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Journal of Bacteriology, September 2000, p. 5097-5104, Vol. 182, No. 18
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Characterization of Vibrio cholerae O1 Antigen as the Bacteriophage K139 Receptor and Identification of IS1004 Insertions Aborting O1 Antigen Biosynthesis

Jutta Nesper,1 Dagmar Kapfhammer,1 Karl E. Klose,2 Hilde Merkert,3 and Joachim Reidl1,*

Zentrum für Infektionsforschung1 and Institut für Molekulare Infektionsforschung,3 Universität Würzburg, 97070 Würzburg, Germany, and Health Science Center, University of Texas, San Antonio, Texas 78284-77582

Received 16 March 2000/Accepted 23 June 2000

Bacteriophage K139 was recently characterized as a temperate phage of O1 Vibrio cholerae. In this study we have determined the phage adsorption site on the bacterial cell surface. Phage-binding studies with purified lipopolysaccharide (LPS) of different O1 serotypes and biotypes revealed that the O1 antigen serves as the phage receptor. In addition, phage-resistant O1 El Tor strains were screened by using a virulent isolate of phage K139. Analysis of the LPS of such spontaneous phage-resistant mutants revealed that most of them synthesize incomplete LPS molecules, composed of either defective O1 antigen or core oligosaccharide. By applying phage-binding studies, it was possible to distinguish between receptor mutants and mutations which probably caused abortion of later steps of phage infection. Furthermore, we investigated the genetic nature of O1-negative strains by Southern hybridization with probes specific for the O antigen biosynthesis cluster (rfb region). Two of the investigated O1 antigen-negative mutants revealed insertions of element IS1004 into the rfb gene cluster. Treating one wbeW::IS1004 serum-sensitive mutant with normal human serum, we found that several survivors showed precise excision of IS1004, restoring O antigen biosynthesis and serum resistance. Investigation of clinical isolates by screening for phage resistance and performing LPS analysis of nonlysogenic strains led to the identification of a strain with decreased O1 antigen presentation. This strain had a significant reduction in its ability to colonize the mouse small intestine.

* Corresponding author. Mailing address: Zentrum für Infektionsforschung, Universität Würzburg, Röntgenring 11, 97070 Würzburg, Germany. Phone: (49) (0) 931 312153. Fax: (49) (0) 931 312578. E-mail: joachim.reidl{at}mail.uni-wuerzburg.de.

Journal of Bacteriology, September 2000, p. 5097-5104, Vol. 182, No. 18
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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