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TRANSCRIPT
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STRATEGIES TO MITIGATE THE ADVERSE EFFECTS OF ENDOTOXINS IN TRANSITION DAIRY COWS
Italian Society of Buiatria, Parma 23-24 Nov, 2017Nov 24, 2017
Burim N. Ametaj, DVM, PhD
University of AlbertaCanada
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Outline of the presentation
¨ Endotoxin-related diseases
¨ What are the sources of endotoxins in a dairy cow?
¨ Does endotoxin translocate into the systemic circulation?
¨ How can we prevent health implications of endotoxins?
¤Grain processing technology
¤Oral vaccination (metritis and mastitis)
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Endotoxin-related Diseases3
START HERE
Eckel & Ametaj, 2016
What are the sources of endotoxins in a dairy cow?
¨ There are three sources of endotoxins in dairy cows
¤ Rumen
¤ Postpartum uterus
¤ Udder
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Proof of endotoxin elevation in the rumen during high grain diets
16-fold increase in endotoxin at 45% barley grain TMR
5
Emanuel et al., 2008
0
2,000
4,000
6,000
8,000
10,000
12,000
1 3 5 7 10
LP
S n
g/m
L
Experimental day
Ruminal fluid LPS in cows fed different amounts of grain
0% Barley grain
15% Barley grain
30% Barley grain
45% Barley grain
Does endotoxin translocate through GI tract: Ussing chamber study
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Endotoxin translocation through rumen increases: in
vitro study7
Emanuel et al., 2007
0
100
200
300
400
500
600
4.5 5.5 6.5
LP
S (µ
g/c
m2/h
)
pH value of perfusate on the mucosal side of rumen tissue
Translocation of LPS from E. coli 055:B5 across rumen
tissue from steers
Trt: P < 0.01
Endotoxin translocation through colon8
Emanuel et al., 2007
0
50
100
150
200
250
5.5 6.5 7.4
LP
S (µ
g/c
m2/h
)
pH value of perfusateon mucosal side of colon tissue
Translocation of LPS from E. coli 055:B5 Across Colon
Tissue of Steers
Trt: P < 0.01
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Rumen pH related to rumen LPS9
Ametaj et al., 2010
Infected mammary as source of endotoxin
¨ Gangrenous mastitis was associated with a plasma
LPS concentration of 85.2 pg/mL, than control cows
at 4.7 pg/mL (18-fold higher) – Hakogi et al.,
(1989)
¨ Intramammary administration of LPS in the udder
enhanced plasma LPS (5-13-fold) – (Dosogne et al.,
2002)
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Mastitis and endotoxin translocation
Hakogi et al., 1989
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Fate of endotoxin in systemic circulation and host response
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Eckel & Ametaj, 2016
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Fate of endotoxin in blood – Macrophage Pathway
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Fate of endotoxin in blood – Lipoprotein Pathway
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q Barley Grain Technology
q Oral Vaccine
New Technologies to mitigate ERD15
Grain and inflammation16
¨ Feeding high amounts of grain triggers
inflammation in dairy cows
¨ High amounts of grain are associated with
increased levels of endotoxin in the rumen fluid
¨ High grain diets is associated with elevated blood
SAA, CRP, and LBP
Emmanuel et al., 2008
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Elevated ruminal fluid LPS with high grain diets
17
0
2,000
4,000
6,000
8,000
10,000
12,000
1 3 5 7 10
LP
S n
g/m
L
Experimental day
Ruminal fluid LPS in cows fed different amounts of grain
0% Barley grain
15% Barley grain
30% Barley grain
45% Barley grain
Emmanuel et al., 2008
Feeding grain is associated with systemic inflammation – Elevated Blood SAA
18
0
5,000
10,000
15,000
20,000
25,000
30,000
35,000
40,000
45,000
50,000
1 3 5 7 10
SA
A (
ng/m
L)
Experimental day
PLasma SAA in cows fed different amounts of grain
15% Concentrate mix
30% Concentrate mix
45% Concentrate mix
60% Concentrate mix
Emmanuel et al., 2008
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Feeding grain is associated with systemic inflammation – Elevated blood CRP
19
0.4
0.6
0.8
1
1.2
1.4
1.6
1.8
1 3 5 7 10
CR
P (
ng/m
L)
Experimental day
Plasma CRP in cows fed different amounts of grain
15% Concentrate mix
30% Concentrate mix
45% Concentrate mix
60% Concentrate mix
Emmanuel et al., 2008
Feeding grain is associated with systemic inflammation – Elevated blood LBP
20
0
2,000
4,000
6,000
8,000
10,000
12,000
14,000
16,000
18,000
1 3 5 7 10
LB
P (
ng/m
L)
Experimental day
Plasma LBP in cows fed different amounts of grain
15% Concentrate mix
30% Concentrate mix
45% Concentrate mix
60% Concentrate mix
Emmanuel et al., 2008
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Barley grain treated with LA prevent elevation of LPS in rumen fluid
21
0
100
200
300
400
500
600
700
1 5 11 14 16 19 21
LP
S (
ng/m
L)
Experimental day
Ruminal LPS in cows fed LA-(0.5) or water-treated barley
LA-Treated Barley
Water-Treated Barley
Adaptation period Measurement period
Iqbal et al., 2012
New barley grain technology lowers the amount of endotoxin in the rumen fluid
¨ Increases resistant starch by 24%
¨ Lowers the amount of endotoxin in the rumen fluid
¨ Lowers the acute phase response as evidenced by lower haptoglobinand serum amyloid A
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0
200
400
600
800
1000
1200
1400
1 5 11
Ru
men
en
do
tox
in (n
g/m
L)
Measurement days (d)
Trt: P < 0.001
Day:P < 0.001Day x Trt: P = 0.019
n = 8SEM = 52.920
400
450
500
550
600
650
700
750
800
850
900
1 5 11
Pla
sma
ha
pto
glo
bin
(µ
g/d
L)
Measurements day
Trt: P = 0.37
Day:P = 0.82Day x Trt: P = 0.94
n = 8SEM = 79.012
a
b
Iqbal et al., 2012
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Development of a vaccine against Endotoxin-related diseases
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Oral vaccination24
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Rationale for the vaccine25
Evaluate health and immunology responses of
repeated oral immunization or intravenous
administration of increasing doses of LPS from
Escherichia coli 0111: B4, two weeks before
and one week after calving
Study 1- Objectives 26
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Materials and methods27
¨ 32 clinically healthy nearing term heifers and
multiparous Holstein cows assigned to four groups
of eight cows each
¨ Two groups of n=8 cows each treated either withoral LPS or oral saline (control) (Experiment 1)
¨ Two groups of n=8 cows each treated withintravenous LPS or saline (control) (Experiment 2)
¨ LPS was administered at increasing doses of
0.01µg, 0.05µg, and 0.1µg/kg BW on day 1 and 3
of each week
Materials and methods28
¨ Blood samples were collected twice per week
during 2 wk and 1 wk before and 1 wk after
parturition as well as once per week during wk 2, 3,
and 4 after parturition.
¨ Measurements
¤ Plasma acute phase proteins: SAA, LBP, haptoglobin
by ELISA using commercially available kits.
¤ Plasma immunoglobulins: IgA, IgG, and IgM by
ELISA kit EndoCab specific to endotoxin
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Blood analyses29
¨ Measurements:
¤ Plasma metabolites: Cortisol, NEFA, BHBA, glucose,lactate, insulin, and cholesterol using commerciallyavailable kits
¤ Plasma minerals: Calcium, iron, zinc, and copper weremeasured by atomic absorption spectrometry
Results: Oral LPS – Plasma SAA30
0
10,000
20,000
30,000
40,000
50,000
60,000
W1D1 W2D1 W3D1 W4 W5 W6
SA
A (
ng/m
L)
Time around parturition (wk)
Plasma SAA in dairy cows around parturition
Oral LPS
Oral saline
P
a
r
t
u
r
i
t
i
o
n
Trt P > 0.05
Week P > 0.05
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Results: Intravenous LPS – Plasma SAA31
0
5,000
10,000
15,000
20,000
25,000
30,000
35,000
40,000
45,000
50,000
W1D1 W2D1 W3D1 W4 W5 W6
SA
A (
ng/m
L)
Time around parturition (wk)
Plasma SAA in dairy cows around parturition
IV Saline IV LPSP
a
r
t
u
r
i
t
i
o
n
Results: Oral LPS – Plasma LBP32
0
10,000
20,000
30,000
40,000
50,000
60,000
70,000
80,000
W1D1 W2D1 W3D1 W4 W5 W6
LB
P (
ng/m
L)
Time around parturition (wk)
Plasma LBP in dairy cows around parturition
Oral LPS
Oral salineP
a
r
t
u
r
i
t
i
o
n
Trt P < 0.05
Week P < 0.05
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Results: Intravenous LPS – Plasma LBP33
0
10,000
20,000
30,000
40,000
50,000
60,000
70,000
80,000
90,000
100,000
W1D1 W2D1 W3D1 W4 W5 W6
LB
P (
ng/m
L)
Time around parturitionj (wk)
Plasma LBP in dairy cows around parturition
IV Saline
IV LPS
P
a
r
t
u
r
i
t
i
o
n
TRT P < 0.01
Results: Oral LPS – Plasma Hp34
0
200
400
600
800
1,000
1,200
W1D1 W2D1 W3D1 W4 W5 W6
Hap
toglo
bin
(µ
g/m
L)
Time around parturition (wk)
Plasma haptoglobin in dairy cows around parturition
Oral LPS
Oral saline
P
a
r
t
u
r
i
t
i
o
n
Trt P > 0.05
Week P < 0.05
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Results: Intravenous LPS – Plasma Hp35
0
200
400
600
800
1,000
1,200
1,400
1,600
W1D1 W2D1 W3D1 W4 W5 W6
Hap
toglo
bin
(µg/m
L)
Time around parturition (wk)
Plasma haptoglobin in cows around parturition
IV Saline
IV LPSP
a
r
t
u
r
i
t
i
o
n
TRT P < 0.01
Results: Oral LPS – Plasma NEFA36
0
200
400
600
800
1,000
1,200
1,400
W1D1 W1D2 W2D1 W2D2 W3D1 W3d2 W4 W5 W6
NE
FA
(m
g/m
L)
Time around parturition (wk)
Plasma NEFA in cows around parturition
Oral LPS
Oral Saline
P
a
r
t
u
r
i
t
i
o
n
TRT P < 0.01
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Results: Intravenous LPS – Plasma NEFA37
0
200
400
600
800
1,000
1,200
1,400
W1D1 W1D2 W2D1 W2D2 W3D1 W3d2 W4 W5 W6
NE
FA
(m
g/m
L)
Time around parturition (wk)
Plasma NEFA in cows around parturition
IV Saline
IV LPS
P
a
r
t
u
r
i
t
i
o
n
TRT P > 0.05
Results: Oral LPS – Plasma BHBA38
0
500
1,000
1,500
2,000
2,500
W1D1 W1D2 W2D1 W2D2 W3D1 W3D2 W4 W5 W6
BH
BA
(m
g/m
L)
Time around parturition (wk)
Plasma BHBA in cows around parturition
Oral LPS
Oral Saline
P
a
r
t
u
r
i
t
i
o
n
TRT P < 0.01
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Results: Intravenous LPS – Plasma BHBA39
0
200
400
600
800
1,000
1,200
1,400
1,600
1,800
W1D1 W1D2 W2D1 W2D2 W3D1 W3D2 W4 W5 W6
BH
BA
(m
g/m
L)
Time around parturition (wk)
Plasma BHBA in cows around parturition
IV Saline
IV LPS
P
a
r
t
u
r
i
t
i
o
n
Results: Oral LPS – Plasma anti-LPS IgA
antibodies40
0.03
0.035
0.04
0.045
0.05
0.055
0.06
0.065
0.07
0.075
W1D1 W2D1 W3D1 W4 W5 W6
IgA
(M
U/m
L)
Time around parturition (wk)
Plasma IgA in cows around parturition
Oral LPS
Oral Saline
P
a
r
t
u
r
i
t
i
o
n
Trt effect P = 0.25