Skip to main content

Leeches as a source of mammalian viral DNA and RNA—a study in medicinal leeches

Abstract

Surveillance of wild vertebrates can be challenging in remote and inaccessible areas such as tropical rainforests. Blood-feeding parasites, such as leeches, can facilitate wild vertebrate monitoring by targeting residual DNA from the animals the leeches feed on. Successes in detecting host DNA from leeches suggest that host viruses may also be detectable. To systematically test this hypothesis, we performed a proof of concept study using quantitative PCR (qPCR) to detect DNA viruses (bovine herpesvirus [BHV], human adenovirus [HAdV]) and RNA viruses (influenza A [InfA] and measles morbillivirus [MeV]) from nucleic acids extracted from medicinal leeches fed with blood spiked with each virus. All viruses except BHV showed a gradual decline in concentration from day 1 to 50, and all except BHV were detectable in at least half of the samples even after 50 days. BHV exhibited a rapid decline at day 27 and was undetectable at day 50. Our findings in medicinal leeches indicate that leeches collected in the wild might be an untapped resource for detecting vertebrate viruses and could provide new opportunities to study wildlife viral diseases of rare species in challenging environments, where capturing and handling of animals is difficult.

This is a preview of subscription content, access via your institution.

Fig. 1
Fig. 2

References

  1. Al-Khleif A, Roth M, Menge C, Heuser J, Baljer G, Herbst W (2011) Tenacity of mammalian viruses in the gut of leeches fed with porcine blood. J Med Microbiol 60:787–792

    Article  PubMed  Google Scholar 

  2. Arez AP, Lopes D, Pinto J, Franco AS, Snounou G, do Rosário VE (2000) Plasmodium sp.: optimal protocols for PCR detection of low parasite numbers from mosquito (Anopheles sp.) samples. Exp Parasitol 94:269–272

    CAS  Article  PubMed  Google Scholar 

  3. Barnes MA, Turner CR (2016) The ecology of environmental DNA and implications for conservation genetics. Conserv Genet 17:1–17

    CAS  Article  Google Scholar 

  4. Borda E, Oceguera-Figueroa A, Siddall ME (2008) On the classification, evolution and biogeography of terrestrial haemadipsoid leeches (Hirudinida: Arhynchobdellida: Hirudiniformes). Mol Phylogenet Evol 46(1):142–154

    Article  PubMed  Google Scholar 

  5. Cleveland WS, Grosse E, Shyu WM (1992) Local regression models. Chapter 8 of Statistical Models in S eds J.M. Chambers and T.J. Hastie, Wadsworth & Brooks/Cole

  6. Elia G, Decaro N, Martella V, Cirone F, Lucente MS, Lorusso E, Trani LD, Buonavoglia C (2006) Detection of canine distemper virus in dogs by real-time RT-PCR. J Virol Methods 136:171–176

    CAS  Article  PubMed  Google Scholar 

  7. EU Environment (2010) Directive 2010/63/EU of the European Parliament and of the council. Official Journal of the European Union, 276/33. (http://eur-lex.europa.eu/legal-content/EN/TXT/?uri=CELEX:32010L0063)

  8. Furuse Y, Suzuki A, Oshitani H (2010) Origin of measles virus: divergence from rinderpest virus between the 11th and 12th centuries. Virol J 7:52

    Article  PubMed  PubMed Central  Google Scholar 

  9. Heard MJ, Smith KF, Ripp KJ, Berger M, Chen J, Dittmeier J, Goter M, McGarvey ST, Ryan E (2013) The threat of disease increases as species move toward extinction. Conserv Biol 27:1378–1388

    Article  PubMed  PubMed Central  Google Scholar 

  10. Hoffmann M, Hilton-Taylor C, Angulo A, Böhm M, Brooks TM, Butchart SHM, Carpenter KE, Chanson J, Collen B, Cox NA, Darwall WRT et al (2010) The impact of conservation on the status of the world’s vertebrates. Science 330:1503–1509

    CAS  Article  PubMed  Google Scholar 

  11. Jensen RH, Mollerup S, Mourier T, Hansen TA, Fridholm H, Nielsen LP, Willerslev E, Hansen AJ, Vinner L (2015) Target-dependent enrichment of virions determines the reduction of high-throughput sequencing in virus discovery. PLoS One 10(4):e0122636

    Article  PubMed  PubMed Central  Google Scholar 

  12. Kutschera U (2003) The feeding strategies of the leech Erpobdella octoculata (L.): a laboratory study. Int Rev Hydrobiol 88(1):94–101

    Article  Google Scholar 

  13. Malele II, Ouma JO, Enyaru JCK, Matovu E, Alibu V, Auma JE, Onyoyo SG, Bateta R, Changasi RE, Mukiria PM, Ndung’u K, Gitonga PK, Mwaniki LM, Nyingilili HS, Lyaruu EA, Kapange LA, Kamau PK, Masiga DK (2013) Comparative diagnostic and analytical performance of PCR and LAMP-based trypanosome detection methods estimated using pooled whole tsetse flies and midguts. Vet Parasitol 197:549–556

    CAS  Article  PubMed  Google Scholar 

  14. Masiga DK, Smyth AJ, Hayes P, Bromidge TJ, Gibson WC (1992) Sensitive detection of trypanosomes in tsetse flies by DNA amplification. Int J Parasitol 22:909–918

    CAS  Article  PubMed  Google Scholar 

  15. McNamara JJ, Laveissiére C, Masiga DK (1995) Multiple trypanosome infections in wild tsetse in Côte d’Ivoire detected by PCR analysis and DNA probes. Acta Trop 59:85–92

    CAS  Article  PubMed  Google Scholar 

  16. Moreno M, Cano J, Nzambo S, Bobuakasi L, Buatiche JN, Ondo M, Micha F, Benito A (2004) Malaria panel assay versus PCR: detection of naturally infected Anopheles melas in a coastal village of Equatorial Guinea. Malar J 3(3):20

    Article  PubMed  PubMed Central  Google Scholar 

  17. Muturi CN, Ouma JO, Malele II, Ngure RM, Rutto JJ, Mithöfer KM, Enyaru J, Masiga DK (2011) Tracking the feeding patterns of tsetse flies (Glossina genus) by analysis of bloodmeals using mitochondrial cytochromes genes. PLoS One 6(2):e17284

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  18. Nehili M, Ilk C, Mehlhorn H, Ruhnau K, Dick W, Njayou M (1994) Experiments on the possible role of leeches as vectors of animal and human pathogens: a light and electron microscopy study. Parasitol Res 80:277–290

    CAS  Article  PubMed  Google Scholar 

  19. Nolan T, Hands RE, Bustin SA (2006) Quantification of mRNA using real-time RT-PCR. Nat Protoc 1(3):1559–1582

    CAS  Article  PubMed  Google Scholar 

  20. Pina S, Puig M, Lucena F, Jofre J, Girones R (1998) Viral pollution in the environment and in shellfish: human adenovirus detection by PCR as an index of human viruses. Appl Environ Microbiol 64(9):3376–3382

    CAS  PubMed  PubMed Central  Google Scholar 

  21. Pirtle EC, Beran GW (1991) Virus survival in the environment. Scientific and Technical Review of the Office International des Epizooties 10(3):733–748

    CAS  Article  Google Scholar 

  22. R Core Team (2015) R: a language and environment for statistical computing. R Foundation for Statistical Computing, Vienna, Austria. URL http://www.R-project.org/

  23. Regnaut S, Lucas FS, Fumagalli L (2006) DNA degradation in avian faecal samples and feasibility of non-invasive genetic studies of threatened capercaillie populations. Conserv Genet 7(3):449–453

    CAS  Article  Google Scholar 

  24. Sawyer RT (1986) Leech biology and behaviour. Volume II Feeding, biology, ecology and systematics. Clarendon Press, Oxford

  25. Schloss L, Falk KI, Skoog E, Brytting M, Linde A, Aurelius E (2009) Monitoring of herpes simplex virus DNA types 1 and 2 viral load in cerebrospinal fluid by real-time PCR in patients with herpes simplex encephalitis. J Med Virol 81:1432–1437

    CAS  Article  PubMed  Google Scholar 

  26. Schnell IB, Thomsen PF, Wilkinson N, Rasmussen M, Jensen LRD, Willerslev E, Bertelsen MF, Gilbert MTP (2012) Screening mammal biodiversity using DNA from leeches. Curr Biol 22(8):R262–R263

    CAS  Article  PubMed  Google Scholar 

  27. Schnell IB, Sollmann R, Calvignac-Spencer S, Siddall ME, Yu DW, Wilting A, Gilbert MTP (2015) iDNA from terrestrial haemophagous leeches as a wildlife surveying and monitoring tool—prospects, pitfalls and avenues to be developed. Frontiers of Zoology 12(1):1

    Article  Google Scholar 

  28. Schunck B, Kraft W, Truyen U (1995) A simple touch-down polymerase chain reaction for the detection of canine parvovirus and feline panleukopenia virus in feces. J Virol Methods 55(3):427–433

    CAS  Article  PubMed  Google Scholar 

  29. Shope RD (1957) The leech as a potential virus reservoir. J Exp Med 105(4):373–382

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  30. Steinhauer DA, Holland JJ (1987) Rapid evolution of RNA viruses. Ann Rev Microbiology 41:409–433

    CAS  Article  Google Scholar 

  31. Taberlet P, Griffin S, Goossens B, Questiau S, Manceau V, Escaravage N, Waits LP, Bouvet J (1996) Reliable genotyping of samples with very low DNA quantities using PCR. Nucleic Acids Res 24(16):3189–3194

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  32. Tischer BK, Osterrieder N (2010) Herpesviruses—a zoonotic threat? Veterinary Micobiology 140:266

    Article  Google Scholar 

  33. To KK, Hung IF, Li IW, Lee KL, Koo CK, Yan WW, Liu R, Ho KY, Chu KH, Watt CL, Luk WK, Lai KY, Chow FL, Mok T, Buckley T, Chan JF, Wong SS, Zheng B, Chen H, Lau CC, Tse H, Cheng VC, Chan KH, Yuen KY, Pandemic H1N1 Study Group (2010) Delayed clearance of viral load and marked cytokine activation in severe cases of pandemic H1N1 2009 influenza virus infection. Clin Infect Dis 50:850–859

    Article  PubMed  Google Scholar 

  34. Verheyen J, Timmen-Wego M, Laudien R, Boussaad I, Sen S, Koc A, Uesbeck A, Mazou F, Pfister H (2009) Detection of adenoviruses and rotaviruses in drinking water sources used in rural areas of Benin, West Africa. Applied Environmental Microbiology 75(9):2798–2801

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  35. Wang J, O’Keefe J, Orr D, Loth L, Banks M, Wakeley P, West D, Card R, Ibata G, Van Maanenc K, Thoren P, Isaksson M, Kerkhofs P (2007) Validation of a real-time PCR assay for the detection of bovine herpesvirus 1 in bovine semen. J Virol Methods 144:103–108

    CAS  Article  PubMed  Google Scholar 

  36. Webster RG, Bean WJ, Gorman OT, Chambers TM, Kawaoka Y (1992) Evolution and ecology of influenza A viruses. Microbiology Reviews 56(1):152–179

    CAS  Google Scholar 

  37. WHO (2009) CDC protocol of realtime RTPCR for swine influenza A(H1N1)

  38. Zerbst-Boroffka I (1973) Osmo- und Volumenregulation bei Hirudo medicinalis nach Nahrungsaufnahme. J Comp Physiol 84:185–204

    Article  Google Scholar 

Download references

Acknowledgements

The authors would like to thank the German Federal Ministry of Education and Research (BMBF FKZ: 01LN1301A) for funding this research. Further thanks are due to Azza Abdelgawad and Karin Hönig (Leibniz Institute for Zoo and Wildlife Research) as well as Bente Andersen (Statens Serum Institut), for help with cultivation of viruses. Stefanie Palczewski (Leibniz Institute for Zoo and Wildlife Research) is also thanked for helping with leech extractions, and Jürgen Niedballa (Leibniz Institute for Zoo and Wildlife Research) has been very helpful with assistance with R.

Author information

Affiliations

Authors

Corresponding author

Correspondence to Andreas Wilting.

Electronic supplementary material

ESM 1

(PDF 37 kb)

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Kampmann, ML., Schnell, I.B., Jensen, R.H. et al. Leeches as a source of mammalian viral DNA and RNA—a study in medicinal leeches. Eur J Wildl Res 63, 36 (2017). https://doi.org/10.1007/s10344-017-1093-6

Download citation

Keywords

  • Medicinal leeches
  • Virus
  • DNA
  • RNA
  • Wildlife monitoring
  • Hirudidae
  • Zoonosis