Brucella melitensis M5-90Δbp26 as a potential live vaccine that allows for the distinction between natural infection and immunization

Brucella is Gram-negative intracellular bacterial pathogen that infects humans and animals and contributes to great economic losses in developing countries. Presently, live attenuated Brucella vaccines (Brucella melitensis M5-90) are the most effective means of brucellosis control and prevention in animals. However, these vaccines have several drawbacks, such as an inability to distinguish between a natural infection and immunization and an association with abortions in pregnant animals. Therefore, this study constructed a Brucella M5-90 Delta bp26 mutant and evaluated its virulence. The survival of the M5-90 Delta bp26 mutant was attenuated in human placenta trophoblastic 8 cells (HPT-8 cells) and in BALB/c mice, with a high immunoprotectivity noted in mice. Furthermore, safety tests showed that the M5-90 Delta bp26 mutant was less virulent than the M5-90 vaccine strain. Additionally, an indirect enzyme-linked immunosorbent assay (ELISA) screening was shown to detect the presence of Brucella protein 26 (BP26) with high sensitivity, with M5-90 Delta bp26 inoculation accompanied with a lack of BP26 expression, and was further confirmed by western blotting. Together, the M5-90 Delta bp26 mutant and the indirect ELISA can be employed to distinguish vaccinated livestock from infected animals.