Skip to main content
Top
Published in: European Journal of Clinical Microbiology & Infectious Diseases 1/2017

01-01-2017 | Original Article

Antimicrobial susceptibility testing of Gram-positive and -negative bacterial isolates directly from spiked blood culture media with Raman spectroscopy

Authors: H. E. Dekter, C. C. Orelio, M. C. Morsink, S. Tektas, B. Vis, R. te Witt, W. B. van Leeuwen

Published in: European Journal of Clinical Microbiology & Infectious Diseases | Issue 1/2017

Login to get access

Abstract

Patients suffering from bacterial bloodstream infections have an increased risk of developing systematic inflammatory response syndrome (SIRS), which can result in rapid deterioration of the patients’ health. Diagnostic methods for bacterial identification and antimicrobial susceptibility tests are time-consuming. The aim of this study was to investigate whether Raman spectroscopy would be able to rapidly provide an antimicrobial susceptibility profile from bacteria isolated directly from positive blood cultures. First, bacterial strains (n = 133) were inoculated in tryptic soy broth and incubated in the presence or absence of antibiotics for 5 h. Antimicrobial susceptibility profiles were analyzed by Raman spectroscopy. Subsequently, a selection of strains was isolated from blood cultures and analyzed similarly. VITEK®2 technology and broth dilution were used as the reference methods. Raman spectra from 67 antibiotic-susceptible strains showed discriminatory spectra in the absence or at low concentrations of antibiotics as compared to high antibiotic concentrations. For 66 antibiotic-resistant strains, no antimicrobial effect was observed on the bacterial Raman spectra. Full concordance with VITEK®2 data and broth dilution was obtained for the antibiotic-susceptible strains, 68 % and 98 %, respectively, for the resistant strains. Discriminative antimicrobial susceptibility testing (AST) profiles were obtained for all bacterial strains isolated from blood cultures, resulting in full concordance with the VITEK®2 data. It can be concluded that Raman spectroscopy is able to detect the antimicrobial susceptibility of bacterial species isolated from a positive blood culture bottle within 5 h. Although Raman spectroscopy is cheap and rapid, further optimization is required, to fulfill a great promise for future AST profiling technology development.
Appendix
Available only for authorised users
Literature
1.
go back to reference Balk RA (2000) Severe sepsis and septic shock. Definitions, epidemiology, and clinical manifestations. Crit Care Clin 16:179–192CrossRefPubMed Balk RA (2000) Severe sepsis and septic shock. Definitions, epidemiology, and clinical manifestations. Crit Care Clin 16:179–192CrossRefPubMed
2.
go back to reference Caliendo AM, Gilbert DN, Ginocchio CC, Hanson KE, May L, Quinn TC, Tenover FC, Alland D, Blaschke AJ, Bonomo RA, Carroll KC, Ferraro MJ, Hirschhorn LR, Joseph WP, Karchmer T, MacIntyre AT, Reller LB, Jackson AF; Infectious Diseases Society of America (IDSA) (2013) Better tests, better care: improved diagnostics for infectious diseases. Clin Infect Dis 57(Suppl 3):S139–S170. doi:10.1093/cid/cit578 CrossRefPubMedPubMedCentral Caliendo AM, Gilbert DN, Ginocchio CC, Hanson KE, May L, Quinn TC, Tenover FC, Alland D, Blaschke AJ, Bonomo RA, Carroll KC, Ferraro MJ, Hirschhorn LR, Joseph WP, Karchmer T, MacIntyre AT, Reller LB, Jackson AF; Infectious Diseases Society of America (IDSA) (2013) Better tests, better care: improved diagnostics for infectious diseases. Clin Infect Dis 57(Suppl 3):S139–S170. doi:10.​1093/​cid/​cit578 CrossRefPubMedPubMedCentral
3.
go back to reference Coelho FR, Martins JO (2012) Diagnostic methods in sepsis: the need of speed. Rev Assoc Med Bras (1992) 58:498–504CrossRef Coelho FR, Martins JO (2012) Diagnostic methods in sepsis: the need of speed. Rev Assoc Med Bras (1992) 58:498–504CrossRef
4.
go back to reference Gaieski DF, Mikkelsen ME, Band RA, Pines JM, Massone R, Furia FF, Shofer FS, Goyal M (2010) Impact of time to antibiotics on survival in patients with severe sepsis or septic shock in whom early goal-directed therapy was initiated in the emergency department. Crit Care Med 38:1045–1053. doi:10.1097/CCM.0b013e3181cc4824 CrossRefPubMed Gaieski DF, Mikkelsen ME, Band RA, Pines JM, Massone R, Furia FF, Shofer FS, Goyal M (2010) Impact of time to antibiotics on survival in patients with severe sepsis or septic shock in whom early goal-directed therapy was initiated in the emergency department. Crit Care Med 38:1045–1053. doi:10.​1097/​CCM.​0b013e3181cc4824​ CrossRefPubMed
5.
go back to reference Kumar A, Roberts D, Wood KE, Light B, Parrillo JE, Sharma S, Suppes R, Feinstein D, Zanotti S, Taiberg L, Gurka D, Kumar A, Cheang M (2006) Duration of hypotension before initiation of effective antimicrobial therapy is the critical determinant of survival in human septic shock. Crit Care Med 34:1589–1596. doi:10.1097/01.CCM.0000217961.75225.E9 CrossRefPubMed Kumar A, Roberts D, Wood KE, Light B, Parrillo JE, Sharma S, Suppes R, Feinstein D, Zanotti S, Taiberg L, Gurka D, Kumar A, Cheang M (2006) Duration of hypotension before initiation of effective antimicrobial therapy is the critical determinant of survival in human septic shock. Crit Care Med 34:1589–1596. doi:10.​1097/​01.​CCM.​0000217961.​75225.​E9 CrossRefPubMed
6.
go back to reference Larché J, Azoulay E, Fieux F, Mesnard L, Moreau D, Thiery G, Darmon M, Le Gall J-R, Schlemmer B (2003) Improved survival of critically ill cancer patients with septic shock. Intensive Care Med 29:1688–1695. doi:10.1007/s00134-003-1957-y CrossRefPubMed Larché J, Azoulay E, Fieux F, Mesnard L, Moreau D, Thiery G, Darmon M, Le Gall J-R, Schlemmer B (2003) Improved survival of critically ill cancer patients with septic shock. Intensive Care Med 29:1688–1695. doi:10.​1007/​s00134-003-1957-y CrossRefPubMed
7.
go back to reference Perez KK, Olsen RJ, Musick WL, Cernoch PL, Davis JR, Peterson LE, Musser JM (2014). Integrating rapid diagnostics and antimicrobial stewardship improves outcomes in patients with antibiotic-resistant Gram-negative bacteremia. J Infect 69:216–225. doi:10.1016/j.jinf.2014.05.005 CrossRefPubMed Perez KK, Olsen RJ, Musick WL, Cernoch PL, Davis JR, Peterson LE, Musser JM (2014). Integrating rapid diagnostics and antimicrobial stewardship improves outcomes in patients with antibiotic-resistant Gram-negative bacteremia. J Infect 69:216–225. doi:10.​1016/​j.​jinf.​2014.​05.​005 CrossRefPubMed
8.
go back to reference van Belkum A, Durand G, Peyret M, Chatellier S, Zambardi G, Schrenzel J, Shortridge D, Engelhardt A, Dunne WM Jr (2013) Rapid clinical bacteriology and its future Impact. Ann Lab Med 33:14–27. doi:10.3343/alm.2013.33.1.14 van Belkum A, Durand G, Peyret M, Chatellier S, Zambardi G, Schrenzel J, Shortridge D, Engelhardt A, Dunne WM Jr (2013) Rapid clinical bacteriology and its future Impact. Ann Lab Med 33:14–27. doi:10.​3343/​alm.​2013.​33.​1.​14
9.
go back to reference Huang AM, Newton D, Kunapuli A, Gandhi TN, Washer LL, Isip J, Collins CD, Nagel JL (2013) Impact of rapid organism identification via matrix-assisted laser desorption/ionization time-of-flight combined with antimicrobial stewardship team intervention in adult patients with bacteremia and candidemia. Clin Infect Dis 57:1237–1245. doi:10.1093/cid/cit498 CrossRefPubMed Huang AM, Newton D, Kunapuli A, Gandhi TN, Washer LL, Isip J, Collins CD, Nagel JL (2013) Impact of rapid organism identification via matrix-assisted laser desorption/ionization time-of-flight combined with antimicrobial stewardship team intervention in adult patients with bacteremia and candidemia. Clin Infect Dis 57:1237–1245. doi:10.​1093/​cid/​cit498 CrossRefPubMed
10.
go back to reference Bork JT, Leekha S, Heil EL, Zhao L, Badamas R, Johnson JK (2015) Rapid testing using the Verigene Gram-negative blood culture nucleic acid test in combination with antimicrobial stewardship intervention against Gram-negative bacteremia. Antimicrob Agents Chemother 59:1588–1595. doi:10.1128/AAC.04259-14 CrossRefPubMedPubMedCentral Bork JT, Leekha S, Heil EL, Zhao L, Badamas R, Johnson JK (2015) Rapid testing using the Verigene Gram-negative blood culture nucleic acid test in combination with antimicrobial stewardship intervention against Gram-negative bacteremia. Antimicrob Agents Chemother 59:1588–1595. doi:10.​1128/​AAC.​04259-14 CrossRefPubMedPubMedCentral
11.
go back to reference Clerc O, Prod’hom G, Senn L, Jaton K, Zanetti G, Calandra T, Greub G (2014) Matrix-assisted laser desorption ionization time-of-flight mass spectrometry and PCR-based rapid diagnosis of Staphylococcus aureus bacteraemia. Clin Microbiol Infect 20:355–360. doi:10.1111/1469-0691.12329 CrossRefPubMed Clerc O, Prod’hom G, Senn L, Jaton K, Zanetti G, Calandra T, Greub G (2014) Matrix-assisted laser desorption ionization time-of-flight mass spectrometry and PCR-based rapid diagnosis of Staphylococcus aureus bacteraemia. Clin Microbiol Infect 20:355–360. doi:10.​1111/​1469-0691.​12329 CrossRefPubMed
13.
go back to reference La Scola B (2011) Intact cell MALDI-TOF mass spectrometry-based approaches for the diagnosis of bloodstream infections. Expert Rev Mol Diagn 11:287–298. doi:10.1586/erm.11.12 PubMed La Scola B (2011) Intact cell MALDI-TOF mass spectrometry-based approaches for the diagnosis of bloodstream infections. Expert Rev Mol Diagn 11:287–298. doi:10.​1586/​erm.​11.​12 PubMed
14.
go back to reference Loonen AJM, Jansz AR, Stalpers J, Wolffs PFG, van den Brule AJC (2012) An evaluation of three processing methods and the effect of reduced culture times for faster direct identification of pathogens from BacT/ALERT blood cultures by MALDI-TOF MS. Eur J Clin Microbiol Infect Dis 31:1575–1583. doi:10.1007/s10096-011-1480-y CrossRefPubMed Loonen AJM, Jansz AR, Stalpers J, Wolffs PFG, van den Brule AJC (2012) An evaluation of three processing methods and the effect of reduced culture times for faster direct identification of pathogens from BacT/ALERT blood cultures by MALDI-TOF MS. Eur J Clin Microbiol Infect Dis 31:1575–1583. doi:10.​1007/​s10096-011-1480-y CrossRefPubMed
15.
go back to reference Hamasha K, Mohaidat QI, Putnam RA, Woodman RC, Palchaudhuri S, Rehse SJ (2013) Sensitive and specific discrimination of pathogenic and nonpathogenic Escherichia coli using Raman spectroscopy—a comparison of two multivariate analysis techniques. Biomed Opt Express 4:481–489. doi:10.1364/BOE.4.000481 CrossRefPubMedPubMedCentral Hamasha K, Mohaidat QI, Putnam RA, Woodman RC, Palchaudhuri S, Rehse SJ (2013) Sensitive and specific discrimination of pathogenic and nonpathogenic Escherichia coli using Raman spectroscopy—a comparison of two multivariate analysis techniques. Biomed Opt Express 4:481–489. doi:10.​1364/​BOE.​4.​000481 CrossRefPubMedPubMedCentral
16.
go back to reference Jung JS, Popp C, Sparbier K, Lange C, Kostrzewa M, Schubert S (2014) Evaluation of matrix-assisted laser desorption ionization-time of flight mass spectrometry for rapid detection of β-lactam resistance in Enterobacteriaceae derived from blood cultures. J Clin Microbiol 52:924–930. doi:10.1128/JCM.02691-13 CrossRefPubMedPubMedCentral Jung JS, Popp C, Sparbier K, Lange C, Kostrzewa M, Schubert S (2014) Evaluation of matrix-assisted laser desorption ionization-time of flight mass spectrometry for rapid detection of β-lactam resistance in Enterobacteriaceae derived from blood cultures. J Clin Microbiol 52:924–930. doi:10.​1128/​JCM.​02691-13 CrossRefPubMedPubMedCentral
17.
go back to reference Wimmer JL, Long SW, Cernoch P, Land GA, Davis JR, Musser JM, Olsen RJ (2012) Strategy for rapid identification and antibiotic susceptibility testing of gram-negative bacteria directly recovered from positive blood cultures using the Bruker MALDI Biotyper and the BD Phoenix system. J Clin Microbiol 50:2452–2454. doi:10.1128/JCM.00409-12 CrossRefPubMedPubMedCentral Wimmer JL, Long SW, Cernoch P, Land GA, Davis JR, Musser JM, Olsen RJ (2012) Strategy for rapid identification and antibiotic susceptibility testing of gram-negative bacteria directly recovered from positive blood cultures using the Bruker MALDI Biotyper and the BD Phoenix system. J Clin Microbiol 50:2452–2454. doi:10.​1128/​JCM.​00409-12 CrossRefPubMedPubMedCentral
18.
go back to reference Espagnon I, Ostrovskii D, Mathey R, Dupoy M, Joly PL, Novelli-Rousseau A, Pinston F, Gal O, Mallard F, Leroux DF (2014) Direct identification of clinically relevant bacterial and yeast microcolonies and macrocolonies on solid culture media by Raman spectroscopy. J Biomed Opt 19:027004. doi:10.1117/1.JBO.19.2.027004 CrossRefPubMed Espagnon I, Ostrovskii D, Mathey R, Dupoy M, Joly PL, Novelli-Rousseau A, Pinston F, Gal O, Mallard F, Leroux DF (2014) Direct identification of clinically relevant bacterial and yeast microcolonies and macrocolonies on solid culture media by Raman spectroscopy. J Biomed Opt 19:027004. doi:10.​1117/​1.​JBO.​19.​2.​027004 CrossRefPubMed
19.
go back to reference Maquelin K, Choo-Smith LP, van Vreeswijk T, Endtz HP, Smith B, Bennett R, Bruining HA, Puppels GJ (2000) Raman spectroscopic method for identification of clinically relevant microorganisms growing on solid culture medium. Anal Chem 72:12–19CrossRefPubMed Maquelin K, Choo-Smith LP, van Vreeswijk T, Endtz HP, Smith B, Bennett R, Bruining HA, Puppels GJ (2000) Raman spectroscopic method for identification of clinically relevant microorganisms growing on solid culture medium. Anal Chem 72:12–19CrossRefPubMed
21.
go back to reference Hutsebaut D, Vandenabeele P, Moens L (2005) Evaluation of an accurate calibration and spectral standardization procedure for Raman spectroscopy. Analyst 130:1204–1214. doi:10.1039/b503624k CrossRefPubMed Hutsebaut D, Vandenabeele P, Moens L (2005) Evaluation of an accurate calibration and spectral standardization procedure for Raman spectroscopy. Analyst 130:1204–1214. doi:10.​1039/​b503624k CrossRefPubMed
22.
go back to reference Loonen AJM, Bos MP, van Meerbergen B, Neerken S, Catsburg A, Dobbelaer I, Penterman R, Maertens G, van de Wiel P, Savelkoul P, van den Brule AJC (2013) Comparison of pathogen DNA isolation methods from large volumes of whole blood to improve molecular diagnosis of bloodstream infections. PLoS One 8, e72349. doi:10.1371/journal.pone.0072349.t002 CrossRefPubMedPubMedCentral Loonen AJM, Bos MP, van Meerbergen B, Neerken S, Catsburg A, Dobbelaer I, Penterman R, Maertens G, van de Wiel P, Savelkoul P, van den Brule AJC (2013) Comparison of pathogen DNA isolation methods from large volumes of whole blood to improve molecular diagnosis of bloodstream infections. PLoS One 8, e72349. doi:10.​1371/​journal.​pone.​0072349.​t002 CrossRefPubMedPubMedCentral
28.
go back to reference Willemse-Erix D, Bakker-Schut T, Slagboom-Bax F, Jachtenberg JW, Lemmens-den Toom N, Papagiannitsis CC, Kuntaman K, Puppels G, van Belkum A, Severin JA, Goessens W, Maquelin K (2012) Rapid typing of extended-spectrum β-lactamase- and carbapenemase-producing Escherichia coli and Klebsiella pneumoniae isolates by use of SpectraCell RA. J Clin Microbiol 50:1370–1375. doi:10.1128/JCM.05423-11 CrossRefPubMedPubMedCentral Willemse-Erix D, Bakker-Schut T, Slagboom-Bax F, Jachtenberg JW, Lemmens-den Toom N, Papagiannitsis CC, Kuntaman K, Puppels G, van Belkum A, Severin JA, Goessens W, Maquelin K (2012) Rapid typing of extended-spectrum β-lactamase- and carbapenemase-producing Escherichia coli and Klebsiella pneumoniae isolates by use of SpectraCell RA. J Clin Microbiol 50:1370–1375. doi:10.​1128/​JCM.​05423-11 CrossRefPubMedPubMedCentral
29.
go back to reference Bernatová S, Samek O, Pilát Z, Šerý M, Ježek J, Jákl P, Šiler M, Krzyžánek V, Zemánek P, Holá V, Dvořáčková M, Růžička F (2013) Following the mechanisms of bacteriostatic versus bactericidal action using Raman spectroscopy. Molecules 18:13188–13199. doi:10.3390/molecules181113188 CrossRefPubMed Bernatová S, Samek O, Pilát Z, Šerý M, Ježek J, Jákl P, Šiler M, Krzyžánek V, Zemánek P, Holá V, Dvořáčková M, Růžička F (2013) Following the mechanisms of bacteriostatic versus bactericidal action using Raman spectroscopy. Molecules 18:13188–13199. doi:10.​3390/​molecules1811131​88 CrossRefPubMed
30.
go back to reference Jones RN, Erwin ME, Croco JL (1996) Critical appraisal of E test for the detection of fluoroquinolone resistance. J Antimicrob Chemother 38:21–25CrossRefPubMed Jones RN, Erwin ME, Croco JL (1996) Critical appraisal of E test for the detection of fluoroquinolone resistance. J Antimicrob Chemother 38:21–25CrossRefPubMed
31.
go back to reference Doern GV, Brueggemann AB, Perla R, Daly J, Halkias D, Jones RN, Saubolle MA (1997) Multicenter laboratory evaluation of the bioMérieux Vitek antimicrobial susceptibility testing system with 11 antimicrobial agents versus members of the family Enterobacteriaceae and Pseudomonas aeruginosa. J Clin Microbiol 35(8):2115–2119PubMedPubMedCentral Doern GV, Brueggemann AB, Perla R, Daly J, Halkias D, Jones RN, Saubolle MA (1997) Multicenter laboratory evaluation of the bioMérieux Vitek antimicrobial susceptibility testing system with 11 antimicrobial agents versus members of the family Enterobacteriaceae and Pseudomonas aeruginosa. J Clin Microbiol 35(8):2115–2119PubMedPubMedCentral
32.
go back to reference Willemse-Erix HFM, Jachtenberg J, Barutçi H, Puppels GJ, van Belkum A, Vos MC, Maquelin K (2010) Proof of principle for successful characterization of methicillin-resistant coagulase-negative staphylococci isolated from skin by use of Raman spectroscopy and pulsed-field gel electrophoresis. J Clin Microbiol 48:736–740. doi:10.1128/JCM.01153-09 CrossRefPubMed Willemse-Erix HFM, Jachtenberg J, Barutçi H, Puppels GJ, van Belkum A, Vos MC, Maquelin K (2010) Proof of principle for successful characterization of methicillin-resistant coagulase-negative staphylococci isolated from skin by use of Raman spectroscopy and pulsed-field gel electrophoresis. J Clin Microbiol 48:736–740. doi:10.​1128/​JCM.​01153-09 CrossRefPubMed
33.
go back to reference Gherardi G, Angeletti S, Panitti M, Pompilio A, Di Bonaventura G, Crea F, Avola A, Fico L, Palazzo C, Sapia GF, Visaggio D, Dicuonzo G (2012) Comparative evaluation of the Vitek-2 Compact and Phoenix systems for rapid identification and antibiotic susceptibility testing directly from blood cultures of Gram-negative and Gram-positive isolates. Diagn Microbiol Infect Dis 72:20–31. doi:10.1016/j.diagmicrobio.2011.09.015 CrossRefPubMed Gherardi G, Angeletti S, Panitti M, Pompilio A, Di Bonaventura G, Crea F, Avola A, Fico L, Palazzo C, Sapia GF, Visaggio D, Dicuonzo G (2012) Comparative evaluation of the Vitek-2 Compact and Phoenix systems for rapid identification and antibiotic susceptibility testing directly from blood cultures of Gram-negative and Gram-positive isolates. Diagn Microbiol Infect Dis 72:20–31. doi:10.​1016/​j.​diagmicrobio.​2011.​09.​015 CrossRefPubMed
34.
go back to reference Orelio CC, Beiboer SHW, Morsink MC, Tektas S, Dekter HE, van Leeuwen WB (2014) Comparison of Raman spectroscopy and two molecular diagnostic methods for Burkholderia cepacia complex species identification. J Microbiol Methods 107:126–132. doi:10.1016/j.mimet.2014.10.002 CrossRefPubMed Orelio CC, Beiboer SHW, Morsink MC, Tektas S, Dekter HE, van Leeuwen WB (2014) Comparison of Raman spectroscopy and two molecular diagnostic methods for Burkholderia cepacia complex species identification. J Microbiol Methods 107:126–132. doi:10.​1016/​j.​mimet.​2014.​10.​002 CrossRefPubMed
Metadata
Title
Antimicrobial susceptibility testing of Gram-positive and -negative bacterial isolates directly from spiked blood culture media with Raman spectroscopy
Authors
H. E. Dekter
C. C. Orelio
M. C. Morsink
S. Tektas
B. Vis
R. te Witt
W. B. van Leeuwen
Publication date
01-01-2017
Publisher
Springer Berlin Heidelberg
Published in
European Journal of Clinical Microbiology & Infectious Diseases / Issue 1/2017
Print ISSN: 0934-9723
Electronic ISSN: 1435-4373
DOI
https://doi.org/10.1007/s10096-016-2773-y

Other articles of this Issue 1/2017

European Journal of Clinical Microbiology & Infectious Diseases 1/2017 Go to the issue