Taxonomic Value of Infrared Spectra.

Efforts to demonstrate species or generic relationships among bacteria throu-h the use of infrared (IR) spectra of whole cells (Norris, J. Hyg. 57:326, 1959) have been increasing despite frequent cautions (Kenner et al., J. B3acteriol. 75:16, 1958) that such procedures are fraught with experimental pitfalls. AIany of the reports concerning bacterial identification by this technique are favorable and indicate a general acceptance of the method. While investigating the balanced growth of Escherichia coli K-12 (Wright, Ph.D. Thesis, Iowa State University, Ames, 1964), IR spectra were obtained from cells grown in a defined medium with either the carbon or nitrogen source as the limiting substrate, and at either a high (0.5 generation per hr) or a low (0.1 generation per hr) growth rate. The medium contained: 0.7% K2HPO4 3H20, 0.3%O KH2PO4, and 0.01% MgSO4 7H20 in deionized water, to which was added 400,ug of (NH4)2SO4 per ml, and either 500,ug of glucose per ml (for carbonlimited cultures) or 4,000 Ag of glucose per ml (for nitrogen-limited cultures). Rate of growth was controlled by the dilution rate in continuous culture. Pressed KBr discs were prepared with cells from each culture, and the absorption spectra between 850 and 1,820 cnml were determined in a Beckman IR-7 spectrophotometer. IR spectra of genetically identical cells, with all environmental factors controlled and differing only in their rate of growth or limiting substrate, show considerably greater differences (Fig. 1) than those previously reported as sionificant in differentiating strains of a species (Scopes, J. Gen. Microbiol. 28:69, 1962) or the species of a genus (Levine et al., J. Infect. Dis. 96:193, 1955). If the IR patterns (Fig. 1) are analyzed by means of base line ratios (Wright, Ind. Eng. Chem. Anal. Ed. 13:1, 1941), it is apparent that the spectra are indeed different, suggesting that each cell type produces a characteristic pattern dependent both on the growth-limiting substrate and the rate of growth. The quantitative relationships among the various absorption bands were found by dividing their absorbancy by the absorbancy at 1,530 cm-'. The ratios from the