Application of long read sequencing to determine expressed antigen diversity in Trypanosoma brucei infections

Jayaraman, S. et al. (2019) Application of long read sequencing to determine expressed antigen diversity in Trypanosoma brucei infections. PLoS Neglected Tropical Diseases, 13(4), e0007262. (doi: 10.1371/journal.pntd.0007262) (PMID:30943202) (PMCID:PMC6464242)

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Abstract

Antigenic variation is employed by many pathogens to evade the host immune response, and Trypanosoma brucei has evolved a complex system to achieve this phenotype, involving sequential use of variant surface glycoprotein (VSG) genes encoded from a large repertoire of ~2,000 genes. T. brucei express multiple, sometimes closely related, VSGs in a population at any one time, and the ability to resolve and analyse this diversity has been limited. We applied long read sequencing (PacBio) to VSG amplicons generated from blood extracted from batches of mice sacrificed at time points (days 3, 6, 10 and 12) post-infection with T. brucei TREU927. The data showed that long read sequencing is reliable for resolving variant differences between VSGs, and demonstrated that there is significant expressed diversity (449 VSGs detected across 20 mice) and across the timeframe of study there was a clear semi-reproducible pattern of expressed diversity (median of 27 VSGs per sample at day 3 post infection (p.i.), 82 VSGs at day 6 p.i., 187 VSGs at day 10 p.i. and 132 VSGs by day 12 p.i.). There was also consistent detection of one VSG dominating expression across replicates at days 3 and 6, and emergence of a second dominant VSG across replicates by day 12. The innovative application of ecological diversity analysis to VSG reads enabled characterisation of hierarchical VSG expression in the dataset, and resulted in a novel method for analysing such patterns of variation. Additionally, the long read approach allowed detection of mosaic VSG expression from very few reads–the earliest in infection that such events have been detected. Therefore, our results indicate that long read analysis is a reliable tool for resolving diverse gene expression profiles, and provides novel insights into the complexity and nature of VSG expression in trypanosomes, revealing significantly higher diversity than previously shown and the ability to identify mosaic gene formation early during the infection process.

Item Type:Articles
Status:Published
Refereed:Yes
Glasgow Author(s) Enlighten ID:Cobbold, Dr Christina and Vaikkinen, Ms Heli and Reeve, Dr Richard and Harris, Claire and Donachie, Ms Anne Marie and McCulloch, Professor Richard
Creator Roles:
Harris, C.Formal analysis, Methodology, Writing – original draft
Donachie, A.-M.Investigation, Methodology
Vaikkinen, H.Investigation, Methodology
McCulloch, R.Conceptualization, Writing – original draft
Cobbold, C.Conceptualization, Formal analysis, Investigation, Methodology, Supervision, Writing – original draft, Writing – review and editing
Reeve, R.Formal analysis, Investigation, Methodology, Supervision, Writing – original draft, Writing – review and editing
Authors: Jayaraman, S., Harris, C., Paxton, E., Donachie, A.-M., Vaikkinen, H., McCulloch, R., Hall, J. P.J., Kenny, J., Lenzi, L., Herts-Fowler, C., Cobbold, C., Reeve, R., Micheol, T., and Morrison, L. J.
College/School:College of Medical Veterinary and Life Sciences
College of Medical Veterinary and Life Sciences > Institute of Biodiversity Animal Health and Comparative Medicine
College of Medical Veterinary and Life Sciences > Institute of Infection Immunity and Inflammation
College of Science and Engineering > School of Mathematics and Statistics
Journal Name:PLoS Neglected Tropical Diseases
Publisher:Public Library of Science
ISSN:1935-2727
ISSN (Online):1935-2735
Published Online:03 April 2019
Copyright Holders:Copyright © 2019 Jayaraman et al.
First Published:First published in PLoS Neglected Tropical Diseases 13(4):e0007262
Publisher Policy:Reproduced under a Creative Commons license

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Project CodeAward NoProject NamePrincipal InvestigatorFunder's NameFunder RefLead Dept
529045The host/parasite interface: trypanosomes as a paradigmLiam MorrisonThe Royal Society (ROYSOC)UF090083III - BACTERIOLOGY
593881The in vivo transcriptome of Trypanosoma brucei as a tool to examine the host-parasite interactionLiam MorrisonThe Royal Society (ROYSOC)RG110378III - BACTERIOLOGY
730011Mathematical Theory and Biological Applications of DiversityRichard ReeveBiotechnology and Biological Sciences Research Council (BBSRC)BB/P004202/1RI BIODIVERSITY ANIMAL HEALTH & COMPMED