滴度及注射方式对2型腺相关病毒载体表达效果的影响

由于腺相关病毒载体(adeno-associated viral vector,AAV)本身的侵染不具有神经元特异性,其在神经系统相关研究中会存在侵染范围不符合试验要求的情况,因此本试验拟研究不同滴度和注射方式对病毒侵染范围的影响。结果显示,在注射较高滴度的AAV2-CMV-EGFP(1.3×10¹¹mg/L)3周后,其侵染范围可由延髓中缝苍白核(raphe pallidus nucleus,RPa)区域延伸至下丘脑室旁核(paraventricular nucleus,PVN)区域;较低滴度(1.3×10⁸mg/L)结合压力注射的方式可以在一定程度上限制AAV的侵染范围,并且载体携带的EGFP荧光基团在637 nm波段不具有明显的假阳性信号。     

Control of insect pests using trunk injection in a newly established apple orchard

We evaluated the potential for using preplant trunk injections of emamectin benzoate in nonbearing apple trees. Trees were evaluated for pest injury and emamectin residues throughout the planting season and into the following year. Injections into the trunk best delivered emamectin benzoate to the canopy compared with injections into the taproot, and the higher rate reduced insect pests more than the lower rate. In the following year, differences in insect control between trunk and root injections were less pronounced, but the higher rate of emamectin benzoate persisted longer and better reduced pests relative to the other treatments.     

Embryo-mediated genome editing for accelerated genetic improvement of livestock

Selecting beneficial DNA variants is the main goal of animal breeding. However, this process is inherently inefficient because each animal only carries a fraction of all desirable variants. Genome editing technology with its ability to directly introduce beneficial sequence variants offers new opportunities to modernize animal breeding by overcoming this biological limitation and accelerating genetic gains. To realize rapid genetic gain, precise edits need to be introduced into genomically-selected embryos, which minimizes the genetic lag. However, embryo-mediated precision editing by homology-directed repair (HDR) mechanisms is currently an inefficient process that often produces mosaic embryos and greatly limits the numbers of available edited embryos. This review provides a summary of genome editing in bovine embryos and proposes an embryo-mediated accelerated breeding scheme that overcomes the present efficiency limitations of HDR editing in bovine embryos. It integrates embryo-based genomic selection with precise multi-editing and uses embryonic cloning with elite edited blastomeres or embryonic pluripotent stem cells to resolve mosaicism, enable multiplex editing and multiply rare elite genotypes. Such a breeding strategy would enable a more targeted, accelerated approach for livestock improvement that allows stacking of beneficial variants, even including novel traits from outside the breeding population, in the most recent elite genetic background, essentially within a single generation.     

单壁碳纳米管载嗜水气单胞菌外膜蛋白亚单位疫苗的研制及免疫效果评价

为了筛选细菌的疫苗候选蛋白,本实验根据课题组前期的研究,挑选了2个嗜水气单胞菌外膜蛋白(A0KLQ6和A0KH89),以A0KHR9(OmpAII)为对照,通过比较外膜蛋白与碳纳米管载外膜蛋白通过腹腔注射与浸泡免疫模式,比较和评价斑马鱼的免疫反应与免疫保护效果。结果显示,腹腔注射组和浸泡组与普通蛋白组对比,碳纳米管载蛋白能够显著提高斑马鱼免疫相关基因表达量。腹腔注射组在5μg剂量下免疫A0KHR9、A0KLQ6和A0KH89蛋白分别获得61.60%、 65.39%和84.96%相对免疫保护率(relative immune protection rate,RPS),在5μg剂量下免疫碳纳米管载蛋白SWCNTs-A0KHR9、SWCNTsA0KLQ6和SWCNTs-A0KH89分别获得68.10%、77.50%和90.43%的RPSs。浸泡组在40 mg/L剂量下免疫A0KHR9、A0KLQ6和A0KH89蛋白分别获得30.80%、25.85%和37.80%的RPSs,在40 mg/L最高剂量下免疫碳纳米管载蛋白疫苗SWCNTs-A0KHR9、SWCNTsA0KLQ6和SWCNTs-A0KH89分别获得63.60%、73.74%和68.49%的RPSs。研究表明,铁离子限制下嗜水气单胞菌外膜蛋白A0KLQ6和A0KH89等能够作为抗嗜水气单胞菌的亚单位疫苗候选蛋白,而单壁碳纳米管载蛋白通过注射免疫与浸泡免疫均能显著提升免疫疗效,是一种疫苗的适宜纳米载体。以上研究为寻找高效的嗜水气单胞菌亚单位疫苗候选蛋白及佐剂提供了理论依据。     

注射多巴胺对中华绒螯蟹打斗行为的影响

选同一家系规格相似、健康有活力的中华绒螯蟹新品种“光合1号”1龄雄蟹100只,分为4组,放入直径20 cm、高40 cm PE管(底部均匀钻12个直径为5 mm的孔洞)中,PE管置于120 L水槽中。用2个不透明烧杯将2只中华绒螯蟹分别倒扣于试验容器中,置于PE管两侧1 h。在中华绒螯蟹第三步足的动脉膜中分别注射20μL生理盐水(对照组),10^-8 mol、10-7 mol和10-6 mol的多巴胺液,利用闭路摄像头记录4 h内2只中华绒螯蟹打斗时长、次数、打斗分数及胜负关系等指标。打斗结束后,迅速取出中华绒螯蟹血淋巴,用ELISA酶联免疫法测定多巴胺含量。试验结果显示,对照组中华绒鳌蟹打斗时间最长,与各注射组差异显著(P<0.05);注射组中,随注射量的增加,打斗时长下降。对照组中华绒螯蟹打斗强度低于10^-8 mol组,高于其他注射组,与各注射组间差异显著(P<0.05)。注射组中,随注射多巴胺量升高,打斗强度下降,10^-8 mol组打斗强度最高,各组间差异显著(P<0.05)。对照组打斗次数与10-7 mol组差异不显著(P>0.05),与其余各组差异显著(P<0.05)。注射组中,随注射量的增加,打斗次数下降,10^-8 mol组与其余各组差异显著(P<0.05),10-7 mol组与10-6 mol组差异不显著(P>0.05)。打斗结束后,胜利方体内多巴胺含量与正常水平差异不显著(P>0.05),但显著高于失败方(P<0.05)。试验结果表明,注射多巴胺极显著影响中华绒螯蟹打斗行为(P<0.01),随多巴胺注射量的升高,中华绒螯蟹打斗行为整体下降。     

硫酸卡那霉素注射液含量测定的不确定度分析

为研究微生物检定法(管碟法)测定硫酸卡那霉素注射液生物效价试验中各因素对结果的影响,采用不确定度分析,将影响因素归纳为称量、稀释、加样、测定、平行试验,量化各不确定度分量,评估各因素的影响因子。结果表明,牛津杯加样体积对测定结果影响最大,提示试验中应尽可能精确控制加样体积。     

Development of piscirickettsiosis in Atlantic salmon (Salmo salar L.) smolts after intraperitoneal and cohabitant challenge using an EM90‐like isolate: A comparative study

Piscirickettsiosis, caused by the intracellular Gram‐negative bacteria Piscirickettsia salmonis, is at present the most devastating disease in the Chilean salmon industry. The aim of this study was to analyse disease development after challenge with a P. salmonis strain (EM90‐like) under a controlled environment by comparing intraperitoneal challenge with cohabitation challenge. The P. salmonis EM90‐like isolate was cultured in a liquid medium for the challenge of 400 Atlantic salmon (Salmo salar) smolts. Cumulative mortality was registered, necropsy was performed, and bacterial distribution in the tissues and histopathological changes were analysed. The results revealed a similar progression of the disease for the two different challenge models. Pathological and histopathological changes became more visible during the development of the clinical phase of the disease. Bacterial DNA was identified in all the analysed tissues indicating a systemic infection. Bacterial tropism to visceral organs was demonstrated by real‐time quantitative PCR and immunohistochemistry. Better knowledge of disease development during P. salmonis infection may contribute to further development of challenge models that mimic the field situation during piscirickettsiosis outbreaks. The models can be used to develop and test future preventive measures against the disease.     

Protective effect of Huangqi injection combined with puerarin injection on myocardium of type 2 diabetes mice

AIM:To investigate the effect of Huangqi injection combined with puerarin injection on the myocardium of the mice with type 2 diabetes. METHODS:Diabetic KKAy mice were randomly divided into model group and treatment group (Huangqi injection combined with puerarin injection). The male KKAy mice of the same age were used as control group. All mice were sacrificed at 21, 24 and 28 weeks. Morphological changes of the myocardium were observed by HE staining. Apoptosis of the cardiomyocytes was measured by TUNEL staining. The mRNA levels of glucose-regulated protein 78(GRP78), C/EBP hoinologous protein (CHOP) and p53 up-regulated modulator of apoptosis (PUMA) were detected by real-time PCR, and the protein levels of GRP78, CHOP, PUMA, caspase-3, cleaved caspase-3, caspase-9, cleaved caspase-9, poly(ADP-ribose) polymerase (PARP) and cleaved PARP were determined by Western blot. RESULTS:Cardiomyocyte hypertrophy, partly dissolved sarcoplasm and necrosis were observed in model group, and these lesion were alleviated in treatment group. Obvious increased apoptosis in model group and significantly decreased apoptosis of cardiomyocytes in treatment group was observed (P<0.05). At 21, 24 and 28 weeks, the mRNA and protein levels of GRP78, CHOP and PUMA and the protein levels of cleaved caspase-3, cleaved caspase-9 and cleaved PARP in model group were increased significantly as compared with control group (P<0.01), and these in treated group were decreased compared with model group. CONCLUSION:Huangqi injection combined with puerarin injection has cardioprotective effects on type 2 diabetes mice and its mechanism of the action was implemented via inhibiting the activation of endoplasmic reticulum stress and caspase pathway, thus resulting in suppressed apoptosis.     

Clinical Therapeutic Effect of Praziquantel Injection on Buffalo Schistosomiasis

To observe the clinical therapeutic effect of 30% praziquantel injection on buffalo schistosomiasis, the sick buffalos naturally infected with Schistosoma japonicum were selected by miracidium hatching method. In experimental clinical trials, sick buffalos were randomly divided into five groups, high-dose (20 mg/kg), middle-dose (10 mg/kg) and low-dose (5 mg/kg) praziquantel injection groups, praziquantel tablet group (30 mg/kg) and positive control group. After the treatment of 30 d, the negative conversion rate of miracidium were 100.0%, 100.0%, 77.8% and 85.7%, respectively, in praziquantel injection groups with the high, middle and low dose and oral medication group. In expanded clinical trials, the sick buffalos were randomly divided into two groups, praziquantel injection group (10 mg/kg, i.m.) and praziquantel tablet group (30 mg/kg). The results showed that, the negative conversion rate of miracidium were all 100% in the former 52 patients and the latter 6 patients after 30 d. The research confirmed that praziquantel injection was significantly effective in the treatment of buffalo schistosomiasis and convenient for administration. It was concluded that 30% praziquantel injection could replace the traditional oral praziquantel tablet for the treatment of buffalo schistosomiasis, and the recommended dose was 10 mg/kg.