Monitoring Bt Susceptibilities in Helicoverpa zea and Heliothis

virescens: Results of 2006 Studies

Ibrahim Ali and R. G. LuttrellUniversity of Arkansas, Fayetteville, AR

Craig A . AbelUSDA-ARS, SIMRU, Stoneville, MS

Collected and reared ~ 400 heliothine colonies.

~200 colonies assayed with Cry1Ac, Cry2Ab and Vip3A insecticidal proteins.

Developed benchmarks for Bt resistance monitoring on cotton.

Previous Studies (2002-2005)

Ali et al. 2003 to 2006 Beltwide Cotton Proc.Ali et al. 2006, J. Econ. Entomol 99: 164-175

2006 Studies: Colony Information

0

20

40

60

80

AR NC GA AL MS LA TX

# C

olle

cte

d

0

5

10

15

20

AR NC GA AL MS LA TX

Cou

nty/

Stat

e

111 colonies, Larvae = 93 , Eggs= 18

2006 Studies: Colony Information

0

10

20

30

40

Co

lon

y (

#)

0

5

10

15

20

25

30

April May June July August Sept.

Colo

ny (

%)

Bioassays

Species Cry1Ac Cry2Ab

H. zea41 colonies

16, 000 larvae0 to 250 ug/ml

33 colonies9,000 larvae0 to 150 ug/ml

H. virescens8 colonies

4,000 larvae0 to 57 ug/ml

7 colonies>4,000 larvae0 to 20 ug/ml

All assays were diet-incorporated, 3-8 rep/dose, 48-128 larvae/rep

Bioassays

Mortality (Larvae those died at 7 days post-treatment).Stunting (Larvae failed to molt to 2nd instars).

Regressions for LC50 (Mortality) and MIC50 (Mortality + stunting) were estimated.

Dose-response of H. zea Exposed to Cry1Ac

For Cry1Ac, 32 field populations had significantly lower mortality than 99.9% LCL for UA susceptible laboratory colony at the highest dose (250 ug/ml).

0

20

40

60

80

100

0 50 100 150 200 250 300

% M

ort

alit

y

H. zea colonies exposed to doses (ug/ml) of Cry1Ac

UALabZA

Dose-response of H. zea Exposed to Cry1Ac

1

10

100

1000

10000

0 20 40 60

Colonies (#) exposed to Cry1Ac

LC

50 (

ug

/ml)

LC50s for 28 field populations > susceptible laboratory colony.LC50s for 8 field populations > 250 ug/ml.

MIC50s for 8 field populations > susceptible laboratory colony, and MIC50s for field populations varied over 100-fold.

UALAbZA

r = 0.463

0

1000

2000

3000

4000

20 40 60 80 100% moratlity of H. zea at 250 ug/ml of Cry1Ac

LC

50 (

ug

/ml)

Correlation of mortalities and regression estimates for H zea exposed to Cry1Ac

LC50 estimates of H. zea were significantly correlated with mortalities at highest tested dose of Cry1Ac (r=0.463, N= 42, P= 0.0174).

The correlation coefficient for MIC50s and mortality +stunting was 0.596 (N= 42, P < 0.0001).

Dose-response of H. zea Exposed to Cry2Ab

0

20

40

60

80

100

0 50 100 150 200

H. zea colonies exposed to doses (ug/ml) of Cry2Ab

%

Mo

rtali

ty

For Cry2Ab, 26 populations had significantly lower mortality than that of susceptible laboratory colony at the highest tested dose of Cry2Ab (150 ug/ml).

UALabZA

Dose-response of H. zea Exposed to Cry2Ab

LC50s for 20 field populations > susceptible laboratory colony.LC50s for 6 field populations > 150 ug/ml.

MIC50s for 18 field populations > susceptible laboratory colony, and MIC50s for field populations varied 69-fold.

1

10

100

1000

10000

0 10 20 30 40

Colonies (#) exposed to Cry2Ab

LC

50 (

ug

/ml)

UALabZA

r = 0.705

-500

0

500

1000

1500

2000

2500

20 40 60 80 100

% mortality of H. zea at 150 ug/ml of Cry2Ab

LC

50 (

ug

/ml)

Correlation of mortalities and regression estimates for H zea exposed to Cry2Ab

LC50 estimates of H. zea were significantly correlated with mortalities at highest tested dose of Cry2Ab (r=0.705, N= 31, P <0001).

The correlation coefficient for MIC50s and mortality +stunting was 0.697 (N= 31, P < 0.0001).

Dose-response of H. virescens Exposed to Cry1Ac

0

1

2

3

4

5

6

0 2 4 6 8

Colonies (#) exposed to Cry1Ac

LC

50 (

ug

/ml)

LC50s for three field populations > UALabVR colony and varied 7-fold. MIC50s varied only two-fold across all colonies.

UALabVR

Dose-response of H. virescens Exposed to Cry2Ab

Susceptibilities of all H. virescens populations exposed to Cry2Ab varied 2- to 3- fold.

0

1

2

3

4

0 2 4 6 8

Colonies (#) exposed to Cry2Ab

LC

50 (

ug

/ml)

UALabVR

LC50s for H zea field populations exposed to Cry1Ac in assays against different generations

in laboratory culture

LC50s of populations exposed to Cry1Ac tended to decrease in laboratory culture.

a

a

b

1

10

100

1000

10000

F0406 F0506 F2606 F7707 F7906 F8006 F8106 F8606

Colony

LC

50 (

ug

/ml)

First Second Third

LC50s for H zea field populations exposed to Cry2Ab in assays against different generations

in laboratory culture

LC50s of field populations exposed to Cry2Ab decreased in laboratory culture.

1

10

100

1000

10000

F7706 F8006 F8606

Colony

LC50

(ug/

ml)

First Second

Conclusion

Susceptibility of field colonies of H. zea to Cry1Ac and Cry2Ab varied significantly compared to the UA susceptible laboratory H. zea colony.

Considerable variability exists in H. zea field colonies exposed to Cry1Ac and Cry2Ab.

Susceptibility of field colonies to Cry1Ac was reduced through laboratory colonization.

LC50 and MIC50 estimates for H. zea for Cry1Ac and Cry2Ab were significantly correlated with mortalities and mortalities + stunting.

Susceptibility of field colonies of H. virescens to Cry1Ac and Cry2Ab varied 2- to 3-fold compared to the UA susceptible laboratory H. virescens colony.

Acknowledgement

This research was supported by a cooperative agreement with the USDA-ARS Southern Insect Management Research Unit, Stoneville, MS.

Supports from Monsanto Company and U.S. EPA are acknowledged.

We thank students and employees of the University of Arkansas for their assistance with field collections, rearing of insects and assays.

AcknowledgementWe like to thank and acknowledge the following research collaborators for providing insects for these studies:

J. R. Bradley, Astrid Groot, North Carolina State University, Raleigh, NC.

Greg Payne, State University of West Georgia, Carrollton, GA.

Bill Moar, Auburn University, AL.

Carlos Blanco, USDA-ARS, SIMRU, Stoneville, MS.

Roger Leonard, Louisiana State University, Winnsboro, LA.