番木瓜环斑病毒山东分离物的全基因组序列分析

Papaya ringspot virus (PRSV) is a major limiting factor to cucurbit production worldwide. One zucchini sample showing symptoms of leaf mosaic and fruit ringspot was collected from Ji’nan city of Shandong province. Primary experiments showed that the sample was infected with PRSV, which was designated as PRSV-SD accordingly. The complete genomic fragment of PRSV-SD was obtained with RT-PCR. The results of sequencing showed that the full genomic sequence of PRSV-SD was 1 0337 nucleotides (nt) excluding the 3′- terminal poly (A) tail. The 5′- and 3′-untranslated regions (UTR) were 90 and 206 nt, respectively. The putative polyprotein was 3 346 amino acids in length. Comparison of the PRSV-SD isolate with 18 other PRSV isolates revealed that they shared nucleotide identities of 82.1%~89.3% at the complete genomic levels and amino acid identities of 90.6%~94.7% for the polyprotein. No recombination was detected throughout the genome of PRSV-SD. Phylogenetic analysis with complete genomic sequences indicated that the PRSV isolates were clustered into two major groups: Asia and America and PRSV-SD was clustered to the Asia group. Selection pressure analysis revealed that all of the 11 proteins of PRSV-SD were under negative selection, but positive selection sites were detected in P1, P3, 6K1, NIa-pro and NIb. Our research results provide a theoretical foundation for the detection and prevention of PRSV. Key words:Papaya ringspot virus; complete genomic sequence; phylogenetic analysis; selection pressure; recombination 中图分类号:S436.429; Q939.46 文献标识码:A 文章编号:0412-0914(2018)02-0285-04 番木瓜环斑病毒(Papaya ringspot virus,PRSV)属于马铃薯Y病毒科(Potyviridae)马铃薯Y病毒属(Potyvirus)^[1]。根据寄主范围可划分为P和W 2个株系,其中P株系侵染番木瓜(Carica papaya L.)和葫芦科(Cucurbitaceae)作物,而W株系只侵染葫芦科作物,两者血清学反应密切相关。PRSV可引起植株叶片褪绿、花叶、卷曲等症状,在果实表面形成环斑。 PRSV于20世纪40年代末在美国首次报道,目前该病毒在巴西、印度、波兰等国均有发生^[2]。2000~2001年,PRSV使我国海南番木瓜损失严重。2005年以来,我国广东、四川先后检测到PRSV侵染罗汉果、苦瓜等葫芦科作物^[3]。2016年从山东西葫芦上检测到PRSV,该分离物属于W株系^[4]。我国关于PRSV进化的研究大多集中于P株系,有关W株系全基因组序列分析的报道相对较少。为了进一步研究我国PRSV-W株系的基因组特性,为PRSV的监测预警提供依据,我们测定了PRSV山东分离物的全基因组序列,并进行了核苷酸和氨基酸序列一致率、重组、系统发育和基因选择压力分析。 2期黄显德,等:番木瓜环斑病毒山东分离物的全基因组序列分析 植物病理学报48卷 1 材料与方法 1.1 材料 发病西葫芦样品采自山东省济阳县。大肠杆菌DH5α由本实验室保存。植物总RNA提取试剂盒TRIzol、DNA凝胶回收试剂盒等均购自北京全式金公司;Taq DNA聚合酶、 pMD18-T克隆载体等购自TaKaRa公司;SuperScript^TM Ⅳ逆转录酶购自Invitrogen公司。其他生化试剂及普通化学试剂均为进口或国产分析纯。 1.2 实验方法 1.2.1 扩增策略及引物合成 根据GenBank中PRSV基因组序列,设计引物分4段扩增基因组序列。根据所得序列设计5′-RACE引物扩增5′-端片段。测序后用SeqMan拼接得到全基因组序列。所用引物详见表1。 1.2.2 植物总RNA提取及RT-PCR扩增 按照RNA提取试剂盒说明书进行植物总RNA提取。以总RNA为模板,用随机引物反转录合成cDNA。通过4次PCR和5′-RACE扩增PRSV-SD基因组片段。 1.2.3 克隆及序列测定 电泳分离PCR产物,切胶回收后连接到pMD18-T载体上,转化E. coli DH5α感受态细胞,挑选阳性克隆进行核苷酸测序。     

云南文山州首次在感病西葫芦上检测到番木瓜环斑病毒

为明确引起云南文山州广南县西葫芦发生蕨叶、斑驳、泡状斑等症状的病原,从采集的西葫芦病样中提取总RNA,利用RT-PCR扩增后经电泳检测及测序获得大小为887bp的片段;经过BLAST序列同源性比对分析,该病原与番木瓜环斑病毒Papaya ringspot virus(PRSV)(KY996464)基因序列同源性达87%。由此可知,侵染文山州西葫芦的病毒病原是番木瓜环斑病毒。     

弱毒黄萎病菌的筛选及诱导向日葵交互保护作用的条件优化

向日葵黄萎病是一种难以防治的土传病害。本研究通过毒力和致病力双重评价标准筛选得到一株弱毒黄萎病菌菌株Vn-1，采用诱导接种和挑战接种方法，明确了弱毒菌株Vn-1可以诱导向日葵产生对黄萎病菌的交互保护作用。本试验进一步对菌株Vn-1诱导间隔期和诱导浓度进行了条件优化，并在最优条件下探究其交互保护作用的持效性以及对向日葵植株生长情况的影响。结果表明，菌株Vn-1诱导向日葵交互保护的最佳诱导间隔为5 d，最佳诱导浓度为1×10⁶孢子/mL；在最佳诱导条件下，防效最大可达72.27%，且在挑战接种44 d后维持防效在30%以上；对比直接接种强毒菌株处理，最佳诱导条件下，向日葵株高和根长显著提高，表明菌株Vn-1诱导能够减小强毒菌株对向日葵植株生长的抑制作用。     

利用病毒弱株系接种防治脐橙茎陷点病

在田间对枳砧森田脐橙利用选出的柑桔衰退病毒(CTV)和脉突病毒(CVEV)弱毒系对CTV强毒系的交叉保护作用进行了测定。结果表明,经CTV M—15A和M—16A预先接种的植株,树势强健,且果大。此外,用和CTV多克隆抗血清无反应的No.145及CVEV NO.1605弱毒系预接种植株生长也很好。     

番木瓜环斑病毒株系的分子生物学方法鉴定

 以PRSV株系特异性引物对PRSV的PRSV126(PRSV日本分离物)、Ys、Vb和Sm等株系进行RT-PCR方法鉴定,引物PR21/PR22能把Ys从Vb和Sm中鉴定出来,PR300/PR301则能把Vb从Ys、Sm和PRSV126中鉴定出来;用限制性内切酶Hae Ⅱ、Sau3A I和Hinf I对PRSV的PRSV126、Ys、Vb和Sm等株系进行单酶切RT-PCR-RFLP分析,Hinf I能把PRSV126与Ys、Vb和Sm鉴别开来,Sau3A I能把Ys与Vb和Sm鉴别开来,Hae Ⅱ则能把Ys与PRSV126、Vb和Sm鉴别开来;以P1/P2为引物,对Vb、Ys和Sm株系进行RT-PCR-RFLP-SSCP分析,结果能一次把三者较好地区别开来。     

番木瓜抗环斑病毒突变体抗性遗传及RAPD标记

 ⁶⁰Coγ-射线处理番木瓜种子,从诱变一代中筛选到1株耐环斑病毒(PRSV)的变异植株(M₁),其侧芽组培后代(VM₁)部分植株也表现出了耐病性,以VM₁为母本进行回交,人工接种病毒鉴定回交后代(BM₂)的抗病性,结果为:在出自部分回交果实的BM₂群体中,包含有对PRSV Ys和Vb株系具抗性的植株,抗感分离比为1:1,但未发现抗Sm株系的植株;BM₂抗病两性株自交或以其为母本进行回交,分别获得诱变第三代(M₃)及回交诱变第三代(BM₃),人工接种PRSVYs株系,结果表明,BM₃抗感分离比也为1:1,因而认为辐射处理突变产生了具PRSV株系专化性的显性抗病基因,命名为Rys;但M₃抗感分离比不符合显性单基因3:1理论值,认为是单倍体选择的结果。运用BSA法,在BM₂中寻找到一个与抗病性密切相关的RAPD标记,经在BM₂、M₃及BM₃抗感群体中检验,可作为抗病育种的辅助选择标记。Rys是番木瓜栽培种中发现的第一个抗PRSV基因。     

FgLEU1在禾谷镰刀菌亮氨酸生物合成及产毒致病中起关键作用

为挖掘新型药剂的潜在靶标,利用靶向基因敲除和互补技术研究赤霉病病原菌禾谷镰刀菌Fusarium graminearum中必需氨基酸亮氨酸合成酶编码基因FgLEU1的功能,并测定禾谷镰刀菌的生物学表型。结果表明,FgLEU1编码亮氨酸合成途径中的3-异丙基苹果酸脱水酶,其敲除突变体表现亮氨酸营养缺陷。生物学表型测定结果显示,与野生型菌株相比,FgLEU1敲除突变体的产孢量和孢子萌发率显著下降,产孢量仅为野生型菌株的20.96%,培养4 h后孢子萌发率下降了49.45%,且合成脱氧雪腐镰刀烯醇（呕吐毒素）能力丧失,在麦穗上的致病力下降,仅能侵染接种小穗,赤霉病症状不能扩展。外源添加一定量的亮氨酸、FgLeu1催化产物或导入含启动子的全长FgLEU1基因可以恢复敲除突变体表型缺陷。表明FgLEU1基因在禾谷镰刀菌亮氨酸合成、菌丝孢子形成及产毒致病过程中发挥着重要作用,可作为新型安全杀菌剂的潜在研发靶标,用于持续有效控制麦类赤霉病和镰刀菌毒素。     

马铃薯Y病毒HC-Pro第182位赖氨酸残基参与引起烟草叶脉坏死

 马铃薯Y病毒(potato virus Y,PVY)是侵染烟草的最重要病毒之一。不同PVY株系侵染烟草可引起不同症状,有些PVY株系可引起烟草叶脉坏死,严重影响烟草的产量和品质。PVY A12分离物属于NTN-NW株系,但侵染珊西烟(Nicotiana tabacum cv. Xanthi)不能引起叶脉坏死。分析发现,PVY分离物A12 HC-Pro第182和245位的氨基酸均为精氨酸(R),而能引起叶脉坏死的其他NTN-NW分离物HC-Pro的这2个位点均为赖氨酸(K)。PVY坏死株系N605的HC-Pro第182位和245位氨基酸也均为K。本研究通过定点突变,将N605侵染性克隆PVY^N605-GFP HC-Pro第245位残基K突变为R,突变体仍然能够引起叶脉坏死,而将其HC-Pro第182位残基K突变为R,突变体不能引起叶脉坏死。Western blot检测发现,2个突变体与野生型病毒CP蛋白在珊西烟中的表达水平没有明显差异。沉默抑制实验结果显示,2个突变体和野生型的HC-Pro抑制RNA沉默能力没有发生变化。初步确定PVY^N605-GFP HC-Pro第182残基K是引起叶脉坏死的关键氨基酸,推测PVY A12分离物不能引起烟草叶脉坏死的原因是其HC-Pro第182残基R引起的。     

番木瓜环斑病毒南瓜分离物外壳蛋白基因的克隆及序列分析

利用电镜和酶联免疫法在云南省采集到的5份南瓜病样中检测到番木瓜环斑病毒(Papayaring spot virus,PRSV)。为了进一步从分子水平确定云南省南瓜病毒病原种类,并为下一步转基因育种提供抗性基因,采用反转录PCR(RT-PCR)方法扩增了5个分离物的外壳蛋白(coat protein,CP)基因片段,并克隆到pGEM-T载体中。核苷酸序列测定表明,番木瓜环斑病毒石屏分离物(PRSV-SP)和番木瓜环斑病毒蒙自分离物(PRSV-MZ)的CP基因长873nt,编码290个氨基酸,番木瓜环斑病毒峨山分离物(PRSV-ES)、番木瓜环斑病毒版纳分离物(PRSV-BN)和番木瓜环斑病毒宾川分离物(PRSV-BC),3个分离物CP基因长867nt,编码288个氨基酸。PRSV5个分离物核苷酸序列的同源性在94%以上,氨基酸序列的同源性在96%以上。与国内外17个分离物相比,核苷酸序列同源性为89.6%~98.7%,氨基酸序列同源性为86.5%~99.6%。其中PRSV-SP和来自于越南分离物PRSV-V47无论是核苷酸序列,还是氨基酸序列同源性都达到了最高,而5个分离物与来自于巴西(PRSV-BR)、美国(PRSV-USA)、墨西哥(PRSV-Y)核苷酸序列同源性均低于90%。     

烟草赤星病菌对嘧菌酯抗性突变体的诱导及其生物学特性

为评价烟草赤星病致病菌链格孢Alternaria alternata对嘧菌酯的抗性风险，以敏感菌株J6为试材，通过菌丝药剂驯化和分生孢子紫外诱变诱导抗性突变体，并对抗性突变体的生物学特性进行了研究，同时对抗性突变体与敏感菌株线粒体的细胞色素b基因 (cyt b) cDNA序列全长进行了测序分析。结果表明:经药剂驯化未获得抗性突变体，而紫外诱变共获得7株抗性突变体，突变频率约为0.007%，抗性水平分别为5.27、8.28、25.28、12.82、6.14、9.28和52.91倍。适合度研究表明，抗性突变体与敏感菌株的分生孢子萌发能力及致病力相当，但分生孢子产生量均高于敏感菌株，菌丝生长速率除突变体6-1外均快于敏感菌株。cyt b基因cDNA序列分析表明:有4株抗性突变体在不同位点上发生了核苷酸突变，其中突变体6-7 cyt b的249位和871位碱基由T突变为C，但其编码的氨基酸未发生突变；突变体6-8 cyt b的734位碱基由T突变为C，引起所编码的245位丙氨酸突变为缬氨酸 (V245A)；突变体6-9 cyt b的510位碱基由T突变为A，所编码的170位由精氨酸替代了丝氨酸 (S170R)；突变体6-11 cyt b的732位碱基由T突变为A，所编码的244位由苯丙氨酸替代了亮氨酸 (L244F)，其776位碱基由T突变为C，所编码的259位由丙氨酸替代了缬氨酸（V259A），其1 156位碱基由A突变为G，所编码的氨基酸未发生变化。研究结果初步表明，烟草赤星病菌对嘧菌酯存在潜在的抗药性风险，其cyt b基因的点突变与其对嘧菌酯的抗药性有关。     

Importance of cucurbits in the epidemiology of Papaya ringspot virus type P

Papaya ringspot virus type P (PRSV‐P) systemically infects Carica papaya and species belonging to the family Cucurbitaceae. Attempts to recover PRSV‐P from naturally infected cucurbit plants grown near or among diseased papaya trees have shown conflicting results worldwide. This study aimed to evaluate the natural infection of cucurbit species grown among and near papaya trees infected with PRSV‐P in Brazil. Natural infection of cucurbits with PRSV‐P occurred in zucchini squash but not in watermelon and cucumber. However, several attempts to recover PRSV‐P from numerous Cucurbita pepo cv. Caserta (zucchini squash) plants grown 5–80 m from diseased papaya trees in the field failed. Mechanical inoculations of Cucurbita pepo cv. Caserta, Cucurbita maxima cv. Exposição (pumpkin), Cucumis sativus cv. Primepack Plus (cucumber) and Citrullus lanatus cv. Crimson Sweet (watermelon) with five Brazilian PRSV‐P isolates showed that zucchini squash was the most susceptible species followed by watermelon and cucumber, while pumpkin was not infected. The results confirmed the variable susceptibility of cucurbit species to experimental and natural PRSV‐P infection. Given these facts, the control of the disease through roguing should focus mainly on diseased papaya plants, as has been practised successfully in Brazil for many years, and on those cucurbits particularly known to be susceptible to natural infection with PRSV‐P.     

Vegetative compatibility groups inVerticillium dahliae isolates from cotton in Turkey

Verticillium dahliae Kleb. with a complicated genetic diversity is a widely distributed major pathogen resulting in cotton wilt, which causes high economic losses in cotton lint production in the cotton belt of Turkey. A collection of 70 TurkishV. dahliae isolates (68 from wilted cotton plants in 28 districts and two from watermelon plants in two districts) were tested for vegetative compatibility by observing heterokaryon formation among complementary nitrate-nonutilizing (nit) mutants. The mutants were tested against international reference tester isolates and also were paired with one another. Thirty-nine isolates were assigned to vegetative compatibility group (VCG) 2B, 19 to VCG2A and three to VCG4B. One isolate was self-incompatible and eight others could not be assigned to any of the identified VCGs because theirnit mutants showed negative reactions with the tester isolates of four VCGs or theirnit mutants reverted back to the wild type. This is the first report of VCGs inV. dahliae from cotton in Turkey.     

Suppression of fusarium wilt of radish by co-inoculation of fluorescentPseudomonas spp. and root-colonizing fungi

In an earlier study, treatment of radish seed with the bacteriumPseudomonas fluorescens WCS374 suppressed fusarium wilt of radish (Fusarium oxysporum f. sp.raphani) in a commercial greenhouse [Leemanet al., 1991b, 1995a]. In this greenhouse, the areas with fusarium wilt were localized or expanded very slowly, possibly due to disease suppressiveness of the soil. To study this phenomenon, fungi were isolated from radish roots collected from the greenhouse soil. Roots grown from seed treated with WCS374 were more abundantly colonized by fungi than were roots from nonbacterized plants. Among these were several species known for their antagonistic potential. Three of these fungi,Acremonium rutilum, Fusarium oxysporum andVerticillium lecanii, were evaluated further and found to suppress fusarium wilt of radish in a pot bioassay. In an induced resistance bioassay on rockwool,F. oxysporum andV. lecanii suppressed the disease by the apparent induction of systemic disease resistance. In pot bioassays with thePseudomonas spp. strains, the pseudobactin-minus mutant 358PSB^– did not suppress fusarium wilt, whereas its wild type strain (WCS358) suppressed disease presumably by siderophore-mediated competition for iron. The wild type strains of WCS374 and WCS417, as well as their pseudobactin-minus mutants 374PSB^– and 417PSB^– suppressed fusarium wilt. The latter is best explained by the fact that these strains are able to induce systemic resistance in radish, which operates as an additional mode of action. Co-inoculation in pot bioassays, ofA. rutilum, F. oxysporum orV. lecanii with thePseudomonas spp. WCS358, WCS374 or WCS417, or their pseudobactin-minus mutants, significantly suppressed disease (except forA. rutilum/417PSB^– and all combinations with 358PSB^–), compared with the control treatment, if the microorganisms were applied in inoculum densities which were ineffective in suppressing disease as separate inocula. If one or both of the microorganism(s) of each combination were applied as separate inocula in a density which suppressed disease, no additional suppression of disease was observed by the combination. The advantage of the co-inoculation is that combined populations significantly suppressed disease even when their individual population density was too low to do so. This may provide more consistent biological control. The co-inoculation effect obtained in the pot bioassays suggests that co-operation ofP. fluorescens WCS374 and indigenous antagonists could have been involved in the suppression of fusarium wilt of radish in the commercial greenhouse trials.Abbreviations CFU colony forming units - KB King's B - PGPR plant growth-promoting rhizobacteria - CQ colonization quotient     

Suppression of <Emphasis Type="Italic">Papaya ringspot virus</Emphasis> infection in <Emphasis Type="Italic">Carica papaya</Emphasis> with CAP-34, a systemic antiviral resistance inducing protein from <Emphasis Type="Italic">Clerodendrum aculeatum</Emphasis>

CAP-34, a protein from Clerodendrum aculeatum inducing systemic antiviral resistance was evaluated for control of Papaya ringspot virus (PRSV) infection in Carica papaya. In control plants (treated with CAP-34 extraction buffer) systemic mosaic became visible around 20 days that intensified up to 30 days in 56% plants. During this period, CAP-34-treated papaya did not show any symptoms. Between 30 and 60 days, 95% control plants exhibited symptoms ranging from mosaic to filiformy. In the treated set during the same period, symptoms appeared in only 10% plants, but were restricted to mild mosaic. Presence of PRSV was determined in induced-resistant papaya at the respective observation times by bioassay, plate ELISA, immunoblot and RT-PCR. Back-inoculation with sap from inoculated resistant plants onto Chenopodium quinoa did not show presence of virus. The difference between control and treated sets was also evident in plate-ELISA and immunoblot using antiserum raised against PRSV. PRSV RNA was not detectable in treated plants that did not show symptoms by RT-PCR. Control plants at the same time showed a high intensity band similar to the positive control. We therefore suggest that the absence/delayed appearance of symptoms in treated plants could be due to suppressed virus replication.     

Molecular characterization of two papaya ringspot virus isolates that cause devastating symptoms in Norte de Santander,Colombia

Papaya ringspot virus is an RNA virus that belongs to the genus Potyvirus, family Potyviridae and affects both papaya and cucurbits, causing great economic losses. PRSV isolates are divided into biotypes P and W; both biotypes naturally infect plants in the family Cucurbitaceae, whereas the P type also naturally infects papaya (Carica papaya L). In the present study, we report the full-length genome sequence of two PRSV-P isolates sampled from the Campo Hermoso (PRSV-CH) and Villa del Rosario (PRSV-VR) localities in Norte de Santander, Colombia. The genomes of these PRSV isolates are 10,326 nt in length and have a predicted ORF of 3344 aa. The identity among Colombian PRSV isolates is 96.9% and 97.3% at the nucleotide and deduced amino acid levels, respectively. PRSV isolates from China had the lowest identity at 78.3% and 89.2% (nucleotide-amino acid), whereas the highest identities were detected in PRSV isolates from Mexico, Venezuela and Hawaii. At the polyprotein level, the amino acid composition surrounding the active polyprotein cleavage sites differ in the Colombian PRSV sequences. The predicted cleavage site in P1/HC-Pro is LEQY/N – LEQY/S instead of MEQY/N. Both of the Colombian PRSV isolates have a putative recombination event in the P1 coding region, which is common in all PRSV isolates from the American continent. The new full-length PRSV sequences from Colombia provide a better understanding of the dynamics of papaya ringspot virus infections in papaya in Colombia and worldwide.     

Genetic analysis of an attenuated <Emphasis Type="Italic">Papaya ringspot virus</Emphasis> strain applied for cross-protection

Papaya ringspot virus (PRSV) HA 5-1, a nitrous acid-induced mild mutant of severe strain HA, widely applied for control of PRSV by cross-protection, was used to study the genetic basis of attenuation. Using infectious clones, a series of recombinants was generated between HA 5-1 and HA and their infectivity was analyzed on the systemic host papaya and the local lesion host Chenopodium quinoa. The recombinants that contained mutations in P1 and HC-Pro genes caused attenuated infection on papaya without conspicuous symptoms, similar to HA 5-1. The recombination and sequence analyses strongly implicated two amino acid changes in the C-terminal region of P1 and two in HC-Pro of HA 5-1 involved in the attenuated infection on papaya. The recombinants that infected C. quinoa plants without local lesions contained the same mutations in the C-terminal region of HC-Pro for attenuated infection on papaya. We conclude that both P1 and HC-Pro bear important pathogenicity determinants for the infection on the systemic host papaya and that the mutations in HC-Pro affecting pathogenicity on papaya are also responsible for the inability to induce hypersensitive reaction on C. quinoa.     

Erwinia amylovora hrpN mutants,blocked in harpin synthesis,express a reduced virulence on host plants and elicit variable hypersensitive reactions on tobacco

Erwinia amylovora is the bacterium responsible for fire blight, a necrotic disease affecting many rosaceous plants and especially pear tree and apple tree. A protein named harpin, secreted through the Hrp secretion pathway and able to elicit an hypersensitive reaction (HR) on tobacco has recently been isolated. Mutants inhrpN, the gene encoding harpin were described as non pathogenic on immature pear fruit and unable to elicit an HR on tobacco [Weiet al., 1992; Wei and Beer, 1993]. In this paper, the phenotype on plant ofhrpN mutants was carefully determined.hrpN mutants expressed a weak but significant virulence on host plants. Furthermore, when infiltrated into tobacco leaf mesophyll, thehrpN mutants elicited varied responses that fluctuated from null reaction to full necrosis of the infiltrated area. These results show that harpin is not absolutely required neither for pathogenicity on host plant nor for elicitation of an hypersensitive reaction on tobacco. Furthermore, in all the tests performed, mutant blocked in harpin secretion remained non pathogenic and unable to elicit an HR on tobacco. This suggests that factor(s), different from harpin, involved both in pathogenicity and HR eliciting ability is (are) secreted through the Hrp secretion pathway.Abbreviations HR hypersensitive reaction - NSI necrosis severity index - CFU colonie forming units     

Differentiation ofVerticillium dahliae populations on the basis of vegetative compatibility and pathogenicity on cotton

Complementary auxotrophic nitrate-nonutilizing (nit) mutants were used to investigate vegetative compatibility within 27 strains ofVerticillium dahliae isolated from several hosts originating from Africa, Asia, Europe and the United States. Using about 500nit mutants generated from these strains, three vegetative compatibility groups, 1, 2 and 4, were identified. Simultaneously, virulence of each strain was assessed on cultivars ofGossypium hirsutum, G. barbadense andG. arboreum, based upon Foliar Alteration Index (FAI) and Browning Index (BI) estimation. The strains in VCG1 were of both the cotton-defoliating pathotype and race 3 (on cotton) but were non pathogenic on tomato; those in VCG2 and VCG4 were of the nondefoliating pathotype and belonged to different races on cotton and on tomato. Hyaline mutants deriving from parental wild-type strain showed differences in pathogenicity but were always assigned to the parental VCG. A relationship was established between VCGs and the taxonomic position of host plants. Data fromnit pairings indicated that the sub-populations ofV. dahliae (VCGs) may not be completely isolated genetically.     

Vegetative Compatibility amongVerticillium dahliae Isolates from Watermelon in Greece

Vegetative compatibility among 17 isolates ofVerticillium dahliae obtained from watermelon, originating from eight regions of Greece, was investigated using complementation tests between nitrate-nonutilizing(nit) mutants. Among 529 chlorate-resistant sectors obtained, only 107 werenit mutants. These mutants were paired with tester strains (from Greece and other countries) of previously described vegetative compatibility groups (VCGs), and also were paired in many combinations among themselves. All isolates were self-compatible. Sixteen isolates were found to belong to VCG2. Only isolate V75 could not be assigned to a VCG, because the threenit mutants obtained from it showed negative reactions with the tester strains of four VCGs and with complementary mutants from other isolates. Based on this sample, we conclude that the population ofV. dahliae from watermelon in Greece is homogeneous in respect to VCG.