Developmental competence of heat stressed oocytes from Holstein and Limousine cows matured in vitro

The negative effects of heat stress on dairy cattle's fertility have been extensively studied, but the relevant knowledge for beef cattle is rather limited. The aims of this study were to investigate the effects of HS during in vitro maturation on the developmental potential of oocytes derived from Limousine and Holstein cows and to estimate the effect of the differential gene expression of important genes in oocytes, cumulus cells and blastocysts in the growth competence between the breeds. In seven replicates, cumulus oocyte complexes from Holstein and Limousine cows were matured for 24 hr at 39°C (controls C; Hol_39, Lim_39) or at 41°C from hour 2 to hour 8 of IVM (treated T; Hol_41, Lim_41), fertilized, and presumptive zygotes were cultured for 9 days at 39°C. Cleavage and embryo formation rates were evaluated 48 hr post-insemination and on days 7, 8 and 9, respectively. From all groups, subsets of cumulus cells, oocytes and blastocysts were analysed for the relative expression of genes related to metabolism, stress, apoptosis and placentation. No difference was detected in cleavage rate or in blastocyst formation rate among the control groups. In both breeds, heat stress reduced blastocyst yield, but at all days the suppression was higher in Limousines. In Holsteins, altered gene expression was detected in cumulus cells (G6PD, GLUT1) and blastocysts (PLAC8), while in Limousines, differences were found in oocytes (G6PD, HSP90AA1), in cumulus cells (CPT1B, HSP90AA1, SOD2) and blastocysts (DNMT, HSP90AA1, SOD2). It appears that Holstein COCs are more tolerant than Limousine COCs, possibly due to compulsory, production driven selection.     

Cytogenetic effects of grape extracts (Vitis vinifera) and polyphenols on mitomycin C-induced sister chromatid exchanges (SCEs) in human blood lymphocytes

In the present study, the effects of extracts and polyphenol-rich fractions as well as monomer polyphenols identified in them, from both red and white grapes, on mitomycin C (MMC) induced sister chromatid exchanges (SCEs) in human peripheral blood lymphocytes were investigated. The grape extracts and two of the three polyphenol-rich fractions promoted MMC-induced SCEs at concentrations from 75 to 300 microg/mL. However, none of the extracts or fractions alone induced SCEs. Thus, these results suggest caution especially with regard to the use of grape extracts as dietary supplements. On the other hand, the fact that these extracts were not genotoxic alone may indicate a selective activity against genetically damaged cells. This is the first study regarding the clastogenic effects of grape extracts in human cells. Moreover, from the tested polyphenols, caffeic acid, gallic acid, and rutin hydrate enhanced MMC-induced clastogenicity, whereas ferulic acid, protocatechuic acid, (+)-catechin, (-)-epicatechin, and trans-resveratrol had no effect at concentrations between 5 and 100 microM. The differences in the chemical structures of the tested polyphenols may account for their differential effects on MMC clastogenicity.     

Comparison of Greek breeds of horses using RAPD markers

To assess genetic markers able to distinguish between five Greek breeds of horses, the random amplified polymorphic DNA (RAPD) technique was used. A total of 40 primers were initially screened in all breeds to identify primers that consistently revealed well-amplified, polymorphic bands. Despite the variability that was found within breeds, no specific RAPD marker for the discrimination of the breeds was detected. This is probably due to the extensive cross-breeding that has taken place over the years, resulting in a mixing of breeds. The high phenotypic divergence of the pony of Skyros from all the other Greek breeds of horses could be explained in terms of micro-evolutionary processes, such as natural selection owing to differential fitness. In addition, to determine the genetic affinities between breeds, all samples were screened with five primers. These primers produced a total of 93 bands, 51 of which were polymorphic. Data analysis for genetic polymorphism revealed a degree of divergence between breeds. According to the discriminant analysis (DA) classification, 100% of the individuals examined in this study could be classified correctly into their breeds. The results of this study show that RAPDs could be a powerful tool for the classification testing of individual horses, as well as for the determination of the genetic variation found among breeds, which is important for a proper management policy designed to protect this species.     

Insecticide resistance status of the codling moth Cydia pomonella (Lepidoptera: Tortricidae) from Greece

The codling moth Cydia pomonella L. is controlled mostly with chemical insecticides in Greece and control failures have been reported. However, there are no insecticide resistance studies in the country as yet. We examined the insecticide resistance status of 33 and 38 populations of fifth-instar non-diapausing and diapausing larvae, respectively by applying bioassays, biochemical and DNA diagnostics. Diagnostic concentrations of azinphos-methyl, phosalone, deltamethrin, thiacloprid, fenoxycarb, tebufenozide, methoxyfenozide and diflubenzuron were used in bioassays. Almost all populations showed reduced susceptibility to at least one insecticide and approximately half of them to all insecticides examined compared to a laboratory susceptible strain used as reference. However, only one out of six populations tested showed reduced susceptibility in ovicidal tests with fenoxycarb. Cross-resistances were observed among most insecticides, except from the pairs fenoxycarb–phosalone and thiacloprid–phosalone, in non-diapausing larvae. The more obvious biochemical marker associated with the reduced susceptibility observed in both larval instars was elevated cytochrome P₄₅₀ polysubstrate monooxygenases activity, followed by elevated glutathione-S-transferase activity and reduced carboxylesterases activity. Neither sodium channel nor AChE known resistance mutations were found in any of the approximately 1000 individuals of each larval instar screened with diagnostic PCR. Actions for Integrated Resistance Management and application of alternative control methods are discussed.     

Genetic diversity and thermotolerance in Holstein cows: Pathway analysis and marker development using whole-genome sequencing

Heat stress causes extensive losses in the dairy sector, due to negative effects on milk production and reproduction. Cows have evolved a series of protective mechanisms, (physiological, biochemical, behavioural) to cope with the thermostressing environments, which have allowed the preservation of productive and reproductive potential of specific animals during summer; these animals are considered thermotolerant and could be used to design programs of selective breeding. These programs, targeting the generations of a population of heat-resistant animals, would increase the frequency of the desired phenotypes, tackling the financial losses on one hand and reducing the carbon footprints of the dairy sector on the other. The development of genomics techniques has enabled genome wide variant calling, to detect SNPs associated with the desired phenotypes. In this study, we used a comparative genomics approach to detect genetic variation associated with thermotolerance and to design molecular markers for characterizing the animals as tolerant/sensitive. A total of 40 cows from each group were split in four sequencing pools and a whole-genome sequencing approach was used. Results and conclusion: Genome-wide genetic variation between groups was characterized and enrichment analysis revealed specific pathways which participate in the adaptive mechanisms of thermotolerance, implicated into systemic and cellular responses, including the immune system functionality, Heat Stress and Unfolded Protein Response. The markers made a promising set of results, as specific SNPs in five genes encoding for Heat Shock Proteins were significantly associated with thermotolerance.     

Seasonal size distribution,condition status and muscle yield of Mediterranean horse mackerel <Emphasis Type="Italic">Trachurus mediterraneus</Emphasis> from the North Aegean Sea,Greece

ABSTRACT:   The seasonal intrinsic variability of the North Aegean Sea Mediterranean horse mackerel Trachurus mediterraneus population has been studied from monthly samples taken from the Greek purse-seine fishery over a 4-year period (2001–2005). The length and weight frequency distributions differed significantly by season, the highest mean lengths and weights attained in spring and the lowest in autumn. Length-weight relationships were determined on a monthly and annual basis, and monthly variations on the allometry coefficient were found. The differences between the coefficient obtained for annual data and the coefficients for each month were not significant in 50% of the cases. The b -values obtained both in this study and some of the previously reported in the Mediterranean Sea and other Greek waters were compared, showing basic similarities and minor spatial variation. The seasonal cycle in Mediterranean horse mackerel condition appears to be related to the reproductive cycle of the fish. Values of muscle yield ranged between 23.33% and 33.07% (mean value 28.48 ± 2.49%), but variations within seasons were not significant.