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BACKGROUND Pseudocercospora fijiensis is the causal agent of the black leaf streak disease (BLSD) of banana. Bananas are important global export commodities and a major staple food. Their susceptibility to BLSD pushes disease management towards excessive fungicide use, largely relying on multisite inhibitors and sterol demethylation inhibitors (DMIs). These fungicides are ubiquitous in plant disease control, targeting the CYP51 enzyme. We examined sensitivity to DMIs in P. fijiensis field isolates collected from various major banana production zones in Colombia, Costa Rica, Dominican Republic, Ecuador, the Philippines, Guadalupe, Martinique and Cameroon and determined the underlying genetic reasons for the observed phenotypes. RESULTS We observed a continuous range of sensitivity towards the DMI fungicides difenoconazole, epoxiconazole and propiconazole with clear cross‐sensitivity. Sequence analyses of PfCYP51 in 266 isolates showed 28 independent amino acid substitutions, nine of which correlated with reduced sensitivity to DMIs. In addition to the mutations, we observed up to six insertions in the Pfcyp51 promoter. Such promoter insertions contain repeated elements with a palindromic core and correlate with the enhanced expression of Pfcyp51 and hence with reduced DMI sensitivity. Wild‐type isolates from unsprayed bananas fields did not contain any promoter insertions. CONCLUSION The presented data significantly contribute to understanding of the evolution and global distribution of DMI resistance mechanisms in P. fijiensis field populations and facilitate the prediction of different DMI efficacy. The overall reduced DMI sensitivity calls for the deployment of a wider range of solutions for sustainable control of this major banana disease. © 2021 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry., Studies on Pseudocercospora fijiensis demethylation inhibitor (DMI) sensitivity shows that Pfcyp51 modulations correlated with fungicide resistance. The importance of amino acid substitutions at positions 137, 311, 378, 379 and 458–461 is strengthened by mathematical modelling. Changes in these positions compromise the three‐dimensional structure of the protein resulting in an affinity change. The presence of repeated elements and insertions in the promotor region of Pfcyp51 was also positively correlated with resistance to DMIs. © 2021 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

International audience; Plants are colonized by diverse fungal and viral communities that influence their growth and survival as well as ecosystem functioning. Viruses interact with both plants and the fungi they host. Our understanding of plant–fungi–virus interactions is very limited, especially in wild plants. Combining metagenomic and culturomic approaches, we assessed the richness, diversity, and composition of leaf‐associated fungal and viral communities from pools of herbaceous wild plants representative of four sites corresponding to cultivated or natural ecosystems. We identified 161 fungal families and 18 viral families comprising 249 RNA‐dependent RNA polymerase‐based operational taxonomic units (RdRp OTUs) from leaves. Fungal culturomics captured 12.3% of the fungal diversity recovered with metagenomic approaches and, unexpectedly, retrieved viral OTUs that were almost entirely different from those recovered by leaf metagenomics. Ecosystem management had a significant influence on both leaf mycobiome and virome, with a higher fungal community richness in natural ecosystems and a higher viral family richness in cultivated ecosystems, suggesting that leaf‐associated fungal and viral communities are under the influence of different ecological drivers. Both the leaf‐associated fungal and viral community compositions showed a strong site‐specificity. Further research is needed to confirm these trends and unravel the factors structuring plant–fungi–virus interactions in wild plant populations.

International audience; In this study, we report the complete genome sequence of a novel luteovirus detected in almonds using high-throughput sequencing. The genome of the new luteovirus comprises 5,047 nucleotides, and its genomic organization is similar to that of the recently described nectarine stem pitting associated virus (NSPaV), with only four open reading frames encoding for replication related proteins, the coat protein (CP) and a CP readthrough protein involved in the aphid transmission of luteovirids. Phylogenic and pairwise distance analyses showed that this virus shares 79% and 57.8% amino acid identities in the P1-P2 fusion protein and the P3-P5 protein, respectively, with the most closely related luteovirus NSPaV, suggesting that it represents a novel species for which the name of Almond associated luteovirus 1 is proposed. To our knowledge, this is the first report of an almond-infecting luteovirus.

International audience; Many recessive resistances against potyviruses are mediated by eukaryotic translation initiation factor 4E (eIF4E). In tobacco, the va resistance gene commonly used to control Potato virus Y (PVY) corresponds to a large deletion affecting the eIF4E-1 gene on chromosome 21. Here, we compared the resistance durability conferred by various types of mutations affecting eIF4E-1 (deletions of various sizes, frameshift or nonsense mutations). The 'large deletion' genotypes displayed the broadest and most durable resistance, whereas frameshift and nonsense mutants displayed a less durable resistance, with rapid and frequent apparition of resistance-breaking variants. In addition, genetic and transcriptomic analyses revealed that resistance durability is strongly impacted by a complex genetic locus on chromosome 14, which contains three other eIF4E genes. One of these, eIF4E-3, is rearranged as a hybrid gene between eIF4E-2 and eIF4E-3 (eIF4E-2-3 ) in the genotypes showing the most durable resistance, while eIF4E-2 is differentially expressed between the tested varieties. RNA-seq and quantitative reverse transcriptase-polymerase chain reaction experiments demonstrated that eIF4E-2 expression level is positively correlated with resistance durability. These results suggest that besides the nature of the mutation affecting eIF4E-1, three factors linked with a complex locus may potentially impact va durability: loss of an integral eIF4E-3, presence of eIF4E-2-3 and overexpression of eIF4E-2. This latter gene might act as a decoy in a non-productive virus-plant interaction, limiting the ability of PVY to evolve towards resistance breaking. Taken together, these results show that va resistance durability can in large part be explained by complex redundancy effects in the eIF4E gene family.

The aim of this study was to develop a set of experiments to screen and decipher the mechanisms of biocontrol agents (BCAs), isolated from commercial formulation, against two major mycotoxigenic fungi in cereals, Fusarium graminearum and Fusarium verticillioides. These two phytopathogens produce mycotoxins harmful to human and animal health and are responsible for the massive use of pesticides, for the protection of cereals. It is therefore essential to better understand the mechanisms of action of alternative control strategies such as the use of BCAs in order to optimize their applications. The early and late stages of interaction between BCAs and pathogens were investigated from germination of spores to the effects on perithecia (survival form of pathogen). The analysis of antagonist activities of BCAs revealed different strategies of biocontrol where chronological, process combination and specialization aspects of interactions are discussed. Streptomyces griseoviridis main strategy is based on antibiosis with the secretion of several compounds with anti-fungal and anti-germination activity, but also a mixture of hydrolytic enzymes to attack pathogens, which compensates for an important deficit in terms of spatial colonization capacity. It has good abilities in terms of nutritional competition. Trichoderma asperellum is capable of activating a very wide range of defenses and attacks combining the synthesis of various antifungal compounds (metabolite, enzymes, VOCs), with different targets (spores, mycelium, mycotoxins), and direct action by mycoparasitism and mycophagy. Concerning Pythium oligandrum, its efficiency is mainly due to its strong capacity to colonize the environment, with a direct action via microbial predation, stimulation of its reproduction at the contact of pathogens and the reduction of perithecia formation.

Phytoplasmas inhabit phloem sieve elements and cause abnormal growth and altered sugar partitioning. However, how they interact with phloem functions is not clearly known. The phloem responses were investigated in tomatoes infected by “Candidatus Phytoplasma solani” at the beginning of the symptomatic stage, the first symptoms appearing in the newly emerged leaf at the stem apex. Antisense lines impaired in the phloem sucrose transporters SUT1 and SUT2 were included. In symptomatic sink leaves, leaf curling was associated with higher starch accumulation and the expression of defense genes. The analysis of leaf midribs of symptomatic leaves indicated that transcript levels for genes acting in the glycolysis and peroxisome metabolism differed from these in noninfected plants. The phytoplasma also multiplied in the three lower source leaves, even if it was not associated with the symptoms. In these leaves, the rate of phloem sucrose exudation was lower for infected plants. Metabolite profiling of phloem sap-enriched exudates revealed that glycolate and aspartate levels were affected by the infection. Their levels were also affected in the noninfected SUT1- and SUT2-antisense lines. The findings suggest the role of sugar transporters in the responses to infection and describe the consequences of impaired sugar transport on the primary metabolism.

While the focus of plant virology has been mainly on horticultural and field crops as well as fruit trees, little information is available on viruses that infect forest trees. Utilization of next-generation sequencing (NGS) methodologies has revealed a significant number of viruses in forest trees and urban parks. In the present study, the full-length genome of a novel Emaravirus has been identified and characterized from sycamore maple (Acer pseudoplatanus) – a tree species of significant importance in urban and forest areas – showing leaf mottle symptoms. RNA-Seq was performed on the Illumina HiSeq2500 system using RNA preparations from a symptomatic and a symptomless maple tree. The sequence assembly and analysis revealed the presence of six genomic RNA segments in the symptomatic sample (RNA1: 7,074 nt-long encoding the viral replicase; RNA2: 2,289 nt-long encoding the glycoprotein precursor; RNA3: 1,525 nt-long encoding the nucleocapsid protein; RNA4: 1,533 nt-long encoding the putative movement protein; RNA5: 1,825 nt-long encoding a hypothetical protein P5; RNA6: 1,179 nt-long encoding a hypothetical protein P6). Two independent NGS sequencing runs from the same symptomatic maple tree detected the same genome segments. For one of these sequencing runs the cDNA library was prepared using a primer targeting the conserved genome terminal region, known to be shared between emaraviruses genome segments. We suggest, therefore, that the six identified genome segments represent the complete genome of a novel emaravirus from maple, which we tentatively name maple mottle-associated virus (MaMaV). Phylogenetic and sequence homology analyses place this virus on the distinct “subgroup a” clade within the Emaravirus genus along with – among others – rose rosette virus, Actinidia emaravirus 2, and fig mosaic virus. Validation RT-PCR assays performed on symptomatic and non-symptomatic trees suggest that MaMaV may be the symptom-inducing virus in the diseased trees. To our knowledge, this is the first time an Emaravirus is described from maple and is fully genetically characterized. With the discovery of MaMaV, the genus Emaravirus comprising negative-sense single-stranded viruses with very divergent genomes – that were until recently overlooked – has substantially increased counting 22 established and putative members.

Alfalfa (Medicago sativa L.), also known as lucerne, is a major forage crop worldwide. In the United States, it has recently become the third most valuable field crop, with an estimated value of over $9.3 billion. Alfalfa is naturally infected by many different pathogens, including viruses, obligate parasites that reproduce only inside living host cells. Traditionally, viral infections of alfalfa have been considered by breeders, growers, producers and researchers to be diseases of limited importance, although they are widespread in all major cultivation areas. However, over the past few years, due to the rapid development of high-throughput sequencing (HTS), viral metagenomics, bioinformatics tools for interpreting massive amounts of HTS data and the increasing accessibility of public data repositories for transcriptomic discoveries, several emerging viruses of alfalfa with the potential to cause serious yield losses have been described. They include alfalfa leaf curl virus (family Geminiviridae), alfalfa dwarf virus (family Rhabdoviridae), alfalfa enamovirus 1 (family Luteoviridae), alfalfa virus S (family Alphaflexiviridae) and others. These discoveries have called into question the assumed low economic impact of viral diseases in alfalfa and further suggested their possible contribution to the severity of complex infections involving multiple pathogens. In this review, we will focus on viruses of alfalfa recently described in different laboratories on the basis of the above research methodologies. Fil: Bejerman, Nicolas. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigaciones Agropecuarias. Unidad de Fitopatología y Modelización Agrícola. Grupo Vinculado Catedra de Estadística y Biometría de la Facultad de Ciencias Agropecuarias de la Universidad Nacional de Córdoba al Ufyma | Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Unidad de Fitopatología y Modelización Agrícola. Grupo Vinculado Catedra de Estadística y Biometría de la Facultad de Ciencias Agropecuarias de la Universidad Nacional de Córdoba al Ufyma; Argentina Fil: Roumagnac, Philippe. Université Montpellier II; Francia Fil: Nemchinov, Lev G.. Fundación para el Estudio de Especies Invasivas; Argentina

To cause disease, diverse pathogens deliver effector proteins into host cells. Pathogen effectors can inhibit defense responses, alter host physiology, and represent important cellular probes to investigate plant biology. However, effector function and localization have primarily been investigated after overexpression in planta. Visualizing effector delivery during infection is challenging due to the plant cell wall, autofluorescence, and low effector abundance. Here, we used a GFP strand system to directly visualize bacterial effectors delivered into plant cells through the type III secretion system. GFP is a beta barrel that can be divided into 11 strands. We generated transgenic Arabidopsis thaliana plants expressing GFP1-10 (strands 1 to 10). Multiple bacterial effectors tagged with the complementary strand 11 epitope retained their biological function in Arabidopsis and tomato (Solanum lycopersicum). Infection of plants expressing GFP1-10 with bacteria delivering GFP11-tagged effectors enabled direct effector detection in planta. We investigated the temporal and spatial delivery of GFP11-tagged effectors during infection with the foliar pathogen Pseudomonas syringae and the vascular pathogen Ralstonia solanacearum. Thus, the GFP strand system can be broadly used to investigate effector biology in planta.

UMR BFP - Equipe Virologie; International audience; We developed an innovative RNAi concept based on two gene constructs built from the capsid gene (CP) cistron of the Plum pox virus (PPV) genome. First, designated as amiCPRNA, a potential molecule interfering with PPV genome translation and the second one is the ami-siCPRNA to target viral genome translation and PPV RNA replication. Following the previous engineering of these constructs in an experimental herbaceous host, they were introduced into Prunus domestica (plum tree) genome. Previously propagated onto a susceptible rootstock, these clones were graft-inoculated with PPV. After four dormancy cycles, and consistent with our experience of PPV infection, some clones showed a common phenomenon of silencing that can differ between the detailed plant phenotypes. Three different phenotypes were developed by the amisiCPRNA clones. First, the high resistance character shown by the amisiCPRNA plum-7 that was similar to the resistance expressed by HoneySweet plum. Secondly, a recovery reaction was developed by the two other amisiCPRNA plum-3 and plum-4 that differed from the rest, characterized as susceptible clones, among these were the amiCPRNA plums. Having assessed the behavior of these plums versus the herbaceous host accumulating the similar form of RNAi: ami-, si-, and ami-siRNA, challenging assays in perennials consistently reflect the natural context of viral genome targeting

Yam (Dioscorea spp.) is an important crop in tropical and subtropical regions. Many viruses have been recently identified in yam, hampering genetic conservation and safe international exchanges of yam germplasm. We report on the implementation of reliable and cost-effective PCR-based detection tools targeting eight different yam-infecting viruses. Viral indexing of the in vitro yam collection maintained by the Biological Resources Center for Tropical Plants (BRC-TP) in Guadeloupe (French West Indies) unveiled a high prevalence of potyviruses, badnaviruses, Dioscorea mosaic associated virus (DMaV) and yam asymptomatic virus 1 (YaV1) and a high level of coinfections. Infected yam accessions were subjected to a combination of thermotherapy and meristem culture. Sanitation levels were monitored using PCR-based and high-throughput sequencing-based diagnosis, confirming the efficacy and reliability of PCR-based detection tools. Sanitation rates were highly variable depending on viruses. Sixteen accessions were successfully sanitized, paving the way to safe yam germplasm exchanges and the implementation of clean seed production programs worldwide.

International audience; The management of genes conferring resistance to plant–pathogens should make it possible to control epidemics (epidemiological perspective) and preserve resistance durability (evolutionary perspective). Resistant and susceptible cultivars must be strategically associated according to the principles of cultivar mixture (within a season) and rotation (between seasons). We explored these questions by modeling the evolutionary and epidemiological processes shaping the dynamics of a pathogen population in a landscape composed of a seasonal cultivated compartment and a reservoir compartment hosting pathogen year-round. Optimal deployment strategies depended mostly on the molecular basis of plant–pathogen interactions and on the agro-ecological context before resistance deployment, particularly epidemic intensity and landscape connectivity. Mixtures were much more efficient in landscapes in which between-field infections and infections originating from the reservoir were more prevalent than within-field infections. Resistance genes requiring two mutations of the pathogen avirulence gene to be broken down, rather than one, were particularly useful when infections from the reservoir predominated. Combining mixture and rotation principles were better than the use of the same mixture each season as (i) they controlled epidemics more effectively in situations in which within-field infections or infections from the reservoir were frequent and (ii) they fulfilled the epidemiological and evolutionary perspectives.

During their lifetime, perennial woody plants are expected to face multiple infection events. Furthermore, multiple genotypes of individual virus species may co-infect the same host. This may eventually lead to a situation where plants harbor complex communities of viral species/strains. Using high-throughput sequencing, we describe co-infection of sweet and sour cherry trees with diverse genomic variants of two closely related viruses, namely prunus virus F (PrVF) and cherry virus F (CVF). Both viruses are most homologous to members of theFabavirusgenus (Secoviridaefamily). The comparison of CVF and PrVF RNA2 genomic sequences suggests that the two viruses may significantly differ in their expression strategy. Indeed, similar to comoviruses, the smaller genomic segment of PrVF, RNA2, may be translated in two collinear proteins while CVF likely expresses only the shorter of these two proteins. Linked with the observation that identity levels between the coat proteins of these two viruses are significantly below the family species demarcation cut-off, these findings support the idea that CVF and PrVF represent two separateFabavirusspecies.

The cAMP-dependent transcriptional regulator Clr of Sinorhizobium meliloti regulates the overall number of infection events on Medicago roots by a so-far unknown mechanism requiring smc02178, a Clr-target gene of unknown function. In order to shed light on the mode of action of Clr on infection and potentially reveal additional biological functions for Clr, we inventoried genomic Clr target genes by transcriptome profiling. We have found that Clr positively controls the synthesis of cAMP-dependent succinoglycan as well as the expression of genes involved in the synthesis of a so-far unknown polysaccharide compound. In addition, Clr activated expression of 24 genes of unknown function in addition to smc02178. Genes negatively controlled by Clr were mainly involved in swimming motility and chemotaxis. Functional characterization of two novel Clr-activated genes of unknown function, smb20495 and smc02177, showed that their expression was activated by the same plant signal as smc02178 ex planta. In planta, however, symbiotic expression of smc02177 proved independent of clr. Both smc02177 and smb20495 genes were strictly required for the control of secondary infection on M. sativa. None of the three smc02177, smc02178 and smb20495 genes were needed for plant signal perception. Altogether this work provides a refined view of the cAMP-dependent Clr regulon of S. meliloti. We specifically discuss the possible roles of smc02177, smc02178, smb20495 genes and other Clr-controlled genes in the control of secondary infection of Medicago roots.

International audience; Virus host range, i.e., the number and diversity of host species of viruses, is an important determinant of disease emergence and of the efficiency of disease control strategies. However, for plant viruses, little is known about the genetic or ecological factors involved in the evolution of host range. Using available genome sequences and host range data, we performed a phylogenetic analysis of host range evolution in the genus Potyvirus, a large group of plant RNA viruses that has undergone a radiative evolution circa 7000 years ago, contemporaneously with agriculture intensification in mid Holocene. Maximum likelihood inference based on a set of 59 potyviruses and 38 plant species showed frequent host range changes during potyvirus evolution, with 4.6 changes per plant species on average, including 3.1 host gains and 1.5 host loss. These changes were quite recent, 74% of them being inferred on the terminal branches of the potyvirus tree. The most striking result was the high frequency of correlated host gains occurring repeatedly in different branches of the potyvirus tree, which raises the question of the dependence of the molecular and/or ecological mechanisms involved in adaptation to different plant species.

Induced plant defenses against either pathogens or herbivore attackers are regulated by phytohormones. These phytohormones are increasingly recognized as important mediators of interactions between organisms associated with plants. In this review, we discuss the role of plant defense hormones in sequential tri-partite interactions among plants, pathogenic microbes, and herbivorous insects, based on the most recent literature. We discuss the importance of pathogen trophic strategy in the interaction with herbivores that exhibit different feeding modes. Plant resistance mechanisms also affect plant quality in future interactions with attackers. We discuss exemplary evidence for the hypotheses that (i) biotrophic pathogens can facilitate chewing herbivores, unless plants exhibit effector-triggered immunity, but (ii) facilitate or inhibit phloem feeders. (iii) Necrotrophic pathogens, on the other hand, can inhibit both phloem feeders and chewers. We also propose herbivore feeding mode as predictor of effects on pathogens of different trophic strategies, providing evidence for the hypotheses that (iv) phloem feeders inhibit pathogen attack by increasing SA induction, whereas (v) chewing herbivores tend not to affect necrotrophic pathogens, while they may either inhibit or facilitate biotrophic pathogens. Putting these hypotheses to the test will increase our understanding of phytohormonal regulation of plant defense to sequential attack by plant pathogens and insect herbivores. This will provide valuable insight into plant-mediated ecological interactions among members of the plant-associated community.

UMR BFP - Equipe Virologie; International audience; With the increased use of high-throughput sequencing methods, new viruses infecting Prunus spp. are being discovered and characterized, especially in the family Betaflexiviridae. Double-stranded RNAs from symptomatic leaves of a Japanese apricot (Prunus mume) tree from Japan were purified and analyzed by Illumina sequencing. Blast comparisons of reconstructed contigs showed that the P. mume sample was infected by a putative novel virus with homologies to Cherry virus A (CVA) and to the newly described Currant virus A (CuVA), both members of genus Capillovirus. Completion of the genome showed the new agent to have a genomic organization typical of capilloviruses, with two overlapping open reading frames encoding a large replication-associated protein fused to the coat protein (CP), and a putative movement protein (MP). This virus shares only, respectively, 63.2% and 62.7% CP amino acid identity with the most closely related viruses, CVA and CuVA. Considering the species demarcation criteria in the family and phylogenetic analyses, this virus should be considered as representing a new viral species in the genus Capillovirus, for which the name of Mume virus A is proposed

Moniliophthora perniciosa, the causal agent of witches’ broom disease of Theobroma cacao, significantly affected cacao production in South America and Caribbean countries. Host colonization by the pathogen exhibits a concerted succession of symptoms, starting with hypertrophic growth and ‘‘broom’’ formation, followed by tissue degeneration and death. To understand mechanisms of host susceptibility, we investigated fungal development during a compatible interaction with a susceptible genotype. Microscopic analysis revealed the initial fungal biotrophic intercellular growth, followed by intracellular growth associated with the presence of an increasing number of host apoptotic nuclei and calcium oxalate crystals, with subsequent accumulation of hydrogen peroxide and cell death. Active oxalate degradation and its possible source of origin were detected in infected tissues. Together, these processes may increase the availability of nutrients for the fungal mycelia and may contribute to the disease cycle in this plant–fungal hemibiotrophic interaction. Based on the histological and gene expression data, a novel role for calcium oxalate in disease susceptibility is proposed. # 2007 Elsevier Ireland Ltd. All rights reserved.

The hemibiotrophic basidiomycete Moniliophthora perniciosa causes witches' broom disease of Theobroma cacao. Analysis of the M. perniciosa draft genome led to the identification of three putative genes encoding necrosis and ethylene-inducing proteins (MpNEPs), which are apparently located on the same chromosome. MpNEP1 and 2 have highly similar sequences and are able to induce necrosis and ethylene emission in tobacco and cacao leaves. MpNEP1 is expressed in both biotrophic and saprotrophic mycelia, the protein be-haves as an oligomer in solution and is very sensitive to temperature. MpNEP2 is expressed mainly in biotrophic mycelia, is present as a monomer in solution at low concentrations (

Populations of Xanthomonas axonopodis pv. manihotis and X. axonopodis pv. vignicola, causal agents of cassava and cowpea bacterial blight, respectively, were quantified in insects. The pathogens were found in the faeces, the intestines, and on the legs and mandibles of Zonocerus variegatus. Additionally, X. axonopodis pv. manihotis was localized in the insect gut by immunofluorescence microscopy. Xanthomonas axonopodis pv. manihotis survived at least 1 week in the insect intestines and at least 5 weeks in faeces kept under controlled conditions, while survival in faeces exposed to sunlight was

International audience; After more than 70 years of chemical pesticide use, modern agriculture is increasingly using biological control products. Resistances to conventional insecticides are wide spread, while those to bio-insecticides have raised less attention, and resistance management is frequently neglected. However, a good knowledge of the limitations of a new technique often provides greater sustainability. In this review, we compile cases of resistance to widely used bio-insecticides and describe the associated resistance mechanisms. This overview shows that all widely used bio-insecticides ultimately select resistant individuals. For example, at least 27 species of insects have been described as resistant to Bacillus thuringiensis toxins. The resistance mechanisms are at least as diverse as those that are involved in resistance to chemical insecticides, some of them being common to bio-insecticides and chemical insecticides. This analysis highlights the specific properties of bio-insecticides that the scientific community should use to provide a better sustainability of these products.

Publication Inra prise en compte dans l'analyse bibliométrique des publications scientifiques mondiales sur les Fruits, les Légumes et la Pomme de terre. Période 2000-2012. http://prodinra.inra.fr/record/256699; International audience; Viruses frequently use insect vectors to effect rapid spread through host populations. In plant viruses, vector transmission is the major mode of transmission, used by nearly 80% of species described to date. Despite the importance of this phenomenon in epidemiology, the specificity of the virus-vector relationship is poorly understood at both the molecular and the evolutionary level, and very limited data are available on the precise viral protein motifs that control specificity. Here, using the aphid-transmitted Cauliflower mosaic virus (CaMV) as a biological model, we confirm that the "noncirculative" mode of transmission dominant in plant viruses (designated "mechanical vector transmission" in animal viruses) involves extremely specific virus-vector recognition, and we identify an amino acid position in the "helper component" (HC) protein of CaMV involved in such recognition. Site-directed mutagenesis revealed that changing the residue at this position can differentially affect transmission rates obtained with various aphid species, thus modifying the spectrum of vector species for CaMV. Most interestingly, in a virus line transmitted by a single vector species, we observed the rapid appearance of a spontaneous mutant specifically losing its transmissibility by another aphid species. Hence, in addition to the first identification of an HC motif directly involved in specific vector recognition, we demonstrate that change of a virus to a different vector species requires only a single mutation and can occur rapidly and spontaneously.

Two economically important characters, starch content and cassava bacterial blight resistance, were targeted to generate a large collection of cassava ESTs. Two libraries were constructed from cassava root tissues of varieties with high and low starch contents. Other libraries were constructed from plant tissues challenged by the pathogen Xanthomonas axonopodis pv.manihotis. We report here the single pass sequencing of 11,954 cDNA clones from the 5' ends, including 111 from the 3' ends. Cluster analysis permitted the identification of a unigene set of 5,700 sequences. Sequence analyses permitted the assignment of a putative functional category for 37% of sequences whereas approximately 16% sequences did not show any significant similarity with other proteins present in the database and therefore can be considered as cassava specific genes. A group of genes belonging to a large multigene family was identified. We characterize a set of genes detected only in infected libraries putatively involved in the defense response to pathogen infection. By comparing two libraries obtained from cultivars contrasting in their starch content a group of genes associated to starch biosynthesis and differentially expressed was identified. This is the first large cassava EST resource developed today and publicly available thus making a significant contribution to genomic knowledge of cassava.

The coffee disease complex known as corky-root, composed of the root-knot nematode #Meloidogyne arabicida# and #Fusarium oxysporum#, causes serious damage to #Coffea arabica# in Costa Rica. Resistance to #Meloidogyne arabicida# alone and to the field population (#M. arabicida#, #M. exigua#, #F. oxysporum#) was evaluated under controlled and field conditions. The studied material consisted of subspontaneous-derived #C. arabica# accessions, #C. canephora# accessions, introgression lines derived from the interspecific Timor Hybrid (#C. arabica# x #C. canephora#) and F1 hybrids from crosses between subspontaneous-derived accessions and cultivars. Accessions resistant to #M. arabicida# and to the field population were identified at varying frequencies in the different groups of study materials. The results showed that resistance to corky-root is heritable and that genetic resistance to #M. arabicida# is an effective strategy against corky-root disease. By using #C. canephora# rootstocks, it was possible to substantially reduce mortality in the field and reduce by half the number of plants with corky-root symptoms. The results were used to define appropriate strategies for the sustainable management of coffee corky-root disease resistance. (Resume d'auteur)

BGPI : équipe 4; International audience; Plant immunity is often triggered by the specific recognition of pathogen effectors by intracellular nucleotide-binding, leucine-rich repeat receptors (NLR). Plant NLRs contain an N-terminal signaling domain that is mostly represented by either a Toll-interleukinl receptor (TIR) domain or a coiled coil (CC) domain. In many cases, single NLR proteins are sufficient for both effector recognition and signaling activation. However, many paired NLRs have now been identified where both proteins are required to confer resistance to pathogens. Recent detailed studies on the Arabidopsis thaliana TIR-NLR pair RRS1 and RPS4 and on the rice CC-NLR pair RGA4 and RGA5 have revealed for the first time how such protein pairs function together. In both cases, the paired partners interact physically to form a hetero-complex receptor in which each partner plays distinct roles in effector recognition or signaling activation, highlighting a conserved mode of action of NLR pairs across both monocotyledonous and dicotyledonous plants. We also describe an "integrated decoy" model for the function of these receptor complexes. In this model, a plant protein targeted by an effector has been duplicated and fused to one member of the NLR pair, where it acts as a bait to trigger defense signaling by the second NLR upon effector binding. This mechanism may be common to many other plant NLR pairs.

The majority of plant disease-resistance genes (R-genes) isolated so far encode a predicted nucleotide-binding site (NBS) domain. NBS domains related to R-genes show a highly conserved backbone of amino acid motifs, which makes it possible to isolate resistance gene analogues (RGAs) by PCR with degenerate primers. Multiple combinations of primers with low degeneracy, designed from two conserved motifs in the NBS regions of R-genes of various plants, were used on genomic DNA from coffee trees, an important perennial tropical crop. Nine distinct classes of RGAs of the NBS-like type, representing a highly diverse sample, were isolated from Coffea arabica and C. canephora species. The analysis of one coffee RGA family suggested point mutations as the primary source of diversity. With one exception, coffee RGA families appeared to be closely related in sequence to at least one cloned R-gene. In addition, deduced amino acid sequences of coffee RGAs were identified that showed strong sequence similarity to almost all known non-TIR (Toll/Interleukin 1 Receptor)-type R-genes. The high degree of similarity between particular coffee RGAs and R-genes isolated from other angiosperm species, such as Arabidopsis, tomato and rice, indicates an ancestral relationship and the existence of common ancestors. The data obtained from coffee species suggests that the evolution of NBS-encoding sequences involves the gradual accumulation of mutations and slow rates of divergence within distinct R-gene families, rather than being a rapid process. Functional inferences drawn from the suggested pattern of evolution of NBS-type R-genes is also discussed.

We evaluated cassava bacterial blight (CBB) infection in a pair-cross population of 150 individuals derived from an intra-specific cross between two non-inbred cassava (#Manihot esculenta$ Crantz) lines. The replicated trials were carried out in the field under high disease pressure over two consecutive crop cycles. Evaluations were conducted at 4 and 7 months after planting for the two cycles. Simple regression analysis and the nonparametric Kruskal-Wallis rank-sum test revealed that eight quantitative trait loci (QTLs) were invovled in resistance. We detected changes in QTLs from crop cycle to crop cycle. The pathogen population (#Xanthomonas axonopodis$ pv. #manihotis$) was also monitored over the period, using a restriction fragment length polymorphism probe and pathogenic tests. Changes in QTL detection over the 2 years could be correlated with changes in pathogen population structure. One QTL, located in linkage group D, was conserved over the two crop cycles, and in field to greenhouse evaluations. This study thus identified molecular markers useful for marker assisted-selection, a technique that can accelerate the long, multiple-season process of breeding for CBB resistance. (Résumé d'auteur)

African cassava mosaic virus (ACMV), vectored by the whitefly, Bermisia tabaci (Gennadius), is considered the most damaging pathogen of any African crop. Information about vector movement is important for understanding the epidemiology of this disease and the experiments reported here were designed to examine B.tabaci flight activity both in and around a cassava crop in relation to time of day, crop growth stage, wind direction and speed, and to the resulting pattern of infected plants within the field at harvest. At wind speeds of −1 adult B. tabaci approached the yellow traps by flying upwind. At greater wind speeds, significantly fewer B. tabaci adults approached the traps from downwind, thus reversing the directionally of the catch. When the direction of the prevailing south-west wind reversed, so did the directionality of the catch. Bemisia tabaci adults were flight active throughout the day and the greatest percentage were caught above the canopy between 06.00–08.00 h, when wind speeds were lowest. Trap height and position significantly affected catch with the greatest numbers caught on the lowest traps. More than three times as many B. tabaci adults were caught on traps situated downwind from the field compared to those upwind. Suggesting that the field was acting as a source of whiteflies. In both years, Africa cassava mosaic disease (ACMD) incidence was highest and lowest, respectively, on the edges and in the middle of the trials, with the highest incidence occuring on the edges facing the prevailing wind direction. These results are discussed in relation to the epidemiology of ACMD and to potential cultural control methods such as the use of ACMD-resistant guard rows to protect a mainly susceptible crop.

Rusts are one of the most severe threats to cereal crops because new pathogen races emerge regularly, resulting in infestations that lead to large yield losses. In 1999, a new race of stem rust, Puccinia graminis f. sp. tritici (Pgt TTKSK or Ug99), was discovered in Uganda. Most of the wheat and barley cultivars grown currently worldwide are susceptible to this new race. Pgt TTKSK has already spread northward into Iran and will likely spread eastward throughout the Indian subcontinent in the near future. This scenario is not unique to stem rust; new races of leaf rust (Puccinia triticina) and stripe rust (Puccinia striiformis) have also emerged recently. One strategy for countering the persistent adaptability of these pathogens is to stack complete- and partial-resistance genes, which requires significant breeding efforts in order to reduce deleterious effects of linkage drag. These varied resistance combinations are typically more difficult for the pathogen to defeat, since they would be predicted to apply lower selection pressure. Genetical genomics or expression Quantitative Trait Locus (eQTL) analysis enables the identification of regulatory loci that control the expression of many to hundreds of genes. Integrated deployment of these technologies coupled with efficient phenotyping offers significant potential to elucidate the regulatory nodes in genetic networks that orchestrate host defense responses. The focus of this review will be to present advances in genetical genomic experimental designs and analysis, particularly as they apply to the prospects for discovering partial disease resistance alleles in cereals.

Publication Inra prise en compte dans l'analyse bibliométrique des publications scientifiques mondiales sur les Fruits, les Légumes et la Pomme de terre. Période 2000-2012. http://prodinra.inra.fr/record/256699; International audience; Plum pox virus (PPV), a member of the genus Potyvirus, is the causal agent of Sharka, the most detrimental disease of stone-fruit trees worldwide. PPV isolates are grouped into seven distinct strains. The minor PPV-W strain was established recently for the divergent W3174 isolate found in Canada. Here, the partial or complete genomic sequences of four PPV-W isolates from Latvia have been determined. The completely sequenced isolates LV-141pl and LV-145bt share 93.1 and 92.1% nucleotide identity, respectively, with isolate W3174, with two regions of higher (>20%) divergence in the P1/HC-Pro and NIa (VPg) regions. Further analyses demonstrated that these two regions correspond to two independent recombination events in the W3174 genome, one involving PPV-M (approximate genome positions 692 to 1424) and the other PPV-D (nucleotides 5672 to 5789). The LV-141pl and LV-145bt isolates appear to be representatives of the “ancestral” PPV-W strain, not affected by recombination. The PPV-W intrastrain variability is substantially higher than that of all other PPV strains, with potential implications for the serological detection of PPV-W isolates. A PPV-W-specific primer pair has been developed, allowing the specific reverse-transcription polymerase chain reaction detection of all five presently available W isolates. The characterization of these new PPV-W isolates sheds light on PPV-W evolutionary history, further supports the hypothesis of its East-European origin, and opens the way for the biological and epidemiological characterization of this poorly known PPV strain.

Aplysia gonad lectin, which binds polygalacturonic acid, was complexed to colloidal gold and used for localizing molecules that contain polygalacturonic acid in rubber tree roots infected with the white-rot root fungus Rigidoporus lignosus. Colonization of root tissues was associated with strong wall alteration of phloem cells and with degradation of the compound middle lamella in both the phloem (10 weeks after inoculation) and the xylem (15 weeks after inoculation). Our data suggest that pectin breakdown during root decay likely occurs after cellulose and lignin breakdown and may result from the fungal pectinase activities that were detected in vitro. Released pectin oligomers may act as inducers of both fungal laccase and of the tree defense system during root invasion. Key words: root rotting fungi, rubber, pectin, cellulose, cytochemistry.

Black Sigatoka is an important fungal disease of bananas and plantains. Dew duration on the upper leaves of these plants (Leaves 0 and 1) is an important factor in the development of the disease. A general dew estimation scheme, based on an energy budget approach, was adapted to the particular case of banana and plantain upper leaves in order to provide a tool for Black Sigatoka management. This scheme allows the estimation of dew duration from standard weather station measurements of air temperature and humidity, wind velocity, solar radiation and cloud cover. Values of dew duration estimated by this scheme were compared with observed values on plantain during 18 nights. On average, the differences between observed and estimated values were less than 1 h.

La nécrose du phloème d'Hévéa est une grave maladie répandue dans les plantations ouest-africaines. A l'échelle ultrastructurale, l'altération des parois cellulaires est la modification la plus fréquemment observée. Une étude cytochimique a été entreprise dans le but de mieux comprendre les mécanismes de la dégradation de la cellulose des parois. Les observations microscopiques ont également montré la présence de vésicules paramurales et de dépôts le long des parois dans les espaces périplasmiques, suggérant la participation directe, ou indirecte, de microorganismes à la nécrose du phloème d'Hévéa. (D'après résumé d'auteur)

Phloem necrosis of rubber tree (Hevea brasiliensis) is characterized by cell wall degradation, alteration of the middle lamella, the vesiculation of endomembranes, the formation of tylosoids and the internal coagulation of rubber

A collection of 51 Xanthomonas campestris strains from throughout the world was studied to detect and assess genetic diversity among pathogens of small grains. Isolates from barley, bread wheat, bromegrass, canary grass, cassava, maize, orchard grass, rice, rough-stalked meadow grass, rye, timothy, and triticale were analyzed by pathogenicity tests on bread wheat cv. Alondra and barley cv. Corona, indirect immunofluorescence, and restriction fragment length polymorphism (RFLP). Three probes were used for the RFLP analysis. They were an acetylaminofluorene-labelled 16S+23S rRNA probe from Escherichia coli and two (sup32)P-labelled restriction fragments from either plasmidic (pBSF2) or chromosomal (pBS8) DNA of X. campestris pv. manihotis. Strains clustered in 9 and 20 groups with the rRNA probe and the pBSF2 DNA probe, respectively. Strains of X. campestris pv. graminis, X. campestris pv. phleipratensis, and X. campestris pv. poae are shown to be related but are also distinguishable by RFLP patterns, serology, and pathogenicity on bread wheat. Strains pathogenic only for barley and not for wheat grouped together. Another group is temporarily designated deviant X. campestris pv. undulosa. These South American isolates from bread wheat did not react by indirect immunofluorescence and produced atypical lesions in pathogenicity tests. The results stress the need to perform pathogenicity tests before strains are named at the pathovar level. The importance of the different probes used for epidemiological studies or phylogenetic studies of closely related strains is underlined.

In spite of numerous aeroplane flights and different interpretations of the infrared colour photographs obtained, the remote sensing of root rotting fungi of rubber tree has failed in practice in the Ivory Coast. The reasons for this failure and conclusions on the efficiency of this method are discussed.

SUMMARY Asystasia mottle virus (AsMV) was detected serologically in samples of Asystasia gangetica with mottle symptoms, from several areas of tropical West Africa. It infected 12 species systemically and induced local lesions in a further four. The virus lost infectivity after dilution to 10-4, after 10 min at 75 °C and after 3 days at 27°C. Purified virus had an A260/A280 ratio of c. 1·2 and a protein subunit mol. wt of c. 33 000. Particles were c. 750 nm long and cytoplasmic inclusions typical of potyviruses were seen in ultrathin sections of infected leaves. The antiserum prepared had a titre of 1/1024 in microprecipitin tests but purified virus failed to react with 31 antisera to known potyviruses. The virus was transmitted in the non-persistent manner by Aphis spiraecola but only very infrequently. On the basis of these properties, AsMV is considered to be a new member of the potyvirus group.

BGPI : équipe 2/7; International audience; During a large scale "non a priori" survey in 2010 of South African plant-infecting single stranded DNA viruses, a highly divergent geminivirus genome was isolated from a wild spurge, Euphorbia caput-medusae. In addition to being infectious in E. caput-medusae, the cloned viral genome was also infectious in tomato and Nicotiana benthamiana. The virus, named Euphorbia caput-medusae latent virus (EcmLV) due to the absence of infection symptoms displayed by its natural host, caused severe symptoms in both tomato and N. benthamiana. The genome organisation of EcmLV is unique amongst geminiviruses and it likely expresses at least two proteins without any detectable homologues within public sequence databases. Although clearly a geminivirus, EcmLV is so divergent that we propose its placement within a new genus that we have tentatively named Capulavirus. Using a set of highly divergent geminiviruses genomes, it is apparent that recombination has likely been a primary process in the genus-level diversification of geminiviruses. It is also demonstrated how this insight, taken together with phylogenetic analyses of predicted coat protein and replication associated protein (Rep) amino acid sequences indicate that the most recent common ancestor of the geminiviruses was likely a dicot-infecting virus that, like modern day mastreviruses and becurtoviruses, expressed its Rep from a spliced complementary strand transcript.

Background Spiroplama citri, the causal agent of citrus stubborn disease, is a bacterium of the class Mollicutes and is transmitted by phloem-feeding leafhopper vectors. In order to characterize candidate genes potentially involved in spiroplasma transmission and pathogenicity, the genome of S. citri strain GII3-3X is currently being deciphered. Results Assembling 20,000 sequencing reads generated seven circular contigs, none of which fit the 1.8 Mb chromosome map or carried chromosomal markers. These contigs correspond to seven plasmids: pSci1 to pSci6, with sizes ranging from 12.9 to 35.3 kbp and pSciA of 7.8 kbp. Plasmids pSci were detected as multiple copies in strain GII3-3X. Plasmid copy numbers of pSci1-6, as deduced from sequencing coverage, were estimated at 10 to 14 copies per spiroplasma cell, representing 1.6 Mb of extrachromosomal DNA. Genes encoding proteins of the TrsE-TraE, Mob, TraD-TraG, and Soj-ParA protein families were predicted in most of the pSci sequences, in addition to members of 14 protein families of unknown function. Plasmid pSci6 encodes protein P32, a marker of insect transmissibility. Plasmids pSci1-5 code for eight different S. citri adhesion-related proteins (ScARPs) that are homologous to the previously described protein P89 and the S. kunkelii SkARP1. Conserved signal peptides and C-terminal transmembrane alpha helices were predicted in all ScARPs. The predicted surface-exposed N-terminal region possesses the following elements: (i) 6 to 8 repeats of 39 to 42 amino acids each (sarpin repeats), (ii) a central conserved region of 330 amino acids followed by (iii) a more variable domain of about 110 amino acids. The C-terminus, predicted to be cytoplasmic, consists of a 27 amino acid stretch enriched in arginine and lysine (KR) and an optional 23 amino acid stretch enriched in lysine, aspartate and glutamate (KDE). Plasmids pSci mainly present a linear increase of cumulative GC skew except in regions presenting conserved hairpin structures. Conclusion The genome of S. citri GII3-3X is characterized by abundant extrachromosomal elements. The pSci plasmids could not only be vertically inherited but also horizontally transmitted, as they encode proteins usually involved in DNA element partitioning and cell to cell DNA transfer. Because plasmids pSci1-5 encode surface proteins of the ScARP family and pSci6 was recently shown to confer insect transmissibility, diversity and abundance of S. citri plasmids may essentially aid the rapid adaptation of S. citri to more efficient transmission by different insect vectors and to various plant hosts.

Wild plant populations may harbour a myriad of unknown viruses. As the majority of research efforts have targeted economically important plant species, the diversity and prevalence of viruses in the wild has remained largely unknown. However, the recent shift towards metagenomics-based sequencing methodologies, especially those targeting small RNAs, is finally enabling virus discovery from wild hosts. Understanding this diversity of potentially pathogenic microbes in the wild can offer insights into the components of natural biodiversity that promotes long-term coexistence between hosts and parasites in nature, and help predict when and where risks of disease emergence are highest. Here, we used small RNA deep sequencing to identify viruses inPlantago lanceolatapopulations, and to understand the variation in their prevalence and distribution across the Åland Islands, South-West Finland. By subsequent design of PCR primers, we screened the five most common viruses from two sets ofP. lanceolataplants: 164 plants collected from 12 populations irrespective of symptoms, and 90 plants collected from five populations showing conspicuous viral symptoms. In addition to the previously reported speciesPlantago lanceolata latent virus(PlLV), we found four potentially novel virus species belonging toCaulimovirus, Betapartitivirus, Enamovirus,andClosterovirusgenera. Our results show that virus prevalence and diversity varied among the sampled host populations. In six of the virus infected populations only a single virus species was detected, while five of the populations supported between two to five of the studied virus species. In 20% of the infected plants, viruses occurred as coinfections. When the relationship between conspicuous viral symptoms and virus infection was investigated, we found that plants showing symptoms were usually infected (84%), but virus infections were also detected from asymptomatic plants (44%). Jointly, these results reveal a diverse virus community with newly developed tools and protocols that offer exciting opportunities for future studies on the eco-evolutionary dynamics of viruses infecting plants in the wild.