See discussions, stats, and author profiles for this publication at: https://www.researchgate.

net/publication/345977436

Allele frequencies and forensic parameters of 22 autosomal STR loci in

population of 983 individuals from Serbia and comparison with 24 other
populations

Article  in  Annals of Human Biology · November 2020

DOI: 10.1080/03014460.2020.1846784

CITATION READS

1 160

5 authors, including:

Dijana Takić Miladinov Perica Vasiljević

Institute of Legal Medicine Niš (Republic of Serbia) University of Niš
8 PUBLICATIONS   25 CITATIONS    94 PUBLICATIONS   775 CITATIONS   

SEE PROFILE SEE PROFILE

Dejan Šorgić Eva Podovšovnik

University of Belgrade University of Primorska
3 PUBLICATIONS   1 CITATION    27 PUBLICATIONS   123 CITATIONS   

SEE PROFILE SEE PROFILE

Some of the authors of this publication are also working on these related projects:

Age-related DNA methylation changes for forensic age-prediction View project

VIHOS - Towards Virtual Physiological Human View project

All content following this page was uploaded by Dijana Takić Miladinov on 23 November 2020.

The user has requested enhancement of the downloaded file.

 Annals of Human Biology

ISSN: (Print) (Online) Journal homepage: https://www.tandfonline.com/loi/iahb20

Allele frequencies and forensic parameters of

22 autosomal STR loci in a population of 983
individuals from Serbia and comparison with 24
other populations

Dijana Takić Miladinov , Perica Vasiljević , Dejan Šorgić , Eva Podovšovnik

Axelsson & Aleksandra Stefanović

To cite this article: Dijana Takić Miladinov , Perica Vasiljević , Dejan Šorgić , Eva Podovšovnik
Axelsson & Aleksandra Stefanović (2020): Allele frequencies and forensic parameters of 22
autosomal STR loci in a population of 983 individuals from Serbia and comparison with 24 other
populations, Annals of Human Biology, DOI: 10.1080/03014460.2020.1846784

To link to this article: https://doi.org/10.1080/03014460.2020.1846784

Published online: 23 Nov 2020.

Submit your article to this journal

Article views: 35

View related articles

View Crossmark data

Full Terms & Conditions of access and use can be found at

https://www.tandfonline.com/action/journalInformation?journalCode=iahb20
ANNALS OF HUMAN BIOLOGY
https://doi.org/10.1080/03014460.2020.1846784

HUMAN BIOLOGICAL SURVEY

Allele frequencies and forensic parameters of 22 autosomal STR loci in a

population of 983 individuals from Serbia and comparison with 24 other
populations
Dijana Taki
c Miladinova , Perica Vasiljevi
cb , Dejan Sorgi
ca, Eva Podovsovnik Axelssonc and
Aleksandra Stefanovi
ca
a
Institute of Legal Medicine, Nis, Republic of Serbia; bFaculty of Science and Mathematics, Department of Biology and Ecology, University of
Nis, Nis, Republic of Serbia; cTuristica – Faculty of Tourism Studies, University of Primorska, Portoroz, Slovenia

ABSTRACT ARTICLE HISTORY

Background: Analysis of allele frequencies of short tandem repeat (STR) loci in ethnically diverse pop- Received 14 February 2020
ulations is essential for forensic reference database construction and studies on population genetics. Revised 12 August 2020
Aim: To analyse genetic polymorphisms of 22 autosomal STR loci in the Serbian population and to Accepted 1 October 2020
compare them with previously published data from some European and Turkish populations.
KEYWORDS
Subjects and methods: The study was conducted among 983 unrelated individuals from Serbia. STR; population data;
Genotyping was performed using the PowerPlexV Fusion amplification kit. Allele frequencies and
R

PowerPlex Fusion; forensic

forensic parameters were calculated using FORSTAT software. Interpopulation comparisons and genetic genetics; Serbia
distance calculations were performed in Arlequin and POPTREEW software.
Results: A total of 280 alleles were detected with corresponding allelic frequencies ranging from
0.0005 to 0.5255. Based on heterozygosity and the polymorphism information content, D1S1656 and
Penta E may be considered as the most informative markers. Both the combined power of discrimin-
ation (CPD) and the combined power of exclusion (CPE) for the 22 analysed loci were higher than
0.999999. The combined match probability (CPM) for all 22 loci was 6.773688e
29.
Conclusion: With respect to the results, the 22 STR loci are highly polymorphic and discriminating in
the Serbian population and could be used for forensic practice and population genetics studies.

Background
Their highly polymorphic nature, short sequence lengths,
and ability to amplify minute quantities of template DNA
make short tandem repeat (STR) loci invaluable markers
widely used in forensic and paternity testing as well as in
studies on population genetics (Butler 2012). To obtain reli-
able results of the biostatistical analysis of DNA profiles, it is
necessary to use allele frequency data obtained from a suffi-
cient number of unrelated healthy individuals of a popula-
tion (Butler 2010). A few published studies have
characterised autosomal STRs in the Serbian population, but
are, however, based on different numbers and sets of STR
loci in comparison to our study. Namely, 14 STR loci were
employed in the study of 296–531 (depending on the locus)
unrelated individuals from Serbia and Montenegro
(Keckarevi
c et al. 2005), 15 AmpFlSTR Identifiler loci in the
analysis of 356 ethnic Serbian individuals from the Republic
of Serbia (Novkovi
c et al. 2010) and 15 AmpFlSTR Identifiler
loci in the analysis of 100 individuals from Novi Sad
(Vojvodina Province, Serbia) (Veselinovi
c et al. 2004). With
the decision of the European Network of Forensic Science
Institutes and the European DNA Profiling Group to extend
the current European Standard Set loci (D3S1358, vWA,
D8S1179, D21S11, D18S51, TH01, and FGA) with five new loci
(D1S1656, D2S441, D10S1248, D12S391, and D22S1045),
there was a need to obtain data for these loci within the
Serbian population (Gill et al. 2006a, 2006b). Besides the
increase in the number of loci, the latest recommendations
regarding the publication and usage of STR population data
in biostatistical calculations for forensic purposes, emphas-
ised the need for increasing the size of population samples
in order to increase the accuracy of calculations (Dupuy et al.
2013; Gusmao et al. 2017).
Here we have reported allele frequencies and forensic
parameters of 22 autosomal STR loci included in the
PowerPlex Fusion PCR amplification kit (PPF), in a sample of
983 unrelated healthy individuals from Serbia. Based on five
dyes, PPF is a 24-locus multiplex system that ensures simul-
taneous detection of 20 autosomal loci in the expanded
CODIS (CSF1PO, FGA, TH01, TPOX, vWA, D1S1656, D2S1338,
D2S441, D3S1358, D5S818, D7S820, D8S1179, D10S1248,
D12S391, D13S317, D16S539, D18S51, D19S433, D21S11, and
D22S1045) and two more loci for better discrimination
(Penta D and Penta E). Additional sex determining loci (ame-
logenin and DYS391) were amplified but were excluded from
the statistical analysis.

CONTACT Dijana Taki
c Miladinov takicdijana@gmail.com Institute of Legal Medicine, Blvd. Zorana Djinjica, 81, Nis 18000, Republic of Serbia
ß 2020 Informa UK Limited, trading as Taylor & Francis Group
2 D. TAKIĆ MILADINOV ET AL.

Table 1. List of populations used for comparative analysis.

Country (population) References
Kosovo Province, Southern Serbia (Albanians) Kubat et al. (2004)
Austria (Austrian Caucasians from Innsbruck area) Ross et al. (2001); Hatzer-Grubwieser et al. (2012)
Belarus (ethnic Belarusians from 73 Belarus’ districts) Zhivotovsky, Veremeichyk, et al. (2009)
Belgium (Belgians from various parts of Belgium) Decorte et al. (2004)
Bosna and Herzegovina Pilav et al. (2017)
Croatia

c et al. (2012)
Proji
c et al. (2007); Curi
Cyprus (Greek Cypriots) Cariolou et al. (2006)
Czech Republic (Caucasian Czechs) Simkov
a et al. (2009)
Denmark Pereira et al. (2015)
Estonia Sadam et al. (2015)
Hungary Rak et al. (2011)
Italy (Italians from Brescia area, Northern Italy) Cerri et al. (2003)
Latvia (Latvians from various parts of Latvia) Jemeljanova et al. (2015)
Moldavia Stanciu, Popescu, et al. (2009)
Montenegro (adults from central part of the Republic of Montenegro) Jeran et al. (2007)
Pomorze Zachodnie region of Poland, North-western Poland Piatek et al. (2008)
Portugal (Individuals from Azores archipelago and Central Portugal) Lopes et al. (2009)
Romania (Romanians from Wallachia region, South Romania) Stanciu, Stoian, et al. (2009)
Russia (Russians from Belgorod city, Pskov city, and Velikiy Novgorod city) Zhivotovsky, Malyarchuk, et al. (2009)
Spain (Caucasians from North-eastern Spain) Paredes et al. (2003)
Sweden Montelius et al. (2008)
The Netherlands Westen et al. (2014)
Turkey Poetsch and von Wurmb-Schwark (2013)
Ukraina (males living in Kyiv) Serga et al. (2017)

Figure 1. (A) Republic of Serbia (SRB) and Populations used for comparisons are from countries labelled with following abbreviations: BIH – Bosnia and
Herzegovina, MNT – Montenegro, HRV – Croatia, HUN – Hungary, ROM – Romania, MDA – Moldavia, UKR – Ukraine, BLR – Belarus, RUS – Russia, EST – Estonia,
LVA – Latvia, AUT – Austria, ITA – Italy, ESP – Spain, PRT – Portugal, BEL – Belgium, NLD – The Netherlands, DNK – Denmark, SWE – Sweden, POL – Poland, CZE –
Republic of Czech, TUR – Turkey, CYP – Cyprus. (B) Districts of Republic of Serbia. Percent show a number of individuals from different districts involved in
our study.

We performed extensive comparisons of data from the
studied Serbian population with that previously published
for 24 other populations. The list and appropriate references
of the selected populations are given in Table 1, while geo-
graphical locations are presented in Figure 1(A).
This is the first study analysing an expanded number of
STR loci in a representative sample of the Serbian population
and the first study of comprehensive comparison with previ-
ously published data for other populations.
The DNA analysis Department of the Institute of Legal
Medicine in Nis, Republic of Serbia, is accredited to ISO/IEC
17025 (accreditation number 01–486), and regularly partici-
pates in quality control proficiency testing provided by the
German DNA Profiling group (GEDNAP).

Sample
Buccal swabs of unrelated individuals processed in casework
analyses, performed between June 2016 and June 2019 at
the Institute for Legal Medicine, were used as the primary
source material to generate frequency data. The Republic of
Serbia has two Provinces (Vojvodina and Kosovo & Metohija)
and 30 districts. Participants were linked to the appropriate
district according to their place of birth, so the structure of
our sample was as follows: 45% of individuals were from
Nisavski/Toplicki/Pirotski regions, 15% from Jablanicki/
Pcinjski, 14% from Borski/Zajecarski, 10% from Macvanski/
Kolubarski/Moravicki/Raski/Rasinski, 4% from Sumadijski/
Pomorovaski, 3% from Kosovo & Metohija Province and the
other 9% of individuals were from all other regions (Figure
1(B)). This work was approved by the Ethics Committee of
the Institute of Legal Medicine, University of Nis, Republic of
Serbia (number 04_2814/19).
Data collection
Buccal swabs were placed in 1.5 ml tubes and immediately
extracted using the QIAamp DNA mini kit (Qiagen, Hilden,
Germany) according to the manufacturer’s instructions. DNA
quantification was conducted using the Quantifiler HP kit
and 7500 RealTime PCR System (Applied Biosystems, Foster
City, CA). All DNA isolates were immediately processed to
the next step or stored at
20  C for a few days. After quan-
tification, DNA was amplified with the PPF kit (Promega
Corporation, Madison, WI), in the ProFlexTM 3  32-well PCR
System (Thermo Fisher Scientific Company, Waltham, MA)
according to the manufacturer’s recommendations. Capillary
electrophoresis was performed on the ABI 3500 Genetic
Analyser (Thermo Fisher Scientific Company). GeneMarker
software version 3 (Soft Genetics, State College, PA) was
used for raw data analysis.
Statistical analysis
Allele frequencies and parameters of forensic interest
(expected and observed heterozygosity (He, Ho), power of
discrimination (PD), power of exclusion (PE), matching prob-
ability (MP), polymorphic information content (PIC) and
paternity index (PI)) were calculated using FORSTAT software
version 1 (Ristow and D’Amato 2017). Interpopulation com-
parisons were conducted in Arlequin software version 3.5.1.3
(Excoffier and Lischer 2010). P values less than 0.05 are con-
sidered as significant. Nei’s genetic distances (Nei 1972)
among the compared populations were derived and used to
generate a Neighbor-Joining (NJ) dendrogram in POPTREEW
software (Takezaki et al. 2014). Graphical representation of
genetic distances was performed based on multidimensional
scaling (MDS) analysis using the Past 3.25 software (Hammer
et al. 2001).
ANNALS OF HUMAN BIOLOGY 3
Results
The allele frequencies and forensic parameters of the 22
autosomal STR loci under study are given in Table 2. A total
of 280 alleles were counted and no rare (off-ladder) alleles
could be found in our samples. FGA possessed the highest
number of alleles (22) and TH01 possessed the lowest num-
ber of alleles (7). Allele 8 at TPOX locus was the most fre-
quent in the entire population. The observed and expected
heterozygosity ranged from 0.619 (TPOX) to 0.895 (D1S1656)
and from 0.633 (TPOX) to 0.895 (Penta E), respectively. The
most polymorphic and discriminatory STR locus in the
studied population was Penta E with respective values of
0.886 and 0.982 for PIC and PD. PIC values, as measures of
informativeness, can theoretically range from 0 to 1. A PIC
value of greater than 0.7 is considered to be highly inform-
ative (Hildebrand et al. 1992). Markers with greater numbers
of alleles tend to have higher PIC values and thus are more
informative, such as D12S391, D1S1656, and Penta E in our
study. The PE values ranged from 0.315 (TPOX) to 0.786
(D1S1656) with a value of greater than 0.999999995 for the
combined power of exclusion (CPE). The PD values ranged
from 0.844 (TPOX) to 0.982 (Penta E), with a value of 1 for
the combined power of discrimination (CPD). The combined
probability of the match (CPM) and combined paternity
index (CPI) for all 22 autosomal STR loci were found to be
6.17e
29 and 3.14e18, respectively. Therefore, the 22 auto-
somal STR loci exhibited high efficacy and informativeness
for forensic investigation and individualisation purposes.
The calculated allele frequencies of the studied Serbian
population were compared with those from 24 other popula-
tions at 12 loci which were common among all populations
(D21S11, D7S820, D5S818, D3S1358, VWA, D8S1179,
D16S539, TH01, D18S51, FGA, D13S317, and D19S433). First,
we performed an exact population differentiation test using
Arlequin software. The results, presented in Table 3, show
that allele frequencies at 12 loci in the studied Serbian popu-
lation are not significantly different from those of the neigh-
bouring Bosnian population and the geographically more
distant country of Spain. It should be taken into account that
the Spanish sample, used for comparison in our study, had a
limited number of participants (204), so in the future, it
would be better to compare a larger number of loci between
populations or use data from a larger Spanish sample when
this becomes available. p-Values from locus-by-locus compar-
isons highlight the significant differences between Serbs and
Kosovo Albanians, Austrians and Moldavians (in 1 out of 12
compared loci), Montenegrins, Croatians, Hungarians, Italians,
Turks, Danes and Ukrainians (in 2 out of 12 compared loci),
Belgians (in 3 out of 12 compared loci), Estonians, Poles and
Czechs (in 4 out of 12 compared loci), Romanians (in 5 out
of 12 compared loci), Latvians (in 6 out of 9 compared loci),
Russians and Swedes (in 6 out of 12 compared loci), Dutch
and Portuguese (in 7 out of 12 compared loci), and
Belarusians (in 8 out of 12 compared loci). The most promin-
ent difference was found between Serbs and Greeks from
Cyprus (in 9 out of 12 compared loci).
Next, we calculated genetic distances between studied
populations (Table 4). A Neighbor-Joining dendrogram was
 4

Table 2. Allele frequencies and forensic parameters of 22 autosomal STR loci in Serbian population (n ¼ 983).
Allele CSF1PO D10S1248 D12S391 D13S317 D16S539 D18S51 D19S433 D1S1656 D21S11 D22S1045 D2S1338 D2S441 D3S1358 D5S818 D7S820 D8S1179 FGA Penta D Penta E TH01 TPOX vWA
6 0.0005 0.074 0.0005
0.0005 0.0005 0.0005 0.271 0.0005
6.2
7 0.002 0.015 0.003 0.147 0.134 0.001
7.3 0.0005 0.004
8 0.002 0.151 0.021 0.002 0.162 0.015 0.014 0.014 0.124 0.525
8.3 0.0005 0.0005
9 0.029 0.076 0.109 0.002 0.001 0.038 0.15 0.019 0.245 0.013 0.206 0.104
9.1 0.001
9.3 0.302 0.001 0.056 0.051 0.012 0.002 0.182 0.089 0.001 0.06 0.104 0.113 0.251 0.055
D. TAKIĆ MILADINOV ET AL.

10 0.266 0.012
10.3 0.0005
11 0.309 0.003 0.331 0.312 0.016 0.008 0.09 0.142 0.311 0.001 0.309 0.224 0.071 0.199 0.095 0.275 0.0005
11.3 0.293 0.017 0.271 0.296 0.111 0.094 0.144 0.016 0.072 0.0005 0.36 0.135 0.128 0.172 0.172 0.033 0.0005
12 0.043
12.1 0.0005
12.2 0.001
12.3 0.002 0.001
13 0.058 0.216 0.077 0.184 0.128 0.235 0.078 0.005 0.002 0.021 0.003 0.185 0.039 0.328 0.167 0.122 0.0005 0.003
13.2 0.021
13.3 0.002
14 0.005 0.317 0.034 0.023 0.169 0.321 0.092 0.054 0.321 0.091 0.011 0.003 0.237 0.064 0.056 0.102
14.2 0.044 0.001
14.3
15 0.001 0.226 0.039 0.003 0.001 0.151 0.143 0.171 0.341 0.002 0.036 0.268 0.001 0.0005 0.113 0.021 0.053 0.105
15.2 0.045
15.3 0.169 0.033 0.0005 0.0005 0.0005 0.0005 0.045 0.332 0.051 0.007 0.248 0.023 0.0005 0.006 0.047 0.193
16 0.159 0.052 0.137
16.2 0.024
16.3 0.047
17 0.045 0.094 0.102 0.002 0.039 0.093 0.25 0.199 0.004 0.003 0.04 0.276
17.2 0.015 0.005 0.092
17.3
18 0.005 0.19 0.069 0.003 0.013 0.102 0.181 0.012 0.028 0.221
18.2 0.001
18.3 0.0005 0.025 0.037 0.05 0.004 0.094 0.007 0.065 0.013 0.084
19 0.106 0.001
19.2 0.0005
19.3 0.007 0.008
20 0.121 0.019 0.138 0.0005 0.127 0.004 0.013
20.2 0.0005
20.3 0.0005 0.0005
21 0.111 0.015 0.029 0.179 0.004 0.001
21.2 0.121 0.006 0.021 0.006 0.001
22 0.176
22.2 0.005
22.3 0.0005 0.001
23 0.087 0.001 0.11 0.139
23.2
24 0.031 0.096 0.01
24.2 0.141
24.3 0.001
25 0.017 0.085 0.0005
(continued)
Table 2. Continued.
Allele CSF1PO D10S1248 D12S391 D13S317 D16S539 D18S51 D19S433 D1S1656 D21S11 D22S1045 D2S1338 D2S441 D3S1358 D5S818 D7S820 D8S1179 FGA Penta D Penta E TH01 TPOX vWA
25.2 0.002 0.002 0.016 0.086
26 0.002
26.2 0.04
27 0.0005 0.021 0.003 0.001
28 0.152 0.005
28.1 0.0005 0.001
28.2 0.0005
29 0.228
29.2 0.006 0.0005
30 0.193
30.2 0.04
31 0.064
31.2 0.114
32 0.01
32.2 0.114
33.2 0.043
34.2 0.008
35.1 0.0005
Forensic parameters
Ho 0.743 0.758 0.884 0.788 0.778 0.871 0.78 0.895 0.841 0.738 0.858 0.779 0.776 0.737 0.805 0.79 0.862 0.801 0.887 0.771 0.619 0.825

He 0.723 0.771 0.89 0.778 0.765 0.877 0.804 0.892 0.853 0.741 0.866 0.758 0.786 0.73 0.81 0.797 0.868 0.827 0.895 0.787 0.633 0.809
PD 0.887 0.913 0.979 0.928 0.908 0.972 0.939 0.977 0.963 0.896 0.969 0.903 0.921 0.897 0.946 0.933 0.968 0.957 0.982 0.926 0.844 0.938
PE 0.498 0.524 0.762 0.577 0.559 0.737 0.599 0.786 0.677 0.489 0.711 0.561 0.556 0.489 0.609 0.581 0.719 0.602 0.769 0.547 0.315 0.646
MP 0.112 0.087 0.021 0.072 0.092 0.028 0.061 0.023 0.037 0.104 0.031 0.097 0.079 0.102 0.054 0.067 0.032 0.043 0.018 0.074 0.156 0.062

PIC 0.671 0.734 0.88 0.747 0.729 0.864 0.78 0.882 0.837 0.7 0.853 0.721 0.751 0.685 0.783 0.771 0.854 0.804 0.886 0.754 0.58 0.782
PI 1.946 2.07 4.297 2.362 2.253 3.889 2.497 4.779 3.148 1.907 3.521 2.263 2.237 1.905 2.568 2.381 3.633 2.52 4.426 2.185 1.313 2.854
No. of 8 10 19 11 10 16 17 17 16 9 14 12 10 9 14 10 22 11 18 7 9 11
alleles
ANNALS OF HUMAN BIOLOGY
5
Table 3. Comparison of allele frequencies on 12 STR loci between Serbian population and previously published population data (exact test ± standard error).
6

Loci/
population D3S1358 D5S818 D7S820 D8S1179 D13S317 D16S539 D18S51 D19S433 D21S11 FGA TH01 vWA
BLR 0.007 ± 0.002 0.181 ± 0.025 0.039 ± 0.011 0.010 ± 0.006 0.006 ± 0.004 0.002 ± 0.001 0.096 ± 0.022 0.517 ± 0.027 0.000 ± 0.000 0.000 ± 0.000 0.000 ± 0.000 0.670 ± 0.024
NLD 0.101 ± 0.019 0.001 ± 0.000 0.025 ± 0.009 0.012 ± 0.007 0.000 ± 0.000 0.350 ± 0.034 0.074 ± 0.017 0.001 ± 0.000 0.000 ± 0.000 0.407 ± 0.045 0.000 ± 0.000 0.626 ± 0.025
LVA 0.015 ± 0.005 0.202 ± 0.017 0.025 ± 0.008 0.001 ± 0.000 0.005 ± 0.001 0.017 ± 0.004 0.273 ± 0.031 0.000 ± 0.000 0.635 ± 0.024
EST 0.005 ± 0.002 0.916 ± 0.008 0.953 ± 0.007 0.304 ± 0.018 0.083 ± 0.009 0.052 ± 0.008 0.050 ± 0.008 0.003 ± 0.001 0.032 ± 0.009 0.577 ± 0.019 0.022 ± 0.004 0.443 ± 0.027
RUS 0.002 ± 0.001 0.671 ± 0.012 0.947 ± 0.009 0.036 ± 0.006 0.102 ± 0.021 0.212 ± 0.020 0.578 ± 0.023 0.241 ± 0.024 0.012 ± 0.006 0.017 ± 0.005 0.003 ± 0.002 0.050 ± 0.008
BIH 0.648 ± 0.027 0.682 ± 0.023 0.512 ± 0.045 0.221 ± 0.017 0.490 ± 0.047 0.852 ± 0.018 0.576 ± 0.035 0.433 ± 0.019 0.761 ± 0.019 0.292 ± 0.040 0.421 ± 0.040 0.589 ± 0.031
MNT 0.710 ± 0.015 0.203 ± 0.014 0.702 ± 0.016 0.627 ± 0.015 0.430 ± 0.013 0.471 ± 0.014 0.965 ± 0.003 0.000 ± 0.000 0.001 ± 0.001 0.724 ± 0.021 0.618 ± 0.015 0.239 ± 0.018
KOS ALB 0.767 ± 0.012 0.898 ± 0.013 0.340 ± 0.023 0.341 ± 0.021 0.594 ± 0.018 0.893 ± 0.007 0.061 ± 0.006 0.411 ± 0.016 0.036 ± 0.004 0.658 ± 0.019 0.457 ± 0.020 0.186 ± 0.012
HRV 0.526 ± 0.016 0.644 ± 0.016 0.762 ± 0.018 0.234 ± 0.019 0.992 ± 0.002 0.983 ± 0.001 0.043 ± 0.005 0.267 ± 0.032 0.977 ± 0.004 0.442 ± 0.024 0.038 ± 0.008 0.363 ± 0.021
HUN 0.428 ± 0.033 0.402 ± 0.038 0.011 ± 0.004 0.403 ± 0.049 0.032 ± 0.013 0.164 ± 0.026 0.973 ± 0.003 0.126 ± 0.028 0.176 ± 0.030 0.097 ± 0.023 0.091 ± 0.022 0.986 ± 0.004
D. TAKIĆ MILADINOV ET AL.

AUT 0.859 ± 0.008 0.780 ± 0.010 0.765 ± 0.016 0.389 ± 0.016 0.985 ± 0.002 0.534 ± 0.015 0.492 ± 0.041 0.284 ± 0.018 0.173 ± 0.016 0.700 ± 0.023 0.002 ± 0.000 0.944 ± 0.004
SWE 0.011 ± 0.002 0.038 ± 0.011 0.623 ± 0.025 0.127 ± 0.022 0.221 ± 0.032 0.824 ± 0.017 0.984 ± 0.003 0.000 ± 0.000 0.000 ± 0.000 0.015 ± 0.009 0.000 ± 0.000 0.930 ± 0.006
NW POL 0.000 ± 0.000 0.131 ± 0.014 0.992 ± 0.002 0.065 ± 0.007 0.493 ± 0.014 0.269 ± 0.020 0.410 ± 0.024 0.354 ± 0.019 0.031 ± 0.007 0.000 ± 0.000 0.001 ± 0.001 0.911 ± 0.009
CZE 0.043 ± 0.008 0.096 ± 0.016 0.798 ± 0.039 0.206 ± 0.022 0.371 ± 0.044 0.066 ± 0.031 0.480 ± 0.018 0.815 ± 0.015 0.000 ± 0.000 0.016 ± 0.004 0.000 ± 0.000 0.999 ± 0.000
BEL 0.112 ± 0.012 0.648 ± 0.025 0.995 ± 0.002 0.751 ± 0.016 0.545 ± 0.018 0.722 ± 0.015 0.812 ± 0.020 0.010 ± 0.004 0.007 ± 0.003 0.637 ± 0.027 0.012 ± 0.003 0.923 ± 0.007
ITA 0.256 ± 0.015 0.369 ± 0.011 0.218 ± 0.016 0.053 ± 0.004 0.250 ± 0.010 0.000 ± 0.000 0.999 ± 0.000 0.062 ± 0.008 0.057 ± 0.005 0.091 ± 0.010 0.000 ± 0.000 0.344 ± 0.011
Greek CPY 0.000 ± 0.000 0.042 ± 0.010 0.000 ± 0.000 0.000 ± 0.000 0.058 ± 0.015 0.023 ± 0.010 0.000 ± 0.000 0.040 ± 0.008 0.106 ± 0.021 0.001 ± 0.001 0.000 ± 0.000 0.199 ± 0.038
TUR 0.090 ± 0.013 0.480 ± 0.023 0.051 ± 0.005 0.051 ± 0.010 0.768 ± 0.018 0.025 ± 0.007 0.315 ± 0.022 0.009 ± 0.004 0.944 ± 0.009 0.399 ± 0.030 0.369 ± 0.019 0.914 ± 0.007
PRT 0.293 ± 0.030 0.000 ± 0.000 0.033 ± 0.012 0.081 ± 0.021 0.000 ± 0.000 0.432 ± 0.028 0.302 ± 0.052 0.004 ± 0.002 0.000 ± 0.000 0.133 ± 0.022 0.000 ± 0.000 0.015 ± 0.003
ESP 0.486 ± 0.019 0.971 ± 0.004 0.556 ± 0.027 0.128 ± 0.013 0.418 ± 0.032 0.968 ± 0.005 0.538 ± 0.018 0.254 ± 0.016 0.373 ± 0.027 0.601 ± 0.018 0.359 ± 0.021 0.491 ± 0.018
DNK 0.134 ± 0.016 0.390 ± 0.013 0.985 ± 0.004 0.245 ± 0.024 0.559 ± 0.015 0.452 ± 0.026 0.427 ± 0.033 0.083 ± 0.015 0.016 ± 0.004 0.605 ± 0.025 0.001 ± 0.000 0.787 ± 0.014
UKR 0.010 ± 0.002 0.485 ± 0.021 0.976 ± 0.004 0.868 ± 0.011 0.837 ± 0.012 0.000 ± 0.000 0.964 ± 0.004 0.547 ± 0.023 0.402 ± 0.032 0.478 ± 0.020 0.313 ± 0.025 0.298 ± 0.019
MDA 0.616 ± 0.022 0.815 ± 0.016 0.514 ± 0.035 0.649 ± 0.029 0.105 ± 0.011 0.000 ± 0.000 0.932 ± 0.011 0.180 ± 0.025 0.177 ± 0.027 0.409 ± 0.049 0.513 ± 0.036 0.480 ± 0.041
ROM 0.106 ± 0.016 0.343 ± 0.036 0.035 ± 0.013 0.200 ± 0.022 0.000 ± 0.000 0.000 ± 0.000 0.000 ± 0.000 0.214 ± 0.016 0.159 ± 0.030 0.225 ± 0.034 0.660 ± 0.019 0.016 ± 0.006
p value of the exact test of population differentiation. Significant differences (p < 0.05) are in bold.
the fourth.

Central Europe.

and might indicate greater genetic isolation than expected

tions. Also, locus-by-locus comparison between those two

literature describing the separation of Italians (Lao et al.

from Czech, Austria, and Croatia, are positioned between
Montenegro, Hungary, Moldavia, and Romania are clustered
Russia, Belarus and Ukraine form another cluster of North-
Sweden, Denmark, The Netherlands, and Belgium form one
Peninsula, namely from Portugal and Spain, formed another
Turkish and Greek Cypriots who share the same territory of

tion on the loci D21S11 and D19S433 were also observed in

frequency for 2 out of 12 compared loci and we calculated
Montenegrins revealed statistically significant differences in
group. Populations from the former Soviet Union countries
Cyprus formed one group. Populations from the Iberian
created according to Nei’s genetic distances (Figure 2.). As

clustering of the studied populations into four groups. The

observed in this study was most likely biased due to the

Europeans, the outlying positioning of these populations
2008) and Albanians (Davidovic et al. 2015) from other
“outliers” in this study. Although there are similar data in the
of Albanians, although these two populations live on the
ously (Pilav et al. 2017). A comparison between Serbs and
allele frequencies. Similar results have been published previ-
populations revealed no statistically significant differences in
distances were not as similar as expected. The lowest genetic
Namely, locus-by-locus comparisons and measured genetic
in a group of South-eastern European countries. Populations
iscent of the geographic map of Europe. At distinct south
the STR-based grouping of European populations was remin-
clustering of the examined populations based on the similar-
The Netherlands, Belgium, Denmark and Sweden formed

(Serbia, Montenegro, Bosnia and Herzegovina, and Croatia).

son between populations from former Yugoslavian Republics
The most surprising results were obtained after compari-
lysis. A two-dimensional plot was constructed to show the
ing Poland formed the third group and populations from
can be seen in Figure 2, the NJ dendrogram showed the

same territory of Kosovo. It should also be considered that

population might be due to the prominent ethnic isolation
populations compared not only to Serbs but also to all the
for ethnically close nations. Based on measured genetic dis-

other populations. This observed deviation from the Serbian

previously published population data (Novkovi
c et al. 2010)
eastern countries. Serbia, Bosnia and Herzegovina,
cluster of North-western European countries and Poland,
are Cyprus and Turkey, and at north Estonia and Latvia.

Kosovo Albanians and Italians might be some kind of

tances, Italians and Kosovo Albanians are the most distant
allele frequency distribution from the Montenegrin popula-
the genetic distance value of 0.014. Deviations of Serbian
distance was observed between Serbian and Bosnian popula-
(Estonia, Latvia, Belarus, Russia, and Ukraine) and neighbour-

south and north, somehow representing a region of

ities/differences among them. As can be seen in Figure 3,
Finally, we performed multidimensional scaling (MDS) ana-
Table 4. Nei genetic distance coefficients between 25 populations.
NW Greek
SRB BLR NLD LVA EST RUS BIH MNT KOS ALB HRV HUN AUT SWE POL CZE BEL ITA CPY TUR PRT ESP DNK UKR MDA ROM
SRB 0.006 0.007 0.02 0.016 0.006 0.002 0.014 0.055 0.008 0.003 0.011 0.01 0.007 0.005 0.009 0.07 0.01 0.008 0.018 0.008 0.008 0.008 0.003 0.004
BLR 0.008 0.015 0.013 0.004 0.002 0.017 0.063 0.007 0.003 0.011 0.009 0.004 0.004 0.01 0.071 0.015 0.012 0.019 0.009 0.008 0.006 0.005 0.007
NLD 0.021 0.016 0.009 0.005 0.019 0.064 0.008 0.005 0.01 0.005 0.008 0.005 0.005 0.071 0.014 0.01 0.015 0.007 0.004 0.012 0.006 0.009
LVA 0.023 0.018 0.017 0.033 0.08 0.02 0.017 0.025 0.022 0.018 0.017 0.022 0.089 0.032 0.027 0.032 0.022 0.022 0.02 0.019 0.022
EST 0.014 0.014 0.028 0.074 0.017 0.013 0.02 0.017 0.014 0.014 0.016 0.08 0.025 0.02 0.026 0.017 0.016 0.015 0.014 0.017
RUS 0.004 0.017 0.063 0.008 0.005 0.012 0.01 0.005 0.004 0.011 0.074 0.015 0.012 0.019 0.01 0.009 0.007 0.006 0.009
BIH 0.012 0.056 0.005 0.002 0.009 0.008 0.003 0.003 0.007 0.069 0.01 0.008 0.015 0.007 0.007 0.006 0.002 0.004
MNT 0.069 0.019 0.014 0.024 0.022 0.017 0.016 0.019 0.069 0.022 0.018 0.027 0.019 0.02 0.02 0.014 0.016
KOS 0.061 0.057 0.067 0.066 0.068 0.062 0.068 0.125 0.063 0.062 0.071 0.06 0.065 0.065 0.054 0.055
ALB
HRV 0.005 0.012 0.009 0.007 0.006 0.01 0.077 0.015 0.013 0.019 0.01 0.009 0.01 0.006 0.009
HUN 0.008 0.007 0.004 0.003 0.007 0.069 0.01 0.008 0.016 0.006 0.005 0.007 0.002 0.003
AUT 0.012 0.01 0.01 0.013 0.076 0.019 0.016 0.021 0.011 0.01 0.013 0.01 0.012
SWE 0.009 0.006 0.007 0.071 0.019 0.014 0.017 0.009 0.005 0.013 0.008 0.011
NW 0.005 0.009 0.074 0.016 0.014 0.018 0.009 0.008 0.007 0.006 0.01
POL
CZE 0.007 0.07 0.014 0.011 0.016 0.007 0.006 0.007 0.004 0.007
BLR 0.072 0.015 0.011 0.017 0.009 0.007 0.013 0.008 0.011
ITA 0.074 0.071 0.079 0.068 0.07 0.073 0.07 0.069
Greek 0.008 0.023 0.014 0.015 0.017 0.008 0.009
CPY
TUR 0.021 0.012 0.011 0.014 0.007 0.008
PRT 0.016 0.015 0.021 0.017 0.02
ESP 0.008 0.011 0.006 0.008
DNK 0.011 0.007 0.01
UKR 0.008 0.01
MDA 0.001
ANNALS OF HUMAN BIOLOGY
7
8 D. TAKIĆ MILADINOV ET AL.

Figure 2. Neighbor-joining tree showing the relationship of Serbian population and 24 others populations. The tree was constructed based on allele frequencies
for 12 autosomal STR loci shared among all populations (D21S11, D19S433, D7S820, D5S818, D3S1358, VWA, D8S1179, D16S539, TH01, D18S51, FGA,
and D13S317).

Figure 3. Multidimensional scaling plot of genetic distances between Serbian population and selected populations. Abbreviations for populations are the same as
in Figure 1.

exceptionally small publicly available samples (136 Albanians
and 72 Italians).
Comments
In summary, the study represents valuable genetic data on
22 autosomal STR loci in the Serbian population. The calcu-
lated forensic parameters demonstrate that the 22 analysed
loci are highly polymorphic and, in combination, can be
applied as a powerful tool for personal identification pur-
poses in paternity testing and forensics. Based on the gen-
etic distance values, it can be concluded that populations
differ according to geographic location and/or traditional
customs. The Serbian population is most similar to the
neighbouring Bosnian population and is quite different from
geographically distant countries such as Latvia and Estonia.
Observed differences from neighbouring Montenegrins and
Kosovo Albanians might be due to traditional customs, but
further research including comparisons with data from
enlarged samples is required to confirm our results.
Disclosure statement
No potential conflict of interest was reported by the author(s).
ORCID
Dijana Taki
c Miladinov http://orcid.org/0000-0003-4848-1918
Perica Vasiljevi
c http://orcid.org/0000-0003-0006-7187
Eva Podovsovnik Axelsson http://orcid.org/0000-0001-7449-7038
References
Butler JM. 2010. Fundamentals of forensic DNA typing. San Diego (CA):
Elsevier Academic Press. Statistical interpretation: evaluating the
strength of forensic DNA evidence; p. 229–258.
Butler JM. 2012. Advanced topics in forensic DNA typing: methodology.
San Diego (CA): Elsevier Academic Press.
Cariolou MA, Manoli P, Demetriou N, Bashiardes E, Karagrigoriou A,
Budowle B. 2006. Allele distribution of 15 STR loci used for human
identity purposes in the Greek Cypriot population of the island of
Cyprus. Forensic Sci Int. 164(1):75–78.
Cerri N, Franchi M, Mascadri S, De Ferrari F. 2003. Allele frequency distri-
bution of 13 STRs in an Italian and immigrant population sample. Int
Con Ser. 1239:137–143.


c G, Gasi
c V, Pluzari
c V, Smiljci
c D. 2012. Genetic parameters of five
Curi
new European Standard Set STR loci (D10S1248, D22S1045, D2S441,
D1S1656, D12S391) in the population of eastern Croatia. Croat Med J.
53(5):409–415.
Davidovic S, Malyarchuk B, Aleksic JM, Derenko M, Topalovic V, Litvinov
A, Stevanovic M, Kovacevic-Grujicic N. 2015. Mitochondrial DNA per-
spective of Serbian genetic diversity. Am J Phys Anthropol. 156(3):
449–465.
Decorte R, Engelen M, Larno L, Nelissen K, Gilissen A, Cassiman JJ. 2004.
Belgian population data for 15 STR loci (AmpFlSTR SGM Plus and
AmpFlSTR profiler PCR amplification kit). Forensic Sci Int. 139(2–3):
211–213.
Dupuy BM, Stenersen M, Kling D. 2013. Frequency data for 35 autosomal
STR markers in a Norwegian, an East African, an East Asian and
Middle Asian population and simulation of adequate database size.
Forensic Sci Int Genet Suppl. 4(1):e378–e379.
ANNALS OF HUMAN BIOLOGY 9
Excoffier L, Lischer HEL. 2010. Arlequin suite ver 3.5: a new series of pro-
grams to perform population genetics analyses under Linux and
Windows. Mol Ecol Resour. 10(3):564–567.
Gill P, Fereday L, Morling N, Schneider PM. 2006a. The evolution of DNA
databases- recommendations for new European STR loci. Forensic Sci
Int. 156(2–3):242–244.
Gill P, Fereday L, Morling N, Schneider PM. 2006b. New multiplexes for
Europe amendments and clarification of strategic development.
Forensic Sci Int. 163(1–2):155–157.
Gusmao L, Butler JM, Linacre A, Parson W, Roewer L, Schneider PM,
Carracedo A. 2017. Revised guidelines for the publication of genetic
population data. Forensic Sci Int Genet. 30:160–163.
Hammer Ø, Harper DAT, Ryan PD. 2001. PAST: paleontological statistics
software package for education and data analysis. Palaeontol
Electron. 4:9.
Hatzer-Grubwieser P, Berger B, Niederwieser D, Steinlechner M. 2012.
Allele frequencies and concordance study of 16 STR loci-including the
new European Standard Set (ESS) loci-in an Austrian population sam-
ple. Forensic Sci Int Genet. 6(1):e50–e51.
Hildebrand CE, Torney D, Wagner RP. 1992. Informativeness of
Polymorphic DNA Marker. Los Alamos Sci. 20:100–102.
Jemeljanova V, Gobrusjonoka O, Bergere O, Latisheva K, Podovsovnik
Axelsson E, Zupanic Pajnic I. 2015. Population data for 15 autosomal
STR loci from Latvia. Int J Legal Med. 129(4):739–740.
Jeran N, Havas D, Ivanovi
c V, Rudan P. 2007. Genetic diversity of 15 STR
loci in a population of Montenegro. Coll Antropol. 31:847–852.
Keckarevi
c D, Savi
c D, Keckarevi
c M, Stevanovi
c M, Tarasjev A, Culjkovi
c
B, Darmati A, et al. 2005. Population data on 14 STR loci from popula-
tion of Serbia and Montenegro (new and renewed data). Forensic Sci
Int. 151(2–3):315–316.
Kubat M, Skavi
c J, Behluli I, Nuraj B, Bekteshi T, Behluli M, Klari
c IM,
Perici
c M. 2004. Population genetics of the 15 AmpF lSTR identifiler
loci in Kosovo Albanians. Int J Legal Med. 118(2):115–118.
Lao O, Lu TT, Nothnagel M, Junge O, Freitag-Wolf S, Caliebe A,
Balascakova M, et al. 2008. Correlation between genetic and geo-
graphic structure in Europe. Curr Biol. 18(16):1241–1248.
Lopes V, Serra A, Gamero J, Sampaio L, Balsa F, Oliveira C, Batista L,
et al. 2009. Allelic frequency distribution of 17 STRs from identifiler
and PowerPlex-16 in Central Portugal area and the Azores archipel-
ago. Forensic Sci Int Genet. 4(1):e1–e7.
Montelius K, Karlsson AO, Holmlund G. 2008. STR data for the AmpFlSTR
Identifiler loci from Swedish population in comparison to European,
as well as with non-European population. Forensic Sci Int Genet. 2(3):
e49–e52.
Nei M. 1972. Genetic distance between populations. Am Nat. 106(949):
283–291.
Novkovi
c T, Pani
c B, Banjac A, Kovac D - eki
c T, Tomisi
c-Kosi
c I, Vuceti
c-
Dragovi
c A, Stamenkovi
c G, et al. 2010. Genetic polymorphisms of 15
AmpFlSTR Identifiler loci in a Serbian population. Forensic Sci Int.
4(5):e149–e150.
Paredes M, Crespillo M, Luque JA, Valverde JL. 2003. STR frequencies for
the PowerPlexV 16 system kit in a population from Northeast Spain.
R
Forensic Sci Int. 135(1):75–78.
Pereira V, Tomas C, Sanchez JJ, Syndercombe-Court D, Amorim A,
Gusm~ao L, Prata MJ, Morling N. 2015. The peopling of Greenland: fur-
ther insights from the analysis of genetic diversity using autosomal
and X-chromosomal markers. Eur J Hum Genet. 23(2):245–251.
Piatek J, Jacewicz R, Ossowski A, Parafiniuk M, Berent J. 2008. Population
genetics of 15 autosomal STR loci in the population of Pomorze
Zachodnie (NW Poland). Forensic Sci Int Genet. 2(3):e41–e43.
Pilav A, Pojski
c N, Ahatovi
c A, Dzehverovi
c M, Cakar  J, Marjanovi
c D.
2017. Allele frequencies of 15 STR loci in Bosnian and Herzegovinian
population. Croat Med J. 58(3):250–256.
Poetsch M, von Wurmb-Schwark N. 2013. Allele frequencies for the 16
short tandem repeats of the Powerplex ESX17 kit in a population
from Turkey. Int J Legal Med. 127(3):591–592.
Proji
c P, Skaro V, Samija I, Pojski
c N, Durmi
c-Pasi
c A, Kovacevi
c L, Bakal
N, et al. 2007. Allele frequencies for 15 short tandem repeat loci in
representative sample of Croatian population. Croat Med J. 48:
473–477.
10 D. TAKIĆ MILADINOV ET AL.
Rak SA,
Zal
an A, Szabados G, Pamjav H. 2011. Population genetic data
on 15 STR loci in the Hungarian population. Forensic Sci Int Genet.
5(5):543–544.
Ristow PG, D’Amato ME. 2017. Forensic statistics analysis toolbox
(FORSTAT): a streamlined workflow for forensic statistics. Forensic Sci
Int Genet Suppl Ser. 6:e52–e54.
Ross J, Parson W, Fura
c I, Kubat M, Holland M. 2001. Multiplex PCR amp-
lification of eight STR loci in Austrian and Croatian Caucasian popula-
tions. Int J Legal Med. 115(1):57–60.
Sadam M, Tasa G, Tiidla A, Lang A, Podovsovnik Axelsson E, Zupanic
Pajnic I. 2015. Population data for 22 autosomal STR loci from
Estonia. Int J Legal Med. 129(6):1219–1220.
Serga SV, Dombrovskyi IV, Maistrenko OM, Ostapchenko LI, Demydov SV,
Krivda RG, Kozeretska IA. 2017. Allele frequencies for 15 STR loci in
the Ukrainian population. Forensic Sci Int Genet. 29:e40–e41.
Simkov
a H, Faltus V, Marvan R, Pexa T, Stenzl V, Broucek J, Hor
ınek A,
et al. 2009. Allele frequency data for 17 short tandem repeats in a
Czech population sample. Forensic Sci Int Genet. 4(1):e15–e17.
Stanciu F, Popescu OR, Stoian IM. 2009. Allele frequencies of 15 STR
loci in Moldavia region (NE Romania). Forensic Sci Int Genet. 4(1):
e39–e40.
View publication stats
Stanciu F, Stoian IM, Popescu OR. 2009. Population data for 15 short tan-
dem repeat loci from Wallachia region, South Romania. Croat Med J.
50(3):321–325.
Takezaki N, Nei M, Tamura K. 2014. POPTREEW: web version of POPTREE
for constructing population trees from allele frequency data and com-
puting some other quantities. Mol Biol Evol. 31(6):1622–1624.
Veselinovi
c I, Kubat M, Furac I, 
Skavi
c J, Martinovi
c Klari
c I, Tasi
c M.
2004. Allele frequencies of the 15 AmpF lSTR Identifiler loci in the
population of Vojvodina Province, Serbia and Montenegro. Int J Legal
Med. 118(3):184–186.
Westen AA, Kraaijenbrink T, Robles de Medina EA, Harteveld J, Willemse
P, Zuniga SB, van der Gaag KJ, et al. 2014. Comparing six commercial
autosomal STR kits in a large Dutch population sample. Forensic Sci
Int Genet. 10:55–63.
Zhivotovsky LA, Malyarchuk BA, Derenko MV, Wozniak M, Grzybowsk T.
2009. Developing STR databases on structured populations: the native
South Siberian population versus the Russian population. Forensic Sci
Int Genet. 3(4):e111–e116.
Zhivotovsky LA, Veremeichyk VM, Kuzub NN, Atramentova LA, Udina IG,
Kartel NA, Tsybovsky IS. 2009. A reference data base on STR allele fre-
quencies in the Belarus population developed from paternity cases.
Forensic Sci Int Genet. 3(3):e107–e109.