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Microbial biomarkers that indicate aspects of an organism's growth conditions are important targets of forensic research. In this study, we examined fatty acid composition as a signature for the types of complex nutrients in the culturing medium. Bacillus cereus T-strain spores were grown in medium formulations supplemented with one of the following: peptone (meat protein), tryptone (casein protein), soy protein, and brain-heart infusion. Cellular biomass was profiled with fatty acid methyl ester (FAME) analysis. Results showed peptone cultures produced spores enriched in straight-chained lipids. Tryptone cultures produced spores enriched in branched-odd lipids when compared with peptone, soy, and brain-heart formulations. The observed FAME variation was used to construct a set of discriminant functions that could help identify the nutrients in a culturing recipe for an unknown spore sample. Blinded classification tests were most successful for spores grown on media containing peptone and tryptone, showing 88% and 100% correct identification, respectively., (© 2015 American Academy of Forensic Sciences.)

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.