A multilocation clinical trial in lactating dairy cows affected with clinical mastitis to compare the efficacy of treatment with intramammary infusions of a lincomycin/neomycin combination with an ampicillin/cloxacillin combination


Deluyker, H.A.; Chester, S.T.; Oye, S.N. van

Journal of Veterinary Pharmacology and Therapeutics 22(4): 274-282

1999


The efficacies of intramammary infusions in quarters affected with clinical mastitis of a formulation containing 330 mg lincomycin and 100 mg neomycin in a 10-ml aqueous solution (LINCOCIN FORTE S.) and a formulation containing 75 mg ampicillin and 200 mg cloxacillin in an oil suspension (AMPICLOX) were compared in lactating cows. This study was designed as a multicentre clinical trial involving investigators in France, Germany and Belgium and carried out according to the European Commission guidelines on Good Clinical Practices. Cows in the herds were monitored for clinical mastitis. When evidence of clinical mastitis was detected in a single quarter, a pretreatment milk sample was collected from the affected quarter. After milk sampling, the cow was assigned to one of the 2 treatment groups at random and treated with an intramammary infusion of one syringe of either LINCOCIN FORTE S or AMPICLOX for 3 successive milkings in the mastitic quarter. At 4-5, 13-15 and 20-22 days after first infusion, the veterinarian returned to the farm to conduct a clinical examination and collect milk samples from the affected quarter. Milk samples were cultured for mastitis organisms and somatic cell count (SCC) was measured. Following a 10-month study period, 256 cases were enrolled in the study. 232 and 189 cases were examined for clinical cure and for clinical-plus-bacteriological cure, respectively. The proportions of cases cured clinically and cured clinically-plus-bacteriologically were compared between the treatment groups. Somatic cell count differences between treatment groups were also tested. The clinical cure rate for LINCOCIN FORTE S (62.5%) was significantly better than for AMPICLOX (51.8%) (P = 0.035). The clinical-plus-bacteriological cure rate was also significantly better for LINCOCIN FORTE S (38.1%) than for AMPICLOX (21.7%) (P = 0.005). Among bacteriologically cured cases, the SCC declined in both treatment groups but the SCC was significantly higher for the AMPICLOX group than for the LINCOCIN FORTE S group (P = 0.036). In conclusion, clinical cure rate, clinical-plus-bacteriological cure rate, and SCC level were significantly better with LINCOCIN FORTE S than for AMPICLOX.

J.
vet.
Pharmacol.
Therap.
22,
274-282,
1999.
THERAPEUTICS
A
multilocation
clinical
trial
in
lactating
dairy
cows
affected
with
clinical
mastitis
to
compare
the
efficacy
of
treatment
with
intramammary
infusions
of
a
lincomycin/neomycin
combination
with
an
ampicillin/cloxacillin
combination
Deluyker,
H.
A.,
Chester,
S.
T.,
Van
Oye,
S.
N.
A
multilocation
clinical
trial
in
lactating
dairy
cows
affected
with
clinical
mastitis
to
compare
the
efficacy
of
treatment
with
intramammary
infusions
of
a
lincomycin/neomycin
combina-
tion
with
an
ampicillin/cloxacillin
combination.
J.
vet.
Pharmacol.
Therap.
22,
274-282.
A
study
was
conducted
to
compare
the
efficacy
in
lactating
dairy
cows
of
intramammary
infusions
in
quarters
affected
with
clinical
mastitis
between
a
formulation
containing
330
mg
lincomycin
and
100
mg
neomycin
in
a
10-mL
aqueous
solution
(LINCOCIN
®
FORTE
S,
Pharmacia
&
Upjohn)
and
a
formulation
containing
75
mg
ampicillin
and
200
mg
cloxacillin
in
an
oil
suspension
(AMPICLOX
Tm
,
Pfizer
Animal
Health).
This
study
was
designed
as
a
multicentre
clinical
trial
involving
investigators
in
France,
Germany
and
Belgium
and
carried
out
according
to
the
European
Commission
guidelines
on
Good
Clinical
Practices.
Cows
in
the
herds
were
monitored
for
clinical
mastitis.
When
evidence
of
clinical
mastitis
was
detected
in
a
single
quarter,
a
pretherapy
milk
sample
was
collected
from
the
affected
quarter.
After
milk
sampling,
the
cow
was
assigned
to
one
of
the
two
treatment
groups
at
random
and
treated
with
an
intramammary
infusion
of
one
syringe
of
either
LINCOCIN
®
FORTE
S
or
AMPICLOX
Tm
for
three
successive
takings
in
the
mastitic
quarter.
At
4-5,
13-15
and
20-22
days
after
first
infusion,
the
veterinarian
returned
to
the
farm
to
conduct
a
clinical
examination
and
collect
milk
samples
from
the
affected
quarter.
Milk
samples
were
cultured
for
the
presence
of
mastitis
organisms
and
somatic
cell
count
(SCC)
was
measured.
Following
a
10-month
study
period,
256
cases
were
enrolled
in
the
study.
A
total
of
232
and
189
cases
were
analysed
for
clinical
cure
and
for
clinical-plus-
bacteriological
cure,
respectively.
The
proportions
of
cases
cured
clinically
and
cured
clinically-plus-bacteriologically
were
compared
between
the
two
treat-
ment
groups.
Somatic
cell
count
differences
between
treatment
groups
were
also
tested.
The
clinical
cure
rate
for
LINCOCIN
®
FORTE
S
(62.5%)
was
significantly
better
than
for
AMPICLOX
Tm
(51.8%)
(P
=
0.035).
The
clinical-plus-bacter-
iological
cure
rate
was
also
significantly
better
for
LINCOCIN
®
FORTE
S
(38.1%)
than
for
AMPICLOX
TM
(21.7%)
(P
=
0.005).
Among
bacteriologically
cured
cases,
the
SCC
declined
in
both
treatment
groups
but
the
SCC
was
significantly
higher
for
the
AMF'ICLOX
Tm
group
than
for
the
LINCOCIN
®
FORTE
S
group
(P
=
0.036).
In
conclusion,
clinical
cure
rate,
clinical-plus-bacteriological
cure
rate,
and
SCC
level
were
significantly
better
with
LINCOCIN
®
FORTE
S
than
for
AMPICLOX
Im
.
(Paper
received
3
April
1998;
accepted
for
publication
19
March
1999)
H.
A.
Deluyker,
Worldwide
Animal
Health
Clinical
Development,
Pharmacia
&
Upjohn,
Rijksweg
12,
2870
Puurs,
Belgium.
H.
A.
DELUYKER*
S.
T.
CHESTERt
&
S.
N.
VAN
OYE*
*Worldwide
Animal
Health
Clinical
Development,
Pharmacia
&
Upjohn,
Rijksweg
12,
2870
Puurs,
Belgium;
-
Worldwide
Animal
Health
Biostatistics
and
Animal
Performance;
Pharmacia
&
Upjohn,
5300
N.
28th
Street,
Richland,
MI
49083,
U.S.A.
C)1999
Blackwell
Science
Ltd
274
Clinical
mastitis
efficacy
275
INTRODUCTION
Clinical
mastitis
is
an
important
disease
in
the
dairy
industry
(Wilesmith
et
al.,
1986;
Deluyker,
1991).
In
any
herd,
clinical
mastitis
can
be
caused
by
a
variety
of
microorganisms.
Epidemiological
surveys
have
identified
Staphylococcus
(Staph.)
aureus,
Streptococcus
(Strep.)
uberis,
and
Escherichia
coli
as
the
most
frequently
isolated
causal
organism
of
clinical
mastitis
(Barkema
et
al.,
1998).
These
encompass
both
Gram
positive
and
Gram
negative
bacteria,
and
the
causal
organism
can
not
be
determined
based
on
clinical
signs.
Often
broad-spectrum
antibacterial
therapy
is
used
to
treat
such
cases
immediately
rather
than
wait
until
culture
results
have
identified
the
causative
organism.
LINCOCIN
®
FORTE,
(Pharmacia
&
Upjohn,
Puurs,
Belgium)
is
a
currently
marketed
intramammary
pathogen-free
infusion
containing
lincomycin
and
neomycin
as
active
ingredients.
In
vitro,
streptococci
are
sensitive
to
lincomycin,
coliforms
to
neomycin,
whereas
Staph.
aureus
is
sensitive
to
lincomycin
and
neomycin.
LINCOCIN
®
FORTE
was
reformulated
so
as
to
meet
European
Pharmacopoeia
manufacturing
standards
on
sterility.
In
addition,
the
quantity
and
the
ratio
of
the
active
ingredients
were
revised.
This
new
formulation
(LINCOCIN
®
FORTE
S)
contains
330
mg
lincomycin
and
100
mg
neomycin
per
syringe.
Compounds
used
for
the
treatment
of
clinical
mastitis
are
allowed
on
the
market
only
after
their
quality,
efficacy
and
safety
for
the
target
animal,
humans,
and
the
environment
have
been
demonstrated.
The
objective
of
this
study
was
to
evaluate
the
efficacy
of
LINCOCIN
®
FORTE
S
in
lactating
dairy
cows
with
clinical
mastitis
through
comparison
of
the
cure
rates
with
those
obtained
with
a
combination
of
ampicillin
and
cloxacillin
(AMPICLOK
lm
,
Pfizer
Animal
Health,
Sandwich,
UK),
a
widely
marketed
combination
for
the
treatment
of
clinical
mastitis.
MATERIALS
AND
METHODS
The
study
was
conducted
according
to
the
European
Commis-
sion
guidelines
on
Good
Clinical
Practices
for
the
conduct
of
clinical
trials
for
veterinary
medicinal
products
(CVMP,
1994)
and
those
on
the
conduct
of
clinical
trials
to
assess
the
efficacy
of
veterinary
medicinal
products
administered
via
the
teat
duct
to
lactating
cows
for
the
treatment
of
clinical
mastitis
(CVMP,
1993).
Herds
with
a
regular
history
of
mastitis
were
included.
They
were
selected
such
that
Staph.
aureus,
Strep.
uberis,
and
E.
coli
mastitis
cases
were
expected
to
occur
during
the
study
period.
Herds
with
a
high
incidence
of
damaged
teats
or
teat
ends
were
not
used.
Enrollment
of
clinical
mastitis
cases
A
quarter
was
considered
to
have
signs
of
clinical
mastitis
if
the
quarter
milk
was
visually
abnormal
(flakes,
clots,
or
watery
secretion)
or
if
swelling,
heat,
or
tenderness/pain
on
palpation
were
present.
In
the
selected
herds,
a
trained
dairyman
or
dairy
employee
monitored
cows
for
abnormal
milk
by
routine
use
of
a
strip
cup
or
any
other
method
used
in
the
herd
to
ascertain
visual
appearance
of
quarter
milk
prior
to
milking
initiation.
When
evidence
of
clinical
mastitis
was
detected
the
veterinarian
was
notified.
Only
cows
with
clinical
mastitis
in
a
single
quarter
were
enrolled.
Also
excluded
were
cases
that
pertained
to
quarters
with
damaged
teats
or
teat
ends;
cows
with
hyperacute
mastitis
(i.e.
gangrenous
mastitis);
cows
with
systemic
illness
(pyrexia,
anorexia,
depression,
or
dehydration);
cows
that
had
calved
less
than
48
h
prior
to
enrollment;
cows
that
had
received
antibacterial
or
anti-inflammatory
therapy
less
than
30
days
prior
to
enrollment;
and
cows
previously
enrolled
in
the
study.
From
the
affected
quarter,
two
pretherapy
milk
samples
were
collected
aseptically
into
sterile
vials
by
the
veterinarian.
One
was
used
for
bacteriological
identification
and
one
for
SCC
determination.
Study
design
and
treatment
Enrolled
cows
in
a
herd
were
grouped
in
blocks
of
two
animals,
by
entry
into
the
study.
Treatments
were
randomly
assigned
within
the
blocks
using
a
predetermined
randomization
sche-
dule.
After
collection
of
the
pretherapy
milk
samples,
the
mastitic
quarter
was
immediately
treated
by
the
veterinarian
with
one
syringe
of
either
LINCOCIN
®
FORTE
S
or
a
combination
of
75
mg
ampicillin
and
200
mg
cloxacillin
(AMPICLOK
Im
).
The
follow-up
treatments
were
administered
by
a
trained
dairyman
or
dairy
employee.
The
infected
quarter
was
infused
with
one
syringe
three
times,
at
the
end
of
three
successive
milkings.
Sterile
alcohol
pads
for
teat
disinfection
were
provided
with
each
syringe.
The
labels
on
the
two
formulations
identified
the
treatment
as
A
and
B
so
as
to
disguise
their
identities.
The
personnel
involved
in
the
bacteriological
diagnoses
and
in
the
SCC
determination
were
blinded
as
to
the
nature
of
the
treatment
the
cow
had
received.
Response
measurement
At
4-5,
13-15
and
20-22
days
after
first
infusion,
the
veterinarian
returned
to
the
farm
to
conduct
a
clinical
examina-
tion
and
to
aseptically
collect
two
milk
samples
from
the
affected
quarter
for
bacteriological
identification
of
mastitis
organisms
and
SCC
measurement.
Generally,
the
milk
samples
for
bacter-
iological
identification
were
refrigerated
and
delivered
to
the
laboratory
within
24
h.
If
this
was
not
possible,
then
the
samples
were
frozen
and
delivered
to
the
laboratory
within
a
week.
For
the
bacteriological
identification,
the
National
Mastitis
Council
Guidelines
(1990)
were
followed.
It
aimed
at
differ-
entiating
the
following
causal
microorganisms:
Staph.
aureus,
non-aureus
staphylococci,
Strep.
agalactiae,
Strep.
dysgalactiae,
Strep.
uberis,
other
Esculine-positive
streptococci,
E.
coli,
Klebsiella
species,
other
enterobacteriaceae,
Actinomyces
pyogenes,
Bacillus
cereus,
and
other
organisms.
If
more
than
two
different
bacterial
species
were
observed
in
the
sample
it
was
considered
contaminated,
unless
there
were
more
than
five
colonies
of
Staph.
aureus,
Strep.
agalactiae,
or
Strep
dysgalactiae.
©1999
Blackwell
Science
Ltd,
J.
vet.
Pharmacol.
Therap.
22,
274-282
276
H.
A.
Deluyker
et
al.
The
SCC
of
each
milk
sample
was
measured
using
a
Fossomatic
counter
(Foss
Electric,
Hillered,
Denmark).
Cure
definition
A
case
was
considered
a
clinical
cure
if,
at
4-5
days
after
first
treatment,
milk
in
the
affected
quarter
was
normal,
other
clinical
signs
of
mastitis
had
disappeared,
and
no
relapse
occurred
during
the
20-22-day
observation
period.
Bacteriological
results
were
interpreted
such
that
quarters
where
the
same
bacterium
species
was
cultured
before
and
at
any
sampling
time
after
treatment
onset
(4-5,
13-15
or
20-22
days)
were
considered
a
treatment
failure
while
those
with
growth
in
the
initial
sample
and
no
growth
or
growth
of
different
bacteria
after
treatment
onset,
were
considered
a
treatment
cure.
A
case
was
considered
cured
clinically-plus-bacteriologically
if
the
enrolled
quarter
was
cured
clinically
as
well
as
bacteriologically.
A
clinically-plus-bacteriologically
cured
case
was
considered
infected
with
another
organism
if
a
bacteria,
different
from
the
originally
isolated
species,
was
isolated
at
any
of
the
three
post-
treatment
sampling
times.
The
rate
of
these
occurrences
was
calculated
by
treatment
group
for
the
species
most
frequently
isolated
pretreatment.
Statistical
analysis
of
cure
rates
The
proportion
of
cured
cases
within
each
herd
by
treatment
group
was
transformed
using
the
Freeman-Tukey
angular
transformation.
These
values
were
subsequently
analysed
using
a
randomized
block
design
with
blocks
(herds)
considered
a
random
effect
in
a
weighted
ANOVA
with
the
weights
n
+
'/2
where
n
=
number
of
cases
treated
within
the
herd
by
treatment
group
(PROC
MIXED,
SAS).
To
compare
the
two
treatments,
the
following
hypothesis
was
tested.
Let
7C11
and
n
a
be
the
cure
rates
for
LINCOCIN
®
FORTE
S
and
AMPICLOX
Tm
,
respectively.
The
figure
below
modelled
the
difference
between
the
cure
rates,
n
ir
n
a
.
Difference
in
cure
rates
between
experimental
compound
and
positive
control
4—AMPICLOX
Tm
better—
—LINCOCIN®
FORTE
S
better—)
4—Not
different
—)1
—LINCOCIN
®
FORTE
S
not
worse
than
AMPICLOX
Tm
—)
-A
0
A
n
rna
The
region to
the
right
of
zero
corresponded
to
those
situations
in
which
LINCOCIN
®
FORTE
S
was
superior
to
AMPICLOX
Tm
,
while
the
region
to
the
left
of
zero
corresponded
to
AMPICLOX
TM
being
superior
to
LINCOCIN
®
FORTE
S.
In
addition,
the
parameter
A
represented
the
'difference'
threshold
such
that
when
the
difference
fell
outside
the
interval
-A
to
A,
then
the
two
treatments
were
considered
to
be
truly
different.
To
test
whether
LINCOCIN
®
FORTE
S
was
comparable
with
AMPICLOX
Tm
,
the
following
two
hypotheses
were
combined
in
a
single
overall
hypothesis:
1
whether
LINCOCIN
®
FORTE
S
was
better
than
AMPICLOX
Tm
;
2
whether
the
two
treatments
were
not
different.
As
hypothesis
testing
machinery
was
designed
to
prove
H
1
,
the
above
figure
suggested
the
hypothesis:
H
o
:
LINCOCIN
®
FORTE
S
was
worse
than
AMPICLOX
Tm
(nit
rca
-A)
vs
[1]
11
1
:
LINCOCIN
®
FORTE
S
was
not
worse
than
AMPICLOX
Tm
(n
if
na
>
-A)
The
difference
threshold,
A,
was
set
to
15%
when
the
cure
rate
was
50%
(Schukken
&
Deluyker,
1995).
This
threshold
was
translated
to
the
angular
scale
of
the
Freeman-Tukey
transfor-
mation
by
the
relationship
A
a
=
2[arcsine
,J('/2
+
A)]-2[arcsine
1
/
2
]
=
0.30.
Thus,
if
i
7
if—t
and
Sd
were
the
estimated
difference
between
the
two
cure
rates
and
its
standard
error,
respectively,
both
in
the
angular
scale,
and
with
the
standard
error
associated
with
f
degrees
of
freedom,
then
the
test
statistic
for
testing
[1]
had
the
form
i
7
y—i
7
a
±
0.30
t
Then,
the
test
would
be
If
t
t
.
95
,
f
then
reject
H
o
and
conclude
that
the
data
supports
H
1
.
In
that
case
the
separate
hypothesis
was
tested
to
determine
whether
the
LINCOCIN
®
FORTE
S
cure
rate
was
significantly
better
than
AMPICLOX
Tm
.
Statistical
analysis
of
somatic
cell
counts
The
design
of
the
study
was
such
that
SCC
measurements
on
a
case
were
stopped
when
the
case
was
deemed
a
treatment
failure,
i.e.
no
clinical
cure.
Thus,
for
the
analysis
of
SCC,
only
cases
that
were
considered
clinical
cures
were
utilized.
The
use
of
the
initial
SCC
was
initially
considered
as
a
covariate.
As
three
post-treatment
SCC
measurements
were
taken,
the
pattern
of
SCC
was
summarized
into
two
orthogonal,
linear
and
quadratic
contrasts.
RESULTS
A
total
of
256
cases
were
enrolled
from
herds
(n
=
56)
in
Belgium
(n
=
15),
France
(n
=
20),
and
Germany
(n
=
21).
From
this
raw
data
set,
24
cases
were
excluded
because
they
did
not
meet
the
criteria
for
study
entry
(n
=
4),
data
sheets
were
incomplete
(n
=
7),
adjunctive
therapy
was
given
during
the
monitoring
period
by
the
farmer
without
the
veterinarian's
agreement
(n
=
5)
or
by
the
veterinarian
but
therapy
was
not
needed
for
animal
survival
(n
=
1),
therapy
was
given
for
mastitis
in
another
quarter
during
the
monitoring
period
(n
=
5),
or
the
cow
was
dried-off
during
the
monitoring
period
(n
=
2).
S
a
©1999
Blackwell
Science
Ltd,
J.
vet.
Pharmacol.
Therap.
22,
274-282
Clinical
mastitis
efficacy
277
Clinical
cure
After
exclusion
there
were
232
cases
for
analysis
of
clinical
cure
rate.
The
percentage
cases
by
parity
group
for
each
treatment
(Fig.
1)
indicates
that
in
the
AMPICLOX
TM
group
there
were
more
second
parity
cows
(25.2%)
than
in
the
LINCOCIN
®
FORTE
S
group
(15.8%).
Conversely,
there
were
more
fourth-and-higher
parity
cows
in
the
LINCOCIN
®
FORTE
S
group
(50.8%)
than
in
the
AMPICLOX
TM
group
(40.5%).
Across
treatments,
clinical
cure
rates
were
lowest
for
the
fourth-and-higher
parity
group
(55.7%)
and
highest
for
the
second
parity
(59.6%),
consistent
with
existing
literature
(Daniel
et
al.,
1982).
Statistical
analysis
of
the
232
cases
showed
that
the
clinical
cure
rate
was
not
worse
than
(P
<
0.0001)
as
well
as
significantly
better
than
(P
=
0.035)
for
LINCOCIN
®
FORTE
S
(62.5%)
than
for
AMPICLOX
TM
(51.8%).
Clinical-plus-bacteriological
cure
From
the
232
cases
used
for
analysis
of
the
clinical
cure
rate
another
43
cases
were
excluded
from
the
analysis
of
clinical-
plus-bacteriological
cure
rate.
This
pertained
to
cases
with
missing
(n
=
1),
negative
(n
=
31),
and
contaminated
(n
=
9)
pretreatment
bacteriology
as
well
as
cases
with
more
than
one
contaminated
post-treatment
bacteriology
result
(n
=
2).
This
left
189
cases
and
208
pretreatment
isolates
(i.e.
19
cases
with
two
isolates
pretreatment)
for
analysis
of
clinical-plus-bacter-
iological
cure
rate.
The
most
frequently
isolated
pathogen
species
in
this
study
were
Staph.
aureus,
Esculine-positive
streptococci
(principally
Streptococcus
uberis),
and
E.
coli
(Fig.
2).
Clinical-
plus-bacteriological
cure
rates
were
lowest
for
the
fourth-and-
higher
parity
group
(26.2%)
and
highest
for
the
first
parity
(39.4%)
(Fig.
3),
consistent
with
existing
literature
(Daniel
et
al.,
1982).
Statistical
analysis
of
the
189
cases
showed
the
clinical-plus-
bacteriological
cure
rate
with
LINCOCIN
®
FORTE
S
(38.1%)
to
be
not
worse
than
(P
<
0.0001)
as
well
as
significantly
better
than
(P
=
0.005)
with
AMPICLOX
TM
(21.7%).
As
in
previous
studies
(Pearson
&
Mackie,
1979;
Wilson
et
al.,
1986)
the
Staph.
aureus
clinical-plus-bacteriological
cure
rate
was
lower
than
for
other
bacterial
species
in
both
treatment
groups.
The
percentage
of
cases
cured
clinically-plus-bacteriolo-
gically
was
higher
with
LINCOCIN
®
FORTE
S
for
every
pathogen
species
found
pretreatment
(Table
1).
The
difference
in
the
clinical-plus-bacteriological
cure
rate
of
LINCOCIN
®
FORTE
S
minus
AMPICLOX
TM
,
expressed
as
a
percentage
difference
from
the
AMPICLOX
TM
cure
rate,
was
+
175%,
+
32%,
and
+
21%
for
Staph.
aureus,
Esculine-positive
streptococci,
and
E.
coli,
respectively.
Somatic
cell
count
The
initial
SCC
values
were
not
significantly
different
between
the
two
treatments
at
the
10%
significance
level.
Also,
at
the
20%
significance
level,
there
was
no
evidence
that
the
slopes
of
the
initial
SCC
were
significantly
different
from
zero.
Thus,
covariance
analysis
was
not
utilized.
SCC
in
cases
that
were
either
cured
bacteriologically
or
had
no
growth
in
the
initial
sample
were
not
significantly
different
from
each
other
(P
=
0.11),
but
both
had
significantly
lower
SCC
than
those
cases
that
failed
to
cure
(P
<
0.0001).
The
overall
linear
effect
was
significant
(P
<
0.0001)
and
there
was
a
significant
two-way
interaction
of
the
linear
effect
with
bacteriological
cure
(P
=
0.006)
but
not
with
treatment
(P
=
0.298).
There
was
no
significant
three-way
interaction
between
linear
effect,
bacter-
iological
cure,
and
treatment
(P
=
0.610).
The
overall
quadratic
effect
(P
=
0.180)
and
interactions
with
the
quadratic
effect
were
not
significant,
though
the
interaction
between
quadratic
effect
and
bacteriological
cure
was
almost
significant
(P
=
0.079).
As
shown
in
Figs
4,
5
and
6,
these
results
are
because
of
a
numerical
decline
in
SCC
over
the
observation
period
for
cases
with
clinical-plus-bacteriological
cure
(Fig.
4)
and
those
with
an
initial
negative
culture
result
(Fig.
5),
while
cases
that
did
not
cure
bacteriologically
showed
little
decline
in
SCC
(Fig.
6).
0
Lincocin
Forte
S
Ampiclox
50.8
40.5
25.2
18.0
16.7
16.2
5
67
4
r
i
30
20
10
2
>=.4
Panty
Group
Fig.
1.
Distribution
of
cases
by
parity
group
(1,
2,
3,
or
4)
for
each
treatment
group.
Data
base:
232
cases
(IINCOC1N
®
FORTE
S
=
120
cases,
AMPICLOX
TM
=
112
cases).
©1999
Blackwell
Science
Ltd,
J.
vet.
Pharmacol.
Therap.
22,
274-282
278
H.
A.
Deluyker
et
al.
4
30
25
7,
20
Lincocin
Forte
S
IMAmpiclox
30.4
26.2
21.8
16.0
6.5
9
5.6
2.8
SA+
STA
STD
EPS
ECO
OEN
OTH
Pathogens
species
Fig.
2.
Distribution
of
pathogen
species
isolated
pretreatment,
by
treatment
group.
Data
base:
208
isolates
from
189
cases
(IINCOON
®
FORTE
S
group
=
107
isolates
from
97
cases,
AMPICLOX
Tm
group
=
101
isolates
from
92
cases).
Pathogen
species:
SA+
=
Staphylococcus
aureus,
SA—
=
Staphylococcus
species,
STA
=
Streptococcus
agalactiae,
STD
=
Streptococcus
dysgalactiae,
EPS
=
Esculine
positive
streptococci,
ECO
=
Escherichia
coli,
OEN
=
other
enterobacteriaceae,
0TH
=
other
microorganisms.
45
40
39.4
35
31.6
30
30.3
26.2
20
5
10
2
3
>=4
Parity
Group
Fig.
3.
Clinical-plus-bacteriological
cure
rates
by
parity
group
(1,
2,
3,
or
4)
across
treatments.
Data
base:
189
cases
(LINCOCIN
®
FORTE
S
group
=
97
cases,
AMPICLOX
Tm
group
=
92
cases).
For
the
cases
that
showed
clinical-plus-bacteriological
cure,
there
was
no
evidence
of
significant
treatment—time
interaction
(P
=
0.390),
but
there
was
an
overall
significant
difference
between
the
two
treatments with
respect
to
SCC
(P
=
0.036)
and
the
SCC
exhibited
significant
linear
(P
<
0.0001)
as
well
as
quadratic
(P
=
0.003)
effects
over
time.
In
Table
2
the
geometric
mean
SCC
and
the
frequency
with
which
another
organism
was
isolated
post-treatment
are
provided
for
cases
cured
clinically
and
bacteriologically
from
Staph.
aureus,
Esculine-positive
streptococci,
and
E.
coli.
The
SCC
appeared
to
decline
with
time,
regardless
of
the
pathogen
species
isolated
prior
to
treatment
onset,
but
less
so
if
the
rate
of
infection
with
another
organism
was
high.
DISCUSSION
Clinical
cure
The
clinical
cure
rates
estimates
of
62.5%
and
51.8%
are
lower
than
those
from
previous
reports.
This
may
be
because
of
the
early
assessment
post-treatment
of
clinical
cure
in
our
study.
For
example,
Wilson
et
al.
(1986)
noted
clinical
cure
rates
of
about
82%
by
day
14
after
treatment
onset.
Guterbock
et
al.
(1993)
however,
observed
values
of
66%
to
68%
clinical
cure
rate
when
making
an
evaluation
sooner,
i.e.
by
4-5
days
after
treatment
initiation.
This
may
be
particularly
relevant
for
the
disappear-
ance
of
clinical
signs
of
inflammation
of
the
udder
which
have
©1999
Blackwell
Science
Ltd,
J.
vet.
Pharmacol.
Therap.
22,
274-282
Clinical
mastitis
efficacy
279
Table
1.
Clinical,
bacteriological
and
clinical-plus-bacteriological
cure
rates
per
pathogen
species
isolated
pretreatment,
by
treatment
group
UNCOCIN®
FORTE
S
AMPICLOX
I
'
m
Pathogen
species
Clinical
Clinical-plus- Clinical-plus-
Bacteriological*
bacteriological
Clinical
Bacteriological*
bacteriological
NI
N
%
Staphylococcus
aureus
17/28
60.7
7/17
41.2
7/28
25.0
13/22
59.1
2/13
15.4
2/22
9.1
Staphylococcus
species
6/7
85.7
3/6
50.0
3/7
42.9
3/3
100.0
1/3
33.3
1/3
33.3
Streptococcus
agalactiae
3/3
100.0
3/3
100.0
3/3
100.0
1/2
50.0
1/1
100.0
1/2
50.0
Streptococcus
dysgalactiae
5/6
83.3
5/5
100.0
5/6
83.3
3/8
37.5
2/3
66.7
2/8
25.0
Esculine-positive
streptococci§
17/33
51.5
10/17
58.8
10/33
30.3
14/35
40.0
8/14
57.1
8/35
22.9
Escherichia
coli
13/18
72.2
8/13
61.5
8/18
44.4
9/19
47.4
7/9
77.8
7/19
36.8
Other
enterobacteriaceae#
3/3
100.0
3/3
100.0
3/3
100.0
3/5
60.0
2/3
66.7
2/5
40.0
Other
microorganisms]
6/9
66.7
5/6
83.3
5/9
55.6
3/7
42.9
0/3
0.0
0/7
0.0
Data
base:
208
isolates
from
189
cases
(LINCOCIN
®
FORTE
S
group
=
107
isolates
from
97
cases,
AMPICLOX
I
'
m
group
=
101
isolates
from
92
cases).
*Calculated
for
clinically
cured
cases.
=
number
cured/Total.
§The
category
'Other
Esculine-positive
cocci'
includes
Strep.
uberis
(n
=
54).
#The
category
'Other
enterobacteriaceae'
includes
Klebsiella
species
(n
=
1).
tThe
category
'Other
microorganisms'
includes
Corynebacterium
bovis
(n
=
5),
Bacillus
species
(n
=
2),
Actinomyces
pyogenes
(n
=
2),
Pasteurella
multocida
(n
=
2),
Clostridium
species
(n
=
2),
yeast
(n
=
2),
and
Moraxella
species
(n
=
1).
0
Lincocin
Forte
S
0Ampiclox
6119
3269
2762
1888
831
612
443
339
7000
6000
5000
4000
0
0
3000
2000
1000
4
to
5
13
to
15
20
to
22
Days
after
first
treatment
Fig.
4.
Geometric
mean
somatic
cell
count
(cells
x
10
-3
/mL)
for
cases
cured
clinically-plus-bacteriologically,
over
time
and
by
treatment.
been
noted
to
persist
longer
than
abnormal
milk
appearance.
The
main
reason
for
choosing
an
early
time
of
veterinary
evaluation
of
clinical
cure
was
to
ensure
maximal
protocol
adherence
by
guaranteeing
the
farmers,
who
were
blinded
to
the
treatments,
that
the
cases
would
be
re-examined
soon
after
treatment
and
that
at
that
time
alternative
therapy
could
be
initiated,
if
deemed
necessary
by
the
veterinarian.
Clinical-plus-bacteriological
cure
Clinical-plus-bacteriological
cure
rate
is
low,
particularly
for
AMPICLOX
Tm
(21.7%),
when
compared
to
previously
observed
figures
(40%
to
70%)
(Wilson
et
al.,
1986;
Guterbock
et
al.,
1993).
Differences
with
previously
reported
clinical-plus-bacter-
iological
cure
rates
may
be
the
result
of
differences
in
the
calculation
of
this
cure
rate.
For
example,
bacteriological
cure
has
been
reported
after
excluding
cases
which
required
additional
treatment
(Guterbock
et
al.,
1993).
In
our
study,
such
cases
were
not
excluded
but
rather
considered
failures.
Also,
in
our
study,
there
were
three
bacteriological
sampling
points
at
which
the
originally
isolated
bacteria
were
not
to
be
found
in
order
to
be
considered
a
cure.
This
is
more
than
in
previous
studies.
The
difference
in
clinical-plus-bacteriological
cure
rate
(16.4%)
between
the
two
treatment
groups
was
higher
than
the
10.7%
difference
observed
with
clinical
cure.
The
numerical
differences
for
Staph.
aureus
between
LINCOCIN
®
FORTE
S
and
AMPICLOX
TM
in
bacteriological-plus-clinical
cure
rates
were
attributable
to
a
higher
bacteriological
cure
in
the
LINCOCIN
®
FORTE
S
treated
cases,
as
the
Staph.
aureus
clinical
cure
rates
were
similar
between
both
treatments.
It
is
noteworthy
that
cases
which
were
clinically
cured
following
infections
with
E.
coli
were
not
always
bacteriologically
negative.
This
is
consistent
with
Guterbock
et
al.
(1993)
and
©1999
Blackwell
Science
Ltd,
J.
vet.
Pharmacol.
Therap.
22,
274-282
1728
--
-
5267
4627
2106
2044
2029
3292
2684
280
H.
A.
Deluyker
et
al.
Lincocin
Forte
S
InAmpiclox
-
2683
-
3079
1125
954
488
607
422
187
4
to
5
13
to
15
20
to
22
7000
6000
5000
4000
0
3000
2000
1000
Days
after
first
treatment
Fig.
5.
Geometric
mean
somatic
cell
count
(cells
x
10
-3
/mL)
for
cases
cured
clinically
and
with
an
initial
negative
culture
result,
over
time
and
by
treatment.
0
Lincocin
Forte
S
53Ampiclox
4
to
5
13
to
15
20
to
22
Days
after
first
treatment
Fig.
6.
Geometric
mean
somatic
cell
count
(cells
x
10
-3
/mL)
for
cases
clinically
cured
but
bacteriologically
not
cured,
over
time
and
by
treatment.
Table
2.
Geometric
mean
somatic
cell
count
for
clinically-plus-bacteriologically
cured
cases
over
time,
by
treatment
group
and
major
pathogen
isolated
pretreatment.
Causative
organism*
Treatment
group
SCC
(cells
x
10
-3
/mL)
Rate
of
new
infection
NI
0
Time
(days)
after
first
infusion
4-5
13-15
20-22
Escherichia
coli
LINCOCIN
®
FORTE
S
6951
23
78
282
243
1/8
12.5
AMPICLOX
TM
8755
5465
764
930
3/7
42.9
Staphylococcus
aureus
LINCOCIN
®
FORTE
S
10
595
2189
431
534
0/6
0.0
AMPICLOX
TM
479
7
1284
726
400
0/2
0.0
Esculine-positive
streptococci
LINCOCIN
®
FORTE
S
4148
1229
451
294
1/10
10.0
AMPICLOX
TM
3417
2225
1009
884
3/7
42.3
*Organism
isolated
pretreatment.
Number
of
cases
with
at
least
one
post-treatment
milk
sample
that
was
bacteriologically
positive
with
an
organism
different
than
the
causative
organism
divided
by
the
total
number
of
clinically-plus-bacteriologically
cured
cases
from
which
the
causative
organism
was
isolated.
7000
6000
5000
4000
0
0
N
3000
2000
1000
©1999
Blackwell
Science
Ltd,
J.
vet.
Pharmacol.
Therap.
22,
274-282
Clinical
mastitis
efficacy
281
suggests
that
disappearance
of
E.
coli
following
clinical
cure
is
not
necessarily
'automatic'.
Somatic
cell
count
While
SCC
was
not
a
cure
criterion,
it
is
negatively
associated
with
milk
yield
and
milk
quality,
and
may
affect
the
payment
the
dairy
farmer
receives
per
kg
milk
sold.
SCC
has
been
found
to
decline
over
an
15-day
period
to
pretreatment
levels
of
150
000
SCC
following
onset
of
clinical
mastitis
caused
by
environ-
mental
mastitis
pathogens
(Deluyker
et
al.,
1993).
The
current
study
confirms
this
pattern
but
shows
that
this
decline
only
occurs
in
cases
that
were
either
bacteriologically
negative
pretherapy
or
were
cured
clinically-plus-bacteriologically.
The
SCC
declined
in
both
treatment
groups
but
was
lower
for
the
LINCOCIN
®
FORTE
S
group
than
for
the
AMPICLOX
Tm
group.
This
is
a
surprising
result,
as
both
groups
represent
cases
that
are
considered
to
be
fully
cured.
One
explanation
could
have
been
possible
differences
in
initial
SCC
between
treatment
groups.
In
this
regard,
Pyorala
&
Syvajarvi
(1987)
reported
cases
with
high
levels
of
antitrypsin
and
NAGase
to
recover
poorly,
except
for
coliform
mastitis
where
it
was
suggested
that
an
intensive
initial
inflammatory
response
was
beneficial
to
the
course
of
the
disease.
In
our
study,
initial
SCC
was
not
different
between
treatment
groups
and
was
not
associated
with
post-
treatment
SCC
level.
It
should
be
noted
that
our
study
population
did
not
include
cases
with
severe
mastitis,
which
required
additional
treatment.
Pyorala
&
Pyorala
(1997)
reported
that
efficiency
of
SCC
to
predict
bacteriological
cure
was
greatest
for
cases
that
had
been
infected
with
Staph.
aureus,
whereas
for
streptococci
and
coliforms
specificity
was
low
because
SCC
remained
high
following
bacteriological
cure.
The
difference
in
SCC
between
treatments
was
observed
when
either
E.
coli
or
Esculine-positive
streptococci
were
initially
diagnosed
(Table
2).
For
Staph.
aureus,
there
was
little
difference
in
SCC
between
treatment
groups
but
the
number
of
cases
cured
following
AMPICLOX
Tm
treatment
was
small.
One
explanation
for
the
differences
in
SCC
observed
between
the
two
treatment
groups
is
that,
for
cases
from
which
E.
coif
or
Esculine-positive
streptococci
had
been
isolated
initially,
the
rate
of
infection
with
another
organism
in
cured
cases
was
higher
following
treatment
with
AMPICLOX
Tm
than
with
LINCOCIN
®
FORTE
S.
The
species
most
frequently
isolated
at
any
post-treatment
sampling
from
such
cases
was
non-aureus
staphylococcus
(LINCOCIN
®
FORTE
S,
n
=
1;
AMPICLOX
Tm
,
n
=
5).
Other
species
were
isolated
only
from
one
case
in
each
treatment
group
(Esculine-positive
streptococci
and
other
organisms)
or
only
from
one
case
in
the
AMPICLOX
Tm
treatment
group
(Staph.
aureus
and
Bacillus
species).
The
purpose
of
the
study
was
to
test
the
hypothesis
whether
cure
with
LINCOCIN
®
FORTE
S
was
lower
than
cure
following
treatment
with
AMPICLOX
TM
.
This
hypothesis
was
rejected,
showing
that
LINCOCIN
®
FORTE
S
cure
was
similar
or
better.
In
fact,
clinical
cure
rate,
clinical-plus-bacteriological
cure
rate,
and
SCC
level
were
significantly
better
following
treatment
with
LINCOCIN
®
FORTE
S
than
with
AMPICLOX
Tm
.
AMPICLOX
Tm
has
similar
characteristics
to
LINCOCIN
®
FORTE
S,
i.e
a
broad
spectrum
(Gram
+
and
Gram
—)
of
activity
and
an
identical
treatment
schedule.
These
characteristics,
along
with
AMPICLOX
Tm
being
registered
in
a
variety
of
countries
worldwide,
made
it
a
pertinent
positive
control
product.
ACKNOWLEDGMENTS
The
authors
wish
to
express
their
gratitude
to
the
investiga-
tors
who
took
part
in
the
various
aspects
of
the
trial:
E.
Bleckmann,
A.
de
ICruif,
S.
Delobel,
L.
Devriese,
B.
Faroult,
C.
Foulcher,
R.
Kramer,
H.
Laevens,
R.
Le
Goas,
P.
Legrand,
G.
Montagut,
K.
Steinbach,
E.
Van
Baar,
L.
Vandevelde,
de
Vlaamse
Rundveeteelt
Vereniging,
M.
Weihe;
K.
Neyrinck
for
manuscript
preparation.
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