一株鸭源禽流感病毒（H9N2）全基因序列分析及其排毒规律的研究

为研究一株鸭源H9N2亚型禽流感病毒(JN1株)全基因组的分子特征和遗传进化情况以及感染雏鸭后的排毒规律,对其生物学特性、全基因组序列以及遗传进化进行了分析,通过点眼滴鼻方式感染3周龄健康雏鸭,利用建立的实时荧光定量PCR(RT-PCR)检测鸭的排毒规律,并测定血清抗体效价。JNI株的鸡胚半数感染量为10~(-7.54)/0.2mL,鸡胚最小致死量的平均死亡时间为72h,静脉致病指数为0.266,抗原相关系数为0.47。系统发育分析表明,JN1株的HA与CK/HK/G9/97和DK/HK/Y280/97在同一分支上,NA、M、NP、PA、PB1、PB2基因与SH/F/98同源性较高,NS基因则与H5N1亚型禽流感病毒进化关系较近。HA的裂解位点为PSRSSR↓GL,存在6个潜在糖基化位点,在234位的氨基酸残基为L,NA基因颈部存在缺失。雏鸭在感染该病毒后,饮食正常,精神良好,3~17d均有排毒,咽拭子和泄殖腔棉拭子排毒规律基本一致,分别在第3天和第13天出现排毒高峰。该H9N2亚型禽流感病毒属欧亚大陆分支,为低致病性禽流感,病毒可通过呼吸道和泄殖腔向外界排毒,排毒时间较长。     

NDV和IBV感染对肉鸡大肠杆菌病发病的影响

为了解新城疫弱毒苗和传染性支气管炎弱毒苗接种对肉鸡大肠杆菌病发病的影响。对8日龄肉仔鸡用不同剂量NDVclone-30株或IBV-H120株进行滴鼻及饮水,48h后相同途径接种2株不同“O”血清型的致病性大肠杆菌。结果表明,致病性大肠杆菌单独接种鸡死亡率在20%-25%之间,而NDVclone-30株或IBVH120株与致病性大肠杆菌联合接种鸡死亡率有明显增加达40%-75%。     

Periplaneta americana peptide decreases apoptosis of pig-ovary granulosa cells induced by H2O2 through FoxO1

Oxidative stress can induce apoptosis of granulosa cells and lead to follicular atresia, thereby reducing the number of pigs giving birth. The aim of this study was to investigate the protective effect of Periplaneta americana peptide (PAP) on the apoptosis of the granulosa cells of pig ovaries (PGCs) induced by hydrogen peroxide (H₂O₂) via FoxO1. PGCs were treated with H₂O₂ to establish a cell apoptosis model. Cell viability was measured using the cell counting kit-8 (CCK-8) assay, and cell apoptosis was detected using flow cytometry. The malondialdehyde (MDA) level and nitric oxide (NO) content were detected to reflect the oxidative stress. Western blotting, qRT-PCR and overexpression were undertaken to determine the expression of FoxO1 and caspase-3, and immunofluorescence was used to detect FoxO1 in the nucleus and cytoplasm. PGCs were treated with 100 μM H₂O₂ for 6 hr, which resulted in oxidative damage and apoptosis and an apoptosis rate for PGCs of 32.95%. Next, PGCs were treated with 400 μg/ml PAP for 24 hr to repair the apoptosis induced by H₂O₂. PAP improved cell viability in H₂O₂-stimulated PGCs, the increased MDA level and NO content caused by H₂O₂ stimulation were reversed and the apoptotic rate of PGCs was reduced. The qRT-PCR and Western blotting results indicated that PAP decreased the H₂O₂-induced apoptosis and the expression of FoxO1 and caspase-3 in PGCs. The effect of PAP was the same following FoxO1 overexpression. FoxO1 was expressed in the nucleus when stimulated by H₂O₂ or overexpression; however, it migrated to the cytoplasm following PAP treatment. PAP decreased the apoptosis of PGCs induced by H₂O₂ by regulating FoxO1 expression and nuclear translocation.     

Immunoadjuvant effects of bacterial genomic DNA and CpG oligodeoxynucleotides on avian influenza virus subtype H5N1 inactivated oil emulsion vaccine in chicken

This study investigated the immunoadjuvant effects of three types of bacterial genomic DNA and CpG oligonucleotides (CpG ODN) on the avian influenza virus (AIV) subtype H5N1 inactivated oil emulsion vaccine under two immunization strategies. The genomic DNA extracted from Escherichia coli O₂, Staphylococcus aureus, Streptococcus faecalis FQ68, and synthetic CpG ODN were used as adjuvants, and their effects on the AIV oil emulsion vaccine were examined in chickens. The results indicated that when administered separately from the vaccine, adjuvants induced lower haemagglutination inhibition (HI) titres and serum IgG titres but resulted in higher concentrations of IFN-γ and IL-10. In contrast, when combined with the oil emulsion vaccine prior to inoculation, CpG ODN induced higher HI, IgG titres and IFN-γ concentration but resulted in lower IL-10 concentration. These data suggest that, depending on the immunization approaches, adjuvants may exert distinct immune effects in chickens receiving AIV H5N1 oil emulsion vaccine: the prior incorporation of CpG ODN into the vaccine may augment both the humoral and Th1 type immune responses, while separate inoculation of adjuvants has not shown better adjuvanticity.     

全球人感染H5N1禽流感病例流行病学特征分析

该文基于近年来国内外报道的H5N1亚型高致病性禽流感病毒感染人的病例,系统分析了人感染H5N1禽流感病毒的流行病学特征,阐述人禽流感暴发流行的可能性及预防控制的紧迫性。     

AIV H5N1亚型高免卵黄抗体的制备及理化特性研究

应用禽流感灭活疫苗(H5N1,Re-1株)免疫20周龄的高产来航蛋鸡,制备抗AIV H5N1高免卵黄抗体(IgY).在比较了3种缓冲液对IgY活性的影响后,确定采用水稀释法提取IgY.同时在模拟的胃肠内环境下对IgY的各种理化性质进行了试验.结果表明,纯化的IgY在pH 2.0和胃蛋白酶终浓度为1.5 mg/mL的环境下,以及在pH 8.0和胰蛋白酶终浓度为2.5 mg/mL的环境下,37℃作用1 h后其ELISA抗体效价均无明显下降;同时,热稳定性试验表明,IgY在60℃以下作用30 min,其ELISA抗体效价也无明显下降;中和试验测得IgY的中和效价为1:640,具有较好的中和H5N1亚型AIV的能力.以上结果表明,本试验所制备的抗禽流感H5N1亚型高免IgY具有较高的抗体效价和中和效价,同时还具有一定的耐蛋白酶消化的作用,可用于H5N1亚型禽流感的预防和治疗.     

两株H6N2亚型鸡源禽流感病毒的分离鉴定

本研究利用血凝抑制试验(HI)、反转录-聚合酶链式反应(RT-PCR)、基因测序等方法,对广东省某活禽交易市场进行流行病学调查时获得的两株非H5、H9亚型禽流感病毒65株和C7株进行了亚型鉴定。结果表明这两株病毒均具有血凝活性,且能被抗H6亚型禽流感病毒标准阳性血清特异性抑制。用针对禽流感病毒的M基因、H6亚型禽流感病毒HA基因、N2亚型禽流感病毒NA基因特异性鉴定引物对65株和C7株进行RT-PCR扩增,分别获得特异性目的片段。测序及BLAST分析表明两株分离株与H6N2亚型禽流感广东分离株的HA基因和NA基因核苷酸序列相似性均高达95%以上。将该两分离株鉴定为H6N2亚型禽流感病毒,并命名为A/Chicken/Guangdong/65/2009、A/Chicken/Guangdong/C7/2009。     

6-硫-6-烷硫基-12H-氯代双苯并[d,g][1,3,2]-二氧磷杂八环的合成

通过双- (2 -羟基 -氯代苯基 )甲烷 2与 P4S10 在三乙胺存在下的关环反应 ,得到了6 -硫 - 6巯基 - 12 H-氯代双苯并 [d,g][1,3,2 ]-二氧磷杂八环三乙胺盐 3,后者与卤代烷酯化得到了一系列 6 -硫 - 6 -烷硫基 - 12 H-氯代双苯并 [d,g][1,3,2 ]-二氧磷杂八环 4 ,测试并讨论了它们的1H和31P NMR数据。初步生物试验结果表明化合物 4对黄瓜霜霉病菌具有较好的抑制活性。     

Activity of H+-K+-ATPase in gastric parietal cells under stress in rats and analysis of electron microscopic enzyme cytochemistry

AIM: To demonstrate the changes of activity and electron microscopic enzyme cytochemistry staining of H⁺-K⁺-ATPase of gastric parietal cells under stress in rats. METHODS: Twenty-four male SD rats were randomly divided into normal group, stress group and stress+omeprazole (OM) group. Water immersion-restraint stress (WRS) model in SD rats was performed. The ulcer index (UI) of gastric mucosa and H⁺-K⁺-ATPase activity of gastric parietal cells were measured. The changes of ultrastructure and electron microscopic enzyme cytochemistry staining of parietal cells were observed under transmission electron microscope (TEM). RESULTS: Compared with control group, the UI of gastric mucosa and H⁺-K⁺-ATPase activity of gastric parietal cells increased (P<0.01 and P<0.05) in stress group. In stress+OM group, both UI and H⁺-K⁺-ATPase activity decreased (P<0.01) compared with stress group. Parietal cells were in a resting state in control group, and became active in stress group, where plenty of intracellular canaliculi were observed under the TEM. In stress+OM group, the dilated intracellular canaliculi lined with rare microvilli were founded. Enzyme cytochemistry staining showed that there was little black punctate enzyme reactive product scatted in intracellular canaliculi and the apical plasma membrane of parietal cells in control group, and there were large amounts of black enzyme reactive product accumulated at the intracellular canaliculi in stress group. Scarcely deposition of enzyme reactive product in intracellular canaliculi was observed in stress+OM group. CONCLUSION: The results indicate that the H⁺-K⁺-ATPase activity of gastric parietal cells increases under WRS, and is in accordance with ultrastructure changes. These findings suggeste that gastric acid might be one of the most important factors that result in stress ulcer.     

大青叶中4（3H）喹唑酮体外抗H1N1型流感病毒的作用研究

目的：探讨大青叶中4（3H）喹唑酮体外抗H1N1型流感病毒的作用。方法：用MTT染色法测定大青叶中4（3H）喹唑酮通过不同加药方式对H1N1型流感病毒的体外抑制作用。结果：大青叶中4（3H）喹唑酮对H1N1型流感病毒具有明显的阻断作用和对细胞内病毒有抑制作用。结论：大青叶中4（3H）喹唑酮具有明显的体外阻止H1N1型流感病毒吸附到靶细胞和抑制H1N1型流感病毒增殖的作用。