小西葫芦黄化花叶病毒罗汉果分离株的抗血清制备及其田间寄主检测
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摘要
罗汉果(Siraitia grosvenori)为葫芦科多年生藤本植物,是广西著名特产,具有重要的医疗和保健价值。长期以来,罗汉果一直受着病毒病的危害。本研究对小西葫芦黄化花叶病毒罗汉果分离株(ZYMV-LHG)进行了分离、繁殖、提纯、体外性质研究、抗血清的制备,并用抗血清检测田间植物的带毒情况。研究结果显示,用西葫芦可以从罗汉果病叶中分离到ZYMV-LHG,ZYMV-LHG在西葫芦上表现明脉、黄化、蕨叶症状;ZYMV-LHG的钝化温度为55℃,稀释限点为10~(-3),体外保毒期为6 d;自繁殖ZYMV-LHG的西葫芦病叶上提纯病毒;利用提纯的病毒免疫BALB/C小鼠制备抗血清,所获抗血清的效价为1∶16000;利用该抗血清检测罗汉果果园附近的作物及杂草,发现11科的14种植物表现阳性,其中包括葫芦科(Cucurbitaceae)的丝瓜(Luffa cylindrica)、南瓜(Cucurbita moschata)、葫芦(Lagenaria siceraria)、茄科(Solanaceae)的辣椒(Capsicum annuum)、豆科(Leguminosae)的四季豆(Amphicarpaea edgeworthii)和杂草中堇菜科(Violaceae)的堇菜(Viola verecunda)、马鞭草科(Verbenaceae)的大青(Clerodendrum cyrtopAyllum)、鸭跖草科(Commelinaceae)的鸭跖草(Commelina communis)、菊科(Compositae)的野茼蒿(Crossocephalumcrepidioides)、蓼科(Polygonaceae)的齿果酸模(Rumex dentatus)、苋科(Amaranthaceae)的莲子草(Alternanther sessilis)、车前草科(Plantaginaceae)的车前(Plantago asiatica)和酢浆草科(Oxalidaceae)的酢浆草(Oxalis corniculata)。温室测定化学药剂对罗汉果病毒病的防治效果,先喷施化学药剂后摩擦接种病毒的处理中,以1.45%三氮唑核苷·烷醇·锌防治效果最好,其防效为58.99%,而5%二(辛基胺乙基)甘氨酸盐的防效最差,其防效为27.19%。在先摩擦接种病毒再喷施化学药剂的处理中,1.45%三氮唑核苷·烷醇·锌、5%盐酸吗啉胍和20%盐酸吗啉胍·乙铜的防效较好,其防效分别为52.82%、50.23%、47.17%,而5%二(辛基胺乙基)甘氨酸盐的防效最差,其防效为22.07%。但整体来说,化学药剂对罗汉果病毒病防治效果不是太理想。
Siraitia grosvenori(Luohanguo)belongs to Cucurbitaceae, a perennial liana especially grown in Guangxi province, which had important medical and healthcare value. However, Siraitia grosvenori viral disease has been a major problem in its production industry. In this paper, ZYMV-LHG separation, propagation, purification, characteristics in vitro and antiserum preparation were conducted, and the nature hosts in the field were detected using ZYMV-LHG antiserum. The results showed that ZYMV-LHG could be separated by Cucurbita pepo from diseased leaves of Siraitia grosvenori. Symptoms of Cucurbita pepo infected by ZYMV-LHG were vein-clearing, etiolation, and strip-like malformation. The thermal inactivation of ZYMV-LHG point was 55°C. The dilution end point was 10"3. The longevity in vitro was 6 days. ZYMV-LHG was purified from diseased leaves of Cucurbita pepo. The antiserum against ZYMV-LHG was prepared by injected BALB/C mice with purified virus particles. The titer of this antiserum was 1:16 000. The crops and weeds in or nearby the Siraitia grosvenori orchard were detected with ZYMV-LHG antiserum. The results showed that 11 families 14 species had positive reaction with ZYMV-LHG antiserum, including Cucurbitaceae (Luffa cylindric, Cucurbita moschata and Lagenaria siceraria), Solanaceae (Capsicum annuum), Leguminosae (Amphicarpaea edgeworthii), Violaceae (Viola verecunda), Verbenaceae (Clerodendrum cyrtophyllum), Commelinaceae (Commelina communis), Compositae (Crassocephalum crepidioides), Polygonaceae (Rumex dentatus), Amaranthaceae (Alternanther sessilis), Plantaginaceae (Plantago asiatica), Oxalidaceae (Oxalis corniculata). The control efficiencies of several virucides to Siraitia grosvenori viral disease were carried out in greenhouse. In the treatment of spraying virucides before inoculating ZYMV-LHG, 1.45%Ribavirin copper. Triacontanol. Zinc had the highest control efficiency (58.99%). With lower control efficiency (27.19%) was 5% Octyl glycine ethyl amine salt. In the treatment of spraying virucides after inoculating ZYMV-LHG, three virucides had higher control efficiency. There were 52.82% (1.45% Ribavirin copper·Triacontanol·Zinc)、50.23% (5% Moroxy dine hydrochioride) and 47.17%(20%Moroxydine hydrochloride·Second copper) respectively. With lower control efficiency (22.07%) was 5%Octyl glycine ethyl amine salt. In brief, the virucides with high efficiency to control Siraitia grosvenori viral disease were not available.
Siraitia grosvenori(Luohanguo)belongs to Cucurbitaceae, a perennial liana especially grown in Guangxi province, which had important medical and healthcare value. However, Siraitia grosvenori viral disease has been a major problem in its production industry. In this paper, ZYMV-LHG separation, propagation, purification, characteristics in vitro and antiserum preparation were conducted, and the nature hosts in the field were detected using ZYMV-LHG antiserum. The results showed that ZYMV-LHG could be separated by Cucurbita pepo from diseased leaves of Siraitia grosvenori. Symptoms of Cucurbita pepo infected by ZYMV-LHG were vein-clearing, etiolation, and strip-like malformation. The thermal inactivation of ZYMV-LHG point was 55°C. The dilution end point was 10"3. The longevity in vitro was 6 days. ZYMV-LHG was purified from diseased leaves of Cucurbita pepo. The antiserum against ZYMV-LHG was prepared by injected BALB/C mice with purified virus particles. The titer of this antiserum was 1:16 000. The crops and weeds in or nearby the Siraitia grosvenori orchard were detected with ZYMV-LHG antiserum. The results showed that 11 families 14 species had positive reaction with ZYMV-LHG antiserum, including Cucurbitaceae (Luffa cylindric, Cucurbita moschata and Lagenaria siceraria), Solanaceae (Capsicum annuum), Leguminosae (Amphicarpaea edgeworthii), Violaceae (Viola verecunda), Verbenaceae (Clerodendrum cyrtophyllum), Commelinaceae (Commelina communis), Compositae (Crassocephalum crepidioides), Polygonaceae (Rumex dentatus), Amaranthaceae (Alternanther sessilis), Plantaginaceae (Plantago asiatica), Oxalidaceae (Oxalis corniculata). The control efficiencies of several virucides to Siraitia grosvenori viral disease were carried out in greenhouse. In the treatment of spraying virucides before inoculating ZYMV-LHG, 1.45%Ribavirin copper. Triacontanol. Zinc had the highest control efficiency (58.99%). With lower control efficiency (27.19%) was 5% Octyl glycine ethyl amine salt. In the treatment of spraying virucides after inoculating ZYMV-LHG, three virucides had higher control efficiency. There were 52.82% (1.45% Ribavirin copper·Triacontanol·Zinc)、50.23% (5% Moroxy dine hydrochioride) and 47.17%(20%Moroxydine hydrochloride·Second copper) respectively. With lower control efficiency (22.07%) was 5%Octyl glycine ethyl amine salt. In brief, the virucides with high efficiency to control Siraitia grosvenori viral disease were not available.
引文
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陈娟,朱永和.病毒必克防治水稻条纹叶枯病研究.江西农业学报,2006,18(6):116-117
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杭玲,苏国秀,夏阳升,等.罗汉果组培苗栽培技术.广西农业科学,2003,6:70-72
郝桂霞,罗汉果提取液对自由基的消除作用.江西化工,2005,4:89-90
黄慧鸣.中国高等植物图鉴(第五版)北京:科技出版社,2002
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黄清汉.罗汉果常见病虫害防治方法.广西园艺,2005,16(6):41-42
黄思良,陈作胜,晏卫红,等.罗汉果芽枯病病因及防治技术研究.中国农业科学,2001,34(4):385-390
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李土生.罗汉果组培苗种植技术.农业科技通讯,2004,9:33
李淑菊,王惠哲,霍振荣,等.利用RT-PCR对黄瓜病毒病毒原种类进行检测.华北农学报,2004,19(3):100-102
梁贵秋.罗汉果组培苗的栽培技术.广西热带农业,2005,1:17-19
廖咏梅,王忠文,韩英,等.广西罗汉果病毒病的发生与危害的初步研究.中国食用菌(增刊),2006,25:35-38
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