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pIII^CTX, a Predicted CTX Minor Coat Protein, Can Expand the Host Range of Coliphage fd To Include Vibrio cholerae

Division of Geographic Medicine/Infectious Disease, New England Medical Center and Tufts University School of Medicine, and Howard Hughes Medical Institute, Boston, Massachusetts 02111

Received 19 August 2002/ Accepted 11 November 2002

CTX is a filamentous bacteriophage that encodes cholera toxin. CTX infection of its host bacterium, Vibrio cholerae, requires the toxin-coregulated pilus (TCP) and the products of the V. cholerae tolQRA genes. Here, we have explored the role of OrfU, a predicted CTX minor coat protein, in CTX infection. Prior to the discovery that it was part of a prophage, orfU was initially described as an open reading frame of unknown function that lacked similarity to known protein sequences. Based on its size and position in the CTX genome, we hypothesized that OrfU may function in a manner similar to that of the coliphage fd protein pIII and mediate CTX infection as well as playing a role in CTX assembly and release. Deletion of orfU from CTX dramatically reduced the number of CTX virions detected in supernatants from CTX-bearing cells. This defect was complemented by expression of orfU in trans, thereby confirming a role for this gene in CTX assembly and/or release. To evaluate the requirement for OrfU in CTX infection, we introduced fragments of orfU into gIII in an fd derivative to create OrfU-pIII fusions. While fd is ordinarily unable to infect V. cholerae, an fd phage displaying the N-terminal 274 amino acids of OrfU could infect V. cholerae in a TCP- and TolA-dependent fashion. Since our findings indicate that OrfU functions as the CTX pIII, we propose to rename OrfU as pIII^CTX. Our data also provide new evidence for a conserved pathway for filamentous phage infection.

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