The Body Condition index and Genetic connectivity among foraging Green Turtle Populations in Indonesia at different scales. A case study from Berau Green Turtle Rookery, East-Kalimantan.
THE BODY CONDITION INDEX AND GENETIC CONNECTIVITY AMONG FORAGING GREEN TURTLE
POPULATIONS IN INDONESIA AT DIFFERENT SCALES :
A CASE STUDY FROM BERAU GREEN TURTLE ROOKERY, EAST-KALIMANTAN
Windia Adnyana
The Faculty of Veterinary Medicine, Udayana University, Bali - Indonesia
SUMMARY
The purpose of this study is to assess the body condition index of green turtles at three different seagrass
meadows of the Berau Isles, assess the genetic composition of green turtles, identify presence or absence of
genetic differentiation among the green turtles reside in the three foraging areas, and to identify links among
these foraging areas and various nesting sites in the Australasian region.
Sampling were carried out between 8 to 23 December 2009 from three main foraging locations within the Berau
Isles. The selected foraging habitats were the water in the northern side of Pulau Derawan, near Pulau Panjang,
and the water of Payung-Payung near Maratua Island. Turtles were captured either by net or rodeo technique,
taken onboard, their biometric values were measured and the skin samples were taken for genetic analysis.
A total of 310 Chelonia mydas were captured and observed. Most of them (50.32%) are medium size turtles with
Curved Carapace Length between 60-80 cm. The proportions of turtle with CCL > 80 cm and with CCL < 60 cm
were 27.10% and 22.58%, respectively. Their bodyweight ranged from 7.70 kg to 158.90 kg. In view of their sex
status which was determined based on the Total Tail Length, 17.42% were defined to be males and 10.32% were
females. Most turtles (72.26%) were sexually undifferentiated by means of external characteristics. Thir Body
Condition Index (BCI) varied between 0.71 – 1.80. Most turtles were in very good (70.0%) and good (11.29%)
conditions, as compared to average (8.06%) and poor conditions (10.65%). Based on their capture locations, the
highest BCI was calculated for Payung-Payung population (1.30±0.21; range=0.80–1.66; n=102), followed by
Pulau Derawan (1.26±0.16; range=0.84–1.56; n=117) and Pulau Panjang population (1.24±0.20; range=0.711.80; n=91). Statistical analysis showed that there was no significant difference between the population of Pulau
Panjang and Pulau Derawan (P=0.443). Significant difference (P=0.017) was found between the population of
Pulau Panjang and Payung-Payung., but not-significant difference was calculated between Payung-Payung and
Pulau Derawan (P=0.078).
A total of 213 mitochondrial (mt) DNA fragments, out of 309 collected samples were amplified by PCR technique.
Screening of polymorphism within 384 bp mtDNA control region fragments identified 34 polymorphic sites and 17
distinct haplotypes. The most frequent haplotype identified in Berau waters was D2 (40.8%), followed by C3
1
(19.7%), A3 (12.2%), C5 (10.8%), and C14 (6.6%). These five variants accounted for 90.1% of the total resident
foraging population. The other haplotypes pesent in relatively small proportion, ranged from 0.5% to 1.9%.
Despite of minor variations, there was no significant difference (P>0.05) found on the genetic structure of resident
populations in the Pulau Panjang, Pulau Derawan, and Payung-Payung foraging habitats. Mixed stock analysis
(MSA) results revealed that the feeding populations of Berau water were mainly composed of green turtles from
the Turtle Islands Heritage Protected Area (TIHPA) (45.49%), the nesting populations of Berau rookery (26.82%),
Micronesia (9.3%) and Papua New Guinea (8.44%) nesting populations. Small proportion of representatives from
the nesting sites in the South China Sea regions and Aru were also found. The finding emphasizes the need to
build a network of turtle based-MPA across SSME - BSSE - and Micronesian regions.
The population structure of green turtles resided in these three foraging sites were similar, but individual
exchange among the foraging sites is unlikely, which implied that each feeding habitat should be managed
separately. The body condition index of green turtles from Pulau Panjang and Pulau Derawan were significantly
lower than their counterpart which were captured in Payung-Payung. This finding, perhaps, associated with the
relative distance of the foraging habitat to the river come from the mainland of Kalimantan. Payung-Payung is
relatively far away from the mainland as compared to the other two, and possibly the sedimentation substrate
which influences the fertility of the sea grass are present in lower concentration in this water. Managing the
cleanliness of the river will help in maintaining the fertility of the seagrass beds in Pulau Panjang and Pulau
Derawan.
2
INTRODUCTION
The Berau Islands complex is an important nesting and foraging area for the green sea turtle (Chelonia mydas) in
Indonesia. This area is located within coordinates 02049’42.6’’ - 0102’0.06’’N; 117059’17.16’’ - 11902’50.30’’S and
belongs to the Berau district in East-Kalimantan (Figure-1). The area encompasses over 1.2 millions ha of
coastal area across 31 small islands, nine of which represent important nesting areas for green turtles. Annual
census surveys since 2002, suggest that the Berau Islands have one of the highest density of nesting green sea
turtles in the South-East Asia region (Adnyana et al, 2007). Additionally to nesting beaches, the Berau Isles also
contains several large seagrass meadows, which provide a critical foraging habitat for green turtles. Recent
surveys, conducted by researchers from the Radboud University in Nijmegen in collaboration with WWF
Indonesia and Udayana University, revealed that the density of green turtles on the seagrass meadows in this
area is among the highest in the world (Christianen in prep). The average green turtle density on the foraging
grounds is 17 ± 1.5 individuals per hectare.
The spatial extend of some of the seagrass meadows in Berau region is declining. The cause of this decline is
believed to be related to increased nutrient and sediment loads as a result of upstream erosion in the Berau river.
A reduced area of available nutrients could potentially lead to a reduced fitness of a green turtle population. The
resilience of a population under stress of e.g. seagrass loss or harvesting depends on the propensity of individual
turtles to switch foraging grounds and on the onset of the switching. Mark-recapture studies of foraging green
turtles from all size classes (Christianen in prep) suggest that subadult green turtles are not switching between
foraging grounds on a small spatial scale (i.e.within the Berau Isles) and thus, individuals associated with the
declining seagrass meadows are at risk of undernourishment. The body condition index (BCI) we be measured to
infer food availability and quality at the different foraging areas within the Berau water.
Mark-recapture studies of adult green turtles have revealed links between the nesting and the foraging
populations within the Sulu-Sulawesi marine Ecoregion. For example, turtles that received a tag while nesting on
the beach of Palau, the Philippines, and Malaysia (Sarawak, and Sabah) were later recaptured while foraging at
Derawan, Panjang or Maratua waters (WWF pers. Comm.). In addition, some individuals were tracked using
satellite telemetry (for a maximum of 155 days) and found to migrate from the Derawan and Sangalaki Islands to
Sabah and the Philippines (Adnyana et al. 2007) after nesting. While these observations provide valuable insight
into individual movement patterns, the relatedness of the Derawan green turtle population to populations in other
parts of the Australasian region, as well as an understanding of genetic differentiation among feeding grounds
within the archipelago remains unresolved. An assessment of the genetic composition of the Berau foraging
population examines the relative contribution of green turtle breeding populations to this assemblage, thus
providing a good understanding of the migratory pathways of green turtles to and from the Berau Archipelago.
3
This study is part of a PhD project on “sea grasses under green turtle grazing and nutrient loads”, conducted by
Marjolijn Christianen, Radboud University Nijmegen. This project primarily builds on previous genetic studies of
Australasian green turtle nesting populations, identifying 17 genetically distinct breeding stocks (Dethmers et al.
2006). This work is critical as the success of green turtle management strategies is contingent on understanding
of their population dynamics.
Objectives of this study
1. Assess the body condition index of green turtles at three different seagrass meadows of the Berau Isles
2. Assess the genetic composition of green turtles at the three main seagrass meadows of the Berau Isles
3. Identify presence or absence of genetic differentiation among the three foraging areas.
4. Identify links among these foraging areas and the Australasian 17 genetic stocks
4
Figure-1: A map showing the locations of three different foraging habitats within the Berau Archipelago. The
approximation of geographic location of this Isles is 02049’42.6’’ - 0102’0.06’’N; 117059’17.16’’ - 11902’ 50.30’’S.
Above is overall picture of the Berau water; below is the exact locations of sampling in Pulau Panjang (left), Pulau
Derawan (middle), and Payung-Payung, Maratua (right).
5
MATERIALS AND METHODS
Sample collection, measurements, maturity and sexual determination
Sampling were done during 15 working days, from 8 to 23 December 2009 by a team led by Marjolijn Christianen,
a PhD candidate from Radboud University Nijmegen. All samples were taken from three main foraging locations
within the Berau Islands complex, i.e. the water in the northern side of Pulau Derawan, near Pulau Panjang, and
the water of Payung-Payung near Maratua Island (see Figure-1). Turtles were captured either by net or rodeo
technique (jumping straightly to the turtles), taken onboard, and mesured for their Curved Carapace Length
(CCL), Total Tail Length (TTL), and their body-weight (BW). All of these measurement were carried out following
procedures provided by Bolten (1999) and the protocol provided by Adnyana and Hitipeuw (2009).
In view of the maturity status, based on their CCL, turtles with a CCL 80 cm were defined as mature. This criteria is decided based on our field experiences while
observing nesting green turtles on Derawan and Sangalaki Islands in which their minimum CCL was 80 cm. The
sex was predicted by looking at the TTL. Turtles of all size classes with TTL >20 cm were defined as males, while
those with TTL up to 20 cm and the CCL measuring more than 80 cm were defined as females. Immature turtles
(CCL up to 80 cm) with TTL less than 20 cm were considered as unsexed or sexually undetermined.
The Body condition index (BCI), a variable to indicate a decline in individual and population health which needed
to inform inshore management strategies, calculated using Equation derived from Bjorndal et al. (2000). BCI
3
quantifies body condition based on a ratio of weight and SCL. The equation is BCI = ([Weight (kg) / SCL(cm) ] x
10000). The straight carapace length (SCL) did not measured during this study. It was estimated from the values
of the CCL by using an equation SCL = 15 + (0.76 X CCL). This equation was made based on field records
obtained from Derawan nesting island monitoring workers. Total data used for linear regression calculation was
285, and the coeficient correlation (R) and R 2 were 0.771 and 0.594, respectively.
Any observed external morphological lesions on the turtle skin, carapace and plastron such as presence of scars,
wound, notch on marginal scutes, algae, barnacles, and fibropapillomatosis were recorded. Presence of metal
tags in the pliffers were noted, and turtles without tag were given a new one following the protocol provided by
Balazs (1999). Prior to be released back to the water, from each captured turtles, skin samples were taken from
flippers or neck region by using a biopsy punch, and immediately stored in either alcohol 70% or a NaCl saturated
solution of 20% DMSO, and transported to Udayana University for subsequent processes.
6
Molecular Methods
Genomic DNA was isolated from 0.1 g of skin tissue using Qiamp™ DNA Mini Kit from Qiagen®, and stored at 20oC for subsequent polymerase chain reaction (PCR). Successful DNA isolation was confirmed by running 2 μL
of genomic DNA in an ethidium bromide added 1% Agarose gel. A 740 bp segments of the mtDNA control region
for all samples were amplified using LTEi9 (5'-AGCGAATAATCAAAAGAGAAGG-3') and H950 (5'GTCTCGGATTTAGGGGTTTG-3') primers (Abreu-Grobois et al, 2006). The first primer binds at the border
between the tRNA-Thr and tRNA-Pro loci, and the second at position 782 near the end of the d-loop, expected to
produce PCR products of about 860 base pairs (bp). The target mitochondrial sequences were amplified by PCR
using approximately 50 ng of template genomic DNA in 25 µl reaction volume containing 14.3 µl H2O, 2.5 µl DNA
genome, 1.5 units ampli Taq gold polimerase (applied biosystem), 1.5 µl PCR buffer (applied biosystem), 2.5 µl
MgCl2 25 Mm, 2 µl dNTP 1 Mm, 1 µl of each primer 10 Mm. The PCR profile comprised an initial denaturation of
5 min at 94°C (to activate the Ampli Taq gold polymerase), followed by 40 cycles of: 94o C for 45 sec
(denaturation), 55o C for 45 sec (annealing), 72o C for 45 sec (extension), ended in a final extension in 72 o C for 4
min. Electrophoresis was performed to confirm the result of amplification and to determine the length of PCR
product. One µl loading dye (bromophenol-blue and cyline cyanol) was added in 2 µl PCR product.
Electrophoresis was run for negative and positive control (marker) for 30 minutes in 50 Volt of 1% agarose gel
media with etidum bromide dye. PCR product was sent to Macrogen Inc. (Korea) for forward and reverse
sequencing.
Statistical Analysis
Biometrics data were analysed using the Statistical Package for Social Sciences (SPSS) version 13.0. Graphical
presentations were completed either by Microsoft EXCEL version 2010 or SPSS version 13.0. Sequences were
aligned using Clustal X (Thompson et al 1997) and the population parameters such as polimorphic sites, the
percentages of each haplotype, haplotype diversity, and nucleotide diversity were analyzed using DNAsp 4.10
(Rozas et al., 2003). Phylogenetic tree was constructed to visualize the relationship among the observed mt-DNA
variants. AMOVA (Excoffier et al 1992), Exact tests of population differentiation (Raymond & Rousset 1995) and
pairwise Fst tests (Slatkin 1991) implemented in the population genetics package Arlequin version 3.01 (Excoffier
et al, 2006) were used to examined genetic structure among surveyed populations. Mixed stock analysis, to
predict the individual contribution from nesting colonies or management unit to the population of green turtles
reside in the foraging grounds of Pulau Panjang, Pulau Derawan, and Payung-Payung were calculated BAYES
program (Pella and Masuda, 2001).
7
RESULTS
Biometrics, maturity and sex status of Green Turtles in Berau Foraging Grounds
There was a total of 310 individual Chelonia mydas captured during 15 days sampling period during 8 December
– 23 December 2009. Their Curved Carapaca Length (CCL) encompassed all size classes (Figure-2) from small
immatures (CCL=40.80 cm) to large adults (CCL=111.30 cm). The mean ± standard deviation of the CCL was
71.69 ± 15.30 cm. When the CCL values were pooled into three categories (Figure-3), the majority of samples
were found to be medium size with CCL between 60-80 cm (50.32%). The proportions of large-adult (CCL > 80
cm) and small immature turtles with CCL < 60 cm were 27.10% and 22.58%, respectively.
Based on their actual capture sites, the percentage of mature-adult turtles (CCL>80 cm) was proportionally
highest in the Payung-Payung foraging ground as compared to the population of Pulau Panjang and Pulau
Derawan (Table-1). Consequently, as shown in Figure-4, the highest mean value for CCL was also found in the
foraging population of Payung-Payung (75.35±16.90 cm; range = 43.50–111.30 cm; n = 102), followed by Pulau
Panjang (70.63±15.02; range = 40.80–102.40 cm; n = 91) and the foraging population of Pulau Derawan
(69.32±13.46; range = 46.80–104.50 cm; n = 91). Statistically, a one way anova test suggested that significant
differences were calculated between the mean value of CCL of Payung-Payung population and the other two
populations, i.e. Pulau Panjang (P=0.031) and Pulau Derawan (P=0.003). However, no significant difference was
calculated between Pulau Panjang and Pulau Derawan (P=0.535).
Table-1: Size classes indicated by the Curved Carapace Length (CCL) of the Chelonia mydas captured in three
foraging grounds of the Berau Isles.
Capture Locations
Pulau Panjang
Pulau Derawan
Payung-Payung
Combined
>80 cm
21 (23.08%)
24 (20.51%)
39 (38.24%)
84 (27.10%)
Size Class
60-80 cm
51 (56.04%)
64 (54.70%)
41 (40.20%)
156 (50.32%)
20 cm. The summary values for maturity and sex
status per capture locations were presented in Table-2.
40
Frequency
30
20
10
Mean = 71.7177
Std. Dev. = 15.31384
N = 310
0
40.00
60.00
80.00
100.00
120.00
CCL
Figure-2: Histogram showing frequency distribution of the curved carapace length (CCL) of 310 green sea turtles
captured, tagged and measured in three foraging grounds of Berau water. Normal curve is displayed.
9
60.0%
50.0%
Percent
40.0%
30.0%
50.32%
20.0%
27.1%
22.58%
10.0%
0.0%
>80 cm
60-80 cm
80 cm)
Immature (CCL < 80 cm)
Male
Female
Male
Female Undetermined
12 (13.19%) 9 (9.89%)
- 70 (76.92%)
16 (13.68%) 8 (6.84%)
- 93 (79.49%)
24 (23.53%) 15 (14.71%) 2 (1.96%)
- 61 (59.80%)
2
(0.65%)
52 (16.77%) 32 (10.32%)
224 (72.26%)
Total
91 (100%)
117 (100%)
102 (100%)
310 (100%)
The Body Condition Index (BCI) of all samples varied between 0.71 – 1.80, normally distributed (Figure-7) with
the smallest and the highest values were both found in turtles captured from Pulau Panjang. Based on the criteria
defined by Bjorndal et al (2000) (Table-3), it was found that (Figure-8) most turtles were in very good (70.0%)
and good (11.29%) conditions, as compared to average (8.06%) and poor conditions (10.65%).
Regarding their size classes, as shown in Figure-9, the highest mean value for BCI (1.44±0.12; range = 1.141.66; n = 84) was noted in mature-adult turtles measuring > 80 cm. The mean values for size classes between 6080 cm and 1.20
Very Good
1
1.11 – 1.20
Good
2
1.00 – 1.10
Average
3
< 1.00
Poor
Determined from data presented by Bjorndal, K. A., Bolten, A. B. and Chaloupka, M. Y. (2000). Green turtle
somatic growth model: evidence for density dependence’. Ecological Applications 10, 269-282.
70.0%
60.0%
50.0%
Percent
40.0%
70.0%
30.0%
20.0%
10.0%
11.29%
10.65%
8.06%
0.0%
Very good
Good
Average
Poor
Body Condition Index
Figure-8: The Body Condition Index (BCI) of all samples captured from three foraging sites in the Berau water.
Note that most turtles were in very good and good conditions, as compared to average and poor conditions.
13
1.5
1.4
95% CI Body Condition Index
1.3
1.2
1.1
1.0
0.9
>80 cm
60-80 cm
0.05). Similar result was also obtained from pairwise Fst tests which showed no significant
different (P>0.05) on the genetic structure among the surveyed resident populations of the Pulau Panjang, Pulau
Derawan, and Payung-Payung foraging habitats.
Figure-13: Proportion (%) of haplotypes identified in the foraging habitat of Pulau Panjang, Berau of East
Kalimantan – Indonesia (N=60).
18
Figure-14: Proportion (%) of haplotypes identified in the foraging habitat of Pulau Derawan, Berau of East
Kalimantan – Indonesia (N=91).
Figure-5: Proportion (%) of haplotypes identified in the foraging habitat of Payung-Payung, Berau of East
Kalimantan – Indonesia (N=62).
Possible Contributing Stocks for the foraging areas of Berau waters
In the foraging ground of Pulau Panjang, apart from the presence of C3 which is not particularly diagnostic given
its wide distribution in the Australasian green turtle rookeries, the other haplotypes are indicative. The
combination of haplotypes D2, C5, and C14 in high frequencies suggested the importance of rookeries in the
Sulu-Sulawesi Seas (e.g. Malaysia – Philippines Turtle Islands, Berau Isles, and probably Sipadan Island) as the
main contributors. Another source of C14, perhaps, Anu Island (Aru rookery). Finding of A3 in moderate
frequency indicated representation from the green turtle management units (MU) of Long Island (PNG), Raja
19
Ampat (Papua), Micronesia, and perhaps Ashmore Reef. C4 is a dominant haplotype for Sarawak breeding
population and found in low frequency in the nesing colonies of Redang Island (Peninsular Malaysia) and the
Northern Great Barrier Reef (Australia). Nevertheless, the absence of haplotype B1, excluded the possibility of
Northern Great Barrier Reef MU as a source of nester. The low frequencies of the haplotypes of C7, A6, and
New3/P1 were only identified in this foraging ground but absent from the feeding habitats of Pulau Derawan and
Payung-Payung. While C7 confirmed the representation of Long Island rookery, A6 and New3/P1 are diagnostic
for the nesting colony of Sangalaki Island (East Kalimantan, Indonesia) and Paka (Peninsular malaysia),
respectively.
Similarly with the population of Pulau Panjang foraging ground, the most frequent variant in Pulau Derawan
feeding population was D2 (41.8%), folowed by C3 (22.0%), C5 (13.2%), and C14 (7.7%), implying that the SuluSulawesi Seas rookeries and Aru nesting colony are a major and strongly represented in this population.
Presence of A3 (5.5%), A1 (2.2%) and E2 (2.2%) indicated contribution from Micronesian, PNG, Raja Ampat,
and Ashmore Reef breeding aggregates. Similarly to Pulau Panjang assemblage, presence of C4 indicated the
contribution from Sarawak and Redang nesting populations. Finding of B4 in low frequency in this feeding site is
intriguing, and possibly represented the nesters from Khram Island (Gulf of Thailand) and/or Lacepede Island of
North West Shelf (Australia). A single haplotype of New1 is found to be similar to Pi41 which is diagnostic for Raja
Ampat nesting population (Velez-Zuazo et al, 2008). The other two new variants, New2 and New, were orphan
haplotypes in which the contributors could not be identified at the present study.
Like the other foraging habitats (Pulau Panjang and Pulau Derawan), the resident population in this water was
dominated by the haplotype of D2 (40.3%), A3 (21.0%), C3 (16.1%), C5 (8.1%), C14 (6.5%), and A1 (3.2%). The
variants, i.e. A4, C4, and New4 were found in very low frequencies (1.6% each). Apart from the orphan haplotype
of New4, the others represented green turtle nesting assemblages from the Sulu-Sulawesi Seas Rookeries as the
primary contributors, as well as, breeding colonies from Micronesia, PNG, Raja Ampat – Papua, Ashmore Reef
(Australia), Aru, Sarawak and/or Redang Island of Peninsular Mlaysia.
Overall, the mixed stock analysis (MSA) results confirmed that the feeding populations of Berau water were
mainly composed of the Turtle Islands Heritage Protected Area (TIHPA) turtles that belongs to Malaysia and
Philippines (45.49%) and the nesting populations of Berau rookery (26.82%). The Micronesia and Papua New
Guinea nesting populations were represented at 9.3% and 8.44%. Other rookeries, such as Sarawak, Aru, and
Pangumbahan of West Java were represented at 1.45%, 2.12%, and 5.41%, respectively (Table-7). The high
proportions of turtles from TIHPA and Berau MUs among the foraging turtles is expected given the proximity of
these areas to the examined foraging grounds.
20
Table-7: Proportions of green turtle nesting populations from various reookeries within the Australasian region
which identified in the feeding grounds of Berau waters are presented in Table-7.
Percentage Contribution (%)
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
Contributing Stocks
(Management Units)
Northern Great Barrier Reefs
Coral Sea
Southern Great Barrier Reefs
New Caledonia
Micronesia
Papua New Guinea
Gulf of Carpentaria
Aru
Berau Islands
South-East Sabah
Sulu Sea
Sarawak
Peninsular Malaysia
Ashmore Reef
Scott Reef
West Java
North West Shelf
Pulau Derawan
Payung-Payung
Pulau Panjang
Overall (Berau)
0
0
0
0
9,88
0
0
3,09
28,5
0,94
47,15
0
0
0
0
9,33
1,11
0
0
0
0
11,52
15,67
0
3,34
18,6
2,84
44,93
1,65
0
0,01
0
1,44
0
0
0
0
0
6,57
12,93
0
0,12
29,6
3,88
41,62
3,65
0
0
0
1,63
0
0
0
0
0
9,3
8,44
0
2,12
26,82
0,6
45,49
1,45
0
0
0
5,41
0,37
21
CONCLUSION AND MANAGEMENT IMPLICATION
This study confirmed that the foraging grounds of Pulau Panjang, Pulau Derawan, and Payung-Payung are
genetically the most diverse sample analysed so far within the Australasian region, indicating the presence of
several green turtle stocks at high and low frequencies. Most turtles represented rookeries from the Malaysian
and Philippines Turtle Islands (45.49%), Berau nesting islands (26.82%), Micronesia (9.3%), and The birdhead of
Papua/Papua New Guinea (8.44%). Small proportion of green turtles were also representing nesting sites in the
South China Sea regions, and Aru. This finding emphasizes the need to build a network of sea turtle management
across SSME - BSSE - and Micronesian regions.
The population structure of green turtles resided in these three foraging sites were similar. Nevertheless, tagrecaptured study indicated that there was no individual exchange among the foraging sites (Christianen in prep).
This implied that each feeding habitat should be managed separately.
The body condition index (BCI) of green turtles from all three feeding sites were relatively 'very good'. however,
when comparation among the three sites was made, the BCI value of green turtles captured in the water of Pulau
Panjang and Pulau Derawan were significantly lower than their counterpart which were captured in PayungPayung. This finding, perhaps, associated with the relative distance of the foraging habitat to the river come from
the mainland of Kalimantan. Payung-Payung is far away from the mainland as compared to the other two, and
possibly the sedimentation substrate which influences the fertility of the sea grass are present in lower
concentration in this water. Managing the cleanliness of the river will help in maintaining the fertility of the
seagrass beds in Pulau Panjang and Pulau Derawan.
ACKNOWLEDGEMENT
This project was strongly supported by many people. We would like to express our sincere thanks to Rusli Andar
and his team, Hidayatun Nisa Purwanasari, and Made Jayaratha for their invaluable help during the field and lab
works. Special thanks is also for Dita Cahyani for her support in analysing the data.
22
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scale of genetic exchange. Molecular Ecology, 15:3931-3946.Excoffier et al 1992
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among DNA haplotypes: application to human mitochondrial DNA restriction data. Genetics 131: 479-491.
Excoffier L, G Laval, and S Schneider (2006). Arlequin ver 3.01 User Manual: An Integrated Software Package for
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Zoology, University of Berne, Baltzerstrasse 6, 3012 Bern, Switzerland. 143 pp.
Pella, J.J. and M. Masuda. 2001. Bayesian Methods for Analysis of Stock Mixtures from Genetic Characters. Fish
Bull 99:151-67
Raymond, M. and F.Rousset 1995 An exact test for population differentiation. Evolution, 49, 1280-1283.
Rozas, J., JC Sánchez-DelBarrio, X Messeguer, and R Rozas (2003). DnaSP, DNA polymorphism analyses by
the coalescent and other methods. Bioinformatics 19: 2496-2497Slatkin 1991
Slatkin, M. 1991. Inbreeding coefficients and coalescent times. Gent. Res. Camb., 58, 167-175.
Thompson, J.D., T.J.Gibson, F.Plewniak, F.Jeanmougin and D.G.Higgins 1997. The Clustal X windows interface:
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Velez-Zuazo X, IGNK Mahardika, F Ballamu, G Gearheart, AS Kendran, CS Nindia, A Suartini, D Cahyani (2006).
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Ampat, Indonesia. A Final Report to Conservation International Indonesia. The Faculty of Veterinary Medicine
Udayana University, Denpasar, Bali, 29 pp
23
ANNEX-1:
Sequences from identified from green turtles captured in three different foraging grounds in
Berau waters during December 2009.
>W10198
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAGTAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10200
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAGTAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10201
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATTCACATAAATATTTTAATAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10202
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAGTTTGCATAAACATTTTAATAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATTGTCACAGTAATTGGTTATTTCT
TAAATAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAAGCCCATTCAATTTGTGGCGTACATAA
TTTGATCTATTCTGGCCTCTGGTTGTTCTTTCAGGCACATATAAATAACGACGTTCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAGATTTATAACCT
*
>W10203
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATTCACATAAATATTTTAATAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10204
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAGTAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10205
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAGTAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10206
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAGTAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
24
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10207
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAGTAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10208
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAGTAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10209
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTAAATTTGCATAAATGTTTTAATAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATTGTTACGGTAATTGGTTATTTCT
TAAATAACTATTCACGAGAAATAAGCAACCCTTGTTGGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
TTTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTTCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10211
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAGTTTACATAAACATTTTAATAACATGAATATTAAGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATTG
TCACAGTAATTGGTTATTTCTTAAATAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAAGCCCA
TTCAATTTGTGACGTACATAATTTGATCTATTCTGGCCTCTGGTTGTTCTTTCAGGCACATATAAATAACGACGTTCATTCGTTCCTCT
TTAAAAGGCCTTTGGTTGAATGAGTTCTATACATTAGATTTATAACCT
*
>W10212
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAATAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAAGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10213
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATTCACATAAATATTTTAATAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10214
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAATAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10215
25
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAATAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAAGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10216
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAATAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTCATTTCT
TAAATAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10217
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATTCACATAAATATTTTAATAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10218
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAATAACATGAATATTAAGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCG
TCACAGTAATTGGTTATTTCTTAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCA
TTTAGTTTATAGCGTACATAACCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCT
TTAAAAGGCCTTTGGTTGAATGAGTTCTATACATTAAATTTATAACCT
*
>W10219
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAATAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10220
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAGTAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10221
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAATAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAAGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10222
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAGTAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
G
POPULATIONS IN INDONESIA AT DIFFERENT SCALES :
A CASE STUDY FROM BERAU GREEN TURTLE ROOKERY, EAST-KALIMANTAN
Windia Adnyana
The Faculty of Veterinary Medicine, Udayana University, Bali - Indonesia
SUMMARY
The purpose of this study is to assess the body condition index of green turtles at three different seagrass
meadows of the Berau Isles, assess the genetic composition of green turtles, identify presence or absence of
genetic differentiation among the green turtles reside in the three foraging areas, and to identify links among
these foraging areas and various nesting sites in the Australasian region.
Sampling were carried out between 8 to 23 December 2009 from three main foraging locations within the Berau
Isles. The selected foraging habitats were the water in the northern side of Pulau Derawan, near Pulau Panjang,
and the water of Payung-Payung near Maratua Island. Turtles were captured either by net or rodeo technique,
taken onboard, their biometric values were measured and the skin samples were taken for genetic analysis.
A total of 310 Chelonia mydas were captured and observed. Most of them (50.32%) are medium size turtles with
Curved Carapace Length between 60-80 cm. The proportions of turtle with CCL > 80 cm and with CCL < 60 cm
were 27.10% and 22.58%, respectively. Their bodyweight ranged from 7.70 kg to 158.90 kg. In view of their sex
status which was determined based on the Total Tail Length, 17.42% were defined to be males and 10.32% were
females. Most turtles (72.26%) were sexually undifferentiated by means of external characteristics. Thir Body
Condition Index (BCI) varied between 0.71 – 1.80. Most turtles were in very good (70.0%) and good (11.29%)
conditions, as compared to average (8.06%) and poor conditions (10.65%). Based on their capture locations, the
highest BCI was calculated for Payung-Payung population (1.30±0.21; range=0.80–1.66; n=102), followed by
Pulau Derawan (1.26±0.16; range=0.84–1.56; n=117) and Pulau Panjang population (1.24±0.20; range=0.711.80; n=91). Statistical analysis showed that there was no significant difference between the population of Pulau
Panjang and Pulau Derawan (P=0.443). Significant difference (P=0.017) was found between the population of
Pulau Panjang and Payung-Payung., but not-significant difference was calculated between Payung-Payung and
Pulau Derawan (P=0.078).
A total of 213 mitochondrial (mt) DNA fragments, out of 309 collected samples were amplified by PCR technique.
Screening of polymorphism within 384 bp mtDNA control region fragments identified 34 polymorphic sites and 17
distinct haplotypes. The most frequent haplotype identified in Berau waters was D2 (40.8%), followed by C3
1
(19.7%), A3 (12.2%), C5 (10.8%), and C14 (6.6%). These five variants accounted for 90.1% of the total resident
foraging population. The other haplotypes pesent in relatively small proportion, ranged from 0.5% to 1.9%.
Despite of minor variations, there was no significant difference (P>0.05) found on the genetic structure of resident
populations in the Pulau Panjang, Pulau Derawan, and Payung-Payung foraging habitats. Mixed stock analysis
(MSA) results revealed that the feeding populations of Berau water were mainly composed of green turtles from
the Turtle Islands Heritage Protected Area (TIHPA) (45.49%), the nesting populations of Berau rookery (26.82%),
Micronesia (9.3%) and Papua New Guinea (8.44%) nesting populations. Small proportion of representatives from
the nesting sites in the South China Sea regions and Aru were also found. The finding emphasizes the need to
build a network of turtle based-MPA across SSME - BSSE - and Micronesian regions.
The population structure of green turtles resided in these three foraging sites were similar, but individual
exchange among the foraging sites is unlikely, which implied that each feeding habitat should be managed
separately. The body condition index of green turtles from Pulau Panjang and Pulau Derawan were significantly
lower than their counterpart which were captured in Payung-Payung. This finding, perhaps, associated with the
relative distance of the foraging habitat to the river come from the mainland of Kalimantan. Payung-Payung is
relatively far away from the mainland as compared to the other two, and possibly the sedimentation substrate
which influences the fertility of the sea grass are present in lower concentration in this water. Managing the
cleanliness of the river will help in maintaining the fertility of the seagrass beds in Pulau Panjang and Pulau
Derawan.
2
INTRODUCTION
The Berau Islands complex is an important nesting and foraging area for the green sea turtle (Chelonia mydas) in
Indonesia. This area is located within coordinates 02049’42.6’’ - 0102’0.06’’N; 117059’17.16’’ - 11902’50.30’’S and
belongs to the Berau district in East-Kalimantan (Figure-1). The area encompasses over 1.2 millions ha of
coastal area across 31 small islands, nine of which represent important nesting areas for green turtles. Annual
census surveys since 2002, suggest that the Berau Islands have one of the highest density of nesting green sea
turtles in the South-East Asia region (Adnyana et al, 2007). Additionally to nesting beaches, the Berau Isles also
contains several large seagrass meadows, which provide a critical foraging habitat for green turtles. Recent
surveys, conducted by researchers from the Radboud University in Nijmegen in collaboration with WWF
Indonesia and Udayana University, revealed that the density of green turtles on the seagrass meadows in this
area is among the highest in the world (Christianen in prep). The average green turtle density on the foraging
grounds is 17 ± 1.5 individuals per hectare.
The spatial extend of some of the seagrass meadows in Berau region is declining. The cause of this decline is
believed to be related to increased nutrient and sediment loads as a result of upstream erosion in the Berau river.
A reduced area of available nutrients could potentially lead to a reduced fitness of a green turtle population. The
resilience of a population under stress of e.g. seagrass loss or harvesting depends on the propensity of individual
turtles to switch foraging grounds and on the onset of the switching. Mark-recapture studies of foraging green
turtles from all size classes (Christianen in prep) suggest that subadult green turtles are not switching between
foraging grounds on a small spatial scale (i.e.within the Berau Isles) and thus, individuals associated with the
declining seagrass meadows are at risk of undernourishment. The body condition index (BCI) we be measured to
infer food availability and quality at the different foraging areas within the Berau water.
Mark-recapture studies of adult green turtles have revealed links between the nesting and the foraging
populations within the Sulu-Sulawesi marine Ecoregion. For example, turtles that received a tag while nesting on
the beach of Palau, the Philippines, and Malaysia (Sarawak, and Sabah) were later recaptured while foraging at
Derawan, Panjang or Maratua waters (WWF pers. Comm.). In addition, some individuals were tracked using
satellite telemetry (for a maximum of 155 days) and found to migrate from the Derawan and Sangalaki Islands to
Sabah and the Philippines (Adnyana et al. 2007) after nesting. While these observations provide valuable insight
into individual movement patterns, the relatedness of the Derawan green turtle population to populations in other
parts of the Australasian region, as well as an understanding of genetic differentiation among feeding grounds
within the archipelago remains unresolved. An assessment of the genetic composition of the Berau foraging
population examines the relative contribution of green turtle breeding populations to this assemblage, thus
providing a good understanding of the migratory pathways of green turtles to and from the Berau Archipelago.
3
This study is part of a PhD project on “sea grasses under green turtle grazing and nutrient loads”, conducted by
Marjolijn Christianen, Radboud University Nijmegen. This project primarily builds on previous genetic studies of
Australasian green turtle nesting populations, identifying 17 genetically distinct breeding stocks (Dethmers et al.
2006). This work is critical as the success of green turtle management strategies is contingent on understanding
of their population dynamics.
Objectives of this study
1. Assess the body condition index of green turtles at three different seagrass meadows of the Berau Isles
2. Assess the genetic composition of green turtles at the three main seagrass meadows of the Berau Isles
3. Identify presence or absence of genetic differentiation among the three foraging areas.
4. Identify links among these foraging areas and the Australasian 17 genetic stocks
4
Figure-1: A map showing the locations of three different foraging habitats within the Berau Archipelago. The
approximation of geographic location of this Isles is 02049’42.6’’ - 0102’0.06’’N; 117059’17.16’’ - 11902’ 50.30’’S.
Above is overall picture of the Berau water; below is the exact locations of sampling in Pulau Panjang (left), Pulau
Derawan (middle), and Payung-Payung, Maratua (right).
5
MATERIALS AND METHODS
Sample collection, measurements, maturity and sexual determination
Sampling were done during 15 working days, from 8 to 23 December 2009 by a team led by Marjolijn Christianen,
a PhD candidate from Radboud University Nijmegen. All samples were taken from three main foraging locations
within the Berau Islands complex, i.e. the water in the northern side of Pulau Derawan, near Pulau Panjang, and
the water of Payung-Payung near Maratua Island (see Figure-1). Turtles were captured either by net or rodeo
technique (jumping straightly to the turtles), taken onboard, and mesured for their Curved Carapace Length
(CCL), Total Tail Length (TTL), and their body-weight (BW). All of these measurement were carried out following
procedures provided by Bolten (1999) and the protocol provided by Adnyana and Hitipeuw (2009).
In view of the maturity status, based on their CCL, turtles with a CCL 80 cm were defined as mature. This criteria is decided based on our field experiences while
observing nesting green turtles on Derawan and Sangalaki Islands in which their minimum CCL was 80 cm. The
sex was predicted by looking at the TTL. Turtles of all size classes with TTL >20 cm were defined as males, while
those with TTL up to 20 cm and the CCL measuring more than 80 cm were defined as females. Immature turtles
(CCL up to 80 cm) with TTL less than 20 cm were considered as unsexed or sexually undetermined.
The Body condition index (BCI), a variable to indicate a decline in individual and population health which needed
to inform inshore management strategies, calculated using Equation derived from Bjorndal et al. (2000). BCI
3
quantifies body condition based on a ratio of weight and SCL. The equation is BCI = ([Weight (kg) / SCL(cm) ] x
10000). The straight carapace length (SCL) did not measured during this study. It was estimated from the values
of the CCL by using an equation SCL = 15 + (0.76 X CCL). This equation was made based on field records
obtained from Derawan nesting island monitoring workers. Total data used for linear regression calculation was
285, and the coeficient correlation (R) and R 2 were 0.771 and 0.594, respectively.
Any observed external morphological lesions on the turtle skin, carapace and plastron such as presence of scars,
wound, notch on marginal scutes, algae, barnacles, and fibropapillomatosis were recorded. Presence of metal
tags in the pliffers were noted, and turtles without tag were given a new one following the protocol provided by
Balazs (1999). Prior to be released back to the water, from each captured turtles, skin samples were taken from
flippers or neck region by using a biopsy punch, and immediately stored in either alcohol 70% or a NaCl saturated
solution of 20% DMSO, and transported to Udayana University for subsequent processes.
6
Molecular Methods
Genomic DNA was isolated from 0.1 g of skin tissue using Qiamp™ DNA Mini Kit from Qiagen®, and stored at 20oC for subsequent polymerase chain reaction (PCR). Successful DNA isolation was confirmed by running 2 μL
of genomic DNA in an ethidium bromide added 1% Agarose gel. A 740 bp segments of the mtDNA control region
for all samples were amplified using LTEi9 (5'-AGCGAATAATCAAAAGAGAAGG-3') and H950 (5'GTCTCGGATTTAGGGGTTTG-3') primers (Abreu-Grobois et al, 2006). The first primer binds at the border
between the tRNA-Thr and tRNA-Pro loci, and the second at position 782 near the end of the d-loop, expected to
produce PCR products of about 860 base pairs (bp). The target mitochondrial sequences were amplified by PCR
using approximately 50 ng of template genomic DNA in 25 µl reaction volume containing 14.3 µl H2O, 2.5 µl DNA
genome, 1.5 units ampli Taq gold polimerase (applied biosystem), 1.5 µl PCR buffer (applied biosystem), 2.5 µl
MgCl2 25 Mm, 2 µl dNTP 1 Mm, 1 µl of each primer 10 Mm. The PCR profile comprised an initial denaturation of
5 min at 94°C (to activate the Ampli Taq gold polymerase), followed by 40 cycles of: 94o C for 45 sec
(denaturation), 55o C for 45 sec (annealing), 72o C for 45 sec (extension), ended in a final extension in 72 o C for 4
min. Electrophoresis was performed to confirm the result of amplification and to determine the length of PCR
product. One µl loading dye (bromophenol-blue and cyline cyanol) was added in 2 µl PCR product.
Electrophoresis was run for negative and positive control (marker) for 30 minutes in 50 Volt of 1% agarose gel
media with etidum bromide dye. PCR product was sent to Macrogen Inc. (Korea) for forward and reverse
sequencing.
Statistical Analysis
Biometrics data were analysed using the Statistical Package for Social Sciences (SPSS) version 13.0. Graphical
presentations were completed either by Microsoft EXCEL version 2010 or SPSS version 13.0. Sequences were
aligned using Clustal X (Thompson et al 1997) and the population parameters such as polimorphic sites, the
percentages of each haplotype, haplotype diversity, and nucleotide diversity were analyzed using DNAsp 4.10
(Rozas et al., 2003). Phylogenetic tree was constructed to visualize the relationship among the observed mt-DNA
variants. AMOVA (Excoffier et al 1992), Exact tests of population differentiation (Raymond & Rousset 1995) and
pairwise Fst tests (Slatkin 1991) implemented in the population genetics package Arlequin version 3.01 (Excoffier
et al, 2006) were used to examined genetic structure among surveyed populations. Mixed stock analysis, to
predict the individual contribution from nesting colonies or management unit to the population of green turtles
reside in the foraging grounds of Pulau Panjang, Pulau Derawan, and Payung-Payung were calculated BAYES
program (Pella and Masuda, 2001).
7
RESULTS
Biometrics, maturity and sex status of Green Turtles in Berau Foraging Grounds
There was a total of 310 individual Chelonia mydas captured during 15 days sampling period during 8 December
– 23 December 2009. Their Curved Carapaca Length (CCL) encompassed all size classes (Figure-2) from small
immatures (CCL=40.80 cm) to large adults (CCL=111.30 cm). The mean ± standard deviation of the CCL was
71.69 ± 15.30 cm. When the CCL values were pooled into three categories (Figure-3), the majority of samples
were found to be medium size with CCL between 60-80 cm (50.32%). The proportions of large-adult (CCL > 80
cm) and small immature turtles with CCL < 60 cm were 27.10% and 22.58%, respectively.
Based on their actual capture sites, the percentage of mature-adult turtles (CCL>80 cm) was proportionally
highest in the Payung-Payung foraging ground as compared to the population of Pulau Panjang and Pulau
Derawan (Table-1). Consequently, as shown in Figure-4, the highest mean value for CCL was also found in the
foraging population of Payung-Payung (75.35±16.90 cm; range = 43.50–111.30 cm; n = 102), followed by Pulau
Panjang (70.63±15.02; range = 40.80–102.40 cm; n = 91) and the foraging population of Pulau Derawan
(69.32±13.46; range = 46.80–104.50 cm; n = 91). Statistically, a one way anova test suggested that significant
differences were calculated between the mean value of CCL of Payung-Payung population and the other two
populations, i.e. Pulau Panjang (P=0.031) and Pulau Derawan (P=0.003). However, no significant difference was
calculated between Pulau Panjang and Pulau Derawan (P=0.535).
Table-1: Size classes indicated by the Curved Carapace Length (CCL) of the Chelonia mydas captured in three
foraging grounds of the Berau Isles.
Capture Locations
Pulau Panjang
Pulau Derawan
Payung-Payung
Combined
>80 cm
21 (23.08%)
24 (20.51%)
39 (38.24%)
84 (27.10%)
Size Class
60-80 cm
51 (56.04%)
64 (54.70%)
41 (40.20%)
156 (50.32%)
20 cm. The summary values for maturity and sex
status per capture locations were presented in Table-2.
40
Frequency
30
20
10
Mean = 71.7177
Std. Dev. = 15.31384
N = 310
0
40.00
60.00
80.00
100.00
120.00
CCL
Figure-2: Histogram showing frequency distribution of the curved carapace length (CCL) of 310 green sea turtles
captured, tagged and measured in three foraging grounds of Berau water. Normal curve is displayed.
9
60.0%
50.0%
Percent
40.0%
30.0%
50.32%
20.0%
27.1%
22.58%
10.0%
0.0%
>80 cm
60-80 cm
80 cm)
Immature (CCL < 80 cm)
Male
Female
Male
Female Undetermined
12 (13.19%) 9 (9.89%)
- 70 (76.92%)
16 (13.68%) 8 (6.84%)
- 93 (79.49%)
24 (23.53%) 15 (14.71%) 2 (1.96%)
- 61 (59.80%)
2
(0.65%)
52 (16.77%) 32 (10.32%)
224 (72.26%)
Total
91 (100%)
117 (100%)
102 (100%)
310 (100%)
The Body Condition Index (BCI) of all samples varied between 0.71 – 1.80, normally distributed (Figure-7) with
the smallest and the highest values were both found in turtles captured from Pulau Panjang. Based on the criteria
defined by Bjorndal et al (2000) (Table-3), it was found that (Figure-8) most turtles were in very good (70.0%)
and good (11.29%) conditions, as compared to average (8.06%) and poor conditions (10.65%).
Regarding their size classes, as shown in Figure-9, the highest mean value for BCI (1.44±0.12; range = 1.141.66; n = 84) was noted in mature-adult turtles measuring > 80 cm. The mean values for size classes between 6080 cm and 1.20
Very Good
1
1.11 – 1.20
Good
2
1.00 – 1.10
Average
3
< 1.00
Poor
Determined from data presented by Bjorndal, K. A., Bolten, A. B. and Chaloupka, M. Y. (2000). Green turtle
somatic growth model: evidence for density dependence’. Ecological Applications 10, 269-282.
70.0%
60.0%
50.0%
Percent
40.0%
70.0%
30.0%
20.0%
10.0%
11.29%
10.65%
8.06%
0.0%
Very good
Good
Average
Poor
Body Condition Index
Figure-8: The Body Condition Index (BCI) of all samples captured from three foraging sites in the Berau water.
Note that most turtles were in very good and good conditions, as compared to average and poor conditions.
13
1.5
1.4
95% CI Body Condition Index
1.3
1.2
1.1
1.0
0.9
>80 cm
60-80 cm
0.05). Similar result was also obtained from pairwise Fst tests which showed no significant
different (P>0.05) on the genetic structure among the surveyed resident populations of the Pulau Panjang, Pulau
Derawan, and Payung-Payung foraging habitats.
Figure-13: Proportion (%) of haplotypes identified in the foraging habitat of Pulau Panjang, Berau of East
Kalimantan – Indonesia (N=60).
18
Figure-14: Proportion (%) of haplotypes identified in the foraging habitat of Pulau Derawan, Berau of East
Kalimantan – Indonesia (N=91).
Figure-5: Proportion (%) of haplotypes identified in the foraging habitat of Payung-Payung, Berau of East
Kalimantan – Indonesia (N=62).
Possible Contributing Stocks for the foraging areas of Berau waters
In the foraging ground of Pulau Panjang, apart from the presence of C3 which is not particularly diagnostic given
its wide distribution in the Australasian green turtle rookeries, the other haplotypes are indicative. The
combination of haplotypes D2, C5, and C14 in high frequencies suggested the importance of rookeries in the
Sulu-Sulawesi Seas (e.g. Malaysia – Philippines Turtle Islands, Berau Isles, and probably Sipadan Island) as the
main contributors. Another source of C14, perhaps, Anu Island (Aru rookery). Finding of A3 in moderate
frequency indicated representation from the green turtle management units (MU) of Long Island (PNG), Raja
19
Ampat (Papua), Micronesia, and perhaps Ashmore Reef. C4 is a dominant haplotype for Sarawak breeding
population and found in low frequency in the nesing colonies of Redang Island (Peninsular Malaysia) and the
Northern Great Barrier Reef (Australia). Nevertheless, the absence of haplotype B1, excluded the possibility of
Northern Great Barrier Reef MU as a source of nester. The low frequencies of the haplotypes of C7, A6, and
New3/P1 were only identified in this foraging ground but absent from the feeding habitats of Pulau Derawan and
Payung-Payung. While C7 confirmed the representation of Long Island rookery, A6 and New3/P1 are diagnostic
for the nesting colony of Sangalaki Island (East Kalimantan, Indonesia) and Paka (Peninsular malaysia),
respectively.
Similarly with the population of Pulau Panjang foraging ground, the most frequent variant in Pulau Derawan
feeding population was D2 (41.8%), folowed by C3 (22.0%), C5 (13.2%), and C14 (7.7%), implying that the SuluSulawesi Seas rookeries and Aru nesting colony are a major and strongly represented in this population.
Presence of A3 (5.5%), A1 (2.2%) and E2 (2.2%) indicated contribution from Micronesian, PNG, Raja Ampat,
and Ashmore Reef breeding aggregates. Similarly to Pulau Panjang assemblage, presence of C4 indicated the
contribution from Sarawak and Redang nesting populations. Finding of B4 in low frequency in this feeding site is
intriguing, and possibly represented the nesters from Khram Island (Gulf of Thailand) and/or Lacepede Island of
North West Shelf (Australia). A single haplotype of New1 is found to be similar to Pi41 which is diagnostic for Raja
Ampat nesting population (Velez-Zuazo et al, 2008). The other two new variants, New2 and New, were orphan
haplotypes in which the contributors could not be identified at the present study.
Like the other foraging habitats (Pulau Panjang and Pulau Derawan), the resident population in this water was
dominated by the haplotype of D2 (40.3%), A3 (21.0%), C3 (16.1%), C5 (8.1%), C14 (6.5%), and A1 (3.2%). The
variants, i.e. A4, C4, and New4 were found in very low frequencies (1.6% each). Apart from the orphan haplotype
of New4, the others represented green turtle nesting assemblages from the Sulu-Sulawesi Seas Rookeries as the
primary contributors, as well as, breeding colonies from Micronesia, PNG, Raja Ampat – Papua, Ashmore Reef
(Australia), Aru, Sarawak and/or Redang Island of Peninsular Mlaysia.
Overall, the mixed stock analysis (MSA) results confirmed that the feeding populations of Berau water were
mainly composed of the Turtle Islands Heritage Protected Area (TIHPA) turtles that belongs to Malaysia and
Philippines (45.49%) and the nesting populations of Berau rookery (26.82%). The Micronesia and Papua New
Guinea nesting populations were represented at 9.3% and 8.44%. Other rookeries, such as Sarawak, Aru, and
Pangumbahan of West Java were represented at 1.45%, 2.12%, and 5.41%, respectively (Table-7). The high
proportions of turtles from TIHPA and Berau MUs among the foraging turtles is expected given the proximity of
these areas to the examined foraging grounds.
20
Table-7: Proportions of green turtle nesting populations from various reookeries within the Australasian region
which identified in the feeding grounds of Berau waters are presented in Table-7.
Percentage Contribution (%)
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
Contributing Stocks
(Management Units)
Northern Great Barrier Reefs
Coral Sea
Southern Great Barrier Reefs
New Caledonia
Micronesia
Papua New Guinea
Gulf of Carpentaria
Aru
Berau Islands
South-East Sabah
Sulu Sea
Sarawak
Peninsular Malaysia
Ashmore Reef
Scott Reef
West Java
North West Shelf
Pulau Derawan
Payung-Payung
Pulau Panjang
Overall (Berau)
0
0
0
0
9,88
0
0
3,09
28,5
0,94
47,15
0
0
0
0
9,33
1,11
0
0
0
0
11,52
15,67
0
3,34
18,6
2,84
44,93
1,65
0
0,01
0
1,44
0
0
0
0
0
6,57
12,93
0
0,12
29,6
3,88
41,62
3,65
0
0
0
1,63
0
0
0
0
0
9,3
8,44
0
2,12
26,82
0,6
45,49
1,45
0
0
0
5,41
0,37
21
CONCLUSION AND MANAGEMENT IMPLICATION
This study confirmed that the foraging grounds of Pulau Panjang, Pulau Derawan, and Payung-Payung are
genetically the most diverse sample analysed so far within the Australasian region, indicating the presence of
several green turtle stocks at high and low frequencies. Most turtles represented rookeries from the Malaysian
and Philippines Turtle Islands (45.49%), Berau nesting islands (26.82%), Micronesia (9.3%), and The birdhead of
Papua/Papua New Guinea (8.44%). Small proportion of green turtles were also representing nesting sites in the
South China Sea regions, and Aru. This finding emphasizes the need to build a network of sea turtle management
across SSME - BSSE - and Micronesian regions.
The population structure of green turtles resided in these three foraging sites were similar. Nevertheless, tagrecaptured study indicated that there was no individual exchange among the foraging sites (Christianen in prep).
This implied that each feeding habitat should be managed separately.
The body condition index (BCI) of green turtles from all three feeding sites were relatively 'very good'. however,
when comparation among the three sites was made, the BCI value of green turtles captured in the water of Pulau
Panjang and Pulau Derawan were significantly lower than their counterpart which were captured in PayungPayung. This finding, perhaps, associated with the relative distance of the foraging habitat to the river come from
the mainland of Kalimantan. Payung-Payung is far away from the mainland as compared to the other two, and
possibly the sedimentation substrate which influences the fertility of the sea grass are present in lower
concentration in this water. Managing the cleanliness of the river will help in maintaining the fertility of the
seagrass beds in Pulau Panjang and Pulau Derawan.
ACKNOWLEDGEMENT
This project was strongly supported by many people. We would like to express our sincere thanks to Rusli Andar
and his team, Hidayatun Nisa Purwanasari, and Made Jayaratha for their invaluable help during the field and lab
works. Special thanks is also for Dita Cahyani for her support in analysing the data.
22
REFERENCES
Abreu-Grobois FA, Horrocks JA, Formia A et al (2006) New mtDNA dloop primers which work for a variety of
marine turtle species may increase the resolution capacity of mixed stock analyses. Poster presented at the 26th
Annual Symposium on Sea Turtle Biology and Conservation, Crete, Greece, 2-8 April 2006. Available from:
http://www.iucnmtsg.org/genetics/meth/primers/abreu_grobois_etal_new_dloop_primers.pdf
Adnyana W, LP Soede, G Gearheart and M Halim (2007). Status of green turtle (Chelonia mydas) nesting and
foraging populations of Berau, East Kalimantan, Indonesia, including results from tagging and telemetry. IOTN
(7), 1-11.
Adnyana W and C Hitipeuw (2009). Panduan Melakukan Pemantauan Populasi Penyu di Pantai Peneluran di
Indonesia. Edisi Mei, WWF Indonesia Marine Program, 31 pp.
Bolten AB (1999). Techniques for Measuring Sea Turtles. In Research and Management Techniques for the
Conservation of Sea Turtles, Eds. KL Eckert, KA Bjorndal, FA Abreu-Grobois, M Donnelly. IUCN/SSC Marine
Turtle Specialists Group Publication No. 4, p 110-114.
Bjorndal, K. A., Bolten, A. B. and Chaloupka, M. Y. (2000). Green turtle somatic growth model: evidence for
density dependence’. Ecological Applications 10, 269-282.
Dethmers K, D Broderick, C Moritz, NN FitzSimmons, CJ Limpus, S Lavery, S Whiting, M Guinea, RIT Prince, R
Kennett (2006). The genetic structure of Australasian green turtles (Chelonia mydas): exploring the geographical
scale of genetic exchange. Molecular Ecology, 15:3931-3946.Excoffier et al 1992
Excoffier, L., P.E.Smouse and J.M.Quattro 1992. Analysis of molecular variance inferred from metric distances
among DNA haplotypes: application to human mitochondrial DNA restriction data. Genetics 131: 479-491.
Excoffier L, G Laval, and S Schneider (2006). Arlequin ver 3.01 User Manual: An Integrated Software Package for
Population Genetics Data Analysis. Computational and Molecular Population Genetics Lab (CMPG). Institute of
Zoology, University of Berne, Baltzerstrasse 6, 3012 Bern, Switzerland. 143 pp.
Pella, J.J. and M. Masuda. 2001. Bayesian Methods for Analysis of Stock Mixtures from Genetic Characters. Fish
Bull 99:151-67
Raymond, M. and F.Rousset 1995 An exact test for population differentiation. Evolution, 49, 1280-1283.
Rozas, J., JC Sánchez-DelBarrio, X Messeguer, and R Rozas (2003). DnaSP, DNA polymorphism analyses by
the coalescent and other methods. Bioinformatics 19: 2496-2497Slatkin 1991
Slatkin, M. 1991. Inbreeding coefficients and coalescent times. Gent. Res. Camb., 58, 167-175.
Thompson, J.D., T.J.Gibson, F.Plewniak, F.Jeanmougin and D.G.Higgins 1997. The Clustal X windows interface:
flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Research,
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Velez-Zuazo X, IGNK Mahardika, F Ballamu, G Gearheart, AS Kendran, CS Nindia, A Suartini, D Cahyani (2006).
Genetic Composition of the Nesting Aggregation of Green Turtles (Chelonia mydas) in the Archipelago of Raja
Ampat, Indonesia. A Final Report to Conservation International Indonesia. The Faculty of Veterinary Medicine
Udayana University, Denpasar, Bali, 29 pp
23
ANNEX-1:
Sequences from identified from green turtles captured in three different foraging grounds in
Berau waters during December 2009.
>W10198
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAGTAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10200
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAGTAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10201
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATTCACATAAATATTTTAATAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10202
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAGTTTGCATAAACATTTTAATAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATTGTCACAGTAATTGGTTATTTCT
TAAATAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAAGCCCATTCAATTTGTGGCGTACATAA
TTTGATCTATTCTGGCCTCTGGTTGTTCTTTCAGGCACATATAAATAACGACGTTCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAGATTTATAACCT
*
>W10203
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATTCACATAAATATTTTAATAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10204
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAGTAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10205
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAGTAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10206
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAGTAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
24
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10207
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAGTAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10208
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAGTAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10209
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTAAATTTGCATAAATGTTTTAATAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATTGTTACGGTAATTGGTTATTTCT
TAAATAACTATTCACGAGAAATAAGCAACCCTTGTTGGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
TTTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTTCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10211
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAGTTTACATAAACATTTTAATAACATGAATATTAAGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATTG
TCACAGTAATTGGTTATTTCTTAAATAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAAGCCCA
TTCAATTTGTGACGTACATAATTTGATCTATTCTGGCCTCTGGTTGTTCTTTCAGGCACATATAAATAACGACGTTCATTCGTTCCTCT
TTAAAAGGCCTTTGGTTGAATGAGTTCTATACATTAGATTTATAACCT
*
>W10212
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAATAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAAGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10213
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATTCACATAAATATTTTAATAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10214
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAATAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10215
25
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAATAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAAGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10216
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAATAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTCATTTCT
TAAATAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10217
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATTCACATAAATATTTTAATAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10218
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAATAACATGAATATTAAGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCG
TCACAGTAATTGGTTATTTCTTAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCA
TTTAGTTTATAGCGTACATAACCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCT
TTAAAAGGCCTTTGGTTGAATGAGTTCTATACATTAAATTTATAACCT
*
>W10219
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAATAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10220
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAGTAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10221
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAATAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAAGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
GAGTTCTATACATTAAATTTATAACCT
*
>W10222
TAGCATATGACCAGTAATGTTAACAGTTGATTTGGCCCTAAACATGAAAATTATTGAATCCACATAAATATTTTAGTAACATGAATATTA
AGCAGAGAATTAAAAGTGAAATGATATAGGACATAAAATTAAACCATTATACTCAACCATGAATATCGTCACAGTAATTGGTTATTTCT
TAAGTAGCTATTCACGAGAAATAAGCAACCCTTGTTAGTAAGATACAACATTACCAGTTTCAGGCCCATTTAGTTTATAGCGTACATAA
CCTGATCTATTCTGGCCTCTGGTTGTCTTTTCAGGCACATACAAATAGTAACGTCCATTCGTTCCTCTTTAAAAGGCCTTTGGTTGAAT
G