Sie sind auf Seite 1von 55

5/13/2010

Bovine embryo transfer recipient synchronization and management in tropical environments


Pietro S. BaruselliA, Roberta M. FerreiraA, Manoel F. S FilhoA, Luiz F. T. NasserB, Carlos A. RodriguesC and Gabriel A. BD
A B C

Departamento de Reproduo Animal, FMVZ-USP, CEP 05508-000 So Paulo, Brazil FIRMASA-IATF, CEP 790020-210, Campo Grande-MS, Brazil SAMVET Embries, CEP 13560-000, So Carlos, SP, Brazil D Instituto de Reproduccin Animal, Zona Rural General Paz, (5145) Crdoba, Argentina

36th Annual Conference of the International Embryo Transfer Society


Cordoba Argentina - 2010

Theme: Back to the Basis, from the Bench to the Field

5/13/2010

6 time increase

75%

25%

IVF

ET

Figure. Brazilian bovine embryo production 2000 to 2008. (ET = in vivo embryo transfer, IVF = in vitro embryo transfer)
Viana - SBTE, 2009

Brazilian ET market (2008)


ET in vivo ET in vitro
N of ET Embr/SOV

= =

70.000 5,0

= =

14.000 SOV

N of ET Embr/OPU*

220.000 5,0

44.000 OPU

* Mean of 15 viable oocyte per OPU with 30% of in vitro embryo production

5/13/2010

300,000 embryos/year

300,000 recipients
with a good CL / year

Recipients play a critical role in the success of embryo transfer (ET) programs

5/13/2010

Brazilian ET market

Bos indicus donnors (80%)

Bos indicus x Bos taurus recipients

Brazilian ET market

Bos taurus donnors

Lactating Bos taurus recipients


To improve genetic an reproductive efficiency

5/13/2010

Main objectives of our research


1. Improve the proportion of recipients receiving an embryo (transferred/treated) 2. Improve conception rate
(pregnant/transferred)

3. Improve pregnancy rate (pregnant/treated) Development of protocols that are easier to perform by farm personnel

1. Transferred/treated rate
Proportion of recipients receiving an embryo

5/13/2010

Transferred/treated rate
PGF treatment + estrus detection

177 recipients
Estrus detection rate (53.1%)

94 estrus
Estrus but rejected (14.0; 14.9%)

Transferred/treated rate (45.2%)

80 transferred Beef data - Brazil Baruselli et al., 2000 (SBTE)

Transferred-to-treated rate
2 x PGF treatment + estrus detection

1554 recipients
Estrus detection rate (71.7%)

863 estrus
Estrus but rejected (414; 47.9%)

Transferred/treated rate (28.9%)

449 transferred Bolivia and Brazil data

Burry, 2001

5/13/2010

Transferred-to-treated rate
PGF treatment + estrus detection

229 recipients
Estrus detection rate (71.7%)

136 estrus
Estrus but rejected (57.0; 41.9%)

Transferred/treated rate (34.5%)

79 transferred Dairy data - Brazil


Rodrigues et al., 2007 (SBTE)

PGF + estrus detection


(Transferred/treated rate)

300 recipients

250 recipients

200 recipients

Transferred/treated rate (30%)

Transferred/treated rate (40%)

Transferred/treated rate (50%)

100 embryos

5/13/2010

PGF + estrus detection


(Transferred/treated rate)

300 recipients

200 recipients

125 recipients

Transferred/treated rate (30%)

Transferred/treated rate (50%)

Transferred/treated rate (80%)

100 embryos

The low efficiency of traditional PGFbased programs for estrus synchronization in recipients limit the widespread application and success of ET technology

5/13/2010

PGF + Estrus detection + ET Vs Fixed-time embryo transfer


Could FTET improve the proportion of recipients able to receive an embryo?

Estrus detection vs FTET


PGF2 + estrus detection (n=177)
PGF2

ET
Estrus detection 7 days
D7 After estrus

D0

D2

D7

Ovsynch for FTET (n=168)


GnRH
(25g)

PGF2

GnRH
(25g)

FTET
7 days

48h
D0 D7 D9

D16
Baruselli et al., 2000 (SBTE)

5/13/2010

90 80 70 60 50 40 30 20 10 0

PGF + Estrus FTET

P < 0.0001

72.6
b

45.2

a (80/177) (122/168)
Transferred/ treated

Figure. Pregnancy rates in beef recipients transferred 7 days after observation of a prostaglandin F-synchronized estrus or on Day 16 of a fixed-time embryo Baruselli et al., 2000 (SBTE) transfer synchronisation protocol

Estrus detection vs FTET


Group PGF2 + Estrus (n=229)
PGF2

ET
Estrus detection 7 days
D7 After estrus

D0

D2

D7

Group FTET (n=208)


BE+P4 400 UI eCG + 0,5 ml ECP + PGF2

FTET

P4
D0 D8 D17
Rodrigues et al. SBTE (2007)

10

5/13/2010

90 80 70 60 50 40 30 20 10 0

PGF + Estrus FTET

P < 0.0001

75.0
b

34.5

(79/229)

(156/208)

Transferred/ treated

Figure. Pregnancy rates in Holstein repeat-breeder cows transferred 7 days after observation of a prostaglandin F-synchronized estrus or on Day 17 of a fixedRodrigues et al SBTE (2007) time embryo transfer synchronization protocol

Transferred/treated rate
The FTET protocols increase the proportion of recipients transferred compared with the use of PGF followed by ET 7 days after estrus in beef and dairy cattle

11

5/13/2010

2. Pregnant/transferred rate
(Conception rate)

Proportion of recipients pregnant after receiving an embryo

90 80 70 60 50 40 30 20 10 0

PGF + Estrus FTET

P > 0.05

42.3

39.1

45/80

60/122

Pregnant/transferred

Figure. Conception rates beef recipients transferred 7 days after observation of a prostaglandin F-synchronized estrus or on Day 17 of a fixed-time embryo transfer synchronisation protocol.
Baruselli et al., 2000 (SBTE)

12

5/13/2010

90 80 70 60 50 40 30 20 10 0

PGF + Estrus FTET

P >0.05

42.3

39.1

(42/79)

(67/156)

Pregnant/transferred

Figure. Conception rates Holstein cows transferred 7 days after observation of a prostaglandin F-synchronized estrus or on Day 17 of a fixed-time embryo transfer synchronisation protocol.
Rodrigues et al., 2007 (SBTE)

3. Pregnant/treated rate
(Pregnacy rate)

Proportion of recipients pregnant at the end of the program

13

5/13/2010

PGF treatment + estrus detection

100 recipients
Estrus detection rate (50%)

50 estrus
Transferred / treated rate (40%)

Pregnant/treated rate (20%)

40 transferred
Pregnant/transferred rate (50%)

Low efficiency!

20 pregnant

90 80 70 60 50 40 30 20 10 0

75.0

PGF - Estrus FTET + CL

46.8 34.5
a b

39.1 29.3 16.2


a b
(61/208)

(79/229)

(156/208)

(42/79)

(67/156)

(37/229)

Transferred/ treated

Pregnant/ transferred

Pregnant/ treated

Figure. Pregnancy rates in Holstein repeat-breeder cows transferred 7 days after observation of a prostaglandin F-synchronized estrus or on Day 17 of a fixedRodrigues et al SBTE (2007) time embryo transfer synchronisation protocol

14

5/13/2010

90 80 70 60 50 40 30 20 10 0

76.6 56.3 45.2


a b

PGF - Estrus FTET + CL

49.2 35.7 25.4


a b

122/168 80/177

60/122 45/80

60/168 45/177

Transferred/ treated

Pregnant/ transferred

Pregnant/ treated

Figure. Pregnancy rates in recipients transferred 7 days after observation of a prostaglandin F-synchronized estrus or on Day 17 of a fixed-time embryo transfer BARUSELLI et al., 2000 synchronization protocol

Main remarks
The incorporation of techniques designed to control follicular wave dynamics and ovulation eliminates the problem of estrus detection and provides possibilities for the application of FTET programs

15

5/13/2010

FTET protocol (IETS 2002)

EB (2mg) + P4 (50mg)

400 IU eCG
+ PGF2

Implant removal

EB
(1mg)

FTET

P4
D0 D5 D8 D9 D17

5 handlings
B et al., 2002

Main objective of the studies


1. Studies were designed to simplify the protocol by reducing the number of days required for treatment

The purpose of these modifications is to reduce the labor and animal handling associated with the original protocol

16

5/13/2010

Refining the FTET protocol using P4 devices, estradiol and equine chorionic gonadotrophin (eCG)
1. Defining eCG dosage (different doses) 2. Replacement of the second EB administration on Day 9 with estradiol cypionate (EC) on Day 8 (i.e. at the time of implant removal) 3. Evaluation of the effect of delaying the administration of eCG and PGF from Day 5 to Day 8 (i.e. at the time of implant removal)

Effect of the eCG treatment on the efficacy of FTET protocol

17

5/13/2010

No eCG (n=50)
EB
(2.0mg)

PGF2

EB (0.5mg)
US

FTET
US + P4

CIDR-B
D0 D7

24h
D8

D16

eCG on Day 5 (n=50)


EB
(2.0mg)

eCG (800IU)

EB (0.5mg)
PGF2

FTET
US + P4

CIDR-B

24h

US

D0

D5

D7

D8

D16

Baruselli et al., 2000 (SBTE)

90 80 70 60 50 40 30 20 10 0

84

eCG (n=50) No eCG (n=50)

55.3 42 34 29.4 10
b a b
Pregnant/treated

Transferred/treated

Pregnant/transferred

Figure. Pregnancy rate in Bos indicus Bos taurus recipients transferred at a fixed-time and treated or not with eCG.
Baruselli et al., 2000 (SBTE)

18

5/13/2010

90 80 70 60 50 40 30 20 10 0

85 81.4 57.6

eCG (n=50) No eCG (n=50)

48.7 41.7 33.9


a b a b

132/156 127/156

76/132

53/127

76/156

53/156

Transferred/treated

Pregnant/transferred

Pregnant/treated

Figure. Pregnancy rate in Bos taurus and Bos taurus x Bos indicus recipients transferred at a fixed-time and treated or not B et al., 2002 with eCG.

Why did eCG improve the efficiency of the FTET protocol?


1. Follicular growth and ovulation rate 2. Progesterone concentration during diestrus following the synchronized ovulation

19

5/13/2010

1. Effect of eCG on follicular growth and ovulation

Effect of eCG treatment on day 5 of FTET protocol on follicular growth


D0 D5 P4 intravaginal device D8 D9 D17

EB

PGF

EB

FTET

D0

D5 P4 intravaginal device

D8

D9

D17

EB

PGF + 400 IU eCG

EB

FTET

Moreno et al., 2002

20

5/13/2010

16 14 12 DIAMETRO FD (mm) 10 8 6 4 2 0 D0 D1 D2 D3 D4 D5 D6 D7 D8 D9 D10 DIAS DE TRATAMIENTO FD GRUPO CONTROL (n=10) FD GRUPO eCG (n=9)

eCG

Effect of eCG treatment on Day 5 of FTET protocol on follicular growth


Moreno et al., 2002

Effect of eCG treatment on ovulation rate


Group No eCG (n=20)
EB
(2,0mg) PGF2

EB
Blood (P4) + US
Ultrasonography (12/12h)

CIDR

D0

D8

D9

D13 D22

Group eCG (n=21)


EB
(2,0mg)

400IU eCG EB
PGF2

Blood (P4) + US

CIDR

Ultrasonography (12/12h)

D0

D8

D9

D13

D22

Baruselli et al., 2004

21

5/13/2010

90 80 70 60

No eCG eCG

76.2

50.0

% 50
40 30 20 10 0

a
(10/20)
No eCG

b
(16/21)
eCG

Figure. Ovulation rate in recipients treated or not treated with eCG at the fixed-time embryo transfer synchronization protocol. Baruselli et al., 2004

5.0mg Estradiol valerate+ 3.0 mg Norgestomet i.m.

eCG (400IU; n=26) No eCG (n=24)

1,30 1,20 1,10 1,00 0,90 Cm 0,80 0,70 0,60 0,50 0,40
D0 D1 D2 D3 D4 D5 D6 D7 D8 D9 D10am D10pm D11am D11pm

Crestar
eCG no eCG

Ovulation rate

*
73.1 (19/26)a

** ** 50.0 (12/24)

Days

Graphic. Follicular dynamics during ovulation synchronization protocols according to treatment with eCG on implant removal in lactating Bos indicus cows. Pirassunugna, 2004
S Filho et al., 2004 (SBTE)

22

5/13/2010

2. Effect of eCG on progesterone concentrations

9 8 7 6 5 4 3 2 1 0

8.6 6.4 4.6

No eCG eCG

Progesterone concetration (ng/ml)

6.6

4.2 2.2 1.4


a b a b a

3.6

Recipients
Baruselli et al., 2000

Bos taurus x indicus cows


Marques et al., 2003

Nelore heifers
Baruselli et al., 2004

Nelore heifers
S Filho et al., 2005

(n=50)

(n=50)

(n=41)

(n=172)

Figure. Plasma progesterone concentration (ng/mL) during the diestrus following synchronized ovulation in animals treated or not treated with eCG.

23

5/13/2010

Figure. Main effect (mean SEM) of eCG on CL volume (A) and on circulating progesterone concentrations (B) during diestrus following synchronized ovulation in lactating Holstein cows (n = 37). Data was normalized to day of ovulation (Day 0). a,b P < 0.05; A,B P < 0.10.
Souza et al., 2006

Summary
eCG treatment stimulated follicular growth and increased ovulation rate and progesterone concentrations during the subsequent luteal phase

24

5/13/2010

Effect of eCG treatment on conception rate after FTET protocol


P < 0.01
65.3
46.1

90 80 70 60

No eCG eCG

90 80 70 60

No eCG eCG

P < 0.01
55.3

% 50
40 30 20 10 0
No eCG eCG

% 50

40 30 20 10 0

29.4

a
No eCG

b
eCG

Fuentes e De La Fuente, 1997


90 80 70

Baruselli et al., 2000


No eCG eCG

P < 0.01
57.6

eCG CR

% 50
40 30 20 10 0

60
41.7

a
No eCG

b
eCG

Trbulo et al., 2002 Figure. Conception rates (pregnant/transferred rate) in recipients treated or not with eCG at the fixed-time embryo transfer synchronization protocol.

25

5/13/2010

What is the effect of CL size and number on progesterone concentrations and conception rates in embryo recipients?

Conflicting data about the effects of P4 concentrations and CL size on pregnancy rates
Positive effect Nishigai et al. 1998 Mann et al. 1999 Thatcher et al. 2001 Binelli et al. 2001 Nasser et al. 2004 No effect Remsen and Roussel, 1982 Hasler et al. 1987 Spell et al. 2001 Nogueira et al. 2004 Looney et al. 2006

26

5/13/2010

Progesterone concetration (ng/mL)

2.5 2 1.5 1 0.5

2.44
Conception rate (%)

60
1.75

58.4 41.5

50 40 30 20 10 0
(n=77)

a b
0.96

a b

31.8

b
(n=21)

c
(n=77) (n=41) (n=21)

(n=41)

0 CL1 CL2 CL3 (> 2,0 cm2) (1,5 a 2,0 cm2) (< 1,5 cm2)

CL1 CL2 CL3 (> 2,0 cm2) (1,5 a 2,0 cm2) (< 1,5 cm2)

Figures. Progesterone concetration (Day 6 after estrus) and conception rate according to CL area in Bos indicus x Bos taurus recipients (n=139).
Baruselli et al., 2000 (SBTE)

Figure Conception rates (no. recipients pregnant/transferred) relative to plasma progesterone concentrations at the time of embryo transfer in Bos Indicus Bos taurus recipients (n=542). Reis et al., 2004 (SBTE)

27

5/13/2010

Retrospective study
Bos indicus Bos taurus beef recipients 8,034 fresh in vitro-produced embryos
Synchronized with a P4 and estradiol-based protocol for FTET (same protocol) Recipients with CL greater than 16mm () received an embryo (performed by the same technician)

Nasser et al., (IRAC 2009)

Figure. Conception rates (no. recipients pregnant/transferred) relative to the diameter of the corpus luteum (CL) in Bos indicus Bos taurus recipients (n=8034) receiving in vitro-produced embryos (P <0.0001). Nasser et al., (IRAC 2009)

28

5/13/2010

70 60

1 CL 2 or more CL

P < 0.01
51.3

45.8
50

% 40
30 20 10 0
(1880/4109)

b
(328/639)
2 or more CL

1 CL

Figure. Conception rates in Bos indicus x Bos tarus recipients with one CL or with two or more CL on Day 17 of a fixed-time embryo transfer synchronization protocol.

Nasser et al., (IRAC 2009)

Fig. 4. Mean (s.e.m.) progesterone concentrations from Day 0 to Day13 in (a) non-superovulated recipients (n=8; ) and (b) superovulated recipients (n=7; ). Each recipient received 10 embryos produced either in vivo or in vitro.

Lonergan et al., Reproduction, Fertility and Development, 2007, 19, 861868

29

5/13/2010

P < 0.001
High progesterone = 3.86 0.45mm2

Low progesterone = 1.66 0.38mm2

Figure. Mean (s.e.m.) embryo area of Day 13 bovine embryos produced following transfer of Day 7 in vivo or in vitro-derived blastocysts to recipients with High or Control concentrations of circulating progesterone (P).
Lonergan et al., Reproduction, Fertility and Development, 2007, 19, 861868

Figure. Serum progesterone concentrations in heifers treated or not treated with a PRID device from Day 3 until slaughter.
Carter et al., Reproduction, Fertility and Development, 2008, 20, 368375

30

5/13/2010

Normal Progesterone

Elevated Progesterone

Length: 5.97 1.18 cm (n=6)

Length: 14.06 1.18 cm (n=9)

Figure. Representative images of Day 16 bovine concepti recovered from inseminated heifers with normal or elevated progesterone concentrations from Day 3.5. Scale bar=5 mm.
Carter et al., Reproduction, Fertility and Development, 20, 368375, 2008

1. Defining eCG dosage


(different doses)
Objective: evaluate the effect of different doses of eCG (400 v. 500 v. 600 IU) on pregnancy rates.
600 B. indicus B. taurus recipients received a fresh in vitro produced embryos

31

5/13/2010

FTET protocol (IETS 2002)


600 IU ? 500 IU ?
EB (2mg) + P4 (50mg) 400 IU eCG
+ PGF2

Implant removal

EB
(1mg)

FTET

P4
D0 D5 D8 D9 D17

B et al., 2002

Defining eCG dosage


400 IU on Day5 (n=101)
EB + P4

eCG (400IU)
PGF2

EB
(1mg)

FTET

P4 Device
D0
EB + P4

24h
D8 D9 D17

D5 eCG (500UI)
PGF2

500IU on Day 5 eCG (n=98)

EB
(1mg)

FTET

P4 Device
D0 D5 eCG (600IU)
PGF2

24h
D8 D9 D17

600IU on Day 5 (n=100)


EB + P4

EB
(1mg)

FTET

P4 Deivice
D0 D5 D8

24h
D9 D17

32

5/13/2010

Defining eCG dosage


400 IU on Day 8 (n=100)
EB + P4

eCG (400IU)
PGF2

EB
(1mg)

FTET

P4 Device
D0

24h
D8 eCG (500IU)
PGF2

500IU on Day 8 (n=99)


BE + P4

D9

D17

BE
(1mg)

FTET

P4 Device
D0 D8

24h
D9 D17

600IU ond Day 8 (n=96)


BE + P4

eCG (600IU)
PGF2

EB
(1mg)

FTET

P4 Device
D0 D8

24h
D9 D17

90 80 70 60
% 50

87.2 82.1 83.8

600 IU eCG 500 IU eCG 400 IU eCG


51.8 49.4 44.1

P > 0,05
42.3 36 42.9

40 30 20 10
(165/201) (165/197) (171/196) (85/164) (71/161) (84/170)

(85/201) (71/197)

(84/196)

Transferred/treated

Pregnant/transferred

Pregnant/treated

Effect of eCG dose on FTET protocol in Bos indicus x Bos taurus recipients. Reis et al., 2004 (ICAR)

33

5/13/2010

90 80 70 60 % 50 40 30 20 10 0

No eCG eCG (400 IU)

65.5 59.1

P > 0.05
36.3 33.6 23.8 19.9

(256/391) (232/393)

(93/256) (78/232)

(93/391) (78/393)

Transferred/ treated

Pregnant/ transferred

Pregnant/ treated
Unpublished data

Figure. Pregnancy rates in high-producing Holstein repeat-breeder recipients transferred at a fixed-time and treated or not treated with 400 IU of eCG.

FTET protocol (IETS 2002)


800 IU ? 600 IU ?
EB (2mg) + P4 (50mg) 400 IU eCG
+ PGF2

Implant removal

EB
(1mg)

FTET

P4
D0 D5 D8 D9 D17

B et al., 2002

34

5/13/2010

2. Replacement of the second EB administration on Day 9 with estradiol cypionate (EC) on Day 8
(i.e. at the time of implant removal)

FTET protocol (IETS 2002)


EC
EB (2mg) + P4 (50mg) 400 IU eCG
+ PGF2

Implant removal

EB
(1mg)

FTET

P4
D0 D5 D8 D9 D17

5 handlings
B et al., 2002

35

5/13/2010

EB vs EC (LH suge and ovulatiion)


LH

BE P4
24h 22h 26h

72h CE P4
LH

Maio et al., 2007

46h

26h

72h

Sales et al., 2007

EB BE(n=20)

Day 9

EC (n=25) CE

Day 8

18
Number of ovulations

15 12

13 8 3 -2 0 24 48 60 72 84 96 Horas aps a retirada (h) Interval device removal and ovulation 0 0 0 0 0 1 1 1 6 7 1 1

Figure Interval P4 device removal and ovulation in Bos indicus cows treated with estradiol benzoate (1mg) on Day 9 and estradio cipionate (1mg) on Day 8 (implant removal) SALES et al., 2008

36

5/13/2010

5 treatments (n=100) 400 IU eCG


+ PGF2

EB

Implant removal

EB

FTET

CRESTAR
D0 D5 D8 D9 D17

4 treatments (n=100) 400 IU eCG


EB + PGF2

EB

FTET

CRESTAR
D0 D5 D8 D9 D17

3 treatments (n=96)
EB 400 IU eCG + 0.5mg EC + PGF2

FTET

CRESTAR
D0 D5 D8 D9 D17

100 90 80 70 60 50 40 30 20 10 0

92

93 96

5 treatments 4 treatments 3 treatments

EC
59 42
b a

61.6 46.6 45.2


b b a

42
b

(92/100) (93/100) (96/100)

42/92

42/93

59/96

(42/100) (42/100) (59/100)

Transferred/treated

Preganat/transferred

Pregnant/treated

Effect of the day of the eCG treatment (Day 5 vs Day 8) and estadiol treatment (EC on Day 8 vs EB on Day 9) on the FTET protocol in Bos indicus x Bos Ferreira et al., 2006 (Animal Reproduction) taurus recipients.

37

5/13/2010

FTET protocol (IETS 2002)


eCG + PGF EB (2mg) + P4 (50mg) 400 IU eCG
+ PGF2

Implant removal

EB
(1mg)

FTET

P4
D0 D5 D8 D9 D17

5 handlings
B et al., 2002

90 80 70 60
% 50

87 81.7

Day 5 Day 8

51.8 45
a b a b

44.1 36.6

40 30 20 10 0
260/299 241/295 132/255 108/240 132/299

108/295

Transferred/treated

Pregnant/transferred

Pregnant/treated

Effect of the day of the eCG treatment (Day 5 vs Day 8) on the FTET protocol in Bos indicus x Bos taurus recipients.
Reis et al., 2004 (ICAR)

38

5/13/2010

EB

PGF2 + eCG

PGF2 + eCG

P4
LH

OR DF CL P4 P4 OR DF CL

14
mm 10

6 2

Day 0

Day 5

Day 8

PGF

Hypothesis
Follicular Diameter (mm) P4 device + EB 12 10 8 6 4
LH

P4

Day 0

Days

39

5/13/2010

Synchronization of ovulation protocol

Group 1: Bos indicus (n=23) Group 2: B. indicus x B. taurus (n=25) Group 3: Bos taurus (n=22)
PGF PGF with, without PGF
EB (2,0mg)

PGF

EB (1mg)

CIDR

D-24 D-12

D0
US 24/24h

D8

D9
US 12/12h

D13

Synchronization ovulation protocol in Bos indicus vs Bos taurus heifers

Nelore heifers (Bos indicus)

Nelore X Angus heifers (Bos indicus x Bos taurus)

Angus heifers (Bos taurus)

Same management

40

5/13/2010

Holstein heifers (Bos taurus)

Girolando heifers (Bos taurus x Bos indicus)

Gir heifers (Bos indicus)

Same management

EB (2mg)

PGF EB (1mg)

EB (2mg)

PGF EB (1mg)

CIDR
8,0 7,0 6,0 P4 (n g /m l) 5,0 4,0 3,0 2,0 1,0 0,0 DIA 0 DIA 3 DIA 6 DIA 8 DIA 10

1,4
com PGF no DO (n=34) 1,2

CIDR
C o m P GF no D 0 (n=34) Sem P GF no D 0 (n=36)

a a a b b b

sem PGF no DO (n=36)

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

1 0,8 0,6 0,4

**

PGF

0,2 0 D0 D1 D2 D3 D4 D5 D6 D7 D8 D9 D10-M

Fig. Serum progesterone concentrations in B. indicus, B. indicus B. taurus, and B. taurus heifers treated with an intravaginal progesterone-releasing , device (CIDR) for 8 days, estradiol benzoate (EB) with or without PGF at . device insertion (Day 0), and PGF at device removal (Day 8).
Carvalho et al. (15th ICAR. v.1, p.112, 2004 )

41

5/13/2010

EB (2mg)

PGF

EB (1mg)

1,4 1,2 1 0,8 0,6

CIDR
*
C o m P GF no D 0 (n=34 ) Sem P GF no D 0 (n=36 )

Ovulation rate
* 79,4% (27/34)a

* * * ** ** ** 52,7% (19/36)b

PGF
0,4 0,2 0 D0 D1 D2 D3 D4 D5

** **

D6

D7

D8

D9

D10-M

Fig. Serum progesterone concentrations and follicular dynamics in B. indicus, B. indicus B. taurus, and B. taurus heifers treated with an CIDR) for 8 days, estradiol benzoate (EB) with or without PGF at device insertion (Day 0), and PGF at device removal (Day 8).

Carvalho et al. (15th ICAR. v.1, p.112, 2004 )

No PGF on Day 0 (n=99)


EB
(2.0mg)

EC (1.0mg)

400 IU eCG
PGF2

FTET + US

CIDR

Ultrasonography (12/12h)

D0

D8
EC (1.0mg)

D13

D17

PGF on Day 0 (n=99)


EB
(2.0mg)

400 IU eCG
PGF2

+ PGF CIDR

FTET + US

Ultrasonography (12/12h)

D0

D8

D13

D17

42

5/13/2010

20 18 16 14 12 % 10 8 6 4 2 0

No PGF on Day 0 PGF on Day 0


14.7 12.6

18.1

19.3

b
P = 0.09

P < 0.05

Diameter of ovulatory follicle

CL diameter at ET

Figure. Embryo recipients treated or not with PGF on Day 0 of FTET protocol. Reis et al., 2008 (SBTE)

100 90 80 70 60 % 50 40 30 20 10 0

83.8 90.9

No PGF Day 0 (n=99) PGF Day 0 (n=99)

P = 0.07
47 55.6 39.4 50.5

a
Transferred/treated Pregnant/transferred

Pregnant/treated

Figure. Embryo recipients treated or not with PGF on Day 0 of FTET protocol. Reis et al., 2008 (SBTE)

43

5/13/2010

FTET protocol (IETS 2002)


PGF
EB (2mg) + P4 (50mg) 400 IU eCG
+ PGF2

eCG
Implant removal EB
(1mg)

FTET

P4
D0 D5 D8 D9 D17

5 handlings
B et al., 2002

80 70 60 50
% 40

79.7 70.5

PGF Day 0 + eCG Day 8 PGF and eCG on Day 5

58.6 50.6

P > 0.05
40.3 41.4

30 20 10 0
Transferred/treated Pregnant/transferred Pregnant/treated

305

312

(123/305) (129/312)

Figure. Embryo recipients treated with PGF on Day 0 and eCG on Day 8 or with PGF and eCG on Day 5.
Peres et al., 2007

44

5/13/2010

FTET protocol (IETS 2002)


P4 i.m. on Day 0?
EB (2mg) + P4 (50mg) 400 IU eCG
+ PGF2

Implant removal

EB
(1mg)

FTET

P4
D0 D5 D8 D9 D17

5 handlings
B et al., 2002

90 80 70 60
%

86.7 87.4

EB on Day 0 EB + P4 on Day 0

52.3 48.5

P > 0.05
45.3 42.4

50 40 30 20
130/150 132/151 68/130

64/132

68/150

64/151

10 0
Transferred/treated Pregnant/transferred Pregnant/treated

Figure. Embryo recipients treated or not with progesterone (50mg i.m.) on Day 0 of FTET protocol.
Nasser et al., 2004 (Theriogenology)

45

5/13/2010

Proposed new protocol for fixed-time embryo transfer (FTET) in cattle (three treatments)

EB (2 mg) + PGF2 P4 device

EC (1.0 mg) + PGF2 + eCG 400 IU

TETF

D0

D8

D17

FTET with P4 + estadiol and eCG


2,525 recipients
Transferred / treated rate (84.9%)

Pregnant/treated rate (42.9%)

2,146 transferred
Pregnant/transferred rate (50.5%)

1,084 pregnant

USP data 2009

46

5/13/2010

Effect of FTET on reproductive efficiency in high-producing Holstein cows

Dairy farms

47

5/13/2010

50 45 40 35 30 25 20 15 10 5 0

Summer

Fall-winter

Spring

30 25
Envroumental temperature (C)

36,3 31,3 24,6 24,3 20,6 20,5 27,5 39,7

38,7

39,6

pregnancy rate (%)

30,5

15 to 20% 24,6

20 15

AI

T (C)

10 J F M A M J J A S O N D
months

Figure. Conception rates in lactating Holstein cows submitted to AI (n=7501) throughout the year (2000 to 2003).
Rodrigues et al., 2004 (ICAR)

Retrospective study
Lactanting Holstein cows (milk production = 28.4 2.3 kg/day) 2,112 in vivo embryo produced 7,501 artificial insemination AI 12 h after estrus detection (farm personal) ET 7 days later estrus detection in cow with CL > 16mm (performed by the same technician)

Rodrigues et al. (ICAR 2004)

48

5/13/2010

Sammer

Fall-winter

Spring

Figure. Conception rates in lactating Holstein cows submitted to AI (n=7501) or embryo transfer (n=2112) throughout the year.

Rodrigues et al. (ICAR 2004)

AI

90 80 70
pregnancy rate (%)

Sammer

ET Fall-winter AI x ET (P<0.05) T (C)

Spring

30 25
temperature (C)

60 50 40 30 20 10 0 J F M A M J J A S O N D
months

20 15 10 5 0

Figure. Conception rates in repeat breeder lactating Holstein cows submitted to AI (n=2803) or embryo transfer (n=2066) throughout the year. Rodrigues et al., 2004 (SBTE)

49

5/13/2010

Group FTET with CL on Day 0 (n=208)

BE+P4

400 UI eCG + 0,5 ml ECP + PGF2

TETF

P4
D0 D8 D17

Group FTET without CL on Day 0 (n=221)

BE+P4

400 UI eCG + 0,5 ml ECP + PGF2

TETF

P4
D0 D8 D17

90 80 70 60 50 40 30 20 10 0

75,0

TETF - CL TETF - No CL
**

61,2

39,1 37,4 29,3 22,9


(156/208) (131/214)

(67/156) (49/131)

(61/208) (49/214)

Transferred/ treated

Pregnant/ transferred

Pregnant/ treated

Effect of the presence of CL at the beginning of the protocol for FTET on fertility rates of high-producing Holstein repeat-breeder recipients
Rodrigues et al SBTE (2007)

50

5/13/2010

Can we replace estradiol with GnRH in the FTET protocol for lactating Holstein cows?
Estradiol is not available for synchronization in some countries

Estradiol vs GnRH on the FTET protocol


N=390 lactanting cows
EB eCG (400 UI) + ECP (1.0 mg) + PGF FTET

Crestar
D0 D8 D17

N=394 lactanting cows


GnRH

eCG (400UI) + PGF GnRH FTET

Crestar
D0 D7 D9 D16

Unpublished data

51

5/13/2010

90 80 70 60 50 % 40 30 20 10 0

Estradiol GnRH

68.0 56.4 36.8 33.6


a b

20.8 22.8
(220/390) (268/394) (81/220) (90/268) (81/390) (90/394

Transferred/ treated

Pregnant/ transferred

Pregnant/ treated
Unpublished data

Figure. Pregnancy rates in high-producing Holstein repeat-breeder recipients transferred at a fixed-time and treated or not treated with 4000IU of eCG.

Main remarks
The FTET protocol increased the proportion of recipients transferred and improved pregnancy rates in repeat breeder high-producing dairy cows

52

5/13/2010

Main conclusions
FTET protocols using progesterone releasing devices and eCG improve the efficiency of the ET programs by: 1. Increasing the number of recipients receiving an embryo (Beef= 85%; Dairy = 70%) 2. Without affecting conception rates 3. Resulting in higher pregnancy rates

Furthermore, the FTET protocol is easy to perform by farm personnel, reducing labor and animal handling and facilitating the use of ET

53

5/13/2010

What was the proportion of recipients transferred using FTET in 2009?

Around 70%

300,000 ET / year

~210,000 FTET / year

Thank you

54

5/13/2010

ACKNOWLEDGMENTS
Graduation Students
Everton Luis Reis
Mrcio de Oliveira Marques Manoel de S Filho Lindsay Gimenes Luis Nasser Roberto Mendes Porto Filho Antnio Jorge Del Rei Moura Cludio Coutinho Bartolomeu Joo Batista de Carvalho Marcelo Trigo Nelcio Antnio Tonizza de Carvalho Rodolfo Cassimiro de A. Berber Henderson Ayres Claudiney Martins Alexandre Henrily Souza Jos Ribamar Torres Junior Mrcio Leo Ferraz Jos Nelio de Sousa Sales Gabriel Armond Crepaldi Alessandra Teixeira Roberta Ferreira

Institutions

FAPESP CNPq Field veterinarians Farm owners Partner companies

55