Early transcriptomic responses of rice leaves to herbivory by Spodoptera frugiperda.

During herbivory, chewing insects deposit complex oral secretions (OS) onto the plant wound. Understanding how plants respond to the different cues of herbivory remains an active area of research. In this study, we used an herbivory-mimick experiment to investigate the early transcriptional response of rice plants leaves to wounding, OS, and OS microbiota from Spodoptera frugiperda larvae. Wounding induced a massive early response associated to hormones such as jasmonates. This response switched drastically upon OS treatment indicating the activation of OS specific pathways. When comparing native and dysbiotic OS treatments, we observed few gene regulation. This suggests that in addition to wounding the early response in rice is mainly driven by the insect compounds of the OS rather than microbial. However, microbiota affected genes encoding key phytohormone synthesis enzymes, suggesting an additional modulation of plant response by OS microbiota.

The endosymbiont and the second bacterial circle of entomopathogenic nematodes.

Single host-symbiont interactions should be reconsidered from the perspective of the pathobiome. We revisit here the interactions between entomopathogenic nematodes (EPNs) and their microbiota. We first describe the discovery of these EPNs and their bacterial endosymbionts. We also consider EPN-like nematodes and their putative symbionts. Recent high-throughput sequencing studies have shown that EPNs and EPN-like nematodes are also associated with other bacterial communities, referred to here as the second bacterial circle of EPNs. Current findings suggest that some members of this second bacterial circle contribute to the pathogenic success of nematodes. We suggest that the endosymbiont and the second bacterial circle delimit an EPN pathobiome.

Diverse Roles for a Conserved DNA-Methyltransferase in the Entomopathogenic Bacterium Xenorhabdus.

In bacteria, DNA-methyltransferase are responsible for DNA methylation of specific motifs in the genome. This methylation usually occurs at a very high rate. In the present study, we studied the MTases encoding genes found in the entomopathogenic bacteria Xenorhabdus. Only one persistent MTase was identified in the various species of this genus. This MTase, also broadly conserved in numerous Gram-negative bacteria, is called Dam: DNA-adenine MTase. Methylome analysis confirmed that the GATC motifs recognized by Dam were methylated at a rate of >99% in the studied strains. The observed enrichment of unmethylated motifs in putative promoter regions of the X. nematophila F1 strain suggests the possibility of epigenetic regulations. The overexpression of the Dam MTase responsible for additional motifs to be methylated was associated with impairment of two major phenotypes: motility, caused by a downregulation of flagellar genes, and hemolysis. However, our results suggest that dam overexpression did not modify the virulence properties of X. nematophila. This study increases the knowledge on the diverse roles played by MTases in bacteria.

The Odilorhabdin Antibiotic Biosynthetic Cluster and Acetyltransferase Self-Resistance Locus Are Niche and Species Specific.

Antibiotic resistance is an increasing threat to human health. A direct link has been established between antimicrobial self-resistance determinants of antibiotic producers, environmental bacteria, and clinical pathogens. Natural odilorhabdins (ODLs) constitute a new family of 10-mer linear cationic peptide antibiotics inhibiting bacterial translation by binding to the 30S subunit of the ribosome. These bioactive secondary metabolites are produced by entomopathogenic bacterial symbiont Xenorhabdus (Morganellaceae), vectored by the soil-dwelling nematodes. ODL-producing Xenorhabdus nematophila symbionts have mechanisms of self-protection. In this study, we cloned the 44.5-kb odl biosynthetic gene cluster (odl-BGC) of the symbiont by recombineering and showed that the N-acetyltransferase-encoding gene, oatA, is responsible for ODL resistance. In vitro acetylation and liquid chromatography-tandem mass spectrometry (LC-MS/MS) analyses showed that OatA targeted the side chain amino group of ODL rare amino acids, leading to a loss of translation inhibition and antibacterial properties. Functional, genomic, and phylogenetic analyses of oatA revealed an exclusive cis-link to the odilorhabdin BGC, found only in X. nematophila and a specific phylogenetic clade of Photorhabdus. This work highlights the coevolution of antibiotic production and self-resistance as ancient features of this unique tripartite complex of host-vector-symbiont interactions without odl-BGC dissemination by lateral gene transfer. IMPORTANCE Odilorhabdins (ODLs) constitute a novel antibiotic family with promising properties for treating problematic multidrug-resistant Gram-negative bacterial infections. ODLs are 10-mer linear cationic peptides inhibiting bacterial translation by binding to the small subunit of the ribosome. These natural peptides are produced by Xenorhabdus nematophila, a bacterial symbiont of entomopathogenic nematodes well known to produce large amounts of specialized secondary metabolites. Like other antimicrobial producers, ODL-producing Xenorhabdus nematophila has mechanisms of self-protection. In this study, we cloned the ODL-biosynthetic gene cluster of the symbiont by recombineering and showed that the N-acetyltransferase-encoding gene, oatA, is responsible for ODL resistance. In vitro acetylation and LC-MS/MS analyses showed that OatA targeted the side chain amino group of ODL rare amino acids, leading to a loss of translation inhibition and antibacterial properties. Functional, genomic, and phylogenetic analyses of oatA revealed the coevolution of antibiotic production and self-resistance as ancient feature of this particular niche in soil invertebrates without resistance dissemination.

A novel semi-selective medium for Pseudomonas protegens isolation from soil samples.

Pseudomonas protegens is a rhizosphere pseudomonad with a high agronomical potential (entomopathogenic and beneficial to plants) and bio-catalytic activities, but no selective medium has been described for its isolation. We developed a semi-selective minimum agar medium for the specific isolation and growth of P. protegens. We searched for both (i) a carbon source allowing the growth of P. protegens but potentially inhibiting the growth of other pseudomonads and (ii) an antimicrobial agent suppressing other members of the bacterial rhizosphere community. The M9-PP-agar medium consists of M9 base agar with adipic acid as the only carbon source and Irgasan® as an anti-bacterial agent. We tested the selectivity and sensitivity of M9-PP-agar by measuring the growth of 68 bacterial strains from 36 different species on this medium. Ten of the species tested were able to grow on M9-PP-agar medium: four species from the Pseudomonadaceae (Pseudomonas aeruginosa, Pseudomonas protegens, Pseudomonas putida, Stenotrophomonas maltophilia) as well as Achromobacter xylosoxidans, Agrobacterium tumefaciens, Brevundimonas sp., Serratia liquefaciens, Serratia marcescens and Variovorax paradoxus. All colonies were white, except for those of P. protegens (12 strains), which were typically brown. We demonstrated the efficiency of the M9-PP agar medium for P. protegens isolation, by inoculating two soils with the reference strain P. protegens CHAOT and then reisolating them. We also developed a fitF-PCR test targeting a regulator gene of the insecticidal P. protegens fit locus, for the rapid molecular detection of P. protegens colonies. We, therefore, developed a highly specific process for the routine isolation of new P. protegens strains from the soil environment, based on the use of a semi-selective medium and the specific color of colonies.

Partner-specific induction of Spodoptera frugiperda immune genes in response to the entomopathogenic nematobacterial complex Steinernema carpocapsae-Xenorhabdus nematophila.

The Steinernema carpocapsae-Xenorhabdus nematophila association is a nematobacterial complex used in biological control of insect crop pests. The infection success of this dual pathogen strongly depends on its interactions with the host's immune system. Here, we used the lepidopteran pest Spodoptera frugiperda to analyze the respective impact of each partner in the induction of its immune responses. First, we used previously obtained RNAseq data to construct the immunome of S. frugiperda and analyze its induction. We then selected representative genes to study by RT-qPCR their induction kinetics and specificity after independent injections of each partner. We showed that both X. nematophila and S. carpocapsae participate in the induction of stable immune responses to the complex. While X. nematophila mainly induces genes classically involved in antibacterial responses, S. carpocapsae induces lectins and genes involved in melanization and encapsulation. We discuss putative relationships between these differential inductions and the pathogen immunosuppressive strategies.

Entomopathogenic nematode-associated microbiota: from monoxenic paradigm to pathobiome.

BACKGROUND: The holistic view of bacterial symbiosis, incorporating both host and microbial environment, constitutes a major conceptual shift in studies deciphering host-microbe interactions. Interactions between Steinernema entomopathogenic nematodes and their bacterial symbionts, Xenorhabdus, have long been considered monoxenic two partner associations responsible for the killing of the insects and therefore widely used in insect pest biocontrol. We investigated this "monoxenic paradigm" by profiling the microbiota of infective juveniles (IJs), the soil-dwelling form responsible for transmitting Steinernema-Xenorhabdus between insect hosts in the parasitic lifecycle. RESULTS: Multigenic metabarcoding (16S and rpoB markers) showed that the bacterial community associated with laboratory-reared IJs from Steinernema carpocapsae, S. feltiae, S. glaseri and S. weiseri species consisted of several Proteobacteria. The association with Xenorhabdus was never monoxenic. We showed that the laboratory-reared IJs of S. carpocapsae bore a bacterial community composed of the core symbiont (Xenorhabdus nematophila) together with a frequently associated microbiota (FAM) consisting of about a dozen of Proteobacteria (Pseudomonas, Stenotrophomonas, Alcaligenes, Achromobacter, Pseudochrobactrum, Ochrobactrum, Brevundimonas, Deftia, etc.). We validated this set of bacteria by metabarcoding analysis on freshly sampled IJs from natural conditions. We isolated diverse bacterial taxa, validating the profile of the Steinernema FAM. We explored the functions of the FAM members potentially involved in the parasitic lifecycle of Steinernema. Two species, Pseudomonas protegens and P. chlororaphis, displayed entomopathogenic properties suggestive of a role in Steinernema virulence and membership of the Steinernema pathobiome. CONCLUSIONS: Our study validates a shift from monoxenic paradigm to pathobiome view in the case of the Steinernema ecology. The microbial communities of low complexity associated with EPNs will permit future microbiota manipulation experiments to decipher overall microbiota functioning in the infectious process triggered by EPN in insects and, more generally, in EPN ecology.

Bacterial community profile after the lethal infection of Steinernema-Xenorhabdus pairs into soil-reared Tenebrio molitor larvae.

The host microbiota may have an impact on pathogens. This is often studied in laboratory-reared hosts but rarely in individuals whose microbiota looks like that of wild animals. In this study, we modified the gut microbiota of the insect Tenebrio molitor by rearing larvae in soil sampled from the field. We showed by high throughput sequencing methods that this treatment modifies the gut microbiota so that it is more diversified than that of laboratory-reared insects, and closely resembled the one of soil-dwelling insects. To describe what the entomopathogenic bacterial symbiont Xenorhabdus (Enterobacteriaceae), vectored by the soil-dwelling nematode Steinernema, might experience in natural conditions, we studied the infestation of the soil-reared T. molitor larvae with three Steinernema-Xenorhabdus pairs. We performed the infestation at 18°C, which delays the emergence of new infective juveniles (IJs), the soil-dwelling nematode forms, but which is a temperature compatible with natural infestation. We analyzed by high throughput sequencing methods the composition of the bacterial community within the insect cadavers before the first emergences of IJs. These bacterial communities were generally characterized by one or two non-symbiont taxa. Even for highly lethal Steinernema-Xenorhabdus pairs, the symbiont does not dominate the bacterial community within the insect cadaver.

rpoB, a promising marker for analyzing the diversity of bacterial communities by amplicon sequencing.

BACKGROUND: Microbiome composition is frequently studied by the amplification and high-throughput sequencing of specific molecular markers (metabarcoding). Various hypervariable regions of the 16S rRNA gene are classically used to estimate bacterial diversity, but other universal bacterial markers with a finer taxonomic resolution could be employed. We compared specificity and sensitivity between a portion of the rpoB gene and the V3 V4 hypervariable region of the 16S rRNA gene. RESULTS: We first designed universal primers for rpoB suitable for use with Illumina sequencing-based technology and constructed a reference rpoB database of 45,000 sequences. The rpoB and V3 V4 markers were amplified and sequenced from (i) a mock community of 19 bacterial strains from both Gram-negative and Gram-positive lineages; (ii) bacterial assemblages associated with entomopathogenic nematodes. In metabarcoding analyses of mock communities with two analytical pipelines (FROGS and DADA2), the estimated diversity captured with the rpoB marker resembled the expected composition of these mock communities more closely than that captured with V3 V4. The rpoB marker had a higher level of taxonomic affiliation, a higher sensitivity (detection of all the species present in the mock communities), and a higher specificity (low rates of spurious OTU detection) than V3 V4. We compared the performance of the rpoB and V3 V4 markers in an animal ecosystem model, the infective juveniles of the entomopathogenic nematode Steinernema glaseri carrying the symbiotic bacteria Xenorhabdus poinarii. Both markers showed the bacterial community associated with this nematode to be of low diversity (< 50 OTUs), but only rpoB reliably detected the symbiotic bacterium X. poinarii. CONCLUSIONS: Our results confirm that different microbiota composition data may be obtained with different markers. We found that rpoB was a highly appropriate marker for assessing the taxonomic structure of mock communities and the nematode microbiota. Further studies on other ecosystems should be considered to evaluate the universal usefulness of the rpoB marker. Our data highlight two crucial elements that should be taken into account to ensure more reliable and accurate descriptions of microbial diversity in high-throughput amplicon sequencing analyses: i) the need to include mock communities as controls; ii) the advantages of using a multigenic approach including at least one housekeeping gene (rpoB is a good candidate) and one variable region of the 16S rRNA gene. This study will be useful to the growing scientific community describing bacterial communities by metabarcoding in diverse ecosystems.

The complete methylome of an entomopathogenic bacterium reveals the existence of loci with unmethylated Adenines.

DNA methylation can serve to control diverse phenomena in eukaryotes and prokaryotes, including gene regulation leading to cell differentiation. In bacteria, DNA methylomes (i.e., methylation state of each base of the whole genome) have been described for several species, but methylome profile variation during the lifecycle has rarely been studied, and only in a few model organisms. Moreover, major phenotypic changes have been reported in several bacterial strains with a deregulated methyltransferase, but the corresponding methylome has rarely been described. Here we report the first methylome description of an entomopathogenic bacterium, Photorhabdus luminescens. Eight motifs displaying a high rate of methylation (>94%) were identified. The methylome was strikingly stable over course of growth, but also in a subpopulation responsible for a critical step in the bacterium's lifecycle: successful survival and proliferation in insects. The rare unmethylated GATC motifs were preferentially located in putative promoter regions, and most of them were methylated after Dam methyltransferase overexpression, suggesting that DNA methylation is involved in gene regulation. Our findings bring key insight into bacterial methylomes and encourage further research to decipher the role of loci protected from DNA methylation in gene regulation.

The insect pathogenic bacterium Xenorhabdus innexi has attenuated virulence in multiple insect model hosts yet encodes a potent mosquitocidal toxin.

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.

Variable virulence phenotype of Xenorhabdus bovienii (γ-Proteobacteria: Enterobacteriaceae) in the absence of their vector hosts.

Xenorhabdus bovienii bacteria have a dual lifestyle: they are mutualistic symbionts to many species of Steinernema nematodes and are pathogens to a wide array of insects. Previous studies have shown that virulence of X.bovienii-Steinernema spp. pairs decreases when the nematodes associate with non-cognate bacterial strains. However, the virulence of the X. bovienii strains alone has not been fully investigated. In this study, we characterized the virulence of nine X. bovienii strains in Galleria mellonella and Spodoptera littoralis and performed a comparative genomic analysis to correlate observed phenotypes with strain genotypes. Two X. bovienii strains were found to be highly virulent against the tested insect hosts, while three strains displayed attenuated insect virulence. Comparative genomic analyses revealed the presence of several clusters present only in virulent strains, including a predicted type VI secretion system (T6SS). We performed intra-species-competition assays, and showed that the virulent T6SS+ strains generally outcompeted the less virulent T6SS- strains. Thus, we speculate that the T6SS in X. bovienii may be another addition to the arsenal of antibacterial mechanisms expressed by these bacteria in an insect, where it could potentially play three key roles: (1) competition against the insect host microbiota; (2) protection of the insect cadaver from necrotrophic microbial competitors; and (3) outcompeting other Xenorhabdus species and/or strains when co-infections occur.

A New Member of the Growing Family of Contact-Dependent Growth Inhibition Systems in Xenorhabdus doucetiae.

Xenorhabdus is a bacterial symbiont of entomopathogenic Steinernema nematodes and is pathogenic for insects. Its life cycle involves a stage inside the insect cadaver, in which it competes for environmental resources with microorganisms from soil and the insect gut. Xenorhabdus is, thus, a useful model for identifying new interbacterial competition systems. For the first time, in an entomopathogenic bacterium, Xenorhabdus doucetiae strain FRM16, we identified a cdi-like locus. The cdi loci encode contact-dependent inhibition (CDI) systems composed of proteins from the two-partner secretion (TPS) family. CdiB is the outer membrane protein and CdiA is the toxic exoprotein. An immunity protein, CdiI, protects bacteria against inhibition. We describe here the growth inhibition effect of the toxic C-terminus of CdiA from X. doucetiae FRM16, CdiA-CTFRM16, following its production in closely and distantly related enterobacterial species. CdiA-CTFRM16 displayed Mg2+-dependent DNase activity, in vitro. CdiA-CTFRM16-mediated growth inhibition was specifically neutralized by CdiIFRM16. Moreover, the cdi FRM16 locus encodes an ortholog of toxin-activating proteins C that we named CdiCFRM16. In addition to E. coli, the cdiBCAI-type locus was found to be widespread in environmental bacteria interacting with insects, plants, rhizospheres and soils. Phylogenetic tree comparisons for CdiB, CdiA and CdiC suggested that the genes encoding these proteins had co-evolved. By contrast, the considerable variability of CdiI protein sequences suggests that the cdiI gene is an independent evolutionary unit. These findings further characterize the sparsely described cdiBCAI-type locus.

Comparative Genomics between Two Xenorhabdus bovienii Strains Highlights Differential Evolutionary Scenarios within an Entomopathogenic Bacterial Species.

Bacteria of the genus Xenorhabdus are symbionts of soil entomopathogenic nematodes of the genus Steinernema. This symbiotic association constitutes an insecticidal complex active against a wide range of insect pests. Within Xenorhabdus bovienii species, the X. bovienii CS03 strain (Xb CS03) is nonvirulent when directly injected into lepidopteran insects, and displays a low virulence when associated with its Steinernema symbiont. The genome of Xb CS03 was sequenced and compared with the genome of a virulent strain, X. bovienii SS-2004 (Xb SS-2004). The genome size and content widely differed between the two strains. Indeed, Xb CS03 had a large genome containing several specific loci involved in the inhibition of competitors, including a few NRPS-PKS loci (nonribosomal peptide synthetases and polyketide synthases) producing antimicrobial molecules. Consistently, Xb CS03 had a greater antimicrobial activity than Xb SS-2004. The Xb CS03 strain contained more pseudogenes than Xb SS-2004. Decay of genes involved in the host invasion and exploitation (toxins, invasins, or extracellular enzymes) was particularly important in Xb CS03. This may provide an explanation for the nonvirulence of the strain when injected into an insect host. We suggest that Xb CS03 and Xb SS-2004 followed divergent evolutionary scenarios to cope with their peculiar life cycle. The fitness strategy of Xb CS03 would involve competitor inhibition, whereas Xb SS-2004 would quickly and efficiently kill the insect host. Hence, Xenorhabdus strains would have widely divergent host exploitation strategies, which impact their genome structure.

Attenuated virulence and genomic reductive evolution in the entomopathogenic bacterial symbiont species, Xenorhabdus poinarii.

Bacteria of the genus Xenorhabdus are symbionts of soil entomopathogenic nematodes of the genus Steinernema. This symbiotic association constitutes an insecticidal complex active against a wide range of insect pests. Unlike other Xenorhabdus species, Xenorhabdus poinarii is avirulent when injected into insects in the absence of its nematode host. We sequenced the genome of the X. poinarii strain G6 and the closely related but virulent X. doucetiae strain FRM16. G6 had a smaller genome (500-700 kb smaller) than virulent Xenorhabdus strains and lacked genes encoding potential virulence factors (hemolysins, type 5 secretion systems, enzymes involved in the synthesis of secondary metabolites, and toxin-antitoxin systems). The genomes of all the X. poinarii strains analyzed here had a similar small size. We did not observe the accumulation of pseudogenes, insertion sequences or decrease in coding density usually seen as a sign of genomic erosion driven by genetic drift in host-adapted bacteria. Instead, genome reduction of X. poinarii seems to have been mediated by the excision of genomic blocks from the flexible genome, as reported for the genomes of attenuated free pathogenic bacteria and some facultative mutualistic bacteria growing exclusively within hosts. This evolutionary pathway probably reflects the adaptation of X. poinarii to specific host.

Draft Genome Sequence and Annotation of the Entomopathogenic Bacterium Xenorhabdus szentirmaii Strain DSM16338.

We report the genome sequence of Xenorhabdus szentirmaii DSM16338 (4.84 Mb), a symbiont of the entomopathogenic nematode Steinernema rarum. This strain produces antimicrobial activity.

Draft Genome Sequence and Annotation of the Entomopathogenic Bacterium Xenorhabdus nematophila Strain F1.

We report the 4.3-Mb genome sequence of Xenorhabdus nematophila strain F1, a Gram-negative bacterium that is a symbiont of the entomopathogenic nematode Steinernema carpocapsae and pathogenic by direct injection for a wide variety of insects.

Comparative analysis of P2-type remnant prophage loci in Xenorhabdus bovienii and Xenorhabdus nematophila required for xenorhabdicin production.

The xnp1 remnant P2-type prophage of Xenorhabdus nematophila produces xenorhabdicin that is active against closely related species. Xenorhabdicin had not been characterized previously in other Xenorhabdus species. Here, we show xenorhabdicin production in six different strains of Xenorhabdus bovienii. The sequenced genome of X. bovienii SS-2004 was found to possess a highly conserved remnant P2-type cluster (xbp1). Inactivation of the xbpS1 sheath gene resulted in loss of bacteriocin activity, indicating that the xbp1 locus was required for xenorhabdicin production. xbp1 and xnp1 contain a CI-type repressor, a dinI gene involved in stabilization of ssDNA-RecA complexes and are inducible with mitomycin C, suggesting that both loci are regulated by cleavage of the CI repressor. Both xnp1 and xbp1 lack typical P2-type lysis genes but contain a predicted endolysin gene (enp) that may be involved in cell lysis. The main tail fibers of xnp1 and xbp1 are mosaic structures with divergent C-terminal regions suggesting they differ in host specificity. Several genes encoding C-terminal tail fiber fragments are present in the same position in xnp1 and xbp1. Recombination between the main fiber genes and the C-terminal fragments could potentially expand the host range specificity of xenorhabdicin in the respective strains.

The entomopathogenic bacterial endosymbionts Xenorhabdus and Photorhabdus: convergent lifestyles from divergent genomes.

Members of the genus Xenorhabdus are entomopathogenic bacteria that associate with nematodes. The nematode-bacteria pair infects and kills insects, with both partners contributing to insect pathogenesis and the bacteria providing nutrition to the nematode from available insect-derived nutrients. The nematode provides the bacteria with protection from predators, access to nutrients, and a mechanism of dispersal. Members of the bacterial genus Photorhabdus also associate with nematodes to kill insects, and both genera of bacteria provide similar services to their different nematode hosts through unique physiological and metabolic mechanisms. We posited that these differences would be reflected in their respective genomes. To test this, we sequenced to completion the genomes of Xenorhabdus nematophila ATCC 19061 and Xenorhabdus bovienii SS-2004. As expected, both Xenorhabdus genomes encode many anti-insecticidal compounds, commensurate with their entomopathogenic lifestyle. Despite the similarities in lifestyle between Xenorhabdus and Photorhabdus bacteria, a comparative analysis of the Xenorhabdus, Photorhabdus luminescens, and P. asymbiotica genomes suggests genomic divergence. These findings indicate that evolutionary changes shaped by symbiotic interactions can follow different routes to achieve similar end points.

The first record of entomopathogenic nematodes (Rhabiditiae: Steinernematidae and Heterorhabditidae) in natural ecosystems in Lebanon: A biogeographic approach in the Mediterranean region.

A survey of entomopathogenic nematodes in Lebanon was conducted for the first time during 2008-2009. Samples were collected on the coastal strip and in nine vegetation types extending from the coastal line to 3088m above sea level. Wooded and herbaceous ecosystems were considered for sampling purposes. A total of 570 samples were taken, out of which 1% were positive for entomopathogenic nematodes. Approximately, 15.8% out of the 19 sites sampled revealed entomopathogenic nematodes presence (representing three samples). Two entomopathogenic nematodes species Heterorhabditis bacteriophora and Steinernema feltiae were recovered, and identification of their symbiotic bacteria revealed the presence of a Xenorhabdus bovienii, Photorhabdus temperata subsp. thracensis, Photorhabdus luminescens subsp. kayaii and Photorhabdus luminescens subsp. Laumondii.