PHYLOGEOGRAPHY OF THE MALLARD (ANAS PLATYRHYNCHOS): HYBRIDIZATION, DISPERSAL, AND LINEAGE SORTING CONTRIBUTE TO COMPLEX GEOGRAPHIC STRUCTURE

Auk, The, Jul 2005 by Kulikova, Irina V, Drovetski, Sergei V, Gibson, Daniel D, Harrigan, Ryan J, Et al

We investigated Mallard mtDNA genetic differentiation in Western Russia, North Asia, the Aleutian Islands, and mainland Alaska. Our objectives were to (1) reconstruct the phylogenetic relationships of Mallard mtDNA haplotypes sampled broadly throughout the Holarctic, (2) test whether haplotype frequencies differ between the Old World and New World (and between these continents and the Aleutian Islands), and (3) further evaluate evidence of interspecific hybridization and incomplete lineage sorting of Mallard mtDNA haplotypes in different geographic regions.

METHODS

We collected 152 Mallards from Western Russia (Arkhangel'skaya, Astrakhan'skaya; n = 13), North Asia (Kazakhstan, Mongolia, Primorye, Khabarovsk, Magadan; n = 91), the Aleutian Islands (n = 17), and mainland Alaska (n = 31). We recorded specific collecting localities and museum catalogue numbers for vouchered specimens and combined specimens into general localities and regional samples (Table 1). DNA was extracted from liver and muscle tissues using a DNeasy Tissue Kit (Qiagen, Valencia, California). We amplified the 5' end of the mtDNA control region: positions 79-773 in the chicken (Callus gallus) mitochondrial genome (Desjardins and Morais 1990). Control region primers included L78 (Sorenson and Fleischer 1996) and H774 (Sorenson et al. 1999). Polymerase chain reactions (PCR) were done in a DNA Engine DYAD thermal cycler (MJ Research, Waltham, Massachusetts) using 50 µL reactions containing template DNA, 2.5 µL of each primer (10 mM), 5 µL of 10 mM dNTPs, 5 µL of 25 mM MgCl^sub 2^, 5 µL of 10× PCR buffer, and 0.25 µL Taq DNA Polymerase. Thermal cycling was as follows: 7 min preheat at 94°C, followed by 45 cycles of 20 s at 94°C, 20 s at 52°C, 1 min at 72°C, and a final extension of 7 min at 72°C. The PCR products were electrophoresed in agarose, excised from the gel, and purified using QIAquick Gel Extraction Kits (Qiagen). Both strands were cycle-sequenced using BigDye Terminator Cycle Sequencing Kits diluted four-fold, followed by electrophoresis on an ABI 3100 automated DNA sequencer (Applied Biosystems, Foster City, California). Sequences from opposite mtDNA strands were reconciled and verified for accuracy using SEQUENCHER 3.1 (Gene Codes, Ann Arbor, Michigan). Sequences are archived in GenBank (accession numbers AY506868-AY506870, AY506873-AY506901, AY506904-AY506908, AY506910-AY506917, AY506919-AY506944, AY506974-AY506984 [Kulikova et al. 2004], AY928831-AY928900 [present study]).

We used PAUP* 4.0b10 (Swofford 1998) and unweighted parsimony to estimate the phylogenetic relationships of each unique Mallard haplotype. Tree searches were performed in two steps: 500 random-addition replicates, each limited to 100 trees, followed by a single search, with no limit, using all minimum-length trees from the first round as starting trees. Searches were heuristic, with tree-bisection-reconnection (TBR) branch-swapping and gaps coded as a fifth character state. Two divergent mtDNA clades were observed, and all trees were rooted at the midpoint. We used the software TCS (Clement et al. 2000) to illustrate unrooted haplotype networks for each clade. Ambiguities in the haplotype network were resolved using the criteria suggested by Crandall and Templeton (1993).