水稻黑条矮缩病抗性评价方法及抗性资源筛选

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王宝祥, 胡金龙, 孙志广, 宋兆强, 卢百关, 周振玲, 樊继伟, 秦德荣, 刘裕强, 江玲, 徐大勇, 万建民. 水稻黑条矮缩病抗性评价方法及抗性资源筛选. 作物学报, 2014, 40(9): 1521-1530[WANG Bao-Xiang, HU Jin-Long, SUN Zhi-Guang, SONG Zhao-Qiang, LU Bai-Guan, ZHOU Zhen-Ling, FAN Ji-Wei, QIN De-Rong, LIU Yu-Qiang, JIANG Ling, XU Da-Yong, WAN Jian-Min. An Evaluation System for Rice Black-Streaked Dwarf Virus Disease and Screening for Resistant Rice Germplasm. Acta Agronomica Sinica, 2014, 40(9): 1521-1530] 复制到剪切板

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水稻黑条矮缩病抗性评价方法及抗性资源筛选

王宝祥¹, 胡金龙², 孙志广², 宋兆强², 卢百关¹, 周振玲¹, 樊继伟¹, 秦德荣², 刘裕强², 江玲², 徐大勇^1,^*, 万建民^2,^3,^*

¹江苏徐淮地区连云港农业科学研究所, 江苏连云港222006

²南京农业大学 / 作物遗传与种质创新国家重点实验室, 江苏南京210095

³ 中国农业科学院作物科学研究所, 北京100081

^* 通讯作者(Corresponding authors): 万建民, E-mail:wanjm@njau.edu.cn, Tel: 025-84396516; 徐大勇, E-mail:xudayong3030@sina.com, Tel: 0518-85589316

收稿日期:2014-06-16

基金:本研究由国家科技支撑计划项目(2013BAD01B02-16), 国家公益性行业(农业)科研专项(201003031), 江苏省自然科学基金项目(BK2011403), 江苏省科技支撑计划项目(BE2013301), 江苏省农业科技自主创新资金(CX[12]5112)和江苏省农业三项工程项目(sx[2011]003)资助;

摘要

水稻黑条矮缩病是水稻主要病毒病害之一。目前由于缺乏规模、高效的黑条矮缩病抗性鉴定体系, 制约了抗黑条矮缩病水稻资源的发掘, 限制了抗黑条矮缩病的育种进程和基础研究。本研究通过分析水稻黑条矮缩病田间鉴定所需灰飞虱的有效接种密度、带毒率及播期等, 提出水稻黑条矮缩病田间鉴定有效接种的灰飞虱密度在800万头 hm^-2左右较为合理, 而带毒率应不低于5%。并进一步对现有黑条矮缩病人工接种鉴定的循回期、接种虫量、接种时间及虫龄等进行了优化。利用上述鉴定体系, 2010年对来源于20个国家的共1240份水稻种质进行黑条矮缩病田间鉴定, 初步获得发病率低于10%的品种34个; 2011、2012连续两年对该34个品种进行多年多点重复抗性鉴定, 发现来自东南亚地区的3个品种Kanyakumari 29、Madurai 25和Vietnam 160连续3年发病率均低于10%, 表现较高的黑条矮缩病的抗性。进一步分期播种鉴定的结果表明, Kanyakumari29在3个播期、3个鉴定点的发病率均低于12%, 而Madurai 25和Vietnam 160发病率均低于9%。此外, 在人工接种条件下Kanyakumari 29、Madurai 25和Vietnam 160的发病率均低于9%。因此, 多年多点田间鉴定和人工室内接种鉴定的结果均表明, Kanyakumari 29、Madurai 25和Vietnam 160稳定、高抗黑条矮缩病。综上所述, 本研究建立的田间鉴定与室内鉴定相结合的黑条矮缩病鉴定体系准确、可靠, 可用于黑条矮缩病的大规模鉴定, 该体系的建立及高抗黑条矮缩病水稻资源的发掘为水稻抗黑条矮缩病基因的鉴定及育种利用提供了重要的方法和材料基础。

关键词: 水稻; 黑条矮缩病; 抗性鉴定

An Evaluation System for Rice Black-Streaked Dwarf Virus Disease and Screening for Resistant Rice Germplasm

WANG Bao-Xiang¹, HU Jin-Long², SUN Zhi-Guang², SONG Zhao-Qiang², LU Bai-Guan¹, ZHOU Zhen-Ling¹, FAN Ji-Wei¹, QIN De-Rong¹, LIU Yu-Qiang², JIANG Ling², XU Da-Yong^1,^*, WAN Jian-Min^2,^3,^*

¹Institute of Lianyungang Agricultural Science of Xuhuai Area, Lianyungang 222006, China

² State Key Laboratory of Crop Genetics and Germplasm Enhancement / Nanjing Agricultural University, Nanjing 210095, China

³ Institute of Crop Science, Chinese Academy of Agricultural Sciences, Beijing 100081, China

Fund:

Abstract

Rice black-streaked dwarf virus disease (RBSDVD) is one of the most serious viral diseases. The absence of efficient resistance evaluation system for RBSDVD restricts the excavation of the resistance resource, thereby the resistance breeding and basal researches are limited. Here, we proposed that the appropriate density of small brown planthopper (SBPH) should be 8 × 10⁴ ha^-1 and the rate of RBSDV-carrying SBPH should be over 5% for RBSDVD evaluation in field condition. Further, we optimized the artificial inoculation evaluation system including circulative period, number of SBPH per plant, infestation time and age of SBPH. We evaluated 1240 varieties from 21 countries for RBSDV resistance by natural inoculation in 2010, 34 cultivars had the disease incidence less than 10%. The 34 cultivars were further evaluated by natural inoculation in three different locations in 2011 and 2012, respectively. Finally, only three varieties, Kanyakumari 29, Madurai 25, and Vietnam 160, which all came from Southeast Asia, had the consistent disease incidence less than 10% in different locations and years. To verify the resistance of the three varieties against RBSDVD, we carried out the interval sowing inoculation experiment in the three different locations in 2012. In the interval sowing experiment, the disease incidence of Kanyakumari 29 was less than 12%, whereas Madurai 25 and Vietnam 160 less than 9%. In addition, in the artificial field inoculation and artificial room inoculation experiments, the disease incidence of the three varieties was less than 9%. The above natural and artificial inoculation experiments showed that Kanyakumari 29, Madurai 25, and Vietnam 160 performed stable and high RBSDVD resistance. Taken together, the RBSDVD evaluation system combining field with artificial infestation identifications used in this study is accurate and reliable. This system could be used to widely screen RBSDVD resistance resources. The construction of RBSDVD evaluation system and the RBSDVD resistance resources identified in this study will provide the useful tools and materials for the identification of RBSDVD resistance gene(s) and development of RBSDVD resistance varieties.

Keyword: Rice (Oryza sativa L.); Rice black-streaked dwarf virus; Resistance identification

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水稻黑条矮缩病(rice black-streaked dwarf virus disease, RBSDVD)是由水稻黑条矮缩病毒(rice black-streaked dwarf virus, RBSDV)引起的, 主要由灰飞虱以持久性不经卵方式传播^{[ 1, 2]}。近年来, 随着耕作制度和栽培方式的变化、冬季气候变暖和感病(虫)品种的大面积推广, 传毒介体灰飞虱发生量日益上升, RBSDVD在江苏、浙江、江西和福建大规模发生, 并迅速上升为生产上最主要的病毒病害, 给水稻的安全生产带来很大威胁^{[ 3, 4, 5, 6]}。RBSDVD的典型症状是病株矮缩, 叶色浓绿僵硬, 叶背、叶鞘和茎秆有早期蜡白色, 后期为黑褐色的短条纹不规则突起, 严重发病株不抽穗。目前主要是依赖化学农药控制介体灰飞虱来防治RBSDVD, 但由于灰飞虱的迁飞习性及传毒的瞬时性, 防治效果有限。应用抗性品种被认为是最经济有效的防治措施。

抗性资源的鉴定与发掘, 是培育抗性品种和开展抗RBSDVD基础研究的先决条件。但由于RBSDVD为虫传病害, 且不经卵传毒, RBSDVD的抗性鉴定, 不仅需要大量的灰飞虱虫源, 而且每一代灰飞虱还需带毒处理, 鉴定较为复杂, 导致现有的鉴定方法准确度不高。虽然周彤等^{[ 7]}建立了黑条矮缩病室内人工接种方法, 对黑条矮缩病的鉴定具有重要参考价值, 但室内鉴定费时、费事, 工作量大, 难以满足规模化抗源筛选的迫切需要。目前对黑条矮缩病的鉴定主要是依赖田间自然接种鉴定, 但田间鉴定受灰飞虱接种密度和带毒率等因素制约, 往往重复性、准确性较差。对于如何优化田间鉴定条件、提高鉴定结果的可靠性和准确性缺乏系统性的研究。

前期的研究结果表明多数水稻品种易感RBSDVD, 目前尚未发现高抗水稻黑条矮缩病的水稻品种^{[ 8]}, 虽然少数水稻资源具有一定的RBSDVD抗性, 并检测到一些微效抗性QTL^{[ 9, 10, 11]}。但抗性水平不高, 且QTL效应也相对较低, 难以在水稻育种中利用。因此, 迫切需要建立高效、准确的抗性鉴定体系, 发掘稳定、高抗RBSDVD的水稻资源。本研究对灰飞虱接种密度及带毒率和接种时期对水稻黑条矮缩病田间鉴定的影响进行了探索, 提出了田间鉴定的最佳条件, 并对现有的室内人工接种鉴定方法进行了优化; 进一步的田间和室内筛选鉴定获得了3份稳定、高抗黑条矮缩病的水稻种质。上述鉴定方法的建立及抗源的获得为后续开展抗水稻黑条矮缩病基因的定位、克隆和育种利用奠定了基础。

1 材料与方法

1.1 供试材料

供试材料为来自21个国家的1240份水稻资源, 其中粳稻品种620份, 籼稻品种620份(表1)。武育粳3号为感病对照品种。

表1 试验材料统计 Table 1 Number of tested materials

1.2 田间鉴定

1.2.1 试验材料的田间种植于江苏省东海县黄川镇、赣榆县土城镇和灌云县东辛农场试验田, 选用四周为麦田的地块种植待鉴定材料, 以保证获得足量虫源。2010年和2011年材料均在5月10日(麦收前3周)分2个重复播种, 每个重复单个品种播80粒, 均匀撒播1行, 行长50 cm, 行距10 cm。6月27日移栽, 单苗栽插, 行株距15 cm × 20 cm, 每个品种栽插60穴。常规肥水管理, 不施用任何农药。2012年于5月5日、5月10日和5月15日3个播期, 每个品种播150粒, 移栽时间分别为6月20日、6月25日和6月30日, 每个品种栽插120穴, 其他种植方式与2010年和2011年相同。

1.2.2 田间发病率调查于7月下旬水稻分蘖盛期统计发病率。发病植株极明显矮缩、心叶突破下叶叶鞘而出、部分植株从下叶枕口呈螺旋状伸出、部分叶片的顶端出现旋状卷曲; 而健康稻株分蘖旺盛, 生命力强, 但尚未拔节, 发病植株和健康植株反差非常明显; 另外, 此时的发病植株很少有中间型病状。以每个试验点2个重复的发病百分率平均值作为品种抗黑条矮缩病表型值进行统计。

1.2.3 田间灰飞虱密度调查首先通过盘拍法调查灰飞虱虫口密度, 将33 cm × 45 cm × 5 cm的搪瓷盆对准秧苗基部轻拍植株, 将灰飞虱拍落于盘中, 然后迅速将落入盘中的灰飞虱倒入高约为60 cm, 直径约为35 cm的塑料桶内。于6月上旬在种植材料田块对角线上定5点取样, 每点拍0.30 m², 记载灰飞虱数量。

1.2.4 水稻黑条矮缩病田间有效接种的灰飞虱密度和带毒率要求试验因为各个试验点灰飞虱带毒率不同, 在各个试验点通过改变灰飞虱接虫量来研究水稻黑条矮缩病田间有效接种的灰飞虱密度和带毒率要求。2011年在连云港市东辛农场、东海县黄川镇和赣榆县土城镇3个试验基地分别选取6块6.67 m²方地, 用防虫网把四周和上面封住, 形成方形笼子, 使灰飞虱无法入内。于5月12日至15日在隔离好的笼子内种植2000粒武育粳3号。于6月10日分别按接种密度200万头 hm^-2、400万头 hm^-2、600万头 hm^-2、800万头 hm^-2、1000万头 hm^-2和1200万头 hm^-2接入灰飞虱。本试验分为2个重复, 取平均值。本试验播种日期、移栽和查病方式同1.2.1。

1.2.5 水稻黑条矮缩病对产量损失的评价 2011年在东海县黄川镇对苗期选择性地遮盖防虫网, 以不同小区间不同遮盖程度导致不同接种量, 最终导致小区间发病率差异。5月15日播种, 6月25日插秧, 行距15 cm, 株距20 cm, 每个小区栽插400株。7月25调查发病率。成熟后按小区测产。

1.3 灰飞虱人工接种鉴定体系

1.3.1 低龄灰飞虱的饲养将饲养灰飞虱的品种武育粳3号(感灰飞虱品种即可)浸种催芽, 均匀播于70.5 cm (长) × 50.5 cm (宽) × 41.5 cm (高)的塑料箱内, 底部接塑料管通水, 保持土层湿润。当箱内秧苗长至2.5~3.0叶时接入田间或实验室饲养的成虫灰飞虱, 在箱内孵化幼虫, 保持室内26℃恒温。因黑条矮缩病病毒不经灰飞虱卵传毒, 所以孵化出的灰飞虱若虫不带毒。

1.3.2 箱内灰飞虱饲毒将田间或实验室内处于分蘖盛期发病的水稻植株移至20.5 cm (长) × 13.5 cm (宽) × 13.5 cm (高)的塑料桶内(底有一小孔, 便于透水), 然后移至塑料箱内, 同时向箱内接入孵化出的1~2龄灰飞虱若虫, 饲毒72 h, 接虫量为鉴定苗数的3倍。

1.4 灰飞虱及其带毒率检测

参照周彤等^{[ 12]}方法随机采集100头灰飞虱检测水稻黑条矮缩病毒, 确定带毒率。

2 结果与分析

2.1 田间灰飞虱数量及带毒率调查

经调查, 2010、2011和2012年黄川、土城及东辛3个试点秧田的虫口密度均在800万头 hm^-2以上。进一步对3个试验点接种时期的灰飞虱带毒率测定表明, 黄川试验田灰飞虱带毒率最高, 3年均在6%和9%之间, 其他2个试验点除2010年土城试验点带毒率达到7.0%外, 均在3.0%和5.0%之间(表2)。

表2 试验秧田灰飞虱密度和带毒率 Table 2 Density and RBSDV carrying rate of the SBPH in the experimental seedling bed

2.2 灰飞虱接种密度及其带毒率对黑条矮缩病田间鉴定效果的影响

在不同带毒率接种条件下, 通过人工补虫或驱虫等措施, 分析了灰飞虱接种密度及其带毒率对黑条矮缩病田间鉴定效果的影响。自然带毒率分别为3%、5%和8%时, 接虫密度从200万头 hm^-2到1200万头 hm^-2时, 武育粳3号发病率分别为15%~41%、16%~67%和19%~79% (表3)。上述结果表明, 田间发病率与灰飞虱密度及其带毒率呈正相关。同时, 随着接虫密度的增加, 死苗率也显著增加, 如接虫量从800万头 hm^-2上升至1200万头 hm^-2时, 武育粳3号死苗率从16%、14%和23%上升到29%、26%和41% (表3), 死苗率的增加将大大影响黑条矮缩病的鉴定结果, 因此在增加发病率的同时还需严格控制接虫密度, 以减少死苗率对黑条矮缩病鉴定结果的影响。分析发现在灰飞虱虫量为800万头 hm^-2条件下, 灰飞虱带毒率为3%时, 武育粳3号发病率仅为26%, 发病较轻; 而带毒率为5%和8%时, 发病率分别达到了48%和64%, 而此时死苗率仅为14%和23%, 能够满足黑条矮缩病的鉴定要求。因此, 水稻黑条矮缩病田间有效接种灰飞虱的密度在800万头 hm^-2左右较为合理, 而带毒率应不低于5%, 期间间隔赶虫, 以增加毒虫的传毒机率, 能进一步提高鉴定效率。

表3 虫量及带毒率对武育粳3号发病率和死苗率的影响 Table 3 Effect of SBPH number and RBSDV carrying rate of SBPH on incidence and mortality rate of Wuyujing 3

2.3 调整播种时期提高黑条矮缩病的鉴定效率

带毒灰飞虱由麦田迁飞到鉴定秧田的时期与适龄稻苗的对接, 是影响田间发病的关键。为寻求最佳播种时期, 分别于5月5日、5月10日和5月15日分期播种, 考察武育粳3号的田间发病率。结果发现, 黄川、土城和东辛3个试点均得到了类似的结果, 5月10日播种的田块武育粳3号的发病率最高, 分别达到了74%、61%和40% (表4)。6月1日、6月5日和6月10日秧田灰飞虱虫量的调查结果表明, 6月1日前后灰飞虱虫口密度最大(表2), 此时, 第一期播种的秧龄较大(约2~3叶), 错过了最佳接种时期; 而由于第3期播种的秧龄相对较小(约0.5~1.0叶), 高密度灰飞虱迁飞后, 其死苗率较高, 可能是影响其发病率降低的主要原因; 5月10日播种的秧龄约为1~2叶, 正处于最佳接种时期, 所以其发病率相对较高。

表4 分期播种条件下武育粳3号的黑条矮缩病发病率 Table 4 Rice black-streaked dwarf virus disease incidence of Wuyujing 3 in different sowing dates

2.4 黑条矮缩病室内鉴定条件的优化

本研究采用的人工室内接种鉴定的操作流程如图1所示, 人工饲养的1~2龄无毒灰飞虱若虫, 移至黑条矮缩病病株饲毒, 将待鉴定水稻品种约10粒播于直径5.8 cm、高6.0 cm的圆形塑料钵中(钵底有一小孔, 便于透水), 一叶期淘汰病弱苗, 每钵保留5株整齐一致的健苗, 每个品种共鉴定100棵苗, 次日将待鉴定材料与带有饲毒灰飞虱的病株移至同一塑料箱内, 间隔12 h驱赶1次灰飞虱, 使带毒灰飞虱能在苗间相互传毒, 增加传毒机率, 持续10 d, 之后将饲毒后的灰飞虱全部赶至鉴定植株上, 移走饲毒病株, 饲毒后的灰飞虱继续在鉴定植株上接种2 d, 之后移走灰飞虱, 此时利用RT-PCR方法检测灰飞虱带毒率。最后将鉴定植株移栽至大田, 待其发病。

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图1 黑条矮缩病人工接种鉴定流程图A: 室内养虫; B: 病苗; C: 虫箱内饲毒; D: 室内接种(7 d); E: 移植到田间。Fig. 1 The flow graph of artificial infestation for RBSDVD evaluationA: rearing SBPH in the room; 2: disease plant; C: virus acquisition in the SBPH rearing box; D: inoculation in the room (7 d);
E: cultivated in the field RBSDVD: rice black-streaked dwarf virus disease; SBPH: small brown planthopper.

为提高人工接种鉴定的准确性, 进一步分析了饲毒时间、接虫密度、接虫虫龄及水稻接种的龄期等因素对人工接种鉴定效果的影响。由图2-A可知, 5~20 d内, 随饲毒时间的延长, 发病率持续上升, 但延长至25 d后, 发病率并无显著提高, 表明人工接种鉴定的最佳饲毒时间为20 d左右。图2-B表明, 发病率随接虫量的增加而增加, 但虫量的加大也会影响植株的长势, 甚至导致死亡, 因此应避免过量接虫。3个不同发育时期接种带毒灰飞虱发现, 一叶期接种的发病率最高, 二叶期和三叶期无显著差异(图2-C)。不同龄期的无毒灰飞虱饲毒处理表明, 随着灰飞虱龄期的增加, 其传毒率呈降低趋势(图2-D)。综上所述, 黑条矮缩病人工室内接种鉴定, 采用1~2龄若虫, 饲毒处理20 d, 接种带毒灰飞虱密度为每株10头, 接种时间在水稻一叶期进行较为适宜。

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图2 不同接种条件对黑条矮缩病人工接种鉴定的效果比较A: 不同饲毒时间对发病率影响; B: 不同接虫密度对发病率影响; C: 水稻不同接种龄期黑条矮缩病发病率;
D: 灰飞虱接种龄期对发病率影响。Fig. 2 Comparison of artificial infestation efficiency in different conditionsA: effects of virus acquisition time on incidence; B: effects of small brown planthopper number on incidence;
C: effects of Leaf age on incidence; D: effects of instar on incidence.

2.5 不同黑条矮缩病发病率造成的产量损失

发病率为0~5%时, 产量损失率低于1%; 发病率6%~11%时, 产量损失0.8%~3.4%; 发病率12%~ 20%时, 产量损失4.1%~8.2%; 发病率23%~62%时, 产量损失10.1%~39.4% (表5)。上述结果表明, 产量损失率随发病率的增加而增加, 但产量损失率均低于发病率, 显示健康植株存在一定的补偿作用。

表5 不同黑条矮缩病发病率对产量的影响 Table 5 Effect of rice black-streaked dwarf virus disease
incidence on rice yield

2.6 抗黑条矮缩病水稻品种的筛选与差异比较分析

2010年3个鉴定试验点, 分别对来自21个国家的1240份水稻品种进行了黑条矮缩病的初步筛选鉴定, 分析了不同地区水稻品种对黑条矮缩病的抗性差异。分析鉴定品种发病率分布发现, 发病率低于10%的品种仅有34个, 主要分布在我国的华南、西南、华中和东南亚水稻种植区(表6), 发病率低于5%的品种主要为东南亚水稻品种, 我国的华北、西北、东北和其他地区(非洲、美洲等地区)的水稻品种发病率较高。另外, 江苏稻区水稻品种发病率主要集中在20%左右, 没有发现发病率低于10%的品种。2011年对2010年初步筛选到的发病率低于10%的水稻品种进行了重复鉴定, 结果发现仅有3个籼稻品种Madurai 25、Vietnam 160和Kanyakumari 29在连续2年6个点的鉴定中发病率均低于10%, 可作为重要种质资源进行进一步鉴定(表6)。

表6 34个水稻品种在不同年份间黑条矮缩病发病率差异 Table 6 Difference of RBSDVD incidence between 34 rice varieties in the different years (%)

品种 Variety	类型 Type	地区 Region	2010年黑条矮缩病发病率 Incidence of RBSDVD in 2010			2011年黑条矮缩病发病率 Incidence of RBSDVD in 2011
品种 Variety	类型 Type	地区 Region	黄川 Huangchuan	土城 Tucheng	东辛 Dongxin	黄川 Huangchuan	土城 Tucheng	东辛Dongxin
Kanyakumari 29	籼 Indica	印度India	0	4	1	10	0	5
Madurai 25	籼 Indica	印度India	5	1	1	7	3	0
Vietnam160	籼 Indica	越南Vietnam	1	0	3	10	5	3
测64 Ce 64	籼 Indica	中国江西Jiangxi, China	9	9	9	19	5	11
献改B Xiangai B	籼 Indica	中国江西Jiangxi, China	8	9	3	31	14	11
过冬青Guodongqing	籼 Indica	中国江西Jiangxi, China	7	9	7	18	5	5
锅底黑 Guodihei	籼 Indica	中国安徽Anhui, China	7	8	10	55	8	10
小红稻 Xiaohongdao	籼 Indica	中国安徽Anhui, China	5	9	4	24	21	21
当育5号 Dangyu 5	粳 Japonica	中国安徽Anhui, China	9	9	6	33	14	6
秋前白 Qiuqianbai	籼 Indica	中国安徽Anhui, China	9	8	7	17	20	16
黄丝桂占Huangsiguizhan	籼 Indica	中国广东Guangdong, China	9	9	6	57	11	22
闷加高 Menjiagao	粳 Japonica	中国海南Hainan, China	6	9	9	54	4	9
中9B Zhong 9B	籼 Indica	中国浙江Zhejiang, China	6	9	10	18	6	10
花谷 Huagu	籼 Indica	中国浙江Zhejiang, China	9	4	9	17	11	9
祥湖301 Xianghu 301	粳 Japonica	中国浙江Zhejiang, China	9	9	3	44	21	34
细麻线 Ximaxian	籼 Indica	中国云南Yunnan, China	9	8	5	66	13	24
剑粳6号 Jianjing 6	粳 Japonica	中国云南Yunnan, China	3	8	8	18	21	8
古154 Gu 154	粳 Japonica	中国云南Yunnan, China	8	9	9	51	0	9
Cambodia 37	籼 Indica	柬埔寨Cambodia	4	8	7	61	8	7
Cambodia 42	籼 Indica	柬埔寨Cambodia	4	0	9	42	9	9
IR2153-14-1-6	籼 Indica	菲律宾Philippines	6	5	8	18	24	9
IR13423-17-1-2-1	籼 Indica	菲律宾Philippines	3	0	4	13	14	22
Burmal47	籼 Indica	老挝Laos	8	0	9	43	34	9
Bp205f-kn-78-1	籼 Indica	印度尼西亚Indonesia	0	7	10	24	14	10
Batutegi 22	籼 Indica	印度尼西亚Indonesia	8	7	6	37	38	10
Gundi1 Ku	籼 Indica	印度尼西亚Indonesia	9	3	7	29	22	7
Lv-16	籼 Indica	越南Vietnam	9	0	9	35	22	9
秀水04 Xiushui 04	粳 Japonica	中国浙江Zhejiang, China	5	5	8	12	8	8
早单8号 Zaodan 8	粳 Japonica	中国江苏Jiangsu, China	8	7	6	44	24	8
云香粳 Yunxiangjing	粳 Japonica	中国云南Yunnan, China	8	5	7	57	17	19
吾别 Wubie	籼 Indica	中国云南Yunnan, China	5	8	7	38	15	7
雪禾矮早Xuehe’aizao	籼 Indica	中国云南Yunnan, China	6	8	6	27	21	6
木邦谷 Mubanggu	籼 Indica	中国云南Yunnan, China	9	8	5	31	19	24
武育粳3号Wuyujing 3	粳 Japonica	中国江苏Jiangsu, China	61	44	55	90	53	42

RBSDVD: rice black-streaked dwarf virus disease.

表6 34个水稻品种在不同年份间黑条矮缩病发病率差异 Table 6 Difference of RBSDVD incidence between 34 rice varieties in the different years (%)

2.7 抗性品种分期播种及人工接种验证

为进一步验证经田间鉴定筛选到的Kanyaku mari 29、Madurai 25和Vietnam 160黑条矮缩病抗性水平, 2012年对它们进行了分期播种(表7)和人工接种鉴定(图3)。在5月10日播期试验中, Kanyakumari 29在土城和东辛鉴定点的发病率分别为12%和11%, Madurai 25在黄川、土城和东辛3个鉴定点的发病率分别为7%、9%和7%, Vietnam 160在黄川和土城2个鉴定点的发病率分别为5%和6%, 除此之外, 其他所有播期和鉴定点中, 3个品种的发病率均低于5%, 这说明4个品种在不同鉴定年份、环境下发病率差异较小, 均表现出稳定的黑条矮缩病抗性。另外, 4个品种在5月10日播种发病率最高, 进一步表明在本地区进行黑条矮缩病鉴定, 5月10日左右是最佳的播种期。

为了排除田间环境和介体昆虫取食偏好对鉴定结果的影响, 进行了人工田间接种(接种虫源直接由田间搜集)和人工室内接种试验。结果表明, 在两种人工接种条件下, 感病对照品种武育粳3号的发病率在34%~48%之间, 表现较感黑条矮缩病, 而Kanyakumari 29、Madurai 25和Vietnam 160的发病率均低于9%, 进一步表明这3个水稻品种稳定、高抗黑条矮缩病(图3)。

表7 分期播种条件下4个抗性品种的抗性表现 Table 7 Resistance reaction of four rice varieties to rice black-streaked dwarf virus disease in different sowing dating

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	图3 人工接种4个品种对RBSDV的抗性表现A: 田间人工接种鉴定; B: 室内人工接种鉴定。Fig. 3 Resistant evaluation of 4 rice varieties to RBSDV by the artificial inoculationA: Artificial infestation in field; B: Artificial infestation in room. ** P< 0.01. 1: Wuyujing 3; 2: Kanyakumari 29; 3: Madurai 25; 4: Vietnam 160.

3 讨论

目前, 黑条矮缩病在江苏、浙江、上海、安徽、山东等长江中下游地区广泛发生, 已成为该地区主要水稻病毒病之一, 造成严重的经济损失^{[ 3, 4, 5, 6]}。培育与应用抗性品种是防治此类病害最为经济、有效的途径之一, 因此, 建立有效的黑条矮缩病鉴定体系, 筛选抗黑条矮缩病水稻种质资源是遏制这一病害流行的首要任务。灰飞虱密度和带毒率是水稻黑条矮缩病田间接种的关键因素, 同一个鉴定点的灰飞虱带毒率是不变的, 在此条件下, 我们在3个鉴定点(2011年发病率分别为3%、5%和8%)通过人为控制灰飞虱接种量研究田间有效接种的灰飞虱密度和带毒率要求。结果表明, 接虫量在1000万头 hm^-2时, 武育粳3号死苗率较高, 3个点的死苗率分别达到29%、26%和40% (表3), 而死苗率上升导致栽插苗减少, 无法对全部鉴定苗进行抗性评价, 影响鉴定结果。另外, 在灰飞虱虫量为800万头 hm^-2条件下, 灰飞虱带毒率为3%时, 武育粳3号发病率为26%, 发病较轻; 而带毒率分别为5%和8%时, 发病率达48%和64%, 满足鉴定感病对照发病率要求。因此, 水稻黑条矮缩病田间接种有效接种的灰飞虱密度在800万头 hm^-2左右较为合理, 而带毒率应不低于5%, 鉴定材料应在麦收前3周左右播种。

本研究在前人研究的基础上^{[ 7]}, 对黑条矮缩病室内人工接种方法进一步优化, 确定了黑条矮缩病人工室内接种鉴定, 以1~2龄若虫, 饲毒处理20 d, 在水稻一叶期每株接种10头较为适宜。虽然人工接种具有受外界环境干扰相对较小、准确性较高等优点, 在一定程度上可以弥补自然接种鉴定的不足, 但由于受到空间及带毒虫源等的影响, 难以实现规模化鉴定的需求。本研究为确保田间鉴定的成功率, 鉴定初期, 对灰飞虱重发区进行虫源调查, 确定最终鉴定田块, 并通过分期播种鉴定, 确定最佳播种时期一般在麦收前3周。在此条件下, 进行多年多点抗病种质资源的筛选与验证, 逐年淘汰感病品种, 最终筛选到的抗病种质再经人工接种鉴定进一步验证。本研究采用的前期大规模田间自然接种与后期室内人工接种相结合的鉴定策略, 既实现了快速、规模化鉴定的需求, 又确保了鉴定结果的可靠性。为今后进行黑条矮缩病资源的筛选提供了借鉴。此外, 本研究以粳稻品种“武育粳3号”为研究对象, 建立了发病率与产量损失之间的关系, 为水稻黑条矮缩病定级评价及产量损失评估提供了依据。

2000年左右灰飞虱传播的条纹叶枯病对我国东部稻区水稻生产造成严重影响^{[ 13, 14, 15]}。育种和植保工作者, 从抗源的搜集与发掘入手, 并相继培育出一系列抗条纹叶枯病水稻新品种, 随着抗性品种的推广, 有效遏制了此类病害对该地区水稻生产的危害。但值得的注意的是, 虽然同为灰飞虱传播的病毒病, 但当前培育和大面积推广应用的抗条纹叶枯病品种, 并不抗黑条矮缩病, 一旦该病大面积爆发, 将造成严重损失。虽然前期有研究者开展了抗黑条矮缩病资源的筛选, 但均未发现高抗抗源。本研究通过多年多点田间鉴定和人工室内接种鉴定, 筛选到的3个稳定、高抗黑条矮缩病水稻品种Kanya-- kumari 29、Madurai 25和Vietnam 160, 为抗水稻黑条矮缩病基因的定位和培育抗水稻黑条矮缩病提供了重要的材料基础。水稻各类病虫害抗源的分布往往存在一定的地理与生态特异性。例如, 对同样由灰飞虱传播的另一类病毒病条纹叶枯病的研究表明, 粳稻品种普遍高感, 而大多数籼稻品种具有一定的抗性^{[ 16, 17, 18, 19]}。褐飞虱和细菌性条斑病的抗源主要存在于东南亚地区的籼稻品种中^{[ 20, 21]}。本研究获得的3个黑条矮缩病高抗种质也均为东南亚地区的籼稻品种。越来越多的研究表明东南亚地区可能是籼稻的起源地, 该地区的籼稻品种存在较多的遗传多样性。而且多种抗病、虫资源筛选的结果也表明, 该地区蕴含丰富的抗性基因资源。上述发现为今后抗黑条矮缩病资源的搜集指明了方向。本研究建立的田间鉴定与室内鉴定相结合的黑条矮缩病鉴定体系准确、可靠, 可用于黑条矮缩病的大规模鉴定, 该体系的建立及高抗黑条矮缩病水稻资源的发掘为水稻抗黑条矮缩病基因的鉴定及育种利用提供了重要的方法和材料基础。

4 结论

水稻黑条矮缩病田间鉴定有效接种的灰飞虱密度在800万头 hm^-2左右较为合理, 而带毒率应不低于5%; 建立田间鉴定与室内鉴定相结合的黑条矮缩病鉴定体系, 该体系可准确、可靠地进行大规模黑条矮缩病鉴定。利用上述鉴定体系, 对来源于20个国家的共1240份水稻种质连续3年进行多点重复抗性鉴定, 发现来自东南亚地区的3个品种Kanya-kumari 29、Madurai 25和Vietnam 160连续3年发病率均低于10%, 表现较高的黑条矮缩病的抗性, 且抗性稳定。该体系的建立及高抗黑条矮缩病水稻资源的发掘为水稻抗黑条矮缩病基因的鉴定及育种利用提供了重要方法和材料基础。

The authors have declared that no competing interests exist.

作者已声明无竞争性利益关系。The authors have declared that no competing interests exist.

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[12]	周彤, 杜琳琳, 范永坚, 周益军. 水稻黑条矮缩病毒RT-LAMP快速检测方法的建立. 中国农业科学, 2012, 45: 1285-1292 Zhou T, Du L L, Fan Y J, Zhou Y J. Development of a RT-LAMP assay for rapid detection of rice black-streaked dwarf virus. Sci Agric Sin, 2012, 45: 1285-1292 (in Chinese with English abstract) [本文引用:1] [CJCR: 1.889]
[13]	周彤, 王磊, 程兆榜, 范永坚, 周益军. 主栽品种镇稻88对水稻条纹叶枯病的抗性特征及其遗传研究. 中国农业科学, 2009, 42: 103-109 Zhou T, Wang L, Cheng Z B, Fan Y J, Zhou Y J. Mechanism and inheritance of resistance to rice stripe disease in the japonica rice cultivar Zhendao 88. Sci Agric Sin, 2009, 42: 103-109 (in Chinese with English abstract) [本文引用:1] [CJCR: 1.889]
[14]	孙黛珍, 江玲, 张迎信, 程遐年, 王春明, 翟虎渠, 万建民. 8个水稻品种的条纹叶枯病抗性特征. 中国水稻科学, 2006, 20: 219-222 Sun D Z, Jiang L, Zhang Y X, Cheng X N, Wang C M, Zhai H Q, Wan J M. Resistance to rice stripe in eight rice varieties. Chin J Ric Sci, 2006, 20: 219-222 (in Chinese with English abstract) [本文引用:1]
[15]	Hibino H. Biology and epidemiology of rice viruses. Annu Rev Phytopathol, 1996, 34: 249-274 [本文引用:1] [JCR: 10.229]
[16]	Yamaguchi T, Yasuo S, Ishi M. Studies on rice stripe disease: III. Study on varietal resistance to stripe disease of rice. Vent Agric Exp Station, 1965, 8: 109-160 [本文引用:1]
[17]	Washio O, Ezuka A, Toriyama K, Sakurai Y. Studies on the breeding of rice varieties resistant to stripe disease: II. Genetic study on resistance to stripe disease in Japanese upland rice. Jpn J Breed, 1968, 18: 96-101 [本文引用:1]
[18]	Washio O, Ezuka A, Toriyama K, Sakurai Y. Studies on the breeding of rice varieties resistant to stripe disease: III. Genetic studies on resistance to stripe disease in foreign varieties. Jpn J Breed, 1968, 18: 167-172 [本文引用:1]
[19]	Washio O, Ezuka A, Toriyama K, Sakurai Y. Testing method for genetics and breeding for resistance to rice stripe disease. Bull Chugoku Agric Exp Station Ser A, 1968, 16: 39-197 [本文引用:1]
[20]	Liu Y, Su C, Jiang L, He J, Wu H, Peng C, Wan J. The distribution and identification of brown planthopper resistance genes in rice. Hereditas, 2009, 146: 67-73 [本文引用:1] [CJCR: 0.928]
[21]	徐建龙, 王汉荣, 林怡滋, 奚永安. 水稻细菌性条斑病和白叶枯病抗性遗传研究. 遗传学报, 1997, 24: 330-335 Xu J L, Wang H R, Lin Y Z, Xi Y A. Study on the inheritance of the resistance of rice to bacterial leaf streak and bacterial leaf blight. Acta Genet Sin, 1997, 24: 330-335 (in Chinese with English abstract) [本文引用:1]

1977

2.844

0.0

. 1977, 21:null-null

Maize rough dwarf and related viruses

Milne R G , Lovisolo O

This chapter shows that the time is ripe for the solution of many of the remaining problems concerning MRDV-like viruses. Recognition of these viruses should no longer be difficult and confusion with other agents is now avoidable. Purification of most members will still not be easy, but at least the objectives and possible methods are reasonably well defined. Biochemical work should also be able to progress rapidly, at least with maize rough dwarf virus (MRDV), fiji disease virus (FDV), and rice black streaked dwarf virus (RBSDV). The MRDV-like viruses are only just becoming recognized as the typical members of the reoviridae that are, however, quite distinct from wound-tumor virus (WTV) and rice dwarf virus (RDV) in a number of characteristics, notably their RNA segmentation, vectors, and capsid structure. A very interesting result to be expected shortly is firm information on the size distribution of the RNA segments of maize wallaby ear virus (MWEV). This should indicate at once whether MWEV resembles WTV or MRDV or perhaps neither, and may clarify the taxonomy of these viruses. Although this uncertainty remains at present, it seems appropriate to look for a suitable name to indicate “MRDV-like virus.” The name ACANTHOVIRUS has been proposed because of the spikes on both the outer and inner capsids, this combination being so far unique to these viruses.

... 水稻黑条矮缩病(rice black-streaked dwarf virus disease, RBSDVD)是由水稻黑条矮缩病毒(rice black-streaked dwarf virus, RBSDV)引起的, 主要由灰飞虱以持久性不经卵方式传播^[1,2] ...

1993

3.127

0.0

1979

0.0

1.088

... 近年来, 随着耕作制度和栽培方式的变化、冬季气候变暖和感病(虫)品种的大面积推广, 传毒介体灰飞虱发生量日益上升, RBSDVD在江苏、浙江、江西和福建大规模发生, 并迅速上升为生产上最主要的病毒病害, 给水稻的安全生产带来很大威胁^[3,4,5,6] ...

... 3 讨论目前, 黑条矮缩病在江苏、浙江、上海、安徽、山东等长江中下游地区广泛发生, 已成为该地区主要水稻病毒病之一, 造成严重的经济损失^[3,4,5,6] ...

2008

2.03

0.0

. 2008, 153:1893-1898 DOI:10.1007/s00705-008-0209-4

A black-streaked dwarf disease on rice in China is caused by a novel fiji virus

1.Zhejiang Academy of Agricultural Sciences Virology Laboratory Hangzhou 310021 China 2.Rothamsted Research Department of Plant Pathology and Microbiology Harpenden Herts AL5 2JQ UK 3.Zhejiang Academy of Agricultural Sciences Institute of Virology and Biotechnology Hangzhou 310021 People’s Republic of China

An isolate of a plant reovirus causing severe stunting and dark leaf symptoms on rice from Guangdong, China, was similar in virion morphology and serologically related to rice black-streaked dwarf virus (RBSDV). The electrophoretic profiles of genome segments of the two viruses in agarose or polyacrylamide gel were indistinguishable. The four genome segments of the new isolate corresponding to RBSDV S7–S10 were amplified by ligation RT-PCR and sequenced. The size and organization of each genome segment was very similar to its counterparts in RBSDV, maize rough dwarf virus (MRDV), and mal de Rio Cuarto virus (MRCV). Sequence identity was greatest to RBSDV and MRDV (ranging from about 60–85% depending on the protein), but identities were always much lower than those between RBSDV and MRDV. These comparisons and phylogenetic analyses suggested that the virus represents a new species in genus Fijivirus group 2, tentatively named Rice black-streaked dwarf virus-2.

2001

1.61

0.0

. 2001, 107:563-567 DOI:10.1023/A:1011204010663

Sequence analysis shows that a dwarfing disease on rice, wheat and maize in China is caused by rice black-steaked dwarf virus

1.Zhejiang University 2.Zhejiang Academy of Agricultural Sciences 3.Zhejiang Academy of Agricultural Sciences 4.IACR-Rothamsted, Harpenden

Isolates of plant reoviruses causing severe stunting and dark leaf symptoms on wheat in Hebei province, on maize in Hubei province and on rice in Zhejiang province, China have been characterized. Four of the ten genome segments corresponding to Rice black-streaked dwarf virus (RBSDV) S7, S8, S9, S10 were amplified by RT-PCR from the Hebei and Zhejiang isolates and sequenced. Sequences of S9 and S10 were also obtained from the Hubei isolate. Sequences of corresponding segments of the Chinese isolates were very similar to each other (94.0–99.0% identical nucleotides and 96.3–100% identical amino acids) and were closer to those of a previously reported Japanese RBSDV isolate (90.0–94.9% identical nucleotides and 91.1–98.6% identical amino acids) than to those of an Italian Maize rough dwarf virus isolate (85.1–88.1% identical nucleotides and 85.5–94.3% identical amino acids). The Chinese isolates should be classified as RBSDV.

2009

2.729

0.0

. 2009, 58(5):815-825

Studies on the epidemiology and yield losses from rice black-streaked dwarf disease in a recent epidemic in Zhejiang province, China

H.-D. Wang 1 ,J.-P. Chen 2,* ,A.-G. Wang 3 ,X.-H. Jiang 1 andM. J. Adams 4

1 Zhejiang Provincial Station of Plant Protection and Plant Quarantine, Hangzhou 310020 2 Institute of Virology and Biotechnology, Zhejiang Academy of Agricultural Sciences, Hangzhou 310021 3 Linghai Station of Plant Protection, Taizhou 317000, China 4 Department of Plant Pathology and Microbiology, Rothamsted Research, Harpenden, Herts, AL5 2JQ, UK * *E-mail: jpchen2001@yahoo.com.cn

> The spread of rice black-streaked dwarf disease, which has emerged as a major problem on winter wheat and the two summer rice crops (early indica and late japonica ) grown in central and southern Zhejiang province, China, is documented from 1995 to 2007. The late japonica crop suffered the most: up to 64 640 ha were affected with estimated losses of c . 120 000 t grain per year. Peak adult numbers of the small brown planthopper vector, Laodelphax striatellus , coincided with the seedling stages of both rice crops and the proportion of the insect population carrying virus increased during 1998–2005. Seedlings with three to four leaves were the most susceptible, whereas plants inoculated after the end of tillering developed few or no symptoms. Disease levels were strongly correlated with numbers of viruliferous vectors. In sowing-date experiments with both rice crops, the earliest sowings had the most disease and suffered the greatest yield losses. With the last sowing date (25 days after the first), there were almost no losses. There were yield losses of 0·80% for every 1% increase in disease incidence in early indica rice and rather more (0·92%) in the late japonica crop. There were large differences in susceptibility between cultivars, indicating the possibility, within currently available germplasm, of using more resistant cultivars to help contain the disease. Changes in cropping practice and in recent winter weather conditions have probably contributed to the emergence of the virus as a major pathogen in eastern China.

2011

0.0

... 虽然周彤等^[7]建立了黑条矮缩病室内人工接种方法, 对黑条矮缩病的鉴定具有重要参考价值, 但室内鉴定费时、费事, 工作量大, 难以满足规模化抗源筛选的迫切需要 ...

... 本研究在前人研究的基础上^[7], 对黑条矮缩病室内人工接种方法进一步优化, 确定了黑条矮缩病人工室内接种鉴定, 以1~2龄若虫, 饲毒处理20 d, 在水稻一叶期每株接种10头较为适宜 ...

2010

0.0

1.667

. 2010, 36:1258-1264 DOI:10.3724/SP.J.1006.2010.01258

The screening of the rice resources against rice black-streaked dwarf virus and mapping of resistant QTL

1 State Key Laboratory of Crop Genetics and Germplasm Enhancement / Jiangsu Plant Gene Engineering Research Center, Nanjing Agricultural University, Nanjing 210095, China; 2 Institute of Lianyungang Agricultural Science of Xuhuai Area , Lianyungang 222006, China; 3 Institute of Crop Sciences, Chinese Academy of Agricultural Sciences, Beijing 100081, China

Rice black-streaked dwarf virus(RBSDV)disease become epidemic in Jiangsu and Zhejiang provinces. To screen resistant germplasms and discover new resistance genes/QTLs against RBSDV, we evaluated the resistance to RBSDV in 311 japonica cultivars in field test. The results showed that no cultivar was immune to RBSDV. The disease rate of 24 main japonica cultivars grown in Jiangsu province at present was 10.0%–29.0% and that of 71.5% cultivars in 287 japonicapopularized in Jiangsu beforewas between 10.0% and 30.0%. Furthermore, quantitative trait loci (QTL) analysis was conducted by using 162 recombinant inbred lines (RILs), which derived from a cross between Guichao 2, a susceptible indica variety, and Koshihikari, a japonica variety with resistance to RBSDV. RBSDV resistances were evaluated using natural infection methods by scoring the disease rating. One putative QTL ( qRBSDV3 ) controlling RBSDV resistance was mapped between the marker RM7 and RM5748 on chromosome 3, which explained 17.1% of the total phenotypic variation with LOD score of 5.4. The positive resistant effect contributed from Koshihikari. Further analysis revealed that the lines harboring the alleles of qRBSDV3 exhibited significantly increased resistance to RBSDV. The results should be very useful for breeding new RBSDV-resistant cultivars by marker-assisted selection (MAS). cultivars

通过水稻黑条矮缩病的田间鉴定发现，分蘖盛期的病状最为显著，是病症观察的最佳时期。对江苏省311份粳稻品种进行重病区田间鉴定试验，未发现对黑条矮缩病毒(RBSDV)免疫的品种。江苏省目前正在推广的24个主栽水稻品种发病率在10.0%~30.0%之间，曾推广的287份粳稻品种中，71.5%的品种发病率在10.0%~29.0%之间，其余品种发病率在29%以上。来源于日本的粳稻品种Koshihikari的黑条矮缩病发病率相对较低，籼稻高产品种桂朝2号为感病品种。利用Koshihikari/桂朝2号RILs群体进行抗RBSDV的基因定位，结果在第3染色体上标记RM7–RM5748之间检测到1个抗黑条矮缩病的QTL，来自Koshihikari的等位基因增强了水稻黑条矮缩病的抗性，携带抗性位点的家系明显提高了对RBSDV的抗性。本研究结果将为研究水稻黑条矮缩病和水稻抗黑条矮缩病分子育种提供重要信息。

... 前期的研究结果表明多数水稻品种易感RBSDVD, 目前尚未发现高抗水稻黑条矮缩病的水稻品种^[8], 虽然少数水稻资源具有一定的RBSDVD抗性, 并检测到一些微效抗性QTL^[9,10,11] ...

2009

0.0

1.667

. 2009, 35:2213-2217 DOI:10.3724/SP.J.1006.2009.02213

Detection of QTL for resistance to rice black-streaked dwarf viral disease

Pan C H , Li A H , Chen Z X , Wu L B , Dai Z Y , Zhang H X , Huang N S , Chen X J , Zhang Y F , Zuo S M , Pan X B

1Key Laboratory of Plant Functional Genomics of Ministry of Education/Key Laboratory of Crop Genetics and Physiology of Jiangsu Province,Yangzhou University,Yangzhou 225009,China；2Lixiahe Agricultural Research Institute of Jiangsu Province/Nanjing Subcenter of National Rice Improvement Center，Yangzhou 225007，China

The rice black-streaked dwarf viral disease (RBSDV), transmitted by the planthopper Laodelphax striatellus Fallèn, was one of the most serious diseases in rice in south China. Breeding of RBSDV resistance variety using marker-assisted selection was effective measure to reduce the disease damage. For achieving this aim, QTL mapping of resistance to RBSDV was studied using a recombinant inbred line (RIL) population from the cross between Zhenshan 97B and Minghui 63. Reactions of 242 RILs to RBSDV were investigated by natural infection method in Agricultural College of Yangzhou University and Lixiahe Agricultural Research Institute, and scored by the incidence of RBSDV. The trait of RBSDV incidence was normally distributed and marked bias towards the resistant parent, which implied that they were controlled by quantitative trait loci. Six QTLs for resistance to RBSDV were detected by WinQTLcart 2.5 software. Two adjacent QTLs were mapped on chromosome 6, two on chromosome 11, one on chromosomes 7 and 9, respectively. Four QTLs on chromosomes 6, 7, and 9 could be detected in two locations, and were stably expressed across the two environments. There had not been any reports about location of QTL for resistance of rice stripe virus and planthopper, which suggested that six QTLs in this study were really resistant to RBSDV. Two QTLs on chromosome 6 were with main effect, and their contributions to the total variation were 20.20% and 18.68% with LOD scores of 12.09 and 9.77 respectively, moreover, they had high value for study on genetics of RBSDV and should be useful in marker-assisted selection for resistance to RBSDV.

利用珍汕97B/明恢63的重组自交系群体，采用自然发病鉴定的方法，以穴发病率为表型值，对各株系进行黑条矮缩病抗性鉴定。群体的穴发病率偏向于抗病亲本，且呈连续性分布，表明水稻黑条矮缩病抗性受数量性状基因控制。利用WinQTLcart 2.5软件对黑条矮缩病抗性QTL进行分析，共检测到6个QTL，其中第6、11染色体上各有2个，第7、9染色体各有1个。第6、7、9染色体上的4个QTL在两个地点都能检测到，是稳定表达的QTL，尤其是第6染色体上的2个QTL，LOD值分别为12.09和9.77，贡献率分别为20.20%和18.68%，是主效QTL，能在分子标记辅助选择育种中加以利用。

2013

3.251

0.0

. 2013, 32:1-13 DOI:10.1007/s11032-012-9807-7

Identification and fine mapping of qRBSDV-6MH, a major QTL for resistance to rice black-streaked dwarf virus disease

1. Lixiahe Agricultural Research Institute, Yangzhou, 225007, Jiangsu Province, People’s Republic of China 2. Key Laboratory of Plant Functional Genomics, Ministry of Education, Yangzhou University, Yangzhou, 225009, People’s Republic of China

Abstract Rice black streaked-dwarf virus (RBSDV) disease is recently expanding in southern China and poses a serious threat to rice crops. Few studies related to the genetics and breeding of RBSDV resistance have been reported. We have previously mapped a number of quantitative trait loci (QTLs) for RBSDV resistance by using a recombinant inbred line population of ‘Zhenshan 97’ (ZS97, susceptible)/‘Minghui 63’ (MH63, resistant) with natural infection data in two locations. In the present study, we confirmed the presence of a number of resistant QTLs on chromosomes 6, 7, and 9 from MH63 by using the same population in four different locations. We then focused on a major QTL, qRBSDV - 6 MH , on chromosome 6 and introduced it into a highly susceptible japonica rice variety, ‘Huaidao 5’, using MH63 as the donor via marker-assisted selection, to generate seven backcross inbred lines (BILs). Natural infection and artificial inoculation-based tests revealed that all of the BILs had a significantly higher resistance to RBSDV than the recurrent parent. These results demonstrate that qRBSDV - 6 MH is a stable major resistance QTL of high breeding value. We also constructed a set of chromosome segment substitution lines (CSSLs) specific to the qRBSDV - 6 MH region and these used as fine mapping population. Combining the genotypes of CSSLs with the phenotypes from natural infection data in a highly RBSDV epidemic area during two different sowing seasons, we were able to precisely map qRBSDV - 6 MH to the markers S18 and S23 at a physical distance of 627.6 kb on the Nipponbare reference genome.

2011

0.0

2012

0.0

1.889

. 2012, 45:1285-1292 DOI:10.3864/j.issn.0578-1752.2012.07.006

Development of a RT-LAMP assay for rapid detection of rice black-streaked dwarf virus

1.江苏省农业科学院植物保护研究所/江苏省植物病毒病诊断检测技术中心，南京 210014. 2.南京农业大学植物保护学院，南京210095

【Objective】The objective of this study is to develop a reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay for rapid and sensitive detection of Rice black-streaked dwarf virus (RBSDV) from host plants and insect vector.【Method】Four primers matching a total of 6 sequences of the S10 of RBSDV were synthesized for the RT-LAMP assay. The concentration of the primer and MgSO4 were optimized, and the best temperature and reaction time for detecting the virus were found, respectively. Detection sensitivity comparisons were performed between the RT-LAMP and RT-PCR assay using a single extraction of total RNA from RBSDV-infected rice leaves which was serially diluted in ten-fold increments in DEPC-treated water. In order to determine the specificity of the RT-LAMP assay, total RNA of rice leaves infected with RBSDV or Southern rice black-streaked dwarf virus (SRBSDV) was applied separately to the reaction system. RBSDV-infected rice plants collected from rice field were detected.【Result】 This method demonstrated a high degree of specificity for RBSDV, which can distinguish RBSDV from SRBSDV. The method was also proved to be extremely sensitive, which was as much as the RT-PCR for RBSDV detection. The detection of amplified products was easily monitored.【Conclusion】The RT-LAMP assay is suitable for rapid detection of RBSDV in host plant and vectors.

【目的】建立一种快速、灵敏的逆转录环介导等温扩增方法（RT-LAMP）检测寄主植物和传毒介体体内的水稻黑条矮缩病毒（Rice black-streaked dwarf virus，RBSDV）。【方法】合成4条针对RBSDV S10核苷酸序列6个位点的特异性引物。分别对引物浓度、MgSO4浓度、反应温度和时间进行优化。将感病水稻总RNA梯度稀释后进行灵敏性检验并与RT-PCR比较分析。选择RBSDV和南方水稻黑条矮缩病毒（SRBSDV）验证该方法的特异性。用本RT-LAMP方法检测田间病株。【结果】RT-LAMP检测方法可排除SRBSDV的干扰而特异地检测植物和飞虱体内的RBSDV，与RT-PCR灵敏性基本一致。检测结果易于判定。【结论】RT-LAMP检测方法适合寄主植物和介体体内RBSDV的快速检测。

... 4 灰飞虱及其带毒率检测参照周彤等^[12]方法随机采集100头灰飞虱检测水稻黑条矮缩病毒, 确定带毒率 ...

2009

0.0

1.889

. 2009, 42:103-109

Mechanism and inheritance of resistance to rice stripe disease in the japonica rice cultivar Zhendao 88

江苏省农业科学院植物保护研究所

【Objective】 The objective of this experiment was to analyze the resistance mechanism and resistance genetic model to rice stripe disease in Zhendao 88, and to screen the SSR marker related to the gene of rice stripe disease resistance in Zhendao 88. 【Method】The resistance to rice stripe virus (RSV) and the vector, small brown planthopper (SBPH) (Laodelphax striatellus Fallèn) was analyzed in Zhendao 88 by using the method of tiller inoculation, inoculation at the seeding stage, non-preference test and antibiosis test. The inheritance of rice stripe disease resistance in F2 lines and F2:3 lines from the cross Zhendao 88/ Wuyujing No. 3 was studied by using tiller inoculation and inoculation at the seeding stage, respectively. The resistance gene was mapped based on SSR analysis.【Result】 Zhendao 88 was highly resistant to rice stripe virus and weakly resistant to vector. Genetic analysis indicated that the resistance to rice stripe disease in Zhendao 88 was inherited as a single dominant trait. The rice stripe disease resistance gene in Zhendao 88 was located on the chromosome 11, with the distance of 4.7cM from a SSR marker RM229. 【Conclusion】 The rice stripe disease resistance in Zhendao 88 was mainly derived from the resistance to virus and controlled by a single major gene. The breeding process for rice stripe disease resistance should be accelerated by using Zhendao 88 as a resistance parent.

【目的】明确镇稻88对水稻条纹叶枯病的抗性特征及其抗性遗传模式，寻找与抗条纹叶枯病基因连锁的分子标记。【方法】利用单分蘖鉴定、苗期接种鉴定、非嗜性测验和抗生性测验分析镇稻88对水稻条纹叶枯病和介体灰飞虱的抗性;分别采用单分蘖鉴定和苗期接种鉴定方法对武育粳3号与镇稻88构建的F2群体、F2:3家系进行水稻条纹叶枯病抗性遗传分析;以SSR标记连锁分析方法对抗病基因进行定位。【结果】镇稻88表现出较强的病毒抗性和较弱的抗虫性;其对水稻条纹叶枯病的抗性表现为显性单基因的遗传模式;SSR标记分析显示该抗病基因位于水稻第11染色体上，与标记RM229间的遗传距离为4.7 cM。【结论】镇稻88对水稻条纹叶枯病的抗性主要来自于对病毒的抗性，由一对主效核基因控制，以镇稻88为抗病亲本结合本研究结果，可望加快水稻条纹叶枯病抗性品种选育的速度。

... 2000年左右灰飞虱传播的条纹叶枯病对我国东部稻区水稻生产造成严重影响^[13,14,15] ...

2006

0.0

. 2006, 20:219-222

Resistance to rice stripe in eight rice varieties

The resistance to rice stripe virus (RSV) and vector, small brown planthopper (SBPH) (Laodelphax striatellus Fallen) was studied and analyzed in eight indica rice varieties by using individual inoculation, mass inoculation, non-preference test and antibiosis test. The mechanism of resistance to RSV and vector SBPH was different in the eight rice varieties. Some rice varieties were highly resistant to RSV and vector SBPH, such as IR36, Kasalath, Zhaiyeqing 8, Daorenqiao and DV85; some others, such as Aichi 97 and Kenta Nakan were only resistant to RSV but susceptible to SBPH; and IR24 was moderately resistant to RSV and SBPH.

利用强迫饲毒、集团接种、非嗜性测验、抗生性测验的方法研究与分析了8个抗条纹叶枯病水稻品种对条纹病毒和介体灰飞虱的抗性,发现不同水稻抗性品种的抗性特征并不完全相同。IR36、Kasalath、窄叶青8号、道人桥、DV85既抗条纹病毒又抗介体灰飞虱，对条纹叶枯病所表现的抗性是条纹病毒和介体灰飞虱抗性共同作用的结果；爱知97和Kenta Nakan抗条纹病毒，但不抗介体灰飞虱，对条纹叶枯病所表现的抗性是品种对条纹病毒的抗性；而IR24对条纹病毒和介体灰飞虱仅表现中等抗性。

... 2000年左右灰飞虱传播的条纹叶枯病对我国东部稻区水稻生产造成严重影响^[13,14,15] ...

1996

10.229

0.0

... 2000年左右灰飞虱传播的条纹叶枯病对我国东部稻区水稻生产造成严重影响^[13,14,15] ...

1965

0.0

... 例如, 对同样由灰飞虱传播的另一类病毒病条纹叶枯病的研究表明, 粳稻品种普遍高感, 而大多数籼稻品种具有一定的抗性^{[16,17,18,19]} ...

1968

0.0

... 例如, 对同样由灰飞虱传播的另一类病毒病条纹叶枯病的研究表明, 粳稻品种普遍高感, 而大多数籼稻品种具有一定的抗性^{[16,17,18,19]} ...

1968

0.0

... 例如, 对同样由灰飞虱传播的另一类病毒病条纹叶枯病的研究表明, 粳稻品种普遍高感, 而大多数籼稻品种具有一定的抗性^{[16,17,18,19]} ...

1968

0.0

... 例如, 对同样由灰飞虱传播的另一类病毒病条纹叶枯病的研究表明, 粳稻品种普遍高感, 而大多数籼稻品种具有一定的抗性^{[16,17,18,19]} ...

2009

0.0

0.928

. 2009, 146(2):67-73

The distribution and identification of brown planthopper resistance genes in rice

Yuqiang Liu 1 ,Changchao Su 1 ,Ling Jiang 1 ,Jun He 1 ,Han Wu 1 ,Cheng Peng 1 andJianmin Wan 1,2

1 National Key Laboratory of Crop Genetics and Germplasm Enhancement, Nanjing Agricultural University, Nanjing, P. R. China 2 Chinese Academy of Agricultural Sciences, Beijing, P. R. China * Jianmin Wan, National Key Laboratory for Crop Genetics and Germplasm Enhancement, Nanjing Agricultural University, CN–210095 Nanjing, P. R. China and Chinese Academy of Agricultural Sciences, CN–100081 Beijing, P. R. China. E-mail: wanjm@njau.edu.cn

> A collection of 515 rice landraces originating from Vietnam and China were screened for the reaction to brown planthopper (BPH) infestation. Most of the resistant landraces were indica types from Vietnam and the Guangxi province in China. An F 2 mapping population was created from the cross between a BPH resistant Vietnamese landrace Yagyaw and the susceptible cultivar Cpslo17. Four quantitative trait loci (QTL) contributing to BPH resistance were mapped on chromosomes 2, 4, 7 and 9, respectively. The individual QTL accounted 5.64% to 12.77% of the phenotypic variance, and three resistant alleles were harbored in the resistant landrace Yagyaw. Two QTL located on chromosomes 2 and 4 were identified with significant additive effects and are useful in breeding new rice inbred lines. One resistant allele was harbored by the susceptible parent Cpslo17. This gene is important in selecting rice inbred lines with stronger resistances to BPH.

... 褐飞虱和细菌性条斑病的抗源主要存在于东南亚地区的籼稻品种中^[20,21] ...

1997

0.0

. 1997, 24:330-335

Study on the inheritance of the resistance of rice to bacterial leaf streak and bacterial leaf blight

(Zhejiang Academy of Agricultural Sciences Hangzhou 310021)

The inheritance of the resistance of three resistant rice varieties, Hashikalmi,Dular and 90IRBBN44, to S-103 isolate of bacterial leaf streak and P1 strain of bacterial leaf blight was analysed.The results indicated that Hashikalmi and Dular had two recessive resistant genes to S-103, and 90IRBBN44 had one recessive gene. According to allelism test, at least one of the two genes of Hashikalmi and Dular was allelic, but they were nonallelic to that of 90IRBBN44. All three resistant varieties possessed a recessive resistant gene to P1,which was allelic to xa-5.One of the two genes of Hashikalmi and Dular resistant to S-103 was link to xa-5.One of the two genes of Hashikalmi and Dular resistant to S-103 was link to xa-5, but the gene of 90IRBBN44 was independent of xa-5. Breeding for resistance to bacterial leaf streak in rice was discussed.

分析了Hashikalmi、 Dular和90IRBBN44三个抗源品种对水稻细菌性条斑病S-103菌株和白叶枯病PI菌系的抗性遗传。结果表明,Hashikalmi和Dul ar对S-103的抗性均由2对隐性基因所控制,90IRBBN44则带有1对隐性抗性基因。经等位性测定表明,Hashikalmi和Dular的2对基因中至少有1对是等位的,但它们与90IRBBN44的1对基因均不等位。 3个抗源品种对P1的抗性都受1对隐性基因控制,该基因与xa-5等位。连锁遗传分析表明,Hashik almi和Dular对S-103的2对抗细条病基因中的1对与xa-5相连锁,而90IRBB N44的1对抗性基因与xa-5呈独立遗传。本文还就开展水稻抗细菌性条斑病育种进行了讨论。

... 褐飞虱和细菌性条斑病的抗源主要存在于东南亚地区的籼稻品种中^[20,21] ...