Further Studies on the Morphology ofEscherichia coliExposed to Penicillin

Abstract

In a previous paper, Shanahan, Eisenstark, and Tanner (1947) reported electron microscope studies of Escherichia coli cells exposed to penicillin. Brief mention was made of light microscope studies in contrast with electron microscope results. Further information gained by light microscope studies as well as other pertinent data is presented in the present paper. MATERIALS AND METHODS Twenty-four strains of gram-negative rods considered to be Escherichia coli were used. Strains 251, 252, 253, 254, and 257 were from the department culture collection. Strains "0," "R," and "U" were kindly supplied by Dr. Alture-Werber of the Jewish Hospital, Brooklyn, New York. Eight other strains were obtained from various laboratories, and the remaining eight cultures were isolated by the senior author from fecal samples. Stock cultures were routinely transferred on nutrient agar with frequent colony isolations made to ensure pure cultures. As suggested by the work of Alture-Werber et al. (1945), MacConkey's agar (Difco) was selected as the test medium. Concentrations of penicillin2 are given in terms of final concentration in the medium. For examination of morphological changes in cells, colonies were selected on MacConkey's agar plates, which were incubated at 37 C. Gram stains and certain other staining procedures-were used to determine the morphology after cells had grown in penicillin-containing media. Slide cultures for the examination of living cells were made as follows: About 0.1 ml of MacConkey's agar containing a desired concentration of penicillin was pipetted aseptically onto a sterile glass slide. A small drop of the test organism, usually diluted 1: 10 with sterile saline or broth, was placed on the hardened agar. A sterile cover slip was quickly placed over the inoculum and agar. Melted paraffin was run around the edges of the cover slip with a wooden applicator to ensure a firm seal. Culture slides were placed in sterile petri dishes and incubated in a moist chamber at 37 C. Slides were removed for examination at intervals, all examinations being made with oil immersion lens. Dienes (1942) noted that large round bodies developing in bacterial cultures were fragile, and he did not recommend direct microscopical examination of unstained cultures. In the present work it was found relatively simple to tease off