Differentiation of penicillinase in Bacillus cereus

Abstract

Agar gel electrophoresis has been used to achieve the separation of beta-lactamases produced by two strains of Bacillus cereus. The detection and Identification of these enzymes was made by meaos of combining microbiological, Chemical and immunological revelation of the electropherograms.

Bacillus cereus NRRL 569/H, constitutive strain, produced five exopenicillinases named P-38, P-52, P-67, P-96 and P-143, according to their electrophoretic mobility relative to that of the human albumin.
Other physico-chemical characteristics (thermolability, sedimentation constant, inactivation by iodine, etc.) were also studied which allowed us to differentiate two isopenicillinase families which are immunologically completely different.

All cell-bound constitutive and induced penicillinases were also found in the corresponding cell-free supernatant fluid.

When the constitutive strain was cultured in presence of sodium benzylpenicillin, 10 ul/ml, no quantitatively significant change was observed in total penicillinase level. Nevertheless the electrophoretic distribution of exopenicillinases was altered in such a manner that it was not possible to recognize some normal consti­tutive ones while another «forced-constitutive» penicillinase (P-78) appeared. Its antigenic community with normal consti­ tutive penicillinases was showed by heterologous absorption of antisera. Only in this forced-constitutive culture one cellbound penicillinase was detected which was not present in the corresponding cell-free fluid.

The inducible strain Bacillus cereus NRRL 569 was also cultured both with and without sodium benzylpenicillin in the media. In the second case no penicilli­ nases were found in the cell extract in the cell-free culture fluid. Nevertheless, when effective induction was performed six isopenicillinases were detected five of which were the very same constitutive ones. The other one (P-80) seems characteristic and exclusive of induced culture. It presents immunological differences with respect to the other five induced penicillinases.