A toxin complex protein from Photorhabdus akhurstii conferred oral insecticidal activity against Galleria mellonella by targeting the midgut epithelium

ABSTRACT Photorhabdus luminescens is a gram-negative enteric bacterium that is found in association with entomopathogenic nematodes of the family Heterorhabditidae. The nematodes infect a variety of soil-dwelling insects. Upon entering an insect host, the nematode releases P. luminescens cells from its intestinal tract, and the bacteria quickly establish a lethal septicemia. When grown in peptone broth, in the absence of the nematodes, the bacteria produce a protein toxin complex that is lethal when fed to, or injected into the hemolymph of, Manduca sexta larvae and several other insect species. The toxin purified as a protein complex which has an estimated molecular weight of 1,000,000 and contains no protease, phospholipase, or hemolytic activity and only a trace of lipase activity. The purified toxin possesses insecticidal activity whether injected or given orally. Analyses of the denatured complex by sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed it to be composed of several protein subunits ranging in size from 30 to 200 kDa. The complex was further separated by native gel electrophoresis into three components, two of which retained insecticidal activity. The purified native toxin complex was found to be active in nanogram concentrations against insects representing four orders of the classInsecta.

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3D structure of the Yersinia entomophaga toxin complex and implications for insecticidal activity

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1111155108 ◽

2011 ◽

Vol 108 (51) ◽

pp. 20544-20549 ◽

Cited By ~ 56

Author(s):

M. J. Landsberg ◽

S. A. Jones ◽

R. Rothnagel ◽

J. N. Busby ◽

S. D. G. Marshall ◽

...

Keyword(s):

Insecticidal Activity ◽

3D Structure ◽

Toxin Complex

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Isolation of Bacillus thuringiensis and its Insecticidal Effect against Galleria mellonella

Himalayan Journal of Science and Technology ◽

10.3126/hijost.v4i0.33918 ◽

2020 ◽

pp. 96-102

Author(s):

Jyoti Limbu ◽

Bijay Kumar Shrestha ◽

Jenish Shakya ◽

Sabin Bahadur Khatri ◽

Hemanta Khanal

Keyword(s):

Bacillus Thuringiensis ◽

Insecticidal Activity ◽

Galleria Mellonella ◽

Geographical Location ◽

Insecticidal Effect ◽

Greater Wax Moth ◽

Effective Level ◽

And Control ◽

Geographical Locations ◽

Wax Moth

Bacillus thuringiensis (Bt) synthesize a large diversity of crystal proteins (Cry and Cyt) during sporulation which exhibit insecticidal activity against insects and protozoa. The main aim of this study was to isolate Bacillus thuringiensis and study its insecticidal effect against Galleria mellonella. Soil samples from four different geographical locations of Koshi Zone viz. Itahari, Tarhara, Dharan and Vedetar of Eastern Nepal were collected. The isolation of Bt was done by acetate selection method. The insect bioassay of Bt isolates were performed against greater wax moth (G. mellonella) by feeding the third instar larvae by extracted crystal spores with three different concentrations. The overall distribution of Bt from the study sample was found to be 30% (30/100). Bt was isolated from all four geographical location with higher incidence; 9 (36%) in Tarhara region followed by Dharan (32%), Itahari (28%) and Vedetar (24%). However, the incidence of Bt with potent insecticidal activity against G. mellonella was reported to be 4% (4/100). The insecticidal activity of isolated Bt between test and control groups was found to be statistically significant (p<0.05). LC50 value of Bt from Tarhara (Tar1) was 388.29μg/mL, Dharan; Drn8 and Drn1 was 416.20μg/mL and 463.15μg/mL respectively and from Vedetar (Vd5) was 476.63μg/mL. In overall study the Bt isolated from Tarhara (Tar1) region exhibited greater incidence, Bt index, efficacy and effective level of LC50 against greater wax moth. Native Bt strains isolated from soil of Eastern Nepal possess effective insecticidal activity and hence can used as biocontrol agent in controlling honeycomb pest like G. mellonella.

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Comparative Genomics and Pathogenicity Analysis of Two Bacterial Symbionts of Entomopathogenic Nematodes: The Role of The GroEL Protein in Virulence

10.21203/rs.3.rs-1032559/v1 ◽

2021 ◽

Author(s):

Abraham Rivera-Ramírez ◽

Rosalba Salgado-Morales ◽

Alfredo Jiménez-Pérez ◽

Rebeca Pérez-Martínez ◽

Blanca Inés García-Gómez ◽

...

Keyword(s):

Insecticidal Activity ◽

Entomopathogenic Nematodes ◽

Core Genome ◽

Galleria Mellonella ◽

Gene Clusters ◽

Gene Repertoire ◽

Amino Acid Residues ◽

Whole Cells ◽

Close Phylogenetic Relationship ◽

Biological Tests

Abstract Bacteria of the genera Xenorhabdus and Photorhabdus are symbionts of entomopathogenic nematodes. Despite their close phylogenetic relationship, they show differences in their pathogenicity and virulence mechanisms in target insects. These differences can be explored by the analysis of the pangenome, as it provides a framework for characterizing and defining the gene repertoire. Here, we report the genome of strain SC 0516. In addition, we performed the first pangenome analysis of 91 strains of Xenorhabdus and Photorhabdus, obtaining a total of 23,603 gene clusters and a core genome of 348 genes. Phylogenetic analysis performed with the core genome showed that our strain belonged to the X. nematophila group. Biological tests showed that whole cells of X. nematophila SC 0516 were more virulent than those of P. luminescens HIM3 when both were injected into Galleria mellonella larvae. In addition, we cloned and expressed the GroEL proteins of both bacteria, as this protein has been previously indicated to show insecticidal activity in the genus Xenorhabdus. Cpn60-Xn was found to be the most toxic at all concentrations tested, with an LC50 value of 102.34 ng/larva. Sequence analysis suggested that the Cpn60-Xn toxin was homologous to Cpn60-Pl; however, Cpn60-Xn contained thirty-five differentially substituted amino acid residues that could be responsible for its insecticidal activity.

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Activity of a Holin-Endolysin System in the Insecticidal Pathogenicity Island ofYersinia enterocolitica

Journal of Bacteriology ◽

10.1128/jb.00180-18 ◽

2018 ◽

Vol 200 (16) ◽

Cited By ~ 2

Author(s):

Katharina Springer ◽

Sandra Reuter ◽

Mandy Knüpfer ◽

Lukas Schmauder ◽

Philipp-Albert Sänger ◽

...

Keyword(s):

Insecticidal Activity ◽

High Specificity ◽

Pathogenicity Island ◽

Open Reading Frames ◽

Content Type ◽

Toxin Complex ◽

Genes Encoding ◽

Conserved Genes ◽

Nematocidal Activity ◽

Lysis Cassette

ABSTRACTYersinia enterocoliticais a pathogen that causes gastroenteritis in humans. Because of its low-temperature-dependent insecticidal activity, it can oscillate between invertebrates and mammals as host organisms. The insecticidal activity of strain W22703 is associated with a pathogenicity island of 19 kb (Tc-PAIYe), which carries regulators and genes encoding the toxin complex (Tc). The island also harbors four phage-related and highly conserved genes of unknown functions, which are polycistronically transcribed. Two open reading frames showed significant homologies to holins and endolysins and exhibited lytic activity inEscherichia colicells upon overexpression. When a set ofYersiniastrains was tested in an equivalent manner, highly diverse susceptibilities to lysis were observed, and some strains were resistant to lysis. If cell lysis occurred (as demonstrated by membrane staining), it was more pronounced when two accessory elements of the cassette coding for an i-spanin and an o-spanin were included in the overexpression construct. The pore-forming function of the putative holin, HolY, was demonstrated by complementation of the lysis defect of a phage λ S holin mutant. In experiments performed with membrane preparations, ElyY exhibited high specificity for W22703 peptidoglycan, with a cleavage activity resembling that of lysozyme. Although the functionality of the lysis cassette from Tc-PAIYewas demonstrated in this study, its biological role remains to be elucidated.IMPORTANCEThe knowledge of how pathogens survive in the environment is pivotal for our understanding of bacterial virulence. The insecticidal and nematocidal activity ofYersiniaspp., by which the bacteria gain access to nutrients and thus improve their environmental fitness, is conferred by the toxin complex (Tc) encoded on a highly conserved pathogenicity island termed Tc-PAIYe. While the regulators and the toxin subunits of the island had been characterized in some detail, the role of phage-related genes within the island remained to be elucidated. Here, we demonstrate that this cassette encodes a holin, an endolysin, and two spanins that, at least upon overexpression, lyseYersiniastrains.

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Gut Bacteria Are Not Required for the Insecticidal Activity of Bacillus thuringiensis toward the Tobacco Hornworm, Manduca sexta

Applied and Environmental Microbiology ◽

10.1128/aem.00966-09 ◽

2009 ◽

Vol 75 (15) ◽

pp. 5094-5099 ◽

Cited By ~ 43

Author(s):

Paul R. Johnston ◽

Neil Crickmore

Keyword(s):

Bacillus Thuringiensis ◽

Manduca Sexta ◽

Insecticidal Activity ◽

Lymantria Dispar ◽

Pieris Rapae ◽

Galleria Mellonella ◽

Sublethal Concentration ◽

Gut Bacteria ◽

First Instar ◽

Vanessa Cardui

ABSTRACT It was recently proposed that gut bacteria are required for the insecticidal activity of the Bacillus thuringiensis-based insecticide, DiPel, toward the lepidopterans Manduca sexta, Pieris rapae, Vanessa cardui, and Lymantria dispar. Using a similar methodology, it was found that gut bacteria were not required for the toxicity of DiPel or Cry1Ac or for the synergism of an otherwise sublethal concentration of Cry1Ac toward M. sexta. The toxicities of DiPel and of B. thuringiensis HD73 Cry− spore/Cry1Ac synergism were attenuated by continuously exposing larvae to antibiotics before bioassays. Attenuation could be eliminated by exposing larvae to antibiotics only during the first instar without altering larval sterility. Prior antibiotic exposure did not attenuate Cry1Ac toxicity. The presence of enterococci in larval guts slowed mortality resulting from DiPel exposure and halved Cry1Ac toxicity but had little effect on B. thuringiensis HD73 Cry− spore/Cry1Ac synergism. B. thuringiensis Cry− cells killed larvae after intrahemocoelic inoculation of M. sexta, Galleria mellonella, and Spodoptera litura and grew rapidly in plasma from M. sexta, S. litura, and Tenebrio molitor. These findings suggest that gut bacteria are not required for B. thuringiensis insecticidal activity toward M. sexta but that B. thuringiensis lethality is reduced in larvae that are continuously exposed to antibiotics before bioassay.

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The Larvicidal and Adulticidal Effects of Selected Plant Essential Oil Constituents on Greater Wax Moths

Journal of Economic Entomology ◽

10.1093/jee/toaa249 ◽

2020 ◽

Author(s):

Mubasshir Sohail ◽

Muhammad Anjum Aqueel ◽

Pingli Dai ◽

James D Ellis

Keyword(s):

Apis Mellifera ◽

Essential Oil ◽

Honey Bee ◽

Insecticidal Activity ◽

Galleria Mellonella ◽

Insect Pests ◽

Important Pest

Abstract Some plant essential oil constituents, such as monoterpenoids and phenylpropanoids, are promising insecticides in some situations and for certain insect pests. They vary in their toxicity, depending on the target insect. Moths (Lepidoptera) appear susceptible to these compounds, making them of promise for use against greater wax moths (Galleria mellonella Fabricius, (Lepidoptera: Pyrallidae), GWM), an important pest of western honey bee (Apis mellifera Linnaeus. (Hymenoptera: Apidae)) colonies. We determined the LC50 for GWM larvae or LD50 for GWM adults of select compounds (thymol, carvacrol, (S)-(+)-carvone, estragole, citral, linalool, (S)-(−)-limonene, and γ-terpinene). Concentrations between 8 and 2,266 µg/cm3 were mixed into the diets of GWM larvae and doses ranging between 0.08 and 70.3 mg per adult were applied topically to the abdomens of GWM adults. Lethal concentrations and doses were calculated after of 72 h of exposure. All eight compounds showed insecticidal activity against all tested stages of GWMs. Thymol (LC50 µg/cm3 (95% CI) = 21 [9–56], carvacrol = 46 [26–79], citral = 63 [30–134], and carvone = 76 [33–201]) had the highest toxicity toward GWM larvae. The hydrocarbons limonene (296 [231–377]), estragole (466 [354–611]), and γ-terpinene (729 [630–857]) had the lowest toxicity toward GWM larvae. Thymol had the highest toxicity towards GWM adults (LD50 (CI) = 0.5 [0.4–0.8] mg/adult). However, carvacrol (11.6 [10.1–13.6]), linalool (12.9 [9.3–17.8]), and limonene (15.8 [13.1–19.2]) were less toxic to GWM larvae. Our data show that select plant essential oil constituents are promising controls of GWM.

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Thermal stable and proteinase-K resistant insecticidal toxins produced by Photorhabdus luminescens

Abasyn Journal Life Sciences ◽

10.34091/ajls.4.1.19 ◽

2021 ◽

pp. 161-167

Author(s):

Ihsan Ullah ◽

M.S. Al- ghamdi

Keyword(s):

Insecticidal Activity ◽

Galleria Mellonella ◽

Proteinase K ◽

Photorhabdus Luminescens ◽

Heat Stable ◽

Insecticidal Toxins ◽

High Thermal Resistance ◽

Culture Extract ◽

Mutualistic Association ◽

High Degree

Photorhabdus is lives in a mutualistic association with nematodes from the family Heterorhabditis. Bacteria of the Photorhabdus can survive independently and cause toxicity in a larger variety of insects. In the present study, insecticidal activity of non-portentous heat-stable metabolites of Photorhabdus luminescens was evaluated against Galleria mellonella. For this purpose, the culture extract of P. luminescens was injected into the G. mellonella larvae, which killed almost 90% of larvae within 48 h. The extract showed 100% insecticidal activity after heat treatment of 70 C for 30 min and even 60% and 40% activity lasted at 80 C and 90 C respectively. The extract also showed a high degree of thermal stability and was 100% actives after 60 min at 70 C. In addition, insecticidal activity was preserved up to 100% after all proteinase-K treatments (0 ?g/mL to 50 ?g/mL). The results revealed that the extracts were non-portentous and showed high thermal resistance and stability. Keywords: Photorhabdus, insecticidal activity, toxins, heat stable non-proteinaceous

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DEVELOPMENT AND PATHOGENICITY OF INFECTIVE JUVENILES ORIGINATING VIA ENDOTOKIA MATRICIDA IN AXENIC STEINERNEMATID NEMATODES

Journal of Biological Researches ◽

10.23869/bphjbr.14.1.20082 ◽

2008 ◽

Vol 14 (1) ◽

pp. 7-13

Author(s):

Yuliantoro Baliadi

Keyword(s):

Insecticidal Activity ◽

Galleria Mellonella ◽

Liver Extract ◽

Axenic Culture ◽

Chicken Liver ◽

Monoxenic Culture ◽

Medium Supplement ◽

Steinernema Glaseri ◽

Extract Medium ◽

Uterine Development

Development and insecticidal activity of axenic infective juveniles (IJs) originating from endotokia matricida in maternal bodies of Steinernema glaseri and S. carpocapsae were investigated with a comparison to IJs developed in monoxenic culture. In comparison with the monoxenic steinernematids, the axenic ones grew slower and produced fewer IJs when they were cultured in a sterile chicken liver extract medium supplement with an autoclaved nematode infected Galleria mellonella larva. The phenomena of endotokia matricida, an intra-uterine development of hatched juvenile, occurred in an axenic culture as did the monoxenic ones. Although it occurred faster in monoxenic culture, the ratio of females bearing endotokia matricida was more numerous in axenic ones. These axenic females also produced IJs through the endotokia matricida phenomenon. Compared to the normal IJs develop in monoxenic culture, the IJs originated via endotokia matricida of axenic nematodes showed lower insecticidal activity and it could not reproduce in G. mellonella cadaver.

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