Calorimetric Analysis of the Growth of Anaerobic Microbes Cultured on Insoluble Carbon Sources

摘要

Methods for the measurement of anaerobic microbial growth are generally complex. Furthermore, aggregation and precipitation make it difficult to estimate bacterial cell counts using turbidity. Recently, we demonstrated that heat evolution is a good indicator of the growth of anaerobic bacteria cultured on soluble carbon sources. In this study, we validated the use of calorimetry to measure growth on insoluble carbon sources using Clostridium spp. When Clostridium cellulovorans and C. thermocellum were cultured on crystalline cellulose, their optical density values over time were irregular. However, the thermograms indicating the heat evolution and the integrated ATP concentrations of the cultures showed highly correlated growth curves. When C. acetobutylicum and C. beijerinckii were cultured on corn starch, peaks were observed at the initial stage based on thermograms and ATP concentration measurements. Thin-layer chromatography revealed that these peaks corresponded to the rapid metabolism of monosaccharides and disaccharides in the culture broth. Thus, the growth of anaerobic microbes cultured on insoluble carbon sources can be measured conveniently and accurately by calorimetry, without confounding effects of aggregation or precipitation. Growth can be measured with high sensitivity, comparable to that of ATP measurements, indicating that calorimetry is a promising method for measuring microbial growth.