Occurrence, Distribution, and Genetic Diversity of Alfalfa (Medicago sativa L.) Viruses in Four Major Alfalfa-Producing Provinces of China

Alfalfa (Medicago sativa L.) is one of the most widely cultivated forage crops in the world. China is the second largest producer of alfalfa in terms of the planting area worldwide, with Gansu, Henan, Inner Mongolia, and Shaanxi provinces being the production hubs. Alfalfa viruses have been reported on a small-scale survey in some of these areas, but they have not been well characterized. In the present study, seven viruses were detected in 12 fields of 10 cities/counties of the four abovementioned provinces by high-throughput sequencing and assembly of small RNA. Their incidence, distribution, and genetic diversity were analyzed by enzyme-linked immunosorbent assay, polymerase chain reaction (PCR)/reverse transcription-PCR and clone sequencing. The results showed that alfalfa mosaic virus (AMV), pea streak virus (PeSV), lucerne transient streak virus (LTSV), alfalfa dwarf virus (ADV), Medicago sativa alphapartitivirus 1 (MsAPV1), MsAPV2, and alfalfa leaf curl virus (ALCV) were the main viruses infecting alfalfa in four examined provinces. AMV and MsAPV1 had the highest incidences in all 4 provinces. SDT analysis of the 7 viruses isolated in China revealed a highly conserved among AMV, LTSV, ADV, MsAPV1, MsAPV2, and ALCV, but the sequence was a high variation between China isolates to abroad isolates in PeSV, ADV, and ALCV. To our knowledge, this is the first report of ADV in Inner Mongolia and Gansu, ALCV in Inner Mongolia, MsAPV1 and MsAPV2 in all 4 provinces, and PeSV and LTSV in China. These findings provide a basis for future research on the genetic evolution of alfalfa viruses in China and on strategies to prevent diseases in alfalfa caused by these viruses.

First Insights Into the Virus and Viroid Communities in Hemp (Cannabis sativa)

Hemp (Cannabis sativa L.) production has increased significantly in recent years; however, the crop has been understudied in the U.S. since its production declined in the late 1950s. Disease identification and management is an increasing challenge for hemp growers across the country. In 2019, beet curly top virus (BCTV) was first reported in hemp in Colorado. Hence, we were motivated to understand the diversity and prevalence of BCTV strains infecting hemp in Colorado. We detected BCTV at high incidence rate (81%) in leaf samples from 12 counties. Two different strains of BCTV, Worland (Wor) and Colorado (CO) were present as a single or mixed infection in hemp leaf samples. Phylogenetic analysis revealed BCTV sequences from hemp formed a distinct group along with BCTV strains CO and Wor. To determine other potential viral and viroid pathogens in hemp, we performed next generation sequencing (NGS). Virome analysis revealed the presence of both virus and viroid sequences that had high nucleotide sequence identity with GenBank accessions for cannabis cryptic virus, cannabis sativa mitovirus, citrus yellow vein associated virus, opuntia-like virus and hop latent viroid. In contrast, tobacco streak virus sequences were highly variable compared to sequences in GenBank suggesting a possible new genotype of this virus. The data presented here has important implications for the epidemiology and management of the various diseases of hemp and will lead to the development of integrated pest management strategies designed to interrupt transmission cycles and facilitate efficient crop production.

Mining the Genome of Bacillus velezensis VB7 (CP047587) for MAMP Genes and Non-Ribosomal Peptide Synthetase Gene Clusters Conferring Antiviral and Antifungal Activity

Chemical pesticides have an immense role in curbing the infection of plant viruses and soil-borne pathogens of high valued crops. However, the usage of chemical pesticides also contributes to the development of resistance among pathogens. Hence, attempts were made in this study to identify a suitable bacterial antagonist for managing viral and fungal pathogens infecting crop plants. Based on our earlier investigations, we identified Bacillus amyloliquefaciens VB7 as a potential antagonist for managing Sclerotinia sclerotiorum infecting carnation, tobacco streak virus infecting cotton and groundnut bud necrosis infecting tomato. Considering the multifaceted action of B. amyloliquefaciens VB7, attempts were made for whole-genome sequencing to assess the antiviral activity against tomato spotted wilt virus infecting chrysanthemum and antifungal action against Fusarium oxysporum f. sp. cubense (Foc). Genome annotation of the isolate B. amyloliquefaciens VB7 was confirmed as B. velezensis VB7 with accession number CP047587. Genome analysis revealed the presence of 9,231,928 reads with an average read length of 149 bp. Assembled genome had 1 contig, with a total length of 3,021,183 bp and an average G+C content of 46.79%. The protein-coding sequences (CDS) in the genome was 3090, transfer RNA (tRNA) genes were 85 with 29 ribosomal RNA (rRNA) genes and 21 repeat regions. The genome of B. velezensis VB7 had 506 hypothetical proteins and 2584 proteins with functional assignments. VB7 genome had the presence of flagellin protein FlaA with 987 nucleotides and translation elongation factor TU (Ef-Tu) with 1191 nucleotides. The identified ORFs were 3911 with 47.22% GC content. Non ribosomal pepide synthetase cluster (NRPS) gene clusters in the genome of VB7, coded for the anti-microbial peptides surfactin, butirosin A/butirosin B, fengycin, difficidin, bacillibactin, bacilysin, and mersacidin the Ripp lanthipeptide. Antiviral action of VB7 was confirmed by suppression of local lesion formation of TSWV in the local lesion host cowpea (Co-7). Moreover, combined application of B. velezensis VB7 with phyto-antiviral principles M. Jalapa and H. cupanioides increased shoot length, shoot diameter, number of flower buds per plant, flower diameter, and fresh weight of chrysanthemum. Further, screening for antifungal action of VB7 expressed antifungal action against Foc in vitro by producing VOC/NVOC compounds, including hexadecanoic acid, linoelaidic acid, octadecanoic acid, clindamycin, formic acid, succinamide, furanone, 4H-pyran, nonanol and oleic acid, contributing to the total suppression of Foc apart from the presence of NRPS gene clusters. Thus, our study confirmed the scope for exploring B. velezensis VB7 on a commercial scale to manage tomato spotted wilt virus, groundnut bud necrosis virus, tobacco streak virus, S. sclerotiorum, and Foc causing panama wilt of banana.

Farmer and Field Survey in Cassava-Growing Districts of Rwanda Reveals Key Factors Associated With Cassava Brown Streak Disease Incidence and Cassava Productivity

Cassava (Manihot esculenta Crantz) is a vital crop in Rwanda where it ranks as the third most consumed staple. However, cassava productivity remains below its yield potential due to several constraints, including important viral diseases, such as cassava brown streak disease (CBSD). Because various factors can be addressed to mitigate the impact of viral diseases, it is essential to identify routes of virus contamination in the cassava agrosystems from the seed system to farmer's practices and knowledge. The present study aimed at (1) assessing the current cassava seed system and farmers' practices and their knowledge of the biotic constraints to cassava production, (2) determining the status of CBSD as well as critical factors associated with its spread through the seed system channels, and (3) determining factors that influence cassava productivity in Rwanda. A cross-sectional study was carried out from May to September 2019 in 13 districts of Rwanda. A total of 130 farmers and cassava fields were visited, and the incidence and severity of CBSD were evaluated. CBSD was detected in all cassava-producing districts. The highest field incidence of CBSD was recorded in the Nyanza district (62%; 95% CI = 56–67%) followed by the Bugesera district (60%; 95% CI = 54–65%), which recorded the highest severity score of 3.0 ± 0.6. RT-PCR revealed the presence of CBSD at the rate of 35.3%. Ugandan cassava brown streak virus was predominant (21.5%) although cassava brown streak virus was 4% and mixed infection was 10%. An informal cassava seed system was dominant among individual farmers, whereas most cooperatives used quality seeds. Cassava production was found to be significantly influenced by the use of fertilizer, size of the land, farming system, cassava viral disease, and type of cassava varieties grown (p < 0.001). Disease management measures were practiced by a half of participants only. Factors found to be significantly associated with CBSD infection (p < 0.05) were the source of cuttings, proximity to borders, age of cassava, and knowledge of CBSD transmission and management.

Transcriptome of the Maize Leafhopper ( Dalbulus maidis ) and Its Transcriptional Response to Maize Rayado Fino Virus (MRFV), Which It Transmits in a Persistent, Propagative Manner

The transcriptome of the corn leafhopper, D. maidis , revealed conserved biochemical pathways for immunity and discovered transcripts responsive to MRFV-infected plants at two time points, providing a basis for functional identification of genes that either limit or promote the virus-vector interaction. Compared to other hopper species and the propagative plant viruses they transmit, D. maidis shared 15 responsive transcripts with S. furcifera (to southern rice black-streaked dwarf virus [SRBSDV]), one with G. nigrifrons (to maize fine streak virus [MFSV]), and one with P. maidis (to maize mosaic virus [MMV]), but no virus-responsive transcripts identified were shared among all four hopper vector species.

Extrachromosomal viral DNA produced by transcriptionally active endogenous viral elements in non-infected banana hybrids impedes quantitative PCR diagnostics of banana streak virus infections in banana hybrids

The main edible and cultivated banana varieties are intra- and interspecific hybrids of the two main Musa species, Musa acuminata and Musa balbisiana, having diploid genomes denoted A and B, respectively. The B genome naturally hosts sequences of banana streak virus (BSV) named endogenous BSV (eBSV). Upon stress, eBSVs are identified as the origin of BSV infection for at least three BSV species, causing banana streak disease. For each of the three species, BSV and eBSV share >99.9 % sequence identity, complicating PCR-based diagnosis of viral infection in the B genome-containing bananas. Here, we designed a quantitative PCR-based method to only quantify episomal BSV particles produced, overcoming the limitation of eBSV also being detected by qPCR by using it as a ‘calibrator’. However, our results revealed unexpected variation of eBSV amplification in calibrator plants composed of a clonal population of 53 replicating virus-free banana hybrids with the same AAB genotype. Our in-depth molecular analyses suggest that this calibrator variation is due to the variable abundance of non-encapsidated extrachromosomal viral DNA, likely produced via the transcription of eBSVs, followed by occasional reverse transcription. We also present evidence that accumulation of viral transcripts in AAB plants is downregulated both at post-transcriptional and transcriptional levels by an RNA interference mechanism that keeps the plants free of virus infection. Finally, we recommend that such eBSV amplification variation be taken into account to establish a quantitative viral diagnostic for banana plants with the B genome.

Virus Diseases of Cereal and Oilseed Crops in Australia: Current Position and Future Challenges

This review summarizes research on virus diseases of cereals and oilseeds in Australia since the 1950s. All viruses known to infect the diverse range of cereal and oilseed crops grown in the continent’s temperate, Mediterranean, subtropical and tropical cropping regions are included. Viruses that occur commonly and have potential to cause the greatest seed yield and quality losses are described in detail, focusing on their biology, epidemiology and management. These are: barley yellow dwarf virus, cereal yellow dwarf virus and wheat streak mosaic virus in wheat, barley, oats, triticale and rye; Johnsongrass mosaic virus in sorghum, maize, sweet corn and pearl millet; turnip yellows virus and turnip mosaic virus in canola and Indian mustard; tobacco streak virus in sunflower; and cotton bunchy top virus in cotton. The currently less important viruses covered number nine infecting nine cereal crops and 14 infecting eight oilseed crops (none recorded for rice or linseed). Brief background information on the scope of the Australian cereal and oilseed industries, virus epidemiology and management and yield loss quantification is provided. Major future threats to managing virus diseases effectively include damaging viruses and virus vector species spreading from elsewhere, the increasing spectrum of insecticide resistance in insect and mite vectors, resistance-breaking virus strains, changes in epidemiology, virus and vectors impacts arising from climate instability and extreme weather events, and insufficient industry awareness of virus diseases. The pressing need for more resources to focus on addressing these threats is emphasized and recommendations over future research priorities provided.

PAL1 gene of the phenylpropanoid pathway increases resistance to the Cassava brown streak virus in cassava

Background The phenylalanine ammonia lyase genes play crucial role in plant response to biotic and abiotic stresses. In this study, we characterized the role of PAL genes in increasing resistance to the Cassava brown streak virus that causes the economically important cassava brown streak disease (CBSD) on cassava in Africa. Methods The whole transcriptomes of eight cassava varieties differing in resistance to CBSD were obtained at 1, 5 and 8 weeks after CBSV infection. Results Analysis of RNA-Seq data identified the overexpression of PAL1, PAL2, cinnamic acid and two chalcone synthase genes in CBSD-resistant cassava varieties, which was subsequently confirmed by RT-qPCR. The exogenous application of Acibenzolar-S-Methyl induced PAL1 gene expression to enhance resistance in the susceptible var. Kalawe. In contrast, the silencing of PAL1 by RNA interference led to increased susceptibility of the resistant var. Kaleso to CBSD. Conclusions PAL1 gene of the phenylpropanoid pathway has a major role in inducing resistance to CBSD in cassava plants and its early induction is key for CBSD resistance.