Molecular diagnosis of Maize dwarf mosaic virus (MDMV) and bio-control with food supplements and Bacillus subtilis on several maize genotypes

A potyvirus (proposed name of Zea mosaic virus [ZeMV]) isolated from maize in Israel was analyzed by serology, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of capsid proteins, symptomatology, and sequencing. Parts of the nuclear inclusion b, coat protein, and 3' regions were sequenced; the amino acid sequence of ZeMV capsid was determined by time-of-flight mass spectrometry (TOFMS). The results of these analyses were compared with those of similar analyses of the following potyviruses: Maize dwarf mosaic virus (MDMV), Sugarcane mosaic virus strain MDB (SCMV-MDB), Johnsongrass mosaic virus(JGMV), Sorghum mosaic virus (SrMV), and an isolate of MDMV from Israel. Indirect enzyme-linked immunosorbent assay using ZeMV antiserum detected only ZeMV, and reciprocal tests using MDMV, JGMV, or SrMV antisera failed to detect ZeMV. ZeMV cross-reacted weakly when SCMV-MDB antiserum was used. The mass of ZeMV capsid was determined to be 36,810 Da by SDS-PAGE and 34,216 Da by TOFMS. The ZeMV systemically infected johnsongrass (Sorghum halepense), but did not infect oat (Avena sativa), pearl millet (Pennisetum glaucum), barley (Hordeum vulgare), or rye (Secale cereale). Necrosis was caused in 19 sorghum lines by SrMV, in 15 by ZeMV, in 14 by MDMV, and in 5 by JGMV and SCMV-MDB. The nucleic acid and amino acid sequences of ZeMV clearly showed that it is not a strain of JGMV, MDMV, SCMV, or SrMV.

Sugarcane mosaic disease caused by sugarcane mosaic virus (SCMV), Johnsongrass mosaic virus (JGMV), maize dwarf mosaic virus (MDMV) and sorghum mosaic Virus (SrMV) is an economically important viral disease of sugarcane worldwide. Field survey was conducted to assess the presence of the viruses involve in mosaic disease of sugarcane in Makarfi Local Government Areas of Kaduna State (Northern Guinea Savannah), Nigeria. A range of symptoms were observed on the  infected land races from the pale green stripes to yellow chlorotic stripes on a dark green background. The purple land race (“Bakarkwama”) was highly susceptible followed by green land race (“Bahausa”) and the least infected was the white land race (“fararkwama”). 63 symptomatic and asymptomatic sugarcane leaves and stem juice extract from 14 villages of Makarfi L.G.A. were screened for the four viruses using DAS and TAS ELISA methods. SCMV, MDMV and SrMV were detected from both symptomatic and asymptomatic sugarcane leaf samples whereas JGMV was not detected in the locations sampled. SCMV isolate has the highest incidence (83%) from all the locations followed by SrMV (10%) and MDMV (5%) isolates. Mixed infections of the three viruses were also detected in some samples. This is the first report of identification of virus isolates inducing sugarcane disease in one of the major sugarcane producing areas of the Northern Guinea Savannah of Nigeria.   Key words: Sugarcane, SCMV, MDMV, SrMV, TAS-ELISA, DAS-ELISA.

SUMMARYIn 1977 a sudden and dramatic epidemic of maize dwarf mosaic virus (MDMV) struck commercial fields of sweet corn in Minnesota causing multi‐million dollar losses. The epidemic was unusual in that MDMV historically had been confined to the southern United States and Ohio River Valley with only occasional occurrences reported from states bordering Canada. An extensive 5‐yr study of MDMV in Minnesota revealed a low incidence of MDMV from 1978 ‐ 81, and commercial fields badly infected in 1977 were not again infected. No evidence of wild host plants was found, neither was seed transmission in maize considered to be important, and MDMV strain ratios changed from year to year as did the distribution of infected fields. Concomitant studies on aphid retention revealed that MDMV could be retained by aphid vectors for more than 19 h, and that the 1977 epidemic was associated with a weather pattern that could be linked to potential aphid transport from the southern Great Plains of North America. All circumstantial evidence led to a long‐distance aphid transport hypothesis. This may be worth considering whenever unexpected epidemics of aphid‐transmitted non‐persistent viruses occur in regions where the particular virus is not endemic, and may also explain the widespread distribution of certain aphid‐transmitted non‐persistent viruses.

In February 2014, a severe disease on maize (Zea mays L.) broke out in the fields of central and southwestern Taiwan and caused yield losses in sweet corn production. Chlorotic spots first appeared at the base of infected leaves and later developed into systemic mottling. Diffused necrotic patches were also found on leaves or husks of the diseased plants. Moreover, severe rosetting and stunting accompanied by abnormalities in ear production were observed on mature plants. Eighteen leaf samples from symptomatic plants were collected and submitted to our Plant Diagnostic Clinic for virus diagnosis. All of the samples were first tested by reverse transcriptase (RT)-PCR to detect Maize stripe virus (MSpV) and by indirect ELISA to detect Maize dwarf mosaic virus (MDMV) or Sugarcane mosaic virus (SCMV), which were endemic to this area (1). Only 2 out of 18 samples were positive for MDMV, SCMV, or mixed infection of both viruses. Sap inoculation tests conducted on seedlings of sweet corn cv. Honey 236 indicated that the MDMV- and SCMV-negative samples still had an unknown pathogen causing original symptoms in the receptor plants. The isolate from Yunlin county reacted only with the antibody to Maize chlorotic mottle virus (MCMV) (AC Diagnostics, Fayetteville, AR) in ELISA. For further identification, the MCMV-specific primers (forward: MCMVg3514F-GGGAACAACCTGCTCCA; reverse MCMVg4014R-GGACACGGAGTACGAGA) were designed from the nucleotide sequence of MCMV coat protein (CP) gene. In RT-PCR using the AccuPower RT/PCR PreMix kit (Bioneer, Daejeon, Korea), an expected 500-bp DNA fragment was observed. This PCR product was cloned and its nucleotide sequence was determined by Mission Biotech Co., Taipei, Taiwan. BLAST analysis of the CP gene of the MCMV-Yunlin revealed the maximum nucleotide identities (99%) with Chinese Sichuan isolates (GenBank Accession No. JQ984270) and 98% identities to four Chinese Yunnan isolates (GU138674, JQ982468, JQ982469, and KF010583) and one Kenya isolate (JX286709), compared with 97% to Kansas isolate (X14736) and 96% to Nebraska isolate (EU358605). Subsequently, the complete nucleotide sequence of the viral genome (KJ782300) was determined from five overlapping DNA fragments obtained from independent RT-PCR amplification. The virus isolate was infectious to sweet corn cultivars Bai-long-wang, Devotion, SC-34, SC2015, and Zheng-zi-mi, on which similar symptoms were developed after mechanical inoculation. During the spring of 2014, a total of 224 sweet corn samples were collected from the epidemic areas of Taichung, Yunlin, Chiayi, and Kaohsiung counties. Samples (n= 161) reacted positive for MCMV in ELISA and/or RT-PCR. In the field survey, more than 20 adult thrips might be observed on an MCMV-infected plant. Two species of Frankliniella were found on maize plants: F. williamsi Hood and F. intonsa Trybom. Maize thrips (F. williamsi), an occasional pest of maize occurring during winter and spring in Taiwan, was characterized by its abdominal sternite II on which 1 or 2 discal setae of equal length with posteromarginal setae were borne (2). Samples with 1, 5, 10, and 30 F. williamsi collected in the field were tested by RT-PCR; MCMV was detectable not only in the pooled crushed bodies but also in a single maize thrips. This is the first report of MCMV occurrence on maize in Taiwan and of the virus transmitted by maize thrips.

Maize dwarf mosaic (MDM) is one of the most important virus diseases of maize worldwide. Caused by the potyviruses maize dwarf mosaic virus (MDMV) or sugarcane mosaic virus (SCMV), MDM can cause up to 90% yield loss in susceptible hybrids. One of the most effective management strategies for MDM is growing potyvirus-resistant corn varieties. However, yield impacts associated with MDM and the corresponding efficacy of genetic resistance present in modern United States commercial hybrid lines is uncharacterized. In this study, we evaluated the disease response of 78 commercial hybrids to MDMV and SCMV and quantified yield losses associated with infection over multiple trials. We determined that while 97% of the hybrids tested were resistant to MDMV, 100% were susceptible to SCMV, with mean disease incidence per line averaging between 45 and 78% across six trial years. Despite only one hybrid displaying visible mosaic symptoms when inoculated with MDMV, MDMV reduced average yields by approximately 5% across all hybrids compared with the mock-inoculated treatment. The yield impact of SCMV was more severe, reducing average yields by 10% across replicated experiments. These results indicate that while most commercial hybrids are resistant to MDMV, possibly due to the presence of the major Scmv1 resistance locus on chromosome 6, additional potyvirus resistance genes are needed to manage SCMV-induced MDM. Pyramiding resistance loci, such as Scmv2 on chromosome 3 or Scmv3 on chromosome 10 in addition to Scmv1, could be an effective strategy to mitigate the yield impact of MDM disease.

Coldberg, K.-B.. and Brakke, M. K. 1987. Concentration of maize chlerotic mottie virus increased in mixed infections with maize dwarf mosaic virus, strain B. Phytopathology 77: 162-167. The concentration of rnaize chlorotic mottle virus (MCMV) was up to chlorophyIl and a lower than normal ratio of chloroplast to cytoplasmic 5.4 times higher in plants infected with both MCMV and maize dwarf rRNA. Purified MCMVhadanextinctioncoefficient of6.7cm2mg' at260 mosaic virus. strain R (MDMV-BE. rhan in plants infected with MCMY nm, an absorption maximum at 258 nm, minimum at 240 nm. and 25% only. The concentration of MDMV-B was the same in doubly and singly RNA. infected plants. Plants infected with both viruses had a reduced level of

Maize dwarf mosaic is the most widespread virus disease affecting corn production in Hungary. In attempts to identify the causal virus by test plant reactions, enzyme‐linked immunosorbent assay (ELISA) and polymerase chain reaction (PCR), only Maize dwarf mosaic virus (MDMV) was detected. To further characterize Hungarian isolates of MDMV, one isolate from each of the sweet corn varieties Dallas, Royalty and GH23‐85 was selected for sequence analysis of its coat protein (CP) gene. The three Hungarian isolates shared CP amino acid sequence similarities of 95–98% not only with one another but also with MDMV isolates from other countries. However, the N‐terminus of the CP of the ‘Dallas’ isolate was unusual in containing a stretch of 13 additional amino acids. This is the first report of variation in the size of the N‐terminus of the MDMV CP.

Sorghum (Sorghum bicolor (L.) Moench that is infected by maize dwarf mosaic virus (MDMV) expresses different symptoms depending on plant genotype and temperature. Infection causes a yellow to dark green pattern (mosaic), and a necrotic reaction (red-leaf) develops on certain genotypes when the temperature drops below approximately 21 ‘C (69°F). Hybrids expressing the red-leaf symptom show g-eater reduction in yield. Immunity to MDMV is not present in commercially available hybrid sorghums.

JMS stylet oil was evaluated for maize dwarf mosaic virus (MDMV) control in Ohio sweet corn. In laboratory tests, oil sprays reduced transmission of MDMV by single green bugs, Schizaphis graminum (Rondani), from 33% in untreated controls to 3% when virus source plants were sprayed, 6% when test plants were sprayed, and 0% when both source and test plants were sprayed. Alate corn leaf aphids, Rhopalosiphum maidis (Fitch), took longer to initiate probes on oil-sprayed than on nonsprayed corn leaves. In 1978, oil failed to control MDMV spread in a field experiment with a high inoculum level (33%). In a 1979 field test with a 1% inoculum level, maize dwarf mosaic (MDM) incidence was 3.2% in a plot receiving weekly oil sprays and 57.9% in an unsprayed control. In 1980, also with a 1% inoculum level, plots receiving weekly and biweekly oil sprays had significantly less MDM spread (19.3 and 14.2%, respectively) than did nontreated controls (25.7%). A simulation experiment indicated that rainfall was not the reason for the poor performance of oil in the field. Until formulations or application techniques are improved, oil sprays are unlikely to control the spread of MDMV in Ohio sweet corn.

Improvement of resistance to maize dwarf mosaic virus mediated by transgenic RNA interference

Maize dwarf mosaic is the most widespread virus disease affecting corn production in Hungary and Bulgaria. Samples from virus infected maize were collected from different part of Bulgaria and employed test plants, ELISA serological method and RT-PCR in order to identify the viral pathogen. Maize dwarf mosaic virus (MDMV) was detected in all tested samples. For further investigation three MDMV isolates were selected and cloned. Cloned cDNAs representing the coat protein gene of the virus have been sequenced. The coat protein genes of Bulgarian and Hungarian isolates of MDMV were compared. The nucleotide sequence identity and amino acid sequence similarity of the coat protein region varied from 88% to 99.1% and from 95.1% to 99.6%, respectively. The N-terminal region of coat protein was compared with other members SCMV subgroup and phylogenetic tree was constructed.

Eastern gamagrass (EGG) (Tripsacum dactyloides) is a high-quality, high-production native forage grass with potential as a high-protein, perennial grain crop. We used enzyme-linked immunosorbent assay to test EGG samples for virus infection. Over a 2-yr period, we found five instances of maize dwarf mosaic virus (MDMV) and a high incidence (12 and 72% in 3-mo-old and 5-yr-old plantings, respectively) of sugarcane mosaic virus strain MDMV-B (SCMV-MDMV-B) in experimental breeding nurseries in Kansas. Incidence of SCMV-MDMV-B was higher in older breeding nurseries. SCMV-MDMV-B, but not MDMV, was found in three of 42 natural EGG stands (.)

This study aimed to detect the harmful effects of Maize dwarf mosaic virus (MDMV) infection, and to demonstrate the potential benefits of S-methylmethionine-salicylate (MMS) pretreatment in infected maize (Zea mays L.) plants. The results of chlorophyll a fluorescence measurements showed that in MDMV-infected plants additional quenchers of fluorescence appear, probably as the result of associations between the virus coat protein and thylakoid membranes. It is important to note that when infected plants were pretreated with MMS, such associations were not formed. MDMV infection and MMS pretreatment resulted in a decrease in ascorbate peroxidase (APX) activity in maize leaves, while infection contributed to an increase in activity in the roots. Infection raised the guaiacol peroxidase (GPX) enzyme activity level, which was reduced by MMS pretreatment. MMS contributed to a decrease in both the RNA and coat protein content of MDMV, to an equal extent in maize leaves and roots. The results showed that MMS pretreatment enhanced the stress response reactions against MDMV infection in maize plants and retarded the spreading of infection.

Maize dwarf mosaic virus: a new causal agent inducing disease in rice plants of the fields of Kurdistan Region of Iraq

The beneficial and protective effects of S-methylmethionine (SMM) were investigated in Maize dwarf mosaic virus (MDMV) infected maize (Zea mays L.). Response reactions and alterations in the physiological state of the plants were monitored by following changes in the values of chlorophyll a fluorescence and chlorophyll content, and differences in the expression patterns of the stress-related genes S-adenosylmethionine synthase (SAMS) and the 14-3-3-like protein gene G-box factor 14-6 (GF14-6). Infection was validated using the ELISA technique. A pronounced decrease in the red to far-red chlorophyll a fluorescence ratio, indicative of chlorophyll content, was observed in infected plants, which was tempered by SMM pretreatment. A noticeable decrease in the photochemical quenching of photosystem II and the thermal dissipation of the antennae was observed, together with a notable increase in other non-photochemical energy dissipation parameters in response to MDMV infection. SMM treatment enhanced blue fluorescence in both uninfected and MDMV-infected plants (probably due to the production of protective phenolic compounds), while infection characteristically increased green fluorescence emission. SMM treatment was found to elevate the rate of gene expressions of SAMS and GF14-6. The results suggest that SMM pretreatment enhances the stress response reactions that protect maize plants against MDMV infection.