The recombinant isolate of cucurbit aphid-borne yellows virus from Brazil is a polerovirus transmitted by whiteflies

The severe yellowing disease (amarelão) on melon plants is a serious problem in Brazil, although the causative agent remained unknown for a long time. Recently, recombinant isolates of cucurbit aphid-borne yellows virus (CABYV) were reported as the possible causative agents of this disease on melon plants. Although aphids are known to be the vectors of the common type of CABYV isolates, almost no aphid colony was observed in the major melon fields in Brazil with high incidence of the severe yellowing disease. In contrast, whiteflies are often abundant. Based on this observation, the hypothesis of the transmission of recombinant CABYV by whiteflies was evaluated. After thorough transmission experiments, we found that this recombinant CABYV isolate was transmitted by the whitefly Bemisia tabaci MEAM1, but not by Aphis gossipii. Furthermore, the host response by whitefly-based inoculation in cucurbits and other indicator plants showed differences in host range when compared to the common type of CABYV. Due to its transmissibility by the whitefly and the distant relationship of the P3/P5 protein to CABYV, the name “cucurbit whitefly-borne yellows virus” is proposed for this recombinant CABYV. This is the second report of polerovirus transmission by the whitefly B. tabaci, following the report of pepper whitefly-borne vein yellows virus.     

Aphid-borne viruses infecting cultivated watermelon and squash in Spain: Characterization of a variant of cucurbit aphid-borne yellows virus (CABYV)

Aphid-borne viruses are responsible for major cucurbit diseases and hamper the sustainability of crop production. Systematic monitoring can reveal the occurrence and distribution of these viruses, in addition to unadvertised viruses, facilitating the control of diseases. For three consecutive (2018–2020) seasons, the presence of aphid-borne viruses was monitored from a total of 292 samples of watermelon and squash plants that showed yellowing symptoms in three major cucurbit-producing areas (Castilla La-Mancha, Alicante, and Murcia) in Spain. We observed that cucurbit aphid-borne yellows virus (CABYV) was the most common virus found (29%) in the plants from both crops. Likewise, except for squash samples from Castilla La-Mancha and Alicante, watermelon mosaic virus (WMV) was also found (23%) with a relatively high frequency. Furthermore, we observed the exacerbation of bright yellowing symptoms in watermelon plants that was often accompanied by considerable fruit abortion. CABYV was the only causative agent for this new yellowing disease, and two infectious cDNA clones (one from watermelon, CABYV-LP63, and another from melon, CABYV-MEC12.1) were constructed to further compare and characterize this CABYV disease. Based on the full-length genome, both isolates were grouped phylogenetically together within the Mediterranean clade. However, the Koch's postulates tests were only successfully completed for the LP63 isolate, which also showed several amino acid changes and two potential recombination events, as compared to MEC12.1. Remarkably, the LP63 isolate caused more severe symptoms and showed higher RNA accumulation than MEC12.1 in five cucurbit plant species. These results suggest that a novel CABYV variant that causes severe yellowing symptoms may be causing outbreaks in cucurbit crops.     

Immunodetection of the replicative complex of <Emphasis Type="Italic">Barley yellow dwarf</Emphasis><Emphasis Type="Italic">virus</Emphasis>-PAV <Emphasis Type="Italic">in vivo</Emphasis>

The genomic fragments of two open reading frames (ORFs) 1 and 2 of German and Canadian PAV isolates of Barley yellow dwarf virus (BYDV-PAV) were sequenced. Sequences only slightly differed from previously published sequences of this virus. Two polyclonal antisera against proteins encoded by ORFs 1 and 2 of a German ASL-1 isolate were developed using recombinant antigens expressed in E. coli as a fusion either to His₆− or thioredoxin-tags. In Western blot analysis with total protein extracts from BYDV infected plants, antisera efficiently recognized the 99 kDa fusion protein expressed from ORF1 and ORF2 (P1–P2 protein). Later in infection the P1–P2 protein disappeared and two smaller proteins, revealing sizes of 39 and 60 kDa, could be detected.     

甜瓜黄斑病毒三亚分离物S RNA的分子特征

 甜瓜黄斑病毒（Melon yellow spot virus, MYSV）首次发生于日本,造成甜瓜和黄瓜的严重损失,Kato等系统地研究了病毒的传播方式、寄主范围、超微结构和基因组特征,认为MYSV应为番茄斑萎病毒属（Tospovirus）的1个新种［1,2］。2006年以来,台湾的西瓜［3］和黄瓜［4］上相继发现MYSV。2009年春季,古勤生在海南三亚的保护地甜瓜上发现一种新发生的病毒病,发病率30%~100%,病株出现系统性黄化坏死斑点,为MYSV侵染的典型症状,结合分子检测结果判定病原为MYSV。     

Phylogeny of Egyptian isolates of <Emphasis Type="Italic">Cucumber mosaic virus</Emphasis> (CMV) and <Emphasis Type="Italic">Tomato mosaic virus</Emphasis> (ToMV) infecting <Emphasis Type="Italic">Solanum lycopersicum</Emphasis>

Four Cucumber mosaic virus (CMV) (CMV-HM 1–4) and nine Tomato mosaic virus (ToMV) (ToMV AH 1–9) isolates detected in tomato samples collected from different governorates in Egypt during 2014, were here characterized. According to the coat protein gene sequence and to the complete nucleotide sequence of total genomic RNA1, RNA2 and RNA3 of CMV-HM3 the new Egyptian isolates are related to members of the CMV subgroup IB. The nine ToMV Egyptian isolates were characterized by sequence analysis of the coat protein and the movement protein genes. All isolates were grouped within the same branch and showed high relatedness to all considered isolates (98–99%). Complete nucleotide sequence of total genomic RNA of ToMV AH4 isolate was obtained and its comparison showed a closer degree of relatedness to isolate 99–1 from the USA (99%). To our knowledge, this is the first report of CMV isolates from subgroup IB in Egypt and the first full length sequencing of an ToMV Egyptian isolate.     

Pink root rot of squash caused by Setophoma terrestris in Japan

Pink root rot of squash (Cucurbita moschata) caused by Setophoma terrestris was found in Maebashi, Gunma Prefecture, Japan in July 2007. Cucumber grafted on the squash first developed wilt and finally blight. These symptoms followed a severe pink root rot of the squash rootstock. The fungal isolates from diseased roots were identified as S. terrestris based on morphological characteristics and nucleotide sequences. One isolate induced a similar pink root rot but not entire wilting of the cucumber vine. We propose the name “pink root rot” (koshoku-negusare-byo in Japanese) of squash for the new disease.     

Molecular and biological characterization of a severe isolate of <Emphasis Type="Italic">Tomato chlorotic dwarf viroid</Emphasis> containing a novel terminal right (T<Subscript>R</Subscript>) domain sequence

Tomato chlorotic dwarf viroid (TCDVd) manually inoculated to transgenic (cv.‘Desiree’) potato plants containing antimicrobial cationic peptides failed to develop symptoms in above ground plant parts, but infected tubers were symptomatic. Plants from the infected tubers (second generation plants) emerged as either severely stunted (bushy stunt isolate, BSI) or tall and symptomless. Molecular characterization of BSI isolates showed TCDVd sequence variants 95 to 98% identical to TCDVd sequences from the database, while a viroid variant identical to TCDVd type isolate (acc # AF162131) was cloned from symptomless plants. The TCDVd BSI variants had novel U165C, GU177-178AA, and UCAC181-184CUUU nucleotide substitutions in the terminal right (T_R) domain of the viroid molecule. The cloned viroid cDNAs of the BSI were infectious to experimental (cv. ‘Sheyenne’) tomato plants causing stunted plants with profuse auxiliary shoots. Visual evaluation of the susceptibility of the BSI to 18 potato and 21 tomato cultivars revealed severe symptoms in most cultivars of both species. The progeny variants accumulating in each potato and tomato cultivar exhibited the same novel T_R domain in most cultivars, with only a slight variation in a few. The severity of the stunting symptoms induced by TCDVd from BSI isolates in both potato and tomato cultivars has not been noted previously with other TCDVd isolates and, as such, it is proposed that this new isolate be recognized as a distinct genotype. Emergence of this type of sequence variant in commercial fields or commercial tomato greenhouses could potentially cause relevant losses in both crops.     

南瓜蚜传黄化病毒甜瓜分离物的侵染性克隆构建

 南瓜蚜传黄化病毒(cucurbit aphid-borne yellows virus,CABYV)近年来发生普遍,严重威胁甜瓜的生产。前期构建了一个CABYV丝瓜分离物(CABYV-QY)的侵染性克隆,但其在甜瓜中的侵染率偏低,不宜用于甜瓜接种。本研究以CABYV甜瓜分离物CABYV-WS为研究对象,通过RT-PCR扩增、拼接获得全基因组序列,通过构建全长基因组cDNA克隆,分析其侵染性。结果显示,该分离物基因组全长为5682 nt,与CABYV-QY(MT943520)的核苷酸序列一致性为88.61%~100.00%,氨基酸为84.94%~100.00%。将cDNA克隆接种分析,发现所用的8个甜瓜品种均能被系统侵染并引起典型的黄化症状,侵染率为70%~100%。其中,甜瓜品种‘新密杂11号’和‘新密25号’感病性较强,接种CABYV后发病周期短且侵染率可达100%。CABYV侵染性克隆的成功构建有助于该病毒的分子致病性和寄主的抗病性等研究。     

Diversity and occurrence of chickpea chlorotic dwarf virus on legumes from Iran

Chickpea chlorotic dwarf virus (CpCDV; genus Mastrevirus, family Geminiviridae) is one of the most important legume-infecting viruses with a wide host range and geographic distribution in Africa and Asia. In Iran, CpCDV is common in chickpea (Cicer arietinum), but there is limited information about diversity and infections in plants of other legume species. In the current study, a total of 1671 leaf samples from different pulse crops with symptoms were collected in nine provinces of Iran, and the CpCDV infection status was tested by PCR and/or rolling circle amplification (RCA), resulting in the detection of CpCDV in samples of chickpea, lentil (Lens culinaris) and faba bean (Vicia faba) from different regions. Sequence analysis of complete genomes of 18 isolates recovered by digestion of RCA products revealed infection with isolates of the strains CpCDV-A and CpCDV-F in chickpea, lentil and faba bean. Phylogenetic analysis showed that the Iranian isolates of CpCDV were closely related to previously sequenced isolates of CpCDV-A and CpCDV-F. To the authors' knowledge, this is the first report of CpCDV-F in Iran. Using agroinoculation with infectious clones for one isolate each of CpCDV-A and CpCDV-F, infectivity was confirmed in both faba bean and chickpea, with plants developing leaf curling and/or yellowing. Both infectious clones also successfully infected Nicotiana benthamiana resulting in mild yellowing and intensive leaf curling for CpCDV-A, and dark-green mosaic, dwarfing and mild leaf curling for CpCDV-F.     

Incidence of potato viruses and characterisation of <Emphasis Type="Italic">Potato virus Y</Emphasis> variability in late season planted potato crops in Northern Tunisia

Surveys were conducted of symptomatic potato plants in late season crops, from the major potato production regions in Northern Tunisia, for infection with six common potato viruses. The presence of Potato leafroll virus (PLRV), Potato virus Y (PVY), Potato virus X (PVX), Potato virus A (PVA), Potato virus S (PVS) and Potato virus M (PVM) was confirmed serologically with virus infection levels up to 5.4, 90.2, 4.3, 3.8, 7.1 and 4.8%, respectively. As PVY was prevalent in all seven surveyed regions, further biological, serological and molecular typing of 32 PVY isolates was undertaken. Only one isolate was shown to induce PVY^O-type symptoms following transmission to tobacco and to react only against anti-PVY^O-C antibodies. Typical vein necrosis symptoms were obtained from 31 samples, six of which reacted against both anti-PVY^N and anti-PVY^O-C antibodies showing they contained mixed isolates, while 25 of them reacted only with anti-PVY^N antibodies. An immunocapture RT-PCR molecular test using a PVY^NTN specific primer pair set in the 5’NTR/P1 genomic region and examination of recombinant points in three genomic regions (HC-Pro/P3, CI/NIa and CP/3’NTR) showed that all 25 serotype-N PVY isolates were PVY^NTN variants with similar recombinations to the standard PVYNTN-H isolate. This is the first report of the occurrence of the PVY^NTN variant and its high incidence in late season potatoes in Tunisia.     

Occurrence of Tomato infectious chlorosis virus (TICV) in Jordan

A new disease on tomato ( Lycopersicon esculentum L.) caused by Tomato infectious chlorosis virus (TICV) has been detected for the first time in Jordan. Disease symptoms consisted of interveinal yellowing areas in older leaves followed by generalized yellowing. Using specific primers, Tomato infectious chlorosis virus was detected in symptomatic plants by RT-PCR. The amplified fragment (416 bp) was cloned and sequenced. Results of sequence analysis showed that the Jordanian isolate of TICV shared high nucleotide similarity (> 98%) with two other isolates from Japan and France. The distribution of TICV has been investigated in four regions in the Jordan Valley by non-radioactive dot blot hybridization. Data from the study showed high incidence of the disease in all surveyed regions. In addition, the expected size of the coat protein gene of TICV could be amplified from two symptomatic weeds species, Chenopodium album and Chenopodium murale , indicating that these weeds are natural hosts for the virus.     

A new yellowing disease of cucurbits caused by a luteovirus, cucurbit aphid-borne yellows virus

Since 1988, a yellowing disease of melon, cucumber and zucchini squash has been frequently observed in summer and autumn crops in France. Infected plants show yellowing and thickening of the older leaves; symptom intensity differs depending upon cultivar and season, and can be easily overlooked when plants are already infected by mosaic-inducing viruses or other pathogens. The disease is associated with the presence of a virus with spherical particles c. 25 nm in diameter, which is readily transmitted in a persistent manner by the aphids Myzus persicae and Aphis gossypii , but not mechanically. Serological analysis, nucleic-acid-hybridization experiments and host-range studies indicate that the virus is distantly related to, but distinct from, beet western yellows virus (BWYV). We propose to name this virus cucurbit aphid-borne yellows virus (CABYV), and to consider it as a tentative new member of the luteovirus group. CABYV was found to reduce significantly the yields of melon and cucumber by decreasing the number of fruit per plant but not by altering the fruit shape or quality. Preliminary investigations of the epidemiology of CABYV indicate that the virus is common in weeds and in cultivated cucurbits. CABYV was frequently detected in various regions of France, suggesting that it is one of the most prevalent viruses infecting cucurbits in this country.     

Host range and molecular analysis of Beet leaf yellowing virus,Beet western yellows virus-JP and Brassica yellows virus in Japan

Beet western yellows virus (BWYV; genus Polerovirus, family Luteoviridae) is one of the most important viruses causing yellowing disease of many field and vegetable crops. This study isolated different poleroviruses from sugar beet, spinach, radish and brassica in Japan, and identified them as BWYV-JP, Beet leaf yellowing virus (BLYV), Brassica yellows virus (BrYV) and BrYV-R (radish strain) based on host range and molecular analysis. Among over 100 plant species from 19 families inoculated with the vector Myzus persicae, about half of the species in 13 families were infected with some of these viruses. BLYV shared a similar host range to Beet mild yellowing virus (BMYV). These had a much more limited host range than BWYV-JP, which resembled BWYV-USA. The host range of BrYV was similar to that of Turnip yellows virus (TuYV). Phylogenetic analyses at the 5′ portion (replication-related gene) of the genome showed that BLYV, BMYV, BWYV (-JP and -USA) and Cucurbit aphid-borne yellows virus (CABYV) formed one large group, whereas BrYV and TuYV were grouped together. BLYV and BWYV were most closely related to each other, and were more closely related to CABYV than to BMYV. However, at the 3′ end (coat protein gene), BLYV and BWYV-JP formed a distinct group, separated from the BrYV group, which in turn was more closely related to BWYV-USA, BMYV, TuYV and Beet chlorosis virus, a group originating from outside Asia. Thus, this study presents host range differences and phylogeographical relationships of BWYV-like poleroviruses that are distributed worldwide.     

Recombination-like sequences in the upstream region of the phage-type DNA polymerase in ‘<Emphasis Type="Italic">Candidatus</Emphasis> Liberibacter asiaticus’

We determined and compared nucleotide sequences in the upstream region of the bacteriophage-type DNA polymerase in 15 Asian isolates of the citrus-greening bacterium ‘Candidatus Liberibacter asiaticus’ using the genomic information of American isolate Psy62. Some isolates had nucleotide changes and a large 156-bp deletion, which were concentrated in the putative bacteriophage genes ORF3 and ORF4 and corresponding intergenic region. Eight partial recombination-like patterns were observed in this variable region. Most were confirmed to have occurred between the original types 1 and 2, but a fragment, only several dozen nucleotides long and possibly derived from a third type, was also identified.     

南瓜蚜传黄化病毒遗传多样性分析

哈密瓜是新疆重要的经济作物之一.南瓜蚜传黄化病毒(cucurbit aphid-borne yellows virus,CABYV)是世界重要的瓜类黄化病毒,在新疆哈密瓜植株上普遍发生,严重威胁新疆哈密瓜的品质与产量.本研究从新疆阿克苏市随机采集120份哈密瓜叶片样品,经过RT-PCR检测证实38个样品为CABYV阳性...     

Sensitivity of<Emphasis Type="Italic">Botrytis cinerea</Emphasis> from greenhouse vegetables to DMIs and fenhexamid

Two hundred isolates ofBotrytis cinerea were collected from greenhouse vegetables between 2003 and 2006 to determine their baseline sensitivity to triadimefone, penconazole, tebuconazole and fenhexamid. Mean values of 50% effective concentrations (EC₅₀) of inhibiting growth were 4.853±5.102, 0.41±0.215, 0.19±0.099 and 0.36±0.891 mgl ⁻¹, respectively (mean±SD). Individuals ofB. cinerea in the population differed by a factor (EC₅₀ of the least sensitive isolate/EC₅₀ of the most sensitive isolate) of 6625, 20, 603 and 1800, respectively. Naturally fenhexamid-resistant isolates were detected with an unexpected high frequency of 10% although the pathogen population had never been exposed to this fungicide. The resistance level (mean EC₅₀ of resistant isolates / mean EC₅₀ of sensitive isolates) was 19.5. These naturally resistant isolates also were resistantin vivo, and there was no significant difference in growth rate, conidial production or pathogenicity ability between naturally resistant and wild sensitive isolates. These results indicated that there was a potential risk of practical resistance if fenhexamid was applied alone. Negative cross-resistance was observed between fenhexamid and tebuconazole in 90% of the naturally resistant isolates. Moreover, an obvious synergism of the antifungal activity of fenhexamid by tebuconazole was demonstrated in some of the naturally fenhexamid-resistant isolates. http://www.phytoparasitica.org posting May 9, 2007.     

Analysis of genetic diversity in <Emphasis Type="Italic">Monilinia fructicola</Emphasis> from the Ebro Valley in Spain using ISSR and RAPD markers

The genetic diversity of Spanish and French field populations of Monilinia fructicola, a quarantine fungal pathogen in Europe, was compared with that of Californian, Uruguayan, and New Zealand M. fructicola populations using inter simple sequence repeat (ISSR) and random amplified polymorphic DNA (RAPD) markers. Unweighted pair-group method with arithmetic average (UPGMA) cluster analysis and principal component analysis (PCA) of the ISSR data set revealed that the Spanish and French M. fructicola isolates were more closely related between themselves than to the non-European isolates. The levels of genetic diversity in the Spanish and French isolates are lower than those of the non-European isolates, indicating that M. fructicola is a recently introduced pathogen. UPGMA cluster analysis and PCA of the combined ISSR + RAPD data set of the European M. fructicola populations revealed that the Spanish isolates were more closely related among themselves than with the French isolates. Analysis of molecular variance partitioned the genetic variance to among the two regions (Spain and France) (20%), among the regional populations (35%), and within the populations in each region (45%) suggesting restricted gene flow between the three European populations. The observed index of association (I_A) in each European M. fructicola populations indicates that the French and Spanish populations of M. fructicola are mainly asexually reproducing, with the Sudanell population potentially having a teleomorphic stage. The present finding of low genetic diversity in the Spanish and French M. fructicola populations is probably due to founder effects and genetic drift.