Effects of pH on Double Stranded RNA Stability in European Corn Borer Nucleases

RNA interference (RNAI) is an immune response that can be exploited to make greener pesticides. It works by inciting suppression of a specific target gene using fed or injected dsRNA. Targeting a specific gene sequence also means RNAi can be used to target a specific organism. However, some insects, such as lepidopterans, have nucleases, called dsRNases, in their gut and hemolymph that sever dsRNA and lower RNAi efficiency (1). Ostrinia nubilalis, the European corn borer, (ECB), is a prime example of a lepidopteran pest which decimates corn supplies across the Midwest and does not respond to RNAi. Comparison of dsRNA stability in dsRNase genes in ECB and western corn rootworm (WCR), a coleopteran pest that has very high RNAi efficiency, indicates that dsRNA is rapidly degraded in ECB tissues, but not WCR tissues, despite similar expression of dsRNase genes in both species. These findings suggest that another variable, such as pH may be influencing dsRNA stability in insects (2)

Evaluation of Several Genes as Targets for RNAi in German Cockroach

Citation: Banister, H. (2017). Evaluation of Several Genes as Targets for RNAi in German Cockroach . 1st Annual Undergraduate Research Experience in Entomology Symposium, November 16, 2016. Manhattam, KS.RNA interference (RNAi) was first discovered in nematodes when exogenous double-stranded RNA (dsRNA) complementary to a specific gene suppressed expression of that gene [1]. Subsequently, much effort has been devoted to developing RNAi as a highly specific tool for therapeutic interventions and control of insect pests [2]. However, there are still many challenges associated with using RNAi to control insects, including efficient delivery and selection of appropriate targets. In this study, we evaluated three genes as potential targets for causing mortality via RNAi in German cockroach, Blatella germanica. German cockroaches are ubiquitous structural pests that can serve as reservoirs for species of pathogenic bacteria, viruses, or fungi in humans [3,4] and is an excellent organism for exploring insect control using RNAi. Injection of dsRNA complementary to either tubulin (Tub, a cytoskeletal structural protein), VATPase subunit 1 (Vha, an integral membrane protein), or Snf7 (an ESCRT III protein) caused decreased survival with Snf7 causing the greatest and fastest mortality (LT50 = 8.2 days). Our results demonstrate that when suppressed with RNAi, these genes could be effective targets for cockroach control. Furthermore, knowing these genes can be effectively used for RNAi, we can now attempt to understand why methods other than injection for RNAi delivery are less efficient in an effort to improve the utility of RNAi in insect control

Evaluation of Several Genes as Targets for RNAi in German Cockroach

Citation: Wilkins, R. (2017). Evaluation of Several Genes as Targets for RNAi in German Cockroach . 1st Annual Undergraduate Research Experience in Entomology Symposium, November 16, 2016. Manhattam, KS.RNA interference (RNAi) was first discovered in nematodes when exogenous double-stranded RNA (dsRNA) complementary to a specific gene suppressed expression of that gene [1]. Subsequently, much effort has been devoted to developing RNAi as a highly specific tool for therapeutic interventions and control of insect pests [2]. However, there are still many challenges associated with using RNAi to control insects, including efficient delivery and selection of appropriate targets. In this study, we evaluated three genes as potential targets for causing mortality via RNAi in German cockroach, Blatella germanica. German cockroaches are ubiquitous structural pests that can serve as reservoirs for species of pathogenic bacteria, viruses, or fungi in humans [3,4] and is an excellent organism for exploring insect control using RNAi. Injection of dsRNA complementary to either tubulin (Tub, a cytoskeletal structural protein), VATPase subunit 1 (Vha, an integral membrane protein), or Snf7 (an ESCRT III protein) caused decreased survival with Snf7 causing the greatest and fastest mortality (LT50 = 8.2 days). Our results demonstrate that when suppressed with RNAi, these genes could be effective targets for cockroach control. Furthermore, knowing these genes can be effectively used for RNAi, we can now attempt to understand why methods other than injection for RNAi delivery are less efficient in an effort to improve the utility of RNAi in insect control

Sino-European Transcontinental Basic and Clinical High-Tech Acupuncture Studies—Part 1: Auricular Acupuncture Increases Heart Rate Variability in Anesthetized Rats

Evidence-based research concerning the effects of high-tech acupuncture on autonomic function was performed by two research teams from China and Austria. This study describes the first transcontinental teleacupuncture measurements in animals. Heart rate (HR) and heart rate variability (HRV) recordings in 10 male Sprague-Dawley anesthetized rats were performed under stable conditions in Beijing, China, and the data analysis was completed in Graz, Austria. The electrocardiograms (ECGs) were recorded by an HRV Medilog AR12 system during acupuncture of the ear and body (PC6 Neiguan, CV12 Zhongwan, ST36 Zusanli). The data were analyzed using specially adapted novel Austrian software. HR did not change significantly during any acupuncture stimulation in anesthetized rats (ear acupuncture, PC6, CV12, or ST36). Total HRV only changed significantly (P = 0.025) during auricular acupuncture (acupoint heart). The low-frequency/high-frequency ratio parameter decreased significantly (P = 0.03) during stimulation of ST36. This change was based on intensification of the related mechanism of blood pressure regulation that has been demonstrated in previous studies in humans. Modernization of acupuncture research performed as a collaboration between China and Austria has also been demonstrated

Effects of Nanoparticles on Double-Stranded RNA Stability in Moth Hemolymph

RNA interference (RNAi) is an immune response in which double-stranded RNA (dsRNA) suppresses a target gene. By designing dsRNA to target genes that are necessary for life, dsRNA can potentially be used as an insecticide. RNAi-based insecticides are badly needed because they are more specific than conventional pesticides and because many insects have developed resistance to pesticides. Unfortunately, some insects produce enzymes that degrade dsRNA and prevent the RNAi response (Cooperet al., 2018). Therefore, RNAi-based insecticides currently cannot be used to control all insects. Here we investigate dsRNA stability when incubated in hemolymph ex vivoto determine if degradation of dsRNA is contributing to the inadequate RNAi response exhibited by lepidopterans, such as the European corn borer (ECB, Ostrinia nubilalis). Our findings indicate that dsRNA is significantly degraded in ECB hemolymph, but encapsulation of dsRNA in chitosan-based nanoparticles (CB-NPs) enhances stability. These findings provide insight into RNAi efficiency limitations in insects, and may provide a method to enhance RNAi efficiency in lepidopterans and other RNAi-refractory pests

Effects of Nanoparticles on Double-Stranded RNA Stability in Corn Soil

Double-stranded RNA (dsRNA) can potentially be used as a pesticide because these molecules trigger an immune response called RNA interference (RNAi). If the expression of essential genes matching the dsRNA sequence are silenced, then the pest dies. New classes of pesticides, including RNAi-based pesticides, are needed to overcome pesticide resistance and reduce the environmental impacts of pesticides. Unfortunately, dsRNA is easily degraded by enzymes in the environment, particularly those produced by microbes in the soil (Dubelmanet al., 2014),severely limiting delivery of dsRNA to cryptic (soil dwelling) species unless transgenic plants are used. Here we investigate dsRNA stability when incubated in corn soil supernatant ex situ to determine if encapsulation of dsRNA in chitosan-basednanoparticles (CB-NPs) enhances stability in corn soil. Interestingly, dsRNA stability was not affected by soil supernatant, possibly due to the time of year when sampling was performed (Icozet al., 2008). Nonetheless, these findings provide insight into dsRNA stability in soil, and in the future may lead to a method for protecting dsRNA from environmental degradation using CB-NPs

Effects of Nanoparticles on Double-Stranded RNA Stability in Corn Soil

Double-stranded RNA (dsRNA) can potentially be used as a pesticide because these molecules trigger an immune response called RNA interference (RNAi). If the expression of essential genes matching the dsRNA sequence are silenced, then the pest dies. New classes of pesticides, including RNAi-based pesticides, are needed to overcome pesticide resistance and reduce the environmental impacts of pesticides. Unfortunately, dsRNA is easily degraded by enzymes in the environment, particularly those produced by microbes in the soil (Dubelmanet al., 2014),severely limiting delivery of dsRNA to cryptic (soil dwelling) species unless transgenic plants are used. Here we investigate dsRNA stability when incubated in corn soil supernatant ex situ to determine if encapsulation of dsRNA in chitosan-basednanoparticles (CB-NPs) enhances stability in corn soil. Interestingly, dsRNA stability was not affected by soil supernatant, possibly due to the time of year when sampling was performed (Icozet al., 2008). Nonetheless, these findings provide insight into dsRNA stability in soil, and in the future may lead to a method for protecting dsRNA from environmental degradation using CB-NPs

Induction of Reproductive Diapause in \u3ci\u3eHabrobracon hebetor\u3c/i\u3e (Hymenoptera: Braconidae) When Reared at Different Photoperiods at Low Temperatures

Development of the parasitoid Habrobracon hebetor (Say) (Hymenoptera: Braconidae) at low temperatures was determined to identify rearing conditions that might result in adults that were in reproductive diapause. Diapausing adults would be expected to survive cold storage longer than nondiapausing adults for use in biological control programs. Only a few eggs were found in the ovaries when H. hebetor females were reared during the immature stages at 17.5 and 20°C with a 16-h photoperiod, and the ovaries were poorly developed and contained no eggs when females were reared with a 10-h photoperiod in these low temperatures. Rearing H. hebetor at 17.5 and 20°C did not result in diapause of immature stages, but did appear to result in possible adult reproductive diapause when the immature stages were reared with a 10-h photoperiod. Females reared during the immature stages at 17.5°C with a 10-h photoperiod lived longer and took longer to lay their first eggs and to lay 50% of their eggs than those females reared at 17.5°C with a 16-h photoperiod. Females reared during the immature stages at 20°C with a 10-h photoperiod took longer to lay their first eggs and to lay 50% of their eggs, and they had a lower respiration rate, than those females reared at 20°C with a 16-h photoperiod. Females that were reared in conditions that appeared to induce reproductive diapause resumed oviposition and their respiration rate increased soon after being transferred to a higher temperature (27.5°C). Thus, females reared at a 10-h photoperiod at 17.5 and 20°C appear to enter reproductive diapause