DE2935285A1 - IMPROVEMENTS RELATING TO ARTIFICIAL INSEMINATION - Google Patents

Description

DESCRIPTION

In modern animal husbandry and rearing, artificial insemination is the most suitable and most economical method of insemination.

Artificial insemination offers numerous beneficial aspects, including the possibility of using semen of a male animal to inseminate a significantly larger number of female animals than by the normal Pairing would be possible. Semen of male animals with particularly good hereditary traits can be frozen and over are kept in stock for a very long period and are available for the insemination of female animals, if good hereditary traits are desirable in the offspring are. Artificial insemination makes it possible for the individual farmer or breeder to produce livestock with the desired properties to "produce" without the risk of inbreeding and without the need to use one or more "unproductive" Male animals available on site for the purpose of breeding to keep.

However, it is known that artificial insemination results in a slightly lower fertility rate compared to that can be achieved through natural insemination. The aim of the present invention is to improve this situation.

For a breeder it would be of great economic importance whether the size of the litter in relation to the normal size for artificial insemination for animals that have multiple births, such as pigs, are enlarged can, or whether the number of offspring carried per

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artificial insemination of animals _for which usually give birth only one offspring such as horse or cow can be increased ,, "

In the field of human medicine it has been in recent years Signs given that a change in fertility may be immunological reasons (Beer a Billingham, The Immunology of Mammalian Reproduction (Prenticehall: Englewood Cliffs, N.J .., 1976), Scott J.S. and Jones W.R., Immunology of Human Reproduction, Academic Press, London 1976).

If this assumption is correct, it is possible with help immunological mechanisms a number of natural phenomena with direct or indirect relevance to fertility to explain. Of these phenomena, the following can be mentioned:

Mating animals from different breeds often leads to However, there is an increased fertility, which is called "Heterose ™" after a term from genetics no physiological explanation of this phenomenon. If heterosis ^, from a physiological point of view, one immunological explanation, then the immunological response should intensify when the animals are not with one another are related and consequently one should theoretically observe a larger heterosis effect, i.e. one Increase in fertility.

As is known, the mating of related animals leads to x In = breeding deterioration and, among other things, % u to reduced fertility. If the physiological level is immunologically conditioned, one should, if the animals are more closely related, a lower immune response and consequently

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expect reduced inbreeding deterioration.

Good results can also be achieved by using a mixture of semen from several male animals same species, which makes the observed percentage of pregnancies higher than when using the semen of the most fertile male in question. However, from a breeding standpoint, it is not It is advantageous to use mixtures of seeds from different males of the same species, since then one cannot predict can determine which male animal will be the biological father of the offspring.

A comparison of the litter size after natural mating and after the use of artificial insemination in pigs shows, as already mentioned, clearly that when using artificial insemination Insemination a smaller litter size is the result. If the ejaculate contains important antigens, the explanation could be this is because the strong thinning of the seed leads to a smaller litter size. Related investigations with hybrid pigs in Norway have shown that when using artificial insemination a very low heterosis is observed for the litter size, while the corresponding heterosis effect is observed in natural mating makes up almost 1.0 piglets. The physiological explanation for this could be the one indicated above. It was also shown that the male pig has a significant influence on the litter size. If there is a correlation between the immune response and litter size (embryo death), then there is good reason to assume that the parents have a Have an influence on the litter size.

An experiment has shown that according to the present Invention can achieve improved fertility when antigens are added to the semen, especially when the

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Semen is diluted with a diluent »The diluent is used to dilute the ejaculate to a large number of insemination doses and it can consist of various buffer mixtures that also contain active ingredients to reduce the thermal shock effect if the dose is to be refrigerated or frozen Antibiotics, nutrients and the like "The diluent can also be based on egg yolk, skimmed milk, dry milk and the like." ESE Hafez _# Reproduction in Farm Animals, pages 153-154, Lea & Felsiger, Philadelphia 1966, reference is also made to Pälson Einarson , Allanbane- Insemination och Insemination Teknik Centraltrykkeriet AB Boras 1972 “From these references it can be seen that one at the production of a g suitable semen diluent to achieve the optimal living conditions for the sperm seed in order to get the greatest possible look! of spenas to enable survival and, consequently, participation in fertilization.

In US Pat. No. 3,718,740, it is described that the motility and fertility of spermatoson in animal semen preparations are increased by adding S-glucurinudase, the motility of the sperm cells being significantly improved after storage. However, this has not yet been mentioned or implied, that seeds or -with a diluent dilute seeds with active ingredients mixed v can ground /, which ensures improve fertility. According DER present invention, seeds or seeds can be mixed plus diluent with antigens and can iaan achieve a larger litter size thereby securely or the chances of getting an excellent

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to achieve carried offspring per fertilization, improve. Suitable antigens can be added to the seed / diluent mixture in the form of leukocytes. However, one can Do not add whole blood as the erythrocytes form agglomerates around the sperm and inactivate them.

In the publication by L.V. Swanson and A.G. Hunter: "Egg Yolk antigens and their effect on fertility in rabbits" (Biology of Reproduction 1, (4), 324-329, 1969) Experiments described to determine which proteins in the egg yolk are antigenic, to define them immunologically, and to determine whether antibodies to egg yolk could affect fertility when mixed with rabbit semen that is in suspended in an egg yolk dilution.

The antigens can also be introduced into the vagina and / or uterus before or after natural or artificial insemination.

In the present invention, antigens are active ingredients to understand that stimulate the female immune system and the antigens can be in the form of cellular antigens or "pure" Antigens "isolated from cells or cell tissue are present. The antigens are preferred in the form of leukocytes or lymphocytes of the same species or different species, or from foreign cells, including from different ones Microorganisms and cells from tissue cultures are used. It is also possible that a synergistic effect is achieved can be made by using different antigens obtained from the same species or from different species had received.

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An antigen can be defined as any substance which stimulates antibody production. Most antigens are proteins, but some have been found to be other large molecules can serve as antigens. For this see, for example, Stanier, Doudoroff and Adelberg: "The Microbial World" (Prentice Hall, Inc. 1958), pp. 594-871.

The following examples, in which leukocytes were used as antigens, illustrate the invention.

example 1

1.73 sows of the Norwegian Lard breed at weaning age and weighing approx. 20 kg were purchased (spring 1978),

2. In the second heat period the sows were divided into three Groups shared:

A. 1/3 of the sows were inseminated in the usual way (control group) .

B. 1/3 of the sows were inseminated with semen, the cellular Antigens (leukocytes) from the same boar used for insemination seeds used was added.

C. 1/3 of the sows were inseminated with semen, the cellular antigens (Leukocytes) from another boar of another Schxveinezucht (Yorkshire) was added.

3. 4-5 weeks after the artificial insemination, the sows were slaughtered and the following parameters recorded:

- Number of corpora lutea

- Number of normal (living) fetuses

- Number of dead fetuses

- size of the fetuses

4. The fetuses were used for later determination of the sex ratio and determining the cause of embryo death.

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The results of this experiment can be summarized as follows:

- The addition of cellular antigens to the semen resulted in significantly larger litters (a higher number of fetuses) in the Compared to using the normal dose of sperm.

- Addition of antigens to the seed resulted in a 12% increase in litter size.

- The probability that the observed difference was due to random fluctuations was less than 2.5%.

The average values for the experiment were:

group number
the
animals 26th Normal fetuses
the number of d. 57 in percent
corp. lut. Relative
Reference
counting A (control) 25th 72, 24 % 100 B. 21st 83, 88 % 114.7 C. 81 112.9

The percentage of normal fetuses in the control group corresponded to normal experience, while the percentages for groups B and C were much higher than usual.

Example 2

To verify the promising results of the previous example, 68 sows (spring 1979) were divided into three Groups divided and inseminated:

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A. Control group , inseminated in the usual way.

B. Inseminated with seeds that are pooled with antigens in the form White blood cells from Yorkshire pigs mixed was.

C- inseminated with seeds that are pooled with antigens in the form Bovine leukocytes of the species Norwegian Red and White was mixed.

The fertility rate was determined as described above «.

group number
the
animals Normal fetuses in percent
the number of d. corp. lut. " Relative
Reference
counting Ä (control) 24
B- 22
C 22 63
79
I 77 100. -
- 116
113

The results of this experiment corroborate those previously obtained Results.

An analysis of the pooled results of the two remote searches yielded the following results:

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group

Number of sows

1978 + 1979

Averages

corpus luteum: normal fetuses X

X ^A

Reference numbers

Control (I) 50 | 13.92

Leukocytes from the sperm donor (II) Leukocytes I from the other

I boar (III) I s

Bovine leukocytes (IV) I.

2.42

9.30

25th

44

I 13.52 2.50 j 10.69

3.42

3.49

14.321 2.23? 10.53 3.53

14.231 2.93

10.27

3.08

100 114.9

113.2 110.4

* Earnings vm varying the number of corpora lutea

corrected

The probability of the existence of an actual sub seriieds in the number of normal fetuses in the control group 'Inue the examined groups yields the following seven ratios!:

! »Ficht corrected for the variation in the number of corpora lutea; 4.73 -

Corrected vsa the variation in the number of corpora lutea: 5.05 **

These results show that there is a highly significant (99%) difference between the results obtained for the control group and the groups examined.

It is known that the fertility is determined by mammals to a great extent by the rate of fetal death. Early fetal death is the main reason why you

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Must breed new animals. For horses, for example, it was shown that 100 matings lead to 40 offspring and 24 prematurely dying fetuses, see E.P. Cunningham ^ Annual Meeting of the European Association of Animal Production (E.A.A.P.) Stockholm 1978.

The addition of antigens to the semen of pigs and cattle led to a significant reduction in premature fetal death, and consequently increased fertility. "This is also to be expected for" horses ".

A summary of the leukocyte isolation can be found in Εφγηη, A. Isolation of Lymphocytes ,, Granulocytes and Macrophages, Scand. J. Immonul -, - Volume 5 _# Suppl. 5, 1976.

1 ml of the leukocytes obtained according to the above work can be added to a semen dose for pigs. About 0 ^ 5 ml of the antigens can be added to a dose of semen for cattle and the insemination can be carried out in a known manner. As can be seen from the test results, the litter size was increased by approx. 12%, which is very important can be" -

According to the F&O, the pig population is the entire World approx. 670 million, of which approx. 60 million are breeding sows

Assuming that 10% of the sows are inseminated and that there is also an increase in fertility corresponding to half a piglet per sow, “ this increase amounts to 3 million additional piglets per year.

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Assuming the value of a piglet at $ 50, so the worldwide total profit amounts to approx. 150 million dollars per year.

In countries where artificial insemination is common, an increase in litter size of the order of 12% to be of great importance.

Many attempts have been made to determine the minimum number of sperm that must be inseminated to achieve a concept. From the accompanying diagram, taken from the book by G.W. Salisbury, N.L. van Denmark and J.R. Lodge, "Physiology of Reproduction and Artificial Insemination of Cattle," W.H. Friedmand & Co, San Francisco 1978, is taken, it can be seen that the probability of conception rapidly with the decrease in the number of inseminated Sperm is falling off. It is generally believed that 8 to 12 million sperm should be inseminated to get one satisfactory degree of conception, i.e. in the area before. 65 to 68% can be achieved.

With a semen dose of approx. 1 million sperm, a maximum of 4 calves per 10 inseminations would be expected. Like from the As can be seen in the diagram, both the X-axis and the Y-axis and the curve itself are in the range below 2 mill. Sperm per dose extrapolated.

Example 3

In order to find out whether the addition of antigens could compensate for a reduction in the number of sperm cells per dose, the The following experiment was carried out: 10 cows of the Norwegian breed Red and White were given a dose of semen containing approx. 1 million sperm and the 3 10 sperm from pigs,

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ie spenas of another species were added, inseminated _for what resulted in 3 calves carried to term. «This is a very good and unexpected result. In the above-mentioned experiment, in which bull semen were mixed with antigens in the form of boar semen, pregnancies and childbearing amounted to about 90% (which is considerably beyond the scope of the above-mentioned diagram). The fact that the fertility rate according to the present Invention can be enlarged, is of great economic importance-, ι * ΐϊ & from the following ersielitlicli is »

Each percent increases the failure rate at Eiadeoi one counts according to UB Lindstrfiki, Efordiske Jordbruksforskeres Congress i Oslo, July 1979 ,. with approx "O, SO US - 0 per EMi tanci year"

According to FAO, the Milsiikulipopulation is the United! States, Europe; r_-cl Russia sea »430 million. In many countries more than 75 0 clsr ICühs IdmsfclicJi are inseminated _ff ηίΈαϊίά, but the» Tmschz-skmiit ösr yt5-iaa.ateB countries with 29 0 aa " so the Eaäs ^ risar, class K = 215 Mi 11 ", cows wsq 3smz would be artificially inseminated"

Sine large ÄnsaLl "ion -küastliclien inseminations X'jisä aaeSi in South America, Australia 'USID. I ^ ^ Sisai conducted esätsliola wird- a large Änsahl 7on Eimäera to FleischerseiagüaEig · annually artificially inseminated Bsreits. Based a'uf ~? Om 200 ■ "• Mill. Inseminations per year, however, would result in a Verfeessarang the conception rate of 2.5 % a profit in the large = - order of 200 million dollars per year.

Further experiments have _gezeigt _f that addition of antigens su to the seed or seeds, which is diluted with a diluent, the percentage being transmitted offspring per

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Increase insemination. Such antigens can be found in the seed as such or in the seed that has been diluted with a diluent is to be added. However, from a practical point of view, the antigen is preferably in the diluent or in a mixture from which the diluent is prepared by adding water. the Antigens can also be introduced into the vagina and / or uterus before or after natural mating or artificial insemination will.

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Claims (12)

"DETJFEL · · SCHON · HERTEL PAT E JS" TA N WÄLT K DR. WOLFGANG MÜLLER-BORE (PATENT ADVOCATE FROM 1927-1973) DR. PAUU DEUPEL. D1PL.-CHEM. DR. ALFRED SCHÖN, DIPL.-CHEM. WERNER HEFiTEL, D1PL.-PHYS. APPROVED REPRESENTATIVES AT THE EUROPEAN PATENT OFFICE REPRESENTATIVES BEFORE THE EUROPEAN ΡΑΤΕΓΉ "OFFICE MANOATAIRES AGRiES PRES!." OFFICE EUROPEEN DES BREVETS S 3219 - dt / n 31 AUG. 1979 HARALD SKJERVOLD, ütveien 15, 1430 As, Norway As, Norway KARE FOSSÜM, Agronomveien 6, Oslo 11, Norway OLA ONSTAD, H ^ gäsveien 23, 1346 Gjettum, Norway "Improvements in connection with artificial insemination" 1. Method of treating seeds, where appropriate is diluted with a diluent, characterized in that the seed is mixed with at least one active ingredient which stimulates the immune system of the female animal. 2. The method according to claim 1, characterized in that that antigens are added to the seed. 3. The method according to claim 1, characterized in that that the semen has cellular antigens such as leukocytes, lymphocytes or other foreign cells, including microorganisms and cells from tissue cultures. " "..- ■ - 030011/0174 MffKOHEJf 86 · SIEBEHTSTR. 4 · POST BOX S60720 ■ KTABEI .: ΜΌΈΒΟΡΑΤ · SBi. (OSOJ 4740OS TEEEX 5-34285 4. The method according to claim 1, characterized in that that the semen is mixed with the semen of an animal of another species. 5. Application of the method according to claim 1 to 4 in a method for increasing the fertility of mammals by artificial insemination, characterized in that the seed dose is mixed with at least one active ingredient, which stimulates the immune system of the female animal. 6. Means for performing the method according to claim in the form of a seed diluent or active ingredient on the Based on buffers, nutrients, emulsifiers and other auxiliary substances and the like, characterized in that it contains at least one active ingredient which the female Stimulates the immune system » 7. seed diluent according to claim 6, characterized in that that it contains antigens. 8. seed diluent according to claim 6, characterized in that that there are cellular antigens such as leukocytes, lymphocytes or other foreign cells including Microorganisms and cells from tissue cultures. 9. Modification of the method for improving fertility according to claim 1 through artificial and natural insemination, characterized in that a liquid is introduced into the vagina and / or uterus which contains at least one active ingredient, which stimulates the female immune system. 10. The method according to claim 9, characterized in that that one introduces an antigen-containing liquid. 030011/0874 293528S 11. The method according to claim 3, characterized / that a liquid is introduced containing cellular antigens, such as leukocytes, lymphocytes or other foreign cells, including microorganisms and cells from tissue cultures. 12. The method according to claim 9, characterized in, that one introduces a liquid containing the seeds of an animal of another species ^ 030011/0874