Citrate catabolism by Lactobacillus casei in ripening cheese.
Citrate catabolism was utilized as a model to design novel protocols for the identification and study of alternate energy sources for the non-starter lactic acid bacteria (NSLAB), Lactobacillus casei , in ripening cheese. Techniques utilized to develop such protocols included the screening of the Lactobacillus casei ATCC334 genomic sequence, metabolic flux analyses, and a cheese model system, Cheddar cheese extract (CCE). Additionally, the influence of selected culture conditions on citrate utilization by Lb. casei ATCC334, and end products formed from its catabolism were evaluated. Citrate utilization by this microorganism was inhibited in the presence of glucose, lactose, and excess carbohydrate. External pH influenced citrate utilization by extending the lag phase for substrate catabolism with increasing acidity. Lb. casei growth was enhanced in modified chemically defined media (mCDM) supplemented with limiting galactose and citrate. Acetic acid was the main catabolic end product detected in mCDM supplemented with limiting galactose, while L-lactate was the main product in the presence of excess galactose. Acetic and L-lactic acid production by Lb. casei in CCE containing approximately 2.56 +/- 0.09 mM hexose equivalents and 14.7 +/- 0.13 mM citrate was observed. Additional end products detected in CCE in trace concentrations include diacetyl, formate, acetoin, and ethanol. Production of 2-butanone by Lb. casei during the late logarithmic phase of growth in CCE was additionally detected. The screening of the Lb. casei ATCC334 genomic sequence suggested that 2-butanone might have been produced from catabolism of butyric acid.