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Biomedical subjects

D Waddington

Publications and source records attributed to D Waddington.

At least 37 records · Page 2Linked to original sources

Maintenance of bovine oocytes in meiotic arrest and subsequent development In vitro: A comparative evaluation of antral follicle culture with other methods.

The frequency of development of bovine embryos produced by maturation, fertilization, and culture in vitro is lower than that observed in vivo. One factor that may affect both the frequency of development and the quality of the embryos produced is the developmental competence of the oocyte. In current in vitro production systems, oocyte maturation, characterized by the resumption of meiosis, occurs after oocyte aspiration from the follicle. The developmental competence of individual oocytes may be improved by inducing maturation after culturing under conditions that inhibit the resumption of meiosis. In order to test this hypothesis, a system has been established in which intact antral follicles (3-8 mm in diameter) are cultured in vitro. During this period the oocytes are maintained at the germinal vesicle (GV) stage under the inhibitory effects of the follicle. Culture of intact antral follicles was compared with two other "physiological" methods for the maintenance of GV arrest: oocytes were cultured attached to a small part of the follicle wall or within hemisections of follicles. It was found that 96.8% of oocytes recovered from intact antral follicles-as compared to 24.6% attached to a small part of the follicle wall and 62.7% within hemisections of follicles-were maintained at the GV stage after 24-h culture. The effects on GV arrest and subsequent maturation of the oocytes were evaluated after longer periods of antral follicle culture (2, 4, and 7 days). As the culture period increased, the number of GV-arrested oocytes decreased; the maximum percentage of GV arrest was observed after 24-h culture. The majority of these oocytes matured to metaphase II. A comparison of blastocyst production was made after fertilization and subsequent development of oocytes obtained following follicle culture and of control oocytes aspirated directly from antral follicles. The cleavage rate and percentage of blastocyst production in these two groups were 54.6 +/- 13.9%, 48.4 +/- 8.4% and 68.6 +/- 8.6%, 32.8 +/- 10.8%, respectively. Statistical analysis showed significant differences in both cleavage rate and blastocyst production between these two groups. Total cell numbers in the control group were 144.6 +/- 7.28 and 152.0 +/- 25.8 after follicle culture. It is concluded that culture of intact antral follicles for 24 h is an alternative method for the maintenance of bovine oocytes in meiotic arrest and that these oocytes acquire a greater developmental competence in vitro.

Animals↗

Chronology of events in the first cell cycle of the polyspermic egg of the domestic fowl (Gallus domesticus).

The nuclear population in the polyspermic egg of the domestic hen was examined in whole-mount preparations of the germinal disc. The numbers of nuclei varied in groups of hens from averages of 5.9 to 26, depending on days from insemination. Changes in development from initial formation of pronuclei to the early mitoses of the zygote nucleus were staged according to the position of the egg in the oviduct. The findings substantiated earlier accounts on the timing of the apposition of the parental pronuclei towards the end of the first cell cycle. Additionally, analysis of the spatial distribution of accessory spermatozoal nuclei showed a slight, but significant, dispersal from a clustered arrangement at this time.

Animals↗

Allograft responses can interfere with the development of immunity against Theileria annulata following vaccination with parasite infected cell lines.

Theileria annulata macroschizont-infected cell lines are successfully used as vaccines in several countries. The inoculated animals produce a strong allogeneic response against the MHC antigens of the immunizing cell line followed by an anti-parasite response. Immunity against the parasite wanes in the absence of challenge and re-immunization is sometimes recommended. However, it is not known if allogeneic responses generated by the first immunization with a T. annulata infected cell line will interfere with the boosting of immunity against the parasite at the time of re-immunization with the same cell line. Animals were primed against MHC antigens by skin grafting, followed by immunization with a T. annulata infected cell line prepared from the skin donor. A strong anti-MHC response was produced. This interfered with parasite transfer and the development of an anti-parasite immune response; the effect was more marked when a low vaccine cell dose was used. There was a negative correlation between the ease of isolating infected cells from the animals after cell line immunization, and the subsequent response to challenge. Where no cell lines could be isolated, the animals were fully susceptible to sporozoite challenge. These observations are of immediate importance in endemic areas where cell lines of T. annulata schizonts are being used as vaccines to control the disease.

Animals↗

Behavioural and anatomical consequences of two beak trimming methods in 1- and 10-d-old domestic chicks.

1. To examine the effects of beak trimming on behaviour, beak anatomy, weight gain, food intake and feather condition 360 ISA Brown chicks were trimmed by hot cut or cold cut at 1 d or 10 d of age or were sham-operated controls. The experiment was a 3 x 2 factorial design, with the chicks housed in littered pens in groups of 10 and observed for 6 weeks after trimming. 2. In the first week after trimming, when trimmed birds were compared with untrimmed controls, they were less active (sat and slept more), fed less, preened less and generally engaged in less beak-related behaviour. 3. These differences waned sharply during week 2 and had disappeared by week 5. There were very few differences between hot- and cold-cut birds. 4. There were also differences in production variables: trimmed birds grew more slowly during the week after trimming, their food intake was depressed for 3 weeks and food conversion efficiency improved for 2 weeks. 5. The only significant effect on feather scores was better plumage condition in the groups trimmed at 1 d and scored at 6 weeks. 6. To examine the anatomical effects 36 ISA Brown chicks trimmed by hot or cold cut at 1 d or 10 d of age were killed at 21 and 42 d after trimming, and their beaks were processed and examined histologically. In all trimmed groups healing was very rapid and no scar tissue was seen but, unlike the controls, the regrown tips contained no afferent nerves or sensory corpuscles. 7. Beak lengths immediately after trimming were 40% to 50% shorter than controls; the anatomical consequences of both methods were identical. 8. Overall, it was judged that the effects on behaviour and beak anatomy were much less severe than previously reported for birds trimmed at older ages. If birds do have to be trimmed then the procedure should be carried out in young birds: from the birds' standpoint 1 d appears to be the most suitable.

Aggression↗

Vitamin C supplementation and fear-reduction in Japanese quail: short-term cumulative effects.

1. Fear-related behaviour was measured in individually-housed Japanese quail which had received either untreated drinking water (UDW) or a vitamin C solution (ascorbyl-2-polyphosphate, APP, 1 g L-ascorbic acid/1 for 1 or 4 d. Behavioural tests were carried out at 24 or 27 d of age. Water consumption was measured daily and plasma ascorbic acid (AA) content was assayed after differential treatment for 1 or 4 d. 2. The quail in the APP treatment groups drank significantly more than did those receiving untreated drinking water. 3. Supplementation with APP raised plasma AA concentrations substantially above those found in UDW quail. The lower AA concentrations found after APP treatment for 4 d rather than one may have reflected either inter-assay variation or limited intestinal absorption following excess ingestion. 4. Quail treated with APP showed less home-cage avoidance of a novel object and shorter tonic immobility fear reactions than did the UDW birds. The UDW quail showed remarkably similar avoidance and TI scores on test days 1 and 4. Conversely, continued treatment with APP further reduced avoidance scores but, although TI durations fell slightly with continued treatment there were no significant day effects. 5. It was concluded that APP supplementation can reduce underlying fearfulness but that it may not be necessary to continue treatment for longer than 24 h immediately prior to a known frightening event.

Analysis of Variance↗

Factors influencing the transmission and incidence of tropical theileriosis (Theileria annulata infection of cattle) in Morocco.

A longitudinal epidemiological investigation of tropical theileriosis was carried out in an endemic region of Morocco during the 1991 disease season. This involved approximately 220 cross-bred cattle on 15 farms. Data were collected on the frequency of existing infections, the size of the vector tick population and the incidences of new infections and clinical disease, and these were then analysed using statistical models. The prevalence of subclinically infected carriers of Theileria annulata, the number of adult Hyalomma detritum, the vector, and the probability of becoming newly infected with T. annulata increased significantly with the age of cattle, although the age effect on new infections may be a result of increased tick numbers on older animals. The probability of clinical disease in newly infected cattle was not significantly influenced by age or by the number of adult ticks, but was significantly positively associated with the cattle population on the farm. The number of H. detritum nymphs counted on cattle in the autumn was related significantly to the previous adult tick count on the same animal.

Animals↗

Overdose rates in lithium-treated versus antidepressant-treated outpatients.

Our clinical experience that lithium is an agent used infrequently for self-poisoning was investigated. The rate at which such overdoses occurred was determined from hospital records. A retrospective case notes study then compared overdose rates in patients taking lithium versus patients on antidepressant medication. A low rate of lithium overdoses was identified, and this appeared to be due to patients on lithium rarely taking overdoses rather than their choice of agent.

Adult↗

Electroblotting proteolytic products from native gel for direct N-terminal sequence analysis: an approach for studying protein-protein interaction.

Proteins which are electroblotted from native gels onto polyvinylidene difluoride (PVDF) membranes are suitable for detailed structural analysis. This method, in conjunction with limited proteolysis and N-terminal sequencing, has been used to study the molecular interactions between native protein molecules. The interaction between recombinant interleukin-2 (rIL-2) and its receptor (rIL-2R alpha) was examined as a model system. The working strategy consists of (i) proteolysis of rIL-2R alpha and rIL-2R alpha/rIL-2 complex, (ii) separation of the major proteolytic products by native polyacrylamide gel electrophoresis followed by electroblotting onto PVDF membrane, and (iii) sequence analysis of the blotted protein bands for the identification of peptide regions sensitive to proteolysis. Results have indicated that the exon 3 encoded region in rIL-2R alpha is sensitive to proteolysis regardless whether it is complexed with rIL-2 or not. This suggests that no major conformational changes occur in rIL-2R alpha during interaction with rIL-2. This electroblotting approach is, therefore, useful for studying protein-protein interaction in solution.

Animals↗

Relationship between clustering of post-ovulatory and hierarchical follicles and their order of ovulation in the mature domestic hen (Gallus gallus domesticus).

Hierarchical (> 8 mm) and post-ovulatory follicles from the ovaries of laying hens at 45 and 86 weeks of age were assigned to clusters defined by the coalescence of their follicular stalks. They were also classified by size to give the succession in which ovulation had occurred, or would occur for the hierarchical follicles. The association between the order of ovulation and the observed clusters was compared with randomness and with dependence on the cluster in which the immediately preceding ovulation occurred. The order of ovulation was associated randomly with clusters, and was independent of the cluster containing the preceding ovulation. Many perceived clusters of successive follicles and post-ovulatory follicles may be compatible with a random order of ovulation. These findings do not support theories of stimulation or inhibition of adjacent follicles, leading to eventual ovulation, by a dominant follicle.

Aging↗

Modification of intra-ovarian follicular distributions in broiler breeder hens by ad libitum or restricted feeding.

1. Ovarian follicles in broiler hens from 3 lines fed ad libitum or food restricted were examined at point of lay and from 2 of these lines at 47 weeks of age. The lines were Ross 1 birds and birds selected to be lean or fat on the basis of their plasma very low density lipoprotein concentrations at 7 weeks of age. 2. Follicles of diameter greater than 1.4 mm were classified into size groups, and the numbers in these groups in the anterior and posterior halves of the ovary were compared for the effects of line and feeding régimen. 3. There were more yellow follicles (> 8 mm diameter) in the anterior half of the ovary, and this difference was greater for ad libitum birds. Ad libitum feeding did not influence the number of yellow follicles in the posterior segment of the ovary. 4. The increase in numbers of yellow follicles in the anterior segment of the ovary differed according to line at point of lay, but not at 47 weeks of age. 5. Birds of 47 weeks of age showed higher numbers of follicles 1.4-6 mm in diameter in the posterior segment of the ovary, and smaller numbers of follicles 6-8 mm in diameter. Thus, although large numbers of white follicles were in the earlier stages of growth, there were fewer ovulable yellow follicles because of higher levels of atresia.

Animals↗

Influence of quantity of litter on nest box selection and nesting behaviour of domestic hens.

1. Twenty two hens (Gallus gallus domesticus) were given a free choice, for egg laying, between 4 nest boxes containing different quantities of wood shavings (litter): 3180 cm3 (large), 1060 cm3 (medium), 350 cm3 (small) and none (zero). 2. There was considerable individual variation in the quantity of litter selected for egg laying, but 17 hens laid 50% or more of their eggs in a single litter quantity. Hens selected the large quantity in preference to the small and zero. More searching, nest selection and nesting behaviours tended to be directed to the boxes with the largest quantities and also to the quantities that the hens finally selected for egg laying. 3. In a second experiment, the hens from experiment 1 which showed a preference for litter, and a further 22 naïve hens were given a choice of three nest boxes containing no litter and one containing one of 6 quantities. Over 30 trials there were 5 replicates using each of the three quantities used in experiment 1 plus 115 cm3, 40 cm3 and 10 cm3. 4. There was a wide range in the proportion of trials in which hens selected litter; for experienced hens the range was 0.43 to 0.97, but only two naïve hens were within this range, all others being between 0 and 0.43. 5. The pattern of choosing litter quantities was comparable but at different levels of preference in experienced and naïve hens and a contour map was derived enabling predictions to be made of the probability of a particular quantity being chosen. 6. The variability of individual responses suggest that no single type of nest box system can cater for the requirements of all hens.

Animals↗

Prevalence of Theileria in the tick Hyalomma detritum detritum in the Doukkala region, Morocco.

The overall prevalence of Theileria species, mainly, if not exclusively, T.annulata, in 901 Hyalomma detritum detritum collected from cattle in the Doukkala region of Morocco over a period of 2 years was 21.5%. The quantity of infection (number of sporoblasts per infected tick) followed the negative binomial distribution with between one and 250 sporoblasts per infected tick. Infected ticks were found in eight of fourteen areas examined whilst T.annulata was present in all fourteen. There were significant differences in both the prevalence and the quantity of infection between ticks collected from different farms, and between nymphs collected in the autumn from these farms, and moulted in the laboratory, and adults collected in the following summer. The prevalence, but not the quantity, of infection was higher in female than in male ticks. No correlations were established between infection of engorged nymphs and the breed, sex and Theileria piroplasm parasitaemia of the host animal. However, calves infected a greater proportion of nymphs than adult cattle and the heavier the infestation of nymphs on an animal, up to a plateau, the higher the prevalence of infection in those nymphs. There were no differences in infection between ticks moulted at 24 degrees C and 37 degrees C, after the engorged nymphs had been stored at 18 degrees C to simulate over-wintering.

Animals↗

Size distribution of adipocytes and variation in adipocyte number in lines of mice selected for high or low body fat.

1. Adipocyte size and number were measured in epididymal fat pads from lines of mice selected for high or low body fat. 2. Epididymal fat pad weight at 10 weeks of age was 4-fold greater in the high fat than the low fat mice, accompanied by a 7-fold greater adipocyte volume but there was no difference in adipocyte number. Smaller differences in fatness and cell volume were observed at younger ages. 3. The high fat mice showed one or two populations of cells up to 5 weeks of age and at 7 weeks 3 days and 10 weeks showed two or more populations. In contrast, the low fat animals at all ages showed either one or two populations. 4. These findings imply that greater lipogenesis and cell filling, not cell proliferation, is the primary mechanism of increased fatness in these selected lines of mice.

Adipose Tissue↗

Changes in ovarian function of female turkeys photostimulated at 18, 24 or 30 weeks of age and fed ad libitum or restricted until point of lay.

1. Females from a medium body weight strain of turkeys were fed ad libitum or restricted to 0.6 of ad libitum-fed body weight at sexual maturity and were photostimulated at 18, 24 or 30 weeks of age. Ovarian follicular structure was assessed in 8 birds from both rearing treatments at photostimulation, after photostimulation for 6 or 12 weeks and at 55 weeks of age. 2. The patterns of small white follicles and of larger yellow follicles were similar in restricted and in ad libitum-fed turkeys for birds in lay at the same age, regardless of the time elapsed since photostimulation. 3. Among birds in lay the number of normal yellow follicles and the proportion of multiple yellow follicles declined with age. A relatively low proportion of restricted birds photostimulated at 18 and 24 weeks were in lay after 12 weeks of photostimulation. 4. For birds in lay the number of white follicles 1.4 to less than 1.8 mm in diameter increased and the number greater than 1.8 mm declined with age. There was a corresponding increase in the rates of atresia in follicles 1.8 to less than 5.0 mm in diameter.

Age Factors↗

Estimation of restriction maps with known site order using a generalized linear model.

A generalized linear model with Gamma errors is used to estimate the coordinates of a restriction map when the site order is known. This can be conveniently programmed in a wide range of statistical packages (e.g. Genstat 5, Minitab, SAS), and gives maximum likelihood estimates with their associated optimal properties. Regression diagnostics allow the checking of assumptions and help to identify mis-specified, influential or discordant fragment lengths. A specific diagnostic for identifying fragment lengths causing reversal of restriction site order is derived. Exact 'fragment' lengths from DNA sequencing can be conveniently included in an approximate manner by giving them a larger weight than observed restriction fragment lengths. Two examples and the Genstat 5 codes used in their analysis are presented.

Algorithms↗