Biochemical composition of amniotic fluid in normal puppies at term of pregnancy: preliminary data

The full knowledge of the normal fetal fluids composition could be useful in the dog for the better management of newborns. The aim of the present study was to define the biochemical composition of amniotic fluid of puppies born by elective Caesarean section (CS) at term of pregnancy. The study enrolled 24 purebred bitches, classified into small size (<10 kg) and large size breeds (>20kg). All the bitches were healthy and clinically monitored from mating until parturition. For all the bitches an elective CS at term of pregnancy was performed [1]. For each puppy, the amniotic fluid was collected, immediately centrifuged and frozen at – 20° C until analysis for ALB, AMY, TB, CHOL, CK, ALP, GGT, AST, ALT, LDH, Mg, Ca, K, Na, Trig, BUN, Glc, TP, CREA, LIP, Cl, and GLOB. Data were analyzed by ANCOVA to verify the possible effects of parity, breed body size and newborn gender on amniotic biochemical composition. A total of 69 amniotic fluid samples were collected. The amniotic mean±SD and min-max values for each parameter were defined. LDH (p<0.01) and CK activity (p<0.05), as well as Glc concentrations (p<0.0001) were negatively influenced by the parity. AMY activity was significantly (p<0.05) higher in large sized (44.2±20.87 U/L) respect to small sized dogs (30.3±19.89 U/L), while lower (p<0.05) CHOL amniotic concentrations were found in small sized (3.0±2.71 mg/dl) as compared to large sized (3.9±2.93 mg/dl) dogs. Gender of the newborn did not influence the amniotic biochemical composition. The preliminary results of this study showed some similarities as well as some differences concerning the biochemical composition of the amniotic fluid in dogs at term of pregnancy if compared to data reported for the cat [2]. Furthermore, the results suggested that, in dogs, some amniotic parameters could be influenced by breed body size and by parity

Physiological pregnancy in a tiger affected by endometritis and benign mesenchymal neoplasm.

In large felids, uncomplicated endometritis and benign mesenchymal uterine tumours usually represent post-mortem findings. These pathologies generally remain clinically asymptomatic, and their impact on reproductive outcome and maternal care is unknown. A 13-year-old female tiger (Panthera tigris) was evaluated by a team of veterinarians from the University of Padova for abnormal behaviour after parturition. The animal spontaneously gave birth to a healthy male cub of 1.460 kg, but after three hours of normal parental care, she showed careless about the cub, occasional abdominal contractions, excessive self-licking and restlessness. After 10 hours of continuous observation, the tiger was anaesthetised for a complete clinical evaluation. The physical examination was unremarkable, but a vaginal bloody/mucous discharge with no obstruction of the birth canal was noted. Abdominal ultrasound revealed an enlarged left uterine horn with heterogeneous echogenicity. Blood analysis showed moderate neutrophilia and leucocytosis. Differential diagnoses included placental retention, uterine inflammation/infection, neoplasia. The tiger was transferred to the Veterinary Teaching Hospital where the explorative laparotomy and the ovario-hysterectomy were performed. The uterine horns appeared asymmetric and two nodular lesions of 1.5-cm-diameter were observed on the left horn. Histological assessment indicated the presence of a multifocal, subacute, moderate neutrophilic endometritis in both horns associated with diffuse, severe oedema. The nodules were consistent with a mesenchymal benign neoplasia. No relevant complication was observed in the following postoperative period and the cub was successfully hand raised. To the best of our knowledge, no report describes the birth of a healthy cub from a tiger affected by endometritis and neoplasia. Considering the normal maternal care observed in the previous twin-pregnancies in this animal, we cannot exclude that the uterine affection caused her abnormal behaviour in this case. Therefore, careful observation of the mother in the post-partum period is always strongly recommended

Contribution to and reconsideration of knowledge in small animal neonatology and gonadectomy in the dog

Part 1: Physiology of the neonate and indicators for its well-being Veterinary neonatology is a rather young field of research. Important differences between the physiology of the neonate compared to the adult have to be taken into consideration in its clinical evaluation. The first topic of the present PhD thesis describes the current knowledge of neonatal physiology and different methods for evaluation of the newborns’ viability. Methods such as the Apgar scoring system and blood gas analysis are successfully used in veterinary medicine. Birth weight and neonatal growth have been evaluated as parameters to estimate a puppy’s risk for neonatal mortality. Although such a correlation has been proven, the populations used for this prove are composed of a high number of different breeds. To evaluate the usefulness of such an approach a comparison with a breed-specific evaluation has been made (Appendix 1). This comparison has shown important differences and the importance of breed-specific investigations of birth weight and growth of the neonate. Part 2: Current knowledge on beneficial and noxious effects of gonadectomy Elective gonadectomy is an important part of the daily veterinary practice. Although beneficial effects of gonadectomy on the dog have been proven, over the last two decades the number of studies reporting an increased risk for neoplastic and other disorders in the castrated animal increased. These reports, although predominantly retrospective, statistical investigations have caused concern and doubt within practitioners and owners. Easy access to information granted by the internet increased this doubt in the group of owners. Results of these investigations have been cited and described in scientific papers, as well as on non-scientific web-sites. In both, although most frequently in the latter, results of studies have been mentioned without describing or considering the study design, the population and the context in which the results have been presented. The lack of this information lead to premature conclusions which have an important impact on many stakeholders. It is therefore that the second part of the present PhD thesis presents a literature review of beneficial and noxious effects of gonadectomy. Prostatic neoplasia in the dog was used as an example of problematic differences in study design, population composition and interpretation of the results (Appendix 2). This part further presents the motivation for the experimental design of Part 3. 5 Part 3 and 4: Immunohistochemical evaluation of steroid hormone receptor expression in extra-genital tissue in the bitch motivated by reports on negative long-term health effects of gonadectomy Hormone receptor expression has proven to be of importance not only in the research of a pathological pathway, but gave also indications on possible treatment options (e.g. human breast cancer). In veterinary medicine the research on hormone receptor expression is not yet as commonly used as in human medicine. The presence of hormone receptors in a tissue may be considered the basis for a direct effect of a hormone on a tissue. Nevertheless, their absence does not exclude the possibility that a certain hormone may not have an impact following a secondary pathway. Reports on effects of gonadectomy on extra-genital tissue gave reason to investigate the expression of Estrogen and Progesterone receptors. Immunohistochemistry is a viable method to evaluate hormone receptor expression. Samples of 14 bitches between the age of 2.1 to 16.7 years were taken within a maximum of 4 hours postmortem. Ten bitches resulted gonadectomized at the moment of death. Collected tissues include the right cardiac ventricle, the splenic hilum and the popliteal and retromandibular lymph nodes. Samples were formalin- fixed, paraffin-embedded and stained with hematoxylin and eosin for histological evaluation. Expression of progesterone receptor (PR) and estrogen receptor alpha (ERa) was evaluated with anti-human antibodies: monoclonal rabbit PR (Clone 1E2) and monoclonal mouse ERa (Clone EP1) using an automatic immunostainer (Ventana BenchMark GX, Roche Diagnostic). Reactivity with canine tissue was confirmed using dog uterus and ovaries showing strong positive nuclear staining for both ERa and PR. Tissues of 11 dogs were considered physiologic or mildly compromised, whereas 3 dogs presented neoplasia in heart (n=2) and spleen (n=1). IHC showed negative immunostaining within investigated tissues for neither ERa not PR. Prior to drawing final conclusions, additional tests have to be performed to confirm the absence of ERa and PR expression. Despite the preliminary results in our study, we consider further research on hormone receptor expression necessary to understand the possible impact of gonadectomy on extra-genital tissue in the dog

Contribution to and reconsideration of knowledge in small animal neonatology and gonadectomy in the dog

Part 1: Physiology of the neonate and indicators for its well-being Veterinary neonatology is a rather young field of research. Important differences between the physiology of the neonate compared to the adult have to be taken into consideration in its clinical evaluation. The first topic of the present PhD thesis describes the current knowledge of neonatal physiology and different methods for evaluation of the newborns’ viability. Methods such as the Apgar scoring system and blood gas analysis are successfully used in veterinary medicine. Birth weight and neonatal growth have been evaluated as parameters to estimate a puppy’s risk for neonatal mortality. Although such a correlation has been proven, the populations used for this prove are composed of a high number of different breeds. To evaluate the usefulness of such an approach a comparison with a breed-specific evaluation has been made (Appendix 1). This comparison has shown important differences and the importance of breed-specific investigations of birth weight and growth of the neonate. Part 2: Current knowledge on beneficial and noxious effects of gonadectomy Elective gonadectomy is an important part of the daily veterinary practice. Although beneficial effects of gonadectomy on the dog have been proven, over the last two decades the number of studies reporting an increased risk for neoplastic and other disorders in the castrated animal increased. These reports, although predominantly retrospective, statistical investigations have caused concern and doubt within practitioners and owners. Easy access to information granted by the internet increased this doubt in the group of owners. Results of these investigations have been cited and described in scientific papers, as well as on non-scientific web-sites. In both, although most frequently in the latter, results of studies have been mentioned without describing or considering the study design, the population and the context in which the results have been presented. The lack of this information lead to premature conclusions which have an important impact on many stakeholders. It is therefore that the second part of the present PhD thesis presents a literature review of beneficial and noxious effects of gonadectomy. Prostatic neoplasia in the dog was used as an example of problematic differences in study design, population composition and interpretation of the results (Appendix 2). This part further presents the motivation for the experimental design of Part 3. 5 Part 3 and 4: Immunohistochemical evaluation of steroid hormone receptor expression in extra-genital tissue in the bitch motivated by reports on negative long-term health effects of gonadectomy Hormone receptor expression has proven to be of importance not only in the research of a pathological pathway, but gave also indications on possible treatment options (e.g. human breast cancer). In veterinary medicine the research on hormone receptor expression is not yet as commonly used as in human medicine. The presence of hormone receptors in a tissue may be considered the basis for a direct effect of a hormone on a tissue. Nevertheless, their absence does not exclude the possibility that a certain hormone may not have an impact following a secondary pathway. Reports on effects of gonadectomy on extra-genital tissue gave reason to investigate the expression of Estrogen and Progesterone receptors. Immunohistochemistry is a viable method to evaluate hormone receptor expression. Samples of 14 bitches between the age of 2.1 to 16.7 years were taken within a maximum of 4 hours postmortem. Ten bitches resulted gonadectomized at the moment of death. Collected tissues include the right cardiac ventricle, the splenic hilum and the popliteal and retromandibular lymph nodes. Samples were formalin- fixed, paraffin-embedded and stained with hematoxylin and eosin for histological evaluation. Expression of progesterone receptor (PR) and estrogen receptor alpha (ERa) was evaluated with anti-human antibodies: monoclonal rabbit PR (Clone 1E2) and monoclonal mouse ERa (Clone EP1) using an automatic immunostainer (Ventana BenchMark GX, Roche Diagnostic). Reactivity with canine tissue was confirmed using dog uterus and ovaries showing strong positive nuclear staining for both ERa and PR. Tissues of 11 dogs were considered physiologic or mildly compromised, whereas 3 dogs presented neoplasia in heart (n=2) and spleen (n=1). IHC showed negative immunostaining within investigated tissues for neither ERa not PR. Prior to drawing final conclusions, additional tests have to be performed to confirm the absence of ERa and PR expression. Despite the preliminary results in our study, we consider further research on hormone receptor expression necessary to understand the possible impact of gonadectomy on extra-genital tissue in the dog

Prevalence of cesarean sections in swiss Bernese Mountain Dogs (2001–2020) and identification of risk factors

Abstract Background Dystocia is an important limiting factor in animal breeding due to its cost, stress for the mother and risk of death for the neonates. Assessment of incidence and characteristics of dystocia and the inherent risk of Cesarean section are of major importance. The aim of the present study was to evaluate the reproductive performance of Bernese Mountain Dogs in Switzerland, with a particular focus on the prevalence of Cesarean sections due to dystocia, and identification of possible risk factors. Results The investigated population included 401 bitches, 207 sires, and 1127 litters. Litter size was significantly influenced by age and parity of the dam. Incidence of Cesarean section was 30.4%, with 2.0% of procedures being elective. History of previous Cesarean section, age of the dam, and a small litter size significantly influenced the risk for Cesarean section. The stillbirth rate was 12.0%, and the number of stillborn pups was significantly higher for litters delivered by Cesarean sections after birth of the first pup. The inbreeding coefficient had a low to non-significant impact on all reproductive parameters (e.g., litter size, number of stillborn pups). Conclusion The sample of Bernese Mountain Dogs of our study had an increased prevalence of Cesarean sections compared to the literature, and advanced age of the dam, litter size and prior Cesarean sections in the dam’s reproductive history was identified as significantly influencing factors. In order to improve pups’ survival rate, elective Cesarean section may be indicated in bitches that have had a previous Cesarean-section/s, are of advanced age, and/or have a small litter

Incidence and concomitant factors of cesarean sections in the bitch: A questionnaire study

Dystocia in the canine species is a common problem, and elective cesarean sections (C-sections) have become more frequent in breeds that are at risk. The aim of this study was to evaluate the incidence of C-section and contributing factors and to compare data on elective and emergency C-sections (e.g., regarding stillbirth). Using a questionnaire, a total of 423 bitches of 80 breeds and their 899 litters were included. The mean number of litters per bitch was 2.1 ± 1.1 litters. The overall rate of stillbirth was 6.7%. Of all litters, 194 were born via C-sections (21.6%), of which 35 were declared as elective and 159 as emergency due to dystocia. Significantly more C-sections were performed in either small litters (1-2 pups) or large litters (>12 pups) (p < 0.001). Bitches that have had prior C-sections had a 4-fold increase in the risk of successive C-sections (RR = 4.54 (95%CI 2.56-7.70; p < 0.001). Furthermore, primiparous bitches of advanced age had a significantly higher incidence of emergency C-sections (p = 0.004). Stillbirth was significantly higher in emergency C-sections compared with that in elective C-sections (p = 0.003). Also, timing of intervention had a significant impact on stillbirth in emergency C-sections (p = 0.025). Within a breed-specific evaluation, significant differences were observed between breeds regarding incidence of C-section and stillbirth. Lesser-known breeds were represented in the population, and the results showed that the Norwich Terrier had the highest (51.6%) and the Gordon Setter had the lowest (4.8%) incidence of C-section (p < 0.001). The inclusion and evaluation of lesser-known breeds regarding incidence of C-section is of importance as it shows that certain breeds without phenotypical traits such as brachycephaly may also have an increased incidence of emergency C-section and stillbirth. We further conclude that more importance may be given to the age at first parturition concerning the occurrence of dystocia and the decision making regarding possible elective C-sections

Bodyweight at Birth and Growth Rate during the Neonatal Period in Three Canine Breeds

Weight at birth (bBW) and early weight gain have been linked to the risk of neonatal mortality. Pups are described to be of low bBW if weighing less than one standard deviation (SD) below the mean. Most studies classified breeds according to their expected adult bodyweight. Our aim was to evaluate the breed specificity of these parameters. We assessed the bBW of 213 puppies of Bernese Mountain Dog (BMD), Tibetan Terrier (TT), and Lhasa Apso (LA) breeds, as well as the neonatal growth rate of 133 puppies of BMD and TT. BMD puppies were born relatively smaller than puppies of TT and LA (p ≤ 0.0001) and gained less weight than TT puppies during the first 14 days (p ≤ 0.05). Litter size had a significant impact on bBW and daily gain until the onset of the third week for BMD (p < 0.0001 and p = 0.0005, respectively) and TT (p = 0.0003 and p = 0.0064, respectively). When using bBW means and SD specifically assessed according to breed, 29 out of the 213 neonates of our study were judged as being of low bBW, whereas, when using the classical criteria (based on breed groups), the number of low bBW pups was 160 of 213. These results suggest that evaluations of bBW and neonatal growth should be performed in a breed-specific manner

Prospective clinical study to evaluate an oscillometric blood pressure monitor in pet rabbits

Abstract Background Rabbits are particularly sensitive to develop hypotension during sedation or anaesthesia. Values of systolic or mean non-invasive arterial blood pressure below 80 or 60 mmHg respectively are common under anaesthesia despite an ongoing surgery. A reliable method of monitoring arterial blood pressure is extremely important, although invasive technique is not always possible due to the anatomy and dimension of the artery. The aim of this study was to evaluate the agreement between a new oscillometric device for non-invasive arterial blood pressure measurement and the invasive method. Moreover the trending ability of the device, ability to identify changes in the same direction with the invasive methods, was evaluated as well as the sensibility of the device in identifying hypotension arbitrarily defined as invasive arterial blood pressure below 80 or 60 mmHg. Results Bland-Altman analysis for repeated measurements showed a poor agreement between the two methods; the oscillometric device overestimated the invasive arterial blood pressure, particularly at high arterial pressure values. The same analysis repeated considering oscillometric measurement that match invasive mean pressure lower or equal to 60 mmHg showed a decrease in biases and limits of agreement between methods. The trending ability of the device, evaluated with both the 4-quadrant plot and the polar plot was poor. Concordance rate of mean arterial blood pressure was higher than systolic and diastolic pressure although inferior to 90%. The sensibility of the device in detecting hypotension defined as systolic or mean invasive arterial blood pressure lower than 80 or 60 mmHg was superior for mean oscillometric pressure rather than systolic. A sensitivity of 92% was achieved with an oscillometric measurement for mean pressure below 65 mmHg instead of 60 mmHg. Non-invasive systolic blood pressure is less sensitive as indicator of hypotension regardless of the cutoff limit considered. Conclusions Although mean invasive arterial blood pressure is overestimated by the device, the sensitivity of this non-invasive oscillometric monitor in detecting invasive mean pressure below 60 mmHg is acceptable but a cutoff value of 65 mmHg needs to be used