Generation of small colony variants in biofilms by Escherichia coli harboring a conjugative F plasmid

A conjugative F plasmid induces mature biofilm formation by Escherichia coli by promoting F-pili-mediated cell-cell interactions and increasing the expression of biofilm-related genes. We herein demonstrated another function for the F plasmid in E. coli biofilms; it contributes to the emergence of genetic and phenotypic variations by spontaneous mutations. Small colony variants (SCVs) were more frequently generated in a continuous flow-cell biofilm than in the planktonic state of E. coli harboring the F plasmid. E. coli SCVs represented typical phenotypic changes such as slower growth, less biofilm formation, and greater resistance to aminoglycoside antibiotics than the parent strain. Genomic and complementation analyses indicated that the small colony phenotype was caused by the insertion of Tn1000, which was originally localized in the F plasmid, into the hemB gene. Furthermore, the Tn1000 insertion was removed from hemB in the revertant, which showed a normal colony phenotype. This study revealed that the F plasmid has the potential to increase genetic variations not only by horizontal gene transfer via F pili, but also by site-specific recombination within a single cell.