Smooth-type Lipopolysaccharide Research Articles

Extraction of lipopolysaccharide from fish-derived Aeromonas hydrophila and their immunostimulatory effect on Qihe crucian carp

Purified lipopolysaccharide (LPS) was first obtained from fish-derived Aeromonas hydrophila (A. hydrophila) by hot phenol-water combined with enzymatic hydrolysis. Subsequently, different concentrations of LPS (0, 0.2, 0.4, 0.8, and 1.2 mg/mL) were injected intraperitoneally into Qihe crucian carp (Carassius auratus var. Qihe) with a dose of 3 μL/g body weight to evaluated their immunostimulatory activity by detecting the expression of immune-related genes in the spleen, head kidney, and hepatopancreas. The results showed that the average extraction rate of LPS was about 1.063% based on the precipitation weight of wet bacteria, of which the polysaccharide content was 9.39%. The silver staining band of LPS after SDS-PAGE electrophoresis showed a ladder distribution typically characterized by smooth-type LPS. Protein and nucleic acid electrophoresis results showed very low amounts of residual protein and nucleic acid, and high-performance liquid chromatography (HPLC) results showed that the purity of the extracted A. hydrophila LPS was similar to that of high-purity commercial LPS. Pro-inflammatory cytokine genes such as IL-1, IL-8, TNF-α, and IL-6 were more sensitive to LPS stimulation than anti-inflammatory factors such as IL-10 and TGF-β. After 2 h of injection, their expression levels in the spleen, head kidney, and hepatopancreas were significantly increased (P < 0.05), with the 0.8 and 1.2 mg/mL injection groups showing the most significant increase. The gene expression of IL-10 and TGF-β showed slight fluctuations in response to A. hydrophila LPS stimulation (P < 0.05). In conclusion, high-purity LPS was extracted from A. hydrophila derived from fish by modified hot phenol-water method, it demonstrating that LPS product could significantly regulate the innate immunity of Qihe crucian carp, with the most effective vaccination dose being 2 mg/kg/BW. These findings provide useful information for future research on endotoxin vaccines.

A chronic strain of the cystic fibrosis pathogen Pandoraea pulmonicola expresses a heterogenous hypo-acylated lipid A

Pandoraea sp. is an emerging Gram-negative pathogen in cystic fibrosis causing severe and persistent inflammation and damage of the lungs. The molecular mechanisms underlying the high pathogenicity of Pandoraea species are still largely unknown. As Gram-negatives, Pandoraea sp. express lipopolysaccharides (LPS) whose recognition by the host immune system triggers an inflammatory response aimed at the bacterial eradication from the infected tissues. The degree of the inflammatory response strongly relies on the fine structure of the LPS and, in particular, of its glycolipid moiety, i.e. the lipid A. Here we report the structure of the lipid A isolated from the LPS of a chronic strain of P. pulmonicola (RL 8228), one of the most virulent identified so far among the Pandoraea species. Our data demonstrated that the examined chronic strain produces a smooth-type LPS with a complex mixture of hypoacylated lipid A species displaying, among other uncommon characteristics, the 2-hydroxylation of some of the acyl chains and the substitution by an additional glucosamine on one or both the phosphate groups.

Identification of MAMP-Responsive Plasma Membrane-Associated Proteins in Arabidopsis thaliana Following Challenge with Different LPS Chemotypes from Xanthomonas campestris.

Lipopolysaccharides (LPS) are recognized as microbe-associated molecular patterns (MAMPs) responsible for eliciting defense-related responses and while the effects have been well-documented in mammals, there is a lack of knowledge regarding the mechanism of perception in plant systems and recognized structural moieties within the macromolecular lipoglycan structure. Thus, identification of the LPS plasma membrane (PM) receptor(s)/receptor complex in Arabidopsis thaliana through proteomics will contribute to a deeper understanding of induced defense responses. As such, structurally characterized LPS chemotypes from Xanthomonas campestris pv. campestris (Xcc) wild-type 8004 (prototypical smooth-type LPS) and mutant 8530 (truncated core with no O–chain) strains were utilized to pre-treat A. thaliana plants. The associated proteomic response/changes within the PM were compared over a 24 h period using mass spectrometry-based methodologies following three variants of LPS-immobilized affinity chromatography. This resulted in the identification of proteins from several functional categories, but importantly, those involved in perception and defense. The distinct structural features between wild-type and mutant LPS are likely responsible for the differential changes to the proteome profiles, and many of the significant proteins were identified in response to the wild-type Xcc LPS where it is suggested that the core oligosaccharide and O-chain participate in recognition by receptor-like kinases (RLKs) in a multiprotein complex and, notably, varied from that of the mutant chemotype.

Inhibitory effects of a novel cationic dodecapeptide [CL(14–25)] derived from cyanate lyase of rice on endotoxic activities of LPSs from Escherichia coli and periodontopathic Aggregatibacter actinomycetemcomitans

ObjectiveCL(14–25), a dodecapeptide of cyanate lyase from rice, is a novel cationic α-helical antimicrobial peptide. In this study, we examined inhibitory ability of CL(14–25) against endotoxic activities of lipopolysaccharides (LPSs) from Escherichia coli and periodontal pathogenic Aggregatibacter actinomycetemcomitans. MethodsEndotoxin-neutralizing activity of CL(14–25) was evaluated by inhibition to induction of cytokine and nitric oxide in human aortic endothelial cells (HAECs) and RAW264 mouse macrophage cells, respectively. Protective effect of CL(14–25) was determined in mice against lethal toxicity of LPS. ResultsIL-6 in HAECs was induced by stimulation with LPS preparations of A. actinomycetemcomitans and E. coli tested in this study, and addition of CL(14–25) to the medium caused inhibition of their induction in a dose-dependent manner. CL(14–25) inhibited NO induction in RAW264 cells by a smooth type LPS of E. coli O55:B5 and an Rc type LPS of E. coli J5 as well as lipid A of E. coli R515 in a dose-dependent manner. Simultaneous injection of E. coli O55:B5 LPS and CL(14–25) in BALB/c mice resulted in prevention of lethal toxicity of the former. The results of a Limulus amebocyte lysate assay and surface plasmon resonance analysis of interaction between CL(14–25) and E. coli LPS or lipid A showed that CL(14–25) specifically binds to a lipid A moiety of LPS. ConclusionThe results of present study suggest that CL(14–25) has a potential to be used as a nutraceutical agent for periodontal therapy.

Presumable role of outer membrane proteins of Salmonella containing sialylated lipopolysaccharides serovar Ngozi, sv. Isaszeg and subspecies arizonae in determining susceptibility to human serum.

BackgroundThe O48 group comprises Salmonella bacteria containing sialic acid in the lipopolysaccharide (LPS). Bacteria with sialylated surface structures are described as pathogens that avoid immunological response of the host by making similar their surface antigens to the host’s tissues (molecular mimicry). It is known that the smooth-type LPS of Salmonella enterica and outer membrane proteins (OMP) PgtE, PagC and Rck mediate serum resistant phenotype by affecting complement system (C). The aim of this study was to investigate C3 component activation by Salmonella O48 LPS and OMP.FindingsIn the present study, we examined C3 component deposition on the three Salmonella O48 strains: S. enterica subspecies enterica serovar Ngozi, S. enterica subsp. enterica sv. Isaszeg, and S.enterica subsp. arizonae containing sialic acid in the O-specific part of LPS. The greatest C3 deposition occurred on Salmonella sv. Isaszeg cells (p < 0.005) as well as on their LPS (low content of sialic acid in LPS) (p < 0.05) after 45 min of incubation in 50% human serum. Weaker C3 deposition ratio on the Salmonella sv. Ngozi (high content of sialic acid in LPS) and Salmonella subsp. arizonae (high content of sialic acid in LPS) cells correlated with the lower C3 activation on their LPS. Immunoblotting revealed that OMP isolated from the tested strains also bound C3 protein fragments.ConclusionsWe suggest that activation of C3 serum protein is dependent on the sialic acid contents in the LPS as well as on the presence of OMP in the range of molecular masses of 35–48 kDa.

Structure of the O-antigen polysaccharide present in the lipopolysaccharide ofCronobacter dublinensis(subspecieslactaridiorlausannensis) HPB 3169

Cronobacter dublinensis (formerly Enterobacter sakazakii) HPB 3169 is a pathogenic Gram-negative bacterium that produces a smooth-type lipopolysaccharide in which the antigenic O-polysaccharide component was determined to be a repeating pentasaccharide unit composed of L-rhamnose; 2-acetamido-2-deoxy-D-glucose; 3,6-dideoxy-3-(R)-3-hydroxybutyramido-D-glucose; and 3-deoxy-manno-oct-2-ulosonic acid in the respective molar ratio 2:1:1:1. Chemical and 2D NMR analyses of the O-polysaccharide and a pentasaccharide derived by the mild acid hydrolysis of the ketosyl linkage of the Kdo (3-deoxy-D-manno-2-octulosonic acid) residue in the O-polysaccharide established that the O-antigen is a high molecular mass unbranched polymer of a repeating pentasaccharide unit and has the structure [see formula in text] where Bu is a (R)-3-hydroxybutanoyl substituent. The O-antigen is structurally similar to that of the recently reported Cronobacter sakazakii strain G706 (designated as serotype O5), except that in strain G706 the d-Qui3N is in its N-acetyl form, in contrast to its presence as a 3-deoxy-3-(R)-3-hydroxybutyramido derivative in the C. sakazakii HPB 3169 strain O-antigen.

Characterization of the lipopolysaccharide O-antigen of Cronobacter turicensis HPB3287 as a polysaccharide containing a 5,7-diacetamido-3,5,7,9-tetradeoxy- d- glycero- d- galacto-non-2-ulosonic acid (legionaminic acid) residue

Cronobacter turicensis, previously known as Enterobacter sakazakii, is a Gram-negative opportunistic food-borne pathogen that has been reported as a cause of life-threatening neonatal infections. From chemical and physical analyses involving composition analysis, methylation, two-dimensional high-resolution nuclear magnetic resonance, and mass spectrometry methods, the antigenic O-polysaccharide in the smooth-type lipopolysaccharide of C. turicensis (strain HPB 3287) was determined to be a high molecular mass polymer of a repeating pentasaccharide unit composed of d-galactose, d-glucose, 2-acetamido-2-deoxy- d-galactose, and 5,7-diacetamido-3,5,7,9-tetradeoxy- d- glycero- d- galacto-non-2-ulosonic acid (legionaminic acid), in a molar ratio 2:1:1:1, and having the structure: ▪

The lipid A of Burkholderia multivorans C1576 smooth-type lipopolysaccharide and its pro-inflammatory activity in a cystic fibrosis airways model

Cystic fibrosis is an autosomal recessive disorder and it is characterised by chronic bacterial airway infection which leads to progressive lung deterioration, sometimes with fatal outcome. Burkholderia multivorans and Burkholderia cenocepacia are the species responsible for most of the infections of cystic fibrosis patients. Lipopolysaccharide endotoxins (LPSs) are among the foremost factors of pathogenesis of Gram-negative infection and, in particular, lipid A is the endotoxic portion of LPS responsible for eliciting host innate immune response. In this work, the complete primary structure of the lipid A from B. multivorans C1576 has been defined and, further, its pro-inflammatory activity in a cystic fibrosis airways model is shown. The structure of B. multivorans lipid A was attained by chemical, mass spectrometry and nuclear magnetic resonance analyses whereas its biological activity was assessed on the intestinal epithelial cell line CACO-2 cells, on the airway epithelial IB3-1 cells, carrying the ΔF508/W1282X CFTR mutation and on an ex vivo model of culture explants of nasal polyps.

Isolation of smooth‐type lipopolysaccharides to electrophoretic homogeneity

The high structural heterogeneity of smooth-type lipopolysaccharides (LPS) enormously complicates the isolation of their constituent molecular species. Proof of concept is given here on the feasibility of using preparative slab-PAGE to isolate highly homogeneous smooth-type LPS glycoforms. LPS species (from 3.6 to 14.2 kDa) from Escherichia coli K-235 were separated by preparative slab-PAGE and recovered by utilizing the combined on-gel LPS reverse staining, extrusion, and passive elution techniques. As a result, 15 electrophoretically pure LPS fractions were obtained. The LPS content in the recovered fractions ranged from 280 ng (intermediate mobility glycoforms) to 411 mug (highest mobility glycoforms). The quantities of LPS fractions were sufficient to allow quantitation of the Limulus amebocyte lysate (LAL) activities of these distinct-molecular-mass LPS species, in the range from (1.1 +/- 0.1)x10(3) to (8.7 +/- 0.3)x10(5) endotoxin units (EU)/mL, by standard LAL assay. We have thus definitively demonstrated that slab-PAGE may be a suitable platform to more selectively purify individual glycoform fractions from smooth-type LPS.

Preparation and characterization of truncated human lipopolysaccharide-binding protein in Escherichia coli

Lipopolysaccharide (LPS) is a component of the outer membrane of Gram-negative bacteria, and is the causative agent of endotoxin shock. LPS induces signal transduction in immune cells when it is recognized by the cell surface complex of toll-like receptor 4 (TLR4) and MD-2. The complex recognizes the lipid A structure in LPS, which is buried in the membrane of the outer envelope. To present the Lipid A structure to the TLR4/MD-2, processing of LPS by LPS-binding protein (LBP) and CD14 is required. In previous studies, we expressed recombinant proteins of human MD-2 and CD14 as fusion proteins with thioredoxin in Escherichia coli, and demonstrated their specific binding abilities to LPS. In this study, we prepared a recombinant fusion protein containing 212 amino terminal residues of human LBP (HLB212) by using the same expression system. The recombinant protein expressed in E. coli was purified as a complex form with host LPS. The binding was not affected by high concentrations of salt, but was prevented by low concentrations of various detergents. Both rough-type LPS lacking the O antigen and smooth-type LPS with the antigen bound to HLBP212. Therefore, oligosaccharide repeats appeared to be unnecessary for the binding. A nonpathogenic penta-acylated LPS also bound to HLBP212, but the binding was weaker than that of the wild type. The hydrophobic interaction between the LBP and acyl chains of lipid A appears to be important for the binding. The recombinant proteins of LPS-binding molecules would be useful for analyzing the defense mechanism against infections.

Liver argininosuccinate synthase binds to bacterial lipopolysaccharides and lipid A and inactivates their biological activities

The liver is known to clear and detoxify circulating lipopolysaccharide (LPS). To characterize the molecules involved in this process in the liver, we attempted to purify mouse liver protein(s) that can interact with lipid A, a biologically active portion of LPS. By partially purifying the inactivating activity against a synthetic lipid A analog, we observed the enrichment of a 45-kDa protein in the active fractions. The internal amino acid sequences of the protein were identical with those of argininosuccinate synthase (EC 6.3.4.5). To examine whether argininosuccinate synthase can interact with lipid A, we purified the enzyme from mouse liver and found the co-elevation of the specific enzyme activity and specific lipid A-inactivating activity, indicating that argininosuccinate synthase is the major lipid A-interacting protein in liver. Argininosuccinate synthase also inhibited the biological activities (macrophage activation and Limulus test) of natural lipid A and rough-type LPS but not smooth-type LPS. The enzyme activity was inhibited by lipid A and rough-type LPS and also by smooth-type LPS. Native gel electrophoresis of a mixture of argininosuccinate synthase and LPS and immunoprecipitation of a mixture of argininosuccinate synthase and [(3)H]-LPS with anti-argininosuccinate synthase antiserum showed that argininosuccinate synthase stably bound lipid A and LPS. These findings, together, indicate that argininosuccinate synthase can effectively bind LPS in the liver.

Liver argininosuccinate synthase binds to bacterial lipopolysaccharides and lipid A and inactivates their biological activities

The liver is known to clear and detoxify circulating lipopolysaccharide (LPS). To characterize the molecules involved in this process in the liver, we attempted to purify mouse liver protein(s) that can interact with lipid A, a biologically active portion of LPS. By partially purifying the inactivating activity against a synthetic lipid A analog, we observed the enrichment of a 45-kDa protein in the active fractions. The internal amino acid sequences of the protein were identical with those of argininosuccinate synthase (EC 6.3.4.5). To examine whether argininosuccinate synthase can interact with lipid A, we purified the enzyme from mouse liver and found the co-elevation of the specific enzyme activity and specific lipid A-inactivating activity, indicating that argininosuccinate synthase is the major lipid A-interacting protein in liver. Argininosuccinate synthase also inhibited the biological activities (macrophage activation and Limulus test) of natural lipid A and rough-type LPS but not smooth-type LPS. The enzyme activity was inhibited by lipid A and rough-type LPS and also by smooth-type LPS. Native gel electrophoresis of a mixture of argininosuccinate synthase and LPS and immunoprecipitation of a mixture of argininosuccinate synthase and [3H]-LPS with anti-argininosuccinate synthase antiserum showed that argininosuccinate synthase stably bound lipid A and LPS. These findings, together, indicate that argininosuccinate synthase can effectively bind LPS in the liver.

Erratum to “Separation of 6-deoxy-heptan from a smooth-type lipopolysaccharide preparation of Burkholderia pseudomallei”

Erratum to “Separation of 6-deoxy-heptan from a smooth-type lipopolysaccharide preparation of Burkholderia pseudomallei”

Comparison of the Antibody Response in Adult Cattle Against Different Epitopes of Brucella abortus Lipopolysaccharide

The comparison of serological responses in a sample of adult, vaccinated and field‐infected bovines with Brucella abortus is reported. Indirect enzyme immunoassay (EIA) titration curves and Western blotting tests for smooth‐type lipopolysaccharide (S‐LPS), rough‐type LPS (R‐LPS) and lipid A were performed. In the initial screening of sera, an overall prevalence of 20.5 % was found, which corresponds to a country with a high incidence of brucellosis. End‐point EIA titres against LPS antigens from vaccinated and field‐infected cows were not significantly different. However, the absorbance values in the titration curves were significantly higher for S‐LPS as compared with the other antigens. A high correlation coefficient (r=0.933) was obtained when the titres to R‐LPS versus lipid A were compared. Western blotting reactions of vaccinated and field‐infected animals were indistinguishable. S‐LPS, R‐LPS and lipid A epitopes were recognized in a heterogeneous manner. In general, the number of bovines that reacted against LPS was higher in the field‐infected group, with a stronger binding to S‐LPS. Based on our observations, the vaccinated and field‐infected bovines are capable of producing similar antibody responses to the Brucella main outer surface antigen, LPS. It should be emphasized that the humoral response of cattle to Brucella LPS contains significant amounts of antibodies to other antigenic moieties of this important surface molecule, which may contribute to the immunity to brucellosis.

Separation of 6-deoxy-heptane from a smooth-type lipopolysaccharide preparation of Burkholderia pseudomallei

Smooth-type lipopolysaccharide (LPS) of Burkholderia pseudomallei has been reported to contain two kinds of O-antigenic polysaccharides, a 1,3-linked homopolymer of 6-deoxy-heptose and a polymer with a repeating unit of →3)-glucose-(1→3)-6-deoxy-talose-(1→ with O-acetyl or O-methyl modifications. A LPS preparation containing these two polysaccharides was separated by gel-permeation chromatography in this study. Chemical analysis of the separated fractions revealed the 6-deoxy-heptane to be a polysaccharide without a lipid portion and the polymer of glucose and 6-deoxy-talose to be an O-antigenic polysaccharide of the LPS. This result was further supported by the assay of these polysaccharide molecules for macrophage activation activity. The 6-deoxy-heptane showed no macrophage activation, indicating that this polysaccharide was not the LPS, but one of the capsular polysaccharides of B. pseudomallei.

Separation of 6-deoxy-heptan [correction of 6-deoxy-heptane] from a smooth-type lipopolysaccharide preparation of Burkholderia pseudomallei.

Smooth-type lipopolysaccharide (LPS) of Burkholderia pseudomallei has been reported to contain two kinds of O-antigenic polysaccharides, a 1,3-linked homopolymer of 6-deoxy-heptose and a polymer with a repeating unit of -->3)-glucose-(1-->3)-6-deoxy-talose-(1--> with O-acetyl or O-methyl modifications. A LPS preparation containing these two polysaccharides was separated by gel-permeation chromatography in this study. Chemical analysis of the separated fractions revealed the 6-deoxy-heptan [corrected] to be a polysaccharide without a lipid portion and the polymer of glucose and 6-deoxy-talose to be an O-antigenic polysaccharide of the LPS. This result was further supported by the assay of these polysaccharide molecules for macrophage activation activity. The 6-deoxy-heptan [corrected] showed no macrophage activation, indicating that this polysaccharide was not the LPS, but one of the capsular polysaccharides of B. pseudomallei.

Diffusion of macrolide antibiotics through the outer membrane of Moraxella catarrhalis

We reported previously that the high susceptibility of Moraxella catarrhalis to macrolide antibiotics and other hydrophobic antimicrobial agents was related to the hydrophobicity of the cell surface. Electrophoretic analysis of lipopolysaccharide (LPS) extracted from M. catarrhalis revealed a deep rough-type profile similar to that of an LPS Re type mutant of Salmonella typhimurium, which also exhibits high susceptibility to macrolides. Moreover, treatment of 32P-labeled cells of M. catarrhalis by phospholipase C induced the release of radioactive materials. These results suggested that hydrophobic agents such as macrolides readily access the cell surface exposed by the deep roughtype LPS and phospholipids, and permeate into the cell interior through the lipid bilayer. In fact, M. catarrhalis cells rapidly accumulated large amounts of the macrolide antibiotics, erythromycin and rokitamycin, whereas no accumulation of the macrolides was observed in cells having smooth-type or Rc type LPS under the same conditions.

Biological activities of lipopolysaccharides extracted from porcine vaccine strains.

Lipopolysaccharides (LPSs) were purified from Actinobacillus pleuropneumoniae serotype 2, Bordetella bronchiseptica and Haemophilus parasuis serotype 5, which were used for vaccine production in Japan, by the phenol-water procedure. In SDS-PAGE analysis, A. pleuropneumoniae LPS, as well as Escherichia coli LPS, demonstrated a typical ladder profile of a smooth-type LPS. On the other hand, B. bronchiseptica and H. parasuis LPSs lacked the ladder profiles. It was found that the biological activity of these LPSs was comparable to those of E. coli LPS in terms of activation of the clotting enzyme of Limulus amoebocyte lysate, mitogenic activity of mouse spleen cells, stimulation of TNF-alpha and nitric oxide production, but IL-6 production could hardly be observed in any LPS.

Serological studies of an acid-labile O-polysaccharide of Proteus vulgaris OX19 lipopolysaccharide using human and rabbit antibodies.

In a Weil-Felix test, sera from patients infected with Rickettsia sp. agglutinate Proteus OX types of bacteria and Proteus lipopolysaccharide (LPS) are responsible for the cross-reaction. Data on the character of LPS of one of the OX group strains, Proteus vulgaris OX19, are contradictory, and it remained unclear whether it has an O-polysaccharide (OPS) and is thus LPS of the smooth type (S) or not (rough-type LPS). Our studies showed that P. vulgaris OX19 (strain PZH-24) produces a smooth-type LPS that contains a long-chain OPS, but it undergoes depolymerization during mild acid hydrolysis conventionally used for LPS delipidation and loses the serological activity. An elucidation of the complete structure of OPS demonstrated the presence of a glycosyl phosphate linkage responsible for the acid-lability of the polysaccharide chain. In ELISA, both IgM type antibodies in a Weil-Felix test with human anti-Rickettsia typhi sera and rabbit anti-P. vulgaris OX19 antibodies reacted with OPS. Rabbit antibodies did not inhibit the cross-reaction with human antibodies and thus bind to different epitopes.

A two-component regulatory system playing a critical role in plant pathogens and endosymbionts is present in Brucella abortus and controls cell invasion and virulence.

Two mutants showing increased sensitivity to polycations and surfactants were obtained by transposon mutagenesis of virulent Brucella abortus 2308 Nalr. These mutants showed no obvious in vitro growth defects and produced smooth-type lipopolysaccharides. However, they hardly multiplied or persisted in mouse spleens, displayed reduced invasiveness in macrophages and HeLa cells, lost the ability to inhibit lysosome fusion and were unable to replicate intracellularly. Subsequent DNA analyses identified a two-component regulatory system [Brucella virulence related (Bvr)] with a regulatory (BvrR) and sensory (BvrS) protein. Cloning of bvrR in the BvrR-deficient mutant restored the resistance to polycations and, in part, the invasiveness and the ability to multiply intracellularly. BvrR and BvrS were highly similar (87-89% and 70-80% respectively) to the regulatory and sensory proteins of the chromosomally encoded Rhizobium meliloti Chvl-ExoS and Agrobacterium tumefaciens Chvl-ChvG systems previously shown to be critical for endosymbiosis and pathogenicity in plants. Divergence among the three sensory proteins was located mostly within a periplasmic domain probably involved in stimulus sensing. As B. abortus, R. meliloti and A. tumefaciens are phylogenetically related, these observations suggest that these systems have a common ancestor that has evolved to sense stimuli in plant and animal microbial environments.