Using Botanical Fumigants to Control Sciarid Flies. Richard Massy

Size: px

Start display at page:

Download "Using Botanical Fumigants to Control Sciarid Flies. Richard Massy"

Randell Pitts
5 years ago
Views:

1 Using Botanical Fumigants to Control Sciarid Flies Richard Massy

2 Background Experiment to investigate semiochemicals for sciarid larvae inadvertently identified potential fumigants -100% mortality from clove oil, spearmint oil and 1-octen-3-ol!

3 F: Sciaridae Dark winged fungus gnats Small (Bradysia adults 4mm) Fast life cycle (18-26 days) Adults lay up to 1000 eggs 9:1 F:M gender ratio 22.5mm Notable species: Lycoriella ingenua Bradysia difformis Pest of: Mushroom farms Nurseries, compost

4 Goal To develop a treatment of cheap and safe, botanically derived fumigants to treat growing media

5 Fumigation commences 2 nd December 2014 Multiple semiochemicals screened for fumigatory activity in 24h petri dish assays 8.5cm dish water saturated compost substrate Chemical added to filter paper on top of glass slide Ten 3 rd instar larvae added Pilot experiment sciarid count: 100

6 Various plant volatiles and essential oils screened 8.5cm petri dish Laboratory: 22 C Dose: 10µl = 0.14µl ml -1 N = 10, 3 replicates Treatment Mortality (%) Significance (p) compared to control 1-hexanol 93 < octen-3-ol 100 < hexanol 100 < Clove oil 7 1 (-) limonene 7 1 (±) linalool Peppermint oil Spearmint oil (-) trans caryophyllene 10 1 β-citronellol 7 1 β-myrcene 17 1 Control (water) 10 n = 360 (460)

7 Various plant volatiles and essential oils screened 8.5cm petri dish Laboratory: 22 C Dose: 10µl = 0.14µl ml -1 N = 10, 3 replicates Treatment Mortality (%) Significance (p) compared to control 1-hexanol 93 < octen-3-ol 100 < hexanol 100 < Clove oil 7 1 (-) limonene 7 1 (±) linalool Peppermint oil Spearmint oil (-) trans caryophyllene 10 1 β-citronellol 7 1 β-myrcene 17 1 Control (water) 10 n = 360 (460)

8 Successive assays reduce the list GLASSHOUSE C Mortality (%) p (compared to control) Control (water) 3 Peppermint oil 90 < Spearmint oil octen-3-ol 100 < hexanol hexanol 97 < (±) linalool 83 < cm petri dish Dose = 2µl, 0.23µl ml -1 : N = 10, 3 replicates LABORATORY 22 C Mortality (%) p (compared to control) Control (water) 3 Peppermint oil 100 < Spearmint oil 97 < octen-3-ol 100 < hexanol 93 < hexanol 100 < (±) linalool 100 < n = 210 * 2 (880)

Successive assays reduce the list GLASSHOUSE 10-20 C Mortality (%) p (compared to control) Control (water) 3 Peppermint oil 90 <0.0005 Spearmint oil 30 0.001 1-octen-3-ol 100 <0.0005 3-hexanol 50 0.

9 Successive assays reduce the list GLASSHOUSE C Mortality (%) p (compared to control) Control (water) 3 Peppermint oil 90 < Spearmint oil octen-3-ol 100 < hexanol hexanol 97 < (±) linalool 83 < cm petri dish Dose = 2µl, 0.23µl ml -1 : N = 10, 3 replicates LABORATORY 22 C Mortality (%) p (compared to control) Control (water) 3 Peppermint oil 100 < Spearmint oil 97 < octen-3-ol 100 < hexanol 93 < hexanol 100 < (±) linalool 100 < n = 210 * 2 (880)

10 More assays 3.4cm petri dish Dose = variable N = 10, 3 replicates GLASSHOUSE C Mort (%) p Mort (%) p Mort (%) Sig, p Dose 2µl 1µl 0.5µl Control (water) Peppermint oil 27 < Spearmint oil octen-3-ol 30 < < < hexanol 29 < (±) linalool 25 < n = 360 (1240)

11 More assays 3.4cm petri dish Dose = variable N = 10, 3 replicates GLASSHOUSE C Mort (%) p Mort (%) p Mort (%) Sig, p Dose 2µl 1µl 0.5µl Control (water) Peppermint oil 27 < Spearmint oil octen-3-ol 30 < < < hexanol 29 < (±) linalool 25 < n = 360 (1240)

12 Yet more assays Mort (%) p Mort (%) p Mort (%) p Mort (%) p Mort (%) Sig, p Dose 2µl 1.3µl 1µl 0.7µl 0.5µl Control (water) octen-3-ol 100 < < < < (±) linalool 83 < < n = 180 (1420)

13 Yet more assays Mort (%) p Mort (%) p Mort (%) p Mort (%) p Mort (%) Sig, p Dose 2µl 1.3µl 1µl 0.7µl 0.5µl Control (water) octen-3-ol 100 < < < < (±) linalool 83 < < octen-3-ol LD50 = 89 (57-111) 1-octen-3-ol LD90 = 157 ( ) Linalool LD50 = 150 ( ) Linalool LD90 = 248 ( ) [Unit = µl chemical per litre of airspace. Brackets = 95% confidence interval.] n = 180 (1420)

14 I ve done bioassays and statistics, but graphing is where I draw the line! Mortality (%) Control 1-octen-3-ol linalool Dose (µl L -1 )

15 A brief interlude: Report on toxicity pathways 1-octen-3-ol = dopamine inhibitor Drosophila study: Reduced dopamine levels, and reduced uptake of dopamine by dopamine neurones (Inamdar et al. 2013)

16 A brief interlude: Report on toxicity pathways Linalool = not fully understood Acetylcholine esterase inhibition(praveena & Sanjayan 2011). Blocking somatic sensory nerves in rats (Leal-Cardoso et al. 2010) Spontaneous activity of sensory nerves, leading to hyperstimulation of motor nerves causing knockdown. (unverified sources)

17 Study 2: emergence assays Emergence assays to test long term exposure Semi-field Doses reduced Fumigation compared to biocontrols (entomopathogenic fungus and nematodes)

18 Study 2: emergence assays Plastic pots 166ml of compost substrate Ten third instar larvae Experimental treatment mixed with 15ml 0.001% Tween 80 Decayed potato added for food

Biocontrol emergence assay Fungus = M. anisopliae 4556 and v275 Three concentrations of each fungus strain tested: 10 5, 10 6 and 10 7.

19 Biocontrol emergence assay Fungus = M. anisopliae 4556 and v275 Three concentrations of each fungus strain tested: 10 5, 10 6 and Nematodes = Steinernema feltiae Three concentrations of nematode tested: 10, 5 and 2.5 IJ* ml -1 (nematodes supplied by e-nema) *IJ = infective juveniles

20 Biocontrol results n = 300*3 (2320)

21 Fumigation emergence assay Cup total volume = 345ml 1-octen-3-ol and linalool tested Treatments of 60, 30 and 15µl per cup (174, 87 and 43µl L-1 of substrate/ airspace)

22 Fumigation E.A. results n = 210*4 (3160)

23 Fumigation E.A. results Chemical and dose Significance, p, compared to control 1-octen-3-ol 60µl < octen-3-ol 30µl < octen-3-ol 15µl Linalool 60µl Linalool 30µl Linalool 15µl 0.863

24 However Relative mammalian toxicities suggest linalool has a higher safe dose. Chemical Oral LD50, rat Dermal LD50, rabbit 1-octen-3-ol Linalool Unit = mg/ kg Extra info: 1-octen-3-ol vapour toxicity = 3.72 mg/ L (4 hours)

25 Fumigation emergence assay 2 Extra compounds identified from literature added to explore synergies with currently identified compounds. Compound Diethyl Maleate Eucalyptol Activity Blocks enzyme pathway (glutathione S- transferase) for detoxifying linalool (Davoudi et al. 2011). Fumigation activity against grain beetles (Lee et al. 2003), and toxicity to house flies (multiple) Activates cold receptors TRPM8 and TRPA1 (Gonzales 2014) Reduced mitochondrial membrane potential, inflammation and ROS in rat PC12 cells (Khan et al. 2014)

26 Fumigation E.A assay 2 results n = 210*3 (3790)

27 Fumigation E.A assay 2 results Chemical and dose Significance, p, compared to control Eucalyptol 60µl < Eucalyptol 30µl < Eucalyptol 15µl Diethyl maleate 60µl Diethyl maleate 30µl Diethyl maleate 15µl 0.644

28 Ongoing work Investigating chemical combinations, and more doses in emergence assays. Sub-lethal petri-dish assays to investigate molecular markers associated with stress and health (HSP70, ITS2 etc). Investigate the efficacy against other insect pests Try not to go mad Ongoing experiment n = 150*3 (4240)

29 Acknowledgements: Professor Tariq Butt - Head of Swansea University BANP group. Project leader and direction Dr Ian Bedford Head of Entomology, John Innes Centre. Supply of sciarid cultures. This project was funded by INBIOSOIL. An EU project for the control of soil pests.

30 References: Davoudi A., Shayesteh N., Shirdel D. and Hosseinzadeh A. (2011). Effect of diethyl maleate on toxicity of linalool against two stored product insects in laboratory condition. Africa J. Biotech. 10(48): Gonzales R. (2014). Painful, hot, and toxic secrets of TRP channels. Eukaryon. 10: Inamdar A., Hossain M., Bernstein A., Miller G., Richardson J. and Bennett J. (2013). Fungalderived semiochemical 1-octen-3-ol disrupts dopamine packaging and causes neurodegeneration. PNAS. 110(48) Khan A. et al. (2014). 1,8-cineole (Eucalyptol) mitigates inflammation in amyloid beta toxicated PC12 cells: relevance to Alzheimer s disease. Neurochem. Res. 39: Leal-Cardoso et al Linalool blocks excitability in peripheral nerves and voltage-dependent Na + current in dissociated dorsal root ganglia neurones. Euro. J. Pharma. 645: Lee B., Annis P. and Tumaalii F. (2003). The potential of 1,8-cineole as a fumigant for stored wheat. Proc. Australian Postharvest Tech. Conf, Canberra, June Praveena A., Sanjayan K. (2011). Inhibition of acetylcholinesterase in three insects of economic importance by Linalool, a monoterpene phytochemical. Insect Pest Manage. 2011:

Insect growth regulators in pest management programs. Frank Arthur USDA-ARS-GMPRC Manhattan, KS 66502

Insect growth regulators in pest management programs Frank Arthur USDA-ARS-GMPRC Manhattan, KS 66502 Reduced-risk Insecticides Seen as replacements for organophosphate and carbamate neurotoxins Examples: