Canine pulmonary clearance during feto-neonatal transition according to the type of delivery.

The success of immediate adaptation to extrauterine life depends on appropriate lung function, however, elective cesarean section can increase the risk of respiratory distress as a result of reduced pulmonary fluid absorption. This study aimed to evaluate the influence of birth mode on pulmonary clearance and respiratory performance of canine neonates in the transition period. For this purpose, 37 neonates were selected according to the obstetric condition: Vaginal Eutocia (n = 17) and Elective C-section (n = 20). Neonates were evaluated for neonatal vitality score, as well as evaluation of heart and respiratory rates, body temperature and body weight, venous hemogasometric evaluation, blood lactate and glucose, pulse oximetry and radiographic evaluation during the first 24 h of life. Additionally, amniotic fluid electrolyte composition of each puppy was evaluated. There was no influence of the type of delivery on electrolyte composition of canine amniotic fluid and neonatal pulmonary liquid content, analyzed by thoracic X-Rays. On the other hand, elective cesarean section delayed pulmonary adaptation, resulting in hypoxemia and less efficient compensatory response to acid-base imbalance and thermoregulation. In conclusion, elective c-section does not delay pulmonary clearance, whilst alters pulmonary adaptation by less efficient gas exchange and lower oxygenation, hindering the compensatory response to acid-base imbalance during the fetal-neonatal transition in dogs.

Maternal and neonatal cortisol and catecholamines throughout vaginal eutocia and C-section in dogs.

Increased fetal cortisol is associated with catecholamine release, a primary factor for neonatal adaptive responses. However, relationship between canine peripartum cortisol and catecholamine is unknown. We aimed to compare serum adrenaline, noradrenaline and cortisol during peripartum in bitches and neonates of distinct obstetric conditions and to assess amniotic fluid cortisol concentration. Twenty females and maximum of three puppies per litter were allocated into Vaginal Eutocia (10 females; 17 neonates) and Elective C-section (10 females; 20 neonates) groups. Amniotic fluid was collected at delivery for cortisol concentration. Maternal and neonatal blood were collected prepartum, intrapartum, postpartum and 1h postpartum, and at birth, 30 and 60min, 12hs and 24hs, respectively, for cortisol, adrenaline and noradrenaline assessment. C-section determined higher noradrenaline throughout delivery and cortisol concentration from intrapartum through 1h postpartum, compared to vaginal birth. C-section maternal cortisol showed progressive increase from intrapartum onwards, while neonatal cortisol remained unchanged. No difference of maternal cortisol concentration occurred along whelping, whereas a significant decrease was verified for vaginal delivery puppies from birth until the 12hs. Puppies delivered vaginally had higher cortisol concentration at birth and 30min, compared to c-section puppies. There was a higher concentration of amniotic fluid cortisol in vaginal eutocia. In conclusion, c-section induces higher maternal stress during and after surgery, whilst vaginal delivery is a more neonatal physiologically stressful condition, contributing to better adaptation during transition.

Frozen-Thawed Sperm Analysis of Benign Prostatic Hyperplasia Dogs Treated With Finasteride.

Benign Prostatic Hyperplasia (BPH) is a pathological condition that directly interferes with the reproductive potential of senile dogs, by leading to prostate enlargement and sperm injury, which in turn may compromise sperm freezeability. Moreover, albeit finasteride treatment reduces prostatic volume and blood supply and maintains seminal quality and testicular integrity, the effects of sperm samples submitted to cryopreservation after the finasteride treatment are still unknown. Thus, the aim of this study was to evaluate frozen-thawed semen of BPH dogs, as well as dogs subjected to BPH pharmacological treatment with finasteride. For such purpose, 20 dogs were previously selected and assigned to three experimental groups, according to BPH diagnosis and treatment with finasteride: Control (n = 9), BPH Group (n = 5) and BPH-Finasteride Group (n = 6). Semen was subjected to one-step cryopreservation protocol with tris-fructose-citric acid extender with 5% glycerol and thawed at 37°C for 30 sec. Fresh and post-thaw sperm samples were evaluated for macroscopic parameters, sperm concentration, sperm motility kinetics, sperm mitochondrial activity and potential, oxidative stress, plasmatic and acrosome membrane integrity, sperm DNA fragmentation and sperm binding test on perivitelic membrane of chicken egg yolk. Regarding fresh semen, BPH-Finasteride group had the lowest ejaculate visual aspect (opacity), higher frequency of sperm flagellar beating (BCF) and percentage of sperm with medium velocity. Control group had the highest percentage of sperm DNA integrity compared to BPH group. For the frozen-thawed semen, Control group presented the highest percentage of spermatozoa with high mitochondrial activity. However, the BPH-Finasteride group showed higher number of sperm bound to the perivitelline membrane of chicken egg yolk compared to the BPH Group. Conversely, BPH group had higher percentage of DNA damage. In conclusion, the ejaculate of BPH dogs has higher susceptibility to cryoinjury, whereas finasteride-treated dogs have increased spermatozoa functional performance, suggesting a promising use of BPH dogs as semen donors in sperm cryopreservation programs.

Prenatal or postnatal corticosteroids favor clinical, respiratory, metabolic outcomes and oxidative balance of preterm lambs corticotherapy for premature neonatal lambs.

Challenges of the extrauterine environment can be life threatening for a premature fetus with inadequate fetal maturity. Maternal corticosteroids therapy is widely employed to induce fetal pulmonary maturation. Nevertheless, whenever therapeutic pregnancy interruption has to be performed in a time manner insufficient to treat the dam, postnatal corticotherapy can be considered an alternative. However, it is not known if antenatal and postnatal corticotherapy can improve similarly neonatal outcomes and pulmonary function. This research aimed to analyze antenatal and postnatal corticotherapy on premature lambs vitality, pulmonary functioning, metabolic and oxidative status. Lambs were evaluated according to the mode of treatment: Prenatal Corticosteroid Group (8 lambs born after maternal betamethasone treatment 48 h prior to birth), Postnatal Corticosteroid Group (9 lambs subjected to betamethasone treatment 10 min after birth) and Control Group (5 lambs remained untreated). Lambs were medically followed-up from birth to 72 h thereafter through a complete physical examination, as well as lactatemia, glycaemia, blood acid-base balance and antioxidant status. Treated lambs had higher vitality score than the Control Group. Heart rate was higher in postnatal therapy compared to prenatal treatment. Respiratory rate and rectal temperature were higher in treated groups. Treated lambs had hyperglycemia, while the Postnatal Group had higher lactatemia than the Control Group. The Prenatal Group had lower and normal pCO2 from 60 min onwards. The Postnatal Group had higher superoxide dismutase activity than untreated lambs. In conclusion, prenatal and postnatal betamethasone treatments favor neonatal clinical outcome, respiratory function, glucose homeostasis and oxidative balance.

Both spontaneous vaginal delivery and elective caesarean section influence neonatal redox status in dogs.

BACKGROUND: The influence of birth mode on redox balance is unknown in dogs. We aimed to compare antioxidant activity and oxidative stress of puppies born by elective c-section or vaginal delivery. METHODS: Twenty-one bitches were elected for spontaneous vaginal delivery (n = 10) and elective cesarean section (n = 11) groups, considering only the first and last puppies born. Neonatal oxygen saturation, lactatemia, superoxide dismutase (SOD), reduced glutathione (GSH), glutathione peroxidase (GPx) and total antioxidant capacity; marker of oxidative stress (thiobarbituric acid reactive substances [TBARS]) and protein oxidation were analysed at birth, 1 h, 12 h, 24 h and 72 h. RESULTS: C-section newborns had lower oximetry at birth and 1 h. Lactatemia was higher in the last vaginal delivery puppies and higher at birth, and 1 h. Regardless of birth mode, last puppies had hyperlactatemia at birth. Vaginal delivery puppies had higher TBARS, while protein oxidation was higher in c-section, mostly in last puppies. C-section puppies had higher GPx activity, whereas GSH concentration decreased overtime. Surgically born puppies had lowering SOD between 12 h and 72 h. CONCLUSIONS: Vaginal delivery enhances lipid peroxidation, counteracted mostly by GPx. Conversely, c-section triggers protein oxidation, acutely scavenged by SOD and GSH, thus puppies are less exposed to the negative consequences of oxidative stress.

A triple stain method in conjunction with an in-depth screening of cryopreservation effects on post-thaw sperm in dogs.

In order to accurately analyze the possible side effects of sperm cryopreservation, an in-depth screening of post-thaw sperm status is necessary. Thus, this study aimed to identify thorough effects of sperm cryopreservation, by evaluating the integrity of all specific structures of the canine spermatozoa. Thirteen (n = 13) mature dogs of different breeds were selected. Six dogs (n = 6) were subjected to sperm cryopreservation, whereas seven dogs (n = 7) were used as semen donors to validate a simultaneous assessment of sperm plasmatic, acrosomal, and mitochondrial membranes (triple stain) by fluorescent probes. Fresh and post-thaw semen samples were evaluated through a computer-assisted analysis of sperm motility, sperm morpho-functional evaluation, triple stain and sperm DNA integrity. Post-thaw semen samples had lower total and progressive motility, as well as higher percentage of minor and major defects. Moreover, post-thaw samples had higher percentage of sperm with plasma membrane and mitochondrial damage but intact acrosome, and also sperm with simultaneous damaged plasma, acrosomal and mitochondrial membranes. Furthermore, post-thaw sperm had higher protamination deficiency and DNA fragmentation. In conclusion, cryopreservation has a broad impact in sperm morphology and function, altering motility patterns, plasma, acrosome and mitochondrial membranes integrity, as well as sperm DNA.

Ageing changes testes and epididymis blood flow without altering biometry and echodensity in dogs.

Senescence leads to deleterious effects in testicular function, sperm quality and fertility in dogs. There, however, are no consistent results of vascular changes in the testes and epididymis during natural ageing in dogs. The aim of this study, therefore, was to compare testes and epididymis blood flow, biometry and echodensity between young and senile dogs. Ten young dogs (1-4 years) and eight senile dogs (over 7 years) were selected and assigned to two experimental groups: Young Group and Senile Group. Dogs were evaluated using testicular and epididymis B-mode (dimensions and echodensity) and Doppler ultrasonography (tissue perfusion parameters and blood flow velocity of the testicular artery). There were no differences between experimental groups for the echographic evaluation of testicular and epididymis parenchyma and biometric variables. The dogs in the Young Group had greater (P = 0.02) testes vascularization score and greater (P = 0.06) testicular artery blood flow velocity than those in the Senile Group. Furthermore, the older dogs had a greater (P = 0.06) pulsatility index of the testicular artery than those in the Young Group. Ageing, therefore, seems to cause natural hemodynamical changes to the testicular artery, resulting in reduced blood flow (ischemia) and tissue damage. Testes and epididymis vascular characteristics, therefore, may represent the causal factors for changes in spermatogenesis and, as a consequence, negatively affect the sperm quality of older dogs. In conclusion, senescence alters testicular artery blood flow and vascularization of the testes, without changing testicular and epididymis ultrasonographic dimensions and echodensity in dogs.

Prostaglandin and antigestagen in pyometra bitches: vascular and stereological effect.

Effects of conservative treatment on uterine blood flow and morphometric findings are still unknown in bitches. Thus, this study aimed to compare uterine changes of pyometra bitches subjected to distinct modes of treatment. Pyometra bitches were assigned to: OHE (ovariohysterectomy immediately after diagnosis), Aglepristone (days 1, 2 and 8) and Associative (aglepristone treatment coupled with cloprostenol for 7 days) groups. After 9 days, bitches were ovariohysterectomized. Before surgery, uterine area was measured ultrasonographically and the uterine artery Doppler velocimetry analyzed blood flow velocity and indexes. Uterine horns were classified according to resistance index (RI) as more compromised and less compromised. Endometrial vasculature was quantitatively evaluated by color flow Doppler. Blood samples were collected to determine nitric oxide (NO) concentrations. Histological uterine structures were quantified by stereology and VEGF-A (vascular endothelial growth factor) and eNOS were (endothelial nitric oxide synthase) immunohistochemically analyzed. Aglepristone and Associative groups had lower uterine area and vascularization, and higher blood flow velocity and indexes compared to OHE group. Less compromised horn of Associative group had higher blood flow velocity compared to OHE group. Aglepristone group presented lower inflammatory infiltrate and larger uterine stroma. Associative group had lower volume density and absolute surface of endometrial cysts and lower VEGF-A expression for glandular epithelium and stromal cells. Blood NO and e-NOS immunostaining were not different among groups. In conclusion, association between aglepristone and prostaglandin is more effective in decreasing uterine vascularization and modulating uterine blood flow. Moreover, associative therapy promotes marked morphological changes. LAY SUMMARY: This research compared two medical protocols of treatment for uterine infection (pyometra) in bitches, using a hormone blocker (anti-progesterone aglepristone) solely or in association with a uterine contraction inducer (prostaglandin; associative therapy). After treatment, bitches were gonadectomized and a microscopic analysis of uterine blood vessel formation and uterine tissue elements were performed as well as uterine blood flow evaluation through Doppler ultrasonography. According to vascular resistance, uterine horns were additionally classified as more compromised and less compromised. Both treatment protocols led to reduction of uterine dimensions and vascularization, and higher blood flow compared to untreated bitches. Less compromised uterine horn of the associative treatment had higher blood flow compared to untreated bitches. The hormone blocker treatment had lower inflammatory cells and larger uterine histological structure, while associative treatment had less uterine pathological cysts and lower blood vessel formation. The associative therapy is effective in decreasing uterine vascularization and modulating uterine blood flow as well as reestablishing endometrium structure in bitches with uterine infection.

Clinical Changes and Uterine Hemodynamic in Pyometra Medically Treated Bitches.

Cystic endometrial hyperplasia-pyometra complex is a canine endometrial disorder, considered the most common and important among uterine diseases. The treatment of choice is ovariohysterectomy, but medical treatments have become an alternative. However, no studies have been performed in order to evaluate uterine hemodynamic changes during medical treatment for pyometra bitches. Therefore, the aim of this study was to compare the effectiveness of two medical protocols by means of clinical (heart rate, respiratory rate and body temperature), medullar response (complete blood count (CBC)) and uterine hemodynamic evaluation in pyometra bitches. For such purpose, 10 bitches with pyometra were assigned to two groups: Aglepristone Group (n = 5) and Associative Group (aglepristone + prostaglandin; n = 5). The Associative Group had altered respiratory rate and decreased body temperature. The Aglepristone Group had decreased plasma protein, however, leukocyte count reduced over time for both therapeutic protocols. Uterine area and vascularization score decreased throughout treatment. There was a significant reduction in uterine perimeter, area and vascularization score. The Associative Group had lower final diastolic velocity and higher systole:diastole ratio (S/D), pulsatility index (PI) and resistance index (RI). In conclusion, both medical treatments are effective in reversing clinical and CBC changes of pyometra, especially aglepristone, however they cannot fully restore local uterine changes in a short-term evaluation. Conversely, associative therapy was more effective in decreasing uterine vascularization and modulating uterine blood flow.

Effect of senescence on morphological, functional and oxidative features of fresh and cryopreserved canine sperm.

The present research aimed to compare the hormonal profile, sperm quality and freezability of young and senile dogs. Dogs were assigned into Young Group (n = 11) and Senile Group (n = 11), additionally divided into Fresh Semen Group and Cryopreserved Semen Group. Males were evaluated for libido score and blood estrogen and testosterone assay. Sperm morphofunctional evaluations were performed based on Computer Assisted Sperm Analysis, morphology, mitochondrial activity, mitochondrial membrane potential, plasma and acrosomal membrane integrity, and DNA fragmentation. Sperm oxidative features were: protein oxidation, lipid peroxidation and production of advanced glycation end-products. Young dogs had higher libido score, sperm velocity average pathway, linearity of motility and mitochondrial activity index and lower percentage of major defects, total defects and proximal cytoplasmic droplet, despite the lack of difference between hormone profile of aged dogs. Fresh semen of senile dogs had increased percentage of spermatozoa with high mitochondrial membrane potential compared to young dogs and to cryopreserved sperm. Cryopreserved semen of young dogs had higher acrosomal membrane integrity compared to the Senile Group. In conclusion, sperm of aged dogs have reduced quality, signaled by higher morphological defects, ultimately altering sperm mitochondrial function and sperm kinetics. Furthermore, spermatozoa from senile dogs are more sensible to cryoinjury.

Haematological and biochemical analysis of healthy neonatal puppies during the immediate foetal-to-neonatal transition.

For the neonatal patient, precocity of diagnosis is crucial for effectiveness of medical approach. However, the newborn has its own physiological peculiarities due to ongoing adaptive mechanism for extrauterine life and deserves special attention in order to underline a specific management or clinical approach. The objective of this work was to verify clinical adaptations and biochemical balance of neonates during immediate period, with special reference to haematological, renal and metabolic functions. Neonatal puppies (n = 51) were physically examined for vitality and rectal temperature at birth, 5 and 60 min post-birth. Blood was collected at birth and 60 min post-birth for analysis of glucose, sodium, potassium, chlorine, blood urea nitrogen (BUN), haematocrit and haemoglobin. Neonatal vitality was lower at birth compared with 5 min and 60 min post-birth. Progressive decline in rectal temperature (36.5 ± 0.3°C, 34.2 ± 0.2°C, 32.3 ± 0.5°C) was observed at birth, 5 min and 60 min post-birth, respectively. Puppies presented slight hyponatremia (140.7 ± 0.5 mmol/L) at birth and hypopotassemia (3.5 ± 0.1 mmol/L) and blood urea nitrogen (13.1 ± 0.7 mg/dl) during the first hour, and high haematocrit (45.1 ± 1.0%) and haemoglobin (15.3 ± 0.3 g/dl) concentration. In conclusion, puppies had rapid evolution of vitality. Marked decrease in rectal temperature occurred at 5 min post-birth. Haematological values of neonates immediately after birth reflected mainly the dam's blood status, not being useful for a blood panel at this time point. The peculiar pattern of BUN, sodium and potassium observed during transition period, suggested that specific reference range should be considered for neonatal puppies.

Relationship between fetal biometric assessment by ultrasonography and neonatal lamb vitality, birth weight and growth.

Ultrasonographic examination of pregnant ewes can enable the identification of perinatal abnormalities and establish prenatal assistance responsible for minimizing morbidity and perinatal mortality. Therefore, we aimed to evaluate the feasibility of a fetal biometric analysis by ultrasonography to predict neonatal vitality and lamb growth during the first month of life. A longitudinal study was conducted with 13 healthy ewes, subjected to ultrasonographic examination every 15 days from 60th day of pregnancy until lambing, evaluating thoracic diameter, abdominal diameter, biparietal diameter, humerus, femur and placentome length. At birth, 22 lambs were assessed through Apgar score at 5 minutes and after 1 hour. Measurement of body weight was also carried out immediately at birth and weekly during 30 days after birth. Thoracic diameter showed a significant increase between 91-105 days and 121-135 days. Conversely, abdominal diameter had a progressive growth until 106-120 days, and then, a steady development was observed. Biparietal diameter showed progressive growth only towards days 91 and 105. For the humerus length, we verified a significant increase between 106-120 days and 121-135 days, remaining unaltered onwards; while femur length continued to grow until lambing. The linear regression analysis between birth weight and biparietal diameter at 60-75 days was high (R2=0.96; P<0.0001; coefficient of variability of 3.3%). In conclusion, ultrasonographic analysis of fetal biparietal diameter at mid-pregnancy can be used as a predictor of lamb weight at birth. Moreover, assessment of femur length at final pregnancy can be employed for fetal and neonatal development estimation.

Calving duration and obstetric assistance influence pulmonary function of Holstein calves during immediate fetal-to-neonatal transition.

Neonatal immediate adaptation to extrauterine life depends mainly on adequate lung function, which is under the influence of previous fetal maturation and obstetrical condition, both acting to stimulate the efficient liquid removal from the pulmonary parenchyma during the immediate transition period. The objective of the present study is to identify chest radiographic changes of neonatal calves born under the influence of different duration of calving and obstetric assistance and correlate with clinical analysis and blood acid-base balance. Experimental groups were determined according to the duration of calving: 2 h (n = 16), 2-4 h (n = 16) and >4 h (n = 12), and additionally by two sub-groups: no-intervention calving (n = 22) and intervention calving (n = 22). Neonatal calves were evaluated for heart and respiratory rate at birth, 5 min, every 10 min until 90 min. Arterial acid-base balance was determined immediately after calving and thoracic radiographs were made at 10 min of life. Lung radiopacity was higher in the 2-4 hr Group compared to the 2 hr Group. When calving duration was greater than 4 hours, a significant respiratory depression was observed. Calving greater than 2 hours slower neonatal pulmonary clearance, 100% and 91.6% of the calves born in the 2-4 hr and >4 hr Groups, respectively, had mild to moderate lung parenchyma opacity. There was a positive correlation between lung radiographic changes and blood TCO2 and negative correlation between pulmonary opacity score and blood PaO2 and SO2. Hence, it is possible to infer that neonatal hypoxia during prolonged calving has an imperative influence on pulmonary fluid absorption in calves. In conclusion, calving greater than 2 hours impacts pulmonary function at birth, leading to altered lung gas exchange, pulmonary clearance, cardiac and respiratory pattern. Conversely, obstetric intervention when calving has duration greater than 4 hours is beneficial for neonatal oxygenation.

Redox, acid-base and clinical analysis of preterm and term neonatal lambs.

During pregnancy, fetal lambs are exposed to low oxygen tension. Thus, an effective antioxidant mechanism is partially developed which sensitizes fetus to oxidative stress. Consequently, term and preterm neonates are susceptible to molecular and cellular injury caused by oxygen species (ROS). This study aimed to evaluate the development of antioxidant enzymes and oxidative profile of preterm (135 days of pregnancy) and term (145 days of pregnancy) neonatal lambs, correlating with clinical analysis. Preterm lambs had significantly (P ≤ 0.05) lower score of vitality (4.00 ± 1.10), bradycardia (99 ± 34 bpm) and bradypnea (13 ± 10 mpm). However, both groups were normothermic and euglycemic. Preterm group had low blood pH (7.07 ± 0.10) and both groups had hypercapnia, more severe in preterm group (85.52 ± 18.65 mmHg). In addition, premature newborns had lower pO2 (10.67 ± 5.65 mmHg) and SO2 (6.17 ± 5.85%) values. No significant difference (P ≥ 0.05) on antioxidant enzymes and oxidative stress were verified among experimental groups, although glutathione peroxidase negatively correlated with Apgar score, heart rate, SO2 and pO2. Our data show that preterm neonates are less adapted to the odds of labor and to overcome the immediate changes of extra-uterine life. Furthermore, we verified an influence of glutathione peroxidase in controlling oxidative stress, which highlights mature enzymatic mechanisms of cell redox, even in premature lambs.

Cystic endometrial hyperplasia-pyometra syndrome in bitches: identification of hemodynamic, inflammatory, and cell proliferation changes.

Cystic endometrial hyperplasia (CEH)-pyometra syndrome is one of the most common diseases of noncastrated female dogs. However, determination of etiological mechanisms and differential diagnosis of CEH-pyometra syndrome are undefined. The aim of this study is to compare immunohistochemically the expression of cyclooxygenase-2 (COX-2) inflammatory mediator, Ki-67 antigen proliferation marker, vascular endothelial growth factor (VEGF-A) angiogenesis mediator and its FLT-1 and KDR receptors, and correlate with Doppler velocimetry of uterine artery and endometrial vascularization in bitches with CEH-pyometra syndrome. Bitches were allocated into CEH-mucometra Group (n = 13), Pyometra Group (n = 11), and Control Group (n = 8). Pyometra Group presented cytoplasmatic staining intensity for COX-2, VEGF-A, and FLT-1 and KDR receptors in luminal epithelium cells significantly higher compared to CEH-mucometra and Control groups. For the glandular epithelium, Pyometra Group had higher immunostaining score for VEGF-A and its receptors (FLT-1 and KDR). Hemodynamic indexes showed negative correlation with VEGF-A and its receptors as well as with COX-2. On the other hand, uterine vascularization score showed positive correlation in relation to immunostaining of COX-2, VEGF-A, and receptors in the endometrium luminal epithelium. In conclusion, uterus of bitches with CEH-pyometra syndrome show inflammatory process characterized by COX-2 expression, resulting in greater expression of proliferative Ki-67 marker as tissue response against the infectious agent. Furthermore, the increased VEGF-A expression and its receptors in CEH-pyometra reflect the increased blood flow and lower vascular resistance. Therefore, canine pyometra is characterized by an inflammatory, proliferative, and vascular disorder.

Sperm-binding to the perivitelline membrane of chicken egg yolk as a functional test for sperm evaluation in dogs / Teste de ligação espermática à membrana perivitelínica da gema de ovo de galinha como avaliação funcional do sêmen de cães

During fertilization, spermatozoa interact with the zona pellucida (ZP) through the binding between the acrosome and proteins 2 and 3 (ZP2 and ZP3). The perivitelline membrane of chicken egg yolk is homologous to the mammalian ZP3, which allows the binding of sperm of several species. The aim of this study was to standardize and evaluate the efficiency of sperm-binding to the perivitelline membrane of chicken eggs as a functional method for canine semen evaluation. For this purpose, nine post-thaw sperm samples were used, which were divided into two aliquots: the first was kept in water bath at 37ºC (live sample) and the second was submitted to cold shock to induce cellular damage (dead sample). The two aliquots were mixed on five proportions, corresponding to 0%, 25%, 50%, 75%, and 100% of viable cells, and the binding test was performed by analyzing the number of spermatozoa bonded to the perivitelline membrane by means of computerized assessment of sperm motility (CASA) or conventional microscopy. Additionally, samples were submitted to sperm motility analysis, evaluation of plasmatic and acrosomal membrane integrity, and sperm mitochondrial activity. The sperm-binding test to the perivitelline membrane of chicken egg yolk was considered a feasible sperm analysis test for both fertilizing capacity and overall sperm attributes evaluation, mainly when the analysis is performed by a conventional microscope, which expands its practicality to the majority of canine reproduction laboratories.(AU)

Durante a fecundação, os espermatozoides interagem com a zona pelúcida (ZP) por meio da ligação entre o acrossomo e as proteínas 2 e 3 (ZP2 e ZP3). A membrana perivitelínica da gema de ovo de galinhas é homóloga à ZP3 de mamíferos, possibilitando a ligação espermática de diversas espécies. Este trabalho padronizou e avaliou a eficiência do teste de ligação espermática à membrana perivitelínica da gema de ovo de galinhas como avaliação funcional do sêmen de cães. Para tal, foram utilizadas nove amostras seminais previamente criopreservadas. Cada amostra foi dividida em duas alíquotas: a primeira foi mantida em banho-maria à 37ºC (vivos) e a segunda submetida a choque térmico com o intuito de induzir dano celular (mortos). As duas alíquotas foram misturadas, correspondendo a 0, 25, 50, 75 e 100% de células viáveis. As amostras foram avaliadas quanto ao número de espermatozoides ligados à membrana perivitelínica por meio da análise computadorizada da motilidade (CASA) ou microscopia convencional. Ademais, as amostras foram avaliadas quanto à motilidade espermática, integridade das membranas acrossomal e plasmática e atividade mitocondrial espermática. O teste de ligação espermática à membrana perivitelínica de ovos de galinha foi considerado um teste de análise seminal exequível tanto para avaliar a capacidade fecundante dos espermatozoides como atributos seminais gerais, especialmente quando realizado em microscopia convencional, expandindo sua praticidade para a maioria dos laboratórios de análise de sêmen canino.(AU)

The influence of the obstetrical condition on canine neonatal pulmonary functional competence.

OBJECTIVE: To compare the influence of the obstetrical condition on neonatal respiratory performance, to estimate surfactant synthesis through lecithin and sphingomyelin ratio (L/S) in amniotic fluid of pregnant bitches and correlate the L/S with the respiratory condition during the first hour of life. DESIGN: Prospective randomized study. SETTING: University veterinary teaching hospital and private referral centers. ANIMALS: In accordance to the condition at birth, puppies from 25 healthy bitches aged 2-6 years were allocated into: Eutocia Group--EUT (n = 19 neonates and 14 bitches); Dystocia Group--DYS (n = 8 neonates and 5 bitches) and Elective Cesarean Section Group--CS (n = 12 neonates and 11 bitches). INTERVENTIONS: Amniotic fluid was drawn from amniotic sac and lecithin (L) and sphingomyelin (S) were measured by high performance liquid chromatography to obtain the L/S ratio. Neonatal physical examination was performed at 1, 5, and 60 minutes after delivery, and included the assessment of respiratory rate (RR) and respiration effort (RE). MEASUREMENTS AND MAIN RESULTS: CS group had significantly lower amniotic L/S ratio when compared to EUT and DYS. There were no significant differences between vaginal groups (EUT versus DYS) in respect to L/S ratios. RE of eutocic neonates improved promptly, while RE of DYS and CS groups improved only 1 hour following birth. Moreover, amniotic L/S ratio positively correlated with RR after 60 minutes. CONCLUSIONS: We identified a possible influence of the delivery method on the final surfactant maturation process. Thus, neonates born by elective CS prior to the onset of expulsive uterine contractions should have their respiratory parameters carefully monitored. Additionally, we propose that assessment of respiratory parameters 1 hour following birth can serve as a practical means to indirectly estimate pulmonary maturation (ie, surfactant synthesis) in puppies.

Estrogen and oxytocin receptors in the canine corpus luteum during pregnancy and parturition / Receptores de estrógeno e ocitocina no corpo lúteo durante a gestação e parto em cadelas

The expression of genes encoding the receptors for estrogen (ERαmRNA) and oxytocin (OTRmRNA) was studied in the corpus luteum during pregnancy and parturition in dogs. Real-time PCR was performed to quantify the levels of ERαmRNA and OTRmRNA in the corpus luteum of bitches during Early (up to 20 days of gestation), Mid (20 to 40 days) and Late Pregnancy (40 to 60 days), and Parturition (first stage of labor). The corpus luteum expressed mRNA for OTR, however ERα mRNA was not detected. There was a reduction of OTR mRNA expression in the corpus luteum from gestational Day 20 onward, which suggests an important role of OTR mRNA in the mechanism of pregnancy recognition in dogs. We concluded that the expression of OTR mRNA in canine corpus luteum vary over time, which support the idea that the sensitivity and response to hormone therapy can vary along the course of pregnancy and labor. Moreover, the canine CL lacks ERα mRNA expression during pregnancy.

A expressão dos genes que codificam os receptores de estrógeno (REα RNAm) e ocitocina (ROT RNAm) foi estudado no corpo lúteo de cadelas durante a gestação e parto. A técnica de PCR em tempo real foi realizada para quantificar a expressão do REα RNAm e ROT RNAm no corpo lúteo de cadelas durante o início (até 20 dias de gestação), meio (20 a 40 dias) e final da gestação (40 a 60 dias), e durante o parto (pródomos do parto). O corpo lúteo apresentou expressão do RNAm para o ROT, entretanto o RNAm para o REα não foi detectado. Houve redução na expressão do ROT RNAm no corpo lúteo a partir de 20 dias da gestação, indicando papel no mecanismo de reconhecimento gestacional em cadelas. Em conclusão, a expressão do ROT RNAm no corpo lúteo de cadelas apresentou variação ao longo do tempo de gestação, sugerindo que a resposta e sensibilidade à terapia hormonal pode variar conforme o momento da gestação e parto. Ademais, o corpo lúteo canino não expressa REα RNAm durante a gestação.

Influenza viruses in adult dogs raised in rural and urban areas in the state of São Paulo, Brazil / Incidência do vírus influenza em cães adultos criados em áreas rural e urbana do estado de São Paulo, Brasil

In 1970, searching for the interspecies transmission of influenza viruses led to the first study on influenza viruses in domestic animals. Birds and mammals, including human beings, are their natural hosts; however, other animals may also play a role in the virus epidemiology. The objective was to investigate the incidence of influenza viruses in adult dogs raised in rural (9, 19.56%) and urban (37, 80.43%) areas in the state of São Paulo, Brazil. Dog serum samples were examined for antibodies to influenza viruses by the hemagglutination inhibition (HI) test using the corresponding antigens from the circulating viruses in Brazil. Dogs from rural areas presented antibodies to influenza A H3N2, and influenza A H7N7 and H3N8. In rural areas, dog sera displayed mean titers as 94.37, 227.88, 168.14, 189.62 HIU/25 µL for subtypes H1N1, H3N2, H7N7, H3N8, respectively. About 84% and 92% of dogs from urban areas exhibited antibodies to human influenza A H1N1 and H3N2, respectively, with statistical difference at p < 0.05 between the mean titers of antibodies to H1N1 and H3N2. About 92% and 100% were positive for H7N7 and H3N8, respectively. In dogs from urban areas, the mean titers of antibodies against influenza A H1N1, H3N2, H7N7 and H3N8, were 213.96, 179.42, 231.76, 231.35 HIU/25 µL respectively. The difference among them was not statistically significant at p > 0.05. In conclusion, these dogs were positive for both human and equine influenza viruses. The present study suggests the first evidence that influenza viruses circulate among dogs in Brazil.

A transmissão interespecífica do vírus influenza é relatada em estudo sobre influenza com animais domésticos desde 1970. Pássaros e mamíferos, incluindo o homem, são seus hospedeiros naturais, porém outros animais podem participar da sua epidemiologia. Foi investigada a incidência do vírus influenza em cães adultos criados em zonas rural (9, 19,56%) e urbana (37, 80,43%), do Estado de São Paulo. Os soros dos cães foram examinados pelo teste de inibição da hemaglutinação (IH), usando antígeno dos vírus influenza circulantes no Brasil. Nos cães rurais foram detectados títulos médios de 94,37, 227,88, 168,14 e 189,62 UIH/25 mL (unidades inibidoras de hemaglutinação/25 mL) para os subtipos H1N1, H3N2, H7N7, H3N8 de vírus influenza A, respectivamente, com diferenças estatisticamente significativas (p<0,05) entre as médias de títulos de anticorpos contra H1N1 e H3N2. Cerca de 84% e 92% dos cães urbanos responderam aos vírus influenza A humano H1N1 e H3N2, respectivamente e destes 92% e 100% foram positivos para os vírus eqüinos H7N7 e H3N8, respectivamente. Para esses cães as médias de títulos de anticorpos para os vírus influenza A H1N1, H3N2, H7N7 e H3N8 foram 213,96, 179,42, 231,76 e 231,35UIH/25 mL, respectivamente. As diferenças entre as médias não foram estatisticamente significativas (p>0,05). Conclui-se que os cães apresentaram positividade para ambos vírus influenza humano e equino. O presente estudo sugere, pela primeira vez, evidências de que há circulação do vírus influenza em cães, no Brasil.

Influenza viruses in adult dogs raised in rural and urban areas in the state of São Paulo, Brazil.

In 1970, searching for the interspecies transmission of influenza viruses led to the first study on influenza viruses in domestic animals. Birds and mammals, including human beings, are their natural hosts; however, other animals may also play a role in the virus epidemiology. The objective was to investigate the incidence of influenza viruses in adult dogs raised in rural (9, 19.56%) and urban (37, 80.43%) areas in the state of São Paulo, Brazil. Dog serum samples were examined for antibodies to influenza viruses by the hemagglutination inhibition (HI) test using the corresponding antigens from the circulating viruses in Brazil. Dogs from rural areas presented antibodies to influenza A H3N2, and influenza A H7N7 and H3N8. In rural areas, dog sera displayed mean titers as 94.37, 227.88, 168.14, 189.62 HIU/25 µL for subtypes H1N1, H3N2, H7N7, H3N8, respectively. About 84% and 92% of dogs from urban areas exhibited antibodies to human influenza A H1N1 and H3N2, respectively, with statistical difference at p < 0.05 between the mean titers of antibodies to H1N1 and H3N2. About 92% and 100% were positive for H7N7 and H3N8, respectively. In dogs from urban areas, the mean titers of antibodies against influenza A H1N1, H3N2, H7N7 and H3N8, were 213.96, 179.42, 231.76, 231.35 HIU/25 µL respectively. The difference among them was not statistically significant at p > 0.05. In conclusion, these dogs were positive for both human and equine influenza viruses. The present study suggests the first evidence that influenza viruses circulate among dogs in Brazil.