Selection of an Attenuated Rifampicin Resistant Mutant of Streptococcus Iniae and Evaluation of its Immunoprotective Effect as Naturally Delivered Vaccines

Streptococcus iniae is a Gram-positive bacterium and a significant pathogen to a wide range of farmed fish. In the present study, we obtained a SF1 derivative, SF1M1, by selecting rifampicin resistance mutants. Compared to the wild type, SF1M1 (i) was slow in growth, (ii) showed less extracellular protease activity and produced a much lower amount of siderophores, and (iii) had a median lethal dose that is more than 100 times higher. In vitro and in vivo studies showed that the attenuated virulence of SF1M1 was stably maintained in the absence of selective pressure. To examine the potential of SF1M1 as a naturally delivered vaccine, Japanese flounder ( Paralichthys olivaceus ) were vaccinated with SF1M1 via bath immersion and oral feeding. At one and two months post-vaccination, the fish were challenged with SF1 or a heterogeneous strain S. iniae 29177, which is of serotype II. The results showed that fish vaccinated with SF1M1 exhibited relative percent survival rates of 54%-70% and produced specific serum antibodies that enhanced complement-mediated bactericidal activity against both SF1 and S. iniae 29177. Taken together, these results indicate that SF1M1 is an effective oral and immersion vaccine that induces protective immunity against serotype I and serotype II S. iniae .