Forensic differentiation of Bacillus cereus spores grown using different culture media using Raman spectroscopy
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Some microorganisms have been shown to retain a chemical signature indicative of the medium used for culturing. However, the repeatability of medium-specific chemical signatures has not been demonstrated from samples of microorganisms produced in the same batch or in different batches by the same sporulation protocol. Here, the variation in Raman spectra of bacterial endospores repeatedly prepared by the same procedure is compared to the variation between Raman spectra of spores prepared using different media. Bacillus cereus T strain (BcT) samples were correctly classified according to the medium used to induce sporulation for 100 % of spores grown in a controlled manner by the same scientist using Raman spectroscopy and multivariate data analysis. The proof-of-concept results from BcT spores produced in 12 different sporulation media showed correct classification by medium for 98 % of samples (with 100 % classification accuracy for all but one sporulation medium in this data set). Spectral differences were discerned between spores that had been freshly prepared or freeze-dried and spores that had been frozen; however, the differences did not impact the classification of the sporulation medium. Latent variables reduced the classification accuracy of BcT sporulated in G medium by different scientists using different media lots and stored for different periods of time and requires further study.
KeywordsBioanalytical methods Bacterial spores Chemometrics/statistics IR spectroscopy/Raman spectroscopy
We are very grateful to Jack Hietpas for independent statistical analysis and confirmation of the results presented here. We would also like to thank Jason Brewer (FBI Laboratory, Chemistry Unit) for providing assistance with the Raman instrument and Lindsay Lundberg for spore preparations. This research was supported in part by an appointment to the Visiting Scientist Program at the FBI Laboratory Division, administered by the Oak Ridge Institute of Science and Education, through an interagency agreement between the US Department of Energy and the FBI.
This is publication number 14-11 of the Laboratory Division of the Federal Bureau of Investigation. Names of commercial manufacturers are provided for information only, and inclusion does not imply endorsement by the FBI or the US Government. The views expressed are those of the authors and do not necessarily reflect the official policy or position of the FBI or the U.S. Government.
- 1.Jackman J (2012) The microbe: the basics of structure, morphology, and physiology as they relate to microbial characterization and attribution. In: Cliff JB, Kreuzer HW, Ehrhardt CJ, Wunschel DS (eds) Chemical and physical signatures for microbial forensics. Springer, New York, pp 13–24CrossRefGoogle Scholar
- 16.Kümmerle M, Scherer S, Seiler H (1998) Appl Environ Microbiol 64:2207–2214Google Scholar
- 21.Church BD, Halvorson H, Halvorson HO (1954) J Bacteriol 68:393–399Google Scholar
- 23.Burke WF (1982) J Gen Microbiol 128:1591–1597Google Scholar
- 27.Morisaki S, Ota C, Matsuda K, Kaku N, Fujiwara H, Oda R, Ishibashi H, Kubo T, Kawata M (2013). J Biomed Opt 18:116011–116018Google Scholar
- 32.Næs T, Isaksson T, Fearn T, Davies T (2002) A user-friendly guide to multivariate calibration and classification. NIR Publications, Chicester, UKGoogle Scholar
- 34.Snow CC, Hartman S, Giles E, Young FA (1979) J Forensic Sci 24:448–460Google Scholar
- 41.Bassi D, Cappa F, Cocconcelli PS (2012) Water and cations flux during sporulation and germination. In: Abel-Santos E (ed) Bacterial spores: current research and applications. Caister Academic Press, NorfolkGoogle Scholar
- 42.Bach ML, Gilvarg C (1966) J Biol Chem 241:4563–4566Google Scholar
- 44.Aronson AI (2012) The structure and composition of the outer layers of the bacterial spores. In: Abel-Santos E (ed) Bacterial spores: current research and applications. pp 57–71Google Scholar
- 45.Dai D, Holder D, Raskin L, Xi C (2011). BMC Microbiol 11 (59)Google Scholar