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Silencing of essential genes by RNA interference in Haemonchus contortus

Published online by Cambridge University Press:  20 February 2012

J. L. ZAWADZKI ,
A. C. KOTZE ,
J.-A. FRITZ ,
N. M. JOHNSON ,
J. E. HEMSWORTH ,
B. M. HINES  and
C. A. BEHM
J. L. ZAWADZKI
Affiliation:
Biosciences Research Division, Department of Primary Industries Victoria, 475 Mickleham Road, Attwood, Victoria, 3049, Australia
A. C. KOTZE
Affiliation:
CSIRO Livestock Industries, 306 Carmody Rd. St Lucia, QLD 4067, Australia
J.-A. FRITZ
Affiliation:
Division of Biomedical Science and Biochemistry, Research School of Biology, The Australian National University, Canberra, A.C.T. 0200, Australia
N. M. JOHNSON
Affiliation:
Division of Biomedical Science and Biochemistry, Research School of Biology, The Australian National University, Canberra, A.C.T. 0200, Australia
J. E. HEMSWORTH
Affiliation:
Biosciences Research Division, Department of Primary Industries Victoria, 475 Mickleham Road, Attwood, Victoria, 3049, Australia
B. M. HINES
Affiliation:
CSIRO Livestock Industries, 306 Carmody Rd. St Lucia, QLD 4067, Australia
C. A. BEHM*
Affiliation:
Division of Biomedical Science and Biochemistry, Research School of Biology, The Australian National University, Canberra, A.C.T. 0200, Australia
*
*Corresponding author: Division of Biomedical Science and Biochemistry, Research School of Biology, The Australian National University, Canberra, A.C.T. 0200, Australia. Tel: +61 2 61252203. Fax: +61 2 61250313. E-mail: [email protected]
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Summary

In this study we assessed three technologies for silencing gene expression by RNA interference (RNAi) in the sheep parasitic nematode Haemonchus contortus. We chose as targets five genes that are essential in Caenorhabditis elegans (mitr-1, pat-12, vha-19, glf-1 and noah-1), orthologues of which are present and expressed in H. contortus, plus four genes previously tested by RNAi in H. contortus (ubiquitin, tubulin, paramyosin, tropomyosin). To introduce double-stranded RNA (dsRNA) into the nematodes we tested (1) feeding free-living stages of H. contortus with Escherichia coli that express dsRNA targetting the test genes; (2) electroporation of dsRNA into H. contortus eggs or larvae; and (3) soaking adult H. contortus in dsRNA. For each gene tested we observed reduced levels of mRNA in the treated nematodes, except for some electroporation conditions. We did not observe any phenotypic changes in the worms in the electroporation or dsRNA soaking experiments. The feeding method, however, elicited observable changes in the development and viability of larvae for five of the eight genes tested, including the ‘essential’ genes, Hc-pat-12, Hc-vha-19 and Hc-glf-1. We recommend the E. coli feeding method for RNAi in H. contortus and provide recommendations for future research directions for RNAi in this species.

Keywords

Haemonchus contortusRNA interferenceRNAidsRNAelectroporationCaenorhabditis elegansgene knockdown
Type
Research Article
Information
Parasitology , Volume 139 , Special Issue 5: Genetic manipulation of parasitic helminths to define gene function , April 2012 , pp. 613 - 629
Copyright
Copyright © Cambridge University Press 2012

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