Chapter Three - Recent advances in functional research in Giardia intestinalis
Introduction
Undertaking comprehensive functional studies has remained a persistent obstacle in parasitological research, outside of a small number of apicomplexans, such as species of Plasmodium and Toxoplasma gondii (Limenitakis and Soldati-Favre, 2011; Meissner et al., 2007). Primary obstacles to genetically tractable parasites include an inability to readily culture them outside of a host, a lack of knowledge of the genetic and regulatory systems of parasites or the unavailability of tools to apply to them. Giardia intestinalis has been in vitro culturable for several decades in complex media, principally Keister's modified TYI-S-33 (Davids and Gillin, 2011) and its regulatory genetics and gene composition have been explored through the publication of reference genomes (Franzen et al., 2009; Jerlstrom-Hultqvist et al., 2010; Morrison et al., 2007) and other “omics” driven research over the past decade (e.g., Ansell et al., 2015a, Ansell et al., 2017; Emery et al., 2018; Franzén et al., 2013; Ma'ayeh et al., 2018; Spycher et al., 2013), albeit with much still to be done. The complex binucleate and multiploidal (2N, 4N and 8N at various stages of the life cycle; Bernander et al., 2001) cellular biology of Giardia has proven a persistent obstacle to functional research, and for much of its post-genomic period efforts to develop functional tools for Giardia have met with limited success (Luján and Svärd, 2011). Because of its importance as a parasite, its scalable culturability in cell-free media, and its deep-branching position within the eukaryotic tree of life (Morrison et al., 2007), Giardia has long been proposed as an intriguing and potentially impactful model organism (Luján and Svärd, 2011), but its recalcitrance to genetic manipulation has proven a persistent road-block to realising this potential.
Here, we review advances in functional research of Giardia, drawing on recent publications in the field, as well as novel advances in other fields that may be applicable for Giardia. Our hope is that this review will act not only as a summary of the progress in research in this field over the last few years, but also as stimulus for renewed thinking on functional research in Giardia and on the potential to exploit these technologies for novel anti-giardial therapies, but to harness this fascinating protist as a model for eukaryotic biology.
Section snippets
Advances in in vitro cultivation
The inability to culture parasites in a laboratory setting, either in vitro or in vivo, presents the primary obstacle to advancing functional research. Fortunately, this obstacle has been overcome for Giardia for several decades. In vitro cultivation of G. intestinalis is readily undertaken in TYI-S-33 media (Davids and Gillin, 2011), with or without nitrogen-sparging for culture-adapted strains isolates. Giardia intestinalis trophozoites (at least of the assemblage A) can be triggered into in
Major knowledge gaps in functional biology in Giardia
Draft genomes have been available for Giardia intestinalis assemblage A (WB: Morrison et al., 2007), B (GS: Franzen et al., 2009) and E (P15: Jerlstrom-Hultqvist et al., 2010) for a number of years. These assemblies have provided the basis for predicting Giardia's ~ 5500 coding gene models. Recent efforts (see current release in GiardiaDB) to curate these models have significantly improved their functional annotation; yet, ~ 60% of these are defined as “conserved” or “hypothetical” proteins.
Computational and “omic” approaches to functional biology research in Giardia
It is clear that the next major breakthroughs in our understanding of the molecular biology of Giardia must make full use of advances in functional research. Conceptually, these advances can be categorized into three major and complimentary themes: (i) improved in silico inference, (ii) direct empirical assessment of function through “omics”, labelling and other approaches and (iii) direct empirical assessment of function through targeted genetic manipulation.
Direct, targeted-based assessment of gene function
As most parasites are unculturable and genetically intractable, much of the functional biology is understood through in-direct inference by in silico modelling and, more recently, “omics” and systems-based research. These are powerful tools, but are most valuable when they are used to guide empirical interrogation of gene function through direct, target-based studies. For many years, Giardia was proposed as a model parasite because of its ready in vitro cultivation, but significant challenges
Concluding remarks
Giardia has proven a tantalizingly close but frustratingly distant model eukaryote and genetically tractable parasitic species. Its ready culturability and early discoveries in its regulatory biology, as well as its status as an early diverging branch of the eukaryotic tree and cause of significant disease in humans and animals, has piqued interest in the species for many years. However, its binucleate cell, tetraploid genome and limited acceptance of seemingly attractive genetic tools, such as
Acknowledgements
A.J. acknowledges funding support from the Australian National Health and Medical Research Council (Career Development Fellowship APP1126395), Melbourne Water and the Victorian State Government Operational Infrastructure Support and Australian Government National Health and Medical Research Council Independent Research Institute Infrastructure Support Scheme.
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