Phylogenetic analysis
To analyze mtDNA sequences, the concatenated 1111-bp Cyt b and CR
sequence data from Jiangsu were aligned with NCBI sequences. One hundred
and seventeen unique haplotypes were identified from the data set
combining NCBI and 78 individuals, and 128 unique haplotypes were
identified from the data set combining NCBI and 173 individuals. Then,
79 unique haplotypes were identified by collapsing the concatenated
sequences from 2015 and 2016. For the 117 haplotypes, the
best-fit
model from Bayesian analysis was the GTR+I+G model. The software
Jmodeltest v2.1.7 identified the TrN+I+G model as most appropriate from
ML analyses. The GTR+I+G and TIM3+I+G models were identified as most
appropriate from BI and ML analyses of 128 haplotypes. The HKY+I+G and
TPM2uf+I+G models were suitable from BI and ML analyses of 79
haplotypes, respectively. The two trees were all paraphyletic and
consensus was from an overall perspective with credible bootstrap values
for all relationships among branches. Dunlins from a stopover in Jiangsu
shared haplotypes or formed sister relationships with haplotypes from
six subspecies, including C . a. actites , C .a. kistchinski , C . a. sakhalina , C. a.
hudsonia , C. a. pacifica , and C. a.
arcticola .
Haplotypes from Jiangsu and the breeding areas combined were clustered
into two
lineages,
Alaskan and Beringian lineages, based on the percentage of speculated
subspecies, which suggested that dunlins staging in Jiangsu migrated
from Beringian and Alaskan areas. The corresponding numbers of
individual dunlins are in parentheses in Fig. 2, and the numbers of
dunlin individuals at the end of each branch are shown in Tables S1 and
S2.
Fifty-six
individuals (78 in total) were inferred into the two lineages in 2015,
and the proportion identified was 71.79%. One hundred and thirty-six
individuals (173 in total) were inferred into two lineages in 2016, and
the proportion identified was 78.61%. Twenty-two individuals in 2015
and 37 individuals in 2016 could not be identified. The proportion of
Alaskan birds increased from 58.97% in 2015 to 63.58% in 2016, and the
proportion of Beringian birds increased from 12.08% in 2015 to 15.02%
in 2016. Simultaneously, the proportion of unknown birds decreased from
28.21% in 2015 to 21.39% in 2016. Three subspecies, C. a.
arcticola , C . a. sakhalina , and C . a.
kistchinski , were more common in Jiangsu than other subspecies (Fig.
2). Moreover, distinct from the phylogeny shown in Fig. 2a, there were
some haplotypes found only in Jiangsu that were assigned to a single
clade of unknown subspecies in Fig. 2b. Notably, some of the dunlin
individuals from Alaskan breeding areas detected at Jiangsu had a close
phylogenetic relationship with C. a. hudsonia.