The autotransporter protein BatA is a protective antigen against lethal aerosol infection with Burkholderia mallei and Burkholderia pseudomallei.

BACKGROUND: Burkholderia mallei and Burkholderia pseudomallei are the causative agents of glanders and melioidosis, respectively. There is no vaccine to protect against these highly-pathogenic and intrinsically antibiotic-resistant bacteria, and there is concern regarding their use as biological warfare agents. For these reasons, B. mallei and B. pseudomallei are classified as Tier 1 organisms by the U.S. Federal Select Agent Program and the availability of effective countermeasures represents a critical unmet need. METHODS: Vaccines (subunit and vectored) containing the surface-exposed passenger domain of the conserved Burkholderia autotransporter protein BatA were administered to BALB/c mice and the vaccinated animals were challenged with lethal doses of wild-type B. mallei and B. pseudomallei strains via the aerosol route. Mice were monitored for signs of illness for a period of up to 40 days post-challenge and tissues from surviving animals were analyzed for bacterial burden at study end-points. RESULTS: A single dose of recombinant Parainfluenza Virus 5 (PIV5) expressing BatA provided 74% and 60% survival in mice infected with B. mallei and B. pseudomallei, respectively. Vaccination with PIV5-BatA also resulted in complete bacterial clearance from the lungs and spleen of 78% and 44% of animals surviving lethal challenge with B. pseudomallei, respectively. In contrast, all control animals vaccinated with a PIV5 construct expressing an irrelevant antigen and infected with B. pseudomallei were colonized in those tissues. CONCLUSION: Our study indicates that the autotransporter BatA is a valuable target for developing countermeasures against B. mallei and B. pseudomallei and demonstrates the utility of the PIV5 viral vaccine delivery platform to elicit cross-protective immunity against the organisms.

Use of Immunohistochemistry to Demonstrate In Vivo Expression of the Burkholderia mallei Virulence Factor BpaB During Experimental Glanders.

Burkholderia mallei causes the highly contagious and debilitating zoonosis glanders, which infects via inhalation or percutaneous inoculation and often culminates in life-threatening pneumonia and sepsis. In humans, glanders is difficult to diagnose and requires prolonged antibiotic therapy with low success rates. No vaccine exists to protect against B. mallei, and there is concern regarding its use as a bioweapon. The authors previously identified the protein BpaB as a potential target for devising therapies due to its role in adherence to host cells and the formation of biofilms in vitro and its contribution to pathogenicity in a mouse model of glanders. In the present study, the authors developed an immunostaining approach to probe tissues of experimentally infected animals and demonstrated that BpaB is produced exclusively in vivo by wild-type B. mallei in target organs from mice and marmosets. They detected the expression of BpaB by B. mallei both extracellularly and within macrophages, neutrophils, and epithelial cells in respiratory tissues (7/10 marmoset; 2/2 mouse). The authors also noted the intracellular expression of BpaB by B. mallei in macrophages in the regional lymph nodes of mice (2/2 tissues) and MALT of marmosets (4/5 tissues). It is interesting that B. mallei bacteria infecting distal organs did not express BpaB (2/2 mice; 3/3 marmosets), suggesting that the protein is not necessary for bacterial fitness in these anatomic locations. These findings underscore the value of BpaB as a target for developing medical countermeasures and provide insight into its role in pathogenesis.

Antibodies against In Vivo-Expressed Antigens Are Sufficient To Protect against Lethal Aerosol Infection with Burkholderia mallei and Burkholderia pseudomallei.

Burkholderia mallei, a facultative intracellular bacterium and tier 1 biothreat, causes the fatal zoonotic disease glanders. The organism possesses multiple genes encoding autotransporter proteins, which represent important virulence factors and targets for developing countermeasures in pathogenic Gram-negative bacteria. In the present study, we investigated one of these autotransporters, BatA, and demonstrate that it displays lipolytic activity, aids in intracellular survival, is expressed in vivo, elicits production of antibodies during infection, and contributes to pathogenicity in a mouse aerosol challenge model. A mutation in the batA gene of wild-type strain ATCC 23344 was found to be particularly attenuating, as BALB/c mice infected with the equivalent of 80 median lethal doses cleared the organism. This finding prompted us to test the hypothesis that vaccination with the batA mutant strain elicits protective immunity against subsequent infection with wild-type bacteria. We discovered that not only does vaccination provide high levels of protection against lethal aerosol challenge with B. mallei ATCC 23344, it also protects against infection with multiple isolates of the closely related organism and causative agent of melioidosis, Burkholderia pseudomallei Passive-transfer experiments also revealed that the protective immunity afforded by vaccination with the batA mutant strain is predominantly mediated by IgG antibodies binding to antigens expressed exclusively in vivo Collectively, our data demonstrate that BatA is a target for developing medical countermeasures and that vaccination with a mutant lacking expression of the protein provides a platform to gain insights regarding mechanisms of protective immunity against B. mallei and B. pseudomallei, including antigen discovery.