Coll. Antropol. 24 (2000) 2: 267–279 UDC 572:575.116(497.5) Original scientific paper MtDNA Haplogroups in the Populations of Croatian Adriatic Islands H. V. Tolk1, M. Peri~i}2, L. Bara}2, I. Martinovi} Klari}2, B. Jani}ijevi}2, I. Rudan3, J. Parik1, R. Villems1 and P. Rudan2 1 Department of Evolutionary Biology – Institute for Molecular and Cell Biology, University of Tartu, Tartu, Estonia 2 Institute for Anthropological Research, Zagreb, Croatia 3 »Andrija [tampar« School of Public Health, Faculty of Medicine, University of Zagreb, Zagreb, Croatia ABSTRACT The number of previous anthropological studies pointed to very complex ethnohis- torical processes that shaped the current genetic structure of Croatian island isolates. The scope of this study was limited to the general insight into their founding popula- tions and the overall level of genetic diversity based on the study mtDNA variation. A to- tal of 444 randomly chosen adult individuals from 32 rural communities of the islands of Krk, Bra~, Hvar and Kor~ula were sampled. MtDNA HVS-I region together with RFLP sites diagnostic for main Eurasian and African mtDNA haplogroups were ana- lysed in order to determine the haplogroup structure. The most frequent haplogroups were »H« (27.8–60.2%), »U« (10.2–24.1%), »J« (6.1–9.0%) and »T« (5.1–13.9%), which is similar to the other European and Near Eastern populations. The genetic drift could have been important aspect in history, as there were examples of excess frequencies of certain haplogroups (11.3% of »I« and 7.5% of »W« in Krk, 10.5% of »HV« in Bra~, 13.9% of »J« in Hvar and 60.2% of »H« in Kor~ula). As the settlements on the islands were formed trough several immigratory episodes of genetically distant populations, this analysis (performed at the level of entire islands) showed greater genetic diversity (0.940–0.972) than expected at the level of particular settlements. Introduction are among the best characterised human isolate resources in the world. As a result The four islands in eastern Adriatic, of the continuous anthropological research Croatia (Krk, Bra~, Hvar and Kor~ula) during the past quarter of century, more Received for publication November 29, 2000. 267 H. V. Tolk et al.: MtDNA Haplogroups in Croatians, Coll. Antropol. 24 (2000) 2: 267–279 than 100 publications refering to these very valuable in the emerging field of mo- populations have been recorded in the in- lecular anthropology due to the three ternational biomedical and anthropologi- important properties of mtDNA: high cal databases to date. The strategy of the mutation rate35, strictly maternal inhe- research was set by Rudan1–3, who recon- ritance36 and the absence of recombina- structed the ethnohistory and demogra- tion. Due to maternal inheritance, mito- phy and analysed the linguistic pecula- chondrial genome as a single locus has rities of these populations with Sujold`i} four times smaller effective population et al.5–7. In addition, the detailed anthro- size than nuclear autosomal loci. This in- pometrical measurements were carried creases the sensitivity of mtDNA diver- out by Rudan et al.8, physiological mea- sity to fluctuations in population size and surements were performed by Smolej- the effect of genetic drift in small isolated -Naran~i} et al.9,10, dermatoglyphics stud- populations. Most studies conducted so ied by Rudan11 and Mili~i} et al.12,13, and far using mtDNA can be considered as the migrational analyses carried out by macro-geographical in their approach (out- Jovanovi} et al.14,15 and [poljar-Vr`ina et -of-Africa, peopling of Americas, Europe al.16–18. Using the obtained data, the pop- etc.) while the micro-geographic studies ulation structure in the studied region are still quite exceptional and only start- was assessed through the holistic appro- ing to emerge37, 38. According to numerous ach relying on the model-bound me- previous studies, human mtDNA varia- thods19,20, the model-free methods21–24 and tion is highly region-specific, giving a original experimental designs 25. good background for more detailed stud- The insight into population structure ies on isolated populations. of these islands has prompted further This paper presents the very first population genetic research. The initial mtDNA study on Croatian population in results were obtained through isonymy order to describe the mtDNA structure in studies26,27 and the analyses of assorta- the context of European haplogroups pro- tive mate choice28. These simple appro- viding an additional explanation to previ- aches were followed by blood sampling ous research conducted on the popula- and the analysis of the distribution of tions of the Adriatic islands. The aim of allelic frequencies of some basic seroge- this study is to determine population ge- netic polymorphysms in collaboration with netic structure in isolated rural popula- the laboratory in Newcastle upon Tyne, tions of four Adriatic islands (Krk, Bra~, UK. The most important findings were Hvar, and Kor~ula) using mtDNA varia- published by Jani}ijevi} et al.29 and Rob- tion, and to find out is there any »non- erts et al.30. Following the emergence of -Caucasoid« mtDNA types as well as to molecular genetic technology, the rese- study the diversity levels. arch has been extended to include the analysis of STR and VNTR DNA poly- morphysms in the papers by Martinovi} Brief genetic history of the studied et al.31–34. populations Recently, the collaboration was estab- In terms of the genetic history of these lished between the Institute of Anthropo- four island populations, the most impor- logical Research in Zagreb, Croatia, and tant characteristic that needs to be em- the Institute of Molecular and Cell Biol- phasised is that the current populations ogy in Tartu, Estonia to study the charac- were formed through several immigra- teristics of mtDNA in Croatian island po- tory episodes of genetically quite distant pulations. This type of study has become populations. 268 H. V. Tolk et al.: MtDNA Haplogroups in Croatians, Coll. Antropol. 24 (2000) 2: 267–279 The archeological evidence recently were influencing the shaping of the cur- reviewed by Forenbaher39 implies that rent genetic structure was recently re- the first settlers of the eastern Adriatic viewed by Rudan et al.40. It is important islands were Mediterranean non-Indo- to note that the founder unit of the popu- -European people who probably came lation is a settlement (village) rather than from the south of today’s Italy and were each island itself, and that any study later replaced by Indo-European Illyri- analysing the diversity at the level of en- ans. The oldest archaeological evidence of tire island populations, such as the pres- human settlements on the Dalmatian is- ent one, is expected to reveal far more di- lands date back to Neolithic, approxima- versity than it would be expected within tely 5,000 years before present (YBP). the single villages. From about the 10th century BC onward the Illyrians ruled the entire region of eastern Adriatic coast. Sample and methods Subsequent migratory episodes in the A) Sample early B.C. period brought Greeks (4th cen- As the scope of this research was lim- tury B.C.) and then Romans (2nd century ited to performing the general overview B.C. onward). Romans organised their of the haplogroups in the populations of settlements at the coastline of the is- interest, and bearing in mind that the lands, as they were used as harbours on previous analyses of the other world pop- the trade routes from the western Adri- ulations were based on very small sample atic coast to Greece, Middle East and sizes, we decided to include 15–20 adult Northern Africa. The extent of the Ro- persons from each village found on each man admixture with preexisting Illyrians of these four islands. The examinees were is difficult to estimate. then randomly chosen from parish regis- The first great influx of Croats onto tries in those villages. The final sample the Adriatic islands occurred between 6th size was 105 in Bra~ island, 108 in Hvar and 8th century. The Croats gradually island, 98 in Kor~ula island and 133 in Slavicised the islands, but the extent of Krk island, i.e. the total of 444 individu- their genetic admixture with Romans, als from 32 rural communities (Figure 1). Greeks and Illyrians is unknown. It is important to emphasise that the sampling was not based on the likely The second large immigration wave founder populations (villages) but merely resulted from migrations from the main- on geography (islands) and that the se- land of the Balkans peninsula during the lected individuals therefore constitute a expansion of the Ottoman empire. The very heterogenous group with respect to greatest influx of the immigrants occur- possible founders. The blood samples th red in the 17 century during the Can- were collected during the field research dian and Morean wars. The Croatian im- by the staff of the Institute for Anthropo- migrants brought their own customs and logical Research in Zagreb, Croatia in the gene pool, but the settlements they foun- late 1980’s and early 1990’s following the ded remained isolated from those of the appropriate approval of the ethical com- preexisting settlers throughout the cen- mitee on this institution. All the subse- turies due the various political, cultural quent molecular analysis of the mtDNA and legal barriers to intermarriage im- was performed in the Estonian Biocentre 19 posed by the governors . and Department of Evolutionary Biology The detailed ethnohistory of these po- at the Institute of Molecular and Cell Bi- pulations and the historic events that ology, University of Tartu, Estonia. 269 H.