Download Ex situ conservation status of an endangered Yangtze finless

ICES Journal of Marine Science, 62: 1711e1716 (2005)
doi:10.1016/j.icesjms.2005.06.002
Ex situ conservation status of an endangered Yangtze
finless porpoise population (Neophocaena phocaenoides
asiaeorientalis) as measured from microsatellites
and mtDNA diversity
Junhong Xia, Jinsong Zheng, and Ding Wang
Xia, J., Zheng, J., and Wang, D. 2005. Ex situ conservation status of an endangered Yangtze
finless porpoise population (Neophocaena phocaenoides asiaeorientalis) as measured from
microsatellites and mtDNA diversity. e ICES Journal of Marine Science, 62: 1711e1716.
The Yangtze finless porpoise, Neophocaena phocaenoides asiaeorientalis, is an endangered
small cetacean that occurs only in the middle and lower reaches of the Yangtze River of
China. The establishment of a breeding population of the porpoise in Tian-e-Zhou Baiji
National Natural Reserve represents the first attempt at ex situ conservation efforts for
a cetacean species. With the goal of effective protection, management, and monitoring of
this preserved population, we examined its genetic diversity using 930 bp of mtDNA
control region sequences and 13 polymorphic microsatellite loci. A very low level of
genetic variation (h Z 0.6010 G 0.0029 s.d.; p Z 0.0007 G 0.0000002 s.d.) in the mtDNA
control region sequences and a moderate genetic diversity (Ho Z 0.5740 G 0.2575 s.d.) in
the microsatellites were detected in the population. It is necessary to introduce more
individuals with representative genetic variations into the reserve in order to form a larger
and healthier group structure for long-term survival of the population. Successful
establishment of the Yangtze finless porpoise population in the Reserve also provides
a useful model for an ex situ conservation programme for other rare and endangered species.
Ó 2005 International Council for the Exploration of the Sea. Published by Elsevier Ltd. All rights reserved.
Keywords: ex situ conservation, genetic diversity, Neophocaena phocaenoides asiaeorientalis.
Received 6 June 2004; accepted 15 June 2005.
J. Xia and J. Zheng: Institute of Hydrobiology, the Chinese Academy of Sciences, Wuhan
430072 Hubei, China; Graduate School of the Chinese Academy of Sciences, Beijing
100039, China. D. Wang: Institute of Hydrobiology, the Chinese Academy of Sciences,
Wuhan 430072 Hubei, China. Correspondence to D. Wang: tel: C86 27 68780178; fax:
C86 27 68780123; e-mail: [email protected].
Introduction
With increasing human activities, large-scale habitats and
their associated environments have been destroyed or have
deteriorated, which consequently can lead to mass species
extinction. At present, there mainly exist two strategies for
conservation of an endangered species, i.e. in situ and ex
situ conservation. Although the best remedy to prevent
species extinction is habitat preservation (in situ conservation), ex situ conservation is very necessary to preserve
endangered or rare species when their natural habitats have
already been completely destroyed or so reduced in size
and so fragmented that the species are in imminent danger
of extinction (Li et al., 2002).
1054-3139/$30.00
The Yangtze finless porpoise, Neophocaena phocaenoides asiaeorientalis (Gao and Zhou, 1995), an endemic
and endangered small cetacean, only occurs in the middle
and lower reaches of the Yangtze River. This porpoise is
listed in the Second Order of Protected Animals in China and
as Endangered by the IUCN Red List of Threatened species.
Surveys from 1978 to 1991 produced an estimate of 2700
individuals in the Yangtze (Zhang et al., 1993). Additional
surveys conducted in some segments of the Yangtze from
1989 to 1999 indicated a drastic decrease (Wang et al.,
2000), with a loss rate of 7.3% per year (Wei et al., 2002b).
Population viability analysis suggested that the Yangtze
finless porpoise would become extinct within 24e94 years if
no protection was given (Zhang and Wang, 1999).
Ó 2005 International Council for the Exploration of the Sea. Published by Elsevier Ltd. All rights reserved.
1712
J. Xia et al.
Since further degradation of the Yangtze River is likely,
Wang et al. (2000) concluded that the establishment of ex
situ breeding colonies would be necessary as a means of
saving the Yangtze finless porpoise population from
extinction. In 1992, the Tian-e-Zhou Oxbow, located in
Shishou, Hubei, China, was approved by the central
government as a National Natural Reserve for the Baiji,
Lipotes vexillifer, a critically endangered cetacean species
also only occurring in the Yangtze River, and for the
Yangtze finless porpoise. This oxbow was formed naturally
when it was cut off from the main channel of the Yangtze
River in 1972. It is 21 km long, 1e1.5 km wide, and has an
average bottom depth of 4.5 m (Wei et al., 2002a). Instead
of the Baiji, the Yangtze finless porpoise was introduced
into the reserve first as early as 1990 for testing the
feasibility of the establishment of the Reserve during
a baseline study. This represents the world’s first attempt at
ex situ conservation efforts for a cetacean species (Zhang
et al., 1995; Wang et al., 2000; Wei et al., 2002a). Because
of escapment and release of Reserve porpoises into the
Yangtze River, there were only four individuals left in early
spring 1997. Thereafter, another three and six individuals
from the Yangtze River were introduced into the Reserve in
1998 and 1999, respectively (Wang et al., 2000; Wei et al.,
2002a). With 1e3 calves born in it each year, there were 22
animals living in the reserve when the present study started
(Wang et al., 2000; Wei et al., 2002a).
Although many ecological and behavioural studies have
been undertaken on this population (e.g. Yang et al., 1995;
Yang and Chen, 1996; Zhang et al., 1996; Akamatsu et al.,
1998, 2000, 2002; Wei et al., 2002a), no molecular data
from the population were available. As small isolated
populations are subject to genetic drift, which will affect
their evolutionary potential through fixation of deleterious
mutations, understanding the patterns of genetic variation
and the efficiency of genetic conservation in an ex situ
strategy is of fundamental importance. This study mainly
aims to explore the genetic variation and its maintenance to
provide guidelines for further conservation strategies and
management of this isolated population, and could be used
as a useful case study of ex situ conservation for other rare
and endangered species, such as the critically endangered
Baiji.
Material and methods
Sample collection
During early June 2002, we captured all 22 porpoises in the
Reserve by net under a special permit of the Department of
Fisheries Management, Hubei Province. Gender was
identified morphologically for each, and 10e20 ml whole
blood was drawn from each porpoise with a hypodermic
syringe. Blood was immediately anti-coagulated with ACD
(acidecitrateedextrose) solution and preserved in liquid
nitrogen until DNA extraction. Another female porpoise,
captured from the Reserve in 1999 and raised in the
Institute of Hydrobiology, the Chinese Academy of
Sciences, was also sampled. Therefore, 23 individuals were
used for analysing genetic diversity of the Reserve
population. Of these, 8 were females and 15 were males.
DNA extraction
Total genomic DNA was extracted using protocols based
on E.Z.N.A.Ò Blood DNA Kit (Omega Bio-tek, Inc.) and
re-hydrated in TE buffer (10 mM Tris, 0.1 mM EDTA, pH
8.0) for future use.
mtDNA control region sequencing
and microsatellite genotyping
A fragment of approximately 1.1 kb of the mtDNA control
region was amplified by PCR with primers (L: 5#-GAA
TTC CCC GGT CTT GTA AAC C-3# and H: 5#-TCT CGA
GAT TTT CAG TGT CTT GCT TT-3#; Hoelzel et al.,
1998). The PCR cycling profile consisted of an initial
denaturation of 3 min at 94(C, followed by 30 cycles of
40 s at 94(C, 1 min annealing at 63(C and 2 min at 72(C,
and with a final step of extension of 7 min at 72(C.
Amplified fragments were purified with QIAquickÒ PCR
Purification Kit and then sequenced from both ends with
primers L and H, respectively.
Thirteen microsatellite loci were amplified for each
individual in reaction, as described in Valsecchi (1996;
EV10pm, EV104Mn), Rosel et al. (1999; PPHO104,
PPHO137, PPHO110, PHO142, PHO130), Rooney et al.
(1999; Texvet2), Waldick et al. (1999; rw410, rw34), and
Berube et al. (2000; GT271, GT509, GT310). PCR
products were run on 6.0% denaturing polyacrylamide gels
and then detected using the silver staining technique.
Microsatellite alleles were sized relative to an internal
DNA size-standard (PBR322/MspI marker) using the
software Labimage (ver. 2.6, programmed by Kapelan,
http://www.labimage.de). Each locus was re-amplified and
run at least twice to ensure accuracy of scoring.
Data analysis
Control region sequences were aligned using the program
Clustal X (Jeffs, 1998), with all parameters set to default
values. After correction by hand, we combined L and H
sequences from the same sample. Population genetic
analyses such as haplotypic diversity (h) and nucleotide
diversity (p) were carried out using the program DnaSP
(ver. 3.15, Rozas and Rozas, 1999).
For microsatellite data, linkage disequilibrium tests
between pairs of loci, allele frequencies, effective number
of alleles (ne), Shannon’s Information Index (I), observed
heterozygosity (Ho), and expected heterozygosity (He) at
each locus were computed with the program POPGENE
(ver. 1.3.1, Yeh et al., 1997).
Conservation status of Yangtze finless porpoise
Results
Table 2. Summary of descriptive statistics for the Reserve
population at 13 microsatellite loci.
mtDNA data
Sequences (930 bp) of mtDNA control region were aligned
among the 23 individual samples. Two polymorphic sites
were detected and three unique haplotypes were defined
(H1, H2, and H3). Both mutations were transitions (Table 1).
The haplotypic diversity (h) was estimated to be
0.6010 G 0.0029 s.d. and nucleotide diversity (p) was
0.0007 G 0.0000002 s.d. among the 23 samples, showing
a very low level of genetic variation in the population for
mtDNA.
The Reserve population consisted primarily of the
individuals with haplotype H1 or H2, in which haplotypic
frequency for H1 was estimated to be 0.4783, and 0.4348
for H2. The male group possessed all the three haplotypes,
while only haplotypes H1 and H2 were detected within the
female group (Table 1).
Microsatellite data
No significant evidence of linkage disequilibrium was
detected among the 13 microsatellite loci used, so all
were kept for further analysis. The average effective
number of alleles (ne) and Shannon’s Information Index
(I) per locus were 2.82 G 1.60 s.d. and 1.05 G 0.45 s.d.,
respectively. The observed single locus heterozygosity
(Ho) of the population ranged from 0.2174 to 0.9565,
with an average of 0.5740 G 0.2575 s.d. at the 13 loci
(Table 2).
Discussion
Genetic status of the Reserve population
Previously, Yoshida et al. (2001) and Yoshida (2002) have
reported the genetic variation of five finless porpoise
populations inhabiting Japanese coastal waters. Their
studies indicated that the p values at mtDNA level ranged
from 0.000 (Omura Bay, n Z 8) to 0.0041 (Ariake SoundTachibana Bay, n Z 65), with an average of 0.0016
(n Z 173). The average value was just slightly higher than
Table 1. mtDNA sequence polymorphic sites and the estimation
and distribution of haplotype frequencies for each group.
Variable sites
Haplotype
H1
H2
H3
1713
143
414
C
T
T
C
Absolute Relative
Female Male frequency frequency
3
5
0
8
5
2
11
10
2
0.4783
0.4348
0.0869
Variable sites were identified with the 930 bp sequence, where a ‘’
represented identity with haplotype H1. Haplotype sequences were
deposited in GenBank (Accession No. AY334099eAY334101).
Loci
K
Ho
He
PPHO104
PPHO137
PPHO110
PPHO142
PPHO130
rw410
rw34
EV10pm
GT271
GT509
GT310
EV104Mn
Texvet2
3
6
7
4
7
2
6
3
3
3
2
5
2
0.4348
0.9444
0.9565
0.6522
0.8696
0.2174
0.4348
0.3000
0.4348
0.5652
0.4348
0.8696
0.3478
0.5952
0.8365
0.8541
0.5729
0.8242
0.3217
0.4995
0.3090
0.5179
0.4792
0.5024
0.6435
0.4870
Mean
s.d.
4.076
1.891
0.5740
0.2575
0.5725
0.1778
The observed number of alleles (K), and the observed and expected
heterozygotsities (Ho and He) for each locus were reported; the
mean and the standard deviation (s.d.) at 13 microsatellite loci were
also reported.
that obtained from the Reserve Yangtze finless porpoise
population in this study (p Z 0.0007, h Z 0.6010, n Z 23).
However, comparison of the mtDNA data of our study with
those detected in other cetacean species, e.g. harbour
porpoise, Phocoena phocoena (p Z 0.0110, h Z 0.93,
n Z 253; Rosel et al., 1999) and Dall’s porpoise,
Phocoenoides dalli (p Z 0.0142, h Z 0.952, n Z 113;
Escorza-Trevino and Dizon, 2000) revealed a very low
level of genetic variation in the Reserve’s Yangtze finless
porpoise population. Additionally, some recent mtDNA
sequence analyses also detected a very low level of genetic
diversity in the wild Yangtze River population (p Z 0,
n Z 4, Yang and Zhou, 1997; p Z 0.0015, h Z 0.60,
n Z 6, Yang et al., 2002; p Z 0.0011, h Z 0.65, n Z 39,
Zheng et al., 2005). All these data, therefore, indicated that
a very low genetic variability at mtDNA level was a feature
of the Yangtze finless porpoise. In addition, the heterozygosity values of many cetacean species measured by
microsatellites were higher than 0.6 (Rooney et al.,
1999), e.g. harbour porpoise (Ho Z 0.834, n Z 253; Rosel
et al., 1999) and Dall’s porpoise (Ho Z 0.864, n Z 119;
Escorza-Trevino and Dizon, 2000). However, for the
endangered North Atlantic right whale, Eubalaena glacialis, Ho was estimated to be only 0.5 (Waldick et al., 1999).
Compared with these studies, the average heterozygosity
(Ho Z 0.5740) reported here for the Yangtze finless
porpoise population in the Reserve was moderate.
Generally, the loss of genetic diversity through a bottleneck occurs over a number of generations. Therefore, being
composed of several stocks caught from different segments
1714
J. Xia et al.
of the Yangtze River and having just a recent population
history (Wang et al., 2000; Wei et al., 2002a), the Reserve
population should have conserved similar genetic variation
of the Yangtze population. However, the finless porpoise
population in the Reserve may poorly represent the natural
population as the founder population is small (Wang et al.,
2000; Wei et al., 2002a). In a recent study on the Yangtze
finless porpoise wild population that included 39 individuals from the middle and lower reaches of the Yangtze
River, seven mtDNA haplotypes were detected using
mtDNA control region sequence analysis (Zheng et al., in
press). By comparison, we found that all the three
haplotypes detected in the Reserve population were
included in the seven haplotypes of the wild population.
This means that only about 42.86% of the mtDNA genetic
variation of the wild Yangtze River population was
conserved in the Reserve population. This implies that the
low genetic diversity of the Reserve population may mainly
be due to the wild population being poorly represented in
the Reserve. One of the main reasons for this poor
representation might be that besides small founder
population, most founders of the Reserve population were
from the middle reaches of the Yangtze River (Wang et al.,
2000; Wei et al., 2002a), where the genetic diversity of the
population is less than in the lower reaches (Zheng et al., in
press). Consequently, the conserved porpoise population in
the reserve could not represent the genetic diversity of the
wild population. Another possibility is that the finless
porpoise were caught in groups in the River, and members
of the groups might be related to each other. However, our
recent relatedness analysis on some porpoise groups in the
Reserve indicates that except for a lactating mother and
child group, the pairwise relatedness among individuals
within a group was relatively random (J. Xia et al.,
unpublished data). Therefore, the possibility of close
relatedness seems to be eliminated. Additionally, the study
also showed that the mtDNA haplotype H3 was only
detected within the males and not within the females in the
Reserve population. As mtDNA is characterized by
inheritance from mother to offspring, if no new females
are introduced, this haplotype H3 is likely to be lost in the
future.
The principal objective of ex situ conservation is to
establish secure and self-sustaining populations, which can
ultimately be released into preserved or restored habitat
(Krauss et al., 2001; Li et al., 2002). For such a conservation programme, successful reproduction is an essential
short-term goal, whereas the maintenance of initial genetic
diversity of wild populations is the long-term objective
(Krauss et al., 2001; Morgan, 2000). Long-term ecological
studies indicated that the Yangtze finless porpoise could
breed successfully in the Reserve, confirming that the
Reserve is an appropriate habitat for the porpoise (Wang
et al., 2000; Wei et al., 2002a). However, the current
conserved genetic diversity in the Reserve is neither
representative nor adequate.
Consequences of the population status
Isolation of a rare and small population will decrease
genetic variation and reduce fitness due to genetic drift and/
or inbreeding, and the extent of inbreeding in a translocated
population is significantly higher than what would be likely
in natural populations, consequently increasing the risk of
population extinction (Mitton and Grant, 1984; Caughley,
1994; Frankham, 1995, 1998; Lacy, 1997; Bosch et al.,
1998; Krauss et al., 2001). In our study, molecular analysis
did detect a highly low level of genetic diversity at mtDNA
level, which showed that the Reserve population would be
highly vulnerable and susceptible to some demographic and
environmentally stochastic factors such as loss of individuals or epidemic diseases. Furthermore, an effective
population size (Ne) of approximately 50 has been used
as a theoretical minimum for long-term self-sustaining of
a population. Generally, Ne is 0.25e0.75n (Nunney and
Elam, 1994; Waite and Parker, 1996), so the Ne of the
Reserve population could be roughly estimated to be
5.5e16.5. As the Reserve Ne was relatively small, and the
genetic variation, as discussed above, was low and cannot
represent the real status of the wild population, there might
be little hope for the long-term survival of this Reserve
population if no further protection measures are taken.
Recommendations for conservation
As the status of the finless porpoise in the Yangtze River is
becoming increasingly serious, ex situ protection will play
a very important role in its conservation (Wang et al., 2000).
For ex situ conservation, knowledge of genetic diversity and
structure, and gene flow is important. However, previous
molecular analyses on this population were few, focused
mainly at the level of mtDNA variation, and most of the
studies were limited by small sample size (e.g. n Z 4, Yang
and Zhou, 1997; n Z 6, Yang et al., 2002). It is, therefore,
crucial to conduct more genetic investigations on this
species at both nuclear and mtDNA levels, to explore
genetic diversity and to distinguish unambiguous management units (MUs) and evolutionarily significant units
(ESUs) throughout its distribution range. In addition, the
long-term viability of a population probably depends on the
genetic variability it retains. For effective conservation, 95%
of the total genetic diversity of a species must be contained
(Li et al., 2002). At present, owing to the lack of systematic
ecological surveys, the actual population size of the Yangtze
finless porpoise is unknown. However, it was roughly
estimated to be 2700 based on survey results from 1978 to
1991, and the current total population may be less than 2000
(Zhang et al., 1993; Wang et al., 2000; Wei et al., 2002b).
Therefore, there still exists a relatively large effective
population size in the wild with much more possibly
available genetic variation to supply the Reserve population.
To maximize the genetic variation included in the Reserve
population and to form a larger and healthier group structure
for long-term self-sustainability, and to achieve the objective
Conservation status of Yangtze finless porpoise
of precluding the loss of rare alleles and decreasing
heterozygosity in the ex situ conserved population, porpoises
should be collected representatively from different source
stocks in the river and translocated into the Reserve.
The Baiji, which occurs sympatrically in the Yangtze
River with the Yangtze finless porpoise, is one of the two
critically endangered cetaceans in the world with fewer
than 100 individuals left (Liu et al., 1996). Many scientists
suggested that it was almost impossible to conserve this
species in the Yangtze River owing to its very small
effective population size and deterioration of its habitat;
most urgent thing for its conservation is to translocate as
many Baiji as possible from the Yangtze River into the
Reserve to establish an ex situ breeding colony (Liu et al.,
1996; Wang et al., 2000). The successful establishment of
the Yangtze finless porpoise Reserve population provided
a useful model of an ex situ conservation programme.
Clearly, such a programme can provide significant guidelines for conservation strategies for other endangered
species.
Acknowledgements
We thank X. Zhang, Q. Zhao, Z. Wei, K. Wang, X. Wang,
and B. Yu of our department and staff of the Tian-e-Zhou
Baiji National Natural Reserve for their support in the
capture operation for this study. Dr Yunhan Hong and two
anonymous reviewers provided valuable and constructive
comments on an earlier version of this manuscript. The
research was supported by National Natural Science
Foundation of China (No. 30170142), the Chinese Academy of Sciences (CAS) (No. KSCX2-SW-118), Ministry of
Science and Technology of China (No. 2004DFB03000),
and the Institute of Hydrobiology, CAS (No. 220103).
References
Akamatsu, T., Wang, D., Nakamura, K., and Wang, K. X. 1998.
Echolocation range of captive and free-ranging baiji (Lipotes
vexillifer), finless porpoise (Neophocaena phocaenoides), and
bottlenose dolphin (Tursiops truncatus). Journal of the Acoustical Society of America, 104: 2511e2516.
Akamatsu, T., Wang, D., Wang, K. X., and Naito, Y. 2000. A
method for individual identification of echolocation signals in
free-ranging finless porpoises carrying data loggers. Journal of
Acoustics Society of America, 108: 1353e1356.
Akamatsu, T., Wang, D., Wang, K. X., Wei, Z., Zhao, Q. Z., and
Naito, Y. 2002. Diving behavior of freshwater finless porpoises
(Neophocaena phocaenoides) in an oxbow of the Yangtze River,
China. ICES Journal of Marine Science, 59: 438e443.
Berube, M., Jorgensen, H., McEwing, R., and Per, J. P. 2000.
Polymorphic di-nucleotide microsatellite loci isolated from the
humpback whale, Megaptera novaeangliae. Molecular Ecology,
9: 2155e2234.
Bosch, M., Simon, J., Molero, J., and Blanché, C. 1998.
Reproductive biology, genetic variation and conservation of
rare endemic dysploid Delphinium bolosii (Ranunculaceae).
Biological Conservation, 86: 57e66.
1715
Caughley, G. 1994. Directions in conservation biology. Journal of
Animal Ecology, 63: 215e244.
Escorza-Trevino, S., and Dizon, A. E. 2000. Phylogeography,
intraspecific structure and sex-based dispersal of Dall’s porpoise,
Phocoenoides dalli, revealed by mitochondrial and microsatellite DNA analyses. Molecular Ecology, 9: 1049e1060.
Frankham, R. 1995. Conservation genetics. Annual Review of
Genetics, 29: 305e327.
Frankham, R. 1998. Inbreeding and extinction: island populations.
Conservation Biology, 12: 665e675.
Gao, A., and Zhou, K. 1995. Geographical variation of external
measurements and three subspecies of Neophocaena phocaenoides in Chinese waters. Acta Theriologica Sinica, 15:
81e92.
Hoelzel, A. R., Natoli, A., Dahlheim, M. E., Olavarria, C., Baird,
R. W., and Black, N. A. 1998. Low genetic variation among
killer whales (Orcinus orca) in the eastern North Pacific and
genetic differentiation between foraging specialists. Journal of
Heredity, 89: 121e128.
Jeffs, P. 1998. Multiple sequence alignment with Clustal X.
Computer Corner, 23: 78e80.
Krauss, S. L., Dixon, B., and Dixon, K. W. 2001. Rapid genetic
decline in a translocated population of the endangered plant
Grevillea scapigera. Conservation Biology, 16: 986e994.
Lacy, R. C. 1997. The importance of genetic variation to the
viability of mammalian populations. Journal of Mammalogy, 78:
320e335.
Li, Q., Xu, Z., and He, T. 2002. Ex situ genetic conservation of
endangered Vatica guangxiensis (Dipterocarpaceae) in China.
Biological Conservation, 106: 151e156.
Liu, R. J., Zhang, X. F., Wang, D., and Yang, J. 1996. Once again
studies on the conservation of Lipotes vexillifer and Neophocaena phocaenoides. Resources and Environment in the
Yangtze Valley, 5: 220e225.
Mitton, J. B., and Grant, M. C. 1984. Associations among protein
heterozygosity, growth rate and developmental homeostasis.
Annual Review of Ecology Systematics, 15: 479e499.
Morgan, J. W. 2000. Reproductive success in reestablished versus
natural populations of a threatened grassland daisy (Rutidosis
leptorrhynchoides). Conservation Biology, 14: 780e785.
Nunney, L., and Elam, D. R. 1994. Estimating the effective
population size of conserved populations. Conservation Biology,
8: 175e184.
Rooney, A. P., Merritt, D. B., and Derr, J. N. 1999. Brief
communications: microsatellite diversity in captive bottlenose
dolphins (Tursiops truncatus). The Journal of Heredity, 90:
228e231.
Rosel, P. E., France, S. C., Wang, J. Y., and Kocher, T. D. 1999.
Genetic structure of harbour porpoise Phocoena phocoena
populations in the northwest Atlantic based on mt and nuclear
markers. Molecular Ecology, 8: 41e54.
Rozas, J., and Rozas, R. 1999. DnaSP version 3: an integrated
program for molecular population genetics and molecular
evolution analysis. Bioinformatics, 15: 174e175.
Valsecchi, E. 1996. Cetacean microsatellites. Genebank (http://
www.ncbi.nlm.nih.gov/; accession number: G27347).
Waite, T. A., and Parker, P. G. 1996. Dimensionless life histories
and effective population size. Conservation Biology, 10:
1456e1462.
Waldick, R. C., Brown, M. W., and White, B. N. 1999.
Characterization and isolation of microsatellite loci from the
endangered North Atlantic right whale. Molecular Ecology, 8:
1753e1768.
Wang, D., Liu, R., Zhang, X., Yang, J., Wei, Z., Zhao, Q., and
Wang, X. 2000. Status and conservation of the Yangtze finless
porpoise. In Biology and Conservation of Freshwater Cetaceans
in Asia, 1st edn, pp. 81e85. Ed. by R. R. Reeves, B. D. Smith,
and T. Kasuya.
1716
J. Xia et al.
Wei, Z., Wang, D., Kuang, X., Wang, K., Wang, X., Xiao, J., Zhao,
Q., and Zhang, X. 2002a. Observations on behavior and ecology
of the Yangtze finless porpoise (Neophocaena phocaenoides
asiaeorientalis) group at Tian-e-Zhou Oxbow of the Yangtze
River. The Raffles Bulletin of Zoology Supplement, 10: 97e103.
Wei, Z., Wang, D., Zhang, X., Zhao, Q., Wang, K., and Kuang, X.
2002b. Population size, behavior, movement pattern and
protection of Yangtze finless porpoise at Balijiang section of
the Yangtze River. Resources and Environment in the Yangtze
Basin, 11: 427e432.
Yang, G., and Zhou, K. 1997. Variability of the mitochondrial control
region in population of finless porpoise, Neophocaena phocaenoides, in Chinese waters. Acta Zoologica Sinica, 43: 411e419.
Yang, G., Zhou, K., Ren, W., Liu, S., Ji, G., and Yan, J. 2002.
Population genetic structure of finless porpoises, Neophocaena
phocaenoides, in Chinese waters, inferred from mitochondrial
control region sequences. Marine Mammal Science, 18: 336e347.
Yang, J., and Chen, P. 1996. Movement and behavior of finless
porpoise (Neophocaena phocaenoides) at Swan Oxbow, Hubei
Province. Acta Hydrobiologica Sinica, 20: 32e40.
Yang, J., Zhang, X., Zhou, W., Wang, X., and Chen, P. 1995. The
preliminary observation on living habits of the experimental
rearing finless porpoise in Swan Oxbow, Hubei Province. Acta
Theriologica Sinica, 15: 254e258.
Yeh, F. C., Yang, R. C., Boyle, T. B. J., Ye, Z. H., and Mao, J. X.
1997. POPGENE, the User-friendly Shareware for Population
Genetic Analysis. Molecular Biology and Biotechnology Centre.
University of Alberta, Canada.
Yoshida, H. 2002. Population structure of finless porpoises
(Neophocaena phocaenoides) in coastal waters of Japan. The
Raffles Bulletin of Zoology Supplement, 10: 35e42.
Yoshida, H., Yoshika, M., Chow, S., and Shirakihara, M. 2001.
Population structure of finless porpoises (Neophocaena phocaenoides) in coastal waters of Japan based on mitochondrial DNA
sequences. Journal of Mammology, 82: 123e130.
Zhang, X., and Wang, K. 1999. Population viability analysis for the
Yangtze finless porpoise. Acta Ecologica Sinica, 9: 529e533.
Zhang, X., Lin, R., Zhao, Q., Zhang, G., Wei, Z., Wang, X., and
Yang, J. 1993. The population of finless porpoise in the middle
and lower reaches of Yangtze River. Acta Theriologica Sianica,
13: 260e270.
Zhang, X., Wang, D., Yang, J., Wei, Z., and Wang, K. 1996. Study
on radio-tracking finless porpoise Neophocaena phocaenoides,
at the Yangtze River. Acta Ecologica Sinica, 16: 490e496.
Zhang, X., Wei, Z., Wang, X., Yang, J., and Chen, P. 1995. Studies
on the feasibility of establishment of a semi-natural reserve at
Tian-e-Zhou (Swan) Oxbow for baiji, Lipotes vexillifer. Acta
Hydrobiologica Sinica, 19: 110e123.
Zheng, J., Xia, J., He, S., and Wang, D. 2005. Population genetic
structure of the Yangtze finless porpoise (Neophocaena
phocaenoides asiaeorientalis): implications for management
and conservation. Biochemical Genetics, 43: 307e320.