Analytical and Bioanalytical Chemistry

, Volume 407, Issue 29, pp 8919–8923 | Cite as

Raman spectroscopic monitoring of the growth of pigmented and non-pigmented mycobacteria

  • Stephan Stöckel
  • Andrei Sebastian Stanca
  • Jonathan Helbig
  • Petra Rösch
  • Jürgen Popp


Raman microspectroscopy has increased in popularity in the field of microbiology because it allows a spectral fingerprinting of bacterial pathogens at an unrivaled speed, which is important for the early treatment of infectious diseases such as tuberculosis. An indispensable prerequisite for the success of this method is a profound knowledge, how the spectral profiles depend on the age of the bacteria. We therefore followed the growth of two rapidly growing Mycobacterium tuberculosis relatives, the pigmented Mycobacteriumaurum, and the non-pigmented Mycobacteriumsmegmatis, by means of Raman microspectroscopy. Both species showed remarkable temporal changes in the single-bacteria Raman spectra: In the signatures of M. aurum, pigment-associated Raman signals could be detected not until 72 h of growth and also remained highly variable thereafter. The Raman spectra of M. smegmatis exhibited lipid signals presumably arising from mycolic acids, which are a hallmark feature of mycobacteria, but only after the bacteria reached the late stationary growth phase (>48 h). A principal component analysis thus classified the Raman spectra according to the cultivation age. In summary, these findings have to be reckoned with in future studies dealing with the identification of mycobacteria via Raman microspectroscopy.

Graphical abstract

Changes in the chemical composition of bacterial cells over growth time may influence the results of Raman spectroscopic studies of bacteria


Raman spectroscopy Microscopy Mycobacteria Growth pigments Analytical methods 



The funding of the research projects Fast-TB (2013FE9057) and BioInter (13022-715) by the Free State of Thuringia and the European Union (EFRE) as well as the Deutsche Forschungsgemeinschaft (DFG) for the Collaborative Research Center ChemBioSys (SFB 1127) is highly acknowledged. The authors also thank Sophie Friedrich for technical assistance.

Conflict of interest

The authors declare that they have no conflict of interest.


  1. 1.
    Lewin A, Sharbati-Tehrani S (2005) Slow growth rate of mycobacteria. Possible reasons and significance for their pathogenicity (Das langsame Wachstum von Mykobakterien. Mögliche Ursachen und Bedeutung fur die Pathogenität.). Bundesgesundhbl Gesundheitsforsch Gesundheitsschutz 48(12):1390–1399CrossRefGoogle Scholar
  2. 2.
    Pahlow S, Meisel S, Cialla-May D, Weber K, Rösch P, Popp J (2015) Isolation and identification of bacteria by means of Raman spectroscopy. Adv Drug Deliv Rev 89:105–120CrossRefGoogle Scholar
  3. 3.
    Gupta A, Bhakta S (2012) An integrated surrogate model for screening of drugs against Mycobacterium tuberculosis. J Antimicrob Chemother 67(6):1380–1391CrossRefGoogle Scholar
  4. 4.
    Brown-Elliott BA, Wallace RJ (2002) Clinical and taxonomic status of pathogenic nonpigmented or late-pigmenting rapidly growing mycobacteria. Clin Microbiol Rev 15(4):716–746CrossRefGoogle Scholar
  5. 5.
    Stöckel S, Meisel S, Elschner M, Melzer F, Rösch P, Popp J (2015) Raman spectroscopic detection and identification of Burkholderia mallei and Burkholderia pseudomallei in feedstuff. Anal Bioanal Chem 407(3):787–794CrossRefGoogle Scholar
  6. 6.
    Viveiros M, Krubasik P, Sandmann G, Houssaini-Iraqui M (2000) Structural and functional analysis of the gene cluster encoding carotenoid biosynthesis in Mycobacterium aurum A+. FEMS Microbiol Lett 187(1):95–101CrossRefGoogle Scholar
  7. 7.
    Jehlička J, Edwards HGM, Oren A (2014) Raman spectroscopy of microbial pigments. Appl Environ Microbiol 80(11):3286–3295CrossRefGoogle Scholar
  8. 8.
    Kumar BNV, Kampe B, Rösch P, Popp J (2015) Characterization of carotenoids in soil bacteria and investigation of their photodegradation by UVA radiation via resonance Raman spectroscopy. Analyst 140(13):4584–4593CrossRefGoogle Scholar
  9. 9.
    Koyama Y, Kito M, Takii T, Saiki K, Tsukida K, Yamashita J (1982) Configuration of the carotenoid in the reaction centers of photosynthetic bacteria—comparison of the resonance Raman-spectrum of the reaction center of Rhodopseudomonas sphaeroides G1C with those of cis-trans isomers of beta-carotene. Biochim Biophys Acta 680(2):109–118CrossRefGoogle Scholar
  10. 10.
    de Oliveira VE, Castro HV, Edwards HGM, de Oliveira LFC (2010) Carotenes and carotenoids in natural biological samples: a Raman spectroscopic analysis. J Raman Spectrosc 41(6):642–650CrossRefGoogle Scholar
  11. 11.
    Walter A, Schumacher W, Bocklitz T, Reinicke M, Rösch P, Kothe E, Popp J (2011) From bulk to single-cell classification of the filamentous growing streptomyces bacteria by means of Raman spectroscopy. Appl Spectrosc 65(10):1116–1125CrossRefGoogle Scholar
  12. 12.
    Marrakchi H, Lanéelle MA, Daffé M (2014) Mycolic acids: structures, biosynthesis, and beyond. Chem Biol 21(1):67–85CrossRefGoogle Scholar
  13. 13.
    Rivera-Betancourt OE, Karls R, Grosse-Siestrup B, Helms S, Quinn F, Dluhy RA (2013) Identification of mycobacteria based on spectroscopic analyses of mycolic acid profiles. Analyst 138(22):6774–6785CrossRefGoogle Scholar
  14. 14.
    Kumar V, Kampe B, Rösch P, Popp J (2015) Classification and identification of pigmented cocci bacteria relevant to the soil environment via Raman spectroscopy. Environ Sci Pollut Res. doi: 10.1007/s11356-015-4593-5 Google Scholar
  15. 15.
    Silge A, Abdou E, Schneider K, Meisel S, Bocklitz T, Lu-Walther H-W, Heintzmann R, Rösch P, Popp J (2015) Shedding light on host niches: label-free in situ detection of Mycobacterium gordonae via carotenoids in macrophages by Raman microspectroscopy. Cell Microbiol 17(6):832–842CrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2015

Authors and Affiliations

  • Stephan Stöckel
    • 1
    • 2
  • Andrei Sebastian Stanca
    • 1
  • Jonathan Helbig
    • 1
  • Petra Rösch
    • 1
    • 2
  • Jürgen Popp
    • 1
    • 2
    • 3
  1. 1.Institute of Physical Chemistry and Abbe School of PhotonicsFriedrich Schiller University JenaJenaGermany
  2. 2.InfectoGnostics Forschungscampus JenaJenaGermany
  3. 3.Leibniz-Institute of Photonic TechnologyJenaGermany

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