Biology subjects

Pybus, O. G.

Publications and source records attributed to Pybus, O. G..

32 records · Page 2Linked to original sources

bioRxiv2019

Oropouche virus cases identified in Ecuador using an optimised rRT-PCR informed by metagenomic sequencing

Oropouche virus (OROV) is responsible for outbreaks of Oropouche fever in parts of South America. We recently identified and isolated OROV from a febrile Ecuadorian patient, however, a previously published rRT-PCR assay did not detect OROV in the patient sample. A primer mismatch to the Ecuadorian OROV lineage was identified from metagenomic sequencing data. We report the optimisation of an rRT-PCR assay for the Ecuadorian OROV lineage, which subsequently identified a further five cases in a cohort of 196 febrile patients. We isolated OROV via cell culture and developed an algorithmically-designed primer set for whole-genome amplification of the virus. Metagenomic sequencing of the patient samples provided OROV genome coverage ranging from 68 - 99%. The additional cases formed a single phylogenetic cluster together with the initial case. OROV should be considered as a differential diagnosis for Ecuadorian patients with febrile illness to avoid mis-diagnosis with other circulating pathogens.

genomics↗

bioRxiv2019

The evolutionary dynamics of Oropouche Virus (OROV) in South America

The Amazon basin is host to numerous arthropod-borne viral pathogens that cause febrile disease in humans. Among these, Oropouche orthobunyavirus (OROV) is a relatively understudied member of the Peribunyavirales that causes periodic outbreaks in human populations in Brazil and other South American countries. Although several studies have described the genetic diversity of the virus, the evolutionary processes that shape the viral genome remain poorly understood. Here we present a comprehensive study of the genomic dynamics of OROV that encompasses phylogenetic analysis, evolutionary rate estimates, inference of natural selective pressures, recombination and reassortment, and structural analysis of OROV variants. Our study includes all available published sequences, as well as a set of new OROV genomes sequences obtained from patients in Ecuador, representing the first set of viral genomes from this country. Our results show that differing evolutionary processes on the three segments that encompass the viral genome lead to variable evolutionary rates and TMRCAs that could be explained by cryptic reassortment. We also present the discovery of previously unobserved putative N-linked glycosylation sites, and codons which evolve under positive selection on the viral surface proteins, and discuss the potential role of these features in the evolution of the virus through a combined phylogenetic and structural approach.

evolutionary biology↗

bioRxiv2019

Repertoire-wide phylogenetic models of B cell molecular evolution reveal evolutionary signatures of aging and vaccination

In order to produce effective antibodies, B cells undergo rapid somatic hypermutation (SHM) and selection for binding affinity to antigen via a process called affinity maturation. The similarities between this process and evolution by natural selection have led many groups to use evolutionary and phylogenetic methods to characterize the development of immunological memory, vaccination, and other processes that depend on affinity maturation. However, these applications are limited by several features of affinity maturation that violate assumptions in standard phylogenetic models. Further, most phylogenetic models are designed to be applied to individual lineages comprising genetically diverse sequences, while B cell repertoires often consist of hundreds to thousands of separate low-diversity lineages. Here, we introduce a hierarchical phylogenetic framework that incorporates the unique features of SHM, and integrates information from all lineages in a repertoire to more precisely estimate model parameters. We demonstrate the power of this approach by characterizing previously un-described phenomena in affinity maturation. First, we find evidence consistent with age related changes in SHM hot-and cold-spot motifs. Second, we identify a consistent relationship between increased tree length and signs of increased negative selection, apparent in the repertoires of both healthy subjects and those undergoing active immune responses. This suggests that B cell lineages shift towards negative selection over time as a general feature of affinity maturation. Our study provides a framework for undertaking repertoire-wide phylogenetic testing of SHM hypotheses, and provides a new means of charactering the process of mutation and selection during affinity maturation.

immunology↗

bioRxiv2019

Emergence of the Zika virus Asian lineage in Angola

Research In ContextO_ST_ABSEvidence before this studyC_ST_ABSWe searched PubMed without language restrictions using the keywords Zika and Africa for papers published to October 2018. We also checked available Situation Report publications from WHO for evidence of Zika virus (ZIKV) or congenital Zika disease in Africa. ZIKV African lineage has been detected within Africa since the mid 20th century, yet evidence for spread of the ZIKV Asian lineage within Africa is limited. Two countries in Africa (Cabo Verde and Angola) have reported ZIKV cases that are believed to be caused by a newly introduced Asian lineage virus. Sequence data are critical for confirming and understanding the spread of ZIKV Asian lineage within Africa, but these data are currently limited to a single 193bp fragment of the ZIKV NS1 gene from Angola. In addition, whilst epidemiological data on ZIKV and suspected microcephaly cases have been reported in detail from Cabo Verde, data from Angola are extremely limited.\n\nAdded value of this studyWe provide a detailed report of detected ZIKV acute cases and suspected microcephaly cases in Angola. We sequence ZIKV genomes from three acutely infected cases. These represent the first three Asian lineage genomes available from Africa, one of which was acquired from a baby with confirmed microcephaly. Analysis of these sequences suggests that ZIKV may have been introduced to Angola between July 2015 and June 2016, after which it likely circulated for at least one year. This provides the first genetic confirmation of autochthonous ZIKV Asian lineage transmission within Africa. We suggest that the virus was more likely introduced to Angola directly from Brazil, rather than from Cabo Verde. Our analyses from Angola, only the second African country to report presence of the Asian virus lineage, therefore improve our understanding of the extent and clinical impact of ZIKV Asian lineage in the continent.\n\nImplications of all the available evidenceThe circulation of ZIKV Asian lineage within parts of sub-Saharan Africa is concerning given the potential for continued viral spread across much of the continent. Available evidence suggests that ZIKV has circulated and caused cases of microcephaly in Cabo Verde and in Angola. Detecting additional ZIKV transmission using only clinical data on suspected microcephaly or clusters of mild illness may be challenging in countries where systems for reporting birth defects are limited and infectious disease burden is high. Further spread of the ZIKV Asian lineage would likely not be detected unless molecular surveillance systems for ZIKV are implemented to routinely monitor ZIKV transmission in Africa. Implementation of such a surveillance system is especially important in countries that are linked by high human mobility to areas that have experienced recent or ongoing outbreaks of ZIKV.\n\nAbstractO_ST_ABSBackgroundC_ST_ABSZika virus (ZIKV) infections and suspected microcephaly cases have been recently reported in Angola, but no data are available on the origins, epidemiology, and diversity of the virus.\n\nMethodsSerum samples from 54 suspected ZIKV cases, 76 suspected microcephaly cases, and 24 mothers of infants with suspected microcephaly were received by the Angolan Ministry of Health. Computed tomographic brain imaging and serological assays (PRNT) were conducted on one microcephalic infant. All sera were tested for ZIKV by RT-qPCR. 349 samples from HIV+ patients and 336 samples from patients suspected of chikungunya virus or dengue virus infection were also tested. Portable sequencing was used to generate Angolan ZIKV genome sequences, including from a ZIKV+ neonate with microcephaly born in Portugal to an Angolan resident. Genetic and mobility data were analysed to investigate the date of introduction and geographic origin of ZIKV in Angola.\n\nFindingsFour autochthonous cases were ZIKV positive via RT-qPCR, with all positive samples collected between December 2016 and June 2017. Viral genomes were generated for two of these cases, and from the neonate with microcephaly identified in Portugal. Genetic analyses and other data indicate that ZIKV was introduced to Angola from Brazil between July 2015 and June 2016. This introduction likely initiated local ZIKV circulation in Angola that continued until June 2017. The scanned microcephaly case showed brain abnormalities consistent with congenital Zika syndrome and serological evidence for maternal ZIKV infection.\n\nInterpretationOur analyses confirm the autochthonous transmission of the ZIKV Asian lineage in continental Africa. Conducting ZIKV surveillance throughout Africa is critical in the light of presented evidence for autochthonous ZIKV transmission in Angola, and associated microcephaly cases.\n\nFundingRoyal Society, Wellcome Trust, CNPq, CAPES, ERC, Oxford Martin School, Global Challenges Research Fund, Africa Oxford, and John Fell Fund.

epidemiology↗

bioRxiv2018

High resolution evolutionary analysis of within-host hepatitis C virus infection

Despite the breakthroughs in the treatment of HCV infection in recent years, we have a limited understanding of how virus diversity generated within individuals impacts the evolution and spread of HCV variants at the population scale. Addressing this gap will be important for building models for molecular epidemiology, which can identify main sources of disease transmission and evaluate the risks of drug-resistance mutations emerging and disseminating in a population. Here, we have undertaken a high-resolution analysis of HCV within-host evolution from four individuals co-infected with HIV. Specifically, we used long-read, deep-sequenced data of the full-length HCV envelope glycoprotein, longitudinally sampled from acute to chronic HCV infection to investigate the underlying viral evolutionary dynamics. In three individuals we found strong statistical support for population structure maintaining within-host HCV genetic diversity. Furthermore, we found significant variation in rates of molecular evolution among different regions of the HCV envelope region, both within and between individuals. Lastly, we report the first estimate of the within-host population genetic rate of recombination for HCV (0.28 x 10-7 recombinations per site per day; interquartile range: 0.13-1.05 x 10-7), which is two orders of magnitude lower than that estimated for HIV-1, and four orders of magnitude lower than the nucleotide substitution rate of the HCV envelope gene. Together, these observations indicate that population structure and strong genetic linkage shapes within-host HCV evolutionary dynamics. These results will guide the future investigation of potential HCV drug resistance adaptation during infection, and at the population scale.

evolutionary biology↗

bioRxiv2018

Parallel Evolution and the Emergence of Highly Pathogenic Avian Influenza A Viruses

Parallel molecular evolution and adaptation are important phenomena commonly observed in viruses. Here we exploit parallel molecular evolution to understand virulence evolution in avian influenza viruses (AIV). Highly-pathogenic AIVs independently evolve from low-pathogenic ancestors via acquisition of a polybasic cleavage sites (pCS). Why some AIV lineages but not others evolve in this way is unknown. We hypothesise that the parallel emergence of highly-pathogenic AIV may be facilitated by permissive or compensatory mutations occurring across the AIV genome. We combined phylogenetic, statistical and structural approaches to discover parallel mutations in AIV genomes associated with the highly-pathogenic phenotype. Parallel mutations were screened using a new statistical test of mutation-phenotype association and further evaluated in the contexts of positive selection and protein function. The mutational panel we present reveals new links between virulence evolution and other viral traits and raises the possibility of predicting aspects of AIV evolution.

evolutionary biology↗

bioRxiv2018

Robust Design for Coalescent Model Inference

--The coalescent process describes how changes in the size of a population influence the genealogical patterns of sequences sampled from that population. The estimation of population size changes from genealogies that are reconstructed from these sequence samples, is an important problem in many biological fields. Often, population size is characterised by a piecewise-constant function, with each piece serving as a population size parameter to be estimated. Estimation quality depends on both the statistical coalescent inference method employed, and on the experimental protocol, which controls variables such as the sampling of sequences through time and space, or the transformation of model parameters. While there is an extensive literature devoted to coalescent inference methodology, there is surprisingly little work on experimental design. The research that does exist is largely simulation based, precluding the development of provable or general design theorems. We examine three key design problems: temporal sampling of sequences under the skyline demographic coalescent model, spatio-temporal sampling for the structured coalescent model, and time discretisation for sequentially Markovian coalescent models. In all cases we prove that (i) working in the logarithm of the parameters to be inferred (e.g. population size), and (ii) distributing informative coalescent events uniformly among these log-parameters, is uniquely robust. Robust means that the total and maximum uncertainty of our estimates are minimised, and are also insensitive to their unknown (true) parameter values. Given its persistence among models, this formally derived two-point theorem may form the basis of an experimental design paradigm for coalescent inference.

bioinformatics↗

bioRxiv2017

Identification of an epitope of limited variability under strong immune selection in the haemagglutinin head domain of H1N1 influenza

Antigenic targets of influenza vaccination are currently seen to be polarised between (i) highly immunogenic (and protective) epitopes of high variability, and (ii) conserved epitopes of low immunogenicity. This requires vaccines directed against the variable sites to be continuously updated, with the only other alternative being seen as the artificial boosting of immunity to invariant epitopes of low natural efficacy. However, theoretical models suggest that the antigenic evolution of influenza is best explained by postulating the existence of highly immunogenic epitopes of limited variability. Here we report the identification of such an epitope of limited variability in the head domain of the H1 haemagglutinin protein. We show that the epitope mediates immunity to historical influenza strains not previously seen by a cohort of young children. Furthermore, vaccinating mice with these epitope conformations can induce immunity to all the human H1N1 influenza strains that have circulated since 1918. The identification of epitopes of limited variability offers a mechanism by which a universal influenza vaccine can be created; these vaccines would also have the potential to protect against newly emerging influenza strains.

epidemiology↗

bioRxiv2017

Phylodynamic assessment of intervention strategies for the West African Ebola virus outbreak

This preprint has been reviewed and recommended by Peer Community In Evolutionary Biology (http://dx.doi.org/10.24072/pci.evolbiol.100046). The recent Ebola virus (EBOV) outbreak in West Africa witnessed considerable efforts to obtain viral genomic data as the epidemic was unfolding. If such data can be deployed in real-time, molecular epidemiological investigations could play a role in complementing contact tracing undertaken by public health agencies. Analysing the EBOV genomes accumulated to date can also deliver insights into epidemic dynamics. Such analyses have been shown that metapopulation dynamics were critical for EBOV dispersal between rural and urban areas during the epidemic, but the implications for specific intervention scenarios remain unclear. Here, we address this issue using a collection of phylodynamic approaches. We show that long-distance dispersal events (between administrative areas >250 km apart) were not crucial for epidemic expansion and that preventing viral lineage movement to any given administrative area would, in most cases, have had little impact. However, urban areas - specifically those encompassing the three capital cities and their suburbs - were critical in attracting and further disseminating the virus: preventing viral lineage movement to all three simultaneously would have contained epidemic size by two-thirds. Using continuous phylogeographic reconstructions we estimate a distance kernel for EBOV spread and reveal considerable heterogeneity in dispersal velocity through time. We also show that announcements of border closures were followed by a significant but transient effect on international virus dispersal. By quantifying the hypothetical impact of different intervention strategies as well as the impact of barriers on dispersal frequency, our study illustrates how phylodynamic analyses can help to address specific epidemiological and outbreak control questions.

epidemiology↗

bioRxiv2017

Reverse immunodynamics: a new method to identifying targets of protective immunity.

Despite a dramatic increase in our ability to catalogue variation among pathogen genomes, we have made far fewer advances in using this information to identify targets of protective immunity. We propose a novel methodology that combines predictions from epidemiological models with phylogenetic and structural analyses to identify such targets. Epidemiological models predict that strong immune selection can cause antigenic variants to exist in non-overlapping combinations. A corollary of this theory is that targets of immunity may be identified by searching for non-overlapping associations among antigenic variants. We applied this concept to the AMA-1 protein of the malaria parasite Plasmodium falciparum and found strong signatures of immune selection among certain regions of low variability which could render them ideal vaccine candidates.

epidemiology↗

bioRxiv2017

Molecular evolution, diversity and adaptation of H7N9 influenza A viruses in China

A novel H7N9 avian influenza virus has caused five human epidemics in China since 2013. The substantial increase in prevalence and the emergence of antigenically divergent or highly pathogenic (HP) H7N9 strains during the current outbreak raises concerns about the epizootic-potential of these viruses. Here, we investigate the evolution and adaptation of H7N9 by combining publicly available data with newly generated virus sequences isolated in Guangdong between 2015-2017. Phylogenetic analyses show that currently-circulating H7N9 viruses belong to distinct lineages with differing spatial distributions. Using ancestral sequence reconstruction and structural modelling we have identified parallel amino-acid changes on multiple separate lineages. Furthermore, we infer mutations in HA primarily occur at sites involved in receptor-recognition and/or antigenicity. We also identify seven new HP strains, which likely emerged from viruses circulating in eastern Guangdong around March 2016 and is further associated with a high rate of adaptive molecular evolution.

evolutionary biology↗

bioRxiv2017

Emergence and persistence of the Chikungunya virus East-Central-South-African genotype in Northeast Brazil

We investigate an outbreak of exanthematous illness in Maceio, Alagoas, using molecular surveillance. Of 273 samples, 76% tested RT-qPCR positive for Chikungunya virus. Phylogenetic analysis reveals that the outbreak was caused by the East-Central-South-African genotype, and that this lineage has likely persisted since mid-2014 in Northeast Brazil.\n\nArticle summary lineTransmission of the Chikungunya virus East-Central-South-African genotype has been ongoing in the Northeast region of Brazil since mid-2014.

epidemiology↗

bioRxiv2017

Epidemiological and ecological determinants of Zika virus transmission in an urban setting.

Zika has emerged as a global public health concern. Although its rapid geographic expansion can be attributed to the success of its Aedes mosquito vectors, local epidemiological drivers are still poorly understood. The city of Feira de Santana played a pivotal role in the early phases of the Chikungunya and Zika epidemics in Brazil. Here, using a climate-driven transmission model, we show that low Zika observation rates and a high vectorial capacity in this region were responsible for a high attack rate during the 2015 outbreak and the subsequent decline in cases in 2016, when the epidemic was peaking in the rest of the country. Our projections indicate that the balance between the loss of herd-immunity and the frequency of viral re-importation will dictate the transmission potential of Zika in this region in the near future. Sporadic outbreaks are expected but unlikely to be detected under current surveillance systems.

epidemiology↗

bioRxiv2017

Multiplex PCR method for MinION and Illumina sequencing of Zika and other virus genomes directly from clinical samples

Genome sequencing has become a powerful tool for studying emerging infectious diseases; however, genome sequencing directly from clinical samples without isolation remains challenging for viruses such as Zika, where metagenomic sequencing methods may generate insufficient numbers of viral reads. Here we present a protocol for generating coding-sequence complete genomes comprising an online primer design tool, a novel multiplex PCR enrichment protocol, optimised library preparation methods for the portable MinION sequencer (Oxford Nanopore Technologies) and the Illumina range of instruments, and a bioinformatics pipeline for generating consensus sequences. The MinION protocol does not require an internet connection for analysis, making it suitable for field applications with limited connectivity. Our method relies on multiplex PCR for targeted enrichment of viral genomes from samples containing as few as 50 genome copies per reaction. Viral consensus sequences can be achieved starting with clinical samples in 1-2 days following a simple laboratory workflow. This method has been successfully used by several groups studying Zika virus evolution and is facilitating an understanding of the spread of the virus in the Americas.

genomics↗