Bacteriophages such as the Escherichia coli phage Lambda have been studied extensively for several decades as model systems for the study of such topics as gene regulation, host-virus interactions, and macromolecular assembly. We have taken advantage of the base of information about Lambda and related phages for two purposes: (1) Detection of mutagens and carcinogens: We developed a novel assay for the detection of mutations in a Lambda transgene contained in mice. The assay selects for phage containing forward mutations only in the Lambda cII gene, using a mutant (hfl) Escherichia colihost. In addition to the relative ease of direct selection, the sensitivity of this assay for both spontaneous and chemically induced mutation was comparable to the widely used mutational target gene lacI. Moreover, our assay costs 80 times less to use than the lacIsystem. Our cII assay system is now being used worldwide as a replacement for the lacIsystem. In a collaboration with Dr. Glenn Merlino, the system is now being used to study the role of mutagenesis in cancer development. It appears that expression of the new gene causes a change in the spectrum of mutations. This project has now been completed, published, and terminated. (2) Bacteriophage therapy: The increased prevalence of multidrug-resistant bacterial pathogens motivated us to attempt to enhance the therapeutic efficacy of bacteriophages. The therapeutic application of phages as antibacterial agents was impeded by the capacity of mammalian host defense systems to remove phage particles from the circulatory system. In our studies involving bacteremic mice, to reduce phage elimination by the host defense system, we previously isolated E. coli phage Lambda and Salmonella phage P22 mutants able to remain in the circulatory system for longer periods of time and also have greater capability as antibaceterial agents in animals infected with lethal doses of bacteria. The mutations in Lambda are in the gene for the major capsid protein. In a collaboration with Dr. Carl Merril and Dr. Dean Scholl, we have continued our phage therapy project with other pathogenic E. coli strains by isolating and characterizing suitable phages. A virulent double-stranded DNA bacteriophage, f K1-5, has been isolated and found to be capable of infecting pathogenic Escherichia coli strains that possess either the K1 or the K5 polysaccharide capsule. Electron micrographs showed that the virion consists of a small icosohedral head with short tail spikes, similar to members of the Podoviridae family. DNA sequence analysis of the region encoding the tail fiber protein showed two open reading frames encoding previously characterized hydrolytic phage tail fiber proteins. The first in the K5 lyase protein gene of Phi K5, which allows this phage to specifically infect K5 E. coli strains. A second open reading frame encodes a protein almost identical in amino acid sequence to the N-acetylneuraminidase (endosialidase) protein of Phi K1E, which allows this phage to specifically infect K1 strains of E. Coli. We provide experimental evidence that mature phage particles contain both tail fiber proteins, and mutational analysis indicates that each protein can be independently inactivated. A comparison of the tail gene regions of Phi K5, Phi K1E, and Phi K1-5 shows that the genes are arranged in a modular or cassette configuration and suggests that this family of phages can broaden host range by horizontal gene transfer.
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