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K. Wieland, J. Kuligowski, D. Ehgartner, G. Ramer, C. Koch, J. Ofner, C. Herwig, B. Lendl:
"Toward a Noninvasive, Label-Free Screening Method for Determining Spore Inoculum Quality of Penicillium chrysogenum Using Raman Spectroscopy";
Applied Spectroscopy, 71 (2017), 12; 2661 - 2669.

We report on a label-free, noninvasive method for determination of spore inoculum quality of Penicillium chrysogenum prior
to cultivation/germination. Raman microspectroscopy providing direct, molecule-specific information was used to extract
information on the viability state of spores sampled directly from the spore inoculum. Based on the recorded Raman
spectra, a supervised classification method was established for classification between living and dead spores and thus
determining spore inoculum quality for optimized process control. A fast and simple sample preparation method consisting
of one single dilution step was employed to eliminate interfering signals from the matrix and to achieve isolation of single
spores on the sample carrier (CaF2). Aiming to avoid any influence of the killing procedure in the Raman spectrum of the
spore, spores were considered naturally dead after more than one year of storage time. Fluorescence staining was used as
reference method. A partial least squares discriminant analysis classifier was trained with Raman spectra of 258 living and
dead spores (178 spectra for calibration, 80 spectra for validation). The classifier showed good performance when being
applied to a 1 mL droplet taken from a 1:1 mixture of living and dead spores. Of 135 recorded spectra, 51% were assigned to
living spores while 49% were identified as dead spores by the classifier. The results obtained in this work are a fundamental
step towards developing an automated, label-free, and noninvasive screening method for assessing spore inoculum quality.

http://dx.doi.org/10.1177/0003702817727728