Identification of Cytokeratins in Bovine Sperm Outer Dense Fibre Fractions

Outer dense fibres (ODF) are important substructures of mammalian sperm tails that are involved in the regulation of sperm motility. In this study, we investigated the identity of several sodium dodecyl sulphate (SDS)‐insoluble ODF proteins. Bovine ODF were purified by separating sperm heads and tails using ultrasound and Percoll^® density gradient centrifugation. Sperm flagella were treated with the detergent cetyltrimethylammonium bromide (CTAB). CTAB‐insoluble material, which reportedly represents the ODF fraction, was collected, and electron microscopy confirmed a highly purified ODF fraction. We found after solubilization of this fraction with SDS that high amounts of insoluble material were retained after centrifugation. SDS‐insoluble material was collected and quantitatively dissolved in 8 M urea. SDS‐gel electrophoresis in the presence of urea revealed polypeptides with apparent molecular masses of approximately 25, 43, and 50 kDa. Subsequent immunoblotting with anti‐cytokeratin antibodies detected two urea‐soluble, SDS‐insoluble proteins with apparent molecular masses of 45 and 66 kDa. The 45‐kDa protein was identified as cytokeratin 19. An antibody reacting with a palette of cytokeratins (CK 1–18 and CK 20), KL1, was the only antibody that reacted with the 66‐kDa polypeptide. We conclude that sperm ODF fractions contain at least one each of type I and type II intermediate filaments. As keratins and intermediate filaments are described as rope‐like structures, we suggest that these intermediate filaments play an important structural or tension‐bearing role in sperm flagella.     

Effects of Confluent, Roscovitine Treatment and Serum Starvation on the Cell-cycle Synchronization of Bovine Foetal Fibroblasts

The present study was designed to examine the effects of cell-cycle synchronization protocols, such as confluent, roscovitine treatment and serum starvation, in bovine foetal fibroblasts on synchronization accuracy at G0/G1, viability, apoptosis, necrosis and ploidy for use as a nuclei donor. The cells in 5-10 passages were randomly allocated into three treated groups. Cells were cultured either in Dulbecco's modified Eagle's medium (DMEM) + 10% foetal bovine serum (FBS) until 90% confluent (group 1, confluent), in DMEM + 10% FBS + 30 microM roscovitine for 12 h (group 2, roscovitine), or in DMEM + 0.5% FBS for 5 days (group 3, serum starvation). Most of the cells (>80%) in all groups were arrested at the G0/G1 stage. Although the rates did not differ, cells in group 1 showed an increased cell population arrested at the G0/G1 phase. Significantly (p < 0.05) higher rates of apoptosis occurred in group 3 than in group 1 and 2 (10% vs 6% and 6%, respectively). No differences in chromosomal abnormality were observed among groups. However, by increasing the number of cell culture passages up to 15, significantly (p < 0.05) higher chromosomal abnormality was observed than in 5 and 10 passages (39% vs 28% and 23%, respectively) in group 1. The results clearly indicated that bovine foetal fibroblasts could be effectively synchronized at G0/G1 stages by all the three different treatments, confluent, roscovitine and serum starvation. However, cells in confluent showed reduced apoptosis and necrosis when they underwent 5-10 passages, exhibiting increased percentage of cells with stable chromosome diversity. Hence, cells in confluent merit further studies before they could be used as nuclear donors.     

Theileria orientalis: a review

Theileria orientalis (also known historically as T. sergenti and T. buffeli) is responsible for benign or non-transforming theileriosis, and exerts its major effect through erythrocyte destruction. The life cycle of T. orientalis is essentially similar to that of other Theileria species, except that the schizonts do not induce transformation and fatal lymphoproliferation. The pathogenesis of anaemia as a result of infection is not clearly established and may be multifaceted. Clinical signs of weakness, reluctance to walk and abortion are early but non-specific indications of disease, particularly if accompanied by a history of cattle being moved. Physical examination may reveal pallor (pale eyes, vaginal mucosa), pyrexia, and elevated heart and respiratory rates. T. orientalis is an economically important parasite of cattle in New Zealand, Australia and Japan, especially where naïve animals are introduced into an endemic area or in animals under stress. Increased awareness of the risks posed by the parasite is required to enable management practices to be implemented to minimise its impact.     

The serological BHV1 status of dams determines the precolostral status of their calves

Precolostral calves and their dams were serologically investigated for the presence of antibodies against Bovine Herpesvirus 1 in diagnostic tests with a very high sensitivity and specificity. Although the syndesmo-chorial type of placenta of ruminants does not transfer gamma globulins, a large number of calves had antibodies, in most cases in a very low concentration. Significant correlations were found between the serological status of the dam, the status of the calf, and the titre of antibodies. Oral intake of maternal blood by the calf at birth or transmission or leakage of maternal antibodies during pregnancy might be possible causes of precolostrally positive calves. From the results it is concluded that to reduce the risk of obtaining BHV1-positive calves, BHV1-negative dams should be selected for breeding purposes.     

含莫纳可林K红曲降低鸡蛋胆固醇效果的研究

48只68周龄的海兰褐蛋鸡分成三组，按0.5％和10％比例分别添加含0．4％莫纳可林K红曲于各组日粮内，饲喂22d，观测其对鸡蛋胆固醇变化的影响。结果表明，投药后5—6d，10％添加组和5％添加组蛋黄胆固醇开始下降；15—22d的胆固醇含量与1～5d相比，10％添加组和5％添加组分别下降了9．02％和5．10％，而对照组升高了5．22％。     

Dispersal of Dictyocaulus viviparus larvae from bovine faeces in Ireland

An involvement of Pilobolus species fungus in the dispersal of Dictyocaulus viviparus third stage larvae from dung to surrounding herbage under Irish conditions was investigated. The presence of Pilobolus kleinii on artificial dung pats containing first stage larvae of D viviparus was associated with a 19-fold increase (P less than 0.05) in numbers of third stage larvae recovered from the surrounding herbage. A subjective examination of natural dung pats showed that the presence of Pilobolus species was significantly correlated with hours of bright sunshine (r = -0.5, P less than 0.01), total rainfall (r = 0.41, P less than 0.05) and the height of herbage surrounding the pats (r = 0.31, P less than 0.001). A multiple regression analysis showed that meteorological parameters and the height of surrounding herbage accounted for 38 per cent of the variation in growth of Pilobolus species on dung pats. The incidence of extensive damage to natural dung pats within five days of deposition, caused by biotic factors, another possible cause of D viviparus third stage larvae dispersal, varied from 0 to 92 per cent of the pats depending on their degree of dryness.     

Aluminium tolerance in wheat: correlating hydroponic evaluations with field and soil performances*

The objective of this study was to correlate root length of wheat (Triticum aestivum L.) genotypes grown in Al-containing, acidic hydroponic solutions, with root weights from acid-soil experiments and field scores from Brazilian acid-field trials. A total of 43 wheat genotypes, primarily from Brazil, were evaluated by growing seedlings for 4 days in hydroponic solutions containing 0.0–4.0 mg/l Al. The root growth rate of all the genotypes was reduced with the addition of Al to the solution and the Al-sensitive and Al-tolerant wheat genotypes were clearly identified. Genotypes with intermediate Al-tolerance levels showed variable root lengths in response to Al stress. Correlations between root length or a root tolerance index (RTI) in the Al solutions versus acid-soil experiments and acid-field trials were highly significant (r = 0.71–0.85, P < 0.01). The most significant correlation was observed among seedlings grown in 1 mg/1 Al. This study presents evidence that this short duration and simple screening technique provides a highly significant correlation with previous acid-soil Al-tolerance evaluations. Furthermore, the data obtained suggest that hydroponic screening of wheat seedlings for Al tolerance may be used in breeding programmes or in screening germplasm collections.     

Estimating the genetic relationship of hybrid bromegrass to smooth bromegrass and meadow bromegrass using RAPD markers

The two most widely grown bromegrass species in North America are smooth bromegrass, a hay type grass, and meadow bromegrass, a pasture type grass. Hybrid populations between these two species have been developed through hybridization and recurrent selection. The objective of this study was to determine the genetic relatedness of the hybrid bromegrass population S‐9073M to its parental populations using random amplified polymorphic DNA (RAPD) markers and to determine genetic variation within and between populations. Individuals from each of the three populations were genotyped at 43 polymorphic RAPD loci. One of the RAPD fragments was meadow bromegrass‐specific. Cluster analysis showed three groups representing the two parental populations and the hybrid population. An analysis of molecular variance (AMOVA), showed that the hybrid population had the highest within‐population variation, followed by meadow bromegrass and smooth bromegrass. The interpopulation genetic distance (phi‐statistic =Φ_st) was highest between meadow and smooth bromegrass and lowest between smooth and hybrid bromegrass. The hybrid population was genetically intermediate to smooth bromegrass and meadow bromegrass, but closer to smooth bromegrass, possibly reflecting the selection criteria used in the development of this hybrid.