There is an urgent need to combat infections caused by Burkholderia pseudomallei, the bacterial agent responsible for the disease melioidosis. Melioidosis is a health threat in tropical endemic regions of the world and is now recognized as an emergent disease in areas considered non-endemic. In addition to causing acute infections that are considered lethal if left untreated, the bacteria can persist in the host and produce very difficult to treat latent infections. Therefore, our long-term goal is to develop effective treatments against melioidosis and prevent latent or chronic disease outcomes. In this project, we will evaluate the function of a subset of toxin-antitoxin systems that have been identified as one of the bacterial switches that sense stress and activate a persistence phenotype. We will determine their association with bacterial persistence both in vitro and in vivo. Our central hypothesis is that selective toxins are upregulated in vivo in response to specific conditions, and loss of their function results in reduced host-associated persistence. It is important to point out that current antibiotics used against B. pseudomallei generate persister frequencies of up to 50%. Because current treatments last months due to the propensity of B. pseudomallei to generate latent infections, the study of toxin-antitoxin systems can help identify new drug targets and therefore improve antibiotic efficacy. Our proposed experimental approach will focus on two specific aims: first, we will evaluate the role of selected toxins in survival and induction of a persistent stage during B. pseudomallei infection of macrophages because this cell type likely facilitates latent infections; secondly, we will evaluate the contribution of the toxins in B. pseudomallei's ability to persist during host-associated in vivo conditions. Our proposal is significant because it is expected to elucidate the role of the specific toxins that have a strong association with the persistence phenotype, and this might result in the development of alternative therapeutic approaches to prevent latent infections caused by B. pseudomallei, a CDC Category B Tier 1 Select Biothreat Agent and an understudied emerging pathogen.
A novel bioinformatics/big data analysis approach will be utilized to determine the participation of toxin-antitoxin systems for the development of a persistence state during infection by the pathogen Burkholderia pseudomallei. The approach has allowed us to identify those toxin-antitoxin systems that are expressed during host infection and to use isogenic mutants to define their contribution in vitro, during infection of macrophages, and in vivo, using a murine model of persistent melioidosis.
