Dataset Information

RNA interference technology to control pest sea lampreys--a proof-of-concept.

ABSTRACT: The parasitic sea lamprey (Petromyzon marinus) has caused extensive losses to commercial fish stocks of the upper Great Lakes of North America. Methods of controlling the sea lamprey include trapping, barriers to prevent migration, and use of a chemical lampricide (3-trifluoromethyl-4-nitrophenol) to kill the filter-feeding larvae. Concerns about the non-specificity of these methods have prompted continued development of species-specific methods to control lampreys outside their native range. In this study, we considered the utility of RNA interference to develop a sea lamprey-specific lampricide. Injection of six different short interfering, double-stranded RNAs (siRNAs) into lamprey embryos first confirmed that the siRNAs could reduce the targeted transcript levels by more than 50%. Two size classes of lamprey larvae were then fed the siRNAs complexed with liposomes, and three of the siRNAs (targeting elongation factor 1?, calmodulin, and ?-actinin) reduced transcript levels 2.5, 3.6, and 5.0-fold, respectively, within the lamprey midsections. This is not only the first demonstration of RNAi in lampreys, but it is also the first example of delivery of siRNAs to a non-mammalian vertebrate through feeding formulations. One of the siRNA treatments also caused increased mortality of the larvae following a single feeding of siRNAs, which suggests that prolonged or multiple feedings of siRNAs could be used to kill filter-feeding larvae within streams, following development of a slow-release formulation. The genes targeted in this study are highly conserved across many species, and only serve as a proof-of-concept demonstration that siRNAs can be used in lampreys. Given that RNA interference is a sequence-specific phenomenon, it should be possible to design siRNAs that selectively target gene sequences that are unique to sea lampreys, and thus develop a technology to control these pests without adversely affecting non-target species.

SUBMITTER: Heath G

PROVIDER: S-EPMC3914985 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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RNA interference technology to control pest sea lampreys--a proof-of-concept.

Heath George G Childs Darcy D Docker Margaret F MF McCauley David W DW Whyard Steven S

PloS one 20140205 2

The parasitic sea lamprey (Petromyzon marinus) has caused extensive losses to commercial fish stocks of the upper Great Lakes of North America. Methods of controlling the sea lamprey include trapping, barriers to prevent migration, and use of a chemical lampricide (3-trifluoromethyl-4-nitrophenol) to kill the filter-feeding larvae. Concerns about the non-specificity of these methods have prompted continued development of species-specific methods to control lampreys outside their native range. In ...[more]

PMID: 24505485

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Project description:IntroductionTight glycemic control (TGC) has shown benefits but has been difficult to achieve consistently. STAR (Stochastic TARgeted) is a flexible, model-based TGC approach directly accounting for intra- and inter- patient variability with a stochastically derived maximum 5% risk of blood glucose (BG) < 4.0 mmol/L. This research assesses the safety, efficacy, and clinical burden of a STAR TGC controller modulating both insulin and nutrition inputs in pilot trials.MethodsSeven patients covering 660 hours. Insulin and nutrition interventions are given 1-3 hourly as chosen by the nurse to allow them to manage workload. Interventions are calculated by using clinically validated computer models of human metabolism and its variability in critical illness to maximize the overlap of the model-predicted (5-95th percentile) range of BG outcomes with the 4.0-6.5 mmol/L band while ensuring a maximum 5% risk of BG < 4.0 mmol/L. Carbohydrate intake (all sources) was selected to maximize intake up to 100% of SCCM/ACCP goal (25 kg/kcal/h). Maximum insulin doses and dose changes were limited for safety. Measurements were made with glucometers. Results are compared to those for the SPRINT study, which reduced mortality 25-40% for length of stay ≥3 days. Written informed consent was obtained for all patients, and approval was granted by the NZ Upper South A Regional Ethics Committee.ResultsA total of 402 measurements were taken over 660 hours (~14/day), because nurses showed a preference for 2-hourly measurements. Median [interquartile range, (IQR)] cohort BG was 5.9 mmol/L [5.2-6.8]. Overall, 63.2%, 75.9%, and 89.8% of measurements were in the 4.0-6.5, 4.0-7.0, and 4.0-8.0 mmol/L bands. There were no hypoglycemic events (BG < 2.2 mmol/L), and the minimum BG was 3.5 mmol/L with 4.5% < 4.4 mmol/L. Per patient, the median [IQR] hours of TGC was 92 h [29-113] using 53 [19-62] measurements (median, ~13/day). Median [IQR] results: BG, 5.9 mmol/L [5.8-6.3]; carbohydrate nutrition, 6.8 g/h [5.5-8.7] (~70% goal feed median); insulin, 2.5 U/h [0.1-5.1]. All patients achieved BG < 6.1 mmol/L. These results match or exceed SPRINT and clinical workload is reduced more than 20%.ConclusionsSTAR TGC modulating insulin and nutrition inputs provided very tight control with minimal variability by managing intra- and inter- patient variability. Performance and safety exceed that of SPRINT, which reduced mortality and cost in the Christchurch ICU. The use of glucometers did not appear to impact the quality of TGC. Finally, clinical workload was self-managed and reduced 20% compared with SPRINT.

Project description:RNA interference (RNAi) is a convenient tool to identify and characterize biological functions in organisms. Recently, it has become an alternative to chemical insecticides as a biologically based control agent. This promising technology has the potential to avoid many problems associated with conventional chemical insecticides. In order for RNAi application to be practical for field use, a major hurdle is the development of a cost-effective system of double-stranded RNA (dsRNA) production for a large quantity of dsRNA. A handful of research reports has demonstrated microbial-based dsRNA production using L4440 vector and HT115 (DE3) Escherichia coli for application to vertebrate and invertebrate systems. However, the dsRNA yield, production efficiency, and biological purity from this in vitro system is still unclear. Thus, our study detailed biochemical and molecular tools for large-scale dsRNA production using the microbial system and investigated the production efficiency and yield of crude and purified dsRNAs. An unrelated insect gene, green fluorescent protein (GFP), and an insect neuropeptide gene, pyrokinin (PK) identified from Drosophila suzukii, were used to construct the recombinant L4440 to be expressed in the HT115 (DE3) cell. A considerable amount of dsRNA, 19.5 µg/mL of liquid culture, was isolated using ultrasonic disruption followed by phenol extraction. The sonication method was further evaluated to extract crude dsRNA without the additional phenol extraction and nuclease treatments and also to reduce potential bacterial viability. The results suggest that the ultrasonic method saved time and costs to isolate crude dsRNA directly from large volumes of cell culture without E coli contamination. We investigated whether the injection of PK dsRNA into flies resulted in increased adult mortality, but it was not statistically significant at 95% confidence level. In this study, the microbial-based dsRNA production has potential for applied RNAi technology to complement current insect pest management practices.

Dataset Information

RNA interference technology to control pest sea lampreys--a proof-of-concept.

Publications

RNA interference technology to control pest sea lampreys--a proof-of-concept.

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