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Title: Caenorhabditis elegans as a model to screen plant extracts and compounds as natural anthelmintics for veterinary use

Author
item KATIKI, LUCIANA - Sao Paulo State University (UNESP)
item Ferreira, Jorge
item ZAJAC, ANNE - Virginia-Maryland Regional College Of Veterinary Medicine (VMRCVM)
item Masler, Carol
item LINDSAY, DAVID - Virginia-Maryland Regional College Of Veterinary Medicine (VMRCVM)
item CHAGAS, ANA CAROLINA - Embrapa
item AMARANTE, ALESSANDRO - Sao Paulo State University (UNESP)

Submitted to: Veterinary Parasitology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/16/2011
Publication Date: 6/13/2011
Citation: Katiki, L., Ferreira, J.F., Zajac, A., Masler, C.A., Lindsay, D., Chagas, A., Amarante, A. 2011. Caenorhabditis elegans as a model to screen plant extracts and compounds as natural anthelmintics for veterinary use. Veterinary Parasitology. 182:264-268.

Interpretive Summary: The most challenging obstacles to testing plant products as vermifuges are: 1) establishing a suitable assay from which results indicate their potential use against a parasitic worm of interest, and 2) preparing plant extracts in a way that, once freeze-dried, they can be redissolved in solvents that are miscible in the test medium. The use of parasitic worms as a screening system is hindered by the difficulty of keeping them alive for long periods outside their host and by the need to keep infected animals as sources of eggs or adults as needed. We developed a method that uses a non-parasitic soil nematode as a model to screen plant extracts for their potential anthelmintic effect against small ruminant gastrointestinal nematodes, including Haemonchus contortus. This modified method uses only liquid sterile medium, instead of agar plates inoculated with bacteria, and two selective sieves to obtain adult nematodes. Enough adults could be produced to screen at least six plant extracts within a week, and extracts could be tested at five concentrations each, every 48 hours. The high availability, ease of culture, and rapid proliferation of this soil free-living nematode make it a useful screening system to test plant extracts and other plant products to investigate their potential anthelmintic activity against parasitic nematodes.

Technical Abstract: The most challenging obstacles to testing plant products for their anthelmintic activity are: 1) establishing a suitable nematode in vitro assay from which results can be indicative of potential use against a parasitic nematode of interest, and 2) preparing the extracts in a way that, once lyophilized, they can be redissolved in solvents that are miscible in the test medium, and at concentrations well tolerated by the nematode system used for screening. The use of parasitic nematodes as a screening system is hindered by the difficulty of keeping them alive for long periods outside their host and by the need to keep infected animals as sources of eggs or adults when needed. This method uses the free-living soil nematode Caenorhabditis elegans as a system to screen plant extracts for their potential anthelmintic effect against small ruminant gastrointestinal nematodes, including Haemonchus contortus. This modified method uses only liquid axenic medium, instead of agar plates inoculated with E. coli, and two selective sieves to obtain adult nematodes. During screening, the use of a balanced salt solution (M-9), instead of distilled water, resulted in 95-100% motile adults and improved results obtained with plant extracts. Enough adults could be produced to screen at least six plant extracts within a week. Extracts could be tested at five concentrations each, every 48 hours. Plant extracts prepared in water, ethanol:water (70:30) and acetone:water (70:30) were easily redissolved in the M-9 medium with 1% DMSO. While adult worms tolerated DMSO, ethanol, methanol, and Tween 80 at 1 and 2%, Labrasol (a bioenhancer with low toxicity to mammals) and Tween 20 were very toxic to C. elegans even at 1%. The high availability, ease of culture, and rapid proliferation of C. elegans make it a useful screening system to test plant extracts and other plant products to investigate their potential anthelmintic activity against parasitic nematodes.