Biology subjects

Hamilton, W. L.

Publications and source records attributed to Hamilton, W. L..

9 recordsLinked to original sources

bioRxiv2025

De novo assembly of complete Plasmodium falciparum isolate genomes using PacBio HiFi sequencing technology

Plasmodium falciparum possesses a highly structured genome with extensive sequence diversity concentrated in Variant Surface Antigen (VSA) families. These genes--var, rif, and stevor--play key roles in immune evasion and pathogenesis and are difficult to assemble using short-read sequencing technologies. Here, we applied PacBio HiFi long-read sequencing to generate high-quality de novo genome assemblies from 43 P. falciparum parasite cultures originating from community cases in The Gambia. Parasites were culture-adapted, cloned by limiting dilution where possible, and sequenced using high molecular weight DNA extracts. Assemblies from single-genotype lineages were constructed using hifiasm, producing complete chromosomal-length scaffolds with high base accuracy without requiring short-read polishing. We recovered full repertoires of var, rif, and stevor genes and classified them into known subgroups. Together, our results demonstrate that PacBio HiFi sequencing enables accurate assembly of complex P. falciparum genomes from natural infections. This work provides a valuable genomic resource for future studies of parasite evolution, transmission dynamics, and antigenic diversity, and suggests that VSA repertoires can serve as reliable proxies of genetic relatedness across infections.

genomics↗

bioRxiv2024

Variable rates of SARS-CoV-2 evolution in chronic infections

An important feature of the evolution of the SARS-CoV-2 virus has been the emergence of highly mutated novel variants, which are characterised by the gain of multiple mutations relative to viruses circulating in the general global population. Cases of chronic viral infection have been suggested as an explanation for this phenomenon, whereby an extended period of infection, with an increased rate of evolution, creates viruses with substantial genetic novelty. However, measuring a rate of evolution during chronic infection is made more difficult by the potential existence of compartmentalisation in the viral population, whereby the viruses in a host form distinct subpopulations. We here describe and apply a novel statistical method to study within-host virus evolution, identifying the minimum number of subpopulations required to explain sequence data observed from cases of chronic infection, and inferring rates for within-host viral evolution. Across nine cases of chronic SARS-CoV-2 infection in hospitalised patients we find that non-trivial population structure is relatively common, with four cases showing evidence of more than one viral population evolving independently within the host. We find cases of within-host evolution proceeding significantly faster, and significantly slower, than that of the global SARS-CoV-2 population, and of cases in which viral subpopulations in the same host have statistically distinguishable rates of evolution. Non-trivial population structure was associated with high rates of within-host evolution that were systematically underestimated by a more standard inference method.

evolutionary biology↗

bioRxiv2024

A complex Plasmodium falciparum cryptotype circulating at low frequency across the African continent

The population structure of the malaria parasite Plasmodium falciparum can reveal underlying demographic and adaptive evolutionary processes. Here, we analyse population structure in 4,376 P. falciparum genomes from 21 countries across Africa. We identified a strongly differentiated cluster of parasites, comprising [~]1.2% of samples analysed, geographically distributed over 13 countries across the continent. Members of this cluster, named AF1, carry a genetic background consisting of a large number of highly differentiated variants, rarely observed outside this cluster, at a multitude of genomic loci distributed across most chromosomes. At these loci, the AF1 haplotypes appear to have common ancestry, irrespective of the sampling location; outside the shared loci, however, AF1 members are genetically similar to their sympatric parasites. AF1 parasites sharing up to 23 genomic co-inherited regions were found in all major regions of Africa, at locations over 7,000 km apart. We coined the term cryptotype to describe a complex common background which is geographically widespread, but concealed by genomic regions of local origin. Most AF1 differentiated variants are functionally related, comprising structural variations and single nucleotide polymorphisms in components of the MSP1 complex and several other genes involved in interactions with red blood cells, including invasion and erythrocyte antigen export. We propose that AF1 parasites have adapted to some as yet unidentified evolutionary niche, by acquiring a complex compendium of interacting variants that rarely circulate separately in Africa. As the cryptotype spread across the continent, it appears to have been maintained mostly intact in spite of recombination events, suggesting a selective advantage. It is possible that other cryptotypes circulate in Africa, and new analysis methods may be needed to identify them.

genomics↗

bioRxiv2023

A fourth locus in the Plasmodium falciparum genome associated with sickle haemoglobin

BackgroundHeterozygosity for sickle haemoglobin (HbS) confers protection against severe malaria caused by the parasite Plasmodium falciparum. Recent work has suggested that this protective effect can depend on the parasite genotype: P. falciparum sickle-associated (Pfsa) variants were found disproportionately in individuals with severe malaria carrying HbS alleles in The Gambia and Kenya. Interactions between the P. falciparum genome and HbS have not previously been investigated in mild malaria cases or in Ghana. MethodsWe performed a genome-wide association analysis of P. falciparum against human {beta}-globin genotypes in a sample of 1,368 people with mild malaria in northern Ghana. ResultsWe replicated the previously identified associations with HbS at two parasite loci (Pfsa1 and Pfsa3). Pfsa2 was absent from this population. A candidate newly identified locus within the serine/ threonine kinase FIKK4.2, which we putatively term Pfsa4, was also associated with HbS; this finding replicated in a published sample from Mali. The Pfsa1-4 mutations vary widely in frequencies across Africa, are absent or very low frequency in Asia, and are highly correlated with each-other across multiple populations. We found no strong associations with haemoglobin C. ConclusionsThis study replicates previously reported sickle-associated loci in the P. falciparum genome and has produced new evidence of a potential association with sickle haemoglobin at a fourth parasite locus. Further research is needed to validate the tentative fourth locus. These findings add new complexity to the emerging picture of association between human and co-evolving malaria parasite genomes, suggesting new avenues for functional exploration.

microbiology↗

bioRxiv2022

Nanopore sequencing for real-time genomic surveillance of Plasmodium falciparum

Malaria is a global public health priority causing over 600,000 deaths annually, mostly young children living in Sub-Saharan Africa. Molecular surveillance can provide key information for malaria control, such as the prevalence and distribution of antimalarial drug resistance. However, genome sequencing capacity in endemic countries can be limited. Here, we have implemented an end-to-end workflow for P. falciparum genomic surveillance in Ghana using Oxford Nanopore Technologies, targeting antimalarial resistance markers and the leading vaccine antigen circumsporozoite protein (csp). The workflow was rapid, robust, accurate, affordable and straightforward to implement, and could be deployed using readily collected dried blood spot samples. We found that P. falciparum parasites in Ghana had become largely susceptible to chloroquine, with persistent sulfadoxine-pyrimethamine (SP) resistance, and no evidence of artemisinin resistance. Multiple Single Nucleotide Polymorphism (SNP) differences from the vaccine csp sequence were identified, though their significance is uncertain. This study demonstrates the potential utility and feasibility of malaria genomic surveillance in endemic settings using Nanopore sequencing.

genomics↗

bioRxiv2022

Integrated pretraining with evolutionary information to improve RNA secondary structure prediction

AO_SCPLOWBSTRACTC_SCPLOWRNA secondary structure prediction is a fundamental task in computational and molecular biology. While machine learning approaches in this area have been shown to improve upon traditional RNA folding algorithms, performance remains limited for several reasons such as the small number of experimentally determined RNA structures and suboptimal use of evolutionary information. To address these challenges, we introduce a practical and effective pretraining strategy that enables learning from a larger set of RNA sequences with computationally predicted structures and in the meantime, tapping into the rich evolutionary information available in databases such as Rfam. Coupled with a flexible and scalable neural architecture that can navigate different learning scenarios while providing ease of integrating evolutionary information, our approach significantly improves upon state-of-the-art across a range of benchmarks, including both single sequence and alignment based structure prediction tasks, with particularly notable benefits on new, less well-studied RNA families. Our source code, data and packaged RNA secondary structure prediction software RSSMFold can be accessed at https://github.com/HarveyYan/RSSMFold.

molecular biology↗

bioRxiv2020

Patterns of within-host genetic diversity in SARS-CoV-2

Monitoring the spread of SARS-CoV-2 and reconstructing transmission chains has become a major public health focus for many governments around the world. The modest mutation rate and rapid transmission of SARS-CoV-2 prevents the reconstruction of transmission chains from consensus genome sequences, but within-host genetic diversity could theoretically help identify close contacts. Here we describe the patterns of within-host diversity in 1,181 SARS-CoV-2 samples sequenced to high depth in duplicate. 95% of samples show within-host mutations at detectable allele frequencies. Analyses of the mutational spectra revealed strong strand asymmetries suggestive of damage or RNA editing of the plus strand, rather than replication errors, dominating the accumulation of mutations during the SARS-CoV-2 pandemic. Within and between host diversity show strong purifying selection, particularly against nonsense mutations. Recurrent within-host mutations, many of which coincide with known phylogenetic homoplasies, display a spectrum and patterns of purifying selection more suggestive of mutational hotspots than recombination or convergent evolution. While allele frequencies suggest that most samples result from infection by a single lineage, we identify multiple putative examples of co-infection. Integrating these results into an epidemiological inference framework, we find that while sharing of within-host variants between samples could help the reconstruction of transmission chains, mutational hotspots and rare cases of superinfection can confound these analyses.

genomics↗

bioRxiv2020

Graph neural representational learning of RNA secondary structures for predicting RNA-protein interactions

AO_SCPLOWBSTRACTC_SCPLOWO_ST_ABSMotivationC_ST_ABSRNA-protein interactions are key effectors of post-transcriptional regulation. Significant experimental and bioinformatics efforts have been expended on characterizing protein binding mechanisms on the molecular level, and on highlighting the sequence and structural traits of RNA that impact the binding specificity for different proteins. Yet our ability to predict these interactions in silico remains relatively poor. ResultsIn this study, we introduce RPI-Net, a graph neural network approach for RNA-protein interaction prediction. RPI-Net learns and exploits a graph representation of RNA molecules, yielding significant performance gains over existing state-of-the-art approaches. We also introduce an approach to rectify particular type of sequence bias present in many CLIP-Seq data sets, and we show that correcting this bias is essential in order to learn meaningful predictors and properly evaluate their accuracy. Finally, we provide new approaches to interpret the trained models and extract simple, biologically-interpretable representations of the learned sequence and structural motifs. AvailabilitySource code can be accessed at https://www.github.com/HarveyYan/RNAonGraph. Contactwlh@cs.mcgill.ca, blanchem@cs.mcgill.ca

molecular biology↗

bioRxiv2019

Evolution and expansion of multidrug resistant malaria in Southeast Asia: a genomic epidemiology study

BackgroundA multidrug resistant co-lineage of Plasmodium falciparum malaria, named KEL1/PLA1, spread across Cambodia c.2008-2013, causing high treatment failure rates to the frontline combination therapy dihydroartemisinin-piperaquine. Here, we report on the evolution and spread of KEL1/PLA1 in subsequent years.\n\nMethodsWe analysed whole genome sequencing data from 1,673 P. falciparum clinical samples collected in 2008-2018 from northeast Thailand, Laos, Cambodia and Vietnam. By investigating genome-wide relatedness between parasites, we inferred patterns of shared ancestry in the KEL1/PLA1 population.\n\nFindingsKEL1/PLA1 spread rapidly from 2015 into all of the surveyed countries and now exceeds 80% of the P. falciparum population in several regions. These parasites maintained a high level of genetic relatedness reflecting their common origin. However, several genetic subgroups have recently emerged within this co-lineage with diverse geographical distributions. Some of these emerging KEL1/PLA1 subgroups carry recent mutations in the chloroquine resistance transporter (crt) gene, which arise on a specific genetic background comprising multiple genomic regions.\n\nInterpretationAfter emerging and circulating for several years within Cambodia, the P. falciparum KEL1/PLA1 co-lineage diversified into multiple subgroups and acquired new genetic features including novel crt mutations. These subgroups have rapidly spread into neighbouring countries, suggesting enhanced fitness. These findings highlight the urgent need for elimination of this increasingly drug-resistant parasite co-lineage, and the importance of genetic surveillance in accelerating elimination efforts.\n\nFundingWellcome Trust, Bill & Melinda Gates Foundation, UK Medical Research Council, UK Department for International Development.\n\nResearch in context O_TEXTBOXEvidence before this studyThis study updates our previous work describing the emergence and spread of a multidrug resistant P. falciparum co-lineage (KEL1/PLA1) within Cambodia up to 2013. Since then, a regional genetic surveillance project, GenRe-Mekong, has reported that markers of dihydroartemisinin-piperaquine (DHA-PPQ) resistance have increased in frequency in neighbouring countries. A PubMed search (terms: \"artemisinin\", \"piperaquine\", \"resistance\", \"southeast asia\") for articles listed since our previous study (from 30/10/2017 to 05/01/2019) yielded 28 results, including reports of a recent sharp decline in DHA-PPQ clinical efficacy in Vietnam; the spread of genetic markers of DHA-PPQ resistance into neighbouring countries by Imwong and colleagues; and multiple reports associating mutations in the crt gene with piperaquine resistance, including newly emerging crt variants in Southeast Asia.\n\nAdded value of this studyWe analysed P. falciparum whole genomes collected up to early 2018 from Eastern Southeast Asia (Cambodia and surrounding regions), describing the fine-scale epidemiology of multiple KEL1/PLA1 genetic subgroups that have spread out from Cambodia since 2015 and taken over indigenous parasite populations in northeastern Thailand, southern and central Vietnam and parts of southern Laos. Several newly emerging crt mutations accompanied the spread and expansion of KEL1/PLA1 subgroups, suggesting an active proliferation of biologically fit, multidrug resistant parasites.\n\nImplications of all the available evidenceThe problem of P. falciparum multidrug resistance has dramatically worsened in Eastern Southeast Asia since previous reports. KEL1/PLA1 has diversified and spread widely across Eastern Southeast Asia since 2015, becoming the predominant parasite group in several regions. This may have been fuelled by continued parasite exposure to DHA-PPQ, resulting in sustained selection after KEL1/PLA1 became established. Continued drug pressure enabled the acquisition of further mutations, resulting in higher levels of resistance. These data demonstrate the value of pathogen genetic surveillance and the urgent need to eliminate these dangerous parasites.\n\nC_TEXTBOX

epidemiology↗