Whole genome analysis of a schistosomiasis-transmitting freshwater snail

Abstract

Biomphalaria snails are instrumental in transmission of the human blood fluke Schistosoma mansoni. With the World Health Organization's goal to eliminate schistosomiasis as a global health problem by 2025, there is now renewed emphasis on snail control. Here, we characterize the genome of Biomphalaria glabrata, a lophotrochozoan protostome, and provide timely and important information on snail biology. We describe aspects of phero-perception, stress responses, immune function and regulation of gene expression that support the persistence of B. glabrata in the field and may define this species as a suitable snail host for S. mansoni. We identify several potential targets for developing novel control measures aimed at reducing snail-mediated transmission of schistosomiasis.

Figure 1. Candidate chemosensory receptors of B. glabrata.

(a) LGUN_random_Scaffold39 contains fourteen candidate chemosensory receptor (CR) genes (BgCRa-n). Most encode seven-transmembrane domain G-protein-coupled receptor-like proteins, BgCRm and BgCRn are truncated to six-transmembrane domains. See Supplementary Data 11 for gene model identifiers. (b) Phylogenetic analysis (neighbour joining, scale bar represents amino-acid substitutions per site) of chemosensory receptors on LGUN_random_Scaffold39 (protein-level). (c) Schematic of receptor showing conserved and invariable amino acids, transmembrane domains I-VII; and location of glycosylation sites. (d) Scanning electron micrograph showing anterior tentacle, with cilia covering the surface. Scale bar, 20 μm (top); 10 μm (bottom). (e) RT–PCR gel showing amplicon for BgCR509a and actin from B. glabrata tentacle. (f,g) In situ hybridization showing sense (negative control) and antisense localization of BgCR509a mRNA in anterior tentacle section (purple). Scale bar (f): 100 μm; (g) 50 μm.

Figure 2. TLR genes in B. glabrata.

(a) Analysis of the (complete) TIR domains from BgTLRs identified seven classes (neighbour-joining tree, scale bar represents amino-acid substitutions per site). Bootstrap values shown for 1,000 replicates. Comparisons included TLRs from A. californica (Ac), L. gigantea (Lg), Mytilus galloprovincialis (Mg) and C. gigas (Cg). The presence or absence of orthologues of each class in each molluscan species is indicated. A representative of the B. glabrata class 1/2/3 clade is present within A. californica, but is independent of the B. glabrata TLR classes (indicated by the large pink box). Grey font indicates pseudogenes or partial genes. (b) B. glabrata has both single cysteine cluster (scc; blue line)- and multiple cysteine cluster (mcc; orange line) TLRs. Domain structures are shown for BgTLR classes. BgTLRs consist of an LRRNT (orange hexagon), a series of LRRs (ovals), a variable region (curvy line), LRRCT (yellow box), and transmembrane domain, and an intracellular TIR domain (hexagon). The dark blue ovals indicate well defined LRRs (predicted by LRRfinder57); light blue ovals are less confident predictions. Each of the two class 7 BgTLRs has a distinct ectodomain structure. The numbers of complete, pseudogenes (Ψ) and partial genes are indicated for each class.

Figure 3. Expression of cardiac genes and actin genes in B. glabrata tissues.

(a) Cardiac regulatory genes. (b) Cardiac structural genes. (c) Relative expression of actin genes in B. glabrata tissues. For (a–c), the score represents gene level aggregate of normalized FPKM counts for de novo assembled tissue transcripts, relative to expression levels in the heart/APO sample. The counts were scaled (with median read count as 0) to indicate expression intensity with red indicating highest, blue lowest. AG, Albumen gland; BUC, buccal mass; CNS, central nervous system; DG, digestive gland; FOOT, headfoot; HAPO, heart/APO; KID, kidney; MAN, mantle edge; OVO, ovotestes; SAL, salivary glands; STO, stomach; TRG, terminal genitalia. (d) Maximum Likelihood tree (Phylogeny.fr, scale bar represents amino-acid substitutions per site) showing phylogenetic relationships of actin genes, based on amino-acid sequence alignment (ClustalW). Biomphalaria -snail; Crassostrea gigas—oyster; Haliotis iris– abalone;, Hirudo medicinalis – leech (all lophotrochozoans); Amphimedon queenslandica, sponge, Prebilateria, ophotrochozoans), Drosophila melanogaster—fruit fly, Ecdysozoa), and the deuterostomes Ciona intestinalis, sea squirt; Homo sapiens, human. See Supplementary Note 31 for accession numbers.

Figure 4. Comparison of molluscan shell forming proteomes.

Circos diagram of 177 mantle-specific, secreted Biomphalaria gene products compared against shell forming proteomes of six other molluscs (BLASTp threshold ≤10e⁻⁶). Protein pairs that share sequence similarity in the top quartile are linked in purple, the second quartile is linked in blue, third quartile has orange links and lowest quartile of similarity has grey links. Species that occupy marine habitats are surrounded by a solid line, A finely dashed line identifies the terrestrial species Cepea nemoralis, the freshwater species B. glabrata has a coarse line. Percentages (and proportions in brackets) indicate the number of proteins that shared similarity with a Biomphalaria shell forming candidate gene. The width of each sector line around the ideogram is proportional to the length of that gene in basepairs. Photographs taken by DJ Jackson, with exception of photograph of C. gigas, by David Monniaux, distributed under a CC-A SA 3.0 license.

Figure 5. Transposable element (TE) landscape of B. glabrata.

(a) Left: proportion (%) of the genome assembly annotated as TE (black). Right: TE composition by class (indicating % of the genome corresponding to each class). (b) Evolutionary view of TE landscape. For each class, cumulative amounts of DNA (in Mb) are shown as function of the percentage of divergence from the consensus (by bins of 1%, first one being ≥0 and <1; see Supplementary Note 32 for Methods). Percentage of divergence from consensus is used as a proxy for age: the older the invasion of the TE is, the more copies will have accumulated mutations (higher percentage of divergence, right of the graph; left of the graph: youngest elements showing little divergence from consensus). Note that the result of this analysis of assembled sequence does not exclude the likelihood that intact transposable elements are present in B. glabrata. Colors are as in a. RC, rolling circle.