The Use of Bacteriophage-Receptor Genes of Lactococcus lactis to Develop Bacteriophage Resistance in Cheddar Cheese Starter Strains

Principal Investigator:

Bruce L. Geller, Associate Professor of Microbiology, Oregon State University

Co-Investigators:

 

Project Summary:

The proposed research examines early steps of bacteriophage infection of L. lactis, which include attachment of the phage to the surface of cells and entry of phage DNA into cells. Both of these steps are required for infection by phage. Our strategy of strain improvement is to prevent to phage from attaching or entering the host in the first place. To do this requires knowledge of the host components required for attachment and phage entry. The outcomes of this proposal will enable the construction of new strains with defined mechanisms of phage-resistance. Host genes required for phage infection of L. lactis will be identified and isolated. We have previously isolated one such gene named pip (an acronym for phage infection protein). The protein encoded by pip (Pip) is a receptor for phage attachment and phage DNA entry into the host. We have constructed phage-resistant strains of L. lactis by replacing the pip gene with a defective version. There is evidence that host components in addition to pip are required for phage attachment and DNA entry. Isolating genes in addition to pip that are required for phage infection will enable the construction of new strains with alterations in two or more different host components. The strategy of combining multiple phage-resistance mechanisms will greatly decrease the chance that the strain will fail after introduction into commercial use. Genes will be isolated that also extend the range of resistance to phages that do not require pip. We propose to isolate genes that encode host receptors for two different types of small isometric-head phage (p335 and 936). Together with the phages that required pip, the p335 and 936 species of phage cause nearly all the starter failures in U.S. cheese factories. Phages of the p335 species are particularly troublesome, as they have only recently emerged as a major problem, and less is known about their mechanism of infection. The isolated receptor genes will be inactivated and used to construct a new commercial strain with a combined phage-resistance defined by each of the inactivated genes (including pip). The phage-resistant strain will be evaluated for physiological characteristics important for making cheese.

Publications:

Kraus, J. and B. L. Geller. 2001. Cloning of Genomic DNA of Lactococcus lactis that restores phage sensitivity to an unusual bacteriophage sk1-resistant mutant. Appl. Environ. Microbiol. 67:791-798.

Dai, G. P. Su, G. E. Alllison, B. L. Geller, P. Zhu, W. S. Kim, and N. W. Dunn. 2001. Molecular Characterization of a New Abortive Infection System (AbiU) from Lactococcus lactis LL 51-1. Appl. Environ. Microbiol. 67:5225-5232.

Theses:

None

Published Abstract:

None

Presentations:

"Characterization of a Receptor for Small Isometric Headed Phages of Lactococcus lactis ". Hang Thu Ngo , Ping Su, Noel Dunn, and Bruce L. Geller. Federation of Microbiology Societies Seventh Symposium on Lactic Acid Bacteria; Genetics, Metabolism and Applications. September 1-5, 2002, Egmond aan Zee, The Netherlands.