Breed variations in the incidence of pyometra and mammary tumours in Swedish dogs.

Dogs enrolled in a Swedish insurance company (during 1995-2006) were studied for development of pyometra and mammary tumours (MTs), with special attention to breed and age. There were over 260,000 female dogs with over 1,000,000 dog-years at risk (DYAR) in the database, using data on bitches up to 10 years of age and 110 breeds with over 1000 DYAR. In total, 20 423 bitches were diagnosed with pyometra and 11,758 with MTs and 30,131 with either or both of the two diseases. The incidence rate (IR) for pyometra was 199 (95% CI 196-202), for MTs 112 (95% CI 110-114) and for either or both of the two diseases 297 (95% CI 294-301) dogs per 10,000 DYAR. The mean age of diagnosis pyometra was 7.0 years (SD ± 2.2), MTs 8.0 years (SD ± 1.6). In all breeds, the overall proportion of the bitches that developed disease by 10 years of age was for pyometra 19%, MTs 13%, and either or both of two diseases 30%. The top 10 breeds diagnosed with either or both of the two diseases were the Leonberger (73%), Irish Wolfhound (69%), Bernese Mountain Dog (69%), Great Dane (68%), Staffordshire Bull Terrier (66%), Rottweiler (65%), Bullterrier (62%), Doberman (62%), Bouvier des Flandres (60%), Airdaleterrier (60%). These data provide information of the combined disease incidence in a large number of different breeds. Breed variations in incidence rate suggests genetic components in disease development. Our study may be valuable in the search for genetic risk-factors or protective factors.

Canine herpesvirus during pregnancy and non-pregnant luteal phase.

Canine herpesvirus (CHV) is a widespread infection among dogs that typically get latently infected after exposure and can reactivate the infection after stress. The aim of the present study was to study the effects of latent CHV infection during pregnancy on pregnancy outcome, and to study if there are signs of genital viral reactivation during pregnancy or during non-pregnant luteal phase. Twelve mated bitches and eight control bitches were followed and sampled regularly during pregnancy or non-pregnant luteal phase. Blood samples were taken for antibody analysis and vaginal swabs for real-time PCR analysis. Three of the pregnant bitches were vaccinated against CHV during pregnancy. All bitches had antibodies to CHV. Two pregnant bitches that were not vaccinated had a twofold or larger increase in CHV titre, with no negative effects detected on pregnancy. Higher titres were not associated with smaller litters or with vaccination. There was no consistent variation in antibody titres due to pregnancy or non-pregnant luteal phase. Vaginal excretion of CHV was not detected from any of the bitches.

Tumour necrosis factor in the canine endometrium: an immunohistochemical study.

Tumour necrosis factor (TNF), a pleiotropic cytokine that regulates cell growth and differentiation as well as the synthesis of other cytokines, has been identified in the uterus of several species describing a cyclic pattern, eventually under ovarian steroid regulation. Information is yet limited on the presence of TNF protein in the canine endometrium during the oestrous cycle and early pregnancy. This study depicts the temporal immunolocalization of TNF in the bitch endometrium along the oestrous cycle and changes associated with the early steps of embryo invasion. TNF immunolabelling was found in both the stromal fibroblasts and epithelial components of the canine endometrium in all stages studied. Stromal immunostaining was more intense than that of the epithelia, in all the stages of the oestrous cycle. In addition, a tendency for a decrease in the surface epithelium intensity score was found in early dioestrus. A positive glandular content was only observed in anoestrus and proestrus stages. In early pregnancy (days 13-16), TNF immunolabelling was detected at the embryo-maternal surface, in the syncytium cords and the trophoblast, as well in the endometrial stroma and the basal endometrial glands, but not in the lacunar epithelium. The overall TNF immunoreactivity was higher in early pregnancy samples in comparison with those of the early dioestrus and dioestrus stages, suggesting it plays a role during implantation.

Mortality of life-insured Swedish cats during 1999-2006: age, breed, sex, and diagnosis.

BACKGROUND: A cat life insurance database can potentially be used to study feline mortality. HYPOTHESIS: The aim was to describe patterns of mortality in life-insured Swedish cats. Cats: All cats (<13 years of age) with life insurance during the period 1999-2006 were included. METHODS: Age-standardized mortality rates (MR) were calculated with respect to sex (males and females), age, breed, and diagnosis. Survival to various ages is presented by time period and breed. RESULTS: The total number of cats insured was 49,450 and the number of cat-years at risk (CYAR) was 142,049. During the period, 6,491 cats died and of these 4,591 cats (71%) had a diagnosis, ie, were claimed for life insurance. The average annual MR was 462 deaths per 10,000 CYAR (95% confidence interval, 431-493). Sex-specific rates did not differ significantly. The overall mortality of the Persian and the Siamese groups was higher than that of several other breeds. Overall and breed-specific (for most breeds) survival increased with time when analyzed by 2-year periods. The 6 most common diagnostic categories (ignoring cats recorded as dead with no diagnosis) were urinary, traumatic, neoplastic, infectious, cardiovascular, and gastrointestinal. The MR within diagnostic categories varied by age and breed. CONCLUSIONS AND CLINICAL IMPORTANCE: In this mainly purebred, insured cat population, the overall mortality varied with age and breed but not with sex. The increase in survival over time is likely a reflection of willingness to keep pet cats longer and increased access to and sophistication of veterinary care.

Estradiol measurement after GnRH-stimulation as a method to diagnose the presence of ovaries in the female domestic cat.

There is currently no method to reliably diagnose the presence of ovarian tissue in inactive intact female cats except laparatomy which is an invasive procedure. All available tests require that the queen is in estrus. Obvious overt symptoms of estrus are, however, not always observed and some queens may have only 2 estrus periods/year. Therefore this study was designed to evaluate if it is possible to diagnose the presence of ovarian tissue by measurement of estradiol before and/or after stimulation with a GnRH-analogue. Twenty-two female cats were divided into two groups; 11 females that were known to have been ovariectomized and 11 females that were known to be intact. From each cat a heparinised blood sample was collected from the cephalic vein for resting estradiol and progesterone measurements. All cats were treated with a GnRH-analogue buserelin (Receptal, 0.4microg/kg im). Two hours later a second blood sample was collected. Median estradiol increased after stimulation with buserelin in intact but not in ovariectomized females (11 range 5-21 vs. 20 range 12-41, P=0.004 and 6 range 4-9 vs. 6 range 5-9 P=0.8) and did not overlap between the two groups. The highest estradiol concentration post-GnRH in the ovariectomized group was 9pmol/L while the lowest in the intact group was 12pmol/L. Progesterone was basal in all cats except one both before and after GnRH-stimulation. In conclusion this study demonstrates that measurement of estradiol concentration in plasma 2h after stimulation with a GnRH-analogue seems to be a reliable method to diagnose the presence of ovarian tissue in the female cat.

Concentrations of anti-Müllerian hormone in the domestic cat. Relation with spay or neuter status and serum estradiol.

Female cats with unknown history can be diagnosed as spayed or intact with a GnRH-stimulation test or an LH test independent of the stage in the estrous cycle. However, although most females are correctly diagnosed with the LH test, the sensitivity and specificity are not 100%. The GnRH-stimulation test, although reliable, requires an injection of buserelin 2 hours before the blood sample is collected. Granulosa cells are the only cell type that produces anti-Müllerian hormone (AMH) in females, whereas Sertoli cells produce AMH in males. Anti-Müllerian hormone has been linked to spay status in dogs and cats and to ovarian and testicular pathology and fertility in different species. Our aim was to evaluate serum AMH concentrations in spayed female cats and in intact female cats of known age and reproductive stage (inactive ovaries or luteal phase). In addition, our aim was to compare serum AMH concentrations in intact and neutered male cats. We analyzed serum AMH concentrations in 15 spayed and 16 intact females and in 15 intact and 12 neutered male cats. Serum AMH was below the lowest standard point (<0.14 ng/mL) in all spayed females and neutered males, ranged between 1.3 and 19.0 ng/mL in the intact females and between 4.8 and 81.3 ng/mL in intact males. Thus, the AMH test had 100% sensitivity and specificity to diagnose the presence or absence of ovaries and testes in this study. In addition, in contrast to serum estradiol, serum AMH was not affected by buserelin stimulation (P = 0.459). Serum AMH was not correlated with serum estradiol before (rs = -0.188, P = 0.519) or after (rs = 0.335, P = 0.242) buserelin stimulation in the intact females. Four 6-month-old intact cats (two females and two males) had the highest AMH concentrations which in the females might represent a prepubertal peak previously described in other species and in males is likely due to high concentrations before puberty. In conclusion, we found that the AMH Gen II ELISA is reliable for diagnosing spay and neuter status of cats and that the domestic cat might be an interesting model for studies on AMH dynamics.

Incidence of Diabetes Mellitus in Insured Swedish Cats in Relation to Age, Breed and Sex.

BACKGROUND: Diabetes mellitus (DM) is a common endocrinopathy in cats. Most affected cats suffer from a type of diabetes similar to type 2 diabetes in humans. An increasing prevalence has been described in cats, as in humans, related to obesity and other lifestyle factors. OBJECTIVES: To describe the incidence of DM in insured Swedish cats and the association of DM with demographic risk factors, such as age, breed and sex. ANIMALS: A cohort of 504,688 individual cats accounting for 1,229,699 cat-years at risk (CYAR) insured by a Swedish insurance company from 2009 to 2013. METHODS: We used reimbursed insurance claims for the diagnosis of DM. Overall incidence rates and incidence rates stratified on year, age, breed, and sex were estimated. RESULTS: The overall incidence rate of DM in the cohort was 11.6 cases (95% confidence interval [CI], 11.0-12.2) per 10,000 CYAR. Male cats had twice as high incidence rate (15.4; 95% CI, 14.4-16.4) as females (7.6; 95% CI, 6.9-8.3). Domestic cats were at higher risk compared to purebred cats. A significant association with breed was seen, with the Burmese, Russian Blue, Norwegian Forest cat, and Abyssinian breeds at a higher risk compared to other cats. No sex predisposition was found among Burmese cats. Several breeds with a lower risk of DM were identified. CONCLUSIONS AND CLINICAL IMPORTANCE: Our results verify that the Burmese breed is at increased risk of developing DM. We also identified several previously unreported breeds with increased or decreased risk of DM.

Comparison of fertility data from vaginal vs intrauterine insemination of frozen-thawed dog semen: a retrospective study.

Fertility data from 327 artificial inseminations (AIs) using frozen-thawed dog semen are presented here. The AIs were performed in 274 bitches using semen from 185 males of 76 breeds. The data cover all AIs conducted during 1983 through 1995 at Cryogenetic Laboratories (CLONE) in the United States with AKC-registered and research bitches, and all AIs carried out at the Department of Obstetrics and Gynecology at the Swedish University of Agricultural Sciences in Uppsala, Sweden, using semen frozen by CLONE, in 0.5-mL straws. Semen was frozen using a standardized, three-step liquid nitrogen vapor freezing method. Whelping rates > 70% were obtained when post-thaw motility was 40% or higher. The inseminations were made either directly into the uterus using transcervical catheterization with the Norwegian catheter (NIU; 167 AIs) or a fiberoptic endoscope (EIU; 19 AIs), or in the cranial vagina (VAG; 141 AIs). Resulting whelping rates were 84.4% (NIU), 58.9% (VAG; P < 0.001), and 57.9% (EIU). Increasing the number of VAG AIs per cycle from 1 to 2 enhanced the whelping rate (P < 0.05). The mean interval from the first AI to whelping was 61.8 +/- 2.4 d, and was longer for VAG AIs (62.7 +/- 2.7 d) than for NIU AIs (61.2 +/- 2.1 d; P < 0.001). The mean interval from the last AI was 60.1 +/- 1.9 d, and did not differ between VAG AIs (60.2 +/- 2.2 d) and NIU AIs (60.0 +/- 1.6 d). Gestation length was not influenced by breed or litter size. A total of 1158 pups resulted from the 327 AIs. Litter size was 5.4 +/- 3.0 (NIU), 4.0 +/- 2.7 (VAG; P < 0.001), and 6.0 +/- 2.1 (EIU). Litter size was also influenced by breed (P = 0.006) and, for VAG AIs, by the number of inseminations performed per cycle (P = 0.009). This study is the largest that has been carried out on frozen-thawed dog semen AI. It shows that using a good method for cryopreservation, together with nonsurgical intrauterine AI employing the Norwegian catheter, can yield whelping rates and litter sizes similar to those reported from well-controlled natural matings. Furthermore, this is the first study to show that intrauterine deposition of frozen-thawed dog semen results in a significantly higher whelping rate and larger litter size than vaginal deposition.

Comparison of the GnRH-stimulation test and a semiquantitative quick test for LH to diagnose presence of ovaries in the female domestic cat.

It is generally recommended that female cats not intended for planned breeding are spayed to reduce the population of feral cats and also because spaying is beneficial for the long-term health of the individual. For female cats of unknown origin or with estrous symptoms after spaying there is a need for a reliable method to diagnose or rule out the presence of ovaries to avoid unnecessary surgery. Methods previously recommended include vaginal cytology, evaluation of serum estradiol concentration during suspected estrus, induction of ovulation and subsequent evaluation of progesterone, or explorative laparotomy. These methods have the disadvantages that an accurate diagnosis only can be made during estrus or that an invasive procedure is required. Previously, the use of a GnRH challenge test and a semiquantitative LH test have been reported. Our aim was to compare these two methods. We therefore divided 31 female cats in two groups: (1) intact nonestrous females (N = 16), and (2) previously ovariohysterectomized females (N = 15). A blood sample was collected (Time 0) and 0.4 µg/kg buserelin (Receptal; Intervet, Danderyd, Sweden) was injected im. A new blood sample was collected 120 min after the injection. A drop of serum from the sample collected at Time 0 was placed on the LH test (Witness LH; Synbiotics, Corp., San Diego, CA, USA) and the result was evaluated as negative or positive. The remaining serum was frozen and analyzed for estradiol in one batch. Serum estradiol before buserelin stimulation ranged between 5 and 45 pmol/L (N = 14) in intact nonestrous queens and between 2 and 6 pmol/L (N = 15) in ovariohysterectomized females. Estradiol in samples collected after 120 min ranged between 12 and 51 pmol/L (N = 16) in intact queens and between 1 and 7 pmol/L (N = 15) in spayed cats giving a sensitivity and specificity of 100% for the buserelin stimulation test at a cutoff value of 11 pmol/L. All intact queens were negative in the semiquantitative LH test while 14/15 spayed cats were positive and one was negative giving a sensitivity of 100% and a specificity of 93.8% to detect the presence of ovaries in nonestrous cats.

Prediction of the oocyte recovery rate in the bitch.

In order to investigate the possibility of predicting the recovery rate of oocytes for use in a sperm-zona pellucida binding assay, ovaries were obtained from 67 bitches of 37 different breeds, and cumulus-oocyte complexes (COCs) were recovered by mincing the ovaries with a scalpel. The mean number of COCs recovered was 37.2 +/- 34.1 (range 0-145) per ovary. Age significantly affected COC recovery rates. From bitches 1-6 years old, 54.2 +/- 35.1 COCs/ovary were recovered, compared to 26.4 +/- 29.0 from bitches 7-13 years old (P = 0.003). The morphology of the uterus or the presence or absence of ovarian structures had no significant effect on COC recovery rates, although there was a tendency for more COCs to be recovered from ovaries with only follicles visible on the surface. There were no significant correlations between body weight or ovarian weight and COC recovery rates. There was a high correlation in the COC recovery rate between the two ovaries of a bitch, enabling an approximate estimation of the COC recovery rate from the second ovary when the COCs from the first ovary have been recovered. The large variation in COC recovery rates between bitches stresses the need for storage of canine oocytes in order to secure a high enough number of oocytes for a homologous sperm-zona pellucida binding assay in the dog.

Sperm binding capacity and ultrastructure of the zona pellucida of stored canine oocytes.

Sperm binding to the zona pellucida is a prerequisite for fertilization, and tests that evaluate this function have been described for several species. When carrying out such tests in the canine species, ovaries or oocytes have to be stored to obtain a sufficient number of oocytes at the time of testing. In the present study, the sperm binding capacities of salt-stored oocytes and oocytes from deep frozen ovaries were measured and compared with that of fresh oocytes. Two different procedures for washing the sperm-oocyte complexes (gentle and tough) were used before evaluating the number of bound spermatozoa. The total number of oocytes that bound spermatozoa was significantly lower for both salt-stored and deep frozen oocytes compared with fresh oocytes. Significantly fewer spermatozoa bound to stored oocytes than to fresh oocytes (P

Evaluation of chilled and frozen-thawed canine spermatozoa using a zona pellucida binding assay.

Zona pellucida binding assays provide information about the fertilizing ability of spermatozoa. A zona-binding assay for canine spermatozoa using intact, denuded homologous oocytes has not been evaluated previously. In the present study, an assay using canine oocytes derived from frozen-thawed ovaries was evaluated using three types of semen: fresh untreated; killed; and a 50:50 mixture of untreated and killed spermatozoa. The assays were performed on 3 x 20 oocytes for each sperm treatment, using semen from pooled ejaculates (0.5 x 10(6) spermatozoa in each 50 microliter droplet containing five oocytes). There was a significant difference (P < 0. 001) between all treatments. Thereafter, the same procedure was used to evaluate methods of chilling and freeze-thawing of canine semen. There was a trend (P = 0.067) for more sperm binding after 1 day of chilling compared with after 4 days of chilling. Semen samples frozen using an extender (with or without the addition of Equex STM paste) were evaluated. Equex had a significant (P = 0.034) positive effect on the capacity of the spermatozoa to bind to the zona pellucida. In conclusion, the addition of a zona pellucida binding assay to established in vitro tests should give a better estimate of the damage caused by the various procedures when developing new techniques for chilling and freeze-thawing. Furthermore, the present study showed that chilling for 4 days tended to reduce the zona-binding capacity of the spermatozoon, and that Equex STM paste had a beneficial effect on the capacity of the frozen-thawed spermatozoon to bind to the zona pellucida.