Construction of a reporter system to study Burkholderia mallei type III secretion and identification of the BopA effector protein function in intracellular survival

Abstract

Burkholderia mallei, the aetiological agent of glanders disease, is a Gram-negative facultative intracellular bacterium. Despite numerous studies, the detailed mechanism of its pathogenesis is almost unknown. The presence of a type III secretion system (TTSS) is one of the known mechanisms associated with virulence. An intact TTSS indicates that B. mallei is able to secrete proteins in response to different environmental conditions, which could play an important role in pathogenesis. Therefore, characterization of the TTSS and identification of the secreted proteins associated with bacterial pathogenesis could provide crucial information for the development of a candidate vaccine. In the current study, we used an enzymatic reporter system to establish some of the conditions enabling TTS. Construction of the TTSS bopA mutant revealed that BopA is important for B. mallei invasion and intracellular survival. Overall, our study elucidates how BopA can aid in the optimization of TTS and defines the function of TTS effectors in bacterial intracellular survival and invasion.

Protective response to subunit vaccination against intranasal Burkholderia mallei and B. pseudomallei challenge

“…Therefore, we assessed flagellin as a potential candidate for inclusion in a Burkholderia vaccine and found it unsuitable (our unpublished data). In contrast, all of the current evidence indicates that other surface-expressed or secreted proteins are immunogenic and structural similarity exists between the proteins in B. pseudomallei and B. mallei (10–11). In this study, we aimed to identify Burkholderia protective proteins that could be administered in vaccines to generate cross-protective immunity against both B. mallei and B. pseudomallei .…”

Analysis of the Prevalence, Secretion and Function of a Cell Cycle-Inhibiting Factor in the Melioidosis Pathogen Burkholderia pseudomallei

“…Only a small number of effectors have been confirmed to be substrates of the Bsa T3SS in B. pseudomallei , including BopC [4] and the guanine nucleotide exchange factor BopE [5]. A further candidate effector (BopA) was demonstrated to be Type III secreted in a surrogate bacterial host [6] and to interfere with LC3-associated phagocytosis [7]. A homologue of an E. coli Type III secreted effector termed Cif (cycle-inhibiting factor) was identified in B. pseudomallei and exhibits 21% amino acid identity and 40% similarity [8], but no evidence has yet been presented that it is secreted via the Bsa apparatus or that it influences pathogenesis during melioidosis.…”

Type III Secretion in the Melioidosis Pathogen Burkholderia pseudomallei

“…The B. pseudomallei homolog of IpgA is BicP, which co-purifies with BopA and helps to prevent its degradation, indicating that it is the chaperone for BopA (Kayath et al, 2010). BopA is secreted by T3SS-3 in a bsaZ -dependant manner (Vander Broek et al, 2015) and the first 58 amino acids of B. mallei BopA fused to the Yersinia enterolitica phospholipase YplA, has been shown to be secreted in a surrogate enteropathogenic E. coli host (Whitlock et al, 2008). …”

“…Another study demonstrated that BopA is important for escape of the bacterium from the phagosome (Gong et al, 2011). A B. mallei ATCC 23344 bopA mutant demonstrated reduced intracellular survival in J774A.1 cells (Whitlock et al, 2008). Interestingly, in the murine alveolar macrophage cell line MH-S, the same B. mallei bopA mutant exhibited increased intracellular survival when compared to the isogenic parental strain, indicating that different cell types may rely on different mechanisms to control intracellular B. mallei (Whitlock et al, 2009).…”