Viral haemorrhagic septicaemia (VHS) caused by the rhabdovirus VHSV is economically the most important viral disease in European rainbow trout farming. Until 1989, this virus was mainly isolated from freshwater salmonids but in the last decade, it has also been isolated from an increasing number of free-living marine fish species. To study the genetic evolution of VHSV, the entire G gene from 74 isolates was analysed. VHSV from wild marine species caught in the Baltic Sea, Skagerrak, Kattegat, North Sea, and English Channel and European freshwater isolates, appeared to share a recent common ancestor. Based on the estimated nucleotide substitution rate, the ancestor of the European fresh water isolates was dated some 50 years ago. This finding fits with the initial reports in the 1950s on clinical observations of VHS in Danish freshwater rainbow trout farms. The study also indicates that European marine VHSV and the North American marine line separated approx. 500 years ago. The codon substitution rate among the freshwater VHSV isolates was found to be 2?5 times faster than among marine isolates. The data support the hypothesis of the marine environment being the original reservoir of VHSV and that the change in host range (to include rainbow trout) may have occurred several times. Virus from the marine environment will therefore continue to represent a threat to the trout aquaculture industry.

Infectious hematopoietic necrosis virus (IHNV) is a rhabdoviral pathogen that infects wild and cultured salmonid fish throughout the Pacific Northwest of North America. IHNV causes severe epidemics in young fish and can cause disease or occur asymptomatically in adults. In a broad survey of 323 IHNV field isolates, sequence analysis of a 303 nucleotide variable region within the glycoprotein gene revealed a maximum nucleotide diversity of 8?6 %, indicating low genetic diversity overall for this virus. Phylogenetic analysis revealed three major virus genogroups, designated U, M and L, which varied in topography and geographical range. Intragenogroup genetic diversity measures indicated that the M genogroup had three-to fourfold more diversity than the other genogroups and suggested relatively rapid evolution of the M genogroup and stasis within the U genogroup. We speculate that factors influencing IHNV evolution may have included ocean migration ranges of their salmonid host populations and anthropogenic effects associated with fish culture. INTRODUCTIONInfectious hematopoietic necrosis virus (IHNV) is a rhabdovirus that causes acute, systemic disease in salmonid fish and also occurs in asymptomatic fish hosts. The virus is currently endemic throughout the Pacific Northwest of North America, with a contiguous range extending from Alaska to California and inland to Idaho. Within this geographical area the host range of IHNV includes five species of Pacific salmon, Atlantic salmon and several trout species (Wolf, 1988;Bootland & Leong, 1999). The first reported epidemics of IHNV occurred in sockeye salmon (Oncorhynchus nerka) fry at Washington and Oregon fish hatcheries during the 1950s (Rucker et al., 1953;Guenther et al., 1959;Wingfield et al., 1969). Surveys indicated that IHNV was endemic in sockeye throughout Alaska by 1974(Grischkowsky & Amend, 1976, but the virus was not widespread in Washington and Oregon through the 1970s (Amend & Wood, 1972; Mulcahy et al., 1980;Pilcher & Fryer, 1980). Subsequently, two virus emergence events occurred in which IHNV became endemic in rainbow trout (O. mykiss) throughout the Hagerman Valley trout farming industry in southern Idaho between 1977(Busch, 1983 and in salmonids of the middle and lower Columbia River basin in the early 1980s (Groberg, 1983;Groberg & Fryer, 1983). In addition to cultured fish, IHNV is endemic in many wild salmonid stocks in the Pacific Northwest (Bootland & Leong, 1999).Due to the extensive economic losses caused by IHNV in fish culture facilities, the virus has been well characterized in biological, immunological and molecular biological studies (for reviews, see Wolf, 1988;Bootland & Leong, 1999). IHNV is the type species of the genus Novirhabdovirus, within the family Rhabdoviridae. Similar to other rhabdoviruses, IHNV has a linear single-stranded, negative-sense RNA genome of approximately 11 000 nucleotides. The IHNV genome contains six genes in the order 39-N-P-M-G-NV-L-59, representing the nucleocapsid, phosphoprotein, matrix protein, glyco...

The rapid identification of antimicrobial resistance is essential for effective treatment of highly resistant Mycobacterium tuberculosis. Whole-genome sequencing provides comprehensive data on resistance mutations and strain typing for monitoring transmission, but unlike for conventional molecular tests, this has previously been achievable only from cultures of M. tuberculosis. Here we describe a method utilizing biotinylated RNA baits designed specifically for M. tuberculosis DNA to capture full M. tuberculosis genomes directly from infected sputum samples, allowing whole-genome sequencing without the requirement of culture. This was carried out on 24 smear-positive sputum samples, collected from the United Kingdom and Lithuania where a matched culture sample was available, and 2 samples that had failed to grow in culture. M. tuberculosis sequencing data were obtained directly from all 24 smear-positive culture-positive sputa, of which 20 were of high quality (>20؋ depth and >90% of the genome covered). Results were compared with those of conventional molecular and culture-based methods, and high levels of concordance between phenotypical resistance and predicted resistance based on genotype were observed. High-quality sequence data were obtained from one smear-positive culture-negative case. This study demonstrated for the first time the successful and accurate sequencing of M. tuberculosis genomes directly from uncultured sputa. Identification of known resistance mutations within a week of sample receipt offers the prospect for personalized rather than empirical treatment of drug-resistant tuberculosis, including the use of antimicrobial-sparing regimens, leading to improved outcomes.T he global incidence of multidrug-resistant (MDR), extensively drug-resistant (XDR), and totally drug-resistant tuberculosis (TB) has risen over the last decade (1), making it increasingly important to rapidly and accurately detect resistance. The gold standard for antimicrobial resistance testing relies on bacterial culture, which can take upwards of several weeks for Mycobacterium tuberculosis. Molecular tests, such as the Xpert (MTB/RIF) and line probe assays, which can be used directly on sputum have improved identification of MDR M. tuberculosis but are able to identify only limited numbers of specific resistance mutations (2, 3).Whole-genome sequencing (WGS) of bacterial genomes allows simultaneous identification of all known resistance mutations as well as markers with which transmission can be monitored (4). WGS of M. tuberculosis provides resolution superior to that of other current methods such as spoligotyping and mycobacterial interspersed repetitive-unit-variable-number tandemrepeat (MIRU-VNTR) analysis for strain genotyping (5), and its usefulness in defining outbreaks has been demonstrated previously (6-9). Currently, however, WGS of M. tuberculosis requires prior bacterial enrichment by culturing and most outbreak studies have therefore been retrospective (6-8). Recently, WGS of M. tuberculosis has been achieved ...

Human cytomegalovirus (HCMV) infects most of the population worldwide, persisting throughout the host's life in a latent state with periodic episodes of reactivation. While typically asymptomatic, HCMV can cause fatal disease among congenitally infected infants and immunocompromised patients. These clinical issues are compounded by the emergence of antiviral resistance and the absence of an effective vaccine, the development of which is likely complicated by the numerous immune evasins encoded by HCMV to counter the host's adaptive immune responses, a feature that facilitates frequent super-infections. Understanding the evolutionary dynamics of HCMV is essential for the development of effective new drugs and vaccines. By comparing viral genomes from uncultivated or low-passaged clinical samples of diverse origins, we observe evidence of frequent homologous recombination events, both recent and ancient, and no structure of HCMV genetic diversity at the whole-genome scale. Analysis of individual gene-scale loci reveals a striking dichotomy: while most of the genome is highly conserved, recombines essentially freely and has evolved under purifying selection, 21 genes display extreme diversity, structured into distinct genotypes that do not recombine with each other. Most of these hyper-variable genes encode glycoproteins involved in cell entry or escape of host immunity. Evidence that half of them have diverged through episodes of intense positive selection suggests that rapid evolution of hyper-variable loci is likely driven by interactions with host immunity. It appears that this process is enabled by recombination unlinking hyper-variable loci from strongly constrained neighboring sites. It is conceivable that viral mechanisms facilitating super-infection have evolved to promote recombination between diverged genotypes, allowing the virus to continuously diversify at key loci to escape immune detection, while maintaining a genome optimally adapted to its asymptomatic infectious lifecycle.

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