海南兰花病毒病调查及病原检测

2007～2008年,对海南主要兰花种植基地进行病毒病危害情况调查。结果表明,海南兰花病毒病症状复杂,以斑驳花叶型和坏死型最为普遍。调查还发现,兰花的病毒病发病率与龄期呈正相关,文心兰、石斛兰的发病率显著高于蝴蝶兰。病原血清学检测表明,海南兰花病原病毒主要为建兰花叶病毒、齿兰环斑病毒,以及二者复合感染,检出率分别为29.5%、2.3%、15.9%。其中文心兰和蝴蝶兰带毒率较高,石斛兰带毒率最低,石斛兰上未检出齿兰环斑病毒。     

两种主要兰花病毒的快速检测初报

采用组织涂抹酶联免疫吸附技术，对本单位近年引入的部分优良洋兰品种进行２种主要病毒（即ＯＲＳＶ和Ｃｙｍｖ）的快速检测。结果表明，蝴蝶兰和心兰极易感染Ｃｙｍｖ病毒，带毒品种分别占被测品种的４５．５％和５７．１％；石斛兰对这２种病毒具有较强的抗性，大花蕙兰较易感染ＯＲＳＶ病毒。     

Control of virus diseases in intensively cultivated vanilla plots of French Polynesia

The results of long term virus surveys in intensively cultivated vanilla plots in the Society Islands (French Polynesia), between 1999 and 2007 are reported here. The data confirmed a potential for high incidence of aphid borne viruses in particular Cucumber mosaic virus (CMV) and Watermelon mosaic virus (WMV) as well as the non vectored Cymbidium mosaic virus (CymMV). CMV had a particularly high prevalence (over 30% of the plots) and could severely damage up to 50% of the vines before blossom. Severe outbreaks of CMV were correlated to the presence of the weed Commelina diffusa as a reservoir of virus and aphid vectors. The application in 2003 of simple prophylactic measures resulted in a sharp reduction of virus incidence in new plantations, compared to levels of virus diseases recorded in the previous decade. Indeed, at the beginning of fruiting, the incidence of aphid borne viruses did not exceeded 1.6% of vines for the 29 shade houses which adopted the virus prophylaxis. The three remaining shade houses were severely (20–50%) infected by WMV or CymMV because of the planting of virus-infected cuttings. These results demonstrate the benefit of implementing prophylaxis at the scale of an archipelago, and widen the possibilities of developing intensive cultivation of vanilla in other areas confronted with similar virus constraints.     

海南岛兰科植物区系的组成及其特征

海南岛是我国兰科植物分布最集中的地区之一,目前已发现有78属214种。其中,中国特有种31种(包括海南特有种14种),非中国特有种但在中国仅见于海南岛的有20种。种数在10种以上的属有羊耳蒜属(Liparis)、石斛属(Dendrobium)、毛兰属(Eria)、石豆兰属(Bulbophyllum)和兰属(Cymbidium)。海南岛兰科植物属的地理成分可划分为12个分布型和变型,其中热带亚洲成分占总属数的51.3%,热带分布属有71属,占绝对优势。海南岛兰科植物的生活型多样,附生兰有43属,地生兰有32属,腐生兰5属(其中羊耳蒜属和兰属中存在附生和地生2种生活型)。多个种在叶型和花色上变异较大,可作为优良育种材料。与邻近地区相比,海南岛兰科植物与西双版纳共有属最多(为56属),其次依序为台湾、广东、广西和福建。虽然海南岛与台湾和西双版纳同处热带北缘,受热带季风气候影响,但其兰科植物在属、物种和特有种的数量上比后两者要少。总的说来,海南岛的兰科植物区系较为多样,与亚洲热带兰科植物区系亲缘关系密切。     

海南岛兰花种质资源考察报告

对海南岛15个县(市)不同生态类型区兰花种质资源进行了全面的考察,共搜集了野生兰57属161种(变种、型),栽培兰14属(剔除与野生种重复)50余个品种,其中海南特有种21个,东亚特有属8个,海南新记录属6个,海南新记录种13个,保存活种质180余种(品种、型),10多种野生兰有观赏价值。通过实地考察,基本摸清了海南岛兰花种质的分布(水平分布和垂直分布)、生态环境、多度、花果期及形态特征等情况。     

Comparative levels of potato viruses S and Y infection of microplants and tuber-propagated plants in the field

The practice of transplanting microplants from tissue-culture to the field was compared to normal tuber propagation with respect to the transmission and translocation of potato virus S (PVS) and potato virus Y (PVY) to the daughter tubers of Red Pontiac, Shepody, Kennebec, and Russet Burbank cultivars in Prince Edward Island in 1984 and 1985. In general, the use of microplants to produce seed stocks appeared to increase the risk of infection with both viruses, although the results for tuber infection with PVY were only significant in one of the years.     

海南番木瓜PRSV和PLDMV病毒发生情况及分子鉴定

对海南昌江、乐东、澄迈、文昌和三亚5个主要番木瓜产区的病毒病发生情况进行调查。对76份番木瓜疑似病毒样品进行检测,检测结果表明海南番木瓜病毒病主要有2种：由番木瓜环斑病毒（Papaya ring spot virus,PRSV）引起的番木瓜环斑病毒病和由番木瓜畸形花叶病毒（Papaya leaf-distortion mosaic virus,PLDMV）引起的番木瓜畸形花叶病毒病。其中,番木瓜环斑病毒病最为常见,检出率达77.63％,是影响海南地区番木瓜产业健康发展的主要病毒病;畸形花叶病毒病发生率很低,仅检出2例番木瓜畸形花叶病毒病样品。以PRSV病毒基因组P1和Hc-Pro作为靶标基因进行分子克隆、测序,结果表明海南地区PRSV病毒可明显分为2个株系：HN-PRSV-1株系和HN-PRSV-2株系。此外,基于CP基因的系统进化树分析结果表明在海南新发现的PLDMV病毒可能源于台湾和日本。本研究可为开展番木瓜花叶病的防控和创制抗花叶病毒番木瓜新种质提供数据参考。     

海南岛花卉种质资源考察报告

1987—1990年考察了海南岛17个县市的花卉资源,共搜集花卉种质859种(变种、品种、型),内含野生种406种,栽培种453种,隶属97科,390属。其中,海南特有种47种,兰科植物东亚特有属3个,海南新记录属5个,新记录种13个;保存活种质713份。这次考察基本摸清了海南岛花卉种质资源的分布、生态、环境、花果期、主要性状和分类名称。最后,推荐十余种观赏价值较高的种类,供研究和利用。     

3个甘蔗与斑茅远缘杂交后代BC1的染色体遗传分析

对3个甘蔗与斑茅远缘杂交后代BC1进行真实性鉴定及染色体核型分析,以探讨甘蔗与斑茅BC1的染色体传递方式。利用2对鉴定斑茅真实杂交后代的特异引物对3个甘蔗与斑茅BC1进行鉴定,采用根尖分生区细胞去壁低渗涂片法制片,显微拍照计数染色体数目,并进行染色体核型分析。3个BC1材料均为斑茅的真实杂交后代,崖城01-69体细胞染色体核型公式为2n=121=120 m+1 sm,其染色体按2n+n方式传递;崖城01-116的体细胞染色体核型公式为2n=122=118 m+4 sm,其染色体传递方式为2n+n;崖城01-134的体细胞染色体核型公式为2n=121=120 m+1 sm,其染色体传递为2n+n。推断甘蔗与斑茅BC1的染色体以2n+n的方式传递。     

海南岛黄瓜病毒病种类鉴定及其发生分布研究

2018年11月—2019年3月,在海南省三亚、乐东、东方、儋州、澄迈、海口、万宁和五指山共8个市县,采集疑似病毒病的叶片样本302份,采用小RNA深度测序和RT-PCR检测发现,黄瓜样品中存在黄瓜花叶病毒（Cucumber mosaic virus, CMV）、甜瓜黄斑病毒（Melon yellow spot virus, MYSV）、黄瓜绿斑驳花叶病毒（Cucumber green mottle mosaic virus, CGMMV）、烟草花叶病毒（Tobacco mosaic virus, TMV)和瓜类褪绿黄化病毒（Cucurbit chlorotic yellows virus, CCYV）共5种病毒,检出率分别为32.79%、31.46%、23.18%、9.61%和2.65%。病毒的复合侵染率为11.28%,有8种复合侵染类型,以其中2种病毒复合侵染为主。明确了海南岛黄瓜病毒病主要种类及发生分布情况。     

Visual inspections of nursery stock fail to protect new plantings from Blueberry scorch virus infection

Blueberry scorch virus (BlScV) is one of the most pervasive pathogens of highbush blueberry. The virus is aphid-vectored and exhibits a latent period between infection and symptom expression in the host plant of up to 5 years. In many cases, we have observed BlScV symptom expression in new fields that appears inconsistent with aphid-vectored introduction and spread. It was, therefore, speculated that the virus may be introduced through infected nursery stock. To examine this possibility, we first surveyed selected nurseries to determine if mother plants, used for propagation by cuttings, were BlScV-infected. Two nurseries were found to harbor symptomless, infected, mother plants (cv. Duke). Cuttings from one nursery were collected from infected and non-infected plants and rooted in propagation beds. The survival and infection of cohorts from each mother plant were determined one year after planting. A significantly greater proportion of cuttings survived from non-infected mother plants (0.7) than from infected mother plants (0.5). Of the cohort from infected mother plants that survived, 40% tested positive for BlScV. We also surveyed the distribution of infected, symptomatic plants in recent ‘Duke’ plantings that originated from nurseries with BlScV-infected mother plants and compared distribution with older plantings with more advanced BlScV outbreaks. In all cases, the distribution of BlScV symptom development in young fields was random, which is consistent with introduction from planting stock. Older plantings showed a strong clustered distribution, which is consistent with aphid transmission. This study identifies infected nursery stock as an important source of BlScV dissemination and underscores the importance of having symptomless mother plants virus tested.     

Distribution of tobacco rattle virus and potato virus X in leaves,roots, and fruits and/or seeds of naturally-infected weeds

Weeds in tobacco rattle virus (TRV)-infested potato fields were examined for TRV and potato virus X (PVX) with the enzyme-linked immunosorbent assay. Questionable positive reactions were evaluated with immunosorbent electron microscopy. Of 615 weeds, representing 28 species, sampled over a two-year period, four species contained only TRV, three were infected only with PVX and eight had both viruses. Tobacco rattle virus was located only in the roots of five species and PVX was found only in the foliage of five species. Seeds and/or fruits of three species contained TRV while two contained PVX. Only one weed species,Solanum sara choides, contained both viruses in all plant portions tested. Potato virus X was not detected in weeds within a field planted with PVX-free potato seed.     

建兰花叶病毒CP基因的原核表达及抗血清制备

通过间接酶联免疫检测和电镜观察对从福建省漳州市采集的卡特兰病样进行检测,证明样品感染了建兰花叶病毒。设计一对特异性引物,扩增并克隆病毒分离物的外壳蛋白基因,随后将目的基因插入pET-29a（＋）中构建相应的原核表达载体。目的蛋白经诱导表达及纯化后免疫家兔并获得了特异性抗血清。Westernblot检测结果表明,抗血清与诱导表达的CyMV外壳蛋白发生特异性反应。间接酶联免疫法检测结果表明,抗血清可检测病汁液的最低稀释度达1∶51200,最佳工作浓度为1∶1000,病汁液灵敏度为0.39mg/mL,而与TMV等11种同源或异源病毒均无明显的血清学交叉反应。     

海南50个甘蔗品种黄叶病毒的分子鉴定

为明确国家甘蔗体系集成示范及区试甘蔗品种在海南省被甘蔗黄叶病毒(SCYLV)侵染情况及病毒基因型类型,从集成示范及区试的50个甘蔗品种上采集带有显著病症或不显病症样品50份,采用特异引物通过RT-PCR方法进行甘蔗黄叶病毒检测和病毒基因型鉴定。结果表明:50个甘蔗品种中有31个被检测到SCYLV,检出率为62%;病毒基因型有古巴(CUP)基因型、巴西-秘鲁(BAR-PER)基因型和留尼汪岛(REU)基因型3种,主要以CUP基因型为主,占58.06%,其次是BAR-PER基因型,占51.61%,REU基因型最少,占29.03%;未发现CHN1、CHN2、CHN3基因型。另外,不同基因型混合侵染现象普遍存在,总混合侵染率达32.26%。可见,海南国家甘蔗体系集成示范及区试甘蔗品种严重感染甘蔗黄叶病毒(SCYLV),且存在不同基因型混合侵染,建议推广种植脱毒种苗来控制甘蔗黄叶病的发生。本研究为在海南蔗区推广和种植甘蔗脱毒种苗及抗病育种提供了理论支持。      

Naturally occurring viral infections inSolanum brevidens andS. fernandezianum

Summary Two species of wild potato,S. brevidens andS. fernandezianum, were surveyed for six potato-infecting viruses in their natural habitats in the Puerto Montt Region and Robinson Crusoe Island of Chile, respectively. Potato viruses S (PVS), M (PVM) and X (PVX) and potato yellowing virus (PYV) were found in some clones of the Chilean Potato Germplasm Collection, whereas only one population ofS. brevidens out of six was virus-infected, namely by PVS. The cultivated potatoes on Robinson Crusoe Island were infected by PVS, PVM and PVX and infested byAulacorthum solani andMyzus persicae, whereasS. fernandezianum was infected by PYV and infested byA. circumflexum, A. solani andAphis spp.     

基于GEE和Landsat时间序列数据的海南岛土地利用分类研究

土地利用/覆盖变化（land use/cover change,LUCC）是当前全球变化研究的核心内容之一。土地利用遥感监测是土地利用变化相关研究的重要技术手段,尤其是高分辨率遥感技术和谷歌地球引擎（Google Earth Engine,GEE）云计算平台的出现,为土地利用空间信息的获取提供了新的途径和方法。本研究基于GEE云平台提供的Landsat-8 OLI时间序列卫星影像数据,采用随机森林（random forest,RF）和支持向量机（support vector machines,SVM）分类算法,对海南岛土地利用类型进行了遥感分类研究。结果表明：RF与SVM算法对海南岛土地利用中水体和建筑用地的分类精度均较高,对耕地、园地和林地分类精度较低。与SVM方法相比,RF分类方法能够更准确识别各类地物信息,更适于海南岛土地利用分类的研究。海南岛林地（包括天然林、橡胶林等）所占比例最大,主要分布在海南岛中部;耕地和园地面积接近,相间分布于海南岛大部分区域;水体和建筑用地面积较小,在海南岛均呈零散的分布状态,以沿海地区为主。GEE平台对于开展大区域土地利用分类与遥感动态监测具有重要的意义。     

建兰花叶病毒RT-LAMP检测方法的建立

建兰花叶病毒（Cymbidium mosaic virus,CyMV）是侵染兰花的主要病毒,严重影响其观赏价值,建立快速、灵敏的检测方法显得尤为重要。根据CyMV的外壳蛋白基因序列设计4对特异性引物,经过优化反应条件,建立该病毒的RT-LAMP 检测方法,并进行LAMP检测的特异性、敏感性检测。该方法能特异扩增CyMV,与其他4种病毒（齿兰环斑病毒、菜豆黄花叶病毒、黄瓜花叶病毒和小苍兰花叶病毒）不发生反应;灵敏度为RT-PCR的10倍。田间检测20份样品中,RT-LAMP和RT-PCR检测结果一致,检出率为60%。在产物中加入荧光染料SYBR GreenⅠ,直接用肉眼观察就可判断样品是否感染CyMV,可省去电泳分析的时间。针对CyMV建立的RT-LAMP方法具有特异性强、灵敏度高、操作简单、快速等特点,适用于在进境检疫及种苗繁育过程中的检测鉴定。     

海南野生疣柄魔芋组培快繁技术

为开发利用海南本地魔芋种质资源,以海南本地野生疣柄魔芋球茎顶芽为外植体,研究其组培快繁技术。结果表明：外植体消毒以在0.1% HgCl2溶液中浸泡20 min为宜;采用MS＋6-BA 3.0 mg/L＋NAA 0.1 mg/L＋蔗糖30 g/L＋琼脂7 g/L培养基,较有利于从顶芽基部直接诱导出丛芽,且增殖系数较高,达4.37;而丛芽分切成单芽后,在MS＋NAA 0.5 mg/L＋蔗糖30 g/L＋琼脂7 g/L＋活性炭1 g/L培养基上的生根率可达100%。