The quality of porcine oocytes is affected by sexual maturity of the donor gilt

Although differences in the quality of oocytes derived from young gilts and adult sows are well documented, evidence concerning gametes of pre-pubertal and cycling gilts is scarce and inconsistent. The aim of this work was to establish whether sexual maturity of gilts affects the quality of their oocytes with the use of the brilliant cresyl blue (BCB) test, oocyte diameter and apoptosis. Ovarian morphology was evaluated, and gonads with corpus luteum or albicans were recognized as originating form cycling gilts (C) and those with follicles as originating form pre-pubertal females (P). Altogether 952 cumulus-oocyte complexes (COCs; group P: 554; group C: 398) were examined, whereas 149 COCs, not subjected to BCB test, served as a control for TUNEL. COCs of proper morphology were evaluated by the BCB test which differentiated two categories of gametes: more competent, BCB+, and less competent BCB- oocytes. The control group comprised oocytes of proper morphology aspirated from ovaries of P and C gilts not subjected to BCB test. Finally five groups of COCs were matured in vitro: 1/P-BCB+, 2/P-BCB-, 3/C-BCB+, 4/ C-BCB- and 5/ control. Significantly more large oocytes (≥ 120 μm), more BCB+ oocytes and more high quality (both BCB+ and ≥ 120 μm) oocytes originated from ovaries of cycling gilts than pre-pubertal gilts (p<0.001). The rate of mature oocytes at the MII stage differed significantly between C-BCB+ (68.5%) and P-BCB+ (32.9%) oocytes. The incidence of apoptosis among BCB-treated oocytes after in vitro maturation was 21.4% and was similar to that observed in control oocytes (17.4%). BCB+ oocytes from cycling gilts showed significantly higher (28.7%) incidence of apoptosis than that of the group P (16.2%). Interestingly, high quality oocytes displayed a similar level of apoptosis regardless of the donor puberty. We demonstrated that C gilts provided more BCB+ oocytes as well as more large oocytes than P gilts, although C-BCB+ oocytes showed higher apoptotic rate. In conclusion, high incidence of apoptosis and a big variation in the diameter of more competent BCB+ oocytes make the BCB test a less effective selection tool than previously reported.     

Apoptotic index within cumulus cells is a questionable marker of meiotic competence of bovine oocytes matured in vitro

Information gained from most human studies indicate a negative correlation between the apoptotic index (AI) in cumulus cells (CC) and the quality of the corresponding oocytes. However, results obtained in other species are not so consistent. The rate of apoptosis-free COCs (cumulus oocytes complexes) subjected to IVM (in vitro maturation) also varies among studies. The aim of the present study was to investigate whether the AI in cumulus cells of post-IVM COCs is related to the morphology of pre-IVM COCs and to meiotic competence of bovine oocytes. COCs of known morphology (four grade scale) obtained from individual follicles were matured in a well-in-drop system. After IVM, the external layers of CC of each COC were analyzed by TUNEL. In order to determine the meiotic stage, oocytes were stained with DAPI. It was found that 25.6% of bovine COCs contained apoptosis-free cumulus cells. Moreover, the majority of COCs with apoptotic cells were characterized by apoptotic index lower than 15%. The level of apoptosis in CC was related neither to COC morphology nor to the oocyte meiotic stage. It is suggested that the extent of apoptosis in cumulus cells is not a reliable quality marker of the corresponding oocyte after IVM.     

Timing of the first zygotic cleavage as a marker of developmental potential of mammalian embryos

Embryo quality related to its developmental potential is now one of the most important issues in modern embryology. It has been demonstrated that some in vitro produced blastocysts fail to hatch and implant after transfer despite a normal morphology. Although embryos are able to adjust to sub-optimal culture conditions, significant changes in expression profiles of developmentally important genes have been noticed. Timing of the first zygotic cleavage is considered a non-invasive marker of embryo developmental potential and has been successfully used in human IVF programs for identifying embryos of superior quality. Early-cleaving zygotes are more likely to develop to the blastocyst stage than their late-cleaving counterparts. The timing of the first zygotic cleavage has been associated with several parameters that may affect developmental potential of the resulting embryos. The mechanism causing variation in the timing of the first zygotic cleavage has not been identified. It may be related to culture environment or to some intrinsic factors within the oocyte, the sperm or both. In this paper we discuss some of the important aspects related to the timing of the first zygotic cleavage and its influence on the developmental competence of resulting embryos.     

Preliminary in vitro study on the effect of xanthohumol on rumen methanogenesis

There is a great interest in reducing the methane emission from ruminants as one possible cause of global warming. The aim of the presented study was to determine the effects of xanthohumol, one of the hop (Humulus lupulus) phytofactors, on methane production, microbial population and basic parameters of ruminal fermentation. The experiment was carried out in a batch culture system. The basic substrate (400 mg) consisting of meadow hay and barley meal (60:40) was supplemented with 0 (Control), 0.1, 0.2 or 1.0 mg of xanthohumol. The basic parameters of rumen fermentation and composition of microbial population were measured after 24 h of incubation. For the first time, the results of this in vitro study have demonstrated that xanthohumol is capable to reduce the methane production, even at the lowest dose applied (0.1 mg/400 mg). The observed reduction in methane production by 12-13% was not accompanied by altering the basic rumen fermentation parameters. However, the practical utility of this supplement needs further investigation under long-term in vivo conditions.     

Gilts and sows produce similar rate of diploid oocytes in vitro whereas the incidence of aneuploidy differs significantly

Oocytes derived from prepubertal gilts show reduced developmental competence when compared to oocytes collected from adult sows. Therefore, the aim of the study was to investigate whether gilts (4-5 months old) and adult sows (average age 3.5 years) of the same breed (Polish Landrace x Polish Large White crossbred) differ with regard to the rate of chromosomally unbalanced oocytes after IVM. COCs derived from individual pairs of slaughterhouse ovaries were matured in vitro and analyzed cytogenetically by conventional staining (Giemsa) and FISH methods (probes corresponding to centromeric regions of pig chromosomes 1 and 10). Altogether, 72 females (31 sows, 41 gilts) and 430 secondary oocytes (194 and 236 oocytes of sows and gilts, respectively) were investigated. Cytogenetic analysis revealed diploid (Giemsa, FISH) and aneuploid (FISH) spreads. The incidence of diploid oocytes was similar for sows (26.0%) and gilts (24.5%) whereas the rate of aneuploid oocytes (nullisomic/disomic) was eight times higher in gilts (10.8%) than in sows (1.3%). Diploid and aneuploid oocytes were observed in 64% of investigated females. Pig chromosome 10 was more frequently disomic/nullisomic compared to chromosome 1 suggesting, that like in human, small porcine chromosomes are often involved in the nondisjunction process. In conclusion, chromosomal imbalance significantly contributes to in vitro embryo production in the pig, since over 60% of females produced diploid or aneuploid gametes. The significantly higher rate of aneuploidy among oocytes derived from gilt ovaries may contribute to the reduced developmental competence of gametes collected from nonmature female pigs.     

Disturbances of nuclear maturation in BCB positive oocytes collected from peri-pubertal gilts

The developmental competence (quality) of oocytes is affected by several factors linked to their intrinsic properties and also to growth and maturation environment. Donor puberty and chromosomal complement are one of the main factors influencing oocyte quality. A high rate of porcine oocytes matured in vitro is chromosomally imbalanced. Moreover, there is no published data on chromosomal aberrations in oocytes selected by the brilliant cresyl blue (BCB) test. Therefore, the aim of this study was to analyze whether BCB positive (BCB+) oocytes derived from ovaries of peripubertal gilts (prepubertal NCL and cyclic CL) differ with respect to the incidence of numerical chromosome aberrations. COCs collected from NCL and CL ovaries were selected by the BCB test. Only BCB+ oocytes were matured in vitro and subjected to FISH analysis using molecular probes for chromosome pairs 1 and 10. The rate of BCB+ oocytes was similar for both groups of ovaries (NCL 80%, CL 92%). Altogether 554 oocytes were fixed and 471 oocytes at the MII stage were analyzed cytogenetically. Diploid (2MII) and aneuploid oocytes were detected. The contribution of MII oocytes was similar for NCL (85%) and CL (90%) group. Chromosomally aberrant BCB+ oocytes accounted for 18.0% and ranged from 13.7% for CL and 22.2% for NCL ovaries. Diploidy was a predominant anomaly observed (11.2%) with a significantly higher frequency in BCB+ oocytes of pre-pubertal (16.7%) than cyclic gilts (5.6%, P < 0.05). Aneuploid oocytes occurred with similar rate in NCL (6.7%) and CL (8.5%) females. The majority of aneuploid spreads (72.2%; P < 0.01) concerned the chromosome pair 10. The overall rate of disomy (56%) and nullisomy (44.4%) was similar. We have shown that donor puberty affects the incidence of chromosomal abnormalities in porcine oocytes matured in vitro. Significantly more diploid oocytes was derived from prepubertal ovaries, whereas the frequency of aneuploidy was similar in NCL and CL gilts.     

The present status and potential distribution of relict populations of Aesculus hippocastanum L. in Greece and the diverse infestation by Cameraria ohridella Deschka & Dimić.

Aesculus hippocastanum is a well-known species, which is popular because of its ornamental value. However, data on the demographic structure and potential distribution of A. hippocastanum are limited. The invasion of Cameraria ohridella into Europe has harmed those trees growing in artificial sites, but the presence of this insect in natural stands has been little studied. Here we aimed to investigate the demographic structure infestation level of natural populations of horse-chestnut. Additionally, Maxent modelling was used to predict the potential range of A. hippocastanum, based on the localities available in the literature. Field data analysis indicated that natural populations of A. hippocastanum are mostly found nearby mountain streams. The populations showed a diverse height structure and large numbers of seedlings, which indicate high population dynamics. The level of infestation by C. ohridella varied greatly and correlated with altitude. Secondary infestation might explain this infestation variability in some natural populations. Other hypotheses, such as environmental resistance factors or different genetic variability, are also discussed. By spatial distribution modelling, we found that the precipitation of the coldest and warmest quarters, as well as altitude, are important factors influencing the potential distribution of A. hippocastanum.     

Rich but not poor conditions determine sex‐specific differences in growth rate of juvenile dioecious plants

Journal of Plant Research - Causes of secondary sexual dimorphism (SSD) in dioecious plants are very poorly understood, especially in woody plants. SSD is shown mainly in mature plants, but little...