ANALYTICAL APPROACHES TO THE CHARACTERIZATION OF SAMPLES OF MICROBIALCOMMUNITIES USING PATTERNS OF POTENTIAL C SOURCE UTILIZATION

Profiles of potential utilization of 95 separate C sources by microbia l communities can be readily generated from direct incubation of envir onmental samples in BIOLOG microplates. Color formation from a redox i ndicator dye is used to quantify the degree of C source utilization. I aimed to examine different analytical approaches for classifying micr obial communities based on these profiles. Specifically, the relative effects of average rate of color development versus the pattern of rel ative C source utilization on the classification of rhizosphere sample s from different crop types were evaluated. The average rate of color development was correlated to the density of total (acridine-orange) b acterial cells (R(2) = 0.52) and active (5-cyano-2,3-ditolyl tetrazoli um chloride) bacterial cells (R(2) = 0.70) inoculated into the plate. Analysis of plates with different rates of color development after a s pecific incubation period resulted in samples with variation in the ov erall extent of color development (expressed as average well color dev elopment, or AWCD). Classification of these samples using principal co mponent analysis was significantly influenced by the variation in AWCD , resulting in the classification of samples based on the density of i noculum rather than the pattern of C source utilization. The effect of variation in AWCD was eliminated by normalizing data prior to ordinat ion, or by using Bn alternative ordination technique, detrended corres pondence analysis. Variation in AWCD can be limited through multiple-p late readings and subsequent selection of plates with a common referen ce point in AWCD. The specific AWCD used for analysis does not appear important for classification purposes;consistent discrimination of rhi zosphere samples from different crop types was apparent for analysis a cross a wide range of AWCD (0.25-1.00 abs. units). The specific differ ences in C source utilization between rhizosphere sample types did dep end on the set point used for analysis due to the differences in the r ate of color formation among wells. Results suggest that single-plate readings can be used to classify samples, but only if potential differ ences in AWCD are accounted for in the data analysis. Repeated plate r eadings will provide a more complete understanding of differences in C source utilization among samples.