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Bacterial ABC toxin complexes (Tcs) comprise three core proteins: TcA, TcB and TcC. The TcA protein forms a pentameric assembly that attaches to the surface of target cells and penetrates the cell membrane. The TcB and TcC proteins assemble as a heterodimeric TcB–TcC subcomplex that makes a hollow shell. This TcB–TcC subcomplex self-cleaves and encapsulates within the shell a cytotoxic `cargo' encoded by the C-terminal region of the TcC protein. Here, we describe the structure of a previously uncharacterized TcC protein from Yersinia entomophaga, encoded by a gene at a distant genomic location from the genes encoding the rest of the toxin complex, in complex with the TcB protein. When encapsulated within the TcB–TcC shell, the C-terminal toxin adopts an unfolded and disordered state, with limited areas of local order stabilized by the chaperone-like inner surface of the shell. We also determined the structure of the toxin cargo alone and show that when not encapsulated within the shell, it adopts an ADP-ribosyltransferase fold most similar to the catalytic domain of the SpvB toxin from Salmonella typhimurium. Our structural analysis points to a likely mechanism whereby the toxin acts directly on actin, modifying it in a way that prevents normal polymerization.

Pine wood nematode disease is currently the most deadly forest disease in China, and the Monochamus alternatus is its primary vector. Controlling the M. alternatus is crucial for managing pine wood nematode disease. This study, based on the selected HasA (pGHKW4) secretory expression vector, used electroporation to combine the genetically modified high-toxicity toxin Cry3Aa-T with the entomopathogenic bacterium Yersinia entomophaga isolated from the gut of the M. alternatus. The SDS-PAGE and Western blotting techniques were employed to confirm the toxin protein's secretion capability. The engineered bacteria's genetic stability and effectiveness in controlling M. alternatus were assessed for their insecticidal activity. The results of the SDSPAGE and Western blotting analyses indicate that the HasA system effectively expresses toxin protein secretion, demonstrates certain genetic stability, and exhibits high insecticidal activity against M. alternatus. This study constructed a highly toxic entomopathogenic engineered bacterial strain against M. alternatus larvae, which holds significant implications for controlling M. alternatus, laying the foundation for subsequent research and application of this strain. [ABSTRACT FROM AUTHOR]

Bacterial ABC toxin complexes (Tcs) comprise three core proteins: TcA, TcB and TcC. The TcA protein forms a pentameric assembly that attaches to the surface of target cells and penetrates the cell membrane. The TcB and TcC proteins assemble as a heterodimeric TcB-TcC subcomplex that makes a hollow shell. This TcB-TcC subcomplex self-cleaves and encapsulates within the shell a cytotoxic `cargo' encoded by the C-terminal region of the TcC protein. Here, we describe the structure of a previously uncharacterized TcC protein from Yersinia entomophaga, encoded by a gene at a distant genomic location from the genes encoding the rest of the toxin complex, in complex with the TcB protein. When encapsulated within the TcB-TcC shell, the C-terminal toxin adopts an unfolded and disordered state, with limited areas of local order stabilized by the chaperone-like inner surface of the shell. We also determined the structure of the toxin cargo alone and show that when not encapsulated within the shell, it adopts an ADP-ribosyltransferase fold most similar to the catalytic domain of the SpvB toxin from Salmonella typhimurium. Our structural analysis points to a likely mechanism whereby the toxin acts directly on actin, modifying it in a way that prevents normal polymerization., (open access.)

The use of microbial insect pathogens as biocontrol agents is a well-established practice, with several bacteria and fungi available in commercial products. However, lack of efficacy, persistence and restricted host ranges are limiting the widespread uptake of insect pathogens. Combining multiple agents into a single product may overcome some limitations, if they are compatible and could provide synergy. In laboratory bioassays and field applications, we examined the combined effect of two insecticidal bacteria, Yersinia entomophaga MH96 and Brevibacillus laterosporus (1951 and 1821) and the entomopathogenic fungus, Beauveria bassiana J18 against larvae of diamondback moth, Plutella xylostella. In laboratory bioassays, effects of combined bacterial agents were greater than additive at lower doses, but less than additive at higher dose. Results of the field trial showed no significant difference between the combined agents and the single treatments in most cases, although there was reduction in insect damage in combined treatments. [ABSTRACT FROM AUTHOR]

Pine wood nematode disease is currently the most deadly forest disease in China, and the Monochamus alternatus is its primary vector. Controlling the M. alternatus is crucial for managing pine wood nematode disease. This study, based on the selected HasA (pGHKW4) secretory expression vector, used electroporation to combine the genetically modified high-toxicity toxin Cry3Aa-T with the entomopathogenic bacterium Yersinia entomophaga isolated from the gut of the M. alternatus . The SDS-PAGE and Western blotting techniques were employed to confirm the toxin protein's secretion capability. The engineered bacteria's genetic stability and effectiveness in controlling M. alternatus were assessed for their insecticidal activity. The results of the SDS-PAGE and Western blotting analyses indicate that the HasA system effectively expresses toxin protein secretion, demonstrates certain genetic stability, and exhibits high insecticidal activity against M. alternatus. This study constructed a highly toxic entomopathogenic engineered bacterial strain against M. alternatus larvae, which holds significant implications for controlling M. alternatus , laying the foundation for subsequent research and application of this strain., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Han, Huang, Qi, Zhu, Hu, Wen, Long, Xu and Zhang.)

BACKGROUND Yersinia entomophaga is an entomopathogenic bacterium that is active against scarab beetles, among other insects. In New Zealand, the African black beetle, Heteronychus arator (Coleoptera: Scarabaeidae), is a major pest of pastures and arable crops but very few control options exist and no insecticides are registered for use in established pastures. RESULTS: In laboratory bioassays, H. arator adults were susceptible to a bait containing Y. entomophaga at low doses. This bait was more effective against H. arator adults during spring than autumn in small‐scale field plots (320 mm diameter). A large‐scale field trial (40 × 40 m plots) reduced adult numbers substantially: approximately twice as many beetles were captured in pitfall traps from untreated plots compared with plots treated with the Y. entomophaga bait at 70 kg ha−1. This single bait application in spring also reduced subsequent larval populations in summer. CONCLUSIONS: Heteronychus arator is a difficult pest to manage using chemical insecticides. This biopesticide with Y. entomophaga as the active ingredient offers a new solution for New Zealand pastures, with potential for application to other crops affected by H. arator and for control of other pests. © 2020 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

BACKGROUND: Porina is the common name for moths and larvae of the genus Wiseana (Lepidoptera: Hepialidae), some of which are significant pasture pests in New Zealand. Because of environmental concerns and the non‐target effects of insecticide control measures, biological alternatives for the control of insect pests such as porina are required. RESULTS: Using a food preference assay and time‐lapse photography, a range of low‐cost food ingredients were assessed for their palatability to porina larvae. Lead candidates were combined into extruded bait variants, allowing assessment of their palatability to porina larvae. A composite bait consisting of palatable ingredients was developed, into which the porina‐active entomopathogen Yersinia entomophaga was incorporated. A 7 day minimum median lethal dose of approximately 6.0 × ±1 × 106Y. entomophaga cells per 0.02 g of bait was defined. Field trials showed that the mean change in larval density over time differed between treatments, with Y. entomophaga bait applied at 87 kg ha−1 resulting in a mean 65% reduction in larval density relative to the control plots, and diflubenzuron treatment resulting in a mean 77% reduction relative to the control plots. The mean dry matter yields over the course of the trial were highest for diflubenzuron (5029 kg ha−1), followed by the Y. entomophaga (4783 kg ha−1) and control (4673 kg ha−1) treatments. CONCLUSIONS: The bacterium Y. entomophaga applied as a composite bait offers an environmentally sustainable approach for porina pest control. © 2019 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • Y. entomophaga is active against Bt Cry1A-, Cry1C-resistant, Plutella xylostella larvae. • Y. entomophaga is active against indoxacarb/pyrethroid, spinosad-resistant P. xylostella. • LD50 of Y. entomophaga in P. xylostella -challenged larvae (5 days) is 2.7 × 103. Abstract The application of the biocontrol bacterium Yersinia entomophaga as a foliar spray was assessed for its efficacy against larvae of the diamondback moth, Plutella xylostella. The bacterium was applied as either a broth suspension, or as a biopolymer-based gel foliar spray and compared with commercial insecticides Dipel (Bacillus thuringiensis) and Spinosad. The performance of Y. entomophaga was comparable with that of Dipel. The gel-based formulation extended leaf persistence over that of the basic broth culture spray, while also providing higher initial foliar deposition rates. The bacterium was found to multiply within the P. xylostella larvae to 5.8 × 105 cells per larva, while the median lethal dose (LD 50) was determined to be 2.69 × 103 cells per larva. Importantly, B. thuringiensis Cry1A-resistant, Cry1C-resistant, indoxacarb/pyrethroid-resistant, and Spinosad-resistant P. xylostella larvae were susceptible to Y. entomophaga. [ABSTRACT FROM AUTHOR]

An O-polysaccharide was isolated from the lipopolysaccharide of an entomopathogenic bacterium Yersinia entomophaga MH96 T by mild acid hydrolysis and studied by 2D NMR spectroscopy. The following structure of the branched tetrasaccharide repeating unit of the polysaccharide was established: Image 2 where Tyv indicates 3,6-dideoxy- d - arabino -hexose (tyvelose). The structure established is consistent with the gene content of the O-antigen gene cluster. The O-polysaccharide structure and gene cluster of Y. entomophaga are related to those of some Y. pseudotuberculosis serotypes. [ABSTRACT FROM AUTHOR]

Here we report the draft genome of Yersinia entomophaga type strain MH96T. The genome shows 93.8% nucleotide sequence identity to that of Yersinia nurmii type strain APN3a-cT, and comprises a single chromosome of approximately 4,275,531 bp. In silico analysis identified that, in addition to the previously documented Y. entomophaga Yen-TC gene cluster, the genome encodes a diverse array of toxins, including two type III secretion systems, and five rhs-associated gene clusters. As well as these multicomponent systems, several orthologs of known insect toxins, such as VIP2 toxin and the binary toxin PirAB, and distant orthologs of some mammalian toxins, including repeats-in-toxin, a cytolethal distending toxin, hemolysin-like genes and an adenylate cyclase were identified. The genome also contains a large number of hypothetical proteins and orthologs of known effector proteins, such as LopT, as well as genes encoding a wide range of proteolytic determinants, including metalloproteases and pathogen fitness determinants, such as genes involved in iron metabolism. The bioinformatic data derived from the current in silico analysis, along with previous information on the pathobiology of Y. entomophaga against its insect hosts, suggests that a number of these virulence systems are required for survival in the hemocoel and incapacitation of the insect host. [ABSTRACT FROM AUTHOR]

Here we report the draft genome of Yersinia entomophaga type strain MH96T. The genome shows 93.8% nucleotide sequence identity to that of Yersinia nurmii type strain APN3a-cT, and comprises a single chromosome of approximately 4,275,531 bp. In silico analysis identified that, in addition to the previously documented Y. entomophaga Yen-TC gene cluster, the genome encodes a diverse array of toxins, including two type III secretion systems, and five rhs-associated gene clusters. As well as these multicomponent systems, several orthologs of known insect toxins, such as VIP2 toxin and the binary toxin PirAB, and distant orthologs of some mammalian toxins, including repeats-in-toxin, a cytolethal distending toxin, hemolysin-like genes and an adenylate cyclase were identified. The genome also contains a large number of hypothetical proteins and orthologs of known effector proteins, such as LopT, as well as genes encoding a wide range of proteolytic determinants, including metalloproteases and pathogen fitness determinants, such as genes involved in iron metabolism. The bioinformatic data derived from the current in silico analysis, along with previous information on the pathobiology of Y. entomophaga against its insect hosts, suggests that a number of these virulence systems are required for survival in the hemocoel and incapacitation of the insect host.

Highlights: [•] The pathology of Yersinia entomophaga against Costelytra zealandica was examined. [•] Post ingestion, the larvae exhibited reduced feeding and expulsion of gut contents. [•] Within 48h the bacteria invaded the haemocoel resulting in death by septicaemia. [•] Mortality is directly proportional to dose and time after infection. [•] The LD50 was determined as 2.9×104 bacteria per C. zealandica larva. [ABSTRACT FROM AUTHOR]

The Yersinia entomophaga toxin complex (Yen-TC) is the bacterium's main virulence determinant. Because of its high insect activity, methods were developed to allow the routine isolation and purification of Yen-TC from an overnight bacterial culture using size exclusion chromatography. Here we outline an overnight purification procedure using a 100-ml culture volume, where approximately 2 mg of Yen-TC, with an approximate purity of 95-98 %, can be routinely obtained.