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Pest Management Science Apr 2024In the perpetual struggle to manage mosquito populations, there has been increasing demand for the development of biopesticides to supplant/complement current products....
BACKGROUND
In the perpetual struggle to manage mosquito populations, there has been increasing demand for the development of biopesticides to supplant/complement current products. The insecticidal potential of Xenorhabdus and Photorhabdus has long been recognized and is of interest for the control of important mosquitoes like Aedes albopictus which vectors over 20 different arboviruses of global public health concern.
RESULTS
The larvicidal effects of cell-free supernatants, cell growth cultures and cell mass of an extensive list of Xenorhabdus and Photorhabdus spp. was investigated. They were quite effective against Ae. albopictus causing larval mortality ranging between 52-100%. Three Photorhabdus spp. and 13 Xenorhabdus spp. release larvicidal compounds in cell-free supernatants. Cell growth culture of all tested species exhibited larvicidal activity, except for Xenorhabdus sp. TS4. Twenty-one Xenorhabdus and Photorhabdus bacterial cells (pellet) exhibited oral toxicity (59-91%) against exposed larvae. The effect of bacterial supernatants on the mosquito eggs were also assessed. Bacterial supernatants inhibited the hatching of mosquito eggs; when unhatched eggs were transferred to clean water, they all hatched. Using the easyPACId approach, the larvicidal compounds in bacterial supernatant were identified as fabclavine from X. szentirmaii and xencoumacin from X. nematophila (causing 98 and 70% mortality, respectively, after 48 h). Xenorhabdus cabanillasii and X. hominickii fabclavines were as effective as commercial Bacillus thuringiensis subsp. israelensis and spinosad products within 5 days post-application (dpa).
CONCLUSION
Fabclavine and xenocoumacin can be developed into novel biolarvicides, can be used as a model to synthesize other compounds or/and can be combined with other commercial biolarvicides. © 2024 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.
PubMed: 38619291
DOI: 10.1002/ps.8127 -
Angewandte Chemie (International Ed. in... Oct 2020Leupeptin is a bacterial small molecule that is used worldwide as a protease inhibitor. However, its biosynthesis and genetic distribution remain unknown. We identified...
Leupeptin is a bacterial small molecule that is used worldwide as a protease inhibitor. However, its biosynthesis and genetic distribution remain unknown. We identified a family of leupeptins in gammaproteobacterial pathogens, including Photorhabdus, Xenorhabdus, and Klebsiella species, amongst others. Through genetic, metabolomic, and heterologous expression analyses, we established their construction by discretely expressed ligases and accessory enzymes. In Photorhabdus species, a hypothetical protein required for colonizing nematode hosts was established as a new class of proteases. This enzyme cleaved the tripeptide aldehyde protease inhibitors, leading to the formation of "pro-pyrazinones" featuring a hetero-tricyclic architecture. In Klebsiella oxytoca, the pathway was enriched in clinical isolates associated with respiratory tract infections. Thus, the bacterial production and proteolytic degradation of leupeptins can be associated with animal colonization phenotypes.
Topics: Animals; Gammaproteobacteria; Leupeptins; Protease Inhibitors
PubMed: 32609431
DOI: 10.1002/anie.202005506 -
Bioorganic & Medicinal Chemistry Letters Oct 2022Hydroxyequols are promising analogues of the biologically active flavonoid, equol. We recently found that the flavin-dependent monooxygenase HpaB of Rhodococcus opacus...
Hydroxyequols are promising analogues of the biologically active flavonoid, equol. We recently found that the flavin-dependent monooxygenase HpaB of Rhodococcus opacus regioselectively synthesizes 3'-hydroxyequol from equol, whereas HpaB of Photorhabdus luminescens synthesizes 6-hydroxyequol. In this study, we investigated the cascade synthesis of a dihydroxyequol compound from equol using these two enzymes. When Escherichia coli cells expressing HpaB and cells expressing HpaB were simultaneously incubated with equol, the cells efficiently synthesized 6,3'-dihydroxyequol (8.7 mM, 2.4 g/L) via 3'- and 6-hydroxyequols in one pot. The antioxidant activity of the equol derivatives increased with an increase in the number of hydroxyl groups on the equol scaffold. 6,3'-Dihydroxyequol exhibited potent antioxidant activity. In addition, 6-hydroxyequol significantly inhibited the growth of E. coli. Cell survival studies suggested that 6-hydroxyequol is a bactericidal rather than bacteriostatic compound. To our knowledge, this is the first report describing the antibacterial activity of hydroxyequols.
Topics: Anti-Bacterial Agents; Antioxidants; Biocatalysis; Equol; Escherichia coli; Isoflavones
PubMed: 35902062
DOI: 10.1016/j.bmcl.2022.128908 -
BMC Microbiology Feb 2021The insect pathogenic bacterium Photorhabdus luminescens exists in two phenotypically different forms, designated as primary (1°) and secondary (2°) cells. Upon yet...
BACKGROUND
The insect pathogenic bacterium Photorhabdus luminescens exists in two phenotypically different forms, designated as primary (1°) and secondary (2°) cells. Upon yet unknown environmental stimuli up to 50% of the 1° cells convert to 2° cells. Among others, one important difference between the phenotypic forms is that 2° cells are unable to live in symbiosis with their partner nematodes, and therefore are not able to re-associate with them. As 100% switching of 1° to 2° cells of the population would lead to a break-down of the bacteria's life cycle the switching process must be tightly controlled. However, the regulation mechanism of phenotypic switching is still puzzling.
RESULTS
Here we describe two novel XRE family transcriptional regulators, XreR1 and XreR2, that play a major role in the phenotypic switching process of P. luminescens. Deletion of xreR1 in 1° or xreR2 in 2° cells as well as insertion of extra copies of xreR1 into 2° or xreR2 into 1° cells, respectively, induced the opposite phenotype in either 1° or 2° cells. Furthermore, both regulators specifically bind to different promoter regions putatively fulfilling a positive autoregulation. We found initial evidence that XreR1 and XreR2 constitute an epigenetic switch, whereby XreR1 represses xreR2 expression and XreR2 self-reinforces its own gene by binding to XreR1.
CONCLUSION
Regulation of gene expression by the two novel XRE-type regulators XreR1 and XreR2 as well as their interplay represents a major regulatory process in phenotypic switching of P. luminescens. A fine-tuning balance between both regulators might therefore define the fate of single cells to convert from the 1° to the 2° phenotype.
Topics: Animals; Bacterial Proteins; Gene Expression Regulation; Insecta; Nematoda; Phenotype; Photorhabdus; Symbiosis; Transcription Factors
PubMed: 33627070
DOI: 10.1186/s12866-021-02116-2 -
Biology Feb 2022(Lepidoptera: Lycaenidae) and (Lepidoptera: Pyralidae) are the key pests of pomegranates in Saudi Arabia that are managed mainly using broad-spectrum pesticides....
Isolation, Identification, and Biocontrol Potential of Entomopathogenic Nematodes and Associated Bacteria against (Lepidoptera: Lycaenidae) and (Lepidoptera: Pyralidae).
(Lepidoptera: Lycaenidae) and (Lepidoptera: Pyralidae) are the key pests of pomegranates in Saudi Arabia that are managed mainly using broad-spectrum pesticides. Interactions between the entomopathogenic nematodes (EPNs) Steinernematids, and Heterorhabditids, and their entomopathogenic bacterial symbionts (EPBs) have long been considered monoxenic 2-partner associations responsible for killing insects and, therefore, are widely used in insect pest biocontrol. However, there are limited reports identifying such organisms in Taif, Saudi Arabia. The current study aimed to identify the EPNs and their associated bacteria isolated from Taif, Saudi Arabia, and evaluate their biocontrol potential on third instar larvae of and under laboratory conditions. A total of 35 EPN isolates belonging to (20) and (15) were recovered from 320 soil samples. Twenty-six isolates of symbiotic or associated bacteria were isolated from EPNs and molecularly identified as (6 isolates), (4 isolates), (7), or (9). A pathogenicity assay revealed that spp. were more virulent than spp. against the two pomegranate insects, with LC values of 18.5 and 13.6 infective juveniles (IJs)/larva of for spp. and 52 and 32.4 IJs/larva of for spp. at 48 and 72 h post-treatment, respectively. Moreover, LC values of 9 and 6.6 IJs/larva ( spp.) and 34.4 and 26.6 IJs/larva ( spp.) were recorded for larvae at 48 and 72 h post-treatment. In addition, the EPB CQ1, isolated from spp., surpassed SJ10, associated with spp., in their ability to kill or larvae within 6 h post-application, resulting in 100% mortality in both insects after 24 and 48 h of exposure. We conclude that either application of EPNs' IJs or their associated EPBs could serve as potential biocontrol agents for and .
PubMed: 35205161
DOI: 10.3390/biology11020295 -
Journal of the American Chemical Society Apr 2021Carminic acid is an aromatic polyketide found in scale insects (i.e., ) and is a widely used natural red colorant. It has long been produced by the cumbersome farming of...
Carminic acid is an aromatic polyketide found in scale insects (i.e., ) and is a widely used natural red colorant. It has long been produced by the cumbersome farming of insects followed by multistep purification processes. Thus, there has been much interest in producing carminic acid by the fermentation of engineered bacteria. Here we report the complete biosynthesis of carminic acid from glucose in engineered . We first optimized the type II polyketide synthase machinery from , enabling a high-level production of flavokermesic acid upon coexpression of the cyclases ZhuI and ZhuJ from sp. R1128. To discover the enzymes responsible for the remaining two reactions (hydroxylation and -glucosylation), biochemical reaction analyses were performed by testing enzyme candidates reported to perform similar reactions. The two identified enzymes, aklavinone 12-hydroxylase (DnrF) from and -glucosyltransferase (GtCGT) from , could successfully perform hydroxylation and -glucosylation of flavokermesic acid, respectively. Then, homology modeling and docking simulations were performed to enhance the activities of these two enzymes, leading to the generation of beneficial mutants with 2-5-fold enhanced conversion efficiencies. In addition, the GtCGT mutant was found to be a generally applicable -glucosyltransferase in , as was showcased by the successful production of aloesin found in . Simple metabolic engineering followed by fed-batch fermentation resulted in 0.63 ± 0.02 mg/L of carminic acid production from glucose. The strategies described here will be useful for the design and construction of biosynthetic pathways involving unknown enzymes and consequently the production of diverse industrially important natural products.
Topics: Carmine; Escherichia coli; Fermentation; Glucosyltransferases; Metabolic Engineering; Polyketides
PubMed: 33797895
DOI: 10.1021/jacs.0c12406 -
PLoS Computational Biology Feb 2021The vast expansion of protein sequence databases provides an opportunity for new protein design approaches which seek to learn the sequence-function relationship...
The vast expansion of protein sequence databases provides an opportunity for new protein design approaches which seek to learn the sequence-function relationship directly from natural sequence variation. Deep generative models trained on protein sequence data have been shown to learn biologically meaningful representations helpful for a variety of downstream tasks, but their potential for direct use in the design of novel proteins remains largely unexplored. Here we show that variational autoencoders trained on a dataset of almost 70000 luciferase-like oxidoreductases can be used to generate novel, functional variants of the luxA bacterial luciferase. We propose separate VAE models to work with aligned sequence input (MSA VAE) and raw sequence input (AR-VAE), and offer evidence that while both are able to reproduce patterns of amino acid usage characteristic of the family, the MSA VAE is better able to capture long-distance dependencies reflecting the influence of 3D structure. To confirm the practical utility of the models, we used them to generate variants of luxA whose luminescence activity was validated experimentally. We further showed that conditional variants of both models could be used to increase the solubility of luxA without disrupting function. Altogether 6/12 of the variants generated using the unconditional AR-VAE and 9/11 generated using the unconditional MSA VAE retained measurable luminescence, together with all 23 of the less distant variants generated by conditional versions of the models; the most distant functional variant contained 35 differences relative to the nearest training set sequence. These results demonstrate the feasibility of using deep generative models to explore the space of possible protein sequences and generate useful variants, providing a method complementary to rational design and directed evolution approaches.
Topics: Algorithms; Computational Biology; Computer Simulation; Escherichia coli; Machine Learning; Neural Networks, Computer; Oxidoreductases; Photorhabdus; Proteins; Recombinant Proteins; Reproducibility of Results; Solubility
PubMed: 33635868
DOI: 10.1371/journal.pcbi.1008736 -
Applied Microbiology and Biotechnology Jul 2021Xenorhabdus and Photorhabdus spp. are enteric bacterial symbionts of Steinernema and Heterorhabditis nematodes, respectively. These bacteria produce an extensive set of...
Antifungal activity of different Xenorhabdus and Photorhabdus species against various fungal phytopathogens and identification of the antifungal compounds from X. szentirmaii.
Xenorhabdus and Photorhabdus spp. are enteric bacterial symbionts of Steinernema and Heterorhabditis nematodes, respectively. These bacteria produce an extensive set of natural products (NPs) with antibacterial, antifungal, antiprotozoal, insecticidal, or other bioactivities when vectored into insect hemocoel by nematodes. We assessed the in vitro activity of different Xenorhabdus and Photorhabdus cell-free supernatants against important fungal phytopathogens, viz., Cryphonectria parasitica, Fusarium oxysporum, Rhizoctonia solani, and Sclerotinia sclerotiorum and identified the bioactive antifungal compound/s present in the most effective bacterial supernatant using the easyPACId (easy promoter-activated compound identification) approach against chestnut blight C. parasitica. Our data showed that supernatants from Xenorhabdus species were comparatively more effective than extracts from Photorhabdus in suppressing the fungal pathogens; among the bacteria assessed, Xenorhabdus szentirmaii was the most effective species against all tested phytopathogens especially against C. parasitica. Subsequent analysis revealed fabclavines as antifungal bioactive compounds in X. szentirmaii, generated by a polyketide synthase (PKS) and non-ribosomal peptide synthetase (NRPS) hybrid system. Fabclavines are broad-spectrum, heat-stable NPs that have great potential as biological control compounds against fungal plant pathogens. More studies are needed to assess the potential phytotoxicity of these compounds and their effects on non-target organisms before commercialization. KEY POINTS: • Chemical fungicides have toxic effects on humans and other non-target organisms. • Alternatives with novel modes of action to supplant current fungicide are needed. • A novel bioactive antifungal compound from Xenorhabdus szentirmaii was identified.
Topics: Animals; Antifungal Agents; Ascomycota; Fusarium; Humans; Photorhabdus; Plant Diseases; Rhizoctonia; Symbiosis; Xenorhabdus
PubMed: 34250572
DOI: 10.1007/s00253-021-11435-3 -
Advanced Science (Weinheim,... Jun 2024Pigments such as anthraquinones (AQs) and melanins are antioxidants, protectants, or virulence factors. AQs from the entomopathogenic bacterium Photorhabdus laumondii...
Pigments such as anthraquinones (AQs) and melanins are antioxidants, protectants, or virulence factors. AQs from the entomopathogenic bacterium Photorhabdus laumondii are produced by a modular type II polyketide synthase system. A key enzyme involved in AQ biosynthesis is PlAntI, which catalyzes the hydrolysis of the bicyclic-intermediate-loaded acyl carrier protein, polyketide trimming, and assembly of the aromatic AQ scaffold. Here, multiple crystal structures of PlAntI in various conformations and with bound substrate surrogates or inhibitors are reported. Structure-based mutagenesis and activity assays provide experimental insights into the three sequential reaction steps to yield the natural product AQ-256. For comparison, a series of ligand-complex structures of two functionally related hydrolases involved in the biosynthesis of 1,8-dihydroxynaphthalene-melanin in pathogenic fungi is determined. These data provide fundamental insights into the mechanism of polyketide trimming that shapes pigments in pro- and eukaryotes.
Topics: Anthraquinones; Polyketides; Melanins; Polyketide Synthases; Photorhabdus; Naphthols; Pigments, Biological
PubMed: 38491909
DOI: 10.1002/advs.202400184 -
Journal of Fungi (Basel, Switzerland) Apr 2022is a phytopathogenic fungus that causes devastating losses in strawberries without effective countermeasures. Members of the genus exhibit antimicrobial capability and...
is a phytopathogenic fungus that causes devastating losses in strawberries without effective countermeasures. Members of the genus exhibit antimicrobial capability and have been found to have the potential for use as biocontrol agents against . species exhibit two phase variations with a differentiated composition of secondary metabolites designated to each phase. In this study, sp. nov. 0813-124 exhibited phase I (PL1) and phase II (PL2); however, only PL1 displayed distinct inhibition of in the confrontation assay. We identified the bioactive ingredients of sp. nov. 0813-124 to be glidobactin A and cepafungin I, with MIC values lower than 1.5 and 2.0 µg/mL, respectively. Furthermore, we revealed the biosynthetic gene cluster (BGC) of corresponding bioactive molecules through genomics analysis and determined its expression level in PL1 and PL2. The expression of glidobactin BGC in PL1 increased rapidly within 24 h, while PL2 was eventually stimulated after 60 h. In summary, we demonstrated that sp. nov. 0813-124 could potentially be used as a biocontrol agent or part of a natural product repertoire for combating .
PubMed: 35448634
DOI: 10.3390/jof8040403