Comparative plant transcriptome profiling of Arabidopsis and Camelina infested with Myzus persicae aphids acquiring circulative and non-circulative viruses reveals virus- and plant-specific alterations relevant to aphid feeding behavior and transmission

Background: Evidence accumulates that plant viruses alter host-plant traits in ways that modify their insect vectors' behavior. These alterations often enhance virus transmission, which has led to the hypothesis that these effects are manipulations caused by viral adaptation. However, the genetic basis of these indirect, plant-mediated effects on vectors and their dependence on the plant host and the mode of virus transmission is hardly known. Results: Transcriptome profiling of Arabidopsis thaliana and Camelina sativa plants infected with turnip yellows virus (TuYV) or cauliflower mosaic virus (CaMV) and infested with the common aphid vector Myzus persicae revealed strong virus- and host-specific differences in the gene expression patterns. CaMV infection caused more severe effects on the phenotype of both plant hosts than did TuYV infection, and the severity of symptoms correlated strongly with the proportion of differentially expressed genes, especially photosynthesis genes. Accordingly, CaMV infection modified aphid behavior and fecundity stronger than did infection with TuYV. Conclusions: Overall, infection with CaMV — relying on the non-circulative transmission mode — tends to have effects on metabolic pathways with strong potential implications for insect-vector / plant-host interactions (e.g. photosynthesis, jasmonic acid, ethylene and glucosinolate biosynthetic processes), while TuYV — using the circulative transmission mode — alters these pathways only weakly. These virus-induced deregulations of genes that are related to plant physiology and defense responses might impact aphid probing and feeding behavior on both infected host plants, with potentially distinct effects on virus transmission. Keywords: Caulimovirus, polerovirus, aphid vector, transmission, feeding behavior, insect-plant interactions, transcriptome profiling, RNA-seq.

Effects of an alphasatellite on life cycle of the nanovirus Faba bean necrotic yellows virus

Nanoviruses are plant viruses with a multipartite single-stranded DNA (ssDNA) genome. Alphasatellites are commonly associated with nanovirus infections, but their putative impact on their helper viruses is unknown. In this study, we investigated the role of subterranean clover stunt alphasatellite 1 (hereafter named SCSA 1) on various important traits of faba bean necrotic yellows virus (FBNYV) in its host plant Vicia faba and aphid vector Acyrthosiphon pisum , including disease symptoms, viral accumulation and transmission. The results indicate that SCSA 1 does not affect the symptom severity nor the overall FBNYV accumulation in V. faba, but changes the relative amounts of its different genomic segments. Moreover, the association of SCSA 1 with FBNYV increases the rate of plant-to-plant transmission by a process seemingly unrelated to simple increase of the viral accumulation in the vector. These results represent the first study on the impact of an alphasatellite on the biology of its helper nanovirus. They suggest that SCSA 1 may benefit FBNYV, but the genericity of this conclusion is discussed and questioned. Importance Alphasatellites are circular single stranded DNA molecules frequently found in association with natural isolates of nanoviruses and some geminiviruse, the two ssDNA plant infecting virus families. While the implications of alphasatellite presence in geminivirus infections are relatively well documented, comparable studies on alphasatellites associated with nanoviruses are not available. Here we confirm that subterranean clover stunt alphasatellite 1 affects different traits of its helper nanovirus, faba bean necrotic yellows virus, both in the host plant and aphid vector. We show that the frequencies of the virus segments change in the presence of alphasatellite, in both plant and vector. We also confirm that while within-plant virus load and symptom are not affected by alphasatellite, the presence of alphasatellite decreases within-aphid virus load, but significantly increases virus transmission rate, so may confer a possible evolutionary advantage for the helper virus.

Alfalfa leaf curl virus is efficiently acquired by its aphid vector Aphis craccivora but inefficiently transmitted

Alfalfa leaf curl virus (ALCV) is the first geminivirus for which aphid transmission was reported. Transmission by Aphis craccivora was determined previously to be highly specific and circulative. Using various complementary techniques, the transmission journey of ALCV was monitored from its uptake from infected plant tissues up to the head of its vector. ALCV was shown to be restricted to phloem tissues using fluorescence in situ hybridization (FISH) and electropenetrography (EPG) monitoring of virus acquisition. Furthermore, the virus is heterogeneously distributed in phloem tissues, as revealed by FISH and quantitative PCR of viral DNA acquired by EPG-monitored aphids. Despite the efficient ingestion of viral DNA, about 106 viral DNA copies per insect in a 15 h feeding period on ALCV-infected plants, the individual maximum transmission rate was 12 %. Transmission success was related to a critical viral accumulation, around 1.6×107 viral DNA copies per insect, a threshold that generally needed more than 48 h to be reached. Moreover, whereas the amount of acquired virus did not decrease over time in the whole aphid body, it declined in the haemolymph and heads. ALCV was not detected in progenies of viruliferous aphids and did not affect aphid fitness. Compared to geminiviruses transmitted by whiteflies or leafhoppers, or to luteoviruses transmitted by aphids, the transmission efficiency of ALCV by A. craccivora is low. This result is discussed in relation to the aphid vector of this geminivirus and the agroecological features of alfalfa, a hardy perennial host plant.

Co-Acquired Nanovirus and Geminivirus Exhibit a Contrasted Localization within Their Common Aphid Vector

Single-stranded DNA (ssDNA) plant viruses belong to the families Geminiviridae and Nanoviridae. They are transmitted by Hemipteran insects in a circulative, mostly non-propagative, manner. While geminiviruses are transmitted by leafhoppers, treehoppers, whiteflies and aphids, nanoviruses are transmitted exclusively by aphids. Circulative transmission involves complex virus–vector interactions in which epithelial cells have to be crossed and defense mechanisms counteracted. Vector taxa are considered a relevant taxonomic criterion for virus classification, indicating that viruses can evolve specific interactions with their vectors. Thus, we predicted that, although nanoviruses and geminiviruses represent related viral families, they have evolved distinct interactions with their vector. This prediction is also supported by the non-structural Nuclear Shuttle Protein (NSP) that is involved in vector transmission in nanoviruses but has no similar function in geminiviruses. Thanks to the recent discovery of aphid-transmitted geminiviruses, this prediction could be tested for the geminivirus alfalfa leaf curl virus (ALCV) and the nanovirus faba bean necrotic stunt virus (FBNSV) in their common vector, Aphis craccivora. Estimations of viral load in midgut and head of aphids, precise localization of viral DNA in cells of insect vectors and host plants, and virus transmission tests revealed that the pathway of the two viruses across the body of their common vector differs both quantitatively and qualitatively.

Route of a Multipartite Nanovirus across the Body of Its Aphid Vector

ABSTRACT Vector transmission plays a primary role in the life cycle of viruses, and insects are the most common vectors. An important mode of vector transmission, reported only for plant viruses, is circulative nonpropagative transmission whereby the virus cycles within the body of its insect vector, from gut to salivary glands and saliva, without replicating. This mode of transmission has been extensively studied in the viral families Luteoviridae and Geminiviridae and is also reported for Nanoviridae. The biology of viruses within these three families is different, and whether the viruses have evolved similar molecular/cellular virus-vector interactions is unclear. In particular, nanoviruses have a multipartite genome organization, and how the distinct genome segments encapsidated individually transit through the insect body is unknown. Here, using a combination of fluorescent in situ hybridization and immunofluorescence, we monitor distinct proteins and genome segments of the nanovirus Faba bean necrotic stunt virus (FBNSV) during transcytosis through the gut and salivary gland cells of its aphid vector Acyrthosiphon pisum. FBNSV specifically transits through cells of the anterior midgut and principal salivary gland cells, a route similar to that of geminiviruses but distinct from that of luteoviruses. Our results further demonstrate that a large number of virus particles enter every single susceptible cell so that distinct genome segments always remain together. Finally, we confirm that the success of nanovirus-vector interaction depends on a nonstructural helper component, the viral protein nuclear shuttle protein (NSP), which is shown to be mandatory for viral accumulation within gut cells. IMPORTANCE An intriguing mode of vector transmission described only for plant viruses is circulative nonpropagative transmission, whereby the virus passes through the gut and salivary glands of the insect vector without replicating. Three plant virus families are transmitted this way, but details of the molecular/cellular mechanisms of the virus-vector interaction are missing. This is striking for nanoviruses that are believed to interact with aphid vectors in ways similar to those of luteoviruses or geminiviruses but for which empirical evidence is scarce. We here confirm that nanoviruses follow a within-vector route similar to that of geminiviruses but distinct from that of luteoviruses. We show that they produce a nonstructural protein mandatory for viral entry into gut cells, a unique phenomenon for this mode of transmission. Finally, noting that nanoviruses are multipartite viruses, we demonstrate that a large number of viral particles penetrate susceptible cells of the vector, allowing distinct genome segments to remain together.

Alfalfa leaf curl virus is efficiently acquired by its aphid vector Aphis craccivora but inefficiently transmitted

ABSTRACTAlfalfa leaf curl virus (ALCV) is the first geminivirus for which an aphid transmission was reported. Transmission by Aphis craccivora was determined previously to be highly specific and circulative. Using various complementary techniques, the transmission journey of ALCV was monitored from its uptake in an infected plant tissue up to the head of its vector. ALCV was shown to be restricted in the phloem using fluorescent in situ hybridization (FISH) and electropenetrography (EPG) monitoring of virus acquisition. Furthermore, the virus is heterogeneously distributed in the phloem as revealed by FISH and qPCR quantification of the viral DNA acquired by aphids monitored by EPG. In spite of the efficient ingestion of viral DNA, about 106 in a 15-hour feeding on ALCV infected plants, the individual transmission rate was at a maximum of 12%. Transmission success was related to a critical viral accumulation, around 1.6×107 viral DNA copies per insect, a threshold that needs generally more than 48 hours to be reached. Moreover, whereas the amount of acquired virus does not decrease over time in the whole aphid body, it decreased in hemolymph and heads. ALCV was not detected in progenies of viruliferous aphids and had no effect on aphid fitness. Compared to geminiviruses transmitted by whiteflies or leafhoppers or to luteovirus transmitted by aphids, the transmission efficiency of ALCV by A. craccivora is low. This result is discussed in relation to the aphid vector of this geminivirus and the agroecological features of alfalfa, a hardy perennial host plant.