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Selective Breeding for High and Low Antibody Responses to Inactivated Newcastle Disease Virus in Japanese Quails1,2
BREEDING AND GENETICS Selective Breeding for High and Low Antibody Responses to Inactivated Newcastle Disease Virus in Japanese Quails1'2 S. TAKAHASHI, 3 S. INOOKA,4 and Y. MIZUMA Department of Animal Science, Faculty of Agriculture, Tohoku University, 1-1 AmamiyamachiTsutsumidori, Sendai, Japan (Received for publication July 16,1982)
1984 Poultry Science 63:595-599 INTRODUCTION
Recent selection research based on antibody producing ability (Biozzi, et al, 1980) demonstrated that this ability was controlled by multiple genes and could be correlated with other immune abilities such as antibody producing ability to unrelated antigens or resistance to several diseases. These facts suggest that genetic regulation systems for antibody production are involved with those of general antibody production or resistance. Further evidence on the relationships between antibody producing ability and natural resistance must be obtained to determine whether it is possible to select animals for high genetic resistance to pathogenic microbes without inducing prophylactic infections by such organisms. The purpose of this paper is to report the development of divergent lines of Japanese quail on the basis of their high and low serum antibody responses to inactivated Newcastle disease virus (NDV) antigen. This paper reports the selective process through the 9 generation during these selection experiments. Subsequent
'Supported in part by Grant-in-Aid, No 456194, from the Scientific Research Fund of the Ministry of Education, Science and Culture, Japan. 2 The quail were sent to the National Institute for Environmental Studies. At this Institute, selection has proceeded under H. Takahashi' direction. 3 The National Institute for Environmental Studies, Yatabemachi, Tsukuba, Ibaraki 300-21, Japan. 4 Reprint requests.
papers will consider the immunological traits of these lines. MATERIALS AND METHODS
Selection Procedures. The foundation stock consisted of 132 birds (T) maintained at the Laboratory of Animal Breeding, the University of Tohoku, and 139 birds (G) acquired from the University of Gifu. The T line had been selected by Phamm (1974) for high and low antibody response to NDV, and his selection experiment was abandoned after eight generations because of low fertility of the population. A base population was obtained by reciprocal matings of the T and G population. Sixty-three matings of T males and G females and 54 matings of T females and G males produced 234 offspring. High and low antibody response lines had been developed by mass selection of the highest responding birds at 7 weeks of age as parents of the high line and the lowest responding birds at the same age as parents of the low line. Selected females (1 to 3) were mated to a single male, and matings between half-sibs or more closely related individuals were avoided. Selection percentages ranged from 13.1 to 45.5% (except the control line) over the nine generations of selection (Table 1). A nonselected control population was also maintained by the same management. Management Procedures. Fertilized eggs from the parent stock were collected daily, cleaned, stored, and incubated. Quail were kept in electrically heated batteries until 7 weeks of
595
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ABSTRACT Two lines of Japanese quail were divergently selected for high and low antibody responses after beii.g twice injected at 4 and 6 weeks of age with inactivated Newcastle disease virus antigen. Afte seven generations of selection, the serum antibody level in the high line was 24% greater than the level in the unselected control lines, whereas the low line antibody level was approximately 37% less than that in the control line. The estimated heritability ( H | + D ) was approximately .12 ± .50. The realized h 2 , calculated from coefficient of regression formula on seven generations (G 3 to G 9 ), was .07. (Key words: selective breeding, Newcastle disease virus, Japanese quail)
TAKAHASHI ET AL.
596
TABLE 1. Outline of selection for high and low antibody responses to inactivated Newcastle disease virus
Percentage of selection
N o . of paired matings
HI of p r o g e n y 2 (No. of progeny tested)
Difference (H-L, R-L, H-R) in HI titer of progeny
33 30 41
6.54 ± 2 . 3 4 ( 1 2 2 ) 4.75 ± 2 . 0 9 ( 1 0 1 ) 5.82 ± 2 . 5 3 ( 1 0 2 )
1.75*** 1.07** .68*** 1
Generation
Line 1
1
H L R
2
H L R
37.7 45.5 54.9
23 23 28
4.78 ± 2.06(159) 3.63 ± 1 . 9 4 ( 1 6 6 ) 4.34 ± 2.19(115)
3
H L R
16.1 19.0 41.8
24 26 32
4.18 ± 1.96(219) 3.65 ± 1 . 8 4 ( 2 0 3 ) 3.28 ± 2.03 ( 98)
.53** -.37 p 0 * **
4
H L R
22.8 24.6 61.2
25 25 30
4.22 ± 1.94(152) 2.54 ± 1.60(157) 3.07 ± 1.73 ( 75)
1.68*** .53* 1.15***
5
H L R
36.8 35.7 80.0
28 28 30
4.10 ± 1.67(175) 2.27 ± 1 . 3 9 ( 1 7 7 ) 3.47 ± 1.65 ( 89)
1.83*** 1.20*** .63**
6
H L R
32.0 31.6 67.4
28 28 30
3.39 ± 1.80 ( 94) 2.09 ± 1 . 6 3 ( 1 2 0 ) 2.83 ± 1.66 ( 94)
1 30* * * 74** .56*
7
H L R
13.1 30.3 63.8
30 30 30
4.61 ± 1.65(156) 2 . 3 4 ± 1.75 (202) 3.71 ± 1 . 6 8 ( 1 1 1 )
2.27*** 1.37*** .90***
8
H L
32.0 24.7
25 25
4.31 ± 1.59(121) 1.99 ± 1 . 6 2 ( 1 2 6 )
2.32***
9
H L
33.0 31.7
20 20
4.63 ± 1 . 9 6 ( 1 0 7 ) 2.63 ± 2.03 (134)
2.00***
H = High line, L = low line, R = control line. Mean hemagglutination inhibition (HI) titer (log 2 ) ± SD.
A 4* * *
*P<.05. **P<.01. ***P<.001.
age and then mated in breeding cages. Fluorescent lighting was provided under a regimen of 14 hr of light and 10 hr of darkness. The quail were fed a commercial feed (23% protein, 2.5% fat, 4.0% fiber, 12.5% ash, 2.5, 2.5% Ca, .55% P) throughout the experiments. Immunization and Hemagglutination Inhibition Assay for Selection. Four-week-old birds were immunized by intraperitoneal injection with .5 ml of inactivated NDV (Kyoto Biken Co., Japan). Two weeks later they were reimmunized with a .5 ml intraperitoneal injection. Seven days after the second injection, blood was collected from the jugular vein and allowed to clot at 37 C for 30 min and at 4 C overnight. Sera were stored at —20 C.
Hemagglutination inhibition (HI) titers to NDV were determined by the microtiter plate method (Beard and Wilkes, 1973). A negative and a high titer positive control were present in all titrations. The titers are expressed as log 2 of the reciprocal of the last dilution showing HI. Fitness Parameters. Comparison between the selected lines were also made for egg weight, egg production, fertility, hatchability, and rearing rate. Eggs produced from 6 to 16 days after mating were collected and weighed. Fertility was calculated as a percentage of fertilized eggs to incubated eggs. Hatchability was calculated as a percentage of hatching to fertilized eggs after 20 days of incubation. Rearing rate was calculated as a percentage of
Downloaded from http://ps.oxfordjournals.org/ at Karolinska Institutet University Library on June 2, 2015
1 2
1 C* * *
.71**
ANTIBODY RESPONSE IN JAPANESE QUAIL
RESULTS Selection for High and Low Antibody Titers. Table 1 shows the means and standard deviation for serum antibody levels in the high, low, and control (random) lines with the number of progeny, percentage of selection, and pairs of mating with generations. The HI values in the base population (G 0 ) were 6.32 ± 3.01 (n = 234). In the G ] , differences in HI antibody levels between the H and L lines were already significant (P<.001). The significance between lines continued through the 9 generations of selection. The HI titers for the control line were intermediate to those for the selected lines excepting G 3 . As shown in Table 1, there is some variation in the actual values for HI titer throughout the generations. However, after 7 generations of selection, the serum antibody level in the high line was 24% greater than the level in the unselected control lines, whereas the low line level was approximately 37% less than that in the control line. Thus, the genetic response was greater in selecting for low than for high antibody production (Fig 1). The regression formula throughout G 7 was y = 2.86 + 108.6 for the high line, and y = - 5 . 1 4 + 102.8 for the low line (X = generation, y = percent deviation from control). Also, the regression formula of the differences in HI titers between lines (y) over generation (X) was calculated as y = .19 X + .67 (Fig 2). Thus, the difference between the lines was separated by .19 for each generation.
FIG. 1. Percentage of hemagglutination inhibition (HI) antibody deviation from control (R) by line and generation. H, high and L, low antibody response lines.
The heritability first was estimated by analysis of variance based upon variance factors of HI titers in the base population (14 pairs of parents) and its progeny (84 birds). The h | was .92 ± .48 and h j , was .67 ± .32. In all, h | + D was .12 ± .37. The realized h 2 calculated from the coefficient of regression formula for nine generations (Gi to G 9 ) and seven generations (G 3 to G 9 ) were .001 and .07, respectively. Comparisons of Other Traits. As shown in Table 2, egg weight appeared to decrease with generation in each line. In the high line, hatchability appeared to decrease with subsequent generations, whereas in the low line it did not. No differences were detected in egg production and feritility. The fitness index was almost the same for all generations. DISCUSSION
Although a previous attempt to conduct a similar selection experiment (Phamm, 1974)
FIG. 2. Differences of mean hemagglutination inhibition (HI) antibody titers between high (H) and low antibody response (L) lines by generation.
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hatchings alive at 7 weeks. Also, egg production was calculated as a percentage (from 6 to 16 days) after the pairing of the parents. A fitness index was calculated from egg production X fertility X hatchability X rearing rate. Statistical Methods. The t test was used to test for significance of differences between H and R, or H and L each generation. Sex differences were not tested for significance, because it has already been demonstrated by Phamm (1974) that sex differences were not significant. In the genetic experiments, heritabilities were calculated by analysis of variance based upon variance factors of HI titers in the base population and its progeny. The parent consisted of 14 pairs (14 males and 28 females), and h 2 was estimated from their progeny (84 birds). The realized h 2 was calculated from coefficient of regression formula to 9 generation (Gx to G 9 ) and 7 generation (G3 to G9).
597
TAKAHASHI ET AL.
598
TABLE 2. Fitness parameters in selection for high and low antibody responses to inactivated Newcastle disease virus Fitness parameter 1 Generation
Fitness index 3
Rearing4 rate
88.6 82.4 88.1
43.2 41.6 40.3
56.5 63.0 58.3
85.0 97.4 92.2
88.9 80.2 88.0
38.8 42.6 32.3
65.3 65.3 46.7
90.0 + 11.3 95.6 ± 10.4 93.7 ± 13.5
95.4 88.4 94.0
78.5 79.1 76.2
47.3 43.8 59.2
70.2 65.6 88.3
10.0 ± 6 9.7 ± 7 10.0 ± 6
89.0 + 15.8 92.9 ± 13.3 91.8 ± 13.7
85.5 92.9 77.6
89.0 79.3 83.8
52.9 51.2 38.6
78.1 74.8 64.6
H L R
10.0 + 7 10.3 ± 8 10.4 ± 7*
97.3 ± 16.1 95.0 ± 10.7 93.3 ± 12.4
93.7 95.1 100.0
86.8 73.5 87.1
60.4 54.7 57.4
76.3 82.4 70.6
H L R
9.8 ± 6 10.3 + 6*** 10.1 ± 7
92.0 ± 11.9 91.8 ± 10.0 90.7 + 11.1
91.0 78.8 98.2
70.3 69.4 87.2
33.9 42.2 53.3
57.7 84.0 68.6
H L R
9.3 ± 8 9.9 ± 8 * * 10.0 ± 8**
84.8 + 18.3 89.7 + 12.2 93.0 ± 11.7
96.7 96.0 97.8
74.0 76.8 82.9
43.8 48.1 61.1
72.2 72.7 81.0
H L
9.9 ± 7 10.0 ± 7
85.8 + 16.1 92.2 ± 10.0
89.4 95.9
78.2 85.2
36.8 40.1
61.4 53.2
89.4 ± 11.4 88.9 ± 11.8
92.4 90.5
70.9 76.7
40.2 49.2
68.6 79.8
Egg wt.
Egg prod.
H L R
(g) 10.7 ± 7 10.8 ± 6 10.7 + 7
87.0 + 12.0 83.0 ± 17.7 80.0 + 21.6
99.2 96.5 98.0
H L R
10.0 ± 8 10.8 ± 9 11.0 ± 8
78.6 ± 15.4 83.6 ± 12.7 85.3 ± 11.8
H L R
9.7 ± 6 10.3 ± 6** 10.5 + 7* *#
H L R
H L
9.0 ± 6 9.5 ± 8*
Fertility
Hatchability
(%)
1
Mean ± SD.
2
H = High line, L = low line, R = control line.
3
Egg prod X fertility X hatchability X rearing rate.
4
Percentage of hatchtings alive at 7 weeks.
-
*P<.05. **P<01. ***P<.001.
failed, the birds in the present experiment diverged quickly into those with high and low responses as shown in Table 1. However, the divergence rate was slow compared with other successful genetic selection experiments. For example, in mice (Biozzi., 1970), the populations were completely separated by the nineth generation. Scheibel (1943) succeeded in separating lives of guinea pigs for six consecutive generations between good and poor lines with respect to the production of diphtheria antitoxin.
In our selective breeding experiments there have been two basic differences, species of animal and test antigen. In chickens, Siegel and Gross (1980) showed the response to divergent selection for high and low 5-day antibody titers was consistent with that noted for mice, which had also received SRBC, during two bidirectional selection experiments involving antibody response to SRBC. In our present experiments, heritability (h£ jO of HI antibody production in G 0 was .12 ± .50. The value was approximately the
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Line 2
ANTIBODY RESPONSE IN JAPANESE QUAIL
complex in quail, compared with other species such as mice or guinea pigs. The fitness index during the present selection remained almost the same throughout all generations. REFERENCES Beard, C. W., and W. J. Wilkes, 1973. A simple and rapid microtest procedure for determining Newcastle hemagglutination-inhibition (HI) antibody titers. Proc. US Anim. Health Assoc. 77:596-600. Benedict, A. A., L. W. Pollard, P. R. Morrow, H. Abplanalp, P. H. Maurer, and W. E. Briles, 1975. Genetic control of immune responses in chickens. 1. Responses to a terpolymer of poly (Glu 60 Ala 30 Tyr 10 ) associated with the major histocompatibility complex. Immunogenetics 2: 313— 324. Biozzi, G., C. Stiffel, D. Mouton, Y. Bouthillier, and C. Decreusefond, 1970. Genetic selection for antibody production in mice. Pages 161—167 in Protides of the Biological Fluids. H. Peeters, ed. Pergamon Press, Oxford. Biozzi, G., M. Siqueira, C. Stiffel, O. M. Ibanez, D. Mouton, and V.C.A. Ferreira, 1980. Genetic selections for relevant immunological functions. Pages 432—457 in Progress in immunology. IV. M. Fougereau and J. Dausset, ed., Academic Press, New York, NY. Scheibel, I. F., 1943. Hereditary differences in the capacity of guinea pigs for the production of diphtheria antitoxin. Acta Pathol. Microbiol. Scand. 20:464-484. Siegel, P. B., and W. B. Gross, 1980. Production and persistence of antibodies in chickens to sheep erythrocytes. 1. Directional selection. Poultry Sci. 5 9 : 1 - 5 . Peleg, B. A., M. SoUer, N. Ron, K. Hornstein, T. Brody, and E. Kalmar, 1976. Familial differences in antibody response of broiler chickens to vaccination with attenuated and inactivated Newcastle disease virus vaccine. Avian Dis. 20: 661-668. Phamm, M. K., 1974. Selective breeding for high and low antibody synthesis of Newcastle disease virus in Japanese quails. Ph.D. diss., Tohoku Univ. Sendai, Japan.
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same, as described in other experiments (Peleg et ah, 1976). However, the h 2 for each generation was very low and variable (data not shown). Such variance, especially in the early stage of selection, may be due to the heterogeneous characteristics of parents used in selection or the interference factors to be described. Thus, the heritability (.07) calculated from the coefficient of regression for seven generations (G 3 to G 9 ) is a more reliable estimate than h 2 for each generation. We have no data as to whether the selection rate was influenced by the test antigen. However, in general, some factors that genetically or environmentally interfere with the immune response may exist depending on the type of antigen. Among these factors, it is well known that passively transmitted maternal antibodies interfere with the immune responses of progeny. Therefore, the first immunization was carried out at the age of 4 weeks of age after the disapearance of the maternal antibodies. The next factor to be considered is whether cell-associated or cell-bound antibodies were still present 4 weeks after immunization. Humoral antibodies were determined; however, no determination of cell-associated or cellbound anti-NDV antibodies as natural antibodies existed or remained. This factor may also be influenced by the phagocytic activities of the antigen. Apart from these environment interference factors, it has been demonstrated (Benedict et ah, 1975) that responsiveness to synthetic antigen is associated with the B-complex, which is known to be a major histocompatibility complex in chickens. Therefore, the fact that the high and low responder lines cannot be completely separated may reflect the genetic complexity of the major histocompatibility
599
1984 Poultry Science 63:595-599 INTRODUCTION
Recent selection research based on antibody producing ability (Biozzi, et al, 1980) demonstrated that this ability was controlled by multiple genes and could be correlated with other immune abilities such as antibody producing ability to unrelated antigens or resistance to several diseases. These facts suggest that genetic regulation systems for antibody production are involved with those of general antibody production or resistance. Further evidence on the relationships between antibody producing ability and natural resistance must be obtained to determine whether it is possible to select animals for high genetic resistance to pathogenic microbes without inducing prophylactic infections by such organisms. The purpose of this paper is to report the development of divergent lines of Japanese quail on the basis of their high and low serum antibody responses to inactivated Newcastle disease virus (NDV) antigen. This paper reports the selective process through the 9 generation during these selection experiments. Subsequent
'Supported in part by Grant-in-Aid, No 456194, from the Scientific Research Fund of the Ministry of Education, Science and Culture, Japan. 2 The quail were sent to the National Institute for Environmental Studies. At this Institute, selection has proceeded under H. Takahashi' direction. 3 The National Institute for Environmental Studies, Yatabemachi, Tsukuba, Ibaraki 300-21, Japan. 4 Reprint requests.
papers will consider the immunological traits of these lines. MATERIALS AND METHODS
Selection Procedures. The foundation stock consisted of 132 birds (T) maintained at the Laboratory of Animal Breeding, the University of Tohoku, and 139 birds (G) acquired from the University of Gifu. The T line had been selected by Phamm (1974) for high and low antibody response to NDV, and his selection experiment was abandoned after eight generations because of low fertility of the population. A base population was obtained by reciprocal matings of the T and G population. Sixty-three matings of T males and G females and 54 matings of T females and G males produced 234 offspring. High and low antibody response lines had been developed by mass selection of the highest responding birds at 7 weeks of age as parents of the high line and the lowest responding birds at the same age as parents of the low line. Selected females (1 to 3) were mated to a single male, and matings between half-sibs or more closely related individuals were avoided. Selection percentages ranged from 13.1 to 45.5% (except the control line) over the nine generations of selection (Table 1). A nonselected control population was also maintained by the same management. Management Procedures. Fertilized eggs from the parent stock were collected daily, cleaned, stored, and incubated. Quail were kept in electrically heated batteries until 7 weeks of
595
Downloaded from http://ps.oxfordjournals.org/ at Karolinska Institutet University Library on June 2, 2015
ABSTRACT Two lines of Japanese quail were divergently selected for high and low antibody responses after beii.g twice injected at 4 and 6 weeks of age with inactivated Newcastle disease virus antigen. Afte seven generations of selection, the serum antibody level in the high line was 24% greater than the level in the unselected control lines, whereas the low line antibody level was approximately 37% less than that in the control line. The estimated heritability ( H | + D ) was approximately .12 ± .50. The realized h 2 , calculated from coefficient of regression formula on seven generations (G 3 to G 9 ), was .07. (Key words: selective breeding, Newcastle disease virus, Japanese quail)
TAKAHASHI ET AL.
596
TABLE 1. Outline of selection for high and low antibody responses to inactivated Newcastle disease virus
Percentage of selection
N o . of paired matings
HI of p r o g e n y 2 (No. of progeny tested)
Difference (H-L, R-L, H-R) in HI titer of progeny
33 30 41
6.54 ± 2 . 3 4 ( 1 2 2 ) 4.75 ± 2 . 0 9 ( 1 0 1 ) 5.82 ± 2 . 5 3 ( 1 0 2 )
1.75*** 1.07** .68*** 1
Generation
Line 1
1
H L R
2
H L R
37.7 45.5 54.9
23 23 28
4.78 ± 2.06(159) 3.63 ± 1 . 9 4 ( 1 6 6 ) 4.34 ± 2.19(115)
3
H L R
16.1 19.0 41.8
24 26 32
4.18 ± 1.96(219) 3.65 ± 1 . 8 4 ( 2 0 3 ) 3.28 ± 2.03 ( 98)
.53** -.37 p 0 * **
4
H L R
22.8 24.6 61.2
25 25 30
4.22 ± 1.94(152) 2.54 ± 1.60(157) 3.07 ± 1.73 ( 75)
1.68*** .53* 1.15***
5
H L R
36.8 35.7 80.0
28 28 30
4.10 ± 1.67(175) 2.27 ± 1 . 3 9 ( 1 7 7 ) 3.47 ± 1.65 ( 89)
1.83*** 1.20*** .63**
6
H L R
32.0 31.6 67.4
28 28 30
3.39 ± 1.80 ( 94) 2.09 ± 1 . 6 3 ( 1 2 0 ) 2.83 ± 1.66 ( 94)
1 30* * * 74** .56*
7
H L R
13.1 30.3 63.8
30 30 30
4.61 ± 1.65(156) 2 . 3 4 ± 1.75 (202) 3.71 ± 1 . 6 8 ( 1 1 1 )
2.27*** 1.37*** .90***
8
H L
32.0 24.7
25 25
4.31 ± 1.59(121) 1.99 ± 1 . 6 2 ( 1 2 6 )
2.32***
9
H L
33.0 31.7
20 20
4.63 ± 1 . 9 6 ( 1 0 7 ) 2.63 ± 2.03 (134)
2.00***
H = High line, L = low line, R = control line. Mean hemagglutination inhibition (HI) titer (log 2 ) ± SD.
A 4* * *
*P<.05. **P<.01. ***P<.001.
age and then mated in breeding cages. Fluorescent lighting was provided under a regimen of 14 hr of light and 10 hr of darkness. The quail were fed a commercial feed (23% protein, 2.5% fat, 4.0% fiber, 12.5% ash, 2.5, 2.5% Ca, .55% P) throughout the experiments. Immunization and Hemagglutination Inhibition Assay for Selection. Four-week-old birds were immunized by intraperitoneal injection with .5 ml of inactivated NDV (Kyoto Biken Co., Japan). Two weeks later they were reimmunized with a .5 ml intraperitoneal injection. Seven days after the second injection, blood was collected from the jugular vein and allowed to clot at 37 C for 30 min and at 4 C overnight. Sera were stored at —20 C.
Hemagglutination inhibition (HI) titers to NDV were determined by the microtiter plate method (Beard and Wilkes, 1973). A negative and a high titer positive control were present in all titrations. The titers are expressed as log 2 of the reciprocal of the last dilution showing HI. Fitness Parameters. Comparison between the selected lines were also made for egg weight, egg production, fertility, hatchability, and rearing rate. Eggs produced from 6 to 16 days after mating were collected and weighed. Fertility was calculated as a percentage of fertilized eggs to incubated eggs. Hatchability was calculated as a percentage of hatching to fertilized eggs after 20 days of incubation. Rearing rate was calculated as a percentage of
Downloaded from http://ps.oxfordjournals.org/ at Karolinska Institutet University Library on June 2, 2015
1 2
1 C* * *
.71**
ANTIBODY RESPONSE IN JAPANESE QUAIL
RESULTS Selection for High and Low Antibody Titers. Table 1 shows the means and standard deviation for serum antibody levels in the high, low, and control (random) lines with the number of progeny, percentage of selection, and pairs of mating with generations. The HI values in the base population (G 0 ) were 6.32 ± 3.01 (n = 234). In the G ] , differences in HI antibody levels between the H and L lines were already significant (P<.001). The significance between lines continued through the 9 generations of selection. The HI titers for the control line were intermediate to those for the selected lines excepting G 3 . As shown in Table 1, there is some variation in the actual values for HI titer throughout the generations. However, after 7 generations of selection, the serum antibody level in the high line was 24% greater than the level in the unselected control lines, whereas the low line level was approximately 37% less than that in the control line. Thus, the genetic response was greater in selecting for low than for high antibody production (Fig 1). The regression formula throughout G 7 was y = 2.86 + 108.6 for the high line, and y = - 5 . 1 4 + 102.8 for the low line (X = generation, y = percent deviation from control). Also, the regression formula of the differences in HI titers between lines (y) over generation (X) was calculated as y = .19 X + .67 (Fig 2). Thus, the difference between the lines was separated by .19 for each generation.
FIG. 1. Percentage of hemagglutination inhibition (HI) antibody deviation from control (R) by line and generation. H, high and L, low antibody response lines.
The heritability first was estimated by analysis of variance based upon variance factors of HI titers in the base population (14 pairs of parents) and its progeny (84 birds). The h | was .92 ± .48 and h j , was .67 ± .32. In all, h | + D was .12 ± .37. The realized h 2 calculated from the coefficient of regression formula for nine generations (Gi to G 9 ) and seven generations (G 3 to G 9 ) were .001 and .07, respectively. Comparisons of Other Traits. As shown in Table 2, egg weight appeared to decrease with generation in each line. In the high line, hatchability appeared to decrease with subsequent generations, whereas in the low line it did not. No differences were detected in egg production and feritility. The fitness index was almost the same for all generations. DISCUSSION
Although a previous attempt to conduct a similar selection experiment (Phamm, 1974)
FIG. 2. Differences of mean hemagglutination inhibition (HI) antibody titers between high (H) and low antibody response (L) lines by generation.
Downloaded from http://ps.oxfordjournals.org/ at Karolinska Institutet University Library on June 2, 2015
hatchings alive at 7 weeks. Also, egg production was calculated as a percentage (from 6 to 16 days) after the pairing of the parents. A fitness index was calculated from egg production X fertility X hatchability X rearing rate. Statistical Methods. The t test was used to test for significance of differences between H and R, or H and L each generation. Sex differences were not tested for significance, because it has already been demonstrated by Phamm (1974) that sex differences were not significant. In the genetic experiments, heritabilities were calculated by analysis of variance based upon variance factors of HI titers in the base population and its progeny. The parent consisted of 14 pairs (14 males and 28 females), and h 2 was estimated from their progeny (84 birds). The realized h 2 was calculated from coefficient of regression formula to 9 generation (Gx to G 9 ) and 7 generation (G3 to G9).
597
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598
TABLE 2. Fitness parameters in selection for high and low antibody responses to inactivated Newcastle disease virus Fitness parameter 1 Generation
Fitness index 3
Rearing4 rate
88.6 82.4 88.1
43.2 41.6 40.3
56.5 63.0 58.3
85.0 97.4 92.2
88.9 80.2 88.0
38.8 42.6 32.3
65.3 65.3 46.7
90.0 + 11.3 95.6 ± 10.4 93.7 ± 13.5
95.4 88.4 94.0
78.5 79.1 76.2
47.3 43.8 59.2
70.2 65.6 88.3
10.0 ± 6 9.7 ± 7 10.0 ± 6
89.0 + 15.8 92.9 ± 13.3 91.8 ± 13.7
85.5 92.9 77.6
89.0 79.3 83.8
52.9 51.2 38.6
78.1 74.8 64.6
H L R
10.0 + 7 10.3 ± 8 10.4 ± 7*
97.3 ± 16.1 95.0 ± 10.7 93.3 ± 12.4
93.7 95.1 100.0
86.8 73.5 87.1
60.4 54.7 57.4
76.3 82.4 70.6
H L R
9.8 ± 6 10.3 + 6*** 10.1 ± 7
92.0 ± 11.9 91.8 ± 10.0 90.7 + 11.1
91.0 78.8 98.2
70.3 69.4 87.2
33.9 42.2 53.3
57.7 84.0 68.6
H L R
9.3 ± 8 9.9 ± 8 * * 10.0 ± 8**
84.8 + 18.3 89.7 + 12.2 93.0 ± 11.7
96.7 96.0 97.8
74.0 76.8 82.9
43.8 48.1 61.1
72.2 72.7 81.0
H L
9.9 ± 7 10.0 ± 7
85.8 + 16.1 92.2 ± 10.0
89.4 95.9
78.2 85.2
36.8 40.1
61.4 53.2
89.4 ± 11.4 88.9 ± 11.8
92.4 90.5
70.9 76.7
40.2 49.2
68.6 79.8
Egg wt.
Egg prod.
H L R
(g) 10.7 ± 7 10.8 ± 6 10.7 + 7
87.0 + 12.0 83.0 ± 17.7 80.0 + 21.6
99.2 96.5 98.0
H L R
10.0 ± 8 10.8 ± 9 11.0 ± 8
78.6 ± 15.4 83.6 ± 12.7 85.3 ± 11.8
H L R
9.7 ± 6 10.3 ± 6** 10.5 + 7* *#
H L R
H L
9.0 ± 6 9.5 ± 8*
Fertility
Hatchability
(%)
1
Mean ± SD.
2
H = High line, L = low line, R = control line.
3
Egg prod X fertility X hatchability X rearing rate.
4
Percentage of hatchtings alive at 7 weeks.
-
*P<.05. **P<01. ***P<.001.
failed, the birds in the present experiment diverged quickly into those with high and low responses as shown in Table 1. However, the divergence rate was slow compared with other successful genetic selection experiments. For example, in mice (Biozzi., 1970), the populations were completely separated by the nineth generation. Scheibel (1943) succeeded in separating lives of guinea pigs for six consecutive generations between good and poor lines with respect to the production of diphtheria antitoxin.
In our selective breeding experiments there have been two basic differences, species of animal and test antigen. In chickens, Siegel and Gross (1980) showed the response to divergent selection for high and low 5-day antibody titers was consistent with that noted for mice, which had also received SRBC, during two bidirectional selection experiments involving antibody response to SRBC. In our present experiments, heritability (h£ jO of HI antibody production in G 0 was .12 ± .50. The value was approximately the
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Line 2
ANTIBODY RESPONSE IN JAPANESE QUAIL
complex in quail, compared with other species such as mice or guinea pigs. The fitness index during the present selection remained almost the same throughout all generations. REFERENCES Beard, C. W., and W. J. Wilkes, 1973. A simple and rapid microtest procedure for determining Newcastle hemagglutination-inhibition (HI) antibody titers. Proc. US Anim. Health Assoc. 77:596-600. Benedict, A. A., L. W. Pollard, P. R. Morrow, H. Abplanalp, P. H. Maurer, and W. E. Briles, 1975. Genetic control of immune responses in chickens. 1. Responses to a terpolymer of poly (Glu 60 Ala 30 Tyr 10 ) associated with the major histocompatibility complex. Immunogenetics 2: 313— 324. Biozzi, G., C. Stiffel, D. Mouton, Y. Bouthillier, and C. Decreusefond, 1970. Genetic selection for antibody production in mice. Pages 161—167 in Protides of the Biological Fluids. H. Peeters, ed. Pergamon Press, Oxford. Biozzi, G., M. Siqueira, C. Stiffel, O. M. Ibanez, D. Mouton, and V.C.A. Ferreira, 1980. Genetic selections for relevant immunological functions. Pages 432—457 in Progress in immunology. IV. M. Fougereau and J. Dausset, ed., Academic Press, New York, NY. Scheibel, I. F., 1943. Hereditary differences in the capacity of guinea pigs for the production of diphtheria antitoxin. Acta Pathol. Microbiol. Scand. 20:464-484. Siegel, P. B., and W. B. Gross, 1980. Production and persistence of antibodies in chickens to sheep erythrocytes. 1. Directional selection. Poultry Sci. 5 9 : 1 - 5 . Peleg, B. A., M. SoUer, N. Ron, K. Hornstein, T. Brody, and E. Kalmar, 1976. Familial differences in antibody response of broiler chickens to vaccination with attenuated and inactivated Newcastle disease virus vaccine. Avian Dis. 20: 661-668. Phamm, M. K., 1974. Selective breeding for high and low antibody synthesis of Newcastle disease virus in Japanese quails. Ph.D. diss., Tohoku Univ. Sendai, Japan.
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same, as described in other experiments (Peleg et ah, 1976). However, the h 2 for each generation was very low and variable (data not shown). Such variance, especially in the early stage of selection, may be due to the heterogeneous characteristics of parents used in selection or the interference factors to be described. Thus, the heritability (.07) calculated from the coefficient of regression for seven generations (G 3 to G 9 ) is a more reliable estimate than h 2 for each generation. We have no data as to whether the selection rate was influenced by the test antigen. However, in general, some factors that genetically or environmentally interfere with the immune response may exist depending on the type of antigen. Among these factors, it is well known that passively transmitted maternal antibodies interfere with the immune responses of progeny. Therefore, the first immunization was carried out at the age of 4 weeks of age after the disapearance of the maternal antibodies. The next factor to be considered is whether cell-associated or cell-bound antibodies were still present 4 weeks after immunization. Humoral antibodies were determined; however, no determination of cell-associated or cellbound anti-NDV antibodies as natural antibodies existed or remained. This factor may also be influenced by the phagocytic activities of the antigen. Apart from these environment interference factors, it has been demonstrated (Benedict et ah, 1975) that responsiveness to synthetic antigen is associated with the B-complex, which is known to be a major histocompatibility complex in chickens. Therefore, the fact that the high and low responder lines cannot be completely separated may reflect the genetic complexity of the major histocompatibility
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