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Applied and Environmental Microbiology, September 2006, p. 6174-6182, Vol. 72, No. 9
0099-2240/06/$08.00+0     doi:10.1128/AEM.00515-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Use of DNA Quantification To Measure Growth and Autolysis of Lactococcus and Propionibacterium spp. in Mixed Populations

Janneke Treimo,1* Gerd Vegarud,1 Thor Langsrud,1 and Knut Rudi2,3*

Department of Chemistry, Biotechnology and Food Science, Norwegian University of Life Sciences, N-1432 Ås, Norway,1 MATFORSK, Norwegian Food Research Institute, Osloveien 1, N-1430 Ås, Norway,2 Hedmark University College, N-2306 Hamar, Norway3

Received 3 March 2006/ Accepted 10 July 2006

Autolysis is self-degradation of the bacterial cell wall that results in the release of enzymes and DNA. Autolysis of starter bacteria, such as lactococci and propionibacteria, is essential for cheese ripening, but our understanding of this important process is limited. This is mainly because the current tools for measuring autolysis cannot readily be used for analysis of bacteria in mixed populations. We have now addressed this problem by species-specific detection and quantification of free DNA released during autolysis. This was done by use of 16S rRNA gene single-nucleotide extension probes in combination with competitive PCR. We analyzed pure and mixed populations of Lactococcus lactis subsp. lactis and three different species of Propionibacterium. Results showed that L. lactis subsp. lactis INF L2 autolyzed first, followed by Propionibacterium acidipropionici ATCC 4965, Propionibacterium freudenreichii ISU P59, and then Propionibacterium jensenii INF P303. We also investigated the autolytic effect of rennet (commonly used in cheese production). We found that the effect was highly strain specific, with all the strains responding differently. Finally, autolysis of L. lactis subsp. lactis INF L2 and P. freudenreichii ISU P59 was analyzed in a liquid cheese model. Autolysis was detected later in this cheese model system than in broth media. A challenge with DNA, however, is DNA degradation. We addressed this challenge by using a DNA degradation marker. We obtained a good correlation between the degradation of the marker and the target in a model experiment. We conclude that our DNA approach will be a valuable tool for use in future analyses and for understanding autolysis in mixed bacterial populations.

* Corresponding author. Mailing address for Knut Rudi: MATFORSK, Osloveien 1, N-1430 Ås, Norway. Phone: 47 64 97 01 00. Fax: 47 64 97 03 33. E-mail: knut.rudi{at}matforsk.no. Mailing address for Janneke Treimo: Department of Chemistry, Biotechnology and Food Science, Norwegian University of Life Sciences, N-1432 Ås, Norway. Phone: 47 64 96 59 00. Fax: 47 64 96 59 01. E-mail: janneke.treimo{at}umb.no.

Applied and Environmental Microbiology, September 2006, p. 6174-6182, Vol. 72, No. 9
0099-2240/06/$08.00+0     doi:10.1128/AEM.00515-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Hojo, K., Watanabe, R., Mori, T., Taketomo, N. (2007). Quantitative Measurement of Tetrahydromenaquinone-9 in Cheese Fermented by Propionibacteria. J DAIRY SCI 90: 4078-4083 [Abstract] [Full Text]  
  • Trosvik, P., Skanseng, B., Jakobsen, K. S., Stenseth, N. C., Naes, T., Rudi, K. (2007). Multivariate Analysis of Complex DNA Sequence Electropherograms for High-Throughput Quantitative Analysis of Mixed Microbial Populations. Appl. Environ. Microbiol. 73: 4975-4983 [Abstract] [Full Text]  


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