A method for selection of antibiotic producing strains of Penicillium chrysogenum on solid media by means of microcup cultivation and image analyzing of inhibition zones

Abstract

The paper describes a method for proper estimation of differences of antibiotic yields produced by selectants of Penicillium chrysogenum on solid media. The reliability of the technique was proved by the use of different mutants isolated after treatment with NTG. By the aid of an inoculator, spores of each strain were transferred onto agar surfaces of so-called microcups which all possessed open bottoms for the determination of antibiotic content by bioassay. These were small cylindrical vessels of polypropylene containing one ml of solid agar medium similar in its composition to fermentation broth. After sufficient incubation, the developed microcultures were stored at 4–6°C for at least 24 h before they were set up on bioassay plates. This procedure permitted equilibration of antibiotic concentration within the cylindrical agar piecces. This was absolutely necessary in order to reproduce the characteristic distinctions between the proved selectants. The yields were then estimated by measuring the areas of the inhibition zones with an image analyzing system (Quantimet 720 M, controlled by a PDP 11 computer). In contrast to microcultures that had not been cooled after growth, the statistical comparison of repeated experiments, calculated by a new method (Horn, M., Bormann, E.J., Harnish, M. (1984), Ein statistisches Verfahren zur Ermittlung der Leistungs-Korrelation antibiotikabildender Mikroorganismen im Labor- und Tecknikumsmaβstab. Allg. Mikrobiol. 24, 7, 443–449), delivered a very good correlation.