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.

Eukaryotic protein kinases (ePKs) are good medical targets for drug development in different biological systems. ePKs participate in many cellular processes, including the p38 MAPK regulation of homeostasis upon oxidative stress. We propose to assess the role of Smp38 MAPK signaling pathway in Schistosoma mansoni development and protection against oxidative stress, parasite survival, and also to elucidate which target genes have their expression regulated by Smp38 MAPK. After a significant reduction of up to 84% in the transcription level by Smp38 MAPK gene knockdown, no visible phenotypic changes were reported in schistosomula in culture. The development of adult worms was tested in vivo in mice infected with the Smp38 knocked-down schistosomula. It was observed that Smp38 MAPK has an essential role in the transformation and survival of the parasites as a low number of adult worms was recovered. Smp38 knockdown also resulted in decreased egg production, damaged adult worm tegument, and underdeveloped ovaries in females. Furthermore, only ~13% of the eggs produced developed into mature eggs. Our results suggest that inhibition of the Smp38 MAPK activity interfere in parasites protection against reactive oxygen species. Smp38 knockdown in adult worms resulted in 80% reduction in transcription levels on the 10th day, with consequent reduction of 94.4% in oviposition in vitro. In order to search for Smp38 MAPK pathway regulated genes, we used an RNASeq approach and identified 1,154 DEGs in Smp38 knockdown schistosomula. A substantial proportion of DEGs encode proteins with unknown function. The results indicate that Smp38 regulates essential signaling pathways for the establishment of parasite homeostasis, including genes related to antioxidant defense, structural composition of ribosomes, spliceosomes, cytoskeleton, as well as, purine and pyrimidine metabolism pathways. Our data show that the Smp38 MAPK signaling pathway is a critical route for parasite development and may present attractive therapeutic targets for the treatment and control of schistosomiasis.

Spliced leader dependent trans-splicing (SLTS) has been described as an important RNA regulatory process that occurs in different organisms, including the trematode Schistosoma mansoni. We identified more than seven thousand putative SLTS sites in the parasite, comprising genes with a wide spectrum of functional classes, which underlines the SLTS as a ubiquitous mechanism in the parasite. Also, SLTS gene expression levels span several orders of magnitude, showing that SLTS frequency is not determined by the expression level of the target gene, but by the presence of particular gene features facilitating or hindering the trans-splicing mechanism. Our in-depth investigation of SLTS events demonstrates widespread alternative trans-splicing (ATS) acceptor sites occurring in different regions along the entire gene body, highlighting another important role of SLTS generating alternative RNA isoforms in the parasite, besides the polycistron resolution. Particularly for introns where SLTS directly competes for the same acceptor substrate with cis-splicing, we identified for the first time additional and important features that might determine the type of splicing. Our study substantially extends the current knowledge of RNA processing by SLTS in S. mansoni, and provide basis for future studies on the trans-splicing mechanism in other eukaryotes.

Schistosomiasis is a neglected tropical disease (NTD) caused by helminthes from the Schistosoma genus. This NTD can cause systemic symptoms induced by the deposition of parasite eggs in the host liver, promoting severe complications. Functional studies to increase knowledge about parasite biology are required for the identification of new drug targets, because the treatment is solely based on praziquantel administration, a drug in which the mechanism of action is still unknown. Protein kinases are important for cellular adaptation and maintenance of many organisms homeostasis and, thus, are considered good drug targets for many pathologies. Accordingly, those proteins are also important for Schistosoma mansoni, as the parasite relies on specific environmental signals to develop into its different stages. However, the specific roles of protein kinases in S. mansoni biology are not well understood. This work aims at investigating the tyrosine-protein kinase FES (Feline Sarcoma) functions in the maintenance of S. mansoni life cycle, especially in the establishment of mammalian and invertebrate hosts' infection. In this regard, the verification of Smfes expression among S. mansoni stages showed that Smfes is more expressed in infective free-living stages: miracidia and cercariae. Schistosomula exposed to SmFES-dsRNA in vitro presented a reduction in movement and size and increased mortality. Mice infected with Smfes-knockeddown schistosomula exhibited a striking reduction in the area of liver granuloma and an increased rate of immature eggs in the intestine. Female adult worms recovered from mice presented a reduced size and changes in the ovary and vitellarium; and males exhibited damage in the gynecophoral canal. Subsequently, miracidia hatched from eggs exposed to SmFES-dsRNA presented changes in its capability to infect and to sense the snail mucus. In addition, the SmFES RNAi effect was stable from miracidia to cercariae. The establishment of infection with those cercariae reproduced the same alterations observed for the knocked-down schistosomula infection. Our findings show that SmFES tyrosine kinase (1) is important in schistosomula development and survival;(2) has a role in adult worms pairing and, consequently, female maturation; (3) might be essential for egg antigen expression, thus responsible for inducing granuloma formation

Dengue is an acute viral disease caused by Dengue virus (DENV) and is considered to be the most common arbovirus worldwide. The clinical characteristics of dengue may vary from asymptomatic to severe complications and severe organ impairment, particularly affecting the liver. Dengue treatment is palliative with acetaminophen (APAP), usually known as Paracetamol, being the most used drug aiming to relieve the mild symptoms of dengue. APAP is a safe and effective drug but, like dengue, can trigger the development of liver disorders. Given this scenario, it is necessary to investigate the effects of combining these two factors on hepatocyte homeostasis. Therefore, this study aimed to evaluate the molecular changes in hepatocytes resulting from the association between DENV infection and treatment with sub-toxic APAP concentrations. Using an in vitro experimental model of DENV-2 infected hepatocytes (AML-12 cells) treated with APAP, we evaluated the influence of the virus and drug association on the transcriptome of these hepatocytes by RNA sequencing (RNAseq). The virus–drug association was able to induce changes in the gene expression profile of AML-12 cells and here we highlight and explore these changes and its putative influence on biological processes for cellular homeostasis.

Schistosomiasis is a parasitic neglected disease with praziquantel (PZQ) utilized as the main drug for treatment, despite its low effectiveness against early stages of the worm. To aid in the search for new drugs to tackle schistosomiasis, computer-aided drug design has been proved a helpful tool to enhance the search and initial identification of schistosomicidal compounds, allowing fast and cost-efficient progress in drug discovery. The combination of high-throughput in silico data followed by in vitro phenotypic screening assays allows the assessment of a vast library of compounds with the potential to inhibit a single or even several biological targets in a more time- and cost-saving manner. Here, we describe the molecular docking for in silico screening of predicted homology models of five protein kinases (JNK, p38, ERK1, ERK2, and FES) of Schistosoma mansoni against approximately 85,000 molecules from the Managed Chemical Compounds Collection (MCCC) of the University of Nottingham (UK). We selected 169 molecules predicted to bind to SmERK1, SmERK2, SmFES, SmJNK, and/or Smp38 for in vitro screening assays using schistosomula and adult worms. In total, 89 (52.6%) molecules were considered active in at least one of the assays. This approach shows a much higher efficiency when compared to using only traditional high-throughput in vitro screening assays, where initial positive hits are retrieved from testing thousands of molecules. Additionally, when we focused on compound promiscuity over selectivity, we were able to efficiently detect active compounds that are predicted to target all kinases at the same time. This approach reinforces the concept of polypharmacology aiming for “one drug-multiple targets”. Moreover, at least 17 active compounds presented satisfactory drug-like properties score when compared to PZQ, which allows for optimization before further in vivo screening assays. In conclusion, our data support the use of computer-aided drug design methodologies in conjunction with high-throughput screening approach.

Mitogen-activated protein kinases (MAPKs) play a regulatory role and influence various biological activities, such as cell proliferation, differentiation, and survival. Our group has demonstrated through functional studies that Schistosoma mansoni c-Jun N-terminal kinase (SmJNK) MAPK is involved in the parasite’s development, reproduction, and survival. SmJNK can, therefore, be considered a potential target for the development of new drugs. Considering the importance of SmJNK in S. mansoni maturation, we aimed at understanding of SmJNK regulated signaling pathways in the parasite, correlating expression data with S. mansoni development. To better understand the role of SmJNK in S. mansoni intravertebrate host life stages, RNA interference knockdown was performed in adult worms and in schistosomula larval stage. SmJNK knocked-down in adult worms showed a decrease in oviposition and no significant alteration in their movement. RNASeq libraries of SmJNK knockdown schistosomula were sequenced. A total of 495 differentially expressed genes were observed in the SmJNK knockdown parasites, of which 373 were down-regulated and 122 up-regulated. Among the down-regulated genes, we found transcripts related to protein folding, purine nucleotide metabolism, the structural composition of ribosomes and cytoskeleton. Genes coding for proteins that bind to nucleic acids and proteins involved in the phagosome and spliceosome pathways were enriched. Additionally, we found that SmJNK and Smp38 MAPK signaling pathways converge regulating the expression of a large set of genes. C. elegans orthologous genes were enriched for genes related to sterility and oocyte maturation, corroborating the observed phenotype alteration. This work allowed an in-depth analysis of the SmJNK signaling pathway, elucidating gene targets of regulation and functional roles of this critical kinase for parasite maturation.

The original version of this Article contained an error in the spelling of the author Leon Di Stefano, which was incorrectly given as Leon di Stephano. This has now been corrected in both the PDF and HTML versions of the Article.