【作者】 彭超；

【导师】 何凡；

【作者基本信息】 海南大学 ， 植物保护， 2016， 硕士

【摘要】 火龙果〔Hylocereus polyrhizus Webb.Britton&Rose〕又称麒麟果、仙蜜果、红龙果等,仙人掌科(Cactaceae)量天尺属(Hylocereus undatus)(红皮)或蛇鞭柱属(Selenicereus)(黄皮),是原产于墨西哥境内和中、南美洲各国的太平洋沿岸地带,的一种典型热带作物。随着近年来集中连片大规模种植火龙果的种植大户越来越多,火龙果的病害发生的严重性瞬间凸显;但是国内火龙果针对火龙果病害的研究还相对较少。本研究通过对海南地区火龙果主产区病害普查,初步确定海南火龙果病害主要有11种,主要针对发病最为严重的溃疡病和海南地区首次发现的茎斑病和灰霉病两种真菌病原进行详细鉴定,同时对海南地区火龙果病毒病进行初步分子鉴定,为今后对火龙果更为深入的研究打下基础。本研究主要得到以下主要结果:1.海南地区火龙果病害主要有10种,包括真菌病害7种,病毒病害1种,生理病害2种。其中溃疡病〔Neoscytalidium dimidiatum.(Penz.)Crous&Slippers〕、茎腐病〔Fusarium solani(Mart.)Sacc.〕发生最为严重,其它炭疽病(Colletotrichum gloeosporioides Penz)、茎斑病(Phomosis sp.)、灰霉病(Fusarium equiseti(Corda)Sacc 〕、果腐病(Bipolaris sp.)、基腐病、病毒病(Cacts virus X、Zygocactus virus X、Pitaya virus X)、寒害和裂果等病害也均有发生,病害的发生严重威胁着海南地区火龙果产业的健康和长远的发展。2.部分病原真菌的生物学特性研究表明:新暗色柱节孢(N.dimidiatum)菌丝的最适生长温度为30℃～35℃,最适产孢温度为30℃,菌丝最适生长pH为7,产孢最适pH为8;PSA培养基为菌丝生长最适宜培养基,CA培养基最适合病原菌产孢;淀粉为病原菌菌丝生长的最适碳源,麦芽糖为碳源最有利于病原菌产孢;酵母粉为病原菌菌丝生长的最适氮源,病原菌不能利用尿素为氮源,蛋白胨作为氮源时最有利于病原菌产孢。拟茎点霉菌(Phomosis sp.)菌丝最适生长温度为30℃,最适pH为8,PSA、PDA和PDA+酵母粉培养基为最适菌丝生长培养基,病原菌菌丝在以淀粉和木糖作为碳源的培养基上生长速度最快,牛肉膏为菌丝生长最适氮源,尿素不能作为病原菌生长的氮源。菌丝生长的最适温度为25℃~28℃,温度为28℃时最适合病原菌产孢,菌丝最适生长pH为8～9,pH为6时产孢量最大,PDA培养基为最适菌丝生长培养基,LB培养最适合病原菌产孢;蔗糖、淀粉、葡萄糖、乳糖为菌丝生长的最适碳源,乳糖和木糖为病原菌产孢最适产孢碳源;酵母粉为菌丝生长的最适氮源,蛋白胨作为氮源时最有利于病原菌产孢。3.采用真菌rDNA-ITS通用引物ITS1和ITS4成功克隆了编号为:1409HSCA1、1410BSZL1、1410SMPF1的ITS1-4序列;经Blast比对和同源性分析结果显示,所获得的序列分别为新暗色柱节孢(N.dimidiatum)、拟茎点霉(Phomosis sp.)和木贼镰刀(F.equiseti)菌的rDNA-ITS序列,这与形态学鉴定的结果相符,说明可以将ITS序列分析作为快速、有效地鉴定火龙果溃疡病、茎斑病和灰霉病病原的辅助手段。4.部分病原真菌室内毒力测定结果表明:多菌灵对火龙果溃疡病病原菌新暗色柱节孢菌(N.dimidiatum)生长的抑菌效果最好,绿妃也表现出很好的抑菌效果,阿米西达对病原菌的抑制效果最差。多菌灵对火龙果茎斑病病原菌拟茎点霉菌(Phomosis sp.)生长的抑制效果最好,扑海因和甲基托布津也表现出较好的抑菌效果,而绿妃的抑菌效果最差。绿妃对火龙果灰霉病病菌木贼镰刀菌(F.equiseti)生长的抑制效果最好,必绿也表现出出色,达克宁的效果最差。5.应用RT-PCR病毒检测技术对海南地区火龙果病毒病疑似样本进行病毒种类检测,经测序分析、同源性比对和构建系统发育树后发现CVX-003 isolate Hainan和CVX004 isolate Hainan 属于 Cactus virus X部分基因序列,ZVX-501 isolate Hainan 属于Zygocactus virus X基因的的部分序列、PVX702 isolate Hainan属于Pitaya virus X的部分序列。由此可证明,海南地区火龙果病毒性病害疑似样本中至少存在Cactus virus X、Zygocactus virus X、Pitaya virus X3种病毒的复合侵染。更多还原

【Abstract】 Dragon fruit Hylocereus polyrhizus Webb.Britton&Rose(Cactaceae)commonly known as pitaya or strawberry pear,is widely cultured in tropical and subtropical regions of China and has become increasingly important fruit in recent years.native to the tropicalforest regions of Mexico and Central and South America Pacific Ocean coast.Prevalent form of cultivation until little more than Several decades was in small-scale family production.However,since ifs cultivation in commercial plantations and further domestication started in earnest,some symptoms of rotting and spots in stems and fruits were being observed.in some cases,but commercial production plots had to be abandoned due to the diseases.There were more and more reachies about dragon fruit diseases had been reported in the word.Aiming in such a situation,a more comprehensive and detailed study on the fungal and viral disease of dragon fruit all over the Hainan province was accomplishen in this program.The main conclusion of this thesis as follows:1 The inverstigation showed that there were 10 kinds of disease been found on dragon fruit,in whitch the canker disease caused by Neoscytalidium dimidiatum.(Penz.)Crous&Slippers,stem rot caused by Fusarium solani(Mart.)Sacc.anthracnose caused by Colletotrichum gloeosporioides Penz,stem spot caused by Phomosis sp.、sooty mould caused by Fusarium equiseti(Corda)Sacc、fruit rot caused by Bipolaris sp.、foot rot、viral disease caused by Cactus virus X、Zygocactus virus X and Pitaya virus X,chilling injury and fruit cracking.2.The study on biological characteristics of N.dimidiatum.showed the optimal conditions for mycelial and sporulation:the temperation was 30℃～35℃,pH was 7,the carbon sources was starch,the nitrogen sources were yeast extract powder,the medium was PSA medium formycelial growth;and the temperation 30℃～35℃,pH was 7,the carbon sources was starch,the nitrogen sources were yeast extract powder,the medium was PSA medium for mycelial growth,for mycelial growth was CA medium.Experiments on Phomosis sp.figured out the optimum conditions for mycelial growth:the temperature was 25 ℃～28℃;the pH was 8;the carbon sources were starch or xlose;the nitogen sourse was yeast extract powder and the fungal could not use the urea as the nitrogen.The related research on F.equiseti showed the optimum condintion for mycelial growth and sporulation:the temperature was 25℃～28℃and 28℃;the pH was 8～9and 6,the medium was PDA and LB,the carbon sources were lactose(for both),starch,glucose,sucrose(for growth only)and xlose(for porulation);the nitrogen sourses were diseaseyeast extract powder and beef extract.3.The PCR results of fungus rDNA-ITS amplificatiom using universal primer pair ITS1/ITS4 suggested that:the sequences of the suspected strains of 1409HSCA1、1410BSZL1、1410SMPF1,which were proved to be lacated in the rDNA-ITS of N.dimidiatum,Phomosis sp.,and F.equiseti after blast and homology analysis.We confirmed that,as a result,the rDNA-ITS analysis should be a dependable method for fungal pathogens of dragon fruit canker disease;stem spot and sooty mould desease.4.Indoor toxicity of six fungicides to N.dimidiatum,Phomosis sp.,and F.equiseti was tested by messuring mycelial growth rate.The result showed that all of the fungicides were able to inhibit the growth of fungal pathogens in our expriments.Duojunling was highliest toxic to N.dimidiatum,Lvfei after it,but Amixida showed very low toxity to the fungal pathogens.Duojunling was highliest toxic to Phomosis sp.,Rovral and Topsin-M after it,but Lvfei showed very low toxity to this fungal pathogens.Howere Lvfei was highliest toxic to F.equiseti,Bilv after it,but Daktarin showed very low toxity to the fungal pathogens.5.The RT-PCR detection showed that:These sequencese and its length were CVX-003 isolate Hainan(354 bp)、SVX-301 isolate Hainan(122 bp)、ZVX-501 isolate Hainan(233 bp)、PVX-701 isolate Hainan(322bp)、CVX004 isolate Hainan(923)、PVX702 isolate Hainan(876).And we known that CVX-003 isolate Hainan and CVX004 isolate Hainan classified as the partial sequences of Cacts virus X,ZVX-501 isolate Hainan classified as the partial sequences of Zygocactus virus X,PVX702 isolate Hainan classified as the partial sequences of Pitaya virus X,after blast and respectively homology analysis.Therefore,we considered that there’re at least 3 kind of virus inhabiting in virus-like symptomatic samples on dragon fruit stems.更多还原