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Genetic studies on fertility in A.I. bulls. I. Age, season and genetic effects on semen characteristics in young bulls
Animal Reproduction Science, 19 (1989) 1-17
1
Elsevier Science Publishers B.V., Amsterdam - - Printed in The Netherlands
Genetic Studies on Fertility in A.I. Bulls. I. Age, S e a s o n and Genetic Effects on S e m e n Characteristics in Y o u n g Bulls E V A - M A R I E S T A L H A M M A R , L E N N A R T J A N S O N and J A N P H I L I P S S O N
Department of Animal Breeding and Genetics, Swedish University of Agricultural Sciences, Box 7023, 750 07 Uppsala (Sweden) (Accepted 17 October 1988)
ABSTRACT St~lhammar, E.-M., Janson, L. and Philipsson, J., 1989. Genetic studies on fertility in A.I. bulls. I. Age, season and genetic effects on semen characteristics in young bulls. Anim. Reprod. Sci, 19: 1-17. Genetic and environmental factors that influence semen quantity and quality were evaluated for 215 young bulls of the two main dual-purpose breeds in Sweden, Swedish Red and White (SRB) and Swedish Friesian breed (SLB). Semen was collected at three bull stations from September 1983 to August 1985, inclusive. The effects of bull's sire, bull, season and year of collection, and bull's age at collection were estimated for ejaculate volume, motility, total no. of sperm cells per ejaculate and no. of semen doses per collection. Heritabilities (h2), repeatabilities (t) and phenotypic correlations were estimated for these traits. Motility and ejaculate volume were significantly affected by bull's sire (P < 0.01 ). The effect of bull, season of collection, and bull's age at collection were highly significant (P < 0.001 ) for most of the traits. The effect of year of collection was only significant for one semen characteristic. The summer months June, July and August were the best months for quantitative traits. Estimates of the heritability for semen characteristics ranged between 0.02 and 0.28, with the highest estimates found for ejaculate volume and semen doses per collection. The h2-estimates differed somewhat between breeds. The repeatabilities ranged between 0.5 and 0.6 for all the traits studied. It was concluded that the h2-estimates were sufficiently high to permit selection on individual bull level for some of the traits. At least 20 ejaculates have to be considered for total no. of sperm cells per ejaculate and post-freezing motility in order to achieve a selection accuracy (rTi) of 0.3-0.5. One must take into account a bull's age and season of collection when evaluating semen quality.
INTRODUCTION
The fertility of bulls is an important consideration, since each bull can be used for a large number of inseminations. In Sweden, it has been the practice hitherto to slaughter bulls once 30 000-45 000 semen doses have been col-
0378-4320/89/$03.50
© 1989 Elsevier Science Publishers B.V.
2 TABLE 1 Review of some parameters, means (x), standard deviations (SD), heritabilities (h 2) of some semen characteristics in the literature Characteristic
2
SD
h2
Pre-freezing motility, %
66 57 59.7 58.6 8.0 2.81
2.4
Hafs et al., 1958 Seidel and Foote, 1972 Everett et al., 1978 Everett and Bean, 1982 Hafs et al., 1958 Hultn~is, 1959 0.10-0.21 Maijala, 1969 Seidel and Foote, 1972 Everett et al., 1978 Everett and Bean, 1982 Jakubiec, 1983
Volume, ml
5.9 7.8 5.80 3.8 7.9 6.3 5.0 Total no. of sperm cells)< 109 per ejaculate
13.6 7.23 8.67 10.3 7.81 3.064 5.8-8.6 10.9 7.4 6.9
Post-freezing motility, 64.1 % 53 60.91 33.06
3.81 9.0 1.52
1.77 0.3 3.1 3.0 2.3
0.16
Taylor et al., 1985
0.12
Taylor and Everett, 1985 Hafs et al., 1958 Hultn~is, 1959 Seidel and Foote, 1972 Everett et al., 1978 Everett and Bean, 1982 Jakubiec, 1983 Jansen, 1984
0.05
Taylor et al., 1985
0.02
Taylor and Everett, 1985 Lasley, 1951 Seidel and Foote, 1972 Uwland, 1984 Chandler et al., 1985
6.0 3.18 3.31 0.829 5.7 5.0 4.5
6.24 7.99
Reference
}
0.21
lected, except at one A.I. station. Now, however, with the i n t r o d u c t i o n of a new system, in w h i c h the bulls are k e p t alive d u r i n g the process of p r o g e n y testing, the A.I. s t a t i o n s will be able to m a k e m o r e intensive use of genetically superior bulls. It is of b o t h genetic a n d e c o n o m i c i n t e r e s t to a s c e r t a i n w h i c h factors influence the q u a n t i t y a n d quality of semen. T h e possible r e l a t i o n s h i p b e t w e e n various s e m e n c h a r a c t e r i s t i c s a n d fertility is of p a r t i c u l a r i m p o r t a n c e . S u c h a r e l a t i o n s h i p has been a s s e r t e d by L i n f o r d et al. (1976) a n d W o o d et al. (1986) a n d will be i n v e s t i g a t e d u s i n g Swedish d a t a in a f o r t h c o m i n g study. T h e r e are i n t e r e s t i n g older studies on s e m e n quality f r o m S w e d e n a n d several o t h e r c o u n t r i e s (Table 1). B a n e (1954) studied m a t i n g b e h a v i o u r a n d
semen characteristics in monozygous bull twins and found differences between the genotypes. Marked similarities between twin brothers and wide disparities between pairs showed that heredity exerted considerable influence on mating ability and libido. There are also appreciable variations in sex drive between bulls kept under similar conditions (Bishop, 1970). Hultn~is (1959) showed that Swedish Red and White bulls in the age interval 15-26 months increased ejaculate volume on average 3% for each month of life. The practices of discarding ejaculates for the sake of maintaining a high semen quality and the culling of bulls with the purpose of achieving a good conception result have been shown to differ considerably between Swedish bull stations (Bernes, 1981 ). The ultimate effects of different culling policies are not well known. The study by Bernes (1981) showed breed differences in bull fertility. Age of bull, season, and the frequency of ejaculation are known to influence semen output (Everett et al., 1978; Almquist, 1982; Everett and Bean, 1982 ). The purpose of the present study was to determine the effects of different environmental factors such as season, year and age of bull on libido and various semen characteristics under routine Swedish A.I. conditions. The intention was also to study genetic variability in these traits and their mutual interrelationships. MATERIALSAND METHODS
Materials Data on semen characteristics from 215 young bulls of the two main dualpurpose breeds in Sweden, Swedish Red and White (SRB) and Swedish Friesian Breed (SLB) were obtained from three A.I. stations. The data were recorded in the normal semen collection routine between September 1983 and August 1985, inclusive. The bull stations were Elitsemin (E), Nordavel {N) (SRB-bulls) and L~glandavel, Falkenberg (F) (SLB-bulls). The bulls were sons of 40 sires with 2 to 14 sons per sire (Table 2 ). The first collection started from 11 to 17 months of age. The oldest bulls in this study were about 3 years TABLE 2 Numberof bulls and sires in the materialsanalysed Bull station
Breed
No. of bulls
No. of sires
Elitsemin Nordavdel L~glandsavel, Falkenberg
SRB SRB SLB
62 70 83
11 14 15
4 TABLE 3 Number of semen collections per season in the materials analysed Bull station
Season
Elitsemin Nordavel L~glandsavel, Falkenberg
Winter
Spring
Summer
Autumn
Total number of collections
1174 685 1089
748 868 1321
790 265 1280
1428 503 972
4140 2321 4662
TABLE 4 Number of collections in the different materials analysed Bull station
Original Collections no. of without collections recorded volume
Elitsemin 4705 Nordavel 2904 L~glandsavel, 8603 Falkenberg
Collections with two ejaculates
No. of collections not included for various reasons
n
%
n
%
n
%
140 30 469
3.0 1.0 5.4
72 253 2571
1.5 8.7 29.9
353 300 901
7.5 10.3 10.5
Maximum no. of collections in the analysis
No. of collections per bull on average
4140 2321 4662
67 33 56
old. In E the majority of semen collections were made in a u t u m n and winter, while in N most were during winter and spring. The collections in F were more evenly spread over the year (Table 3). Table 4 shows the restrictions placed on the materials and the total number of collections eventually analysed. Information on each ejaculate in this study included (1) bull's identity; (2) collection code, including date; (3) ejaculate order number; (4) libido; (5) original volume of ejaculate; (6) total no. of sperm cells per ejaculate; (7) motility, both before and after freezing; (8) no. of semen doses per collection; and (9) any discarding, either before or after freezing. The bull's birth date and sire were added to this information. If a bull was the only son of a sire, it was deleted from the genetic analyses in order to reduce confounding with sire and bull within sire in the analyses. Collections without recorded volume were excluded. W h e n a bull ejaculated more t h a n once on the same collection day it was not possible to distinguish between the ejaculates after freezing, as the two ejaculates were pooled when frozen and were assigned the same collection code. Due to the possible effects of taking two ejaculates per collection day more frequently in F than at the other bull stations, the analyses of pre-freezing characteristics were based on only 1st ejaculates.
Collection routines Young bulls were semen collected once a week. Bulls older than 15-18 months i.e. bulls in production, were ejaculated twice a week. In F, a second ejaculate was taken if the preset limit of 450 semen doses from the first ejaculate was not reached. The bulls were stimulated prior to collection by having another bull waiting at the same time in the collection room. The bulls were permitted one false mount and then restrained before mounting and ejaculating on the teaser bull. F used the same stimulation procedure before taking the second ejaculate. In station F bulls were kept grazing during the summer and they were taken to the collection room in pairs, stimulating each other on the way. All the A.I. stations allowed the bulls with initial sexual disturbance or poor semen to have a trial and learning period of 2-3 months before they were culled.
Recording and evaluation procedures Libido was recorded on a scale from 1-5, with 1 = no sex drive, 3 = good sex drive, and 5 = abnormally vigorous sex drive. The fresh semen was evaluated immediately after each collection. This procedure included measuring the volume (ml) and determining motility subjectively under a phase-contrast microscope. Motility was evaluated as the percentage of forward-motile sperm cells. Spermatozoal concentration was determined spectrophotometrically. The turbidity was compared with a known standard. Total no. of sperm cells per ejaculate was calculated by multiplying the spermatozoal concentration per ml by the volume. From the total no. of sperm cells per ejaculate, the number of semen doses was calculated automatically. The stations had different set standards for the concentration, giving different numbers of sperm cells per semen dose (0.25 ml). The standards were 80 (E), 88 (N) and 70 (F)×106 sperm cells per ml respectively, giving 20 (E), 22 (N) and 17.5 (F) × 106 sperm cells per semen dose. The assessment of motility after freezing and thawing was normally made on 1-2 semen doses per collection. The observer did not know from which bull the sample being evaluated had been obtained. The established minimum standards for motility varied slightly between stations. Minimum pre-freezing motility was 60-70% forward-motile sperm cells and for post-freezing it was 4550%. The recording of the different characteristics, both subjectively and objectively was not always made by the same person.
Statistical methods Data were analysed by using Henderson's method III for estimation of variance components and several procedures within the Statistical Analysis System (SAS, 1982 ). The coefficient of determination (R 2) was calculated as the
sums of squares for the model, divided by the total sums of squares (SS model/ SS total). Adjustments for the degrees of freedom had no effect on the R 2values, due to the small size of the model and large number of observations. Since semen was not collected from each bull every month season rather than month of collection was assigned, each spanning 3 months, starting with winter in December. Age of bull at collection was divided into twelve classes, starting at an age of ~ 12 months and ending with bulls more than 30 months old. The age classes each covered 2 months, beginning from 15 months of age, while ~<12, 13 and 14 months of age were each considered as one class. Two-factor interactions between year, season, and bull's age at collection respectively were analysed, b u t were not found significant ( P > 0.05). These interaction effects were therefore excluded from the final model. The three A.I. stations were analysed separately. The model chosen as the most appropriate for estimation of variance components and correlations for libido and semen characteristics at bull station F was: = tt + si + bij + sek + at +yrm + eijh~mn
Yijklmn gijklmn
-
-
8i bo
sek al yrm
eijklrnn
-
-
observed semen characteristic least squares means effect of ith sire effect o f j t h bull within ith sire effect of kth season of collection (k-- 1...4) effect o f / t h age of bull at collection (/--1...12) effect of mth year of collection ( m = 1...3) random error element with expected mean value zero and variance 2 O" e
The model chosen for E and N, was the same as above, except for the year effect. The sire and bull components were considered random and all other elements in the models were assumed fixed. Preferably the model for SRB also should have included A.I. station, but the sires in common were too few. The above described model was chosen, because of the structure of the materials, with sires neither strictly nested within station nor strictly cross-classified with station. The heritability (h 2) of the semen characteristics studied was estimated through the sire, a~, and the bull within sire, a~:s, component of variance, as h2=4o /(o + ab:s . 2 + de) 2 on individual ejaculates. The repeatability coefficient (t) of the semen characteristics in the full model was calculated within 2 2 2 bull as, t = as2 + ab:s/(as + a~:s + a e2) . To demonstrate the heritability of n ejaculates, an estimate ofh2n was calculated as h2n = n ' h 2 / 1 + ( n - 1 )t. The phenotypic correlations were calculated by using the paternal half-sib method according to Harvey (1977).
RESULTS AND DISCUSSION
The overall means and standard deviations for libido and the semen characteristics are given in Table 5. The overall means and especially the variation in most of the traits being studied varied considerably between the two SRBstations. The SD for total no. of sperm cells was much higher at N. One reason for this difference was the bull's age. This will be discussed later. Motility was judged subjectively and it is therefore uncertain whether differences in motility were real, or merely a reflection of differences in judgement between bull stations. Furthermore the SD both absolutely and relatively was greater for motility after freezing than before freezing. F differed from the other stations especially as regards no. of semen doses per collection. This may largely be due to the high proportion of double ejaculates taken and the lower standards of 17.5× 108 sperm cells per semen dose applied. Although only 1st ejaculates were analysed, a possible effect of a second ejaculate could be discerned in F. In general, the overall means and SD in this investigation were consistent with many of the results found in the literature (Table 1 ). Motility was expressed differently in the literature, both as progressive motile and as percentage living spermatozoa. The evaluation of post-freezing motility was done as a 'blind-test' at all stations, as recommended by Amann (1981), which would minimize the bias of the observer. Compared to the results of Hultn~is (1959) the semen volume of young SRB bulls has almost been doubled. Mean volume of first ejaculate was 2.8 ml versus 5.1 to 5.3 ml in the present study. Although the collection routines were different, these results could indicate a genetic improvement, since one selection criterion has been an increased minimum volume of semen. Both volume and total no. of sperm cells per ejaculate are TABLE5 Overall means (2) and standard deviations (SD) for the semen characteristics analysed Breed
SRB (E)
No. of 1st ejaculates
4140 3870 a SRB (N) 2321 2177 a SLB (F) 4662 4176 a
Pre-freezing motility, %
Volume, ml
Total no. of sperm cells per ejaculate, × 106
No. of semen doses per collection
£
SD
£
2
£
SD
75
4
5.2 1.8
6930 2478 340
114
52 8
69
7
5.5 2.2
6961 4828 328
213
48 7
69
5
5.1 1.8
6466 2911 500 b 155
53 6
SD
SD
aNo. of observations after freezing and thawing. bDouble ejaculates to a greater extent than in E and N.
Post-freezing motility, %
SD
TABLE
6
Significance levels for libido and semen characteristics from least-square analyses of variance and c o e f f i c i e n t s o f d e t e r m i n a t i o n ( R 2)a f o r t h e t h r e e b u l l s t a t i o n s Source of variation
Sire
Bull
Season of collection Age of bull at collection Year of collection R 2, %
A.I. stud
Significance levels Libido
Prefreezing motility
Volume
Total no. of sperm cells per ejaculate × 10 ~
No. of semen doses per collection
Postfreezing motility
E N F E N F E N F E N F F
ns ns ns *** *** *** *** ** ns *** ** *** *
*** ns ns *** *** *** *** ** *** *** *** *** ns
ns ns ** *** *** *** ** ** *** *** *** *** ns
* ns ns *** *** *** *** *** *** *** *** *** *
** ns * *** *** *** *** *** *** *** ** *** ns
** ** ns *** *** *** *** *** ns *** ** ns ns
E N F
33.1 16.8 14.0
17.6 21.6 10.2
30.3 42.3 38.2
41.5 36.7 30.8
38.0 36.3 33.1
14.1 16.0 20.1
ns = not significant; *P < 0.05; **P < 0.01; ***P < 0.001. aR 2 is c a l c u l a t e d a s SSmode~.
SStotal
closely dependent upon the sexual preparation of the bull before collection. Large differences in no. of sperm cells per ejaculate may well be attributable to differences in collection frequency (Everett and Bean, 1982).
Analyses of variance Results from the analyses of variance are shown in Table 6. The bull effect was highly significant ( P < 0.001 ) for all traits at all stations. In E, sire of bull was significant for all traits except libido and volume. At the other stations, a significant effect of the bulls's sire was found for volume and no. of semen doses in F and for post-freezing motility in N. Bull's age at collection was significant for all traits, except for post-freezing motility in F. Less than 5% of the variation of libido was due to bull's age at collection. The frequency distribution of libido showed that more than 90% of the ejaculates were scored a minimum of 3. Even so it was found important to
evaluate the variation in libido in this study, since it is decisive for a successful semen collection. Season of collection was significant for most of the characteristics. Between 10.2 and 42.3% of the variation in the semen characteristics was explained by causes considered in the models used. W i t h few exceptions there were only minor differences in R2-values for the same traits between A.I. stations. The highest coefficients of determination were obtained for ejaculate volume, total no. of sperm cells per ejaculate, and no. of semen doses per collection.
Effect of age Bull's age at first collection differed slightly between A.I. stations. Usually test collections started at an age of 12-13 months (Fig. 1 ). The distribution of age at first collection reflects the variation in sexual maturity and managemental factors. At N, bulls were slightly older at the start, because one group of bulls were raised on a performance testing station in another area. Age of bull at collection affected ejaculate volume, total no. of sperm cells and no. of semen doses per collection. For the quantitative traits, 5-15% of the variation was due to bull's age at collection. Figure 2 shows the increase in total no. of sperm cells per collection day. This parameter included both single and double ejaculates (i.e., the total output of sperm cells per collection da5). Total no. of sperm cells collected per day doubled from 12 months to 30 months of age and older. These results were consistent with those of Hultn~s (1959) and Everett and Bean (1982). The effect of bull's age at collection on the two motility characteristics was significant except for post-freezing motility in F (Fig. 3 ). The age effect accounted for < 5% of the variation in motility. The increase in ejaculate volume with increasing age varied in size between bull stations. It
Percentage
lOOJ 904 i 80~
[ ] SRB, Elitsernin • SRB, Nordavel [ ] SLB, L~g[andsavel, Fatkenberg
7oJ
JI !
11.6 12.0 12./* 12.8 1t2 13.6 l&0 1/,./, 1/,.8 15.2 15.6 15.0 l&t, 16.8 Age at 1sf collection, months
Fig. 1. Percentage distribution of age of bull at 1st collection. No. of 1st ejaculates were: 4140 in E
with 2=12.3 and SD=0.3; 2321 in N with 2=14.1 and SD=0.8; 4662 in F with 2=12.5 and SD = 0.3, respectively.
10 11000 ~.I 0000 o
9 000 8000
u 7000 E 6 000 tn
5000
~ooo
o--o
SR8 (E) SRB (N) [ 3 - - [ 3 SLB(F)
~ooo 2000
~ 1 000 0
J.i
.
J.L
. i . i . l . i
. i . t . l .
i,
J.i
, I . ,
,i
. ~ i l
12 13 I&15-16 17-18 19-20 21-22 23-2& 25-26 27-28 29 30:-30 Age,monfhs
Fig. 2. Effect of age or bull at collection on total no. of sperm cells- 106/day.
77 76
7& 73 72 71 70-
~
69 6g
67 66 ~ -
)
O SRB (El ~. SRB IN) {3 SLB IF)
6s
ss ~c
v, 54
52 51 50 49 ~8
&7
•
SRB (El
•
SLB
• SR8(N)
z.6 ~]
.l.
[
. I . I . l .
[
.I.
I
.l
. ~ , I
, i , I .
12 13 14 15-16 17-18 19-20 21-22 23-2& 25-26 Age,monfhs
I
(F)
J i l l
.101
27-28 29-30 >30
Fig. 3. Effect of age of bull at collection on motility before and after freezing.
11 is important to notice that not all the bulls in this study were collected in all age classes. The effect of age at collection was considerable in comparison with the results of Chandler et al. (1985). Age effects on semen-producing capacity are difficult to separate from effects of body size changes in growing bulls. As maturity is reached, other factors such as stress or infections are likely to be of greater importance than age (Salisbury et al., 1978). This causes the nonlinear relationship between semen production and age to disappear.
Effect of season and year There was a significant seasonal effect on the semen characteristics. Semen characteristics seemed better when obtained in June, July and August than during the remaining months of the year. The differences between seasons were greater in the northernmost station. This could be an effect of more appreciable climatic differences in the north of Sweden due mainly to seasonal differences in day length. Station F is situated just south of the 57th latitude and the northernmost station, N, is on the 63rd latitude. Hultniis (1959) divided the year into three seasons, and season was not found significant for any of the analysed traits. Chandler et al. (1985), who assigned two seasons, found that season accounted for 1.9% of the total variation in post-freezing motility and that summer collections had better motility than winter ones. Amann et al. (1966) found the highest percentage of ejaculates with the largest semen volume from July to September. The superiority of the summer months was also in agreement with Taylor et al. (1985). Lodge and Salisbury (1970) concluded that seasonal effects on the male play a contributory role in the variability found in fertility. The effect of year on ~emen characteristics was evaluated in F. The material consisted of 3 years, 1983-1985. Between these years, no significant difference was found for any semen trait, except total no. of sperm cells per ejaculate ( P < 0.05). However, collections were not made from the same bulls in all 3 years. Hultniis (1959) reported differences in years and possibly between short periods in the same feed-year for initial motility and total no. of sperm cells per ejaculate. No significant annual variations in ejaculate volume were found by Hultn~is (1959).
Heritability estimates Heritabilities (h 2) and repeatabilities (t) with standard errors estimated for different semen characteristics are shown in Tables 7 and 8. Heritability and repeatability estimates for volume in N were not published because of negative values of the sire variance components. The zero variance was possibly due to a small no. of sires and a great influence of random error factors. In general, the estimated h2-values were low to medium. Due to the limited sets of data,
12 TABLE 7 Estimated heritabilities with standard errors for individual records (h 2) and for 20 observations per bull (h ~0) on libido and some semen characteristics Breed
Pre-freezing
h~
No. of Libido 1st ejaculates SRB (E) h 2
4140
SRB (N) h 2
2231
h~o SLB (F) h 2
4662
hgo
Post-freezing Motility
Volume
0.08_+0.04 0.18_+0.08 0.13 0.32 0.05__+0.03 0.08_+0.04 0.09 0.14 0.03_+0.02 0.02±0.01 0.05 0.04
Total no. of sperm cells per ejaculate × l06
0.02±0.01 0.17_+0.07 0.03 0.29 -0.04±0.03 0.07 0.28±0.09 0.06-+0.03 0.46 0.10
No. of 1st ejaculates
No. of semen dosesper collection
Motility
3870
0.26±0.10 0.43 0.07±0.04 0.12 0.12±0.05 0.20
0.12_+0.06 0.22 0.18_+0.07 0.31 0.06±0.03 0.10
2177 4176
TABLE 8 E s t i m a t e d r e p e a t a b i l i t i e s w i t h i n bull for libido a n d s o m e s e m e n c h a r a c t e r i s t i c s Breed
SRB (E) SRB (N) SLB (F)
Pre-freezing
Post-freezing
No. of 1st ejaculates
Libido
Motility
Volume
T o t a l no. of s p e r m cells p e r ejaculate × 106
No. of 1st ejaculates
No. of s e m e n doses p e r collection
Motility
4140 2231 4662
0.59 0.53 0.53
0.54 0.55 0.52
0.57 -0.59
0.57 0.56 0.56
3870 2177 4176
0.58 0.56 0.57
0.53 0.55 0.55
S t a n d a r d errors o f e s t i m a t e d r e p e a t a b i l i t i e s r a n g e d b e t w e e n 0.005 a n d 0.011.
the s.e. was relatively wide. The estimates of h 2 ranged from 0.02 to 0.28. The repeatabilities were very similar for all traits as well as between bull stations. The estimated values were between 0.52 and 0.59, i.e. much higher than those reported by Taylor et al. (1985) (t: 0.20-0.49) and Chandler et al. (1985) (t: 0.31-0.44) on similar traits. The rather low heritability indicated a comparatively large influence of environmental effects being associated with the individual bull. Therefore the heritability was also estimated based on 20 ejaculates (h2,n= 20). The estimates were then nearly doubled for all traits (Table 7). The h~-estimates for post-freezing motility in E and N were within the estimate given for percentage progressive motility immediately after thawing, in the study by Chandler et al. (1985).
Phenotypic correlations The phenotypic correlations between the semen characteristics are shown in Table 9. The materials used in these calculations consisted of collection
13 TABLE9 Phenotypic correlations between libido and some semen characteristicsa Bullstation
Characteristic
(E) (N)
1. Libido 2. Pre-freezing
motility 3. Volume 4. Total no. of sperm cells per ejaculate 5. No. of semen doses per collection 6. Post-freezing motility
1
2 0.03 0.01
-0.3
3
4
5
6
-0.04 0.06 -0.08 -0.02
-0.01 0.01 -0.05 0.36 0.74 0.63
0.01 0.02 -0.04 0.33 0.70 0.58 0.96 0.96
-0.02 0.01 0.13 0.08 -0.05 0.01 - 0.07 -0.07 -0.07 -0.07
0.07
-0.04
0.07
- 0.02
0.77
0.04
-0.03
0.51
0.75
-0.08
-0.23
-0.04
0.15
-0.21
~SRB (E) 3870 ejaculates and SRB (N) 2177 ejaculates above the diagonal; SLB 4176 ejaculates. records where post-freezing motility was recorded, i.e. ejaculates which were not discarded before freezing. Calculation of correlations in materials like these will be biased downwards due to t he continuous selection of ejaculates, but it is not possible to use unselected semen in the normal A.I. routine. T h e p h en o ty p i c correlations were generally low or close to zero and not always consistent between one breed and another. Ejaculate volume, total no. of sperm cells per ejaculate, and no. of semen doses per collection, being the exceptions since these correlations were pa r t l y due to autocorrelations. T h e low correlation between the two motility characteristics seems to indicate t h a t they are p ar tly two separate traits. Noticeable was t h a t motility was limited in variation, due to discarding ejaculates with low pre-freezing motility. E v e r e t t et al. (1978) r e por t ed t h a t the pr o port i on of motile sperm cells appeared to increase as the c o n c e n t r a t i o n of sperm cells increased (r=0.107). T h is indicated a par t l y over e s t i m a t e d motility in more c o n c e n t r a t e d samples. In a large material investigated by T a y l o r et al. (1985), genetic correlations between ejaculate volume and total no. of sperm cells per ejaculate were rep o r ted to be 0.51.
Discarded semen and culling routines A bull can be culled either during the p e r f o r m a n c e test or during the semen collection period. Culling due to fertility problems among bulls which had passed the p e r f o r m a n c e test was r e por t ed by Bernes (1981) and S t ~ l h a m m a r (1984) to be about 30%. T h r o u g h o u t the period of semen collection the bulls were continuously monitored. P o o r quality ejaculates can be discarded either before
14
or after freezing. Table 10 shows the discarded semen collections for the different bull stations. F had an almost twice as high a discarding rate before freezing (10.4%) compared with the other bull stations. Regarding the discarding frequency after the freeze-thaw procedure it can be seen that E and N compensated by discarding more collections than F at this stage. Total discarding before and after freezing was however almost identical in the three bull studs. According to the annual report of N for 1983-1984, the frequency of discarded ejaculates after freezing was 4.4%. During the time period under investigation there were disturbances at this bull station due to possible virus infections, which resulted in lowered freezability and consequently higher discarding rate (8.1%). The most common culling/discarding remarks for all A.I. stations in the present study were poor foreleg grasp by the bull and free heads of the spermatozoa. In an investigation made by Jansen ( 1984 ) into efficiency in semen production at ten A.I. centres in The Netherlands, it was shown that the frequency of ejaculates discarded on the basis of semen characteristics varied between 2.4 and 26.8% per year. The author reported several possible reasons, the most important affecting semen production being the preparation of the bull prior to collection. Jansen (1984) also found that the main reasons for differences in efficiency in semen production between A.I. centres were the total no. of sperm cells produced per ejaculate and the proportion of discarded ejaculates. An important question arises when differences in efficiency between A.I. stations are discussed i.e. if discarding ejaculates within bull is more efficient than culling bulls with the lowest fertility. According to Oltenacu et al. (1980), gains in fertility can be achieved by discarding the less fertile ejaculates within bulls on the basis of available semen quality tests, or by discarding all semen from bulls with a low ( <75% 59-day) NR rate (i.e. culling bulls), or a combination of these two programs. Culling of bulls requires a less rigorous culling of semen and is therefore less expensive. According to Oltenacu et al. (1980), discarding of ejaculates will achieve little improvement in fertility unless reliable predictors of ejaculate fertility are developed. Goffaux (1984) concluded T A B L E 10 Discarded semen collections Bull station
Elitsemin Nordavel L~glandsavel, Falkenberg
Total no. of collections 4140 2321 4662
Discarded pre-freezing
Discarded post-freezing
n
To
n
%
250 141 484
6.0 6.1 10.4
449 188 341
10.8 8.1 7.3
Total discarded, %
16.8 14.2 17.7
15 that a significant improvement in fertility by selection can only be achieved by sacrificing a great number of ejaculates or bulls. One breeding consequence of culling low fertility bulls would be to raise the NR level in the population in the longer term. The discarding of ejaculates maintains a high NR rate in the short term.
Concluding remark There is generally a genetic variation in the semen characteristics studied and the repeatability between ejaculates is considerable. Heritability estimates on repeated evaluations of the semen are high enough to select on the individual bull level. Temporary environmental effects, illness and other disturbances will affect the phenotypic variation of semen characteristics. Therefore there is a certain risk in selecting individual young bulls, for example on 20 ejaculates during one season. To spread the risk one might have to prolong the test period or repeat the test. Evaluation of bulls on the basis of 20 ejaculates ought to be practicable and would increase the heritability considerably, giving an accuracy (rw!) in selection of 0.3-0.5 for post-freezing motility and total no. of sperm cells per ejaculate. In an evaluation of A.I. bulls for semen characteristics, age of bull and season of collection will have to be considered. The ultimate value of selection based on semen characteristics applied at either ejaculate or bull level could not be decided solely on the basis of the results presented here. Semen characteristics can only be used to predict the fertility, when the correlation between a semen characteristic (or a combination of traits) and the true fertility is assessed. Until this is done it is difficult to choose between a system where ejaculates within bulls are discarded or a system where bulls with low values for semen quality and quantity or fertility are culled. However, the study does show that there are possibilities for genetic selection based on semen characteristics. But the effects on true fertility would still need to be investigated. ACKNOWLEDGEMENTS The authors wish to thank chief veterinarians Lars Drejare, Nordavel, GSran Malmberg, Elitsemin and Kjell Wass, Lfiglandsavel and Professor Kjell Larsson, Dept. of Obstetrics and Gynaecology, Swedish University of Agricultural Sciences, Uppsala for valuable information and advice. The study was supported financially by the Farmers Research Council for Information and Development, which is gratefully acknowledged.
16 REFERENCES Almquist, J.O., 1982. Effect of long term ejaculation at high frequency on output of sperm, sexual behaviour, and fertility of Holstein bulls; relation of reproductive capacity to high nutrient allowance. J. Dairy Sci., 65: 814-823. Amann, R., 1981. A critical review of methods for evaluation of spermatogenesis from seminal characteristics. J. Androl., 2: 37-58. Amann, R.P., Almquist, J.O. and Lambiase, J.T., 1966. Seasonal cycles in bull semen characteristics. J. Anim. Sci., 25: 916. Bane, A., 1954. Studies on monozygous cattle twins. XV. Sexual functions of bulls in relation to heredity, rearing intensity and somatic conditions. Acta Agric. Scand., 2: 95-208. Bernes, G., 1981. Utslagsorsaker och fertilitet hos ungtjurar i semin. Swedish University of Agricultural Sciences, Department of Animal Breeding and Genetics, Box 7023, 750 07 Uppsala, examensarbete nr. 95. Bishop, M.W.H., 1970. Ageing and reproduction in the male. J. Reprod. Fertil. Suppl., 12: 65-87. Chandler, J.E., Adkinson, R.W., Hay, G.M. and Crain, R.L., 1985. Environmental and genetic sources of variation for seminal quality in mature Holstein bulls. J. Dairy Sci., 68: 1270-1279. Everett, R.W. and Bean, B., 1982. Environmental influences on semen output. J. Dairy Sci., 65: 1303-1310. Everett, R.W., Bean, B. and Foote, R.H., 1978. Sources of variation of semen output. J. Dairy Sci., 61: 90-95. Goffaux, M., 1984. Cost of further selecting bulls or ejaculates in order to improve the mean N.R. rate of the A.I. station. Paper presented at the 10th Int. Congr. on Anim. Reprod. and A.I. University of Illinois at Urbana-Champaign, IL, 3 pp. Hafs, H.D., Bratton, R.W., Henderson, C.R. and Foote, R.H., 1958. Estimation of some variance components of bovine semen criteria and their use in the design of experiments. J. Dairy Sci., 41: 96-104. Harvey, W.R., 1977. User's guide for LSML 76. Mixed model least-squares and maximum likelihood computer program. Ohio State University, OH, 76 pp. Hultn~is, C.A., 1959. Studies on variation in mating behaviour and semen picture in young bulls of the Swedish red-and-white breed and on causes of this variation. Acta Agric. Scand. Suppl. 6, 82 pp. Jakubiec, J., 1983. Testicular size and consistency in bulls as a criterion of evaluation and selection with regard to fertility in cattle. Paper presented at the 34th E.A.A.P. session of the Commission on Cattle Production, Madrid, 4 pp. Jansen, H.B., 1984. The efficiency of bull semen production on 10 A.I. centres in The Netherlands. In: M. Courot (Editor), The Male in Farm Animal Reproduction. Martinus Nijhoff, Boston, MA, pp. 264-268. Lasley, J.F., 1951. Spermatozoan motility as a measure of semen quality. J. Animal Sci., 10: 211218. Linford, E., Glover, F.A., Bishop, C. and Stewart, D.L., 1976. The relationship between semen evaluation methods and fertility in the bull. J. Reprod. Fertil., 47: 283-291. Lodge, J.R. and Salisbury, G.W., 1970. Seasonal variation and male reproductive efficiency. In: A.D. Johnson, W.R. Gomes and M.L. Vandemark (Editors), The Testis. Academic Press, New York, NY, pp. 139-167. Maijala, K., 1969. Ober die Erblichkeit von Spermaeigenschaften yon KB-Bullen. Der Tierztichter, 3: 62-65. Oltenacu, E.A.B., Foote, R.H. and Bean, B., 1980. Effects of two simulated semen cullingprograms on predicted fertility in an artificially inseminated cow population. J. Dairy Sci., 63: 13511355.
17 Salisbury, G.W., Van Demark, N.L. and Lodge, J.R., 1978. Physiology of Reproduction and Artificial Insemination of Cattle. W.H. Freeman, San Francisco, CA, 574 pp. SAS, 1982. SAS User's Guide. Statistics, Statistical Analysis Systems Institute, Cary, NC, 584 pp. Seidel, G.E. and Foote, R.H., 1972. Variance components of semen criteria from bulls ejaculated frequently and their use in experimental design. J. Dairy Sci., 56: 399-405. St~lhammar, E-M., 1984. En uppf~ljning av utslagsorsaker och fertilitet hos ungtjurar i semin. Swedish University of Agricultural Sciences, Department of Animal Breeding and Genetics, Box 7023, 750 07 Uppsala, mimeo, 8 pp. Taylor, J.F. and Everett, R.W., 1985. Estimation of variance components by the expectationmaximization algorithm for restricted maximum likelihood in a repeatability model for semen production. J. Dairy Sci., 68: 2948-2953. Taylor, J.F., Bean, B., Marshall, C.E. and Sullivan, J.J., 1985. Genetic and environmental components of semen production traits of artificial insemination of Holstein bulls. J. Dairy Sci., 68: 2703-2722. Uwland, J., 1984. Possibilities and limitations of semen evaluation from the prognosis of male fertility. In: A. Courot (Editor), The Male in Farm Animal Reproduction. Martinus Nijhoff, Boston, MA, pp. 269-289. Wood, P.D.P., Foulkes, J.A., Shaw, R.C. and Melrose, D.R., 1986. Semen assessment, fertility and the selection of Hereford bulls for use in A.I.J. Reprod. Fertil., 76: 783-795.
1
Elsevier Science Publishers B.V., Amsterdam - - Printed in The Netherlands
Genetic Studies on Fertility in A.I. Bulls. I. Age, S e a s o n and Genetic Effects on S e m e n Characteristics in Y o u n g Bulls E V A - M A R I E S T A L H A M M A R , L E N N A R T J A N S O N and J A N P H I L I P S S O N
Department of Animal Breeding and Genetics, Swedish University of Agricultural Sciences, Box 7023, 750 07 Uppsala (Sweden) (Accepted 17 October 1988)
ABSTRACT St~lhammar, E.-M., Janson, L. and Philipsson, J., 1989. Genetic studies on fertility in A.I. bulls. I. Age, season and genetic effects on semen characteristics in young bulls. Anim. Reprod. Sci, 19: 1-17. Genetic and environmental factors that influence semen quantity and quality were evaluated for 215 young bulls of the two main dual-purpose breeds in Sweden, Swedish Red and White (SRB) and Swedish Friesian breed (SLB). Semen was collected at three bull stations from September 1983 to August 1985, inclusive. The effects of bull's sire, bull, season and year of collection, and bull's age at collection were estimated for ejaculate volume, motility, total no. of sperm cells per ejaculate and no. of semen doses per collection. Heritabilities (h2), repeatabilities (t) and phenotypic correlations were estimated for these traits. Motility and ejaculate volume were significantly affected by bull's sire (P < 0.01 ). The effect of bull, season of collection, and bull's age at collection were highly significant (P < 0.001 ) for most of the traits. The effect of year of collection was only significant for one semen characteristic. The summer months June, July and August were the best months for quantitative traits. Estimates of the heritability for semen characteristics ranged between 0.02 and 0.28, with the highest estimates found for ejaculate volume and semen doses per collection. The h2-estimates differed somewhat between breeds. The repeatabilities ranged between 0.5 and 0.6 for all the traits studied. It was concluded that the h2-estimates were sufficiently high to permit selection on individual bull level for some of the traits. At least 20 ejaculates have to be considered for total no. of sperm cells per ejaculate and post-freezing motility in order to achieve a selection accuracy (rTi) of 0.3-0.5. One must take into account a bull's age and season of collection when evaluating semen quality.
INTRODUCTION
The fertility of bulls is an important consideration, since each bull can be used for a large number of inseminations. In Sweden, it has been the practice hitherto to slaughter bulls once 30 000-45 000 semen doses have been col-
0378-4320/89/$03.50
© 1989 Elsevier Science Publishers B.V.
2 TABLE 1 Review of some parameters, means (x), standard deviations (SD), heritabilities (h 2) of some semen characteristics in the literature Characteristic
2
SD
h2
Pre-freezing motility, %
66 57 59.7 58.6 8.0 2.81
2.4
Hafs et al., 1958 Seidel and Foote, 1972 Everett et al., 1978 Everett and Bean, 1982 Hafs et al., 1958 Hultn~is, 1959 0.10-0.21 Maijala, 1969 Seidel and Foote, 1972 Everett et al., 1978 Everett and Bean, 1982 Jakubiec, 1983
Volume, ml
5.9 7.8 5.80 3.8 7.9 6.3 5.0 Total no. of sperm cells)< 109 per ejaculate
13.6 7.23 8.67 10.3 7.81 3.064 5.8-8.6 10.9 7.4 6.9
Post-freezing motility, 64.1 % 53 60.91 33.06
3.81 9.0 1.52
1.77 0.3 3.1 3.0 2.3
0.16
Taylor et al., 1985
0.12
Taylor and Everett, 1985 Hafs et al., 1958 Hultn~is, 1959 Seidel and Foote, 1972 Everett et al., 1978 Everett and Bean, 1982 Jakubiec, 1983 Jansen, 1984
0.05
Taylor et al., 1985
0.02
Taylor and Everett, 1985 Lasley, 1951 Seidel and Foote, 1972 Uwland, 1984 Chandler et al., 1985
6.0 3.18 3.31 0.829 5.7 5.0 4.5
6.24 7.99
Reference
}
0.21
lected, except at one A.I. station. Now, however, with the i n t r o d u c t i o n of a new system, in w h i c h the bulls are k e p t alive d u r i n g the process of p r o g e n y testing, the A.I. s t a t i o n s will be able to m a k e m o r e intensive use of genetically superior bulls. It is of b o t h genetic a n d e c o n o m i c i n t e r e s t to a s c e r t a i n w h i c h factors influence the q u a n t i t y a n d quality of semen. T h e possible r e l a t i o n s h i p b e t w e e n various s e m e n c h a r a c t e r i s t i c s a n d fertility is of p a r t i c u l a r i m p o r t a n c e . S u c h a r e l a t i o n s h i p has been a s s e r t e d by L i n f o r d et al. (1976) a n d W o o d et al. (1986) a n d will be i n v e s t i g a t e d u s i n g Swedish d a t a in a f o r t h c o m i n g study. T h e r e are i n t e r e s t i n g older studies on s e m e n quality f r o m S w e d e n a n d several o t h e r c o u n t r i e s (Table 1). B a n e (1954) studied m a t i n g b e h a v i o u r a n d
semen characteristics in monozygous bull twins and found differences between the genotypes. Marked similarities between twin brothers and wide disparities between pairs showed that heredity exerted considerable influence on mating ability and libido. There are also appreciable variations in sex drive between bulls kept under similar conditions (Bishop, 1970). Hultn~is (1959) showed that Swedish Red and White bulls in the age interval 15-26 months increased ejaculate volume on average 3% for each month of life. The practices of discarding ejaculates for the sake of maintaining a high semen quality and the culling of bulls with the purpose of achieving a good conception result have been shown to differ considerably between Swedish bull stations (Bernes, 1981 ). The ultimate effects of different culling policies are not well known. The study by Bernes (1981) showed breed differences in bull fertility. Age of bull, season, and the frequency of ejaculation are known to influence semen output (Everett et al., 1978; Almquist, 1982; Everett and Bean, 1982 ). The purpose of the present study was to determine the effects of different environmental factors such as season, year and age of bull on libido and various semen characteristics under routine Swedish A.I. conditions. The intention was also to study genetic variability in these traits and their mutual interrelationships. MATERIALSAND METHODS
Materials Data on semen characteristics from 215 young bulls of the two main dualpurpose breeds in Sweden, Swedish Red and White (SRB) and Swedish Friesian Breed (SLB) were obtained from three A.I. stations. The data were recorded in the normal semen collection routine between September 1983 and August 1985, inclusive. The bull stations were Elitsemin (E), Nordavel {N) (SRB-bulls) and L~glandavel, Falkenberg (F) (SLB-bulls). The bulls were sons of 40 sires with 2 to 14 sons per sire (Table 2 ). The first collection started from 11 to 17 months of age. The oldest bulls in this study were about 3 years TABLE 2 Numberof bulls and sires in the materialsanalysed Bull station
Breed
No. of bulls
No. of sires
Elitsemin Nordavdel L~glandsavel, Falkenberg
SRB SRB SLB
62 70 83
11 14 15
4 TABLE 3 Number of semen collections per season in the materials analysed Bull station
Season
Elitsemin Nordavel L~glandsavel, Falkenberg
Winter
Spring
Summer
Autumn
Total number of collections
1174 685 1089
748 868 1321
790 265 1280
1428 503 972
4140 2321 4662
TABLE 4 Number of collections in the different materials analysed Bull station
Original Collections no. of without collections recorded volume
Elitsemin 4705 Nordavel 2904 L~glandsavel, 8603 Falkenberg
Collections with two ejaculates
No. of collections not included for various reasons
n
%
n
%
n
%
140 30 469
3.0 1.0 5.4
72 253 2571
1.5 8.7 29.9
353 300 901
7.5 10.3 10.5
Maximum no. of collections in the analysis
No. of collections per bull on average
4140 2321 4662
67 33 56
old. In E the majority of semen collections were made in a u t u m n and winter, while in N most were during winter and spring. The collections in F were more evenly spread over the year (Table 3). Table 4 shows the restrictions placed on the materials and the total number of collections eventually analysed. Information on each ejaculate in this study included (1) bull's identity; (2) collection code, including date; (3) ejaculate order number; (4) libido; (5) original volume of ejaculate; (6) total no. of sperm cells per ejaculate; (7) motility, both before and after freezing; (8) no. of semen doses per collection; and (9) any discarding, either before or after freezing. The bull's birth date and sire were added to this information. If a bull was the only son of a sire, it was deleted from the genetic analyses in order to reduce confounding with sire and bull within sire in the analyses. Collections without recorded volume were excluded. W h e n a bull ejaculated more t h a n once on the same collection day it was not possible to distinguish between the ejaculates after freezing, as the two ejaculates were pooled when frozen and were assigned the same collection code. Due to the possible effects of taking two ejaculates per collection day more frequently in F than at the other bull stations, the analyses of pre-freezing characteristics were based on only 1st ejaculates.
Collection routines Young bulls were semen collected once a week. Bulls older than 15-18 months i.e. bulls in production, were ejaculated twice a week. In F, a second ejaculate was taken if the preset limit of 450 semen doses from the first ejaculate was not reached. The bulls were stimulated prior to collection by having another bull waiting at the same time in the collection room. The bulls were permitted one false mount and then restrained before mounting and ejaculating on the teaser bull. F used the same stimulation procedure before taking the second ejaculate. In station F bulls were kept grazing during the summer and they were taken to the collection room in pairs, stimulating each other on the way. All the A.I. stations allowed the bulls with initial sexual disturbance or poor semen to have a trial and learning period of 2-3 months before they were culled.
Recording and evaluation procedures Libido was recorded on a scale from 1-5, with 1 = no sex drive, 3 = good sex drive, and 5 = abnormally vigorous sex drive. The fresh semen was evaluated immediately after each collection. This procedure included measuring the volume (ml) and determining motility subjectively under a phase-contrast microscope. Motility was evaluated as the percentage of forward-motile sperm cells. Spermatozoal concentration was determined spectrophotometrically. The turbidity was compared with a known standard. Total no. of sperm cells per ejaculate was calculated by multiplying the spermatozoal concentration per ml by the volume. From the total no. of sperm cells per ejaculate, the number of semen doses was calculated automatically. The stations had different set standards for the concentration, giving different numbers of sperm cells per semen dose (0.25 ml). The standards were 80 (E), 88 (N) and 70 (F)×106 sperm cells per ml respectively, giving 20 (E), 22 (N) and 17.5 (F) × 106 sperm cells per semen dose. The assessment of motility after freezing and thawing was normally made on 1-2 semen doses per collection. The observer did not know from which bull the sample being evaluated had been obtained. The established minimum standards for motility varied slightly between stations. Minimum pre-freezing motility was 60-70% forward-motile sperm cells and for post-freezing it was 4550%. The recording of the different characteristics, both subjectively and objectively was not always made by the same person.
Statistical methods Data were analysed by using Henderson's method III for estimation of variance components and several procedures within the Statistical Analysis System (SAS, 1982 ). The coefficient of determination (R 2) was calculated as the
sums of squares for the model, divided by the total sums of squares (SS model/ SS total). Adjustments for the degrees of freedom had no effect on the R 2values, due to the small size of the model and large number of observations. Since semen was not collected from each bull every month season rather than month of collection was assigned, each spanning 3 months, starting with winter in December. Age of bull at collection was divided into twelve classes, starting at an age of ~ 12 months and ending with bulls more than 30 months old. The age classes each covered 2 months, beginning from 15 months of age, while ~<12, 13 and 14 months of age were each considered as one class. Two-factor interactions between year, season, and bull's age at collection respectively were analysed, b u t were not found significant ( P > 0.05). These interaction effects were therefore excluded from the final model. The three A.I. stations were analysed separately. The model chosen as the most appropriate for estimation of variance components and correlations for libido and semen characteristics at bull station F was: = tt + si + bij + sek + at +yrm + eijh~mn
Yijklmn gijklmn
-
-
8i bo
sek al yrm
eijklrnn
-
-
observed semen characteristic least squares means effect of ith sire effect o f j t h bull within ith sire effect of kth season of collection (k-- 1...4) effect o f / t h age of bull at collection (/--1...12) effect of mth year of collection ( m = 1...3) random error element with expected mean value zero and variance 2 O" e
The model chosen for E and N, was the same as above, except for the year effect. The sire and bull components were considered random and all other elements in the models were assumed fixed. Preferably the model for SRB also should have included A.I. station, but the sires in common were too few. The above described model was chosen, because of the structure of the materials, with sires neither strictly nested within station nor strictly cross-classified with station. The heritability (h 2) of the semen characteristics studied was estimated through the sire, a~, and the bull within sire, a~:s, component of variance, as h2=4o /(o + ab:s . 2 + de) 2 on individual ejaculates. The repeatability coefficient (t) of the semen characteristics in the full model was calculated within 2 2 2 bull as, t = as2 + ab:s/(as + a~:s + a e2) . To demonstrate the heritability of n ejaculates, an estimate ofh2n was calculated as h2n = n ' h 2 / 1 + ( n - 1 )t. The phenotypic correlations were calculated by using the paternal half-sib method according to Harvey (1977).
RESULTS AND DISCUSSION
The overall means and standard deviations for libido and the semen characteristics are given in Table 5. The overall means and especially the variation in most of the traits being studied varied considerably between the two SRBstations. The SD for total no. of sperm cells was much higher at N. One reason for this difference was the bull's age. This will be discussed later. Motility was judged subjectively and it is therefore uncertain whether differences in motility were real, or merely a reflection of differences in judgement between bull stations. Furthermore the SD both absolutely and relatively was greater for motility after freezing than before freezing. F differed from the other stations especially as regards no. of semen doses per collection. This may largely be due to the high proportion of double ejaculates taken and the lower standards of 17.5× 108 sperm cells per semen dose applied. Although only 1st ejaculates were analysed, a possible effect of a second ejaculate could be discerned in F. In general, the overall means and SD in this investigation were consistent with many of the results found in the literature (Table 1 ). Motility was expressed differently in the literature, both as progressive motile and as percentage living spermatozoa. The evaluation of post-freezing motility was done as a 'blind-test' at all stations, as recommended by Amann (1981), which would minimize the bias of the observer. Compared to the results of Hultn~is (1959) the semen volume of young SRB bulls has almost been doubled. Mean volume of first ejaculate was 2.8 ml versus 5.1 to 5.3 ml in the present study. Although the collection routines were different, these results could indicate a genetic improvement, since one selection criterion has been an increased minimum volume of semen. Both volume and total no. of sperm cells per ejaculate are TABLE5 Overall means (2) and standard deviations (SD) for the semen characteristics analysed Breed
SRB (E)
No. of 1st ejaculates
4140 3870 a SRB (N) 2321 2177 a SLB (F) 4662 4176 a
Pre-freezing motility, %
Volume, ml
Total no. of sperm cells per ejaculate, × 106
No. of semen doses per collection
£
SD
£
2
£
SD
75
4
5.2 1.8
6930 2478 340
114
52 8
69
7
5.5 2.2
6961 4828 328
213
48 7
69
5
5.1 1.8
6466 2911 500 b 155
53 6
SD
SD
aNo. of observations after freezing and thawing. bDouble ejaculates to a greater extent than in E and N.
Post-freezing motility, %
SD
TABLE
6
Significance levels for libido and semen characteristics from least-square analyses of variance and c o e f f i c i e n t s o f d e t e r m i n a t i o n ( R 2)a f o r t h e t h r e e b u l l s t a t i o n s Source of variation
Sire
Bull
Season of collection Age of bull at collection Year of collection R 2, %
A.I. stud
Significance levels Libido
Prefreezing motility
Volume
Total no. of sperm cells per ejaculate × 10 ~
No. of semen doses per collection
Postfreezing motility
E N F E N F E N F E N F F
ns ns ns *** *** *** *** ** ns *** ** *** *
*** ns ns *** *** *** *** ** *** *** *** *** ns
ns ns ** *** *** *** ** ** *** *** *** *** ns
* ns ns *** *** *** *** *** *** *** *** *** *
** ns * *** *** *** *** *** *** *** ** *** ns
** ** ns *** *** *** *** *** ns *** ** ns ns
E N F
33.1 16.8 14.0
17.6 21.6 10.2
30.3 42.3 38.2
41.5 36.7 30.8
38.0 36.3 33.1
14.1 16.0 20.1
ns = not significant; *P < 0.05; **P < 0.01; ***P < 0.001. aR 2 is c a l c u l a t e d a s SSmode~.
SStotal
closely dependent upon the sexual preparation of the bull before collection. Large differences in no. of sperm cells per ejaculate may well be attributable to differences in collection frequency (Everett and Bean, 1982).
Analyses of variance Results from the analyses of variance are shown in Table 6. The bull effect was highly significant ( P < 0.001 ) for all traits at all stations. In E, sire of bull was significant for all traits except libido and volume. At the other stations, a significant effect of the bulls's sire was found for volume and no. of semen doses in F and for post-freezing motility in N. Bull's age at collection was significant for all traits, except for post-freezing motility in F. Less than 5% of the variation of libido was due to bull's age at collection. The frequency distribution of libido showed that more than 90% of the ejaculates were scored a minimum of 3. Even so it was found important to
evaluate the variation in libido in this study, since it is decisive for a successful semen collection. Season of collection was significant for most of the characteristics. Between 10.2 and 42.3% of the variation in the semen characteristics was explained by causes considered in the models used. W i t h few exceptions there were only minor differences in R2-values for the same traits between A.I. stations. The highest coefficients of determination were obtained for ejaculate volume, total no. of sperm cells per ejaculate, and no. of semen doses per collection.
Effect of age Bull's age at first collection differed slightly between A.I. stations. Usually test collections started at an age of 12-13 months (Fig. 1 ). The distribution of age at first collection reflects the variation in sexual maturity and managemental factors. At N, bulls were slightly older at the start, because one group of bulls were raised on a performance testing station in another area. Age of bull at collection affected ejaculate volume, total no. of sperm cells and no. of semen doses per collection. For the quantitative traits, 5-15% of the variation was due to bull's age at collection. Figure 2 shows the increase in total no. of sperm cells per collection day. This parameter included both single and double ejaculates (i.e., the total output of sperm cells per collection da5). Total no. of sperm cells collected per day doubled from 12 months to 30 months of age and older. These results were consistent with those of Hultn~s (1959) and Everett and Bean (1982). The effect of bull's age at collection on the two motility characteristics was significant except for post-freezing motility in F (Fig. 3 ). The age effect accounted for < 5% of the variation in motility. The increase in ejaculate volume with increasing age varied in size between bull stations. It
Percentage
lOOJ 904 i 80~
[ ] SRB, Elitsernin • SRB, Nordavel [ ] SLB, L~g[andsavel, Fatkenberg
7oJ
JI !
11.6 12.0 12./* 12.8 1t2 13.6 l&0 1/,./, 1/,.8 15.2 15.6 15.0 l&t, 16.8 Age at 1sf collection, months
Fig. 1. Percentage distribution of age of bull at 1st collection. No. of 1st ejaculates were: 4140 in E
with 2=12.3 and SD=0.3; 2321 in N with 2=14.1 and SD=0.8; 4662 in F with 2=12.5 and SD = 0.3, respectively.
10 11000 ~.I 0000 o
9 000 8000
u 7000 E 6 000 tn
5000
~ooo
o--o
SR8 (E) SRB (N) [ 3 - - [ 3 SLB(F)
~ooo 2000
~ 1 000 0
J.i
.
J.L
. i . i . l . i
. i . t . l .
i,
J.i
, I . ,
,i
. ~ i l
12 13 I&15-16 17-18 19-20 21-22 23-2& 25-26 27-28 29 30:-30 Age,monfhs
Fig. 2. Effect of age or bull at collection on total no. of sperm cells- 106/day.
77 76
7& 73 72 71 70-
~
69 6g
67 66 ~ -
)
O SRB (El ~. SRB IN) {3 SLB IF)
6s
ss ~c
v, 54
52 51 50 49 ~8
&7
•
SRB (El
•
SLB
• SR8(N)
z.6 ~]
.l.
[
. I . I . l .
[
.I.
I
.l
. ~ , I
, i , I .
12 13 14 15-16 17-18 19-20 21-22 23-2& 25-26 Age,monfhs
I
(F)
J i l l
.101
27-28 29-30 >30
Fig. 3. Effect of age of bull at collection on motility before and after freezing.
11 is important to notice that not all the bulls in this study were collected in all age classes. The effect of age at collection was considerable in comparison with the results of Chandler et al. (1985). Age effects on semen-producing capacity are difficult to separate from effects of body size changes in growing bulls. As maturity is reached, other factors such as stress or infections are likely to be of greater importance than age (Salisbury et al., 1978). This causes the nonlinear relationship between semen production and age to disappear.
Effect of season and year There was a significant seasonal effect on the semen characteristics. Semen characteristics seemed better when obtained in June, July and August than during the remaining months of the year. The differences between seasons were greater in the northernmost station. This could be an effect of more appreciable climatic differences in the north of Sweden due mainly to seasonal differences in day length. Station F is situated just south of the 57th latitude and the northernmost station, N, is on the 63rd latitude. Hultniis (1959) divided the year into three seasons, and season was not found significant for any of the analysed traits. Chandler et al. (1985), who assigned two seasons, found that season accounted for 1.9% of the total variation in post-freezing motility and that summer collections had better motility than winter ones. Amann et al. (1966) found the highest percentage of ejaculates with the largest semen volume from July to September. The superiority of the summer months was also in agreement with Taylor et al. (1985). Lodge and Salisbury (1970) concluded that seasonal effects on the male play a contributory role in the variability found in fertility. The effect of year on ~emen characteristics was evaluated in F. The material consisted of 3 years, 1983-1985. Between these years, no significant difference was found for any semen trait, except total no. of sperm cells per ejaculate ( P < 0.05). However, collections were not made from the same bulls in all 3 years. Hultniis (1959) reported differences in years and possibly between short periods in the same feed-year for initial motility and total no. of sperm cells per ejaculate. No significant annual variations in ejaculate volume were found by Hultn~is (1959).
Heritability estimates Heritabilities (h 2) and repeatabilities (t) with standard errors estimated for different semen characteristics are shown in Tables 7 and 8. Heritability and repeatability estimates for volume in N were not published because of negative values of the sire variance components. The zero variance was possibly due to a small no. of sires and a great influence of random error factors. In general, the estimated h2-values were low to medium. Due to the limited sets of data,
12 TABLE 7 Estimated heritabilities with standard errors for individual records (h 2) and for 20 observations per bull (h ~0) on libido and some semen characteristics Breed
Pre-freezing
h~
No. of Libido 1st ejaculates SRB (E) h 2
4140
SRB (N) h 2
2231
h~o SLB (F) h 2
4662
hgo
Post-freezing Motility
Volume
0.08_+0.04 0.18_+0.08 0.13 0.32 0.05__+0.03 0.08_+0.04 0.09 0.14 0.03_+0.02 0.02±0.01 0.05 0.04
Total no. of sperm cells per ejaculate × l06
0.02±0.01 0.17_+0.07 0.03 0.29 -0.04±0.03 0.07 0.28±0.09 0.06-+0.03 0.46 0.10
No. of 1st ejaculates
No. of semen dosesper collection
Motility
3870
0.26±0.10 0.43 0.07±0.04 0.12 0.12±0.05 0.20
0.12_+0.06 0.22 0.18_+0.07 0.31 0.06±0.03 0.10
2177 4176
TABLE 8 E s t i m a t e d r e p e a t a b i l i t i e s w i t h i n bull for libido a n d s o m e s e m e n c h a r a c t e r i s t i c s Breed
SRB (E) SRB (N) SLB (F)
Pre-freezing
Post-freezing
No. of 1st ejaculates
Libido
Motility
Volume
T o t a l no. of s p e r m cells p e r ejaculate × 106
No. of 1st ejaculates
No. of s e m e n doses p e r collection
Motility
4140 2231 4662
0.59 0.53 0.53
0.54 0.55 0.52
0.57 -0.59
0.57 0.56 0.56
3870 2177 4176
0.58 0.56 0.57
0.53 0.55 0.55
S t a n d a r d errors o f e s t i m a t e d r e p e a t a b i l i t i e s r a n g e d b e t w e e n 0.005 a n d 0.011.
the s.e. was relatively wide. The estimates of h 2 ranged from 0.02 to 0.28. The repeatabilities were very similar for all traits as well as between bull stations. The estimated values were between 0.52 and 0.59, i.e. much higher than those reported by Taylor et al. (1985) (t: 0.20-0.49) and Chandler et al. (1985) (t: 0.31-0.44) on similar traits. The rather low heritability indicated a comparatively large influence of environmental effects being associated with the individual bull. Therefore the heritability was also estimated based on 20 ejaculates (h2,n= 20). The estimates were then nearly doubled for all traits (Table 7). The h~-estimates for post-freezing motility in E and N were within the estimate given for percentage progressive motility immediately after thawing, in the study by Chandler et al. (1985).
Phenotypic correlations The phenotypic correlations between the semen characteristics are shown in Table 9. The materials used in these calculations consisted of collection
13 TABLE9 Phenotypic correlations between libido and some semen characteristicsa Bullstation
Characteristic
(E) (N)
1. Libido 2. Pre-freezing
motility 3. Volume 4. Total no. of sperm cells per ejaculate 5. No. of semen doses per collection 6. Post-freezing motility
1
2 0.03 0.01
-0.3
3
4
5
6
-0.04 0.06 -0.08 -0.02
-0.01 0.01 -0.05 0.36 0.74 0.63
0.01 0.02 -0.04 0.33 0.70 0.58 0.96 0.96
-0.02 0.01 0.13 0.08 -0.05 0.01 - 0.07 -0.07 -0.07 -0.07
0.07
-0.04
0.07
- 0.02
0.77
0.04
-0.03
0.51
0.75
-0.08
-0.23
-0.04
0.15
-0.21
~SRB (E) 3870 ejaculates and SRB (N) 2177 ejaculates above the diagonal; SLB 4176 ejaculates. records where post-freezing motility was recorded, i.e. ejaculates which were not discarded before freezing. Calculation of correlations in materials like these will be biased downwards due to t he continuous selection of ejaculates, but it is not possible to use unselected semen in the normal A.I. routine. T h e p h en o ty p i c correlations were generally low or close to zero and not always consistent between one breed and another. Ejaculate volume, total no. of sperm cells per ejaculate, and no. of semen doses per collection, being the exceptions since these correlations were pa r t l y due to autocorrelations. T h e low correlation between the two motility characteristics seems to indicate t h a t they are p ar tly two separate traits. Noticeable was t h a t motility was limited in variation, due to discarding ejaculates with low pre-freezing motility. E v e r e t t et al. (1978) r e por t ed t h a t the pr o port i on of motile sperm cells appeared to increase as the c o n c e n t r a t i o n of sperm cells increased (r=0.107). T h is indicated a par t l y over e s t i m a t e d motility in more c o n c e n t r a t e d samples. In a large material investigated by T a y l o r et al. (1985), genetic correlations between ejaculate volume and total no. of sperm cells per ejaculate were rep o r ted to be 0.51.
Discarded semen and culling routines A bull can be culled either during the p e r f o r m a n c e test or during the semen collection period. Culling due to fertility problems among bulls which had passed the p e r f o r m a n c e test was r e por t ed by Bernes (1981) and S t ~ l h a m m a r (1984) to be about 30%. T h r o u g h o u t the period of semen collection the bulls were continuously monitored. P o o r quality ejaculates can be discarded either before
14
or after freezing. Table 10 shows the discarded semen collections for the different bull stations. F had an almost twice as high a discarding rate before freezing (10.4%) compared with the other bull stations. Regarding the discarding frequency after the freeze-thaw procedure it can be seen that E and N compensated by discarding more collections than F at this stage. Total discarding before and after freezing was however almost identical in the three bull studs. According to the annual report of N for 1983-1984, the frequency of discarded ejaculates after freezing was 4.4%. During the time period under investigation there were disturbances at this bull station due to possible virus infections, which resulted in lowered freezability and consequently higher discarding rate (8.1%). The most common culling/discarding remarks for all A.I. stations in the present study were poor foreleg grasp by the bull and free heads of the spermatozoa. In an investigation made by Jansen ( 1984 ) into efficiency in semen production at ten A.I. centres in The Netherlands, it was shown that the frequency of ejaculates discarded on the basis of semen characteristics varied between 2.4 and 26.8% per year. The author reported several possible reasons, the most important affecting semen production being the preparation of the bull prior to collection. Jansen (1984) also found that the main reasons for differences in efficiency in semen production between A.I. centres were the total no. of sperm cells produced per ejaculate and the proportion of discarded ejaculates. An important question arises when differences in efficiency between A.I. stations are discussed i.e. if discarding ejaculates within bull is more efficient than culling bulls with the lowest fertility. According to Oltenacu et al. (1980), gains in fertility can be achieved by discarding the less fertile ejaculates within bulls on the basis of available semen quality tests, or by discarding all semen from bulls with a low ( <75% 59-day) NR rate (i.e. culling bulls), or a combination of these two programs. Culling of bulls requires a less rigorous culling of semen and is therefore less expensive. According to Oltenacu et al. (1980), discarding of ejaculates will achieve little improvement in fertility unless reliable predictors of ejaculate fertility are developed. Goffaux (1984) concluded T A B L E 10 Discarded semen collections Bull station
Elitsemin Nordavel L~glandsavel, Falkenberg
Total no. of collections 4140 2321 4662
Discarded pre-freezing
Discarded post-freezing
n
To
n
%
250 141 484
6.0 6.1 10.4
449 188 341
10.8 8.1 7.3
Total discarded, %
16.8 14.2 17.7
15 that a significant improvement in fertility by selection can only be achieved by sacrificing a great number of ejaculates or bulls. One breeding consequence of culling low fertility bulls would be to raise the NR level in the population in the longer term. The discarding of ejaculates maintains a high NR rate in the short term.
Concluding remark There is generally a genetic variation in the semen characteristics studied and the repeatability between ejaculates is considerable. Heritability estimates on repeated evaluations of the semen are high enough to select on the individual bull level. Temporary environmental effects, illness and other disturbances will affect the phenotypic variation of semen characteristics. Therefore there is a certain risk in selecting individual young bulls, for example on 20 ejaculates during one season. To spread the risk one might have to prolong the test period or repeat the test. Evaluation of bulls on the basis of 20 ejaculates ought to be practicable and would increase the heritability considerably, giving an accuracy (rw!) in selection of 0.3-0.5 for post-freezing motility and total no. of sperm cells per ejaculate. In an evaluation of A.I. bulls for semen characteristics, age of bull and season of collection will have to be considered. The ultimate value of selection based on semen characteristics applied at either ejaculate or bull level could not be decided solely on the basis of the results presented here. Semen characteristics can only be used to predict the fertility, when the correlation between a semen characteristic (or a combination of traits) and the true fertility is assessed. Until this is done it is difficult to choose between a system where ejaculates within bulls are discarded or a system where bulls with low values for semen quality and quantity or fertility are culled. However, the study does show that there are possibilities for genetic selection based on semen characteristics. But the effects on true fertility would still need to be investigated. ACKNOWLEDGEMENTS The authors wish to thank chief veterinarians Lars Drejare, Nordavel, GSran Malmberg, Elitsemin and Kjell Wass, Lfiglandsavel and Professor Kjell Larsson, Dept. of Obstetrics and Gynaecology, Swedish University of Agricultural Sciences, Uppsala for valuable information and advice. The study was supported financially by the Farmers Research Council for Information and Development, which is gratefully acknowledged.
16 REFERENCES Almquist, J.O., 1982. Effect of long term ejaculation at high frequency on output of sperm, sexual behaviour, and fertility of Holstein bulls; relation of reproductive capacity to high nutrient allowance. J. Dairy Sci., 65: 814-823. Amann, R., 1981. A critical review of methods for evaluation of spermatogenesis from seminal characteristics. J. Androl., 2: 37-58. Amann, R.P., Almquist, J.O. and Lambiase, J.T., 1966. Seasonal cycles in bull semen characteristics. J. Anim. Sci., 25: 916. Bane, A., 1954. Studies on monozygous cattle twins. XV. Sexual functions of bulls in relation to heredity, rearing intensity and somatic conditions. Acta Agric. Scand., 2: 95-208. Bernes, G., 1981. Utslagsorsaker och fertilitet hos ungtjurar i semin. Swedish University of Agricultural Sciences, Department of Animal Breeding and Genetics, Box 7023, 750 07 Uppsala, examensarbete nr. 95. Bishop, M.W.H., 1970. Ageing and reproduction in the male. J. Reprod. Fertil. Suppl., 12: 65-87. Chandler, J.E., Adkinson, R.W., Hay, G.M. and Crain, R.L., 1985. Environmental and genetic sources of variation for seminal quality in mature Holstein bulls. J. Dairy Sci., 68: 1270-1279. Everett, R.W. and Bean, B., 1982. Environmental influences on semen output. J. Dairy Sci., 65: 1303-1310. Everett, R.W., Bean, B. and Foote, R.H., 1978. Sources of variation of semen output. J. Dairy Sci., 61: 90-95. Goffaux, M., 1984. Cost of further selecting bulls or ejaculates in order to improve the mean N.R. rate of the A.I. station. Paper presented at the 10th Int. Congr. on Anim. Reprod. and A.I. University of Illinois at Urbana-Champaign, IL, 3 pp. Hafs, H.D., Bratton, R.W., Henderson, C.R. and Foote, R.H., 1958. Estimation of some variance components of bovine semen criteria and their use in the design of experiments. J. Dairy Sci., 41: 96-104. Harvey, W.R., 1977. User's guide for LSML 76. Mixed model least-squares and maximum likelihood computer program. Ohio State University, OH, 76 pp. Hultn~is, C.A., 1959. Studies on variation in mating behaviour and semen picture in young bulls of the Swedish red-and-white breed and on causes of this variation. Acta Agric. Scand. Suppl. 6, 82 pp. Jakubiec, J., 1983. Testicular size and consistency in bulls as a criterion of evaluation and selection with regard to fertility in cattle. Paper presented at the 34th E.A.A.P. session of the Commission on Cattle Production, Madrid, 4 pp. Jansen, H.B., 1984. The efficiency of bull semen production on 10 A.I. centres in The Netherlands. In: M. Courot (Editor), The Male in Farm Animal Reproduction. Martinus Nijhoff, Boston, MA, pp. 264-268. Lasley, J.F., 1951. Spermatozoan motility as a measure of semen quality. J. Animal Sci., 10: 211218. Linford, E., Glover, F.A., Bishop, C. and Stewart, D.L., 1976. The relationship between semen evaluation methods and fertility in the bull. J. Reprod. Fertil., 47: 283-291. Lodge, J.R. and Salisbury, G.W., 1970. Seasonal variation and male reproductive efficiency. In: A.D. Johnson, W.R. Gomes and M.L. Vandemark (Editors), The Testis. Academic Press, New York, NY, pp. 139-167. Maijala, K., 1969. Ober die Erblichkeit von Spermaeigenschaften yon KB-Bullen. Der Tierztichter, 3: 62-65. Oltenacu, E.A.B., Foote, R.H. and Bean, B., 1980. Effects of two simulated semen cullingprograms on predicted fertility in an artificially inseminated cow population. J. Dairy Sci., 63: 13511355.
17 Salisbury, G.W., Van Demark, N.L. and Lodge, J.R., 1978. Physiology of Reproduction and Artificial Insemination of Cattle. W.H. Freeman, San Francisco, CA, 574 pp. SAS, 1982. SAS User's Guide. Statistics, Statistical Analysis Systems Institute, Cary, NC, 584 pp. Seidel, G.E. and Foote, R.H., 1972. Variance components of semen criteria from bulls ejaculated frequently and their use in experimental design. J. Dairy Sci., 56: 399-405. St~lhammar, E-M., 1984. En uppf~ljning av utslagsorsaker och fertilitet hos ungtjurar i semin. Swedish University of Agricultural Sciences, Department of Animal Breeding and Genetics, Box 7023, 750 07 Uppsala, mimeo, 8 pp. Taylor, J.F. and Everett, R.W., 1985. Estimation of variance components by the expectationmaximization algorithm for restricted maximum likelihood in a repeatability model for semen production. J. Dairy Sci., 68: 2948-2953. Taylor, J.F., Bean, B., Marshall, C.E. and Sullivan, J.J., 1985. Genetic and environmental components of semen production traits of artificial insemination of Holstein bulls. J. Dairy Sci., 68: 2703-2722. Uwland, J., 1984. Possibilities and limitations of semen evaluation from the prognosis of male fertility. In: A. Courot (Editor), The Male in Farm Animal Reproduction. Martinus Nijhoff, Boston, MA, pp. 269-289. Wood, P.D.P., Foulkes, J.A., Shaw, R.C. and Melrose, D.R., 1986. Semen assessment, fertility and the selection of Hereford bulls for use in A.I.J. Reprod. Fertil., 76: 783-795.