EvaluationEvaluation ofof thethe effectivenesseffectiveness ofof fumagillinfumagillin andand alternativealternative therapiestherapies forfor thethe controlcontrol ofof NosemaNosema ceranaeceranae
Stephen F. Pernal, Abdullah Ibrahim and Adony Melathopoulos AAFC Beaverlodge Research Farm Beaverlodge, Alberta, Canada OverwinteringOverwintering MortalityMortality inin CanadaCanada 20092009 Wintering Losses Province # Colonies Dead (% Provincial Total) British Columbia 8,778 24 Alberta 98,700 44 Saskatchewan 24,130 25 Manitoba 24,000 30 Ontario 24,800 31 Quebec 10,800 32 Nova Scotia 5,694 29 New Brunswick 4,455 43 PEI 1,620 40 34% CANADA 202,977 (of National Total) NosemaNosema –– microsporidiamicrosporidia Highly specialized parasitic fungi
Nosema Reduces:
Nursing
Lifespan
Pollen collection
Queen fertility
Queen acceptance
Leads to wintering mortality, poor spring build-up and loss of honey production. WhatWhat isis NosemaNosema ceranae?ceranae? • Isolated from Apis cerana in China in 1996.
• Reported from Apis mellifera in Taiwan 2005 and later confirmed in Spain. Later confirmations from Germany and France.
• In Spain, N. ceranae characterized by gradual depopulation, higher fall/winter mortality, low honey production but NOT dysentery or crawling bees typically seen with N. apis.
• Above symtoms similar to “dry nosemosis” seen in France.
• Widepsread in the U.S. and present in all Canadian Provinces. N. ceranae
N. ceranae N. apis
N. apis
Fries et al. 2006 Seasonal shift in Nosema incidence
Martín-Hernández et al. 2007 Nosema apis A problem in the winter and spring
3.0 18.5 x x 37.6 15 May (Ottawa, 69 Furgala) 21 April (Beaverlodge, 87 Szabo)
2.5
2.0 1.6 x 1.5 12 April (Langley, 87 Wyborn) (millions) spores / bee 1.0
0.5
0.0 Jan Feb Mar Apr May Jun Jul Aug Sep Oct Nov Dec
Fries from Bailey and Ball What is unknown about N. ceranae?
• Interactions / competition with N. apis.
• Transmission
• Survival / Infectivity of spores.
• Temporal phenology of symptoms.
• Most effective methods of management.
• Use of current or new antibiotics. Objectives:Objectives:
• To evaluate methods for disinfecting N. ceranae spores on comb.
• To evaluate different formulations of spring- applied fumagillin treatments for suppressing N. ceranae infections in overwintered colonies. CombComb DisinfectionDisinfection ExperimentExperiment
• Treatments: –Inoculated, Irradiated – Inoculated, Acetic Acid Fumigated – Inoculated, Heat Treated – Inoculated, Untreated – Non-Inoculated, Untreated
12 Replicate colonies / treatment CombComb DisinfectionDisinfection ExperimentExperiment Inoculating Comb
4.51 × 108 N. ceranae spores per colony CombComb DisinfectionDisinfection ExperimentExperiment Electron Beam Irradiation
FWT-602P Radiachromic film dosimeters CombComb DisinfectionDisinfection ExperimentExperiment Electron Beam Irradiation
10 kGy exposure, on tops and bottoms of boxes. CombComb DisinfectionDisinfection ExperimentExperiment Acetic acid Fumigation
Peter Jessing’s Yard 22-29 April 2009
4.52 × 1.78 × 2.42 m high Comb Disinfection Experiment Acetic acid Fumigation
480 mL 80% acetic acid per four brood chambers
Joints between boxes sealed. Empty box placed on top of stack. Comb Disinfection Experiment Acetic acid Fumigation
200 - 380 ppm; 7 day exposure at 30° C CombComb DisinfectionDisinfection ExperimentExperiment Heat Treatment
49 ± 1° C for 24 h
Melting Point of Beeswax: 62-65° C 2 May 2009 CombComb DisinfectionDisinfection ExperimentExperiment Sampling Bees for Spores
Weekly samples of 100 foragers (2 May - 4 June), then bi-weekly until end of October.
Spores enumerated microscopically at 400x. Colony assessments 3 July, 31 Aug. Comb Disinfection Experiment Hived with New Zealand Packages: 2 May 2009
5 Acetic Acid Fumigation Heat Irradiation
) Inoculated (No Disinfection)
6 4 * Non-Inoculated a 3 * a * Inoculated ab a ab Ht, N-Inoc ab * 2 a Inoculated b Fum, Irrad ab Ht, Fum, N-Inoc b Irradiation bc
Spores (xSpores / Bee 10 1 c b 0 09 09 09 09 09 09 09 y- y- y- n- l- l- g- a a a u Ju Ju u -M -M -M -J 6- 0- -A 02 15 21 16 1 3 12 Fumagillin Efficacy Experiment N. ceranae-infected overwintered singles: 22 April 2009
Pre-experimental infection level: 4.3 ± 0.5 x 106 spores per bee FumagillinFumagillin EfficacyEfficacy ExperimentExperiment Treatments (All containing 50 mg a.i. fumagillin and applied 2×):
– Sucrose Syrup (2 L) – Sucrose Syrup (250 mL - Drench) – Icing Sugar (20 g) – Pollen Patties (100 g) – Sucrose Syrup (2 L, unmedicated)
– 12 Replicate colonies / treatment
Fumagillin dicyclohexylammonium FumagillinFumagillin EfficacyEfficacy ExperimentExperiment Treatment Formulations – Spring 2009
DUSTING PATTY Icing Sugar Dust Pollen, Soy Flour, Syrup
SYRUP 1:1 Sucrose Drench vs. Full Colonies assessed Volume 30 June, 26 Aug 09. FumagillinFumagillin EfficacyEfficacy ExperimentExperiment SamplingSampling ResiduesResidues
15 g honey samples
Development of LC-MS/MS technique to determine residue levels of Fumagillin and degradation products. Fumagillin Efficacy Experiment Treatments applied 22April, 6 May 2009; Total a.i. = 100 mg
8 Drench Dust 7 Patty
) Syrup 6 6 Untreated 5 a a 4 a 3 a
2 *a Untreated
Spores / Spores (xBee 10 b ab Patty, Syrup b b Dust, Drench 1 Patty, Syrup ab b Drench, Dust b 0 9 9 9 9 9 9 9 9 9 9 -0 -0 -0 -0 -0 -0 -0 l-0 l-0 -0 pr pr y y y y n u u g A A a a a a Ju -J -J u 2- 9- -M -M -M -M - 5 9 -A 2 2 06 13 20 27 03 1 2 11 FumagillinFumagillin EfficacyEfficacy ExperimentExperiment Honey Production
80 70 60 50 40 30 20 Honey Yield (kg) Honey Yield 10 0 h ty d nc Dust e Pat Syrup eate Dr r Unt ConclusionsConclusions • Irradiation is the most effective method of disinfecting comb contaminated with N. ceranae.
• Applications of 100 mg a.i. fumagillin are effective at suppressing active infections of N. ceranae. over spring and summer months.
• In northern Alberta, N. ceranae spore levels appear to naturally decline during mid-summer, similar to patterns historically seen for N. apis. Acknowledgements
• Medivet Pharmaceuticals • Alberta Beekeepers Commission • Canadian Bee Research Fund • Bee Maid Honey • Matching Investment Initiative (AAFC) • Iotron Industries Canada ltd.