Abstract: Background: Transposable elements (TEs) have been recognized as potentially powerful dr... more Abstract: Background: Transposable elements (TEs) have been recognized as potentially powerful drivers of genomic evolutionary change, but factors affecting their mobility and regulation remain poorly understood. Chaperones such as Hsp90 buffer environmental perturbations by regulating protein conformation, but are also part of the PIWI-interacting RNA pathway, which regulates genomic instability arising from mobile TEs in the germline. Stress-induced mutagenesis from TE movement could thus arise from functional trade-offs in the dual roles of Hsp90. Results: We examined the functional constraints of Hsp90 and its role as a regulator of TE mobility by exposing nematodes (Caenorhabditis elegans and C. briggsae) to environmental stressors, with and without RNAi-induced silencing of Hsp90. TE excision rates increased with environmental stress intensity at multiple loci in several strains of each species. These effects were compounded by RNAi-induced knockdown of Hsp90. Mutation frequen...
Daphnia spp. cycle between asexual and sexual reproduction depending on environmental conditions.... more Daphnia spp. cycle between asexual and sexual reproduction depending on environmental conditions. Under favorable conditions, female offspring are produced, resulting in rapid population expansion; in less favorable conditions, males are produced, initiating the sexual portion of their lifestyle. Although mediated by environmental cues, sex determination is ultimately dependent on the expression of Doublesex genes, Dsx1 and Dsx2, in Daphnia. We exposed Daphnia magna to juvenile hormone III (JHIII), an insect hormone often mimicked by agricultural insecticides. Juvenile hormone (JH) and JH analogues disrupt the insect endocrine system, inhibiting reproduction; these chemicals pose an environmental threat to non-target organisms by leaching into surrounding waters. When Daphnia are exposed to JH or JH analogues, it causes a premature shift from asexual to sexual reproduction. We showed that Dsx1 is upregulated throughout vitellogenesis in female D. magna producing male offspring and Dsx2 is upregulated during late vitellogenesis, suggesting that both Dsx1 and Dsx2 contribute to male sex determination. In addition, Dsx2 expression increases during sexual maturation in male D. magna, suggesting that it also plays a role in spermatogenesis. In addition, exposure to JHIII decreased reproductive output, slowed growth and prematurely shifted D. magna reproductive strategy to the sexual portion of their lifecycle.
Estimated direct and indirect losses of grains and grain-based products caused by stored-product ... more Estimated direct and indirect losses of grains and grain-based products caused by stored-product insects range from about 10% in temperate regions to almost 50% in humid tropical areas. Pest management strategies in bulk grains include the use of fumigants such as phosphine and sulfuryl fluoride, and grain protectants, which are sprayed directly on commodities as they are loaded into storage. Fumigants, aerosols, and contact sprays are also used as structural treatments in mills, processing plants, and food warehouses. Some older organophosphate protectants and contact sprays have been phased out worldwide and have been replaced by safer insecticides, including pyrethroids and insect growth regulators (IGRs). These IGRs include juvenile hormone analogues (JHAs), ecdysteroids and chitin synthesis inhibitors, and are considered safe due to their insect specificity. Methoprene is the JHA that has been used most extensively in stored-product pest management. The formulations of methoprene originally introduced into the stored-product market in the 1980s contained the racemic mixture with both R-and S-forms, but now only the purified S-methoprene isomer is used. Methoprene has received broad attention and has been tested over decades for its direct lethal effects, but many recent studies focus more on sub-lethal effects. Although methoprene has been used for more than four decades, there has not been a recent and comprehensive synopsis or review of this IGR on stored-product insects. This review addresses the history and present use of methoprene with special emphasis on stored-product protection.
Tissue-specific mRNA profiling of the Brassica napus-Sclerotinia sclerotiorum interaction uncover... more Tissue-specific mRNA profiling of the Brassica napus-Sclerotinia sclerotiorum interaction uncovers novel regulators of plant immunity
Drosophila melanogaster has been an important model organism in the field of genetics for almost ... more Drosophila melanogaster has been an important model organism in the field of genetics for almost a century. Since the first release of its entire genome sequence in 2000, the challenge that Drosophila researchers face is to use reverse genetics to ascribe functions to the many newly identified genes. There are a variety of reverse genetics tools available for Drosophila researchers, such as chemical mutagenesis, transposon mutagenesis and gene replacement technologies, but none are as easy to use or as amenable to high throughput reverse genetic screens as RNA interference (RNAi). The simplest approaches to high-throughput RNAi have been conducted in Drosophila cell cultures however, the intent of this chapter is not to focus on high throughput RNAi techniques, but rather to provide protocols useful to the wider community of researchers who seek to validate the functions of a smaller subset of genes. Many of the protocols described are also readily adaptable to other insects, and readers are encouraged to adapt these protocols to the study of their favourite model species.
Biliary atresia (BA) is a rare neonatal disease with unknown causes. Approximately 10% of BA case... more Biliary atresia (BA) is a rare neonatal disease with unknown causes. Approximately 10% of BA cases develop in utero with other congenital defects that span a large spectrum of disease variations, including degeneration of the gall bladder and bile duct as well as malformation of the liver, intestines, and kidneys. Similar developmental alterations are manifested in a unique animal model, the sea lamprey (Petromyzon marinus), in which BA occurs naturally during metamorphosis. With the likelihood of conserved developmental mechanisms underlying organogenesis and degeneration, lamprey developmental BA may be a useful model to infer mechanisms underlying human embryonic BA. We reasoned that hepatobiliary transcriptomes regulate the transition between landmark stages of BA. Therefore, we examined sea lamprey hepatobiliary transcriptomes at four stages (M0, metamorphic stage 0 or larval stage, no BA; M2, metamorphic stage 2, onset of BA; M5, metamorphic stage 5, BA, and heightened hepatocyte proliferation and reorganization; and JV, juvenile, completion of BA) using messenger RNA sequencing and Kyoto Encyclopedia of Genes and Genomes pathway analyses. We found gene-expression patterns associated with the transition between these stages. In particular, transforming growth factor β (TGF-β), hedgehog, phosphatidylinositol-4,5-bisphosphate 3-kinase-Akt, Wnt, and mitogen-activated protein kinase pathways were involved during biliary degeneration. Furthermore, disrupting the TGF-β signaling pathway with antagonist or small interfering RNA treatments at the onset of BA delayed gall bladder and bile duct degeneration. Conclusion: Distinctive gene-expression patterns are associated with the degeneration of the biliary system during developmental BA. In addition, disrupting TGF-β signaling pathway at the onset of BA delayed biliary degeneration. (Hepatology Communications 2020;4:219-234). B iliary atresia (BA), a rare infant disease with unknown causes, is characterized by fibrosclerosing cholangiopathy, resulting in obstruction or obliteration of extrahepatic bile ducts (EHBDs) and intrahepatic bile ducts (IHBDs). (1) In the United States, BA affects 1 in 8,000 to 20,000 neonates, with 250 to 400 new cases reported every year. (2) Approx imately 10% of BA cases are embryonic form
Canadian Journal of Fisheries and Aquatic Sciences, 2021
Successful integrated management of the invasive predatory sea lamprey (Petromyzon marinus) in th... more Successful integrated management of the invasive predatory sea lamprey (Petromyzon marinus) in the Laurentian Great Lakes of North America is owed largely to the long history of beneficial use of two lampricides: 3-trifluoromethyl-4-nitrophenol (TFM) and 2′,5-dichloro-4′-nitrosalicylanilide (niclosamide). Ensuring continued successful sea lamprey control necessitates consideration of possible next-generation lampricides to supplement or replace current lampricides. This review identifies fifteen hallmarks of success for current lampricides to be used as design criteria in a search for next-generation lampricides. A three-stage research approach is outlined. Targeted research using omics, computer modelling, and high-throughput technology to define molecular mechanisms and high probability molecular targets for sea lamprey selective toxic action is crucial to prioritizing chemical candidates. Targeted delivery or identifying synergists to existing or new lampricides can provide incre...
Sclerotinia sclerotiorum is a pathogenic fungus that infects hundreds of crop species, causing ex... more Sclerotinia sclerotiorum is a pathogenic fungus that infects hundreds of crop species, causing extensive yield loss every year. Chemical fungicides are used to control this phytopathogen, but with concerns about increasing resistance and impacts on non-target species, there is a need to develop alternative control measures. In the present study, we engineered Brassica napus to constitutively express a hairpin (hp)RNA molecule to silence ABHYRDOLASE-3 in S. sclerotiorum. We demonstrate the potential for Host Induced Gene Silencing (HIGS) to protect B. napus from S. sclerotiorum using leaf, stem and whole plant infection assays. The interaction between the transgenic host plant and invading pathogen was further characterized at the molecular level using dual-RNA sequencing and at the anatomical level through microscopy to understand the processes and possible mechanisms leading to increased tolerance to this damaging necrotroph. We observed significant shifts in the expression of gene...
... General Esterase, Malathion Carboxylesterase, and Malathion Resistance in Culex tarsalis R. Z... more ... General Esterase, Malathion Carboxylesterase, and Malathion Resistance in Culex tarsalis R. ZIEGLER,' S. WHYARD, AER DOWNS, GR WYATT, AND VK ... In some other insects, includinghouse flies and blow flies, a decrease in general esterase activity has been correlated ...
Deformed wing virus (DWV) is a serious pathogen of the honey bee, Apis mellifera L., vectored by ... more Deformed wing virus (DWV) is a serious pathogen of the honey bee, Apis mellifera L., vectored by the parasitic mite Varroa destructor. The virus is associated with wing deformity in symptomatic bees, and premature death and reduced colony performance in asymptomatic bees. In the present study we reduced DWV infection by feeding both first instar larvae and adult A. mellifera with a double-stranded (ds) RNA construct, DWV-dsRNA, which is specific to DWV in DWV-inoculated bees, by mixing it with their food. We showed that feeding DWV to larvae causes wing deformity in adult bees in the absence of varroa mites and decreases survival rates of adult bees relative to bees not fed DWV. Feeding larvae with DWV-dsRNA in advance of inoculation with virus reduced the DWV viral level and reduced wing deformity relative to larvae fed DWV or DWV with green fluorescent protein-dsRNA (probably a result of RNA silencing), but did not affect survival to the adult stage. Feeding DWV-dsRNA did not affect larval survival rates, which suggests that dsRNA is non-toxic to larvae. Feeding adult workers with DWV-dsRNA in advance of inoculation with virus increased their longevity and reduced DWV concentration relative to controls.
Abstract: Background: Transposable elements (TEs) have been recognized as potentially powerful dr... more Abstract: Background: Transposable elements (TEs) have been recognized as potentially powerful drivers of genomic evolutionary change, but factors affecting their mobility and regulation remain poorly understood. Chaperones such as Hsp90 buffer environmental perturbations by regulating protein conformation, but are also part of the PIWI-interacting RNA pathway, which regulates genomic instability arising from mobile TEs in the germline. Stress-induced mutagenesis from TE movement could thus arise from functional trade-offs in the dual roles of Hsp90. Results: We examined the functional constraints of Hsp90 and its role as a regulator of TE mobility by exposing nematodes (Caenorhabditis elegans and C. briggsae) to environmental stressors, with and without RNAi-induced silencing of Hsp90. TE excision rates increased with environmental stress intensity at multiple loci in several strains of each species. These effects were compounded by RNAi-induced knockdown of Hsp90. Mutation frequen...
RNAi-mediated knockdown of nuclease genes in <i>B. tryoni</i> adults (50:50 males and... more RNAi-mediated knockdown of nuclease genes in <i>B. tryoni</i> adults (50:50 males and females) through hemocoel injections. qRT-PCR analyses were performed on adult whole bodies collected at 24 hours (a,b) and 48 hours (c,d) post-injection. The values represent the means and standard errors for 8 biological replicates. Asterisks indicate significant differences (<i>p</i><0.05).
Knockdown of <i>dsRNase1</i> and <i>dsRNase2</i> following different dsRN... more Knockdown of <i>dsRNase1</i> and <i>dsRNase2</i> following different dsRNA delivery methods. A) DsRNAs were injected into the hemolymph of adults (mixed sexes) and 6 days later, RNA was extracted to assess levels of <i>dsRNase1</i> and <i>dsRNase2</i> using qRT-PCR. Negative controls were injected with <i>gfp</i>-dsRNA, while experimental treatments were injected with a 50:50 mixtures of dsRNAs targeting both nuclease gene transcripts. Values represent the means and standard errors of 8 biological replicates; different letters indicate significant differences (ANOVA with Tukey's <i>post hoc</i> test, <i>p</i><0.05).
Identity matrix of <i>yellow</i> genes at the nucleotide level. Sequence identity ana... more Identity matrix of <i>yellow</i> genes at the nucleotide level. Sequence identity analyses were performed using the BLAST program of the National Center for Biotechnology Information (NCBI) (http://blast.ncbi.nlm.nih.gov/Blast.cgi).
Percentages of nucleotide similarity between the genome of <i>Bactrocera tryoni</i> a... more Percentages of nucleotide similarity between the genome of <i>Bactrocera tryoni</i> and nuclease genes in <i>B. dorsalis</i> and <i>Drosophila melanogaster</i>. Sequence identity analyses were performed using the BLAST program of the National Center for Biotechnology Information (NCBI) (http://blast.ncbi.nlm.nih.gov/Blast.cgi).
Persistence of knockdown of <i>dsRNase1</i> and <i>dsRNase2</i> in 5 day ... more Persistence of knockdown of <i>dsRNase1</i> and <i>dsRNase2</i> in 5 day old adults (mixed sexes) that had been fed dsRNA during their larval development. Larvae were fed either <i>gfp</i>-dsRNA (negative controls) or were fed mixtures of <i>yellow</i> dsRNA and dsRNAs targeting both <i>dsRNase1</i> and <i>dsRNase2</i>. A) Knockdown of <i>dsRNase1</i> and B) knockdown of <i>dsRNase2</i>. Values represent the means and standard errors of 8 biological replicates; different letters indicate significant differences (ANOVA with Tukey's <i>post hoc</i> test, <i>p</i><0.05).
Neighbour-joining phylogenetic trees of a) dsRNase1 and b) dsRNase2 with evolutionary distance co... more Neighbour-joining phylogenetic trees of a) dsRNase1 and b) dsRNase2 with evolutionary distance computed using the maximum composite likelihood method shown on the x-axis. Bootstrap values of 500 replicates are shown at nodes. Alignments were done using CLUSTAL O (1.2.4) and phylogenetic trees constructed and tested using MEGA X.
Abstract: Background: Transposable elements (TEs) have been recognized as potentially powerful dr... more Abstract: Background: Transposable elements (TEs) have been recognized as potentially powerful drivers of genomic evolutionary change, but factors affecting their mobility and regulation remain poorly understood. Chaperones such as Hsp90 buffer environmental perturbations by regulating protein conformation, but are also part of the PIWI-interacting RNA pathway, which regulates genomic instability arising from mobile TEs in the germline. Stress-induced mutagenesis from TE movement could thus arise from functional trade-offs in the dual roles of Hsp90. Results: We examined the functional constraints of Hsp90 and its role as a regulator of TE mobility by exposing nematodes (Caenorhabditis elegans and C. briggsae) to environmental stressors, with and without RNAi-induced silencing of Hsp90. TE excision rates increased with environmental stress intensity at multiple loci in several strains of each species. These effects were compounded by RNAi-induced knockdown of Hsp90. Mutation frequen...
Daphnia spp. cycle between asexual and sexual reproduction depending on environmental conditions.... more Daphnia spp. cycle between asexual and sexual reproduction depending on environmental conditions. Under favorable conditions, female offspring are produced, resulting in rapid population expansion; in less favorable conditions, males are produced, initiating the sexual portion of their lifestyle. Although mediated by environmental cues, sex determination is ultimately dependent on the expression of Doublesex genes, Dsx1 and Dsx2, in Daphnia. We exposed Daphnia magna to juvenile hormone III (JHIII), an insect hormone often mimicked by agricultural insecticides. Juvenile hormone (JH) and JH analogues disrupt the insect endocrine system, inhibiting reproduction; these chemicals pose an environmental threat to non-target organisms by leaching into surrounding waters. When Daphnia are exposed to JH or JH analogues, it causes a premature shift from asexual to sexual reproduction. We showed that Dsx1 is upregulated throughout vitellogenesis in female D. magna producing male offspring and Dsx2 is upregulated during late vitellogenesis, suggesting that both Dsx1 and Dsx2 contribute to male sex determination. In addition, Dsx2 expression increases during sexual maturation in male D. magna, suggesting that it also plays a role in spermatogenesis. In addition, exposure to JHIII decreased reproductive output, slowed growth and prematurely shifted D. magna reproductive strategy to the sexual portion of their lifecycle.
Estimated direct and indirect losses of grains and grain-based products caused by stored-product ... more Estimated direct and indirect losses of grains and grain-based products caused by stored-product insects range from about 10% in temperate regions to almost 50% in humid tropical areas. Pest management strategies in bulk grains include the use of fumigants such as phosphine and sulfuryl fluoride, and grain protectants, which are sprayed directly on commodities as they are loaded into storage. Fumigants, aerosols, and contact sprays are also used as structural treatments in mills, processing plants, and food warehouses. Some older organophosphate protectants and contact sprays have been phased out worldwide and have been replaced by safer insecticides, including pyrethroids and insect growth regulators (IGRs). These IGRs include juvenile hormone analogues (JHAs), ecdysteroids and chitin synthesis inhibitors, and are considered safe due to their insect specificity. Methoprene is the JHA that has been used most extensively in stored-product pest management. The formulations of methoprene originally introduced into the stored-product market in the 1980s contained the racemic mixture with both R-and S-forms, but now only the purified S-methoprene isomer is used. Methoprene has received broad attention and has been tested over decades for its direct lethal effects, but many recent studies focus more on sub-lethal effects. Although methoprene has been used for more than four decades, there has not been a recent and comprehensive synopsis or review of this IGR on stored-product insects. This review addresses the history and present use of methoprene with special emphasis on stored-product protection.
Tissue-specific mRNA profiling of the Brassica napus-Sclerotinia sclerotiorum interaction uncover... more Tissue-specific mRNA profiling of the Brassica napus-Sclerotinia sclerotiorum interaction uncovers novel regulators of plant immunity
Drosophila melanogaster has been an important model organism in the field of genetics for almost ... more Drosophila melanogaster has been an important model organism in the field of genetics for almost a century. Since the first release of its entire genome sequence in 2000, the challenge that Drosophila researchers face is to use reverse genetics to ascribe functions to the many newly identified genes. There are a variety of reverse genetics tools available for Drosophila researchers, such as chemical mutagenesis, transposon mutagenesis and gene replacement technologies, but none are as easy to use or as amenable to high throughput reverse genetic screens as RNA interference (RNAi). The simplest approaches to high-throughput RNAi have been conducted in Drosophila cell cultures however, the intent of this chapter is not to focus on high throughput RNAi techniques, but rather to provide protocols useful to the wider community of researchers who seek to validate the functions of a smaller subset of genes. Many of the protocols described are also readily adaptable to other insects, and readers are encouraged to adapt these protocols to the study of their favourite model species.
Biliary atresia (BA) is a rare neonatal disease with unknown causes. Approximately 10% of BA case... more Biliary atresia (BA) is a rare neonatal disease with unknown causes. Approximately 10% of BA cases develop in utero with other congenital defects that span a large spectrum of disease variations, including degeneration of the gall bladder and bile duct as well as malformation of the liver, intestines, and kidneys. Similar developmental alterations are manifested in a unique animal model, the sea lamprey (Petromyzon marinus), in which BA occurs naturally during metamorphosis. With the likelihood of conserved developmental mechanisms underlying organogenesis and degeneration, lamprey developmental BA may be a useful model to infer mechanisms underlying human embryonic BA. We reasoned that hepatobiliary transcriptomes regulate the transition between landmark stages of BA. Therefore, we examined sea lamprey hepatobiliary transcriptomes at four stages (M0, metamorphic stage 0 or larval stage, no BA; M2, metamorphic stage 2, onset of BA; M5, metamorphic stage 5, BA, and heightened hepatocyte proliferation and reorganization; and JV, juvenile, completion of BA) using messenger RNA sequencing and Kyoto Encyclopedia of Genes and Genomes pathway analyses. We found gene-expression patterns associated with the transition between these stages. In particular, transforming growth factor β (TGF-β), hedgehog, phosphatidylinositol-4,5-bisphosphate 3-kinase-Akt, Wnt, and mitogen-activated protein kinase pathways were involved during biliary degeneration. Furthermore, disrupting the TGF-β signaling pathway with antagonist or small interfering RNA treatments at the onset of BA delayed gall bladder and bile duct degeneration. Conclusion: Distinctive gene-expression patterns are associated with the degeneration of the biliary system during developmental BA. In addition, disrupting TGF-β signaling pathway at the onset of BA delayed biliary degeneration. (Hepatology Communications 2020;4:219-234). B iliary atresia (BA), a rare infant disease with unknown causes, is characterized by fibrosclerosing cholangiopathy, resulting in obstruction or obliteration of extrahepatic bile ducts (EHBDs) and intrahepatic bile ducts (IHBDs). (1) In the United States, BA affects 1 in 8,000 to 20,000 neonates, with 250 to 400 new cases reported every year. (2) Approx imately 10% of BA cases are embryonic form
Canadian Journal of Fisheries and Aquatic Sciences, 2021
Successful integrated management of the invasive predatory sea lamprey (Petromyzon marinus) in th... more Successful integrated management of the invasive predatory sea lamprey (Petromyzon marinus) in the Laurentian Great Lakes of North America is owed largely to the long history of beneficial use of two lampricides: 3-trifluoromethyl-4-nitrophenol (TFM) and 2′,5-dichloro-4′-nitrosalicylanilide (niclosamide). Ensuring continued successful sea lamprey control necessitates consideration of possible next-generation lampricides to supplement or replace current lampricides. This review identifies fifteen hallmarks of success for current lampricides to be used as design criteria in a search for next-generation lampricides. A three-stage research approach is outlined. Targeted research using omics, computer modelling, and high-throughput technology to define molecular mechanisms and high probability molecular targets for sea lamprey selective toxic action is crucial to prioritizing chemical candidates. Targeted delivery or identifying synergists to existing or new lampricides can provide incre...
Sclerotinia sclerotiorum is a pathogenic fungus that infects hundreds of crop species, causing ex... more Sclerotinia sclerotiorum is a pathogenic fungus that infects hundreds of crop species, causing extensive yield loss every year. Chemical fungicides are used to control this phytopathogen, but with concerns about increasing resistance and impacts on non-target species, there is a need to develop alternative control measures. In the present study, we engineered Brassica napus to constitutively express a hairpin (hp)RNA molecule to silence ABHYRDOLASE-3 in S. sclerotiorum. We demonstrate the potential for Host Induced Gene Silencing (HIGS) to protect B. napus from S. sclerotiorum using leaf, stem and whole plant infection assays. The interaction between the transgenic host plant and invading pathogen was further characterized at the molecular level using dual-RNA sequencing and at the anatomical level through microscopy to understand the processes and possible mechanisms leading to increased tolerance to this damaging necrotroph. We observed significant shifts in the expression of gene...
... General Esterase, Malathion Carboxylesterase, and Malathion Resistance in Culex tarsalis R. Z... more ... General Esterase, Malathion Carboxylesterase, and Malathion Resistance in Culex tarsalis R. ZIEGLER,' S. WHYARD, AER DOWNS, GR WYATT, AND VK ... In some other insects, includinghouse flies and blow flies, a decrease in general esterase activity has been correlated ...
Deformed wing virus (DWV) is a serious pathogen of the honey bee, Apis mellifera L., vectored by ... more Deformed wing virus (DWV) is a serious pathogen of the honey bee, Apis mellifera L., vectored by the parasitic mite Varroa destructor. The virus is associated with wing deformity in symptomatic bees, and premature death and reduced colony performance in asymptomatic bees. In the present study we reduced DWV infection by feeding both first instar larvae and adult A. mellifera with a double-stranded (ds) RNA construct, DWV-dsRNA, which is specific to DWV in DWV-inoculated bees, by mixing it with their food. We showed that feeding DWV to larvae causes wing deformity in adult bees in the absence of varroa mites and decreases survival rates of adult bees relative to bees not fed DWV. Feeding larvae with DWV-dsRNA in advance of inoculation with virus reduced the DWV viral level and reduced wing deformity relative to larvae fed DWV or DWV with green fluorescent protein-dsRNA (probably a result of RNA silencing), but did not affect survival to the adult stage. Feeding DWV-dsRNA did not affect larval survival rates, which suggests that dsRNA is non-toxic to larvae. Feeding adult workers with DWV-dsRNA in advance of inoculation with virus increased their longevity and reduced DWV concentration relative to controls.
Abstract: Background: Transposable elements (TEs) have been recognized as potentially powerful dr... more Abstract: Background: Transposable elements (TEs) have been recognized as potentially powerful drivers of genomic evolutionary change, but factors affecting their mobility and regulation remain poorly understood. Chaperones such as Hsp90 buffer environmental perturbations by regulating protein conformation, but are also part of the PIWI-interacting RNA pathway, which regulates genomic instability arising from mobile TEs in the germline. Stress-induced mutagenesis from TE movement could thus arise from functional trade-offs in the dual roles of Hsp90. Results: We examined the functional constraints of Hsp90 and its role as a regulator of TE mobility by exposing nematodes (Caenorhabditis elegans and C. briggsae) to environmental stressors, with and without RNAi-induced silencing of Hsp90. TE excision rates increased with environmental stress intensity at multiple loci in several strains of each species. These effects were compounded by RNAi-induced knockdown of Hsp90. Mutation frequen...
RNAi-mediated knockdown of nuclease genes in <i>B. tryoni</i> adults (50:50 males and... more RNAi-mediated knockdown of nuclease genes in <i>B. tryoni</i> adults (50:50 males and females) through hemocoel injections. qRT-PCR analyses were performed on adult whole bodies collected at 24 hours (a,b) and 48 hours (c,d) post-injection. The values represent the means and standard errors for 8 biological replicates. Asterisks indicate significant differences (<i>p</i><0.05).
Knockdown of <i>dsRNase1</i> and <i>dsRNase2</i> following different dsRN... more Knockdown of <i>dsRNase1</i> and <i>dsRNase2</i> following different dsRNA delivery methods. A) DsRNAs were injected into the hemolymph of adults (mixed sexes) and 6 days later, RNA was extracted to assess levels of <i>dsRNase1</i> and <i>dsRNase2</i> using qRT-PCR. Negative controls were injected with <i>gfp</i>-dsRNA, while experimental treatments were injected with a 50:50 mixtures of dsRNAs targeting both nuclease gene transcripts. Values represent the means and standard errors of 8 biological replicates; different letters indicate significant differences (ANOVA with Tukey's <i>post hoc</i> test, <i>p</i><0.05).
Identity matrix of <i>yellow</i> genes at the nucleotide level. Sequence identity ana... more Identity matrix of <i>yellow</i> genes at the nucleotide level. Sequence identity analyses were performed using the BLAST program of the National Center for Biotechnology Information (NCBI) (http://blast.ncbi.nlm.nih.gov/Blast.cgi).
Percentages of nucleotide similarity between the genome of <i>Bactrocera tryoni</i> a... more Percentages of nucleotide similarity between the genome of <i>Bactrocera tryoni</i> and nuclease genes in <i>B. dorsalis</i> and <i>Drosophila melanogaster</i>. Sequence identity analyses were performed using the BLAST program of the National Center for Biotechnology Information (NCBI) (http://blast.ncbi.nlm.nih.gov/Blast.cgi).
Persistence of knockdown of <i>dsRNase1</i> and <i>dsRNase2</i> in 5 day ... more Persistence of knockdown of <i>dsRNase1</i> and <i>dsRNase2</i> in 5 day old adults (mixed sexes) that had been fed dsRNA during their larval development. Larvae were fed either <i>gfp</i>-dsRNA (negative controls) or were fed mixtures of <i>yellow</i> dsRNA and dsRNAs targeting both <i>dsRNase1</i> and <i>dsRNase2</i>. A) Knockdown of <i>dsRNase1</i> and B) knockdown of <i>dsRNase2</i>. Values represent the means and standard errors of 8 biological replicates; different letters indicate significant differences (ANOVA with Tukey's <i>post hoc</i> test, <i>p</i><0.05).
Neighbour-joining phylogenetic trees of a) dsRNase1 and b) dsRNase2 with evolutionary distance co... more Neighbour-joining phylogenetic trees of a) dsRNase1 and b) dsRNase2 with evolutionary distance computed using the maximum composite likelihood method shown on the x-axis. Bootstrap values of 500 replicates are shown at nodes. Alignments were done using CLUSTAL O (1.2.4) and phylogenetic trees constructed and tested using MEGA X.
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Papers by Steven Whyard