Characterization of a plasmid-cured variant isolated from the lysogenic Lactobacillus helveticus CRL 1179.

GORRITI, M.; VILLENA, J.C.; ESPECHE TURBAY, B.; DYEN, M.; HEBERT, E.M.; RAYA, R.R.

San Miguel de Tucumán

Simposio; II SIMPOSIO INTERNACIONAL DE BACTERIAS LACTICAS; 2006

CERELA-CONICET

Lactobacillus helveticus is a homofermentative, thermophilic lactic acid bacterium used commercially for the manufacture of hard cheeses. In this work, strain CRL1179 was chosen as a model microorganisms to optimize an electroporation protocol for L. helveticus, since there is as yet no a generalized method for transforming this bacterium. It was found that strain CRL 1179 is lysogenic, possesses two plasmids of approximately 7 y 10 kilobases (Kb), and shows an Fmc- phenotype, since it coagulates nonfat milk after 16 h. The temperate phage of CRL1179, named φLhR3, could be induced to the lytic cycle with mitomycin C (0,2 µg/ml). Phage lysates could inhibit cell growth of sensitive cells, but not phage plaques could be observed. A plasmid-free derivative of CRL1179, named C4, was isolated and characterized. Strain C4 has lost the 7 kb plasmid, and shows a phenotype of strong agglutination and an extended lag phase (2-4 h) with regard to the original strain CRL1179. In both strains the phenotype Fmc- could be attributed to a deficiency in the biosynthesis of purines, but not to the absence of a proteinase activity. All intents to electroporate CRL1179 y C4 with plasmids pGK12 and pSA3 has been so far unsatisfactory.