These authors have contributed equally to this work.
Genetic diversity in the Maremmano horse and its relationship with other European horse breeds
Article first published online: 10 NOV 2010
© 2010 The Authors, Journal compilation © 2010 Stichting International Foundation for Animal Genetics
Special Issue: Horse Genomics and the Dorothy Russell Havemeyer Foundation
Volume 41, Issue Supplement s2, pages 53–55, December 2010
How to Cite
Felicetti, M., Lopes, M. S., Verini-Supplizi, A., Da Câmara Machado, A., Silvestrelli, M., Mendonça, D. and Distl, O. (2010), Genetic diversity in the Maremmano horse and its relationship with other European horse breeds. Animal Genetics, 41: 53–55. doi: 10.1111/j.1365-2052.2010.02102.x
- Issue published online: 10 NOV 2010
- Article first published online: 10 NOV 2010
- Accepted for publication 20 September 2010
- clustering analysis;
- Equus caballus;
The Maremmano is an Italian warmblood horse breed from central Italy. We characterized the genetic diversity and the degree of admixture in Maremmano in comparison to 14 other European horse breeds using 30 microsatellites. Between-breed diversity explained about 9 per cent of the total genetic diversity. Cluster analysis, genetic distances and genetic differentiation coefficients showed a close relationship of Maremmano with Hanoverian and Lusitano in accordance with breed history.
The Maremmano is an Italian warmblood horse mostly bred in the provinces of Grosseto and Viterbo (Central Italy). It is believed that the origin of this horse breed goes back to local Etruscan horse populations which were crossbred with modern breeds in the last centuries. Pedigree analysis of the Maremmano horses revealed four male lines contributing 11.1% to the genetic diversity of all Maremmano stallions (Silvestrelli 1991). In 1980, Maremmano breeders established a stud book for Maremmano.
The objective of this study was to investigate the genetic structure and the degree of admixture of Maremmano together with the Italian nucleus of Lipizzan and Lusitano as well as twelve further horse breeds previously characterized by Aberle et al. (2004). This data set consisted of Hanoverian, Arabian, Exmoor, Icelandic, Przewalski, Sorraia and all German coldblood horse breeds. We used the same marker set of 30 autosomal microsatellites and the same PCR conditions and reference samples as described elsewhere (Aberle et al. 2004).
A total of 146 animals that were not closely related were genotyped in this study: Maremmano (n = 50), the Italian nucleus of Lipizzan (n = 49) and Lusitano (n = 47) from Portugal. DNA was extracted from EDTA-blood using standard methods.
Genetic variability of the breeds genotyped in this study was determined by the mean number of alleles, observed and expected heterozygosities, breed-specific alleles and the molecular variance (amova) using GENALEX 6 (Peakall & Smouse 2006). Excess and deficiency of heterozygotes, which are deviations from Hardy-Weinberg equilibrium, were estimated using GENEPOP (Raymond & Rousset 1995). Molecular genetic relationships among populations were estimated using Wright’s FST and Nei’s standard genetic distance (GST; Nei 1973) by bootstrapping 1000 replicates using MICROSAT (Minch 1997). Phenograms based on Nei’s GST and genetic distances among all 549 animals were drawn using the unweighted pair group method with arithmetic mean algorithm (UPGMA) by PHYLIP (Felsenstein 1989) and displayed by TREEVIEW (Page 1996). The Bayesian clustering procedure of STRUCTURE was employed to investigate the genetic structure and the degree of admixture of the 15 horse breeds (Pritchard et al. 2000). A 20 000 initial burn-in was used, followed by 100 000 MCMC iterations as recommended by Falush et al. (2007) with 10 independent replicates each. All runs used an admixture model with correlated frequencies and the parameter of individual admixture alpha.
The mean number of alleles was 4.7 for the Lipizzan, 6.7 for the Lusitano and 7.3 for the Maremmano. For Maremmano and Lipizzan breeds the total observed heterozygosity was higher than the expected, whereas for Lusitano breed 19 out of the 30 loci showed observed heterozygosity values lower than the expected ones (Table S1).
The amova indicated that for Maremmano, Lipizzan and Lusitano 9% of the total genetic variability is attributed to significant differences between the horse breeds, whereas 91% of the observed variation was from within the breeds. The amova performed for the five riding horse breeds (Maremmano, Lipizzan, Lusitano, Arabian, Hanoverian), considered separately from the others, showed 11% of the genetic variability due to breed differences. When the German coldblood breeds were included, the variation among breeds increased only slightly to 12%; with all 15 breeds the total genetic variability rose to 14.6%.
Maremmano showed the lowest genetic differentiation with the Hanoverian (5.5%); in addition, the Lipizzan and Lusitano, when matched with the Maremmano, showed the lowest FST values (11.4% and 6%, respectively). Among the three breeds genotyped here, the Maremmano showed the least differentiation when compared to the coldblood breeds (Table S2). Similar results were obtained when the genetic differentiation based on Nei’s GST among breed pairs was used. The only exception was for the Maremmano, which showed lowest genetic differentiation (14.7%) with the Lusitano.
The phylogenetic tree based on Nei’s GST (Fig. 1) and the dendrogram based on the proportion of shared alleles (Fig. S1) displayed three main clusters representing riding horses, the Exmoor, Przewalski and Sorraia group and the German draught horses. The Icelandic horses did not cluster with any of the other breeds.
Clustering using STRUCTURE separated for K = 3 horses into riding horse breeds, ancient and isolated breeds, and German draught horses. For K = 9, Maremmano clustered together with Lusitano, but Hanoverian and Arabian were separate clusters. When K = 14, Maremmano and Lusitano also clustered in their own pre-defined populations (Fig. S2; Table S3).
From the three breeds genotyped in this study, both the Maremmano and the Lusitano showed a high level of genetic variability and similar to that observed for the same breeds in other studies (Luís et al. 2007; Zuccaro et al. 2008). The high level of genetic differentiation observed for the Maremmano may partly reflect contributions from several breeds, although a significant proportion of the Maremmano is descending from a reduced number of male lines. The Italian nucleus of Lipizzan horses showed low levels of variation similar to those observed by Achmann et al. (2004). The small number of founders resulting in a small effective population size and the traditional purebreeding system within a nucleus without any crossbreeding may explain the reduced variability within the Lipizzan. In agreement with the results of Luís et al. (2007) in the Lusitano, levels of observed heterozygosity were lower than their expected counterpart.
Pairwise FST values among Maremmano, Lipizzan, Lusitano and Hanoverian were in a similar range as those among closely related coldblood breeds (Aberle et al. 2004; Druml et al. 2007). Inclusion of Hanoverian and German coldblood increased the proportion of variance among breeds only slightly, indicating a close relationship of these breeds with Maremmano, Lipizzan and Lusitano in contrast to Arabian, pony and primitive horse breeds. The two genetic differentiation measures and the two model-based clustering approaches also revealed a genetic proximity of Maremmano with the Hanoverian and the Lusitano. Two male lines founded by thoroughbred stallions (Aiace and Ingres) and a Trakehner stallion might have created the relationship between Maremmano and Hanoverian, as thoroughbred and Trakehner stallions have been intensely used in the Hanoverian warmblood (Hamann & Distl 2008). Regarding the close genetic proximity between Maremmano and Lusitano, it is believed that Iberian horses, the ancestors of the Lusitano and the Andalusian, were in the Stato dei Reali Presidi di Spagna (1557–1800), located between Stato Pontificio and Granducato di Toscana, and therefore might have influenced the founder lines of the Maremmano breed.
In conclusion, the Maremmano retained a high genetic diversity and the results reported here can be used to prevent genetic erosion of the Maremmano breed.
Thanks to Dr Luca Buttazzoni for technical support and Mr Gianluca Alunni for expert technical assistance. This work was supported by SELMOL-MIPAF (Silvestrelli) and by FTC and FEDER within the projects POCTI/CVT/41890/2001 and POCI2010 Ref. GG/GGP/ME611. CBA-UAç was supported by FCT and DRCT and Lopes by DRCT M3.1.1/I/017A/2005.
Conflict of interest
The authors have declared no potential conflicts.
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Table S1 Number of alleles, observed and expected heterozygosity for Maremmano, Lusitano and Italian Lipizzan horses (146 horses) based on 30 microsatellite loci.
Table S2 Nei’s standard genetic distance and Wright′s distance among 15 horse breeds.
Table S3 Estimated memberships to inferred clusters obtained by STRUCTURE and individual assignments of 549 horses according to their own predefined breed or to another breed as obtained using GeneClass2.
Figure S1 UPGMA dendrogram constructed from allele-sharing distances among 549 animals from 15 different horse breeds.
Figure S2 Graphical presentation of the population structure analyses for a sample of 549 horses from 15 different horse breeds obtained by STRUCTURE.
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