensley DM, Tapia R et al (2009) An evaluation of the advandx Staphylococcus aureus/CNS PNA FISH assay. Clin Lab Sci 22(1):30–33PubMedGoogle Scholar
  45. Hill CE (2011) Nucleic acid isolation: overview of sample preparation methods. In: Persing DH, Tenover FC, Tang Y et al (eds) Molecular microbiology: diagnostic principles and practice, vol 2. ASM Press, Washington, DC, pp 119–126Google Scholar
  46. Hillemann D, Rusch-Gerdes S et al (2011) Rapid molecular detection of extrapulmonary tuberculosis by the automated GeneXpert MTB/RIF system. J Clin Microbiol 49(4):1202–1205PubMedGoogle Scholar
  47. Hofstadler SA, Sampath R et al (2005) TIGER: the universal biosensor. Int J Mass Spectrom 242:23–41Google Scholar
  48. Holland RD, Wilkes JG et al (1996) Rapid identification of intact whole bacteria based on spectral patterns using matrix-assisted laser desorption/ionization with time-of-flight mass spectrometry. Rapid Commun Mass Spectrom 10:1227–1232PubMedGoogle Scholar
  49. Ioannidis P, Papaventsis D et al (2011) Cepheid GeneXpert MTB/RIF assay for Mycobacterium tuberculosis detection and rifampin resistance identification in patients with substantial clinical indications of tuberculosis and smear-negative microscopy results. J Clin Microbiol 49(8):3068–3070PubMedGoogle Scholar
  50. Jordan JA, Butchko AR et al (2005) Use of pyrosequencing of 16 S rRNA fragments to differentiate between bacteria responsible for neonatal sepsis. J Mol Diagn 7(1):105–110PubMedGoogle Scholar
  51. Jordan JA, Jones-Laughner J et al (2009) Utility of pyrosequencing in identifying bacteria directly from positive blood culture bottles. J Clin Microbiol 47(2):368–372PubMedGoogle Scholar
  52. Kaleta EJ, Clark AE et al (2011) Use of polymerase chain reaction coupled to electrospray ionization mass spectrometry for rapid identification of bacteria and yeast bloodstream pathogens from blood culture bottles. J Clin Microbiol 49(1):345–353PubMedGoogle Scholar
  53. Kato H, Arakawa Y (2011) Use of the loop-mediated isothermal amplification method for identification of PCR ribotype 027 Clostridium difficile. J Med Microbiol 60(Pt 8):1126–1130PubMedGoogle Scholar
  54. Knepp JH, Geahr MA et al (2003) Comparison of automated and manual nucleic acid extraction methods for detection of enterovirus RNA. J Clin Microbiol 41(8):3532–3536PubMedGoogle Scholar
  55. Kwoh DY, Davis GR et al (1989) Transcription-based amplification system and detection of amplified human immunodeficiency virus type 1 with a bead-based sandwich hybridization format. Proc Natl Acad Sci USA 86(4):1173–1177PubMedGoogle Scholar
  56. Lalande V, Barrault L et al (2011) Evaluation of a loop-mediated isothermal amplification assay for diagnosis of Clostridium difficile infections. J Clin Microbiol 49(7):2714–2716PubMedGoogle Scholar
  57. Lanotte P, Plouzeau C et al (2011) Evaluation of four commercial real-time PCR assays for detection of Bordetella spp. in nasopharyngeal aspirates. J Clin Microbiol 49(11):3943–3946PubMedGoogle Scholar
  58. Laudat P, Demondion E et al (2011) Detection of Staphylococcus aureus resistant to methicillin (MRSA) by molecular biology (Cepheid GeneXpert IL, GeneOhm BD, Roche LightCycler, Hyplex Evigene I2A) versus screening by culture: economic and practical strategy for the laboratory. Pathol Biol (Paris) 60(3):208–213Google Scholar
  59. Lawn SD (2011) Pre-screening with GeneXpert(R) MTB/RIF may increase use of isoniazid preventive therapy in antiretroviral programmes. Int J Tuberc Lung Dis 15(9):1272–1273PubMedGoogle Scholar
  60. Lay JO Jr (2001) MALDI-TOF mass spectrometry of bacteria. Mass Spectrom Rev 20(4):172–194PubMedGoogle Scholar
  61. Loeffelholz MJ, Pong DL et al (2011) Comparison of the FilmArray respiratory panel and prodesse real-time PCR assays for detection of respiratory pathogens. J Clin Microbiol 49(12):4083–4088PubMedGoogle Scholar
  62. Maier T, Kostrzewa M (2007) Fast and reliable MALDI-TOF MS-based microorganism identification. Chemistry Today 25:68–71Google Scholar
  63. Marklein G, Josten M et al (2009) Matrix-assisted laser desorption ionization-time of flight mass spectrometry for fast and reliable identification of clinical yeast isolates. J Clin Microbiol 47(9):2912–2917PubMedGoogle Scholar
  64. Marlowe EM, Wolk DM (2006) Pathogen detection in the genomic era. In: Tang YW, Stratton CW (eds) Advanced techniques in diagnostic microbiology. Springer, New YorkGoogle Scholar
  65. Marlowe EM, Novak-Weekley SM et al (2011) Evaluation of the Cepheid Xpert MTB/RIF assay for direct detection of Mycobacterium tuberculosis complex in respiratory specimens. J Clin Microbiol 49(4):1621–1623PubMedGoogle Scholar
  66. Marner ES, Wolk DM et al (2011) Diagnostic accuracy of the Cepheid GeneXpert vanA/vanB assay ver. 1.0 to detect the vanA and vanB vancomycin resistance genes in Enterococcus from perianal specimens. Diagn Microbiol Infect Dis 69(4):382–389PubMedGoogle Scholar
  67. Massire C, Ivy CA et al (2011) Simultaneous identification of mycobacterial isolates to the species level and determination of tuberculosis drug resistance by PCR followed by electrospray ionization mass spectrometry. J Clin Microbiol 49(3):908–917PubMedGoogle Scholar
  68. Miller MB, Popowitch EB et al (2011) Performance of Xpert MTB/RIF RUO assay and IS6110 real-time PCR for Mycobacterium tuberculosis detection in clinical samples. J Clin Microbiol 49(10):3458–3462PubMedGoogle Scholar
  69. Morgan M, Marlowe E et al (2010) Multicenter evaluation of a new shortened peptide nucleic acid fluorescence in situ hybridization procedure for species identification of select Gram-negative bacilli from blood cultures. J Clin Microbiol 48(6):2268–2270PubMedGoogle Scholar
  70. Morgan MA, Marlowe E et al (2011) A 1.5 hour procedure for identification of Enterococcus Species directly from blood cultures. J Vis Exp (48):e2616, doi:10.3791/2616Google Scholar
  71. Mori Y, Nagamine K et al (2001) Detection of loop-mediated isothermal amplification reaction by turbidity derived from magnesium pyrophosphate formation. Biochem Biophys Res Commun 289(1):150–154PubMedGoogle Scholar
  72. Moure R, Martin R et al (2011a) Effectiveness of an integrated real-time PCR method for the detection of Mycobacterium tuberculosis complex in smear-negative extrapulmonary samples in a low prevalence area. J Clin Microbiol 50(2):513–515PubMedGoogle Scholar
  73. Moure R, Munoz L et al (2011b) Rapid detection of Mycobacterium tuberculosis complex and rifampin resistance in smear-negative clinical samples by use of an integrated real-time PCR method. J Clin Microbiol 49(3):1137–1139PubMedGoogle Scholar
  74. Moussaoui W, Jaulhac B et al (2010) Matrix-assisted laser desorption ionization time-of-flight mass spectrometry identifies 90 % of bacteria directly from blood culture vials. Clin Microbiol Infect 16(11):1631–1638PubMedGoogle Scholar
  75. Muresu R, Rubino S et al (1994) A new method for identification of Trichomonas vaginalis by fluorescent DNA in situ hybridization. J Clin Microbiol 32(4):1018–1022PubMedGoogle Scholar
  76. Nagamine K, Watanabe K et al (2001) Loop-mediated isothermal amplification reaction using a nondenatured template. Clin Chem 47(9):1742–1743PubMedGoogle Scholar
  77. Noren T, Alriksson I et al (2011) Rapid and sensitive loop-mediated isothermal amplification test for Clostridium difficile detection challenges cytotoxin B cell test and culture as gold standard. J Clin Microbiol 49(2):710–711PubMedGoogle Scholar
  78. Notomi T, Okayama H et al (2000) Loop-mediated isothermal amplification of DNA. Nucleic Acids Res 28(12):E63PubMedGoogle Scholar
  79. Novak-Weekley SM, Marlowe EM et al (2010) Clostridium difficile testing in the clinical laboratory by use of multiple testing algorithms. J Clin Microbiol 48(3):889–893PubMedGoogle Scholar
  80. Parta M, Goebel M et al (2009) Identification of methicillin-resistant or methicillin-susceptible Staphylococcus aureus in blood cultures and wound swabs by GeneXpert. J Clin Microbiol 47(5):1609–1610PubMedGoogle Scholar
  81. Pierce VM, Elkan M et al (2011) Comparison of the Idaho Technology FilmArray system to real-time PCR for detection of respiratory pathogens in children. J Clin Microbiol 50(2):364–371PubMedGoogle Scholar
  82. Poritz MA, Blaschke AJ et al (2011) FilmArray, an automated nested multiplex PCR system for multi-pathogen detection: development and application to respiratory tract infection. PLoS One 6(10):e26047PubMedGoogle Scholar
  83. Prod'hom G, Bizzini A et al (2010) Matrix-assisted laser desorption ionization-time of flight mass spectrometry for direct bacterial identification from positive blood culture pellets. J Clin Microbiol 48(4):1481–1483PubMedGoogle Scholar
  84. Quiles-Melero I, Garcia-Rodriguez J et al (2011) Rapid identification of yeasts from positive blood culture bottles by pyrosequencing. Eur J Clin Microbiol Infect Dis 30(1):21–24PubMedGoogle Scholar
  85. Rand KH, Rampersaud H et al (2011) Comparison of two multiplex methods for detection of respiratory viruses: FilmArray RP and xTAG RVP. J Clin Microbiol 49(7):2449–2453PubMedGoogle Scholar
  86. Rossney AS, Herra CM et al (2008) Evaluation of the Xpert methicillin-resistant Staphylococcus aureus (MRSA) assay using the GeneXpert real-time PCR platform for rapid detection of MRSA from screening specimens. J Clin Microbiol 46(10):3285–3290PubMedGoogle Scholar
  87. Saffert RT, Cunningham SA et al (2011) Comparison of Bruker Biotyper matrix-assisted laser desorption ionization-time of flight mass spectrometer to BD Phoenix automated microbiology system for identification of gram-negative bacilli. J Clin Microbiol 49(3):887–892PubMedGoogle Scholar
  88. Scanvic A, Courdavault L et al (2011) Interest of real-time PCR Xpert MRSA/SA on GeneXpert((R)) DX System in the investigation of staphylococcal bacteremia. Pathol Biol (Paris) 59(2):67–72Google Scholar
  89. Shore AC, Rossney AS et al (2008) Detection of staphylococcal cassette chromosome mec-associated DNA segments in multiresistant methicillin-susceptible Staphylococcus aureus (MSSA) and identification of Staphylococcus epidermidis ccrAB4 in both methicillin-resistant S. aureus and MSSA. Antimicrob Agents Chemother 52(12):4407–4419PubMedGoogle Scholar
  90. Shore AC, Deasy EC et al (2011) Detection of staphylococcal cassette chromosome mec type XI carrying highly divergent mecA, mecI, mecR1, blaZ, and ccr genes in human clinical isolates of clonal complex 130 methicillin-resistant Staphylococcus aureus. Antimicrob Agents Chemother 55(8):3765–3773PubMedGoogle Scholar
  91. Spencer DH, Sellenriek P et al (2011) Validation and implementation of the GeneXpert MRSA/SA blood culture assay in a pediatric setting. Am J Clin Pathol 136(5):690–694PubMedGoogle Scholar
  92. Stender H (2003) PNA FISH: an intelligent stain for rapid diagnosis of infectious diseases. Expert Rev Mol Diagn 3(5):649–655PubMedGoogle Scholar
  93. Stevenson LG, Drake SK et al (2010) Rapid identification of bacteria in positive blood culture broths by matrix-assisted laser desorption ionization-time of flight mass spectrometry. J Clin Microbiol 48(2):444–447PubMedGoogle Scholar
  94. Tang YW, Kilic A et al (2007) StaphPlex system for rapid and simultaneous identification of antibiotic resistance determinants and Panton-Valentine leukocidin detection of staphylococci from positive blood cultures. J Clin Microbiol 45(6):1867–1873PubMedGoogle Scholar
  95. Taylor LH, Latham SM et al (2001) Risk factors for human disease emergence. Philos Trans R Soc Lond B Biol Sci 356(1411):983–989PubMedGoogle Scholar
  96. Templeton K, Roberts J et al (2001) The detection of Chlamydia trachomatis by DNA amplification methods in urine samples from men with urethritis. Int J STD AIDS 12(12):793–796PubMedGoogle Scholar
  97. Tenover FC, Rasheed JK (2004) Detection of antimicrobial resistance genes and mutations associated with antimicrobial resistance in microorganisms. In: Persing DH, Tenover FC, Versalovic J et al (eds) Molecular microbiology: diagnostic principles and practice. ASM Press, Washington, DC, pp 391–406Google Scholar
  98. Tomita N, Mori Y et al (2008) Loop-mediated isothermal amplification (LAMP) of gene sequences and simple visual detection of products. Nat Protoc 3(5):877–882PubMedGoogle Scholar
  99. Touati A, Benard A et al (2009) Evaluation of five commercial real-time PCR assays for detection of Mycoplasma pneumoniae in respiratory tract specimens. J Clin Microbiol 47(7):2269–2271PubMedGoogle Scholar
  100. Whitehouse CA, Baldwin C et al (2010) Identification of pathogenic Vibrio species by multilocus PCR-electrospray ionization mass spectrometry and its application to aquatic environments of the former soviet republic of Georgia. Appl Environ Microbiol 76(6):1996–2001PubMedGoogle Scholar
  101. Wittwer CT, Kusukawa N (2011) Real-time PCR and melting analysis. In: Persing DH, Tenover FC, Tang Y, Nolte FS, Hayden RT, van Belkim A (eds) Molecular microbiology: diagnostic principle and practice. ASM Press, Washington, DC, pp 63–82Google Scholar
  102. Wolk DM, Dunne WM Jr (2011) New technologies in clinical microbiology. J Clin Microbiol 49(9 Suppl):S62–S67Google Scholar
  103. Wolk DM, Hayden RT (2011) Quantitative molecular methods. In: Persing DH, Tenover FC, Tang Y et al (eds) Molecular microbiology: diagnostic principles and practice, vol 2. ASM Press, Washington,D.C, pp 83–106Google Scholar
  104. Wolk D, Mitchell S et al (2001) Principles of molecular microbiology testing methods. Infect Dis Clin North Am 15(4):1157–1204PubMedGoogle Scholar
  105. Wolk DM, Blyn LB et al (2009a) Pathogen profiling: rapid molecular characterization of Staphylococcus aureus by PCR/electrospray ionization-mass spectrometry and correlation with phenotype. J Clin Microbiol 47(10):3129–3137PubMedGoogle Scholar
  106. Wolk DM, Marx JL et al (2009b) Comparison of MRSASelect Agar, CHROMagar Methicillin-Resistant Staphylococcus aureus (MRSA) Medium, and Xpert MRSA PCR for detection of MRSA in Nares: diagnostic accuracy for surveillance samples with various bacterial densities. J Clin Microbiol 47(12):3933–3936PubMedGoogle Scholar
  107. Wolk DM, Picton E et al (2009c) Multicenter evaluation of the Cepheid Xpert methicillin-resistant Staphylococcus aureus (MRSA) test as a rapid screening method for detection of MRSA in nares. J Clin Microbiol 47(3):758–764PubMedGoogle Scholar
  108. Wolk DM, Struelens MJ et al (2009d) Rapid detection of Staphylococcus aureus and methicillin-resistant S. aureus (MRSA) in wound specimens and blood cultures: multicenter preclinical evaluation of the Cepheid Xpert MRSA/SA skin and soft tissue and blood culture assays. J Clin Microbiol 47(3):823–826PubMedGoogle Scholar
  109. Zabicka D, Strzelecki J et al (2011) Efficiency of the Cepheid Xpert vanA/vanB assay for screening of colonization with vancomycin-resistant enterococci during hospital outbreak. Antonie Van Leeuwenhoek 101(3):671–675PubMedGoogle Scholar
  110. Zeka AN, Tasbakan S et al (2011) Evaluation of the GeneXpert MTB/RIF assay for rapid diagnosis of tuberculosis and detection of rifampin resistance in pulmonary and extrapulmonary specimens. J Clin Microbiol 49(12):4138–4141PubMedGoogle Scholar
  111. Zhang H, Parameswaran P et al (2011) Integrating high-throughput pyrosequencing and quantitative real-time PCR to analyze complex microbial communities. Methods Mol Biol 733:107–128PubMedGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2013

Authors and Affiliations

  1. 1.University of Arizona Medical CenterTucsonUSA
  2. 2.Department of PathologyUniversity of Arizona BIO5 InstituteTucsonUSA

Personalised recommendations