Annual Review of Virology - Volume 10, 2023

From a farming family of 13 children in New Zealand, I graduated with a Master of Science degree in microbiology from the University of Otago (Dunedin, Otago, New Zealand). I established the first veterinary virology laboratory at Wallaceville Animal Research Station. I subsequently completed my PhD degree at Australian National University (Canberra, Australia) and a postdoctoral fellowship at the University of Michigan (Ann Arbor, Michigan). While in New South Wales, Australia, a walk on a beach littered with dead mutton birds (shearwaters) with Dr. Graeme Laver led to the surveillance of influenza in seabirds on the Great Barrier Reef Islands and my lifelong search for the origin of pandemic influenza viruses. Subsequent studies established that (a) aquatic birds are a natural reservoir of influenza A viruses, (b) these viruses replicate primarily in cells lining the intestinal tract, (c) reassortment in nature can lead to novel pandemic influenza viruses, and (d) live bird markets are one place where transmission of influenza virus from animals to humans occurs.

The 1918 Spanish influenza pandemic was one of the deadliest infectious disease events in recorded history, resulting in approximately 50–100 million deaths worldwide. The origins of the 1918 virus and the molecular basis for its exceptional virulence remained a mystery for much of the 20th century because the pandemic predated virologic techniques to isolate, passage, and store influenza viruses. In the late 1990s, overlapping fragments of influenza viral RNA preserved in the tissues of several 1918 victims were amplified and sequenced. The use of influenza reverse genetics then permitted scientists to reconstruct the 1918 virus entirely from cloned complementary DNA, leading to new insights into the origin of the virus and its pathogenicity. Here, we discuss some of the advances made by resurrection of the 1918 virus, including the rise of innovative molecular research, which is a topic in the dual use debate.

Humans have battled viruses for millennia. However, directly linking the symptomatology of disease outbreaks to specific viral pathogens was not possible until the twentieth century. With the advent of the genomic era and the development of advanced protocols for isolation, sequencing, and analysis of ancient nucleic acids from diverse human remains, the identification and characterization of ancient viruses became feasible. Recent studies have provided invaluable information about past epidemics and made it possible to examine assumptions and inferences on the origin and evolution of certain viral families. In parallel, the study of ancient viruses also uncovered their importance in the evolution of the human lineage and their key roles in shaping major events in human history. In this review, we describe the strategies used for the study of ancient viruses, along with their limitations, and provide a detailed account of what past viral infections have revealed about human history.

Understanding the factors that shape viral evolution is critical for developing effective antiviral strategies, accurately predicting viral evolution, and preventing pandemics. One fundamental determinant of viral evolution is the interplay between viral protein biophysics and the host machineries that regulate protein folding and quality control. Most adaptive mutations in viruses are biophysically deleterious, resulting in a viral protein product with folding defects. In cells, protein folding is assisted by a dynamic system of chaperones and quality control processes known as the proteostasis network. Host proteostasis networks can determine the fates of viral proteins with biophysical defects, either by assisting with folding or by targeting them for degradation. In this review, we discuss and analyze new discoveries revealing that host proteostasis factors can profoundly shape the sequence space accessible to evolving viral proteins. We also discuss the many opportunities for research progress proffered by the proteostasis perspective on viral evolution and adaptation.

Two decades of metagenomic analyses have revealed that in many environments, small (∼5 kb), single-stranded DNA phages of the family Microviridae dominate the virome. Although the emblematic microvirus phiX174 is ubiquitous in the laboratory, most other microviruses, particularly those of the gokushovirus and amoyvirus lineages, have proven to be much more elusive. This puzzling lack of representative isolates has hindered insights into microviral biology. Furthermore, the idiosyncratic size and nature of their genomes have resulted in considerable misjudgments of their actual abundance in nature. Fortunately, recent successes in microvirus isolation and improved metagenomic methodologies can now provide us with more accurate appraisals of their abundance, their hosts, and their interactions. The emerging picture is that phiX174 and its relatives are rather rare and atypical microviruses, and that a tremendous diversity of other microviruses is ready for exploration.

The large genetic and structural divergences between plants and fungi may hinder the transmission of viruses between these two kingdoms to some extent. However, recent accumulating evidence from virus phylogenetic analyses and the discovery of naturally occurring virus cross-infection suggest the occurrence of past and current transmissions of viruses between plants and plant-associated fungi. Moreover, artificial virus inoculation experiments showed that diverse plant viruses can multiply in fungi and vice versa. Thus, virus cross-infection between plants and fungi may play an important role in the spread, emergence, and evolution of both plant and fungal viruses and facilitate the interaction between them. In this review, we summarize current knowledge related to cross-kingdom virus infection in plants and fungi and further discuss the relevance of this new virological topic in the context of understanding virus spread and transmission in nature as well as developing control strategies for crop plant diseases.

There are at least 21 families of enveloped viruses that infect mammals, and many contain members of high concern for global human health. All enveloped viruses have a dedicated fusion protein or fusion complex that enacts the critical genome-releasing membrane fusion event that is essential before viral replication within the host cell interior can begin. Because all enveloped viruses enter cells by fusion, it behooves us to know how viral fusion proteins function. Viral fusion proteins are also major targets of neutralizing antibodies, and hence they serve as key vaccine immunogens. Here we review current concepts about viral membrane fusion proteins focusing on how they are triggered, structural intermediates between pre- and postfusion forms, and their interplay with the lipid bilayers they engage. We also discuss cellular and therapeutic interventions that thwart virus-cell membrane fusion.

Phase separation of viral biopolymers is a key factor in the formation of cytoplasmic viral inclusions, known as sites of virus replication and assembly. This review describes the mechanisms and factors that affect phase separation in viral replication and identifies potential areas for future research. Drawing inspiration from studies on cellular RNA-rich condensates, we compare the hierarchical coassembly of ribosomal RNAs and proteins in the nucleolus to the coordinated coassembly of viral RNAs and proteins taking place within viral factories in viruses containing segmented RNA genomes. We highlight the common characteristics of biomolecular condensates in viral replication and how this new understanding is reshaping our views of virus assembly mechanisms. Such studies have the potential to uncover unexplored antiviral strategies targeting these phase-separated states.

Bacteriophages are enigmatic entities that defy definition. Classically, they are specialist viruses that exclusively parasitize bacterial hosts. Yet this definition becomes limiting when we consider their ubiquity in the body coupled with their vast capacity to directly interact with the mammalian host. While phages certainly do not infect nor replicate within mammalian cells, they do interact with and gain unfettered access to the eukaryotic cell structure. With the growing appreciation for the human virome, coupled with our increased application of phages to patients within clinical settings, the potential impact of phage-mammalian interactions is progressively recognized. In this review, we provide a detailed mechanistic overview of how phages interact with the mammalian cell surface, the processes through which said phages are internalized by the cell, and the intracellular processing and fate of the phages. We then summarize the current state-of-the-field with respect to phage-mammalian interactions and their associations with health and disease states.

The nonsegmented, negative-strand RNA viruses (nsNSVs), also known as the order Mononegavirales, have a genome consisting of a single strand of negative-sense RNA. Integral to the nsNSV replication cycle is the viral polymerase, which is responsible for transcribing the viral genome, to produce an array of capped and polyadenylated messenger RNAs, and replicating it to produce new genomes. To perform the different steps that are necessary for these processes, the nsNSV polymerases undergo a series of coordinated conformational transitions. While much is still to be learned regarding the intersection of nsNSV polymerase dynamics, structure, and function, recently published polymerase structures, combined with a history of biochemical and molecular biology studies, have provided new insights into how nsNSV polymerases function as dynamic machines. In this review, we consider each of the steps involved in nsNSV transcription and replication and suggest how these relate to solved polymerase structures.

Protein synthesis by the ribosome is the final stage of biological information transfer and represents an irreversible commitment to gene expression. Accurate translation of messenger RNA is therefore essential to all life, and spontaneous errors by the translational machinery are highly infrequent (∼1/100,000 codons). Programmed −1 ribosomal frameshifting (−1PRF) is a mechanism in which the elongating ribosome is induced at high frequency to slip backward by one nucleotide at a defined position and to continue translation in the new reading frame. This is exploited as a translational regulation strategy by hundreds of RNA viruses, which rely on −1PRF during genome translation to control the stoichiometry of viral proteins. While early investigations of −1PRF focused on virological and biochemical aspects, the application of X-ray crystallography and cryo–electron microscopy (cryo-EM), and the advent of deep sequencing and single-molecule approaches have revealed unexpected structural diversity and mechanistic complexity. Molecular players from several model systems have now been characterized in detail, both in isolation and, more recently, in the context of the elongating ribosome. Here we provide a summary of recent advances and discuss to what extent a general model for −1PRF remains a useful way of thinking.

Human and simian immunodeficiency viruses (HIVs and SIVs, respectively) encode several small proteins (Vif, Vpr, Nef, Vpu, and Vpx) that are called accessory because they are not generally required for viral replication in cell culture. However, they play complex and important roles for viral immune evasion and spread in vivo. Here, we discuss the diverse functions and the relevance of the viral protein U (Vpu) that is expressed from a bicistronic RNA during the late stage of the viral replication cycle and found only in HIV-1 and closely related SIVs. It is well established that Vpu counteracts the restriction factor tetherin, mediates degradation of the primary viral CD4 receptors, and inhibits activation of the transcription factor nuclear factor kappa B. Recent studies identified additional activities and provided new insights into the sophisticated mechanisms by which Vpu enhances and prolongs the release of fully infectious viral particles. In addition, it has been shown that Vpu prevents superinfection not only by degrading CD4 but also by modulating DNA repair mechanisms to promote degradation of nuclear viral complementary DNA in cells that are already productively infected.

Negative-stranded RNA viruses are a large group of viruses that encode their genomes in RNA across multiple segments in an orientation antisense to messenger RNA. Their members infect broad ranges of hosts, and there are a number of notable human pathogens. Here, we examine the development of reverse genetic systems as applied to these virus families, emphasizing conserved approaches illustrated by some of the prominent members that cause significant human disease. We also describe the utility of their genetic systems in the development of reporter strains of the viruses and some biological insights made possible by their use. To conclude the review, we highlight some possible future uses of reporter viruses that not only will increase our basic understanding of how these viruses replicate and cause disease but also could inform the development of new approaches to therapeutically intervene.

N- and O-glycans are both important constituents of viral envelope glycoproteins. O-linked glycosylation can be initiated by any of 20 different human polypeptide O-acetylgalactosaminyl transferases, resulting in an important functional O-glycan heterogeneity. O-glycans are organized as solitary glycans or in clusters of multiple glycans forming mucin-like domains. They are functional both in the viral life cycle and in viral colonization of their host. Negatively charged O-glycans are crucial for the interactions between glycosaminoglycan-binding viruses and their host. A novel mechanism, based on controlled electrostatic repulsion, explains how such viruses solve the conflict between optimized viral attachment to target cells and efficient egress of progeny virus. Conserved solitary O-glycans appear important for viral uptake in target cells by contributing to viral envelope fusion. Dual roles of viral O-glycans in the host B cell immune response, either epitope blocking or epitope promoting, may be exploitable for vaccine development. Finally, specific virus-induced O-glycans may be involved in viremic spread.

Nuclear egress of herpesvirus capsids across the intact nuclear envelope is an exceptional vesicle-mediated nucleocytoplasmic translocation resulting in the delivery of herpesvirus capsids into the cytosol. Budding of the (nucleo)capsid at and scission from the inner nuclear membrane (INM) is mediated by the viral nuclear egress complex (NEC) resulting in a transiently enveloped virus particle in the perinuclear space followed by fusion of the primary envelope with the outer nuclear membrane (ONM). The dimeric NEC oligomerizes into a honeycomb-shaped coat underlining the INM to induce membrane curvature and scission. Mutational analyses complemented structural data defining functionally important regions. Questions remain, including where and when the NEC is formed and how membrane curvature is mediated, vesicle formation is regulated, and directionality is secured. The composition of the primary enveloped virion and the machinery mediating fusion of the primary envelope with the ONM is still debated. While NEC-mediated budding apparently follows a highly conserved mechanism, species and/or cell type–specific differences complicate understanding of later steps.

High-risk human papillomaviruses (HPVs) are associated with several human cancers. HPVs are small, DNA viruses that rely on host cell machinery for viral replication. The HPV life cycle takes place in the stratified epithelium, which is composed of different cell states, including terminally differentiating cells that are no longer active in the cell cycle. HPVs have evolved mechanisms to persist and replicate in the stratified epithelium by hijacking and modulating cellular pathways, including the DNA damage response (DDR). HPVs activate and exploit DDR pathways to promote viral replication, which in turn increases the susceptibility of the host cell to genomic instability and carcinogenesis. Here, we review recent advances in our understanding of the regulation of the host cell DDR by high-risk HPVs during the viral life cycle and discuss the potential cellular consequences of modulating DDR pathways.

Respiratory viruses, such as influenza viruses, cause significant morbidity and mortality worldwide through seasonal epidemics and sporadic pandemics. Influenza viruses transmit through multiple modes including contact (either direct or through a contaminated surface) and inhalation of expelled aerosols. Successful human to human transmission requires an infected donor who expels virus into the environment, a susceptible recipient, and persistence of the expelled virus within the environment. The relative efficiency of each mode can be altered by viral features, environmental parameters, donor and recipient host characteristics, and viral persistence. Interventions to mitigate transmission of influenza viruses can target any of these factors. In this review, we discuss many aspects of influenza virus transmission, including the systems to study it, as well as the impact of natural barriers and various nonpharmaceutical and pharmaceutical interventions.