兔核移植胚胎的克隆研究

本研究改进了兔受体卵母细胞的去核程序及供体卵裂球的处理方法，并对核移植胚的体外克隆、继代克隆及克隆胚的体内发育进行了试验。结果表明：卵龄对受体卵母细胞的去核具有显著的影响，卵龄为１５～１８ｈ的去核率为１００％（４５／４５），显著高于１３～１４ｈ的４６．６％（７／１５），Ｐ＜０．０１。ＤＮＡ合成抑制剂Ａｐｈｉｄｉｃｏｌｉｎ处理１６－细胞期卵裂球后，重组胚的囊胚发育率５４％（２０／３７）高于未处理组的４５％（１４／３１），Ｐ＜０．０５。从２枚１６－细胞供体胚分别获得来自一个供体胚的９枚和７枚核移植克隆囊胚，囊胚发育率为５１．６（１６／３１）。然而第一次继代核移植胚的囊胚发育率仅为１１．５％（６／５２）。１６－及３２－细胞期卵裂球的重组胚可发育至产仔，共获２窝８只仔兔。其中１只、２只、２只和３只仔兔分别来自４个供体胚。     

BFF,rbGH对牛卵泡卵母细胞体外受精后发育的影响

利用屠宰黄牛卵巢，对２～５ｍｍ卵泡卵母细胞的体外成熟（ＩＶＭ）、体外受精（ＩＶＦ）、受精卵的体外培养（ＩＶＣ）进行了系列研究。结果表明，在成熟培养液中单独添加１０％Ｄ０ＥＣＳ（发情当天牛血清）获得了卵泡卵母细胞受精后较高的卵裂率（６４．７％）、桑椹胚发育率（３９．９％）和囊胚发育率（２４．８％），说明在成熟培养液中单独添加Ｄ０ＥＣＳ是可行的；在含１０％Ｄ０ＥＣＳ的成熟培养液中再添加１０％或２０％ＢＦＦ（牛卵泡液），均能提高受精卵的卵裂率以及桑椹胚和囊胚的发育率，以添加２０％ＢＦＦ效果较好，其囊胚发育率（３５．０％）显著高于对照组（２４．８％，Ｐ＜０．０５）；在卵泡卵母细胞成熟培养系统和受精卵共同培养系统中添加１０μｇ／Ｌ重组牛生长因子（ｒｂＧＨ），虽对体外受精卵的卵裂率无显著影响（Ｐ＞０．０５），但能显著提高卵裂胚的囊胚发育率（Ｐ＜０．０５）     

电激活对完全体外化牛细胞核移植的影响

牛卵母细胞在体外成熟２３～２４ｈ时去核，去核卵用８０Ｖ／ｍｍ４０μｓ两次电脉冲激活（Ⅰ组）或不激活（Ⅱ组），然后将体外受精、发育的８～３２细胞期胚胎的单个卵裂球注入卵周隙，用８０Ｖ／ｍｍ４０μｓ两次电脉冲诱导卵裂球与去核卵母细胞融合。Ⅰ组操作后存活率和融合率（８８．４％和５５．０％）显著低于Ⅱ组（９５．９％和６５．１％，Ｐ＜０．０５）。融合卵体外培养２４ｈ和５～８ｄ后，Ⅰ组核移植胚胎的卵裂率和桑椹／囊胚发育率（６３．０％和１５．２％）与Ⅱ组（５０．５％和７．８％）无显著差异，但Ⅰ组结果均好于Ⅱ组。将来自两组的２５枚桑椹和囊胚移植到１６头同期受体，在已检查过的８头受体中２头受体妊娠，其中１头于妊娠后４个多月流产，１头于妊娠期满后产出一雄性牛犊。研究结果表明：去核卵母细胞的电激活尽管对重组卵的存活与融合不利，但可改善核移植胚的卵裂和发育。本研究在我国首次获得牛细胞核移植的成功，证明用ＩＶＭ卵母细胞和ＩＶＦ胚胎进行完全体外化的牛细胞核移植是可行的。     

兔卵母细胞的电刺激孤雌激活及体外发育

本文分析了影响兔卵母细胞电激活及发育的主要因素，建立了激活及体外发育的实验程序。试验表明，激活率随着卵龄的增加而升高，采用０．３Ｍ甘露醇为电激液时２０ｈ卵龄的激活率为４０％，显著高于１６及１８ｈ的１４％（Ｐ＜０．０１）和２０％（Ｐ＜０．０５）。含电解质成分的电激液的激活率显著高于非电解质液。随着电激次数及场强的增加，激活率亦显著升高，以０．３Ｍ甘露醇＋１００μＭ ＣａＣｌ２＋１００μＭ ＭｇＣｌ２     

牛性机能发育阶段与卵泡、黄体因素对卵母细胞体外成熟及体外受精的影响

利用屠宰母牛卵巢．对来自不同性机能发育阶段母牛的卵巢卵母细胞．不同状态卵巢卵泡的卵母细胞的体外成熟（ＩＶＭ）、体外受精（ＩＶＦ）进行了系列研究。结果表明．来自初情期、性成熟母牛卵巢的卵母细胞受精后的卵裂率（ ７４． ７％和 ８１． ５％）、囊胚率（ ２３．０％和 ２６． ８％）显著高于来自初情期前母牛卵巢卵母细胞的卵裂率（１８．８％）和囊胚率（１．８％）,Ｐ＜０．０５;有黄体卵巢的卵母细胞体外受精后的发育能力显著低于无黄体卵巢卵母细胞（卵裂率５８．０％：７９． ９％,囊胚率１３．％：２７．１％,Ｐ＜０．０５）;优势卵泡的有无对卵母细胞的卵裂率和囊胚率无明显影响。     

两种蔗糖浓度对切割取样鼠胚冷冻解冻效果的研究

小鼠晚期桑椹胚、早期囊胚和扩大囊胚，经切割取样分别采用０．３６５Ｍ和０．２５Ｍ蔗糖浓度进一步细管和分步脱除甘油解冻后体外培养发育率分别为６５．５％、４６．４％和７５．４％，两种浓度间差异显著（Ｐ＜０．０５）；分步脱除甘油解冻瑟前者差异不显著（Ｐ＞０．０５），与后者差异极显著（Ｐ＜０．０１）。     

牛体外受精胚冷冻保存的研究

对牛卵泡卵母细胞体外受精（ＩＶＦ）１６８ｈ的致密桑椹胚、囊胚用常规快速冷冻法、预冷和不预冷的超快速冷冻法进行了冷冻保存试验。结果表明：ＩＶＦ囊胚采用含１０％甘油的常规快速冷冻法、含２．１ｍｏｌ／Ｌ甘油和０．２５ｍｏｌ／Ｌ蔗糖预冷５ｍｉｎ的一步冷冻法及含２５％甘油和２５％乙二醇预冷５ｍｉｎ的玻璃化冷冻法等３种方法进行冷冻保存，解冻后的继续发育率（６８．０％，５９．０％，６５．７％）均无显著差异（Ｐ＞０．０５），可用快速、简便、预冷的一步冷冻法或玻璃化冷冻法替代常规快速冷冻法；ＩＶＦ致密桑椹胚可用一步冷冻法（２．１ｍｏｌ／Ｌ甘油＋０．２５ｍｏｌ／Ｌ蔗糖）和玻璃化冷冻法（２５％甘油＋２５％乙二醇或２５％甘油＋２５％１，２－丙二醇）进行预冷的超快速冷冻保存；冻前预冷（５ｍｉｎ）能显著提高ＩＶＦ囊胚的冻后形态正常率和继续发育率（Ｐ＜０．０５）；ＩＶＦ囊胚冷冻—解冻后的继续发育率高于ＩＶＦ致密桑椹胚。     

猪2-细胞胚胎细胞的电融合

为了探索猪胚胎细胞核移植技术中细胞融合参数，进行了猪２－细胞胚胎细胞的电融合。电融合时，直流电场强度为２ｋＶ／ｃｍ，脉冲时程选４０μｓ时，可获得７０．４％的融合率和６８．４％的发育率。非电解质融合液（甘露醇）对于融合率和融合胚的发育均优于电解质融合液（ＤＰＢＳ）。在直流电脉冲前的交流脉冲是必要的，可以显著提高融合率（Ｐ＜０．０５）。     

牛体外受精胚胎卵裂阶段对其体外发育能力影响的研究

将经过体外成熟培养（ＩＶＭ）、体外受精（ＩＶＦ）后获得的牛早期胚胎，按其所处卵裂阶段的不同，划分为２细胞、３～４细胞和５细胞以上３个试验组，在体外条件下继续培养到发育囊胚和孵化囊胚阶段，以观察在胚胎早期，受精卵裂发育的快慢与其后胚胎所获得的继续发育能力的关系。结果显示，牛早期胚胎体外囊胚发育率的高低，与其在体外培养（ＩＶＣ）前卵裂发育的快慢成正比。在ＩＶＦ后４２～４６ｈ处于２细胞、３～４细胞和５细胞以上卵裂阶段的早期胚胎，其囊胚发育率分别为６．５％、２１．４％和４４．９％，差异非常显著（Ｐ＜０．００１）；而经过７～９ｄ的ＩＶＣ后仍滞留在ＩＶＣ前原卵裂阶段，丧失继续发育下去能力的胚胎比率则分别为５１．６％、３２．７％和２５．７％，与其ＩＶＣ前卵裂发育的快慢成反比。在ＩＶＦ后６６～７０ｈ仍处于２细胞和３～４细胞卵裂阶段的早期胚胎，其体外发育能力明显降低，囊胚发育率只有１．３％和８．２％；而滞留胚胎率则分别高达９０．９％和５９．４％。结果表明，牛体外受精早期胚胎卵裂发育的快慢，直接影响到其后体外发育的能力。     

牛胚胎体外生产技术的简化研究

利用屠宰牛卵巢分离的卵母细胞，以卵裂率和囊胚发育率为标准，对牛胚胎体外生产过程进行了简化试验。结果显示：不加卵丘细胞的高密度卵母细胞体外成熟（１００￣２００枚／平皿）可代替加卵丘细胞的标准密度（５０枚／平皿）培养，其卵裂率（５７．６％比６２．５％）和囊胚发育率（２３．７％比２９．１％）没有显著差异（Ｐ〉０．０５），体外授精时间可从成熟培养后２２ｈ延长至２７ｈ，卵裂率和囊胚发育率均没有显著变化，卵母     

Phase I evaluation of CCNU (lomustine) in tumor-bearing cats

1-(2-Chloroethyl)3-cyclohexyl-1-nitrosourea (CCNU) is an alkylating agent in the nitrosourea subclass. A prospective evaluation of CCNU was done to determine the maximally tolerated dosage of CCNU in tumor-bearing cats. Response data were obtained when available. Twenty-five cats were treated with CCNU at a dosage of 50-60 mg/m3 body surface area. Complete hematologic data were available for 13 cats. Neutropenia was the acute dose-limiting toxicity. The median neutrophil count at the nadir was 1,000 cells/microL (mean, 2,433 cells/microL; range, 0-9,694 cells/microL). The time of neutrophil nadir was variable, occurring 7-28 days after treatment, and counts sometimes did not return to normal for up to 14 days after the nadir. Based on these findings, a 6-week dosing interval and weekly hematologic monitoring after the 1st treatment with CCNU are recommended. The nadir of the platelet count may occur 14-21 days after treatment. The median platelet count at the nadir was 43,500 cells/microL. No gastrointestinal, renal, or hepatic toxicities were observed after a single CCNU treatment, and additional studies to evaluate the potential for cumulative toxicity should be performed. Five cats with lymphoma and 1 cat with mast cell tumor had measurable responses to CCNU. Phase II studies to evaluate antitumor activity should be completed with a dosing regimen of 50-60 mg/m3 every 6 weeks.     

Prevalence of gastric lesions in racing Alaskan sled dogs

Human and equine athletes are reported to have a high prevalence of gastric disease, and anecdotal evidence suggests a similar phenomenon applies to racing sled dogs. To investigate the prevalence of gastric disease in racing sled dogs, we conducted 2 gastroscopy studies on dogs competing in the annual Iditarod Sled Dog Race. A pilot study of dogs that were either dropped from the 2000 Iditarod Sled Dog Race because of illness or that finished the race indicated that, approximately 5 days after competing, 10 of 28 dogs (35%) had endoscopic evidence of gastric ulceration, erosion, or hemorrhage. The next year, an endoscopic study of 73 dogs participating in the 2001 Iditarod race was performed in order to evaluate a larger population of dogs. Data from 70 of these dogs could be used; 34 (48.5%) had ulceration, erosion, gastric hemorrhage, or some combination of these findings. When this group of 70 dogs was compared retrospectively to a control group of 87 dogs presented to the Texas A&M University (TAMU) Veterinary Medical Teaching Hospital, the Iditarod sled dogs had a significantly higher prevalence (P = .049) of gastric lesions. These findings suggest that, similar to athletes of other species, elite canine athletes have an increased prevalence of gastric disease compared to the canine population at large.     

Furosemide continuous rate infusion in the horse: evaluation of enhanced efficacy and reduced side effects

Continuous rate infusion (CRI) of furosemide in humans is considered superior to intermittent administration (IA). This study examined whether furosemide CRI, compared with IA, would increase diuretic efficacy with decreased fluid and electrolyte fluctuations and activation of the renin-angiotensin-aldosterone system (RAAS) in the horse. Five mares were used in a crossover-design study. During a 24-hour period, each horse received a total of 3 mg/kg furosemide by either CRI (0.12 mg/kg/h preceded by a loading dose of 0.12 mg/kg IV) or IA (1 mg/kg IV q8h). There was not a statistically significant difference in urine volume over 24 hours between methods; however, urine volume was significantly greater after CRI compared with IA during the first 8 hours ([median 25th percentile, 75th percentile]: 9.6 L [8.9, 14.4] for CRI versus 5.9 L [5.3, 6.0] for IA). CRI produced a more uniform urine flow, decreased fluctuations in plasma volume, and suppressed renal concentrating ability throughout the infusion period. Potassium, Ca, and Cl excretion was greater during CRI than IA (1,133 mmol [1.110, 1,229] versus 764 mmol [709, 904], 102.7 mmol [96.0, 117.2] versus 73.3 mmol [65.0, 73.5], and 1,776 mmol [1,657, 2.378] versus 1,596 mmol [1,457, 1,767], respectively). Elimination half-lives of furosemide were 1.35 and 0.47 hours for CRI and IA, respectively. The area under the excretion rate curve was 1,285.7 and 184.2 mL x mg/mL for CRI and IA, respectively. Furosemide CRI (0.12 mg/kg/h) for 8 hours, preceded by a loading dose (0.12 mg/kg), is recommended when profound diuresis is needed acutely in horses.     

Internal Parasites and Haematological Values in Cattle Slaughtered in Buea Subdivision of Cameroon

Tropical Animal Health and Production -     

The Epidemiology of Gastrointestinal Nematodes of Dairy Cattle in Central Kenya

The epidemiology of H. placei and of other gastrointestinal nematodes in yearling dairy cattle was examined on two farms in Kiambu District, central Kenya during each of 13 one-month periods from April 1993 to April 1994. On each farm, 32 newly weaned dairy calves were given a single dose of albendazole and then placed on experimental pastures. Twelve of the animals were designated for bi-monthly slaughter (n = 2) and analysis of worm population characteristics and 20 were designated for blood and faecal collection and for weighing. Two parasite-free tracer calves were grazed alongside the weaner calves each month throughout the study period and were also slaughtered for analysis of worm populations. Faecal egg counts, haematological and serum pepsinogen determinations, herbage larval counts, and animal live weight changes were recorded monthly. The study revealed that Haemonchus placei, Trichostrongylus axei, Cooperia spp. and Oesophagostomum radiatum were responsible for parasitic gastroenteritis and that H. placei was the predominant nematode present in the young cattle on both farms. Faecal egg counts from resident cattle and necropsy worm counts revealed that pasture larval levels were directly related to the amount of rainfall. The total worm burdens in the animals were highest during the rainy season (March–June and October–December) and lowest during the dry seasons (July–September and January–February). The very low recovery of immature larvae of H. placei from the tracer calves indicated that arrested development is not a feature of the life cycle of this parasite in central Kenya. The maintenance of the parasite population depended on continuous cycling of infection between the host and the pasture. The agroclimatic conditions of the study area were such that, in general, favourable weather conditions for the development and survival of the free-living stages of gastrointestinal nematodes existed all year round.     

Diversity of Ectoparasites in Sheep Flocks in Sa~o Paulo,Brazil

The occurrence of ectoparasites in sheep flocks is frequently reported but seldom quantified. Sheep production used to be a predominantly family activity in the state of Sa~o Paulo (Brazil), but it began to become a commercial activity in the past decade. Thus, information about the ectoparasites existing in sheep flocks has become necessary. The present data were obtained by means of questionnaires sent to all sheep breeders belonging to the `Associaça~o Paulista de Criadores de Ovinos' (ASPACO; Sa~o Paulo State Association of Sheep Breeders). Response reliability was tested by means of random visits paid to 10.6% of the respondents. Most of the properties (89.5%) reported the presence of one or more ectoparasites. Screw-worm (Cochliomyia hominivorax) was the most frequent ectoparasite (72.5%), followed by bot fly larvae (Dermatobia hominis, 45.0%), ticks (Amblyomma cajennense) and Boophilus microplus, 31.3%) and finally lice (Damalinia ovis, 13.8%). Combined infestations also occurred, the most common one being screw-worm with bot fly larvae (36.0%) followed by bot fly larvae with ticks (13.9%), screw-worm with ticks (9.3%), bot fly larvae with lice (6.9%), and ticks with lice (5.0%). The most common triple combination was screw-worm, bot fly larvae and ticks (12.8%). Breeds raised for meat or wool were attacked by bot fly larvae and ticks more often than other breeds. Lice were only absent from animals of indigenous breeds. The relationships among these ectoparasites are discussed in terms of sheep breeds, flock size, seasonality and the ectoparasitic combinations on the host.     

Chemotaxis of macrophage in inflammation

Our particular attention in this article was given to natural mediators for macrophages isolated from the sites of tissue injury. A number of chemotactic factors, which may satisfy many criteria making them acceptable as inflammatory leucocyte chemotactic factors, has been separated. Among them, our laboratory has isolated three macrophage (monocyte) chemotactic factors (MCF-a, -b and -c). Their purification, characterization and functional specificity are discussed.     

Evaluation of biohydrogenation rate of canola vs. soya bean seeds as unsaturated fatty acids sources for ruminants in situ

An experiment was conducted to study disappearance of C14 to C18 fatty acids, lag times and biohydrogenation (BH) rates of C18 fatty acids of ground soya bean and canola seeds in situ. Three ruminally fistulated Dallagh sheep were used to determine ruminal BH of unsaturated fatty acids (UFAs). Differences in the disappearance of fatty acids through the bags and lag times were observed between the oilseeds. We saw that the longer the incubation time of the oilseeds in the rumen, the lower the content of C18:2 and C18:3. Significantly higher lag times for both C18:2 and C18:3 were observed in ground canola compared to ground soya bean. BH rates of C18:2 and C18:3 fatty acids in soya bean were three times higher than those of canola. These results suggest that the fatty acid profile of fat source can affect the BH of UFAs by rumen micro‐organisms. So that UFAs of canola had higher ability to escape from ruminal BH. It seems that fatty acid profile of ruminant products is more affected by canola seed compared to soya bean seed.     

Serum biochemical values and mineral element contents of tissues in yaks

Serum biochemical values and the wet weight to dry weight ratios of tissues were determined in yaks in Shandan county of Gansu province. The liver, kidney, heart and muscle contents of seven elements in yaks were also analysed. Most serum biochemical values were similar to those of cattle, camels and sheep, but the calcium concentration was considerably above the normal range for other ruminants. The liver contained the highest concentrations of copper, manganese and cobalt and the kidney of selenium, iron and calcium.Abbreviations Alb albumin - ALT alanine aminotransferase - AST aspartate aminotransferase - BUN blood urea nitrogen - Chol cholesterol - Glob globulin - -GT -glutamyl transpeptidase - IP inorganic phosphorus - TP total protein