<p>Pupal weights (A) <i>n</i> = at least 52. Larval development time (B) (from ... more <p>Pupal weights (A) <i>n</i> = at least 52. Larval development time (B) (from egg hatching to onset of pupation) <i>n</i> = at least 52 (data presented as mean +/− SEM). a-P<0.05 compared with the NS insects.</p
<p>Expression of antimicrobial peptide genes and other putative immunity/stress-management ... more <p>Expression of antimicrobial peptide genes and other putative immunity/stress-management genes in the integument and fat body of non-selected (NS) and selected line (S) larvae, under basal (uninfected) conditions (A), 24 h after topical <i>B. bassiana</i> infection (B) and 24 h after <i>M. anisopliae</i> topical infection (C). Basal expression in uninfected S larvae (A) is illustrated as a fold change relative to NS uninfected larvae and the x-axis represents basal expression in NS larvae (i.e. 1-fold). Fold induction in infected NS larvae is also calculated relative to NS uninfected larvae (B, C). Fold induction in infected S larvae is calculated relative to the S uninfected expression (B, C). The mean ΔΔCt value of 3 independent experiments (each with a minimum of 5 insects per treatment) and the SEM are reported. *-P<0.05, **-P<0.01, compared with the corresponding induced change in NS line insects. Na = not assayed in fat body tissue.</p
Three types of modified silicon dioxide nanoparticles (SiO2, 10–20 nm) with additives of epoxy, s... more Three types of modified silicon dioxide nanoparticles (SiO2, 10–20 nm) with additives of epoxy, silane and amino groups, used independently and in combination with the entomopathogenic bacteria Bacillus thuringiensis subsp. morrisoni and fungus Metarhizium robertsii were tested against Colorado potato beetle (Leptinotarsa decemlineata) and cabbage beetles (Phyllotreta spp.). All three nanoparticles were found to have an entomocidal effect on Colorado potato beetle larvae and crucifer flea beetles when ingested. Increased susceptibility of insects to B. thuringiensis or M. robertsii blastospores and their metabolites was shown after exposure to the modified silicon dioxide nanoparticles. The potential of modified silicon dioxide nanoparticles to enhance the efficiency of biopesticides based on the bacteria B. thuringiensis and fungi M. robertsii is considered in the paper.
<p>Mortality rate of selected line and non-selected line of <i>G. mellonella</i>... more <p>Mortality rate of selected line and non-selected line of <i>G. mellonella</i> larvae following topical treatment with the fungus <i>B. bassiana</i> (A) and <i>M.anisopliae</i> (B). (a-P<0.01 compared with non-selected line larvae. n = 140–190 per line per treatment).</p
<p>Cuticular phenoloxidase (PO) activity (A), hemolymph phenoloxidase (B), lysozyme-like (C... more <p>Cuticular phenoloxidase (PO) activity (A), hemolymph phenoloxidase (B), lysozyme-like (C) activity and encapsulation responses (D) in hemolymph of <i>G. mellonella</i> larvae from non-selected (NS) and selected (S) lines at 24 h following topical application of <i>B. bassiana</i> (Bb) and <i>M. anisopliae</i> (Ma) (data presented as mean +/− SEM; a-P<0.05, b-P<0.01, c-P<0.001 compared with uninfected larvae from the same line; d-P<0.05 e-P<0.01 f-P<0.001 compared with NS larvae with the same treatment).</p
In order for entomopathogenic fungi to colonise an insect host, they must first attach to, and pe... more In order for entomopathogenic fungi to colonise an insect host, they must first attach to, and penetrate, the cuticle layers of the integument. Herein, we explored the interactions between the fungal pathogen <i>Metarhizium brunneum</i> ARSEF 4556 and two immunologically distinct morphs, melanic (M) and non-melanic (NM), of the greater wax moth <i>Galleria mellonella</i>. We first interrogated the cuticular compositions of both insect morphs to reveal substantial differences in their physiochemical properties. Enhanced melanin accumulation, fewer hydrocarbons, and higher <sub>L</sub>-dihydroxyphenylalanine (DOPA) decarboxylase activity were evident in the cuticle of the M larvae. This "hostile" terrain proved challenging for <i>M. brunneum –</i> reflected in poor conidial attachment and germination, and elevated expression of stress-associated genes (<i>e.g., Hsp30, Hsp70</i>). Lack of adherence to the cuticle imp...
The co-evolutionary arms race between disease-causing agents and their insect victims is ancient ... more The co-evolutionary arms race between disease-causing agents and their insect victims is ancient and complex - leading to the development of specialised attack and defence strategies. Among such strategies is the capacity of fungal and oomycete pathogens to deploy degradative enzymes, notably proteases, to facilitate infection directly across the integument. To counter these proteases, insects such as the greater wax moth Galleria mellonella release metalloprotease inhibitors and other immune factors to thwart the invading fungus. To date, molecular-based confirmation of insect metalloprotease inhibitor's incontrovertible role in antifungal defence has been lacking. We targeted the IMPI gene for suppression using RNAi and exposed those insects to the entomopathogenic fungus Metarhizium brunneum ARSEF4556. Levels of IMPI were reduced significantly in the integument (10-fold) and fat body (5-fold) of RNAi-treated insects when compared to control larvae, and displayed a significantly higher mortality rate. We also surveyed candidate immune/detoxification gene expression levels (e.g., DOPA decarboxylase, galiomycin) in three tissues (integument, midgut, fat body) in order to gauge any potential non-target effects of RNAi. The loss of IMPI via RNAi compromises antifungal defences and leaves G. mellonella vulnerable to infection.
Microevolutionary mechanisms of resistance to a bacterial pathogen were explored in a population ... more Microevolutionary mechanisms of resistance to a bacterial pathogen were explored in a population of the Greater wax moth, Galleria mellonella, selected for an 8.8-fold increased resistance against the entomopathogenic bacterium Bacillus thuringiensis (Bt) compared with a non-selected (suspectible) line. Defence strategies of the resistantand susceptible insect lines were compared to uncover mechanisms underpinning resistance, and the possible cost of those survival strategies. In the uninfected state, resistant insects exhibited enhanced basal expression of genes related to regeneration and amelioration of Bt toxin activity in the midgut. In addition, these insects also exhibited elevated activity of genes linked to inflammation/stress management and immune defence in the fat body. Following oral infection with Bt, the expression of these genes was further elevated in the fat body and midgut of both lines and to a greater extent some of them in resistant line than the susceptible li...
<p>Pupal weights (A) <i>n</i> = at least 52. Larval development time (B) (from ... more <p>Pupal weights (A) <i>n</i> = at least 52. Larval development time (B) (from egg hatching to onset of pupation) <i>n</i> = at least 52 (data presented as mean +/− SEM). a-P<0.05 compared with the NS insects.</p
<p>Expression of antimicrobial peptide genes and other putative immunity/stress-management ... more <p>Expression of antimicrobial peptide genes and other putative immunity/stress-management genes in the integument and fat body of non-selected (NS) and selected line (S) larvae, under basal (uninfected) conditions (A), 24 h after topical <i>B. bassiana</i> infection (B) and 24 h after <i>M. anisopliae</i> topical infection (C). Basal expression in uninfected S larvae (A) is illustrated as a fold change relative to NS uninfected larvae and the x-axis represents basal expression in NS larvae (i.e. 1-fold). Fold induction in infected NS larvae is also calculated relative to NS uninfected larvae (B, C). Fold induction in infected S larvae is calculated relative to the S uninfected expression (B, C). The mean ΔΔCt value of 3 independent experiments (each with a minimum of 5 insects per treatment) and the SEM are reported. *-P<0.05, **-P<0.01, compared with the corresponding induced change in NS line insects. Na = not assayed in fat body tissue.</p
Three types of modified silicon dioxide nanoparticles (SiO2, 10–20 nm) with additives of epoxy, s... more Three types of modified silicon dioxide nanoparticles (SiO2, 10–20 nm) with additives of epoxy, silane and amino groups, used independently and in combination with the entomopathogenic bacteria Bacillus thuringiensis subsp. morrisoni and fungus Metarhizium robertsii were tested against Colorado potato beetle (Leptinotarsa decemlineata) and cabbage beetles (Phyllotreta spp.). All three nanoparticles were found to have an entomocidal effect on Colorado potato beetle larvae and crucifer flea beetles when ingested. Increased susceptibility of insects to B. thuringiensis or M. robertsii blastospores and their metabolites was shown after exposure to the modified silicon dioxide nanoparticles. The potential of modified silicon dioxide nanoparticles to enhance the efficiency of biopesticides based on the bacteria B. thuringiensis and fungi M. robertsii is considered in the paper.
<p>Mortality rate of selected line and non-selected line of <i>G. mellonella</i>... more <p>Mortality rate of selected line and non-selected line of <i>G. mellonella</i> larvae following topical treatment with the fungus <i>B. bassiana</i> (A) and <i>M.anisopliae</i> (B). (a-P<0.01 compared with non-selected line larvae. n = 140–190 per line per treatment).</p
<p>Cuticular phenoloxidase (PO) activity (A), hemolymph phenoloxidase (B), lysozyme-like (C... more <p>Cuticular phenoloxidase (PO) activity (A), hemolymph phenoloxidase (B), lysozyme-like (C) activity and encapsulation responses (D) in hemolymph of <i>G. mellonella</i> larvae from non-selected (NS) and selected (S) lines at 24 h following topical application of <i>B. bassiana</i> (Bb) and <i>M. anisopliae</i> (Ma) (data presented as mean +/− SEM; a-P<0.05, b-P<0.01, c-P<0.001 compared with uninfected larvae from the same line; d-P<0.05 e-P<0.01 f-P<0.001 compared with NS larvae with the same treatment).</p
In order for entomopathogenic fungi to colonise an insect host, they must first attach to, and pe... more In order for entomopathogenic fungi to colonise an insect host, they must first attach to, and penetrate, the cuticle layers of the integument. Herein, we explored the interactions between the fungal pathogen <i>Metarhizium brunneum</i> ARSEF 4556 and two immunologically distinct morphs, melanic (M) and non-melanic (NM), of the greater wax moth <i>Galleria mellonella</i>. We first interrogated the cuticular compositions of both insect morphs to reveal substantial differences in their physiochemical properties. Enhanced melanin accumulation, fewer hydrocarbons, and higher <sub>L</sub>-dihydroxyphenylalanine (DOPA) decarboxylase activity were evident in the cuticle of the M larvae. This "hostile" terrain proved challenging for <i>M. brunneum –</i> reflected in poor conidial attachment and germination, and elevated expression of stress-associated genes (<i>e.g., Hsp30, Hsp70</i>). Lack of adherence to the cuticle imp...
The co-evolutionary arms race between disease-causing agents and their insect victims is ancient ... more The co-evolutionary arms race between disease-causing agents and their insect victims is ancient and complex - leading to the development of specialised attack and defence strategies. Among such strategies is the capacity of fungal and oomycete pathogens to deploy degradative enzymes, notably proteases, to facilitate infection directly across the integument. To counter these proteases, insects such as the greater wax moth Galleria mellonella release metalloprotease inhibitors and other immune factors to thwart the invading fungus. To date, molecular-based confirmation of insect metalloprotease inhibitor's incontrovertible role in antifungal defence has been lacking. We targeted the IMPI gene for suppression using RNAi and exposed those insects to the entomopathogenic fungus Metarhizium brunneum ARSEF4556. Levels of IMPI were reduced significantly in the integument (10-fold) and fat body (5-fold) of RNAi-treated insects when compared to control larvae, and displayed a significantly higher mortality rate. We also surveyed candidate immune/detoxification gene expression levels (e.g., DOPA decarboxylase, galiomycin) in three tissues (integument, midgut, fat body) in order to gauge any potential non-target effects of RNAi. The loss of IMPI via RNAi compromises antifungal defences and leaves G. mellonella vulnerable to infection.
Microevolutionary mechanisms of resistance to a bacterial pathogen were explored in a population ... more Microevolutionary mechanisms of resistance to a bacterial pathogen were explored in a population of the Greater wax moth, Galleria mellonella, selected for an 8.8-fold increased resistance against the entomopathogenic bacterium Bacillus thuringiensis (Bt) compared with a non-selected (suspectible) line. Defence strategies of the resistantand susceptible insect lines were compared to uncover mechanisms underpinning resistance, and the possible cost of those survival strategies. In the uninfected state, resistant insects exhibited enhanced basal expression of genes related to regeneration and amelioration of Bt toxin activity in the midgut. In addition, these insects also exhibited elevated activity of genes linked to inflammation/stress management and immune defence in the fat body. Following oral infection with Bt, the expression of these genes was further elevated in the fat body and midgut of both lines and to a greater extent some of them in resistant line than the susceptible li...
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