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Diagnosing genetically diverse avian malarial infections using mixed-sequence analysis and TA-cloning

Published online by Cambridge University Press:  07 March 2005

J. PÉREZ-TRIS  and
S. BENSCH
J. PÉREZ-TRIS
Affiliation:
Department of Animal Ecology, Lund University, Ecology Building, SE-22362 Lund, Sweden
S. BENSCH
Affiliation:
Department of Animal Ecology, Lund University, Ecology Building, SE-22362 Lund, Sweden
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Abstract

Birds harbouring several malarial parasites are common in the wild, and resolving such multiple infections is important for our understanding of host–parasite relationships. We propose a simple and reasonably accurate method for detecting and resolving multiple infections, based on the analysis of parasite cytochrome b DNA sequences: genetically mixed infections are first identified by double nucleotide peaks on sequence electropherograms, and later retrieved by TA-cloning. We applied this method to wild birds, and to experimentally created mixes with varying proportion of two parasites (Plasmodium spp. and Haemoproteus spp.). In general, the method was very efficient in detecting and resolving multiple infections, but some problems were encountered. Several multiple infections were erroneously scored as simple, either because one of the parasite lineages was a better target for the primers used, or because it was much more abundant in the mix. On the other hand, single nucleotide substitutions and template switching during PCR produced artificial sequences in some clones. We discuss the utility of the method, and propose a framework for its use when screening for genetically diverse avian malarial parasites.

Keywords

cytochrome bhost–parasite relationshipswithin-host interactionsmultiple parasite infectionsnested PCR
Type
Research Article
Information
Parasitology , Volume 131 , Issue 1 , July 2005 , pp. 15 - 23
Copyright
© 2005 Cambridge University Press

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