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Scientific Reports Jun 2022Natural products have been proven to be important starting points for the development of new drugs. Bacteria in the genera Photorhabdus and Xenorhabdus produce...
Natural products have been proven to be important starting points for the development of new drugs. Bacteria in the genera Photorhabdus and Xenorhabdus produce antimicrobial compounds as secondary metabolites to compete with other organisms. Our study is the first comprehensive study screening the anti-protozoal activity of supernatants containing secondary metabolites produced by 5 Photorhabdus and 22 Xenorhabdus species against human parasitic protozoa, Acanthamoeba castellanii, Entamoeba histolytica, Trichomonas vaginalis, Leishmania tropica and Trypanosoma cruzi, and the identification of novel bioactive antiprotozoal compounds using the easyPACId approach (easy Promoter Activated Compound Identification) method. Though not in all species, both bacterial genera produce antiprotozoal compounds effective on human pathogenic protozoa. The promoter exchange mutants revealed that antiprotozoal bioactive compounds produced by Xenorhabdus bacteria were fabclavines, xenocoumacins, xenorhabdins and PAX peptides. Among the bacteria assessed, only P. namnaoensis appears to have acquired amoebicidal property which is effective on E. histolytica trophozoites. These discovered antiprotozoal compounds might serve as starting points for the development of alternative and novel pharmaceutical agents against human parasitic protozoa in the future.
Topics: Antiprotozoal Agents; Entamoeba histolytica; Humans; Photorhabdus; Trypanosoma cruzi; Xenorhabdus
PubMed: 35750682
DOI: 10.1038/s41598-022-13722-z -
BMC Genomics May 2008Photorhabdus luminescens is a Gram-negative luminescent enterobacterium and a symbiote to soil nematodes belonging to the species Heterorhabditis bacteriophora....
BACKGROUND
Photorhabdus luminescens is a Gram-negative luminescent enterobacterium and a symbiote to soil nematodes belonging to the species Heterorhabditis bacteriophora. P.luminescens is simultaneously highly pathogenic to insects. This bacterium exhibits a complex life cycle, including one symbiotic stage characterized by colonization of the upper nematode gut, and a pathogenic stage, characterized by release from the nematode into the hemocoel of insect larvae, resulting in rapid insect death caused by bacterial toxins. P. luminescens appears to sense and adapt to the novel host environment upon changing hosts, which facilitates the production of factors involved in survival within the host, host-killing, and -exploitation.
RESULTS
A differential fluorescence induction (DFI) approach was applied to identify genes that are up-regulated in the bacterium after infection of the insect host Galleria mellonella. For this purpose, a P. luminescens promoter-trap library utilizing the mCherry fluorophore as a reporter was constructed, and approximately 13,000 clones were screened for fluorescence induction in the presence of a G. mellonella larvae homogenate. Since P. luminescens has a variety of regulators that potentially sense chemical molecules, like hormones, the screen for up-regulated genes or operons was performed in vitro, excluding physicochemical signals like oxygen, temperature or osmolarity as variables. Clones (18) were obtained exhibiting at least 2.5-fold induced fluorescence and regarded as specific responders to insect homogenate. In combination with a bioinformatics approach, sequence motifs were identified in these DNA-fragments that are similar to 29 different promoters within the P. luminescens genome. By cloning each of the predicted promoters upstream of the reporter gene, induction was verified for 27 promoters in vitro, and for 24 promoters in viable G. mellonella larvae. Among the validated promoters are some known to regulate the expression of toxin genes, including tccC1 (encoding an insecticidal toxin complex), and others encoding putative toxins. A comparably high number of metabolic genes or operons were observed to be induced upon infection; among these were eutABC, hutUH, and agaZSVCD, which encode proteins involved in ethanolamine, histidine and tagatose degradation, respectively. The results reflect rearrangements in metabolism and the use of other metabolites available from the insect. Furthermore, enhanced activity of promoters controlling the expression of genes encoding enzymes linked to antibiotic production and/or resistance was observed. Antibiotic production and resistance may influence competition with other bacteria, and thus might be important for a successful infection. Lastly, several genes of unknown function were identified that may represent novel pathogenicity factors.
CONCLUSION
We show that a DFI screen is useful for identifying genes or operons induced by chemical stimuli, such as diluted insect homogenate. A bioinformatics comparison of motifs similar to known promoters is a powerful tool for identifying regulated genes or operons. We conclude that signals for the regulation of those genes or operons induced in P. luminescens upon insect infection may represent a wide variety of compounds that make up the insect host. Our results provide insight into the complex response to the host that occurs in a bacterial pathogen, particularly reflecting the potential for metabolic shifts and other specific changes associated with virulence.
Topics: Animals; Base Sequence; Computational Biology; DNA, Bacterial; Fluorescence; Gene Expression; Genes, Bacterial; Genes, Reporter; Host-Pathogen Interactions; Larva; Luminescent Proteins; Moths; Photorhabdus; Promoter Regions, Genetic; Recombinant Fusion Proteins; Red Fluorescent Protein
PubMed: 18489737
DOI: 10.1186/1471-2164-9-229 -
BMC Genomics Nov 2022Nematodes are a major group of soil inhabiting organisms. Heterorhabditis nematodes are insect-pathogenic nematodes and live in a close symbiotic association with...
BACKGROUND
Nematodes are a major group of soil inhabiting organisms. Heterorhabditis nematodes are insect-pathogenic nematodes and live in a close symbiotic association with Photorhabdus bacteria. Heterorhabditis-Photorhabdus pair offers a powerful and genetically tractable model to study animal-microbe symbiosis. It is possible to generate symbiont bacteria free (axenic) stages in Heterorhabditis. Here, we compared the transcriptome of symbiotic early-adult stage Heterorhabditis nematodes with axenic early-adult nematodes to determine the nematode genes and pathways involved in symbiosis with Photorhabdus bacteria.
RESULTS
A de-novo reference transcriptome assembly of 95.7 Mb was created for H. bacteriophora by using all the reads. The assembly contained 46,599 transcripts with N50 value of 2,681 bp and the average transcript length was 2,054 bp. The differentially expressed transcripts were identified by mapping reads from symbiotic and axenic nematodes to the reference assembly. A total of 754 differentially expressed transcripts were identified in symbiotic nematodes as compared to the axenic nematodes. The ribosomal pathway was identified as the most affected among the differentially expressed transcripts. Additionally, 12,151 transcripts were unique to symbiotic nematodes. Endocytosis, cAMP signalling and focal adhesion were the top three enriched pathways in symbiotic nematodes, while a large number of transcripts coding for various responses against bacteria, such as bacterial recognition, canonical immune signalling pathways, and antimicrobial effectors could also be identified.
CONCLUSIONS
The symbiotic Heterorhabditis nematodes respond to the presence of symbiotic bacteria by expressing various transcripts involved in a multi-layered immune response which might represent non-systemic and evolved localized responses to maintain mutualistic bacteria at non-threatening levels. Subject to further functional validation of the identified transcripts, our findings suggest that Heterorhabditis nematode immune system plays a critical role in maintenance of symbiosis with Photorhabdus bacteria.
Topics: Animals; Photorhabdus; Rhabditoidea; Symbiosis; Sequence Analysis, RNA; RNA
PubMed: 36344922
DOI: 10.1186/s12864-022-08952-4 -
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 -
PloS One 2021Xenorhabdus and Photorhabdus are gram negative bacteria that can produce several secondary metabolites, including antimicrobial compounds. They have a symbiotic...
Xenorhabdus and Photorhabdus are gram negative bacteria that can produce several secondary metabolites, including antimicrobial compounds. They have a symbiotic association with entomopathogenic nematodes (EPNs). The aim of this study was to isolate and identify Xenorhabdus and Photorhabdus species and their associated nematode symbionts from Northeastern region of Thailand. We also evaluated the antibacterial activity of these symbiotic bacteria. The recovery rate of EPNs was 7.82% (113/1445). A total of 62 Xenorhabdus and 51 Photorhabdus strains were isolated from the EPNs. Based on recA sequencing and phylogeny, Xenorhabdus isolates were identified as X. stockiae (n = 60), X. indica (n = 1) and X. eapokensis (n = 1). Photorhabdus isolates were identified as P. luminescens subsp. akhurstii (n = 29), P. luminescens subsp. hainanensis (n = 18), P. luminescens subsp. laumondii (n = 2), and P. asymbiotica subsp. australis (n = 2). The EPNs based on 28S rDNA and internal transcribed spacer (ITS) analysis were identified as Steinernema surkhetense (n = 35), S. sangi (n = 1), unidentified Steinernema (n = 1), Heterorhabditis indica (n = 39), H. baujardi (n = 1), and Heterorhabditis sp. SGmg3 (n = 3). Antibacterial activity showed that X. stockiae (bMSK7.5_TH) extract inhibited several antibiotic-resistant bacterial strains. To the best of our knowledge, this is the first report on mutualistic association between P. luminescens subsp. laumondii and Heterorhabditis sp. SGmg3. This study could act as a platform for future studies focusing on the discovery of novel antimicrobial compounds from these bacterial isolates.
Topics: Animals; Anti-Bacterial Agents; Drug Resistance, Bacterial; Gram-Negative Bacteria; Gram-Positive Bacteria; Larva; Microbial Sensitivity Tests; Nematoda; Photorhabdus; Phylogeny; RNA, Ribosomal, 16S; RNA, Ribosomal, 28S; Soil; Soil Microbiology; Symbiosis; Xenorhabdus
PubMed: 34383819
DOI: 10.1371/journal.pone.0255943 -
BMC Microbiology May 2010Photorhabdus are Gram-negative nematode-symbiotic and insect-pathogenic bacteria. The species Photorhabdus asymbiotica is able to infect humans as well as insects. We...
BACKGROUND
Photorhabdus are Gram-negative nematode-symbiotic and insect-pathogenic bacteria. The species Photorhabdus asymbiotica is able to infect humans as well as insects. We investigated the secreted proteome of a clinical isolate of P. asymbiotica at different temperatures in order to identify proteins relevant to the infection of the two different hosts.
RESULTS
A comparison of the proteins secreted by a clinical isolate of P. asymbiotica at simulated insect (28 degrees C) and human (37 degrees C) temperatures led to the identification of a small and highly abundant protein, designated Pam, that is only secreted at the lower temperature. The pam gene is present in all Photorhabdus strains tested and shows a high level of conservation across the whole genus, suggesting it is both ancestral to the genus and probably important to the biology of the bacterium. The Pam protein shows limited sequence similarity to the 13.6 kDa component of a binary toxin of Bacillus thuringiensis. Nevertheless, injection or feeding of heterologously produced Pam showed no insecticidal activity to either Galleria mellonella or Manduca sexta larvae. In bacterial colonies, Pam is associated with an extracellular polysaccharide (EPS)-like matrix, and modifies the ability of wild-type cells to attach to an artificial surface. Interestingly, Surface Plasmon Resonance (SPR) binding studies revealed that the Pam protein itself has adhesive properties. Although Pam is produced throughout insect infection, genetic knockout does not affect either insect virulence or the ability of P. luminescens to form a symbiotic association with its host nematode, Heterorhabditis bacteriophora.
CONCLUSIONS
We studied a highly abundant protein, Pam, which is secreted in a temperature-dependent manner in P. asymbiotica. Our findings indicate that Pam plays an important role in enhancing surface attachment in insect blood. Its association with exopolysaccharide suggests it may exert its effect through mediation of EPS properties. Despite its abundance and conservation in the genus, we find no evidence for a role of Pam in either virulence or symbiosis.
Topics: Adhesins, Bacterial; Animals; Bacillus thuringiensis; Bacterial Adhesion; Bacterial Proteins; Bacterial Toxins; Electrophoresis, Gel, Two-Dimensional; Gram-Negative Bacterial Infections; Humans; Lepidoptera; Nematoda; Photorhabdus; Polysaccharides, Bacterial; Proteome; Sequence Homology, Amino Acid; Surface Plasmon Resonance; Symbiosis; Temperature; Virulence
PubMed: 20462430
DOI: 10.1186/1471-2180-10-141 -
Journal of Invertebrate Pathology Mar 2022Bacterial symbionts associated with entomopathogenic nematodes (EPNs) play an important role in terms of the insecticidal properties of nematodes in pest control....
Bacterial symbionts associated with entomopathogenic nematodes (EPNs) play an important role in terms of the insecticidal properties of nematodes in pest control. Galleria mellonella larvae, shortly after being infected with three different strains of Heterorhabditis zealandica, which were isolated from South African soil, changed from pale white to steel grey-blue (blue), bright red, and yellow with a green tint (green), respectively. The genetic relatedness of the bacterial symbionts that were isolated from the three strains of H. zealandica was determined by means of comparing the 16S rRNA, recA, gyrB, dnaN, gltX and infB gene sequences. Subsequently, comparing the concatenated sequences revealed the presence of three distinct Photorhabdus species. The H. zealandica strain SF41, associated with Photorhabdus heterorhabditis, produced 'blue' G. mellonella larvae. The H. zealandica strain MJ2C, associated with Photorhabdus thracensis, yielded 'green' G. mellonella larvae, while the H. zealandica strain LLM associated with Photorhabdus laumondii subsp. laumondii yielded red larvae. The colour changes in G. mellonella larvae were found to have been instigated by a particular Photorhabdus species associated with H. zealandica. The red and 'green' phenotypes of G. mellonella larvae were found to represent new combinations of Heterorhabditis and Photorhabdus. In future studies, the colour of infected G. mellonella larvae needs to be reported as a phenotypic character, as it indicates the different bacterial species associated with the same nematode host, as shown in the case of H. zealandica.
Topics: Animals; Color; Larva; Moths; Nematoda; Photorhabdus; RNA, Ribosomal, 16S; Strongyloidea
PubMed: 35124069
DOI: 10.1016/j.jip.2022.107729 -
Applied and Environmental Microbiology Aug 2020The number of sustainable agriculture techniques to improve pest management and environmental safety is rising, as biological control agents are used to enhance disease...
The number of sustainable agriculture techniques to improve pest management and environmental safety is rising, as biological control agents are used to enhance disease resistance and abiotic stress tolerance in crops. Here, we investigated the capacity of the secondary variant to react to plant root exudates and their behavior toward microorganisms in the rhizosphere. is known to live in symbiosis with entomopathogenic nematodes (EPNs) and to be highly pathogenic toward insects. The -EPN relationship has been widely studied, and this combination has been used as a biological control agent; however, not much attention has been paid to the putative lifestyle of in the rhizosphere. We performed transcriptome analysis to show how responds to plant root exudates. The analysis highlighted genes involved in chitin degradation, biofilm regulation, formation of flagella, and type VI secretion system. Furthermore, we provide evidence that can inhibit growth of phytopathogenic fungi. Finally, we demonstrated a specific interaction of with plant roots. Understanding the role and the function of this bacterium in the rhizosphere might accelerate the progress in biocontrol manipulation and elucidate the peculiar mechanisms adopted by plant growth-promoting rhizobacteria in plant root interactions. Insect-pathogenic bacteria are widely used in biocontrol strategies against pests. Very little is known about the life of these bacteria in the rhizosphere. Here, we show that can specifically react to and interact with plant roots. Understanding the adaptation of in the rhizosphere is highly important for the biotechnological application of entomopathogenic bacteria and could improve future sustainable pest management in agriculture.
Topics: Biological Control Agents; Chemotaxis; Exudates and Transudates; Fungi; Gene Expression Profiling; Genes, Bacterial; Photorhabdus; Plant Roots; RNA-Seq; Rhizosphere
PubMed: 32591378
DOI: 10.1128/AEM.00891-20 -
PLoS Pathogens 2012The Toxin Complex (TC) is a large multi-subunit toxin first characterized in the insect pathogens Photorhabdus and Xenorhabdus, but now seen in a range of pathogens,...
The Toxin Complex (TC) is a large multi-subunit toxin first characterized in the insect pathogens Photorhabdus and Xenorhabdus, but now seen in a range of pathogens, including those of humans. These complexes comprise three protein subunits, A, B and C which in the Xenorhabdus toxin are found in a 4:1:1 stoichiometry. Some TCs have been demonstrated to exhibit oral toxicity to insects and have the potential to be developed as a pest control technology. The lack of recognisable signal sequences in the three large component proteins hinders an understanding of their mode of secretion. Nevertheless, we have shown the Photorhabdus luminescens (Pl) Tcd complex has been shown to associate with the bacteria's surface, although some strains can also release it into the surrounding milieu. The large number of tc gene homologues in Pl make study of the export process difficult and as such we have developed and validated a heterologous Escherichia coli expression model to study the release of these important toxins. In addition to this model, we have used comparative genomics between a strain that releases high levels of Tcd into the supernatant and one that retains the toxin on its surface, to identify a protein responsible for enhancing secretion and release of these toxins. This protein is a putative lipase (Pdl1) which is regulated by a small tightly linked antagonist protein (Orf53). The identification of homologues of these in other bacteria, linked to other virulence factor operons, such as type VI secretion systems, suggests that these genes represent a general and widespread mechanism for enhancing toxin release in gram negative pathogens.
Topics: Animals; Bacterial Proteins; Bacterial Secretion Systems; Bacterial Toxins; Escherichia coli; Larva; Lipase; Manduca; Membrane Proteins; Photorhabdus; Xenorhabdus
PubMed: 22615559
DOI: 10.1371/journal.ppat.1002692 -
Journal of Invertebrate Pathology Oct 2022Photorhabdus insect related proteins A & B (PirA, PirB) from Photorhabdus and Xenorhabdus bacteria exhibit both oral and injectable toxicity against lepidopteran and...
Photorhabdus insect related proteins A & B (PirA, PirB) from Photorhabdus and Xenorhabdus bacteria exhibit both oral and injectable toxicity against lepidopteran and dipteran insect pest. The pirA, pirAt (encoding 6 N-terminal truncated PirA), pirB genes, pirA-pirB (with ERIC sequences), pirA-pirB-mERIC (modified pirA-pirB with mutated ERIC sequences) and polycistronic-pirAB were cloned and expressed in Escherichia coli. However, PirA protein was expressed in insoluble form and therefore the pirA gene was modified to produce PirAt. Moreover, pirA-pirB-mERIC, polycistronic-pirAB and co-transformed pirA/pirB genes were not expressed in the studied prokaryotic expression systems. None of the single purified proteins or mixtures of the individually expressed and purified proteins were toxic to mosquito larvae of Aedes aegypti and Culex quinquefasciatus. However, PirA-PirB protein mixtures purified from pirA-pirB operon plasmid were toxic to A. aegypti and C. quinquefasciatus larvae with LC values of 991 and 614 ng/ml, respectively. The presence of ERIC sequences between the two orfs of the pirA-pirB operon could help to obtain the proteins in biologically active form. Further, results confirm that PirA-PirB proteins of P. akhurstii subsp. akhurstii K-1 are binary insecticidal toxins and ERIC sequences could play an important role in expression of Pir proteins. Reports of biophysical characterization of individually purified PirAt, PirB and expressed PirA-PirB toxin mixture could provide the structural insight into these proteins.
Topics: Animals; Bacterial Proteins; Bacterial Toxins; Escherichia coli; Insect Proteins; Larva; Photorhabdus; Xenorhabdus
PubMed: 36167186
DOI: 10.1016/j.jip.2022.107829