Manipulation of the oestrous cycle of the bitch-what works for now.

Manipulation of the oestrous cycle of the bitch is a common request of pet owners for different reasons. On one side, clients may be interested in temporally preventing oestrus until the bitch has finished the performance career, or to manage the kennel in the presence of a stud dog and to avoid undesirable breeding. Additionally, clients with no interest to use their animal for breeding, may, for ethical reasons, prefer to avoid elective surgery such as ovariectomy, to control reproduction. On the other side, clients may want to induce oestrus to better control the time of birth, to use a male with limited availability during a specific time, or to reduce the duration of the interoestrous interval in bitches with a long oestrous cycle (>8 months). Both these options for oestrous manipulation are based on hormonal treatments. For oestrous prevention, synthetic progesterone has been used since the 1960s. Induction of oestrus has been established with reliable results using dopamine agonists in the late 1980s and more recently using GnRH agonists. This review is focusing on the current and recently available treatments to manipulate the oestrous cycle in the bitch.

A fluorescent quantum dot conjugate to probe the interaction of Enterolobium contortisiliquum trypsin inhibitor with cancer cells.

Serine proteases play crucial biological roles and have their activity controlled by inhibitors, such as the EcTI, a serine protease inhibitor purified from Enterolobium contortisiliquum seeds, which has anticancer activity. This study aimed to conjugate EcTI with quantum dots (QDs), fluorophores with outstanding optical properties, and investigate the interaction of QDs-EcTI nanoprobe with cancer cells. The conjugation was evaluated by fluorescence correlation spectroscopy (FCS) and fluorescence microplate assay (FMA). EcTI inhibitory activity after interaction with QDs was also analyzed. From FCS, the conjugate presented a hydrodynamic diameter about 4× greater than bare QDs, suggesting a successful conjugation. This was supported by FMA, which showed a relative fluorescence intensity of ca. 3815% for the nanosystem, concerning bare QDs or EcTI alone. The EcTI inhibitory activity remained intact after its interaction with QDs. From flow cytometry analyses, approximately 62% of MDA-MB-231 and 90% of HeLa cells were labeled with the QD-EcTI conjugate, suggesting that their membranes have different protease levels to which EcTI exhibits an affinity. Concluding, the QD-EcTI represents a valuable nanotool to study the interaction of this inhibitor with cancer cells using fluorescence-based techniques with the potential to unravel the intricate dynamics of interplays between proteases and inhibitors in cancer biology.

The View of the French Dog Breeders in Relation to Female Reproduction, Maternal Care and Stress during the Peripartum Period.

In France, as in many other western countries, dogs are an important part of the society as pets or working animals. The exact demand for puppies in France is unknown, as is the proportion of dogs coming from different breeding sources. Nevertheless, the origin of puppies is important since young dogs from puppy mills and pet stores appear to be more likely to develop behavioral disorders. Puppies coming from responsible breeders, on the other hand, tend to be better adapted. In well-managed kennels, an explanation for these behavioral differences may be associated with greater contact of litters with the dam and humans. Another factor that might influence a dog's temperament and character is maternal behavior. Although recent studies have shown that the quality of maternal care in dogs is important, direct effects on the development of behavioral problems such as fearfulness or noise sensitivity are still controversial. To better understand the view of breeders, due to an increased interest in maternal care of dogs, an online questionnaire was developed to assess the dog breeders' profiles and to explore their perception of normal maternal and stress-related behaviors during the peripartum period. In addition, the management of the female during this critical time was queried. Three-hundred and forty-five French dog breeders, representing 91 breeds, completed the online survey. Considering the demographics of the responders, breeding activity in France is mostly family-based with 76% raising two breeds of dogs that produce, on average, five litters/year. Around 60% of the breeders use progesterone levels to determine breeding date. The whelping date is estimated using multiple criteria and most labors and deliveries happen under human supervision. The majority of behaviors associated to good maternal care are defined by the vast majority as being related to more attention of the bitch towards the puppies with the frequency of nursing and licking being important clues. Globally, the peripartum is perceived as a stressful period and to minimize stress and reassure the bitch the favored method used is increasing human presence. Problems related to maternal behavior were described, notably with primiparous bitches.

Influence of ADAPTIL® during the Weaning Period: A Double-Blinded Randomised Clinical Trial.

ADAPTIL®, a dog-appeasing pheromone, was shown to modify the dam-puppies' interactions during the neonatal period but could also influence the weaning period. Fourteen bitch/litter dyads continuously exposed to ADAPTIL® from the third/fourth weeks until the seventh/eighth weeks postpartum were compared to 11 dyads exposed to a placebo. Maternal and puppy behaviours were video-recorded, and at three time points (weeks three/four, weeks five/six and weeks seven/eight) after parturition. The well-being of the puppies and the overall relationship with the bitch were assessed using visual analogue scales (VAS) completed by the caregivers. All mothering behaviours, such as time of contact, licking and the amount of time dedicated to nursing puppies, decreased gradually from weeks three/four to weeks seven/eight. A switch in nursing position was observed over time: the use of the standing position increased compared to the lying position. The treatment had an effect on the nursing position: bitches in the ADAPTIL® group nursed more often in a lying (p = 0.007) or sitting position (p = 0.037), whereas for the placebo group, they favoured the standing position (p = 0.011). Once the puppies became more demanding for suckling, the bitches started showing rejection signs or aggressive growling, with a peak at weeks seven/eight. The pheromone seemed to reduce the intensity of avoidance in bitches exposed to ADAPTIL® at all time points. The score of all events combined as a sign of frustration showed a difference over the full period (p = 0.003), with the placebo group having a significantly higher score. From the caregiver perspective (through the VAS), under ADAPTIL®, the bitches were calmer when puppies tried to suckle (p = 0.001), more tolerant towards pups (p = 0.025), showed a greater motherly attitude (p = 0.016), the puppies cried less when left alone (p < 0.001) and interactions amongst pups were more harmonious (p = 0.055). Under ADAPTIL®, the bitches were less annoyed by the puppies, who seemed to cope better with frustration. The breeders perceived a benefit of the pheromone during the weaning period.

Influence of dog-appeasing pheromone on canine maternal behaviour during the peripartum and neonatal periods.

BACKGROUND: Parturition and the initial postpartum period are important moments in the reproductive cycle of dogs. METHODS: A study assessed the effect of ADAPTIL, a dog-appeasing pheromone, on maternal behaviour during peripartum. Bitches were continuously exposed to ADAPTIL (n=20) or placebo (n=21) in double-blinded conditions from an average of about seven days before parturition up to 21 days postpartum. Differences in maternal behaviour in relation to the treatment were evaluated by the observation of specific activities through video recordings, such as the time spent by the bitch in close contact with the puppies, oronasal interaction and nursing duration and position. Videos were recorded at four time points (W0: within the first 48 hours of whelping; W1: one week after parturition; W2: two weeks after parturition; and W3: three weeks after parturition). In addition, the perception of breeders in relation to the quality of maternal care, puppies' wellbeing and overall relationship between the bitches and the puppies was evaluated using Visual Analogue Scale at the same time points. Moreover, the daily activity of the bitches was measured by using an electronic device (FitBark dog activity trackers, Kansas City, Missouri). RESULTS: For all observed maternal behaviours, there was a steady decrease in levels as the puppies developed, independently of treatment. However, bitches exposed to ADAPTIL tended to nurse significantly more in lying position, while those exposed to the placebo nursed more in a seated position, especially at W1 (P=0.06) and W3 (P=0.005). According to the breeders, the attention scores of bitches towards puppies were significantly higher in ADAPTIL than in the placebo group at each time point (P=0.01). Moreover, a difference according to parity was observed (P=0.004), with greater attention score displayed by primiparous bitches exposed to ADAPTIL compared with placebo on W0 (P=0.02), W1 and W3 (P<0.001). The global mother-puppies relationship was also perceived as significantly better (P=0.0002) by breeders of bitches exposed to ADAPTIL, with significant differences at W2 (P=0.01) and W3 (P=0.001). The bitches' daily activity increased starting two days before the whelp, peaked during parturition and then gradually declined up until four days postpartum. There was a trend towards a difference in the activity level according to the treatment during the full study period (P=0.09) and at two days before parturition (P=0.07). Bitches exposed to ADAPTIL were more active compared with placebo in relation to the FitBark data. CONCLUSION: The use of ADAPTIL in maternity modulated maternal behaviours. Concerning the caregiver's view, bitches under the influence of ADAPTIL had greater and extended attention towards the puppies and they were eager to stay with the puppies for a longer time.

Bauhinia monandra leaf lectin (BmoLL) conjugated with quantum dots as fluorescent nanoprobes for biological studies: application to red blood cells.

Carbohydrates perform important physiological functions in eukaryotic and prokaryotic cells. Indeed, alterations in glycan patterns may be associated with disorders. The analysis of these sugars can be reached using nanoprobes composed by lectins associated with fluorescent nanoparticles. This study reports the conjugation of a galactose-binding lectin (BmoLL) isolated from Bauhinia monandra leaves with quantum dots (QDs) by adsorption. QDs-BmoLL conjugates showed bright fluorescence and the hemagglutination assay revealed that the lectin preserved its carbohydrate-binding ability after the conjugation. To evaluate the efficiency/specificity of the bioconjugate, ABO human red blood cells (RBCs) were used as biological models and the labeling was analyzed by flow cytometry. Among ABO blood groups, higher labeling (71.7 ± 5.9%) was detected for B-type RBCs, whose antigens have galactose in their structure. The specificity of labeling was confirmed since A- and O-types RBCs incubated with QDs-BmoLL, as well as B-type cells incubated with previously galactose-inhibited conjugates, were labeled below 6%. In AB-type RBCs, which simultaneously have B and A (N-acetylgalactosamine) antigens on their membrane, the labeling was ca. 14.1 ± 4.8%. Therefore, a successful conjugation was reached and QDs-BmoLL conjugates can be considered promising fluorescent nanoprobes for biological investigations.

Evaluating glucose and mannose profiles in Candida species using quantum dots conjugated with Cramoll lectin as fluorescent nanoprobes.

Glycoconjugates found on cell walls of Candida species are fundamental for their pathogenicity. Laborious techniques have been employed to investigate the sugar composition of these microorganisms. Herein, we prepared a nanotool, based on the fluorescence of quantum dots (QDs) combined with the specificity of Cramoll lectin, to evaluate glucose/mannose profiles on three Candida species. The QDs-Cramoll conjugates presented specificity and bright fluorescence emission. The lectin preserved its biological activity after the conjugation process mediated by adsorption interactions. The labeling of Candida species was analyzed by fluorescence microscopy and quantified by flow cytometry. Morphological analyses of yeasts labeled with QDs-Cramoll conjugates indicated that C. glabrata (2.7 µm) was smaller when compared to C. albicans (4.0 µm) and C. parapsilosis sensu stricto (3.8 µm). Also, C. parapsilosis population was heterogeneous, presenting rod-shaped blastoconidia. More than 90% of cells of the three species were labeled by conjugates. Inhibition and saturation assays indicated that C. parapsilosis had a higher content of exposed glucose/mannose than the other two species. Therefore, QDs-Cramoll conjugates demonstrated to be effective fluorescent nanoprobes for evaluation of glucose/mannose constitution on the cell walls of fungal species frequently involved in candidiasis.

Expression of genes associated with immunity in the endometrium of cattle with disparate postpartum uterine disease and fertility.

BACKGROUND: Contamination of the uterine lumen with bacteria is ubiquitous in cattle after parturition. Some animals develop endometritis and have reduced fertility but others have no uterine disease and readily conceive. The present study tested the hypothesis that postpartum cattle that develop persistent endometritis and infertility are unable to limit the inflammatory response to uterine bacterial infection. METHODS: Endometrial biopsies were collected several times during the postpartum period from animals that were subsequently infertile with persistent endometritis (n = 4) or had no clinical disease and conceived to first insemination (n = 4). Quantitative PCR was used to determine the expression of candidate genes in the endometrial biopsies, including the Toll-like receptor (TLR 1 to 10) family of innate immune receptors, inflammatory mediators and their cognate receptors. Selected proteins were examined by immunohistochemistry. RESULTS: The expression of genes encoding pro-inflammatory mediators such as interleukins (IL1A, IL1B and IL6), and nitric oxide synthase 2 (NOS2) were higher during the first week post partum than subsequently. During the first week post partum, there was higher gene expression in infertile than fertile animals of TLR4, the receptor for bacterial lipopolysaccharide, and the pro-inflammatory cytokines IL1A and IL1B, and their receptor IL1R2. The expression of genes encoding other Toll-like receptors, transforming growth factor beta receptor 1 (TGFBR1) or prostaglandin E2 receptors (PTGER2 and PTGER4) did not differ significantly between the animal groups. Gene expression did not differ significantly between infertile and fertile animals after the first week postpartum. However, there were higher ratios of IL1A or IL1B mRNA to the anti-inflammatory cytokine IL10, during the first week post partum in the infertile than fertile animals, and the protein products of these genes were mainly localised to the epithelium of the endometrium. CONCLUSION: Cattle may maintain fertility by limiting the inflammatory response to postpartum bacterial infection in the endometrium during the first week after parturition.

Dynamics of postpartum endometrial cytology and bacteriology and their relationship to fertility in dairy cows.

Endometrial samples were obtained from 56 consecutively calving dairy cows examined for endometrial cytology and for aerobic and anaerobic bacterial growth. Changes over time, correlations between different cell types and between cell and bacterial populations and with fertility measures were calculated. The proportion of neutrophils in cytologic preparations decreased with time postpartum. Other cell types did not change significantly with time. The proportion of neutrophils early (Day 0 and 7) postpartum was negatively correlated with neutrophil proportion at 5 or 7 weeks postpartum and positively correlated with fertility. Cows with high proportion of neutrophils at 7 days postpartum (>40%) were significantly more likely to become pregnant than those with lower proportions of neutrophils. Escherichia coli were the bacteria most frequently isolated at 0 or 7 days postpartum but were uncommon after that. Trueperella pyogenes were most prevalent at 3 weeks postpartum and were more likely to infect cows that had previously been infected with E coli. The presence of T pyogenes at 3 weeks postpartum increased the risk of concomitant or later infection with gram-negative anaerobes. The presence of T pyogenes at 3 weeks postpartum significantly reduced the risk of pregnancy at 150 days in milk. The presence of alpha-hemolytic Streptoccus spp. at 7 days postpartum was associated with improved reproductive performance. The proportion of neutrophils at 5 and 7 weeks postpartum was related to concomitant bacterial infection. These findings suggest that rapid mobilization of neutrophils to the postpartum uterus is a beneficial response for uterine health in dairy cows.