title: Two-tiered biological containment strategy for Lactococcus lactis-based vaccine or immunotherapy vectors.
abstract: The concept of biological containment was developed as a strategy to prevent environmental dissemination of engineered live vaccine or drug delivery vehicles. A mutation in the gene encoding thymidylate synthase (thyA), a key enzyme in the pyrimidine biosynthetic pathway, has previously been shown to limit growth of L. lactis vectors under restrictive conditions. We hypothesized that further mutations in the pyrimidine biosynthetic pathway might enhance the stability and safety of live L. lactis vectors. We show that a double mutation in the genes encoding ThyA and CTP synthase (PyrG) in L. lactis confers double auxotrophy for both thymidine and cytidine. However, the combination of two mutations failed to enhance the biological containment phenotype of the engineered strain. In the absence of thymine/thymidine, the thyA mutant exhibited a strong bactericidal phenotype. However, creation of the double mutant caused the loss of this phenotype, though survival in the mouse GI tract was enhanced. The implications for biological containment of live L. lactis based delivery vectors are discussed.
authors.affiliation: Alimentary Pharmabiotic Centre; University College Cork; Cork, Ireland; Department of Microbiology; University College Cork; Cork, Ireland.
authors.forename: Aurelie
authors.initials: A
authors.lastname: Hanin
authors.affiliation: Alimentary Pharmabiotic Centre; University College Cork; Cork, Ireland; Department of Microbiology; University College Cork; Cork, Ireland.
authors.forename: Eamonn P
authors.initials: EP
authors.lastname: Culligan
authors.affiliation: Alimentary Pharmabiotic Centre; University College Cork; Cork, Ireland; Department of Microbiology; University College Cork; Cork, Ireland.
authors.forename: Pat G
authors.initials: PG
authors.lastname: Casey
authors.affiliation: Department of Pharmaceutical Microbiology; Faculty of Pharmacy; Alexandria University; Alexandria, Egypt.
authors.forename: Mohammed
authors.initials: M
authors.lastname: Bahey-El-Din
authors.affiliation: Alimentary Pharmabiotic Centre; University College Cork; Cork, Ireland; Department of Microbiology; University College Cork; Cork, Ireland.
authors.forename: Colin
authors.initials: C
authors.lastname: Hill
authors.affiliation: Alimentary Pharmabiotic Centre; University College Cork; Cork, Ireland; Department of Microbiology; University College Cork; Cork, Ireland.
authors.forename: Cormac G M
authors.initials: CG
authors.lastname: Gahan
mesh_terms: D000818:Animals; D019731:Carbon-Nitrogen Ligases / Q000172:deficiency; D003264:Containment of Biohazards; D004337:Drug Carriers; D041981:Gastrointestinal Tract / Q000382:microbiology; D007167:Immunotherapy / Q000379:methods*; D013294:Lactococcus lactis / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development*; D051379:Mice; D013940:Thymidylate Synthase / Q000172:deficiency; D014612:Vaccines / Q000008:administration & dosage / Q000009:adverse effects* / Q000235:genetics / Q000276:immunology*
chemical_list: D004337:Drug Carriers; D014612:Vaccines; D013940:Thymidylate Synthase; D019731:Carbon-Nitrogen Ligases; C019951:CTP synthetase
keywords: Lactococcus lactis; biological containment; pyrG; thyA; vaccine