Viruses that infect plants are responsible for reduction in both yield and quality of crops around the world, and thus are of great economic importance. This has provided the impetus for the extensive research into the molecular and cellular biology of these pathogens and into their interaction with their plant hosts and their vectors. However, interest in plant viruses extends beyond their ability to damage crops. Many plant viruses — for example, tobacco mosaic virus — have been used as model systems to provide basic understanding of how viruses express genes and replicate. Others permitted the elucidation of the processes underlying RNA silencing, now recognized as a core epigenetic mechanism underpinning numerous areas of biology. This book attests to the huge diversity of research in plant molecular virology.

Written by world authorities in the field, the book opens with two chapters on the translation and replication of viral RNA. Following chapters cover topics such as viral movement within and between plants, plant responses to viral infection, antiviral control measures, virus evolution, and newly emerging plant viruses. The book concludes with two chapters on biotechnological applications of plant viruses. Throughout, the focus is on the most recent, cutting-edge research, making this book essential reading for everyone working with plant viruses.

Recent Advances in Plant Virology

Chapter 1: Roles of Cis-acting Elements in Translation of Viral RNAs

W. Allen Miller , Jelena Kraft, Zhaohui Wang and Qiuling Fan

Cis-acting signals regulate translation of viral RNAs to produce viral proteins at the appropriate levels and timing to maximize virus replication. Here we describe the cis-acting sequences that achieve this translational control via processes such as cap-dependent translation, leaky scanning to initiate translation at more than one start codon, ribosomal shunting, cap-independent translation initiation controlled from the 5' and/or 3' untranslated region, poly(A) tail-independent translation initiation, stop codon readthrough, and ribosomal frameshifting. The secondary structures and, in some cases, tertiary structures of the RNA sequences that control these events are described. We also discuss the mechanisms of the translation events facilitated by the cis-acting signals, and how they mesh with the overall replication strategies of the diverse viruses that employ these mechanisms.

Chapter 2: Replication of Plant RNA viruses

Peter D. Nagy and Judit Pogany

Among plant viruses, the positive-stranded RNA [(+)RNA] viruses are the largest group, and the most widespread. The central step in the infection cycle of (+)RNA viruses is RNA replication, which is carried out by virus-specific replicase complexes consisting of viral RNA-dependent RNA polymerase, one or more auxiliary viral replication proteins, and a number of co-opted host factors. Viral replicase complexes assemble in specialized membranous compartments in infected cells. Sequestering the replicase complexes is not only helpful for rapid production of a large number of viral (+)RNA progeny, but it also facilitates avoiding recognition by the hostÕs anti-viral surveillance system, and it provides protection from degradation of the viral RNA. Successful viral replication is followed by cell-to-cell and long-distance movement throughout the plant, as well as encapsidation of the (+)RNA progeny to facilitate transmission to new plants. This chapter provides an overview of our current understanding of the molecular mechanisms in plant (+)RNA virus replication. Recent significant progress in this research area is based on development of powerful in vivo and in vitro methods, including replicase assays, reverse genetic approaches, intracellular localization studies, genome-wide screens for co-opted host factors and the use of plant or yeast model hosts.

Chapter 3: Plasmodesmata as Active Conduits for Virus Cell-to-Cell Movement

Lourdes Fernandez-Calvino, Christine Faulkner and Andy Maule

It has been known for many decades that viruses need to exploit plasmodesmata as channels of cytoplasmic connectivity through plant cell walls. However, we do not yet understand the molecular mechanisms involved in moving a single infectious entity from cell to cell, although it is clear that virus-encoded movement proteins play a central role. Major progress has been made in identifying movement proteins, their associations with subcellular structures/organelles, and their biochemical properties with respect to nucleic acid-binding and physical associations with host and other viral proteins. These studies reveal a specificity in functional evolution where viruses share some similarities in their movement strategies with near and far phylogenetic groups but show few examples of processes that might apply to all or many individual viruses. Plasmodesmata also provide channels for cellular communication essential for plant growth, development and defense. As such, there is increasing attention aimed at resolving their constituent components necessary for structure and function. With the limited success of genetic screens, proteomic analysis of biochemically-enriched plasmodesmal fractions has also been pursued. Through the identification of plasmodesmal proteins we will have the opportunity to understand how movement proteins bring about the massive changes in the physical behaviour of plasmodesmata that result in the translocation of the macromolecular complexes responsible for virus infectivity.

Chapter 4: Systemic Movement of Viruses Via the Plant Phloem

Vicente Pallás, Ainhoa Genovés, M. Amelia Sánchez-Pina and José Antonio Navarro

The incorporation of non invasive techniques has allowed remarkable progress in our understanding of the vascular transport of plant viruses. Indeed, approximately seventy-five percent of reports about this topic have been published after the first use of the jellyfish green fluorescent protein (GFP) in plant virology. In the last two decades, a very detailed picture of the viral determinants involved in phloem transport of plant viruses has been obtained. However, we realize that most virus-host interactions are pathosystem-specific and, consequently, the identification of common host factors involved in phloem transport of plant viruses is the exception rather than the rule. In addition, we are still far from obtaining a clear picture of how environmental factors influence the vascular invasion of plants by these pathogens. In this chapter, we review the progress made in understanding the viral determinants involved in vascular transport of viruses and the pathways followed by viruses during systemic movement, and we do so mainly by focusing on host and environmental conditions that influence the final distribution of viruses in the plant.

Chapter 5: Functions of Virus and Host Factors During Vector-mediated Transmission

Stéphane Blanc and Martin Drucker

Most plant viruses are transmitted by living vectors that transport viruses to a new host plant. One discriminates between circulative transmission, where viruses must pass through the vector interior and are usually inoculated with the saliva on a healthy plant, and non-circulative transmission, where viruses do not need to pass through the vector interior but are directly inoculated from the mouth parts into a new host. Especially transmission of non-circulative viruses has been regarded as a simple process where a vector more or less accidentally transports the virus. However, it becomes more and more evident that this scenario is unlikely, because transmission constitutes a dramatic bottleneck of the virus life cycle, where only very few viral genomes pass to a new host, and where a given virus must do everything to ensure successful transmission. We will show and discuss in this chapter that viruses - also in non-circulative transmission - deliberately manipulate their hosts and vectors in often very unexpected ways to optimise their transmission.

Chapter 6: RNA Silencing and the Interplay Between Plants and Viruses

Lourdes Fernández-Calvino, Livia Donaire and César Llave

In eukaryotes, RNA silencing controls gene expression to regulate development, genome stability and stress-induced responses. In plants, this process is also recognized as a major immune system targeted against plant viruses. Plant viruses stimulate RNA silencing responses though formation of viral RNA with double-stranded features that are subsequently processed into functional small RNAs (sRNAs). Recent studies highlight the complexity of the viral sRNA populations and their potential to associate with multiple silencing effector complexes. This fact has profound implications in the cross-talk interactions between plants and viruses since both virus genomes and host genes are putative targets of viral sRNAs. In this chapter, the concept of RNA silencing is discussed as an elegant natural antiviral mechanism in plants, assessing the contribution of viral sRNA-mediated regulation of gene expression in the frame of compatible interactions between plants and viruses.

Chapter 7: Mechanism of Action of Viral Suppressors of RNA Silencing

József Burgyán

RNA silencing is an evolutionarily conserved sequence-specific gene-inactivation system that also functions as an antiviral mechanism in higher plants and insects. To overcome this defence system, viruses encode suppressors of RNA silencing, which can counteract the host silencing-based antiviral process. In the past, more than 50 individual viral suppressors have been identified from almost all plant virus genera, underlining their crucial role in successful virus infection. Viral suppressors are considered to be of recent evolution, and they are surprisingly diverse within and across kingdoms, exhibiting no obvious sequence similarity. Virus-encoded silencing suppressors can target several key components in the silencing machinery, such as silencing-related RNA structures and essential effector proteins and complexes. This chapter reviews the most recent progress in our understanding of the mechanism and function of viral suppressors of antiviral RNA silencing in plants.

Chapter 8: NB-LRR Immune Receptors in Plant Virus Defense

Patrick Cournoyer and Savithramma P. Dinesh-Kumar

Resistance genes protect plants from infection by viruses and many other classes of pathogens. The dominant, anti-viral R genes that have been cloned thus far encode NB-LRR immune receptors that detect a single viral protein and trigger defense. Many different types of viral proteins are known to elicit defense by corresponding NB-LRRs. Defense often results in a type of localized programmed cell death at the site of attempted pathogen infection known as the hypersensitive response (HR-PCD), but some NB-LRRs confer resistance to viruses without HR-PCD. The activation of NB-LRRs triggers manifold signaling events including reactive oxygen species (ROS) production, nitric oxide (NO) production, calcium (Ca2+) influx, activation of mitogen activated protein kinases (MAPKs), and production of the plant hormones salicylic acid (SA), jasmonic acid (JA), and ethylene. After a successful NB-LRR-mediated defense event, the plant exhibits heightened resistance to future pathogen challenge in a state called systemic acquired resistance.

Chapter 9: Plant Resistance to Viruses Mediated by Translation Initiation Factors

Olivier Le Gall, Miguel A. Aranda and Carole Caranta

Host resistance to viruses can show dominant or recessive inheritance. Remarkably, recessive resistance genes are much more common for viruses than for other plant pathogens. Recessive resistances to viruses are especially well documented within the dicotyledons, and have been described for various viruses that belong to very different viral genera, although clearly they predominate among viruses belonging to the genus Potyvirus. The elucidation of the molecular nature of this particular class of resistance genes is recent, but has so far only revealed a group of proteins linked to the translation machinery, chiefly the eukaryotic translation initiation factors (eIF) 4E and 4G, which are the subject of this chapter. Thus, we will briefly review how translation initiation is performed in eukaryotes, to then describe the features and mechanisms of eIF4E- and 4G-mediated resistances to potyviruses and viruses belonging to other genera, such as carmoviruses. We will bring the chapter to a close by summarizing conclusions and offering potential research perspectives in this field.

Chapter 10: Advanced Breeding for Virus Resistance in Plants

Alain Palloix and Frank Ordon

Breeding for virus resistance was successful in the past years using conventional breeding methods since many virus resistant cultivars have been delivered for a wide range of crops. Genome mapping provided molecular markers for many resistance loci (i.e., major genes or Quantitative Trait Loci) that were introgressed into cultivars e.g., through backcross breeding schemes. Molecular mapping also delivered much information on the genomic architecture of polygenic and quantitative resistances. However, marker assisted selection for such complex traits is difficult so that the combination of quantitative resistance factors from multiallelic origins commonly relies on sophisticated phenotyping procedures. The cloning of resistance genes and the rapid development of high throughput molecular technologies increased the access to functional markers and multiallelic markers, promoting the applicability of marker assisted selection for complex traits at the whole genome scale in the near future. In parallel, the advances in the identification of molecular determinants of plant/virus interactions and in genetics and evolution of virus populations provide new selection criteria for breeders to choose the most durable resistance genes and gene combinations, so that breeding for durable virus resistance becomes an accessible quest.

Chapter 11: Sustainable Management of Plant Resistance to Viruses

Benoît Moury, Alberto Fereres, Fernando García-Arenal and Hervé Lecoq

Although viruses are among the parasites which induce the most severe damages on cultivated plants, few control methods have been developed against them. Notably, no curative methods can be applied against virus diseases in crops. In view of this major economic problem, the development of resistant cultivars has become a critical factor of competitiveness for breeders. However, plant - virus interactions are highly dynamic and the selective pressure exerted by plant resistance frequently favours the emergence of adapted virus populations. Given the scarcity of resistance genes, there is consequently an urgent need to increase the sustainability of these genetic resources. In this chapter, we will review the biological mechanisms which allow the emergence of virus populations adapted to plant resistances and how we can use this knowledge to explain the relative durability of different resistance genes, to built predictors of resistance durability and to combine the use of resistances with other control methods to increase their sustainability.

Chapter 12: Integrated Control Measures Against Viruses and Their Vectors

Alberto Fereres and Aranzazu Moreno

Viruses and their vectors produce severe damage to crops worldwide. This chapter focuses on the strategies and tactics often used to manage vectors of plant viruses, with special attention to insects, by far the most important type of vector. The philosophy and principles of Integrated Pest Management (IPM) developed long ago can still provide an effective and sustainable way to manage insect vectors of virus diseases. Preventive strategies such as the development of models that forecast virus disease outbreaks together with host plant resistance, cultural and physical tactics are the most effective ways to control nonpersistently-transmitted viruses. A reduction in vector numbers using conventional systemic insecticides or innundative biological control agents can also provide effective control of persistently-transmitted viruses. Recent advances on understanding of the mode of transmission of plant viruses are also a very promising way to develop molecules to block putative virus binding sites within the vector and to avoid virus retention and transmission. Also, the characterization of aphidÕs salivary components that is underway may facilitate the development of new tools to interfere with the process of transmission of plant viruses.

Chapter 13: Population Dynamics and Genetics of Plant Infection by Viruses

Fernando García-Arenal and Aurora Fraile

During the last thirty years, progress in understanding the mechanistic aspects of virus-plant interactions has been remarkable, notably in aspects such as genome replication, movement within the infected host or pathogenesis and resistance. Progress in understanding the population dynamics and genetics of plant infection by viruses has not been as great. However, understanding the kinetics of plant colonisation and the genetic structure of the within-host virus population is necessary for addressing many issues of plant-virus interaction and of virus evolution. The quantitative aspects of plant infection and colonisation by viruses were mostly addressed during the early period of plant virology, when many detailed studies were published that often incorporated mathematical modelling. These issues have not been thoroughly re-examined using molecular techniques. Recent work has focussed on the description of the genetic structure of the virus population at the organ and the plant level. Data suggest that in spite of huge fecundity, the effective numbers of the within-host virus population may be small due to severe population bottlenecks at each stage of plant infection and colonisation, which results in a spatially structured population.

Chapter 14: Evolutionary Constraints on Emergence of Plant RNA Viruses

Santiago F. Elena

Over the recent years, agricultural activity in many regions has been compromised by a succession of devastating epidemics caused by new viruses that switched host species, or by new variants of classic viruses that acquired new virulence factors or changed their epidemiological patterns. Although viral emergence has been classically associated with ecological change or with agronomical practices that brought in contact reservoirs and crop species, it has become obvious that the picture is much more complex, and results from an evolutionary process in which the main players are the changes in ecological factors, the tremendous genetic plasticity of viruses, the several host factors required for virus replication, and a strong stochastic component. The present chapter puts emergence of RNA viruses into the framework of evolutionary genetics and reviews the basic notions necessary to understand emergence, stressing that viral emergence begins with a stochastic process that involves the transmission of a pre-existing viral strain with the right genetic background into a new host species, followed by adaptation to the new host during the early stages of infection.

Chapter 15: Emergence of Begomovirus Diseases

Enrique Moriones, Jesus Navas-Castillo and Juan-Antonio Díaz-Pendón

Begomoviruses (genus Begomovirus, family Geminiviridae) rank among the top of the most important plant viruses causing disease of severe consequences in economically and socially relevant crops. From the early 1990s, a rapid emergence and geographic expansion of begomoviruses has occurred worldwide. As a result, these viruses have become the most destructive group of plant viruses in tropical and subtropical regions of the world. Their emergence is associated with the emergence of populations of the insect vector, the whitefly Bemisia tabaci, probably due to increased plant trading between distantly separated geographical regions and changes in agricultural practices. Human activity seems to have been a major factor promoting emergence of begomoviruses. Other factors driving emergence are discussed in this contribution, and examples of emergence of begomovirus diseases in economically important crops are provided.

Chapter 16: Genomic Approaches to Discovery of Viral Species Diversity of Non-cultivated Plants

Ulrich Melcher and Veenita Grover

Outbreaks of newly emerging and re-emerging animal and plant viruses pose a constant threat to public health and food security and emphasize the need to develop efficient methods for viral detection and identification. Ongoing studies for discovery of viral species in non-cultivated plants utilize genomic approaches for systematic unbiased searches for viruses related to known viruses. Genomic approaches use various combinations of methods for sampling the environment, enriching samples for content of viral genomes, amplifying nucleic acids, and detecting virus-related sequences among the amplified nucleic acids. These methods include particularly array hybridization to macroarrays and microarrays, and various megasequencing approaches. In all cases, relatives of known viruses are discovered. However, the identification of a novel plant virus completely unrelated to known ones remains a challenge. Despite a growing list of viruses infecting wild plants, virus infections in wild plant communities are often underestimated relative to cultivated systems, since viruses in wild plants are generally considered not to harm the host. Viruses may not be explicitly damaging wild plants, but their biodiversity and abundance suggest an important role of these viruses in ecosystems. These roles should not be under-rated just because they are under-researched.

Chapter 17: Endogenous Viral Sequences in Plant Genomes

Pierre-Yves Teycheney and Andrew D.W. Geering

Endogenous viral sequences from members of two virus families, the Caulimoviridae and Geminiviridae, have been discovered in several monocotyledonous and dicotyledonous plant species. For the most part, these sequences are replication-defective but those capable of causing infection have been discovered in tobacco (Nicotiana edwardsonii), petunia (Petunia hybrida) and banana and plantain (Musa spp.). Activation of endogenous caulimovirid sequences is one of the major impediments to international banana and plantain breeding efforts. Research on endogenous viral sequences in plants is still in its infancy, with little known about the contributions of these sequences to host and virus evolution, nor even a classification system adopted. On a practical note, problems still exist with differentially detecting viral genomic DNA in a host genetic background containing endogenous viral sequences, and a solution to the problem of activation of endogenous viral sequences in banana is still far away. In this review, answers to some of these questions are sought by drawing on research from the related fields of endogenous retroviruses in animals and LTR-retrotransposons in eukaryotes in general.

Chapter 18: Virus Particles and the Uses of Such Particles in Bio- and Nanotechnology

George P. Lomonossoff

The capsids of most plant viruses are simple and robust structures consisting of multiple copies of one or a few types of protein subunit arranged with either icosahedral or helical symmetry. The capsids can be produced in large quantities either by the infection of plants or by the expression of the subunit(s) in a variety of heterologous systems. In view of their relative simplicity and ease of production, plant virus particles or virus-like particles (VLPs) have attracted much interest over the past 20 years for applications in both bio- and nanotechnology. As result, plant virus particles have been subjected to both genetic and chemical modification, have been used to encapsulate foreign material and have, themselves, been incorporated into supramolecular structures. This chapter reviews the various ways in which have been exploited over the past two decades.

Chapter 19: Plant Viral Vectors for Protein Expression

Yuri Y. Gleba and Anatoli Giritch

Plant-virus-driven transient expression of heterologous proteins is the basis of several mature manufacturing processes that are currently being used for the production of multiple proteins including vaccine antigens and antibodies. Viral vectors have also become useful tools for research. In recent years, advances have been made both in the development of first-generation vectors (those that employ the 'full virus' strategy) as well as second-generation vectors designed using the 'deconstructed virus' approach. This second strategy relies on Agrobacterium as a vector to deliver DNA copies of one or more viral RNA replicons. Among the most often used viral backbones are those of Tobacco mosaic virus, Potato virus X, and Cowpea mosaic virus. Prototypes of industrial processes that provide for high-yield, rapid scale-up, and fast manufacturing have been recently developed using viral vectors, with several manufacturing facilities compliant with good manufacturing practices (GMP) in place, and a number of pharmaceutical proteins currently in pre-clinical and clinical trials.

"...an extensive overview of recent developments in plant virus research...chapters are well-written and on the cutting edge of research...an excellent piece of work for a specialized audience such as graduate students, postdoctoral and senior researchers or lecturers...each institutional library should stock a copy for reference."
—Microbiology Today