<i>Salmonella</i> effectors within a single pathogenicity island are differentially expressed and translocated by separate type III secretion systems

<jats:title>Summary</jats:title><jats:p>Pathogenicity islands (PAIs) are large DNA segments in the genomes of bacterial pathogens that encode virulence factors. Five PAIs have been identified in the Gram‐negative bacterium <jats:italic>Salmonella enterica</jats:italic>. Two of these PAIs, <jats:italic>Salmonella</jats:italic> pathogenicity island (SPI)‐1 and SPI‐2, encode type III secretion systems (TTSS), which are essential virulence determinants. These ‘molecular syringes’ inject effectors directly into the host cell, whereupon they manipulate host cell functions. These effectors are either encoded with their respective TTSS or scattered elsewhere on the <jats:italic>Salmonella</jats:italic> chromosome. Importantly, SPI‐1 and SPI‐2 are expressed under distinct environmental conditions: SPI‐1 is induced upon initial contact with the host cell, whereas SPI‐2 is induced intracellularly. Here, we demonstrate that a single PAI, in this case SPI‐5, can encode effectors that are induced by distinct regulatory cues and targeted to different TTSS. SPI‐5 encodes the SPI‐1 TTSS translocated effector, SigD/SopB. In contrast, we report that the adjacently encoded effector PipB is part of the SPI‐2 regulon. PipB is translocated by the SPI‐2 TTSS to the <jats:italic>Salmonella</jats:italic>‐containing vacuole and <jats:italic>Salmonella</jats:italic>‐induced filaments. We also show that regions of SPI‐5 are not conserved in all <jats:italic>Salmonella</jats:italic> spp. Although <jats:italic>sigD/sopB</jats:italic> is present in all <jats:italic>Salmonella</jats:italic> spp., <jats:italic>pipB</jats:italic> is not found in <jats:italic>Salmonella bongori</jats:italic>, which also lacks a functional SPI‐2 TTSS. Thus, we demonstrate a functional and regulatory cross‐talk between three chromosomal PAIs, SPI‐1, SPI‐2 and SPI‐5, which has significant implications for the evolution and role of PAIs in bacterial pathogenesis.</jats:p>