鸡传染性支气管炎病毒QX基因型S1蛋白重组乳酸菌的构建及小鼠免疫应答

为探究鸡传染性支气管炎病毒(IBV)S1蛋白重组乳酸菌对小鼠免疫应答情况,本研究将IBV CH/LN/2019毒株的S1基因进行密码子优化,构建重组质粒pNZ8149-S1,电转至乳酸乳球菌NZ3900中,应用Western blot及间接免疫荧光试验评价重组乳酸乳球菌表达水平,免疫SPF小鼠,通过间接ELISA方法分析小鼠特异性抗体、CD4⁺、CD8⁺和细胞因子(IL-4和IFN-γ)分泌情况;结果显示,成功构建重组乳酸乳球菌pNZ8149-S1/NZ3900并表达目的蛋白;小鼠免疫试验结果显示,免疫组小鼠IgG抗体水平均高于pNZ8149/NZ3900、NZ3900和PBS组(P<0.05);特异性sIgA抗体水平显著高于对照组(P<0.01);此外,重组乳酸乳球菌能诱导小鼠产生较高水平的IL-4和IFN-γ(P<0.05)。结果表明,重组乳酸乳球菌pNZ8149-S1/NZ3900可以有效刺激小鼠机体产生体液免疫和细胞免疫,为IBV新型口服疫苗的研制提供了理论参考。     

表达牛乳铁蛋白肽的重组乳酸乳球菌对雏鸡抗鸡白痢沙门氏菌感染的效果研究

为探索表达牛乳铁蛋白肽的重组乳酸乳球菌pAMJ399-LFBA/MG1363对雏鸡抗鸡白痢沙门氏菌感染效果的影响,本实验将该重组乳酸乳球菌饲喂1日龄雏鸡,连续饲喂14 d,并设立空白对照组,第7 d时经灌服鸡白痢沙门氏菌同时感染实验组和对照组,连续观察7 d;分别于攻菌前以及攻菌后的不同天数采集血清及盲肠黏液,检测结果显示攻菌后饲喂重组乳酸乳球菌组雏鸡血清中IgG以及黏液中SIgA的含量极显著高于空白对照组。对各组雏鸡的心脏、肝脏、脾脏及十二指肠进行病理组织学检查,结果显示饲喂重组乳酸乳球菌组各器官的病变情况相对较轻,而且雏鸡盲肠扁桃体中非特异免疫信号通路中相关分子TLR4、My D88、NF-κB、TNF-α、IL-8、IL-6等的m RNA表达相对较高。以上结果表明,表达牛乳铁蛋白肽的重组乳酸乳球菌在抗鸡白痢沙门氏菌感染的过程中可以起到保护雏鸡免受感染的作用。     

产肠毒素大肠杆菌STa突变体、LTB和STb融合蛋白在乳酸菌中组成型分泌表达及其免疫原性分析

为获得产肠毒素大肠杆菌(ETEC)抗原STa突变体(mSTa)、LTb和STb融合蛋白在乳酸菌表达系统中组成型分泌表达,本研究将mSTa、LTb、STb三价抗原融合基因,克隆于p23启动子-USP45分泌信号肽之后,插入到乳酸乳球菌表达载体pTX8048中,构建了重组表达载体pTX-sls,将其电转化到宿主菌乳酸乳球菌L.lactisNZ9000中进行表达,应用westernblot、ELISA方法鉴定蛋白表达情况。将重组乳酸乳球菌pTX-sls/NZ9000口服免疫BALB/c小鼠,分别测定了免疫后不同时间血清中特异性IgG、粪便中特异性的sIgA水平以及血清的中和活性;采用MTT法检测免疫小鼠脾淋巴细胞增殖情况,流式细胞术检测Th细胞免疫类型。结果显示,目的蛋白mSTa-LTB-STb以分泌形式组成型表达,可被阳性血清所识别。在免疫小鼠血清和粪便中均可检测到特异性IgG、sIgA,血清抗体具有一定的中和毒素作用。结果表明,该重组乳酸乳球菌具有作为口服疫苗的潜在应用价值,本研究为研制ETEC乳酸菌活载体口服疫苗奠定了基础。     

鸭不同肠段内菌群结构分析与拟杆菌分布研究

旨在分析健康鸭十二指肠、空肠、回肠和盲肠内容物菌群组成、多样性特征以及拟杆菌分布。本研究选择健康高邮鸭20只,公母各半,70日龄时无菌采集十二指肠、空肠、回肠和盲肠内容物,提取肠道内容物细菌基因组,利用IonS5TMXL平台进行高通量测序,分析肠道内容物菌群结构与丰度特征以及拟杆菌的分布。结果表明,十二指肠、空肠内容物菌群丰度显著高于回肠和盲肠内容物（P<0.05）,回肠内容物菌群多样性最低;十二指肠和空肠内菌群群落结构较为相似,与回肠,特别是盲肠的相似度较小。健康鸭肠内优势菌门为拟杆菌门（Bacteroidetes）、厚壁菌门（Firmicutes）、变形菌门（Proteobacteria）和梭杆菌门（Fusobacteria）,上述菌门在各肠段内容物中相对丰度不同;不同肠段内容物中定植了不同差异微生物物种,十二指肠、空肠、回肠和盲肠内容物中差异菌门分别是变形菌门、放线菌门（Actinobacteria）、厚壁菌门和拟杆菌门。鸭肠道中共分析到28种拟杆菌种,其中B.acidifaciens、B.barnesiae、B.caccae、B.caecicola、B.coprocola、B.sp和B.luti在盲肠内容物中显著聚类,且B.caecigallinarum、B.plebeius和B.barnesiae在鸭盲肠中优势定植。结果显示,鸭肠段空间显著影响了其内容物中菌群丰度与多样性,不同肠段内定植了差异的优势微生物物种,这可能与肠段小环境以及功能一致,在盲肠内容物中优势定植拟杆菌,推测可能与鸭盲肠生理生化功能相关。     

蒲公英和板蓝根对LPS诱导的小鼠肠黏膜屏障损伤的调理机制研究

抗生素杀死细菌释放的内毒素常对动物消化道造成损伤，为筛选能有效净化肠道内毒素从而减轻肠道损伤的中草药，本研究对小鼠腹腔注射脂多糖(LPS)建立肠黏膜屏障损伤模型，随后将小鼠分为蒲公英处理组(P组，80 mg/kg,n=7)，板蓝根处理组(B组，210 mg/kg,n=7)和PBS处理组(G组，0.01 mol/L PBS,n=7)，分别于处理后0、3 d和7 d收集各组小鼠的十二指肠、空肠、回肠和结肠组织，利用小鼠LPS检测试剂盒检测各组小鼠各个时间点肠道组织中的LPS，结果显示，与治疗前比较，蒲公英治疗3 d和7 d小鼠的十二指肠、回肠和结肠组织中LPS的含量显著降低(P<0.05)；板蓝根治疗7 d小鼠的十二指肠、回肠和空肠LPS的含量显著降低(P<0.05)。采用ELISA方法检测各组小鼠各肠道组织中分泌型IgA (sIgA)抗体水平各时间点的变化，结果显示，与治疗前比较，治疗3 d和7 d蒲公英和板蓝根组小鼠空肠和回肠组织s IgA抗体的水平显著升高(P<0.05)。采集治疗0、3 d和7 d各组小鼠肠道组织进行组织病理学观察，结果显示，对照组LPS诱导小鼠主...     

乳酸片球菌的抑菌能力及其对断奶仔猪肠道菌群及黏膜免疫的影响

体外抑菌试验采用牛津杯法,动物试验选用90头21日龄断奶仔猪(杜长大),随机分为空白组、菌粉组、上清液组和发酵液组,试验期49 d,旨在研究乳酸片球菌SKL-03的抑菌性能及其对断奶仔猪肠道菌群及黏膜免疫的影响。试验结果显示:乳酸片球菌SKL-03菌粉、上清液、发酵液对3种常见致病菌均有显著的抑菌效果;动物试验结果显示:相比空白组,饲粮中添加乳酸片球菌SKL-03菌粉、上清液、发酵液可降低盲肠、结肠中的大肠杆菌和沙门氏菌的数量,提高乳杆菌数量和十二指肠、空肠黏膜中SIgA含量。综上所述,乳酸片球菌SKL-03具有改善断奶仔猪肠道菌群,提高小肠黏膜免疫功能的效果。     

鸭瘟病毒强毒感染鸭肠道菌群的分子解析

应用ERIC-PCR法对鸭瘟病毒强毒株经皮下和口服感染20日龄鸭的十二指肠、空肠、回肠、盲肠和直肠中细菌菌群的结构多样性和动态变化进行了检测.结果表明:对照鸭肠道内的菌群结构相对稳定,其中十二指肠和空肠的ERIC-PCR条带最少,盲肠最多;皮下感染24 h和口服48 h前,各肠段ERIC-PCR扩增条带没有明显变化,皮下感染48 h的鸭盲肠、口服感染72 h的鸭盲肠和回肠条带数量明显增加,随着感染时间的延长,条带数呈逐渐减少的趋势,死亡鸭各肠段的条带最少.对ERIC-PCR扩增条带中主带的测序结果表明,大肠杆菌、志贺菌、沙门杆菌等需氧菌或兼性厌氧菌成为皮下感染鸭的直肠、口服感染鸭的盲肠(回肠和直肠)的优势菌群;口服感染鸭的回肠中一个未知菌成为优势菌群.     

斯哒米纳微生态制剂对肉仔鸡生长性能和肠道环境的影响

试验主要探讨微生态制剂(斯哒米纳)对肉仔鸡生产性能和肠道酶活、肠道pH值、肠道微生物的影响。试验结果表明,在生长性能方面,1~42日龄日采食量提高5.2%,日增重提高7.5%,料肉比降低2.4%,死淘率降低23.2%;21日龄检测显示:肠道酶活方面,十二指肠中,蛋白酶、淀粉酶和脂肪酶分别高于对照组22.1%、57.8%和64.8%;空肠中,蛋白酶、淀粉酶和脂肪酶高于对照组26.4%、41.2%和22.6%;肠道pH值方面:十二指肠、空肠、回肠和盲肠pH值分别降低4.2%、1.4%、2.3%和3.7%;肠道微生物方面:空肠中大肠杆菌含量降低14.9%,乳酸杆菌含量升高23.1%,双歧杆菌含量增加18.5%;盲肠中大肠杆菌含量降低14.9%,乳酸杆菌含量升高18.5%,双歧杆菌含量增加17.2%。42日龄检测显示:肠道酶活方面:十二指肠中,蛋白酶、淀粉酶和脂肪酶分别高于对照组24.9%、69.6%和61.8%;空肠中,蛋白酶、淀粉酶和脂肪酶分别高于对照组30.6%、59.9%和43.3%;肠道pH值方面:十二指肠、空肠、回肠和盲肠pH值分别降低1.9%、3.2%、2.1%和3.1%;肠道微生物方面:空肠中大肠杆菌含量降低29.8%,乳酸杆菌含量升高20.3%,双歧杆菌含量增加37.6%;盲肠中大肠杆菌含量降低15.9%,乳酸杆菌含量升高15.4%,双歧杆菌含量增加16.8%。     

肉鸡和肉鸭消化道食糜的状态特性研究

为研究肉鸡和肉鸭各消化道（十二指肠、空肠、回肠、盲肠）食糜状态特性,试验选用体重相似的1日龄肉鸡和肉鸭各120只,分别随机分为10笼,每笼12只进行饲喂,并分别使用普通日粮,试验时间为5 w,之后分别采集各段消化道食糜检测。结果表明：肉鸡和肉鸭盲肠中食糜干物质含量分别高于十二指肠28.9%和10.6%,高于空肠70.7%和19.1%,高于回肠44.3%和9.4%(P <0.05);肉鸡和肉鸭回肠中食糜pH分别高于盲肠8.2%和15.6%(P <0.05);肉鸡盲肠中大肠杆菌含量高于十二指肠20.0%,空肠49.4%,回肠30.3%(P <0.05),盲肠中乳酸菌含量高于十二指肠86.8%,空肠36.1%(P <0.05),肉鸭盲肠中大肠杆菌含量高于空肠66.8%,回肠31.5%(P <0.05),盲肠中乳酸菌高于十二指肠27.1%,空肠63.5%,回肠21.8%(P <0.05);对食糜流变学特性测定发现,各消化道中食糜均表现出黏度随着剪切速率的不断增大先逐渐变小后基本不变的剪切稀化趋势,经过拟合Cross-Williamson模型得到的决定系数R-...     

PEDV COE基因重组乳酸菌在小鼠体内定植规律研究

选用pNZ8149-COE NZ3900重组乳酸菌作为绿色荧光蛋白（green fluorescent protein,GFP）的表达载体,制成活菌制剂,口服免疫小鼠,以GFP基因作为标记跟踪重组菌在小鼠体内的定位,并利用荧光定量PCR技术对各部位菌定量分析。结果表明,研究构建的重组乳酸乳球菌PEDV-COE-GFP NZ3900可较长时间定植于胃肠黏膜表面,为基因工程重组乳酸菌口服疫苗的研究奠定了基础。     

癸氧喹酯干混悬剂含量测定的改进

采用高效液相色谱法测定癸氧喹酯干混悬剂的含量,在2-250μg/mL范围内,峰面积的常用对数与进样量浓度的常用对数呈良好的线性关系,R^2=1(n=5),平均回收率为99.24%~99.51%,RSD在0.05%~0.28%。此方法分析时间短,样品前处理简便、定量结果准确,重现性好,结果满意,为其质量控制提供了依据。     

猪毛色遗传的研究进展

本文概述了猪的毛色类型、猪的毛色遗传模式,着重综述了猪毛色基因分子基础的研究进展,指出存在问题并就未来发展方向做了思考。     

商会自治的基石——商会自治规范研究

在现代法律秩序中,商会自治规范是制定法的基础和必要的补充,甚至在某些方面替代了制定法;商会自治规范主要包括商会组织规范、行为规范、惩罚规范以及争端解决规范等;其效力仅及于其内部成员;商会自治规范和制定法之间存在冲突,但也存在整合的基础。     

獭兔睾丸的组织学观察

家兔作为一种实验动物 ,推动了繁殖技术的发展。本试验通过对不同年龄公獭兔的睾丸进行组织学观察、测定 ,研究精子的发生规律 ,为系统地进行繁殖生理工作提供依据。1　材料与方法选 60日龄、75日龄、90日龄 3个年龄公獭兔各5只 ,用外科手术法摘取两侧睾丸 ,放入 Bouin氏液中固定 ,二甲苯透明 ,石蜡包埋 ,切成 5～ 8μm切片 ,H.E.染色。在显微镜下观察 ,并进行定量组织学指标测定及差异性比较。2　结果和讨论2 .1　睾丸定量组织学指标的测定结果　见表 1。表 1　獭兔睾丸定量组织学指标　　　μm,个 /精细管60日龄 75日龄 90日龄曲细精管…     

中华人民共和国农业部公告 第267号

为贯彻落实《兽药生产质量管理规范》(简称《兽药GMP》),进一步推动兽药GMP实施进程,我部制定了《兽药生产质量管理规范检查验收办法》,现予公告。本公告自2003年6月1日起施行。附件:兽药生产质量管理规范检查验收办法二○○三年四月十日第一章 总则 第一条 为推动《兽药生产质量管理规范》(以下简称兽药GMP)的实施,规范兽药GMP检查验收工作,制定本办法。 第二条 农业部负责全国兽药GMP管理和检查验收工作;负责制修订兽药GMP检查验收管理规定;负责兽药GMP检查员队伍建设和监督管理工作,负责国际兽药贸易中GMP互认工作。 …     

鸡败血支原体垂直传播模式及其阻断方法

以国际标准强毒Ｒ株人工感染非免疫产蛋鸡,定时扑杀,分别从鼻窦、眶下孔、气管、肺、气囊、卵巢和输卵管分离ＭＧ,并收集感染鸡所产蛋分离ＭＧ。结果表明,人工感染４８小时后上、下呼吸道及肺已被全面感染,９６小时气囊已被感染,１２０小时输卵管已能分离到ＭＧ,卵巢始终分离不到ＭＧ。人工感染鸡自１４４小时便能在其所产蛋中分离出ＭＧ。药物治疗能在７２小时内消除感染,油乳剂苗则需２４天后逐渐降低蛋内ＭＧ分离率,药物卵内注射、种蛋药浴、高温处理均能杀死卵内ＭＧ,但以研制的种蛋浸泡剂药浴效果为最好。     

Observations of fracture of the anconeal process of the ulna of swine

Fractures of the anconeal process of 5 pigs ranging in age from 4 to 8 months were studied radiographically and histologically. Clinically, animals with a fracture of the anconeal process had a "tight," restricted gait. In pigs at 4.5 months of age, a radiolucent line through the base of the anconeal process was composed of fibrocartilage, fibrous connective tissue, and hyaline cartilage. Subperiosteal proliferation of woven bone was located along the cranial surface of the olecranon, adjacent to the base of the anconeal process. In older animals, the radiolucent line through the anconeal process contained variable amounts of fibrous connective tissue and fibrocartilage. The proliferation of subperiosteal bone at the base of the anconeal process formed a "buttress callus" which retained a radiolucent area between the callus and the proximal surface of the anconeal process. The latter region of radiolucency was continuous with the transversely oriented line that traversed the base of the anconeal process.     

Effects of size of ingestively masticated fragments of plant tissues on kinetics of digestion of NDF

Ingestively masticated fragments were collected and sized via sieving. Different sizes of esophageal masticate and ruminal digesta fragments, and ground fragments of larger masticated pieces were incubated in vitro, and undigested NDF remaining at intervals of up to 168 h of incubation was determined. The ruminal age-dependent time delay (tau) for onset of digestion of NDF was positively correlated (P < 0.004) with the mean sieve aperture estimated to retain 50% of the fragments between successive sieve apertures (MRA). Degradation rate of potentially degradable NDF (PDF) and level of indigestible NDF were not related (P > 0.10) to MRA of masticated and ground fragments. Estimates of tau were positively related to MRA, with slopes of bermudagrass < corn silage < ruminal fragments of corn silage. It was concluded that fragment size-, and consequently, ruminal age-dependent onset of PDF degradation of a mixture of different fragment sizes results in an age-dependent rate of degradation of the more rapidly degrading of two subentities of PDF. Models are proposed that assume a tau before onset of simultaneous degradation of PDF from two pools characterized as having gamma-modeled age-dependency and age-constant rates. The ruminal age-dependent pool seems to be associated with the faster-degrading pool, and its rate parameter increases with range in MRA in the population of fragments. Conceptually, the ruminal age-dependent rate parameter for PDF degradation seems to represent a composite of several effects: 1) effects of the size-dependent tau; 2) range in MRA of the population of ingestively masticated fragments; and 3) subentities of PDF that degrade via more rapid age-dependent rates compared with subentities of PDF that degrade via age-constant rates. The estimated fractional rates of ruminative comminution of ingestively masticated fragments (0.060 to 0.075/h) were of a magnitude similar to the mean fractional rates of PDF digestion (0.030 to 0.085/h), which implies that ruminative comminution may be first-limiting to fractional rate of PDF digestion. The in vivo roles of ingestive and ruminative mastication of fragments on PDF degradation must be considered in any kinetic system for estimating PDF digestion in the rumen. These results and others in the literature suggest that the rate of surface area exposure rather than intrinsic chemical attributes of PDF may be first-limiting to degradation rate of PDF in vivo.     

Comparison of 2 methods of centesis of the bursa of the biceps brachii tendon of horses

REASONS FOR PERFORMING STUDY: Centesis of the bicipital bursa using an 8.9 cm long spinal needle has been reported but the alternative of employing a 3.8 cm long hypodermic needle requires validation. OBJECTIVE: To compare the efficacy of 2 different methods of centesis of the bicipital bursa and to evaluate the usefulness of ultrasonographic imaging to determine the location of solution administered when centesis of the bursa is attempted. METHODS: For Trial 1, 6 clinicians, who had no previous experience of centesis of the bicipital bursa, attempted to inject a solution composed of an aqueous radiopaque contrast medium and physiological saline solution (PSS) into the bicipital bursae of 2/12 horses using the previously described distal approach to inject one bursa and a proximal approach to inject the contralateral bursa. The bicipital tendon and bursa were examined ultrasonographically before and after injection; and both shoulders were examined radiographically to identify the location of the medium. In Trial 2, another 6 clinicians, also with no previous experience of centesis, repeated Trial 1, using 6 horses, but the radiopaque contrast medium was mixed with air instead of PSS. RESULTS: Accuracy of centesis using the proximal approach was 39% and that of the distal approach 28%. Ultrasonographic examination of the shoulder allowed the location of solution and air to be accurately predicted in all 12 shoulders examined. CONCLUSIONS: Clinicians who have had no previous experience performing centesis of the bicipital bursa are unlikely to be successful in centesis using either approach. Radiographic examination after injecting a radiopaque contrast medium may be necessary to assess the success of centesis especially if bursal fluid is not obtained during centesis. Injecting air along with the radiopaque contrast medium provides more accurate ultrasonographic confirmation of centesis and better radiographic definition than does injection without air.