Anaerobic susceptibility testing slight differences in inoculum size can make a difference in minimum inhibitory concentrations

Abstract

In the present study, we compared the actual inoculum density from inoculated broth microdilution wells to the targeted inoculum size (10 5 CFU/well) when the inoculum was prepared using a McFarland nephelometer to achieve the standard density. Three target inoculum sizes (10 5, 5 × 10 5, and 10 6 CFU/well) were used to compare the effect of slight inoculum size increases of both ATCC and clinical strains of anaerobes on MICs of various antimicrobials. Actual colony counts of Bacteroides fragilis, Bacteroides thetaiotaomicron, Eubacterium lentum, and Veillonella parvula ranged from 0.7 × 10 5 to 1.4 × 10 5 CFU/well. As the inoculum size rose above the desired 10 5 CFU/well level, the MICs of certain antimicrobials became elevated. Ceftizoxime, cefotaxime, and ceftriaxone MICs rose 4- to 16-fold with as little as 0.5 log 10 increase in inoculum size. Other increases were also noted with E. lentum and Clostridium perfringens, but were primarily between the low and high inoculum sizes. Results with cefoxitin, cefotetan, mezlocillin, and imipenem did not show an appreciable increased inoculum effect. This study demonstrates that the variation in organism size among anaerobes (both between species and within species) does make a difference in actual inoculum size and certain anaerobes may require special adjustment to ensure proper MIC results from susceptibility testing.