GSN0对E.tenella内LDH、G-6-PD、ACO和SOD活性的影响

亚硝基谷胱甘肽(GSNO)对鸡柔嫩艾美耳球虫(E.tenella)卵囊的孢子生殖具有不可逆地抑制作用,但其作用机理尚不清楚.本研究采用聚丙烯酰胺凝胶电泳来分析GSNO对E.tenella卵囊内与糖代谢有关的乳酸脱氢酶(LDH)、葡萄糖-6-磷酸脱氢酶(G-6-PD)、乌头酸酶(ACO)以及GSNO对卵囊内与抗氧化有关的超氧化物歧化酶(SOD)活性的影响,电泳后经特异染色显示酶活性.结果表明:GSNO处理的未孢子化与孢子化卵囊中均有LDH、G-6-PD、ACO及SOD活性,这提示外源性NO对球虫孢子生殖的抑制并没有灭活卵囊中LDH、G-6-PD、ACO和SOD的活性或NO对这些酶的作用是可逆的.     

GSNO对鸡球虫LDH和G-6-PD活性的影响

资料显示,GSNO对鸡球虫卵囊有较强的抑制作用,可以完全阻止E.tenella的孢子生殖,球虫卵囊的孢子生殖是一有氧呼吸过程,为了探索GSNO对球虫卵囊呼吸有关酶活性的影响,本实验利用PAGE法分析了与呼吸关系密切的乳酸脱氢酶(LDH)和葡萄糖-6-磷酸脱氢酶(G-6-PD)。电泳后活性染色结果表明,GSNO处理的未孢子化和孢子化卵囊的LDH和G-6-PD条带与对照组的没有明显区别,这提示LDH和G-6-PD的活性没有受到GSNO的影响。     

巨型艾美耳球虫(Eimeria maxima)地理株研究Ⅲ.11株巨型艾美耳球虫的同工酶分析

采用聚丙烯酰胺凝胶垂直板电泳(PAGE)技术,对11株Ei meria maxima和1株E.tenella的孢子化卵囊,进行了乳酸脱氢酶(LDH)、葡萄糖磷酸异构酶(GPI)、葡萄糖-6-磷酸脱氢酶(G6PD)、葡萄糖磷酸变位酶(PGM)和苹果酸脱氢酶(MDH)的同工酶分析。试验结果显示,E.maxima和E.tenella在LDH、GPI、G6PD、PGM和MDH的酶谱上有明显差异,而在11株E.maxima之间无差异,表明E.maxima的酶变异相当保守。     

柔嫩艾美耳球虫孢子发育三种阶段的同工酶研究

采用聚丙烯酰胺凝胶电泳(PAGE)研究柔嫩艾美耳球虫(Eimeria tenella)未孢子化卵囊,孢子化卵囊、孢子囊的乳酸脱氢酶(LDH)、葡萄糖-6-磷酸脱氢酶(G6PD)、异柠檬酸脱氢酶(IDH)、6-磷酸萄萄糖酸脱氢酶(6PGD)、磷酸葡萄糖变位酶(PGM)、碱性磷酸酶(ALP)和葡萄糖磷酸异构酶(GPI)等7种酶的同工酶,除IDH外,6种酶均显示出初步结果,孢子化卵囊和孢子囊6种酶的同工酶完全一致,未孢子化卵囊的LDH、G6PD、PGM、ALP、GPI 5种酶同工酶与孢子化卵囊、孢子囊一致,未孢子化卵囊的6PGD同工酶与孢子化卵囊、孢子囊呈现明显差异、讨论了球虫在孢子化过程中,一些酶的活性或同工酶发生变化的情况。     

一氧化氮对球虫(E.tenella)卵囊孢子生殖的抑制特性

本研究利用NO供体以及几种抗氧化剂和NO清除剂来分析外源性NO对E.tenella卵囊孢子生殖的抑制特性.试验结果表明:S-亚硝基硫醇类NO供体(SNAP和GSNO)对卵囊的抑制作用具有明显的剂量-效应荚系,8 mmol/L的SNAP和GSNO均可完全抑制新鲜卵囊的孢子生殖;KCN对卵囊的抑制呈现为剂量-效应关系,但NaN3没有抑制作用.维生素C、甘露醇和水杨酸钠等抗氧化物质以及FeSO4和DTT均不能消除GSNO对卵囊孢子生殖的抑制作用.K2Cr2O7和KMnO4均会明显抑制GSNO对卵囊的作用.NO清除剂血红蛋白(Hb)能够消除NO供体对卵囊的抑制作用,并呈现剂量-效应关系.     

妥曲珠利对柔嫩艾美耳球虫作用峰期的初步研究

E．tenella球虫又称盲肠球虫，主要侵害鸡的盲肠，一般感染15～35日龄的雏鸡。一旦鸡吞食了E．tenella球虫卵囊，由于球虫的高繁殖性能，随之有众多的卵囊排出并污染环境，导致球虫病暴发。E．tenella球虫发育过程包括3个生殖阶段，孢子生殖在外界环境中进行，其它两种生殖阶段在鸡体内完成。随鸡粪便排到外界环境中的球虫卵囊称为未孢子化卵囊，不具有感染力。     

亚硝基谷胱甘肽对孢子化卵囊、孢子囊活力及致病力的影响

以亚硝基谷胱甘肽(S-nitroso-glutathione,GSNO)处理柔嫩艾美耳球虫(Eimeria tenella)卵囊和孢子囊,通过检测子孢子的脱囊情况以及卵囊和孢子囊的致病力来探讨GSNO对E.tenella卵囊和孢子囊活力的影响。体外脱囊试验结果表明,GSNO处理孢子囊的脱囊率只有1.01%~1.48%,而GSNO处理的孢子化卵囊的脱囊率为45.73%~63.90%。接种试验表明,各接种组鸡均发生了明显的临床球虫病,但与攻毒对照组相比,只有GSNO处理的孢子囊组鸡的发病情况有所减轻,这说明GSNO能明显抑制孢子囊的脱囊,并能在一定程度上减轻其致病力,而对孢子化卵囊无明显抑制作用。     

外源性一氧化氮对鸡柔嫩艾美尔球虫的作用

用酸化亚硝酸钠作为一氧化氮(NO)供体处理新收取的柔嫩艾美尔球虫未孢子化和已孢子化卵囊,来探讨外源性NO对球虫卵囊的抑杀作用。另外通过内服亚硝基谷胱甘肽(GSNO)试验来观察外源性NO对球虫病的防治作用。结果表明,酸化亚硝酸钠溶液(pH=1)对球虫卵囊的孢子生殖具有明显的抑制作用,20mmol/L酸化亚硝酸钠溶液对卵囊孢子生殖的抑制率可达100%;但不经酸化的亚硝酸钠溶液和相同pH的盐酸溶液对卵囊的孢子生殖没有明显影响。经20mmol/L以上浓度酸化亚硝酸钠处理的孢子化卵囊也失去了对雏鸡的致病力,但直接内服GSNO溶液则没有明显的抗球虫作用。这提示外源性NO对鸡柔嫩艾美尔球虫未孢子化和孢子化卵囊均有抑杀作用,但直接内服没有明显的抗球虫病作用。     

新疆阿拉尔某鸡场球虫种类调查

为了了解阿拉尔市某鸡场鸡球虫种类及感染情况,采用群体采样法分别采集新鲜粪便并检出阳性粪便,对检出的阳性粪便采用饱和盐水漂浮法和离心沉淀法进行卵囊分离;显微镜下观察卵囊、最短孢子化时间、最小潜在期和病理变化,进行虫种鉴定。最后得出该鸡场感染有堆型艾美耳球虫（Eimeria acervulina）、巨型艾美耳球虫（E．maxima）和柔嫩艾美耳球虫（E．tenella）,该鸡场球虫感染严重,且为混合感染。     

柔嫩艾美耳球虫Etron2基因在不同发育阶段转录水平差异的分析

为研究柔嫩艾美耳球虫不同发育时期Etron2基因的转录水平差异,选取E.tenella生活史中的未孢子化卵囊、孢子化卵囊、子孢子、第二代裂殖子,分别提取总RNA,反转录为cDNA;选择Etactin作为参照基因,Etron2为目的基因,设计实时荧光定量PCR的引物,分析在E.tenella生活史中不同发育阶段Etron2转录水平的差异。结果显示Etron2在未孢子化卵囊中转录水平最高,然后分别是裂殖子、孢子化卵囊和子孢子。在柔嫩艾美耳球虫发育过程中,Etron2基因可能在未孢子化卵囊阶段被转录储备,随着宿主细胞的刺激,得以大量表达,在入侵过程中发挥作用。     

The effects of nitric oxide donors on the sporulation of Eimeria tenella oocysts

Nitric oxide (NO) is involved in many physiological processes including regulation of blood pressure, immune response, and neural communication. The effects of NO donors on the sporogony of Eimeria tenella oocysts and the blocking effects of NO scavenger, hemoglobin (Hb), against NO donors were investigated in this study. The results showed that the sporulation of oocysts could be completely inhibited by acidic sodium nitrite (NaNO2), S-nitroso-glutathione (GSNO) or S-nitroso-N-acetyl-dl-penicillamine (SNAP) in profound dose-dependent and time-dependent manners, and the inhibiting process was irreversible and not affected by feces of chickens. However, not all NO donors in this study, such as sodium nitroprusside (SNP), demonstrated above-mentioned effect. The NO scavenger Hb could significantly prevent the detrimental effects of NO donors on the sporogony of oocysts in a dose-dependent manner. At present, the inhibiting mechanism of NO donors on the sporulation of oocysts was still unclear.     

柔嫩艾美耳球虫耐药株与鸡3种球虫的同工酶的研究

对不同离子载体抗生素具有抗药性的柔嫩艾美耳球虫和药物敏感的柔嫩区美耳球虫、布氏匀美耳球虫和堆型艾耳球虫的孢子化卵囊,采用聚安凝胶垂直板电泳,进行了乳酸脱氢酶（ＬＤＨ）、葡萄糖磷酸异构酶（ＧＰＩ）和葡萄糖－６－磷酸脱氢酶（Ｇ６ＰＤ）等同工酶华不同离子载体抗生素具有抗药性的柔嫩艾美耳球虫和药物同工酶酶谱可以反应出球虫种间差异;而柔嫩艾耳球虫抗性虫株的ＬＤＨ酶谱是２条带,敏感虫株是３条带,抗性株比敏感株     

环境因素对球虫卯囊孢子化的影响

探讨环境因素对球虫卵囊孢子化的影响,为切断鸡球虫病的感染途径提供依据。本实验将未孢子化球虫卵囊分别置于高温干燥、高温潮湿、室温三个不同环境中处理后,再按常规方法培养,并测其孢子化率。结果表明:80℃高温干燥环境中放置15 min以上,80℃高温潮湿环境中放置30 min以上,室温环境中放置1 d以上,卵囊均已完全丧失孢子化能力。     

Anticoccidial activity of 9-(2-chloro-6-fluorobenzyl)-6-methylaminopurine and 9-(2-chloro-6-fluorobenzyl)-6-dimethylaminopurine against Eimeria tenella, E. acervulina, and E. maxima in battery trials

In a series of battery experiments utilizing 9-day-old White Leghorn male chicks, 9-(2-chloro-6-fluorobenzyl)-6-methylaminopurine (VM 6387) and 9-(2-chloro-6-fluorobenzyl)-6-dimethylaminopurine (VM 6736) showed remarkable anticoccidial activity against Eimeria tenella, E. acervulina, and E. maxima. The effective dose range was estimated from the results of the efficacy test against E. tenella on the basis of improvement of body weight gain and clinical signs of infection. The tests included dietary levels of 60-100 ppm of VM 6387 and 70-110 ppm of VM 6736. Both compounds gave good protection against E. acervulina and E. maxima at the same dose range. The sporulation of oocysts obtained from cecal contents of birds fed lower levels of VM 6387 was inhibited. Studies of the effects of VM 6387 on stages of the E. tenella life cycle demonstrated that the compound possesses remarkable activity at 1-5 days post-infection corresponding to the period of schizogony, as well as prolonged activity thereafter, when the parasite was undergoing gametogenesis.     

Evaluation of interspecific hybrids of the chicken, guinea fowl, and Japanese quail for innate resistance to coccidia

Experimental chicken/guinea fowl hybrids, guinea fowl, and chickens were orally inoculated with Eimeria acervulina or E. tenella, which are specific for chickens, or with E. grenieri, which is specific for guinea fowl. No intact oocysts were found in feces within 24 hr of inoculation, suggesting that excystation occurred in the normal and abnormal hosts. No oocysts were found in the feces of hybrids during a 9-day postinoculation period. The guinea fowl passed oocysts of guinea fowl coccidia (E. grenieri) but not those of chicken coccidia, and the chickens passed oocysts of chicken coccidia (E. acervulina and E. tenella) but not those of guinea fowl coccidia. Some asexual development (schizogony) occurred in hybrids inoculated with E. tenella, but sexual development (gametogony) did not. In contrast, quail/chicken hybrids became infected with oocysts of chicken coccidia (E. acervulina, E. tenella, and E. maxima) and quail coccidia (E. bateri) and passed a few oocysts during the normal patent period; control chickens and quails became heavily infected with oocysts of chicken and quail coccidia, respectively.     

青蒿素与柳氮磺吡啶对柔嫩艾美尔球虫感染的疗效观察

为了探讨青蒿素（artemisinin,ART）和柳氮磺吡啶（sulfasalazine,SASP）单独给药与联合给药对鸡球虫病的防治效果。试验选用50只1日龄黄羽肉鸡,正常饲养至1周龄时分为5组,2周龄时除空白组外,ART组、SASP组、ART＋SASP组和感染对照组分别以105个/羽的剂量接种柔嫩艾美尔球虫（Eimeria tenella,E.tenella）孢子化卵囊,各给药组于攻虫前3d开始给药至试验结束。试验结束后对各组的抗球虫指数（anticoccidial index,ACI）、卵囊的孢子化率、血清NO含量、脏器指数及血清生化指标等数据进行统计。结果表明,ART、SASP以及两者联合用药均能显著提高鸡群的ACI,并使卵囊的孢子化率降低,缓解了球虫感染所致的NO水平升高,且联合用药效果更明显。这提示ART与SASP联合应用在E.tenella感染的防治中具有增效作用。各组鸡群之间血清生化参数无显著性差异,说明该2种药物均无明显毒副作用。     

球虫消毒剂的筛选及氨水对艾美耳球虫卵囊抑杀条件的优化

为了筛选有效的球虫消毒剂,本研究将柔嫩艾美耳球虫卵囊分别与16种常用消毒剂和1种高效抗球虫药物溶液（250 mg/L妥曲珠利溶液）混合作用2 h然后在2.5%重铬酸钾溶液中孢子化培养,或混合作用4 h后接种14日龄雏鸡,根据卵囊孢子化率、孢子化抑制率、每克粪便卵囊数量（oocysts per gram of feces,OPG）、病变计分和血便情况评价消毒剂和抗球虫药物抑杀球虫卵囊效果。结果显示,8%氨水对未孢子化卵囊和孢子化卵囊均表现为100%的抑杀效果;250 mg/L妥曲珠利、3%甲醛、4%苯酚、0.5%过氧乙酸、5% NaOH+10% NaCl和5% NaOH对球虫卵囊活性具有一定的抑制作用,孢子化抑制率超过10%,但是对孢子化卵囊抑杀作用不明显;而剩余10种常用消毒剂对球虫卵囊没有明显的抑杀作用。氨水抑杀球虫卵囊条件优化的试验结果显示,1%~8%氨水与未孢子化卵囊作用1 h以上,均可100%抑杀球虫卵囊活性。粪便干扰试验结果显示,粪便不影响2%和4%氨水对球虫卵囊的抑杀效果。本研究结果说明常用消毒剂和抗球虫药物不能有效杀灭球虫卵囊的活性,而氨水是一种高效球虫消毒剂。本研究结果可为开发新型高效球虫消毒剂和球虫病控制提供参考。     

Three Enzymes Newly Identified from the Genus Eimeria and Two More Newly Identified from E. maxima,Leading to the Discovery of Some Aliphatic Acids with Activity against Coccidia of the Domesticated Fowl

Nine enzymes were detected in sporulated oocysts of Eimeria tenella and E. maxima, parasites of the domesticated fowl (Gallus gallus). Three enzymes, hydroxybutyrate dehydrogenase, alanine aminotransferase and -glutamyltransferase, all identified for the first time in Eimeria of fowl, occurred both in E. tenella and in E. maxima. The remaining enzymes assayed had previously been found in various Eimeria species of fowl, although creatine kinase and glutamate dehydrogenase were hitherto unknown from E. maxima. The three enzymes newly recorded from Eimeria of fowl are of interest as potential genetic markers, and also as potential chemotherapeutic targets. The discovery of hydroxybutyrate dehydrogenase led to the demonstration of anticoccidial activity by some aliphatic acids. The paper also includes a list of the enzymes detected in Eimeria of fowl in previous studies.     

Effects of clopidol on sporulation and infectivity of Eimeria tenella oocysts

Feed additive anticoccidials currently used in Japan were examined for possible effects on oocyst sporulation of Eimeria tenella. Monensin, salinomycin, lasalocid, amprolium plus ethpabate, amporolium plus ethopabate plus sulfaquinoxaline, clopidol, or nicarbazin were given to chickens continuously via the feed at the recommended use level or one-half of that level. Oocysts discharged in feces 7-8 days post inoculation (PI) were collected and aerated for sporulation. Low sporulation rate was noted, when clopidol at 62.5 mg kg-1 was given from 4 to 7 days PI. These oocysts were as infective as oocysts from controls, based on weight gain, feed efficiency, gross lesion score of cecae, and oocyst count 7 days PI. The results of the study indicated that the second schizogony and gametogony are vulnerable to clopidol, as evidenced by oocyst sporulation, but infectivity of these sporulated oocysts was not affected.