Forensic Science International: Genetics 8 (2014) 126–131

Forensic Population Genetics – Original Research

Northern Slavs from Serbia do not show a founder effect at autosomaland Y-chromosomal STRs and retain their paternal genetic heritage

Krzysztof Rebała a,*, Igor Veselinovic b, Daniela Sivakova c, Erika Patskun d,Sergey Kravchenko e, Zofia Szczerkowska a

a Department of Forensic Medicine, Medical University of Gdansk, Gdansk, Polandb Clinical Centre of Vojvodina, Medical Faculty, University of Novi Sad, Novi Sad, Serbiac Department of Anthropology, Comenius University in Bratislava, Bratislava, Slovakiad Transcarpathian Regional Clinical Hospital, Uzhhorod, Ukrainee Institute of Molecular Biology and Genetics, National Academy of Sciences, Kyiv, Ukraine

A R T I C L E I N F O

Article history:

Received 20 June 2013

Received in revised form 7 August 2013

Accepted 26 August 2013

Keywords:

Slavs

Serbia

Autosomal STRs

Y-STRs

Population genetics

A B S T R A C T

Studies on Y-chromosomal markers revealed significant genetic differentiation between Southern and

Northern (Western and Eastern) Slavic populations. The northern Serbian region of Vojvodina is

inhabited by Southern Slavic Serbian majority and, inter alia, Western Slavic (Slovak) and Eastern Slavic

(Ruthenian) minorities. In the study, 15 autosomal STR markers were analysed in unrelated Slovaks,

Ruthenians and Serbs from northern Serbia and western Slovakia. Additionally, Slovak males from Serbia

were genotyped for 17 Y-chromosomal STR loci. The results were compared to data available for other

Slavic populations. Genetic distances for autosomal markers revealed homogeneity between Serbs from

northern Serbia and Slovaks from western Slovakia and distinctiveness of Serbian Slovaks and

Ruthenians. Y-STR variation showed a clear genetic departure of the Slovaks and Ruthenians inhabiting

Vojvodina from their Serbian neighbours and genetic similarity to the Northern Slavic populations of

Slovakia and Ukraine. Admixture estimates revealed negligible Serbian paternal ancestry in both

Northern Slavic minorities of Vojvodina, providing evidence for their genetic isolation from the Serbian

majority population. No reduction of genetic diversity at autosomal and Y-chromosomal markers was

found, excluding genetic drift as a reason for differences observed at autosomal STRs. Analysis of

molecular variance detected significant population stratification of autosomal and Y-chromosomal

microsatellites in the three Slavic populations of northern Serbia, indicating necessity for separate

databases used for estimations of frequencies of autosomal and Y-chromosomal STR profiles in forensic

casework. Our results demonstrate that regarding Y-STR haplotypes, Serbian Slovaks and Ruthenians fit

in the Eastern European metapopulation defined in the Y chromosome haplotype reference database.

� 2013 Elsevier Ireland Ltd. All rights reserved.

Contents lists available at ScienceDirect

Forensic Science International: Genetics

jou r nal h o mep ag e: w ww .e lsev ier . co m / loc ate / fs ig

1. Introduction

The Slavic group of languages, the fourth largest Indo-Europeansubgroup, is one of the major language families of the modernworld with approximately 297 million speakers [1]. Slaviclanguages are traditionally divided into three main branches:Southern Slavic, Western Slavic and Eastern Slavic, with the lattertwo often grouped into Northern Slavic. The Southern Slavs inhabita geographically coherent area in Southeastern Europe, and havebeen separated from Northern (Western and Eastern) Slavs of

* Corresponding author at: Department of Forensic Medicine, Medical University

of Gdansk, ul. Marii Skłodowskiej-Curie 3A, 80-210 Gdansk, Poland.

Tel.: +48 58 3491745; fax: +48 58 3410485.

E-mail address: k.rebala@gumed.edu.pl (K. Rebała).

1872-4973/$ – see front matter � 2013 Elsevier Ireland Ltd. All rights reserved.

http://dx.doi.org/10.1016/j.fsigen.2013.08.011

Central and Eastern Europe since the invasion of Finno-UgricHungarians in the late 9th century.

Contacts between the southern and northern Slavdom becamefacilitated in the 18th and 19th centuries in the HabsburgMonarchy and its successor states, Austrian and Austro-HungarianEmpires, which covered territories inhabited by Southern, Westernand Eastern Slavs and which promoted colonisation from the northto its southern borderland. These migration waves involved, interalia, Northern Slavic-speaking Slovaks and Ruthenians who settledin present-day northern Serbia. The latter term refers to EasternSlavic speakers who used to be Austro-Hungarian subjectsinhabiting mainly present-day western Ukraine (Uzhorod andLviv regions) and did not adopt the ethnonym ‘‘Ukrainian’’ in theearly 20th century. According to the 2011 census, there are 53,000Slovaks and 14,000 Ruthenians in Serbia, most of them inhabitingthe northern Serbian region of Vojvodina with 66.8% Serbian

K. Rebała et al. / Forensic Science International: Genetics 8 (2014) 126–131 127

majority [2]. Slovak and Ruthenian are two out of six officiallanguages of the region.

The aim of the study was to assess genetic variation ofautosomal and Y-chromosomal STR markers commonly used inforensic genetics in the Slovak and Ruthenian minorities ofnorthern Serbia. Their genetic links with linguistically andculturally related Slavic populations were determined. Gene poolsof both ethnic groups were tested for the founder effect and geneflow with the neighbouring Serbian majority population.

2. Materials and methods

2.1. Autosomal markers

A total of 938 unrelated adults were enrolled in the study: 774individuals from Vojvodina (231 Slovaks, 322 Ruthenians and 221Serbs, including 100 individuals analysed in our previous study [3])and 164 Slovaks from western Slovakia as a reference population[4]. A total of 15 autosomal STR markers (CSF1PO, D2S1338,D3S1358, D5S818, D7S820, D8S1179, D13S317, D16S539, D18S51,D19S433, D21S11, FGA, TH01, TPOX, VWA) were analysed with theuse of an AmpFlSTR Identifiler PCR Amplification Kit (AppliedBiosystems). Since no data on forensically used autosomal STRswere published for the Ukrainian population, data for theBelarusian population (n = 176) [5] were used as a proxy (amongSlavic languages, Belarusian is the closest to Ukrainian [1] and bothvernaculars were historically referred to as Ruthenian). Parametersevaluating usefulness of the tested markers in forensic casework(power of discrimination, polymorphism information content,power of exclusion, typical paternity index) were calculated withthe aid of PowerStats 1.2 [6]. Observed and expected heterozygos-ity values as well as P-values testing for Hardy-Weinbergequilibrium at each marker were obtained with the use of Arlequin3.1 software [7]. A non-parametric Wilcoxon signed-rank test wascarried out using STATISTICA 10 software (StatSoft) to test fordifferences in observed and expected heterozygosity. Homogenei-ty at autosomal STRs in the Slavic populations of Vojvodina wasassessed by analysis of molecular variance (AMOVA). Pairwiseweighted average FST values over loci [8] were calculated inArlequin 3.1, linearised [9] and applied in multidimensionalscaling (MDS) analysis with the use of STATISTICA 10.

2.2. Y-chromosomal markers

DNA samples of 200 unrelated Slovak males from Vojvodinawere analysed. A total of 17 Y-STR loci (DYS19, DYS389I, DYS389II-I, DYS390, DYS391, DYS392, DYS393, DYS437, DYS438, DYS439,DYS448, DYS456, DYS458, DYS635, GATA H4.1, DYS385 a/b) weregenotyped by means of an AmpFlSTR Yfiler PCR Amplification Kit(Applied Biosystems). Results have been deposited to the Ychromosome haplotype reference database [10] (accession num-ber: YA003632). Yfiler profiles of the other Northern Slavicminority from Vojvodina, Ruthenians (n = 200), were obtainedfrom our previous study [11]. As reference populations, Yfilerhaplotypes of Serbs from Vojvodina (n = 185) [12], Montenegrins(n = 404) [13], Slovaks from western Slovakia (Bratislava region,n = 164) [14] and Ukrainians from western Ukraine (Lviv region,n = 154) [15] were involved in the study. Minimal haplotypes usedin pairwise comparisons additionally included Serbs from centralSerbia (Belgrade region, n = 114) [16], Slovaks from easternSlovakia (n = 629) [17] as well as Ukrainians from western(Uzhorod region, n = 81) [YHRD accession number: YA003547]and northern Ukraine (Kyiv region, n = 82) [18]. In order to test forgenetic homogeneity in Vojvodina by means of AMOVA, conven-tional FST estimates based on haplotype frequencies werecomputed, as far as they are more meaningful for evaluation of

the weight of evidence in forensic casework than FST estimates[19], which take distances between haplotypes into account andwere computed in the study for pairwise comparisons assessingmutual relations between the populations. Calculations of geneticdistances, estimations of corresponding P-values based on 10,000permutations and AMOVA were performed with the use ofArlequin 3.1 software. MDS based on linearised distances wascarried out with the use of STATISTICA 10 software (StatSoft).Frequencies of predicted haplogroups [20] for Yfiler data were usedin admixture analysis based on mR estimators, which werecomputed with the use of Admix 2.0 [21]. After the prediction,J2a1 and its subclades (J2a1b and J2a1h) were pooled, since theyoften cannot be discriminated with the use of Yfiler profiles [22]. Inaddition, haplogroup prediction accuracy was validated with theuse of the western Slovak population sample, for which high-resolution typing results of Y-chromosomal binary and microsat-ellite markers were available [14]. For each Y-STR marker, locusdiversity (corresponding to expected heterozygosity at autosomalmarkers) and discrimination capacity contribution [23] wereobtained. For Yfiler profiles, haplotype diversity, discriminationcapacity, mean pairwise difference (MPD) and weighted meanintralineage MPD (WIMP) [24] were calculated. A non-parametricWilcoxon signed-rank test was carried out to test for interpopula-tion differences in Y-STR locus diversity.

3. Results

Allele frequencies of the autosomal STR markers in the fourstudied populations are presented in Table S1. Values ofparameters evaluating usefulness of the tested STRs in forensicgenetics and paternity testing revealed D2S1338 and D18S51 to bethe most informative markers, whereas TPOX consistently showedthe lowest values of power of discrimination, polymorphisminformation content, power of exclusion and typical paternityindex in all four populations (Table 1). Several markers exhibiteddeviations from Hardy-Weinberg equilibrium (VWA in Slovaksfrom Vojvodina, CSF1PO in Ruthenians from Vojvodina, D3S1358and D21S11 in Slovaks from western Slovakia) with P < 0.05, butthey appeared to be statistically insignificant after Bonferronicorrection. Wilcoxon signed-rank test revealed no statisticallysignificant differences between observed and expected heterozy-gosity values in all the studied populations, as well as in all but oneinterpopulation comparisons between the Northern Slavic minori-ties of Vojvodina and non-isolated majority populations of Serbia,Slovakia and Belarus. The only dissimilarity was noticed betweenobserved heterozygosity values in Slovaks from Vojvodina andBelarusians (P = 0.01), with higher values of observed heterozy-gosity found in the former. AMOVA in the three Slavic populationsof Vojvodina revealed genetic structure in the region(FST = 0.00298, P < 0.00001). All pairwise FST distances appearedto be statistically significant except for genetic homogeneityobserved between Southern Slavic-speaking Serbs and NorthernSlavic-speaking Slovaks from western Slovakia (Table S2), whichwas also visible in the MDS plot (Fig. 1).

Y-STR haplotypes observed in Slovak males from Vojvodina areprovided in Table S3. Regarding locus diversity, DYS19 and two-locus DYS385 appeared to be the most polymorphic Y-STRmarkers, whereas the lowest variability was observed forDYS392. Examination of discrimination capacity contributionvalues revealed that DYS19 was also the most valuable markerin discrimination of similar Y-STR haplotypes. On the other hand,the least valuable markers in discrimination of similar Y-STRhaplotypes were DYS393 and DYS448 (Table 2). Comparison of theSlovak and Ruthenian Y chromosomes from Vojvodina with Yfilerdata from non-isolated majority populations of Serbia, Slovakiaand Ukraine showed no statistically significant differences

Table 1Values of power of discrimination (PD), polymorphism information content (PIC), power of exclusion (PE) and typical paternity index (TPI) as well as observed (Ho) and

expected (He) heterozygosity with corresponding P-values testing for Hardy-Weinberg equilibrium in the studied populations.

PD PIC PE TPI Ho He P PD PIC PE TPI Ho He P

Slovaks from Vojvodina (n = 231) Ruthenians from Vojvodina (n = 322)

CSF1PO 0.880 0.694 0.546 2.2 0.771 0.741 0.940 0.870 0.685 0.518 2.0 0.755 0.732 0.019

D2S1338 0.966 0.860 0.779 4.6 0.892 0.875 0.276 0.971 0.867 0.784 4.7 0.894 0.880 0.329

D3S1358 0.913 0.741 0.561 2.3 0.779 0.777 0.065 0.919 0.755 0.596 2.5 0.798 0.790 0.159

D5S818 0.875 0.681 0.508 2.0 0.749 0.729 0.861 0.876 0.685 0.486 1.9 0.736 0.732 0.524

D7S820 0.924 0.767 0.658 3.0 0.831 0.796 0.527 0.930 0.769 0.550 2.2 0.773 0.800 0.580

D8S1179 0.926 0.762 0.561 2.3 0.779 0.793 0.294 0.933 0.777 0.625 2.7 0.814 0.805 0.681

D13S317 0.921 0.759 0.577 2.4 0.788 0.790 0.509 0.901 0.703 0.471 1.8 0.727 0.738 0.778

D16S539 0.903 0.713 0.437 1.7 0.706 0.753 0.395 0.881 0.695 0.534 2.1 0.764 0.738 0.575

D18S51 0.969 0.862 0.796 5.0 0.900 0.877 0.947 0.966 0.858 0.734 3.8 0.870 0.873 0.190

D19S433 0.930 0.775 0.569 2.3 0.784 0.802 0.197 0.918 0.747 0.512 2.0 0.752 0.778 0.409

D21S11 0.949 0.811 0.650 2.9 0.827 0.832 0.795 0.957 0.835 0.721 3.7 0.863 0.853 0.441

FGA 0.965 0.852 0.682 3.2 0.843 0.869 0.271 0.955 0.829 0.703 3.4 0.854 0.848 0.474

TH01 0.902 0.737 0.601 2.5 0.801 0.774 0.213 0.910 0.732 0.562 2.3 0.780 0.768 0.970

TPOX 0.797 0.557 0.283 1.2 0.593 0.605 0.723 0.763 0.513 0.222 1.1 0.537 0.560 0.950

VWA 0.923 0.771 0.666 3.0 0.836 0.802 0.009 0.927 0.760 0.507 2.0 0.748 0.789 0.631

Serbs from Vojvodina (n = 221) Slovaks from Slovakia (n = 164)

CSF1PO 0.868 0.674 0.504 2.0 0.747 0.726 0.079 0.866 0.670 0.520 2.1 0.756 0.724 0.713

D2S1338 0.968 0.865 0.750 4.1 0.878 0.879 0.409 0.966 0.851 0.739 3.9 0.872 0.867 0.728

D3S1358 0.915 0.740 0.551 2.2 0.774 0.778 0.720 0.924 0.771 0.552 2.2 0.774 0.804 0.016

D5S818 0.851 0.661 0.504 2.0 0.747 0.712 0.486 0.870 0.656 0.411 1.6 0.689 0.709 0.951

D7S820 0.930 0.771 0.543 2.2 0.769 0.803 0.337 0.928 0.784 0.666 3.0 0.835 0.813 0.577

D8S1179 0.929 0.771 0.568 2.3 0.783 0.801 0.077 0.924 0.762 0.631 2.7 0.817 0.794 0.944

D13S317 0.919 0.754 0.559 2.3 0.778 0.786 0.228 0.916 0.738 0.542 2.2 0.768 0.773 0.893

D16S539 0.907 0.729 0.535 2.1 0.765 0.767 0.404 0.883 0.710 0.586 2.4 0.793 0.752 0.437

D18S51 0.966 0.863 0.806 5.3 0.905 0.878 0.536 0.966 0.862 0.813 5.5 0.909 0.878 0.401

D19S433 0.930 0.769 0.551 2.2 0.774 0.796 0.266 0.927 0.769 0.666 3.0 0.835 0.797 0.810

D21S11 0.950 0.824 0.714 3.6 0.860 0.844 0.277 0.948 0.815 0.520 2.1 0.756 0.838 0.026

FGA 0.958 0.839 0.750 4.1 0.878 0.858 0.594 0.957 0.834 0.763 4.3 0.884 0.854 0.714

TH01 0.913 0.738 0.527 2.1 0.760 0.776 0.646 0.908 0.733 0.531 2.1 0.762 0.770 0.511

TPOX 0.782 0.546 0.383 1.5 0.670 0.596 0.278 0.797 0.570 0.334 1.4 0.634 0.630 0.664

VWA 0.939 0.794 0.696 3.3 0.851 0.820 0.915 0.930 0.776 0.574 2.3 0.787 0.805 0.348

K. Rebała et al. / Forensic Science International: Genetics 8 (2014) 126–131128

between locus diversity values in all the compared populations.There were also no significant differences in haplotype diversity,MPD and WIMP values between the Northern Slavic minorities ofthe region and titular populations of Serbia, Slovakia and Ukraine(Table 3).

AMOVA in the three Slavic populations of Vojvodina revealedpaternal genetic stratification in the region (FST = 0.00075,P < 0.00001). Analysis of pairwise FST distances for Yfiler profiles(excluding two-locus DYS385) showed statistically significantdifferentiation of Northern Slavs inhabiting Vojvodina from theirSerbian neighbours (Table S2), which was particularly evident in

Fig. 1. MDS analysis based on weighted average FST distances for 15 autosomal STR

markers. SloVo, SloW Slovaks from Vojvodina, western Slovakia; RuthVo Ruthenians

from Vojvodina; SeVo Serbs from Vojvodina; Be Belarusians.

the MDS plot (Fig. 2). Similar clear-cut differentiation was alsonoticeable in the first dimension of MDS analysis of FST distancesfor minimal haplotypes (excluding DYS385) between Northern andSouthern Slavic populations, with Slovaks and Ruthenians fromVojvodina grouping together with Slovaks and Ukrainians fromtheir nation states (Table S2 and Fig. 2). Validation showed that 163out of 164 (99.4%) western Slovak Yfiler profiles were correctlyallocated to a haplogroup and only one assignment (0.6%) waserroneous, but concerned a haplogroup which was likely notincluded in the prediction software: J2a-M410*(xM67) chromo-some was called J2b. Scrutiny of predicted haplogroups revealedhigh incidence of haplogroup R1a in both Northern Slavic

Table 2Locus diversity (LD) and discrimination capacity (DC) contribution values for Y-STR

haplotypes in the Slovak population of Vojvodina.

LD DC contribution (%)

DYS19 0.792 4.3

DYS389I 0.497 1.1

DYS389II-I 0.661 1.1

DYS390 0.666 1.1

DYS391 0.517 0.5

DYS392 0.328 0.5

DYS393 0.379 0.0

DYS437 0.561 0.5

DYS438 0.656 0.5

DYS439 0.745 2.2

DYS448 0.490 0.0

DYS456 0.755 2.2

DYS458 0.742 0.5

DYS635 0.566 1.6

GATA H4.1 0.600 0.5

DYS385 0.851 0.5

Table 3Haplotype diversity (HD), discrimination capacity (DC), mean pairwise difference (MPD) and weighted mean intralineage MPD (WIMP) values for 17-locus Y-STR haplotypes

in Slovaks and Ruthenians from Vojvodina and in non-isolated majority populations of Serbia, Slovakia and Ukraine.

HD DC (%) MPDa WIMPa

Slovaks from Vojvodina (n = 200) 0.9991 � 0.0006 92.0 8.95 � 4.14 5.67 � 1.37

Ruthenians from Vojvodina (n = 200) 0.9988 � 0.0007 90.0 8.68 � 4.02 5.11 � 1.31

Serbs from Vojvodina (n = 185) 0.9994 � 0.0007 95.1 9.08 � 4.20 4.97 � 1.02

Slovaks from Slovakia (n = 164) 0.9997 � 0.0007 97.6 8.95 � 4.14 5.47 � 1.40

Ukrainians (n = 154) 0.9993 � 0.0008 94.8 8.70 � 4.04 5.41 � 1.40

a Calculated after exclusion of two-locus DYS385.

K. Rebała et al. / Forensic Science International: Genetics 8 (2014) 126–131 129

minorities inhabiting Serbia (42.0% and 44.0% in Slovaks andRuthenians, respectively), which was comparable to its prevalencein the two Northern Slavic reference populations (45.1% and 43.5%in Slovaks and Ukrainians, respectively), but considerably higherthan the one observed in Southern Slavic Serbs (15.1%, Table S4).Admixture analysis of the predicted haplogroups revealed onlyminor Serbian paternal genetic ancestry among the Slovak andRuthenian males from northern Serbia (Table 4).

4. Discussion

Unlike standard autosomal STR markers regarded as sufficientlyhomogeneous for the purpose of forensic DNA typing in Europe, Y-chromosomal haplotypes show a marked geographic differentia-tion within the continent [10,25]. Studies on Y-STR variationamong the Slavs revealed significant differentiation between

Fig. 2. MDS analysis based on FST distances for (a) Yfiler profiles and (b) minimal Y-

STR haplotypes. SloVo, SloW, SloE Slovaks from Vojvodina, western Slovakia, eastern

Slovakia; RuthVo Ruthenians from Vojvodina; UkrLv, UkrUz, UkrKy Ukrainians from

Lviv region, Uzhorod region, Kyiv region; SeVo, SeBe Serbs from Vojvodina, Belgrade

region; Mo Montenegrins.

paternal gene pools of Northern and Southern Slavs, which is likelyowing to distinct processes, which contributed the most to theformation of the Northern and Southern Slavic populations. WhileNorthern Slavic Y chromosomes show a signal of a recentdemographic expansion prior to the divergence of geographicallydistant populations with only minor admixture with neighbouringnon-Slavic populations [14,26], their Southern Slavic counterpartsreveal significant paternal genetic contribution of peoples whosettled in the region before the Slavic expansion [27]. Consequent-ly, populations which were established as a result of contactsbetween the northern and southern Slavdom seem to be ofparticular interest in population genetics. These include NorthernSlavic-speaking minorities of Slovaks and Ruthenians inhabitingpresent-day northern Serbia.

Both Northern Slavic communities of Vojvodina were estab-lished by relatively few settlers from the north and thus are notnumerous. However, although Y-STR haplotype discriminationcapacity values seemed lower in Slovaks and Ruthenians inhabit-ing Vojvodina, our analysis of observed and expected hetero-zygosities, locus and haplotype diversities, as well as MPDs andWIMPs failed to detect reduction in genetic diversity at bothautosomal and Y-chromosomal microsatellites in comparison tonon-isolated majority populations of Serbia, Slovakia, Ukraine andBelarus. Moreover, paternal gene pools of both Serbian Slovaks andRuthenians (including predicted haplogroup patterns) weregenetically indistinguishable from their homeland populationsin Slovakia and Ukraine and clearly separated from their Serbianneighbours with only minor traces of paternal gene flow with theSerbian majority population in the Slovaks and hardly detectableSerbian paternal ancestry in the Ruthenians. Although ouradmixture results based on Y-chromosomal haplogroups assignedwith the use of prediction software are subject to a certain errorrate due to recurrent mutations or database biases affectingassociation probabilities, especially when a low number of Y-STRsis implemented [28], a higher number of markers (e.g. a full Yfilerprofile) enables accurate assignment to a haplogroup in most cases[22]. Our validation confirmed very low haplogroup predictionerror rate for the Yfiler haplotypes, so its effect on the finalinferences should remain negligible.

Our previous study on autosomal STR markers in Finno-Ugric-speaking Hungarians from Vojvodina revealed their geneticsimilarity to the Slavic-speaking Serbian majority population[29]. This observation was not repeated for the two Northern Slavicminorities of Vojvodina, which appeared to be genetically distinctfrom their Serbian neighbours at both autosomal and Y-chromo-somal STRs. It may be owing to relatively short period of

Table 4Least-squares paternal ancestry estimates (mR) for Slovak and Ruthenian

population samples from Vojvodina.

Hybrid population Homeland populationa Serbian population

Slovaks from Vojvodina 0.911 (�0.151) 0.089 (�0.151)

Ruthenians from Vojvodina 1.017 (�0.212) �0.017 (�0.212)

a Slovaks from western Slovakia (Bratislava region) and Ukrainians from western

Ukraine (Lviv region) for Slovaks and Ruthenians from Vojvodina, respectively.

K. Rebała et al. / Forensic Science International: Genetics 8 (2014) 126–131130

interethnic contacts between the three Slavic populations since thearrival of the settlers from the north as well as to their geneticisolation reflected in their typically Northern Slavic paternalgenetic heritage without significant genetic admixture with theSerbian majority population.

An interesting observation was noted for autosomal STRmarkers, which showed homogeneity only between SouthernSlavic Serbs from Vojvodina and Northern Slavic Slovaks fromwestern Slovakia. The revealed genetic proximity clearly contraststhe previously observed differentiation between Southern andNorthern Slavic populations detected at Y-chromosomal markers[27] and may reflect regional homogeneity of autosomal micro-satellites, as far as northern Serbia (Vojvodina) and westernSlovakia are not geographically distant and are separated fromeach other by a territory inhabited by non-Slavic Hungarians.Moreover, the revealed homogeneity seems to be driven bygeography rather than by language, since it involves also theneighbouring Hungarians, who were additionally shown to begenetically diverse from linguistically related but geographicallydistant Finno-Ugric populations [29]. Differences observed atautosomal STRs in pairwise comparisons of Serbian Slovaks andRuthenians with their homeland populations may be simply owingto local genetic differentiation of the northern Slavdom, inheritedby the settlers from the north, rather than owing to genetic drift,given that our results do not provide evidence for a founder effectat forensically used autosomal markers. The local geneticdifferentiation of the autosomal STR gene pools of the NorthernSlavic populations is supported by a notable dissimilarity betweenSlovaks from western Slovakia and Belarusians, which were shownto be genetically indistinguishable regarding Y-STR haplotypes[27].

Developing a forensic DNA database for a region inhabited bylocal ethnic groups separated by physical and cultural barriers is achallenge since the general population of the region can begenetically subdivided [30]. Where the population structure isobserved, local databases should be built up for forensic purposesafter sampling each of the appropriate subpopulations rather thanrandom individuals [31]. Such local databases should be usedwhenever possible in order to estimate frequencies of geneticprofiles and paternity index values in statistical interpretation offorensic genetic evidence [32]. Our AMOVA results indicate thatseparate forensic DNA databases should be developed and used forthe Slavic populations of Vojvodina for both autosomal and Y-chromosomal STR markers.

Since estimates of the frequency of a particular Y-STR haplotypedepend upon the size of the database used, large databases of Y-chromosomal profiles need to be generated to maximise theprobity of forensic genetic evidence [10,25]. Willuweit and Roewer[10] used pairwise FST estimates of a large pan-European study[18] to assign European populations to one out of three geneticallydefined metapopulations: Southeastern, Eastern and WesternEuropean, which facilitated statistical evaluation of haplotypematches in forensic casework with the use of the YHRD owing tosignificant enlargement of the sample sizes. In the abovemen-tioned pan-European study, Roewer et al. [18] found no Y-STRdifferentiation between Western (Poles) and Eastern Slavs(Ukrainians, Belarusians, Russians), but clear genetic departureof Northern (Western and Eastern) Slavic peoples from SouthernSlavic Bulgarians. This observation was confirmed in a subsequentcomprehensive analysis of Y-STR variation in the Slavic popula-tions [27] and was employed to allocate Southern and NorthernSlavs to Southeastern and Eastern European metapopulations,respectively [10]. Since Serbian Slovaks and Ruthenians analysedin our study inhabit the predominantly Southern Slavic territory,but speak Northern Slavic dialects, their assignment to anappropriate metapopulation required accurate investigation.

Pairwise FST distances revealed genetic proximity betweenWestern Slavic Slovaks and Eastern Slavic Ruthenians inhabitingnorthern Serbia and clear-cut differentiation between Northernand Southern Slavic Y chromosomes of Vojvodina, thus confirmingthe previously detected Y-STR variation pattern among the Slavs.Comparison with other Slavic populations demonstrates thatregarding Y-STR haplotypes, both Northern Slavic minorities fromSerbia fit in the Eastern European metapopulation defined in theYHRD.

The paper follows guidelines for publication of population datarequested by the journal [33].

Appendix A. Supplementary data

Supplementary material related to this article can be found, in the

online version, at doi:10.1016/j.fsigen.2013.08.011.

References

[1] R. Sussex, P. Cubberley, The Slavic Languages, Cambridge University Press,Cambridge, 2006.

[2] Statistical Office of the Republic of Serbia, 2011 Census of Population, Householdsand Dwellings in the Republic of Serbia: Population: Ethnicity: Data by Munici-palities and Cities, Belgrade, 2012.

[3] I. Veselinovic, M. Kubat, I. Furac, J. Skavic, I. Martinovic-Klaric, M. Tasic, Allelefrequencies of the 15 AmpFlSTR Identifiler loci in the population of VojvodinaProvince, Serbia and Montenegro, Int. J. Legal Med. 118 (2004) 184–186.

[4] J. Wysocka, K. Rebała, E. Kapinska, L. Cybulska, A.I. Mikulich, I.S. Tsybovsky, D.Sivakova, Z. Dzupinkova, Z. Szczerkowska, Similarities and differences betweenthree Slavic populations on the basis of allelic frequencies for 15 STR loci, ForensicSci. Int. Genet. Suppl. Ser. 1 (2008) 386–388.

[5] K. Rebała, J. Wysocka, E. Kapinska, L. Cybulska, A.I. Mikulich, I.S. Tsybovsky, Z.Szczerkowska, Belarusian population genetic database for 15 autosomal STR loci,Forensic Sci. Int. 173 (2007) 235–237.

[6] A. Tereba, Tools for analysis of population statistics, Profiles DNA 2 (1999) 14–16.[7] L. Excoffier, G. Laval, S. Schneider, Arlequin (version 3.0): an integrated software

package for population genetics data analysis, Evol. Bioinform. Online 1 (2005)47–50.

[8] B.S. Weir, C.C. Cockerham, Estimating F-statistics for the analysis of populationstructure, Evolution 38 (1984) 1358–1370.

[9] M. Slatkin, A measure of population subdivision based on microsatellite allelefrequencies, Genetics 139 (1995) 457–462.

[10] S. Willuweit, L. Roewer, Y chromosome haplotype reference database (YHRD):update, Forensic Sci. Int. Genet. 1 (2007) 83–87.

[11] I. Veselinovic, G. Petric, D. Vapa, Genetic polymorphism of 17 Y chromosomalSTRs in the Rusyn population sample from Vojvodina Province, Serbia, Int. J. LegalMed. (2013), http://dx.doi.org/10.1007/s00414-013-0877-9.

[12] I.S. Veselinovic, D.M. Zgonjanin, M.P. Maletin, O. Stojkovic, M. Djurendic-Brenesel,R.M. Vukovic, M.M. Tasic, Allele frequencies and population data for 17 Y-chromosome STR loci in a Serbian population sample from Vojvodina province,Forensic Sci. Int. 176 (2008) e23–e28.

[13] S. Mirabal, T. Varljen, T. Gayden, M. Regueiro, S. Vujovic, D. Popovic, M. Djuric, O.Stojkovic, R.J. Herrera, Human Y-chromosome short tandem repeats: a tale ofacculturation and migrations as mechanisms for the diffusion of agriculture in theBalkan Peninsula, Am. J. Phys. Anthropol. 142 (2010) 380–390.

[14] K. Rebała, B. Martınez-Cruz, A. Tonjes, P. Kovacs, M. Stumvoll, I. Lindner, A.Buttner, H.E. Wichmann, D. Sivakova, M. Sotak, L. Quintana-Murci, Z. Szczer-kowska, D. Comas, Genographic Consortium, Contemporary paternal geneticlandscape of Polish and German populations: from early medieval Slavic expan-sion to post-World War II resettlements, Eur. J. Hum. Genet. 21 (2013) 415–422.

[15] M. Mielnik-Sikorska, P. Daca, M. Wozniak, B.A. Malyarchuk, J. Bednarek, T. Dobosz,T. Grzybowski, Genetic data from Y chromosome STR and SNP loci in Ukrainianpopulation, Forensic Sci. Int. Genet. 7 (2013) 200–203.

[16] L. Barac-Lauc, M. Pericic, I. Martinovic-Klaric, A. Sijacki, D. Popovic, B. Janicijevic,P. Rudan, Y chromosome STR polymorphisms in a Serbian population sample,Forensic Sci. Int. 150 (2005) 97–101.

[17] E. Petrejcıkova, M. Sotak, J. Bernasovska, I. Bernasovsky, K. Rebała, A. Sovicova, I.Boronova, A. Bozikova, D. Gabrikova, P. Svıckova, S. Macekova, J. Carnogurska, R.Lohaj, D. Vlcek, Allele frequencies and population data for 11 Y-chromosome STRsin samples from Eastern Slovakia, Forensic Sci. Int. Genet. 5 (2011) e53–e62.

[18] L. Roewer, P.J.P. Croucher, S. Willuweit, T.T. Lu, M. Kayser, R. Lessig, P. de Knijff,M.A. Jobling, C. Tyler-Smith, M. Krawczak, Signature of recent historical events inthe European Y-chromosomal STR haplotype distribution, Hum. Genet. 116(2005) 279–291.

[19] J.U. Palo, M. Hedman, I. Ulmanen, M. Lukka, A. Sajantila, High degree of Y-chromosomal divergence within Finland – forensic aspects, Forensic Sci. Int.Genet. 1 (2007) 120–124.

[20] T.W. Athey, Haplogroup prediction from Y-STR values using a Bayesian-allele-frequency approach, J. Genet. Geneal. 2 (2006) 34–39.

K. Rebała et al. / Forensic Science International: Genetics 8 (2014) 126–131 131

[21] I. Dupanloup, G. Bertorelle, Inferring admixture proportions from molecular data:extension to any number of parental populations, Mol. Biol. Evol. 18 (2001) 672–675.

[22] L.J. Martınez-Gonzalez, E. Martınez-Espın, J.C. Alvarez, F. Albardaner, O. Rickards,C. Martınez-Labarga, F. Calafell, J.A. Lorente, Surname and Y chromosome inSouthern Europe: a case study with Colom/Colombo, Eur. J. Hum. Genet. 20 (2012)211–216.

[23] K. Rebała, Z. Szczerkowska, Polish population study on Y chromosome haplotypesdefined by 18 STR loci, Int. J. Legal Med. 119 (2005) 303–305.

[24] M.E. Hurles, J. Nicholson, E. Bosch, C. Renfrew, B.C. Sykes, M.A. Jobling, Y-chromosomal evidence for the origins of Oceanic-speaking peoples, Genetics160 (2002) 289–303.

[25] J. Ballantyne, L. Fatolitis, L. Roewer, Creating and managing effective Y-STRdatabases, Profiles DNA 9 (2006) 10–13.

[26] O. Balanovsky, S. Rootsi, A. Pshenichnov, T. Kivisild, M. Churnosov, I. Evseeva, E.Pocheshkhova, M. Boldyreva, N. Yankovsky, E. Balanovska, R. Villems, Twosources of the Russian patrilineal heritage in their Eurasian context, Am. J.Hum. Genet. 82 (2008) 236–250.

[27] K. Rebała, A.I. Mikulich, I.S. Tsybovsky, D. Sivakova, Z. Dzupinkova, A. Szczer-kowska-Dobosz, Z. Szczerkowska, Y-STR variation among Slavs: evidence for theSlavic homeland in the middle Dnieper basin, J. Hum. Genet. 52 (2007) 406–414.

[28] D. Taylor, N. Nagle, K.N. Ballantyne, R.A.H. van Oorschot, S. Wilcox, J.Henry, R. Turakulov, R.J. Mitchell, An investigation of admixture in an AustralianAboriginal Y-chromosome STR database, Forensic Sci. Int. Genet. 6 (2012)532–538.

[29] G. Petric, D. Draskovic, D. Zgonjanin-Bosic, B. Budakov, I. Veselinovic,Genetic variation at 15 autosomal STR loci in the Hungarian population ofVojvodina Province, Republic of Serbia, Forensic Sci. Int. Genet. 6 (2012) e163–e165.

[30] L.A. Zhivotovsky, B.A. Malyarchuk, M.V. Derenko, M. Wozniak, T. Grzybowski,Developing STR databases on structured populations: the native South Siberianpopulation versus the Russian population, Forensic Sci. Int. Genet. 3 (2009) e111–e116.

[31] L.A. Zhivotovsky, S. Ahmed, W. Wang, A.H. Bittles, The forensic DNA implicationsof genetic differentiation between endogamous communities, Forensic Sci. Int.119 (2001) 269–272.

[32] U. Toscanini, M. Garcia-Magarinos, G. Berardi, T. Egeland, E. Raimondi, A. Salas,Evaluating methods to correct for population stratification when estimatingpaternity indexes, PLoS ONE 7 (2012) e49832.

[33] A. Carracedo, J.M. Butler, L. Gusmao, A. Linacre, W. Parson, L. Roewer, P.M.Schneider, New guidelines for the publication of genetic population data, ForensicSci. Int. Genet. 7 (2013) 217–220.