Comparative mitogenomics of freshwater snails of the genus Bulinus, obligatory vectors of Schistosoma haematobium, causative agent of human urogenital schistosomiasis.

Among the snail genera most responsible for vectoring human-infecting schistosomes, Bulinus, Biomphalaria, and Oncomelania, the former is in many respects the most important. Bulinid snails host the most common human blood fluke, Schistosoma haematobium, responsible for approximately two-thirds of the estimated 237 million cases of schistosomiasis. They also support transmission of schistosomes to millions of domestic and wild animals. Nonetheless, our basic knowledge of the 37 Bulinus species remains incomplete, especially with respect to genome information, even including mitogenome sequences. We determined complete mitogenome sequences for Bulinus truncatus, B. nasutus, and B. ugandae, and three representatives of B. globosus from eastern, central, and western Kenya. A difference of the location of tRNA-Asp was found between mitogenomes from the three species of the Bulinus africanus group and B. truncatus. Phylogenetic analysis using partial cox1 sequences suggests that B. globosus is a complex comprised of multiple species. We also highlight the status of B. ugandae as a distinct species with unusual interactions with the S. haematobium group parasites deserving of additional investigation. We provide sequence data for potential development of genetic markers for specific or intraspecific Bulinus studies, help elucidate the relationships among Bulinus species, and suggest ways in which mitogenomes may help understand the complex interactions between Schistosoma and Bulinus snails and their relatives.

Examining adaptive evolution of immune activity: opportunities provided by gastropods in the age of 'omics'.

Parasites threaten all free-living organisms, including molluscs. Understanding the evolution of immune defence traits in natural host populations is crucial for predicting their long-term performance under continuous infection risk. Adaptive trait evolution requires that traits are subject to selection (i.e. contribute to organismal fitness) and that they are heritable. Despite broad interest in the evolutionary ecology of immune activity in animals, the understanding of selection on and evolutionary potential of immune defence traits is far from comprehensive. For instance, empirical observations are only rarely in line with theoretical predictions of immune activity being subject to stabilizing selection. This discrepancy may be because ecoimmunological studies can typically cover only a fraction of the complexity of an animal immune system. Similarly, molecular immunology/immunogenetics studies provide a mechanistic understanding of immunity, but neglect variation that arises from natural genetic differences among individuals and from environmental conditions. Here, we review the current literature on natural selection on and evolutionary potential of immune traits in animals, signal how merging ecological immunology and genomics will strengthen evolutionary ecological research on immunity, and indicate research opportunities for molluscan gastropods for which well-established ecological understanding and/or 'immune-omics' resources are already available. This article is part of the Theo Murphy meeting issue 'Molluscan genomics: broad insights and future directions for a neglected phylum'.

Molluscan mitochondrial genomes break the rules.

The first animal mitochondrial genomes to be sequenced were of several vertebrates and model organisms, and the consistency of genomic features found has led to a 'textbook description'. However, a more broad phylogenetic sampling of complete animal mitochondrial genomes has found many cases where these features do not exist, and the phylum Mollusca is especially replete with these exceptions. The characterization of full mollusc mitogenomes required considerable effort involving challenging molecular biology, but has created an enormous catalogue of surprising deviations from that textbook description, including wide variation in size, radical genome rearrangements, gene duplications and losses, the introduction of novel genes, and a complex system of inheritance dubbed 'doubly uniparental inheritance'. Here, we review the extraordinary variation in architecture, molecular functioning and intergenerational transmission of molluscan mitochondrial genomes. Such features represent a great potential for the discovery of biological history, processes and functions that are novel for animal mitochondrial genomes. This provides a model system for studying the evolution and the manifold roles that mitochondria play in organismal physiology, and many ways that the study of mitochondrial genomes are useful for phylogeny and population biology. This article is part of the Theo Murphy meeting issue 'Molluscan genomics: broad insights and future directions for a neglected phylum'.

Sticky problems: extraction of nucleic acids from molluscs.

Traditional molecular methods and omics-techniques across molluscan taxonomy increasingly inform biology of Mollusca. Recovery of DNA and RNA for such studies is challenged by common biological properties of the highly diverse molluscs. Molluscan biomineralization, adhesive structures and mucus involve polyphenolic proteins and mucopolysaccharides that hinder DNA extraction or copurify to inhibit enzyme-catalysed molecular procedures. DNA extraction methods that employ the detergent hexadecyltrimethylammoniumbromide (CTAB) to remove these contaminants importantly facilitate molecular-level study of molluscs. Molluscan pigments may stain DNA samples and interfere with spectrophotometry, necessitating gel electrophoresis or fluorometry for accurate quantification. RNA can reliably be extracted but the 'hidden break' in 28S rRNA of molluscs (like most protostomes) causes 18S and 28S rRNA fragments to co-migrate electrophoretically. This challenges the standard quality control based on the ratio of 18S and 28S rRNA, developed for deuterostome animals. High-AT content in molluscan rRNA prevents the effective purification of polyadenylated mRNA. Awareness of these matters aids the continuous expansion of molecular malacology, enabling work also with museum specimens and next-generation sequencing, with the latter imposing unprecedented demands on DNA quality. Alternative methods to extract nucleic acids from molluscs are available from literature and, importantly, from communications with others who study the molecular biology of molluscs. This article is part of the Theo Murphy meeting issue 'Molluscan genomics: broad insights and future directions for a neglected phylum'.

Transcriptome profiling of Lymnaea stagnalis (Gastropoda) for ecoimmunological research.

BACKGROUND: Host immune function can contribute to numerous ecological/evolutionary processes. Ecoimmunological studies, however, typically use one/few phenotypic immune assays and thus do not consider the complexity of the immune system. Therefore, "omics" resources that allow quantifying immune activity across multiple pathways are needed for ecoimmunological models. We applied short-read based RNAseq (Illumina NextSeq 500, PE-81) to characterise transcriptome profiles of Lymnaea stagnalis (Gastropoda), a multipurpose model snail species. We used a genetically diverse snail stock and exposed individuals to immune elicitors (injury, bacterial/trematode pathogens) and changes in environmental conditions that can alter immune activity (temperature, food availability). RESULTS: Immune defence factors identified in the de novo assembly covered elements broadly described in other gastropods. For instance, pathogen-recognition receptors (PRR) and lectins activate Toll-like receptor (TLR) pathway and cytokines that regulate cellular and humoral defences. Surprisingly, only modest diversity of antimicrobial peptides and fibrinogen related proteins were detected when compared with other taxa. Additionally, multiple defence factors that may contribute to the phenotypic immune assays used to quantify antibacterial activity and phenoloxidase (PO)/melanisation-type reaction in this species were found. Experimental treatments revealed factors from non-self recognition (lectins) and signalling (TLR pathway, cytokines) to effectors (e.g., antibacterial proteins, PO enzymes) whose transcription depended on immune stimuli and environmental conditions, as well as components of snail physiology/metabolism that may drive these effects. Interestingly, the transcription of many factors (e.g., PRR, lectins, cytokines, PO enzymes, antibacterial proteins) showed high among-individual variation. CONCLUSIONS: Our results indicate several uniform aspects of gastropod immunity, but also apparent differences between L. stagnalis and some previously examined taxa. Interestingly, in addition to immune defence factors that responded to immune elicitors and changes in environmental conditions, many factors showed high among-individual variation across experimental snails. We propose that such factors are highly important to be included in future ecoimmunological studies because they may be the key determinants of differences in parasite resistance among individuals both within and between natural snail populations.

Genomic and transcriptional analysis of genes containing fibrinogen and IgSF domains in the schistosome vector Biomphalaria glabrata, with emphasis on the differential responses of snails susceptible or resistant to Schistosoma mansoni.

Achieving a deeper understanding of the factors controlling the defense responses of invertebrate vectors to the human-infecting pathogens they transmit will provide needed new leads to pursue for control. Consequently, we provide new genomic and transcriptomic insights regarding FReDs (containing a fibrinogen domain) and FREPs (fibrinogen domain and one or two IgSF domains) from the planorbid snail Biomphalaria glabrata, a Neotropical vector of Schistosoma mansoni, causative agent of human intestinal schistosomiasis. Using new bioinformatics approaches to improve annotation applied to both genome and RNA-Seq data, we identify 73 FReD genes, 39 of which are FREPs. We provide details of domain structure and consider relationships and homologies of B. glabrata FBG and IgSF domains. We note that schistosome-resistant (BS-90) snails mount complex FREP responses following exposure to S. mansoni infection whereas schistosome-susceptible (M line) snails do not. We also identify several coding differences between BS-90 and M line snails in three FREPs (2, 3.1 and 3.2) repeatedly implicated in other studies of anti-schistosome responses. In combination with other results, our study provides a strong impetus to pursue particular FREPs (2, 3.1, 3.2 and 4) as candidate resistance factors to be considered more broadly with respect to schistosome control efforts, including involving other Biomphalaria species vectoring S. mansoni in endemic areas in Africa.

RNA-seq: The early response of the snail Physella acuta to the digenetic trematode Echinostoma paraensei.

To analyze the response of the snail Physella acuta to Echinostoma paraensei, a compatible digenetic trematode, Illumina RNA-seq data were collected from snails with early infection (5 snails at 2 days post-exposure [DPE]) and established infection (4 snails, 8 DPE), and 7 control (unexposed) snails. A reference transcriptome (325,563 transcripts, including 98% of eukaryotic universal single-copy orthologs; BUSCO) and a draft P. acuta genome (employing available genomic Illumina reads; 799,945 scaffolds, includes 88% BUSCO genes) were assembled to guide RNA-seq analyses. Parasite exposure of P. acuta led to 10,195 differentially expressed (DE) genes at 2 DPE and 8,876 DE genes at 8 DPE with only 18% of up-regulated and 22% of down-regulated sequences shared between these time points. Gene ontology (GO) analysis yielded functional annotation of only 1.2% of DE genes but did not indicate major changes in biological activities of P. acuta between 2 and 8 DPE. Increased insights were achieved by analysis of expression profiles of 460 immune-relevant DE transcripts, identified by BLAST and InterProScan. Physella acuta has expanded gene families that encode immune-relevant domains, including CD109/TEP, GTPase IMAP, Limulus agglutination factor (dermatopontin), FReD (≥82 sequences with fibrinogen-related domains), and transcripts that combine C-type lectin (C-LECT) and C1q domains, novel among metazoa. Notably, P. acuta expressed sequences from these immune gene families at all time points, but the assemblages of unique transcripts from particular immune gene families differed between 2 and 8 DPE. The shift in profiles of DE immune genes, from early exposure to parasite establishment, suggests that compatible P. acuta initially respond to infection but switch to express immune genes that likely are less effective against E. paraensei but counter other types of (opportunistic) pathogens and parasites. We propose that the latter expression profile is part of an extended phenotype of E. paraensei, imposed upon P. acuta through parasite manipulation of the host, following successful parasite establishment in the snail after 2 DPE.

Comparative immunological study of the snail Physella acuta (Hygrophila, Pulmonata) reveals shared and unique aspects of gastropod immunobiology.

The freshwater snail Physella acuta was selected to expand the perspective of comparative snail immunology. Analysis of Physella acuta, belonging to the Physidae, taxonomic sister family to Planorbidae, affords family-level comparison of immune features characterized from Biomphalaria glabrata, the model snail often used to interpret general gastropod immunity. To capture constitutive and induced immune sequences, transcriptomes of an individual Physella acuta snail, 12 h post injection with bacteria (Gram -/+) and one sham-exposed snail were recorded with 454 pyrosequencing. Assembly yielded a combined reference transcriptome containing 24,288 transcripts. Additionally, genomic Illumina reads were obtained (∼15-fold coverage). Recovery of transcripts for two macin-like antimicrobial peptides (AMPs), 12 aplysianins, four LBP/BPIs and three physalysins indicated that Physella acuta shares a similar organization of antimicrobial defenses with Biomphalaria glabrata, contrasting a modest AMP arsenal with a diverse set of antimicrobial proteins. The lack of predicted transmembrane domains in all seven Physella acuta PGRP transcripts supports the notion that gastropods do not employ cell-bound PGRP receptors, different from ecdysozoan invertebrates yet similar to mammals (vertebrate deuterostomes). The well-documented sequence diversification by Biomphalaria glabrata FREPs (immune lectins comprising immunoglobulin superfamily domains and fibrinogen domains), resulting from somatic mutations of a large FREP gene family is hypothesized to be unique to Planorbidae; Physella acuta revealed just two bonafide FREP genes and these were not diversified. Furthermore, the flatworm parasite Echinostoma paraensei, confirmed here to infect both snail species, did not evoke from Physella acuta the abundant expression of FREP proteins at 2, 4 and 8 days post exposure that was previously observed from Biomphalaria glabrata. The Physella acuta reference transcriptome also revealed 24 unique transcripts encoding proteins consisting of a single fibrinogen-related domain (FReDs), with a short N-terminal sequence encoding either a signal peptide, transmembrane domain or no predicted features. The Physella acuta FReDs are candidate immune genes based on implication of similar sequences in immunity of bivalve molluscs. Overall, comparative analysis of snails of sister families elucidated the potential for taxon-specific immune features and investigation of strategically selected species will provide a more comprehensive view of gastropod immunity.

The mitochondrial genome of the planorbid snail Planorbella duryi.

The complete mitochondrial genome of a freshwater planorbid snail, Planorbella duryi (Mollusca, Gastropoda) was recovered from de novo assembly of genomic sequences generated with the Illumina NextSeq500 platform. The P. duryi mitogenome (14,217 base pairs) is AT rich (72.69%) and comprises 13 protein-coding genes, two ribosomal subunit genes, and 22 transfer RNAs. The gene order is identical to that of Biomphalaria glabrata and other snail species in the family Planorbidae. This is the first full characterization of a mitochondrial genome of the genus Planorbella.

Corrigendum: Whole genome analysis of a schistosomiasis-transmitting freshwater snail.

This corrects the article DOI: 10.1038/ncomms15451.

Whole genome analysis of a schistosomiasis-transmitting freshwater snail.

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.

Comparative immunogenomics of molluscs.

Comparative immunology, studying both vertebrates and invertebrates, provided the earliest descriptions of phagocytosis as a general immune mechanism. However, the large scale of animal diversity challenges all-inclusive investigations and the field of immunology has developed by mostly emphasizing study of a few vertebrate species. In addressing the lack of comprehensive understanding of animal immunity, especially that of invertebrates, comparative immunology helps toward management of invertebrates that are food sources, agricultural pests, pathogens, or transmit diseases, and helps interpret the evolution of animal immunity. Initial studies showed that the Mollusca (second largest animal phylum), and invertebrates in general, possess innate defenses but lack the lymphocytic immune system that characterizes vertebrate immunology. Recognizing the reality of both common and taxon-specific immune features, and applying up-to-date cell and molecular research capabilities, in-depth studies of a select number of bivalve and gastropod species continue to reveal novel aspects of molluscan immunity. The genomics era heralded a new stage of comparative immunology; large-scale efforts yielded an initial set of full molluscan genome sequences that is available for analyses of full complements of immune genes and regulatory sequences. Next-generation sequencing (NGS), due to lower cost and effort required, allows individual researchers to generate large sequence datasets for growing numbers of molluscs. RNAseq provides expression profiles that enable discovery of immune genes and genome sequences reveal distribution and diversity of immune factors across molluscan phylogeny. Although computational de novo sequence assembly will benefit from continued development and automated annotation may require some experimental validation, NGS is a powerful tool for comparative immunology, especially increasing coverage of the extensive molluscan diversity. To date, immunogenomics revealed new levels of complexity of molluscan defense by indicating sequence heterogeneity in individual snails and bivalves, and members of expanded immune gene families are expressed differentially to generate pathogen-specific defense responses.

The Planorbid Snail Biomphalaria glabrata Expresses a Hemocyanin-Like Sequence in the Albumen Gland.

The parasitic flatworm Schistosoma mansoni, causative agent of human intestinal schistosomiasis in South America, relies importantly on the freshwater snail Biomphalaria glabrata as intermediate host to achieve development of cercariae that infect humans. The recommendation from the World Health Organization (WHO) to integrate snail control in efforts to counter schistosomiasis transmission provides impetus for in depth study of B. glabrata biology. Our analysis indicates that two distinct hemocyanin-like genes (hcl-1 and hcl-2) are present in B. glabrata, a snail that uses hemoglobin for oxygen transport. Characterization of BAC clones yielded the full length hcl-1 gene, which is comprised of three functional unit (FU) domains at the amino acid level. Database searches and in silico analyses identified the second hcl gene (hcl-2), composed of six FU domains. Both genes are unusual for lacking canonical residues and having fewer FU domains than typical molluscan hemocyanins that contain 7-8 FUs. Reverse transcription PCR demonstrated that Hcl-1 is expressed in a manner that correlates with reproductive maturity in the albumen gland (AG), an immune- and reproduction-relevant organ. Immune cross-reactivity with anti-keyhole limpet hemocyanin (α-KLH) antiserum and tandem-mass spectrometry validated the presence of Hcl-1 protein in the AG and egg mass fluid (EMF). The evolutionary conservation of hemocyanin-like sequences in B. glabrata in the presence of the oxygen carrier hemoglobin, combined with our results, suggest that the Hcl-1protein has a functional role in general and/or reproductive biology. Further investigations are needed to explore Hcl-1 as a potential target for snail control.

Fibrinogen-Related Proteins (FREPs) in Mollusks.

Anti-parasite responses of the snail Biomphalaria glabrata involve antigen-reactive plasma lectins termed fibrinogen-related proteins (FREPs) comprising a C-terminal fibrinogen (FBG) domain and one or two upstream immunoglobulin domains. FREPs are highly polymorphic; they derive from several gene families with multiple loci and alleles that are diversified by exon loss, alternative splicing, and random somatic mutation (gene conversion and point mutations). Individual B. glabrata snails have dynamically distinct FREP sequence repertoires. The immune relevance of B. glabrata FREPs is indicated by FREP binding to polymorphic antigens of (snail-specific) digenean parasites and altered resistance of B. glabrata to digeneans following RNAi knockdown of FREPs. The compatibility polymorphism hypothesis proposes that FREP mutation increases the range of germline-encoded immune recognition in B. glabrata to counter antigenically-varied parasites. Somatic mutation may result from sequence exchange among tandemly arranged FREP genes in the genome, and analysis of sequence variants also suggests involvement of cytidine deaminase-like activity or epigenetic regulation. Without current indications of selection or retention of effective sequence variants toward immunological memory, FREP diversification is thought to afford B. glabrata immunity that is anticipatory but not adaptive. More remains to be learned about this system; other mollusks elaborate diversified lectins consisting of single FBG domains, and bona fide FREPs were reported from additional gastropod species, but these may not be diversified. Future comparative immunological studies and gene discovery driven by next-generation sequencing will further clarify taxonomic distribution of FREP diversification and the underlying mutator mechanisms as a component of immune function in mollusks.

Altered Gene Expression in the Schistosome-Transmitting Snail Biomphalaria glabrata following Exposure to Niclosamide, the Active Ingredient in the Widely Used Molluscicide Bayluscide.

In view of the call by the World Health Organization (WHO) for elimination of schistosomiasis as a public health problem by 2025, use of molluscicides in snail control to supplement chemotherapy-based control efforts is likely to increase in the coming years. The mechanisms of action of niclosamide, the active ingredient in the most widely used molluscicides, remain largely unknown. A better understanding of its toxicology at the molecular level will both improve our knowledge of snail biology and may offer valuable insights into the development of better chemical control methods for snails. We used a recently developed Biomphalaria glabrata oligonucleotide microarray (31K features) to investigate the effect of sublethal exposure to niclosamide on the transcriptional responses of the snail B. glabrata relative to untreated snails. Most of the genes highly upregulated following exposure of snails to niclosamide are involved in biotransformation of xenobiotics, including genes encoding cytochrome P450s (CYP), glutathione S-transferases (GST), and drug transporters, notably multi-drug resistance protein (efflux transporter) and solute linked carrier (influx transporter). Niclosamide also induced stress responses. Specifically, six heat shock protein (HSP) genes from three super-families (HSP20, HSP40 and HSP70) were upregulated. Genes encoding ADP-ribosylation factor (ARF), cAMP response element-binding protein (CREB) and coatomer, all of which are involved in vesicle trafficking in the Golgi of mammalian cells, were also upregulated. Lastly, a hemoglobin gene was downregulated, suggesting niclosamide may affect oxygen transport. Our results show that snails mount substantial responses to sublethal concentrations of niclosamide, at least some of which appear to be protective. The topic of how niclosamide's lethality at higher concentrations is determined requires further study. Given that niclosamide has also been used as an anthelmintic drug for decades and has been found to have activity against several types of cancer, our findings may be of relevance in understanding how both parasites and neoplastic cells respond to this compound.

A family of variable immunoglobulin and lectin domain containing molecules in the snail Biomphalaria glabrata.

Technical limitations have hindered comprehensive studies of highly variable immune response molecules that are thought to have evolved due to pathogen-mediated selection such as fibrinogen-related proteins (FREPs) from Biomphalaria glabrata. FREPs combine upstream immunoglobulin superfamily (IgSF) domains with a C-terminal fibrinogen-related domain (FreD) and participate in reactions against trematode parasites. From RNAseq data we assembled a de novo reference transcriptome of B. glabrata to investigate the diversity of FREP transcripts. This study increased over two fold the number of bonafide FREP subfamilies and revealed important sequence diversity within FREP12 subfamily. We also report the discovery of related molecules that feature one or two IgSF domains associated with different C-terminal lectin domains, named C-type lectin-related proteins (CREPs) and Galectin-related protein (GREP). Together, the highly similar FREPs, CREPs and GREP were designated VIgL (Variable Immunoglobulin and Lectin domain containing molecules).

Physella acuta: atypical mitochondrial gene order among panpulmonates (Gastropoda).

Mitochondrial (mt) sequences are frequently used for phylogenetic reconstruction and for identification of species of molluscs. This study expands the phylogenetic range of Hygrophila (Panpulmonata) for which such sequence data are available by characterizing the full mt genome of the invasive freshwater snail Physella acuta (Physidae). The mt genome sequences of two P. acuta isolates from Stubblefield Lake, New Mexico, USA, differed in length (14,490 vs 14,314 bp) and showed 11.49% sequence divergence, whereas ITS1 and ITS2 sequences from the nuclear genome differed by 1.75%. The mt gene order of P. acuta (cox1, P, nad6, nad5, nad1, D, F, cox2, Y, W, nad4L, C, Q, atp6, R, E, rrnS, M, T, cox3, I, nad2, K, V, rrnL, L1, A, cytb, G, H, L2, atp8, N, nad2, S1, S2, nad4) differs considerably from the relatively conserved gene order within Panpulmonata. Phylogenetic trees show that the 13 protein-encoding mt gene sequences (equivalent codons) of P. acuta group according to gastropod phylogeny, yet branch lengths and dN/dS ratios for P. acuta indicate elevated amino acid substitutions relative to other gastropods. This study indicates that mt sequences of P. acuta are phylogenetically informative despite a considerable intraspecific divergence and the atypical gene order in its mt genome.

Early differential gene expression in haemocytes from resistant and susceptible Biomphalaria glabrata strains in response to Schistosoma mansoni.

The outcome of infection in the host snail Biomphalaria glabrata with the digenean parasite Schistosoma mansoni is determined by the initial molecular interplay occurring between them. The mechanisms by which schistosomes evade snail immune recognition to ensure survival are not fully understood, but one possibility is that the snail internal defence system is manipulated by the schistosome enabling the parasite to establish infection. This study provides novel insights into the nature of schistosome resistance and susceptibility in B. glabrata at the transcriptomic level by simultaneously comparing gene expression in haemocytes from parasite-exposed and control groups of both schistosome-resistant and schistosome-susceptible strains, 2 h post exposure to S. mansoni miracidia, using an novel 5K cDNA microarray. Differences in gene expression, including those for immune/stress response, signal transduction and matrix/adhesion genes were identified between the two snail strains and tests for asymmetric distributions of gene function also identified immune-related gene expression in resistant snails, but not in susceptible. Gene set enrichment analysis revealed that genes involved in mitochondrial electron transport, ubiquinone biosynthesis and electron carrier activity were consistently up-regulated in resistant snails but down-regulated in susceptible. This supports the hypothesis that schistosome-resistant snails recognize schistosomes and mount an appropriate defence response, while in schistosome-susceptible snails the parasite suppresses this defence response, early in infection.