Elsevier

Biotechnology Advances

Volume 28, Issue 6, November–December 2010, Pages 859-870
Biotechnology Advances

Research review article
Progress on the transcriptomics of carcinogenic liver flukes of humans—Unique biological and biotechnological prospects

https://doi.org/10.1016/j.biotechadv.2010.07.006Get rights and content

Abstract

Liver flukes, such as Clonorchis sinensis and Opisthorchis viverrini, are food-borne parasites that have a major impact on the health of humans and animals, particularly in Asia. However, the impact of C. sinensis and O. viverrini, in particular, is exacerbated in that these parasites can induce a malignant, untreatable cancer (cholangiocarcinoma, CCA) in chronically infected people. As a result, these flukes are classified as Group 1 carcinogens. Despite their substantial socio-economic importance, little is known about these parasites and their relationship with the definitive hosts at the molecular level. Here, we provide a background on these two carcinogenic flukes and review recent progress on characterizing their transcriptomes using next-generation technologies. We also describe the prospects that the transcriptomes of C. sinensis and O. viverrini provide as a resource for future -omic explorations and efforts to develop improved methods of intervention and control against these important pathogens and CCA, leading to biotechnological outcomes.

Introduction

Compounded by a massive global food shortage, parasitic worms are one of the world's greatest challenges. Literally billions of people are infected with worms, which have a comparable socio-economic burden to that of diabetes or lung cancer in disability adjusted life years (DALYs) (World Health Organization, 2004). Liver flukes (Platyhelminthes: Trematoda: Digenea) are particularly important food-borne worms of humans (Keiser and Utzinger, 2009). Clonorchis sinensis and Opisthorchis viverrini (Opisthorchiidae) cause the diseases called clonorchiasis and opisthorchiasis, respectively, which have a substantial public health impact mainly in countries of the Asia Pacific (Lun et al., 2005, Sripa et al., 2007). Both parasites severely affect tens of millions of people, and ~ 600 million people are at risk of infection (Keiser and Utzinger, 2005, Sripa et al., 2007). Despite control efforts, their prevalence can be as high as 70% in some Asian countries (Sithithaworn and Haswell-Elkins, 2003, Lun et al., 2005). Chronic infections are mainly linked to cholangitis and associated problems in the liver, most importantly, bile duct cancer (= cholangiocarcinoma, CCA) (Lun et al., 2005, Sripa et al., 2007, Shin et al., 2010). Although, theoretically, infections could be prevented by persuading people to consume cooked fish, clonorchiasis and opisthorchiasis persist in endemic regions (Sripa et al., 2007, Keiser and Utzinger, 2009). Control relies mainly on treating infected people with praziquantel (an anthelmintic compound). It appears that humans develop only a limited degree of immunity against opisthorchiid liver flukes (see Wongratanacheewin et al., 2003), such that they frequently become re-infected (Sithithaworn et al., 2002, Lun et al., 2005). Although there has also been a focus on developing alternative control methods (Keiser et al., 2006, Keiser et al., 2009), no vaccines or new drugs are yet available.

Fundamental molecular biological investigations are needed to underpin the development of novel methods of diagnosis, treatment and control. However, to date, most molecular studies of flukes have focused on human blood flukes (schistosomes). Complementing the nuclear genome sequencing of schistosomes (Berriman et al., 2009, Liu et al., 2009) have been sustained efforts to establish in vitro systems for functional genomic analyses (Brindley and Pearce, 2007, Kalinna and Brindley, 2007, Ndegwa et al., 2007, Morales et al., 2008, Rinaldi et al., 2009). This is in stark contrast to the situation for other flukes, for which the potential to explore transcriptomes and gene function is only now being realized (McGonigle et al., 2008, Rinaldi et al., 2008). Despite their major socio-economic impact, no draft or complete nuclear genomic sequence is available for liver flukes, and transcriptomic data have been scant. However, recent investigations provide exciting new prospects to explore biological pathways in various socio-economically important liver flukes, with a view toward biotechnological outcomes. In the present article, we provide a concise account of the biology of C. sinensis and O. viverrini and the diseases that these parasites cause; refer to the current treatment in humans; indicate the need for novel intervention strategies; and, importantly, report on recent progress on the transcriptomics of these carcinogenic liver flukes using next-generation sequencing and bioinformatic technologies. We also emphasize the prospects that transcriptomes provide for future -omic explorations and efforts toward developing improved methods for the control of these flukes and the diseases that they cause.

Section snippets

Biology of carcinogenic liver flukes and associated diseases

The life histories of C. sinensis and O. viverrini are similar (Yoshitaka and Dawes, 1967, Kaewkes, 2003, Upatham and Viyanant, 2003), involving an aquatic snail (order Mesogastropoda), in which asexual reproduction takes place, and freshwater cyprinid fishes or palaemonid shrimps (for C. sinensis only) as intermediate hosts (Fig. 1). Piscivorous mammals, including humans, canids and felids, act as definitive hosts, in which sexual reproduction occurs. Infections are prevalent in geographical

Cholangiocarcinoma and the need for novel intervention strategies

Chronic clonorchiasis or opisthorchiasis is associated with hepatobiliary damage, inflammation, periductal fibrosis and/or cellular responses to antigens from the infecting fluke (Sripa et al., 2007) and predisposes to CCA (Thamavit et al., 1978, Kim, 1984, Lee et al., 1993). Chronic hepatobiliary damage is considered to be multi-factorial, arising from a continued irritation of the bile duct epithelium by adult flukes, particularly their suckers, metabolites and excreted/secreted molecules (

Next-generation approaches for the sequencing and assembly/annotation of the transcriptomes of carcinogenic flukes

Recently, Young et al. (2010a) utilized 454 sequencing (Roche) from normalized cDNA libraries and a semi-automated bioinformatic platform to assemble and annotate non-redundant sequence datasets representative of the adult transcriptomes of C. sinensis and O. viverrini. Briefly, nucleotide sequences (> 500,000 per species) were assembled de novo and specifically clustered with similar protein coding domains. Redundancy was reduced by clustering sequences, if their predicted proteins aligned over

Major prospects for future -omic research of carcinogenic flukes — biological and biotechnological implications

Two studies conducted recently (Young et al., 2010a, Young et al., 2010b) provide a first, deep insight into the transcriptomes of carcinogenic liver flukes. For C. sinensis and O. viverrini, 50,000 unique sequences were identified. Comparative analyses revealed that most sequences (> 85%) produced for each species were new, thus expanding current databases (Cho et al., 2006, Laha et al., 2007, Cho et al., 2008, Ju et al., 2009) and providing a resource to research groups investigating molecular

Acknowledgements

Current research was supported by the Australian Research Council (RBG) and an Endeavour Fellowship (NDY).

References (155)

  • F. Hu et al.

    Clonorchis sinensis: expression, characterization, immunolocalization and serological reactivity of one excretory/secretory antigen-LPAP homologue

    Exp Parasitol

    (2007)
  • S. Kaewkes

    Taxonomy and biology of liver flukes

    Acta Trop

    (2003)
  • B.H. Kalinna et al.

    Manipulating the manipulators: advances in parasitic helminth transgenesis and RNAi

    Trends Parasitol

    (2007)
  • J.M. Kang et al.

    A family of cathepsin F cysteine proteases of Clonorchis sinensis is the major secreted proteins that are expressed in the intestine of the parasite

    Mol Biochem Parasitol

    (2010)
  • S. Kang et al.

    Molecular identification and phylogenetic analysis of nuclear rDNA sequences among three opisthorchid liver fluke species (Opisthorchiidae: Trematoda)

    Parasitol Int

    (2008)
  • S.Y. Kang et al.

    Clonorchis sinensis: molecular cloning and characterization of 28-kDa glutathione S-transferase

    Exp Parasitol

    (2001)
  • J. Keiser et al.

    Effect of artesunate and artemether against Clonorchis sinensis and Opisthorchis viverrini in rodent models

    Int J Antimicrob Agents

    (2006)
  • A. Krogh et al.

    Predicting transmembrane protein topology with a hidden Markov model: application to complete genomes

    J Mol Biol

    (2001)
  • S. Kurathong et al.

    Opisthorchis viverrini infection in rural and urban communities in northeast Thailand

    Trans Roy Soc Trop Med Hyg

    (1987)
  • T. Laha et al.

    Asparaginyl endopeptidase from the carcinogenic liver fluke, Opisthorchis viverrini, and its potential for serodiagnosis

    Int J Infect Dis

    (2008)
  • N. Laoprom et al.

    Microsatellite loci in the carcinogenic liver fluke, Opisthorchis viverrini and their application as population genetic markers

    Infect Genet Evol

    (2010)
  • T.H. Le et al.

    Clonorchis sinensis and Opisthorchis viverrini: development of a mitochondrial-based multiplex PCR for their identification and discrimination

    Exp Parasitol

    (2006)
  • J.Y. Lee et al.

    Hemolytic activity and developmental expression of pore-forming peptide, clonorin

    Biochem Biophys Res Commun

    (2002)
  • J.Y. Lee et al.

    Use of a recombinant Clonorchis sinensis pore-forming peptide, clonorin, for serological diagnosis of clonorchiasis

    Parasitol Int

    (2003)
  • I. Letunic et al.

    iPath: interactive exploration of biochemical pathways and networks

    Trends Biochem Sci

    (2008)
  • Z.R. Lun et al.

    Clonorchiasis: a key foodborne zoonosis in China

    Lancet Infect Dis

    (2005)
  • L. McGonigle et al.

    The silencing of cysteine proteases in Fasciola hepatica newly excysted juveniles using RNA interference reduces gut penetration

    Int J Parasitol

    (2008)
  • M.E. Morales et al.

    RNA interference of Schistosoma mansoni cathepsin D, the apical enzyme of the hemoglobin proteolysis cascade

    Mol Biochem Parasitol

    (2008)
  • B.K. Na et al.

    CsCF-6, a novel cathepsin F-like cysteine protease for nutrient uptake of Clonorchis sinensis

    Int J Parasitol

    (2008)
  • D. Ndegwa et al.

    Protocols for gene silencing in schistosomes

    Exp Parasitol

    (2007)
  • K. Ando et al.

    Nucleotide sequence of mitochondrial CO I and ribosomal ITS II genes of Opisthorchis viverrini in northeast Thailand

    Southeast Asian J Trop Med Public Health

    (2001)
  • Y.A. Bae et al.

    Evolutionary course of CsRn1 long-terminal-repeat retrotransposon and its heterogeneous integrations into the genome of the liver fluke, Clonorchis sinensis

    Korean J Parasitol

    (2003)
  • Y.A. Bae et al.

    CsRn1, a novel active retrotransposon in a parasitic trematode, Clonorchis sinensis, discloses a new phylogenetic clade of Ty3/gypsy-like LTR retrotransposons

    Mol Biol Evol

    (2001)
  • J.D. Bendtsen et al.

    Improved prediction of signal peptides: SignalP 3.0

    J Mol Biol

    (2004)
  • M. Berriman et al.

    The genome of the blood fluke Schistosoma mansoni

    Nature

    (2009)
  • M. Cancela et al.

    Survey of transcripts expressed by the invasive juvenile stage of the liver fluke Fasciola hepatica

    BMC Genomics

    (2010)
  • P.Y. Cho et al.

    Gene expression profile of Clonorchis sinensis metacercariae

    Parasitol Res

    (2008)
  • P.Y. Cho et al.

    Molecular cloning and characterization of WD40-repeat protein from Clonorchis sinensis

    Parasitol Res

    (2007)
  • P.Y. Cho et al.

    Expressed sequence tag analysis of adult Clonorchis sinensis, the Chinese liver fluke

    Parasitol Res

    (2006)
  • B.I. Choi et al.

    Clonorchiasis and cholangiocarcinoma: etiologic relationship and imaging diagnosis

    Clin Microbiol Rev

    (2004)
  • Y.B. Chung et al.

    Molecular cloning and immunolocalization of the 17 kDa myoglobin of Clonorchis sinensis

    Parasitol Res

    (2003)
  • V. Eursitthichai et al.

    Molecular cloning and characterization of a glutathione S-transferase encoding gene from Opisthorchis viverrini

    Asian Pac J Allergy Immunol

    (2004)
  • P.G. Fallon et al.

    Drug-resistant schistosomiasis: resistance to praziquantel and oxamniquine induced in Schistosoma mansoni in mice is drug specific

    Am J Trop Med Hyg

    (1994)
  • A.M. Feist et al.

    Reconstruction of biochemical networks in microorganisms

    Nat Rev Microbiol

    (2009)
  • D.J. Flavell et al.

    Promotion of N-nitrosodimethylamine-initiated bile duct carcinogenesis in the hamster by the human liver fluke, Opisthorchis viverrini

    Carcinogenesis

    (1983)
  • A. Giubellino et al.

    Grb2 signaling in cell motility and cancer

    Expert Opin Ther Targets

    (2008)
  • Z.G. Han et al.

    Schistosoma genomics: new perspectives on schistosome biology and host–parasite interaction

    Annu Rev Genomics Hum Genet

    (2009)
  • T. Harinasuta et al.

    Opisthorchis viverrini infection: pathogenesis and clinical features

    Arzneimittelforschung

    (1984)
  • M.R. Haswell-Elkins et al.

    Cross-sectional study of Opisthorchis viverrini infection and cholangiocarcinoma in communities within a high-risk area in northeast Thailand

    Int J Cancer

    (1994)
  • S.J. Hong

    Clonorchis sinensis tropomyosin: cloning and sequence of partial cDNA amplified by PCR

    Korean J Parasitol

    (1993)
  • Cited by (24)

    • Genomic and transcriptomic analyses of Clonorchis sinensis infection

      2023, Molecular Medical Microbiology, Third Edition
    • Harnessing model organism genomics to underpin the machine learning-based prediction of essential genes in eukaryotes – Biotechnological implications

      2022, Biotechnology Advances
      Citation Excerpt :

      Such a focus is particularly pertinent, given the major problems associated with the treatment, prevention and control of socioeconomically parasitic worms and arthropods, including the inefficacy or toxicity of some anti-parasitic compounds, and the emergence and spread of drug resistance in some parasite species (Sangster et al., 2018). Genomic, transcriptomic and proteomic data sets are becoming increasingly available (Goodwin et al., 2016), allowing the prediction and prioritisation of potential molecular targets for novel interventions (e.g., Young et al., 2010; Anstead et al., 2016; Emery et al., 2016; Korhonen et al., 2016a, 2016b; Ma et al., 2018; Stroehlein et al., 2018; Wang et al., 2018). The appraisal of current literature (Sections 1–4) reveals that significant efforts have been made to experimentally characterise essential genes in select organisms, supported by genome sequencing and functional genomics.

    • A comprehensive review of omics and host-parasite interplays studies, towards control of Opisthorchis viverrini infection for prevention of cholangiocarcinoma

      2019, Acta Tropica
      Citation Excerpt :

      Transcriptomic studies of adult O. felineus parasites has found at least four highly encoded proteins; myoglobin (15.1%), vitelline precursor protein (8.6%), cysteine proteases (6.8%), and glutathione S-transferase (4.6%). The transcriptomes of O. viverrini and Clonorchis sinensis have also been compared and reviewed by Young et al. (Young et al., 2010b). Analysis of predicted protein-encoding genes from a draft genome of O. viverrini from recent studies have also given credence to the fact that the trematode has a genome that is very divergent from other flukes, including C. sinensis, Schistosoma haematobium, S. japonicum, and S. mansoni (Young et al., 2014; Gasser et al., 2017).

    • Genomics of worms, with an emphasis on Opisthorchis viverrini — opportunities for fundamental discovery and biomedical outcomes

      2017, Parasitology International
      Citation Excerpt :

      Moreover, exploring selected groups of molecules, such as kinases, phosphatases, GPCRs and the complex array of peptidases, and understanding their roles in host–parasite interactions might also support applied research focused on combatting this fluke and the insidious disease complex that it causes. In conclusion, we hope that the genomic/transcriptomic resources established for O. viverrini [19–21] will be useful to scientists around the world working on the fluke, opisthorchiasis and/or CCA and also to those working on translational research focused on biomedical and biotechnological outcomes to prevent and control this insidious disease problem. We concede that there are some challenges in relation to skills required for the handling and bioinformatic analyses of digital data sets, and also regarding research funding for this area.

    View all citing articles on Scopus
    View full text