Method for the selection of Lactococcus lactis mutants producing excess carbon dioxide
One of the characteristics of Roquefort cheese is the presence of irregularly shaped openings. Although many factors affect the development of opening in blue-veined cheeses (Martley & Crow, 1996), the limiting step in the case of Roquefort cheese is the production of CO2 by lactic acid bacteria (J.-P. Reverbel, pers. comm.). Most of the opening occurs after moulding; the process is difficult to control and many manufacturing runs result in cheeses with an insufficient opening. Concentrated suspensions of Leuconostoc strains are used to increase the production of CO2 (Devoyod & Muller, 1969), but it would clearly be useful to have microorganisms that produce larger quantities of the gas. McKay & Baldwin (1974) isolated a spontaneous mutant of Lactococcus lactis that produced more acetoin and CO2 than the parent strain. This mutant was lactate dehydrogenase (LDH)-deficient, which favoured the conversion of pyruvate into end products other than lactate. Following nitrosoguanidine mutagenesis, we isolated three Lc. lactis mutants whose LDH activities were reduced to varying extents and which produced varying amounts of CO2 (Boumerdassi et al. 1997). Subsequent work showed that these mutants were unstable on successive subculture in milk or synthetic broth (El Attar et al. 2000).
The aim of our current work was to select a large number of Lc. lactis mutants producing excess CO2. This would increase the probability of selecting stable mutants and also provide a collection of strains with differing gas production activities. Currently available screening methods are, however, unsuitable for processing large numbers of mutants, which is why we have developed an improved screening method.(Received October 4 1999)
(Accepted June 8 2000)
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p1 Present address: Nestlé Product Technology Centre, F-14100 Lisieux, France.