Efficacy of monensin or amprolium in the prevention of hepatic coccidiosis in rabbits


Fitzgerald, P.R.

Journal of Protozoology 19(2): 332-334

1972


New Zealand White rabbits were fed monensin (Coban® Premix) or amprolium in pelleted feed as a prophylaxis against infection with Eimeria stiedai. Rabbits receiving monensin at 0.005, 0.01, or 0.02% concentrations in the feed did not become infected but ate only small amounts of pellets when the drug concentration was 0.02%. All rabbits given 0.02% amprolium in pelleted feed developed severe infections. Rabbits on "limited" (8 gm pellets per day) diets were susceptible to infection.

332
TREATMENT
OF
HEPATIC
COCCIDIOSIS
IN
RABBITS
and
qualitative
(4)
fluorescent
antibody
technics.
Among
the
previous
investigators
who
used
gel
diffusion
methods
for
antigenic
comparisons
of
E.
invadens
and
E.
histolytica
(for
pertinent
references,
see
2),
only
Talis
(12)
and
AliKhan
&
Meerovitch
(1)
were
able
to
demonstrate
any
cross
-reactivity
between
these
2
species.
According
to
Talis
(12),
a
few
lines
were
formed
in
reactions
involving
anti
-E.
invadens
serum
and
E.
histolytica
antigen;
no
lines
were
noted
in
reciprocal
re-
actions.
AliKhan
&
Meerovitch
(1),
however,
observed
a
few
precipitin
lines
in
both
of
the
above
reactions.
The
present
study
revealed
relatively
more
numerous
common
antigens
in
the
2
amebas.
The
greatest
significance
of
the
gel
diffusion
results
reported
in
this
paper
is
their
agreement
with
those
derived
previously
from
the
fluorescent
antibody
analysis
of
the
5
protozoa
(4).
The
conclusions
with
regard
to
the
antigenic
and
presumably
evolu-
tionary
relationships
among
Trichomonas,
Histomonas,
Dienta-
moeba
and
Entamoeba
adduced
on
the
basis
of
the
fluorescent
antibody
studies
are
fully
confirmed
by
the
present
investiga-
tion.
Moreover,
the
degree
of
cross
-reactivity
among
the
5
organisms
reflects
the
relative
numbers
of
antigens
they
share.
REFERENCES
1.
AliKhan,
Z.
&
Meerovitch,
E.
1968.
A
comparative
study
of
the
antigens
of
some
of
the
"histolytica-type"
strains
of
Entamoeba.
A
qualitative
and
quantitative
evaluation
of
antigens
by
indirect
hemagglutination,
gel
-precipitation,
and
immuno-
electrophoresis.
Am.
J.
Trop.
Med.
Hyg.
17,
528-39.
J.
PROIOZOOL.
19(2),
332-3M
(1972).
2.
Balamuth,
W.
&
Siddiqui,
W.
A.
1970.
Amoebas
and
other
intestinal
protozoa,
in
Jackson,
G.
J.,
Herman,
R.
&
Singer,
I.,
eds.,
Immunity
to
Parasitic
Animals,
Appleton
-Century
-Crofts,
New
York
2,
439-68.
3.
Dwyer,
D.
M.
1971.
Immunologic
analysis
by
gel
diffusion
of
effects
of
prolonged
cultivation
on
Histomonas
nzeleagridis
(Smith).
J.
Protozool.
18,
372-7.
4.
1972.
Analysis
of
the
antigenic
relationships
among
Trichomonas,
Histomonas,
Dientamoeba,
and
Entamoeba.
I.
Quan-
titative
fluorescent
antibody
methods.
J,
Protozool.
19,
316-25.
5.
Goldman,
M.
1954.
Use
of
fluorescein
-tagged
antibody
to
identify
cultures
of
Endamoeba
histolytica
and
Endamoeba
coli.
Am.
J.
Hyg.
59,
318-25.
6.
&
Honigberg,
B.
M.
1968.
Immunologic
analysis
by
gel
diffusion
technics
of
the
effects
of
prolonged
cultivation
on
Trichomonas
gallinae.
J.
Protozool.
15,
350-2.
7.
Honigberg,
B.
M.,
Friedman,
H.
&
Stepkowski,
S.
1971.
Effect
of
different
immunization
procedures
on
agglutination
and
precipitation
reactions
of
Trichomonas
gallinae.
J.
Parasit.
57,
363-9.
8.
Lang,
C.
A.
1958.
Simple
microdetermination
of
Kjeldahl
nitrogen
in
biological
materials.
Anal.
Chem.
30,
1692-4.
9.
Lowry,
0.
H.,
Rosebrough,
N.
J.,
Farr,
A.
L.
&
Randall,
R.
J.
1951.
Protein
measurement
with
the
folin
phenol
reagent.
J.
Biol.
Chem.
193,
265-75.
10.
Siddiqui,
W.
A.
&
Balamuth,
W.
1965.
Serological
com-
parison
of
selected
parasitic
and
free-living
amoebae
in
vitro,
using
diffusion
-precipitation
and
fluorescent
-antibody
technics.
J.
Protozool.
12,
175-82.
11.
Talis,
B.
1964.
Immunological
investigation
of
amoebiasis
by
the
immunofluorescent
method.
J.
Protozool.
11
(Suppl.),
44.
12.
1967.
Antigenic
relationships
among
strains
of
Entamoeba
histolytica,
Dientamoeba
fragilis,
and
Entamoeba
in-
vadens.
J.
Protozool.
14
(Suppl.),
44.
Efficacy
of
Monensin
or
Amprolium
in
the
Prevention
of
Hepatic
Coccidiosis
in
Rabbits
PAUL
R.
FITZGERALD
College
of
Veterinary
Medicine,
University
of
Illinois,
Urbana,
Ill.
61801
SYNOPSIS.
New
Zealand
White
rabbits
were
fed
monensin
(Coban®
Premix)
or
amprolium
in
pelleted
feed
as
a
prophylaxis
against
infection
with
Eimeria
stiedai.
Rabbits
receiving
monensin
at
0.005,
0.01,
or
0.02%
concentrations
in
the
feed
did
not
become
infected
but
ate
only
small
amounts
of
pellets
when
the
drug
concentration
was
0.02%.
All
rabbits
given
0.02%
amprolium
in
pelleted
feed
developed
severe
infections.
Rabbits
on
"limited"
(8
gm
pellets
per
day)
diets
were
susceptible
to
infection.
Index
Key
Words:
Eimeria;
stiedai;
coccidiosis
in
rabbits,
treatment
of;
monensin;
amprolium.
A
LTHOUGH
several
drugs
have
been
used
in
attempts
to
control
coccidial
infections
caused
by
E.
stiedai
in
ex-
perimental
rabbits,
none
has
been
completely
successful
(3,
6).
This
protozoon
is
a
frequent
cause
of
a
significant
disease
prob-
lem
in
rabbitries.
A
logical
method
for
controlling
the
disease
is
to
incorporate
an
effective
drug
in
the
feed.
This
has
been
done
commercially,
but
the
drugs
usually
incorporated
are
not
effective
against
E.
stiedai.
The
present
paper
describes
the
re-
sults
obtained
by
incorporating
a
well
known
drug,
effective
against
other
coccidia
(amprolium-Merck
&
Co.),
or
a
new,
less
well
known
antibiotic
(monensin,
Coban®
Premix
-Eli
Lilly
&
Co.),
in
the
rabbits'
food.
MATERIALS
AND
METHODS
Fifty
10
-week-old
New
Zealand
white
rabbits
were
used
in
2
experiments.
During
the
experimental
period
the
animals
were
kept
individually
in
stainless
steel,
self
-flushing
cages
or
in
galvanized
cages
with
dropping
pans.
Inoculated
rabbits
were
given
50,000
or
225,000
sporulated
oocysts
of
E.
stiedai
by
intubation.
Individual
fecal
examinations
using
McMaster
counting
chambers
were
made
weekly
for
2
weeks
after
inocula-
tion,
and
then
every
other
day
until
the
animals
were
sacrificed.
All
rabbits
were
weighed
weekly
and
at
the
end
of
the
experi-
ments.
Liver
weights
were
determined
to
ascertain
the
extent
of
enlargement.
In
each
experiment
rabbits
of
similar
age
and
weight,
raised
in
our
rabbitry,
were
separated
into
5
groups
of
5
each.
In
the
1st
experiment,
rabbits
in
groups
1
and
2
were
given
monensin
in
pelleted
feed
at
0.01
or
0.02%
(w/w)
concentrations.
Those
in
group
3
were
given
amprolium
in
pelleted
feed
at
a
concentration
of
0.02%
w/w.
Rabbits
in
control
group
4
were
infected
with
the
parasites
but
not
medicated,
while
those
in
control
group
5
were
neither
infected
nor
medicated.
Each
inoculated
rabbit
was
given
50,000
sporulated
oocysts
by
in-
TREATMENT
OF
HEPATIC
COCCIDIOSIS
IN
RABBITS
333
4,000
3,500-
3,00
Weight
in
grams
2,500-
2,000-
1,500
1
I
I
I
Groups
I,
2,
3
Medicated
Feed
225,00
I
I
45,00
20
60,000
100,0
45,00
a
100,000
Inoculation,
50,000
Oocysts
50,00
1
1
1
1
1
0
I
2
3
4
5
Weeks
25,000
6
Gr.
5
Gr.
4
Gr.
3
Gr.
2
Gr.
I
Group
1=
Inoculated
+
monensin
@
0.01%
in
feed
Group
2=
Inoculated
+
monensin
@
0.02%
in
feed
Group
3=
Inoculated
+
amprolium
@
0.02%
in
feed,
severe
hepatic
infections.
Group
4=
Inoculated,
unmedicated,
severe
hepatic
infections
Group
5=
Uninoculated,
unmedicoted,
controls
Fig.
1.
The
effect
of
Eimeria
stiedai
infections
on
weight
and
oocyst
discharge
in
rabbits
fed
monensin
or
amprolium
in
pelleted
feed.
Average
oocysts
per
gram
of
feces
for
all
rabbits
in
group.
Experiment
1.
tubation.
The
2nd
experiment
was
similar
except
that
crystal-
line
monensin
was
used
in
the
pelleted
feed
at
0.005
or
0.01%
(w/w)
concentrations.
Since
amprolium
was
not
effective
in
the
1st
experiment
it
was
not
used
in
the
second;
instead,
the
3rd
group
was
given
a
limited
amount
of
unmedicated
feed
per
day
(about
8
gm
of
pellets)
so
that
the
rabbits
were
main-
tained
at
a
negative
metabolic
level.
Inoculated,
unmedicatecl
and
uninoculated,
unmedicated
controls
were
similar
to
those
of
experiment
1.
Each
inoculated
rabbit
was
given
225,000
sporulated
oocysts
by
intubation.
Pelleted
commercial
rabbit
feed
(
Ralston
-Purina
rabbit
chow,
unmedicated)
was
finely
ground;
the
drugs,
in
appropriate
portions,
were
thoroughly
mixed
into
the
ground
feed
that
was
then
repelleted
into
5
mm
diameter
pellets.
All
rabbits
given
drug
were
started
on
medicated
feed
2
or
7
days
before
in-
oculation.
RESULTS
In
the
1st
experiment,
rabbits
given
pellets
containing
0.01%
monensin
ate
a
moderate
number
of
pellets,
but
those
given
pellets
with
0.02%
monensin
ate
only
small
quantities.
A
marked
decline
in
the
total
average
weight
of
these
animals
was
noted
(Fig.
1).
None
of
them,
however,
discharged
coc-
cidia
in
the
feces
at
any
time.
Rabbits
given
0.02%
amprolium
in
the
pelleted
feed,
ate
the
medicated
feed
well
and
increased
slightly
in
weight.
All
of
them,
however,
developed
severe
infections
and
discharged
large
numbers
of
oocysts
in
the
feces.
Inoculated,
unmedicated
controls
and
uninoculated,
unmedi-
cated
controls
had
similar
growth
rates
but
the
inoculated,
un-
medicated
controls
discharged
large
numbers
of
oocysts
in
the
feces,
went
off
feed
and,
were
beginning
to
have
clinical
signs
of
infection
when
killed
(
Fig.
1).
The
rabbits
given
amprolium,
as
well
as
the
inoculated,
unmedicated
controls,
4,000--1
3,500
Groups
1
2
Medicated
Feed
3,000
Weight
in
grams
2,500
2,000
I
I
I
1,500
0
I
2
3
2000
Inoculation
225,000
Oocysts
>3,000,000
Gr
5
Or
I
m0,000.00
,00
Gr
4
Gr
2
200,000
.10,000,000
Gr
3
Necropsied
I
I
I
I I
4
5
6
7
6
9
Weeks
Group
1.
Inoculated,
@
0.005
%
monensin
in
pelleted
food
Group
2.
Inoculated,
@
0.01%
monensin
in
pelleted
food
Group
3.
Inoculated,
starvation
diet,
severe
hepatic
infections
Group
4.
Inoculated,
unmedicoted,
severe
hepatic
infections
Group
5.
Uninoculated,
unmedicated,
controls
Fig.
2.
The
effect
of
Eimeria
stiedai
infections
upon
weight
and
oocyst
discharge
in
rabbits
fed
monensin
in
pelleted
feed
or
maintained
on
a
limited
diet
(8
g
of
unmedicated
pellets
per
day).
Average
oocysts
per
gram
of
feces
for
all
rabbits
in
group.
Ex-
periment
2.
had
severe
liver
infections
and
on
necropsy
their
gall
bladders
were
found
full
of
oocysts.
Uninoculated,
unmedicated
controls
were
free
of
infections.
The
average
number
of
oocysts
dis-
charged
in
feces
(on
a
weekly
basis)
is
indicated
in
Fig.
1.
Rabbits
given
monensin
at
the
0.005
or
(1.01%
concentrations
(experiment
2)
gained
slightly
or
even
lost
weight,
while
all
those
on
the
limited
diet
lost
weight
(Fig.
2).
Infected
rabbits
that
received
no
medication
maintained
their
weight
for
the
first
2
weeks,
but
then
lost
weight
as
infections
developed.
Un-
inoculated,
untreated
rabbits
continued
to
gain
weight
through-
out
the
experimental
period.
Severe
infections
developed
in
all
infected,
unmedicated
animals
of
groups
3
and
4,
but
no
infections
developed
in
medicated
or
uninoculated
animals
of
groups
1,
2,
or
5.
The
average
numbers
of
oocysts
dis-
charged
(on
a
weekly
basis)
are
shown
in
Fig.
2.
TABLE
1.
Liver
and
total
body
weights
of
monensin-
or
amprolium-
treated,
and
of
untreated
rabbits
6
weeks
after
inoculation
with
50,000
sporulated
oocysts
of
Eimeria
stiedai,
and
of
uninfected
rabbits.
Rabbit
Number
1
2
3
Group
Number*
4
5
1
82f(2925)$
29(1220)1
113(3720)
2
66
(2245)
58(2325)
114(3482)
3
53
(2130)
57(2388)
121(3378)
4
30
(1325)
58(2525)
121(3548)
5
75
(3395)
84(2778)
120(3674)
150(4325)11
99(3650)
117(3700)
125(3580)
"140(3980)
138(4130)
105(3430)
Mean
61
(2404)
57(2247)
118(3560) 118(3827)
130(3829)
*
Groups
1-4,
inoculated
with
E.
stiedai;
groups
1
and
2,
treated
with
0.01
or
0.02%
monensin,
respectively;
group
3,
treated
with
0.02%
amprolium
(both
drugs
fed
in
pelleted
feed)
;
group
4,
untreated;
group
5,
uninfected
and
untreated.
t
Liver weight
in
grams.
t
Total
body
weight
in
grams.
§Necropsied
early;
no
evidence
of
hepatic
coccidiosis.
Necropsied
early;
weights
not
significant.
**
Died
10
days
before
end
of
experiment;
moderate
hepatic
coccidiosis.
334
TREATMENT
OF
HEPATIC
COCCIDIOSIS
IN
RABBITS
TABLE
2.
Liver
and
total
body
weights
of
monensin-treated
and
untreated
rabbits
4
weeks
after
inoculation
with
225,000
sporulated
oocysts
of
Eimeria
stiedai,
and
of
uninfected
rabbits.
Rabbit
Number
Group
Number*
1
2
3
4
5
1
98t(2998)*
44(2349)
142(1939)
357(2957)
108(3628)
2
104
(3224)
84(2891)
205(2289)
553(2496)
93(3399)
3
68
(2943)
57(2640)
130(1816)
437(2975)
83(3605)
4
118
(3668)
60(2483)
176(2398)
227(2488)11
83(3605)
5
110
(1284)/
71(2547)
147(1650)
221(2537)**83(3670)
Mean
97
(2823)
63(2582)
160(2018)
359(2690)
90(3573)
*
Groups
1-4,
inoculated
with
E.
stiedai;
groups
1
and
2,
treated
with
0.01
and
0.005%
monensin
in
pelleted
feed;
group
3,
untreated
and
kept
on
a
limited
diet
(8
grams
of
unmedicated
pellets
per
day)
;
group
4,
untreated;
group
5,
uninfected
and
untreated.
t
Liver
weight
in
grams.
t
Total
body
weight
in
grams.
§
Died
20
days
after
inoculation;
no
gross
pathology
or
coccid-
iosis
detected
at
necropsy.
II
Died
28
days
after
inoculation;
severe
hepatic
coccidiosis
found
at
necropsy.
**
Died
21
days
after
inoculation;
severe
hepatic
coccidiosis
found
at
necropsy.
Tables
1
and
2
summarize
the
extent
of
enlargement
of
the
livers
caused
by
severe
infections
with
E.
stiedai
in
both
ex-
periments.
The
livers
enlarged
from
a
normal
of
2-3%
to
22%
of
the
total
body
weight
in
some
severely
infected
animals.
DISCUSSION
In
the
1st
experiment
the
question
arose
if
partial
starvation
(because
of
refusal
by
the
animals
of
pelleted
feed
containing
monensin)
or
extremely
small
amounts
of
the
antibiotic
pre-
vented
infections
in
rabbits
of
2
of
the
groups.
The
2nd
experi-
ment
was
designed
to
determine
whether
partial
starvation
had
any
significant
effect
upon
susceptibility
to
infection.
The
results
indicated
that
greatly
reduced
food
intake
had
little
or
no
effect
upon
establishment
of
coccidia
in
the
liver.
All
of
the
inoculated,
unmedicated
rabbits
became
infected
and
were
found
to
have
enlarged
livers
on
necropsy.
The
results
of
these
experiments
indicate
that
monensin,
at
the
levels
used,
is
highly
effective
in
controlling
liver
coccidiosis.
Both
experiments
revealed
also
that
pellets
containing
monensin
were
poorly
to
moderately
acceptable
to
rabbits
which
ate
few
medicated
pellets
containing
0.02%
but
accepted
moderately
well
pellets
containing
0.01%
or
0.005%
of
the
antibiotic.
Weight
gains
and
the
amount
of
food
ingested
were
proportional
to
the
amount
of
monensin
in
the
feed.
No
information
was
obtained
about
what
stages
of
the
para-
site
were
affected
by
monensin.
Shumard
&
Callender
(9)
found
the
antibiotic
to
be
highly
effective
against
poultry
coccidia
and
suggested
that
the
sporozoites,
trophozoites,
or
early
schizonts
are
affected
(8).
Estrada
-0
et
al
(4)
reported
that
monensin
interferes
with
ion
(particularly
potassium
ions)
transport
in
mitochondria.
One
can
only
speculate
that
monensin
affects
similarly
translocation
of
cations
in
the
parasites.
The
failure
of
amprolium
to
prevent
infections
in
the
livers
of
rabbits
was
surprising.
It
has
been
found
to
be
highly
effec-
tive
against
intestinal
coccidia
of
poultry
(1,
7,
11).
Hammond
et
al.
(5)
and
Slater
et
al
(10)
have
summarized
recent
infor-
mation
on
the
drug's
activity
in
cattle.
Cvetkovic
&
Tomanovic
(2)
claimed
it
was
effective
against
intestinal
forms
of
rabbit
coccidia
when
fed
in
granular
feed.
They
mentioned
E.
stiedai
in
their
paper,
but
did
not
indicate
whether
it
was
affected
by
amprolium.
Pelleting
involves
a
certain
amount
of
heating
and
pressure
which
may
have
caused
loss
or
alteration
of
its
activity
against
this
parasite.
REFERENCES
1.
Bond,
D.
S.
&
Edgar,
S.
A.
1961.
Efficacy
of
a
new
coccidio-
static
drug
for
the
control
of
coccidiosis
in
chickens
and
turkeys.
Abst.
So.
Agric.
Workers
Mtg.
1961,
1-2.
2.
Cvetkovic,
L.
&
Tomanovic,
B.
1967.
[Investigation
of
Amprolium
in
prophylaxis
of
intestinal
coccidiosis
in
rabbits.]
Veterinarski
Glasnik.
7,
607-12.
3.
Dfirr,
U.
&
Schrecke,
W.
1970.
Sammelreferat:
Die
Chemo-
therapie
der
Kaninchenkokzidiosen.
Deut.
Tieraerztl.
Wochschr.
77,
435-41.
4.
Estrada
-O,
S.,
Rightmire,
B.
&
Lardy,
H.
A.
1967.
Anti-
biotics
as
tools
for
metabolic
studies.
XI.
Specific
inhibition
of
ion
transport
in
mitochondria
by
the
Monensins.
Antimicrobial
Agents
Chemotherapy
1967,
279-88.
5.
Hammond,
D.
M.,
Fayer,
R.
&
Miner,
M.
L.
1966.
Am-
prolium
for
control
of
experimental
coccidiosis
in
cattle.
Am.
J.
Vet.
Res.
27,
199-206.
6.
Lammler,
G.
&
Diirr,
U.
1970.
Zur
Chemotherapie
der
Leberkokzidiose
des
Kaninchens
mit
Sulfadimethoxin-Diaveridin.
Berl.
Muench.
Tieraerztl.
Wochschr.
82,
480-4.
7.
Morrison,
W.
D.,
Ferguson,
A.
E.,
Connell,
M.
C.
&
Mc-
Gregor,
J.
K.
1961.
The
efficacy
of
certain
coccidiostats
against
mixed
avian
coccidial
infections.
Avian
Dis.
5,
222-8.
8.
Shumard,
R.
F.
Personal
Communication.
1971.
9.
&
Callender,
M.
E.
1967.
Monensin,
a
new
bio-
logically
active
compound.
VI.
Anticoccidial
activity.
Antimicrobial
Agents
Chemotherapy
1967,
367-77.
10.
Slater,
R.
L.,
Hammond,
D.
M.
&
Miner,
M.
L.
1970.
Eimeria
bovis:
Development
in
calves
treated
with
thiamine
met-
abolic
antagonist
(Amprolium)
in
feed.
Trans.
Am.
Micr.
Soc.
89,
55-65.
11.
Strout,
R.
1961.
Amprolium
for
replacements.
Tenth
Ann.
Univ.
New
Hampshire
Poultry
Health
Conf.
1961,
43-7.
THIRD
INTERNATIONAL
CONGRESS
OF
PARASITOLOGY
(1974)
The
Third
International
Congress
of
Parasitology
will
be
held
at
Munich,
Germany
(Kongress-Zentrum,
Messegelande),
25-31
August,
1974.
The
Congress
is
sponsored
by
the
World
Federation
of
Parasitologists,
of
which
the
Society
of
Proto-
zoologists
is
a
member.
The
Congress
is
being
organized
by
a
committee
of
the
Deutsche
Gesellschaft
filr
Parasitologie.
Dr.
G.
Piekarski
is
the
President,
Dr.
G.
Lammler
is
the
Secretary
General,
and
Drs.
H.
E.
Krampitz
and
J.
Lamina
constitute
the
Local
Committee;
the
Program
Committee
includes
Drs.
J.
Boch,
F.
Kuhlow,
and
E.
Scholtyseck.
The
Committee
has
asked
that
anyone
wishing
further
information
write
to
Dr.
G.
Liimmler,
Secretary
General,
Institute
for
Parasitology
of
the
University,
R.
Buchheim-Str.
4,
D
6300
Giessen,
Germany.
D.
M.
HAMMOND,
Representative
of
the
Society
of
Protozoologists
to
the
World
Federation
of
Parasitologists.