Rapid differentiation of Bacillus strains using hydrophobic grid membranes and attenuated total reflectance Infrared microspectroscopy.
Bacillus species may be resistant to processing and sanitation procedures, making their control an important issue in the food industry. The objective of this study was to develop a rapid method for the differentiation of Bacillus cells at the strain level using infrared microspectroscopy and multivariate pattern recognition techniques. Aliquots (10 ml) of vegetative cells (approximately 10(3) CFU/ml) from four strains of each of three Bacillus species (B. cereus, B. mycoides, and B. thuringiensis) were filtered onto hydrophobic grid membranes. The membranes were placed on tryptic soy agar and incubated at 42 degrees C for 24 h and then removed from the agar and dried, and the biomass of individual vegetative colonies was directly measured by attenuated total reflectance infrared (ATR-IR) microspectroscopy. Soft independent modeling of class analogy models generated from second derivative transformed spectra in the 1,300 to 900 cm(-1) region exhibited clusters that permitted accurate strain-level classification of all isolates. Major discrimination was related to the signal from phosphate-containing compounds, likely phospholipids. Results indicate that a simple ATR-IR microspectroscopy technique combined with multivariate analysis could provide the food industry with a rapid and reagent-free screening procedure to complement more elaborate molecular identification methods.
Department of Food Science and Technology, The Ohio State University, 2015 Fyffe Road, Parker Food Science Building, Columbus, Ohio 43210, USA.
Elizabeth M
EM
GrasSo
Ahmed E
AE
Yousef
Luis A
LA
Rodriguez-Romo
Luis E
LE
Rodriguez-Saona
D001407:Bacillus / Q000145:classification* / Q000302:isolation & purification; D015169:Colony Count, Microbial / Q000379:methods*; D005506:Food Contamination / Q000032:analysis*; D005516:Food Microbiology; D015591:Microspectrophotometry; D015999:Multivariate Analysis; D010802:Phylogeny; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013045:Species Specificity; D013055:Spectrophotometry, Infrared