Beet curly top virus affects vector biology: the first transcriptome analysis of the beet leafhopper.

Curly top disease, caused by beet curly top virus (BCTV), is among the most serious viral diseases affecting sugar beets in western USA. The virus is exclusively transmitted by the beet leafhopper (BLH, Circulifer tenellus) in a circulative and non-propagative manner. Despite the growing knowledge on virus-vector interactions, our understanding of the molecular interactions between BCTV and BLH is hampered by limited information regarding the virus impact on the vector and the lack of genomic and transcriptomic resources for BLH. This study unveils the significant impact of BCTV on both the performance and transcriptome response of BLHs. Viruliferous BLHs had higher fecundity than non-viruliferous counterparts, which was evident by upregulation of differentially expressed transcripts (DETs) associated with development, viability and fertility of germline and embryos in viruliferous insects. Conversely, most DETs associated with muscle movement and locomotor activities were downregulated in viruliferous insects, implying potential behavioural modifications by BCTV. Additionally, a great proportion of DETs related to innate immunity and detoxification were upregulated in viruliferous insects. Viral infection also induced notable alterations in primary metabolisms, including energy metabolism, namely glucosidases, lipid digestion and transport, and protein degradation, along with other cellular functions, particularly in chromatin remodelling and DNA repair. This study represents the first comprehensive transcriptome analysis for BLH. The presented findings provide new insights into the multifaceted effects of viral infection on various biological processes in BLH, offering a foundation for future investigations into the complex virus-vector relationship and potential management strategies for curly top disease.

First report of Pepper yellow dwarf strain of Beet curly top virus and Spinach curly top Arizona virus in red table beet in Idaho, United States.

In the fall 2021, red table beet plants (Beta vulgaris L. cv 'Eagle') exhibiting stunted growth with shorter petioles were observed at an incidence of 10 to 15 percent in a production field in Payette County, Idaho, United States. In addition to stunting, beet leaves displayed yellowing and mild curling and crumpling, and the roots exhibited hairy root symptoms (sFig.1). To identify potential causal viruses, total RNA was isolated from the leaf and root tissue using RNeasy Plant Mini Kit (Qiagen, Valencia, CA) and subjected to high-throughput sequencing (HTS). Two libraries were prepared, one for the leaf sample and another for the root sample using a ribo-minus TruSeq Stranded Total RNA Library Prep kit (Illumina, San Diego, CA). HTS was performed with 150 bp paired-end sequencing on a NovaSeq 6000 (Novogene, Sacramento, CA). Following adapter trimming and removal of host transcripts, 5.9 and 16.2 million reads were obtained from the leaf and root samples, respectively. These reads were de novo assembled using the SPAdes assembler (Bankevitch et al., 2012; Prjibelski et al., 2020). The assembled leaf sample contigs were aligned to the NCBI non-redundant database to identify contigs matching known viruses. A single contig of 2845 nts that shared 96% coverage and 95.6% sequence identity to the pepper yellow dwarf strain of beet curly top virus (BCTV-PeYD, EU921828; Varsani et al., 2014), and 98% coverage and 98.39% identity with an isolate of BCTV-PeYD (KX529650) from Mexico, was identified in the leaf sample (GenBank Accession OP477336). To validate the HTS detection of BCTV-PeYD, total DNA was isolated from the leaf sample and a 454 bp fragment of the C1 gene (replication-associate protein) was PCR amplified and Sanger sequencing of the amplicon revealed 99.7% identity to the HTS assembled BCTV-PeYD sequence. In addition to the PeYD strain of BCTV, the Worland strain of BCTV (BCTV-Wor) was detected as a single 2930 nt contig with 100% coverage and 97.3% identity to the BCTV-Wor isolate CTS14-015 (KX867045) known to infect sugar beet in Idaho. Of note, there are 11 strains of BCTV and among those, the BCTV-Wor strain induces mild symptoms in sugar beet (Strausbaugh et al., 2017), whereas BCTV-PeYD was found only in pepper from New Mexico. Further, two contigs of 2201 nts and 523 nts were assembled generating a nearly complete genome of spinach curly top Arizona virus (SpCTAV) in the leaf sample with 99% coverage and 99.3% identity (GenBank Accession OQ703946) to the reference genome of SpCTAV (HQ443515; Hernandez-Zepeda et al., 2013). To validate the HTS results, total DNA was isolated from the leaf tissue and PCR amplified a 442 bp fragment that overlaps the V1, V2, and V3 ORFs and its sequence revealed 100% identity with the HTS assembled SpCTAV. The roots sample also showed HTS reads corresponding to BCTV-PeYD and SpCTAV. In addition, beet necrotic yellow vein virus (BNYVV) was detected in the root sample with 30% coverage, but no sequence reads matching to BNYVV was detected in the leaf sample. BNYVV is known to infect sugar beet causing rhizomania (Tamada et al., 1973; Schirmer et al., 2005). To further confirm the BNYVV HTS results, total RNA was extracted separately from the root and leaf tissue, and RT-PCR was performed with primers that were designed to amplify portions of BNYVV RNAs (Weiland et al., 2020). RT-PCR analysis generated the appropriate amplicons with expected sequences corresponding to the RNA-1, RNA-2, RNA-3, and RNA-4 of BNYVV as determined by Sanger sequencing implying BNYVV the causal agent of hairy root symptoms. Similar to observations seen for BNYVV infection in conventional sugar beet varieties, no amplification was detected for BNYVV in the RNA extracted from leaf tissue, indicating that the RT-PCR results are consistent with the HTS analysis. This is the first report of BCTV-PeYD and SpCTAV observed naturally infecting red table beet in Idaho suggesting the geographical expansion of these viruses. The co-existence of BCTV-PeYD and SpCTAV with limited host range needs to be investigated to determine the actual cause of the observed foliar symptoms. This report provides the basis for further research to understand the pathogenic nature of these viruses and their potential threat to red table beet and sugar beet production in Idaho.

Influence of Harvest Timing, Fungicides, and Beet necrotic yellow vein virus on Sugar Beet Storage.

Root rots in sugar beet storage can lead to multimillion dollar losses because of reduced sucrose recovery. Thus, studies were conducted to establish additional fungicide treatments for sugar beet storage and a greater understanding of the fungi involved in the sugar beet storage rot complex in Idaho. A water control treatment and three fungicides (Mertect [product at 0.065 ml/kg of roots; 42.3% thiabendazole {vol/vol}], Propulse [product at 0.049 ml/kg of roots; 17.4% fluopyram and 17.4% prothioconazole {vol/vol}], and Stadium [product at 0.13 ml/kg of roots; 12.51% azoxystrobin, 12.51% fludioxonil, and 9.76% difenoconozole {vol/vol}]) were investigated for the ability to control fungal rots of sugar beet roots held up to 148 days in storage during the 2012 and 2013 storage seasons. At the end of September into October, roots were harvested weekly for 5 weeks from each of two sugar beet fields in Idaho, treated with the appropriate fungicide, and placed on top of a commercial indoor sugar beet storage pile until early February. Differences (P < 0.0001 to 0.0150) among fungicide treatments were evident. Propulse- and Stadium-treated roots had 84 to 100% less fungal growth versus the control roots, whereas fungal growth on Mertect-treated roots was not different from the control roots in 7 of 12 comparisons for roots harvested each of the first 3 weeks in both years of this study. The Propulse- and Stadium-treated roots also reduced (P < 0.0001 to 0.0146; based on weeks 1, 3, and 4 in 2012 and weeks 1, 3, 4, and 5 in 2013) sucrose loss by 14 to 46% versus the control roots, whereas roots treated with Mertect did not change sucrose loss compared with the control roots in 7 of 10 evaluations. The predominant fungi isolated from symptomatic roots were an Athelia-like sp., Botrytis cinerea, Penicillium spp., and Phoma betae. If Propulse and Stadium are labeled for use on sugar beet in storage, these fungicides should be considered for root rot control in commercial sugar beet storage and on roots held for vernalization for seed production of this biennial plant species.

Effects of Strip Tillage in Sugar Beet on Density and Richness of Predatory Arthropods.

Strip tillage, in which tillage and seedbed preparation are limited to a narrow band where the subsequent crop is planted, provides many potential agronomic benefits, including reduced fuel and labor costs, reduced erosion, and improved soil tilth. Lower soil disturbance and enhanced water retention associated with strip tillage also may affect density and diversity of predatory arthropods, which have been little studied in sugar beet. We examined the effects of tillage (conventional versus strip) on the predatory epigeal arthropod fauna in sugar beet. Studies were conducted over three growing seasons (2010-2012) in Idaho using both fenced and unfenced pitfall traps to sample arthropods. Unfenced pitfall traps often captured a greater activity density and richness of predators, and showed no bias of higher captures in conventionally tilled plots as has been shown elsewhere. Total density of predators was higher in strip tillage only during 2011. Density and species richness of carabid beetles did not differ between tillage treatments during the course of the study. Density of the other major taxa (staphylinid beetles, spiders, and Opiliones) was higher under strip tillage during some years, especially early in the season, but richness showed little or no relationship with tillage. Predaceous arthropods might be favored by enhanced ground cover, higher humidity, more moderate temperatures, and/or less habitat disturbance associated with strip-tilled plots. The results suggest that certain groups of soil-dwelling predatory arthropods can be favored by strip tillage in sugar beet, which further adds to the benefits of conservation tillage in this system.