Using whole genome sequencing to reveal malaria parasite genetic diversity and drug resistance in Vietnam
Lead Research Organisation:
London School of Hygiene & Tropical Medicine
Department Name: Infectious and Tropical Diseases
Abstract
Malaria, a mosquito-borne disease caused by Plasmodium parasites, is an important public health problem causing an estimated 212 million cases and 429,000 deaths annually. In the WHO Western Pacific Region, 10 out of 37 countries, including Viet Nam, are endemic for malaria. In Viet Nam, there is heterogeneity in malaria prevalence, with the Southern and Central regions with the highest morbidity (7% of population with >1 case per 1000 people). After successfully reducing the malaria burden to pre-elimination levels over the past two decades, the National Malaria Control Programme (NMCP) has recently switched from control to elimination by 2030. Whilst the number of confirmed cases has more than halved between 2012 and 2016 to ~4,200 (P. falciparum 56%, P. vivax 42%), there is now evidence that P. falciparum malaria parasites resistant to frontline treatments have spread to five Vietnamese provinces. Very little data is available for P. vivax drug resistance. The imminent spread of drug-resistant parasites can undermine progress made towards elimination.
Drug resistance is caused by mutations in the parasite genomes. Therefore to provide biological insights, we propose to characterise the genetic profiles of ~1,000 Pf and Pv samples sourced from endemic provinces in Viet Nam using cutting edge whole genome sequencing (WGS) technologies. We will investigate genetic differences between samples, within province, across time, and detect known and potentially novel drug resistance markers. Our findings will provide data to the NMCP on drug resistant parasites to assist policy development, and inform the development of diagnostics, vaccines and treatments for malaria. The work will lead to new insights into the genomes of Vietnamese malaria parasites, as well as establish a publically available genetic data resource to facilitate future malaria research. Further, the application of WGS technologies in Viet Nam will build capacity for future genomic investigations.
Drug resistance is caused by mutations in the parasite genomes. Therefore to provide biological insights, we propose to characterise the genetic profiles of ~1,000 Pf and Pv samples sourced from endemic provinces in Viet Nam using cutting edge whole genome sequencing (WGS) technologies. We will investigate genetic differences between samples, within province, across time, and detect known and potentially novel drug resistance markers. Our findings will provide data to the NMCP on drug resistant parasites to assist policy development, and inform the development of diagnostics, vaccines and treatments for malaria. The work will lead to new insights into the genomes of Vietnamese malaria parasites, as well as establish a publically available genetic data resource to facilitate future malaria research. Further, the application of WGS technologies in Viet Nam will build capacity for future genomic investigations.
Technical Summary
Malaria, caused by Plasmodium parasites, remains a barrier to social and economic development in resource poor areas of Viet Nam. Between 2012 and 2016, Viet Nam achieved a 52% reduction in malaria cases to ~4,200 (P. falciparum 56%, P. vivax 42%). The campaign against malaria by the Vietnamese Government is now being refocused from control to elimination by year 2030. However, this campaign is challenged by the threat of parasites, which are resistant to existing frontline artemisinin combination treatments and spreading in the Greater Mekong Sub-region (GMS); five Vietnamese provinces have been identified to harbour these drug-resistant parasites. Compared to P. falciparum, very little is known about drug resistance in Pv, but is driven by genetic mutations in the parasite genomes. New molecular tools to characterise circulating parasites are warranted. Here we propose to characterise the genetic diversity and the molecular basis of drug resistance of malaria parasites in Viet Nam using whole genome sequencing (WGS). By examining the genomic variation from both Pf and Pv sourced from 5 malaria endemic provinces, we will be able to detect crucial mutations in drug resistance genes (e.g. PfKelch, PfPlasmepsin2, Pfmdr1 and related genes in Pv), and potentially identify new loci by using population genetic approaches to uncover "selective sweeps". The genomic variation will allow an assessment of differences in population structure across time, between the geographical areas, and within the GMS context and beyond. We will also perform comparative Pf and Pv species analysis to highlight any differential effects of control practice.
As part of this application, it is also proposed to build capacity in Viet Nam in the area of malaria genomics and its applications through on-site training and workshops, as well as exchange of researchers between the UK and Viet Nam. These activities will assist Vietnamese scientists to undertake genomics research studies in the future.
As part of this application, it is also proposed to build capacity in Viet Nam in the area of malaria genomics and its applications through on-site training and workshops, as well as exchange of researchers between the UK and Viet Nam. These activities will assist Vietnamese scientists to undertake genomics research studies in the future.
Planned Impact
The economy
Advances in sequencing technology now allow the genomic characterisation of malaria parasites on an unprecedented scale, and have the potential to greatly accelerate research aimed at understanding the biology of the parasite, its geographical spread, the underlying causes of its drug resistance and transmission, and the epidemiology of the disease. Genomic technologies have the ability to generate vast amounts of data, and there is a need to translate this information into knowledge useable by other research scientists and industry. The proposed work will provide tools useful for genomic data analysis and modelling. The knowledge generated in the project and application of the research could benefit the scientific community, including control programs looking at the spread of disease and drug resistance. Those in the pharmaceutical industry, particularly those developing malaria diagnostics, treatments and vaccines will also benefit from the research. The methods used in this project could have application beyond malaria, to support more widely in the control and prevention of infectious diseases with associated economic benefits.
The results will provide evidence to guide future research in academia and industry and policy decisions in government. It is envisaged that the findings can be used to inform malaria control activities and mitigate the spread of drug resistance strains, enabling Viet Nam to reach (and maintain) elimination status in 2030. Ultimately, the knowledge gained in this study could improve the health and wealth of the country, leading to a working population with lower burden of malarial disease. The NIMPE will facilitate the reach and potential uptake of research findings in Viet Nam through the National Malaria Control Programme (NMCP) and the MOH, as well as links at the national, regional and international levels.
Academic and industrial organisations
New high throughput genomic technologies have the ability to generate vast amounts of data, but there is a need to translate this information into knowledge useable by other research scientists and industry, especially in countries where malaria is endemic. Our work will provide tools useful for genomic data analysis and visualisation, which can be utilized across infectious diseases and in different settings. An understanding of genomic variation could lead to follow-up laboratory experiments, improved diagnostic tests for detecting pan-parasite species, and reveal new parasite gene targets for anti-malarial treatments and vaccines. Scientific developments arising would enhance the commercial private sector for the production of diagnostics, vaccines and other control measures, as well as academics involved in policy formulation. The LSHTM technology transfer office has processes to ensure pipelines to vaccine or other translation tool production and exploitation. Any technologies developed may have enormous implications for policy makers for future disease outbreaks and elimination of infectious diseases.
Training opportunities
The proposal will employ, train and develop a scientist with diverse experience and an 'omic mentality that can be applied in academia, the public sector and industry. The researchers working on the project will develop team working and project management skills, which they can apply in all employment sectors. The researchers employed will have unique opportunities for engagement with experts (e.g. in the LSHTM - Malaria Centre, The Applied Genomics Centre) in malaria biology, epidemiology, clinical care, genomic and public health. There will also be opportunities to train researchers and public health stakeholders in genomic data analysis, including the next generation of Vietnamese researchers. Thus, our proposal will impact on the creation of human resources that could subsequently be employed in challenging interdisciplinary projects in industry, academia and government, across countries.
Advances in sequencing technology now allow the genomic characterisation of malaria parasites on an unprecedented scale, and have the potential to greatly accelerate research aimed at understanding the biology of the parasite, its geographical spread, the underlying causes of its drug resistance and transmission, and the epidemiology of the disease. Genomic technologies have the ability to generate vast amounts of data, and there is a need to translate this information into knowledge useable by other research scientists and industry. The proposed work will provide tools useful for genomic data analysis and modelling. The knowledge generated in the project and application of the research could benefit the scientific community, including control programs looking at the spread of disease and drug resistance. Those in the pharmaceutical industry, particularly those developing malaria diagnostics, treatments and vaccines will also benefit from the research. The methods used in this project could have application beyond malaria, to support more widely in the control and prevention of infectious diseases with associated economic benefits.
The results will provide evidence to guide future research in academia and industry and policy decisions in government. It is envisaged that the findings can be used to inform malaria control activities and mitigate the spread of drug resistance strains, enabling Viet Nam to reach (and maintain) elimination status in 2030. Ultimately, the knowledge gained in this study could improve the health and wealth of the country, leading to a working population with lower burden of malarial disease. The NIMPE will facilitate the reach and potential uptake of research findings in Viet Nam through the National Malaria Control Programme (NMCP) and the MOH, as well as links at the national, regional and international levels.
Academic and industrial organisations
New high throughput genomic technologies have the ability to generate vast amounts of data, but there is a need to translate this information into knowledge useable by other research scientists and industry, especially in countries where malaria is endemic. Our work will provide tools useful for genomic data analysis and visualisation, which can be utilized across infectious diseases and in different settings. An understanding of genomic variation could lead to follow-up laboratory experiments, improved diagnostic tests for detecting pan-parasite species, and reveal new parasite gene targets for anti-malarial treatments and vaccines. Scientific developments arising would enhance the commercial private sector for the production of diagnostics, vaccines and other control measures, as well as academics involved in policy formulation. The LSHTM technology transfer office has processes to ensure pipelines to vaccine or other translation tool production and exploitation. Any technologies developed may have enormous implications for policy makers for future disease outbreaks and elimination of infectious diseases.
Training opportunities
The proposal will employ, train and develop a scientist with diverse experience and an 'omic mentality that can be applied in academia, the public sector and industry. The researchers working on the project will develop team working and project management skills, which they can apply in all employment sectors. The researchers employed will have unique opportunities for engagement with experts (e.g. in the LSHTM - Malaria Centre, The Applied Genomics Centre) in malaria biology, epidemiology, clinical care, genomic and public health. There will also be opportunities to train researchers and public health stakeholders in genomic data analysis, including the next generation of Vietnamese researchers. Thus, our proposal will impact on the creation of human resources that could subsequently be employed in challenging interdisciplinary projects in industry, academia and government, across countries.
Publications
Phelan JE
(2023)
Rapid profiling of Plasmodium parasites from genome sequences to assist malaria control.
in Genome medicine
Description | We have now shared clinical sample processing and sequencing protocols with our collaborators, so that they can perform whole genome sequencing onsite. Analysis pipelines have been shared too, further building capacity for bioinformatics and genome investigations. Plasmodium DNA has been sequenced and is allowing us to assess the population structure, drug resistance mutations, and transmission patterns of Vietnamese parasites, as well as compare the genome diversity to a wider Asian regional setting. |
Exploitation Route | Insights into genomic diversity will facilitate the development of much needed molecular diagnostics. Further, the ability to perform genomic investigations will assist with outbreak investigations, including those involving other pathogens. |
Sectors | Environment Healthcare Government Democracy and Justice Pharmaceuticals and Medical Biotechnology |
Description | The use of whole genome sequencing as a diagnostic, surveillance and outbreak investigation tool has gained traction with policy makers, and is part of the governments infection control strategy. We are working closely with clinicians and infection control policy makers, and our data is providing supporting evidence for the utility of whole genome sequencing, and the related successful implementation of associated technologies and analytical pipelines. |
First Year Of Impact | 2021 |
Sector | Healthcare,Government, Democracy and Justice,Pharmaceuticals and Medical Biotechnology |
Impact Types | Cultural Societal Economic Policy & public services |
Title | DNA processing and genome analysis pipelines |
Description | Within the NIMPE laboratories, we have established selective whole genome amplification protocols for the processing of DNA . Further, bioinformatic pipelines for the analysis of genomic sequence data have been installed and implemented in Vietnam too. |
Type Of Material | Improvements to research infrastructure |
Year Produced | 2020 |
Provided To Others? | No |
Impact | This will give NIMPE the ability to conduct genomic investigations, from sample collection, DNA processing and sequencing, to analysis. |
Title | Malaria-Profiler |
Description | This informatics tool profiles malaria parasite sequence data to infer species, drug resistance and geographical source. It has been implemented on Vietnamese data, and the software is being used in Vietnam. |
Type Of Material | Improvements to research infrastructure |
Year Produced | 2023 |
Provided To Others? | Yes |
Impact | It strengthens capacity for the analysis of sequence data, and is useful for clinical and surveillance applications. |
URL | https://bioinformatics.lshtm.ac.uk/malaria-profiler/ |
Title | Protocols for DNA processing and sequencing |
Description | We have developed and shared tools and protocols for the selective whole genome amplification of DNA, so that low parasitaemia clinical samples may be sequenced. We have also shared protocols for the sequencing of DNA on Oxford MinION and Illumina platforms, where the latter technology is now available in-house at our collaborators. |
Type Of Material | Improvements to research infrastructure |
Year Produced | 2021 |
Provided To Others? | Yes |
Impact | Our collaborator can now whole genome sequence plasmodium parasite DNA processed from clinical samples. |
Title | Database of malaria genetic mutations |
Description | This consists of a set of one million genetic mutations across P. falciparum, P. vivax, P. malariae and P. knowlesi parasite genomes, including drug resistance and geographical associated barcoding markers. This database will provide a look up table to compare to generated Vietnamese sequences, as well as to assist with population structure analysis. |
Type Of Material | Database/Collection of data |
Year Produced | 2020 |
Provided To Others? | No |
Impact | It will assist with population genomic analysis and positioning of the Vietnamese malaria samples in a global context. |
Title | Plasmodium sequencing data |
Description | We have whole genome sequenced Plasmodium species DNA, and curated genomic variation, which will be used for downstream analysis. |
Type Of Material | Database/Collection of data |
Year Produced | 2021 |
Provided To Others? | No |
Impact | The genomic variation will provide insights into the diversity of circulating parasites, drug resistance and transmission. It can also be used to inform on populations structure and the design of diagnostics. |
Title | Whole genome sequencing dataset of Plasmodium parasites and Mosquito vectors |
Description | Plasmodium falciparum and vivax malaria sequencing data (n>600) generated as part of the project, has been combined with publicly available data (n>10,000) within a database for look-up and profiling analysis as part of ongoing sequencing. A similar database is being constructed for Anopheles sequence that has been generated too. |
Type Of Material | Database/Collection of data |
Year Produced | 2022 |
Provided To Others? | No |
Impact | The database is being used to develop new diagnostics and track drug resistance. We have developed a MalariaProfiler software tool that links with the database and profiles whole genome sequenced isolates for drug resistance and likely geographical source, thereby strengthening informatics capacity. |
Description | Malaria genomics with NIMPE |
Organisation | National Institute of Malariology, Parasitology and Entomology |
Country | Viet Nam |
Sector | Public |
PI Contribution | Building capacity in genomic sequencing and data analysis |
Collaborator Contribution | Malaria samples and insights in disease control. |
Impact | DNA samples for sequencing. Bioinformatic and statistical pipelines for the analysis of genomic and epidemiological data. |
Start Year | 2020 |
Description | Plasmodium genomics workshop (October 2022) |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | No |
Geographic Reach | National |
Primary Audience | Professional Practitioners |
Results and Impact | NIMPE hosted an 'omics data analysis workshop in October 2022, which was attended by ~35 malaria researchers and field team members. |
Year(s) Of Engagement Activity | 2022 |
Description | Training on malaria genomics |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | No |
Geographic Reach | Local |
Primary Audience | Professional Practitioners |
Results and Impact | Members of NIMPE came to the LSHTM in June/July 2022 for training in plasmodium DNA amplification and sequencing. |
Year(s) Of Engagement Activity | 2022 |