Association of MTNR1A and GDF9 gene allelles with the reproductive performance, response to oestrus induction treatments and prolificacy, in improved and non-improved local indigenous sheep breeds.

Sheep farming plays a crucial role for Mediterranean countries, wherefrom a plethora of dairy products is produced. Yet, numerous indigenous sheep breeds in temperate latitudes such as the Mediterranean present a serious drawback that milk production is impaired by the seasonality of reproduction. Towards the efforts for reduction of this phenomenon, the purpose of this study was to evaluate and compare different oestrus synchronization treatments, as well as to associate two genes alleles', namely the MTNR1A and the GDF9, with the response to those treatments. Three indigenous breeds were investigated (Florina, Chios and Karagouniko sheep) and inside each breed three different oestrus synchronization treatments were applied (A: intravaginal sponges, B: GNRH use and C: male effect). In group A, Florina ewes expressed oestrus at 90% in July and fecundity was 85%. Karagouniko and Chios ewes exhibited an oestrus expression of 100% with fecundity rates at 95% and 99%, respectively. In group B, Florina ewes expressed oestrus at 60% with fecundity at 57%, Karagouniko ewes expressed oestrus at 65% with fecundity at 54%, whereas Chios breed animals expressed oestrus at 87% with fecundity rate at 85%. In group C, 68% of the Florina breed expressed oestrus 20-25 days post ram induction, whereas this proportion was 84% and 94%, for Karagouniko and Chios breed, respectively. For the molecular analysis, partial segments of the two genes were sequenced and analysed, whereas alleles were scored based on the detected SNPs. All frequencies of the four detected SNPs in MTNR1A gene were statistically and significantly different in ewes that expressed oestrus in comparison with ewes that did not express oestrus in Florina and Karagouniko breeds concerning all treatments. Two SNP's were detected in GDF9 gene, G1 and FecG , from which, only the FecG mutation exhibited statistically significant difference in twins and triplets than in singles in Florina and Karagouniko breeds.

Origin, demographics, inbreeding, phylogenetics, and phenogenetics of Karamaniko breed, a major common ancestor of the autochthonous Greek sheep.

Greece has a long history in autochthonous sheep, the genetic ancestry of which has been associated with four subtypes known to inhabit Greece at the end of the nineteenth century. Among them, the Karamaniko breed is still surviving, however endangered. This study was designed in order to (a) determine the phylogenetic status, (b) to evaluate the levels of inbreeding, and (c) to assess the genetic basis of coat color of Karamaniko breed. For these purposes, the mitochondrial cyt b gene was sequenced, the AFLP methodology was applied, and the MC1R was genotyped, respectively, in 72 female sheep from the Karamaniko breed. Four different novel cyt b haplotypes were defined and three MC1R genotypes were scored, whereas inbreeding levels estimated using AFLPs by the means of relatedness coefficient (r) were 0.287, with gene diversity at the levels of 0.105. Phylogenetic analysis indicated an eastern Asian tropical and subtropical origin of the Karamaniko breed, close with breeds originating from central Turkey, or a clustering within western European or Mediterranean sheep, mirroring a recent genetic divergence, with a non-random spread towards the formation of lowland breeds. The MC1R genotypes were all associated with the white coat color, in which selective breeding has probably been based on traditional morphological characters. Finally, levels of inbreeding do not constitute an indication for a particular mating plan to prevent unpleasant phenomena such as inbreeding depression, probably because of the special attention paid by the farmers towards the avoidance of relative recurrent mating.

High Frequencies of TNC and COL5A1 Genotypes Associated With Low Risk for Superficial Digital Flexor Tendinopathy in Greek Indigenous Horse Breeds Compared With Warmblood Horses.

Tendinopathies constitute a very common category of musculoskeletal disorders, causing economic losses in the equine industry and animal welfare issues in horse populations. Sport and racehorses are in general sensitive to tendinopathies, whereas local indigenous horse populations are often more tolerant to various disorders. Particularly, indigenous Greek horse breeds have evolved and adapted in the rough topographic features of mountainous and semimountainous Greek terrain and are less prone to develop tendinopathies. Susceptibility to tendinopathy has been proposed to be associated with three specific variants in the tenascin-C and collagen type 5 α-1 genes. The present study was designed to analyze these genes and estimate the frequency of the aforementioned variants in three indigenous Greek horse populations, namely the pinia (peneia) breed, the Messara breed, and the nondescript Macedonian pacer, in comparison with a population of warmbloods. Results revealed high frequencies of genotypes with high risk for tendinopathy in the warmblood population and high frequency of low risk genotypes in the Greek breeds. Apart from the three variants, two novel polymorphisms were detected, one of which may also be associated with risk for tendinopathy. Our findings (a) are in agreement with previous ones demonstrating that risk for tendinopathies is associated with particular gene variants and also (b) highlight the value of Greek indigenous horse breeds as a genetic resource that (c) may be used in selective breeding schemes for decreasing the risk for tendinopathies in genetically improved breeds.

Polymorphism of the melatonin receptor 1A (MNTR1A) gene and association with seasonality of reproductive activity in a local Greek sheep breed.

BACKGROUND: Sheep's reproductive physiology in temperate latitudes (such as Greece), is characterized by seasonality and is also regulated by photoperiodic exposure. Melatonin is the key hormone involved in this regulation. However, the melatonin secretion and therefore the ewes reproductive activity underlies variation, proposed to be linked with the melatonin receptor subtype 1A (MNTR1A) gene structure. This study was designed to investigate the polymorphism of the MNTR1A gene in a local Greek sheep breed and to determine its potential association with reproductive seasonality. RESULTS: Two groups of farmed ewes, each consisted of 30 individuals, were chosen. Males were introduced in both groups in spring (April). The first group consisted of ewes that showed reproductive activity in spring (May), while the second of ewes that showed reproductive activity 3 months later, in summer. The PCR-RFLP methodology was carried out on a 824-bp DNA fragment of the MTNR1A exon 2 using the RsaI restriction endonuclease. The electrophoretic procedure revealed three genotypes, C/C, C/T and T/T. Specifically, 44 animals showed the C/C genotype (28 from the first group and 16 from the second), 14 the C/T genotype (2 from the first and 12 from the second) and 2 animals had the T/T genotype (both from the second group). CONCLUSIONS: Statistical analysis indicated a positive correlation between genotype and reproductive seasonality, with C/C genotype playing a crucial role in out-of-season reproduction activity.

Study of Toll-like receptor and B-defensins genes expression pattern in porcine reproductive organs.

Toll-like receptors (TLRs) and b-defensins (BD) molecules are group of molecules that recognize various microbial components and play a crucial role in the activation of the innate immune system in vertebrate species. Although TLRs gene expression has been studied in various pig tissues, little is known about their expression in porcine reproductive tract. Concerning b-defensins genes, only BD1, 2 and 3 counterparts have been well studied in pigs' reproductive organs. The aim of this study was to investigate the expression pattern of both gene families in pigs' male and female reproductive organs, and embryos, as potential tool for further association studies in respect to immunity and disease resistance. RT-PCR analysis revealed that all of the examined TLR genes were expressed in the reproductive organs of male and female pigs, with TLR3 and TLR5 showing the higher levels and TLR9 the lowest, in all analyzed tissues. BD genes showed a different expression pattern in respect to the examined tissue. In embryos, TLR1 revealed high expression levels, while only BD3, BD108, and BD123 were found to be expressed.

Temporal changes in the expression of avian ß-defensins in the chicken vagina during sexual maturation and Salmonella infection.

Avian ß-defensins (AvßDs) constitute a family of antimicrobial peptides that are critical to innate immunity in chickens, providing protection against microbial pathogens including Salmonella Enteritidis (SE). As apart from the digestive tract another main route of SE colonization in birds is via infection of the oviduct and specifically of the vagina, the aim of this study was to investigate the expression of the complete family of AvßDs, in the chicken vagina in vivo, to determine whether sexual maturation affects their mRNA abundance and to investigate whether SE infection alters the vaginal AvßDs expression. Expression analysis revealed that 11 members of the AvßD family were expressed in the chicken vagina. Quantitative real-time PCR analysis revealed that the mRNA abundance of five AvßDs was up regulated and of one AvßD was down regulated with respect to sexual maturation. In addition SE infection resulted in a significant induction of AvßD5, 7, 10, 11, 12 and 14 in the vagina of sexually mature birds, and in a significant induction of AvßD5 and 11 in the vagina of aged birds. These findings provide strong evidence to suggest that an AvßD-mediated immune response mechanism exists in the chicken vagina providing protection against bacterial pathogens including Salmonella species.

Expression of chicken zygote arrest 1 (Zar1) and Zar1-like genes during sexual maturation and embryogenesis.

Maternal mRNAs, which are expressed in oocytes, play an important role in the success of early embryo development, as they allow the first cleavages to occur. Zygote arrest 1 (Zar1) is an oocyte-specific maternal-effect gene that functions at the oocyte-to-embryo transition in many vertebrate species including human, pig, cattle, sheep, mouse, rat, frog and zebrafish. Recently, through in silico studies, a gene structurally related to Zar1, called Zar1-like has been identified in many vertebrates, including the chicken. The objectives of this study were to investigate the expression of the chicken Zar1 and Zar1-like genes in chicken tissues and embryos and to determine whether sexual maturation affects their mRNA abundance. RNA was extracted from various organs of chickens aged from one month up to two years old and from chicken embryos until day ten of embryonic development. Expression analysis of the genes was performed using RT-PCR and real-time PCR. RT-PCR analysis revealed that both genes were preferentially expressed in chicken oocytes, ovary and testes and in embryos during embryonic development. Quantitative real-time PCR analysis revealed a significant up regulation of Zar1 in the mature ovary, and also a significant up regulation of Zar1 and Zar1-like genes in the testes of sexually mature roosters, suggesting a key role of these genes in the chicken fertility. In contrast, expression of Zar1-like was not affected by age in the chicken ovary. Our results indicate that the chicken Zar1 and Zar1-like transcripts are co-expressed in high levels in the chicken gonads. In addition their expression beyond the stage of embryonic genome activation suggests an embryonic and not only a maternal origin of these transcripts.

Seasonal variation in testicular volume and sexual behavior of Chios and Serres rams.

The seasonality of testicular volume and sexual behavior in two Greek breeds of sheep, Chios and Serres, was studied using 10 mature rams of each breed. The rams were kept out of breeding during 2 consecutive years. Live weight was recorded at monthly intervals. Testicular volume was measured at 2-week intervals using an orchidometer. Sexual behavior was recorded at 4-week intervals by exposing rams to oestradiol-treated ovariectomized ewes. The total number of mounts and matings, and the reaction time to first mount and mating were recorded. Chios rams were heavier than Serres rams (87.4 +/- 2.0 kg versus 76.4 +/- 1.9 kg, respectively). All traits varied significantly with month but only testicular volume exhibited a clear seasonal trend in both breeds, with a maximum volume in July-August and a minimum in February-April. Number of mounts showed clear seasonal trend in the Chios breed, with maximum in November. Number of mounts was significantly correlated with length of daylight (-0.18 and -0.21 for Chios and Serres, respectively; P < 0.01) and relative humidity (0.14 and 0.17 for Chios and Serres, respectively; P < 0.05). Testicular volume was positively correlated with air temperature (0.28; P < 0.01) and relative humidity (-0.23; P < 0.01) in the Serres breed only. Results of this study suggest that the testicular volume and number of mounts exhibit seasonal variation in the Chios and Serres rams.

Sources of variation and genetic profile of spontaneous, out-of-season ovulatory activity in the Chios sheep.

Organising the breeding plan of a seasonally breeding species, such as sheep, presents a challenge to farmers and the industry as a whole, since both economical and biological considerations need to be carefully balanced. Understanding the breeding activity of individual animals becomes a prerequisite for a successful breeding program. This study set out to investigate the sources of variation and the genetic profile of the spontaneous, out-of-season ovulatory activity of ewes of the Chios dairy sheep breed in Greece. The definition of the trait was based on blood progesterone levels, measured before exposing the ewes to rams, which marks the onset of the usual breeding season. Data were 707 records, taken over two consecutive years, of 435 ewes kept at the Agricultural Research Station of Chalkidiki in northern Greece. When all available pedigree was included, the total number of animals involved was 1068. On average, 29% of all ewes exhibited spontaneous, out-of-season ovulatory activity, with no substantial variation between the years. Significant sources of systematic variation were the ewe age and live weight, and the month of previous lambing. Older, heavier ewes, that had lambed early the previous autumn, exhibited more frequent activity. Heritability estimates were 0.216 (+/-0.084) with a linear and 0.291 with a threshold model. The latter better accounts for the categorical nature of the trait. The linear model repeatability was 0.230 (+/-0.095). The results obtained in this study support the notion that spontaneous out-of-season ovulatory activity can be considered in the development of a breeding plan for the Chios sheep breed.