Mr Black comes from an agricultural background, where artificial insemination AI is common, especially for dairy cows and pigs, so the idea wasn't as alien to him as it might be to most. He says: "Obviously your dog has only got a five-year breeding life. The problem of diminished gene pools is one that has been troubling breeders for years.
In , a documentary broadcast by the BBC revealed the health problems suffered by some pedigree dogs after years of inbreeding. It claimed the problems were caused by elite breeders prizing looks over health. The Kennel Club - which operates the national register for pedigree dogs in the UK - came under significant criticism.
The BBC dropped Crufts from its schedules. The programme didn't just criticise breed standards. It also highlighted the dangers of small gene pools. The six years since have seen a rise in the number of dogs conceived by AI - a process in which semen is extracted from the male dog and inseminated into a bitch separately. For many breeders, the practice is about expanding the gene pool and creating healthier dogs.
But critics have argued that that bypassing natural mating methods could create animals that can't breed by themselves - and that AI makes it even easier for one dog to father many pups, which is itself a genetic risk.
Figures from the Kennel Club show approximately 1, puppies have been born through AI in the past 17 years. In , there was just one AI puppy - compared to 82 in The figures peaked last year, with registered AI puppies. The rise is small, but steady. For breeders, the practice can be fruitful - though not without significant effort and expense on their part.
They can sell the semen of their best dogs on to willing buyers. Certain savvy breeders advertise the semen of their prize dogs on their websites - price on request. Sandra Brownlie, a Lanark-based breeder of Boerboel dogs, owns an "enviable collection of frozen semen", according to her website, which she started collecting as she was also concerned about the small gene pool of Boerboels in the UK.
Slowly pull the plunger outwards while inside the artificial vagina, and this will collect the seminal fluids into the barrel of the syringe. Insert the 1. Breed and size of the dog will determine how much of the tube should be inserted. For dogs weighing less than 5 pounds, it is best for a veterinarian to perform the procedure.
For small breeds such as Pomeranians and Chihuahuas, insert the tube only 2 inches. For larger breeds such as Great Danes and German Shepherds, it is recommended that the tube be inserted only 5 inches. An incision is made into the abdomen, and the uterus is found.
The semen is injected either into the uterine body or at the base of either horn. This procedure can be done laparoscopically, but because this requires additional equipment, training, and cost, it is generally not preferred.
Conception rates can be as high as percent when the estrus cycle has been appropriately managed. Artificial insemination has become a popular means of mating, and, with the development of these different techniques, it can be performed easily and with relatively good conception rates and litter sizes. The important aspect with any of these techniques is proper cycle management.
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On table 5 , the most frequent indications for routine semen evaluation are presented. Semen evaluation is also frequently performed in the absence of known reproductive pathology, upon request of the owner. In addition, it can be performed at a predetermined moment after the diagnosis of a clinical disease that may have negative reflects on the potential fertility of a male dog.
It should be notice that reliable in vitro estimation of the real fertilizing ability of sperm cells is not always possible. Usually, in males with aspermic no ejaculate , azoospermic no spermatozoa , or necrospermic no motile spermatozoa semen, the fertilizing potential may be excluded.
When the quality of semen in a dog with history of unsuccessful matings is low, premises exist to exclude such male from the breeding programme. However, it should always be remembered that the semen characteristics should be recheck times at weeks intervals, to confirm the male infertility.
On the other hand, good in vitro semen quality does not always prove the fertilizing potential of a particular dog. After a prolonged sexual rest dogs may ejaculate many dead, immotile spermatozoa of abnormal morphology;.
In young inexperienced males and dogs which mated earlier only naturally, without experience on semen collection, the obtained semen sample may contain only the part of sperm-rich fraction. Different approaches are available to assess the quality of the dog semen that can be grouped in conventional and advanced techniques. The later, usually requires more sophisticated means for the semen assessment and the support of a technical equipment, while the former may be performed in an inhouse lab.
The conventional approaches to semen evaluation include macroscopical evaluation of the semen volume and colour , but also the microscopical assessment, which will give the concentration and the number of viable cells in the ejaculate. The volume of the ejaculate may be assessed in the calibrated tubes used for semen collection. It mainly dependends on the size of the dog, the size of the prostate gland, the animal age, the frequency of semen collection, the level of erotisation, and the volume of 3 rd fraction collected.
A decrease of semen volume is observed in cases of benign prostatic hyperplasia, prostatic cysts, inflammatory lesions of prostate and testicles, inflammation of epididymis, vas deferens or urethra and at weak libido. The colour of whole ejaculate depends on the volume of third fraction of ejaculate collected, on the concentration of spermatozoa per mL and the potential presence of non-germ cells in the ejaculate. When analysing the colour, one should be aware of the method of collection, as colour varies with the fraction to be analysed and the fact that analysis may been performed on the whole semen or on fractioned semen.
The normal colour of whole ejaculate is greyish-white. Pathological colours include: green-greyish typical for the presence of the pus in semen; red or pink-specific for erythrocytes contamination haemorrhages from urethra or corpora cavernosa, prostatitis ; yellow specific for urine contamination; and brown, if in the presence of blood.
Any kind of semen contamination, such as hair or mud, exclude the specimen from further procedures including artificial insemination or semen preservation. It is therefore important to check the region of praeputial opening before semen collection and to clean it. The presence of sediment consisting of sperm cells at the bottom of the tube is a normal feature if the semen is left for several minutes.
One of the most important step of conventional semen assessment is the subjective evaluation of progressively motile spermatozoa Spz under contrast-phase microscope. The evaluation is performed under the objective of x20 to x If the highly concentrated sperm-rich fraction is collected separately, the semen should be extended with saline or Tris-buffer to a concentration allowing the observation of particular, single sperm cells. The assessment is based on the evaluation of the average percentage of progressively motile spermatozoa in a few different fields of the specimen.
A decrease in the percentage of motile spermatozoa may results from temperature shock, contamination with water, urine, blood or lubricants but also from long sexual abstinence and systemic or infectious diseases, such as brucellosis. Sperm agglutination is always pathological and is frequently found in cases of infectious diseases.
Concentration and total sperm count. In order to find the sperm count per mL, the number of spermatozoa in the one or four large squares depending of the chamber is multiplied by For the assessment of sperm concentration more sophisticated equipment could also be used, such as the spectrophotometer, flow cytometer or computer assisted semen analyser Rijsselaere et al.
A large variety in the total number of spermatozoa per ejaculate is observed in different breeds. It varies between 50 x10 6 up to x 10 6 Spz Linde-Forsberg, ; Oettle Small breeds do not produce as many spermatozoa as large breeds, as sperm cell production is related to the weight of the testicular tissue.
The number of spermatozoa per ejaculate also varies according to age, testicular weight, sexual activity and the size of the dog Amann, The total number of spermatozoa in the ejaculate may be decreased in young and older dogs and in inbred males. Apprehension, absence of the teaser bitch, painful prostate, spine rear limbs may also negatively influence the number of spermatozoa ejaculated.
Sperm morphology. The morphology may be assessed under contrast-phase microscope, but usually the evaluation is performed under light microscope on stained slides. Smears of undiluted or diluted ejaculate are examined microscopically for the presence of structural abnormalities of spermatozoa. The semen is smeared on a glass slide in a similar manner to that of blood, air dried and stained.
The semen may be also stained with a nigrosin-eosin stain. A drop of this stain is gently mixed with a drop of semen on a pre-warmed slide before being smeared, and allowed to air dry. Evaluation of sperm morphology should be completed microscopically using oil immersion, using an objective of x or x A minimum number of spermatozoa should be counted and evaluated for the presence of abnormalities.
The percentage of cells with particular morphological defects and of normal cells are calculated. Traditionally sperm cells abnormalities are divided into primary defects - originating from abnormalities of spermatogenesis and secondary defects - originating from abnormalities of semen maturation, transit through the ductal system and specimen preparation. According to another classification sperm abnormalities may be divided into major defects, negatively correlated with fertility, and minor defects, unassociated with fertility Table 6 Oettle, When a spermatozoon presents more than one abnormality, it should be classified according to the most important abnormality or with the most prevalent one, if they have equal significance Oettle, The assessment of the percentage of live and dead spermatozoa is based on the assumption that dead spermatozoa possess disintegrated plasma membrane allowing eosin penetration.
Thus the percentage of eosin positive cells stained with nigrosin-eosin stain is considered as percentage of dead cells. The evaluation of the percentage of live and dead spermatozoa and the percentage of morphological defects may be performed on the same nigrosin-eosin stained slides. In the past 2 to 3 decades, several strategies were developed to escape the subjectivity in the semen evaluation, related to the experience and skills of the observer, the method of specimen preparation, staining technique and number of cells evaluated, and wich is particularly important when the fertility potential of preserved sperm cells has to be ascertain.
Moreover, implementation of such methodologies, not routinely usable in the small to median veterinary clinics due to their costs, allows accurate comparisons between laboratories worlwide and minimizes occurence of large errors.
Furthermore, advanced semen assessment is essential whenever the semen has to be preserved, in particular for freezing. Advanced semen assessment techniques are sumarized on table 7. In general, the results obtained with these methods are better correlated with the AI outcome than the results of traditional semen evaluation. The use of adequate number of viable sperm cells per dose. On next sections the major issues on timing the AI and available techniques of semen deposition on the bitch genital tract will be discussed.
Obtaining successful pregnancies and adequate number of offspring per litter depends upon the correct timing for mating, as well as for insemination, particularly because bitches are mono-estrous, presenting usually one to two reproductive cycles per year.
Although relationship among behavioral, hormonal and physiological events for the average bitch exists, considerable individual variation are also currently found on what concerns the duration of the estrogenic and early luteal stages proestrus and estrus and of the anestrus Concannon et al.
The bitch usually presents a relatively long follicular phase and considerable variability exists in the onset of estrous behavior and acceptance of the male, making it difficult to determine occurrence of the LH surge and onset of ovulation in this species unless specific methods for timing the ovulation and estimating the fertile period are used Linde-Forsberg, Furthermore, in this species, ovulation of immature oocytes primary oocytes, before extrusion of the first polar body determines the need for a maturation period in distal oviducts that may last for hours Concannon, , ; Tsuitsui, ; Tsutsui et al.
Those particularities in the reproductive physiology may explain why the major cause for infertility in the bitch is the inappropriate breeding management Goodman, ; Linde-Forsberg, Consequently, careful planning of mating time by timing ovulation is a key step in canine artificial insemination.
These evaluations should be performed in sequence and with days intervals for the majority of females if the bitch has been reported to present short heat period, of about 6 to 9 days, is possible that daily evaluations may be needed. On the vaginal cytology, epithelial cells of the vagina change their form in response to estrogen impregnation, and passes from small round cells with a clearly visible cytoplasm in non-estrogenic stages, to larger, cornified, angular shaped-cells with small pyknotic nucleus, almost to the point of disappearing, under the influence of estrogens Figure 1.
Progesterone semi-quantitative immunoenzymatic assays are available for clinical routines, but although rapids, these test lack accuracy. A recent study showed that, in dogs, semi-quantitative methods for progesterone determination are less accurate than the quantitative methods, in particular at intermediate plasma progesterone concentrations Moxon et al. According to this study, the tested semi-quantitative assay estimated higher progesterone concentration than.
RIA radioimmunoassay , which could suggest that the fertilization period had commenced earlier than it was actually the case. In addition to those assays, quantitative radio or chemiluminescent assays can also be used, even if not always available in the house lab, since cross-reactivity exist to the molecule between different species, for example with human progesterone. Schematic representation of the major morphological changes of the predominant epithelial cell in the vaginal cytology during the dog estrous cycle.
Although ovarian ultrasound examination is a reliable and accurate method to determine ovulation in most domestic females, in the bitch fat accumulation in ovarian bursa that encloses the ovaries may difficult the value of the technique. Consequently, follicular dynamics evaluation through ultrasonography US in dogs is still experimental and must follow a very precise protocol, which accuracy increase with the use of frequent examinations.
In a recent study, Fontbonne reported that US was accurate enough to detect the occurrence of ovulation and obtain comparable numbers of ovarian structures between US examinations and macroscopic visual count on the surface of the ovaries after surgical removal, even if only one daily examination was performed. However, that author accepts that features of ovulation may be difficult to visualize in large breeds and in overweight animals. Pre-ovulatory follicles may present different aspects at US.
Usually they appear as round to slightly triangular anechoic structures, sometimes slightly flattened, giving a honeycomb aspect to the ovary Figure 2. Persistence of non-ovulatory follicular structures was perceived in US images after ovulation.
Also, in the immediate post-ovulation period, until 24 hours after US changes of the ovaries at ovulation, hypoechoic structures were observed in most cases Figure 2. Ultrasonographic scans of canine ovaries before and after LH surge and ovulation. US are compared with images of longitudinal sections of canine ovaries of similar stages of follicle developement. It is possible to use vaginal endoscopy to determine the fertile period although it does not allow accurate timing of ovulation.
However, this method requires expensive equipment. Nevertheless, it may give a huge contribution to the vaginal evaluation and detection of anatomical abnormalities that may impair proper reproductive performance. The fluctuation of estrogen and progesterone concentrations in the blood at consecutive stages of estrous cycle in the bitch results in specific morphologic changes of the vaginal mucosa. The observation of the cranial part of the vagina is performed for this purpose.
The deep introduction of the tip of endoscope into the narrow part of the vagina close to the cervix dorsal median postcervical fold or paracervix, is of less diagnostic value Pineda et al.
Vaginoscopic examination is performed using a rigid endoscope mm in diameter, with diagnostic sheath and a length of cm or longer. The examination should be done on the standing animal. Usually there is no need of administration of sedatives. When the tip of the optics reaches the vagina it should be repositioned at horizontal axis. During proestrus the increase of the estrogen concentration results in the oedema of the vaginal mucosa. Vaginoscopy reveals rounded folds in the vagina.
The mucosa of the folds is turgid, pink in colour and with a smooth surface. Normally the bloody discharge is also visible in the vagina. Sometimes, periodic blood outflow from the cervix, through the paracervix may be observed. The lumen of the vagina is narrow, which can be appreciated when the endoscope is advanced cranially.
At the last days of proestrus and at beginning of estrus, the decrease of estrogen concentration and increase of progesterone P 4 level is noted. It results in the collapse of vaginal folds.
Formerly turgid and smooth, the mucosa, becomes wrinkled and shrunked. Vaginal folds become smaller. Maximal intensity of shrinkage of vaginal mucosa is observed between 3 and days of estrous cycle.
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