Increased Sample Size Proved More Insights for the Population Structure of Mediterranean Loggerhead Sea Turtles

Increased Sample Size Proved More Insights for the Population Structure of Mediterranean Loggerhead Sea Turtles | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Increased Sample Size Proved More Insights for the Population Structure of Mediterranean Loggerhead Sea Turtles Arzu Kaska, Gizem Koç, Dogan Sözbilen, Salih Diryaq, Ashraf Glidan, and 23 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-1649861/v2 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 13 May, 2023 Read the published version in Conservation Genetics Resources → Version 2 posted 7 You are reading this latest preprint version Show more versions Abstract The loggerhead sea turtle ( Caretta caretta ), has the broadest distribution among sea turtle species in the Mediterranean and requires regional and international collaborations in addition to local efforts to better inform conservation actions. Molecular techniques are powerful tools to assess population dynamics at large scales, especially by determining the connectivity among different nesting and foraging sites, and genetic diversity. In this study, a large sample was collected synchronously in the nesting areas located in the north, south and east of the Mediterranean. Recently described nesting sites from Albania and other nesting sites represented by lower sample size were also included in order to fully assess the genetic composition of the region’s rookeries. Samples from 710 individuals were collected and the longer (815 bp) mtDNA D-loop fragment of these samples was amplified. We recorded 15 haplotypes, three of which were novel. In addition, our results show that some haplotypes, considered of Atlantic origin, have a wider dispersal in the Mediterranean than previously thought, albeit with low levels of representation. Our results, which also contribute to determining the likely origin of haplotypes that were previously known only from foraging sites, highlight the utility of broad-scale sampling, with increased sample number and longer mtDNA sequence to determine genetic diversity and connectivity. This study also demonstrates that it is important to continue to monitor the contribution of Atlantic origin haplotypes to the Mediterranean population, and the resident Mediterranean population, which is expected to expand its geographical range for reproduction with the effect of climate change and climate change in the long term. This work is important for, among other things, mixed stock analyses (MSA) that seek to localize the origin of stranded or accidentally caught sea turtles or those purposefully obtained from foraging sites to better understand the migratory distribution for conservation purposes. Caretta caretta Genetic structuring mtDNA Phylogeography Conservation Figures Figure 1 1. Introduction The loggerhead turtle ( Caretta caretta ) is the most common sea turtle species in the Mediterranean Sea (Casale et al., 2018 ). Nesting occurs mainly in the Eastern Mediterranean basin, with the highest number of clutches in Greece, Turkey, Libya and Cyprus (Casale et al., 2018 ), with lower nesting numbers in Egypt, Israel, Italy, Lebanon, Syria and Tunisia. Minor nesting also occurs in the western basin in Malta, Albania, Spain, France and Italy (Casale et al., 2018 ). Globally, sea turtle populations are under many anthropogenic pressures. However, as a result of conservation efforts, the Mediterranean loggerhead population has recovered substantially and is currently classified as Least Concern by the International Union for Conservation of Nature (IUCN) due to increasing nest numbers of the species at major nesting sites of the Mediterranean Sea. Furthermore, it has been considered as Conservation Depended since the threats still exist (Casale, 2015). It is worth noting that the loggerhead turtle is a highly mobile species and migrates between nesting and foraging sites. Recent studies showed that the Mediterranean loggerhead turtle population represents a wide dispersal from nesting sites to different foraging sites (Cerritelli et al. , 2022, Haywood et al. , 2020, Schofield et al., 2010 ). Despite conservation efforts at the local levels, studies based on international cooperation and the determination of area-based conservation measures have been adopted recently (Kot et al. , 2022). Consequently, this step will provide high benefits for sea turtle conservation studies. Sea turtles exhibit a behavior known as philopatry, defined as a tendency to return to their natal beaches (Bowen and Karl, 2007). Accordingly, this behavior results in certain regions forming populations with specific genetic structures over time. Those populations are defined as Management Units (Moritz, 1994 ). The Mediterranean loggerhead turtle population is identified as one of the 10 Regional Management Units (RMU) based on studies of nesting sites, population abundances and trends, population genetics, and satellite telemetry (Wallace et al. , 2010). Although the Mediterranean loggerhead turtle population is defined as a single RMU, there is genetic diversity in both nesting and foraging areas at haplotype level (Carreras et al. , 2006, Yilmaz et al. , 2011, Saied et al., 2012 , Carreras et al., 2014 , Clusa et al., 2014 , Rees et al., 2017 , Clusa et al. , 2018, Tolve et al., 2018 ). However, the Mediterranean loggerhead meta-population has been proved to be composed of three independent RMUs (1 Mediterranean RMU and 2 Atlantic RMUs) (Casale et al., 2018 ). Recently, loggerhead sporadic nesting events have become frequent in the western Mediterranean, and the Atlantic RMUs contributed to these nesting events (Carreras et al. , 2019). Therefore, genetic diversity may differ between foraging and nesting sites. Mixed-stock analysis (MSA) based on mitochondrial DNA (mtDNA) sequences are used to assess haplotype diversity among different nesting sites and estimate the origins of individuals at foraging sites (Bowen et al. , 2007). It has been suggested that the knowledge about the metapopulation structure may have been obtained via available genetic markers (Casale et al., 2018 ). However, due to unsampled breeding sites and incomplete sampling of large, well-known sites, complete genetic characterization was impossible for the Mediterranean. Furthermore, mainly when “orphaned haplotypes” exist (i.e. haplotypes recorded from the population but not reported at any breeding site), the MSA has low power in some foraging sites (Tolve et al., 2018 ). Therefore, to have more comprehensive knowledge on the loggerhead turtle population structure, there is a need for simultaneous studies with an increased number of samples from areas that were previously represented with few samples or were not sampled. In this context, and with the contribution of researchers from seven countries in the Mediterranean, and to have a more robust knowledge, this study aims to answer the following questions: Did we reach the limit on the knowledge about the loggerhead turtle population structure? Is there further strong connectivity among different breeding sites? Do the Atlantic loggerhead turtles contribute to the breeding population in the eastern Mediterranean? What is the origin of the individuals at new nesting sites in the Mediterranean? Do studies with low sample numbers mask the extent of genetic diversity in the Mediterranean? Finally, increasing the sample size of rookeries from different countries contributes to better understanding of the genetic structure of Caretta caretta from Mediterranean rookeries. 2. Material And Method Sample collection A total of 710 samples were collected between 2018 and 2019 along the beaches of Cyprus, Libya, Lebanon, Tunisia, Greece, and Turkey. The sites from Libya and Lebanon have not been previously sampled, the other samples were to replace the findings for them with a larger sample size. Samples from Albania and Egypt were included to identify their haplotypes in the analyses. Skin biopsies were taken from adult females, and hatchlings were preserved in 70% alcohol until further laboratory procedures by researchers in each country. Only nests laid within 15 days, which is the optimum nesting interval for the loggerhead turtles, were used for the genetic analysis to prevent the risk of pseudo-replication. The sample size in Turkey was 384, Libya 134, Greece 104, Tunisia 41, Cyprus 27, Albania and Lebanon eight. Six Albanian samples were collected from live specimens of Caretta caretta captured as bycatch at Ishmi stavnik, Patok area (Drini bay), while the other two were collected from two dead Caretta caretta hatchlings from the first officially documented nest in Albania in Divjaka beach area, in 2018 (Piroli and Haxhi, 2020). Albanian samples and two samples from Egypt were included as non-nesting individuals to show also a case for mixed stock analysis can be performed for strandings. Laboratory analysis The DNA extraction was performed using a standard phenol–chloroform protocol (Kaska et al., 2001 ) or a QuickGene DNA tissue kit (KURABO). Representatives of each research team trained for the same protocol for extraction of DNA and PCR amplification at DEKAMER Lab in Turkey. An approximately 815 base pair (bp) long fragment of the non-coding mitochondrial DNA (mtDNA) control region was amplified by Polymerase Chain Reaction (PCR). The primer pair used was LCM15382 (5′-GCTT AACCCTAAGCATTGG-3′) and H950 (5′-GTCTCGG ATTTAGGGGTTT-3′) (Abreu-Grobois et al. 2006 ). Polymerase Chain Reaction (PCR) was performed in a total volume of 30-µL mastermix, 0.5 µM of each primer, and 2 µL of DNA. Thermal conditions consisted of an initial denaturation at 95°C for 3 minutes (min), followed by 34 cycles of 30 seconds (s) at 95°C, 1 min at 55°C and 30 s at 72°C, with a final extension step at 72°C for 10 minutes. PCR products were visualised on a 1% agarose gel stained with Safeview™ for amplification evaluation. After completing the DNA extraction and the successful PCR products by each partner, the PCR products were sent to Genartek (Istanbul, Turkey) for DNA purification and Sanger sequencing with both forward and reverse primers. The obtained sequences were analysed by researchers from DEKAMER Lab, Turkey. Data analysis Obtained sequences were, if needed, edited in Chromas (v.2.6.6) and aligned in Bioedit (v.7.2.5) (Hall, 1999 ). Haplotype classification was conducted through comparing the sequences with haplotypes already presented in the Archie Carr Center for Sea Turtle Research database (ACCSTR; http://accstr.uf.edu/fles/cclongmtdn a.pdf ) and the sequence comparison tool, GenBank BLAST ( http://ncbi.nlm.nih.gov/Blast.cgi ). Novel haplotypes found were submitted to ACCSTR for assigning the international nomenclature and their sequences were sent to GenBank. To highlight the relationship between different haplotypes identified, haplotype networks based on the median-joining algorithm were created using the program POPART version 1.7 (Leigh and Bryant 2015 ). Polymorphism data was obtained by estimating the haplotype diversity (h), nucleotide diversity (π), number of haplotypes (k) and number of variable sites (p) in DnaSP version 5.10.01 (Librado and Rozas 2009 ). Haplotype networks of mtDNA for loggerhead turtles from different Mediterranean countries were created with the connecting lines between haplotypes representing single mutations. The circle areas of the haplotypes are proportional to the sample size carrying the specific haplotype were also shown in Fig. 1 . The software MEGA v. 7 (Kumar et al. 2016 ) was used to create a phylogenetic tree (Supplement, Fig. 2). The best-fit substitution model (ML) was chosen on the basis of the lowest Bayesian information criterion (BIC) value which was the Tamura 3-parameter with Gamma distribution (T92 + G). For the phylogenetic analysis the neighbour-joining (NJ) method was used including the selected substitution model. To obtain valid results for the nodes the number of bootstrap replications were 1000, bootstrap values greater than or equal to 50% were considered statistically significant (Margush & McMorris 1981 ). For node calibration, D-loop sequences from two olive ridleys ( Lepidochelys olivacea , GenBank AM258984 and JX454991) and Kemp’s ridley ( Lepidochelys kempii , GenBank JX454981) were included in the alignment as outgroups. 3. Results Among the 710 mtDNA sequences analysed 14 variable sites were observed defining 16 haplotypes of which three were previously undescribed. The haplotypes found in the literature and found in this study were presented in Table 1 . Both, CC-A2.1 and CC-A3.1 are the most common haplotypes at different Mediterranean rookeries. The highest number of haplotypes were found in Libya (9), followed by Greece (6) and Turkey (5) (see Table 2 ). The haplotype diversity was highest in Libya-Tunisia Management Unit, followed by western Turkey and Crete, Greece. The most frequently found haplotypes were CC-A2.1 (67.0%) and CC-A3.1 (19.8%), both present at all nesting sites, followed by CC-A2.9 (6.7%) most found in Libya and with 3.2% haplotype CC-A26.1, also originating from Libya. The genetic polymorphism measures data (Table 2 ) and frequencies of haplotypes (Table 3 ) were given according to Management Units. Table 1 Number (n) of Caretta caretta haplotypes sampled at different Mediterranean rookeries and their rookery of origin (Clusa et al.2014). Haplotype n Source rookeries found in this study Rookery CC-A2.1 468 DLM (7), DLY (83), TKE (46), TKM (35), TKW (81), MIS (4), SIR (49), CRT (22), KOR (18), KOT (8), LAK (26), ZAK (19), TUN (39), CYP (26), LEB (5) Mediterranean: MIS, SIR, ISR, LEB, CYP, ETU, MTU, DLM, DLY, CRE, WGR, CAL Atlantic: CEF, SEF, SAL, DRT, QMX, SWF, CWF, NWF, CPV CC-A2.8 5 CRT (4), LAK (1) Mediterranean: CRE Atlantic: – CC-A2.9 47 SIR (45), MIS (1), TUN (1) Mediterranean: MIS, SIR, ISR Atlantic: – CC- A3.1 138 DLM (36), DLY (55), TKE (6), TKW (30) (128), SIR (4), CRT (1), TUN (1), CYP (1), LEB(3) Mediterranean: MIS, SIR, LEB, ETU, WTU, WGR, DLM, DLY Atlantic: CEF, SEF, SAL, QMX, SWF, CWF, NWF CC-A6.1 2 CRT (1), KOT (1) Mediterranean: WGR Atlantic: – CC- A10.4 1 MIS (1) Mediterranean: Tuscany a , Campania a Atlantic: CEF CC-A26.1 22 MIS (1), SIR (21) Mediterranean: SIR Atlantic: – CC-A29.1 1 TKW (1) Mediterranean: ISR Atlantic: – CC-31.1 2 CRT (1), KOR (1) Mediterranean: CAL, WGR, Sicily a Atlantic: – CC-A32.1 1 CRT (1) Mediterranean: WGR Atlantic: – CC-A50.1 1 TKE (1) Mediterranean: CYP CC-A68.1 2 SIR (2) Mediterranean: SIR CC-A71.1 1 SIR (1) Unknown CC-A77.1 3 SIR (3) Novel haplotype CC-A2.16 2 SIR (2) Novel haplotype CC-A3.4 2 DLY (2) Novel haplotype Mediterranean rookeries: MIS (Misurata, Libya), SIR (Sirte, Libya), ISR (Israel), LEB (Lebanon), CYP (Cyprus), ETU (Eastern Turkey), MTU (Middle Turkey), WTU (Western Turkey), DLM (Dalaman, Turkey), DLY( Dalyan, Turkey), CRE and CRT (Crete, Greece), WGR (Western Greece), KOR (Koroni, Western Greece), KOT (Kotychi, Western Greece), CAL (Calabria, Italy). Table 2 Genetic polymorphism measures of loggerhead sea turtles from different management units. Management unit n k p S Hd π WGRC 74 4 3 3 0.0800 0.00007 CRT 30 6 4 4 0.4552 0.00053 DLYDLM 184 3 2 2 0.5132 0.00064 TKW 112 3 2 2 0.4088 0.00051 LIBY_TUN 175 9 8 8 0.6362 0.00107 EMED 123 3 2 2 0.1655 0.00020 Total_Med 698 16 14 14 0.5077 0.00078 n number of turtles sampled, k number of haplotypes, p number of polymorphic (segregation) sites, S Number of variable sites, Hd haplotype diversity, π nucleotide diversity Table 3. Haplotype number and frequencies (%) found in different Management Units. Haplotype WGRC CRT DLYDAL TKW LIBYTUN EMED Overall CC-A2.1 71 (95.9%) 22 (73.3%) 90 (48.9%) 81 (72.3%) 92 (52.6%) 112 (91.1%) 468 (67.0%) CC-A2.8 1 (1.4%) 4 (13.3%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 5 (0.7%) CC-A2.9 0 (0.0%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 47 (26.9%) 0 (0.0%) 47 (6.7%) CC-A3.1 0 (0.0%) 1 (3.3%) 92 (50.0%) 30 (26.8%) 5 (2.9%) 10 (8.1%) 138 (19.8%) CC-A6.1 1 (1.4%) 1 (3.3%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 2 (0.3%) CC-A10.4 0 (0.0%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 1 (0.6%) 0 (0.0%) 1 (0.1%) CC-A26.1 0 (0.0%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 22 (12.6%) 0 (0.0%) 22 (3.2%) CC-A29.1 0 (0.0%) 0 (0.0%) 0 (0.0%) 1 (0.9%) 0 (0.0%) 0 (0.0%) 1 (0.1%) CC-31.1 1 (1.4%) 1 (3.3%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 2 (0.3%) CC-A32.1 0 (0.0%) 1 (3.3%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 1 (0.1%) CC-A50.1 0 (0.0%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 1 (0.8%) 1 (0.1%) CC-A68.1 0 (0.0%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 2 (1.1%) 0 (0.0%) 2 (0.3%) CC-A71.1 0 (0.0%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 1 (0.6%) 0 (0.0%) 1 (0.1%) CC-A77.1 0 (0.0%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 3 (1.7%) 0 (0.0%) 3 (0.4%) CC-A2.16 0 (0.0%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 2 (1.1%) 0 (0.0%) 2 (0.3%) CC-A3.4 0 (0.0%) 0 (0.0%) 2 (1.1%) 0 (0.0%) 0 (0.0%) 0 (0.0%) 2 (0.3%) One new haplotype detected in Sirte, Libya differed by one substitution from CC-A2.9 which is an exclusively Mediterranean haplotype (Tolve et al. 2018 ) and common in the Libyan nesting sites (Splendiani et al. 2017 ). The novel sequence was named CC-A77.1 after the nomenclature rules published on the Archie Carr Center for Sea Turtle Research (ACCSTR) website. The second new haplotype identified at a nesting site in Sirte, Libya differed one substitution from CC-A2.1, which is found in the Mediterranean and Atlantic populations (Shamblin et al. 2014 ) and was named CC-A2.16. The third previously undiscovered haplotype, namely CC-A3.4, was detected at a Turkish nesting site (Dalyan) and is one substitution separated from CC-A3.1, which is also found both in the Mediterranean and the Atlantic (Shamblin et al. 2014 ). All novel haplotypes were deposited in ACCSTR and Blast. The phylogenetic tree of C. caretta generated with the NJ method clearly shows the two different mitochondrial lineages (known as Haplogroup I and Haplogroup II) highlighted in Shamblin et al. ( 2014 ). The three new haplotypes (CC-A2.16, CC-A3.4 and CC-A77.1) are representatives of Haplogroup II. The haplotype network of mtDNA for loggerhead turtles from different Mediterranean management units were also shown in Fig. 1 , the phylogenetic trees were also provided (Supplement, Fig. 2). 4. Discussion Molecular genetic investigation for differentiated populations has been characterised as a powerful tool for conservation purposes (Crandall et al. 2000 ; Moritz, 1994 ). However, length of the sequences, selected markers and sample size play an important role in differentiating populations (Monzón-Argüello et al. 2010 ; Clusa et al. 2013 ). Using extended mtDNA 815bp haplotype sequences, even independent management units (MUs) have been identified within the Mediterranean RMU (Shamblin et al. 2014 ), namely: (i) Calabria-Italy (CAL), (ii) western Greece (WGRC), (iii) Crete-Greece (CRT), (iv) Libya (LIBY), (v) Dalyan-Dalaman-Turkey (DLYDAL), (vi) western Turkey (TKW), (vii) eastern Mediterranean (EMED, including middle and eastern Turkey, Cyprus, Israel, and Lebanon). According to our knowledge so far, the highest diversity with seven haplotypes was reported from nesting sites was Turkey (Yilmaz et al. 2011, Carreras et al. 2014 , Clusa et al. 2014 and references therein). It is followed by Libya with five haplotypes (Said et al. 2012), Greece with four haplotypes (Yilmaz et al. 2011, Carreras et al. 2014 ), Cyprus (Clusa et al. 2013 ) and Lebanon (Yilmaz et al. 2011) with two haplotypes, and Tunisia with one haplotype (Chaieb et al. 2010 ). Although recent sporadic loggerhead turtle nest records were available from Albania (Piroli and Haxhi, 2020), genetic characterization was not available to date. Libyan coasts and information about the nesting sites are identified as the major knowledge gap in the Mediterranean (Casale et al. 2018 ). In addition, Libya is a significant place as it is the area where the loggerhead turtle first colonised in the Mediterranean (Clusa et al. 2013 ). The results show that more effort needs to be put into sampling, especially along the largely unexplored Libyan coasts as two novel haplotypes have been found in this area. In total nine haplotypes were found. The second most frequent haplotype found in Misratah and Sirte, Libya was CC-A2.9 which occurs frequently in Israel and Libyan rookeries (Saied et al. 2012 ; Clusa et al. 2014 ). One specimen from a nesting site in Misratah, Libya was detected carrying the rare haplotype CC-A10.4. This haplotype was previously only recorded occasionally at Tyrrhenian nesting sites (Garofalo et al. 2016a ; Mafucci et al. 2016 ). It derives from the CC-A10 haplotype (380 bp mtDNA sequence) which is previously observed only once in Greece (Laurent et al. 1998 ). For CC-A10.4 it was also predicted that this haplotype probably originated not only from Tyrrhenian rookeries but also from Mediterranean colonies (Tolve et al. 2018 ). The origin of the specimens found in the Adriatic (Yilmaz et al. 2012 , Tolve et al. 2018 , Bertuccio et al. 2019 ) could be in Libya. Both haplotypes, CC-A29.1 and CC-A10.4 were only found once in Turkey and Libya, respectively which is why the sampling size in these rookeries needs to be increased. Another very rare haplotype is CC-A68.1 which was found only once at the nesting location in Sirte, Libya in 2009. In this study two individuals carried CC-A68.1 from Sirte, Libya. Splendiani et al. ( 2017 ), who used samples of rescued Caretta caretta found along the Southern Adriatic coast of Italy, described the haplotype CC-A71.1 for the first time which is why they were not able to determine the origin of it. However, by creating a phylogenetic tree it was possible for them to see the phylogenetic relatedness to CC-A26.1 which is exclusive to Libya (Shamblin et al. 2014 ). For this reason, Splendiani et al. ( 2017 ) suggested that CC-A71.1 could be from a Libyan rookery. Our findings support this proposition since one specimen from Sirte, Libya carrying this haplotype was detected. The 104 samples analysed from Greece, including the Management Unit of Western Greece and Crete, included all known haplotypes in these rookeries. In Western Greece 71 (95.9%) and in Crete 22 (73.3%) of the loggerheads carried the haplotype CC-A2.1. Four individuals from Chania, Crete carried CC-A2.8 while one individual from Mavrovouni, Lakonikos Bay carried this haplotype, which was known to be endemic to the Cretan rookery. One individual from Crete carried a haplotype CC-A3.1, another one CC-A32.1. Haplotype CC-A6.1 was carried by loggerheads from Kotychi and Crete. Both haplotypes, CC-A6.1 and CC-A32.1 were known to be endemic to Western Greece. One specimen from Crete and one from Koroni had haplotype CC-A31.1 originating from Greek and Calabrian rookeries but also found in sporadic Sicilian nesting sites (Garofalo et al. 2016 b). In this study in total five haplotypes were detected at main Turkish nesting sites of which two individuals belong to the same, new haplotype. The most frequent haplotype in Turkey, which is also the most common in the Mediterranean in general, was CC-A2.1 followed by CC-A3.1 also a common haplotype. The discovery of the CC-A29.1 haplotype at a nesting beach in Turkey, more precisely in western Turkey (Çıralı) shows, as has been proposed before (Tolve et al. 2018 ), that the origin of this haplotype must not only be in Israel, but also in other, poorly sampled or unknown nesting sites. This finding reinforces the results of MSA from the Adriatic Sea (Tolve et al. 2018 ): Western Turkish rookeries, which despite the fact that they are much more abundant and closer to the Adriatic Sea, showed a medium contribution probability to the Adriatic stock (Tolve et al. 2018 ). Two samples from Slovenia (northern Adriatic) carried the haplotype CC-A29.1. If CC-A29.1 would also be included as a haplotype of Western Turkish origin, the posterior probability would probably increase, and the contribution would not be similar or smaller than the ones of Israeli nesting areas. Haplotype CC-A50.1 is another very rare haplotype which was identified for the first time in Cyprus (Clusa et al. 2013 ). We are not aware that this haplotype was detected again afterwards. One specimen from a Turkish rookery (Kazanlı) carried this haplotype. Turkey is not only a foraging site for loggerhead sea turtles originating from Cyprus but also a new nesting site. A review of general migratory routes of 63 adult loggerhead turtles released mainly from Greece and Cyprus showed that only a few of them oriented to Turkish coasts (Luschi and Casale, 2014). The tracking studies from Northern Cyprus showed that only early nesters visited other Turkish rookeries (Snape et al. 2016). MSA of an eastern Turkish foraging ground made by Türkozan et al. (2018) showed a local contribution (62%) of Cyprus to the western subdivision. For Tunisian rookeries (Kuriat islands), only the short mtDNA control region fragment (500 bp) has been used so far, and only CC-A2.1 has been found (Chaieb et al. 2010 ). Analysis using the short fragment have shown no significant differences between Libyan and Tunisian rookeries, but due to several hundred kilometres of separation Shamblin et al. ( 2014 ) suggested that these might be demographically isolated nesting populations. A distinction, with the shorter D-loop sequence, between the haplotype CC-A2.9 and the widely distributed haplotype CC-A2.1 is not possible (Splendiani et al. 2017 ), and given the fact that CC-A2.9 is common in Libya, it has been proposed that reanalysing the Tunisian samples using the longer D-loop fragment is crucial (Shamblin et al. 2014 ). So far only for MSA the long fragment was used (Sami et al. 2011 ). In this study the long fragment was used showing that most loggerheads from the Tunisian rookery carry the common haplotype CC-A2.1 (39), however one specimen also carried haplotype CC-A2.9, and another one was CC-A3.1. Based on the detection of the CC-A2.9 haplotype at a Tunisian rookery, classification of Tunisia and Libya as one Management Unit (MU) is reasonable (see Shamblin et al. 2014 ). In Albania, evidence of the occurrence of nesting was apparent with infrequent reports of hatchling sightings along the coastline, but it was not until 2018 that the first official loggerhead nest was confirmed (Piroli and Haxhi, 2020) which is why, until now, no molecular genetic analysis was done. In this study all eight samples analysed carried the cosmopolitan haplotype CC-A2.1. We have included both one hatchling and the other samples into the analyses to see if there is any different haplotype. We have also included 2 samples from Egypt in the same line to see the presence of additional haplotypes present in the Mediterranean as project partners provided samples. Eight loggerheads were of Lebanese origin. The haplotypes they carried are consistent with previous findings, as haplotype CC-A2.1 and CC-A3.1 were found (Saied et al. 2012 ; Clusa et al. 2014 ). Differences in polymorphism measures of different Mediterranean countries were observed. While these differences may reflect a discrepancy in sample size for most comparisons, higher values were measured in Libya than in Turkey, even though fewer loggerhead sea turtle samples were used there. Furthermore, two new haplotypes were discovered in Libya, thus our findings support Saied et al. ( 2012 ) claim that Libya has an important with wide range of haplotypes for loggerhead sea turtle which is why the protection of this assembly is essential in order to conserve the Mediterranean stock. This study has provided new insights into the population structure of the loggerhead sea turtle in the Mediterranean Sea. Haplotypes previously thought to be endemic to certain nesting sites, but now found at other rookeries as well, and due to the assignment of an ''orphaned'' haplotype to a nesting area, it is suggested that MSA should be repeated, as these new findings could change the contribution of some rookeries to certain stocks (e.g. Adriatic Sea). Declarations Acknowledgements : We would like to thank the MAVA Foundation for supporting the project of Conservation of Marine Turtles in the Mediterranean. This paper is significant for the loggerhead turtle in the Mediterranean, and for this reason all authors agreed to collaborate by providing data. We all thank our volunteers in collecting the samples in the field. Author contributions: All authors contributed to the study conception and design. The first draft of the manuscript was written by corresponding author and his co-authors from the same affiliation and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript. References Abreu-Grobois FA, Horrocks J, Formia A, Dutton P, LeRoux R, Vélez-Zuazo X, Soares L, Meylan P (2006) New mtDNA D-loop primers which work for a variety of marine turtle species may increase the resolution of mixed stock analysis. In: Frick M, Panagopoulou A, Rees AF, Williams K (ed) Proceedings of the 26th annual symposium on sea turtle biology Island of Crete, Greece, ISTS, pp 179 Bertuccio V, Costantini F, Angelini V, Furii G, Gobic K, Abbiati M (2019) Haplotype and biometric patterns in loggerhead turtles from the Adriatic foraging ground. J Sea Res 147:1–9. https://doi.org/10.1016/j.seares.2019.02.002 Bolten AB, Crowder LB, Dodd MG, MacPherson SL, Musick JA, Schroeder BA, Witherington BE, Long KJ, Snover ML (2011) Quantifying multiple threats to endangered species: an example of loggerhead sea turtles. Front Ecol Environ 9(5):295–301. https://doi.org/10.1890/090126 Carreras C, Rees AF, Broderick AC, Godley BJ, Margaritoulis D (2014) Mitochondrial DNA markers of loggerhead marine turtles ( Caretta caretta ) (Testudines: Cheloniidae) nesting at Kyparissia Bay, Greece, confirm the western Greece unit and regional structuring. Sci Mar 78:115–124. doi: http://dx.doi.org/10.3989/scimar.03865.27B Casale P, Broderick AC, Camiñas JA, Cardona L et al (2018) Mediterranean sea turtles: current knowledge and priorities for conservation and research. Endanger Species Res 36:229–267. https://doi.org/10.3354/esr00901 Chaieb O, El Ouaer A, Mafucci F, Bradai MN, Bentivegna F, Said K, Chatti N (2010) Genetic survey of loggerhead turtle Caretta caretta nesting population in Tunisia. Mar Biodivers Rec 3:e20. doi: 10.1017/S175526721000014X Clusa M, Carreras C, Pascual M, Demetropoulos A, Margaritoulis D, Rees AF, Hamza AA, Khalil M, Aureggi M, Levy Y, Turkozan O, Marco A, Aguilar A, Cardona L (2013) Mitochondrial DNA reveals Pleistocenic colonisation of the Mediterranean by loggerhead turtles (Caretta caretta). J Exp Mar Biol Ecol 439:15–24. https://doi.org/10.1016/j.jembe.2012.10.011 Clusa M, Carreras C, Pascual M, Gaughran SJ, Piovano S, Giacoma C, Fernández G, Levy Y, Tomás J, Raga JA, Maffucci F, Hochscheid S, Aguilar A, Cardona L (2014) Fine-scale distribution of juvenile Atlantic and Mediterranean loggerhead turtles ( Caretta caretta ) in the Mediterranean Sea. Mar Biol 161:509–519. https://doi.org/10.1007/s00227-013-2353-y Crandall KA, Bininda-Emonds ORP, Mace GM, Wayne RK (2000) Considering evolutionary processes in conservation biology. Trends Ecol Evol 15(7):290–295. https://doi.org/10.1016/S0169-5347(00)01876-0 Ferrera A, Formia A, Ciofi C, Natali C, Agyekumhene A, Allman P (2021) Genetic structure of olive ridley sea turtles ( Lepidochelys olivacea ) in Ghana, West Africa. J Exp Mar Biol Ecol 544:151614. https://doi.org/10.1016/j.jembe.2021.151614 Garofalo L, Mingozzi T, Mico A, Novelletto A (2009) Loggerhead turtle ( Caretta caretta ) matrilines in the Mediterranean: further evidence of genetic diversity and connectivity. Mar Biol 156:2085–2095. https://doi.org/10.1007/s00227-009-1239-5 Garofalo L, Marsili L, Terracciano G, Baini M, Senese M, Mancusi C, Lorenzini R (2016a) Welcome to the North! Genetic makeup of loggerhead Caretta caretta females from the northernmost Italian nesting site. In: Menegon M, Rodriguez-Prieto A, Deforian MC (eds) Atti XI Congresso Nazionale Societas Herpetologica Italica. Ianieri Edizioni, Trento, Italy Garofalo L, Insacco G, Scaravelli D, Lorenzini R (2016) On the connectivity between nesting colonies of the loggerhead turtle Caretta caretta in the Ionian Region. In: Menegon M, Rodriguez-Prieto A, Deflorian MC (eds) Atti XI Congresso Nazionale Societas Herpetologica Italica. Ianieri Edizioni, Trento, Italy, pp 135–136 Groombridge B (1994) Marine turtles in the Mediterranean: distribution, population status, conservation. Council of Europe Press, Nature and environment, Strasbourg, pp 18–48 Hall TA (1999) BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucl Acids Symp Ser 41:95–98 Hamza A (2010) Libya. In: Casale P, Margaritoulis D (eds) Sea turtles in the Mediterranean: distribution, threats and conservation priorities. IUCN, Gland, pp 157–170 Kaska A, Bagcı H, Kaska Y (2001) Genetic Material Collection of Sea Turtles and PCR Amplifications. In: Margaritoulis D, Demetropoulos A (ed) Proceedings of the First Mediterranean Conference on Sea Turtles. Barcelona Convention-Bern Convention-Bonn Convention (CMS), Nicosia, Cyprus, 270 pp 147–150 Kumar S, Stecher G, Tamura K (2016) MEGA7: Molecular evolutionary genetics analysis version 7.0 for bigger datasets. Mol Biol Evol 33:1870–1874. doi: 10.1093/molbev/msw054 Laurent L, Casale P, Bradai MN, Godley BJ, Broderick GG, Schroth W, Schierwater B, Levy AM, Freggi D, Abd El-Mawla EM (1998) Molecular resolution of marine turtle stock composition in fishery bycatch: a case study in the Mediterranean. Mol Ecol 7:1529–1542. https://doi.org/10.1046/j.1365-294x.1998.00471.x Leigh JW, Bryant D (2015) Popart: full-feature software for haplotype network construction. Methods Ecol Evol 6:1110–1116. doi: 10.1111/2041-210X.12410 Librado P, Rozas J (2009) DnaSP v5: a software for comprehensive analysis of DNA polymorphism data. Bioinform 25:1451–1452. https://doi.org/10.1093/bioinformatics/btp187 Mafucci F, Corrado R, Palatella L, Borra M, Marullo S, Hochscheid S, Lacorata G, Iudicone D (2016) Seasonal heterogeneity of ocean warming: a mortality sink for ectotherm colonizers. Sci Rep 6(1):1–9. https://doi.org/10.1038/srep23983 Margush T, McMorris FR (1981) Consensus n -trees. Bull Math Biol 43:239–244. https://doi.org/10.1007/BF02459446 Monzón-Argüello C, Rico C, Naro-Maciel E, Varo-Cruz N, López P, Marco A, López-Jurado LF (2010) Population structure and conservation implications for the loggerhead sea turtle of the Cape Verde Islands. Conserv Genet 11:1871–1884. https://doi.org/10.1007/s10592-010-0079-7 Moritz C (1994) Applications of mitochondrial DNA analysis in conservation: a critical review. Mol Ecol 3:401–411. https://doi.org/10.1111/j.1365-294X.1994.tb00080.x Piroli V, Haxhiu I (2020) Nesting of the loggerhead turtle ( Caretta Caretta ) in the southeast Adriatic confirmed. Nat Croat 29:23–30. https://hrcak.srce.hr/249247[CS1] Rees A, Carreras C, Broderick A, Margaritoulis D, Stringell T, Godley B (2017) Linking loggerhead locations: using multiple methods to determine the origin of sea turtles in feeding grounds. Mar Biol 164(2):1–14. https://doi.org/10.1007/s00227-016-3055-z Saied A, Maffucci F, Hochscheid S, Dryag S, Swayeb B, Borra M, Oureghi A, Procaccini G, Bentivegna F (2012) Loggerhead turtles nesting in Libya: an important management unit for the Mediterranean stock. Mar Ecol Prog Ser 450:207–218. https://doi.org/10.3354/meps09548 Sami K, Maffucci F, Imed J, Abderrahmen B, Bradai M, Bentivegna F (2011) Demographic composition of the Tunisian loggerhead foraging aggregation: does the analysis of a longer mtDNA segment provide additional information? In: Bentivegna F, Maffucci F, Mauriello V (ed) Book of Abstracts, 4th Mediterranean conference on marine turtles, Naples, Italy, pp 74 Schofield G, Hobson V, Fossette S, Lilley M, Katselidis KA, Hays GC (2010) Biodiversity research: Fidelity to foraging sites, consistency of migration routes and habitat modulation of home range by sea turtles. Divers Distrib 16(5):840–853. https://doi.org/10.1111/j.1472-4642.2010.00694.x Shamblin BM, Bolten AB, Abreu-Grobois FA, Bjorndal KA, Cardona L, Carreras C, Clusa M, Monzón-Argüello C, Nairn CJ, Nielsen JT et al (2014) Geographic patterns of genetic variation in a broadly distributed marine vertebrate: new insights into loggerhead turtle stock structure from expanded mitochondrial DNA sequences. PLoS ONE 9(1). https://doi.org/10.1371/journal.pone.0085956 Splendiani A, Fioravanti T, Giovannotti M, D'Amore A, Furii G, Totaro G et al (2017) Mitochondrial DNA reveals the natal origin of Caretta caretta (Testudines: Cheloniidae) stranded or bycaught along the Southwestern Adriatic coasts. Eur Zool J 84(1):566–574. https://doi.org/10.1080/24750263.2017.1400597 Tolve L, Casale P, Formia A, Garofalo L, Lazar B, Natali C et al (2018) A comprehensive mitochondrial DNA mixed-stock analysis clarifies the composition of loggerhead turtle aggregates in the Adriatic Sea. Mar Biol 165(4):1–14. https://doi.org/10.1007/s00227-018-3325-z Yilmaz C, Türkozan O, Bardakci F, White M, Kararaj E (2012) Loggerhead turtles ( Caretta caretta ) foraging at Drini Bay in Northern Albania: Genetic characterisation reveals new haplotypes. Acta Herpetol 7(1):155–162 Additional Declarations No competing interests reported. Supplementary Files floatimage2.jpeg Supplement, Figure 2. The phylogenetic tree of the haplotypes detected in this study. Cite Share Download PDF Status: Published Journal Publication published 13 May, 2023 Read the published version in Conservation Genetics Resources → Version 2 posted Editorial decision: Major revision 03 Apr, 2023 Reviews received at journal 03 Feb, 2023 Reviewers agreed at journal 02 Feb, 2023 Reviewers invited by journal 30 Jan, 2023 Submission checks completed at journal 05 Jul, 2022 Editor assigned by journal 05 Jul, 2022 First submitted to journal 04 Jul, 2022 You are reading this latest preprint version Show more versions Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. 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13:59:23","currentVersionCode":2,"declarations":"","doi":"10.21203/rs.3.rs-1649861/v2","doiUrl":"https://doi.org/10.21203/rs.3.rs-1649861/v2","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1007/s12686-023-01303-9","type":"published","date":"2023-05-13T20:49:15+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":24359953,"identity":"cfe701b6-7392-4143-a509-e417c4e4c827","added_by":"auto","created_at":"2022-07-26 18:09:40","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":159140,"visible":true,"origin":"","legend":"\u003cp\u003eHaplotype network of mtDNA for loggerhead sea turtles from different management units. Connecting lines between haplotypes represent single mutations. The circle areas of the haplotypes are proportional to the sample size carrying the specific haplotype. New haplotypes were given as bold.\u0026nbsp;\u003c/p\u003e","description":"","filename":"floatimage1.png","url":"https://assets-eu.researchsquare.com/files/rs-1649861/v2/c7cf32c820db19e9f70afe86.png"},{"id":44729730,"identity":"d6952915-2fd5-438c-a155-319cfe5d8ca8","added_by":"auto","created_at":"2023-10-16 21:20:38","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":445275,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-1649861/v2/9f4131fc-239a-4073-8cbe-081fe063d282.pdf"},{"id":24360477,"identity":"46d4e131-ce22-4ae7-aeb8-17093cd448b5","added_by":"auto","created_at":"2022-07-26 18:14:40","extension":"jpeg","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":73023,"visible":true,"origin":"","legend":"\u003cp\u003eSupplement, Figure\u0026nbsp;2. The phylogenetic tree of the haplotypes detected in this study.\u003c/p\u003e","description":"","filename":"floatimage2.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-1649861/v2/9ae4e02b6586b61aaed1cab9.jpeg"}],"financialInterests":"No competing interests reported.","formattedTitle":"\u003cp\u003eIncreased Sample Size Proved More Insights for the Population Structure of Mediterranean Loggerhead Sea Turtles\u003c/p\u003e","fulltext":[{"header":"1. Introduction","content":"\u003cp\u003eThe loggerhead turtle (\u003cem\u003eCaretta caretta\u003c/em\u003e) is the most common sea turtle species in the Mediterranean Sea (Casale et al., \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). Nesting occurs mainly in the Eastern Mediterranean basin, with the highest number of clutches in Greece, Turkey, Libya and Cyprus (Casale et al., \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2018\u003c/span\u003e), with lower nesting numbers in Egypt, Israel, Italy, Lebanon, Syria and Tunisia. Minor nesting also occurs in the western basin in Malta, Albania, Spain, France and Italy (Casale et al., \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2018\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eGlobally, sea turtle populations are under many anthropogenic pressures. However, as a result of conservation efforts, the Mediterranean loggerhead population has recovered substantially and is currently classified as \u003cem\u003eLeast Concern\u003c/em\u003e by the International Union for Conservation of Nature (IUCN) due to increasing nest numbers of the species at major nesting sites of the Mediterranean Sea. Furthermore, it has been considered as \u003cem\u003eConservation Depended\u003c/em\u003e since the threats still exist (Casale, 2015).\u003c/p\u003e \u003cp\u003eIt is worth noting that the loggerhead turtle is a highly mobile species and migrates between nesting and foraging sites. Recent studies showed that the Mediterranean loggerhead turtle population represents a wide dispersal from nesting sites to different foraging sites (Cerritelli \u003cem\u003eet al.\u003c/em\u003e, 2022, Haywood \u003cem\u003eet al.\u003c/em\u003e, 2020, Schofield et al., \u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e2010\u003c/span\u003e). Despite conservation efforts at the local levels, studies based on international cooperation and the determination of area-based conservation measures have been adopted recently (Kot \u003cem\u003eet al.\u003c/em\u003e, 2022). Consequently, this step will provide high benefits for sea turtle conservation studies.\u003c/p\u003e \u003cp\u003eSea turtles exhibit a behavior known as philopatry, defined as a tendency to return to their natal beaches (Bowen and Karl, 2007). Accordingly, this behavior results in certain regions forming populations with specific genetic structures over time. Those populations are defined as Management Units (Moritz, \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e1994\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThe Mediterranean loggerhead turtle population is identified as one of the 10 Regional Management Units (RMU) based on studies of nesting sites, population abundances and trends, population genetics, and satellite telemetry (Wallace \u003cem\u003eet al.\u003c/em\u003e, 2010). Although the Mediterranean loggerhead turtle population is defined as a single RMU, there is genetic diversity in both nesting and foraging areas at haplotype level (Carreras \u003cem\u003eet al.\u003c/em\u003e, 2006, Yilmaz \u003cem\u003eet al.\u003c/em\u003e, 2011, Saied et al., \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e2012\u003c/span\u003e, Carreras et al., \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2014\u003c/span\u003e, Clusa et al., \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e2014\u003c/span\u003e, Rees et al., \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e2017\u003c/span\u003e, Clusa \u003cem\u003eet al.\u003c/em\u003e, 2018, Tolve et al., \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). However, the Mediterranean loggerhead meta-population has been proved to be composed of three independent RMUs (1 Mediterranean RMU and 2 Atlantic RMUs) (Casale et al., \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). Recently, loggerhead sporadic nesting events have become frequent in the western Mediterranean, and the Atlantic RMUs contributed to these nesting events (Carreras \u003cem\u003eet al.\u003c/em\u003e, 2019). Therefore, genetic diversity may differ between foraging and nesting sites.\u003c/p\u003e \u003cp\u003eMixed-stock analysis (MSA) based on mitochondrial DNA (mtDNA) sequences are used to assess haplotype diversity among different nesting sites and estimate the origins of individuals at foraging sites (Bowen \u003cem\u003eet al.\u003c/em\u003e, 2007). It has been suggested that the knowledge about the metapopulation structure may have been obtained via available genetic markers (Casale et al., \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). However, due to unsampled breeding sites and incomplete sampling of large, well-known sites, complete genetic characterization was impossible for the Mediterranean. Furthermore, mainly when \u0026ldquo;orphaned haplotypes\u0026rdquo; exist (i.e. haplotypes recorded from the population but not reported at any breeding site), the MSA has low power in some foraging sites (Tolve et al., \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). Therefore, to have more comprehensive knowledge on the loggerhead turtle population structure, there is a need for simultaneous studies with an increased number of samples from areas that were previously represented with few samples or were not sampled.\u003c/p\u003e \u003cp\u003eIn this context, and with the contribution of researchers from seven countries in the Mediterranean, and to have a more robust knowledge, this study aims to answer the following questions:\u003c/p\u003e \u003cp\u003e \u003col\u003e \u003cspan\u003e \u003cli\u003e \u003cp\u003eDid we reach the limit on the knowledge about the loggerhead turtle population structure?\u003c/p\u003e \u003c/li\u003e \u003c/span\u003e \u003cspan\u003e \u003cli\u003e \u003cp\u003eIs there further strong connectivity among different breeding sites?\u003c/p\u003e \u003c/li\u003e \u003c/span\u003e \u003cspan\u003e \u003cli\u003e \u003cp\u003eDo the Atlantic loggerhead turtles contribute to the breeding population in the eastern Mediterranean?\u003c/p\u003e \u003c/li\u003e \u003c/span\u003e \u003cspan\u003e \u003cli\u003e \u003cp\u003eWhat is the origin of the individuals at new nesting sites in the Mediterranean?\u003c/p\u003e \u003c/li\u003e \u003c/span\u003e \u003cspan\u003e \u003cli\u003e \u003cp\u003eDo studies with low sample numbers mask the extent of genetic diversity in the Mediterranean?\u003c/p\u003e \u003c/li\u003e \u003c/span\u003e \u003c/ol\u003e \u003c/p\u003e \u003cp\u003eFinally, increasing the sample size of rookeries from different countries contributes to better understanding of the genetic structure of \u003cem\u003eCaretta caretta\u003c/em\u003e from Mediterranean rookeries.\u003c/p\u003e"},{"header":"2. Material And Method","content":"\u003cp\u003e \u003cb\u003eSample collection\u003c/b\u003e \u003c/p\u003e \u003cp\u003eA total of 710 samples were collected between 2018 and 2019 along the beaches of Cyprus, Libya, Lebanon, Tunisia, Greece, and Turkey. The sites from Libya and Lebanon have not been previously sampled, the other samples were to replace the findings for them with a larger sample size. Samples from Albania and Egypt were included to identify their haplotypes in the analyses. Skin biopsies were taken from adult females, and hatchlings were preserved in 70% alcohol until further laboratory procedures by researchers in each country. Only nests laid within 15 days, which is the optimum nesting interval for the loggerhead turtles, were used for the genetic analysis to prevent the risk of pseudo-replication. The sample size in Turkey was 384, Libya 134, Greece 104, Tunisia 41, Cyprus 27, Albania and Lebanon eight. Six Albanian samples were collected from live specimens of \u003cem\u003eCaretta caretta\u003c/em\u003e captured as bycatch at Ishmi stavnik, Patok area (Drini bay), while the other two were collected from two dead \u003cem\u003eCaretta caretta\u003c/em\u003e hatchlings from the first officially documented nest in Albania in Divjaka beach area, in 2018 (Piroli and Haxhi, 2020). Albanian samples and two samples from Egypt were included as non-nesting individuals to show also a case for mixed stock analysis can be performed for strandings.\u003c/p\u003e \u003cp\u003e \u003cb\u003eLaboratory analysis\u003c/b\u003e \u003c/p\u003e \u003cp\u003eThe DNA extraction was performed using a standard phenol\u0026ndash;chloroform protocol (Kaska et al., \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2001\u003c/span\u003e) or a QuickGene DNA tissue kit (KURABO). Representatives of each research team trained for the same protocol for extraction of DNA and PCR amplification at DEKAMER Lab in Turkey. An approximately 815 base pair (bp) long fragment of the non-coding mitochondrial DNA (mtDNA) control region was amplified by Polymerase Chain Reaction (PCR). The primer pair used was LCM15382 (5\u0026prime;-GCTT AACCCTAAGCATTGG-3\u0026prime;) and H950 (5\u0026prime;-GTCTCGG ATTTAGGGGTTT-3\u0026prime;) (Abreu-Grobois et al. \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2006\u003c/span\u003e). Polymerase Chain Reaction (PCR) was performed in a total volume of 30-\u0026micro;L mastermix, 0.5 \u0026micro;M of each primer, and 2 \u0026micro;L of DNA. Thermal conditions consisted of an initial denaturation at 95\u0026deg;C for 3 minutes (min), followed by 34 cycles of 30 seconds (s) at 95\u0026deg;C, 1 min at 55\u0026deg;C and 30 s at 72\u0026deg;C, with a final extension step at 72\u0026deg;C for 10 minutes. PCR products were visualised on a 1% agarose gel stained with Safeview\u0026trade; for amplification evaluation. After completing the DNA extraction and the successful PCR products by each partner, the PCR products were sent to Genartek (Istanbul, Turkey) for DNA purification and Sanger sequencing with both forward and reverse primers. The obtained sequences were analysed by researchers from DEKAMER Lab, Turkey.\u003c/p\u003e \u003cp\u003e \u003cb\u003eData analysis\u003c/b\u003e \u003c/p\u003e \u003cp\u003eObtained sequences were, if needed, edited in Chromas (v.2.6.6) and aligned in Bioedit (v.7.2.5) (Hall, \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e1999\u003c/span\u003e). Haplotype classification was conducted through comparing the sequences with haplotypes already presented in the Archie Carr Center for Sea Turtle Research database (ACCSTR; \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttp://accstr.uf.edu/fles/cclongmtdn\u003c/span\u003e\u003cspan address=\"http://accstr.uf.edu/fles/cclongmtdn\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e \u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003ea.pdf\u003c/span\u003e) and the sequence comparison tool, GenBank BLAST (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttp://ncbi.nlm.nih.gov/Blast.cgi\u003c/span\u003e\u003cspan address=\"http://ncbi.nlm.nih.gov/Blast.cgi\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003e).\u003c/span\u003e Novel haplotypes found were submitted to ACCSTR for assigning the international nomenclature and their sequences were sent to GenBank. To highlight the relationship between different haplotypes identified, haplotype networks based on the median-joining algorithm were created using the program POPART version 1.7 (Leigh and Bryant \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). Polymorphism data was obtained by estimating the haplotype diversity (h), nucleotide diversity (π), number of haplotypes (k) and number of variable sites (p) in DnaSP version 5.10.01 (Librado and Rozas \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e2009\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eHaplotype networks of mtDNA for loggerhead turtles from different Mediterranean countries were created with the connecting lines between haplotypes representing single mutations. The circle areas of the haplotypes are proportional to the sample size carrying the specific haplotype were also shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e. The software MEGA v. 7 (Kumar et al. \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2016\u003c/span\u003e) was used to create a phylogenetic tree (Supplement, Fig.\u0026nbsp;2). The best-fit substitution model (ML) was chosen on the basis of the lowest Bayesian information criterion (BIC) value which was the Tamura 3-parameter with Gamma distribution (T92\u0026thinsp;+\u0026thinsp;G). For the phylogenetic analysis the neighbour-joining (NJ) method was used including the selected substitution model. To obtain valid results for the nodes the number of bootstrap replications were 1000, bootstrap values greater than or equal to 50% were considered statistically significant (Margush \u0026amp; McMorris \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e1981\u003c/span\u003e). For node calibration, D-loop sequences from two olive ridleys (\u003cem\u003eLepidochelys olivacea\u003c/em\u003e, GenBank AM258984 and JX454991) and Kemp\u0026rsquo;s ridley (\u003cem\u003eLepidochelys kempii\u003c/em\u003e, GenBank JX454981) were included in the alignment as outgroups.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e"},{"header":"3. Results","content":"\u003cp\u003eAmong the 710 mtDNA sequences analysed 14 variable sites were observed defining 16 haplotypes of which three were previously undescribed. The haplotypes found in the literature and found in this study were presented in Table \u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e. Both, CC-A2.1 and CC-A3.1 are the most common haplotypes at different Mediterranean rookeries. The highest number of haplotypes were found in Libya (9), followed by Greece (6) and Turkey (5) (see Table \u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003e). The haplotype diversity was highest in Libya-Tunisia Management Unit, followed by western Turkey and Crete, Greece. The most frequently found haplotypes were CC-A2.1 (67.0%) and CC-A3.1 (19.8%), both present at all nesting sites, followed by CC-A2.9 (6.7%) most found in Libya and with 3.2% haplotype CC-A26.1, also originating from Libya. The genetic polymorphism measures data (Table \u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003e) and frequencies of haplotypes (Table \u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003e) were given according to Management Units.\u0026nbsp;\u003c/p\u003e\n\u003ctable border=\"1\" id=\"Tab1\"\u003e\n \u003ccaption language=\"En\"\u003e\n \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e\n \u003cdiv class=\"CaptionContent\"\u003e\n \u003cp\u003eNumber (n) of \u003cem\u003eCaretta caretta\u003c/em\u003e haplotypes sampled at different Mediterranean rookeries and their rookery of origin (Clusa et al.2014).\u003c/p\u003e\n \u003c/div\u003e\n \u003c/caption\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eHaplotype\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003en\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eSource rookeries found in this study\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eRookery\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A2.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e468\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eDLM (7), DLY (83), TKE (46), TKM (35), TKW (81), MIS (4), SIR (49), CRT (22), KOR (18), KOT (8), LAK (26), ZAK (19), TUN (39), CYP (26), LEB (5)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMediterranean: MIS, SIR, ISR, LEB, CYP, ETU, MTU, DLM, DLY, CRE, WGR, CAL\u003c/p\u003e\n \u003cp\u003eAtlantic: CEF, SEF, SAL, DRT, QMX, SWF, CWF, NWF, CPV\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A2.8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCRT (4), LAK (1)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMediterranean: CRE\u003c/p\u003e\n \u003cp\u003eAtlantic: \u0026ndash;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A2.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e47\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eSIR (45), MIS (1), TUN (1)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMediterranean: MIS, SIR, ISR\u003c/p\u003e\n \u003cp\u003eAtlantic: \u0026ndash;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC- A3.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e138\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eDLM (36), DLY (55), TKE (6), TKW (30) (128), SIR (4), CRT (1), TUN (1), CYP (1), LEB(3)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMediterranean: MIS, SIR, LEB, ETU, WTU, WGR, DLM, DLY\u003c/p\u003e\n \u003cp\u003eAtlantic: CEF, SEF, SAL, QMX, SWF, CWF, NWF\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A6.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCRT (1), KOT (1)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMediterranean: WGR Atlantic: \u0026ndash;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC- A10.4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMIS (1)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMediterranean: Tuscany \u003csup\u003ea\u003c/sup\u003e, Campania \u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003cp\u003eAtlantic: CEF\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A26.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e22\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMIS (1), SIR (21)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMediterranean: SIR\u003c/p\u003e\n \u003cp\u003eAtlantic: \u0026ndash;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A29.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eTKW (1)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMediterranean: ISR\u003c/p\u003e\n \u003cp\u003eAtlantic: \u0026ndash;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-31.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCRT (1), KOR (1)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMediterranean: CAL, WGR, Sicily \u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003cp\u003eAtlantic: \u0026ndash;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A32.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCRT (1)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMediterranean: WGR\u003c/p\u003e\n \u003cp\u003eAtlantic: \u0026ndash;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A50.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eTKE (1)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMediterranean: CYP\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A68.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eSIR (2)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMediterranean: SIR\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A71.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eSIR (1)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eUnknown\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A77.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eSIR (3)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eNovel haplotype\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A2.16\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eSIR (2)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eNovel haplotype\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A3.4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eDLY (2)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eNovel haplotype\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003ctfoot\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"4\"\u003eMediterranean rookeries: MIS (Misurata, Libya), SIR (Sirte, Libya), ISR (Israel), LEB (Lebanon), CYP (Cyprus), ETU (Eastern Turkey), MTU (Middle Turkey), WTU (Western Turkey), DLM (Dalaman, Turkey), DLY( Dalyan, Turkey), CRE and CRT (Crete, Greece), WGR (Western Greece), KOR (Koroni, Western Greece), KOT (Kotychi, Western Greece), CAL (Calabria, Italy).\u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tfoot\u003e\n\u003c/table\u003e\n\u003cdiv class=\"gridtable\"\u003e\u0026nbsp;\u003ctable border=\"1\" id=\"Tab2\"\u003e\n \u003ccaption language=\"En\"\u003e\n \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e\n \u003cdiv class=\"CaptionContent\"\u003e\n \u003cp\u003eGenetic polymorphism measures of loggerhead sea turtles from different management units.\u003c/p\u003e\n \u003c/div\u003e\n \u003c/caption\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eManagement\u003c/p\u003e\n \u003cp\u003eunit\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003en\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003ek\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003ep\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eS\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eHd\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003e\u0026pi;\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eWGRC\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e74\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.0800\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.00007\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCRT\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e30\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.4552\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.00053\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eDLYDLM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e184\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.5132\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.00064\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eTKW\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e112\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.4088\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.00051\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eLIBY_TUN\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e175\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.6362\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.00107\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eEMED\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e123\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.1655\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.00020\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eTotal_Med\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e698\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e16\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e14\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e14\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.5077\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.00078\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003c/table\u003e\n\u003c/div\u003e\n\u003cdiv align=\"char\"\u003en number of turtles sampled, k number of haplotypes, p number of polymorphic (segregation) sites, S Number of variable sites, Hd haplotype diversity, \u0026pi; nucleotide diversity\u003c/div\u003e\n\u003cp\u003eTable 3. Haplotype number and frequencies (%) found in different Management Units.\u003c/p\u003e\n\u003cdiv class=\"gridtable\"\u003e\n \u003ctable border=\"1\" id=\"Tabb\"\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eHaplotype\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eWGRC\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eCRT\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eDLYDAL\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eTKW\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eLIBYTUN\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eEMED\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eOverall\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A2.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e71 (95.9%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e22 (73.3%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e90 (48.9%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e81 (72.3%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e92 (52.6%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e112 (91.1%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e468 (67.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A2.8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e1 (1.4%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e4 (13.3%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e5 (0.7%)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A2.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e47 (26.9%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e47 (6.7%)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A3.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e1 (3.3%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e92 (50.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e30 (26.8%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e5 (2.9%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e10 (8.1%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e138 (19.8%)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A6.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e1 (1.4%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e1 (3.3%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e2 (0.3%)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A10.4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e1 (0.6%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e1 (0.1%)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A26.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e22 (12.6%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e22 (3.2%)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A29.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e1 (0.9%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e1 (0.1%)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-31.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e1 (1.4%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e1 (3.3%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e2 (0.3%)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A32.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e1 (3.3%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e1 (0.1%)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A50.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e1 (0.8%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e1 (0.1%)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A68.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e2 (1.1%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e2 (0.3%)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A71.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e1 (0.6%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e1 (0.1%)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A77.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e3 (1.7%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e3 (0.4%)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A2.16\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e2 (1.1%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e2 (0.3%)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCC-A3.4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e2 (1.1%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0 (0.0%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e2 (0.3%)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003c/table\u003e\n\u003c/div\u003e\n\u003cp\u003e\u003cbr\u003e\u003c/p\u003e\n\u003cp\u003eOne new haplotype detected in Sirte, Libya differed by one substitution from CC-A2.9 which is an exclusively Mediterranean haplotype (Tolve et al. \u003cspan class=\"CitationRef\"\u003e2018\u003c/span\u003e) and common in the Libyan nesting sites (Splendiani et al. \u003cspan class=\"CitationRef\"\u003e2017\u003c/span\u003e). The novel sequence was named CC-A77.1 after the nomenclature rules published on the Archie Carr Center for Sea Turtle Research (ACCSTR) website. The second new haplotype identified at a nesting site in Sirte, Libya differed one substitution from CC-A2.1, which is found in the Mediterranean and Atlantic populations (Shamblin et al. \u003cspan class=\"CitationRef\"\u003e2014\u003c/span\u003e) and was named CC-A2.16. The third previously undiscovered haplotype, namely CC-A3.4, was detected at a Turkish nesting site (Dalyan) and is one substitution separated from CC-A3.1, which is also found both in the Mediterranean and the Atlantic (Shamblin et al. \u003cspan class=\"CitationRef\"\u003e2014\u003c/span\u003e). All novel haplotypes were deposited in ACCSTR and Blast. The phylogenetic tree of \u003cem\u003eC. caretta\u003c/em\u003e generated with the NJ method clearly shows the two different mitochondrial lineages (known as Haplogroup I and Haplogroup II) highlighted in Shamblin et al. (\u003cspan class=\"CitationRef\"\u003e2014\u003c/span\u003e). The three new haplotypes (CC-A2.16, CC-A3.4 and CC-A77.1) are representatives of Haplogroup II. The haplotype network of mtDNA for loggerhead turtles from different Mediterranean management units were also shown in Fig.\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e, the phylogenetic trees were also provided (Supplement, Fig.\u0026nbsp;2).\u003c/p\u003e"},{"header":"4. Discussion","content":"\u003cp\u003eMolecular genetic investigation for differentiated populations has been characterised as a powerful tool for conservation purposes (Crandall et al. \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e2000\u003c/span\u003e; Moritz, \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e1994\u003c/span\u003e). However, length of the sequences, selected markers and sample size play an important role in differentiating populations (Monz\u0026oacute;n-Arg\u0026uuml;ello et al. \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2010\u003c/span\u003e; Clusa et al. \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2013\u003c/span\u003e). Using extended mtDNA 815bp haplotype sequences, even independent management units (MUs) have been identified within the Mediterranean RMU (Shamblin et al. \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e2014\u003c/span\u003e), namely: (i) Calabria-Italy (CAL), (ii) western Greece (WGRC), (iii) Crete-Greece (CRT), (iv) Libya (LIBY), (v) Dalyan-Dalaman-Turkey (DLYDAL), (vi) western Turkey (TKW), (vii) eastern Mediterranean (EMED, including middle and eastern Turkey, Cyprus, Israel, and Lebanon).\u003c/p\u003e \u003cp\u003eAccording to our knowledge so far, the highest diversity with seven haplotypes was reported from nesting sites was Turkey (Yilmaz et al. 2011, Carreras et al. \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2014\u003c/span\u003e, Clusa et al. \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e2014\u003c/span\u003e and references therein). It is followed by Libya with five haplotypes (Said et al. 2012), Greece with four haplotypes (Yilmaz et al. 2011, Carreras et al. \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2014\u003c/span\u003e), Cyprus (Clusa et al. \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2013\u003c/span\u003e) and Lebanon (Yilmaz et al. 2011) with two haplotypes, and Tunisia with one haplotype (Chaieb et al. \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2010\u003c/span\u003e). Although recent sporadic loggerhead turtle nest records were available from Albania (Piroli and Haxhi, 2020), genetic characterization was not available to date.\u003c/p\u003e \u003cp\u003eLibyan coasts and information about the nesting sites are identified as the major knowledge gap in the Mediterranean (Casale et al. \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). In addition, Libya is a significant place as it is the area where the loggerhead turtle first colonised in the Mediterranean (Clusa et al. \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2013\u003c/span\u003e). The results show that more effort needs to be put into sampling, especially along the largely unexplored Libyan coasts as two novel haplotypes have been found in this area. In total nine haplotypes were found. The second most frequent haplotype found in Misratah and Sirte, Libya was CC-A2.9 which occurs frequently in Israel and Libyan rookeries (Saied et al. \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e2012\u003c/span\u003e; Clusa et al. \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). One specimen from a nesting site in Misratah, Libya was detected carrying the rare haplotype CC-A10.4. This haplotype was previously only recorded occasionally at Tyrrhenian nesting sites (Garofalo et al. \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2016a\u003c/span\u003e; Mafucci et al. \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). It derives from the CC-A10 haplotype (380 bp mtDNA sequence) which is previously observed only once in Greece (Laurent et al. \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e1998\u003c/span\u003e). For CC-A10.4 it was also predicted that this haplotype probably originated not only from Tyrrhenian rookeries but also from Mediterranean colonies (Tolve et al. \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). The origin of the specimens found in the Adriatic (Yilmaz et al. \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e2012\u003c/span\u003e, Tolve et al. \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2018\u003c/span\u003e, Bertuccio et al. \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2019\u003c/span\u003e) could be in Libya. Both haplotypes, CC-A29.1 and CC-A10.4 were only found once in Turkey and Libya, respectively which is why the sampling size in these rookeries needs to be increased. Another very rare haplotype is CC-A68.1 which was found only once at the nesting location in Sirte, Libya in 2009. In this study two individuals carried CC-A68.1 from Sirte, Libya. Splendiani et al. (\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e2017\u003c/span\u003e), who used samples of rescued \u003cem\u003eCaretta caretta\u003c/em\u003e found along the Southern Adriatic coast of Italy, described the haplotype CC-A71.1 for the first time which is why they were not able to determine the origin of it. However, by creating a phylogenetic tree it was possible for them to see the phylogenetic relatedness to CC-A26.1 which is exclusive to Libya (Shamblin et al. \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). For this reason, Splendiani et al. (\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e2017\u003c/span\u003e) suggested that CC-A71.1 could be from a Libyan rookery. Our findings support this proposition since one specimen from Sirte, Libya carrying this haplotype was detected.\u003c/p\u003e \u003cp\u003eThe 104 samples analysed from Greece, including the Management Unit of Western Greece and Crete, included all known haplotypes in these rookeries. In Western Greece 71 (95.9%) and in Crete 22 (73.3%) of the loggerheads carried the haplotype CC-A2.1. Four individuals from Chania, Crete carried CC-A2.8 while one individual from Mavrovouni, Lakonikos Bay carried this haplotype, which was known to be endemic to the Cretan rookery. One individual from Crete carried a haplotype CC-A3.1, another one CC-A32.1. Haplotype CC-A6.1 was carried by loggerheads from Kotychi and Crete. Both haplotypes, CC-A6.1 and CC-A32.1 were known to be endemic to Western Greece. One specimen from Crete and one from Koroni had haplotype CC-A31.1 originating from Greek and Calabrian rookeries but also found in sporadic Sicilian nesting sites (Garofalo et al. \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e2016\u003c/span\u003eb).\u003c/p\u003e \u003cp\u003eIn this study in total five haplotypes were detected at main Turkish nesting sites of which two individuals belong to the same, new haplotype. The most frequent haplotype in Turkey, which is also the most common in the Mediterranean in general, was CC-A2.1 followed by CC-A3.1 also a common haplotype. The discovery of the CC-A29.1 haplotype at a nesting beach in Turkey, more precisely in western Turkey (\u0026Ccedil;ıralı) shows, as has been proposed before (Tolve et al. \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2018\u003c/span\u003e), that the origin of this haplotype must not only be in Israel, but also in other, poorly sampled or unknown nesting sites. This finding reinforces the results of MSA from the Adriatic Sea (Tolve et al. \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2018\u003c/span\u003e): Western Turkish rookeries, which despite the fact that they are much more abundant and closer to the Adriatic Sea, showed a medium contribution probability to the Adriatic stock (Tolve et al. \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). Two samples from Slovenia (northern Adriatic) carried the haplotype CC-A29.1. If CC-A29.1 would also be included as a haplotype of Western Turkish origin, the posterior probability would probably increase, and the contribution would not be similar or smaller than the ones of Israeli nesting areas. Haplotype CC-A50.1 is another very rare haplotype which was identified for the first time in Cyprus (Clusa et al. \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2013\u003c/span\u003e). We are not aware that this haplotype was detected again afterwards. One specimen from a Turkish rookery (Kazanlı) carried this haplotype.\u003c/p\u003e \u003cp\u003eTurkey is not only a foraging site for loggerhead sea turtles originating from Cyprus but also a new nesting site. A review of general migratory routes of 63 adult loggerhead turtles released mainly from Greece and Cyprus showed that only a few of them oriented to Turkish coasts (Luschi and Casale, 2014). The tracking studies from Northern Cyprus showed that only early nesters visited other Turkish rookeries (Snape et al. 2016). MSA of an eastern Turkish foraging ground made by T\u0026uuml;rkozan et al. (2018) showed a local contribution (62%) of Cyprus to the western subdivision.\u003c/p\u003e \u003cp\u003eFor Tunisian rookeries (Kuriat islands), only the short mtDNA control region fragment (500 bp) has been used so far, and only CC-A2.1 has been found (Chaieb et al. \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2010\u003c/span\u003e). Analysis using the short fragment have shown no significant differences between Libyan and Tunisian rookeries, but due to several hundred kilometres of separation Shamblin et al. (\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e2014\u003c/span\u003e) suggested that these might be demographically isolated nesting populations. A distinction, with the shorter D-loop sequence, between the haplotype CC-A2.9 and the widely distributed haplotype CC-A2.1 is not possible (Splendiani et al. \u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e2017\u003c/span\u003e), and given the fact that CC-A2.9 is common in Libya, it has been proposed that reanalysing the Tunisian samples using the longer D-loop fragment is crucial (Shamblin et al. \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). So far only for MSA the long fragment was used (Sami et al. \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e2011\u003c/span\u003e). In this study the long fragment was used showing that most loggerheads from the Tunisian rookery carry the common haplotype CC-A2.1 (39), however one specimen also carried haplotype CC-A2.9, and another one was CC-A3.1. Based on the detection of the CC-A2.9 haplotype at a Tunisian rookery, classification of Tunisia and Libya as one Management Unit (MU) is reasonable (see Shamblin et al. \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e2014\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eIn Albania, evidence of the occurrence of nesting was apparent with infrequent reports of hatchling sightings along the coastline, but it was not until 2018 that the first official loggerhead nest was confirmed (Piroli and Haxhi, 2020) which is why, until now, no molecular genetic analysis was done. In this study all eight samples analysed carried the cosmopolitan haplotype CC-A2.1. We have included both one hatchling and the other samples into the analyses to see if there is any different haplotype. We have also included 2 samples from Egypt in the same line to see the presence of additional haplotypes present in the Mediterranean as project partners provided samples.\u003c/p\u003e \u003cp\u003eEight loggerheads were of Lebanese origin. The haplotypes they carried are consistent with previous findings, as haplotype CC-A2.1 and CC-A3.1 were found (Saied et al. \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e2012\u003c/span\u003e; Clusa et al. \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e2014\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eDifferences in polymorphism measures of different Mediterranean countries were observed. While these differences may reflect a discrepancy in sample size for most comparisons, higher values were measured in Libya than in Turkey, even though fewer loggerhead sea turtle samples were used there. Furthermore, two new haplotypes were discovered in Libya, thus our findings support Saied et al. (\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e2012\u003c/span\u003e) claim that Libya has an important with wide range of haplotypes for loggerhead sea turtle which is why the protection of this assembly is essential in order to conserve the Mediterranean stock.\u003c/p\u003e \u003cp\u003eThis study has provided new insights into the population structure of the loggerhead sea turtle in the Mediterranean Sea. Haplotypes previously thought to be endemic to certain nesting sites, but now found at other rookeries as well, and due to the assignment of an ''orphaned'' haplotype to a nesting area, it is suggested that MSA should be repeated, as these new findings could change the contribution of some rookeries to certain stocks (e.g. Adriatic Sea).\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e:\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eWe would like to thank the MAVA Foundation for supporting the project of Conservation of Marine Turtles in the Mediterranean. This paper is significant for the loggerhead turtle in the Mediterranean, and for this reason all authors agreed to collaborate by providing data. We all thank our volunteers in collecting the samples in the field.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor contributions:\u003c/strong\u003e\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eAll authors contributed to the study conception and design. The first draft of the manuscript was written by corresponding author and his co-authors from the same affiliation and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eAbreu-Grobois FA, Horrocks J, Formia A, Dutton P, LeRoux R, V\u0026eacute;lez-Zuazo X, Soares L, Meylan P (2006) New mtDNA D-loop primers which work for a variety of marine turtle species may increase the resolution of mixed stock analysis. In: Frick M, Panagopoulou A, Rees AF, Williams K (ed) Proceedings of the 26th annual symposium on sea turtle biology Island of Crete, Greece, ISTS, pp\u0026nbsp;179\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eBertuccio V, Costantini F, Angelini V, Furii G, Gobic K, Abbiati M (2019) Haplotype and biometric patterns in loggerhead turtles from the Adriatic foraging ground. J Sea Res 147:1\u0026ndash;9. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1016/j.seares.2019.02.002\u003c/span\u003e\u003cspan address=\"10.1016/j.seares.2019.02.002\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eBolten AB, Crowder LB, Dodd MG, MacPherson SL, Musick JA, Schroeder BA, Witherington BE, Long KJ, Snover ML (2011) Quantifying multiple threats to endangered species: an example of loggerhead sea turtles. Front Ecol Environ 9(5):295\u0026ndash;301. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1890/090126\u003c/span\u003e\u003cspan address=\"10.1890/090126\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCarreras C, Rees AF, Broderick AC, Godley BJ, Margaritoulis D (2014) Mitochondrial DNA markers of loggerhead marine turtles (\u003cem\u003eCaretta caretta\u003c/em\u003e) (Testudines: Cheloniidae) nesting at Kyparissia Bay, Greece, confirm the western Greece unit and regional structuring. Sci Mar 78:115\u0026ndash;124. doi: \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttp://dx.doi.org/10.3989/scimar.03865.27B\u003c/span\u003e\u003cspan address=\"10.3989/scimar.03865.27B\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCasale P, Broderick AC, Cami\u0026ntilde;as JA, Cardona L et al (2018) Mediterranean sea turtles: current knowledge and priorities for conservation and research. Endanger Species Res 36:229\u0026ndash;267. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.3354/esr00901\u003c/span\u003e\u003cspan address=\"10.3354/esr00901\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eChaieb O, El Ouaer A, Mafucci F, Bradai MN, Bentivegna F, Said K, Chatti N (2010) Genetic survey of loggerhead turtle Caretta caretta nesting population in Tunisia. Mar Biodivers Rec 3:e20. doi:\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1017/S175526721000014X\u003c/span\u003e\u003cspan address=\"10.1017/S175526721000014X\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eClusa M, Carreras C, Pascual M, Demetropoulos A, Margaritoulis D, Rees AF, Hamza AA, Khalil M, Aureggi M, Levy Y, Turkozan O, Marco A, Aguilar A, Cardona L (2013) Mitochondrial DNA reveals Pleistocenic colonisation of the Mediterranean by loggerhead turtles (Caretta caretta). J Exp Mar Biol Ecol 439:15\u0026ndash;24. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1016/j.jembe.2012.10.011\u003c/span\u003e\u003cspan address=\"10.1016/j.jembe.2012.10.011\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eClusa M, Carreras C, Pascual M, Gaughran SJ, Piovano S, Giacoma C, Fern\u0026aacute;ndez G, Levy Y, Tom\u0026aacute;s J, Raga JA, Maffucci F, Hochscheid S, Aguilar A, Cardona L (2014) Fine-scale distribution of juvenile Atlantic and Mediterranean loggerhead turtles (\u003cem\u003eCaretta caretta\u003c/em\u003e) in the Mediterranean Sea. Mar Biol 161:509\u0026ndash;519. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1007/s00227-013-2353-y\u003c/span\u003e\u003cspan address=\"10.1007/s00227-013-2353-y\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCrandall KA, Bininda-Emonds ORP, Mace GM, Wayne RK (2000) Considering evolutionary processes in conservation biology. Trends Ecol Evol 15(7):290\u0026ndash;295. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1016/S0169-5347(00)01876-0\u003c/span\u003e\u003cspan address=\"10.1016/S0169-5347(00)01876-0\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eFerrera A, Formia A, Ciofi C, Natali C, Agyekumhene A, Allman P (2021) Genetic structure of olive ridley sea turtles (\u003cem\u003eLepidochelys olivacea\u003c/em\u003e) in Ghana, West Africa. J Exp Mar Biol Ecol 544:151614. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1016/j.jembe.2021.151614\u003c/span\u003e\u003cspan address=\"10.1016/j.jembe.2021.151614\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGarofalo L, Mingozzi T, Mico A, Novelletto A (2009) Loggerhead turtle (\u003cem\u003eCaretta caretta\u003c/em\u003e) matrilines in the Mediterranean: further evidence of genetic diversity and connectivity. Mar Biol 156:2085\u0026ndash;2095. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1007/s00227-009-1239-5\u003c/span\u003e\u003cspan address=\"10.1007/s00227-009-1239-5\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGarofalo L, Marsili L, Terracciano G, Baini M, Senese M, Mancusi C, Lorenzini R (2016a) Welcome to the North! Genetic makeup of loggerhead \u003cem\u003eCaretta caretta\u003c/em\u003e females from the northernmost Italian nesting site. In: Menegon M, Rodriguez-Prieto A, Deforian MC (eds) Atti XI Congresso Nazionale Societas Herpetologica Italica. Ianieri Edizioni, Trento, Italy\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGarofalo L, Insacco G, Scaravelli D, Lorenzini R (2016) On the connectivity between nesting colonies of the loggerhead turtle \u003cem\u003eCaretta caretta\u003c/em\u003e in the Ionian Region. In: Menegon M, Rodriguez-Prieto A, Deflorian MC (eds) Atti XI Congresso Nazionale Societas Herpetologica Italica. Ianieri Edizioni, Trento, Italy, pp 135\u0026ndash;136\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGroombridge B (1994) Marine turtles in the Mediterranean: distribution, population status, conservation. Council of Europe Press, Nature and environment, Strasbourg, pp 18\u0026ndash;48\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHall TA (1999) BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucl Acids Symp Ser 41:95\u0026ndash;98\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHamza A (2010) Libya. In: Casale P, Margaritoulis D (eds) Sea turtles in the Mediterranean: distribution, threats and conservation priorities. IUCN, Gland, pp 157\u0026ndash;170\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKaska A, Bagcı H, Kaska Y (2001) Genetic Material Collection of Sea Turtles and PCR Amplifications. In: Margaritoulis D, Demetropoulos A (ed) Proceedings of the First Mediterranean Conference on Sea Turtles. Barcelona Convention-Bern Convention-Bonn Convention (CMS), Nicosia, Cyprus, 270 pp\u0026nbsp;147\u0026ndash;150\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKumar S, Stecher G, Tamura K (2016) MEGA7: Molecular evolutionary genetics analysis version 7.0 for bigger datasets. Mol Biol Evol 33:1870\u0026ndash;1874. doi:\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1093/molbev/msw054\u003c/span\u003e\u003cspan address=\"10.1093/molbev/msw054\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLaurent L, Casale P, Bradai MN, Godley BJ, Broderick GG, Schroth W, Schierwater B, Levy AM, Freggi D, Abd El-Mawla EM (1998) Molecular resolution of marine turtle stock composition in fishery bycatch: a case study in the Mediterranean. Mol Ecol 7:1529\u0026ndash;1542. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1046/j.1365-294x.1998.00471.x\u003c/span\u003e\u003cspan address=\"10.1046/j.1365-294x.1998.00471.x\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLeigh JW, Bryant D (2015) Popart: full-feature software for haplotype network construction. Methods Ecol Evol 6:1110\u0026ndash;1116. doi:\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1111/2041-210X.12410\u003c/span\u003e\u003cspan address=\"10.1111/2041-210X.12410\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLibrado P, Rozas J (2009) DnaSP v5: a software for comprehensive analysis of DNA polymorphism data. Bioinform 25:1451\u0026ndash;1452. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1093/bioinformatics/btp187\u003c/span\u003e\u003cspan address=\"10.1093/bioinformatics/btp187\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMafucci F, Corrado R, Palatella L, Borra M, Marullo S, Hochscheid S, Lacorata G, Iudicone D (2016) Seasonal heterogeneity of ocean warming: a mortality sink for ectotherm colonizers. Sci Rep 6(1):1\u0026ndash;9. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1038/srep23983\u003c/span\u003e\u003cspan address=\"10.1038/srep23983\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMargush T, McMorris FR (1981) Consensus\u003cem\u003en\u003c/em\u003e-trees. Bull Math Biol 43:239\u0026ndash;244. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1007/BF02459446\u003c/span\u003e\u003cspan address=\"10.1007/BF02459446\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMonz\u0026oacute;n-Arg\u0026uuml;ello C, Rico C, Naro-Maciel E, Varo-Cruz N, L\u0026oacute;pez P, Marco A, L\u0026oacute;pez-Jurado LF (2010) Population structure and conservation implications for the loggerhead sea turtle of the Cape Verde Islands. Conserv Genet 11:1871\u0026ndash;1884. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1007/s10592-010-0079-7\u003c/span\u003e\u003cspan address=\"10.1007/s10592-010-0079-7\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMoritz C (1994) Applications of mitochondrial DNA analysis in conservation: a critical review. Mol Ecol 3:401\u0026ndash;411. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1111/j.1365-294X.1994.tb00080.x\u003c/span\u003e\u003cspan address=\"10.1111/j.1365-294X.1994.tb00080.x\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ePiroli V, Haxhiu I (2020) Nesting of the loggerhead turtle (\u003cem\u003eCaretta Caretta\u003c/em\u003e) in the southeast Adriatic confirmed. Nat Croat 29:23\u0026ndash;30. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://hrcak.srce.hr/249247[CS1]\u003c/span\u003e\u003cspan address=\"https://hrcak.srce.hr/249247[CS1]\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eRees A, Carreras C, Broderick A, Margaritoulis D, Stringell T, Godley B (2017) Linking loggerhead locations: using multiple methods to determine the origin of sea turtles in feeding grounds. Mar Biol 164(2):1\u0026ndash;14. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1007/s00227-016-3055-z\u003c/span\u003e\u003cspan address=\"10.1007/s00227-016-3055-z\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSaied A, Maffucci F, Hochscheid S, Dryag S, Swayeb B, Borra M, Oureghi A, Procaccini G, Bentivegna F (2012) Loggerhead turtles nesting in Libya: an important management unit for the Mediterranean stock. Mar Ecol Prog Ser 450:207\u0026ndash;218. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.3354/meps09548\u003c/span\u003e\u003cspan address=\"10.3354/meps09548\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSami K, Maffucci F, Imed J, Abderrahmen B, Bradai M, Bentivegna F (2011) Demographic composition of the Tunisian loggerhead foraging aggregation: does the analysis of a longer mtDNA segment provide additional information? In: Bentivegna F, Maffucci F, Mauriello V (ed) Book of Abstracts, 4th Mediterranean conference on marine turtles, Naples, Italy, pp\u0026nbsp;74\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSchofield G, Hobson V, Fossette S, Lilley M, Katselidis KA, Hays GC (2010) Biodiversity research: Fidelity to foraging sites, consistency of migration routes and habitat modulation of home range by sea turtles. Divers Distrib 16(5):840\u0026ndash;853. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1111/j.1472-4642.2010.00694.x\u003c/span\u003e\u003cspan address=\"10.1111/j.1472-4642.2010.00694.x\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eShamblin BM, Bolten AB, Abreu-Grobois FA, Bjorndal KA, Cardona L, Carreras C, Clusa M, Monz\u0026oacute;n-Arg\u0026uuml;ello C, Nairn CJ, Nielsen JT et al (2014) Geographic patterns of genetic variation in a broadly distributed marine vertebrate: new insights into loggerhead turtle stock structure from expanded mitochondrial DNA sequences. PLoS ONE 9(1). \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1371/journal.pone.0085956\u003c/span\u003e\u003cspan address=\"10.1371/journal.pone.0085956\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSplendiani A, Fioravanti T, Giovannotti M, D'Amore A, Furii G, Totaro G et al (2017) Mitochondrial DNA reveals the natal origin of \u003cem\u003eCaretta caretta\u003c/em\u003e (Testudines: Cheloniidae) stranded or bycaught along the Southwestern Adriatic coasts. Eur Zool J 84(1):566\u0026ndash;574. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1080/24750263.2017.1400597\u003c/span\u003e\u003cspan address=\"10.1080/24750263.2017.1400597\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eTolve L, Casale P, Formia A, Garofalo L, Lazar B, Natali C et al (2018) A comprehensive mitochondrial DNA mixed-stock analysis clarifies the composition of loggerhead turtle aggregates in the Adriatic Sea. Mar Biol 165(4):1\u0026ndash;14. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1007/s00227-018-3325-z\u003c/span\u003e\u003cspan address=\"10.1007/s00227-018-3325-z\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eYilmaz C, T\u0026uuml;rkozan O, Bardakci F, White M, Kararaj E (2012) Loggerhead turtles (\u003cem\u003eCaretta caretta\u003c/em\u003e) foraging at Drini Bay in Northern Albania: Genetic characterisation reveals new haplotypes. Acta Herpetol 7(1):155\u0026ndash;162\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"conservation-genetics-resources","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"cogr","sideBox":"Learn more about [Conservation Genetics Resources](https://www.springer.com/journal/12686)","snPcode":"12686","submissionUrl":"https://submission.nature.com/new-submission/12686/3","title":"Conservation Genetics Resources","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"Caretta caretta, Genetic structuring, mtDNA, Phylogeography, Conservation ","lastPublishedDoi":"10.21203/rs.3.rs-1649861/v2","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-1649861/v2","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eThe loggerhead sea turtle (\u003cem\u003eCaretta caretta\u003c/em\u003e), has the broadest distribution among sea turtle species in the Mediterranean and requires regional and international collaborations in addition to local efforts to better inform conservation actions. Molecular techniques are powerful tools to assess population dynamics at large scales, especially by determining the connectivity among different nesting and foraging sites, and genetic diversity. In this study, a large sample was collected synchronously in the nesting areas located in the north, south and east of the Mediterranean. Recently described nesting sites from Albania and other nesting sites represented by lower sample size were also included in order to fully assess the genetic composition of the region\u0026rsquo;s rookeries. Samples from 710 individuals were collected and the longer (815 bp) mtDNA D-loop fragment of these samples was amplified. We recorded 15 haplotypes, three of which were novel. In addition, our results show that some haplotypes, considered of Atlantic origin, have a wider dispersal in the Mediterranean than previously thought, albeit with low levels of representation. Our results, which also contribute to determining the likely origin of haplotypes that were previously known only from foraging sites, highlight the utility of broad-scale sampling, with increased sample number and longer mtDNA sequence to determine genetic diversity and connectivity. This study also demonstrates that it is important to continue to monitor the contribution of Atlantic origin haplotypes to the Mediterranean population, and the resident Mediterranean population, which is expected to expand its geographical range for reproduction with the effect of climate change and climate change in the long term. This work is important for, among other things, mixed stock analyses (MSA) that seek to localize the origin of stranded or accidentally caught sea turtles or those purposefully obtained from foraging sites to better understand the migratory distribution for conservation purposes.\u003c/p\u003e","manuscriptTitle":"Increased Sample Size Proved More Insights for the Population Structure of Mediterranean Loggerhead Sea Turtles","msid":"","msnumber":"","nonDraftVersions":[{"code":"","date":"2023-04-12 07:44:19","doi":"","editorialEvents":[{"type":"decision","content":"Accepted","date":"2023-04-23T06:02:05+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2023-04-13T02:43:29+00:00","index":"","fulltext":""},{"type":"submitted","content":"Conservation Genetics Resources","date":"2023-04-12T07:35:22+00:00","index":"","fulltext":""}],"status":"private","journal":{"display":true,"email":"[email protected]","identity":"conservation-genetics-resources","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"cogr","sideBox":"Learn more about [Conservation Genetics Resources](https://www.springer.com/journal/12686)","snPcode":"12686","submissionUrl":"https://submission.nature.com/new-submission/12686/3","title":"Conservation Genetics Resources","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false}},{"code":2,"date":"2022-07-26 18:09:38","doi":"10.21203/rs.3.rs-1649861/v2","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Major revision","date":"2023-04-04T03:47:21+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2023-02-03T11:07:26+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"6740a3d0-7372-4d9a-9a33-3e6ba0004a16","date":"2023-02-02T09:37:38+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2023-01-30T09:30:23+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2022-07-05T10:14:25+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2022-07-05T10:14:25+00:00","index":"","fulltext":""},{"type":"submitted","content":"Conservation Genetics Resources","date":"2022-07-04T21:28:57+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"conservation-genetics-resources","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"cogr","sideBox":"Learn more about [Conservation Genetics Resources](https://www.springer.com/journal/12686)","snPcode":"12686","submissionUrl":"https://submission.nature.com/new-submission/12686/3","title":"Conservation Genetics Resources","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"33360d19-7563-4fba-8530-4d7cfe690d46","owner":[],"postedDate":"July 26th, 2022","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2023-10-16T21:06:52+00:00","versionOfRecord":{"articleIdentity":"rs-1649861","link":"https://doi.org/10.1007/s12686-023-01303-9","journal":{"identity":"conservation-genetics-resources","isVorOnly":false,"title":"Conservation Genetics Resources"},"publishedOn":"2023-05-13 20:49:15","publishedOnDateReadable":"May 13th, 2023"},"versionCreatedAt":"2022-07-26 18:09:38","video":"","vorDoi":"10.1007/s12686-023-01303-9","vorDoiUrl":"https://doi.org/10.1007/s12686-023-01303-9","workflowStages":[]},"version":"v2","identity":"rs-1649861","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-1649861","identity":"rs-1649861","version":["v2"]},"buildId":"wLkW0s4AflPzk-lpfg-fK","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}