Synergistic and Sublethal Effects

of Pesticides on Honey Bees

Pesticide/Pollinator Symposium

Alfred, NY

Roger Simonds and

staff

USDA-AMS National

Science Laboratory

Pesticide Database

Program (PDP)

PSU; Pesticide

Investigating GroupPSU; Cox-Foster Lab

CCD working team

Exposure to Pesticides

Samples Analyzed 2007-’08

• > 900 samples

• From

– CCD study

– Apple orchard study

– Migratory study

– Beekeeper submitted samples

• Cost-sharing program

Honey Bees - Excellent Indicator

of Environmental PesticidesAcephate

Acetamiprid

Aldicarb sulfone

Aldicarb sulfoxide

Allethrin

Amicarbazone

Amitraz (2,4-DMA)

Amitraz (2,4-DMPF)

Atrazine

Azinophos-methyl

Azoxystrobin

Bendiocarb

Bifenthrin

Boscalid

Captan

Carbaryl

Carbary l (1-Naphthol)

Carbendazim

Carbofuran , 3-hydroxy

Carfentrazone

Chlorfenapyr

Chlorothalonil

Chlorpyrifos

Coumaphos

Coumaphos -Chlorferone

Coumaphos oxon

Cyfluthrin

Cyhalothr in (total)

Cype rmethrin

Cyprodi nil

Deltamethrin

Diazinon

Dicofol

Difenoconazole

Diflubenzuron

Dimethomorph

Diphenylamine

Endos ulfan 1

Endos ulfan 2

Endos ulfan sulfate

Esfenval erate

Etoxazole

Fenbuconazole

Fenhexamid

Fenpropath rin

Fluva linate

Heptac hlor

Hexachlorobenzene

Imidacloprid

Imidacloprid olefin

Imidacloprid, 5-hydroxy

Indoxacarb

Malathion

Methidathion

Methoxyfeno zide

Methyl par athion

Metolachlor

Metribuzin

Myclobuta nil

Norflurazon

Oxamyl

Oxyfluorfen

p,p' -DDE

Pendi methalin

Permethrin

Phosmet

Pyraclostrobin

Pyrethrins

Pyrimethanil

Quintozene (PCNB)

Sethoxydim

Simazine

Tebuconazole

Tebufenozide

Tebuthiuron

Tetramethrin

Thiabendazole

Thiacloprid

Tribufos

Trifloxystrobin

Trifluralin

Vinclozolin

Multiresidue Pesticide Analysis on US Wax,

Pollen and Bees Samples 2007-08

• Very few samples lacked detections

• Found 121 different pesticides and metabolites

pyrethroids, organophosphates, carbamates, neonicotinoids

insect growth regulators, organochlorines, chlorinated cyclodienes,

20 fungicides, 12 herbicides, 2 acaricides, 1 synergist,

At least 14 of these are systemic pesticides

• On average 6 different pesticides per pollen sample

- Up to 31 in a single sample

Pesticide or

Metabolite Class LOD

Samples

Analyzed

Total

Detections

% of

Samples

Mean

(ppb)

SEM

(ppb)

Lowest

Value

Highest

Value

LD50

(ppb)

Carbaryl

PS

CARB 5 836 44 5.3 170.2 9.9 4.5 1010.0 10500

Coumaphos OP 1 887 668 75.3 1454.6 174.8 1.0 91900.0 46300

Fluvalina te PYR 1 887 749 84.4 2947.6 330.7 1.1 204000.0 15860

Imidacloprid NEO 2 836 14 1.7 77.5 5.9 2.4 646.0 280

Imidacloprid

olefin NEO 25 836 2 0.2 597.0 2.1 554.0 640.0 280

Imidacloprid,

5-hydroxy NEO 25 836 2 0.2 151.0 0.0 150.0 152.0 280

Chlorpyrifos OP 0.1 886 396 44.7 40.1 4.0 0.1 890.0 1220

Chlorothalonil FUNG 1 797 316 39.6 1860.4 286.3 1.0 98900.0 1110000

Selected Pesticides fond in WAX, POLLEN and BEES

Pesticide or

Metabolite Class LOD

Samples

Analyzed

Total

Detections

% of

Samples

Mean

(ppb)

SEM

(ppb)

Lowest

Value

Highest

Value

LD50

(ppb)

Carbaryl

PS

CARB 5 836 44 5.3 170.2 9.9 4.5 1010.0 10500

Coumaphos OP 1 887 668 75.3 1454.6 174.8 1.0 91900.0 46300

Fluvalina te PYR 1 887 749 84.4 2947.6 330.7 1.1 204000.0 15860

Imidacloprid NEO 2 836 14 1.7 77.5 5.9 2.4 646.0 280

Imidacloprid

olefin NEO 25 836 2 0.2 597.0 2.1 554.0 640.0 280

Imidacloprid,

5-hydroxy NEO 25 836 2 0.2 151.0 0.0 150.0 152.0 280

Chlorpyrifos OP 0.1 886 396 44.7 40.1 4.0 0.1 890.0 1220

Chlorothalonil FUNG 1 797 316 39.6 1860.4 286.3 1.0 98900.0 1110000

Selected Pesticides fond in WAX, POLLEN and BEES

Pesticide or

Metabolite Class LOD

Samples

Analyzed

Total

Detections

% of

Samples

Mean

(ppb)

SEM

(ppb)

Lowest

Value

Highest

Value

LD50

(ppb)

Carbaryl

PS

CARB 5 836 44 5.3 170.2 9.9 4.5 1010.0 10500

Coumaphos OP 1 887 668 75.3 1454.6 174.8 1.0 91900.0 46300

Fluvalina te PYR 1 887 749 84.4 2947.6 330.7 1.1 204000.0 15860

Imidacloprid NEO 2 836 14 1.7 77.5 5.9 2.4 646.0 280

Imidacloprid

olefin NEO 25 836 2 0.2 597.0 2.1 554.0 640.0 280

Imidacloprid,

5-hydroxy NEO 25 836 2 0.2 151.0 0.0 150.0 152.0 280

Chlorpyrifos OP 0.1 886 396 44.7 40.1 4.0 0.1 890.0 1220

Chlorothalonil FUNG 1 797 316 39.6 1860.4 286.3 1.0 98900.0 1110000

Selected Pesticides fond in WAX, POLLEN and BEES

Pesticide or

Metabolite Class LOD

Samples

Analyzed

Total

Detections

% of

Samples

Mean

(ppb)

SEM

(ppb)

Lowest

Value

Highest

Value

LD50

(ppb)

Carbaryl

PS

CARB 5 836 44 5.3 170.2 9.9 4.5 1010.0 10500

Coumaphos OP 1 887 668 75.3 1454.6 174.8 1.0 91900.0 46300

Fluvalina te PYR 1 887 749 84.4 2947.6 330.7 1.1 204000.0 15860

Imidacloprid NEO 2 836 14 1.7 77.5 5.9 2.4 646.0 280

Imidacloprid

olefin NEO 25 836 2 0.2 597.0 2.1 554.0 640.0 280

Imidacloprid,

5-hydroxy NEO 25 836 2 0.2 151.0 0.0 150.0 152.0 280

Chlorpyrifos OP 0.1 886 396 44.7 40.1 4.0 0.1 890.0 1220

Chlorothalonil FUNG 1 797 316 39.6 1860.4 286.3 1.0 98900.0 1110000

Selected Pesticides fond in WAX, POLLEN and BEES

Pesticide or

Metabolite Class LOD

Samples

Analyzed

Total

Detections

% of

Samples

Mean

(ppb)

SEM

(ppb)

Lowest

Value

Highest

Value

LD50

(ppb)

Carbaryl

PS

CARB 5 836 44 5.3 170.2 9.9 4.5 1010.0 10500

Coumaphos OP 1 887 668 75.3 1454.6 174.8 1.0 91900.0 46300

Fluvalina te PYR 1 887 749 84.4 2947.6 330.7 1.1 204000.0 15860

Imidacloprid NEO 2 836 14 1.7 77.5 5.9 2.4 646.0 280

Imidacloprid

olefin NEO 25 836 2 0.2 597.0 2.1 554.0 640.0 280

Imidacloprid,

5-hydroxy NEO 25 836 2 0.2 151.0 0.0 150.0 152.0 280

Chlorpyrifos OP 0.1 886 396 44.7 40.1 4.0 0.1 890.0 1220

Chlorothalonil FUNG 1 797 316 39.6 1860.4 286.3 1.0 98900.0 1110000

Selected Pesticides fond in WAX, POLLEN and BEES

Outcomes of Honey Bee Pesticide Analysis

• No “commodity” has had as many detections at such high amounts in so few samples over

such a short time as has bee pollen

• Highest detections were in-house miticides, fluvalinate and coumaphos, but well over 100

other pesticides and metabolites found

• Pyrethroids dominate: known to impact foraging behavior,

• No individual chemical is likely to explain CCD

• Systemic or other fungicides occur at levels that may synergize with pyrethroids,

organophosphates or neonicotinoids.

• Role of pesticides and diseases like IAPV in CCD remains to be reconstituted in lab

bioassays at relevant doses

• Impacts of multiple pesticide residues in bee food most likely will be via synergistic

interactions at sublethal levels on key behaviors/physiology

Key Concerns

• Exposure (kinds, amounts, combinations)

• Systemic pesticides

• Fungicides

• Adjuvents/synergists

• Metabolites

• Labeling - lack of adequate warning

Herbicides • Reduce forage

• Reduce nesting habitat

Insecticides• Lethal/Toxic

– Depending on Class/Mode of Action

– Toxic effect (multiple pesticides) (Sara Ashcraft)

• Sublethal– Behavioral effects

• Learning and retention (Daniel Schmehl)

• Foraging Behavior

– Physiological effects• Immune system function (Diana Cox-Foster)

• Longevity

- Larval development - adult behavior and physiology

• Acute vs Chronic Toxicity

• similar (additive) - combining effects (same or similar target site)

• independent - no interaction (unrelated sites of action)

• synergistic - one chemical increases the biological activity of the

other (> sum of each effect)

• antagonistic - one chemical decreases the biological activity of

the other (< sum of each)

Interactions (joint action) -

Adult Bee Toxicity Bioassay

5 day bioassays

Oral ingestion in sugar

syrup 24 hrs

Assail-Nova Probit at 24 Hours

0

2

4

6

8

10

0 1 2 3

Dose*

Pro

bit

Un

its

Assail

Nova

Assail/Nova

Assail 1: 10ppm

2: 100ppm

3: 200ppm

Nova 1: 50ppm

2: 500ppm

3: 1000ppm

Assail/Nova 1: 5/25ppm

2: 50/250ppm

3: 100/500ppm

Dose*:

0

10

20

30

40

50

60

70

80

90

100

1 2 3 4

Days Since Ingestion

Pe

rce

nt

Mo

rta

lity

treatment

Solvent Control

Delayed Mortality Feeding on Pesticide at 95% Pollen Level

0

10

20

30

40

50

60

70

80

90

100

1 2 3 4

Days Since Ingestion

Pe

rce

nt

Mo

rta

lity

Solvent Control

Fluvalinate

Coumaphos

Delayed Mortality Feeding on Pesticide at 95% Pollen Level

0

10

20

30

40

50

60

70

80

90

100

1 2 3 4

Days Since Ingestion

Pe

rce

nt

Mo

rta

lity

Solvent Control

Fluvalinate

Coumaphos

Chlorpyrifos

Delayed Mortality Feeding on Pesticide at 95% Pollen Level

0

10

20

30

40

50

60

70

80

90

100

1 2 3 4

Days Since Ingestion

Pe

rce

n M

orta

lity

Fluvalinate/Coumaphos

Delayed Mortality Feeding on Pesticide at 95% Pollen Level

0

10

20

30

40

50

60

70

80

90

100

1 2 3 4

Days Since Ingestion

Pe

rce

nt

Mo

rta

lity

Fluvalinate/Coumaphos Fluvalinate/Chlorothalonil

Delayed Mortality Feeding on Pesticide at 95% Pollen Level

0

10

20

30

40

50

60

70

80

90

100

1 2 3 4

Days Since Ingestion

Pe

rce

nt

Mo

rta

lity

Fluvalinate/Coumaphos Fluvalinate/Chlorothalonil Coumaphos/Chlorpyrifos

Delayed Mortality Feeding on Pesticide at 95% Pollen Level

0

10

20

30

40

50

60

70

80

90

100

1 2 3 4

Days Since Ingestion

Pe

rce

nt

Mo

rta

lity

Fluvalinate/Coumaphos Fluvalinate/Chlorothalonil

Coumaphos/Chlorpyrifos Fluvalinate/Coumaphos/Chlorpyrifos/Chlorothalonil

Delayed Mortality Feeding on Pesticide at 95% Pollen Level

UDSA/PDP

• Honey added as a commodity to PDP

• 2007-2008

• 744 samples

• Processed and bottled

– May mitigate residues

found in raw honey

Pesticide Name

# of

Samples

# of

Detects

% of

Detects

Min

Concen

Max

Concen

LOD

(PPB)EPA

Tolerance

Coumaphos 744 263 35.3 1 8.2 1 100

Fluvalinate 744 88 11.8 1 14.5 1 50

2,4-dimethylphenyl formamide (2,4-DMPF) 744 81 10.9 4.1 85.1 4 1000

Dicofol p,p' 744 35 4.7 1 90 1 NT

Piperonyl butoxide 744 10 1.3 6.3 9.6 6 NT

Vinclozolin 744 5 0.67 1.3 2.1 1 NT

Azoxystrobin 744 3 0.4 3.3 4 2 NT

Dicloran 744 3 0.4 1 1.6 1 NT

Carbendazim (MBC) 744 1 0.13 27.3 27.3 5 NT

Chlorpyrifos 744 1 0.13 1.3 1.3 1 NT

Methoxyfenozide 744 1 0.13 2.8 2.8 2 NT

Permethrin Total 744 1 0.13 11 11 10 NT

HONEY

USDA

Pesticide Data Base Program

• http://www.ams.usda.gov/AMSv1.0/ams.fetchTemplateDat

a.do?template=TemplateC&navID=ScienceandLaboratorie

s&leftNav=ScienceandLaboratories&page=PesticideDataP

rogram&description=Pesticide%20Data%20Program&acct

=pestcddataprg

http://www.nhb.org/

SUMMARY

• Honey (processed) for human

consumption contains few residues,

most below allowable EPA tolerance

• Pesticide residues in wax and

pollen in general are numerous and

some are at high levels, including

- cappings wax

- tapped pollen

• Beekeepers should consider

having hive products tested,

especially pollen and wax, if selling for human consumption

Recommendations• Monitor and control varroa mites using “soft” chemicals.

– formic acid (Mite-Away II®, Apiguard®, and Apilife var®.

• Reduce pathogen (and pesticide) build-up by regularly

culling old comb, recycling comb and/or irradiation of old

comb.

– Especially for dead-out colonies.

• Based on wide-spread resistance and these findings

fluvalinate is not recommended.

– Use of off-label product should NOT be considered.

Recommendations

• If coumphos must be used, only the registered product, CheckMite+® should be considered.

• Communicate with growers where bees are used for pollination to minimize colony exposure to agricultural-use pesticides.

• Monitor and control Nosema disease using fumagilin.

Pesticide Analysis

Cost-sharing Program

• Funding from PAm and Foundations for the Preservation of Honey Bees

• Grant pays half the cost of analysis ($134) and beekeepers pays half ($134)

• Results are confidential – Data added to our over all data base

• To participate:

MAAREC.psu.edu and/or email: Mfrazier@psu.edu

Date 9/17/08 9/17/08

Compound name 75030 89543

Pollen Pollen Average* Range*

2,4 Dimethylphenyl formamide (DMPF)ND 40 54 42 -66

Chlorothalonil ND 13.1 534.1 1.1 -4200

Chlorpyrifos ND 6.7 51.5 0.1 -830

Coumaphos 193 8.1 249.9 2.4 -5828

Cyfluthrin ND 6 4.5 4.5

Fenpropathrin 9 9.2

Fluvalinate 13.6 ND 139.6 2.4 -2670

Pendimethalin 11.656 ND 9.4 2.8 -16

FUNDING SOURCES National Honey Board

USDA

Haagen-Dazs

Wyman’s Growers

Penn State University College of Agricultural Sciences

PA Department of Agriculture

PAm

Foundation for the Preservation of Honey Bees

Florida Department of Agriculture

NC508 Sustainable Solutions to Problems Affecting Honey Bee Health

Beekeeping Associations (FL state and Tampa Bay)

MAAREC.psu.edu