Spontaneous Deletion of a 209-Kilobase-Pair Fragment from the Escherichia coli Genome Occurs with Acquisition of Resistance to an Assortment of Infectious Phages

• Published in: Applied and Environmental Microbiology Vol. 74; no. 14; pp. 4256 - 4263
• Main Authors: Tanji, Yasunori, Hattori, Kenji, Suzuki, Kohichi, Miyanaga, Kazuhiko
• Format: Journal Article
• Language: English
• Published: Washington, DC American Society for Microbiology 01.07.2008; American Society for Microbiology (ASM)
• Subjects: Bacterial Outer Membrane Proteins - analysis; Base Sequence; Biological and medical sciences; Cell culture; Chemostats; Coliphages - pathogenicity; DNA, Bacterial - genetics; Drug resistance; E coli; Escherichia coli; Escherichia coli - genetics; Escherichia coli - growth & development; Escherichia coli - virology; Escherichia coli Proteins - analysis; Fundamental and applied biological sciences. Psychology; Genes, Bacterial; Genetic Complementation Test; Genetic recombination; Genome, Bacterial; Genomics; Microbial Ecology; Microbiology; Mutation; Plasmids; Porins - analysis; Proteins; Recombinant Proteins - analysis; Sequence Analysis, DNA; Sequence Deletion; Sewage - virology; Viral Plaque Assay; Virus; Infection; Resistance; Escherichia coli; Deletion; Bacteria; Genome; Fragment; Enterobacteriaceae; Phage
• ISSN: 0099-2240; 1098-5336; 1098-6596
• DOI: 10.1128/AEM.00243-08

To breed resistance to an assortment of infectious phages, continuous cultures of Escherichia coli JM109 grown in a chemostat were exposed to phage mixtures prepared from sewage influent. Four sequential chemostat-grown cultures were each infected with a different phage mixture. At the end of a chemostat run, one phage-resistant colony was isolated and used to inoculate the subsequent culture. This process was repeated, and increased phage resistance of the input bacterial strain resulted from the successive challenges with different phage cocktails. Multiple mutations apparently accumulated progressively. A mutant isolated at the end of the four runs, designated D198, showed resistance to 38 of 40 phages that infect the parent strain, JM109. D198 produced less outer membrane protein C (OmpC) than JM109. However, restoration of the OmpC protein by plasmid-mediated complementation did not completely restore the susceptibility of D198 to the 38 phages. Therefore, alterations beyond the level of OmpC protein production contribute to the phage resistance of D198. PCR-based genetic analysis revealed that D198 has a genome that is 209 kbp (about 200 genes) smaller than JM109. The deletion includes the chromosomal section from ompC to wbbL that encodes the rhamnosyl transferase involved in lipopolysaccharide biosynthesis. Strains D198 and JM109 were comparable in their growth characteristics and their abilities to express a recombinant protein.