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Molecular Microbiology May 2005Photorhabdus are bacteria found colonizing the gut of a specialized stage of the nematode Heterorhabditis, called the infective juvenile (IJ). The IJ is a free-living...
Photorhabdus are bacteria found colonizing the gut of a specialized stage of the nematode Heterorhabditis, called the infective juvenile (IJ). The IJ is a free-living stage of the nematode that seeks out and infects insect larvae. Once inside the insect the IJ release Photorhabdus into the haemolymph where the bacteria rapidly proliferate, killing the insect within 48-72 h. The nematodes grow and reproduce in the insect cadaver by feeding on the Photorhabdus biomass. In this study we use Photorhabdus temperata K122 to show that genes involved in iron acquisition play a key role during the course of the tripartite bacteria-nematode-insect interaction. We show that a strain carrying a mutation in a gene with homology to exbD, encoding a component of the TonB complex, is unable to grow well in conditions where iron is not freely available. In addition, this mutant, BMM417, requires a longer time to kill the insect larvae than the wild-type bacteria and this defect in pathogenicity is complemented by the co-injection of iron. Moreover, the increase in LT(50) observed with BMM417 is correlated with a significantly slower in vivo growth rate suggesting that iron is limiting in the insect. We also show that BMM417 is unable to support the growth and development of the Heterorhabditis nematode. Addition of exogenous iron to the growth media restores nematode growth and development on BMM417, suggesting that aspects of iron metaboism in Photorhabdus are important during the symbiosis with the nematode.
Topics: Animals; Escherichia coli Proteins; Gene Expression Regulation, Bacterial; Genetic Complementation Test; Homeostasis; Host-Parasite Interactions; Hydroxybenzoates; Insecta; Iron; Larva; Mutation; Parasitic Diseases, Animal; Photorhabdus; Rhabditida Infections; Rhabditoidea; Symbiosis; Virulence
PubMed: 15819630
DOI: 10.1111/j.1365-2958.2005.04574.x -
Chembiochem : a European Journal of... May 2021The glidobactin-like natural products (GLNPs) glidobactin A and cepafungin I have been reported to be potent proteasome inhibitors and are regarded as promising...
The glidobactin-like natural products (GLNPs) glidobactin A and cepafungin I have been reported to be potent proteasome inhibitors and are regarded as promising candidates for anticancer drug development. Their biosynthetic gene cluster (BGC) plu1881-1877 is present in entomopathogenic Photorhabdus laumondii but silent under standard laboratory conditions. Here we show the largest subset of GLNPs, which are produced and identified after activation of the silent BGC in the native host and following heterologous expression of the BGC in Escherichia coli. Their chemical diversity results from a relaxed substrate specificity and flexible product release in the assembly line of GLNPs. Crystal structure analysis of the yeast proteasome in complex with new GLNPs suggests that the degree of unsaturation and the length of the aliphatic tail are critical for their bioactivity. The results in this study provide the basis to engineer the BGC for the generation of new GLNPs and to optimize these natural products resulting in potential drugs for cancer therapy.
Topics: Bacterial Proteins; Drug Design; Escherichia coli; Multigene Family; Peptides, Cyclic; Photorhabdus; Proteasome Inhibitors; Structure-Activity Relationship
PubMed: 33452852
DOI: 10.1002/cbic.202100014 -
Insects May 2021The meadow spittlebug (Hemiptera: Aphrophoridae) is the primary vector of (Proteobacteria: Xanthomonadaceae) in Europe, a pest-disease complex of economically relevant...
The meadow spittlebug (Hemiptera: Aphrophoridae) is the primary vector of (Proteobacteria: Xanthomonadaceae) in Europe, a pest-disease complex of economically relevant crops such as olives, almonds, and grapevine, managed mainly through the use of broad-spectrum pesticides. Providing environmentally sound alternatives to reduce the reliance on chemical control is a primary challenge in the control of and, hence, in the protection of crops against the expansion of its associated bacterial pathogen. Entomopathogenic nematodes (EPNs) are well-known biocontrol agents of soil-dwelling arthropods. Recent technological advances in field applications, including improvements in obtaining cell-free supernatant from their symbiotic bacteria, allow their successful implementation against aerial pests. Thus, this study aimed to evaluate, for the first time, the efficacy of EPN applications against nymphal instars of We tested four EPN species and the cell-free supernatant of their corresponding symbiotic bacteria: -, -, -, and - subsp. . First, we showed that 24 and 72 h exposure to the foam produced by nymphs did not affect virulence. The direct application of steinernematid EPNs provided promising results, reaching 90, 78, and 53% nymphal mortality rates after five days of exposure for , , and , respectively. Conversely, the application of the cell-free supernatant from resulted in nymphal mortalities of 64%, significantly higher than observed for species after five days of exposure. Overall, we demonstrated the great potential of the application of specific EPNs and cell-free supernatant of their symbiont bacteria against , introducing new opportunities to develop them as biopesticides for integrated management practices or organic vineyard production.
PubMed: 34068952
DOI: 10.3390/insects12050448 -
PloS One 2015Quorum sensing is a typical communication system among Gram-negative bacteria used to control group-coordinated behavior via small diffusible molecules dependent on cell...
Quorum sensing is a typical communication system among Gram-negative bacteria used to control group-coordinated behavior via small diffusible molecules dependent on cell number. The key components of a quorum sensing system are a LuxI-type synthase, producing acyl-homoserine lactones (AHLs) as signaling molecules, and a LuxR-type receptor that detects AHLs to control expression of specific target genes. Six conserved amino acids are present in the signal-binding domain of AHL-sensing LuxR-type proteins, which are important for ligand-binding and -specificity as well as shaping the ligand-binding pocket. However, many proteobacteria possess LuxR-type regulators without a cognate LuxI synthase, referred to as LuxR solos. The two LuxR solos PluR and PauR from Photorhabdus luminescens and Photorhabdus asymbiotica, respectively, do not sense AHLs. Instead PluR and PauR sense alpha-pyrones and dialkylresorcinols, respectively, and are part of cell-cell communication systems contributing to the overall virulence of these Photorhabdus species. However, PluR and PauR both harbor substitutions in the conserved amino acid motif compared to that in AHL sensors, which appeared to be important for binding the corresponding signaling molecules. Here we analyze the role of the conserved amino acids in the signal-binding domain of these two non-AHL LuxR-type receptors for their role in signal perception. Our studies reveal that the conserved amino acid motif alone is essential but not solely responsible for ligand-binding.
Topics: Acyl-Butyrolactones; Amino Acid Motifs; Bacterial Proteins; Conserved Sequence; Gene Expression Regulation, Bacterial; Ligands; Photorhabdus; Quorum Sensing; Repressor Proteins; Trans-Activators; Transcription Factors
PubMed: 25923884
DOI: 10.1371/journal.pone.0124093 -
Molecular and Cellular Probes Feb 2019Acute hepatopancreatic necrosis disease (AHPND), also known as Early mortality syndrome (EMS), is a recently emerged lethal disease that has caused major economic losses...
Acute hepatopancreatic necrosis disease (AHPND), also known as Early mortality syndrome (EMS), is a recently emerged lethal disease that has caused major economic losses in shrimp aquaculture. The etiologic agents are Vibrio spp. that carry Photorhabdus Insect-Related (Pir) toxin genes pirA and pirB. A multiplex SYBR Green real-time PCR was developed that detects pirA, pirB, and two internal control genes, the shrimp 18S rRNA and the bacterial 16S rRNA genes in a single reaction. The pirB primers amplify the 3'-end of the pirB gene allowing the detection of Vibrio spp. mutants that contain a complete deletion of pirA and the partial deletion of pirB. The assay also detects mutants that contain the entire pirA gene and the deletion of the pirB gene. Since both toxin genes are needed for disease development, this assays can distinguish between pathogenic strains of Vibrio spp. that cause AHPND in shrimp and mutants that do not cause disease. The amplicons for pirA, pirB, 18S rRNA and 16S rRNA showed easily distinguishable melting temperatures of 78.21 ± 0.18, 75.20 ± 0.20, 82.28 ± 0.34 and 85.41 ± 0.21 °C respectively. Additionally, a duplex real-time PCR assay was carried out by designing TaqMan probes for the pirA and pirB primers. The diagnostic sensitivity and specificity was compared between the SYBR Green and TaqMan assays. Both assays showed similar sensitivity with a limit of detection being 10 copies for pirA and pirB, and neither assays showed any cross reaction with other known bacterial and viral pathogens in shrimp. The high sensitivity of both assays make them suitable for the detection of low copies of the pirA and pirB genes in AHPND causing Vibrio spp. as well as for detecting non-pathogenic mutants.
Topics: Animals; Bacterial Toxins; Gene Deletion; Genes, Bacterial; Insecta; Photorhabdus; Real-Time Polymerase Chain Reaction; Reference Standards; Vibrio parahaemolyticus
PubMed: 30576786
DOI: 10.1016/j.mcp.2018.12.004 -
Applied and Environmental Microbiology Jul 2004Photorhabdus sp. strain Az29 is symbiotic with an Azorean nematode of the genus Heterorhabditis in a complex that is highly virulent to insects even at low temperatures....
Photorhabdus sp. strain Az29 is symbiotic with an Azorean nematode of the genus Heterorhabditis in a complex that is highly virulent to insects even at low temperatures. The virulence of the bacteria is mainly attributed to toxins and bacterial enzymes secreted during parasitism. The bacteria secrete proteases during growth, with a peak at the end of the exponential growth phase. Protease secretion was higher in cultures growing at lower temperatures. At 10 degrees C the activity was highest and remained constant for over 7 days, whereas at 23 and 28 degrees C it showed a steady decrease. Two proteases, PrtA and PrtS, that are produced in the growth medium were purified by liquid chromatography. PrtA was inhibited by 1,10-phenantroline and by EDTA and had a molecular mass of 56 kDa and an optimal activity at pH 9 and 50 degrees C. Sequences of three peptides of PrtA showed strong homologies with alkaline metalloproteases from Photorhabdus temperata K122 and Photorhabdus luminescens W14. Peptide PrtA-36 contained the residues characteristic of metzincins, known to be involved in bacterial virulence. In vitro, PrtA inhibited antibacterial factors of inoculated Lepidoptera and of cecropins A and B. PrtS had a molecular mass of 38 kDa and was inhibited by 1,10-phenanthroline but not by EDTA. Its activity ranged between 10 and 80 degrees C and was optimal at pH 7 and 50 degrees C. PrtS also destroyed insect antibacterial factors. Three fragments of PrtS showed homology with a putative metalloprotease of P. luminescens TTO1. Polyclonal antibody raised against PrtA did not recognize PrtS, showing they are distinct molecules.
Topics: Amino Acid Sequence; Animals; Antimicrobial Cationic Peptides; Hemolymph; Metalloendopeptidases; Molecular Sequence Data; Photorhabdus; Temperature
PubMed: 15240252
DOI: 10.1128/AEM.70.7.3831-3838.2004 -
Virulence Nov 2017Previous and recent investigations on the innate immune response of Drosophila have identified certain mechanisms that promote pathogen elimination. However, the...
Previous and recent investigations on the innate immune response of Drosophila have identified certain mechanisms that promote pathogen elimination. However, the function of Thioester-containing proteins (TEPs) in the fly still remains elusive. Recently we have shown the contribution of TEP4 in the antibacterial immune defense of Drosophila against non-pathogenic E. coli, and the pathogens Photorhabdus luminescens and P. asymbiotica. Here we studied the function of Tep genes in both humoral and cellular immunity upon E. coli and Photorhabdus infection. We found that while Tep2 is induced after Photorhabdus and E. coli infection; Tep6 is induced by P. asymbiotica only. Moreover, functional ablation of hemocytes results in significantly low transcript levels of Tep2 and Tep6 in response to Photorhabdus. We show that Tep2 and Tep6 loss-of-function mutants have prolonged survival against P. asymbiotica, Tep6 mutants survive better the infection of P. luminescens, and both tep mutants are resistant to E. coli and Photorhabdus. We also find a distinct pattern of immune signaling pathway induction in E. coli or Photorhabdus infected Tep2 and Tep6 mutants. We further show that Tep2 and Tep6 participate in the activation of hemocytes in Drosophila responding to Photorhabdus. Finally, inactivation of Tep2 or Tep6 affects phagocytosis and melanization in flies infected with Photorhabdus. Our results indicate that distinct Tep genes might be involved in different yet crucial functions in the Drosophila antibacterial immune response.
Topics: Animals; Cytokines; Drosophila Proteins; Drosophila melanogaster; Escherichia coli; Hemocytes; Immunity, Innate; Phagocytosis; Photorhabdus; Serpins
PubMed: 28498729
DOI: 10.1080/21505594.2017.1330240 -
Microbes and Infection Feb 2004The three currently recognised Photorhabdus species are bioluminescent bacteria that are pathogenic to insects. P. luminescens and P. temperata form a symbiotic... (Review)
Review
The three currently recognised Photorhabdus species are bioluminescent bacteria that are pathogenic to insects. P. luminescens and P. temperata form a symbiotic relationship with nematodes that infect insects. P. asymbiotica, on the other hand, has only been isolated from human clinical specimens from the USA and Australia. The bacterium has been associated with locally invasive soft tissue and disseminated bacteraemic infections. An invertebrate vector for P. asymbiotica has not yet been identified.
Topics: Communicable Diseases, Emerging; Humans; Luminescent Measurements; Photorhabdus; Rhabditida Infections
PubMed: 15049334
DOI: 10.1016/j.micinf.2003.10.018 -
The Journal of Antibiotics Aug 2016Photorhabdus luminescens is a bioluminescent entomopathogenic bacterium that undergoes phenotypic variation and lives in mutualistic association with nematodes of the...
Photorhabdus luminescens is a bioluminescent entomopathogenic bacterium that undergoes phenotypic variation and lives in mutualistic association with nematodes of the family Heterorhabditidae. The pair infects and kills insects, and during their coordinated lifecycle, the bacteria produce an assortment of specialized metabolites to regulate its mutualistic and pathogenic roles. As part of our search for new specialized metabolites from the Photorhabdus genus, we examined organic extracts from P. luminescens grown in an amino-acid-rich medium based on the free amino-acid levels found in the circulatory fluid of its common insect prey, the Galleria mellonella larva. Reversed-phase HPLC/UV/MS-guided fractionation of the culture extracts led to the identification of two new pyrazinone metabolites, lumizinones A (1) and B (2), together with two N-acetyl dipeptides (3 and 4). The lumizinones were produced only in the phenotypic variant associated with nematode development and insect pathogenesis. Their chemical structures were elucidated by analysis of 1D and 2D NMR and high-resolution ESI-QTOF-MS spectral data. The absolute configurations of the amino acids in 3 and 4 were determined by Marfey's analysis. Compounds 1-4 were evaluated for their calpain protease inhibitory activity, and lumizinone A (1) showed inhibition with an IC50 (half-maximal inhibitory concentration) value of 3.9 μm.
Topics: Amino Acids; Animals; Chromatography, High Pressure Liquid; Inhibitory Concentration 50; Lepidoptera; Magnetic Resonance Spectroscopy; Mass Spectrometry; Photorhabdus; Protease Inhibitors; Pyrazines; Spectrometry, Mass, Electrospray Ionization
PubMed: 27353165
DOI: 10.1038/ja.2016.79 -
Insects Nov 2022The codling moth, (L.) (Lepidoptera: Tortricidae), is one of the major pests in pome fruit production worldwide. Heavy treatment of the larvae of with insecticides...
Identification and Biocontrol Potential of Entomopathogenic Nematodes and Their Endosymbiotic Bacteria in Apple Orchards against the Codling Moth, (L.) (Lepidoptera: Tortricidae).
The codling moth, (L.) (Lepidoptera: Tortricidae), is one of the major pests in pome fruit production worldwide. Heavy treatment of the larvae of with insecticides triggered the development of resistance to many groups of insecticides. In addition, the increasing concern about the adverse effects of synthetic insecticides on human health and the environment has led to the development of sustainable and eco-friendly control practices for . The entomopathogenic nematodes (EPNs) ( and spp.) and their endosymbionts ( and spp.) represent a newly emerging approach to controlling a wide range of insect pests. In the present study, field surveys were conducted in apple orchards to isolate and identify EPNs and their endosymbionts and evaluate their insecticidal efficacy on the larvae of . EPNs were isolated from 12 of 100 soil samples (12%). Seven samples were identified as (Filipjev, 1934) (Rhabditida: Steinernematidae), whereas five samples were assigned to (Poinar, 1976) (Rhabditida: Heterorhabditidae). The pathogenicity of the EPN species/isolates was screened on the last instar larvae of . The two most pathogenic isolates from each EPN species were tested against fifth instar larvae of under controlled conditions. The maximum mortality (100%) was achieved by all EPN species/isolates at a concentration of 100 IJs/larva 96 h after treatment. The endosymbionts of selected and species were identified as subsp. and , respectively. The mortality rates ranged between 25 and 62% when the fifth larval instar larvae of were exposed to the treatment of cell-free supernatants of symbiotic bacteria. In essence, the present survey indicated that EPNs and their symbiotic bacteria have good potential for biological control of .
PubMed: 36554995
DOI: 10.3390/insects13121085