Integrating High throughput Sequencing into Survey Design Reveals Turnip Yellows Virus and Soybean Dwarf Virus in Pea (Pisum Sativum) in the United Kingdom

There is only limited knowledge of the presence and incidence of viruses in peas within the United Kingdom, therefore high-throughput sequencing (HTS) in combination with a bulk sampling strategy and targeted testing was used to determine the virome in cultivated pea crops. Bulks of 120 leaves collected from twenty fields from around the UK were initially tested by HTS, and presence and incidence of virus was then determined using specific real-time reverse-transcription PCR assays by testing smaller mixed-bulk size samples. This study presents the first finding of turnip yellows virus (TuYV) in peas in the UK and the first finding of soybean dwarf virus (SbDV) in the UK. While TuYV was not previously known to be present in UK peas, it was found in 13 of the 20 sites tested and was present at incidences up to 100%. Pea enation mosaic virus-1, pea enation mosaic virus-2, pea seed-borne mosaic virus, bean yellow mosaic virus, pea enation mosaic virus satellite RNA and turnip yellows virus associated RNA were also identified by HTS. Additionally, a subset of bulked samples were re-sequenced at greater depth to ascertain whether the relatively low depth of sequencing had missed any infections. In each case the same viruses were identified as had been identified using the lower sequencing depth. Sequencing of an isolate of pea seed-borne mosaic virus from 2007 also revealed the presence of TuYV and SbDV, showing that both viruses have been present in the UK for at least a decade, and represents the earliest whole genome of SbDV from Europe. This study demonstrates the potential of HTS to be used as a surveillance tool, or for crop-specific field survey, using a bulk sampling strategy combined with HTS and targeted diagnostics to indicate both presence and incidence of viruses in a crop.

Genomic Characterization of a New Enamovirus Infecting Common Bean

A novel enamovirus was identified from bean plants with disease symptoms. Its genome of 5,781 nucleotides (nt) encodes five open reading frames. The virus and other species of the genus Enamovirus share identities of 50.4%-68.4% at the complete genome, and 19.9%-51.9% of P0, 24.9%-52.5% of P1, 33.4%-62.9% of P1-P2, 30.6%-81.1% of P3, 32.3%-74.2% of P3-P5 at amino acid sequence level, respectively. Phylogenetic analysis showed that the virus is most closely related to Alfalfa enamovirus 1 and Pea enation mosaic virus 1 in the genus Enamovirus within family Solemoviridae. These results suggest that the virus should be considered as a novel species in the genus Enamovirus and tentatively named as “bean enamovirus 1”.

Plant responses to multiple antagonists are mediated by order of attack and phytohormone crosstalk

Plants are often attacked by multiple antagonists, and traits of the attacking organisms, and their order of arrival onto hosts, may affect plant defenses. However, few studies have assessed how multiple antagonists, and varying attack order, affect plant defense or nutrition. To address this, we assessed defensive and nutritional responses of Pisum sativum plants after attack by a vector herbivore (Acrythosiphon pisum), a non-vector herbivore (Sitona lineatus), and a pathogen (Pea enation mosaic virus, PEMV). We show PEMV-infectious A. pisum induced several pathogen-specific plant defense signals, but these defenses were inhibited when S. lineatus was present in peas infected with PEMV. In contrast, feeding by S. lineatus induced anti-herbivore defense signals, but these defenses were enhanced by PEMV. Sitonalineatus also increased abundance of plant amino acids, but only when they attacked after PEMV-infectious A. pisum. Our results suggest that diverse communities of biotic antagonists alter defense and nutritional traits of plants through complex pathways that depend on the identity of attackers and their order of arrival onto hosts. Moreover, we show interactions among a group of biotic stressors can vary along a spectrum from antagonism to enhancement/synergism based on the identity and order of attackers, and these interactions are mediated by a multitude of phytohormone pathways.

Plant responses to multiple antagonists are mediated by order of attack and phytohormone crosstalk

Plants are often attacked by multiple antagonists, and traits of the attacking organisms, and their order of arrival onto hosts, may affect plant defenses. However, few studies have assessed how multiple antagonists, and varying attack order, affect plant defense or nutrition. To address this, we assessed defensive and nutritional responses of Pisum sativum plants after attack by a vector herbivore (Acrythosiphon pisum), a non-vector herbivore (Sitona lineatus), and a pathogen (Pea enation mosaic virus, PEMV). We show PEMV-infectious A. pisum induced several pathogen-specific plant defense signals, but these defenses were inhibited when S. lineatus was present in peas infected with PEMV. In contrast, feeding by S. lineatus induced anti-herbivore defense signals, but these defenses were enhanced by PEMV. Sitona lineatus also increased abundance of plant amino acids, but only when they attacked after PEMV-infectious A. pisum. Our results suggest that diverse communities of biotic antagonists alter defense and nutritional traits of plants through complex pathways that depend on the identity of attackers and their order of arrival onto hosts. Moreover, we show interactions among a group of biotic stressors can vary along a spectrum from antagonism to enhancement/synergism based on the identity and order of attackers, and these interactions are mediated by a multitude of phytohormone pathways.

The Multifunctional Long-Distance Movement Protein of Pea Enation Mosaic Virus 2 Protects Viral and Host Transcripts from Nonsense-Mediated Decay

ABSTRACT The nonsense-mediated decay (NMD) pathway presents a challenge for RNA viruses with termination codons that precede extended 3′ untranslated regions (UTRs). The umbravirus Pea enation mosaic virus 2 (PEMV2) is a nonsegmented, positive-sense RNA virus with an unusually long 3′ UTR that is susceptible to NMD. To establish a systemic infection, the PEMV2 long-distance movement protein p26 was previously shown to both stabilize viral RNAs and bind them for transport through the plant’s vascular system. The current study demonstrated that p26 protects both viral and nonviral messenger RNAs from NMD. Although p26 localizes to both the cytoplasm and nucleolus, p26 exerts its anti-NMD effects exclusively in the cytoplasm independently of long-distance movement. Using a transcriptome-wide approach in the model plant Nicotiana benthamiana, p26 protected a subset of cellular NMD target transcripts, particularly those containing long, structured, GC-rich 3′ UTRs. Furthermore, transcriptome sequencing (RNA-seq) revealed that the NMD pathway is highly dysfunctional during PEMV2 infection, with 1,820 (48%) of NMD targets increasing in abundance. Widespread changes in the host transcriptome are common during plant RNA virus infections, and these results suggest that, in at least some instances, virus-mediated NMD inhibition may be a major contributing factor. IMPORTANCE Nonsense-mediated decay (NMD) represents an RNA regulatory pathway that degrades both natural and faulty messenger RNAs with long 3′ untranslated regions. NMD targets diverse families of RNA viruses, requiring that viruses counteract the NMD pathway for successful amplification in host cells. A protein required for long-distance movement of Pea enation mosaic virus 2 (PEMV2) is shown to also protect both viral and host mRNAs from NMD. RNA-seq analyses of the Nicotiana benthamiana transcriptome revealed that PEMV2 infection significantly impairs the host NMD pathway. RNA viruses routinely induce large-scale changes in host gene expression, and, like PEMV2, may use NMD inhibition to alter the host transcriptome in an effort to increase virus amplification.