1
Bollgard
and
Bollgard
II
Cotton
Bollworm
Resistance
Management
Issues
Sharlene
R.
Matten,
Ph.
D.,
Biologist
USEPA/
OPP
Biopesticides
and
Pollution
Prevention
Division
Matten.
sharlene@
epa.
gov
703­
605­
0514
Bollgard
and
Bollgard
II
Cotton
Bollworm
Resistance
Management
Issues
Sharlene
R.
Matten,
Ph.
D.,
Biologist
USEPA/
OPP
Biopesticides
and
Pollution
Prevention
Division
Matten.
sharlene@
epa.
gov
703­
605­
0514
2
Purpose

Highlight
EPA's
assessment
of
the
Helicoverpa
zea
(
cotton
bollworm,
CBW)
insect
resistance
management
(
IRM)
data
submitted
by
Monsanto
required
as
part
of
the
terms
and
conditions
of
the
Bollgard
and
Bollgard
II
registrations

Bollgard
expresses
moderate
dose
of
Cry1Ac
against
CBW

CBW
resistance
risk
considered
the
greatest

5%
external,
unsprayed,
structured
refuge
option
poses
greatest
concern;
option
expires
Sept.
30,
2004

Specific
data
requirements

North­
south
movement

Pyrethroid
oversprays

Alternative
hosts

Bollgard
and
Bollgard
II
registrations
expire
Sept.
2006
3
EPA
Decision

EPA
will
determine
whether
the
scientific
data
are
adequate
to
support
continuation
of
the
5%

external,
unsprayed,
structured
refuge
for
CBW
resistance
management
after
Sept.
30,
2004.
4
Outline

Current
IRM
refuge
requirements

Major
issues
for
SAP

North­
south
movement

Synthetic
pyrethroid
oversprays

Alternative
hosts
as
natural
refugia

Gustafson
et
al.
CBW
resistance
management
model

Overall
conclusion

SAP
questions
5
Current
IRM
Requirements

5%
external,
unsprayed
refuge:
>
150
ft.
wide
(>
300ft.
preferred),
<
1/
2
mile
(
1/
4
mile
or
adjacent
preferred),
expires
9/
30/
04

20%
external,
sprayed
refuge:
<
1
mile
(
1/
2
mile
or
closer
preferred)
7
Acres
7
Acres
50
Acres
50
Acres
10
Acres
10
Acres
20
Acres
20
Acres
20
Acres
20
Acres
20
Acres
20
Acres
45
Acres
45
Acres
5%
Untreated
Refuge
with
Distance
Requirement
5%
Untreated
Refuge
with
Distance
Requirement
75
Acres
75
Acres
Refuge
Refuge
Refuge
Refuge
30
Acres
30
Acres
50
Acres
50
Acres
<
0.5
Mile
<
0.5
Mile
<
0.5
Mile
<
0.5
Mile
<

0.5
Mile
<

0.5
Mile
<
0.5
Mile
<
0
.5
Mile
<
0.5
Mile
<
0.5
Mile
<
0.5
Mile
<
0.5
Mile
<
0.5
Mile
<
0.5
Mile
<

0.5
Mile
<

0.5
Mile
Road
Road
**********

***********

Refuge
Field
***********
***********
**********

***********

Refuge
Field
***********
***********
One
mile
or
less
On
e
mile
or
less
20%
Sprayed
Refuge
with
Distance
Requirement
20%
Sprayed
Refuge
with
Distance
Requirement
Bollgard
Bollgard
Bollgard
Bollgard
Bollgard
Bollgard
Bollgard
Bollgard
Bollgard
Bollgard
Bollgard
Bollgard
Bollgard
Bollgard
Bollgard
Bollgard
Bollgard
Bollgard
(
20%)
Sprayed
(
20%)
Sprayed
6
Current
IRM
Requirements

5%
embedded
refuge:
>
150
ft.

wide
(>
300ft.
preferred),
refuge
can
only
be
treated
if
entire
field
treated

5%
embedded
refuge
for
PBW:

embedded
refuge
must
be
at
least
1
non­
Bt
cotton
row
for
every
6­
10
rows
of
Bt
cotton.


Community
refuge:
use
either
5%

external,
unsprayed
and/
or
20%

external,
sprayed
refuge
A=
20
A=
20
5%
Embedded
Refuge
(
Field
Unit)

5%
Embedded
Refuge
(
Field
Unit)

One
Mile
One
Mile
One
Mile
One
Mile
Field
Road
Field
Road
50
Acres
Bollgard
50
Acres
Bollgard
100
Acres
Bollgard
100
Acres
Bollgard
50
Acres
Bollgard
50
Acres
Bollgard
40
Acres
Bollgard
40
Acres
Bollgard
40
Acres
Bollgard
40
Acres
Bollgard
100
Acres
Bollgard
100
Acres
Bollgard
10
Acre
Refuge
10
Acre
Refuge
10
Acre
Refuge
10
Acre
Refuge
400
Total
Acres
5%
=
20
Total
Refuge
Acres
Needed
400
Total
Acres
5%
=
20
Total
Refuge
Acres
Needed
A=
7
A=
7
B=
50
B=
50
C=
10
C=
10
A=
20
A=
20
A=
20
A=
20
A=
20
A=
20
B=
45
B=
45
Community
Refuge
Plan
­
20%
Sprayed
Community
Refuge
Plan
­
20%
Sprayed
Grower
A
=
117A
Grower
A
=
117A
Grower
B
=
125A
Grower
B
=
125A
Grower
C
=
85A
Grower
C
=
85A
327A
327A
Refuge
=
20%

Refuge
=
20%

65.4
Acres
Needed
65.4
Acres
Needed
C=
75
C=
75
B=
30
B=
30
A=
50
A=
50
Road
Road
20%
Sprayed
Refuge
20%
Sprayed
Refuge
**********

***********
***********
**********

***********
***********

********************
One
Mile
7
Bollgard
and
Bollgard
II
CBW
IRM
Issues:
Focal
Points
for
SAP

North­
south
movement.
Evaluate
reverse
migration
impact
on
CBW
adaptation
to
Bt
crops.


Pyrethroid
oversprays.
Evaluate
impact
of
pyrethroid
oversprays
on
CBW
survivors
in
Bollgard
fields
and
the
effect
on
resistance
management.


Alternative
hosts.
Evaluate
the
effectiveness
of
alternative
hosts
as
natural
refuges
in
comparison
to
the
5%
external,
unsprayed
structured
refuge

Gustafson
et
al.
model.
Evaluate
impact
of
pyrethroid
oversprays
and
alternative
hosts
on
resistance
evolution.
8
North­
South
(
N­
S)
Movement

CEW/
CBW
adults
move
from
corn
to
cotton
over
the
course
of
the
season.


Feed
on
both
Bt
corn
and
Bt
cotton

May
undergo
selection
for
adaptation
to
Bt
toxins
for
several
generations
each
year.
9
N­
S
Movement

Previously
thought
there
was
only
"
one­
way"
CBW
migration

Gould
et
al.
2002

Indirect
evidence
for
reverse
migration
(
Bossier
Parish,
LA
and
College
Station,
TX)
 
50­
60%

CBW
migrating
from
the
North

Timing
and
extent
of
reverse
migration
appears
to
vary
considerably
from
year
to
year

Postulated
high
Bt
corn
deployment
could
impact
CBW
resistance
evolution
to
Bt
cotton
10
N­
S
Movement
­
Modeling

Storer
et
al.'
s
spatially­
explicit,
stochastic
CBW
model
(
Storer
et
al.,
2003)
was
adapted
to
incorporate
southnorth
migration
in
the
spring
and
north­
south
migration
in
the
summer.


Quantify
how
migration
may
impact
adaptation
rates
under
a
range
of
different
circumstances.


Two
models
were
run
in
parallel
for
the
summer
generations:


cotton­
growing
region

corn­
growing
region.
11
N­
S
Movement
 
Modeling
Assumptions

Corn,
cotton
and
soybean
are
crop
hosts
for
CBW;


Weed
hosts
are
available
at
the
start
of
the
season
and
end
of
the
season;


Complete
cross­
resistance
among
the
three
Cry1
toxins
(
Cry1Ab,
Cry1Ac,
Cry1F);


Additive
functional
dominance
of
allele
for
adaptation
(
h
=
0.5);


Migration
occurs
pre­
mating
(
as
indicated
by
field
studies
on
mating
and
flight
behavior
in
Helicoverpa
spp.);
and

Migration
occurs
to
the
same
extent
every
year.
12
N­
S
Movement
 
Model
Output

Model
output
was
the
resistance
(
r)
allele
frequency
after
15
years
of
deploying
Bt
corn
and
Bt
cotton
13
N­
S
Movement
 
Modeling
Results

Two
parameters
increased
the
r­
allele
frequency
after
15
years

Increasing
the
percentage
of
Bt
corn
(
BtcrS)


Increasing
the
percentage
of
Bt
cotton
(
Btct)


Magnitude
of
significant
effects
or
interactions
ln(
Q+
15/
Q0)/
ln(
Q­
15/
Q0)
­
1

Q+
15
=
r­
allele
frequency
after
15
years
with
reverse
migration

Q0
=
initial
r­
allele
frequency

Q­
15
=
r­
allele
frequency
after
15
years
without
reverse
migration
14
Impact
of
Reverse
Migration
on
Adaptation
Rate
­
Modeling
Impact
of
reverse
migration
on
adaptation
rate
after
15
years
Bt
Cotton
Bt
Corn
Increase
10%

60%

80%

Extreme
Conditions
 
Selection
is
greatest
in
the
North,

Lowest
in
the
South
(
i.
e.,

50%
return
migration,

75%
survival
of
N­
S
migrants)
Increase
2%

60%

80%

High
Bt
Corn
Slow
4%

60%

30%

Current
Deployment
Levels
15
N­
S
Movement
Study
Conclusion

Based
on
the
modeling
studies
submitted
using
the
data
in
Gould
et
al.
(
2002),
CBW
(
also
called
corn
earworm
in
corn)
reverse
migration
has
no
significant
impact
(
0.05<
P)
on
CBW
adaptation
to
Bt
corn
and
cotton.
16
Pyrethroid
Overspray
Studies

Field
studies:
NC,
LA,
MS,
SC
(
Natural
infestations)


Laboratory
studies:
CBW
survivor
susceptibility

Purpose:
Determine
the
impact
of
pyrethroid
oversprays
on
CBW
control
in
Bollgard
fields
and
evolution
of
resistance.
17
North
Carolina
Field
Studies
 
2001­
02
North
Carolina
Bollgard
Non­
Bollgard
78.2
93.7
Adults
73.8
92.1
Pupae
62.1
89.3
Large
larvae
Jackson
et
al.,
2003b
2002
72.1
77.5
%
Damaged
Bolls
70.4
87.9
%
Damaged
Squares
69.7
86.7
%
Infested
Bolls
80.5
92.3
%
Infested
Squares
Jackson
et
al.,
2003a
2001­
02
%
Reduction
Due
to
Spray
What
was
measured
Study
and
Year
Conducted
18
LA
and
MS
Field
Studies
 
2001­
02
Mississippi
Louisiana
55.4
58.3
%
Damaged
Bolls
47.9
59.8
Mean
#
Larvae
per
Sample*
­
9.4
70.0
Mean
#
Larvae
per
25
plants
Harris
et
al.,
2002
2001
40.4
63.6
%
Infested
Bolls
Leonard,
2003
2002
Non­
Bollgard
Bollgard
%
Reduction
Due
to
Spray
What
was
measured
Study
and
Year
19
South
Carolina
 
1999
Field
Study
Irrigated*
83.1
58.5
0.015
93.1
87.1
0.028
Brickle
et
al.,

2001
71.9
85.7
0.015
86.4
94.7
0.028
Brickle
et
al.,

2001
Non­
irrigated
Non­
Bollgard
Bollgard
%
Reduction
Due
to
Spray
Application
Rate
(
kg
ai/
ha)

Study
*
Excessive
soil
moisture
or
vegetative
growth
or
intrinsic
differences
in
Bt
expression
20
Pyrethroid
Oversprays
for
CBW
in
Bollgard
vs.
Non­
Bollgard
Fields:
Conclusions

Field
studies
NC,
MS,
LA,
SC
(
non­
irrigated)

show
pyrethroid
oversprays
are
more
effective
in
Bollgard
fields
than
non­
Bollgard
fields.


Percent
insecticide
control
values
and
total
number
of
adults
(
insecticide­
treated
vs.

nontreated
can
be
used
to
develop
reliable
parameters
for
mathematical
models.
21
Pyrethroid
Oversprays
for
CBW
in
Bollgard
II
Fields:
Conclusions

Percentage
squares
and
bolls
infested
and
percentage
damaged
squares
and
bolls
were
not
statistically
different
in
Bollgard
treated
and
Bollgard
II
untreated
and
treated
plots.


Results
from
North
Carolina
and
Mississippi
field
studies
suggest
that
pyrethroid
oversprays
will
likely
not
be
necessary
for
CBW
in
Bollgard
II
cotton
fields
as
they
were
for
Bollgard
cotton
fields
22
Bollworm
Survivor
Susceptibility:
NC
2003
0.069
0.191
0.150
F2
Bollgard
0.137
0.217
0.179
F2
Non­
Bollgard
12.8
19.6
15.6
F2
Bollgard
25.5
37.9
30.6*

F1
Bollgard
Bollgard
Non­
Bollgard
Non­
Bollgard
Non­
Bollgard
Cotton
Type
Cypermethrin
(
technical
grade)

Cry1Ac
7.41
27.9
13.2
F2
0.528
2.82
0.806
F1
0.504
0.819
0.623
F1
12.3
21.8
16.3
F1
Lower
Upper
95%
Confidence
Limits
LC
50
(
mg/
ml)

Generation
(
Marcus
et
al.
2004)
23
Laboratory
Study
Bollworm
Susceptibility
Conclusions

Tolerance
in
Cry1Ac
in
the
F1
generation
was
not
statistically
significant
and
may
not
truly
exist

No
difference
in
susceptibility
to
pyrethroids

Authors
are
working
to
select
for
a
CBW
strain
that
has
very
high
levels
of
Cry1Ac
resistance
to
provide
more
rigorous
results
24
Pyrethroid
Overspray
Studies:
Overall
Conclusions

Pyrethroid
oversprays
in
Bollgard
cotton
fields
will
increase
the
level
of
control
of
CBW
and
delay
the
evolution
of
resistance.


Findings
can
be
used
to
develop
reliable
parameters
in
mathematical
models

Pyrethroid
oversprays
for
Bollgard
II
cotton
 
no
significant
difference

Laboratory
studies
have
not
uncovered
a
reason
for
the
fitness
difference
(
Bollgard
vs.
non­
Bollgard).
25
Alternative
Host
Studies

In
2001,
EPA
required
that
alternative
host
data
be
generated.


Purpose:
Demonstrate
whether
CBW
alternative
hosts
provide
a
large
and
effective
unstructured
refuge
for
Bollgard
and
Bollgard
II
cotton
in
the
U.
S
to
allow
continuation
of
the
5%
external,

unsprayed
structured
refuge
option.
26
Outline
of
Alternative
Hosts
Studies

Two­
year
study
(
2002­
2003)
conducted
in
LA,
AR,
MS,
GA,
NC

100
sampling
locations:
Bollgard
cotton
field
and
an
adjacent
alternate
host
crop

Bollgard:
corn,
soybean,
sorghum
(
MS,
LA,
AR
­
mid­
South
locations)


Bollgard:
peanuts
(
NC,
GA
­
Southeast
locations)


Bollgard:
non­
Bollgard

Bollgard
cotton:
Bollgard
cotton

Larval
productivity

Adult
productivity:
pheromone
traps

C3/
C4
carbon
isotope
analyses:
crop
source
of
adults

Assessment
of
the
spatial
and
temporal
distribution
of
alternative
hosts
using
satellite
imaging
and
USDA/
NASS
(
2002)
database
27
Literature
Review
(
Benedict,
2004)


Comprehensive
review
of
biology
and
ecology
of
TBW
and
CBW

CBW

Sections:
alternative
hosts,
sequential
host
utilization,
larval
and
adult
productivity
on
each
host,
movement
and
dispersal
properties,
gene
flow

130
crop
and
non­
crop
hosts;
typically
4
generations;
ability
to
move
amongst
hosts
>
10
mi;
little
genetic
variation

Identify
corn,
peanuts,
sorghum,
and
soybeans
as
alternative
hosts
for
CBW

Few,
if
any,
studies
that
have
determined
the
numbers
of
adult
heliothines
produced
per
acre
by
any
host
plant
and
determined
the
number
of
adults
produced
per
acre
by
all
host
plant
species
for
a
crop
production
system

Field
studies
designed
to
prove
whether
alternative
hosts
provide
a
large
and
effective
unstructured
CBW
refuge
28
Cropping
Pattern
Analysis

Corn,
peanuts,
sorghum,

and
soybeans
present
over
substantial
areas
in
all
of
the
major
cotton
growing
areas
at
all
spatial
scales
examined
(
1
mile,
10
mile)


Data
from
USDA/
NASS,

2002

Acreages
remained
stable
from
1995­
2002
in
each
state
or
county
1a.
Arkansas
cropping
patterns
0
40000
80000
120000
160000
1995
1997
1999
2001
Cumulative
acreage
Soybean
Sorghum
Corn
Cotton
1d.
Georgia
cropping
patterns
0
20000
40000
60000
80000
1995
1996
1997
1998
1999
2000
2001
2002
Cumulative
acreage
Tobacco
Soybean
Sorghum
Peanuts
Corn
Cotton
1b.
Louisiana
cropping
patterns
0
45000
90000
135000
180000
1995
1997
1999
2001
Cumulative
acreage
Soybean
Sorghum
Corn
Cotton
1c.
Mississippi
cropping
patterns
0
60000
120000
180000
240000
1995
1997
1999
2001
Cumulative
acreage
Soybean
Sorghum
Corn
Cotton
1e.
North
Carolina
cropping
patterns
0
25000
50000
75000
100000
1995
1996
1997
1998
1999
2000
2001
2002
Cumulative
acreage
Tobacco
Soybean
Sorghum
Peanuts
Corn
Cotton
Tobacco
Soybean
Sorghum
Peanuts
Corn
Cotton
(
Head
and
Voth,
2004)
29
Larval
Productivity
Studies

Larval
distributions
over
time
in
five
states
in
2002
and
2003

Larval
surveys
used
to
measure
productivity
on
each
host
and
predict
when
relevant
crop
fields
will
produce
CBW
adults
and

Time
to
adult
emergence
is
approx.
14
days

Adult
lifespan
is
approx.
7
days
30
3a.
Larval
abundance
in
Arkansas
2003
0
1
2
3
6/
2
6/
16
6/
30
7/
14
7/
28
8/
11
8/
25
9/
8
Abundance
Cor
n
Cotton
Sor
ghum
Soybean
3d.
Larval
abundance
in
Georgia
2003
0
1
2
3
6/
2
6/
16
6/
30
7/
14
7/
28
8/
11
8/
25
9/
8
Abundance
Cor
n
Cotton
Peanut
Soybean
3b.
Larval
abundance
in
Louisiana
2003
0
1
2
3
6/
2
6/
16
6/
30
7/
14
7/
28
8/
11
8/
25
9/
8
Abundance
Cor
n
Cotton
Sor
ghum
Soybean
3c.
Larval
abundance
in
Mississippi
2003
0
1
2
3
6/
2
6/
16
6/
30
7/
14
7/
28
8/
11
8/
25
9/
8
Abundance
Cor
n
Cotton
Sor
ghum
Soybean
3e.
Larval
abundance
in
North
Carolina
2003
0
1
2
3
6/
2
6/
16
6/
30
7/
14
7/
28
8/
11
8/
25
9/
8
Abundance
Cor
n
Cotton
Peanut
Soybean
Larval
Productivity
­
2003
Index:
1
=
20­
200
larvae/
acre;
2
=
200­
2000
larvae/
acre;
3
=

>
2000
larvae/
acre
(
and
generally
>
10,000).

(
Head
and
Voth,
2004)
1.
Order:
corn>>>
sorghum>
cotton
(
peanuts)
>
soybean
2.
NC:
Populations
are
synchronous
Corn
Sorghum
(
AR,
LA,
MS)

Cotton
Soybean
Peanut
(
GA,
NC)
31
Larval
Productivity
Studies
­
Conclusions

Larval
populations
were
high
on
the
alternative
host
crops
and
were
equal
to
or
greater
than
populations
on
cotton

Phenology
of
these
populations
overlapped
with
those
populations
in
cotton
(=
Synchrony
of
production).


Predict
CBW
adults
in
corn
by
mid­
June
and
dominate
the
landscape
until
early
August

Later
adult
generations
are
predicted
to
dominate
in
mid­
August
on
sorghum,
cotton,
peanuts,
and
soybeans
(
depending
on
the
region)


Difficult
to
predict
adult
productivity
from
larval
productivity
studies
32
Adult
Productivity
Studies
 
Pheromone
Trapping
Data
in
2002­
03

Comparable
numbers
of
adult
CBW
were
captured
at
all
the
different
crop
interfaces
in
a
given
state

Adults
moving
broadly
across
the
landscape

No
statistical
differences
except
for
a
few
early
season
samples
in
Arkansas
and
a
few
other
sporadic
cases

In
2003,
30
out
of
878
pairs
of
means
(
3.4%)
were
statistically
significant
at
the
95%
confidence
level
33
Carbon
Isotope
Analysis:
Sources
of
Adult
Moths
(
Semi­
quantitative)


Distinguish
moths
that
feed
on
C3
hosts
(
soybean,
cotton,

peanuts)
from
C4
hosts
(
corn,
sorghum)
(
method
described
in
Gould
et
al.,
2002).


Moths
that
feed
on
C3
hosts
will
have
a
different
13C
to
12C
ratio
than
moths
that
feed
on
C4
hosts
based
on
carbon
assimilation
physiology
of
the
host
plant.


Wings
from
up
to
100
adults
from
a
given
trapping
location
and
date
were
pooled

Percentage
of
C4
plants
was
calculated
by
comparing
the
value
obtained
to
a
standard
curve

ANOVA
run
on
%
moths
from
C4
plants
and
pair­
wise
comparisons
were
performed
for
each
state
and
week
in
both
2002
and
2003
34
Fig.
5.
Percentage
of
C4
Adults
by
State
in
2003
0
20
40
60
80
100
5/
19
6/
9
6/
30
7/
21
8/
11
9/
1
9/
22
10/
13
%

C4
Arkansas
Georgia
Louisiana
Mississippi
North
Carolina
Sources
of
Adult
Moths:

Percentage
of
C4
Adults
by
State
(
Head
and
Voth,
2004)

0
20
40
60
80
100
5/
19
6/
9
6/
30
7/
21
8/
11
9/
1
9/
22
10/
13
%

C4
C4
alternative
hosts
C3
hosts
other
than
cotton
C3
hosts
including
cotton,

soybeans
and
peanuts
35
Sources
of
Moths:
Conclusions

Total
adult
production
from
C4
alternative
host
of
CBW,

averaged
across
all
states,
is
at
least
20%
and
reaches
a
peak
of
80­
100%
from
early
July
through
the
end
of
August.


Exact
source
and
proximity
of
these
alternative
hosts
cannot
be
determined
from
the
data.


Source
of
late
season
CBW
adults
may
be
southern
migrants
from
the
Midwest
corn
belt
(
Gould
et
al.,

2002).
Findings
of
Head
and
Voth
(
2004)
are
the
same
as
Gould
et
al.
(
2002).
36
7a.
%
C4
in
Arkansas
2002
0
20
40
60
80
100
6/
3
6/
17
7/
1
7/
15
7/
29
8/
12
8/
26
9/
9
%

C4
Obser
ved
County
10
mi
le
1
mil
e
7b.
%
C4
in
Louisiana
2002
0
20
40
60
80
100
6/
10
6/
24
7/
8
7/
22
8/
5
8/
19
9/
2
9/
16
%

C4
Obser
ved
County
10
mi
le
1
mi
l
e
7e.
%
C4
in
North
Carolina
2002
0
20
40
60
80
100
6/
17
7/
1
7/
15
7/
29
8/
12
8/
26
9/
9
%

C4
Obser
ved
County
10
mi
le
1
mi
l
e
7c.
%
C4
in
Mississippi
2002
0
20
40
60
80
100
6/
24
7/
8
7/
22
8/
5
8/
19
9/
2
9/
16
%

C4
Obser
ved
County
10
mi
le
1
mi
le
7d.
%
C4
in
Georgia
2002
0
20
40
60
80
100
6/
24
7/
8
7/
22
8/
5
8/
19
9/
2
%

C4
Obser
ved
County
Observed
and
Predicted
Percentage
of
CBW
Moths
from
C4
Hosts
at
Different
Spatial
Scales
°
Predicted
curves:
satellite
imaging
for
1­
mi
and
10­

mi;
USDA/
NASS
for
county
level
at
10­
mi
scale
and
larval
abundances
translated
to
adult
emergence
°
Predicted
underestimate
the
importance
of
C4
contribution.

°
Scale
of
adult
CBW
movement
is
>
10
miles
Observed
County
10
mile
1
mile
*
37
CBW
Alternative
Hosts
Studies
 
Specific
Conclusions

Based
on
carbon
isotope
analyses,
both
C3
and
C4
alternative
hosts
serve
as
unstructured
refugia
throughout
the
growing
season
and
throughout
the
landscape.


Alternative
hosts
contribute
greater
numbers
of
CBW
moths
than
the
local
5%
external,

unsprayed
structured
non­
Bt
cotton
refuge.


C4
alternative
hosts
contribute
20­
95%
of
the
CBW
moths
in
and
around
cotton
fields
throughout
the
growing
season.
38
CBW
Alternative
Hosts
Studies
 
Specific
Conclusions

Corn
is
the
greatest
producer
of
CBW
adults
in
the
early
season;
while,
sorghum,
peanuts,

soybeans,
and
cotton
are
producers
of
CBW
adults
in
the
later
generations.


CBW
adults
are
dispersing
long
distances
(>
10
miles)
in
search
of
suitable
oviposition
and
feeding
sites.
39
CBW
Alternative
Hosts
Studies
 
Overall
Conclusions

CBW
moths
are
produced
on
alternative
hosts
in
sufficient
numbers
throughout
the
cotton
growing
season
to
mate
with
any
putative
resistant
CBW
moths
emerging
in
Bollgard
or
Bollgard
II
cotton
fields
and
dilute
resistance.


Susceptible
CBW
moths
coming
from
alternative
hosts
(
immigrants)
will
reduce
the
intensity
of
Cry1Ac
and
Cry2Ab2
resistance
selection
in
CBW
and
lower
the
likelihood
of
resistance
evolution.


Results
of
the
alternative
host
studies
support
the
continuation
of
the
5%
external,
unsprayed
structured
non­
Bt
cotton
refuge.
40
Gustafson
et
al.
(
2001)
CBW
Model*


Modified
Caprio's
(
1998a)
ILSI
two­
patch,

deterministic,
non­
random,
population
genetics
model

Included
insecticidal
oversprays
of
Bollgard
cotton
fields
and
the
utilization
of
alternative
hosts
as
natural
refugia
as
parameters

*
Same
study
submitted
in
2001
and
2004.
41
Gustafson
et
al.
(
2001)
CBW
model
 
"
Effective
Refuge
Size"


Calculated
by
county

Total
acres
of
non­
Bt
cotton,
Bt
cotton,
soybean,

alfalfa,
sorghum
(
USDA/
NASS,
1999)


Wild
and
weedy
plants
=
10%
(
Caprio
and
Benedict,
1996)


Corn
not
included

Effective
refuge
size:
43­
100%
42
Gustafson
et
al.
(
2001)
CBW
Model

Model
output:
number
of
generations
until
the
rallele
frequency
exceeds
0.5

Marginal
sensitivity
analysis:
one
parameter
alone
was
varied
while
all
other
parameters
were
held
at
the
"
mid"
value
43
Gustafson
et
al.,
(
2001)
CBW
Model:

Sensitivity
Analysis
Results
86
86
47
>
5000
84
68
42
47
High
91
91
85
147
86
86
92
90
Mid
26
26
26
36
25
25
26
27
Mid
27
26
36
13
25
26
>
5000
>
5000
Low
CBW
M
R
q
(
t­
0)

P
rr
Sr
Sr
SS
101
25
Move
outside
habitat
before
ovipositing
101
25
Move
outside
habitat
before
mating
127
16
Resistance
Allele
Initial
Frequency*
25
553
Refuge*
89
25
123
24
Survival
on
Conventional
Cotton
>
5000
15
>
5000
15
Survival
on
Bollgard
Cotton
Low
High
Parameter
CBW
Sprayed*
44
Gustafson
et
al.
(
2001)
CBW
Model
Results
CBW
Untreated
BG
CBW
Treated
BG
Impact
of
effective
refuge
size
and
pyrethroid
oversprays
on
model
results
using
"
mid"
parameter
values
45
Gustafson
et
al.
(
2001)
CBW
Model
 
Issues
with
"
Effective
Refuge
Size"


Monsanto's
calculation
assumes
that
all
hosts
are
equally
productive
and
synchronous,
as
well
as
are
generation
independent.


EPA
disagrees
with
Monsanto's
assumptions.
Same
type
of
calculation
used
in
Head
and
Voth
(
2004).


There
is
no
sequential
host
utilization
by
generation.


Different
generations
will
feed
on
different
hosts.


Larval
and
moth
production
on
each
host
varies
considerably.


Corn
not
included
in
2001.
46
Gustafson
et
al.
(
2001)
CBW
Model
 
Issues
with
"
Effective
Refuge
Size"


EPA
recommends
that
the
"
effective
refuge
size"

be
a
weighted
average
of
the
proportion
of
moths
coming
from
each
alternative
host
for
each
CBW
generation
(
4
to
6
generations)
in
each
cotton
production
system.
47
Gustafson
et
al.
(
2001)
CBW
Model
 
Conclusions

Model
output
is
very
sensitive
to
effective
refuge
size
and
use
of
insecticide
sprays
on
Bollgard
fields
to
control
CBW.


There
is
a
10­
fold
increase
in
time
to
resistance
for
CBW
when
Bollgard
cotton
is
sprayed
with
pyrethroids.


Model
is
limited,
there
is
no
spatial
or
temporal
dynamic
and
"
effective
refuge
size"
should
be
recalculated.
48
Gustafson
et
al.
(
2001)
CBW
Model
 
Conclusions

Despite
the
issues
associated
with
the
calculation
of
"
effective
refuge
size,"
inclusion
of
alternative
hosts
in
the
model
will
delay
resistance
much
longer
than
a
5%

unsprayed,
structured
non­
Bt
cotton
refuge.


Model
was
not
rerun
with
parameter
values
derived
from
the
results
of
the
synthetic
pyrethroid
overspray
studies
(
Greenplate,
2004)
and
the
alternate
host
studies
(
Head
and
Voth,
2004).
If
model
had
been
updated
to
include
study
results,
the
results
would
likely
have
supported
the
continuation
of
the
5%
external,

unsprayed,
structured
refuge
option.
49
Overall
EPA
Conclusion:
Bollgard
and
Bollgard
II
CBW
Resistance
Management

Data
from
the
N­
S
movement
study,
pyrethroid
overspray
studies,
alternative
host
studies
support
the
continuation
of
the
5%
external,
unsprayed,

structured
non­
Bt
cotton
refuge
option
for
CBW
resistance
management.
50
Acknowledgements

Alan
Reynolds,
M.
S.,
OPP/
BPPD

Hilary
Hill,
M.
S.,
OPP/
BPPD

Tessa
Milofsky,
M.
S.,
OPP/
BPPD
51
Bollgard
and
Bollgard
II
CBW
Resistance
Management
SAP
Questions
1.
North­
south
movement.
The
Agency
requests
that
the
SAP
comment
on
whether
CBW
reverse
migration
is
expected
to
have
any
significant
impact
on
CBW
adaptation
to
Bt
crops.
52
Bollgard
and
Bollgard
II
CBW
Resistance
Management
SAP
Questions
2.
Pyrethroid
Oversprays.

a.
The
Agency
requests
that
the
SAP
comment
on
whether
pyrethroid
oversprays
in
Bollgard
cotton
fields
are
likely
to
increase
the
level
of
control
of
CBW,

delay
the
evolution
of
resistance,
and
increase
the
relative
effectiveness
of
the
5%
external,
unsprayed,

structured
refuge.

b.
The
Agency
also
requests
that
the
SAP
comment
on
EPA's
recommendation
that
pyrethroid
oversprays
not
be
included
as
a
parameter
in
the
Gustafson
et
al.

(
2004)
model
for
Bollgard
II.
53
Bollgard
and
Bollgard
II
CBW
Resistance
Management
SAP
Questions
2.
Cont.

c.
The
Agency
requests
the
SAP
comment
on
whether
the
cotton
bollworm
larvae
coming
from
Bollgard
fields
are
more
tolerant
to
the
Cry1Ac
protein
than
those
larvae
coming
from
the
non­
Bollgard
fields.
What,
if
any,
additional
genetic
work
should
be
conducted
to
better
understand
the
nature
of
this
Cry1Ac
tolerance.

d.
The
Agency
requests
the
SAP
to
comment
on
the
value
of
using
a
Cry1Ac­
resistant
CBW
colony
to
investigate
the
genetic
basis
for
CBW
survival
on
Bollgard
cotton.
54
Bollgard
and
Bollgard
II
CBW
Resistance
Management
SAP
Questions
3.
Alternative
Hosts.

a.
Based
on
the
larval
productivity
analyses,
adult
productivity
analyses,
and
satellite
imaging
analysis,

the
Agency
asks
the
SAP
to
comment
on
the
relative
contribution
of
the
C3
and
C4
alternate
hosts
as
unstructured
refugia
to
dilute
CBW
resistance.

b.
Based
on
the
data,
the
Agency
also
asks
the
SAP
to
comment
on
the
spatial
and
temporal
scale
across
the
landscape,
e.
g.,
1
mile,
10
mile
etc.,
in
which
CBW
adult
production
should
be
evaluated.
55
Bollgard
and
Bollgard
II
CBW
Resistance
Management
SAP
Questions
3.
Cont.

c.
The
Agency
asks
the
SAP
to
comment
on
how
to
quantitatively
or
semi­
quantitatively
calculate
"
effective
refuge
size"
locally
and
regionally
using
available
data
(
see
above).
56
Bollgard
and
Bollgard
II
CBW
Resistance
Management
SAP
Questions
4.
Gustafson
et
al.
CBW
Model.

a.
The
Agency
asks
the
SAP
to
comment
on
the
"
effective
refuge
size"
calculation.
Does
the
SAP
agree
with
the
Agency's
conclusion
that
"
effective
refuge
size"
is
a
weighted
average
of
the
proportion
of
moths
coming
from
each
alternate
host
for
each
CBW
generation
(
5
to
6
generations)
in
each
cotton
production
system
(
geography)?

.
57
Bollgard
and
Bollgard
II
CBW
Resistance
Management
SAP
Questions
4.
Cont.

b.
The
Agency
requests
the
SAP
to
comment
on
the
strengths
and
weaknesses
of
the
Gustafson
et
al.
(
2004)
model
and
its
utility
with
regard
to
the
effective
contribution
of
alternate
hosts
as
natural
refuge
per
generation.
How
would
the
model
output
be
altered
if
the
calculation
of
"
effective
refuge
size"
is
changed
(
see
a.
above).
What
are
the
SAP's
recommendations
for
refining
the
Gustafson
et
al.
(
2004)
CBW
resistance
management
model
or
using
a
different
CBW
resistance
management
model
to
more
appropriately
consider
the
spatial
and
temporal
dynamics
of
CBW
utilization
of
alternative
hosts
by
generation
based
on
the
data
in
Head
and
Voth
(
2004)?
58
Bollgard
and
Bollgard
II
CBW
Resistance
Management
SAP
Questions
4.
Cont.

c.
The
Agency
requests
the
SAP
to
comment
on
validity
of
using
the
average
pyrethroid
efficacy
value
against
CBW
based
on
all
the
field
studies
conducted
in
all
four
states
(
North
Carolina,
Louisiana,
Mississippi,

and
South
Carolina)
as
the
parameter
value
in
the
Gustafson
et
al.
(
2004)
model
rather
than
just
the
Brickle
et
al.
(
2001)
data
from
South
Carolina.
