Dynamics of eastern equine encephalitis virus during the 2019 outbreak in the Northeast United States.

Eastern equine encephalitis virus (EEEV) causes a rare but severe disease in horses and humans and is maintained in an enzootic transmission cycle between songbirds and Culiseta melanura mosquitoes. In 2019, the largest EEEV outbreak in the United States for more than 50 years occurred, centered in the Northeast. To explore the dynamics of the outbreak, we sequenced 80 isolates of EEEV and combined them with existing genomic data. We found that, similar to previous years, cases were driven by multiple independent but short-lived virus introductions into the Northeast from Florida. Once in the Northeast, we found that Massachusetts was important for regional spread. We found no evidence of any changes in viral, human, or bird factors which would explain the increase in cases in 2019, although the ecology of EEEV is complex and further data is required to explore these in more detail. By using detailed mosquito surveillance data collected by Massachusetts and Connecticut, however, we found that the abundance of Cs. melanura was exceptionally high in 2019, as was the EEEV infection rate. We employed these mosquito data to build a negative binomial regression model and applied it to estimate early season risks of human or horse cases. We found that the month of first detection of EEEV in mosquito surveillance data and vector index (abundance multiplied by infection rate) were predictive of cases later in the season. We therefore highlight the importance of mosquito surveillance programs as an integral part of public health and disease control.

Dynamics of Eastern equine encephalitis virus during the 2019 outbreak in the Northeast United States.

Eastern equine encephalitis virus (EEEV) causes a rare but severe disease in horses and humans, and is maintained in an enzootic transmission cycle between songbirds and Culiseta melanura mosquitoes. In 2019, the largest EEEV outbreak in the United States for more than 50 years occurred, centered in the Northeast. To explore the dynamics of the outbreak, we sequenced 80 isolates of EEEV and combined them with existing genomic data. We found that, like previous years, cases were driven by frequent short-lived virus introductions into the Northeast from Florida. Once in the Northeast, we found that Massachusetts was important for regional spread. We found no evidence of any changes in viral, human, or bird factors which would explain the increase in cases in 2019. By using detailed mosquito surveillance data collected by Massachusetts and Connecticut, however, we found that the abundance of Cs. melanura was exceptionally high in 2019, as was the EEEV infection rate. We employed these mosquito data to build a negative binomial regression model and applied it to estimate early season risks of human or horse cases. We found that the month of first detection of EEEV in mosquito surveillance data and vector index (abundance multiplied by infection rate) were predictive of cases later in the season. We therefore highlight the importance of mosquito surveillance programs as an integral part of public health and disease control.

Early Genomic Surveillance and Phylogeographic Analysis of Getah Virus, a Reemerging Arbovirus, in Livestock in China.

Getah virus (GETV) mainly causes disease in livestock and may pose an epidemic risk due to its expanding host range and the potential of long-distance dispersal through animal trade. Here, we used metagenomic next-generation sequencing (mNGS) to identify GETV as the pathogen responsible for reemerging swine disease in China and subsequently estimated key epidemiological parameters using phylodynamic and spatially-explicit phylogeographic approaches. The GETV isolates were able to replicate in a variety of cell lines, including human cells, and showed high pathogenicity in a mouse model, suggesting the potential for more mammal hosts. We obtained 16 complete genomes and 79 E2 gene sequences from viral strains collected in China from 2016 to 2021 through large-scale surveillance among livestock, pets, and mosquitoes. Our phylogenetic analysis revealed that three major GETV lineages are responsible for the current epidemic in livestock in China. We identified three potential positively selected sites and mutations of interest in E2, which may impact the transmissibility and pathogenicity of the virus. Phylodynamic inference of the GETV demographic dynamics identified an association between livestock meat consumption and the evolution of viral genetic diversity. Finally, phylogeographic reconstruction of GETV dispersal indicated that the sampled lineages have preferentially circulated within areas associated with relatively higher mean annual temperature and pig population density. Our results highlight the importance of continuous surveillance of GETV among livestock in southern Chinese regions associated with relatively high temperatures. IMPORTANCE Although livestock is known to be the primary reservoir of Getah virus (GETV) in Asian countries, where identification is largely based on serology, the evolutionary history and spatial epidemiology of GETV in these regions remain largely unknown. Through our sequencing efforts, we provided robust support for lineage delineation of GETV and identified three major lineages that are responsible for the current epidemic in livestock in China. We further analyzed genomic and epidemiological data to reconstruct the recent demographic and dispersal history of GETV in domestic animals in China and to explore the impact of environmental factors on its genetic diversity and its diffusion. Notably, except for livestock meat consumption, other pig-related factors such as the evolution of live pig transport and pork production do not show a significant association with the evolution of viral genetic diversity, pointing out that further studies should investigate the potential contribution of other host species to the GETV outbreak. Our analysis of GETV demonstrates the need for wider animal species surveillance and provides a baseline for future studies of the molecular epidemiology and early warning of emerging arboviruses in China.

Comparative Epidemiology of Rabbit Haemorrhagic Disease Virus Strains from Viral Sequence Data.

Since their introduction in 1859, European rabbits (Oryctolagus cuniculus) have had a devastating impact on agricultural production and biodiversity in Australia, with competition and land degradation by rabbits being one of the key threats to agricultural and biodiversity values in Australia. Biocontrol agents, with the most important being the rabbit haemorrhagic disease virus 1 (RHDV1), constitute the most important landscape-scale control strategies for rabbits in Australia. Monitoring field strain dynamics is complex and labour-intensive. Here, using phylodynamic models to analyse the available RHDV molecular data, we aimed to: investigate the epidemiology of various strains, use molecular data to date the emergence of new variants and evaluate whether different strains are outcompeting one another. We determined that the two main pathogenic lagoviruses variants in Australia (RHDV1 and RHDV2) have had similar dynamics since their release, although over different timeframes (substantially shorter for RHDV2). We also found a strong geographic difference in their activities and evidence of overall competition between the two viruses.

On the Use of Phylogeographic Inference to Infer the Dispersal History of Rabies Virus: A Review Study.

Rabies is a neglected zoonotic disease which is caused by negative strand RNA-viruses belonging to the genus Lyssavirus. Within this genus, rabies viruses circulate in a diverse set of mammalian reservoir hosts, is present worldwide, and is almost always fatal in non-vaccinated humans. Approximately 59,000 people are still estimated to die from rabies each year, leading to a global initiative to work towards the goal of zero human deaths from dog-mediated rabies by 2030, requiring scientific efforts from different research fields. The past decade has seen a much increased use of phylogeographic and phylodynamic analyses to study the evolution and spread of rabies virus. We here review published studies in these research areas, making a distinction between the geographic resolution associated with the available sequence data. We pay special attention to environmental factors that these studies found to be relevant to the spread of rabies virus. Importantly, we highlight a knowledge gap in terms of applying these methods when all required data were available but not fully exploited. We conclude with an overview of recent methodological developments that have yet to be applied in phylogeographic and phylodynamic analyses of rabies virus.

YMrCA: Improving Y-chromosomal ancestor time estimation for DNA kinship research.

The Y-chromosome is a valuable kinship indicator in family history and forensic research. To reconstruct genealogies, the time to the most recent common ancestor (tMRCA) between paternal relatives can be estimated through Y-STR analysis. Existing models are the stepwise mutation model (SMM, only one-step Y-STR changes) and the infinite allele model (IAM, new allele per Y-STR change). In this study, these mutation models and all existing tMRCA calculators were validated through a genetic-genealogy database containing 1,120 biologically related genealogical pairs confirmed by 46 Y-STRs with known tMRCA (18,109 generations). Consistent under- and overestimation and broad confidence intervals were observed, leading to dubious tMRCA estimates. This is because they do not include individual mutation rates or multi-step changes and ignore hidden multiple, back, or parallel modifications. To improve tMRCA estimation, we developed a user-friendly calculator, the "YMrCA", including all previously mentioned mutation characteristics. After extensive validation, we observed that the YMrCA calculator demonstrated a promising performance. The YMrCA yields a significantly higher tMRCA success rate (96%; +20%) and a lower tMRCA error (7; -3) compared to the mutation models and all online tMRCA calculators. Therefore, YMrCA offers the next step towards more objective tMRCA estimation for DNA kinship research.

Untangling introductions and persistence in COVID-19 resurgence in Europe.

After the first wave of SARS-CoV-2 infections in spring 2020, Europe experienced a resurgence of the virus starting in late summer 2020 that was deadlier and more difficult to contain1. Relaxed intervention measures and summer travel have been implicated as drivers of the second wave2. Here we build a phylogeographical model to evaluate how newly introduced lineages, as opposed to the rekindling of persistent lineages, contributed to the resurgence of COVID-19 in Europe. We inform this model using genomic, mobility and epidemiological data from 10 European countries and estimate that in many countries more than half of the lineages circulating in late summer resulted from new introductions since 15 June 2020. The success in onward transmission of newly introduced lineages was negatively associated with the local incidence of COVID-19 during this period. The pervasive spread of variants in summer 2020 highlights the threat of viral dissemination when restrictions are lifted, and this needs to be carefully considered in strategies to control the current spread of variants that are more transmissible and/or evade immunity. Our findings indicate that more effective and coordinated measures are required to contain the spread through cross-border travel even as vaccination is reducing disease burden.

Dispersal dynamics of SARS-CoV-2 lineages during the first epidemic wave in New York City.

During the first phase of the COVID-19 epidemic, New York City rapidly became the epicenter of the pandemic in the United States. While molecular phylogenetic analyses have previously highlighted multiple introductions and a period of cryptic community transmission within New York City, little is known about the circulation of SARS-CoV-2 within and among its boroughs. We here perform phylogeographic investigations to gain insights into the circulation of viral lineages during the first months of the New York City outbreak. Our analyses describe the dispersal dynamics of viral lineages at the state and city levels, illustrating that peripheral samples likely correspond to distinct dispersal events originating from the main metropolitan city areas. In line with the high prevalence recorded in this area, our results highlight the relatively important role of the borough of Queens as a transmission hub associated with higher local circulation and dispersal of viral lineages toward the surrounding boroughs.

SARS-CoV-2 European resurgence foretold: interplay of introductions and persistence by leveraging genomic and mobility data.

Following the first wave of SARS-CoV-2 infections in spring 2020, Europe experienced a resurgence of the virus starting late summer that was deadlier and more difficult to contain. Relaxed intervention measures and summer travel have been implicated as drivers of the second wave. Here, we build a phylogeographic model to evaluate how newly introduced lineages, as opposed to the rekindling of persistent lineages, contributed to the COVID-19 resurgence in Europe. We inform this model using genomic, mobility and epidemiological data from 10 West European countries and estimate that in many countries more than 50% of the lineages circulating in late summer resulted from new introductions since June 15th. The success in onwards transmission of these lineages is predicted by SARS-CoV-2 incidence during this period. Relatively early introductions from Spain into the United Kingdom contributed to the successful spread of the 20A.EU1/B.1.177 variant. The pervasive spread of variants that have not been associated with an advantage in transmissibility highlights the threat of novel variants of concern that emerged more recently and have been disseminated by holiday travel. Our findings indicate that more effective and coordinated measures are required to contain spread through cross-border travel.

Relax, Keep Walking - A Practical Guide to Continuous Phylogeographic Inference with BEAST.

Spatially explicit phylogeographic analyses can be performed with an inference framework that employs relaxed random walks to reconstruct phylogenetic dispersal histories in continuous space. This core model was first implemented 10 years ago and has opened up new opportunities in the field of phylodynamics, allowing researchers to map and analyze the spatial dissemination of rapidly evolving pathogens. We here provide a detailed and step-by-step guide on how to set up, run, and interpret continuous phylogeographic analyses using the programs BEAUti, BEAST, Tracer, and TreeAnnotator.

Rivers and landscape ecology of a plant virus, Rice yellow mottle virus along the Niger Valley.

To investigate the spread of Rice yellow mottle virus (RYMV) along the Niger River, regular sampling of virus isolates was conducted along 500 km of the Niger Valley in the Republic of Niger and was complemented by additional sampling in neighbouring countries in West Africa and Central Africa. The spread of RYMV into and within the Republic of Niger was inferred as a continuous process using a Bayesian statistical framework applied previously to reconstruct its dispersal history in West Africa, East Africa, and Madagascar. The spatial resolution along this section of the Niger River was the highest implemented for RYMV and possibly for any plant virus. We benefited from the results of early field surveys of the disease for the validation of the phylogeographic reconstruction and from the well-documented history of rice cultivation changes along the Niger River for their interpretation. As a prerequisite, the temporal signal of the RYMV data sets was revisited in the light of recent methodological advances. The role of the hydrographic network of the Niger Basin in RYMV spread was examined, and the link between virus population dynamics and the extent of irrigated rice was assessed. RYMV was introduced along the Niger River in the Republic of Niger in the early 1980s from areas to the southwest of the country where rice was increasingly grown. Viral spread was triggered by a major irrigation scheme made of a set of rice perimeters along the river valley. The subsequent spatial and temporal host continuity and the inoculum build-up allowed for a rapid spread of RYMV along the Niger River, upstream and downstream, over hundreds of kilometres, and led to the development of severe epidemics. There was no evidence of long-distance dissemination of the virus through natural water. Floating rice in the main meanders of the Middle Niger did not contribute to virus dispersal from West Africa to Central Africa. RYMV along the Niger River is an insightful example of how agricultural intensification favours pathogen emergence and spread.

Markov-Modulated Continuous-Time Markov Chains to Identify Site- and Branch-Specific Evolutionary Variation in BEAST.

Markov models of character substitution on phylogenies form the foundation of phylogenetic inference frameworks. Early models made the simplifying assumption that the substitution process is homogeneous over time and across sites in the molecular sequence alignment. While standard practice adopts extensions that accommodate heterogeneity of substitution rates across sites, heterogeneity in the process over time in a site-specific manner remains frequently overlooked. This is problematic, as evolutionary processes that act at the molecular level are highly variable, subjecting different sites to different selective constraints over time, impacting their substitution behavior. We propose incorporating time variability through Markov-modulated models (MMMs), which extend covarion-like models and allow the substitution process (including relative character exchange rates as well as the overall substitution rate) at individual sites to vary across lineages. We implement a general MMM framework in BEAST, a popular Bayesian phylogenetic inference software package, allowing researchers to compose a wide range of MMMs through flexible XML specification. Using examples from bacterial, viral, and plastid genome evolution, we show that MMMs impact phylogenetic tree estimation and can substantially improve model fit compared to standard substitution models. Through simulations, we show that marginal likelihood estimation accurately identifies the generative model and does not systematically prefer the more parameter-rich MMMs. To mitigate the increased computational demands associated with MMMs, our implementation exploits recent developments in BEAGLE, a high-performance computational library for phylogenetic inference. [Bayesian inference; BEAGLE; BEAST; covarion, heterotachy; Markov-modulated models; phylogenetics.].

A Phylodynamic Workflow to Rapidly Gain Insights into the Dispersal History and Dynamics of SARS-CoV-2 Lineages.

Since the start of the COVID-19 pandemic, an unprecedented number of genomic sequences of SARS-CoV-2 have been generated and shared with the scientific community. The unparalleled volume of available genetic data presents a unique opportunity to gain real-time insights into the virus transmission during the pandemic, but also a daunting computational hurdle if analyzed with gold-standard phylogeographic approaches. To tackle this practical limitation, we here describe and apply a rapid analytical pipeline to analyze the spatiotemporal dispersal history and dynamics of SARS-CoV-2 lineages. As a proof of concept, we focus on the Belgian epidemic, which has had one of the highest spatial densities of available SARS-CoV-2 genomes. Our pipeline has the potential to be quickly applied to other countries or regions, with key benefits in complementing epidemiological analyses in assessing the impact of intervention measures or their progressive easement.

Epidemiological hypothesis testing using a phylogeographic and phylodynamic framework.

Computational analyses of pathogen genomes are increasingly used to unravel the dispersal history and transmission dynamics of epidemics. Here, we show how to go beyond historical reconstructions and use spatially-explicit phylogeographic and phylodynamic approaches to formally test epidemiological hypotheses. We illustrate our approach by focusing on the West Nile virus (WNV) spread in North America that has substantially impacted public, veterinary, and wildlife health. We apply an analytical workflow to a comprehensive WNV genome collection to test the impact of environmental factors on the dispersal of viral lineages and on viral population genetic diversity through time. We find that WNV lineages tend to disperse faster in areas with higher temperatures and we identify temporal variation in temperature as a main predictor of viral genetic diversity through time. By contrasting inference with simulation, we find no evidence for viral lineages to preferentially circulate within the same migratory bird flyway, suggesting a substantial role for non-migratory birds or mosquito dispersal along the longitudinal gradient.

Hamiltonian Monte Carlo sampling to estimate past population dynamics using the skygrid coalescent model in a Bayesian phylogenetics framework.

Nonparametric coalescent-based models are often employed to infer past population dynamics over time. Several of these models, such as the skyride and skygrid models, are equipped with a block-updating Markov chain Monte Carlo sampling scheme to efficiently estimate model parameters. The advent of powerful computational hardware along with the use of high-performance libraries for statistical phylogenetics has, however, made the development of alternative estimation methods feasible. We here present the implementation and performance assessment of a Hamiltonian Monte Carlo gradient-based sampler to infer the parameters of the skygrid model. The skygrid is a popular and flexible coalescent-based model for estimating population dynamics over time and is available in BEAST 1.10.5, a widely-used software package for Bayesian pylogenetic and phylodynamic analysis. Taking into account the increased computational cost of gradient evaluation, we report substantial increases in effective sample size per time unit compared to the established block-updating sampler. We expect gradient-based samplers to assume an increasingly important role for different classes of parameters typically estimated in Bayesian phylogenetic and phylodynamic analyses.

Evolution and epidemic spread of SARS-CoV-2 in Brazil.

Brazil currently has one of the fastest-growing severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) epidemics in the world. Because of limited available data, assessments of the impact of nonpharmaceutical interventions (NPIs) on this virus spread remain challenging. Using a mobility-driven transmission model, we show that NPIs reduced the reproduction number from >3 to 1 to 1.6 in São Paulo and Rio de Janeiro. Sequencing of 427 new genomes and analysis of a geographically representative genomic dataset identified >100 international virus introductions in Brazil. We estimate that most (76%) of the Brazilian strains fell in three clades that were introduced from Europe between 22 February and 11 March 2020. During the early epidemic phase, we found that SARS-CoV-2 spread mostly locally and within state borders. After this period, despite sharp decreases in air travel, we estimated multiple exportations from large urban centers that coincided with a 25% increase in average traveled distances in national flights. This study sheds new light on the epidemic transmission and evolutionary trajectories of SARS-CoV-2 lineages in Brazil and provides evidence that current interventions remain insufficient to keep virus transmission under control in this country.

Online Bayesian Phylodynamic Inference in BEAST with Application to Epidemic Reconstruction.

Reconstructing pathogen dynamics from genetic data as they become available during an outbreak or epidemic represents an important statistical scenario in which observations arrive sequentially in time and one is interested in performing inference in an "online" fashion. Widely used Bayesian phylogenetic inference packages are not set up for this purpose, generally requiring one to recompute trees and evolutionary model parameters de novo when new data arrive. To accommodate increasing data flow in a Bayesian phylogenetic framework, we introduce a methodology to efficiently update the posterior distribution with newly available genetic data. Our procedure is implemented in the BEAST 1.10 software package, and relies on a distance-based measure to insert new taxa into the current estimate of the phylogeny and imputes plausible values for new model parameters to accommodate growing dimensionality. This augmentation creates informed starting values and re-uses optimally tuned transition kernels for posterior exploration of growing data sets, reducing the time necessary to converge to target posterior distributions. We apply our framework to data from the recent West African Ebola virus epidemic and demonstrate a considerable reduction in time required to obtain posterior estimates at different time points of the outbreak. Beyond epidemic monitoring, this framework easily finds other applications within the phylogenetics community, where changes in the data-in terms of alignment changes, sequence addition or removal-present common scenarios that can benefit from online inference.

Transmission dynamics of re-emerging rabies in domestic dogs of rural China.

Despite ongoing efforts to control transmission, rabies prevention remains a challenge in many developing countries, especially in rural areas of China where re-emerging rabies is under-reported due to a lack of sustained animal surveillance. By taking advantage of detailed genomic and epidemiological data for the re-emerging rabies outbreak in Yunnan Province, China, collected between 1999 and 2015, we reconstruct the demographic and dispersal history of domestic dog rabies virus (RABV) as well as the dynamics of dog-to-dog and dog-to-human transmission. Phylogeographic analyses reveal a lower diffusion coefficient than previously estimated for dog RABV dissemination in northern Africa. Furthermore, epidemiological analyses reveal transmission rates between dogs, as well as between dogs and humans, lower than estimates for Africa. Finally, we show that reconstructed epidemic history of RABV among dogs and the dynamics of rabid dogs are consistent with the recorded human rabies cases. This work illustrates the benefits of combining phylogeographic and epidemic modelling approaches for uncovering the spatiotemporal dynamics of zoonotic diseases, with both approaches providing estimates of key epidemiological parameters.

A Relaxed Directional Random Walk Model for Phylogenetic Trait Evolution.

Understanding the processes that give rise to quantitative measurements associated with molecular sequence data remains an important issue in statistical phylogenetics. Examples of such measurements include geographic coordinates in the context of phylogeography and phenotypic traits in the context of comparative studies. A popular approach is to model the evolution of continuously varying traits as a Brownian diffusion process acting on a phylogenetic tree. However, standard Brownian diffusion is quite restrictive and may not accurately characterize certain trait evolutionary processes. Here, we relax one of the major restrictions of standard Brownian diffusion by incorporating a nontrivial estimable mean into the process. We introduce a relaxed directional random walk (RDRW) model for the evolution of multivariate continuously varying traits along a phylogenetic tree. Notably, the RDRW model accommodates branch-specific variation of directional trends while preserving model identifiability. Furthermore, our development of a computationally efficient dynamic programming approach to compute the data likelihood enables scaling of our method to large data sets frequently encountered in phylogenetic comparative studies and viral evolution. We implement the RDRW model in a Bayesian inference framework to simultaneously reconstruct the evolutionary histories of molecular sequence data and associated multivariate continuous trait data, and provide tools to visualize evolutionary reconstructions. We demonstrate the performance of our model on synthetic data, and we illustrate its utility in two viral examples. First, we examine the spatiotemporal spread of HIV-1 in central Africa and show that the RDRW model uncovers a clearer, more detailed picture of the dynamics of viral dispersal than standard Brownian diffusion. Second, we study antigenic evolution in the context of HIV-1 resistance to three broadly neutralizing antibodies. Our analysis reveals evidence of a continuous drift at the HIV-1 population level towards enhanced resistance to neutralization by the VRC01 monoclonal antibody over the course of the epidemic. [Brownian Motion; Diffusion Processes; Phylodynamics; Phylogenetics; Phylogeography; Trait Evolution.].

Understanding Past Population Dynamics: Bayesian Coalescent-Based Modeling with Covariates.

Effective population size characterizes the genetic variability in a population and is a parameter of paramount importance in population genetics and evolutionary biology. Kingman's coalescent process enables inference of past population dynamics directly from molecular sequence data, and researchers have developed a number of flexible coalescent-based models for Bayesian nonparametric estimation of the effective population size as a function of time. Major goals of demographic reconstruction include identifying driving factors of effective population size, and understanding the association between the effective population size and such factors. Building upon Bayesian nonparametric coalescent-based approaches, we introduce a flexible framework that incorporates time-varying covariates that exploit Gaussian Markov random fields to achieve temporal smoothing of effective population size trajectories. To approximate the posterior distribution, we adapt efficient Markov chain Monte Carlo algorithms designed for highly structured Gaussian models. Incorporating covariates into the demographic inference framework enables the modeling of associations between the effective population size and covariates while accounting for uncertainty in population histories. Furthermore, it can lead to more precise estimates of population dynamics. We apply our model to four examples. We reconstruct the demographic history of raccoon rabies in North America and find a significant association with the spatiotemporal spread of the outbreak. Next, we examine the effective population size trajectory of the DENV-4 virus in Puerto Rico along with viral isolate count data and find similar cyclic patterns. We compare the population history of the HIV-1 CRF02_AG clade in Cameroon with HIV incidence and prevalence data and find that the effective population size is more reflective of incidence rate. Finally, we explore the hypothesis that the population dynamics of musk ox during the Late Quaternary period were related to climate change. [Coalescent; effective population size; Gaussian Markov random fields; phylodynamics; phylogenetics; population genetics.