Viruses and mobile genetic elements are molecular parasites or symbionts that coevolve with nearly all forms of cellular life. The route of virus replication and protein expression is determined by the viral genome type. Comparison of these routes led to the classification of viruses into seven "Baltimore classes" (BCs) that define the major features of virus reproduction. However, recent phylogenomic studies identified multiple evolutionary connections among viruses within each of the BCs as well as between different classes. Due to the modular organization of virus genomes, these relationships defy simple representation as lines of descent but rather form complex networks. Phylogenetic analyses of virus hallmark genes combined with analyses of gene-sharing networks show that replication modules of five BCs (three classes of RNA viruses and two classes of reverse-transcribing viruses) evolved from a common ancestor that encoded an RNA-directed RNA polymerase or a reverse transcriptase. Bona fide viruses evolved from this ancestor on multiple, independent occasions via the recruitment of distinct cellular proteins as capsid subunits and other structural components of virions. The single-stranded DNA (ssDNA) viruses are a polyphyletic class, with different groups evolving by recombination between rolling-circle-replicating plasmids, which contributed the replication protein, and positive-sense RNA viruses, which contributed the capsid protein. The double-stranded DNA (dsDNA) viruses are distributed among several large monophyletic groups and arose via the combination of distinct structural modules with equally diverse replication modules. Phylogenomic analyses reveal the finer structure of evolutionary connections among RNA viruses and reverse-transcribing viruses, ssDNA viruses, and large subsets of dsDNA viruses. Taken together, these analyses allow us to outline the global organization of the virus world. Here, we describe the key aspects of this organization and propose a comprehensive hierarchical taxonomy of viruses.

https://mmbr.asm.org/content/mmbr/84/2/e00061-19.full-text.pdf

Global Organization and Proposed Megataxonomy of the Virus World

Viruses are a significant player in many biosphere and human ecosystems, but most signals remain “hidden” in metagenomic/metatranscriptomic sequence datasets due to the lack of universal gene markers, database representatives, and insufficiently advanced identification tools. Here, we introduce VirSorter2, a DNA and RNA virus identification tool that leverages genome-informed database advances across a collection of customized automatic classifiers to improve the accuracy and range of virus sequence detection. When benchmarked against genomes from both isolated and uncultivated viruses, VirSorter2 uniquely performed consistently with high accuracy (F1-score > 0.8) across viral diversity, while all other tools under-detected viruses outside of the group most represented in reference databases (i.e., those in the order Caudovirales). Among the tools evaluated, VirSorter2 was also uniquely able to minimize errors associated with atypical cellular sequences including eukaryotic genomes and plasmids. Finally, as the virosphere exploration unravels novel viral sequences, VirSorter2’s modular design makes it inherently able to expand to new types of viruses via the design of new classifiers to maintain maximal sensitivity and specificity. With multi-classifier and modular design, VirSorter2 demonstrates higher overall accuracy across major viral groups and will advance our knowledge of virus evolution, diversity, and virus-microbe interaction in various ecosystems. Source code of VirSorter2 is freely available (
 https://bitbucket.org/MAVERICLab/virsorter2
 ), and VirSorter2 is also available both on bioconda and as an iVirus app on CyVerse (
 https://de.cyverse.org/de
 ).

/pdf/virsorter2-a-multi-classifier-expert-guided-approach-to-17ci6sjy50.pdf

VirSorter2: a multi-classifier, expert-guided approach to detect diverse DNA and RNA viruses.

Increasing reports of antimicrobial resistance and limited new antibiotic discoveries and development have fuelled innovation in other research fields and led to a revitalization of bacteriophage (phage) studies in the Western world. Phage therapy mainly utilizes obligately lytic phages to kill their respective bacterial hosts, while leaving human cells intact and reducing the broader impact on commensal bacteria that often results from antibiotic use. Phage therapy is rapidly evolving and has resulted in cases of life-saving therapeutic use and multiple clinical trials. However, one of the biggest challenges this antibiotic alternative faces relates to regulations and policy surrounding clinical use and implementation beyond compassionate cases. This review discusses the multi-drug resistant Gram-negative pathogens of highest critical priority and summarizes the current state-of-the-art in phage therapy targeting these organisms. It also examines phage therapy in humans in general and the approaches different countries have taken to introduce it into clinical practice and policy. We aim to highlight the rapidly advancing field of phage therapy and the challenges that lie ahead as the world shifts away from complete reliance on antibiotics.

/pdf/bacteriophage-therapy-clinical-trials-and-regulatory-hurdles-2r41bwka06.pdf

Bacteriophage Therapy: Clinical Trials and Regulatory Hurdles.

The COVID-19 pandemic has severely impacted public health and the worldwide economy Converging evidence from the current pandemic, previous outbreaks and controlled experiments indicates that SARS-CoVs are present in wastewater for several days, leading to potential health risks via waterborne and aerosolized wastewater pathways Conventional wastewater treatment provides only partial removal of SARS-CoVs, thus safe disposal or reuse will depend on the efficacy of final disinfection This underscores the need for a risk assessment and management framework tailored to SARS-CoV-2 transmission via wastewater, including new tools for environmental surveillance, ensuring adequate disinfection as a component of overall COVID-19 pandemic containment Converging evidence indicates that SARS-CoVs are present in wastewater for several days with potential health risks This Review analyses knowledge about such risks as well as the potential spread of SARS-CoVs in waterborne, waterborne–aerosolized and waterborne–foodborne pathways during a pandemic

/pdf/rethinking-wastewater-risks-and-monitoring-in-light-of-the-26vv4l5zrl.pdf

Rethinking wastewater risks and monitoring in light of the COVID-19 pandemic

Bacteriophage (phage) taxonomy has been in flux since its inception over four decades ago. Genome sequencing has put pressure on the classification system and recent years have seen significant changes to phage taxonomy. Here, we reflect on the state of phage taxonomy and provide a roadmap for the future, including the abolition of the order Caudovirales and the families Myoviridae, Podoviridae, and Siphoviridae. Furthermore, we specify guidelines for the demarcation of species, genus, subfamily and family-level ranks of tailed phage taxonomy.

https://www.mdpi.com/1999-4915/13/3/506/pdf

A Roadmap for Genome-Based Phage Taxonomy.

The first sequenced genome was that of the 3569-nucleotide single-stranded RNA (ssRNA) bacteriophage MS2. Despite the recent accumulation of vast amounts of DNA and RNA sequence data, only 12 representative ssRNA phage genome sequences are available from the NCBI Genome database (June 2019). The difficulty in detecting RNA phages in metagenomic datasets raises questions as to their abundance, taxonomic structure, and ecological importance. In this study, we iteratively applied profile hidden Markov models to detect conserved ssRNA phage proteins in 82 publicly available metatranscriptomic datasets generated from activated sludge and aquatic environments. We identified 15,611 nonredundant ssRNA phage sequences, including 1015 near-complete genomes. This expansion in the number of known sequences enabled us to complete a phylogenetic assessment of both sequences identified in this study and known ssRNA phage genomes. Our expansion of these viruses from two environments suggests that they have been overlooked within microbiome studies.

Expansion of known ssRNA phage genomes: From tens to over a thousand

Bacteriophages (phages) or bacterial viruses have long been proposed as an alternative therapy against antibiotic-resistant bacteria such as Escherichia coli Even though poorly documented in the scientific literature, a long clinical history of phage therapy in countries such as Russia and Georgia suggests potential value in the use of phages as antibacterial agents. Escherichia coli is responsible for a wide range of diseases, intestinal (diarrhoea) and extraintestinal (UTI, septicaemia, pneumoniae, meningitis), making it an ideal target for phage therapy. This review discusses the latest research focusing on the potential of phage therapy to tackle E. coli-related illnesses. No intact phages are approved in EU or USA for human therapeutic use, but many successful in vitro and in vivo studies have been reported. However, additional research focused on in vivo multispecies models and human trials are required if phage therapy targeting E. coli pathotypes can be a story with happy end.

Phage therapy targeting Escherichia coli-a story with no end?

The number of novel bacteriophage sequences has expanded significantly as a result of many metagenomic studies of phage populations in diverse environments. Most of these novel sequences bear little or no homology to existing databases (referred to as the "viral dark matter"). Also, these sequences are primarily derived from DNA-encoded bacteriophages (phages) with few RNA phages included. Despite the rapid advancements in high-throughput sequencing, few studies enrich for RNA viruses, i.e., target viral rather than cellular fraction and/or RNA rather than DNA via a reverse transcriptase step, in an attempt to capture the RNA viruses present in a microbial communities. It is timely to compile existing and relevant information about RNA phages to provide an insight into many of their important biological features, which should aid in sequence-based discovery and in their subsequent annotation. Without comprehensive studies, the biological significance of RNA phages has been largely ignored. Future bacteriophage studies should be adapted to ensure they are properly represented in phageomic studies.

RNA Phage Biology in a Metagenomic Era

The human gut is colonised by a vast array of microbes that include bacteria, viruses, fungi, and archaea. While interest in these microbial entities has largely focused on the bacterial constituents, recently the viral component has attracted more attention. Metagenomic advances, compared to classical isolation procedures, have greatly enhanced our understanding of the composition, diversity, and function of viruses in the human microbiome (virome). We highlight that viral extraction methodologies are crucial in terms of identifying and characterising communities of viruses infecting eukaryotes and bacteria. Different viral extraction protocols, including those used in some of the most significant human virome publications to date, have introduced biases affecting their a overall conclusions. It is important that protocol variations should be clearly highlighted across studies, with the ultimate goal of identifying and acknowledging biases associated with different protocols and, perhaps, the generation of an unbiased and standardised method for examining this portion of the human microbiome.

Biases in Viral Metagenomics-Based Detection, Cataloguing and Quantification of Bacteriophage Genomes in Human Faeces, a Review

The vast majority of described prokaryotic viruses have double-stranded or single-stranded DNA or double-stranded RNA genomes. Until 2020, a mere four prokaryotic single-stranded, positive-sense RNA viruses have been classified in two genera (Riboviria; Lenarviricota; Allassoviricetes; Leviviridae). Several recent metagenomic and metatranscriptomic studies revealed a vastly greater diversity of these viruses in prokaryotic soil communities than ever anticipated. Phylogenetic analysis of these newly discovered viruses prompted the reorganization of class Allassoviricetes, now renamed Leviviricetes, to include two orders, Norzivirales and Timlovirales, and a total of six families, 428 genera and 882 species. Here we outline the new taxonomy of Leviviricetes, approved and ratified in 2021 by the International Committee on Taxonomy of Viruses, and describe open-access hidden Markov models to accommodate the anticipated identification and future classification of hundreds, if not thousands, of additional class members into this new taxonomic framework.

Julie Callanan

Papers

Expansion of known ssRNA phage genomes: From tens to over a thousand

Phage therapy targeting Escherichia coli-a story with no end?

RNA Phage Biology in a Metagenomic Era

Biases in Viral Metagenomics-Based Detection, Cataloguing and Quantification of Bacteriophage Genomes in Human Faeces, a Review

Leviviricetes: expanding and restructuring the taxonomy of bacteria-infecting single-stranded RNA viruses.