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A STUDY OF THE RESULTS OF AN EMB RYO TRANSFER PROG RAMME
CONDUCTED DURI NG TWO SEASONAL PERIODS
USI NUFIVE IMPORTED BREEDS OF SHEEP
A thesis presented in partial fulfilment of the requirements for the degree of
Master of Agriculture Science in Animal Science at Massey U niversity
KENNETH BARRY GREANEY 1 99 1
ABSTRACT
The overall objective of this trial was to compare embryo transfer results from two seaso nal periods (out-of-season vs in-season) in order to determine the effect of the season on the different parameters of reproductive performance such as ; the incidence of oestrus, ovarian response to exogenous gonadotrophins, fertilisation rates and the nu mber of lambs born per donor ewe programmed and flushed.
Five breeds of imported sheep (Danish Texel {DT}, Finnish Texel {FT}, Gotland Pelt {GOT}, Oxford Down {OXD}, and the White Headed Marsh {WH M}) consisting of two age groups (14-16 month-old and 26-28 month-old) in a commercial embryo transfer programme (LambXL, Manawatu) were used.
A total of 553 ewes out-of-season and 234 ewes in-season were studied. Two data sets were selected from these seasonal groups ;
(1) The random data set.
(2) The repeat data set, which consisted of the same donor ewes in each seasonal group.
Oestrous synchronisation was attempted with a double CIDR-G regime and a super-ovulatory treatment consisting of an initial PMSG injection (200-300 1U) and a seri es of six descending doses of FSH-P (total dose 24-36mg). The ewes were inseminated intra-uterine with fresh diluted semen from a ram of the same breed on the basis of oestrous detection. Embryo recovery was attempted on day 6.5-7 after oestrous detection using a laparoscope-aided uterine flush technique. Two e mbryos were transferred to each synchronised recipient ewe within two hours of recovery.
The i ncidence of oestrus for the out-of-season and i n-season g roups was 93.3%
and 1 00%, for the random data set compared to 93.9% and 1 00% for the repeat data set, respectively. The ovulatory response to the super-ovulatory treatments
iii was significantly affected by the interaction of the breed and age of the donor in the random data set, but the repeat data set ovulation rate was not significantly affected by any of the variables recorded in this study. This interaction was attributable to the GOT breed having a higher ovulation rate in the older age g roup relative to the you nger age g roup which was the reverse trend exhibited by the remaining breeds. However, there was an overall tendency for the out-of
season ovulation rate to be higher than that in-season, 7.64CL vs 6.60CL for the random data set and a difference (out-of-season - in-season) of +2.86CL was recorded for the repeat data set. The embryo recovery rates were 53.4% out-of
season and 53.5% in-season for the random data set and a difference of -0.7 percentage points was recorded for the repeat data set. The fertilisation rate was not significantly affected by the season with 75.5% out-of-season and 65.7% in
season from the random data set and a difference of -4.5 percentage points was recorded for the repeat data set. The yield of good quality transferable embryos was significantly affected by the season with 78.2% out-of-season and 83.7% in
season from the random data set but the repeat data set was not significantly affected by the season with a difference of -1 1 .0 percentage points. The embryo survival rate to birth was not significantly different for the two seasonal periods with 66.3% out-of-season and 52.4% in-season of the embryos surviving to birth for the random data set and a difference of +9. 7 percentage points was recorded for the repeat data set. This resulted in an average of 1 .66 lambs born per donor ewe programmed out-of-season , which was not significantly different from 1 .00 lambs born in-season for the random data set compared to a difference of +0.1 1 lambs born per donor ewe programmed in the repeat data set.
This work clearly demonst rates the inter-dependence of several factors affecting the number of lambs born per donor ewe in an embryo transfer programme.
However it is concluded that out-of-season embryo transfer is as effective as that conducted in-season, u nder these embryo transfer conditions.
ACKNOWLEDGEMENTS
I wish to thank my supervisor Or M F McDonald.
My sincere gratitude goes to Or H William Vivanco for his patience and efforts throughout the entire duration of this study. He taught me most of the technical skills required in embryo transfer procedures as well as the theoretical knowledge which enabled me to conduct the appropriate analysis and compile this thesis.
Thanks also must be given to the technical staff at LambXL for their direct assistance with the experimental work. The financial support given to me by LambXL is also gratefully acknowledged.
The assistance and helpful suggestions of Raymondo Rangei-Santos are acknowledged.
I am indebted to Or Huge Varela and Or D J Garrick for statistical advice and extreme patience, especially i n the latter case.
The helpful suggestions and support of the Animal Science Department are gratefully acknowledged.
Finally very special thanks to my family, in particular my fiance, Catheri ne, without whose support and encouragement this thesis would not have been possible.
V
TAB LE OF CONTENTS
ABSTRACT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ii
ACKNOWLEDGEMENTS . . . . . . . . . . . . . . . . . . . . . . . . . iv
TABLE OF CONTENTS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . v
CHAPTER ONE . . . 2
INTRODUCTION . . ... . .. . . .. . .. . .. . . . ... . . .. . . ... . . .. 2
CHAPTER TWO . . . 5
LITE RATU R E REVIEW . . . 5
2.1 Oestrous synchronisation . . . 6
2.2 Number of ovulations . . . 9
2.2.1 Physiology of the oestrous cycle and ovulation . . . 9
2.2.2 The seasonality of reproduction . . . 13
2.2.3 Manipulation of the number of ovulations using e xogenous gonadotrophins . . .. . . . 18
2.2.4 Out-of-season hormonal regimes . . . 21
2.3 Recovery rate . . . 23
2.3.1 Embryo development rate . . . 24
Table 2.1 Embryo development rate . . . 25
2.3.2 Day of flush . . . 25
2.3.3 N umber of ovulations . . . 26
2.3.4 Embryo recovery technique . . . 26
2.3.5 N u mber of previous flushes . . . 27
2.3.6 Out-of-season embryo recovery . . . 27
2.4. Fertilisation rate . . . 29
2.4.1 Artificial insemination tech nique . . . 29
2.4.2 AI time relative to oestrus and progesterone removal . . . 30
2.4.3 Semen processing . . . 32
2.5 The yield of good quality transferable embryos . . . 33
2.6 E mbryo survival rate . . . 34
2.6.1 Embryo quality and stage of development . . . 34
2.6.2 Number of embryos transferred per recipient . 35 2.6.3 Recipient synchrony treatment . . . 36
2.6.4 Recipient synchrony . . . 36
2.6.5 Site of transfer . . . 37
2.6.6 Interval between collection and transfer . . . 38
2.6.7 Number of corpora lutea present in the recipient . . . 38
2.6.8 The effect of supplementary progesterone . . . 39
2.6.9 Recipient breed . . . 41
2.6.1 0 Recipient nutrition 41 CHAPTER THREE . . . 44
MATERIALS AND METHODS . . . 44
3.1 Experi mental animals . . . 44
3.1 .1 Donor ewes . . . 44
Table 3.1 The nu mber and age of the donor ewes . . . . 45
Table 3.2 Body weights of the 14-16 month-old donor ewes 45 3.1.2 Rams . . . 46
Table 3.3 The numbers and ages of the rams used . . . 46
3.1.3 Recipient ewes . . . 46
3. 1 .4 Teaser rams . . . . 47
3.2 Expe ri mental plan . . . 47 Table 3.4 The numbers and ages of all of the donor ewes
vii
programmed within regime and age groups . . . . 48
Table 3.5 The number of times the donor ewes were program med . . . . 49
3.3 Synchro nisation of oestrus . . . 50
3.4 Super-ovulatory hormonal regimes . . . 50
Table 3.6 Super-ovulatory hormonal regimes . . . . 51
3.5 Oestrous detection . . . 51
3.6 Timing of i nsemination . . . . 52
Table 3. 7 Timing of insemination . . . . 52
3. 7 Artificial i nsemination technique ........ . . . ·. . . . 52
3.8 E mbryo recovery technique . . . . 53
3.9 E mbryo classification . . . . 54
Table 3.8 E mbryo classifications . . . . 54
3.1 0 Transfer to recipient . . . . 54
3.11 Manage ment of the recipient ewe after transfer . . . . . 55
3.12 Pregnancy testing and lambing . . . 55
3.13 Analysis of the data . . . . 56
3.13.1 I ncidence and time of oestrous detection . . . 58
3.13.2 Ovulation rate . . . . 60
3.13.3 Recovery and fertilisation rates, and the yield of good quality transferable embryos . . . . 61
3.13.4 Embryo survival to scanning or bi rth, and the number of lambs born per donor ewe programmed and flushed . . . 64
CHAPTER FOUR . . . . 67
RESULTS . . . 67
4.1 Incidence and time of oestrous detection . . . . 67
4.1.1 The incidence of oestrus . . . . 67
Table 4.1.1 The effect of season on the incidence of oestru s for the random data set . . . 68
Table 4.1.2 The effect of season on the incidence of oestrus for the repeat data set . . . 68
4.1.2 The time of oestrous detection . . . 69 Table 4.1.3 The effect of season on the time of oestrous
detection for the random data set . . . 69 Table 4.1.4 The effect of season on the time of oestrous
detection for the repeat data set . . . 69 4.2 Ovulation rate . . . 71 Table 4.2.1 Overall ovulation rate for the two seasonal
periods for the random data set . . . 72 Table 4.2.2 Ovulation rate for the donor breed by donor age
interaction for the random data set . . . 72 Table 4.2.3 Ovulation rate within season and donor breed
for the repeat data set . . . 72 4.3 Recovery rate . . . 73 Table 4.3.1 Recovery rate for the two seasonal periods for
the random data set . . . 7 4 Table 4.3.2 The effect of the time of oestrous detection on
the recovery rate for the random data set . . . 7 4 Table 4.3.3 The effect of donor surgeon on the recovery
rate for the random data set . . . 7 4 Table 4.3.4 Recovery rate for the two seasonal periods for
the repeat data set . . . 75 Table 4.3.5 The effect of donor surgeon on the recove ry
rate for the repeat data set . . . 75 4.4 Fertilisation rate . . . 76 Table 4.4. 1 Fertilisation rate within season and breed for
the random data set . . . 77 Table 4.4.2 The effect of interval from CIDR-G removal to
oestrous detection on the fertilisation rate for the
random data set . . . . 77 Table 4.4.3 Fertilisation rate for the two seasonal periods
tor the repeat data set . . . 77 4.5 Embryo quality . . . . 78 Table 4.5.1 The yield of good quality transferable embryos
ix for each seasonal period for the random data set . . 79 Table 4.5.2 The yield of good quality transferable embryos
for each donor breed for the random data set . . . 79 Table 4.5.3 The effect of interval from C l DR-G removal to
oestrous detection on the yield of good quality
transferable embryos for the random data set . . . 79 Table 4.5.4 The yield of good quality transferable embryos
the two seasonal periods for the repeat data set . . . 80 4.6 Survival of the embryos to day 50-55 after transfer . . . 81 Table 4.6.1 Embryo survival rate to scanning for the
random data set . . . 82 Table 4. 6.2 Embryo survival rate to scanning for the two
seasons for the repeat data set . . . 82 4. 7 Embryo survival to birth . . . 83 Table 4.7.1 Embryo survival rate to birth for the random
data set . . . 84 Table 4.7.2 Embryo survival rate to birt h for each season
for the repeat data set . . . 84 4.8 Lambs born . . . 85 Table 4.8.1 N u mber of lambs born for the random data
set . . . 86 Table 4.8.2 N u mber of lambs born for the repeat data set . 86 4.9 Summary of the overall MOET programme . . . 87 Table 4.9.1 S u mmary of the results of the overall MOET
prog ramme for the random data set . . . 89 Table 4.9.2 S ummary of the results of the overall MOET
programm e for the repeat data set . . . 89 Table 4.9.3 S um mary of the difference in results of the
overall MOET programme for the repeat data set . . 90
CHAPTER FIVE . . . 92 D ISCUSSION AND CONCLUSIONS . . . 92 5. 1 I ncidence and the time of oestrous detection . . . 93
5.1.1 The incidence of oestrus . . . 93 5.1.2 The time of oestrous detection . . . 95 5.1.3 Summary of the factors affecting the incidence
and tim e of oestrous detection . . . 96 5.2 Ovulation rate . . . 97
5.2.1 The interaction between the donor ewe breed
and age . . . 97 5.2.2 The effect of the season on the ovulation rate 99 5.2.3 The effect of the hormonal regime on the
ovulation rate . . . 1 02 5.2.4 Summary of the facto rs affecting the ovulation
rate . . . 103 5.3 Recovery rate . . . 1 04 5.3.1 The effect of the season on the recovery rate 1 04 5.3.2 The effect of the donor surgeon used on the
recovery rate . . . 104 5.3.3 The effect of the time of oestrous detection on
the recovery rate . . . 105 5.3.4 The recovery rate and the non-sig nificant
factors . . . 1 06 5.3.5 Summary of the factors affecting the recovery
rate . . . 109 5.4 Fertilisation rate . . . 110
5.4.1 The effect of the season on the fertilisation
rate . . . 110 5.4.2 The effect of the time of oestrous detection on
the fertilisation rate . . . 11 0 5.4.3 The fertilisation rate and the non-significant
factors . . . 111 5.4.4 Summary of the factors affecting the fertilisation
rate . . . 113 5.5 E mbryo quality . . . 114
5.5. 1 The effect of the season on the yield of good
xi quality transferable embryos .. . . 1 1 4 5.5.2 The effect of the donor breed on the yield of
good quality transferable embryos .. .. .. . . 1 1 5 5.5.3 The effect of the time of oestrous detection
after CIDR-G removal on the yi eld of good
quality transferable embryos . . . .. . .. ... . 1 1 5 5.5.4 The yield of good quality transferable embryos
and the non-significant factors . . . 1 1 6 5.5.5 Summary of the factors affecting the yield of
good quality transferable embryos . .. . . 1 1 6 5.6 E mbryo survival to scanning and birth . . . . .. . . 1 1 7
5.6. 1 The effect of the season on embryo survival to
scanning and birth ... . .. . . . ... . .... 1 1 7 5. 6.2 The e mbryo survival to scanning and bi rth and
the non-significant factors . . . .. ... . . . .. . 1 1 7 5.6.3 Summary of the factors affecting the embryo
survival to scanning and birth . . . . . . . 1 1 9 5.7 Number of lambs born . . . . .. . . 1 20 5.8 Overall conclusions ... . .. . . .. .. . . .. .. . . . 1 22
CHAPTER SIX . . . . 1 24 R E FERENCES . . . 1 24
CHAPTER SEVEN . . . 1 43 APPEN D ICES . . . 1 43 APPE N D IX ONE - RANDOM DATA . . . . 1 44 Table 7 . 1 .1 The number of donor ewes in the random
g roup
Table 7 . 1 .2 The number of donor ewes in the random
1 45
group for each breed, age and regime combination 1 46 Table 7 . 1 .3 The effect of season on the incidence of
oestrus for each breed . . . . 1 46 Table 7 . 1 .4 The effect of season on the time of oestrous
detection for each breed . . . 147 Table 7 .1.5 The effect of season and ovulation rate class
on the time of oestrous detection . . . 148 Table 7 .1.6 Ovulation rate within season and donor breed 148 Table 7 .1.7 Ovulation rate withi n donor breed and donor
age . . . 149 Table 7 .1. 8 Ovulation rate for each breed, age, hormonal
regi me and season combination . . . 150 Table 7 .1.9 Recover:y- rate within season and breed . . . . 151 Table 7 .1.1 0 Fertilisation rate within season and breed . . 151 Table 7 .1.11 The yield of good quality transferable embryos
within season and breed . . . 152 Table 7 .1.12 Embryo survival rate to scanning . . . 153 Table 7 .1.13 Embryo su rvival rate to birth for each season
and breed . . . 1 54 Table 7 .1.14 Number of lambs born . . . 155 Table 7 .1.15 Summary of the results of the overall MOET
programme for the random data . . . . 156 APPEN D IX TWO - REPEAT DATA . . . . 157 Table 7 .2.1 The number of donor ewes in the repeat
group
Table 7 .2.2 The nu mber of donor ewes in the repeat g roup
158
for each breed, age and regime combi nation . . . 158 Table 7 .2.3 The effect of season on the incidence of oestrus . . . 159 Table 7 .2.4 The effect of season on the time of oestrous
detection . . . 159 Table 7 .2.5 The effect of season and ovulation rate class
on the time of oestrous detection . . . 160 Table 7 .2.6 Ovulation rate within season and donor breed 161 Table 7 .2.7 Ovulation rate for each breed, age, hormonal
regi me and season combination . . . 162 Table 7 .2.8 Recovery rate within season and breed . . . . 163 Table 7 .2.9 Fertilisation rate within season and breed . . 164
xiii Table 7.2. 1 0 The yield of good quality transferable
embryos within breed and season . . . 1 65 Table 7 .2. 1 1 Embryo survival rate to scanning within
season and breed . . . 1 66 Table 7 .2. 1 2 Embryo survival rate to birth within season
and breed . . . 1 67 Table 7 .2. 1 3 Number of lambs born . . . 1 68 Table 7 .2. 1 4 Sum mary of the results of the overall MOET
programm e . . . 1 69 Table 7 .2. 1 5 Su mmary of the difference in results of the
overall MOET programme . . . . 1 70
