Fig 1.
A map of the United States and Canada depicting the expansion of E. multilocularis over the last several decades.
Each color represents a different decade wherein a peer-reviewed study first confirmed the presence of E. multilocularis morphologically, molecularly, or both. In cases where only one or a few isolated reports exist (e.g., the western U.S.), polygons have been expanded slightly beyond the actual range of detection for visibility. Methods for creating this map can be found in S1 Text. A similar map of Europe and Asia can be found in S1 Fig. The base map for this figure is available via Wikipedia Commons at the following link: https://commons.wikimedia.org/wiki/File:North_America_blank_map_with_state_and_province_boundaries.png.
Fig 2.
Typical life cycle of Echinococcus multilocularis, with definitive host (canids), intermediate host (rodent), and an aberrant intermediate host (human) pictured.
Fig 3.
Study area map showing locations of coyote carcasses and field-collected scats in this study, and the infection status of each sample.Points are jittered to better represent multiple individuals collected from the same location.
The study area is broken up into two sub-areas, north and south, for ease of map viewing. Note that no field-collected coyote scats tested positive for E. multilocularis, but several carcasses from the same region where scats were collected did test positive. This map was created using program R with a basemap from OpenStreetMap: https://www.openstreetmap.org/copyright.
Table 1.
Cestode primer pair used in this study to sequence E. multilocularis with next-generation sequencing. The forward primer was designed for this study. Reverse primer is from 62 (Cest 5).
Table 2.
Summary of E. multilocularis prevalence in our study, split by sample type (carcass or field-collected scat).
Table 3.
Table depicting each sample type, sample sizes, and number and percent of Echinococcus multilocularis infected individuals (carcasses only) in this study. Note that some individuals are represented across multiple sample types, and therefore the total number of individuals is less than the total number of samples. To calculate the number and percent of infected individuals, an individual was considered infected if at least one of their sample types was positive.
Fig 4.
Haplotype network constructed using the pegas package in R with the cob, cox1, and nad2 genes, showing some of the global published haplotypes of E. multilocularis and using the haplotype grouping system recently published in[61].
GenBank accession numbers for each haplotype from the literature are available in Table 1 of 64. Each circle represents one haplotype with names as listed in [61]; full names in Lallemand et al. include the prefix “mtG”). Country or region names below haplotype names indicate where the specimen that was sequenced came from. Tick marks between circles represent mutational steps (i.e., single nucleotide polymorphisms) that separate each haplotype from its relatives. The haplotype network is color-coded using haplotype groups from [61]. The haplotype from our study (GenBank accession numbers PX737897, PX737898, PX737899; labeled h28 WA & BC) is indicated in white and was first published in [28,62] from a dog in Quesnel, British Columbia, Canada (100% base pair match between our sequences and that from [62]. This haplotype is referred to as BC1 in some publications [2,32] and is most closely related to the mtG h1 haplotype from France (only two base pairs apart). Another haplotype from Canada (unknown region) in Haplogroup 2 is marked with an asterisk, first published in [63].
Fig 5.
Representative specimens of E. multilocularis with photos of an unstained whole worm (A), a stained gravid proglottid (B), and a second stained gravid proglottid (C).
These photos demonstrate identifying characteristics of Echinococcus multilocularis, especially in comparison to Echinococcus granulosus, a close relative ([54], see [60] for direct comparisons between the two species). Scale bars are included for each image. In A, the gravid proglottid is less than half the length of the total body length (it is more than half in E. granulosus). The total length of the specimen in A is 2.22 mm (2,221.356 µm) and its gravid proglottid is 1.07 mm (1,072.179 µm). An arrow points to the genital pore locations in B. The genital pore is anterior to the midpoint of the proglottid (it is posterior to the midpoint in E. granulosus). The sac-like uterus shape can also be seen in A, B, and C (E. granulosus has a laterally branching uterus). Photo backgrounds have been edited and arrows added to improve clarity of photos (unmanipulated versions of A, B, and C can be found in S2, S3, and S4 Figs, respectively).