scholarly journals First report of Cucurbit chlorotic yellows virus infecting cantaloupe (Cucumis melo L.) in Texas

Plant Disease ◽  
2021 ◽  
Author(s):  
Regina Nicole Hernandez ◽  
Thomas Isakeit ◽  
Maher Al Rwahnih ◽  
Rick Hernandez ◽  
Olufemi Joseph Alabi
Keyword(s):  
Nucleic Acid ◽  
Coat Protein ◽  
Cucumis Melo ◽  
Composite Sample ◽  
Acid Extraction ◽  
Total Nucleic Acid ◽  
First Report ◽  
Chlorotic Mottle ◽  
Cucumis Melo L ◽  
Cucurbit Chlorotic Yellows Virus

Texas is a major producer of cucurbits such as cantaloupe (Cucumis melo L.), but outbreaks of virus-like diseases often adversely affect yields. Little is known about the identity of the causal or associated viruses. During studies conducted in fall 2020 to explore the virome of cucurbit fields in Texas, a commercial cantaloupe field (~4.1 ha) in Cameron County was observed with virus-like symptoms of interveinal chlorotic mottle and foliar chlorosis and disease incidence was estimated at 100%. Virus-like symptoms including mosaic and leaf curl were also observed in six additional fields across five south and central Texas counties of Atascosa, Hidalgo, Fort Bend, Frio, and Wharton. Forty-six plants, which included 32 symptomatic and 14 non-symptomatic, were sampled from these fields for virus diagnosis and each sample was subjected to total nucleic acid extraction according to Dellaporta et. al. (1983). Initially, equal amounts of nucleic acids from 14 symptomatic plants (two/field) were pooled into one composite sample for preliminary diagnosis by high throughput sequencing (HTS). The cDNA library obtained from the composite sample with a TruSeq Stranded Total RNA with Ribo-Zero Plant Kit (Illumina) was sequenced on the Illumina NextSeq 500 platform, generating ~26.3 M single-end HTS reads (75 nucleotides [nt] each). Analyses of the reads according to Al Rwahnih et al. (2018) revealed several virus-like contigs; among them 23 contigs (206 to 741 nt) shared 98 to 100% nt identities to isolates of cucurbit chlorotic yellows virus (CCYV), genus Crinivirus, family Closteroviridae. Three pairs of CCYV-specific primers were designed from the HTS contigs with primers CCYV-v1330: 5′-AGTCCCTTACCCTGAGATGAA/CCYV-c2369: 5′-CGGAGCATTCGACAACTGAATA targeting ~1 kb fragment of the ORF1a (RNA1), primers CCYV-v4881: 5′-ATAAGGCGGCGACCTAATC/CCYV-c5736: 5′-GATCACTTGACCATCTCCTTCT targeting a ~0.9 kb fragment encompassing the coat protein (CP) cistron of CCYV (RNA2), and primers CCYV-v6362: 5′-CACCTCTTCCAGAACCAGTTAAA/CCYV-c7423: 5′-TGGGAACAACTTATTTCTCCTAGC targeting ~1 kb spanning partial minor coat protein (CPm) and p26 sequences (RNA2). Total nucleic acid extracts of each of the 46 samples from the seven fields were tested by two-step reverse transcription polymerase chain reaction using all three CCYV-specific primer pairs and they yielded amplicons of expected sizes from all five symptomatic cantaloupe samples from the Cameron County field and one additional symptomatic butternut squash sample from a field in Hidalgo County. The DNA bands from three randomly chosen cantaloupe samples were cloned and sequenced as previously described (Oke et al. 2020). In pairwise comparisons, the obtained 1,040 nt ORF1a (MW584332-334), 753 nt complete CP (MW584335-337), and 1,062 nt CPm/p26 (MW584338-340) gene specific sequences from the three samples shared 100% nt identity with each other, and 99-100% nt identities with corresponding RNA1 (AB523788) and RNA2 (AB523788) sequences of the exemplar isolate of CCYV. This is the first report of CCYV in Texas, thus expanding the current geographical range of the virus in the U.S. that includes California (Wintermantel et al. 2019) and Georgia (Kavalappara et al. 2021). The abundance of whiteflies of the Bemisia tabaci species complex in south Texas and other major U.S. cucurbit production areas presents additional challenges to virus disease management.

scholarly journals First report of cucurbit chlorotic yellows virus infecting cucumber in South Korea

Plant Disease ◽  
2021 ◽  
Author(s):  
Hae-Ryun Kwak ◽  
Hui-Seong Byun ◽  
Hong-Soo Choi ◽  
Jong-Woo Han ◽  
Chang-Seok Kim ◽  
...  
Keyword(s):  
Coat Protein ◽  
South Korea ◽  
East Asia ◽  
Rna Extraction ◽  
Rt Pcr ◽  
Specific Primers ◽  
Total Rna ◽  
First Report ◽  
Chlorotic Mottle ◽  
Cucurbit Chlorotic Yellows Virus

In October 2018, cucumber plants showing yellowing and chlorotic mottle symptoms were observed in a greenhouse in Chungbuk, South Korea. The observed symptoms were similar to those caused by cucurbit aphid-borne yellows virus (CABYV), which has been detected on cucumber plants in the region since it was reported on melon in Korea in 2015 (Lee et al 2015). To identify the potential agents causing these symptoms, 28 samples from symptomatic leaves and fruit of cucumber plants were subjected to total RNA extraction using the Plant RNA Prep Kit (Biocubesystem, Korea). Reverse transcription polymerase chain (RT-PCR) was performed on total RNA using CABYV specific primers and protocols (Kwak et al. 2018). CABYV was detected in 17 of the 28 samples, while 11 symptomatic samples tested negative. In order to identify the cause of the symptoms, RT-PCR was performed using cucurbit chlorotic yellows virus (CCYV) and cucurbit yellow stunting disorder virus (CYSDV) specific primers (Wintermantel et al. 2019). Eight of the 28 samples were positive using the CCYV specific primers while seven samples were infected with only CCYV and one contained a mixed infection of CABYV with CCYV. None of the samples tested positive for CYSDV. The expected 373 nt amplicons of CCYV were bi-directionally sequenced, and BLASTn analysis showed that the nucleotide sequences shared 98 to 100% identity with CCYV isolates from East Asia, including NC0180174 from Japan. Two pairs of primers for amplification of the complete coat protein and RNA-dependent RNA polymerase (RdRp) genes (Wintermantel et al., 2019) were used to amplify the 753bp coat protein and 1517bp RdRp genes, respectively. Amplicons of the expected sizes were obtained from a CCYV single infection and ligated into the pGEM T- Easy vector (Promega, WI, USA). Three clones from each amplicon were sequenced and aligned using Geneious Prime and found to have identical sequences (Genbank accession nos. MW033300, MW033301). The CP and RdRp sequences demonstrated 99% nucleotide and 100% amino acid identity with the respective genes and proteins of the CCYV isolates from Japan. This study documents the first report of CCYV in Korea. Since CCYV was first detected on melon in Japan, it has been reported in many other countries including those in East Asia, the Middle East, Southern Europe, North Africa, and recently in North America. CCYV has the potential to become a serious threat to production of cucurbit crops in Korea, particularly due to the increasing prevalence of the whitefly, Bemisia tabaci, in greenhouse production systems. It will be important to continue monitoring for CCYV and determine potential alternate hosts in the region to manage and prevent further spread of CCYV in Korea.

scholarly journals First report of Cucurbit chlorotic yellows virus infecting Cucumis melo (muskmelon and oriental melon) in Korea

Plant Disease ◽  
2021 ◽  
Author(s):  
In Sook Cho ◽  
Tae-Bok Kim ◽  
Ju-Yeon Yoon ◽  
Bong Nam Chung ◽  
John Hammond ◽  
...  
Keyword(s):  
Mosaic Virus ◽  
Cucumis Melo ◽  
Korean Isolate ◽  
Rt Pcr ◽  
Genome Database ◽  
Total Rna ◽  
First Report ◽  
Oriental Melon ◽  
Cucumis Melo L ◽  
Cucurbit Chlorotic Yellows Virus

In December 2018, virus-like symptoms (yellowing, vein clearing) were observed on 2% of muskmelon (Cucumis melo L.) plants in plastic houses on a farm in Gyeongsang province, Korea Total RNA from two symptomatic and two asymptomatic plants was extracted using RNeasy Plant Mini Kit (Qiagen, Germany) for high throughput sequencing (HTS). After pre-processing and Ribo-Zero rRNA removal, a cDNA library was prepared (Illumina TruSeq Stranded Total RNA kit) and sequenced (Illumina NovaSeq 6000 system: Macrogen Inc. Korea). De novo assembly of 88,222,684 HTS reads with Trinity software (r20140717) yielded 146,269 contigs of 201-28,442 bp, which were screened against the NCBI viral genome database by BLASTn. Contigs from cucumber mosaic virus (CMV), melon necrotic spot virus (MNSV), tobacco mosaic virus (TMV) and watermelon mosaic virus (WMV) were identified, all previously reported in Korea. Two contigs (8,539 and 8,040 bp) with 99.9% sequence identity to distinct cucurbit chlorotic yellows virus (CCYV) isolates (JN641883, RNA1, Taiwan; MH819191, RNA2, China) were also identified. The ten sequences most closely related to each RNA of the Korean isolate (≥99% coverage, ≥99.6% nt identity) were from Japan, China, Taiwan, or Israel. CCYV presence was confirmed by reverse transcription-PCR (RT-PCR) using newly designed specific primers, RdRp-F/RdRp-R (5’-ACCGAACACTTGGCTATCCAA-3’/5’-CTTAATGCCGCGTATGAACTCA-3’) span style="font-family:'Times New Roman'; letter-spacing:-0.5pt">and HSP-F/HSP-R (5’-TGAACGACACTGAGTTCATTCCTA-3’/5’-CGCCAAGATCGTACATGAGGAA-3’), against RNA dependent RNA polymerase (RdRp; RNA1) and the heat shock protein 70 homolog (HSP70h; RNA2). Symptomatic samples yielded products of expected sizes (RdRp,450 bp; HSP70h, 510 bp) while asymptomatic samples did not. The amplicons were cloned, and two clones of each were sequenced (BIONEER, Korea; GenBank acc. nos. LC592226 and LC592227) showing 100% and 99.2% nt identity with RdRp and HSP70h genes of Chinese CCYV isolate SD (MH819190 and MH819191, respectively) and other Asian isolates. Primers specific for CMV, WMV, beet pseudo-yellows virus (BPYV) (Okuda et al., 2007), TMV (Kim et al., 2018), MNSV (F/R, 5ʹ-ATCTCGCATTTGGCATTACTC-3ʹ/5ʹ-ATTTGTAGAGATGCCAACGTA-3ʹ), cucurbit yellow stunting disorder virus (CYSDV; Zeng et al., 2011) and cucurbit aphid-borne yellows virus (CABYV; F/R, 5ʹ-CGGTCTATTGTCTGCAGTACCA-3ʹ/5ʹ- GTAGAGGATCTTGAATTGGTCCTCA-3ʹ) were also used. None of these viruses were detected in the symptomatic samples, but both asymptomatic plants were positive for CMV and WMV, and one also for MNSV. In June and September 2020, muskmelon and oriental melon (Cucumis melo L. var. makuwa) plants with yellowing disease (incidence 80-90%) and whiteflies were observed in all investigated plastic houses of one muskmelon and one oriental melon farm in Gyeonggi and Jeolla provinces. Symptomatic samples (14 muskmelon; 6 oriental melon) were collected and RT-PCR tested as above; 19/20 samples were positive for CCYV, but none for the other viruses. The oriental melon sequence (LC592895, LC592230) showed 99.7% and 100% nt identity with the RdRp and HSP70h genes of Chinese isolate SD, respectively. CCYV was first reported in Japan (Okuda et al., 2010), Taiwan, and China (Huang et al., 2010; Gu et al., 2011); to our knowledge, this is the first report of CCYV infecting muskmelon and oriental melon in Korea. Whitefly-transmitted CCYV could present a serious threat of yield losses to cucurbit crops in Korea, requiring control of vector populations to prevent spread of CCYV.

scholarly journals First report of squash vein yellowing virus naturally infecting butternut squash (Cucurbita moschata) in Texas

Plant Disease ◽  
2021 ◽  
Author(s):  
Regina Nicole Hernandez ◽  
Thomas Isakeit ◽  
Maher Al Rwahnih ◽  
Rick Hernandez ◽  
Olufemi Joseph Alabi
Keyword(s):  
Nucleic Acid ◽  
High Throughput Sequencing ◽  
Composite Sample ◽  
Cucurbita Moschata ◽  
Total Nucleic Acid ◽  
Cdna Synthesis ◽  
First Report ◽  
Two Samples ◽  
Hidalgo County ◽  
Yellowing Virus

Virus diseases are major constraints to the production of cucurbits in the Texas Lower Rio Grande Valley. In September 2020, a ~8.1 ha butternut squash (Cucurbita moschata) field in Hidalgo County, Texas, was observed with virus-like symptoms of vein yellowing, leaf curl, mosaic, and foliar chlorosis. The proportion of plants with virus-like symptoms in this field was estimated at 30% and seven samples (symptomatic = 5; non-symptomatic = 2) were collected randomly for virus diagnosis. Initially, equimolar mixtures of total nucleic acid extracts (Dellaporta et. al. 1983) from two symptomatic samples from this field and extracts from 12 additional symptomatic samples from six other fields across south and central Texas was used to generate one composite sample for diagnosis by high throughput sequencing (HTS). The TruSeq Stranded Total RNA with Ribo-Zero Plant Kit (Illumina) was used to construct cDNA library from the composite sample, which was then sequenced on the Illumina NextSeq 500 platform. More than 26 million single-end HTS reads (75 nt each) were obtained and their bioinformatic analyses (Al Rwahnih et al. 2018) revealed several virus-like contigs belonging to different species (data not shown). Among them, 6 contigs that ranged in length from 429 to 3,834 nt shared 96 to 100% identities with isolates of squash vein yellowing virus (SqVYV), genus Ipomovirus, family Potyviridae. To confirm the HTS results, total nucleic acid extracts from the cucurbit samples from all seven fields (n = 46) were used for cDNA synthesis with random hexamers and the PrimeScript 1st strand cDNA Synthesis Kit (Takara Bio). A 1-μL aliquot of cDNA was used in 12.5-μL PCR reaction volumes with PrimeSTAR GXL DNA Polymerase (Takara Bio) and two pairs of SqVYV-specific primers designed based on the HTS derived contigs. The primer pairs SqYVV-v4762: 5′-CTGGATTCTGCTGGAAGATCA & SqYVV-c5512: 5′-CCACCATTAAGGCCATCAAAC and SqYVV-v8478: 5′-TTTCTGGGCAAACAAACATGG & SqYVV-c9715: 5′-TTCAGCGACGTCAAGTGAG targeted ~0.75 kb and ~1.2 kb fragments of the cylindrical inclusion (CI) and the complete coat protein (CP) gene sequences of SqVYV, respectively. The expected DNA band sizes were obtained only from the five symptomatic butternut squash samples from the Hidalgo Co. field. Two amplicons per primer pair from two samples were cloned into pJET1.2/Blunt vector (Life Technologies) and bidirectionally Sanger sequenced, generating 753 nt partial CI specific sequences (MW584341-342) and 1,238 nt that encompassed the complete CP (MW584343-344) of SqVYV. In pairwise comparisons, the partial CI sequences shared 100% nt/aa identity with each other and 98-99% nt/aa identity with corresponding sequences of SqVYV isolate IL (KT721735). The CP cistron of TX isolates shared 100% nt/aa identity with each other and 90-98% nt (97-100% aa) identities with corresponding sequences of several SqVYV isolates in GenBank, with isolates IL (KT721735) and Florida (EU259611) being at the high and low spectrum of nt/aa identity values, respectively. This is the first report of SqVYV in Texas, naturally occurring in butternut squash. SqVYV was first discovered in Florida (Adkins et al. 2007) and subsequently reported from few other states in the U.S. (Adkins et al. 2013; Egel and Adkins 2007; Batuman et al. 2015), Puerto Rico (Acevedo et al. 2013), and locations around the world. The finding shows an expansion of the geographical range of SqVYV and adds to the repertoire of cucurbit-infecting viruses in Texas. Further studies are needed to determine the prevalence of SqVYV in Texas cucurbit fields and an assessment of their genetic diversity.

QTL mapping for resistance to Cucurbit chlorotic yellows virus in melon (Cucumis melo L.)

Euphytica ◽  
2018 ◽  
Vol 214 (12) ◽  
Author(s):  
Yoichi Kawazu ◽  
Koichiro Shimomura ◽  
Shoichi Maeda ◽  
Yoichi Yamato ◽  
Shigenori Ueda ◽  
...  
Keyword(s):  
Qtl Mapping ◽  
Cucumis Melo ◽  
Cucumis Melo L ◽  
Cucurbit Chlorotic Yellows Virus

scholarly journals First Report of Pythium splendens Associated with Severe Wilt of Muskmelon (Cucumis melo) in Oman

Plant Disease ◽  
2008 ◽  
Vol 92 (2) ◽  
pp. 313-313 ◽  
Author(s):  
A. M. Al-Sa'di ◽  
M. L. Deadman ◽  
F. A. Al-Said ◽  
I. Khan ◽  
M. Al-Azri ◽  
...  
Keyword(s):  
Cucumis Melo ◽  
Morphological Characteristics ◽  
Fruit Production ◽  
Maturation Stage ◽  
Vegetable Crops ◽  
First Report ◽  
Sultan Qaboos University ◽  
Pythium Splendens ◽  
Pathogenicity Tests ◽  
Cucumis Melo L

Muskmelon (Cucumis melo L.) is one of the most important vegetable crops in Oman. In the fall of 2004, sudden wilt was observed in muskmelon grown in a field at Sultan Qaboos University, Muscat. The disease was characterized by rapid collapse of vines and muskmelon plants at the fruit production to maturation stage, associated with brown-to-dark brown rotted primary and secondary roots. The disease resulted in death of more than 85% of muskmelon plants in that field. On potato dextrose agar (PDA), with published methods (1), Pythium spp. were consistently isolated from crowns and roots of plants showing wilt symptoms. Further identification of five isolates of Pythium with sequences of the internal transcribed spacer (ITS) of the ribosomal DNA (1) using ITS1 and ITS4 primers produced a nucleotide sequence 806 bp long, which was identical among all isolates. Comparison with sequences deposited at the National Center for Biotechnology Information revealed 100% nucleotide similarity to a previously published sequence (Accession No. DQ381808) of isolate P091 of P. splendens from cucumber from Oman, for which identification has also been confirmed by morphological characteristics. The sequence of one isolate of P. splendens (P222) was assigned GenBank Accession No. EF546436 and deposited at CBS under Accession No. CBS121855. In pathogenicity tests conducted in a greenhouse, P. splendens induced damping-off symptoms on 7-day-old muskmelon seedlings and also reproduced the same wilt symptoms observed in the field when 2-month-old muskmelon plants were inoculated with 3-day-old P. splendens grown in PDA. To our knowledge, this is the first report of association of P. splendens with wilt of muskmelon in Oman. Reference: (1) A. M. Al-Sa'di et al. Plant Pathol. 56:140, 2007.

scholarly journals First Report of Cucurbit chlorotic yellows virus Infecting Melon in China

Plant Disease ◽  
2011 ◽  
Vol 95 (3) ◽  
pp. 354-354 ◽  
Author(s):  
R. Zeng ◽  
F. M. Dai ◽  
W. J. Chen ◽  
J. P. Lu
Keyword(s):  
Cucumis Melo ◽  
Specific Primers ◽  
Virus Diagnosis ◽  
High Tunnel ◽  
First Report ◽  
Reverse Transcription Pcr ◽  
Yellow Color ◽  
Large Populations ◽  
Pcr Products ◽  
Cucurbit Chlorotic Yellows Virus

In October 2007, symptoms of chlorosis on the upper leaves and a bright yellow color on the lower leaves were observed sporadically on hami melon (Cucumis melo cv. Xuelihong) in a high tunnel in Nanhui of Shanghai, China. Disease progresses from initial mottling of leaves into leaves that are completely yellow with the veins remaining green. The oldest leaves develop symptoms first, so these leaves have a pronounced even yellow color. In October 2009, these symptoms were found in all melons produced in the suburbs of Shanghai. These symptoms were similar to those caused by Cucurbit yellow stunting disorder virus (CYSDV) and Cucurbit chlorotic yellows virus (CCYV) (1–3). Twelve samples from symptomatic melons were collected in the Jiading, Nanhui, Fengxian, and Chongming districts of Shanghai for virus diagnosis. Large populations of whiteflies were observed in association with the diseased cucurbit crops. Total RNA was extracted with Trizol reagents (Invitrogen, Carlsbad, CA). We used random primers (9-mer) for reverse transcription-PCR. Extracts were for CYSDV using specific primers CYSDV-CP-F (5′-ATGGCGAGTTCGAGTGAGAA-3′) and CYSDV-CP-R (5′-TCAATTACCACAGCCACCTG-3′) to amplify a 756-bp fragment of coat protein gene and CCYV using specific primers CCYV-HSP-F1 (5′-TGCGTATGTCAATGGTGTTATG-3′) and CCYV-HSP-R1 (5′-ATCCTTCGCAGTGAAAAACC-3′) to amplify a 462-bp fragment of the HSP gene (1). CYSDV was not found in all samples. The expected 462-bp target fragment of CCYV was obtained in all samples but not from any of the healthy controls. All the 462-bp PCR products were cloned to pGEM-T vector (Promega, Madison, WI) and sequenced. All sequences obtained were homologous. A comparison of the submitted sequence (GenBank Accession No. HQ148667) with those in GenBank showed that the sequence had 100% nucleotide identity to the Hsp70h sequences of (CCYV) isolates from Japan (Accession Nos. AB523789 and AB457591) (1,4), Taiwan (Accession No. HM120250) (2), and mainland of China (Accession Nos. GU721105, GU721108, and GU721110). CCYV is a new member of the genus Crinivirus, first discovered in Japan in 2004 (4) and reported in Taiwan in 2009 (2). To our knowledge, this is the first report of CCYV on melon in China. References: (1) Y. Gyoutoku et al. Jpn. J. Phytopathol. 75:109, 2009. (2) L.-H. Huang et al. Plant Dis. 94:1168, 2010. (3) L. Z. Liu et al. Plant Dis.94:485, 2010. (4) M. Okuda et al. Phytopathology 100:560, 2010.

scholarly journals First Report of Strawberry necrotic shock virus in China

Plant Disease ◽  
2011 ◽  
Vol 95 (9) ◽  
pp. 1198-1198 ◽  
Author(s):  
L. Li ◽  
H. Yang
Keyword(s):  
Nucleic Acid ◽  
Home Garden ◽  
Tobacco Streak Virus ◽  
Chinese Isolate ◽  
Streak Virus ◽  
Total Nucleic Acid ◽  
Rt Pcr ◽  
Viral Pathogen ◽  
First Report ◽  
Strawberry Vein Banding Virus

Strawberry necrotic shock virus (SNSV) is an economically important viral pathogen that infects Fragaria and Rubus spp. SNSV was first identified in the 1950s and early studies indicated that SNSV was a strain of Tobacco streak virus (TSV). Recently, it was shown that SNSV was a distinct virus based on molecular characterization (2). Currently, SNSV is a tentative member of the Ilarvirus genus in the Bromoviridae family. In 2008, a small sampling survey for SNSV was done in Heilongjiang Province of China, and 15 strawberry samples were collected from symptomless strawberries in a home garden that had more than 5 years of strawberry cultivation history. Total nucleic acid was extracted from strawberry leaflets by modified cetyltrimethylammoniumbromide methods (3). Reverse transcription (RT)-PCR was operated with the published primer pair CPbeg F/CPend R (2). Amplified DNA fragments with the predicted size were obtained only in one strawberry sample, which was further cloned and sequenced. The sequence (GenBank Accession No. HQ830017) was closely related and highly homologous (89.7 to 98.5% identity) to that of viral isolates (GenBank Accession Nos. AY363228-AY363242) from Fragaria and Rubus spp. Phylogenetic analysis based on nucleotide sequence of the coat protein gene was done with the neighbor-joining method of MEGA 4.0 software. The result showed that all the isolates of SNSV fell into two distinct clades. The Chinese isolate formed one small clade with Japanese isolate 1291. The isolate was also transmitted to Chenopodium quinoa by mechanical inoculation in the greenhouse, and the symptom of chlorotic mottling could be found in C. quinoa and detected by RT-PCR. To determine whether the sample was infected by other strawberry viruses, RT-PCR assays with the published primer pairs SVBVdeta/SVBVdetb, SMoVdeta/SMoVdetb, and SMYEVdeta/SMYEVdetb were also performed for detection of Strawberry vein banding virus, Strawberry mottle virus, and Strawberry mild yellow edge virus using total nucleic acid extracted from the SNSV-positive sample as a template (1). The result indicated that it had been also infected by Strawberry mild yellow edge virus, although no visible symptoms were observed. To our knowledge, this is the first report of SNSV in strawberry in China. Additional work is needed to elucidate the biological characterization and significance of the finding. References: (1) J. R. Thompson et al. J. Virol. Methods 111:85, 2003. (2) I. E. Tzanetakis et al. Arch. Virol. 149:2001, 2004. (3) H. Y. Yang et al. Acta Hortic. 764:127, 2007.

scholarly journals First Report of Fusarium incarnatum-equiseti Species Complex Causing Leaf Spot on Rockmelon (Cucumis melo L.) in Malaysia

Plant Disease ◽  
2020 ◽  
Author(s):  
Siti Izera Ismail ◽  
Nur Ainina Noor Asha ◽  
Dzarifah Zulperi
Keyword(s):  
Leaf Spot ◽  
Cucumis Melo ◽  
Species Complex ◽  
Fruit Rot ◽  
Peat Moss ◽  
Distilled Water ◽  
Conidial Suspension ◽  
Isolation Technique ◽  
First Report ◽  
Cucumis Melo L

Rockmelon, (Cucumis melo L.) is an economically important crop cultivated in Malaysia. In October 2019, severe leaf spot symptoms with a disease incidence of 40% were observed on the leaves of rockmelon cv. Golden Champion at Faculty of Agriculture, Universiti Putra Malaysia (UPM). Symptoms appeared as brown necrotic spots, 10 to 30 mm in diameter, with spots surrounded by chlorotic halos. Pieces (5 x 5 mm) of diseased tissue were sterilized with 0.5% NaOCl for 1 min, rinsed three times with sterile distilled water, plated onto potato dextrose agar (PDA) and incubated at 25°C for 7 days with a 12-h photoperiod. Nine morphologically similar isolates were obtained by using single spore isolation technique and a representative isolate B was characterized further. Colonies were abundant, whitish aerial mycelium with orange pigmentation. The isolates produced macroconidia with 5 to 6 septa, a tapered with pronounced dorsiventral curvature and measured 25 to 30 μm long x 3 to 5 μm wide. Microconidia produced after 12 days of incubation were single-celled, hyaline, ovoid, nonseptate, and 1.0 to 3.0 × 4.0 to 10.0 µm. Morphological characteristics of the isolates were similar to the taxonomic description of Fusarium equiseti (Leslie and Summerell 2006). Genomic DNA was extracted from fresh mycelium using DNeasy Plant Mini kit (Qiagen, USA). To confirm the identity of the fungus, two sets of primers, ITS4/ITS5 (White et al. 1990) and TEF1-α, EF1-728F/EF1-986R (Carbone and Kohn 1999) were used to amplify complete internal transcribed spacer (ITS) and partial translation elongation factor 1-alpha (TEF1-α) genes, respectively. BLASTn search in the NCBI database using ITS and TEF-1α sequences revealed 99 to 100% similarities with species of both F. incarnatum and F. equiseti. BLAST analysis of these in FUSARIUM-ID database showed 100% and 99% similarity with Fusarium incarnatum-F. equiseti species complex (FIESC) (NRRL34059 [EF-1α] and NRRL43619 [ITS]) respectively (Geiser et al. 2004). The ITS and TEF1-α sequences were deposited in GenBank (MT515832 and MT550682). The isolate was identified as F. equiseti, which belongs to the FIESC based on morphological and molecular characteristics. Pathogenicity was conducted on five healthy leaves of 1-month-old rockmelon cv. Golden Champion grown in 5 plastic pots filled with sterile peat moss. The leaves were surface-sterilized with 70% ethanol and rinsed twice with sterile-distilled water. Then, the leaves were wounded using 34-mm-diameter florist pin frog and inoculated by pipetting 20-μl conidial suspension (1 × 106 conidia/ml) of 7-day-old culture of isolate B onto the wound sites. Control leaves were inoculated with sterile-distilled water only. The inoculated plants were covered with plastic bags for 5 days and maintained in a greenhouse at 25 °C, 90% relative humidity with a photoperiod of 12-h. After 7 days, inoculated leaves developed necrotic lesions similar to the symptoms observed in the field while the control treatment remained asymptomatic. The fungus was reisolated from the infected leaves and was morphologically identical to the original isolate. F. equiseti was previously reported causing fruit rot of watermelon in Georgia (Li and Ji 2015) and China (Li et al. 2018). This pathogen could cause serious damage to established rockmelon as it can spread rapidly in the field. To our knowledge, this is the first report of a member of the Fusarium incarnatum-F.equiseti species complex causing leaf spot on Cucumis melo in Malaysia.

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