Photorhabdus is a virulent pathogen that kills its insect host by overcoming immune responses. The bacterium also secretes a range of antibiotics to suppress the growth of other invading microorganisms. Here we show that Photorhabdus produces a small-molecule antibiotic (E)-1,3-dihydroxy-2-(isopropyl)-5-(2-phenylethenyl)benzene (ST) that also acts as an inhibitor of phenoloxidase (PO) in the insect host Manduca sexta. The Photorhabdus gene stlA encodes an enzyme that produces cinnamic acid, a key precursor for production of ST, and a mutation in stlA results in loss of ST production and PO inhibitory activity, which are both restored by genetic complementation of the mutant and also by supplying cinnamic acid. ST is produced both in vitro and in vivo in sufficient quantities to account for PO inhibition and is the only detectable solvent-extractable inhibitor. A Photorhabdus stlA ؊ mutant is significantly less virulent, proliferates slower within the host, and provokes the formation of significantly more melanotic nodules than wild-type bacteria. Virulence of the stlA ؊ mutant is also rescued by supplying cinnamic acid. The proximate cause of the virulence effect, however, is the inhibition of PO, because the effect of the stlA ؊ mutation on virulence is abolished in insects in which PO has been knocked down by RNA interference (RNAi). Thus, ST has a dual function both as a PO inhibitor to counter host immune reactions and also as an antibiotic to exclude microbial competitors from the insect cadaver.Photorhabdus luminescens ͉ RNA interference ͉ stilbene ͉ virulence

Bacterial pathogens and symbionts must suppress or negate host innate immunity. However, pathogens release conserved oligomeric and polymeric molecules or MAMPs (Microbial Associated Molecular Patterns), which elicit host defenses [1], [2] and [3]. Extracellular polysaccharides (EPSs) are key virulence factors in plant and animal pathogenesis, but their precise function in establishing basic compatibility remains unclear [4], [5], [6] and [7]. Here, we show that EPSs suppress MAMP-induced signaling in plants through their polyanionic nature [4] and consequent ability to chelate divalent calcium ions [8]. In plants, Ca2+ ion influx to the cytosol from the apoplast (where bacteria multiply [4], [5] and [9]) is a prerequisite for activation of myriad defenses by MAMPs [10]. We show that EPSs from diverse plant and animal pathogens and symbionts bind calcium. EPS-defective mutants or pure MAMPs, such as the flagellin peptide flg22, elicit calcium influx, expression of host defense genes, and downstream resistance. Furthermore, EPSs, produced by wild-type strains or purified, suppress induced responses but do not block flg22-receptor binding in Arabidopsis cells. EPS production was confirmed in planta, and the amounts in bacterial biofilms greatly exceed those required for binding of apoplastic calcium. These data reveal a novel, fundamental role for bacterial EPS in disease establishment, encouraging novel control strategies.

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Peptidoglycan (PGN) is a unique and essential structural part of the bacterial cell wall. PGNs from two contrasting Gram-negative plant pathogenic bacteria elicited components characteristic of the innate immune system in Arabidopsis thaliana, such as transcription of the defense gene PR1, oxidative burst, medium alkalinization, and formation of callose. Highly purified muropeptides from PGNs were more effective elicitors of early defense responses than native PGN. Therefore, PGN and its constituents represent a Microbe-Associated Molecular Pattern (MAMP) in plant-bacterial interactions. PGN and muropeptides from aggressive Xanthomonas campestris pv. campestris were significantly more active than those from Agrobacterium tumefaciens, which must maintain host cell viability during infection. The structure of muropeptide components and the distinctive differences are described. Differing defense-eliciting abilities appear to depend on subtle structural differences in either carbohydrate or peptide groups.