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BMC Genomics Nov 2018Photorhabdus luminescens is an enteric bacterium, which lives in mutualistic association with soil nematodes and is highly pathogenic for a broad spectrum of insects. A... (Comparative Study)
Comparative Study
BACKGROUND
Photorhabdus luminescens is an enteric bacterium, which lives in mutualistic association with soil nematodes and is highly pathogenic for a broad spectrum of insects. A complete genome sequence for the type strain P. luminescens subsp. laumondii TT01, which was originally isolated in Trinidad and Tobago, has been described earlier. Subsequently, a rifampicin resistant P. luminescens strain has been generated with superior possibilities for experimental characterization. This strain, which is widely used in research, was described as a spontaneous rifampicin resistant mutant of TT01 and is known as TT01-Rif.
RESULTS
Unexpectedly, upon phenotypic comparison between the rifampicin resistant strain and its presumed parent TT01, major differences were found with respect to bioluminescence, pigmentation, biofilm formation, haemolysis as well as growth. Therefore, we renamed the strain TT01-Rif to DJC. To unravel the genomic basis of the observed differences, we generated a complete genome sequence for strain DJC using the PacBio long read technology. As strain DJC was supposed to be a spontaneous mutant, only few sequence differences were expected. In order to distinguish these from potential sequencing errors in the published TT01 genome, we re-sequenced a derivative of strain TT01 in parallel, also using the PacBio technology. The two TT01 genomes differed at only 30 positions. In contrast, the genome of strain DJC varied extensively from TT01, showing 13,000 point mutations, 330 frameshifts, and 220 strain-specific regions with a total length of more than 300 kb in each of the compared genomes.
CONCLUSIONS
According to the major phenotypic and genotypic differences, the rifampicin resistant P. luminescens strain, now named strain DJC, has to be considered as an independent isolate rather than a derivative of strain TT01. Strains TT01 and DJC both belong to P. luminescens subsp. laumondii.
Topics: Anti-Bacterial Agents; Base Sequence; Biofilms; DNA Transposable Elements; Drug Resistance, Bacterial; Genome, Bacterial; Genomics; Hemolysis; Mutation; Open Reading Frames; Phenotype; Photorhabdus; Prophages; Rifampin; Sequence Analysis, DNA; Symbiosis
PubMed: 30497380
DOI: 10.1186/s12864-018-5121-z -
Heliyon Mar 2023For decades, transcription of -operon was considered being constitutive. Therefore, this -operon has been used for measurements in non-specific bacterial luminescent...
For decades, transcription of -operon was considered being constitutive. Therefore, this -operon has been used for measurements in non-specific bacterial luminescent biosensors. Here, the expression of -operon under high temperature was studied. The expression was researched in the natural strain and in the heterologous system of . FV2201 bacterium was isolated from soil in the Moscow region (growth optimum 28 °C). We showed that its luminescence significantly increases when the temperature rises to 34 °C. The increase in luminescence is associated with an increase in the transcription of genes, which was confirmed by RT-PCR. The promoter of the -operon of the related bacterium ZM1 from the forests of Moldova, being cloned in the heterologous system of , is activated when the temperature rises from room temperature to 42 °C. When heat shock is caused by ethanol addition, transcription of -operon increases only in the natural strain of , but not in the heterologous system of cells. In addition, the activation of the -operon of persists in strains deficient in both the and genes. These results indicate the presence of sigma 32 and sigma 24 independent heat-shock-like mechanism of regulation of the -operon of in the heterologous system.
PubMed: 36950606
DOI: 10.1016/j.heliyon.2023.e14527 -
BMC Genomics Jan 2008Photorhabdus luminescens and Yersinia enterocolitica are both enteric bacteria which are associated with insects. P. luminescens lives in symbiosis with soil nematodes... (Comparative Study)
Comparative Study Review
BACKGROUND
Photorhabdus luminescens and Yersinia enterocolitica are both enteric bacteria which are associated with insects. P. luminescens lives in symbiosis with soil nematodes and is highly pathogenic towards insects but not to humans. In contrast, Y. enterocolitica is widely found in the environment and mainly known to cause gastroenteritis in men, but has only recently been shown to be also toxic for insects. It is expected that both pathogens share an overlap of genetic determinants that play a role within the insect host.
RESULTS
A selective genome comparison was applied. Proteins belonging to the class of two-component regulatory systems, quorum sensing, universal stress proteins, and c-di-GMP signalling have been analysed. The interorganismic synopsis of selected regulatory systems uncovered common and distinct signalling mechanisms of both pathogens used for perception of signals within the insect host. Particularly, a new class of LuxR-like regulators was identified, which might be involved in detecting insect-specific molecules. In addition, the genetic overlap unravelled a two-component system that is unique for the genera Photorhabdus and Yersinia and is therefore suggested to play a major role in the pathogen-insect relationship. Our analysis also highlights factors of both pathogens that are expressed at low temperatures as encountered in insects in contrast to higher (body) temperature, providing evidence that temperature is a yet under-investigated environmental signal for bacterial adaptation to various hosts. Common degradative metabolic pathways are described that might be used to explore nutrients within the insect gut or hemolymph, thus enabling the proliferation of P. luminescens and Y. enterocolitica in their invertebrate hosts. A strikingly higher number of genes encoding insecticidal toxins and other virulence factors in P. luminescens compared to Y. enterocolitica correlates with the higher virulence of P. luminescens towards insects, and suggests a putative broader insect host spectrum of this pathogen.
CONCLUSION
A set of factors shared by the two pathogens was identified including those that are involved in the host infection process, in persistence within the insect, or in host exploitation. Some of them might have been selected during the association with insects and then adapted to pathogenesis in mammalian hosts.
Topics: Animals; Bacterial Proteins; Biological Evolution; Genes, Bacterial; Genome, Bacterial; Humans; Insecta; Photorhabdus; Signal Transduction; Species Specificity; Virulence; Yersinia enterocolitica
PubMed: 18221513
DOI: 10.1186/1471-2164-9-40 -
Pathogens (Basel, Switzerland) Mar 2023Symbiotic bacteria form a mutualistic relationship with nematodes and are pathogenic to many insect pests. They kill insects using various strategies to evade or...
Symbiotic bacteria form a mutualistic relationship with nematodes and are pathogenic to many insect pests. They kill insects using various strategies to evade or suppress their humoral and cellular immunity. Here we evaluate the toxic effects of these bacteria and their secondary metabolites on the survival and phenoloxidase (PO) activation of larvae using biochemical and molecular methods. The results show H06 and All treatments caused significant reductions in the number of larvae in a dose-dependent manner. Secondly, the immune system recognizes symbiotic bacteria at early and late stages of infection via the induction of C-type lectin. Live symbiotic bacteria significantly inhibit PO activity in whereas heat-treated bacteria strongly increase PO activity. Additionally, expression levels of four proPhenoloxidase genes following treatment with H06 and All were compared. We found that the expression levels of all proPhenoloxidase genes were significantly down-regulated at all-time points. Similarly, treatments of larvae with metabolites benzylideneacetone and oxindole significantly down-regulated the expression of the PPO gene and inhibited PO activity. However, the addition of arachidonic acid to metabolite-treated larvae restored the expression level of the PPO gene and increased PO activity. Our results provide new insight into the roles of symbiotic bacteria in countering the insect phenoloxidase activation system.
PubMed: 37111392
DOI: 10.3390/pathogens12040506 -
PLoS Biology Nov 2020Lifeact is a short actin-binding peptide that is used to visualize filamentous actin (F-actin) structures in live eukaryotic cells using fluorescence microscopy....
Lifeact is a short actin-binding peptide that is used to visualize filamentous actin (F-actin) structures in live eukaryotic cells using fluorescence microscopy. However, this popular probe has been shown to alter cellular morphology by affecting the structure of the cytoskeleton. The molecular basis for such artefacts is poorly understood. Here, we determined the high-resolution structure of the Lifeact-F-actin complex using electron cryo-microscopy (cryo-EM). The structure reveals that Lifeact interacts with a hydrophobic binding pocket on F-actin and stretches over 2 adjacent actin subunits, stabilizing the DNase I-binding loop (D-loop) of actin in the closed conformation. Interestingly, the hydrophobic binding site is also used by actin-binding proteins, such as cofilin and myosin and actin-binding toxins, such as the hypervariable region of TccC3 (TccC3HVR) from Photorhabdus luminescens and ExoY from Pseudomonas aeruginosa. In vitro binding assays and activity measurements demonstrate that Lifeact indeed competes with these proteins, providing an explanation for the altering effects of Lifeact on cell morphology in vivo. Finally, we demonstrate that the affinity of Lifeact to F-actin can be increased by introducing mutations into the peptide, laying the foundation for designing improved actin probes for live cell imaging.
Topics: Actins; Animals; Bacterial Toxins; Binding Sites; Binding, Competitive; Cofilin 1; Cryoelectron Microscopy; Fluorescent Dyes; HEK293 Cells; Humans; Hydrophobic and Hydrophilic Interactions; In Vitro Techniques; Microfilament Proteins; Microscopy, Confocal; Models, Molecular; Myosins; Peptide Fragments; Protein Engineering; Protein Interaction Domains and Motifs; Rabbits; Recombinant Fusion Proteins; Saccharomyces cerevisiae Proteins
PubMed: 33216759
DOI: 10.1371/journal.pbio.3000925 -
Experimental Parasitology Sep 2019Only two drugs are currently available for the treatment of Chagas disease and their effectiveness are unsatisfactory. Photorhabdus luminescens and Xenorhabdus...
Only two drugs are currently available for the treatment of Chagas disease and their effectiveness are unsatisfactory. Photorhabdus luminescens and Xenorhabdus nematophila, two enteric bacteria highly pathogenic to a broad range of insects, have been studied as potential source for bioactive metabolites against protozoa causing neglected tropical diseases. Therefore, we tested the in vitro anti-Trypanosoma cruzi activity of secreted metabolites from these bacteria. The conditioned medium of X. nematophila and P. luminescens showed significant parasiticidal activity in a concentration-dependent manner (ICXN = 0.34 mg/mL, ICPL = 1.0 mg/mL). The parasiticidal compound was identified as a small molecule stable to heating and pH changes ranging from 2 to 12. Moreover, anti-Trypanosoma molecules secreted by both bacteria stimulate the trypanocidal activity of macrophages by a mechanism independent of nitric oxide. Summarizing, our studies reveal that P. luminescens and X. nematophila are potential sources of putative novel drugs against Chagas disease.
Topics: Analysis of Variance; Animals; Bacterial Proteins; Biological Assay; Chagas Disease; Culture Media, Conditioned; Endopeptidase K; Humans; Hydrogen-Ion Concentration; Inhibitory Concentration 50; Photorhabdus; Temperature; Trypanocidal Agents; Trypanosoma cruzi; Xenorhabdus
PubMed: 31279930
DOI: 10.1016/j.exppara.2019.107724 -
International Journal of Molecular... Jun 2022Due to its essential role in cellular processes, actin is a common target for bacterial toxins. One such toxin, TccC3, is an effector domain of the ABC-toxin produced by...
Due to its essential role in cellular processes, actin is a common target for bacterial toxins. One such toxin, TccC3, is an effector domain of the ABC-toxin produced by entomopathogenic bacteria of spp. Unlike other actin-targeting toxins, TccC3 uniquely ADP-ribosylates actin at Thr-148, resulting in the formation of actin aggregates and inhibition of phagocytosis. It has been shown that the fully modified F-actin is resistant to depolymerization by cofilin and gelsolin, but their effects on partially modified actin were not explored. We found that only F-actin unprotected by tropomyosin is the physiological TccC3 substrate. Yet, ADP-ribosylated G-actin can be produced upon cofilin-accelerated F-actin depolymerization, which was only mildly inhibited in partially modified actin. The affinity of TccC3-ADP-ribosylated G-actin for profilin and thymosin-β4 was weakened moderately but sufficiently to potentiate spontaneous polymerization in their presence. Interestingly, the Arp2/3-mediated nucleation was also potentiated by T148-ADP-ribosylation. Notably, even partially modified actin showed reduced bundling by plastins and α-actinin. In agreement with the role of these and other tandem calponin-homology domain actin organizers in the assembly of the cortical actin network, TccC3 induced intense membrane blebbing in cultured cells. Overall, our data suggest that TccC3 imposes a complex action on the cytoskeleton by affecting F-actin nucleation, recycling, and interaction with actin-binding proteins involved in the integration of actin filaments with each other and cellular elements.
Topics: ADP Ribose Transferases; Actin Cytoskeleton; Actin Depolymerizing Factors; Actins; Adenosine Diphosphate; Photorhabdus
PubMed: 35806028
DOI: 10.3390/ijms23137026 -
MBio Jun 2022With the overmining of actinomycetes for compounds acting against Gram-negative pathogens, recent efforts to discover novel antibiotics have been focused on other groups...
With the overmining of actinomycetes for compounds acting against Gram-negative pathogens, recent efforts to discover novel antibiotics have been focused on other groups of bacteria. Teixobactin, the first antibiotic without detectable resistance that binds lipid II, comes from an uncultured , a betaproteobacterium; odilorhabdins, from , are broad-spectrum inhibitors of protein synthesis, and darobactins from target BamA, the essential chaperone of the outer membrane of Gram-negative bacteria. and are symbionts of the nematode gut microbiome and attractive producers of secondary metabolites. Only small portions of their biosynthetic gene clusters (BGC) are expressed To access their silent operons, we first separated extracts from a small library of isolates into fractions, resulting in 200-fold concentrated material, and then screened them for antimicrobial activity. This resulted in a hit with selective activity against Escherichia coli, which we identified as a novel natural product antibiotic, 3'-amino 3'-deoxyguanosine (ADG). Mutants resistant to ADG mapped to and , kinases of guanosine. Biochemical analysis shows that ADG is a prodrug that is converted into an active ADG triphosphate (ADG-TP), a mimic of GTP. ADG incorporates into a growing RNA chain, interrupting transcription, and inhibits cell division, apparently by interfering with the GTPase activity of FtsZ. Gsk of the purine salvage pathway, which is the first kinase in the sequential phosphorylation of ADG, is restricted to E. coli and closely related species, explaining the selectivity of the compound. There are probably numerous targets of ADG-TP among GTP-dependent proteins. The discovery of ADG expands our knowledge of prodrugs, which are rare among natural compounds. Drug-resistant Gram-negative bacteria have become the major problem driving the antimicrobial resistance crisis. Searching outside the overmined actinomycetes, we focused on , gut symbionts of enthomopathogenic nematodes that carry up to 40 biosynthetic gene clusters coding for secondary metabolites. Most of these are silent and do not express . To gain access to silent operons, we first fractionated supernatant from and then tested 200-fold concentrated material for activity. This resulted in the isolation of a novel antimicrobial, 3'-amino 3'-deoxyguanosine (ADG), active against E. coli. ADG is an analog of guanosine and is converted into an active ADG-TP in the cell. ADG-TP inhibits transcription and probably numerous other GTP-dependent targets, such as FtsZ. Natural product prodrugs have been uncommon; discovery of ADG broadens our knowledge of this type of antibiotic.
Topics: Animals; Anti-Bacterial Agents; Bacterial Outer Membrane Proteins; Biological Products; Deoxyguanosine; Escherichia coli; Escherichia coli Proteins; Gram-Negative Bacteria; Guanosine; Guanosine Triphosphate; Nematoda; Operon; Photorhabdus; Prodrugs; Xenorhabdus
PubMed: 35575547
DOI: 10.1128/mbio.00700-22 -
Applied and Environmental Microbiology Jun 2022Phytopathogens represent a large agricultural challenge. The use of chemical pesticides is harmful to the environment, animals, and humans. Therefore, new sustainable...
Phytopathogens represent a large agricultural challenge. The use of chemical pesticides is harmful to the environment, animals, and humans. Therefore, new sustainable and biological alternatives are urgently needed. The insect-pathogenic bacterium Photorhabdus luminescens, already used in combination with entomopathogenic nematodes (EPNs) as a biocontrol agent, is characterized by two different phenotypic cell forms, called primary (1°) and secondary (2°). The 1° cells are symbiotic with EPNs and are used for biocontrol, and the 2° cells are unable to undergo symbiosis with EPNs, remain in the soil after insect infection, and specifically interact with plant roots. A previous RNA sequencing (RNAseq) analysis showed that genes encoding the exochitinase Chi2A and chitin binding protein (CBP) are highly upregulated in 2° cells exposed to plant root exudates. Here, we investigate Chi2A and CBP functions and demonstrate that both are necessary for P. luminescens 2° cells to inhibit the growth of the phytopathogenic fungus Fusarium graminearum. We provide evidence that Chi2A digests chitin and thereby inhibits fungal growth. Furthermore, we show that 2° cells specifically colonize fungal hyphae as one of the first mechanisms to protect plants from fungal phytopathogens. Finally, soil pot bioassays proved plant protection from F. graminearum by 2° cells, where Chi2A and CPB were essential for this process. This work gives molecular insights into the new applicability of as a plant-growth-promoting and plant-protecting organism in agriculture. The enteric enterobacterium Photorhabdus luminescens is already being used as a bioinsecticide since it is highly pathogenic toward a broad range of insects. However, the bacteria exist in two phenotypically different cell types, called 1° and 2° cells. Whereas only 1° cells are symbiotic with their nematode partner to infect insects, 2° cells were shown to remain in the soil after an insect infection cycle. It was demonstrated that 2° cells specifically interact with plant roots. Here, we show that the bacteria are beneficial for the plants by protecting them from phytopathogenic fungi. Specific colonization of the fungus mycelium as well as chitin-degrading activity mediated by the chitin binding protein (CBP) and the chitinase Chi2A are essential for this process. Our data give evidence for the novel future applicability of as a plant-growth-promoting organism and biopesticide.
Topics: Animals; Chitin; Fusarium; Insecta; Nematoda; Photorhabdus; Soil; Symbiosis
PubMed: 35604230
DOI: 10.1128/aem.00645-22 -
SpringerPlus 2014The Gram-negative bacterium Photorhabdus luminescens which symbiotically associates with the entomopathogenic nematode Heterorhabditis bacteriophora, has a broad...
The Gram-negative bacterium Photorhabdus luminescens which symbiotically associates with the entomopathogenic nematode Heterorhabditis bacteriophora, has a broad insecticidal and nematicidal activity. The virulence of P. luminescens toward the non-mutualistic nematode Caenorhabditis elegans has not been described. We showed that when fed on P. luminescens, the intestinal cells of C. elegans worms become delicate and some crystal-like structure was developed within the intestinal lumen. Next, we examined the requirement of the p38 mitogen-activated protein kinase (MAPK) and insulin/IGF-1 signaling pathway against P. luminescens. Depletion of pmk-1 by RNAi enhances susceptibility to P. luminescens, and numerous downstream targets regulated by the p38 MAPK pathway were induced when fed on P. luminescens. On the other hand, knockdown of daf-16 has no effects on C. elegans lifespan, but knockdown of daf-2 dramatically increased resistance to P. luminescens in a daf-16-dependent manner. We also revealed one of the daf-2 ligands ins-7 was induced and ins-7 deletion mutant survived longer when fed on P. luminescens. These results suggest the p38 MAPK pathway is activated and required for the host defense against P. luminescens. Insulin/IGF-1 signaling pathway is inactivated by P. luminescens through the overexpression of insulin-like gene.
PubMed: 25279274
DOI: 10.1186/2193-1801-3-274