Veterinary Parasitology 190 (2012) 461–466 Contents lists available at SciVerse ScienceDirect Veterinary Parasitology jo u rn al hom epa ge : www.elsevier.com/locate/vetpar Real-time PCR evaluation of Strongylus vulgaris in horses on farms in Denmark and Central Kentucky M.K. Nielsen a,∗ , S.N. Olsen b , E.T. Lyons a , J. Monrad c , S.M. Thamsborg c a M.H. Gluck Equine Research Center, Department of Veterinary Science, University of Kentucky, Lexington, KY, USA b Department of Large Animal Sciences, Faculty of Health and Medical Sciences, University of Copenhagen, Denmark c Danish Center for Experimental Parasitology, Department of Veterinary Disease Biology, Faculty of Health and Medical Sciences, University of Copenhagen, Denmark a r t i c l e i n f o Article history: Received 11 June 2012 Received in revised form 12 July 2012 Accepted 13 July 2012 Keywords: Strongylus vulgaris Real-time PCR Prevalence Larval culture Treatment intensity a b s t r a c t Strongyle parasites are ubiquitous in grazing horses, and the large strongyle Strongylus vul- garis is considered the most pathogenic helminth parasite of horses. Recent investigations have suggested an association between occurrence of this parasite and usage of selective therapy based on regular fecal egg counts. The established diagnostic method for S. vulgaris involves larval culture and subsequent morphological identification of third stage larvae under the microscope. Recently, a real-time PCR assay was developed and validated for the detection and semi-quantification of S. vulgaris eggs in equine fecal samples. The purposes of the present study were (a) to determine the presence of S. vulgaris by real-time PCR in Danish and American horses on farms using vastly different anthelmintic treatment regi- mens and (b) to evaluate the association between larval culture results and the PCR. A total of 991 horses representing 53 different horse farms in Denmark and Central Kentucky were studied. Fresh fecal samples were collected from all horses, and strongyle eggs retrieved for DNA extraction and subsequent real-time PCR analysis. Individual larval cultures were performed on the Danish part of the data set (663 horses on 42 farms). On the Danish farms, the S. vulgaris PCR prevalence was found to be 9.2% on farms not basing parasite control on fecal egg counts, and 14.1% on farms using selective therapy. No horses were PCR positive in the American part of the study (328 horses on 11 farms). Kappa-values indi- cated a moderate agreement between PCR and larval culture results, while McNemar tests revealed no statistical difference between the paired proportions. Significant associations were found between PCR cycle of threshold (Ct) value groups and larval culture counts. Results indicate that both diagnostic methods can be useful for determining the occur- rence of S. vulgaris on horse farms, but that they both are affected by potential sources of error. The PCR results confirmed previous findings suggesting that S. vulgaris can reemerge under selective therapy regimens. © 2012 Elsevier B.V. All rights reserved. 1. Introduction Strongyle type parasites are ubiquitous in grazing horses, and species infecting the horse comprise more than ∗ Corresponding author. Tel.: +1 859 218 1103; fax: +1 859 257 8542. E-mail address: martin.nielsen@uky.edu (M.K. Nielsen). 50 in total (Lichtenfels et al., 2008). Of these, the large strongyle, Strongylus vulgaris, has always been considered the most pathogenic of all helminth parasites infecting the horse (Drudge, 1979). Unlike other strongyle parasites, the third stage larvae (L 3 ) of S. vulgaris invade the intestinal mucosa and make their way to the intestinal blood vessels, where they migrate subendothelially to their predilection site in the Cranial Mesenteric Artery at the root of the 0304-4017/$ – see front matter © 2012 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.vetpar.2012.07.018