Complete Genome Sequence of a Novel Potyvirus Infecting Miscanthus Sinensis (Silver Grass)

Here, we describe the full-length genome sequence of a novel potyvirus, tentatively named “miscanthus sinensis mosaic virus” (MsiMV), isolated from Miscanthus sinensis (silver grass) held in a post entry quarantine facility following its initial import into Western Australia, Australia. The MsiMV genome encompasses 9604 nucleotides (nt) encoding a 3071 amino acids (aa) polyprotein with conserved sequence motifs. The MsiMV genome is most closely related to sorghum mosaic virus (SrMV) with 74% nt and 78.5% aa sequence identity to the SrMV polyprotein region. Phylogenetic analysis based on the polyprotein grouped MsiMV with SrMV, sugarcane mosaic virus (SCMV) and maize dwarf mosaic virus (MDMV). This is the first report of a novel monopartite ssRNA virus in Miscanthus sinensis related to members of the genus Potyvirus in the family Potyviridae.

Identification and Severity Monitoring of Maize Dwarf Mosaic Virus Infection Based on Hyperspectral Measurements

Prompt monitoring of maize dwarf mosaic virus (MDMV) is critical for the prevention and control of disease and to ensure high crop yield and quality. Here, we first analyzed the spectral differences between MDMV-infected red leaves and healthy leaves and constructed a sensitive index (SI) for measurements. Next, based on the characteristic bands (Rλ) associated with leaf anthocyanins (Anth), we determined vegetation indices (VIs) commonly used in plant physiological and biochemical parameter inversion and established a vegetation index (VIc) by utilizing the combination of two arbitrary bands following the construction principles of NDVI, DVI, RVI, and SAVI. Furthermore, we developed classification models based on linear discriminant analysis (LDA) and support vector machine (SVM) in order to distinguish the red leaves from healthy leaves. Finally, we performed UR, MLR, PLSR, PCR, and SVM simulations on Anth based on Rλ, VIs, VIc, and Rλ + VIs + VIc and indirectly estimated the severity of MDMV infection based on the relationship between the reflection spectra and Anth. Distinct from those of the normal leaves, the spectra of red leaves showed strong reflectance characteristics at 640 nm, and SI increased with increasing Anth. Moreover, the accuracy of the two VIc-based classification models was 100%, which is significantly higher than that of the VIs and Rλ-based models. Among the Anth regression models, the accuracy of the MLR model based on Rλ + VIs + VIc was the highest (R2c = 0.85; R2v = 0.74). The developed models could accurately identify MDMV and estimate the severity of its infection, laying the theoretical foundation for large-scale remote sensing-based monitoring of this virus in the future.

First Report of Sugarcane Mosaic Virus in Zea mays L. in Ukraine

Maize viral diseases especially maize dwarf mosaic disease (MDMD), which is caused by potyviruses, lead to significant crop losses worldwide. Aim. The aim of this work was to identify the causal agent of mosaic symptoms, observed on maize plants during 2018—2020 in Kyiv region. Methods. Enzyme-linked immunosorbent assay in the DAS-ELISA modification using commercial Loewe Biochemica test systems for Maize dwarf mosaic virus (MDMV), Sugarcane mosaic virus (SCMV), Wheat streak mosaic virus (WSMV) were applied to identify the causal agent of maize disease in collected samples. Transmission electron microscopy was used in order to direct viral particle visualisation. Aphids, which are natural vectors of plant viruses, were found on diseased plants. Results. Plants with typical mosaic symptoms were observed in corn crops of the Kyiv region in early June 2018. The pathogen was transmitted by mechanical inoculation to maize and sweet maize plants with the manifestation of mosaic symptoms. Electron microscopy of the sap from diseased plants revealed the presence of flexible filamentous virions 750 nm long and 13 nm in diameter, typical for the genus Potyvirus. In August, mosaic symptoms and aphids Rhopalosiphum padi were found on previously healthy plants in the same maize crop. In 2020, in the same sown area, maize plants were free of viral infection during inspection in June, but a re-inspection in September revealed mosaic symptoms on maize crop and the presence of aphids in the leaf axils. The presence of SCMV in maize samples collected in June and August/September 2018 and 2020, as well as in inoculated maize and sweet maize plants, was confirmed by ELISA using a commercial test system. The obtained data allow suggesting that Rhopalosiphum padi is a natural vector of SCMV in agrocenoses of Ukraine. It should be noted that co-infection with MDMV and WSMV in the affected plants was not detected. Conclusions. This study presents the first report of SCMV in maize in Ukraine.

Evolutionary study of maize dwarf mosaic virus using nearly complete genome sequences acquired by next-generation sequencing

Next-generation sequencing is a robust approach to sequence plant virus genomes in a very short amount of time compared to traditional sequencing methods. Maize dwarf mosaic virus (MDMV) is one of the most important plant viruses worldwide and a significant threat to maize production. In this study, we sequenced 19 MDMV isolates (10 from Johnsongrass and 9 from maize) collected in Oklahoma and Missouri during 2017–2019 using Illumina sequencing and determined the genetic diversity. Sequence reads were assembled and 19 nearly complete genome sequences of MDMV isolates were obtained. Phylogenetic analysis based on complete genomes nucleotide and amino acid sequences revealed two main clusters and a close evolutionary relationship among 19 MDMV isolates. Statistical analysis of individual genes for site-specific selection revealed that all genes are under negative selection. The fixation index (FST) analysis of the MDMV isolates revealed no gene flow between the two main phylogenetic clusters, which emphasizes the divergence of MDMV isolates from the USA. Among the USA MDMV isolates, the mean genetic distance (d) and nucleotide diversity ((π) were highest in the P1 gene coding region. This is the first detailed study on the evolutionary relationship of MDMV isolates based on the nearly complete genome analysis from maize and Johnsongrass.

Simultaneous Gene Expression and Multi-Gene Silencing in Zea Mays Using Maize Dwarf Mosaic Virus

Background: Maize dwarf mosaic virus (MDMV), a member of the genus Potyvirus, infects maize and is non-persistently transmitted by aphids. Several plant viruses have been developed as tools for gene expression and gene silencing in plants. The capacity of MDMV for both gene expression and gene silencing were examined.Results: Infectious clones of an Ohio isolate of MDMV, MDMV-OH5, were obtained, and engineered for gene expression only, and for simultaneous marker gene expression and virus-induced gene silencing (VIGS) of three endogenous maize target genes. Single gene expression in single insertion constructs and simultaneous expression of green fluorescent protein (GFP) and silencing of three maize genes in a double insertion construct was demonstrated. Constructs with GFP inserted in the N-terminus of HCPro were more stable than those with insertion at the N-terminus of CP in our study. Unexpectedly, the construct with two insertion sites also retained insertions at a higher rate than single-insertion constructs. Engineered MDMV expression and VIGS constructs were transmissible by aphids (Rhopalosiphum padi). Conclusions: These results demonstrate that MDMV-based vector can be used as a tool for simultaneous gene expression and multi-gene silencing in maize.

Status of Maize Lethal Necrosis Disease in Zambia

Maize is a staple food in Zambia and contributes immensely to food security for smallholder farmers. Disease outbreaks such as Maize Lethal Necrosis Disease (MLND) can be a key constraint to maize production. This disease is caused by synergistic co-infection with Maize Chlorotic Mottle Virus (MCMV) and any virus from the family Potyviridae, particularly, Sugarcane Mosaic Virus (SCMV), Maize Dwarf Mosaic Virus (MDMV) or Wheat Streak Mosaic Virus (WSMV). In 2011, an outbreak of MLND affecting almost all of the currently grown commercial varieties posed a challenge to maize production in Kenya and it has since been reported in DR Congo, Ethiopia, Kenya, Rwanda, Tanzania, and Uganda causing yield losses of up to 100%. Despite MLND having been reported in some neighboring countries, there is no information on the status of the disease in Zambia. Additionally, there is a lot of grain and seed trade between Zambia and other countries among which MLND has been reported. The aim of this study was to establish: (a) the status of MCMV; (b) agricultural practices used by farmers and (c) insect vectors associated with MLND. A survey was conducted in nine (9) provinces of Zambia during 2014/2015 and 2015/ 2016 cropping seasons. Farmers’ maize fields were sampled at every five to ten-kilometer interval and tested using rapid diagnostic kits capable of detecting MCMV. Four hundred and nineteen samples collected all tested negative for MCMV. Zambian Agricultural Research Institute (ZARI), with all stakeholders in the maize value chain should continue implementing measures aimed at preventing the introduction of MLND in Zambia. Key words: Survey, MLND, Losses, food security

Host plant selection and virus transmission by Rhopalosiphum maidis are conditioned by potyvirus infection in Sorghum bicolor

Many plant viruses are significant pathogens that are able to utilize arthropod vectors to infect a vast range of host plants, resulting in serious economic damage to world food crops. One such crop is Sorghum bicolor, grain sorghum, which is the fifth most important global cereal crop, it is grown for human consumption, animal feed, and biofuel. In this study, the Potyviruses Johnsongrass mosaic virus (JGMV), Maize dwarf mosaic virus (MDMV), Sugarcane mosaic virus (SCMV), and Sorghum mosaic virus (SRMV) were tested for their rates of transmission into tissues of S. bicolor by the corn leaf aphid, Rhopalosiphum maidis. In addition, virus infected and non-infected S. bicolor plants were assessed for their effects on R. maidis host plant selection behavior. Further, the propagation of each virus (viral ssRNA copy number) in infected plants was determined using qPCR amplification of viral coating protein gene fragments. The mean rate of JGMV transmission into S. bicolor plants by R. maidis was significantly lower than transmission of MDMV, SCMV, and/or SRMV. Sorghum bicolor plants infected with MDMV, SCMV or SRMV also attract significantly more R. maidis than non-infected plants. JGMV-infected plants do not effect a similar change in R. maidis plant choice preference. The preference of non-viruliferous R. maidis toward S. bicolor plants infected with MDMV, SCMV or SRMV, and lack of such attraction by JGMV-infected plants may play a role in virus transmission strategy and efficiency by the vector.

Complete Genome Characterization and Coat Protein Genealogy of Isolates of Maize dwarf mosaic virus from Johnsongrass and Maize in Oklahoma and Missouri

Maize dwarf mosaic virus (MDMV) significantly affects maize production worldwide, including the United States. This study describes the distribution and biological and molecular characterization of MDMV isolates from Johnsongrass and maize. A total of 262 samples (symptomatic = 214, asymptomatic = 48) were collected in Oklahoma and Missouri during 2016, 2017, and 2019 growing seasons. Based on a dot-immunobinding assay (DIBA), the average incidence of maize dwarf mosaic disease varied from ∼71% (79/111) in 2016, ∼76% (81/106) in 2017, and 62% (28/45) in 2019. Sixty-five DIBA-positive samples for MDMV were further confirmed by RT-PCR, and the complete coat protein (CP) gene was cloned and sequenced. Phylogenetic analysis of 132 isolates (This study = 65; GenBank = 67) revealed two main groups (G1 and G2) of MDMV isolates. All 65 MDMV isolates contained a 39-nucleotide insertion in the N-terminal region of CP genes and clustered in G1 which were different from the isolates in G2, without 39-nucleotide insertion. The first complete genome (9,563 nucleotides) of a MDMV isolate (Bixby1) from Johnsongrass was sequenced, which was distantly related to eight previously reported MDMV isolates from maize. The dN/dS ratio showed mostly purifying selection on each of cistrons except 6K1 being subjected to the diversifying selection. Further analyses revealed three putative recombination events between MDMV-Bixby1 and MDMV isolates from other countries. The successful mechanical and aphid transmission of MDMV-Bixby1 onto maize cultivars was achieved. Altogether, this information showed that Johnsongrass harbors genetically diverse MDMV isolates, which could pose a threat to cultivated crops such as maize and sorghum.

Selecting for Coupling Phase Recombination between Potyvirus Resitance and White Endosperm Colour in Maize Preferred by Farmers in Sub-Saharan Africa (SSA)

Maize lethal necrosis (MLN) disease caused by a combined infection of Maize chlorotic mottle virus (MCMV) and any cereal infecting potyvirus is a threat to food security in Sub-Saharan Africa (SSA). Resistance to potyvirus has been extensively studied and Mdm1 gene for potyvirus resistance on chromosome 6 of maize is linked to Y1 gene for maize endosperm colour. This study is aimed at se- lecting for coupling-phase recombination of potyvirus resistance and white endosperm colour. White susceptible maize lines CML333 and CML277 were crossed with a yellow resistant line, Pa405, to produce F1 and F2 progenies. Progenies were screened using molecular markers to recover 22 white endosperm recombinants. 22 selections were advanced to F3 recombinant families, and 10 were as- sayed for their responses to Maize dwarf mosaic virus (MDMV) and Sugarcane mosaic virus (SCMV). Four families segregated for SCMV resistance, selection of homozygous recombinants within these families will provide lines appropriate for improving lines with resistance to SCMV and MLN resistance in SSA. KEY WORDS: MAIZE LETHAL NECROSIS (MLN), WHITE MAIZE, POTYVIRUS RESISTANCE, GENETIC RECOMBINATION, SUB-SAHARAN AFRICA.