Characterisation of a Temperate Phage Residing in the Ggenome of the Anaerobic Bacteria Fusobacterium nucleatum polymorphum ATCC 10953

Objectives: Characterisation of a prophage residing in the genome of Fusobacterium nucleatum polymorphum ATCC 10953 and an assessment of its presence in periodontal patient plaque DNA. In addition, we are investigating three potential lysin proteins via recombinant production to assess potential antimicrobial activity as therapeutics.
Methods: Mitomycin was used to induce the 10953 prophage with visualisation by TEM. Three prophage genes with potential lysis activity were identified using the PHAge Search Tool (PHAST) server: FNP_1707 (amiC), FNP_1699 and FNP_1700. These genes were commercially synthesised for expression in E. coli. Cloning of the genes and overexpression of its protein were attempted in BL21 and C41. Furthermore, plaque samples from chronic periodontitis patients in Sheffield were screened for F. polymorphum and the occurrence of its prophage using PCR primers.
Results: Despite good expression of amiC-pGEX (FNP_1707 protein), it was found to be mainly insoluble. Cloning and overexpression of FNP_1699 and FNP_1700 are ongoing alongside attempts to further improve production AmiC. 23/45 patients tested screened positively for the presence of Fusobacterium nucleatum polymorphum ATCC10953 strain and its prophage, which represent 405 plaque samples, tested. In addition induction of the prophage has proved inconsistent with visualisation of a tailed phage in some samples.
Conclusions: We have identified that the prophage of FNP10953 is present in a large number of patients, indicating that it is present not only in a lab strain but also in the general population. Attempts to isolate this prophage and its potential lysins are ongoing but present potential novel antimicrobials that would target Fusobacterium nucleatum spp, and might aid in the treatment of gum disease.