ContentsThe apparent lack of classical mechanisms for maternal recognition of pregnancy is one of the most intriguing features of canine reproduction. Consequently, similar levels of circulating luteal steroids are observed in pregnant and non-pregnant dogs.However, the early pre-implantation canine embryo locally modulates uterine responses to its presence, facilitating the successful onset of pregnancy. As a part of this interaction, the canine uterus undergoes a species-specific decidualization. revealed upregulated gene levels of selected major decidualization markers (e.g. PRLR, PGR, IGF1, PTGES). Additionally, the basic decidualization capability of PGE2 was demonstrated, revealing increased levels of, for example, PGR and PRLR gene expression, thereby implying its involvement in the progesterone-dependent decidualization in the canine uterus. In summary, our in vitro model with immortalized DUS cell line could serve as an ideal and unique model to study the underlying molecular and endocrine mechanisms of canine decidualization.

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Hypospadias is rarely reported in dogs. In this study we pre-sent 2 novel cases of this disorder of sexual development and, in addition, a case of hereditary sex reversal in a female with an enlarged clitoris. The first case was a male Moscow watchdog with a normal karyotype (78,XY) and the presence of the SRY gene. In this dog, perineal hypospadias, bilateral inguinal cryptorchidism and testes were observed. The second case, representing the Cocker spaniel breed, had a small penis with a hypospadic orifice of the urethra, bilateral cryptorchidism, testis and a rudimentary gonad inside an ovarian bursa, a normal female karyotype (78,XX) and a lack of the SRY gene. This animal was classified as a compound sex reversal (78,XX, SRY-negative) with the hypospadias syndrome. The third case was a Cocker spaniel female with an enlarged clitoris and internally located ovotestes. Cytogenetic and molecular analyses revealed a normal female karyotype (78,XX) and a lack of the SRY gene, while histology of the gonads showed an ovotesticular structure. This case was classified as a typical hereditary sex reversal syndrome (78,XX, SRY-negative). Molecular studies were focused on coding sequences of the SRY gene (case 1) and 2 candidates for monogenic hypospadias, namely MAMLD1 (mastermind-like domain containing 1) and SRD5A2 (steroid-5-alpha-reductase, alpha polypeptide 2). Sequencing of the entire SRY gene, including 5′- and 3′-flanking regions, did not reveal any mutation. The entire coding sequence of MAMLD1 and SRD5A2 was analyzed in all the intersexes, as well as in 4 phenotypically normal control dogs (3 females and 1 male). In MAMLD1 2 SNPs, including 1 missense substitution in exon 1 (c.128A>G, Asp43Ser), were identified, whereas in SRD5A2 7 polymorphisms, including 1 missense SNP (c.358G>A, Ala120Thr), were found. None of the identified polymorphisms cosegregated with the intersexual phenotype, thus, we cannot confirm that hypospadias may be associated with polymorphism in the coding sequence of the studied genes.

Most donkey and local horse breeds are vulnerable to extinction as mechanization of agriculture progress throughout the world. The present study analyzed the pedigree and herd records of the donkey Asinina de Miranda breed (RAM), identifying genealogical and human factors that may affect the breed genetic diversity in the future and suggesting suitable strategies to breed preservation, early on the conservation program. The breeding rate was very low, with a ratio of foaling/live animals of 0.23 (178/760). The estimated number of founders and ancestors contributing to the reference population was 128 and 121. The number of founder herds in the reference population was 64, with an effective number of founder herds for the reference population of 7.6. The mean age of herd owners was 65.50 ± 0.884 years, with a negative association among the herd size and owner's age ( P < 0.001). In contrast, the size of the herd and the ownership of a male were both positively associated ( P < 0.001) with the herd number of in-born foals. Both the owners' age and the herd location (RAM home region v. dispersal region) were negatively associated with the foaling number ( P < 0.001). The main identified risk factors were: low breeding rates; low number of males and their unequal contribution to the genetic pool; unequal contribution of the herds to genetic pool; and advanced age of herd owners.

Uterine neoplasms of epithelial origin are rare in cats and most often are described in older females. Yet, in less than 2 years, four ovariohysterectomy specimens were submitted from different practices to the Laboratory of Histology and Anatomical Pathology, at UTAD (Vila Real, Portugal), that emitted a diagnosis of feline endometrial adenocarcinoma. Untypically, all the females were aged <1 year old at the surgery. Access to the clinical files was requested to document the clinical features of the four cases, including any complementary data available, to construct the present case reports. The clinical situation developed with discrete signs, but vulvar discharge was present in three cases, ranging from bloody to brownish or colourless, and from purulent to mucous. The females were in dioestrus, although the oestrus remained unperceived in most cases. In this study, the four clinical situations are described and discussed on the basis of available literature, highlighting the aspects that may impair an early diagnosis and that may favour the progression of the disease and also that age should not be an excluding criteria when analysing the differential diagnosis list.

Although the disorder of sex development in dogs with female karyotype (XX DSD) is quite common, its molecular basis is still unclear. Among mutations underlying XX DSD in mammals are duplication of a long sequence upstream of the SOX9 gene (RevSex) and duplication of the SOX9 gene (also observed in dogs). We performed a comparative analysis of 16 XX DSD and 30 control female dogs, using FISH and MLPA approaches. Our study was focused on a region harboring SOX9 and a region orthologous to the human RevSex (CanRevSex), which was located by in silico analysis downstream of SOX9. Two highly polymorphic copy number variable regions (CNVRs): CNVR1 upstream of SOX9 and CNVR2 encompassing CanRevSex were identified. Although none of the detected copy number variants were specific to either affected or control animals, we observed that the average number of copies in CNVR1 was higher in XX DSD. No copy variation of SOX9 was observed. Our extensive studies have excluded duplication of SOX9 as the common cause of XX DSD in analyzed samples. However, it remains possible that the causative mutation is hidden in highly polymorphic CNVR1.

The current study was conducted to analyse the effect of a mixture of probiotic cultures and enzymes (Probio Enzyme, XVET GmbH, Germany) on the immune response and weight of central lymphoid organs and liver in broilers. A total of 270 male chickens were randomly divided into nine groups, with three replicates of 10 birds each. Treatment groups were fed for either 22 or 42 days with different levels of Probio Enzyme 250, 500, 750 and 1000 g/ton, whereas the control group fed a basal diet without Probio Enzyme. To analyse the effects of dietary supplementation on broilers humoral immune response, the antibodies titres for avian influenza (AI) and Newcastle disease virus (NDV) and to the sheep red blood cells (SRBC) challenge were assayed in birds from each experimental unit, along with the assessment of the weight of the main lymphoid organs and liver. The addition of the Probio Enzyme mixture did not significantly affect the titres of the antibodies against AI and NDV at day 42, despite the wide individual variation observed specially on the antibody titres at day 33. Treatments affected the production of IgG after the second challenge with SRBC (P = 0.003), which was transposed to the correspondent total Ig titres (P = 0.044). Conversely, a lower birds' body weight (BW) was found in the majority of treated groups compared to control (P = 0.031). The spleen was the only lymphoid organ showing differences in the absolute and relative weight (P = 0.003 and P = 0.001, respectively). No differences were found in thymus and Bursa of Fabricius weights. In conclusion, broilers treated with Probio Enzyme showed a satisfactory immune response compared with control, despite the wide variation found after the first vaccine challenge against AI. Moreover, the probiotic mixture dose and duration modulated differently the immune response and the spleen weight, unaffecting the central lymphoid organs weight.