Profiling early infection responses:Pseudomonas aeruginosaeludes host defenses by suppressing antimicrobial peptide gene expression

Abstract

Insights into the host factors and mechanisms mediating the primary host responses after pathogen presentation remain limited, due in part to the complexity and genetic intractability of host systems. Here, we employ the model Drosophila melanogaster to dissect and identify early host responses that function in the initiation and progression of Pseudomonas aeruginosa pathogenesis. First, we use immune potentiation and genetic studies to demonstrate that flies mount a heightened defense against the highly virulent P. aeruginosa strain PA14 when first inoculated with strain CF5, which is avirulent in flies; this effect is mediated via the Imd and Toll signaling pathways. Second, we use wholegenome expression profiling to assess and compare the Drosophila early defense responses triggered by the PA14 vs. CF5 strains to identify genes whose expression patterns are different in susceptible vs. resistant host-pathogen interactions, respectively. Our results identify pathogenesis-and defense-specific genes and uncover a previously undescribed mechanism used by P. aeruginosa in the initial stages of its host interaction: suppression of Drosophila defense responses by limiting antimicrobial peptide gene expression. These results provide insights into the genetic factors that mediate or restrict pathogenesis during the early stages of the bacterial-host interaction to advance our understanding of P. aeruginosa-human infections.innate immunity ͉ pathogenesis ͉ Drosophila melanogaster ͉ immune potentiation T he innate immunity system, which first evolved in lower animals, is ancestral and, unlike adaptive immunity, occurs throughout invertebrate and vertebrate species. Current knowledge of this system remains limited, especially with regard to the host defense mechanisms used upon initial pathogen presentation. Innate immunity must act quickly to mount a first line of defense to hold the pathogen in check before the adaptive response matures (1). Although defense strategies are diverse for different pathogens, many of them are evolutionarily conserved, including production of an array of antimicrobial peptides (AMPs), activation of phagocytic cells, and production of toxic metabolites. AMPs, the best-studied defense effectors, are rapidly elicited after microbe presentation. These ancient weapons play crucial roles in combating microbial infections in invertebrates, vertebrates, and plants (2).Drosophila has emerged as an ideal model organism to study the genetic control of immune recognition and response, because of the high degree of conservation between the fly and mammalian innate immune systems (3), along with its genetic tractability and simplicity. The Drosophila genome contains at least 15 AMP genes that encode both broad-spectrum antibiotic peptides and more specialized activities against Gram-negative bacteria or fungi (4). Expression of many of these defense effectors is mediated via activation of the Toll and͞or Imd innate immunity signaling pathways (5).We (6) and others (7-9) have used Drosophila to explore th...

The Host Defense of Drosophila melanogaster

“…A prior injection with nonpathogenic variants partially protects against a subsequent infection by P. aeruginosa, and this effect is Toll-and Imd-dependent. Mutants deficient for Toll and Imd pathways are more susceptible to P. aeruginosa than are wild-type flies (224,225). This indicates that P. aeruginosa is somewhat sensitive to the fly humoral response, but can ultimately overcome it.…”

“…This indicates that P. aeruginosa is somewhat sensitive to the fly humoral response, but can ultimately overcome it. A microarray analysis indicated that AMP gene expression is delayed when flies are challenged with a pathogenic strain of P. aeruginosa as compared to nonpathogenic bacteria (225). Altogether these data point to the existence of a complex set of strategies used by P. aeruginosa to suppress and escape the immune response.…”

Bacterial strategies to overcome insect defences

“…The humoral response can also be suppressed by repressing AMP gene expression through a mechanism that remains to be characterized 57 . In a similar way, Pseudomonas aeruginosa can suppress D. melanogaster defence responses by limiting AMP gene expression when injected into the fly haemolymph 58 , using a mechanism that also remains to be characterized. Additionally, X. nematophila and Photorhabdus temperata can inhibit phospholipase A2 (PLA2) 59 , which catalyses the first step of eicosanoid biosynthesis.…”

DrosophilaRel proteins are central regulators of a robust, multi-organ immune network

“…Although we previously demonstrated that Dif dl mutant larvae induce all of the major antimicrobial peptide genes (Matova and Anderson, 2006), Defensin (Def) was the only antimicrobial peptide whose induction after septic injury was significantly reduced in the double mutants, and was expressed at about 30% of the levels present in the wild type. Among the antimicrobial peptides, Def appears to have the most potent activity: constitutive expression of Def confers resistance to Gram-positive bacteria in immunocompromised adults (Tzou et al, 2002) and to Pseudomonas, a Gram-negative bacterium in wild-type adult Drosophila (Apidianakis et al, 2005). To test whether Def could protect Dif dl larvae from infection, we used the fat body-specific Lsp2-Gal4 driver and the hemocyte-specific He-Gal4 driver to express UAS-Def.…”