Tommy Tsan-Yuk Lam

The genesis and source of the H7N9 influenza viruses causing human infections in China.

A novel H7N9 influenza A virus first detected in March 2013 has since caused more than 130 human infections in China, resulting in 40 deaths. Preliminary analyses suggest that the virus is a reassortant of H7, N9 and H9N2 avian influenza viruses, and carries some amino acids associated with mammalian receptor binding, raising concerns of a new pandemic. However, neither the source populations of the H7N9 outbreak lineage nor the conditions for its genesis are fully known. Using a combination of active surveillance, screening of virus archives, and evolutionary analyses, here we show that H7 viruses probably transferred from domestic duck to chicken populations in China on at least two independent occasions. We show that the H7 viruses subsequently reassorted with enzootic H9N2 viruses to generate the H7N9 outbreak lineage, and a related previously unrecognized H7N7 lineage. The H7N9 outbreak lineage has spread over a large geographic region and is prevalent in chickens at live poultry markets, which are thought to be the immediate source of human infections. Whether the H7N9 outbreak lineage has, or will, become enzootic in China and neighbouring regions requires further investigation. The discovery here of a related H7N7 influenza virus in chickens that has the ability to infect mammals experimentally, suggests that H7 viruses may pose threats beyond the current outbreak. The continuing prevalence of H7 viruses in poultry could lead to the generation of highly pathogenic variants and further sporadic human infections, with a continued risk of the virus acquiring human-to-human transmissibility.

Exploring the temporal structure of heterochronous sequences using TempEst (formerly Path-O-Gen)

Gene sequences sampled at different points in time can be used to infer molecular phylogenies on a natural timescale of months or years, provided that the sequences in question undergo measurable amounts of evolutionary change between sampling times. Data sets with this property are termed heterochronous and have become increasingly common in several fields of biology, most notably the molecular epidemiology of rapidly evolving viruses. Here we introduce the cross-platform software tool, TempEst (formerly known as Path-O-Gen), for the visualization and analysis of temporally sampled sequence data. Given a molecular phylogeny and the dates of sampling for each sequence, TempEst uses an interactive regression approach to explore the association between genetic divergence through time and sampling dates. TempEst can be used to (1) assess whether there is sufficient temporal signal in the data to proceed with phylogenetic molecular clock analysis, and (2) identify sequences whose genetic divergence and sampling date are incongruent. Examination of the latter can help identify data quality problems, including errors in data annotation, sample contamination, sequence recombination, or alignment error. We recommend that all users of the molecular clock models implemented in BEAST first check their data using TempEst prior to analysis.

The ever-expanding diversity of porcine reproductive and respiratory syndrome virus

Porcine reproductive and respiratory syndrome (PRRS) virus appeared 20 years ago as the cause of a new disease in swine. Today PRRS is the most significant swine disease worldwide in spite of intensive immunological interventions. The virus showed remarkable genetic variation with two geographically distinct genotypes at the time of its discovery, indicating the possibility of prolonged evolutionary divergence prior to its appearance as a swine pathogen. Since then, both type 1 and type 2 have spread geographically, radiated genetically, and acquired new phenotypic characteristics, especially increased virulence. Here, we explore various hypotheses that might account for rapid expansion and diversification of PRRSV, including mechanisms specific to PRRSV and other arteriviruses, cellular modification processes, and immunological selection. Phylogenetic analysis of PRRSV has provided a broadly applicable means to relate diverse isolates, but it does not explain biological variation in virulence or immunological cross-protection. We present other methods of classification and review their limitations. Major questions about PRRSV remain unanswered despite intensive investigation, suggesting that the interaction of PRRSV with pigs involves novel biological processes that may be relevant to other RNA virus and host interactions.

Evolutionary Genetics of Human Enterovirus 71: Origin, Population Dynamics, Natural Selection, and Seasonal Periodicity of the VP1 Gene

ABSTRACT Human enterovirus 71 (EV-71) is one of the major etiologic causes of hand, foot, and mouth disease (HFMD) among young children worldwide, with fatal instances of neurological complications becoming increasingly common. Global VP1 capsid sequences (n = 628) sampled over 4 decades were collected and subjected to comprehensive evolutionary analysis using a suite of phylogenetic and population genetic methods. We estimated that the common ancestor of human EV-71 likely emerged around 1941 (95% confidence interval [CI], 1929 to 1952), subsequently diverging into three genogroups: B, C, and the now extinct genogroup A. Genealogical analysis revealed that diverse lineages of genogroup B and C (subgenogroups B1 to B5 and C1 to C5) have each circulated cryptically in the human population for up to 5 years before causing large HFMD outbreaks, indicating the quiescent persistence of EV-71 in human populations. Estimated phylogenies showed a complex pattern of spatial structure within well-sampled subgenogroups, suggesting endemicity with occasional lineage migration among locations, such that past HFMD epidemics are unlikely to be linked to continuous transmission of a single strain of virus. In addition, rises in genetic diversity are correlated with the onset of epidemics, driven in part by the emergence of novel EV-71 subgenogroups. Using subgenogroup C1 as a model, we observe temporal strain replacement through time, and we investigate the evidence for positive selection at VP1 immunogenic sites. We discuss the consequences of the evolutionary dynamics of EV-71 for vaccine design and compare its phylodynamic behavior with that of influenza virus.

Molecular epidemiology of PRRSV: a phylogenetic perspective.

Since its first discovery two decades ago, porcine reproductive and respiratory syndrome virus (PRRSV) has been the subject of intensive research due to its huge impact on the worldwide swine industry. Thanks to the phylogenetic analyses, much has been learned concerning the genetic diversity and evolution history of the virus. In this review, we focused on the evolutionary and epidemiological aspects of PRRSV from a phylogenetic perspective. We first described the diversity and transmission dynamics of Type 1 and 2 PRRSV, respectively. Then, we focused on the more ancient evolutionary history of PRRSV: the time of onset of all existing PRRSV and an origin hypothesis were discussed. Finally, we summarized the results from previous recombination studies to assess the potential impact of recombination on the virus epidemiology.

ggtree: an R package for visualization and annotation of phylogenetic trees with their covariates and other associated data

Summary We present an r package, ggtree, which provides programmable visualization and annotation of phylogenetic trees. ggtree can read more tree file formats than other softwares, including newick, nexus, NHX, phylip and jplace formats, and support visualization of phylo, multiphylo, phylo4, phylo4d, obkdata and phyloseq tree objects defined in other r packages. It can also extract the tree/branch/node-specific and other data from the analysis outputs of beast, epa, hyphy, paml, phylodog, pplacer, r8s, raxml and revbayes software, and allows using these data to annotate the tree. The package allows colouring and annotation of a tree by numerical/categorical node attributes, manipulating a tree by rotating, collapsing and zooming out clades, highlighting user selected clades or operational taxonomic units and exploration of a large tree by zooming into a selected portion. A two-dimensional tree can be drawn by scaling the tree width based on an attribute of the nodes. A tree can be annotated with an associated numerical matrix (as a heat map), multiple sequence alignment, subplots or silhouette images. The package ggtree is released under the artistic-2.0 license. The source code and documents are freely available through bioconductor (http://www.bioconductor.org/packages/ggtree).

Phylogeny-Based Evolutionary, Demographical, and Geographical Dissection of North American Type 2 Porcine Reproductive and Respiratory Syndrome Viruses

ABSTRACT Type 2 (or North American-like) porcine reproductive and respiratory syndrome virus (PRRSV) was first recorded in 1987 in the United States and now occurs in most commercial swine industries throughout the world. In this study, we investigated the epidemiological and evolutionary behaviors of type 2 PRRSV. Based on phylogenetic analyses of 8,624 ORF5 sequences, we described a comprehensive picture of the diversity of type 2 PRRSVs and systematically classified all available sequences into lineages and sublineages, including a number of previously undescribed lineages. With the rapid growth of sequence deposition into the databases, it would be technically difficult for veterinary researchers to genotype their sequences by reanalyzing all sequences in the databases. To this end, a set of reference sequences was established based on our classification system, which represents the principal diversity of all available sequences and can readily be used for further genotyping studies. In addition, we further investigated the demographic histories of these lineages and sublineages by using Bayesian coalescence analyses, providing evolutionary insights into several important epidemiological events of type 2 PRRSV. Moreover, by using a phylogeographic approach, we were able to estimate the transmission frequencies between the pig-producing states in the United States and identified several states as the major sources of viral spread, i.e., “transmission centers.” In summary, this study represents the most extensive phylogenetic analyses of type 2 PRRSV to date, providing a basis for future genotyping studies and dissecting the epidemiology of type 2 PRRSV from phylogenetic perspectives.

Dissemination, divergence and establishment of H7N9 influenza viruses in China

Since 2013 the occurrence of human infections by a novel avian H7N9 influenza virus in China has demonstrated the continuing threat posed by zoonotic pathogens. Although the first outbreak wave that was centred on eastern China was seemingly averted, human infections recurred in October 2013 (refs 3, 4, 5, 6, 7). It is unclear how the H7N9 virus re-emerged and how it will develop further; potentially it may become a long-term threat to public health. Here we show that H7N9 viruses have spread from eastern to southern China and become persistent in chickens, which has led to the establishment of multiple regionally distinct lineages with different reassortant genotypes. Repeated introductions of viruses from Zhejiang to other provinces and the presence of H7N9 viruses at live poultry markets have fuelled the recurrence of human infections. This rapid expansion of the geographical distribution and genetic diversity of the H7N9 viruses poses a direct challenge to current disease control systems. Our results also suggest that H7N9 viruses have become enzootic in China and may spread beyond the region, following the pattern previously observed with H5N1 and H9N2 influenza viruses.

Molecular Characterization of the Fecal Microbiota in Patients with Nonalcoholic Steatohepatitis - A Longitudinal Study

Background The human gut microbiota has profound influence on host metabolism and immunity. This study characterized the fecal microbiota in patients with nonalcoholic steatohepatitis (NASH). The relationship between microbiota changes and changes in hepatic steatosis was also studied. Methods Fecal microbiota of histology-proven NASH patients and healthy controls was analyzed by 16S ribosomal RNA pyrosequencing. NASH patients were from a previously reported randomized trial on probiotic treatment. Proton-magnetic resonance spectroscopy was performed to monitor changes in intrahepatic triglyceride content (IHTG). Results A total of 420,344 16S sequences with acceptable quality were obtained from 16 NASH patients and 22 controls. NASH patients had lower fecal abundance of Faecalibacterium and Anaerosporobacter but higher abundance of Parabacteroides and Allisonella. Partial least-square discriminant analysis yielded a model of 10 genera that discriminated NASH patients from controls. At month 6, 6 of 7 patients in the probiotic group and 4 of 9 patients in the usual care group had improvement in IHTG (P = 0.15). Improvement in IHTG was associated with a reduction in the abundance of Firmicutes (R2 = 0.4820, P = 0.0028) and increase in Bacteroidetes (R2 = 0.4366, P = 0.0053). This was accompanied by corresponding changes at the class, order and genus levels. In contrast, bacterial biodiversity did not differ between NASH patients and controls, and did not change with probiotic treatment. Conclusions NASH patients have fecal dysbiosis, and changes in microbiota correlate with improvement in hepatic steatosis. Further studies are required to investigate the mechanism underlying the interaction between gut microbes and the liver.

Full-Genome Deep Sequencing and Phylogenetic Analysis of Novel Human Betacoronavirus

A novel betacoronavirus associated with lethal respiratory and renal complications was recently identified in patients from several countries in the Middle East. We report the deep genome sequencing of the virus directly from a patient’s sputum sample. Our high-throughput sequencing yielded a substantial depth of genome sequence assembly and showed the minority viral variants in the specimen. Detailed phylogenetic analysis of the virus genome (England/Qatar/2012) revealed its close relationship to European bat coronaviruses circulating among the bat species of the Vespertilionidae family. Molecular clock analysis showed that the 2 human infections of this betacoronavirus in June 2012 (EMC/2012) and September 2012 (England/Qatar/2012) share a common virus ancestor most likely considerably before early 2012, suggesting the human diversity is the result of multiple zoonotic events.