Inter-virus relationships in mixed infections and virus-drought relationships in plants: a quantitative review.

Inter-virus relationships in mixed infections and virus-drought relationships are important in agriculture and natural vegetation. In this quantitative review, we sampled published factorial experiments to probe for relationships against the null hypothesis of additivity. Our sample captured antagonistic, additive and synergistic inter-virus relationships in double infections. Virus-drought relationships in our sample were additive or antagonistic, reinforcing the notion that viruses have neutral or positive effects on droughted plants, or that drought enhances plant tolerance to viruses. Both inter-virus and virus-drought relationships vary with virus species, host plant to the level of cultivar or accession, timing of infection, plant age and trait and growing conditions. The trait-dependence of these relationships has implications for resource allocation in plants. Owing to lagging theories, more experimental research in these fields is bound to return phenomenological outcomes. Theoretical work can advance in two complementary directions. First, the effective theory models the behaviour of the system without specifying all the underlying causes that lead to system state change. Second, mechanistic theory based on a nuanced view of the plant phenotype that explicitly considers downward causation; the influence of the plant phenotype on inter-virus relations and vice versa; the impact of timing, intensity and duration of drought interacting with viruses to modulate the plant phenotype; both the soil (moisture) and atmospheric (vapour pressure deficit) aspects of drought. Theories should scale in time, from short term to full growing season, and in levels of organisation up to the relevant traits: crop yield in agriculture and fitness in nature.

A synchronized, large-scale field experiment using Arabidopsis thaliana reveals the significance of the UV-B photoreceptor UVR8 under natural conditions.

This study determines the functional role of the plant ultraviolet-B radiation (UV-B) photoreceptor, UV RESISTANCE LOCUS 8 (UVR8) under natural conditions using a large-scale 'synchronized-genetic-perturbation-field-experiment'. Laboratory experiments have demonstrated a role for UVR8 in UV-B responses but do not reflect the complexity of outdoor conditions where 'genotype × environment' interactions can mask laboratory-observed responses. Arabidopsis thaliana knockout mutant, uvr8-7, and the corresponding Wassilewskija wild type, were sown outdoors on the same date at 21 locations across Europe, ranging from 39°N to 67°N latitude. Growth and climatic data were monitored until bolting. At the onset of bolting, rosette size, dry weight, and phenolics and glucosinolates were quantified. The uvr8-7 mutant developed a larger rosette and contained less kaempferol glycosides, quercetin glycosides and hydroxycinnamic acid derivatives than the wild type across all locations, demonstrating a role for UVR8 under field conditions. UV effects on rosette size and kaempferol glycoside content were UVR8 dependent, but independent of latitude. In contrast, differences between wild type and uvr8-7 in total quercetin glycosides, and the quercetin-to-kaempferol ratio decreased with increasing latitude, that is, a more variable UV response. Thus, the large-scale synchronized approach applied demonstrates a location-dependent functional role of UVR8 under natural conditions.

Novel program for automatic calculation of EPG variables.

The electrical penetration graph (EPG) technique is the most powerful tool for studying the feeding behavior of pierce-sucking insects. However, calculating EPG variables is often very time-consuming, and consequently, several software programs have been developed for the automatic calculation of EPG variables. Here we present a new user-friendly Excel Workbook that uses a standardized list of EPG variables and follows expert guidelines for calculating them. The program developed in Visual Basic for Applications (VBA) is a step up from the existing software and allows easy data analysis and interpretation. It also includes a novel option for dealing with the common problem of "truncated"-waveforms artificially terminated by the end of recording. The only requirement to run the program is Microsoft Excel software running under a PC environment. The Workbook was validated by calculating variables from EPG recordings of aphids and psyllids and the results obtained were compared with those of existing software such as the Sarria Workbook. Our EPG Workbook provides researchers with a reliable and standardized tool for the automatic calculation of up to 127 EPG variables from phloem-sap-sucking insects.

Epidemiology and Management of Plant Viruses Under a Changing Climate.

Plant viruses are an ever-present threat to agricultural production and provide a wide array of symptoms resulting in economic losses throughout the world. Diseases can be transmitted by insect vectors, as well as through pollen, seed, and other means. With the increased globalization of agriculture, the introduction of new viruses from exotic locations and their establishment in new production regions and even new crops is a growing concern. Advancing knowledge of the epidemiology of plant viruses including development of new diagnostic methods, virus surveillance, and modeling, virus ecology and evolution, virus interactions with insect vectors, and other factors are important toward reducing the spread of plant viruses and managing virus diseases.

Pectin Methylesterases Modulate Plant Homogalacturonan Status in Defenses against the Aphid Myzus persicae.

Because they suck phloem sap and act as vectors for phytopathogenic viruses, aphids pose a threat to crop yields worldwide. Pectic homogalacturonan (HG) has been described as a defensive element for plants during infections with phytopathogens. However, its role during aphid infestation remains unexplored. Using immunofluorescence assays and biochemical approaches, the HG methylesterification status and associated modifying enzymes during the early stage of Arabidopsis (Arabidopsis thaliana) infestation with the green peach aphid (Myzus persicae) were analyzed. Additionally, the influence of pectin methylesterase (PME) activity on aphid settling and feeding behavior was evaluated by free choice assays and the Electrical Penetration Graph technique, respectively. Our results revealed that HG status and HG-modifying enzymes are significantly altered during the early stage of the plant-aphid interaction. Aphid infestation induced a significant increase in total PME activity and methanol emissions, concomitant with a decrease in the degree of HG methylesterification. Conversely, inhibition of PME activity led to a significant decrease in the settling and feeding preference of aphids. Furthermore, we demonstrate that the PME inhibitor AtPMEI13 has a defensive role during aphid infestation, since pmei13 mutants are significantly more susceptible to M. persicae in terms of settling preference, phloem access, and phloem sap drainage.

Impact of the nicotinic acetylcholine receptor mutation R81T on the response of European Myzus persicae populations to imidacloprid and sulfoxaflor in laboratory and in the field.

Sulfoxaflor (Isoclast™ active) is a sulfoximine insecticide that is active on a broad range of sap-feeding insects, including species that exhibit reduced susceptibility to currently available insecticides. Colonies of Myzus persicae (green peach aphid) were established from aphids collected in the field from peach (Prunus persica) and nectarine (Prunus persica var. nucipersica) orchards in France, Italy and Spain. The presence of the nicotinic acetylcholine receptor (nAChR) point mutation R81T was determined for all the colonies. Eight of the 35 colonies collected were susceptible relative to R81T (i.e., R81T absent), three of the colonies were found to be homozygous for R81T while 24 colonies had R81T present in some proportion (heterozygous). Sulfoxaflor and imidacloprid were tested in the laboratory against these M. persicae field colonies, which exhibited a wide range of susceptibilities (sulfoxaflor RR = 0.6 to 61, imidacloprid RR = 0.7 to 986) (resistance ratios, RR) to both insecticides. Although sulfoxaflor was consistently more active than imidacloprid against these field collected M. persicae, there was a statistically significant correlation across all colonies between the RRs for imidacloprid and sulfoxaflor (Pearson's r = 0.939, p < 0.0001). However, when a larger group of the colonies from Spain possessing R81T were analyzed, there was no correlation observed for the RRs between imidacloprid and sulfoxaflor (r = 0.2901, p = 0.3604). Thus, consistent with prior studies, the presence of R81T by itself is not well correlated with altered susceptibility to sulfoxaflor. In field trials, sulfoxaflor (24 and 36 gai/ha) was highly effective (~avg. 88-96% control) against M. persicae, demonstrating similar levels of efficacy as flonicamid (60-70 gai/ha) and spirotetramat (100-180 gai/ha) at 13-15 days after application, in contrast to imidacloprid (110-190 gai/ha) and acetamiprid (50-75 gai/ha) with lower levels of efficacy (~avg. 62-67% control). Consequently, sulfoxaflor is an effective tool for use in insect pest management programs for M. persicae. However, it is recommended that sulfoxaflor be used in the context of an insecticide resistance management program as advocated by the Insecticide Resistance Action Committee involving rotation with insecticides possessing other modes of action (i.e., avoiding rotation with other Group 4 insecticides) to minimize the chances for resistance development and to extend its future utility.

Role of Acrostyle Cuticular Proteins in the Retention of an Aphid Salivary Effector.

To avoid the activation of plant defenses and ensure sustained feeding, aphids are assumed to use their mouthparts to deliver effectors into plant cells. A recent study has shown that effectors detected near feeding sites are differentially distributed in plant tissues. However, the precise process of effector delivery into specific plant compartments is unknown. The acrostyle, a cuticular organ located at the tip of maxillary stylets that transiently binds plant viruses via its stylin proteins, may participate in this specific delivery process. Here, we demonstrate that Mp10, a saliva effector released into the plant cytoplasm during aphid probing, binds to the acrostyles of Acyrthosiphon pisum and Myzus persicae. The effector probably interacts with Stylin-03 as a lowered Mp10-binding to the acrostyle was observed upon RNAi-mediated reduction in Stylin-03 production. In addition, Stylin-03 and Stylin-01 RNAi aphids exhibited changes in their feeding behavior as evidenced by electrical penetration graph experiments showing longer aphid probing behaviors associated with watery saliva release into the cytoplasm of plant cells. Taken together, these data demonstrate that the acrostyle also has effector binding capacity and supports its role in the delivery of aphid effectors into plant cells.

Barley yellow dwarf virus Can Be Inoculated During Brief Intracellular Punctures in Phloem Cells Before the Sieve Element Continuous Salivation Phase.

The distinguished intracellular stylet puncture called phloem-pd (potential drop [pd]) produced by Myzus persicae has been associated with the transmission of the semipersistently transmitted, phloem-limited Beet yellows virus (BYV, Closterovirus). However, the production of intracellular punctures in phloem cells (phloem-pd) by other aphid species and their role in the transmission of persistently transmitted, phloem-limited viruses are still unknown. Previous studies revealed that inoculation of the persistently transmitted, phloem-limited Barley yellow dwarf virus (BYDV, Luteovirus) is associated mainly with the sieve element continuous salivation phase (E1 waveform). However, the role of brief intracellular punctures that occur before the E1 phase in the inoculation of BYDV by aphids is unknown. We aimed to investigate whether the bird cherry-oat aphid Rhopalosiphum padi (Hemiptera: Aphididae) produced a stereotypical phloem-pd and to study its role in the inoculation of BYDV. The feeding behavior of viruliferous R. padi individuals in barley (Hordeum vulgare) was monitored via the electrical penetration graph (EPG) technique. The feeding process was artificially terminated after the observation of specific EPG waveforms: standard-pds, phloem-pd, and E1. Analysis of the EPG recordings revealed the production of a phloem-pd pattern by R. padi, in addition to a short, distinct E1-like pattern (short-E1), both resulting in successful inoculation of BYDV. Also, the transmission efficiency of BYDV was directly proportional to the time spent by aphids in intracellular salivation in phloem cells. Finally, we discussed the main differences between the inoculation process of semipersistent and persistently transmitted phloem-limited viruses by aphids.

Assessing the Impact on Virus Transmission and Insect Vector Behavior of a Viral Mixed Infection in Melon.

Mixed viral infections in plants are common, and can result in synergistic or antagonistic interactions. Except in complex diseases with severe symptoms, mixed infections frequently remain unnoticed, and their impact on insect vector transmission is largely unknown. In this study, we considered mixed infections of two unrelated viruses commonly found in melon plants, the crinivirus cucurbit yellow stunting disorder virus (CYSDV) and the potyvirus watermelon mosaic virus (WMV), and evaluated their vector transmission by whiteflies and aphids, respectively. Their dynamics of accumulation was analyzed until 60 days postinoculation (dpi) in mixed-infected plants, documenting reduced titers of WMV and much higher titers of CYSDV compared with single infections. At 24 dpi, corresponding to the peak of CYSDV accumulation, similar whitefly transmission rates were obtained when comparing either individual or mixed-infected plants as CYSDV sources, although its secondary dissemination was slightly biased toward plants previously infected with WMV, regardless of the source plant. However, at later time points, mixed-infected plants partially recovered from the initially severe symptoms, and CYSDV transmission became significantly higher. Interestingly, aphid transmission rates both at early and late time points were unaltered when WMV was acquired from mixed-infected plants despite its reduced accumulation. This lack of correlation between WMV accumulation and transmission could result from compensatory effects observed in the analysis of the aphid feeding behavior by electrical penetration graphs. Thus, our results showed that mixed-infected plants could provide advantages for both viruses, directly favoring CYSDV dissemination while maintaining WMV transmission.

Newly Distinguished Cell Punctures Associated with Transmission of the Semipersistent Phloem-Limited Beet Yellows Virus.

Here we report on plant penetration activities (probing) by the aphid Myzus persicae (Sulzer, 1776) in association with the transmission, acquisition, and inoculation of the semipersistent Beet yellows virus (BYV; Closterovirus) in sugar beet. During electrical penetration graph (EPG) recording of stylet pathways, standard intracellular stylet punctures occur which are called potential drop (pd) waveforms. In addition to the standard pd, there also appeared to be a unique type of intracellular stylet puncture that always preceded the phloem salivation phase (waveform E1). This type of pd, the phloem-pd, showed properties distinct from those of the standard pds and has never been described before. We manually ended EPG recordings during the acquisition and inoculation tests by removing aphids from the source or test plant after specific waveforms were recorded. Inoculation of BYV occurred at the highest rate when probing was interrupted just after a single or various phloem-pds. In contrast, BYV acquisition showed an intimate association with sustained phloem sap ingestion from phloem sieve elements (SEs) (E2 waveform). Our work shows for the first time that the inoculation of a phloem-limited virus occurs during specific intracellular stylet punctures and before phloem salivation (waveform E1). Further studies are needed to establish in what cells this novel phloem-pd occurs: phloem parenchyma, companion, or SE cells. The role of the different stylet activities in the acquisition and inoculation of BYV by M. persicae is discussed.IMPORTANCE We discovered the specific feeding activities of Myzus persicae (Sulzer, 1776) associated with the transmission of Beet yellows virus (BYV; Closterovirus). Our work strongly suggests that aphids can insert their stylets into the membranes of phloem cells-visualized as a unique type of waveform that is associated with the inoculation of BYV. This intracellular puncture (3 to 5 s) occurs just before the phloem salivation phase and can be distinguished from other nonvascular stylet cell punctures. This is the first time that the transmission of a phloem-limited semipersistent virus has been shown to be associated with a unique type of intracellular puncture. Our work offers novel information and strongly contributes to the existing literature on the transmission of plant viruses. Here we describe a new kind of aphid behavioral pattern that could be key in further works, such as studying the transmission of other phloem-limited viruses (e.g., luteoviruses).

Identification of Plant Virus Receptor Candidates in the Stylets of Their Aphid Vectors.

Plant viruses transmitted by insects cause tremendous losses in most important crops around the world. The identification of receptors of plant viruses within their insect vectors is a key challenge to understanding the mechanisms of transmission and offers an avenue for future alternative control strategies to limit viral spread. We here report the identification of two cuticular proteins within aphid mouthparts, and we provide experimental support for the role of one of them in the transmission of a noncirculative virus. These two proteins, named Stylin-01 and Stylin-02, belong to the RR-1 cuticular protein subfamily and are highly conserved among aphid species. Using an immunolabeling approach, they were localized in the maxillary stylets of the pea aphid Acyrthosiphon pisum and the green peach aphid Myzus persicae, in the acrostyle, an organ earlier shown to harbor receptors of a noncirculative virus. A peptide motif present at the C termini of both Stylin-01 and Stylin-02 is readily accessible all over the surface of the acrostyle. Competition for in vitro binding to the acrostyle was observed between an antibody targeting this peptide and the helper component protein P2 of Cauliflower mosaic virus Furthermore, silencing the stylin-01 but not stylin-02 gene through RNA interference decreased the efficiency of Cauliflower mosaic virus transmission by Myzus persicae These results identify the first cuticular proteins ever reported within arthropod mouthparts and distinguish Stylin-01 as the best candidate receptor for the aphid transmission of noncirculative plant viruses.IMPORTANCE Most noncirculative plant viruses transmitted by insect vectors bind to their mouthparts. They are acquired and inoculated within seconds when insects hop from plant to plant. The receptors involved remain totally elusive due to a long-standing technical bottleneck in working with insect cuticle. Here we characterize the role of the two first cuticular proteins ever identified in arthropod mouthparts. A domain of these proteins is directly accessible at the surface of the cuticle of the acrostyle, an organ at the tip of aphid stylets. The acrostyle has been shown to bind a plant virus, and we consistently demonstrated that one of the identified proteins is involved in viral transmission. Our findings provide an approach to identify proteins in insect mouthparts and point at an unprecedented gene candidate for a plant virus receptor.

Fasting alters aphid probing behaviour but does not universally increase the transmission rate of non-circulative viruses.

A fasting period prior to non-circulative virus acquisition has been shown to increase the rate of transmission by aphids. However, this effect has only been studied for a few virus-vector combinations, and there are contradictory results in the literature as to the role of fasting on virus acquisition. We analysed the influence of fasting on the transmission of three non-circulative viruses, Cucumber mosaic virus, Zucchini yellow mosaic virus and Cauliflower mosaic virus, by two aphid vector species: Myzus persicae Sulzer (Hemiptera: Aphididae) and Aphis gossypii Glover (Hemiptera: Aphididae). All variables tested, including the virus species and isolate, and the species of aphid, influenced the effect of a fasting period on virus transmission efficiency. Furthermore, when aphids were subjected to an overnight feeding period on a sucrose solution, the fasting effect disappeared and the probing behaviour of these aphids was markedly different to plant-reared aphids. The electrical penetration graph (EPG) technique revealed that fasting altered the probing behaviour of M. persicae and A. gossypii, with fasted aphids beginning to feed sooner and having a significantly longer first intracellular puncture, measured as a potential drop. Significantly longer sub-phase II-3 of the potential drop and more archlets during this sub-phase were also observed for fasted aphids of both species. However, these behavioural changes were not predictive of increasing virus transmission following a fasting period. The impacts of pre-acquisition fasting on aphid probing behaviour and on the mechanisms of non-circulative virus transmission are discussed.

Stylet penetration activities of the whitefly Bemisia tabaci associated with inoculation of the crinivirus Tomato chlorosis virus.

Bemisiatabaci is an important vector of numerous plant viruses, including the emergent semi-persistently transmitted crinivirus Tomato chlorosis virus (ToCV). Its vector feeding behaviour is complex, with important implications for virus transmission, epidemiology and control. Thus, the objective of this study was to investigate the role of the stylet penetration activities of B. tabaci in the inoculation of ToCV in tomatoes by using the electrical penetration graph (EPG) technique. EPG recordings were classified into six categories depending on the waveforms observed. The results showed that ToCV inoculation is mainly associated with stylet activities in phloem sieve elements (E1 waveform), as there was a significant increase in the rate of transmission when whiteflies performed waveform E1. The precise stylet activities - either salivation or egestion - associated with virion release, presumably from retention sites in the foregut, need further investigation.

The relationship between host lifespan and pathogen reservoir potential: an analysis in the system Arabidopsis thaliana--cucumber mosaic virus.

Identification of the determinants of pathogen reservoir potential is central to understand disease emergence. It has been proposed that host lifespan is one such determinant: short-lived hosts will invest less in costly defenses against pathogens, so that they will be more susceptible to infection, more competent as sources of infection and/or will sustain larger vector populations, thus being effective reservoirs for the infection of long-lived hosts. This hypothesis is sustained by analyses of different hosts of multihost pathogens, but not of different genotypes of the same host species. Here we examined this hypothesis by comparing two genotypes of the plant Arabidopsis thaliana that differ largely both in life-span and in tolerance to its natural pathogen Cucumber mosaic virus (CMV). Experiments with the aphid vector Myzus persicae showed that both genotypes were similarly competent as sources for virus transmission, but the short-lived genotype was more susceptible to infection and was able to sustain larger vector populations. To explore how differences in defense against CMV and its vector relate to reservoir potential, we developed a model that was run for a set of experimentally-determined parameters, and for a realistic range of host plant and vector population densities. Model simulations showed that the less efficient defenses of the short-lived genotype resulted in higher reservoir potential, which in heterogeneous host populations may be balanced by the longer infectious period of the long-lived genotype. This balance was modulated by the demography of both host and vector populations, and by the genetic composition of the host population. Thus, within-species genetic diversity for lifespan and defenses against pathogens will result in polymorphisms for pathogen reservoir potential, which will condition within-population infection dynamics. These results are relevant for a better understanding of host-pathogen co-evolution, and of the dynamics of pathogen emergence.

A Differential Role of Volatiles from Conspecific and Heterospecific Competitors in the Selection of Oviposition Sites by the Aphidophagous Hoverfly Sphaerophoria rueppellii.

The selection of oviposition sites by syrphids and other aphidophagous insects is influenced by the presence of con- and heterospecific competitors. Chemical cues play a role in this selection process, some of them being volatile semiochemicals. Yet, little is known about the identity and specificity of chemical signals that are involved in the searching behavior of these predators. In this study, we used olfactometer bioassays to explore the olfactory responses of gravid females and larvae of the syrphid Sphaerophoria rueppellii, focussing on volatiles from conspecific immature stages, as well as odors from immature stages of the competing coccinellid Adalia bipunctata. In addition, a multiple-choice oviposition experiment was conducted to study if females respond differently when they can also sense their competitors through visual or tactile cues. Results showed that volatiles from plants and aphids did not affect the behavior of second-instars, whereas adult females strongly preferred odors from aphid colonies without competitors. Odors from conspecific immature stages had a repellent effect on S. rueppellii adult females, whereas their choices were not affected by volatiles coming from immature heterospecific A. bipunctata. The results imply that the syrphid uses odors to avoid sites that are already occupied by conspecifics. They did not avoid the odor of the heterospecific competitor, although in close vicinity they were found to avoid laying eggs on leaves that had traces of the coccinellid. Apparently adult syrphids do not rely greatly on volatile semiochemicals to detect the coccinellid, but rather use other stimuli at close range (e. g., visual or non-volatile compounds) to avoid this competitor.

Resistance to Cucurbit aphid-borne yellows virus in Melon Accession TGR-1551.

The genetic control of resistance to Cucurbit aphid-borne yellows virus (CABYV; genus Polerovirus, family Luteoviridae) in the TGR-1551 melon accession was studied through agroinoculation of a genetic family obtained from the cross between this accession and the susceptible Spanish cultivar 'Bola de Oro'. Segregation analyses were consistent with the hypothesis that one dominant gene and at least two more modifier genes confer resistance; one of these additional genes is likely present in the susceptible parent 'Bola de Oro'. Local and systemic accumulation of the virus was analyzed in a time course experiment, showing that TGR-1551 resistance was expressed systemically as a significant reduction of virus accumulation compared with susceptible controls, but not locally in agroinoculated cotyledons. In aphid transmission experiments, CABYV inoculation by aphids was significantly reduced in TGR-1551 plants, although the virus was acquired at a similar rate from TGR-1551 as from susceptible plants. Results of feeding behavior studies using the DC electrical penetration graph technique suggested that viruliferous aphids can salivate and feed from the phloem of TGR-1551 plants and that the observed reduction in virus transmission efficiency is not related to reduced salivation by Aphis gossypii in phloem sieve elements. Since the virus is able to accumulate to normal levels in agroinoculated tissues, our results suggest that resistance of TGR-1551 plants to CABYV is related to impairment of virus movement or translocation after it reaches the phloem sieve elements.

Are Mummies and Adults of Eretmocerus mundus (Hymenoptera: Aphelinidae) Compatible With Modern Insecticides?

The parasitic wasp Eretmocerus mundus (Mercet) is an important natural enemy of the widespread key pest Bemisia tabaci (Gennadius). The toxicity of 11 modern insecticides applied at their maximum field recommended rate in Spain was tested in two life stages of E. mundus: adults and mummies. Laboratory and persistence tests were conducted and effects assessed not only in terms of mortality but on reproductive performance as well. Some insecticides caused the same effect to both life stages: flubendiamide, methoxyfenozide, spiromesifen, and flonicamid were harmless, while spinosad and sulfoxaflor were harmful. The last two insecticides cannot be used jointly with E. mundus under any condition because they were highly persistent. The rest of tested insecticides (spirotetramat, metaflumizone, deltamethrin, abamectin, and emamectin) caused some direct mortality to one or both life stages and/or affected reproduction of the parasitic wasp and their harmful effect in the field ranged from short lived (metaflumizone and spirotetramat) to slightly persistent (emamectin) and to moderately persistent (abamectin and deltamethrin). Therefore, they could be recommended for use in integrated pest management programs together with the natural enemy if appropriate safety intervals after insecticide application are observed.

Circulating virus load determines the size of bottlenecks in viral populations progressing within a host.

For any organism, population size, and fluctuations thereof, are of primary importance in determining the forces driving its evolution. This is particularly true for viruses--rapidly evolving entities that form populations with transient and explosive expansions alternating with phases of migration, resulting in strong population bottlenecks and associated founder effects that increase genetic drift. A typical illustration of this pattern is the progression of viral disease within a eukaryotic host, where such demographic fluctuations are a key factor in the emergence of new variants with altered virulence. Viruses initiate replication in one or only a few infection foci, then move through the vasculature to seed secondary infection sites and so invade distant organs and tissues. Founder effects during this within-host colonization might depend on the concentration of infectious units accumulating and circulating in the vasculature, as this represents the infection dose reaching new organs or "territories". Surprisingly, whether or not the easily measurable circulating (plasma) virus load directly drives the size of population bottlenecks during host colonization has not been documented in animal viruses, while in plants the virus load within the sap has never been estimated. Here, we address this important question by monitoring both the virus concentration flowing in host plant sap, and the number of viral genomes founding the population in each successive new leaf. Our results clearly indicate that the concentration of circulating viruses directly determines the size of bottlenecks, which hence controls founder effects and effective population size during disease progression within a host.

The African Psyllid Trioza erytreae Del Guercio (1918) Is Very Sensitive to Low Relative Humidity and High Temperatures.

The African citrus psyllid, Trioza erytreae, is one of the two vectors of Huanglongbing, the most serious citrus disease worldwide. The first detection of T. erytreae in the European mainland was on the northwest of the Iberian Peninsula in 2014. Since then, the pest has spread throughout northern Spain (Galicia, Asturias, Cantabria, País Vasco) and along the western Atlantic coast of Portugal (from the Douro e Minho region to the Algarve). We conducted a series of laboratory experiments on lemon plants at different temperatures (from 8 to 34 °C) and humidity conditions (from 40 to 90%) to find out the influence of extreme temperatures and relative humidities (RHs) on the mortality, development and reproduction of T. erytreae. Our results show that temperatures above 30 °C and below 10 °C are very detrimental for nymphal development and nymphs were unable to reach the adult stage. Furthermore, eggs were unable to hatch under temperatures above 33 °C and below 8 °C. Adult mortality was highest at 34 °C and killed more than 50% of the population. We also found that relative humidity is crucial for the development and survival of T. erytreae. Nymphs were unable to reach the adult stage at an RH of 90% and 40%. Also, fecundity was significantly reduced at 90 and 40% RH, and fertility was lowest at 40% RH. Nymphal mortality was highest at an RH of 40%, which was the most detrimental humidity among all tested for the survival and development of T. erytreae. Our work concludes that T. erytreae establishment and spread will be maximum in regions with a temperate and humid climate, being rare in regions where dry and hot weather conditions predominate.

Aphidius colemani Behavior Changes Depending on Volatile Organic Compounds Emitted by Plants Infected with Viruses with Different Modes of Transmission.

Natural enemies are an additional component that may interact directly with the plant-virus-vector association, affecting viral dispersion. In our study, we conducted olfactometry assays to explore how single and mixed infections with CMV or/and CABYV modify the attractiveness of A. colemani to aphid-free and aphid-infested melon plants using two melon genotypes. Subsequently, we investigated the influence of CABYV-infected plants infested by A. gossypii on the parasitism rate and emergence of A. colemani in a dual-choice assay under greenhouse conditions. Our study demonstrates that males showed no preference for either infected or non-infected plants. Female parasitoids exhibit a preference for volatiles emitted by CMV and mixed-infected melon plants over clean air but not over mock-inoculated plants, suggesting a response influenced by plant genotype. Female parasitoid responses to CABYV and its interactions with aphids revealed a preference for mock-inoculated plants over CABYV-infected plants and a parasitism rate slightly higher (7.12%) on non-infected plants. Our study revealed that (1) parasitoids may reject olfactory cues from CABYV-infected plants, potentially interfering with the plant's "cry for help" response; (2) in the case of CMV, whether in single or mixed infections, non-infected plants are as attractive as infected ones to parasitoids. Our findings suggest that persistent viruses manipulate aphid parasitoid behavior to their advantage, promoting virus disease in melon crops.