rmlA 基因缺失影响禽致病性大肠杆菌的生物被膜形成

【目的】构建禽致病性大肠杆菌(Avian pathogenic Escherichia coli,APEC)rmlA基因缺失株,研究该缺失株的生物学特性。【方法】利用Red重组系统构建rmlA缺失株;比较野生株与缺失株在生长特性、运动性和生物被膜形成能力等方面的差异;运用Real-time PCR技术,比较野生株与rmlA缺失株对APEC部分毒力基因转录的影响。【结果】rmlA缺失株,不影响APEC的生长和运动特性,但生物被膜形成能力显著增强,且使luxS、irp2基因转录水平分别上调2倍、1.8倍,iucD、fyuA则下调25倍。【结论】APEC的rmlA基因可以影响禽致病性大肠杆菌的生物被膜形成能力及部分毒力基因的转录水平;而对APEC的生长、运动特性没有影响。     

双组分系统rcsC基因影响禽致病性大肠杆菌的致病性及相关生物学特性

【目的】双组分系统Rcs感受外界环境变化,并调控细菌的适应性及生存等。本文探讨Rcs双组分系统传感器激酶RcsC对禽致病性大肠杆菌(avian pathogenic Escherichia coli,APEC)相关生物学特性及致病性的影响。【方法】采用Red同源重组的方法构建rcsC基因缺失株,并利用互补质粒构建互补株,然后比较野生株、基因缺失株与互补株的生长特性、运动性、生物被膜、凝集沉淀能力、致病力及毒力基因转录水平的差异。【结果】rcsC基因缺失不影响APEC的生长速度,然而,缺失RcsC导致APEC的运动能力升高、生物被膜形成能力降低和凝集能力增强。凝集试验结果显示rcsC基因有助于APEC的凝集沉降。细胞黏附入侵结果表明,rcsC在APEC侵袭DF-1细胞过程中发挥作用,而对黏附能力无影响。动物感染试验结果表明rcsC基因缺失能显著降低APEC的毒力。荧光定量PCR检测结果表明,rcsC基因缺失株中ompA、aatA、fyuA和luxS基因的转录水平均显著降低,而fimC和tsh基因的转录水平显著升高。【结论】RcsC参与调控APEC的运动性、生物被膜形成、凝集沉降和致病力。     

wzzE不影响禽致病性大肠杆菌雏鸭分离株LPS的表型北大核心CSCD

【目的】wzz参与很多革兰氏阴性菌O抗原的合成,并对细菌的致病性具有调控作用。在禽致病性大肠杆菌（Avian pathogenic Escherichia coli,APEC）中,存在Wzz蛋白超家族基因wzz E,目前尚未开展该基因对APEC脂多糖（LPS）的合成及致病作用研究,因此本研究开展了wzz E对APEC LPS合成以及生物学特性影响的研究。【方法】通过构建APEC DE17株的wzz E缺失株,开展对野生型和缺失株的LD50、黏附与入侵DF-1细胞、脂多糖表型及内毒素毒价等相关生物学特性的影响研究。【结果】结果表明,缺失wzz E的APEC,不影响细菌的生长特性。野生型、缺失株及回复株的LPS表型无明显差异。DE17Δwzz E与DE17的黏附与入侵结果无显著差异。血清型鉴定结果表明,缺失wzz E,不影响细菌的血清型。内毒素毒力检测结果DE17、DE17Δwzz E及CΔwzz E内毒素的毒价一致,为4×10~5 EU/mg。对7日龄樱桃谷鸭进行攻毒,测定各菌株的LD50,结果表明,与野生型相比,DE17Δwzz E缺失株毒力下降了10倍。【结论】wzz E缺失对LPS的表型无显著影响,但wzz E参与APEC的致病过程,然而wzz E对APEC毒力的调控机制仍有待进一步研究。     

VI型分泌系统2核心组分VgrG对禽致病性大肠杆菌致病性的影响

【目的】构建禽致病性大肠杆菌(Avian pathogenic Escherichia coli,APEC)VI型分泌系统2(Type VI secretion system 2,T6SS2)结构基因vgrG缺失株,研究其对APEC生物学特性及致病性的影响。【方法】通过Red同源重组方法构建DE719菌株vgrG基因缺失株,并利用低拷贝质粒pSTV28构建互补株。比较分析野生株、缺失株与互补株的生长特性、运动性、生物被膜形成能力、黏附侵袭能力、动物致病力等差异。【结果】vgr G基因缺失不影响DE719的生长速度、运动能力及生物被膜形成能力。致病性试验表明缺失vgrG导致体内定殖能力及致病力显著下降,然而对DF-1细胞的黏附能力增强。【结论】T6SS2核心组分VgrG在APEC感染过程中发挥重要作用,为了解APEC的致病作用提供参考。     

大肠杆菌三型分泌系统2转录调节子EtrA对禽致病性大肠杆菌致病性的影响

【背景】禽致病性大肠杆菌(avian pathogenic Escherichia coli,APEC)是禽类主要病原菌之一,大肠杆菌三型分泌系统2 (Escherichia coli type III secretion system 2,ETT2)可通过转录调节子调控其致病性,但在APEC中转录调节子EtrA对其致病性的影响目前尚不清楚。【目的】研究ETT2中转录调节子EtrA对APEC致病性的影响。【方法】利用Red同源重组技术构建ETT2-etrA基因缺失株及回复株。比较生长性能、生物被膜形成、运动性及对血清敏感性的差异,基于RNA-Seq测序及Real-timePCR技术比较野生株和缺失株中与生物被膜形成、运动性以及毒力因子相关基因的转录水平。【结果】与野生株相比,缺失株及回复株生长特性无显著变化(P0.05),但APEC40-ΔetrA生物被膜形成能力和对血清敏感性明显增强(P0.001),运动性较野生株明显下降(P0.01),回复株的表型有所回复。转录组学筛选出7个毒力差异基因,生物被膜形成相关基因显著上调,参与影响细菌运动性的基因显著下调。qRT-PCR验证与转录组学结果一致。【结论】etrA缺失可以显著影响APEC的生物被膜形成、运动性及对血清的敏感性,这可为进一步探讨ETT2对APEC的致病作用提供参考。     

酰基高丝氨酸内脂合成酶LasI对禽致病性大肠杆菌生物学特性的影响

【背景】禽致病性大肠杆菌(Avian pathogenic Escherichia coli, APEC)是禽类主要病原菌之一,群体感应(Quorumsensing,QS)系统可通过信号分子调控其生物学特性。在APEC中信号分子AHL对其生物学特性的影响目前尚不清楚。【目的】研究信号分子AHL对APEC生物学特性的影响。【方法】将含铜绿假单胞菌酰基高丝氨酸内脂合成酶(Acyl-homoserine-lactone synthase,lasI)基因的表达质粒转化至APEC菌株DE17中,构建重组菌株DE17-lasI,利用LasI在DE17中合成AHL。比较野生株和重组菌株产生AHL信号分子、生长特性、生物被膜形成能力、运动性以及耐药性等生物学特性的差异;运用Real-timePCR技术,比较野生株和重组菌株中与生物被膜形成、运动性以及毒力因子相关基因的转录水平。【结果】对重组菌株AHL信号分子检测表明,DE17-lasI能够产生AHL信号分子,与野生株DE17相比,DE17-lasI生物被膜形成能力和运动性显著降低(P0.01),但其生长特性和耐药性无显著变化(P0.05);Real-time PCR检测结果表明,重组菌株的毒力因子fimH转录水平上调了58.8倍,而ompA、iss分别下调了95.4%、77.3%。与生物被膜形成相关基因agn43下调了75%,鞭毛合成基因flhA下调了80.8%。此外,AHL受体sdiA的转录水平上调了19.8倍。【结论】转化lasI至APEC中,能促进其在APEC中合成信号分子AHL,并显著影响APEC的部分生物学特性,为进一步探讨AHL型群体感应系统对APEC的调控作用提供参考。     

pagP基因对禽致病性大肠杆菌抗菌肽抗性和致病性的影响

【目的】禽致病性大肠杆菌(Avian pathogenic Escherichia coli,APEC)不仅严重影响全球的养禽业,对人类公共健康也造成巨大的潜在威胁。pag P基因在细菌的抗菌肽抗性和致病性方面发挥重要作用,但关于pag P基因在APEC中的功能尚不清楚。本文构建禽致病性大肠杆菌pag P基因缺失株,对缺失株的抗菌肽抗性和致病性进行研究。【方法】利用Red重组系统构建APEC的pag P基因缺失株,然后利用回复质粒构建回复株。研究pag P基因对细胞黏附与入侵、生物被膜形成能力、外膜渗透性、抗菌肽敏感性、致病性等方面的影响。【结果】成功构建pag P基因缺失株和回复株,抗菌肽抗性试验发现pag P基因缺失株对多粘菌素B、鸡β-防御素2(AVBD2)的敏感性显著增加(P0.01),致病性试验结果表明pag P基因缺失株的毒力显著降低(P0.01)。【结论】APEC的pag P基因对AVBD2的敏感性和APEC的致病性密切相关,为深入研究pag P基因的功能及调控作用奠定了基础。     

kpsE和kpsD双基因缺失显著降低肠道外致病性大肠杆菌的毒力

【目的】探究荚膜对肠道外致病性大肠杆菌致病作用的影响。【方法】选取负责荚膜多糖转运的基因kpsE和kpsD,利用λRed重组系统构建APEC E058和UPEC U17荚膜缺失株E058ΔkpsED和U17ΔkpsED,并通过一系列的体内及体外试验对其生物学特性及致病性进行研究。【结果】双基因缺失株的生长速度较野生株没有明显差异,但缺失株抗血清补体杀菌能力和抗鸡巨噬细胞HD-11细胞吞噬能力显著下降。1日龄雏鸡LD50致病性试验结果显示,缺失株E058ΔkpsED和U17ΔkpsED对鸡失去致病力,而回复株毒力恢复至野生株水平;35日龄SPF鸡体内动态分布和竞争试验显示ΔkpsED缺失株在鸡体内定殖能力和竞争性生长能力显著下降,表明kpsED双基因的缺失能显著降低APEC E058和UPEC U17的致病力。【结论】荚膜与肠道外致病性大肠杆菌的致病性相关,是其重要的毒力因子。     

毒力基因yqeH功能分析及对禽致病性大肠杆菌致病性的影响

【背景】禽致病性大肠杆菌(avian pathogenic Escherichia coli,APEC)可引起禽类急性或亚急性感染,在近年新发现的大肠杆菌Ⅲ型分泌系统2 (Escherichia coli type III secretion system 2,ETT2)中,毒力基因yqeH对其致病性的影响尚不明确。【目的】探究yqeH在APEC致病过程中的作用,为后期深入研究ETT2致病机制奠定基础。【方法】利用Red同源重组技术构建yqeH缺失株ΔyqeH及其回复株CΔyqeH,通过运动性、生物被膜形成能力、抗逆性、抗血清杀菌能力等试验分析yqeH对APEC生物学功能的影响,并通过细胞黏附、侵袭试验、致病力测定及荧光定量PCR检测细胞炎性因子转录水平,探究yqeH对APEC感染宿主的影响。【结果】构建了缺失株ΔyqeH和回复株CΔyqeH;生物学特性试验结果表明,与野生株APEC81相比,缺失株ΔyqeH生物被膜形成能力、运动能力降低,对酸、碱、渗透压、氧化休克的耐受力降低,抗血清杀菌能力及致病力显著降低;与野生株APEC81相比,缺失株ΔyqeH对鸡气管黏膜上皮细胞的黏附及侵袭能...     

ygeG基因对禽致病性大肠杆菌生物学特性及致病性的影响

【目的】本研究构建禽致病性大肠杆菌(avian pathogenic Escherichia coli, APEC) yge G基因缺失株,并对其进行生物学特性及致病性分析,探究yge G在APEC致病过程中的作用,为后期深入研究其所在的Ⅲ型分泌系统2 (type three secretion systems 2,ETT2)毒力岛的致病机制奠定基础。【方法】利用Red同源重组技术构建yge G缺失株APEC81-Δyge G及其回复株APEC81-CΔyge G,通过运动性、生物被膜形成能力、抗逆性、抗血清杀菌能力等试验分析yge G对APEC致病性相关的生物学功能的影响,并通过细胞黏附、侵袭试验及荧光定量PCR检测细胞炎性因子转录水平探究yge G对APEC感染宿主过程的影响。【结果】成功构建了缺失株APEC81-Δyge G和回复株APEC81-CΔyge G;与野生株APEC81相比,缺失株APEC81-Δyge G生长特性无显著变化(P>0.05),生物被膜形成能力显著降低(P<0.01),运动能力极显著升高(P<0.001),对酸休克和氧化休克的耐受力极显...     

六种藓类植物茎的比较解剖学研究

通过对6种藓类植物,即褶叶青藓(Brachythecium salebrosum(Web.et Mohr.)B.S.G.)、湿地匐灯藓(Plagiomnium acutum(Lindb.)Kop.)、侧枝匐灯藓(Plagiomnium maximoviczii(Lindb.)Kop.)、大凤尾藓(Fissidensnobilis Griff.)、大羽藓(Thuidium cymbifolium(Doz.et Molk.)B.S.G.)和大灰藓(Hypnum plumaeforme Wils.)嫩茎和老茎的石蜡切片和显微观察发现,同一藓类植株的嫩茎和老茎,茎结构稳定,不同种藓类植物茎横切面具有不同特征.植物体茎横切面形状、表层细胞的层数、细胞大小和细胞壁厚薄、皮层细胞大小和形状、中轴的有无以及比例等特征可以作为藓类植物的分科分类依据之一.     

Cause of Senescence of Nine Sorts of Flowers

The levels of endogenous phytohormones and respiratory rate in nine sorts of flowers such as Cymbidium faberi Rolfe, Nopalxochia ackermannii Kunth and others were investigated both at full bloom and senescence and meanwhile the effect of exogenous phytohormones on prolonging the blossoms and promoting ethylene production were tested. There is a high content of endogenous ethylene in all the long-lived flowere, about 3–16 folds higer than the short-lived ones. There is a high level of ABA at full blooming flowers of short-lived flowers, in which there is no or only some cytokinins in it, but the ratio of CTK (6BA+zeatin)/ABA is smaller(l.7). The endogenous ABA reached a much higher level at senescence in all nine sorts of flowers, so it is reasonable to consider that it is ABA which plays an important role of regulation in controlling flower's senescence. There is a much higher level of GA3 and zeatin in the long-lived flowers which is not demonstrated in the shortlived ones. The respiratory rate is one of the factors controtling the longevity of flowers, but it does not play a decided role. Application of 6BA and zeatin prolongs distinctly orchid’s longevity, however exogenous IAA through the promotive action on ethylene production, evidently extends the longevity of the flowers of the Nopalxochia ackermannii Kunth.     

龙胆科药用植物化学成分的研究现状

龙胆科植物在我国的分布范围很广,且多数为药用植物,其多数种属的药用植物,至今其化学成分尚未被系统研究。综述了目前龙胆科药用植物的化学成分的研究现状及一般提取方法,对近年来发现的环烯醚萜及裂环烯醚萜类化合物进行了总结,为本科药用植物的更深入研究提供了参考。     

The effects of glutamine of the maintenance of embryogenic cultures of Cryptomeria japonica

Summary Embryogenic tissues of sugi (Cryptomeria japonica) were induced on a modified Campbell and Durzan (CD) medium containing 1 μM 2,4-dichlorophenoxyacetic acid (2,4-D) and 600 mg l⁻¹ glutamine, and subcultured in the medium of the same composition for over 1 yr. This resulted in a mixed culture of embryogenic and non-embryogenic cells. When embryogenic cells were isolated and cultured independently, their capacity to form embryogenic aggregates was lost. Thus, the non-embryogenic cells present within a mixed culture system were essential to the formation of embryogenic aggregates. When embryogenic tissues were isolated and cultured independently on a high glutamine-containing (2400 mg l⁻¹) medium, dry weights and endogenous levels of glutamine increased, and the tissue could generate a large number of embryogenic aggregates. Amino acid analysis of embryogenic and non-embryogenic cells from the maintenance culture indicated a higher level of glutamine was present in the latter. The high endogenous level of glutamine in the non-embryogenic portion of mixed cell masses may be the supplier of glutamine for maintaining the embryogenic property of the tissues.     

Specificity of action of piperidylcholine derivatives as substrates of cholinesterases of various origin

The review deals with study of enzymologic properties of a novel highly specific acetylcholinesterase substrate, N-(β-acetoxyethyl) piperidinium iodomethylate (“piperidylcholine”), and its 30 derivatives that were tested as effectors of cholinesterases of mammals and various species of Pacific squids. It was proven for the first time that responsible for specificity of action was structure of cyclic ammonium grouping of the alcohol part of molecule of the ester substrate. Analysis of specificity is performed based on enzymatic hydrolysis parameters—activity of catalytic center of cholinesterases and bimolecular constant of the reaction rate that are determined at optimal and low substrate concentrations. Among the specially synthesized group of thioester compounds there is revealed one more highly specific acetylcholinesterase substrate—N-(β-acetoxyethyl) piperidinium.     

真菌类遗传学分析的知识结构教学

本文以认知结构理论为指导,讨论了真菌类遗传分析与高等动植物遗传分析的内在联系,认为利用这种内在联系进行教学可收到好的效果并说明了作者的具体教学过程。 Abstract:In the paper, the relationship between genetic analysis of Fungi and genetic analysis of high animal and plant was discussed.A good results were obtained when we adopted this method in the teaching.     

Photoregulation of seed germination of wild-type and of an aurea-mutant of tomato

Seed germination of an aurea mutant of tomato ( Lycopersicon esculentum Mill.) is promoted by continuous irradiation with red, far-red or long-wavelength far-red (758 nm) light as well as by cyclic irradiations (5 min red or 5 min far-red/25 min darkness). Far-red light applied immediately after each red does not change the germination behaviour. Seed germination of the isogenic wild-type, cv. UC-105, is promoted by continuous and cyclic red light while it is inhibited by continuous and cyclic far-red light and by continious 758 nm irradiation. Far-red irradiation reverses almost completely the promoting effect of red light. The promoting effect (in the aurea mutant) and the inhibitory effect (in the wild-type) of continuous far-red light do not show photon fluence rate dependency above 20 nmol m⁻² s⁻¹. It is concluded that phytochrome controls tomato seed germination throgh low energy responses in both the wild type and the au mutant. The promoting effect of continuous and cyclic far-red light in the au mutant can be attributed to a greater sensitivity to P_fr.     

The mechanism of hatching of eggs of Haemonchus contortus

Fluid collected from hatching eggs of Haemonchus contortus contained a lipase which hydrolysed 2-naphthyl laurate (about 0·7 μmol naphthol freed /h/10⁶ eggs). The fluid also hydrolysed l-leucinamide (about 2·3 μmol leucine freed/h/10⁶ eggs). The fluid when added to normal or heated eggs caused ‘hatching’. ‘Hatching’ also occurred in exsheathing fluid from infective juveniles and in a preparation of pancreatic lipase containing leucine aminopeptidase. A purified mammalian leucine aminopeptidase in combination with several different lipases did not attack egg shells.The ‘spontaneous’ hatching of eggs of H. contortus was strongly inhibited by 1,10-phenanthroline, 10^?3M, and this inhibition was reversed by Zn²⁺. However, the inhibition of ‘hatching’ of eggs in externally applied hatching fluid, or the hydrolysis of leucinamide in hatching fluid was generally less marked.