Seed-borne Virus Research Articles

Development of simplex and quintuplex RT-PCR for simultaneous detection of soybean viruses

Five simplex and a multiplex-RT-PCR (m-RT-PCR) protocols were developed for detection and differentiation of bean pod mottle virus (BPMV), cherry leaf roll virus (CLRV), raspberry ringspot virus (RpRSV), soybean mosaic virus (SMV) and tomato ringspot virus (ToRSV) infecting soybean. The simplex RT-PCR protocols produced virus-specific amplicons of 538 bp for BPMV, 139 bp for CLRV, 298 bp for RpRSV, 403 bp for SMV, and 282 bp for ToRSV, with sensitivity down to 10−4 diluted cDNA. Further, to detect all the five viruses simultaneously in a single tube a quintuplex RT-PCR protocol was optimized with as low as 10−3 diluted cDNA and 0.05 µM primer. To validate the reliability of the simplex RT-PCR protocol, imported soybean samples were tested by ELISA as well as RT-PCR. The results revealed that the developed protocol could detect the viruses in imported soybean, and found to be efficient than ELISA in resolving ambiguity in detection of seed borne viruses. The developed simplex and quintuplex RT-PCR protocol will be quite helpful for the diagnosis of soybean germplasm co-infected with viruses during the quarantine processing for ensuring virus free long term seed conservation in the National Gene Bank as well as for quarantine certification.

Australian Cool-Season Pulse Seed-Borne Virus Research: 1. Alfalfa and Cucumber Mosaic Viruses and Less Important Viruses.

Here, we review the research undertaken since the 1950s in Australia's grain cropping regions on seed-borne virus diseases of cool-season pulses caused by alfalfa mosaic virus (AMV) and cucumber mosaic virus (CMV). We present brief background information about the continent's pulse industry, virus epidemiology, management principles and future threats to virus disease management. We then take a historical approach towards all past investigations with these two seed-borne pulse viruses in the principal cool-season pulse crops grown: chickpea, faba bean, field pea, lentil, narrow-leafed lupin and white lupin. With each pathosystem, the main focus is on its biology, epidemiology and management, placing particular emphasis on describing field and glasshouse experimentation that enabled the development of effective phytosanitary, cultural and host resistance control strategies. Past Australian cool-season pulse investigations with AMV and CMV in the less commonly grown species (vetches, narbon bean, fenugreek, yellow and pearl lupin, grass pea and other Lathyrus species) and those with the five less important seed-borne pulse viruses also present (broad bean stain virus, broad bean true mosaic virus, broad bean wilt virus, cowpea mild mottle virus and peanut mottle virus) are also summarized. The need for future research is emphasized, and recommendations are made regarding what is required.

The Effects of Some Different Cultural Techniques on the Transmission and Infectious Development of Pepper Yellow Leaf Curl Indonesia Virus on Red Chili

<p>An experiment is conducted to investigate the effects of cultural techniques on pepper yellow leaf curl disease caused by <em>Pepper yellow leaf curl Indonesia virus</em> (PepYLCIV). The investigation is conducted in the area where the disease has been endemic and <em>Bemisia tabaci</em> is abundant. Four cultural techniques are applied in separate lands and cannot interfere with each other. The methods applied are seed treatment, intercropping, trap cropping, and physical barrier. Seeds harvested from infected plants are used for seed treatment experiments, and local farmers use commercial sources for other experiments. The results confirmed that PepYLCIV was a seed-borne virus affected by hot water treatment at 65<sup>o</sup>C for 30 minutes. Turmeric crude extract could reduce the incidence and severity of the disease. The tomato is a better intercrop than eggplant, mung bean, and soybean in reducing disease incidence, but their effects on disease severity and yield reduction were not significantly different. Basil and marigolds were better barrier crops compared to cosmos and zinnia. A 125 cm high physical barrier using 50 mesh cheesecloth could reduce the disease incidence, but not the lower ones. Under different cultural techniques, PepYLCIV causes a 40.00–52.32% chili yield reduction.</p>

First Report of Cucumber Green Mottle Mosaic Virus Infecting Zucchini (Cucurbita pepo) and Wax Gourd (Benincasa hispida) in China.

Cucumber green mottle mosaic virus (CGMMV) was first discovered in China in 2003 and caused an epidemic in 2005. In China, the virus has been reported in gourd crops including watermelons, cucumbers, melons, etc (Sui et al. 2019). In Shandong Province, China from September 2014 to 2017, approximately 30% of zucchini (Cucurbita pepo) and wax gourd (Benincasa hispida) plants in commercial cucurbit fields, the two most important cash crops, exhibited chlorosis, mosaic, and mottling symptoms suspected to be caused by a tobamovirus. To identify the causative pathogens, ten zucchini and 15 wax gourd samples were collected from the commercial cucurbit fields. Total RNA was extracted and all samples were tested using reverse transcription PCR (RT-PCR) with TobamodF/TobamodR and TobamodF2/TobamodR2 (Li et al. 2018a). Five common Cucurbitaceae viruses were also tested: cucumber mosaic virus, papaya ringspot virus, squash mosaic virus, watermelon mosaic virus, and zucchini yellow mosaic virus (Ali et al. 2012). All samples generated positive results using tobamovirus generic primers but were negative for the five common Cucurbitaceae viruses. Amplification products (880 bp) from all samples were inserted into pMD19-T and recombinant clones were selected for Sanger sequencing. The results showed that zucchini green mottle mosaic virus, CGMMV, and tobacco mosaic virus (TMV) were detected in zucchini samples. CGMMV and TMV were detected in the wax gourd samples. To confirm the presence of these viruses, RT-PCR was performed using specific primer pairs, including CGMMV-cpf/CGMMV-cpr (Chen et al. 2006), ZG-F/ZG-R (Li et al. 2018b), and TMV-CP-F/TMV-CP-R (Srivastava et al. 2015). CGMMV was detected in all samples, with four zucchini and nine wax gourds only containing CGMMV. Zucchini (n=4; CGZ1-CGZ4) and wax gourd (n=4; CGWX1-CGWX4) isolates were cloned into pMD19-T and sequenced bidirectionally. The BLASTn results confirmed the presence of CGMMV, and the sequencing results were processed using DNAMAN Version (Lynnon Biosoft, San Ramon, CA, USA) and submitted to the GenBank database (https://www.ncbi.nlm.nih.gov/). A phylogenetic tree based on the CGMMV coat protein (CP) was constructed using CGZ1-CGZ4 (OP779762-OP779765), CGWX1-CGWX4 (OP779766-OP779769), and representative CGMMV sequences from GenBank. Sequence analysis of the CP demonstrated that CGMMV-zucchini and -wax gourd isolates belonged to an independent branch of the Chinese muskmelon AH-FT197 isolate (KU175639) and had 100% identity with the AH-FT197 isolate. To confirm their infectivity, leaf sap extract of CGZ4 and CGWX4 in phosphate buffer (0.1 M, pH 7.0) was mechanically inoculated on leaves of virus-free zucchini seedlings (Cucurbita pepo cv. Zaoqingyidai, 4-leaf-stage, n = 10) or virus-free wax gourd seedlings (Benincasa hispida cv. Tiezhu 2, n = 10). Ten days after inoculation, all plants exhibited symptoms (systemic chlorosis, mosaic, and mottling) similar to those of diseased plants in the field. Control seedlings inoculated with phosphate buffer remained symptomless. RT-PCR analysis using the CGMMV-cpf/CGMMV-cpr primer confirmed that all ten zucchini or wax gourd seedlings were infected with CGMMV, and all the control plants were free from CGMMV. To the best of our knowledge, this is the first report on zucchini and wax gourd as natural hosts for CGMMV in China. CGMMV is a highly contagious seed-borne virus and further attention should be paid to its spread in cucurbit crops.

Kyuri green mottle mosaic virus detected for the first time in Turkey

Turkey is among the top 10 producers of cucumber, melon, watermelon, and squash in the world. Lately, seed-borne viruses have become a major issue in greenhouse and field-grown cucurbits. In this study, the incidence of kyuri green mottle mosaic virus (KGMMV) was determined in seeds from various species (cucumber, melon, watermelon, summer squash, bottle gourd, winter squash) of Cucurbitaceae. KGMMV detection in a total of 20 seed lots of each cucurbit species was done by enzyme-linked immunosorbent assay (ELISA) and RT-PCR. The highest virus incidence was 45% in melon followed by 25% in cucumber and 10% in squash. To the best of our knowledge, this is the first report of KGMMV in cucurbit crops in Turkey and this highlights the potential risk of KGMMV to commercial cucurbit seed lots.

Characterization of the seed virome of alfalfa (Medicago sativa L)

BackgroundSeed transmission of plant viruses can be important due to the role it plays in their dissemination to new areas and subsequent epidemics. Seed transmission largely depends on the ability of a virus to replicate in reproductive tissues and survive during the seed maturation process. It occurs through the infected embryo or mechanically through the contaminated seed coat. Alfalfa (Medicago sativa L.) is an important legume forage crop worldwide, and except for a few individual seedborne viruses infecting the crop, its seed virome is poorly known. The goal of this research was to perform initial seed screenings on alfalfa germplasm accessions maintained by the USDA ARS National Plant Germplasm System in order to identify pathogenic viruses and understand their potential for dissemination.MethodsFor the detection of viruses, we used high throughput sequencing combined with bioinformatic tools and reverse transcription-polymerase chain reactions.ResultsOur results suggest that, in addition to common viruses, alfalfa seeds are infected by other potentially pathogenic viral species that could be vertically transmitted to offspring.ConclusionsTo the best of our knowledge, this is the first study of the alfalfa seed virome carried out by HTS technology. This initial screening of alfalfa germplasm accessions maintained by the NPGS showed that the crop’s mature seeds contain a broad range of viruses, some of which were not previously considered to be seed-transmitted. The information gathered will be used to update germplasm distribution policies and to make decisions on the safety of distributing germplasm based on viral presence.

Seed Transmission of Begomoviruses: A Potential Threat for Bitter Gourd Cultivation.

Bitter gourd (Momordica charantia L.), one of the valued vegetable crops in India, is severely affected by yellow mosaic disease caused by two begomoviruses, tomato leaf curl New Delhi virus (ToLCNDV) and bitter gourd yellow mosaic virus (BgYMV). The symptoms are yellowing, distortion of leaf, puckering, and malformed fruits. Increased incidence of the disease and appearance of symptoms even in young emerging seedling stage were suggestive of seed transmission of the viruses, which was examined in detail. To study the seed transmission, two sources-seeds of elite hybrids H1, H2, H3, H4, and Co1 procured from a seed market; and seeds from infected plants in the farmer's field were tested. Detection of the virus by DAS-ELISA using polyclonal antibody indicated embryo infection up to 63%, 26%, 20%, and 10% in hybrids H1, H2, H3, and H4, respectively, for market-procured seeds. In PCR analysis with primers specific for ToLCNDV and BgYMV, infection by ToLCNDV was as high as 76% and mixed infection was 24%. In contrast, in seeds derived from field-infected plants, the percentage detection was less. Grow-out tests with market-procured seeds revealed no transmission for BgYMV compared with 5% transmission for ToLCNDV. Whether seed-borne inocula could serve as an inoculum for new infection in a field and further progress of the disease was investigated in a microplot study. The study clearly revealed variation in seed transmission between different sources, lots, cultivars, and viruses. The virus present in symptomatic and asymptomatic plants was easily transmitted by whitefly. In another microplot experiment, the potential of seed-borne virus as inoculum was proved. There was 43.3% initial seed transmission in the microplot, increasing to 70% after release of 60 whiteflies.

Development of a reverse-transcription droplet digital PCR method for quantitative detection of Cucumber green mottle mosaic virus

Cucumber green mottle mosaic virus (CGMMV) is a re-emerging threat to the production of greenhouse cucumber and other Cucurbitaceae crops worldwide. This seed-borne virus can easily spread from a contaminated seed to seedlings and adjacent plants by mechanical contact between the foliage of diseased and healthy plants, causing extensive yield losses. An accurate method for detecting and quantifying this virus is urgently needed to ensure the safety of the global seed trade. Here, we report the development of a reverse-transcription droplet digital polymerase chain reaction (RT-ddPCR)-based method for specific and high-sensitive detection of CGMMV. By testing three primer–probe sets and optimizing reaction conditions, we showed that the newly developed RT-ddPCR method is highly specific and sensitive, with a detection limit of 1 fg/μL (0.39 copy/μL). The sensitivity of the RT-ddPCR method was compared with that of real-time fluorescence quantitative RT-PCR (RT-qPCR) using a series of plasmid dilutions and total RNAs extracted from infected cucumber seeds, and the detection limit of RT-ddPCR was 10 times higher than RT-qPCR with plasmid dilutions and 100 times higher than RT-qPCR for detecting CGMMV from infected cucumber seeds. The RT-ddPCR method was further assessed for detecting CGMMV from a total of 323 samples of Cucurbitaceae seeds, seedlings, and fruits as compared with the RT-qPCR method. We found that the infection rate of CGMMV on symptomatic fruits was as high as 100%, whereas infection rates were lower for seeds and lowest for seedlings. Notably, the results of two methods in detecting CGMMV from different cucurbit tissues showed the high consistency with Kappa value from 0.84 to 1.0, demonstrating that the newly developed RT-ddPCR method is highly reliable and practically useful for large-scale CGMMV detection and quantification.

DETEKSI BEGOMOVIRUS PADA BENIH PEPAYA DAN PENGENDALIANNYA DENGAN METODE HOT WATER TREATMENT

Begomovirus is a virus that has been reported as the cause of yellow leaf curl disease on papaya plants in Bengkulu. Incidence of yellow mosaic disease on papaya plants in Bengkulu up to 100% in papaya cultivation centers. So that it strengthens the suspicion of the potential for seed transmission Begomovirus in papaya plants. Seed-borne viruses by Begomovirus are important because : can be a source of initial inoculum in the planted area. Then the control of the virus is carried by the seeds. It is important to suppress the initial source of inoculum in the field. This research aims to Begomovirus detection in papaya seeds was carried out by PCR method using SPG1 . Primer and SPG 2, and control of virus in seeds by controlling Hot Water Treatment. Seed Papaya plants infected with Begomovirus were given hot water treatment which consisted of two treatments temperature is 56℃ for 60 minutes and 58℃ for 40 minutes in a water bath. positive control are seeds that were confirmed positive for begomovirus without Hot Water Treatment (HWT) while the negative control seeds were confirmed negative for begomovirus. The results of this study showed that Begomovirus was not detected at the age of 7 (primary leaves) days after sowing, however detected in plants aged 14 days after sowing (first leaf) using primers universal SPG 1 and SPG 2. After the papaya seeds were treated (HWT with a temperature of 56℃ for 60 minutes and 58℃ for 40 minutes showed negative results in plants aged 7 days after seedlings and 14 days after sowing. This shows that the HWT treatment is able to suppres Begomovirus in papaya seeds.

Actinidia seed-borne latent virus (ASbLV, Betaflexiviridae) is transmitted paternally and maternally at high rates

Actinidia seed-borne latent virus (ASbLV, Betaflexiviridae, genus Prunevirus) was detected at high frequency in healthy seedlings grown from lines of imported seed in a New Zealand post-entry quarantine facility. To determine the route and efficiency of transmission of ASbLV in this dioecious crop species, we developed a rapid molecular protocol and identified a reliable progeny plant tissue to determine paternal and maternal transmission rates. The virus was detected at a high incidence (98%) in individual seeds, but cotyledon testing of seedlings from selected crosses confirmed staminate (male) transmission at high frequency (~ 60%), and pistillate (female) transmission at even higher frequency (~ 80%). The use of cotyledons allows non-destructive detection of ASbLV in very young seedlings that enables early screening of kiwifruit plants in nurseries to manage its spread to orchards. The high ASbLV transmission rates, whether from infected pollen or ovules, facilitate bulk testing of seed lots that could quickly detect infected parent plants (fruit bearing female or male pollinator) already in an orchard. The dioecious nature of Actinidia may provide a useful biological tool to further investigate ASbLV movement, transmission biology, and ultimately its impact on infected Actinidia plants.

Economical and Epidemiological Consequences of Seed Transmitted Plant Viruses. Review

Plant virus diseases greatly influence man’s economy by reducing the yield and quality of plant products. About one-quarter of the known plant viruses are transmitted through the seed. Seed transmission of plant viruses is an important means for the introduction of plant viruses into new localities where they may become established, spread rapidly, and can cause epidemics in the presence of suitable vectors and host species. Viruses may persist in seed for long periods so that commercial distribution of a seed-borne virus over long distances may occur. The location of the virus in seed determines the transmissibility of viruses through seed. The virus is considered to be externally seed transmitted when it is outside the functional seed and internally seed transmitted when it is within the tissue of the seed. Seed infection by the virus is epidemiologically important for the reason that it is the primary source of inoculums and forms the starting point for the initiation of the disease. Avoidance of virus inoculum from infected seeds; chemical seed disinfection, implementing the cultural practices like field sanitation, rouging, and crop rotation can reduce the virus disease incidence. The role of quarantines, resistant cultivars, healthy seed production, and certification schemes for healthy seed production are also critical measures for seed transmitted virus management. Keywords: Inoculum source; Management; Plant virus; Seed transmission DOI: 10.7176/RHSS/12-7-03 Publication date: April 30 th 2022

Localization and Mechanical Transmission of Tomato Brown Rugose Fruit Virus in Tomato Seeds.

Tomato brown rugose fruit virus (ToBRFV), belonging to the genus Tobamovirus, is a highly virulent emerging virus, causing disease outbreaks and significant crop losses worldwide. The growing number of ToBRFV epidemic episodes prompted the investigation of the role of seeds in the dissemination of the virus as an important aspect in the overall disease management. Therefore, the objectives of this study were to determine the localization of ToBRFV within tomato seeds and to evaluate its seed transmission characteristics. Seeds extracted from naturally ToBRFV-infected tomato fruits were tested for the presence of the virus using serological, molecular, and biological assays. Three immunolocalization techniques were used to determine the localization and distribution of ToBRFV within the different tissues and parts of tomato seeds. To evaluate seed transmission of ToBRFV, two grow-out experiments were conducted to assess the rate of both vertical (seeds to progeny seedlings) and possible horizontal transmission (plant to plant) based on serological and molecular assays. Seeds extracted from ToBRFV-infected fruits had a 100% contamination rate. The localization of ToBRFV in tomato seeds is only external on the seed coat (testa). Seed transmission rate from seeds to their seedlings was very low (0.08%), while no transmission was recorded from plants to plants in a small-scale greenhouse experimental setup. In conclusion, ToBRFV is a seedborne virus located externally on tomato seed coat and transmitted mechanically from ToBRFV-contaminated tomato seeds to seedlings, which could initiate a disease foci and eventually drive further dissemination and spread of the disease in a new growing area.

Polycistronic Artificial microRNA-Mediated Resistance to Cucumber Green Mottle Mosaic Virus in Cucumber.

Cucumber green mottle mosaic virus (CGMMV), as a typical seed-borne virus, causes costly and devastating diseases in the vegetable trade worldwide. Genetic sources for resistance to CGMMV in cucurbits are limited, and environmentally safe approaches for curbing the accumulation and spread of seed-transmitted viruses and cultivating completely resistant plants are needed. Here, we describe the design and application of RNA interference-based technologies, containing artificial microRNA (amiRNA) and synthetic trans-acting small interfering RNA (syn-tasiRNA), against conserved regions of different strains of the CGMMV genome. We used a rapid transient sensor system to identify effective anti-CGMMV amiRNAs. A virus seed transmission assay was developed, showing that the externally added polycistronic amiRNA and syn-tasiRNA can successfully block the accumulation of CGMMV in cucumber, but different virulent strains exhibited distinct influences on the expression of amiRNA due to the activity of the RNA-silencing suppressor. We also established stable transgenic cucumber plants expressing polycistronic amiRNA, which conferred disease resistance against CGMMV, and no sequence mutation was observed in CGMMV. This study demonstrates that RNA interference-based technologies can effectively prevent the occurrence and accumulation of CGMMV. The results provide a basis to establish and fine-tune approaches to prevent and treat seed-based transmission viral infections.

A one-step reverse-transcription loop-mediated isothermal amplification assay optimized for the direct detection of cucumber green mottle mosaic virus in cucurbit seeds

Cucumber green mottle mosaic virus (CGMMV) is a seed-borne virus that causes significant economic losses in farms cultivating cucurbit plants. With the increase in global trade of cucurbit seeds, it is essential to develop a rapid, reliable, and convenient diagnostic method for the direct detection of CGMMV in these seeds for prevention and management of the disease. Here, we developed a one-step reverse-transcription loop-mediated isothermal amplification (RT-LAMP) assay for the direct detection of CGMMV in cucurbit seeds. To improve the efficiency of the one-step RT-LAMP assay, six primers were designed to target the most conserved regions of the gene encoding the movement protein of CGMMV. Our one-step RT-LAMP assay was optimized to improve specificity and sensitivity for CGMMV detection in individual seeds. A comparison of the detection sensitivity revealed that our one-step RT-LAMP assay was 100-fold more sensitive than the current reverse transcription-polymerase chain reaction assay used for CGMMV quarantine in Korea. Collectively, the one-step RT-LAMP assay developed in the present study is appropriate for the direct detection of CGMMV in individual cucurbit seeds.

Variation in concentration of Shallot virus X in shallot plants at different developmental stages and its implication in diagnosis

Shallot is one of the major herbs extensively being used in global cuisine. Shallot, like onion and garlic, is infected by various viruses. Shallot virus X (ShVX) infects shallot and cause significant yield loss. In the present study for the first time, effort has been made to examine the load and distribution of ShVX in various parts of shallot plants. Our study indicates that there is uneven distribution of the virus in the plant which vary with different tissues as well as the age of the plant. Accordingly, choice of tissue and age of the selected tissue was found to be a determining factor in indexing process. It was found that bulb is the most suitable tissue for detection of ShVX followed by young leaf. This study also for the first time proved that ShVX is not a seed borne virus.

Recent emergence of seed-borne viruses and viroids on tomato, seed health tests and their implications in global seed trade

ISHS VI International Symposium on Tomato Diseases: Managing Tomato Diseases in the Face of Globalization and Climate Change Recent emergence of seed-borne viruses and viroids on tomato, seed health tests and their implications in global seed trade

Exploring New Sources of Viral Resistance in Cowpea Germplasm and Its Elaboration through Multivariate Analysis

Background: Cowpea is a major food legume rich in protein but its production has been dwindling by several factors including viral infection due to various virus strains in all agro-ecological zones.Methods: Sixty eight cowpea genotypes were screened against qualitative traits (leaf shape, seed surface, twinning tendency, anthocynin pigment, plant type, fodder type and cream color) and four seed borne viruses viz. cucumber mosaic virus (CMV), cowpea aphid borne mosaic virus (CABMV), black eye cowpea mosaic virus (BlCMV) and bean common mosaic virus (BCMV) under both in situ and laboratory conditions using DAC-ELISA.Result: Based on in situ screening, 16 genotypes were found resistant to all the four viruses, whereas for ELISA, 13 genotypes (27005, 27041, 27075, 27141, 27145, 27146, 27147, 27158, 27160, 27167, 27172, IT85F-1380 and IT86D-719) were found resistant to all. Twelve clusters were obtained from UPGMA based on disease severity. Genotype 27008 (Pakistan) was present in cluster VI and was susceptible to all antisera CMV, CABMV, BICMV and BCMV. Whereas 13 genotypes were present in cluster VIII which were found resistant to all the four antisera applied. Therefore, 13 genotypes suggested for safe use in any breeding program at developing resistant cultivars. First two factors obtained through PCA with eigen-values greater than 1 contributed greater than 80 per cent variability. Twelve distinct groups were observed and these were in coordination with cluster analysis.

Molecular Identification of Begomovirus Infecting Angled Luffa

Begomovirus was reported as one of the most aggressive and destructive viruses on several commercial crops, including cucurbits in Indonesia. Plants that infected with Begomovirus show the mosaic symptom on the leaves, change in leaf shape, stunts, change in color and shape of fruit. It was recently observed in cultivated angled luffa [Luffa acutangula (L.) Roxb] around Yogyakarta and Central Java. The aim of this research was to identify the virus by using Polymerase chain reaction (PCR). The result of Begomovirus amplification from the total DNA samples amplification using primer Krusty-Homer showed that DNA of Begomovirus from angled luffa was amplified at ~580bp. The DNA sequencing of angled luffa’s leaf isolate GD1 had 97.8% homology with SCLV-China isolate MC1. However, amplification of DNA seed samples using the same primer showed negative result. It was concluded that Begomovirus was not a seed borne virus. This is the first molecular report on the occurence of Begomovirus in angled luffa in Yogyakarta.

Preliminary Study on the Control of Cucumber Green Mottle Mosaic Virus in Commercial Greenhouses Using Agricultural Disinfectants and Resistant Cucumber Varieties

Cucumber green mottle mosaic virus (CGMMV) is a re-emerging threat to greenhouse cucumber and other Cucurbitaceae crop production worldwide. This seed-borne virus can easily spread from a contaminated seeds to seedlings and to adjacent plants through mechanical contact of the foliage of diseased and healthy plants causing extensive yield losses. Additionally, infection may not be limited to the current crop but may also affect subsequent crops due to the long-term persistence of the virus on contaminated crop residues, greenhouse hard surfaces and soil or soil-less greenhouse substrates. In the present work, three greenhouse experiments were conducted to develop an integrated pest management strategy towards controlling CGMMV in commercial cucumber greenhouses, by implementing an effective sanitization program and using resistant and grafted cucumber varieties. Results of sanitization highlighted that pressure washing and cleansing with an alkaline foam cleanser eliminated CGMMV on some of the most heavily infested areas. However, three successive applications of cleanser and disinfectants were essential to completely eliminate CGMMV on porous and uneven surfaces, such as cement alleyway, tray gutters and floor mats. The screening of 15 cucumber varieties revealed that one Mini (‘Khassib’) and three Long English (‘Sepire’, ‘Bomber’ and ‘LC13900′) had reduced or delayed CGMMV infection spread in the greenhouse but were intermediate in yield. The most resistant Mini variety was ‘Katrina’. This variety showed low CGMMV infection level and high fruit yield. The varieties ‘Jawell’, ‘RZ 22-551′, ‘Sunniwell’, ‘Bonbon’ and ‘Dee Lite’ were the most tolerant to CGMMV. They showed a high CGMMV infection level without compromising yield. These results proved the need for new productive cucumber varieties with CGMMV resistance. Grafting experiments showed a yield increase only in the case of grafted ‘Picowell’ over ‘Bonbon’ but no CGMMV resistance, which is a much more desirable result of grafting experiments in order to have economic potential. In all, the current study revealed unique methods of CGMMV management in commercial greenhouses that are recommended to growers for reducing crop losses and improving economic returns.

Assessment of Yam mild mosaic virus coat protein gene sequence diversity reveals the prevalence of cosmopolitan and African group of isolates in Ghana and Nigeria

This study analyzed the genetic diversity of 18 Yam mild mosaic virus (YMMV, genus Potyvirus) isolates collected from field surveys in Ghana (N = 8) and Nigeria (N = 10) in 2012−13. The full coat protein (CP) encoding region of the virus genome was sequenced and used for comparison and phylogenetic analysis of the YMMV isolates available in the NCBI nucleotide database. The mean nucleotide (nt) diversity was 13.4% among the 18 isolates (17 from D. alata and one from D. rotundata), 11.4% within the isolates of Ghana and 7.4% within the isolates of Nigeria. The phylogenetic clustering of the 18 YMMV isolates did not show correlation with the country of origin, and they aligned with the reference sequences of four of the 11 YMMV monophyletic groups representing the cosmopolitan group and the African group of YMMV isolates. High sequence homology of 99% between the YMMV sequence from Nigeria (CP12-DaN6-1) and a previously reported sequence from Togo (GenBank Accession Number AF548514) suggests a prevalence of seed-borne virus spread within the region. Understanding YMMV sequence diversity in West Africa aid in the improvement of diagnostic assays necessary for virus indexing and seed certification.