Original Research ARTICLE
Microsatellite analysis of geographically close isolates of Cystoisospora suis
- 1University of Veterinary Medicine Vienna, Austria
Microsatellites are short repetitive DNA sequences of two to six repeats interspersed in the genome that display a rapid mutation rate and consequently show high variation between individuals or populations. They have been used to characterise population diversity and structure and the level of variation between different isolates of a number of different organisms, including apicomplexan protozoa. Currently nothing is known about the genetic variability and population structure of Cystoisospora suis (Apicomplexa: Coccidia), the causative agent of piglet coccidiosis, and we made use of the recently available genome of C. suis (strain Wien-I) to amplify microsatellite regions (ca. 300-550 bp) and evaluate the applicability of fluorescence-labelled primers to investigate amplicon length variation at high resolution using capillary electrophoresis (CE). Two phenotypically characterised isolates (Wien-I, toltrazuril susceptible; Holl-1 toltrazuril resistant) and six field isolates from Europe were compared by conventional PCR followed by agar-gel electrophoresis, Sanger sequencing and CE (fluorescence labelling and fragment length analysis) to evaluate the applicability of the method. Four primer pairs could be identified that amplified bands of the expected size and were labelled for CE analysis. High resolution CE for size determination of PCR amplicons proved to be a reliable and simple method. It revealed high diversity of the analysed strains, with marked differences even between two strains from neighbouring swine farms. In follow-up studies, adaptation of the PCR assay to multiplexing and amplification of small DNA quantities will provide a cost-effective tool to analyse field strains to reveal geographic diversity that could be mapped to phenotypic traits
Keywords: Swine, Coccidia, Population Genetics, capillary electrophoresis, short repetitive elements, Isospora suis
Received: 28 Aug 2018;
Accepted: 08 Mar 2019.
Edited by:Luís P. Gondim, Federal University of Bahia, Brazil
Reviewed by:Jan Slapeta, University of Sydney, Australia
Arwid Daugschies, Leipzig University, Germany
Guo-Hua Liu, Hunan Agricultural University, China
Copyright: © 2019 Joachim, Ruttkowski and Palmieri. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
* Correspondence: Prof. Anja Joachim, University of Veterinary Medicine Vienna, Vienna, Austria, Anja.Joachim@vetmeduni.ac.at