旋毛虫肌幼虫总RNA提取方法研究初报

【研究目的】探索高质量的旋毛虫(Trichinella spiralis)肌幼虫总RNA提取方法;【方法】利用Trizol试剂采用三种方法提取旋毛虫肌幼虫总RNA。(1)液氮预冷的研钵中研磨新鲜虫体至样品快融化时,加入适量的Trizol,继续研磨至液状后转移到离心管中;(2)A将新分离的虫体放到-20℃预冷的研钵中,在研磨过程中不断地添加液氮以保持虫体冷冻,之后对冷冻状的样品粉末继续操作;B把液氮冻存的新鲜旋毛虫肌幼虫取出,放到-20℃预冷的研钵中重复A的操作;(3)新鲜虫体放入匀浆器中,加适量Trizol,在冰水混合物中研磨20分钟。虫体破碎后按Trizol说明书继续抽提。最后通过凝胶电泳和紫外吸收光度值检测。【结果】用液氮和Trizol结合的方法,使用新鲜样品提取旋毛虫肌幼虫总RNA,在纯度和得率上都较为理想;【结论】使用新鲜的样品,且液氮与裂解液结合,是获得高质量RNA的可行方法。     

Evaluation of glycoproteins purified from adult and larval camel ticks (Hyalomma dromedarii) as a candidate vaccine

In order to identify antigens that can help prevent camel tick infestations, three major glycoproteins (GLPs) about 97, 66 and 40 kDa in size were purified from adult and larval Egyptian ticks, Hyalomma (H.) dromedarii, using a single-step purification method with Con-A sepharose. The purified GLPs were evaluated as vaccines against camel tick infestation in rabbits. The rabbits received three intramuscular inoculations of GLPs (20 µg/animal) on days 0, 14, and 28. In the immunoblot analysis, Sera from the immunized rabbits recognized the native GLPs and other proteins from larval and adult H. dromedarii ticks along with those from other tick species such as Rhipicephalus sanguineus but not Ornithodoros moubata. The effects of immunity induced by these GLPs were determined by exposing rabbits to adult H. dromedarii ticks. These results demonstrated that GLP immunization led to a slightly decreased reproductive index and significantly reduced rates of egg hatchability. These results demonstrated that immunization with the purified GLPs can provide protection against infestation by H. dromedarii and some other tick species. Further studies are needed to confirm the effectiveness of immunization with GLPs against other tick species.     

基于DNA条形码技术的泰国番石榴中实蝇幼虫分子鉴定研究

实蝇是世界重要的检疫性害虫之一,对果蔬生产及其国际贸易具有很大的影响。而以线粒体DNA细胞色素C氧化酶亚单位I(mtDNACOI)基因的部分序列作为实蝇的DNA条形码能减少对实蝇成虫形态特征的依赖,可检测其任何虫态的样品,有助于实蝇样品的快速鉴定。本研究采用DNA条形码技术,针对采自泰国四色菊市番石榴烂果中的5头实蝇幼虫进行COI扩增测序,与生命条形码数据库(BOLD)中的序列进行比对并利用PAUP4.0软件构建了其系统进化树。根据序列分析和系统进化关系分析的结果,将5头实蝇幼虫样品鉴定为番石榴实蝇(Bactrocera correctaBezzi),并将本研究获得的2条COI序列在GenBank中注册,GenBank的登录号为HM590450和HM590451。     

苏云金芽胞杆菌工程菌BIOT185对金龟子幼虫的田间防效及其安全性评价

为评价苏云金芽胞杆菌(Bacillus thuringiensis,Bt)工程菌BIOT185对金龟子幼虫的田间防治效果以及其中间试验水平的安全性,在河北省定州市花生田开展了两年的田间小区试验。调查结果表明,每mL含100亿芽胞的工程菌BIOT185悬浮剂稀释10倍和100倍处理对金龟子幼虫具有较好的防治效果,虫口减退率可达52.08%～70.51%,保果效果为50.40%～66.57%,说明高浓度的BIOT185可以有效控制金龟子幼虫的危害。利用平板菌落计数法结合PCR检测发现,在施药后的365 d中BIOT185在土壤中的菌量总体呈下降趋势;调查田间各小区的昆虫种群发现,工程菌BIOT185处理区与野生菌HBF-1、BT185以及对照区的昆虫种群差异不显著。说明BIOT185不会在土壤环境中大量残留,也不会影响花生田的昆虫种群。     

丁鱥鱼胚胎及仔鱼发育的观察

【目的】研究丁鱥鱼在南方较高水温条件下的生长发育情况,解决丁鱥鱼在南方推广养殖中苗种欠缺的瓶颈问题。【方法】对丁鱥亲鱼进行人工繁殖,然后对其胚胎及仔鱼发育过程进行观察记录。【结果】丁鱥鱼催产效应时间11h,受精率为75.3%,孵化率为82.7%,仔鱼成活率59.3%,育成规格为1.5～2.0cm鱼苗20.0万尾。丁鱥鱼受精卵呈卵圆形,大部分呈青灰色,直径0.50～0.77mm,具粘性;在27.0～31.0℃的温度下,受精卵历时1936min发育后孵出,有效积温为908.89℃·h;胚胎发育可划分为6个阶段30个发育时期。刚孵出仔鱼全长3.84～3.87mm,出膜后第3d卵黄囊吸收60%~80%,约有30%仔鱼开口摄食;第4d卵黄囊吸收90%,约有90%仔鱼开口;出膜第5d卵黄囊基本吸收完,但出现少许死鱼苗。【结论】丁鱥鱼适宜在南方进行繁殖及鱼苗培育,水温应稳定在20.0～27.0℃,不宜超过30.0℃,在孵化及鱼苗培育过程中注意清除敌害生物及保持清新水质,并根据水温情况及时投喂开口饵料,提高仔鱼成活率。     

斜纹夜蛾幼虫存活过程的神经网络模型的建立

利用BP神经网络对斜纹夜蛾幼虫的存活过程进行建模分析,给出了BP神经网络建模的Matlab算法程序。结果表明,BP神经网络能充分逼近斜纹夜蛾幼虫的存活过程曲线,使用BP神经网络可对斜纹夜蛾幼虫的存活过程进行有效的建模。利用所建立的模型,得到了关于斜纹夜蛾幼虫存活过程的模拟预测结果。不同温度下斜纹夜蛾幼虫种群的死亡过程有所不同。在20,24,36和40℃温度下,斜纹夜蛾幼虫种群的死亡高峰分别出现在卵孵化后20d左右,5～20d,5～10d及3～6d。最后,对神经网络建模的特点,以及影响BP神经网络建模效果的主要因素进行了分析讨论。     

Potential effect of increasing the water content in the digestibility of microdiets for fish larvae

This study aimed to find a way for increasing the water content after rehydration of microdiet particles for fish larvae and to assess whether such increase contributes to a better disaggregation and digestion of particles within the gut of gilthead seabream larvae (4–15 days after hatching, dph). Four microdiets were prepared with increasing amounts of carboxymethylcellulose (CMC): 0, 20, 40 and 80 g kg^?1. The inclusion of CMC resulted in an effective increase of the hydration capacity of particles. Water content after immersion ranged from 800 g kg^?1 in the microdiet without CMC to 900 g kg^?1 with 80 g kg^?1 CMC. Larvae of different ages were fed during 10 h with these microdiets. The microparticles containing 80 g kg^?1 CMC were more degraded in the gut of the younger larvae. No clear differences were observed from 10 dph onwards. Differences in enzymatic activities were not evident except for α‐amylase in 6 dph larvae. Gene expression of the enzyme precursors did not show any relation to the water content of microdiets. Overall, the results indicate that increasing the water content up to 900 g kg^?1 may enhance the disintegration of food particles within the gut of younger larvae and to a lesser extent affect the response of digestive enzyme activities.     

基于形态与分子数据鉴定苹果异胫小卷蛾

采用DNA条形码技术,对珠海出入境检验检疫局从南非进口的鲜橙和鲜葡萄中截获的3头卷蛾幼虫进行分子鉴定,采用通用引物LC01490/HC02198对其线粒体C0I基因的部分序列进行PCR扩增和测序,并与GenBank数据库和DNA条形码数据库BOLDSYSTEMS v3中的已知序列比对,结果与GenBank中20条苹果异胫小卷蛾的序列同源性达99％以上,与BOLDSYSTEMS v3中46条苹果异胫小卷蛾的序列同源性达99％.结合幼虫的形态特征,综合判定该幼虫为苹果异胫小卷蛾Thaumatotibia leucotreta.     

绵羊乳突类圆线虫病的诊断及虫种鉴定

对河南省新乡市某羊场疑似乳突类圆线虫病的绵羊进行了临床检查,取其粪便采用饱和盐水漂浮法检查,斯陶尔氏虫卵计数法计数,幼虫进行培养与鉴定等。结果表明,病羊粪便中有大量疑似乳突类圆线虫虫卵,对虫卵培养后,根据幼虫大小、虫体形态学观察确诊为绵羊乳突类圆线虫病。     

Analysis of sinking death using video images of the swimming performance of Pacific Bluefin tuna (Thunnus orientalis) larvae

In this study, we aimed to clarify the mechanism of sinking death during the larval stage of Pacific Bluefin tuna Thunnus orientalis by investigating the effects of swimming performance on sinking death, using a behavioral approach. Swimming performance was examined 3–9 days after hatching (DAH) under day and night light conditions in cuboid experimental tanks. Swimming behavior variables such as swimming speed and swimming angle were measured under both light conditions. Larvae in the daytime experiment and larvae with inflated swim bladders at night were distributed on the surface layer of the water column. In contrast, larvae with uninflated swim bladders at night were frequently observed swimming vertically or sinking to the bottom of the tank. Larvae with inflated swim bladders at night were always distributed beneath the surface until the next morning (survival rates were 100 %). However, larvae with uninflated swim bladders at night swam upward repeatedly and later sank to the bottom of the tank (survival rates were 60 % and 38 % at 5 and 9 DAH, respectively). Larvae with uninflated swim bladders were not always able to maintain their swimming depth by swimming until the next morning. Additionally, their swimming speed and vertical swimming frequency (ratio) depended on the illumination and swim bladder conditions. Our findings show that larvae with uninflated swim bladders at night were associated with a higher risk of sinking death. The swimming energy capacity of Pacific Bluefin tuna larvae, which indicates the total amount of the energy that enables individuals to swim throughout the night without feeding, was found to be linked to sinking death.