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NilB is a membrane-localized protein and is surface exposed. (A) X. nematophila cells expressing NilB-FLAG26 in a nilR mutant background (HGB1200) were pelleted, resuspended, and sonicated. Soluble and membrane fractions were separated by ultracentrifugation. The crude extract (lane 1), cleared lysate (lane 2), soluble fraction (lane 3), and membrane fraction (lane 4) were electrophoresed on a 12% SDS-polyacrylamide gel and transferred to a membrane. NilB protein (arrowhead) was detected using anti-FLAG antibody. (B) Whole X. nematophila nilR mutant cells expressing NilB-FLAG26 (HGB1200) were incubated with 38.4 μg/ml proteinase K (PK) (final concentration) for 1 to 30 min at 37°C. At various time points (indicated in minutes above each lane), samples were removed and reactions were stopped with the addition of 5 mM PMSF (final concentration) and kept on ice. SDS-PAGE loading dye was added to all the reaction mixtures, which were heated at 95°C for 10 min and electrophoresed on 12% SDS-PAGE gels. NilB (top panel) (anti-NilB) or NilC (bottom panel) (anti-NilC) was detected in Western blots using anti-FLAG or anti-NilC (15) antibody, respectively. Full-length NilB-FLAG26 protein (black arrowheads, No PK lanes) was present in the no-proteinase K control but not in cells lacking nilB (Δnil) (HGB1251). This species decreases in intensity after the addition of proteinase K, while smaller-molecular-mass proteins (black arrowheads, lanes 1 and 10) appear. M, Thermo Scientific Pierce prestained protein molecular mass marker (sizes in kDa are noted on the left).
Source publication
The gammaproteobacterium Xenorhabdus nematophila is a mutualistic symbiont that colonizes the intestine of the nematode Steinernema carpocapsae. nilB (nematode intestine localization) is essential for X. nematophila colonization of nematodes and is predicted to encode an integral outer membrane beta-barrel protein, but evidence supporting
this pred...
Citations
... In the complexes of S. carpocapsae and X. nematophila, nematode intestine localization (nil) factors A, B, and C are identified as molecular components that explain the specificity between the nematode and bacteria [10]. NilB and NilC are an outer membrane beta barrel protein and periplasmic lipoprotein, respectively [11,12], suggesting their roles in the molecular interactions with host gut epithelium. ...
... The different letters above standard deviation bars indicate significant differences among means at Type I error = 0.05 (LSD test). 12 ...
An entomopathogenic nematode, Oscheius tipulae, was isolated from a soil sample. The identification of this species was supported by morphological and molecular markers. The nematode isolate exhibited pathogenicity against different target insects including lepidopteran, coleopteran, and dipteran insects. The virulence of this nematode was similar to that of a well-known entomopathogenic nematode, Steinernema carpocapsae, against the same insect targets. A comparative metagenomics analysis of these two nematode species predicted the existence of a combined total of 272 bacterial species in their intestines, of which 51 bacterial species were shared between the two nematode species. In particular, the common gut bacteria included several entomopathogenic bacteria including Xenorhabdus nematophila, which is known as a symbiotic bacterium to S. carpocapsae. The nematode virulence of O. tipulae to insects was enhanced by an addition of dexamethasone but suppressed by an addition of arachidonic acid, suggesting that the immune defenses of the target insects against the nematode infection is mediated by eicosanoids, which would be manipulated by the symbiotic bacteria of the nematode. Unlike S. carpocapsae, O. tipulae showed high virulence against dipteran insects including fruit flies, onion flies, and mosquitoes. O. tipulae showed particularly high control efficacies against the onion maggot, Delia platura, infesting the Welsh onion in the rhizosphere in both pot and field assays.
... In contrast, the N-terminal ligand-binding domain is specific to hemophilin homologs, making these a distinct structural subgroup of T11SS cargo proteins. We established that, like the hemophilins X. nematophila HrpC and A. baumannii HphA (8,34,35), two other hemophilin homologs, H. haemolyticus Hpl and X. cabanillasii CrpC, rely on a T11SS secretor to reach the extracellular milieu. This establishes T11SS-dependence across all four hemophilin sequence subclusters tested and indicates T11SS-dependent secretion is a hallmark of the entire family. ...
... All strains, plasmids, and primers utilized in this study are described in Supplemental File 3. All cultures were grown in glucose minimal media (34), LB stored in the dark to prevent the formation of oxidative radicals (henceforth dark LB), or glucose minimal media supplemented with 1% dark LB. Plate-based cultures were grown on either LB supple mented with pyruvate to prevent the formation of reactive oxygen radicals (henceforth, LBP or glucose minimal plates) (34). ...
... All cultures were grown in glucose minimal media (34), LB stored in the dark to prevent the formation of oxidative radicals (henceforth dark LB), or glucose minimal media supplemented with 1% dark LB. Plate-based cultures were grown on either LB supple mented with pyruvate to prevent the formation of reactive oxygen radicals (henceforth, LBP or glucose minimal plates) (34). For plasmid-based expression, chemically competent E. coli strain BL21-DE3 (C43) were chosen for ease of transformation and their ability to tolerate expression of membrane proteins (52,53). ...
Cellular life relies on enzymes that require metals, which must be acquired from extracellular sources. Bacteria utilize surface and secreted proteins to acquire such valuable nutrients from their environment. These include the cargo proteins of the type eleven secretion system (T11SS), which have been connected to host specificity, metal homeostasis, and nutritional immunity evasion. This Sec-dependent, Gram-negative secretion system is encoded by organisms throughout the phylum Proteobacteria, including human pathogens Neisseria meningitidis, Proteus mirabilis, Acinetobacter baumannii, and Haemophilus influenzae. Experimentally verified T11SS-dependent cargo include transferrin-binding protein B (TbpB), the hemophilin homologs heme receptor protein C (HrpC), hemophilin A (HphA), the immune evasion protein factor-H binding protein (fHbp), and the host symbiosis factor nematode intestinal localization protein C (NilC). Here, we examined the specificity of T11SS systems for their cognate cargo proteins using taxonomically distributed homolog pairs of T11SS and hemophilin cargo and explored the ligand binding ability of those hemophilin cargo homologs. In vivo expression in Escherichia coli of hemophilin homologs revealed that each is secreted in a specific manner by its cognate T11SS protein. Sequence analysis and structural modeling suggest that all hemophilin homologs share an N-terminal ligand-binding domain with the same topology as the ligand-binding domains of the Haemophilus haemolyticus heme binding protein (Hpl) and HphA. We term this signature feature of this group of proteins the hemophilin ligand-binding domain. Network analysis of hemophilin homologs revealed five subclusters and representatives from four of these showed variable heme-binding activities, which, combined with sequence-structure variation, suggests that hemophilins are diversifying in function.
IMPORTANCE
The secreted protein hemophilin and its homologs contribute to the survival of several bacterial symbionts within their respective host environments. Here, we compared taxonomically diverse hemophilin homologs and their paired Type 11 secretion systems (T11SS) to determine if heme binding and T11SS secretion are conserved characteristics of this family. We establish the existence of divergent hemophilin sub-families and describe structural features that contribute to distinct ligand-binding behaviors. Furthermore, we demonstrate that T11SS are specific for their cognate hemophilin family cargo proteins. Our work establishes that hemophilin homolog-T11SS pairs are diverging from each other, potentially evolving into novel ligand acquisition systems that provide competitive benefits in host niches.
... The mechanisms by which certain classes of proteins, including lipoproteins, are targeted to and oriented within the outer-membrane are still largely unknown. The newly described type XI secretion system (TXISS), comprising an outer membrane protein (OMP) containing a DUF560 (a domain of unknown function 560), is broadly distributed among proteobacteria and mediates translocation of lipoprotein and a soluble protein cargo across the outer membrane (Heungens et al., 2002;Bhasin et al., 2012;Hooda et al., 2017;Grossman et al., 2021). ...
... An X. nematophila TXISS OMP , NilB, is encoded near an outer membrane lipoprotein NilC on a locus known as Symbiosis Region 1 (SR1) (Heungens et al., 2002;Cowles and Goodrich-Blair, 2004;Bhasin et al., 2012). In X. nematophila, the SR1 locus, which encodes both nilB and nilC, is necessary and sufficient for normal levels of colonization of S. carpocapsae intestines (Heungens et al., 2002;Cowles and Goodrich-Blair, 2008;Chaston et al., 2013). ...
... Previous whole-cell protease-digestion data demonstrated periplasmic orientation of the lipoprotein NilC, based on the observation of protease resistance of NilC in the whole cell but not lysate samples of wild type X. nematophila (Cowles and Goodrich-Blair, 2004). Later, another protease digestion experiment to detect surface NilC was performed on an X. nematophila nilR mutant, in which the absence of the transcription factor NilR causes nilB and nilC expression to be derepressed (Cowles and Goodrich-Blair, 2006;Bhasin et al., 2012). In this analysis, slight shaving of NilC was detected in whole cells, indicating some surface exposure (Bhasin et al., 2012). ...
The only known required component of the newly described Type XI secretion system (TXISS) is an outer membrane protein (OMP) of the DUF560 family. TXISSOMPs are broadly distributed across proteobacteria, but properties of the cargo proteins they secrete are largely unexplored. We report biophysical, histochemical, and phenotypic evidence that Xenorhabdus nematophila NilC is surface exposed. Biophysical data and structure predictions indicate that NilC is a two-domain protein with a C-terminal, 8-stranded β-barrel. This structure has been noted as a common feature of TXISS effectors and may be important for interactions with the TXISSOMP. The NilC N-terminal domain is more enigmatic, but our results indicate it is ordered and forms a β-sheet structure, and bioinformatics suggest structural similarities to carbohydrate-binding proteins. X. nematophila NilC and its presumptive TXISSOMP partner NilB are required for colonizing the anterior intestine of Steinernema carpocapsae nematodes: the receptacle of free-living, infective juveniles and the anterior intestinal cecum (AIC) in juveniles and adults. We show that, in adult nematodes, the AIC expresses a Wheat Germ Agglutinin (WGA)-reactive material, indicating the presence of N-acetylglucosamine or N-acetylneuraminic acid sugars on the AIC surface. A role for this material in colonization is supported by the fact that exogenous addition of WGA can inhibit AIC colonization by X. nematophila. Conversely, the addition of exogenous purified NilC increases the frequency with which X. nematophila is observed at the AIC, demonstrating that abundant extracellular NilC can enhance colonization. NilC may facilitate X. nematophila adherence to the nematode intestinal surface by binding to host glycans, it might support X. nematophila nutrition by cleaving sugars from the host surface, or it might help protect X. nematophila from nematode host immunity. Proteomic and metabolomic analyses of wild type X. nematophila compared to those lacking nilB and nilC revealed differences in cell wall and secreted polysaccharide metabolic pathways. Additionally, purified NilC is capable of binding peptidoglycan, suggesting that periplasmic NilC may interact with the bacterial cell wall. Overall, these findings support a model that NilB-regulated surface exposure of NilC mediates interactions between X. nematophila and host surface glycans during colonization. This is a previously unknown function for a TXISS.
... The DUF560 homolog NilB is a host association and species specificity factor in the nematode symbiont Xenorhabdus nematophila, a proteobacterium in the family Morganellaceae (12)(13)(14). A screen for X. nematophila mutants defective in colonizing Steinernema carpocapsae intestines revealed the nematode intestinal localization (nil) locus (14,15). The nil locus contains the genes nilB and nilC, each of which is independently necessary for colonization of nematodes. ...
... The nil locus contains the genes nilB and nilC, each of which is independently necessary for colonization of nematodes. Biochemical and bioinformatic analyses have established that NilC is an outer membrane-associated lipoprotein, and NilB is an outer membrane b-barrel in the DUF560 family with an ;140-amino-acid periplasmic N-terminal domain that contains tetratricopeptide repeats (15)(16)(17)(18). ...
... TXISS cluster according to environment. Using homology to NilB or Slam proteins, previous work identified a wide distribution of DUF560 proteins within mucosa-associated bacteria (9,14,15). To quantifiably delineate subfamilies within the TXISS, we generated a sequence similarity network (SSN) using the Enzyme Function Initiative toolset (EFI) (21)(22)(23) and annotated it to highlight environmental source or taxonomic grouping of microbes containing DUF560 homologs ( Fig. 2; see also Table S1 in the supplemental material). ...
The microbial constituency of a host-associated microbiome emerges from a complex physical and chemical interplay of microbial colonization factors, host surface conditions, and host immunological responses. To fill unique niches within a host, bacteria encode surface and secreted proteins that enable interactions with and responses to the host and cooccurring microbes.
... The globular domain and surface loop 6 play a crucial role in the nematode colonization. Epifluorescence microscopy of these mutants revealed that NilB is necessary at early stages of colonization (Bhasin et al., 2012). (ii) Cj0561c (DUF2860) is a probable membrane fusion protein and contributes to intestinal colonization. ...
OMPdb ( www.ompdb.org ) was introduced as a database for β-barrel outer membrane proteins from Gram-negative bacteria in 2011 and then included 69,354 entries classified into 85 families. The database has been updated continuously using a collection of characteristic profile Hidden Markov Models able to discriminate between the different families of prokaryotic transmembrane β-barrels. The number of families has increased ultimately to a total of 129 families in the current, second major version of OMPdb. New additions have been made in parallel with efforts to update existing families and add novel families. Here, we present the upgrade of OMPdb, which from now on aims to become a global repository for all transmembrane β-barrel proteins, both eukaryotic and bacterial.
... DUF560 family proteins are present throughout Proteobacteria, and only a few members have been characterized [13,14,16,17]. One characterized DUF560 homolog, NilB, was identified as a host-association and species-specificity factor in the entomopathogenic nematode symbiont To begin to understand the range of functions of DUF560 proteins in biology, we assessed their distribution, genomic context, and relatedness. ...
... Using either NilB or Slam proteins as bait, previous work had identified a wide distribution of DUF560 proteins in Gram-negative bacteria, seemingly enriched in those associated with animal mucosal surfaces [14, 16,17]. We sought to gain more quantifiable information about whether sub-families of DUF560 homologs exist and whether DUF560 are enriched among mucosal symbionts. ...
... One example is X. nematophila XNC1_0075, encoded adjacent to the DUF560 homolog XNC1_0074 in a TonB-dependent heme-receptor protein/TonB genomic context (Hrp locus). XNC1_0075 is encoded adjacent to XNC1_0074 and in the same orientation, as previously described [17] and it includes a TbpB_B_D "lipoprotein-5" class domain (Fig. 5) [14,17]. However, XNC1_0075 has a SPI-type (rather than SPII) signal sequence and lacks the canonical lipobox necessary for lipidation so is predicted to be a secreted soluble protein. ...
In host-associated bacteria, surface and secreted proteins mediate acquisition of nutrients, interactions with host cells, and specificity of tissue-localization. In Gram-negative bacteria, the mechanism by which many proteins cross or become tethered to the outer membrane remains unclear. The d omain of u nknown function (DUF)560 occurs in outer membrane proteins throughout Proteobacteria and has been implicated in host-bacteria interactions and lipoprotein surface exposure. We used sequence similarity networking to reveal three subfamilies of DUF560 homologs. One subfamily includes those DUF560 proteins experimentally characterized to date: NilB, a host-range determinant of the nematode-mutualist Xenorhabdus nematophila , and the s urface lipoprotein a ssembly m odulators Slam1 and Slam2, which facilitate msurface exposure of lipoproteins in Neisseria meningitidis (1, 2). We show that DUF560 proteins from a second subfamily facilitate secretion of soluble, non-lipidated proteins across the outer membrane. Using in silico analysis, we demonstrate that DUF560 gene complement correlates with bacterial environment at a macro level and host association at a species level. The DUF560 protein superfamily represents a newly characterized Gram-negative secretion system capable of lipoprotein surface exposure and soluble protein secretion with conserved roles in facilitating symbiosis. In light of these data, we propose that it be titled the type eleven s ecretion s ystem (TXISS).
Importance
The microbial constituents of a host associated microbiome are decided by a complex interplay of microbial colonization factors, host surface conditions, and host immunological responses. Filling such niches requires bacteria to encode an arsenal of surface and secreted proteins to effectively interact with the host and co-occurring microbes. Bioinformatic predictions of the localization and function of putative bacterial colonization factors are essential for assessing the potential of bacteria to engage in pathogenic, mutualistic, or commensal activities. This study uses publicly available genome sequence data, alongside experimental results from representative gene products from Xenorhabdus nematophila , to demonstrate a role for DUF560 family proteins in the secretion of bacterial effectors of host interactions. Our research delineates a broadly distributed family of proteins and enables more accurate predictions of the localization of colonization factors throughout Proteobacteria.
... The pBlueXIS1_460109UpDn or pBlueXIS1_460115UpDn construct was cloned into a pKR100 suicide vector; the resulting pKRXIS1_460115 and pKRXIS1_460109 constructs (Table 1) were separately conjugated into the WT X. innexi using E. coli S-17 λpir donor strain. The resulting mutants were first verified by PCR amplification of nilB, which is a Xenorhabdus-specific gene [118]. The position of mutation was also confirmed by PCR amplification of the flanking regions of the inserted kanamycin cassette. ...
Background: Xenorhabdus innexi is a bacterial symbiont of Steinernema scapterisci nematodes, which is a cricket specialist parasite and together the nematode and bacteria infect and kill crickets. Curiously, X. innexi expresses a potent extracellular mosquitocidal toxin activity in culture supernatants. We sequenced a draft genome of X. innexi and compared it to the genomes of related pathogens to elucidate the nature of specialization.
Results: Using green fluorescent protein-expressing X. innexi we confirm previous reports using culture-dependent techniques that X. innexi colonizes its nematode host at low levels (~3–8 cells per nematode), relative to other Xenorhabdus-Steinernema associations. We found that compared to the well-characterized entomopathogenic
nematode symbiont X. nematophila, X. innexi fails to suppress the insect phenoloxidase immune pathway and is attenuated for virulence and reproduction in the Lepidoptera Galleria mellonella and Manduca sexta, as well as the dipteran Drosophila melanogaster. To assess if, compared to other Xenorhabdus spp., X. innexi has a reduced capacity to
synthesize virulence determinants, we obtained and analyzed a draft genome sequence. We found no evidence for several hallmarks of Xenorhabdus spp. toxicity, including Tc and Mcf toxins. Similar to other Xenorhabdus genomes, we found numerous loci predicted to encode non-ribosomal peptide/polyketide synthetases. Anti-SMASH predictions of
these loci revealed one, related to the fcl locus that encodes fabclavines and zmn locus that encodes zeamines, as a likely candidate to encode the X. innexi mosquitocidal toxin biosynthetic machinery, which we designated Xlt. In support of this hypothesis, two mutants each with an insertion in an Xlt biosynthesis gene cluster lacked the mosquitocidal compound based on HPLC/MS analysis and neither produced toxin to the levels of the wild type parent.
Conclusions: The X. innexi genome will be a valuable resource in identifying loci encoding new metabolites of interest, but also in future comparative studies of nematode-bacterial symbiosis and niche partitioning among bacterial pathogens.
... To create low-and high-Lrp-expressing X. nematophila strains, pMYC4 (low-lrp donor plasmid) and pMYC5 (high-lrp donor plasmid) were inserted into the kefA gene of an lrp-2::kan mutant via biparental conjugation and homologous recombination, respectively. Plasmid integration at the kefA gene locus does not interfere with nematode colonization (4,16,56,57). The Lrp-dependent fluorescence reporter pMYC1 (PfliC-gfp/P lac -rfp) was introduced into the attTn7 site of the genomes in both of the low-Lrp-and high-Lrp-expressing X. nematophila strains described above by triparental conjugation and Tn7 transposition (58,59), creating low-lrp in-genome (low-lrp/Tn7-PfliC-gfp/P lac -rfp) and high-lrp in-genome (high-lrp/Tn7-PfliC-gfp/P lac -rfp) strains. ...
Xenorhabdus nematophila bacteria are mutualistic symbionts of Steinernema carpocapsae nematodes and pathogens of insects. The X. nematophila global regulator Lrp controls the expression of many genes involved in both mutualism and pathogenic activities, suggesting a role in the transition between the two host organisms. We previously reported that natural populations of X. nematophila exhibit various levels of Lrp expression and that cells expressing relatively low levels of Lrp are optimized for virulence in the insect Manduca sexta. The adaptive advantage of the high-Lrp-expressing state was not established. Here we used strains engineered to express constitutively high or low levels of Lrp to test the model in which high-Lrp-expressing cells are adapted for mutualistic activities with the nematode host. We demonstrate that high-Lrp cells form more robust biofilms in laboratory media than do low-Lrp cells, which may reflect adherence to host tissues. Also, our data showed that nematodes cultivated with high-Lrp strains are more frequently colonized than are those associated with low-Lrp strains. Taken together, these data support the idea that high-Lrp cells have an advantage in tissue adherence and colonization initiation. Furthermore, our data show that high-Lrp-expressing strains better support nematode reproduction than do their low-Lrp counterparts under both in vitro and in vivo conditions. Our data indicate that heterogeneity of Lrp expression in X. nematophila populations provides diverse cell populations adapted to both pathogenic (low-Lrp) and mutualistic (high-Lrp) states.
IMPORTANCE
Host-associated bacteria experience fluctuating conditions during both residence within an individual host and transmission between hosts. For bacteria that engage in evolutionarily stable, long-term relationships with particular hosts, these fluctuations provide selective pressure for the emergence of adaptive regulatory mechanisms. Here we present evidence that the bacterium Xenorhabdus nematophila uses various levels of the transcription factor Lrp to optimize its association with its two animal hosts, nematodes and insects, with which it behaves as a mutualist and a pathogen, respectively. Building on our previous finding that relatively low cellular levels of Lrp are optimal for pathogenesis, we demonstrate that, conversely, high levels of Lrp promote mutualistic activities with the Steinernema carpocapsae nematode host. These data suggest that X. nematophila has evolved to utilize phenotypic variation between high- and low-Lrp-expression states to optimize its alternating behaviors as a mutualist and a pathogen.
... This lipoprotein, named NilC has been previously shown to be lipidated in vivo, present in the outer membrane and is important for host colonization (Cowles and Goodrich-Blair, 2004). The gene encoding NilB, a putative Slam homolog, is present next to the gene encoding NilC, and is shown to be required for host colonization (Bhasin et al., 2012). Our analysis suggests that NilC is a surface lipoprotein (SLP) that is dependent on the Slam homolog, NilB, for surface display. ...
The surfaces of many Gram-negative bacteria are decorated with soluble proteins anchored to the outer membrane via an acylated N-terminus; these proteins are referred to as surface lipoproteins or SLPs. In Neisseria meningitidis, SLPs such as transferrin-binding protein B (TbpB) and factor-H binding protein (fHbp) are essential for host colonization and infection because of their essential roles in iron acquisition and immune evasion, respectively. Recently, we identified a family of outer membrane proteins called Slam (Surface lipoprotein assembly modulator) that are essential for surface display of neisserial SLPs. In the present study, we performed a bioinformatics analysis to identify 832 Slam related sequences in 638 Gram-negative bacterial species. The list included several known human pathogens, many of which were not previously reported to possess SLPs. Hypothesizing that genes encoding SLP substrates of Slams may be present in the same gene cluster as the Slam genes, we manually curated neighboring genes for 353 putative Slam homologs. From our analysis, we found that 185 (~52%) of the 353 putative Slam homologs are located adjacent to genes that encode a protein with an N-terminal lipobox motif. This list included genes encoding previously reported SLPs in Haemophilus influenzae and Moraxella catarrhalis, for which we were able to show that the neighboring Slams are necessary and sufficient to display these lipoproteins on the surface of Escherichia coli. To further verify the authenticity of the list of predicted SLPs, we tested the surface display of one such Slam-adjacent protein from Pasteurella multocida, a zoonotic pathogen. A robust Slam-dependent display of the P. multocida protein was observed in the E. coli translocation assay indicating that the protein is a Slam-dependent SLP. Based on multiple sequence alignments and domain annotations, we found that an eight-stranded beta-barrel domain is common to all the predicted Slam-dependent SLPs. These findings suggest that SLPs with a TbpB-like fold are found widely in Proteobacteria where they exist with their interaction partner Slam. In the future, SLPs found in pathogenic bacteria can be investigated for their role in virulence and may also serve as candidates for vaccine development.
... Each X. bovienii strain genomes contained homologs predicted to encode all of these regulators except NilR (Additional file 4). This is consistent with the fact that in X. nematophila NilR functions synergistically to negatively regulate the nilA, B, and C genes, nematode-host range specificity determinants that are not present in the X. bovienii genomes [17,31,58]. ...
Background
Xenorhabdus bacteria engage in a beneficial symbiosis with Steinernema nematodes, in part by providing activities that help kill and degrade insect hosts for nutrition. Xenorhabdus strains (members of a single species) can display wide variation in host-interaction phenotypes and genetic potential indicating that strains may differ in their encoded symbiosis factors, including secreted metabolites.
Methods
To discern strain-level variation among symbiosis factors, and facilitate the identification of novel compounds, we performed a comparative analysis of the genomes of 10 Xenorhabdus bovienii bacterial strains.
Results
The analyzed X. bovienii draft genomes are broadly similar in structure (e.g. size, GC content, number of coding sequences). Genome content analysis revealed that general classes of putative host-microbe interaction functions, such as secretion systems and toxin classes, were identified in all bacterial strains. In contrast, we observed diversity of individual genes within families (e.g. non-ribosomal peptide synthetase clusters and insecticidal toxin components), indicating the specific molecules secreted by each strain can vary. Additionally, phenotypic analysis indicates that regulation of activities (e.g. enzymes and motility) differs among strains.
Conclusions
The analyses presented here demonstrate that while general mechanisms by which X. bovienii bacterial strains interact with their invertebrate hosts are similar, the specific molecules mediating these interactions differ. Our data support that adaptation of individual bacterial strains to distinct hosts or niches has occurred. For example, diverse metabolic profiles among bacterial symbionts may have been selected by dissimilarities in nutritional requirements of their different nematode hosts. Similarly, factors involved in parasitism (e.g. immune suppression and microbial competition factors), likely differ based on evolution in response to naturally encountered organisms, such as insect hosts, competitors, predators or pathogens. This study provides insight into effectors of a symbiotic lifestyle, and also highlights that when mining Xenorhabdus species for novel natural products, including antibiotics and insecticidal toxins, analysis of multiple bacterial strains likely will increase the potential for the discovery of novel molecules
