Ethylene oxide for the control of american foulbrood in honey bees


Shimanuki, H.; Lehnert, T.

Journal of Economic Entomology 61(5): 1456-1457

1968


Since the insecticidal properties of ethylene oxide were 1st determined by Cotton and Roark (1928), this gas has been widely used to sterilize heat-sensitive materials, and numerous nonflammable formulations have been made available.

1456
JOURNAL
OF
ECONOMIC
ENTOMOLOGY
Vol.
61,
No.
5
inheritance
is
involved
in
the
resistance
of
alfalfa
to
the
spotted
alfalfa
aphid,
the
average
number
of
aphids
on
the
F,
plants
arising
from
the
cross
of
susceptible
female
X
resistant
male
should
have
been
significantly
greater
than
the
average
number
of
the
reciprocal
cross.
The
average
numbers
of
aphids
on
the
F,
plants
from
both
crosses
were
essentially
the
same.
Cytoplasmic
inheritance
of
resistance
in
alfalfa
to
the
spotted
alfalfa
aphid
ap-
parently
does
not
occur.
REFERENCES
CITED
Allard,
R.
W.
1960.
Principles
of
Plant
Breeding.
John
Wiley
&
Sons,
Inc.,
New
York.
485
p.
Srb,
A.
M.,
R.
D.
Owen,
and
R.
S.
Edgar.
1965.
General
Genetics.
W.
H.
Freeman
&
Co.,
San
Francisco
&
Lon-
don.
557
p.
Tysdal,
H.
M.,
and
J.
R.
Garl.
1940.
A
new
method
for
alfalfa
emasculation.
Agron.
J.
32
(5)
:
405-7.
Ethylene
Oxide
for
the
Control
of
American
Foulbrood
in
Honey
Beest,
2
'
s
H.
SHIMANUKI
and
T.
LEHNERT
Entomology
Research
Division,
Agr.
Res.
Serv.,
USDA,
Beltsville,
Maryland
20705
Since
the
insecticidal
properties
of
ethylene
oxide
were
1st
determined
by
Cotton
and
Roark
(1928)
,
this
gas
has
been
widely
used
to
sterilize
heat
-sensitive
materials,
and
numerous
nonflammable
formulations
have
been
made
available.
American
foulbrood
(AFB)
is
a
disease
of
larval
honey
bees,
Apis
mellif
era
L.,
caused
by
the
bacterium
Bacillus
larvae
White.
This
disease
causes
the
degeneration
of
larvae
and
pupae
into
a
spore
-laden
mass
that
dries
to
a
scale.
Because
this
scale
adheres
rigidly
to
the
cell
of
the
honeycomb
and
is
difficult
to
remove,
it
becomes
the
chief
source
of
reinfection
of
other
larvae.
Also,
the
stores
of
honey
can
be
contaminated
and
become
a
source
of
the
pathogen.
Claims
of
successful
disinfection
of
honeycombs
were
made
by
Hutzelman
(1922)
for
formalin
and
alcohol;
by
Wilson
(1924)
for
sodium
hyporchlorite;
by
Hutson
(1927)
for
gaseous
chlorine;
by
Katznelson
et
al.
(1952)
for
high
velocity
electrons;
and
Studier
(1958)
for
gamma
radiation,
but
none
of
them
have
proved
prac-
tical.
The
1st
report
on
the
effect
of
ethylene
oxide
as
a
fumigant
for
the
control
of
parasites
and
pests
of
honey
bees
was
made
by
Michael
(1964)
.
He
demonstrated
that
spores
of
B.
larvae
could
be
killed
by
fumigation
with
ethylene
oxide.
Later,
Shimanuki
(1967)
reduced
the
number
of
viable
spores
of
B.
larvae
in
colonies
of
honey
bees
and
suggested
that
a
higher
degree
of
success
could
be
attained
by
improving
the
fumigating
chamber
and
increasing
doses.
The
present
tests
were
made
therefore
to
determine
the
concentration
of
ethylene
oxide
required
to
prevent
reinfection.
MATERIALS
AND
METHODS.
--A
fumigation
chamber
was
constructed
on
a
4X8
-ft
sheet
of
plywood
placed
on
the
ground
to
form
a
flat
surface
on
which
contaminated
equipment
could
be
laid.
Then
a
fumigating
chamber
was
made
on
the
board
by
enclosing
the
contaminated
equipment
with
either
1
or
2
thicknesses
of
black
6
-mil
polyethylene.
Fifty-six
combs
that
each
contained
more
than
100
AFB
scales
were
selected
for
the
test
and
arranged
into
28
hive
bodies
with
2
combs
and
7
sheets
of
foundation
each.
The
28
hive
bodies
were
then
divided
at
random
into
2
groups.
In
the
1st
group,
3
sets
of
4
hive
bodies
were
fumigated
for
24
hr
with
1/2
,
1,
or
2
lb
of
Oxyfume-12®
(ethylene
oxide)
per
hive
body
(available
from
Union
Carbide
Corporation,
Linde
Division)
.
In
the
2nd
group,
3
sets
of
4
hive
bodies
were
fumigated
with
similar
doses
and
then
treated
a
2nd
time
with
the
same
dose
8
hr
after
the
1st
treatment.
The
other
4
hive
bodies
(2
in
each
group)
were
not
fumigated
and
served
as
controls.
After
the
fumigated
hive
bodies
had
aired
24
hr,
each
was
stocked
with
a
3
-lb
package
of
Italian
bees.
Weekly
inspections
for
AFB
were
then
made
of
all
colonies.
A
colony
was
considered
diseased
even
if
only
1
larvae
had
an
active
Hymenoptera:
Apidac.
2
Accepted
for
publication
May
3,
1968.
3
Mention
of
a
proprietary
product
does
not
necessarily
imply
its
endorsement
by
the
USDA.
case
of
AFB.
RESULTS
AND
DiscusstoN.—A
high
concentration
of
ethyl-
ene
oxide
can
be
obtained
in
the
chamber
by
introducing
it
into
a
vacuum
or
by
fl
ushing
out
the
air.
Since
our
goal
was
to
devise
an
inexpensive
portable
system,
we
chose
to
fl
ush
out
the
air.
The
1st
experiment
was
made
to
determine
an
effective
concentration
of
gas
that
would
prevent
recurrence
of
AFB.
All
4
unfumigated
control
colonies
had
active
AFB
(Table
1)
All
colonies
fumigated
with
1/2
or
1
lb
of
Oxyfume-12
had
active
AFB;
when
2
lb
were
used,
3
of
4
colonies
were
diseased.
The
2nd
treatment
effectively
reduced
the
incidence
of
disease
(Table
1)
.
When
1
/
2
lb
of
gas
was
used
twice,
2
of
4
colonies
had
AFB.
However,
when
1
lb
of
gas
was
used
twice,
only
1
colony
developed
AFB,
and
of
the
4
colonies
that
received
the
2
-lb
treatment
twice,
only
1
had
a
dis-
eased
larva.
Two
other
colonies
that
showed
infection
(1
that
received
a
single
1/4
-lb
treatment
and
1
that
received
two
1
/
2
-lb
treatments
)
seemed
to
recover
spontaneously.
In
16
colonies
that
became
infected
early
in
the
experi-
ment,
the
disease
was
allowed
to
become
severe
so
we
would
be
provided
with
equipment
and
stores
contami-
nated
with
AFB
in
all
stages.
Each
such
colony
was
then
treated
twice
with
2
lb
of
gas/hive
body
and
restocked
with
bees.
Three
months
later,
15
of
the
16
had
no
sign
of
infection.
No
reluctance
by
the
bees
in
the
acceptance
of
the
fumigated
combs
was
observed.
The
worker
bees
cleaned
the
scales
out
of
the
cells,
and
the
queen
began
to
lay
eggs
in
less
than
1
week.
Also,
no
evidence
of
toxicity
to
ethylene
oxide
was
apparent
in
any
colony.
Of
the
28
original
colonies
that
would
otherwise
have
been
de-
stroyed,
only
1
has
AFB.
Our
degree
of
success
might
be
improved
in
many
ways.
For
example,
a
3rd
introduction
of
gas
may
be
needed
to
maintain
the
level
in
our
present
chamber,
though
im-
provements
in
the
chamber
might
eliminate
this
need.
Table
1.
—Effect
of
concentrations
of
ethylene
oxide
on
the
incidence
of
AFB
4
months
after
treatment
(all
treat-
ments
replicated
4
times).
Treatment
(lb/hive
body)
"
No.
of
disease
-free
colonies
No.
of
diseased
colonies
Initial
Second
0.0
0.0
0
4
.5
.0
0
4b
1.0
.0
0
4
2.0
.0
1
3
.5
.5
2
2b
1.0
1.0
3
1
2.0 2.0
3
b
Oxyhtme-128.
b
I
diseased
colony
showed
an
apparently
spontaneous
recovery.
October
1968
SCIENTIFIC
NOTES
1457
Also,
controlled
temperature
and
humidity
would
un-
doubtedly
increase
the
efficiency
of
the
fumigation.
ACKNOWLEDGMENTS.
—We
thank
Messrs.
David
Knox
and
Elton
Herbert
of
this
laboratory
for
their
valuable
assist.
ance.
We
are
especially
grateful
to
Mr.
Victor
Thompson
of
the
Ohio
State
University
Apiculture
Laboratory
for
providing
some
of
the
bee
equipment.
REFERENCES
CITED
Cotton,
R.
T.,
and
R.
C.
Roark.
1928.
Ethylene
oxide
as
a
fumigant.
Ind.
Eng.
Chem.
20:
805.
Hutson,
R.
1927.
Gaseous
chlorine
as
a
disinfectant
for
American
foulbrood
infected
combs.
J.
Econ.
Ento-
mol.
20:
516-20.
Hutzelman,
J.
C.
1922.
Can
the
combs
be
saved?
New
treatment
for
American
foulbrood
by
immersion
in
disinfectant
solution.
Gleanings
Bee
Cult.
50:
764-6.
Katznelson,
H.,
C.
A.
Jamieson,
E.
J.
Lawton,
and
W.
D.
Bellamy.
1952.
Studies
on
the
treatment
of
con-
taminated
combs
and
honey
with
high
velocity
elec-
trons.
Can.
J.
Technol.
30:
95-103.
Michael,
A.
S.
1964.
Ethylene
oxide,
a
fumigant
for
con-
trol
of
pests
and
parasites
of
the
honeybee.
Gleanings
Bee
Cult.
92:
102-4.
Shimanuki,
H.
1967.
Ethylene
oxide
and
control
of
American
foulbrood—a
progress
report.
Amer.
Bee
J.
107
(8)
:
290-1.
Studier,
H.
1958.
The
sterilization
of
American
foul
-
brood
by
irradiation
with
gamma
rays.
Amer.
Bee
J.
98:
192.
Wilson,
H.
F.
1924.
Sodium
hypochlorite
treatment;
the
germicidal
action
of
this
disinfectant
on
the
spores
of
American
foulbrood.
Cleanings
Bee
Cult.
52:
648-9.
Field
Studies
of
Sex
Attraction
in
the
-Boll
Weevil'
J.
R.
BRADLEY,
pt.,`
D.
F.
CLOWER,
and
J.
B.
GRAVES
Department
of
Entomology,
Louisiana
State
University,
Baton
Rouge
70803
Laboratory
investigations
(Keller
et
al.
1964,
Hardee
et
al.
1967a,
b)
have
shown
that
the
female
boll
weevil,
Anthonotntes
grandis
Boheman,
responds
to
an
airborne
sex
attractant
emitted
by
the
male.
Cross
and
Mitchell
(1966)
reported
observations
of
mating
behavior
of
the
boll
weevil
in
the
fi
eld.
Males
were
not
observed
respond-
ing
to
females.
However,
females
often
sought
males,
especially
from
downwind
positions,
from
distances
of
more
than
9
m.
This
study
was
conducted
to
acquire
further
informa-
tion
on
fi
eld
behavior
of
the
boll
weevil
in
response
to
the
sex
pheromone.
METHODS
AND
MATERIALS.
—A
trapping
technique
was
used
in
this
study.
The
trap
designed
(Fig.
1)
was
com-
posed
of
a
15X15
-cm
backboard
of
1.3
cm
plywood
placed
on
a
15X10
-cm
stage
of
0.63
-cm
plywood.
This
provided
a
stage
of
4.5
cm
projecting
at
a
right
angle
to
the
back-
board
on
each
side.
The
stage
trapped
any
weevils
that
were
able
to
free
themselves
from
the
backboard
and
would
have
otherwise
fallen
to
the
ground.
A
holding
chamber
was
provided
for
the
caged
weevil
by
drilling
a
2
-cm
hole
through
the
main
board.
The
captive
weevil
was
held
in
the
chamber
by
a
5
-cm
square
of
14
-mesh
screen
stapled
on
each
side
of
the
hole.
The
screen
al-
lowed
movement
of
air
through
the
cage
in
which
the
weevil
was
held,
thereby
facilitating
the
spread
of
any
volatile
material
into
the
environment.
Traps
were
mounted
on
1.3X5
-cm
stakes
which
varied
in
length
ac-
cording
to
cotton
-plant
height.
All
traps
were
coated
with
a
thin
layer
of
Stickem)
(97%
polymerized
butene,
iso-
butene,
and
butane;
Michel
and
Felton
Co.,
Emeryville,
Calif.)
.
Boll
weevils
used
in
this
study
were
obtained
from
lab-
oratory
cultures
of
Louisiana
origin.
The
weevils
were
reared
on
a
synthetic
medium
and
procedures
slightly
modified
from
those
described
by
Earle
et
al.
(1959)
.
Upon
emergence
from
pupal
cells
adults
were
sexed
and
held
in
individual
containers
(10-dr
shell
vials)
.
They
were
fed
fresh
debracted
cotton
squares
for
a
minimum
of
2
days
before
being
placed
in
traps
in
the
fi
eld.
The
study
was
conducted
at
several
locations
within
2
cotton
-growing
areas
of
Louisiana.
Three
trap
treatments
were
used;
boll
weevil
male,
boll
weevil
female,
and
a
control
which
did
not
contain
a
weevil.
At
each
location
a
randomized
arrangement
of
the
3
treatments
was
repli-
cated
several
times.
Traps
were
examined
and
caged
wee-
vils
were
replaced
once
or
twice
each
week.
Boll
weevils
1
Accepted
for
publication
May
26,
1968.
,
Present
address:
Department
of
Entomology,
North
Carolina
State
University,
Raleigh
27607.
collected
on
traps
were
taken
to
the
laboratory
for
dis-
section
and
sex
determination.
All
collections
from
cotton
prior
to
fruiting
were
made
during
April
and
May
of
1965, 1966,
and
1967,
when
overwintered
weevils
only
were
present
in
the
fi
eld
popu-
lations.
Collections
from
fruiting
cotton
were
made
dur-
ing
June,
July,
and
August,
with
I
exception.
In
April
1967,
traps
were
placed
in
fruiting
cotton
which
had
been
transferred
from
a
greenhouse
and
established
in
the
field.
Simultaneously,
traps
were
placed
in
a
fi
eld
of
seed
-
FIG.
1.
—Adhesive
-type
trap
utilized
to
study
sex
attrac-
tion.