Genetic differences between autochthonous and breeding populations of common cypress (Cupressus sempervirens L.) in Greece

Individuals were selected for resistance against the imperfect fungus Seiridium cardinale Wag in six common cypress populations (Cupressus sempervirens L.) in Greece. A total of 109 resistant clones and 140 trees from their base populations were surveyed at several allozyme loci. The comparison yielded information on changes in genetic variation due to artificial selection. The genetic structure of the breeding populations was similar to their base populations. Nevertheless, the number of rare alleles among the resistant clones had decreased, showing a similar trend at most loci and for most comparisons. Possible implications for breeding strategies are discussed.     

Effect of insecticide injection on pyruvate kinase of the insect Tenebrio molitor (Coleopteran)

In the present study, we report on the changes observed in the kinetic properties of pyruvate kinase (PK) from whole body homogenate in the three developmental stages of the insect Tenebrio molitor after insecticide or acetone injection, and after injury and 24 h starvation. The apparent K_m value of the enzyme towards phosphoenolpyruvate was higher in the pupae stage than in the rest developmental stages although the same enzyme type appears to be present throughout the life span of the insect as judged by its V_max and sensitivity towards ATP and Alanine. The highest specific activity, twice as high as that of larvae and pupae, was observed in adults. Starvation resulted to activation of the enzyme by increasing the K_m value in the two feeding stages, namely larvae and adult, while injury had the opposite effect. Acetone injection did not have any significant effect at any stage. Insecticide injection had varying effects, depending upon the developmental stage, the dose and the type of the insecticide. In larvae, low doses of methyl paraoxon and methyl parathion activated the enzyme but as the dose increased the K_m value returned to normal levels. Low dose of decamethrin activated the enzyme also but at high dose it caused a severe inactivation as judged by a fivefold increase of the observed K_m. In pupae, decamethrin had no effect, methyl parathion had similar effects to those observed in larvae, and methyl paraoxon caused activation of the enzyme at any dose given. In the adult stage none of the injected insecticides had any effect on the K_m value although V_max and specific activity were significantly altered.     

Testing Usability of Butylated Hydroxytoluene in Conservation of Goat Semen

The objective of this study was to investigate whether butylated hydroxytoluene (BHT) could be used as a suitable supporter or alternative of egg yolk during preservation of goat spermatozoa. Three in vitro experiments and a fertility test were conducted to evaluate the effect of BHT on viability of chilled‐stored semen as well as motility and kidding rate of frozen‐thawed spermatozoa. In the first two experiments, ejaculates (n = 30/experiment) were collected from 10 bucks, split, diluted with egg yolk‐based and egg yolk‐free extenders supplemented with or without 0.3, 0.6, 2, 5 and 8 mm BHT and stored at 5°C for 168 h. In the third experiment, 30 ejaculates were collected from the above‐mentioned bucks, split and diluted with egg yolk‐free extenders supplemented with or without 0.3, 0.6 and 0.9 mm BHT and egg yolk‐based extenders supplemented with or without 5 mm BHT. Diluted semen was cooled to 5°C over a period of 4 h, frozen and thawed in the form of 0.3‐ml pellets. In the fertility test, 75 ejaculates were collected from two proven fertile bucks, split, diluted with egg yolk‐free extenders containing 0.6 mm BHT and egg yolk‐based extenders supplemented with or without 5 mm BHT, frozen and thawed as described above. An insemination volume of 0.6 ml containing 120–140 × 10⁶ progressively motile spermatozoa was used for a single cervical insemination of cloprostenol‐synchronized does (n = 230). The results showed that addition of 5 mm BHT to egg yolk‐deficient (2.5%) extenders significantly improved viability of chilled‐stored semen together with motility (48.5%) and fertility (62.5%) of frozen‐thawed spermatozoa. Replacement of egg yolk in semen extenders by 0.6 mm BHT could sustain not only viability of chilled‐stored semen but also post‐thaw motility (47.5%) and fertility (53.75%) of frozen‐thawed spermatozoa. In conclusion, supplementation of semen diluents with BHT can ameliorate preservability of goat sperm.     

In vitro Effect of Zearalenone and α-Zearalenol on Boar Sperm Characteristics and Acrosome Reaction

This study was conducted to determine the in vitro effects of three different concentrations (125, 187.5 and 250 microM in diluted semen) of zearalenone (zen) and alpha-zearalenol (alpha-zen) on boar sperm. Semen parameters such as motility, viability and spontaneous acrosome reaction were evaluated. From the results it was shown that both zen and alpha-zen affected the sperm characteristics significantly (p < 0.05), except for alpha-zen at the low concentration which did not decrease the percentage of live reacted spermatozoa significantly. In conclusion, zen and alpha-zen are directly toxic when they affect boar semen in vitro and consequently decrease the fertilization ability of the sperm. The higher the concentration of mycotoxin tested, the greater the decline of sperm parameters noticed. The influence of mycotoxins was found to be time- and dose-dependent.     

Influence of nitrogen nutrition on yield and growth of an everbearing strawberry cultivar (cv. Evie II)

Strawberry plants are relatively unresponsive to nitrogen (N) fertilization. Supraoptimal N application also results in excessive vegetative growth, which competes to reproductive growth. Two strawberry field experiments were conducted for two consecutive years using fertigation to investigate the effect of different nitrogen (N) application rates on yield and growth of an everbearing strawberry (cv. Evie II). N was injected weekly into the drip irrigation system at 0, 0.5, 1, 3, and 6 kg N/ha/week the first year and 0, 1, 2, and 3 kg N/ha/week the next year. Fruit yield and fruit number were not affected the first year, but were increased the second year due to N application. However, there was no effect of N nutrition on average berry weight for both years. It was concluded that N fertilization may increase yield and fruit number, but average berry weight remains unaffected.     

The Impact of Cl ?? Cd Interrelationship on Planning Wastewater Reuse in Cabbage

Two greenhouse pot experiments were conducted in Agrinion, Greece, using a randomized block design in four replications, respectively, as follows: The first one included five levels of treated municipal wastewater (TMWW), being used as an irrigation water source. The second one, five levels of applied Cl at a constant soil applied Cd level of 10.36?mg/kg soil, the plants being irrigated with fresh well water. The purpose of these experiments was to study the impact of the Cl ?? Cd interrelationship on planning TMWW reuse, for the irrigation of Brassica oleracea var. Capitata (cabbage) cv F1 Gloria, ehich was used as test plant, in both of these experiments. It was found that the TMWW Cl content, originating mainly from the procedure of wastewater chlorination, was synergistically interrelated with the toxic heavy metal Cd, increasing its soil availability and cabbage plant leaf uptake (edible plant part). As this increase is directly associated with the consumer??s health, it was suggested that the TMWW be subjected to dechlorination process or the disinfection be made by ozonation or UV, which do not include Cl.     

Novel expression of Haemonchus contortus vaccine candidate aminopeptidase H11 using the free-living nematode Caenorhabditis elegans

With the problem of parasitic nematode drug resistance increasing, vaccine development offers an alternative sustainable control approach. For some parasitic nematodes, native extracts enriched for specific proteins are highly protective. However, recombinant forms of these proteins have failed to replicate this protection. This is thought to be due to differences in glycosylation and/or conformation between native and recombinant proteins. We have exploited the free-living nematode Caenorhabditis elegans to examine its suitability as an alternative system for recombinant expression of parasitic nematode vaccine candidates. We focussed on Haemonchus contortus aminopeptidase H11 glycoprotein, which is enriched in a gut membrane fraction capable of inducing significant protection against this important ovine gastrointestinal nematode. We show that H. contortus H11 expressed in C. elegans is enzymatically active and MALDI mass spectrometry identifies similar di- and tri-fucosylated structures to those on native H11, with fucose at the 3- and/or 6-positions of the proximal GlcNAc. Some glycan structural differences were observed, such as lack of LDNF. Serum antibody to native H11 binds to C. elegans recombinant H11 and most of the antibody to rH11 or native H11 is directed to glycan moieties. Despite these similarities, no reduction in worm burden or faecal egg count was observed following immunisation of sheep with C. elegans-expressed recombinant H11 protein. The findings suggest that the di- and tri-fucosylated N-glycans expressed on rH11 do not contribute to the protective effect of H11 and that additional components present in native H11-enriched extract are likely required for enhancing the antibody response necessary for protection.     

Sperm Chromatin Stability During In Vitro Manipulation of Beef Bull Semen

Seven experiments were conducted to study the effect of freezing extenders, antioxidants, motility stimulants, thawing temperature, incubation temperature and time, centrifugation and capacitation on sperm chromatin instability (CI) as well as the influence of sperm CI on pregnancy rates of heifers (n = 360) after AI with frozen semen. Semen was collected once a week from Blonde d’Aquitaine and Limousine bulls (n = 3/breed) via an artificial vagina and only individual ejaculates (n = 300) of >0.3 × 10⁹ sperm/ml and ≥ 70% progressive motility were used. Sperm CI was evaluated by nuclear DNA susceptibility to acid‐induced denaturation using acridine orange fluorescence and by chromatin susceptibility to decondensation using quantitative transmission electron microscopy. Bioxcell extender was better than AndroMed and egg yolk extenders in terms of low incidence of sperm CI in one bull (p < 0.05). Neither antioxidants (EDTA–2Na, Na‐pyruvate and albumin) nor motility stimulants (caffeine and blood serum) had any significant effect on sperm CI. Thawing of frozen semen at 45°C for 30 s decreased (p < 0.025) CI in one bull. Incubation of frozen sperm at 25 and 39°C for 240 min increased sperm CI percentages from 3.47 ± 0.48 and 4.50 ± 0.41% to 6.70 ± 0.36 and 9.71 ± 0.53%, respectively (p < 0.001). Although centrifugation and removal of extracellular milieu increased CI of cooled sperm, it decreased CI of frozen–thawed sperm (p < 0.025). Follicular fluid as a capacitating agent destabilized chromatin structure (p < 0.001). Sperm vulnerability to CI had a negative impact (r² = 0.37–0.77, p < 0.001) on fertility of frozen ejaculates. In conclusion, in vitro manipulation of bovine semen can influence incidence of sperm CI, whereas integrity of sperm chromatin contributes significantly to heifers’ fertility. We would recommend selection of the appropriate extender and thawing temperature for each bull together with careful manipulation of frozen semen to minimize damage of sperm chromatin.     

Influence of nitrogen nutrition on nitrate levels of strawberry leaf blades and petioles

Hydroponic and field experiments were conducted to assess the optimum nitrogen (N) levels in strawberry leaves for the appropriate management of its N nutrition under field or protected cultivation. Unlike the common view that leaf petiole is the right plant organ to sample for strawberries, the study showed that the leaf blade, of the most recently fully expanded leaf, is more responsive and therefore a better indicator of the N supply changes in the strawberry plant. This was attributed to the distinctive characteristic of strawberry plants to accumulate high concentrations of nitrates in leaf blades when there is luxury consumption of N.     

Method agreement between three different chambers for comparative boar semen computer‐assisted sperm analysis

The computer‐assisted sperm analysis (CASA) has become a standard laboratory tool. Although it contributes a lot to the objective sperm motility assessment, its measurements may be affected by many factors. The aim of the study was to evaluate the effect of chamber on boar semen CASA results. Totally, 100 extended (30 × 10⁶ sperm/ml) boar semen samples were analysed by CASA. Each sample was evaluated using Makler, Leja 4 chamber 20 μm and conventional glass slide/coverslip chambers (MC, LC and GSC, respectively). The differences in values between MC and LC and between MC and GSC were significantly positive (higher values for MC compared with LC and GSC) for total motility, progressive, rapid movement, VCL, VSL, VAP, STR and hyperactive, thus indicating a systematic effect. Between LC and GSC, the differences in many parameters (non‐progressive, progressive, slow, LIN, STR, hyperactive) were evenly distributed around zero, while in all other parameters the differences were significantly positive (higher values for LC compared with GSC), except for medium movement. Based on the estimated intraclass correlation coefficients, the method agreement between MC and LC and between LC and GSC was overall moderate to good, depending on the parameter; nonetheless, it was poor between MC and GSC. The limits of agreement between methods can vary considerably depending on the parameter and should be considered when comparisons between CASA measurements of different andrology laboratories or studies have to be performed.