Abstract
BackgroundEchinococcus tapeworms cause a severe helminthic zoonosis called echinococcosis. The genus comprises various species and genotypes, of which E. granulosus (sensu stricto) represents a significant global public health and socioeconomic burden. Mitochondrial (mt) genomes have provided useful genetic markers to explore the nature and extent of genetic diversity within Echinococcus and have underpinned phylogenetic and population structure analyses of this genus. Our recent work indicated a sequence gap (> 1 kb) in the mt genomes of E. granulosus genotype G1, which could not be determined by PCR-based Sanger sequencing. The aim of the present study was to define the complete mt genome, irrespective of structural complexities, using a long-read sequencing method.MethodsWe extracted high molecular weight genomic DNA from protoscoleces from a single cyst of E. granulosus genotype G1 from a sheep from Australia using a conventional method and sequenced it using PacBio Sequel (long-read) technology, complemented by BGISEQ-500 short-read sequencing. Sequence data obtained were assembled using a recently-developed workflow.ResultsWe assembled a complete mt genome sequence of 17,675 bp, which is > 4 kb larger than the complete mt genomes known for E. granulosus genotype G1. This assembly includes a previously-elusive tandem repeat region, which is 4417 bp long and consists of ten near-identical 441–445 bp repeat units, each harbouring a 184 bp non-coding region and adjacent regions. We also identified a short non-coding region of 183 bp, which includes an inverted repeat.ConclusionsWe report what we consider to be the first complete mt genome of E. granulosus genotype G1 and characterise all repeat regions in this genome. The numbers, sizes, sequences and functions of tandem repeat regions remain to be studied in different isolates of genotype G1 and in other genotypes and species. The discovery of such ‘new’ repeat elements in the mt genome of genotype G1 by PacBio sequencing raises a question about the completeness of some published genomes of taeniid cestodes assembled from conventional or short-read sequence datasets. This study shows that long-read sequencing readily overcomes the challenges of assembling repeat elements to achieve improved genomes.
Highlights
Echinococcus tapeworms cause a severe helminthic zoonosis called echinococcosis
From totals of 2757 PacBio long-reads equating to 41 megabase or megabases (Mb), we assembled a complete mt genome for E. granulosus genotype G1 at an average sequencing depth of 2268, resulting in a contig of 17,675 bp, which is > 4 kb larger than all published mt genomes representing genotype G1 (~ 13,600 bp) [16, 21, 22], but with the same order of protein-coding genes (Fig. 1)
Our results are consistent with the previous observations, in terms of the location of non-coding region 1 (NR1) and non-coding region 2 (NR2), and the length and sequence of NR2; the tandem replication of NR2 and its adjacent sequences are unique features not previously reported for mt genomes of cestodes
Summary
Echinococcus tapeworms cause a severe helminthic zoonosis called echinococcosis. The genus comprises various species and genotypes, of which E. granulosus (sensu stricto) represents a significant global public health and socioeconomic burden. Echinococcus is a diverse cestode group, currently consisting of 10 species [4,5,6,7]: E. multilocularis, E. oligarthra, E. vogeli, E. shiquicus, E. granulosus (sensu stricto; genotypes G1 and G3), E. equinus (genotype G4), E. ortleppi (genotype G5), E. intermedius (species name is being debated [6, 8,9,10]; comprising genotypes G6 and G7), E. canadensis (genotypes G8 and G10) and E. felidis These species are distinctly different from one another in their ecology (e.g. infectivity to humans, prevalence, distribution and host ranges) [1]; exploring the extent of genetic variation within the genus Echinococcus is central to understanding disease transmission patterns. Echinococcus granulosus genotype G1 is recognised as the most wide-spread of all Echinococcus taxa, and is, of particular importance [2, 11]
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