A novel staining method for detecting phytase activity

Differential agar media for the detection of microbial phytase activity use the disappearance of precipitated calcium or sodium phytate as an indicati on of enzyme activity. When this technique was applied to the study of rumi nal bacteria, it became apparent that the method was unable to differentiat e between phytase activity and acid production. Strong positive reactions ( zones of clearing around microbial colonies) observed for acid producing, a naerobic bacteria, such as Streptococcus bovis, were not corroborated by su bsequent quantitative assays. Experimentation revealed that acidic solution s generated false positive results on the selected differential medium. Emp irical studies undertaken to find a solution to this Limitation determined the false positive results could be eliminated through a two step counterst aining treatment (cobalt chloride and ammonium molybdate/ammonium vanadate) which reprecipitates acid solubilized phytate. This report discusses the a pplication of the developed two step counterstaining treatment for the scre ening of phytase producing ruminal bacteria as well as its use in phytase z ymogram assays. (C) 1999 Published by Elsevier Science B.V.