The pathophysiology of schistosomiasis is linked to the formation of fibrous granulomas around eggs that become trapped in host tissues, particularly the intestines and liver, during their migration to reach the lumen of the vertebrate gut. While the development of Schistosoma egg-induced granulomas is the result of finely regulated crosstalk between egg-secreted antigens and host immunity, evidence has started to emerge of the likely contribution of an additional player-the host gut microbiota-to pathological processes that culminate with the formation of these tissue lesions. Uncovering the role(s) of schistosome-mediated changes in gut microbiome composition and function in granuloma formation and, more broadly, in the pathophysiology of schistosomiasis, will shed light on the mechanisms underlying this three-way parasite-host-microbiome interplay. Such knowledge may, in turn, pave the way towards the discovery of novel therapeutic targets and control strategies.
With the exception of Aelurostrongylus abstrusus, feline lungworms have been poorly studied. Information on their distribution is patchy and mostly limited to case reports. In this study, the occurrence of feline lungworms and co-infecting gastrointestinal parasites has been investigated in 12 European countries (i.e. Austria, Belgium, Bulgaria, France, Greece, Hungary, Italy, Portugal, Romania, Spain, Switzerland and the United Kingdom). An average of 10 domestic cats, with regular outdoor access, was sampled each month for 12months, and freshly passed faeces were collected. Stools were processed using a McMaster assay and a quantitative Baermann-Wetzel method. Animals positive for lungworms and/or gastrointestinal parasites were treated with a formulation containing fipronil, (S)-methoprene, eprinomectin, and praziquantel (Broadline®, Merial), and re-sampled 28days post-treatment. The association between lungworm infection and risk factors was analysed using statistical medians/means and the efficacy of the treatment against each lungworm species was assessed. Of 1990 cats sampled, 613 (30.8%) were positive for at least one parasite, while 210 (10.6%) were infected by lungworms. The prevalence of lungworm infection varied between the sampled sites, with the highest recorded in Bulgaria (35.8%) and the lowest in Switzerland (0.8%). None of the cats from Austria or the United Kingdom were infected by lungworms. Aelurostrongylus abstrusus was the species most frequently detected (78.1%), followed by Troglostrongylus brevior (19.5%), Eucoleus aerophilus (14.8%) and Oslerus rostratus (3.8%). The overall efficacy of the treatment was 99% for A. abstrusus and 100% for T. brevior, O. rostratus and E. aerophilus. Data presented provide a comprehensive account of the diagnosis, epidemiology and treatment of feline lungworms in Europe, as well as of the occurrence of co-infections by gastrointestinal parasites.
Parasitic liver flukes of the family Fasciolidae are responsible for major socioeconomic losses worldwide. However, at present, knowledge of the fundamental molecular biology of these organisms is scant. Here, we characterize, for the first time, the transcriptome and secreted proteome of the adult stage of the "giant liver fluke," Fascioloides magna, using Illumina sequencing technology and one-dimensional SDS-PAGE and OFFGEL protein electrophoresis, respectively. A total of ∼54,000,000 reads were generated and assembled into ∼39,000 contiguous sequences (contigs); ∼20,000 peptides were predicted and classified based on homology searches, protein motifs, gene ontology, and biological pathway mapping. From the predicted proteome, 48.1% of proteins could be assigned to 384 biological pathway terms, including "spliceosome," "RNA transport," and "endocytosis." Putative proteins involved in amino acid degradation were most abundant. Of the 835 secreted proteins predicted from the transcriptome of F. magna, 80 were identified in the excretory/secretory products from this parasite. Highly represented were antioxidant proteins, followed by peptidases (particularly cathepsins) and proteins involved in carbohydrate metabolism. The integration of transcriptomic and proteomic datasets generated herein sets the scene for future studies aimed at exploring the potential role(s) that molecules might play at the host–parasite interface and for establishing novel strategies for the treatment or control of parasitic fluke infections. Parasitic liver flukes of the family Fasciolidae are responsible for major socioeconomic losses worldwide. However, at present, knowledge of the fundamental molecular biology of these organisms is scant. Here, we characterize, for the first time, the transcriptome and secreted proteome of the adult stage of the "giant liver fluke," Fascioloides magna, using Illumina sequencing technology and one-dimensional SDS-PAGE and OFFGEL protein electrophoresis, respectively. A total of ∼54,000,000 reads were generated and assembled into ∼39,000 contiguous sequences (contigs); ∼20,000 peptides were predicted and classified based on homology searches, protein motifs, gene ontology, and biological pathway mapping. From the predicted proteome, 48.1% of proteins could be assigned to 384 biological pathway terms, including "spliceosome," "RNA transport," and "endocytosis." Putative proteins involved in amino acid degradation were most abundant. Of the 835 secreted proteins predicted from the transcriptome of F. magna, 80 were identified in the excretory/secretory products from this parasite. Highly represented were antioxidant proteins, followed by peptidases (particularly cathepsins) and proteins involved in carbohydrate metabolism. The integration of transcriptomic and proteomic datasets generated herein sets the scene for future studies aimed at exploring the potential role(s) that molecules might play at the host–parasite interface and for establishing novel strategies for the treatment or control of parasitic fluke infections. Parasitic liver flukes (Platyhelminthes: Trematoda) of livestock, such as Fasciola hepatica and Fasciola gigantica, are responsible for major economic losses worldwide, estimated at USD ∼3 billion, due to morbidity, mortality, and decreased productivity (1Boray J.C. Chemotherapy of infections with Fasciolidae.in: Boray J.C. Immunology, Pathobiology and Control of Fasciolosis. MSD AGVET, Rahway, NJ1997: 83-97Google Scholar, 2Kaplan R.M. Fasciola hepatica: a review of the economic impact in cattle and considerations for control.Vet. 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The penetration of Fascioloides magna miracidia into the snail host Fossaria bulimoides. A scanning electron microscope study.Zeitschrift für Parasitenkunde. 1977; 52: 53-59Crossref PubMed Scopus (7) Google Scholar). In the intermediate host, the parasite develops through the stages of sporocysts, rediae, and cercariae; the latter larval stage emerges from the snail within ∼40–58 days (4Pybus M.J. Liver flukes.in: Samuel W.M. Pybus M.J. Kocan A.A. Parasitic Diseases in Wild Mammals. Iowa State Press, Iowa City, IA2001: 121-149Google Scholar). The cercariae encyst as metacercariae on submerged or emergent vegetation and are then ingested by a mammalian host. Once in the host, the metacercariae excyst and the juvenile flukes penetrate the intestinal walls and migrate to the liver, where they are encapsulated (in pairs) in pseudocysts formed by the hepatic parenchyma and then develop into mature flukes (4Pybus M.J. Liver flukes.in: Samuel W.M. Pybus M.J. Kocan A.A. Parasitic Diseases in Wild Mammals. Iowa State Press, Iowa City, IA2001: 121-149Google Scholar). The migration of the immature stages through the hepatic tissues, together with the large number of pseudocysts and the size of mature flukes (up to 8 cm in length (4Pybus M.J. Liver flukes.in: Samuel W.M. Pybus M.J. Kocan A.A. Parasitic Diseases in Wild Mammals. Iowa State Press, Iowa City, IA2001: 121-149Google Scholar)), can result in liver fibrosis. Clinical signs associated with infection by F. magna include lethargy, anorexia, depression, and weight loss, with sudden death occurring in heavily infected animals (4Pybus M.J. Liver flukes.in: Samuel W.M. Pybus M.J. Kocan A.A. Parasitic Diseases in Wild Mammals. Iowa State Press, Iowa City, IA2001: 121-149Google Scholar). In livestock, the control of liver fluke infections has relied predominantly on treatment with anthelmintic drugs, such as closantel, oxyclozanide, and triclabendazole (6Fairweather I. 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Elucidating the transcriptome of Fasciola hepatica——a key to fundamental and biotechnological discoveries for a neglected parasite.Biotechnol. Adv. 2010; 28: 222-231Crossref PubMed Scopus (113) Google Scholar, 24Robinson M.W. Menon R. Donnelly S.M. Dalton J.P. Ranganathan S. An integrated transcriptomics and proteomics analysis of the secretome of the helminth pathogen Fasciola hepatica: proteins associated with invasion and infection of the mammalian host.Mol. Cell. Proteomics. 2009; 8: 1891-1907Abstract Full Text Full Text PDF PubMed Scopus (231) Google Scholar, 25Young N.D. Jex A.R. Cantacessi C. Hall R.S. Campbell B.E. Spithill T.W. Tangkawattana S. Tangkawattana P. Laha T. Gasser R.B. A portrait of the transcriptome of the neglected trematode, Fasciola gigantica—biological and biotechnological implications.PLoS Negl. Trop. 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Dis. 2011; 5: e1004Crossref PubMed Scopus (81) Google Scholar) and of the composition of the excretory/secretory (ES) 1The abbreviations used are:BLASTBasic Local Alignment Search ToolESexcretory/secretoryGOgene ontologyKEGGKyoto Encyclopedia of Genes and GenomesDTTdithiothreitolIAAiodoacetamideNCBINational Center for Biotechnology InformationOGEOFF-GEL electrophoresisPBSphosphate-buffered salineSDS-PAGEsodium dodecyl-sulfate polyacrylamide gel electrophoresis.1The abbreviations used are:BLASTBasic Local Alignment Search ToolESexcretory/secretoryGOgene ontologyKEGGKyoto Encyclopedia of Genes and GenomesDTTdithiothreitolIAAiodoacetamideNCBINational Center for Biotechnology InformationOGEOFF-GEL electrophoresisPBSphosphate-buffered salineSDS-PAGEsodium dodecyl-sulfate polyacrylamide gel electrophoresis. products from both the juvenile and adult stages of Fa. hepatica (24Robinson M.W. Menon R. Donnelly S.M. Dalton J.P. Ranganathan S. An integrated transcriptomics and proteomics analysis of the secretome of the helminth pathogen Fasciola hepatica: proteins associated with invasion and infection of the mammalian host.Mol. Cell. Proteomics. 2009; 8: 1891-1907Abstract Full Text Full Text PDF PubMed Scopus (231) Google Scholar). In these studies, proteolytic enzymes (e.g. cathepsins, asparaginyl endopeptidase cysteine proteases, and trypsin-like serine proteases) were identified as key molecules in both the transcriptome and ES products, which are likely to play key roles in parasite migration through tissues and in the modulation of immune responses in the mammalian host. Given the potential of proteolytic enzymes as vaccine candidates against trematodiases (18Dalton J.P. Mulcahy G. Parasite vaccines—a reality?.Vet. Parasitol. 2001; 98: 149-167Crossref PubMed Scopus (106) Google Scholar, 19Dalton J.P. Brindley P.J. Knox D.P. Brady C.P. Hotez P.J. Donnelly S. O'Neill S.M. Mulcahy G. Loukas A. 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Advances in "-omic" and computational technologies for the preprocessing, assembly, and annotation of sequence data (27DeMarco R. Verjovski-Almeida S. Schistosomes—proteomics studies for potential novel vaccines and drug targets.Drug Discov. Today. 2009; 14: 472-478Crossref PubMed Scopus (56) Google Scholar, 28Chuan J. Feng Z. Brindley P.J. McManus D.P. Han Z. Jianxin P. Hu W. Our wormy world: genomics, proteomics and transcriptomics in East and southeast Asia.Adv. Parasitol. 2010; 73: 327-371Crossref PubMed Scopus (20) Google Scholar, 29Cantacessi C. Campbell B.E. Gasser R.B. Key strongylid nematodes of animals—impact of next-generation transcriptomics on systems biology and biotechnology.Biotechnol. Adv. 2012; 30: 469-488Crossref PubMed Scopus (35) Google Scholar) are substantially assisting studies of the transcriptomes and proteomes of parasitic helminths (3Young N.D. Hall R.S. Jex A.R. Cantacessi C. Gasser R.B. 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This insight into the molecular biology of F. magna offers unprecedented opportunities for comparative investigations of various economically important liver flukes and the design of new interventions against these parasites. Adult F. magna were collected from naturally infected livers of red deer (Cervus elaphus) (from the Brdy mountains, Czech Republic) and washed in 0.1 m phosphate-buffered saline (PBS), pH 7.2, at 37 °C. Total RNA was extracted from three individual adults of F. magna using TriPure reagent (Roche, Switzerland) and DNase I-treated (25Young N.D. Jex A.R. Cantacessi C. Hall R.S. Campbell B.E. Spithill T.W. Tangkawattana S. Tangkawattana P. Laha T. Gasser R.B. A portrait of the transcriptome of the neglected trematode, Fasciola gigantica—biological and biotechnological implications.PLoS Negl. Trop. Dis. 2011; 5: e1004Crossref PubMed Scopus (81) Google Scholar). 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Balasubramanian S. Swerdlow H.P. George D. Gietzen K.J. Goddard C.P. Golda G.S. Granieri P.A. Green D.E. Gustafson D.L. Hansen N.F. Harnish K. Haudenschild C.D. Heyer N.I. Hims M.M. Ho J.T. Horgan A.M. Hoschler K. Hurwitz S. Ivanov D.V. Johnson M.Q. James T. Huw Jones T.A. Kang G.D. Kerelska T.H. Kersey A.D. Khrebtukova I. Kindwall A.P. Kingsbury Z. Kokko-Gonzales P.I. Kumar A. Laurent M.A. Lawley C.T. Lee S.E. Lee X. Liao A.K. Loch J.A. Lok M. Luo S. Mammen R.M. Martin J.W. McCauley P.G. McNitt P. Mehta P. Moon K.W. Mullens J.W. Newington T. Ning Z. Ling Ng B. Novo S.M. O'Neill M.J. Osborne M.A. Osnowski A. Ostadan O. Paraschos L.L. Pickering L. Pike A.C. Pike A.C. Chris Pinkard D. Pliskin D.P. Podhasky J. Quijano V.J. Raczy C. Rae V.H. Rawlings S.R. Chiva Rodriguez A. Roe P.M. Rogers J. Rogert Bacigalupo M.C. Romanov N. Romieu A. Roth R.K. Rourke N.J. Ruediger S.T. Rusman E. Sanches-Kuiper R.M. Schenker M.R. Seoane J.M. Shaw R.J. Shiver M.K. Short S.W. Sizto N.L. Sluis J.P. Smith M.A. Ernest Sohna Sohna J. Spence E.J. Stevens K. Sutton N. Szajkowski L. Tregidgo C.L. Turcatti G. Vandevondele S. Verhovsky Y. Virk S.M. Wakelin S. Walcott G.C. Wang J. Worsley G.J. Yan J. Yau L. Zuerlein M. Rogers J. Mullikin J.C. Hurles M.E. McCooke N.J. West J.S. Oaks F.L. Lundberg P.L. Klenerman D. Durbin R. Smith A.J. Accurate whole human genome sequencing using reversible terminator chemistry.Nature. 2008; 456: 53-59Crossref PubMed Scopus (2519) Google Scholar), according to the manufacturer's protocols. The 100 bp single-read sequences generated from the non-normalized cDNA library representing the adult stage of F. magna were assembled using the program Oases v0.1.22 (http://www.ebi.ac.uk/∼zerbino/oases/; (34Schulz M.H. Zerbino D.R. Vingron M. Birney E. Oases: robust de novo RNA-seq assembly across the dynamic range of expression levels.Bioinformatics. 2012; 28: 1086-1092Crossref PubMed Scopus (1044) Google Scholar)). Adapter sequences and sequences with suboptimal read quality (i.e. PHRED score < 32.0) were eliminated. The remaining sequences (99%) were used to construct a de Bruijn graph using a k-mer value of 43 bp. Because de novo transcriptome assemblies of short-read sequence libraries can potentially lead to an underrepresentation of members of large families of molecules characterized by a high degree of sequence similarity, such as the cathepsins (25Young N.D. Jex A.R. Cantacessi C. Hall R.S. Campbell B.E. Spithill T.W. Tangkawattana S. Tangkawattana P. Laha T. Gasser R.B. A portrait of the transcriptome of the neglected trematode, Fasciola gigantica—biological and biotechnological implications.PLoS Negl. Trop. 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Hall R.S. Campbell B.E. Spithill T.W. Tangkawattana S. Tangkawattana P. Laha T. Gasser R.B. A portrait of the transcriptome of the neglected trematode, Fasciola gigantica—biological and biotechnological implications.PLoS Negl. Trop. Dis. 2011; 5: e1004Crossref PubMed Scopus (81) Google Scholar, 37Cantacessi C. Jex A.R. Hall R.S. Young N.D. Campbell B.E. Joachim A. Nolan M.J. Abubucker S. Sternberg P.W. Ranganathan S. Mitreva M. Gasser R.B. A practical, bioinformatic workflow system for large data sets generated by next-generation sequencing.Nucleic Acids Res. 2010; 38: e171Crossref PubMed Scopus (62) Google Scholar). Briefly, assembled contigs were compared (using BLASTn and BLASTx algorithms) with (i) sequences available in public databases, including that of NCBI (www.ncbi.nlm.nih.gov) and the EMBL-EBI Parasite Genome Blast Server (www.ebi.ac.uk); (ii) transcriptomic sequence data available for the related liver flukes Fa. hepatica, Fa. gigantica (3Young N.D. Hall R.S. Jex A.R. 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Ascaridoid nematodes are widespread in marine fishes. Despite their major socioeconomic importance, mechanisms associated to the fish-borne zoonotic disease anisakiasis are still obscure. RNA-Seq and de-novo assembly were herein applied to RNA extracted from larvae and dissected pharynx of Hysterothylacium aduncum (HA), a non-pathogenic nematode. Assembled transcripts in HA were annotated and compared to the transcriptomes of the zoonotic species Anisakis simplex sensu stricto (AS) and Anisakis pegreffii (AP). Approximately 60,000,000 single-end reads were generated for HA, AS and AP. Transcripts in HA encoded for 30,254 putative peptides while AS and AP encoded for 20,574 and 20,840 putative peptides, respectively. Differential gene expression analyses yielded 471, 612 and 526 transcripts up regulated in the pharynx of HA, AS and AP. The transcriptomes of larvae and pharynx of HA were enriched in transcripts encoding collagen, peptidases, ribosomal proteins and in heat-shock motifs. Transcripts encoding proteolytic enzymes, anesthetics, inhibitors of primary hemostasis and virulence factors, anticoagulants and immunomodulatory peptides were up-regulated in AS and AP pharynx. This study represents the first transcriptomic characterization of a marine parasitic nematode commonly recovered in fish and probably of negligible concern for public health.
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