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STUDY OF BIRD ECOLOGY, GENETICS AND CONSERVATION
TANIMBAR CATTLE (Cacatua goffiniana) ON TANIMBAR ISLAND,
MALUKU
ARIZONA STATE UNIVERSITY
ABS 370 – ECOLOGY
SPRING 2024
Introduction:
Indonesia is known as a megabiodiversity country due to its high biodiversity at the
ecosystem, species and genetic levels. Indonesia has 19 types of natural ecosystems with 74
types of vegetation. The diversity of marine biota species is recorded at 6396 species
(BAPPENAS 2016), 80,000 species of plant species, 8157 species of vertebrate fauna, 1900
species of butterflies and other flora and fauna (Widjaja et al. 2014). This diversity also holds
a wealth of genetic information, including a wealth of traditional knowledge. One of
Indonesia's vertebrate wealth is the diversity of bird species, currently around 1883 species
(PUSYANTEK 2022). In addition to having a high wealth of biodiversity, Indonesia also
faces the threat of species loss and extinction.
Biodiversity loss can be caused by several things, namely forest conversion, the entry
of invasive foreign species, ecosystem and habitat damage, and over-exploitation of
biological resources including flora and fauna (Widjaja et al. 2014). Local, regional and
global extinction of a species is also caused by fragmentation and loss of its natural habitat
(Whitmore and Sayer 1992). Extinction is also caused by loss of genetic variation resulting in
a decrease in the ability of individuals to respond to changes such as climate change and
disease (Whittaker 1998).
Among the biodiversity that is under threat of species loss and extinction are birds. In
2019 as many as 168 bird species were declared endangered, this figure increased from 2018
which previously was
163 species. Based on the threat status of the International Union for Conservation of Nature
(IUCN), 30 species are categorized as critical, 44 as endangered, 94 as vulnerable and 244 as
near threatened (Burung Indonesia 2019). Birds are in high demand as pets, with more than
2600 wild bird species traded internationally, dominated by Passeriformes and
Psittaciformes (FAO 2008a). Passeriformes birds are traded because of their excellence in
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singing, while Psittaciformes have beautiful feather colors and the ability to imitate sounds.
Passeriformes in Indonesia have around 814 species from 41 families, while Psittaciformes
have two families namely Cacatuidae (seven species) and Psittacidae (81 species). One of the
traded bird species from the Cacatuidae family is the tanimbar cockatoo with the scientific
name C. goffiniana (Roselaar and Michels 2004).
Conservation activities carried out by the government for endemic birds, especially C.
goffiniana, have not shown maximum results. Indicators include declining bird populations,
illegal trade continues, the fulfillment of bird hobby needs has not been met from captive
breeding, and conservation areas have not been able to protect and preserve this bird species.
To improve the optimization of conservation activities, the development of conservation
action plan policies requires comprehensive and complete scientific data related to the species
and its ecology.
This research conducted various studies on the C. goffiniana species through
ecological, genetic and conservation approaches. This study is expected to be the basis for
scientific considerations in the preparation of various policies in the evaluation of habitats
and conservation areas, law enforcement, re-release into nature of confiscated and captive-
bred birds, as well as strengthening and developing captive breeding to meet market needs.
Appropriate policies in developing conservation strategies and actions as well as proper
implementation can realize the conservation goal of preserving biological natural resources,
especially C. goffiniana.
1.1 Problem Formulation
Excessive hunting and exploitation are also threats to population decline in the wild.
The tanimbar cockatoo (C. goffiniana), one of Indonesia's seven cockatoo species, has been a
trade commodity for decades, causing population declines. To meet market demand, captive
breeders have not been able to provide enough birds, so illegal trade still takes place despite
the bird being protected. Captive breeding has not been well developed in terms of the
number of breeders and the number of broodstock bred. In 2005, 10,560 C. goffniana were
reportedly taken and traded (Burung Indonesia 2018).
C. goffiniana has a limited natural distribution in the Tanimbar Islands. The region also
faces the threat of loss of biological resources including flora and fauna due to
overexploitation and habitat destruction due to land use change. Habitat fragmentation has
resulted in small population sizes that can lead to inbreeding and decreased genetic diversity.
Conservation activities of C. goffiniana have been carried out by the government
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through protection, preservation and utilization. Government Regulation No. 7 of 1999
included C. goffiniana in the list of protected bird species, which was updated through
Permenhut LHK No. 106 of 2018. Conservation efforts can be carried out by maintaining a
balanced population in its natural habitat and breeding outside its habitat The government
issued Forest Ministerial Decree No. 149/Kpts-II/1999 to establish the Tanimbar Wildlife
Sanctuary which functions as an area for preservation and protection.
C. goffiniana in its natural habitat. Ex-situ conservation for this species involves breeding in
conservation institutions or captive breeding activities (Kepmenhut 1999).
The government also supervises and enforces the law against illegal capture and
distribution of protected birds. Reintroduction to nature is a program to increase and
strengthen the population of species to maintain their sustainability in nature. Legal efforts,
rehabilitation and release of confiscated animals are conservation actions to preserve wildlife
to maintain ecosystem stability (Prayudi 2015). In terms of captive breeding development,
especially for C. goffiniana, scientific supporting data is needed in the provision of
broodstock and in the return of captive bred animals to natural habitats (restocking). The
acquisition of captive broodstock can be obtained from natural habitats, captive breeding,
overseas, booy, surrender, and other sources from communities, findings and conservation
organizations in accordance with Permenhut No P.19/Menhut-II/2005 (Permenhut 2005).
Captive breeding results are expected to increase and restore natural populations by
releasing at least 10% of captive breeding production back into natural habitats after meeting
qualification standards. To carry out breeding properly, genetic information on C. goffiniana
birds from their natural habitat origin, confiscated birds and existing breeding broodstock is
needed. This genetic information is also needed when breeders want to add broodstock to
avoid inbreeding, which can cause the quality of the breeding stock to decline. Research on
species identification, bird origin and distribution, and habitat suitability are very important
when releasing birds (LIPI 2016).
Law No. 5 of 1990 stipulates the grouping of protected and unprotected wild plants and
animals, and further regulates the provisions on what is allowed and prohibited. Law
enforcement against the implementation of this law is faced with problems in identifying
which species are protected or not, as well as the name of the species so that there are no
errors in the prosecution process. Two or more species often have similar morphological
characteristics such as C. goffiniana and C. sanguinea, and identification difficulties increase
when the birds are still chicks or even eggs. Government Regulation No. 8/1999 stipulates
that the utilization of protected animals for trade purposes is allowed when the animals are
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the result of second generation captive breeding and so on. Law enforcement related to the
implementation of this regulation is faced with the problem of distinguishing between natural
or captive-bred confiscated animals and in more detail the order of their generations. In
addition to this distinction, information on the origin of natural populations is essential in
handling animals when the choice is to return or release them back to nature. Confiscated
animals must be returned to their location of origin, as the information obtained from the
perpetrators is often inaccurate or even unknown. Therefore, accurate data is needed to
determine the origin of the confiscated animal so that when returning it can be done correctly,
errors in determining the location can make new populations invasive, possibly disrupting the
balance of the ecosystem and the purity of the species. To underpin conservation strategies
and actions, scientific data is needed through several studies. Therefore, the research
formulated the problem as shown in Figure 1.
The Indonesian government has developed conservation programs, strategies and action
plans (SRAK) to prevent the extinction of several species under threat of extinction,
including conservation of helmeted hornbills 2018 to 2028 (KLHK 2018), Sumatran tigers
2007 to 2017.
(Soehartono et al. 2007), sea turtles 2016 to 2020 (KKP 2015), pig deer 2013
to 2022 (Permenhut 2013), orangutan 2019 to 2029 (KLHK 2019). However, there is no
specific conservation strategy and action plan for C. goffiniana. However, the SRAK for
ivory-billed hornbills can be used as a reference in carrying out conservation efforts for
C
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goffiniana because have the same threat of extinction. There are five strategies for ivory-
billed hornbill conservation, namely (1) good population and habitat management, (2)
strengthening rules and policies, (3) increasing cooperation between various parties, (4)
communication and public awareness for conservation 5) funding. Some of the conservation
programs are research, species and habitat monitoring, law enforcement, trade and population
monitoring, and population increase (MoEF 2018). Based on these strategies, conservation
actions that can be taken to manage the tanimbar cockatoo include evaluating habitats and
conservation areas, law enforcement, releasing confiscated birds back to nature, strengthening
and developing captive breeding.
Several studies of C. goffiniana have been conducted both outside the habitat and in its
natural habitat. Research on population status and its impact on agriculture was conducted by
Cahyadin et al. (1994a; 1994b); while Jepson et al. (2001) conducted research on its
conservation status. Behavioral research
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C. goffiniana in the laboratory was conducted by Auersperg et al. (2012; 2013a; 2013b;
2014). Astuti (2001) has analyzed genetic diversity in conservation institutions. Research by
O'Hara et al. (2018) and Mioduszewska et al. (2018) conducted studies on food and socio-
ecology.
However, research on C. goffiniana from ecological, genetic and conservation aspects
is still very limited and more studies need to be conducted. Complete and comprehensive
scientific data are needed to realize the preservation of Indonesian birds and biodiversity.
There are several questions in the formulation of the problem for this study, namely:
a) What are the ecological characteristics of C. goffiniana birds in Tanimbar Islands.
b) What are the genetic characteristics of C. goffiniana birds
c) Whether there are differences in genetic characteristics of C. goffiniana between wild and
breeding populations,
d) Are there population differences between islands in Tanimbar Islands.
e) Can a genetic approach be used to estimate the origin and range of C. goffiniana birds?
f) How to implement the conservation of C. goffiniana.
This research will use ecology, genetics, and conservation implementation approaches
for C. goffiniana in Tanimbar Islands so that the results can answer these research questions.
1.4 Research hypothesis
This research was conducted to answer the problems in the problem formulation and
get novelty. Therefore, several hypotheses were formulated.
1.4.1 Ecological Characteristics of C. goffiniana
Yamdena Island is the main and largest island in the Tanimbar Islands. Species richness
is determined by the ecological viability of the island, the size and amount of habitat, or the
availability of ecological niches, with larger islands generally having a higher diversity of
habitat types (Brose 2001). The results of Newton and Dale (2001) explain that unlike other
species, in birds there is no clear relationship between the size of each zoogeographic region
and species diversity. High habitat diversity leads to high diversity in bird taxa. The study will
also update bird diversity in Tanimbar Islands, which has several ecological interactions with
other species.
C. goffiniana. Various human activities through development and fulfillment of needs cause
changes in land use and availability of habitat and other resources. Research hypothesis of
ecological characteristics
C. goffiniana is as follows:
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H0: Land cover does not affect the genetic diversity of C. goffiniana birds
in the Tanimbar Islands.
H1: Land cover affects bird genetic diversity
C. goffiniana in the Tanimbar Islands.
1.4.2 Genetic Characteristics of C. goffiniana
Molecular approaches can be used to assess and understand geological evidence.
Phylogenetic and phylogeographic studies can provide patterns of species distribution that
were largely established in pre-Pleistocene times. The changing connectivity of islands in the
archipelago has affected the distribution of intraspecific variation (Hisheh et al. 1998). For
conservation purposes, genetic approaches can be further developed to understand
phylogeographic structure, past and present gene flow, demographic history, and genetic
diversity within species (Rocha et al. 2014). Genetic markers (mtDNA, microsatellites, single
nucleotide polymorphisms) are used to reveal genetic variation in a population (Manel et al.
2005). This genetic aspect research was conducted to answer the question of genetic
characteristics of C. goffiniana with several hypotheses.
a) Hypothesized differences in natural and breeding populations of C. goffiniana
H0: There is no difference in genetic diversity between wild and breeding populations
H1: There are differences in genetic diversity between wild and breeding populations
b) Hypothesis of differences between populations of C. goffiniana in Tanimbar Islands H0:
There is no separation of populations between P. Yamdena, P. Selaru, and
P.Larat
H1: There is a population split of P. Yamdena with Selaru Island, and Larat Island
1.4.3 Conservation Implementation of C. goffiniana
The utilization of wild plant and animal species has been regulated in Law 5 of 1990
Article 36, including for non-commercial activities such as assessment, research and
development, as well as commercial activities such as breeding in captivity; hunting, trade,
demonstration; exchange; cultivation of medicinal plants; and pleasure. Utilization of
protected plants and animals must be approved by the minister, except for some species that
must be approved by the president. According to Minister of Forestry Regulation No.
P.19/Menhut-II/2005 on Captive Breeding of Wild Plants and Animals, parent animals for
breeding purposes can be obtained from natural habitats, captive breeding, foreign countries,
booty, surrender from the public, and conservation institutions. Furthermore, it is also
stipulated that every breeder who conducts captive breeding is obliged to return to their
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natural habitat captive-bred plant and animal specimens of protected species that have met the
breeding qualification standards of at least 10% of the breeding results (Permenhut 2005).
Laws and regulations have regulated clearly and in detail in terms of utilization, but illegal
activities still continue to take place that can threaten the extinction of a species of wild plants
and animals such as poaching, illegal logging, and trade. This is evidenced by the findings of
the results of law enforcement confiscated by law enforcers.
Determining locations for releasing confiscated birds, restocking captive-bred birds,
and collecting broods from natural populations is not easy. This is due to the distribution of
natural populations of C. goffiniana on several islands (Yamdena, Selaru, Larat and other
islands in Tanimbar Islands), the presence of other introduced populations (Kai Islands,
Puerto Rico and Singapore), and the unclear origin of the broodstock (confiscation and trade).
Incorrect placement of release sites will result in introductions that can disrupt existing bird
populations. Knowledge of the origin of captive-bred broodstock is necessary for the addition
of broodstock and restocking to natural populations. Therefore, this study was conducted to
find out how the implementation of conservation activities that have been carried out so far to
preserve C. goffiniana.
All data from the study will be used as a scientific basis for the management of C.
goffiniana so that conservation efforts can be carried out appropriately and comprehensively.
Data on genetic variation and gene flow provide clues to population viability that can be used
to guide translocation in population management (Woodruff 2001). Therefore, the
identification of species, populations, origins, distribution, range and habitat suitability is
very important when carrying out various conservation efforts and conservation actions to
release C. goffiniana birds. Thus, conservation activities carried out through protection,
preservation and utilization can be carried out properly and achieve optimal results. The
implementation of C. goffiniana conservation, especially in law enforcement, is faced with
the problem of resolving confiscated birds, therefore two hypotheses were developed.
a) Estimating the origin of confiscated C. goffiniana birds
H0: Genetic characters cannot be used to determine the origin of confiscated C. goffiniana
birds.
H1: Genetic characters can be used to estimate the origin of confiscated C. goffiniana birds.
b) Range estimation of C. goffiniana bird
H0: Genetic characters cannot be used to estimate the home range of confiscated C.
goffiniana birds.
H1: Genetic characters can be used to estimate the home range of confiscated C. goffiniana
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birds.
Tanimbar Islands Maluku
The Tanimbar Islands are a small island group in the Tanimbar Regency of Maluku
Province, Indonesia, located between 6o 35'24" - 8o 24'36" N and 130o 37'47" - 132o 4'12"
East, bordered by the Banda Sea (north), East Sea and Arafura Sea (south), while the west is
bordered by Southwest Maluku Regency, and the east is bordered by the Arafura Sea (BPS
2021). Tanimbar Regency has an area of approximately 52,995.2 km2 with a land area of
10,102.92 km2 and waters of 42,892.28 km2. The land area is in the form of lowlands and
hills with the highest peak on Labobar Island around 300 m above sea level (masl). Tanimbar
Regency has two peaks of rainfall, the 2020 data shows the highest rainfall in May (395 mm)
and November (413 mm), while the lowest rainfall in August was 10 mm, the average
temperature was 26.4-29.7 °C, and the humidity was 75-85% (BPS 2021), so that this area is
included as a dry area with annual rainfall only ranging from 1500-200 mm (PemerintahKab
MTB 2012).
Yamdena Island is the largest of the 81 islands in the Tanimbar Islands with an area of
approximately 325,725 ha. The northern part of Yamdena is a lowland of less than 50 meters
above sea level, the southern part is hilly with an altitude of up to 200 meters above sea level,
and the southeastern part is hilly with an altitude of up to 260 meters above sea level (BPS
2021). The capital of Tanimbar Regency is Saumlaki, located on Yamdena Island. The
distance to Timor is about 500 km and 500 km north of Australia (Charlton et al. 1991).
Access to Saumlaki from Jakarta can be reached by airplane via Ambon for about 2 hours 40
minutes, then changing to a smaller plane for 1 hour 35 minutes with daily flights. Sea
transportation using PELNI ships can take about 30-44 hours from Ambon to Saumlaki with
an uncertain schedule, so regular schedule checks must be carried out.
The climate in the Tanimbar Islands often changes rapidly due to the influence of
climate change that occurs in the Banda Sea, Arafura Sea, and the Indonesian Ocean, due to
the circulation of seasonal winds that move from and towards the equator. Therefore,
Tanimbar has two seasons, namely the west season and the east season. The west season
occurs around October-February which usually coincides with the rainy season, while the
east season occurs around April-October which coincides with the dry season. The alternation
of the western season to the eastern season or vice versa is referred to as the transition season
which takes place in March-April and October-November (Purwanto et al. 2004).
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The community life system is governed by government administration and customary
institutions. Customary leadership is hereditary and has the authority to determine several
aspects of community life, social, cultural, economic, and natural resource management
(Purwanto et al. 2004). The population in 2020 was around 123,572 people with a growth
rate of 1.61%, and a density of 12 people per km2 (BPS 2021).
1.1 Biogeography of the Tanimbar Islands
The Wallacea region is the most interesting zoogeographic region because it lies
between the Oriental region to the west and the Australo-Papuan region to the west to the
east. This region is a transition zone for elements of the Oriental and Australo-Papuan
avifaunas that are divided into three sub-regions, namely Sulawesi, Lesser Sunda, and
Maluku (Monk et al. 1997; Coates and Bishop 1997). The Indomalaya-Australasian
Archipelago is a geologically dynamic region that is characterized by species richness and
endemicity (Lohman et al. 2011). The Maluku subregion is also a center of bird endemicity,
with 64 endemic species and 6 genera in the South Maluku (Coates and Bishop 1997). A total
of 336 species are recorded as endemic to the Wallacea region, including 150 species from
Sulawesi, 126 species from Lesser Sunda and 90 species from Maluku (Prawiradilaga 2019).
Although the fauna of the Wallacea region is highly endemic, hierarchical patterns of
relationships are evident within islands and between adjacent islands or continents (Michaux
2010).
The Tanimbar Islands, as one part of the Wallacea biogeography, are geographically
located between the Moluccas to the north, Australia to the south, Sulawesi to the northwest,
the small Sunda Islands to the west, and Papua to the east, and between the Lydekker and
Weber lines, which are considered to have never united with the Sahul shelf (Voris 2000).
However, White and Bruce (1986) included the Tanimbar Islands biogeographically as part
of South Maluku. Based on this geographical location, Tanimbar Islands is an area with high
species diversity and endemicity. One of them has a bird diversity of around 131 species,
including 10 endemic species and 16 endemic subspecies (Bishop and Brickle 1998).
BirdLife International (2020) designated Tanimbar Islands as an endemic bird area of the
Banda Sea Islands (EBA165).
Based on the geological history of the Tanimbar Islands, they were formed from
sedimentary, metamorphic and igneous rocks dating back to Quaternary times, which share a
common geological origin with the southern Moluccas. During the Pleistocene, many of the
present islands merged to form a larger land mass (Morley and Flenley 1987). Tanimbar Kep.
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belongs to the non-volcanic outer Banda arc along with Timor, Kai Besar, Seram and Buru
while the volcanic inner Banda arc consists of Solor, Alor, Wetar and Banda. The non-
volcanic outer Banda arc was formed by sedimentary, metamorphic and igneous rocks since
the quarter ages (Hall 2002). According to Audley-Charles (1986), the outer Banda arc
islands have never merged with islands of different geologic origin. Charlton et al. (1991)
explained that the Tanimbar Islands were geologically formed through deformation in the
Post-Pleiocene period characterized by the separation of the main island of Yamdena from
the western islands of the Tanimbar Islands (Figure 2).
The oldest rocks in Tanimbar are of Triassic age, which are dark gray and flake-shaped
volcanic mudflows. It is further explained that in the late Cretaceous and Palaeogene,
Tanimbar originated from deep sea waters, as the sandstone Ungar formation. The
Tangustabun formation formed in the late Miocene consists of gray clay, gray limestone, and
calcareous sandstone. The eastern part of Yamdena was formed by the Batimafudi formation
with a thickness of 700 to 1000 m in the Miocene era in the form of limestone. Furthermore,
in the Quaternary era, the Batilembuti formation was formed which consisted of sedimentary
rocks of clay and limestone and the Batilembuti formation Saumlaki is formed from coral
reefs less than 50 m thick (Charlton et al. 1991).
Tanimbar Kep. is a low island with an altitude of <300 m covered by marine deposits of
the Pleistocene era, indicating that the archipelago probably rose above sea level about 1
million years ago. The period of emergence is supported by coral beds up to 200m above sea
level. Furthermore, Yamdena is the first major island of the Tanimbar Islands to emerge
during the Pleistocene (De Smet et al. 1989). The geological history has a major influence on
the distribution pattern of fauna in the island region (Hisheh et al. 1998). The unique geology
leads to high endemicity (BAPPENAS 2016). One of the endemic birds with a distribution in
the Tanimbar Islands is C. goffiniana (Coates and Bishop 1997).
1.2 Tanimbar Cockatoo (Cacatua goffiniana)
Cockatoos are large parrots that have a distribution in Australia-Papua, almost all have
a distinctive crest, with a small distribution area, and most are endemic to the island.
Cacatuidae comprises 7 genera and 21 species, of which 7 species (33%) are threatened, i.e.
Near Threatened, Vulnerable, Endangered and Critically Endangered (Olah) et al. 2016;
Heinsohn et al. 2018; Winkler et al. 2020). Indonesia has 7 species of Cacatuidae, namely the
white cockatoo (C. alba), chef cockatoo (C. galerita), tanimbar cockatoo (C. goffiniana),
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Moluccan cockatoo (C. moluccensis), swamp cockatoo (C. sanguiena), yellow-crested
cockatoo (C. sulphurea), and king cockatoo (Probosciger atterimus). Phylogenetic
relationships within the Cacatuidae family (Figures 3, 4) show that C. goffiniana is closely
related to C. sanguinea, which is distributed in Papua and Australia (White et al. 2011;
Rowley and Kirwan 2020).
One of the smallest of the seven cockatoo species is C. goffiniana, characterized by
white plumage and a reddish bridle with a total length of 30-32 cm (Coates and Bishop 1997;
Eaton et al. 2016), a body weight of about 300 g and slightly smaller females (Rowley and
Kirwan 2020). Its closest relative, C. sanguinea, has a total length ranging from 36-39 cm
and a body weight of about 430-580 g. A notable difference is that the periophthalmic ring in
C. goffiniana is pale whitish blue while in C. sanguinea it is blue-gray and wider under the
eye (Rowley and Boesman 2020).
The taxonomic identity of cockatoos from Tanimbar Islands (also known as Timor-
Laut, Timur-Laut, Tenimber, Tenimbar or Tanimber Islands in older literature) was confused
during the 19th century. The Tanimbar cockatoo has a complex history, initially specimens in
Amsterdam Zoo, for which there is no data on the location of origin, was described by Otto
Finsch in 1863 as Lophochroa goffini. However, Schlegel in 1863 referred to it as C. goffini.
Roselaar and Michels (2004) mentioned that Roselaar and Prins in 2000 conducted a study on
specimens of Lophochroa goffini Finsch, 1863 and found that the species is as follows
C. ducorpsii Pucheran, 1853 which has a distribution in the Solomon Islands, because of this
misidentification, the name Cacatua goffini is proposed to be a synonym of C. ducorpsii.
Thus, cockatoos from the Tanimbar Islands do not yet have a valid official species name.
Roselaar and Michels (2004) proposed the new name C. goffiniana based on the description
of specimen RMNH 90750 collected by Felix Kopstein on April 22, 1923 from Saumlaki,
Tanimber a n d specimen RMNH 90751 from Timor Laut collected in 1882 by a hunter
named Riedel. The name was chosen after Andreas Leopold Goffin and Otto Finsch
originally intended to dedicate the species name in honor of his friend.
The natural distribution of C. goffiniana is P. Yamdena, P. Larat, and P. Selaru in
Tanimbar Islands of Maluku Province, where it was probably introduced to Kai Islands
(Roselaar and Michels 2004); other introduced populations were reported in Puerto Rico and
Singapore (Pérez-Rivera and Claudio 1997; Juniper and Parr 1998). The estimated range of
C. goffiniana is only 9200 km2 in the Tanimbar Islands (Birdlife International 2018).
Tanimbar cockatoos inhabit dry and moist tropical deciduous forests (Jepson et al. 2001).
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Population studies of C. goffiniana by (Cahyadin et al. (1994a; 1994b) ranged from 255,000-
347,088 individuals, while Jepson et al. (2001) estimated 231,476 ± SE 33,068 individuals.
Overexploitation and forest conversion leading to loss of important habitat threaten the
population decline of C. goffiniana.
In terms of social structure behavior, most cockatoo species exhibit complex social
structures (Seibert 2006), so observations of tanimbar cockatoos in the wild suggest high
sociability and little variation in social structure (Mulawka 2014). The first knowledge of
social structure in tanimbar cockatoos in the wild was that individuals behaved in sometimes
solitary, paired, or group behaviors (Cahyadin et al. 1994a; O'Hara et al. 2018). A seemingly
similar social structure has been reported for its close relative, C. pastinator (White et al.
2011). There are three levels of social organization in this species that correspond to different
movement patterns (Smith and Moore 1992). First, adult breeding pairs. Second, family
groups consisting of young still in the care of their mothers, and finally nomadic immature
flocks. Both pairs and families appear to settle in fixed areas of forest habitat, while nomadic
immature flocks will venture into other areas in search of suitable mates, food and habitat.
Groups of tanimbar cockatoos in the forest consist of one to six individuals, while
flocks of two to 305
individuals have been
found on agricultural
land. (Cahyadin et al. 1994a). Habitat
C. goffiniana in Yamdena Island are primary and secondary forests, mangrove forests,
freshwater swamp forests, agricultural fields, plantations, and grasslands (Jepson et al. 2001;
Roselaar and Michels 2004; Mioduszewska et al. 2018). Birds favor agricultural areas with
abundant food (Cahyadin et al. 1994a). Cahyadin et al. (1994b) and some local farmers
reported that tanimbar cockatoos during the rainy season will eat corn (Zea mays) on
farmland.
The Cacatuidae family, including Cacatua goffiniana, interacts with its environment
through a strong beak that is used for a variety of activities including manipulating objects,
foraging, climbing, and even acting as an additional limb with a strong tongue and dexterous
feet (O'Hara et al. 2018; Lambert et al. 2018), hence the functional versatility of the
cockatoo's beak, which is often referred to as a multipurpose tool (Huber and Gajdon 2006;
Hansell and Ruxton 2008). C. goffiniana has been observed using its beak to dig into loose
soil, shovel hardened clods of soil, tear off thick layers of fruit, peel tree bark, and
disassemble rotting tree trunks (O'Hara et al. 2018). C. goffiniana has long zygodactyl toes
that allow them to grip well, grasp objects, and even dig holes (O'Hara et al. 2018).
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Qualitatively, the main difference in foraging behavior between forest and farmland is
the level of individual numbers. Trees in forests provide a wide variety of fruits,
inflorescences, and seeds, most of which are located on At higher altitudes (>10 m), the
number of individuals or groups of foraging birds is smaller (<10 birds). On the other hand,
agricultural fields that provide a large number of food sources in one area result in a large
number of individuals (>15 birds) coming to feed at the same time. As birds forage in the
fields, a small number of individuals (1-3) remain perched on higher ground and likely act as
sentinels, as is also the case with Cacatua ducorpsii (Mulawka 2014). Similar foraging
behavior has been observed in wild C. goffiniana in Singapore, which typically forages in
groups of 15 or more where food resources are abundant (Neo 2012).
1.3 Habitat of Cacatua goffiniana
Yamdena Island supports an area of monsoon forest and semi-evergreen forest to the
east of Sumbawa. The monsoon forest is the most threatened tropical forest type in Wallacea,
and Tanimbar Island plays an important role in protecting and conserving this unique forest.
However, the threat of forest conversion and forest destruction continues to this day, so that
land cover continues to be altered by human interests. A timber company had exploited the
forest since 1991 until its license was revoked in 2007, but subsequently through
recommendations from the Regent and Governor, a new license was issued to a different
company to utilize timber products. Until 1998, P. Yamdena was still dominated by primary
forest, but over the next ten years there has been a shrinkage of 14.48% or 9,473 ha/year due
to logging (Rosantika 2016).
Land cover classification in P. Yamdena according to Jepson et al. (2001) divides into
eight main habitats, namely moist deciduous forest, mosaic forest, dry deciduous forest,
logged forest, mangrove forest, swamp forest, agricultural land, and grassland (Figure 5). The
presence of Cacatua goffiniana is generally found in agricultural land, forests and mangroves
but not in coconut plantations and grasslands (Cahyadin et al. 1994). Moist deciduous forests
are located along the hills of the southeastern coast and the southern part of the island, which
receives more rainfall. Drier deciduous forest and less complex vegetation predominate in the
western part of the island, which gradually merges with moist/dry mosaic forest with more
diverse vegetation on the relatively wetter east coast. Logging forests in much of southern
Yamdena form low forest stands. Swamp and mangrove forests are present along the west
coast, especially in the southern part of the island. The availability of freshwater has limited
settlement on the east coast, causing most of the coastal habitat to be converted to agricultural
14
land. Some people have also built settlements on the edge of the forest so that in the forest
area between tall vegetation they have made fields for agriculture (Cahyadin et al. 1994).
The highest population density of C. goffiniana was 105 individuals/km2 in moist
deciduous forest (Jepson et al. 2001). In contrast, Lambert (1993) found that the highest
density of Cacatua alba in North Maluku was in logged-over forest. Tanimbar cockatoos
were observed foraging in cornfields, dry rice fields, peanuts, and green beans (Jepson et al.
2001). The Tanimbar Islands with their diverse habitat types are recommended by Birdlife
International as one of the endemic bird centers that are also relatively rich in bird diversity.
As the Tanimbar Islands are located close to Australia, they are the destination of 15 species
of migrants from Australia (Bishop and Brikle 1998).
1.4 Conservation of Cacatua goffiniana
1.4.1 Local people's perception of C. goffiniana
The people of Tanimbar Islands realize their high dependence on biological resources
for their livelihoods. Since long ago, the community has had a traditional institutional order
to manage and utilize its natural resources, which can now be referred to as conservation.
Customary leadership holds the reins in regulating all aspects of life, namely social norms,
culture, economy, and biological resource management. The concept built by traditional
communities in managing biological resources includes the concept of petuanan to manage
natural resources spatial management and the concept of sacredness in the management of its
species. Petuanan is the concept of control of the area by a clan that exclusively controls the
full natural resources in the area, so that in its utilization must obtain permission from the
customary chairman of the petuanan (Purwanto et al. 2004). Sacredness is the concept of
protecting natural resources by making an area or type of resource as something sacred or
sacred which is associated with religious views or local people's belief in the role of ancestors
in their lives. Values in traditional societies have experienced changes in perspective along
with the times, caused by the influence of modern culture, education levels, and
psychological influences due to ancient and unrealistic assumptions (Purwanto et al. 2004).
The goal of managing biological resources that considers ecology and ethics has shifted to the
main economic goal, resulting in the exploitation of biological resources such as logging and
hunting of wildlife for trade. This includes the Tanimbar cockatoo (C. goffiniana), which is
locally called "manik tilngoi" in Tanimbar language. To protect this bird, their ancestors told
them that the white bird came from a cursed human child, and before flying away from its
15
family, the bird gave a message that it would eat garden plants before humans harvested
them. This myth, which has been passed down from generation to generation, aims to protect
the bird as it is a part of their ancestors.
Changes in people's views that are more concerned with economic goals have also
changed the views of local people towards the tanimbar cockatoo, especially in the 1980s.
The community considers this bird as an agricultural pest that damages and reduces crop
yields, causing losses. Since then, people have hunted and captured the bird to deal with the
pest and even make a profit by selling/trading the bird. Birdlife International (2018) estimates
that the trade of C. goffiniana in t h e 1980s was very high at around 10,000 birds per year,
and in 2005 illegal capture and trade activities of C. goffiniana still occurred, reaching 10,560
individuals (Burung Indonesia 2018). However, along with various approaches by various
parties both government and institutions to provide awareness and understanding of the status
of the bird species, the perception and actions of the community have also changed. The
c o m m u ni t y no longer captures birds that come t o farmland t o eat corn and other
agricultural products, but only guards and evicts them from the land.
1.4.2 Conservation Efforts by the Government
The main population threats to the bird are caused by capture for the wildlife trade and
habitat alteration through logging and conversion to agriculture (BirdLife International 2018;
Winkler et al. 2020). Conservation efforts to protect C. goffiniana have included conservation
efforts are carried out both nationally and internationally. Based on Law No. 5 of 1990, the
government makes conservation efforts through protection, preservation, and sustainable
utilization. C. goffiniana has been designated as a protected animal through Government
Regulation No. 7 Year 1999, which was updated with Minister of Environment and Forestry
Regulation No. P.106/MENLHK/SETJEN/KUM.1/12/2018. To conserve and protect C.
goffiniana in its natural habitat, the Minister of Forestry Decree No. 149/Kpts-II/1999
established the Tanimbar Wildlife Sanctuary in Tanimbar Islands. Other conservation efforts
include law enforcement, ex-situ captive breeding, and release of confiscated birds into the
wild. Internationally, a convention called the Convention on International Trade in
Endangered Species of Wild Fauna and Flora (CITES) has listed C. goffiniana as Appendix I
since 1992 (CITES 2019). The IUCN redlist categorizes C. goffiniana as Near Threatened
(NT), which means that C. goffiniana meets the criteria of near threatened (Birdlife
International 2018).
16
1.5 Biodiversity in Tanimbar Islands
Based on the geographical location and geological history of the formation of Tanimbar
Islands, this area has high species diversity and endemicity, both flora and fauna. This is
supported by the diversity of habitat types that form an ecosystem in Tanimbar Islands, so
that one of the interactions between C. goffiniana birds and other flora and fauna or with
other bird communities in fulfilling their needs for food, nests and becoming part of the food
chain. The Tanimbar Islands are home to hundreds of bird species with high levels of
endemism. The first ornithological expedition recorded in Tanimbar was conducted by
Forbes in 1882 on P. Yamdena, P. Moeloe and P. Kirimoen. The first list of bird species in
Tanimbar Islands was compiled by Bishop and Brickle (1998) from data provided by various
experts and expeditions between 1882 and 1997 recording 131 species.
Flora diversity has been identified around 804 local names of plant species in Tanimbar
Islands from various families including Poaceae, Solanaceae, Bignomiaceae, Annonaceae,
Urticaceae, Asclepiadaceae, Sapindaceae, Fabaceae, Malvaceae, Asteraceae, Tiliaceae,
Meliceae, Mimosaeae, Sapotaceae, Moracaceae, Piperaceae, and many other families
(Purwanto et al. 2004). IPH LIPI research (2018) identified biodiversity in Kep. Tanimbar,
namely 157 species of moth insects (Lepidoptera), 14 species of dragonflies (odonata), five
species of amphibians (Hilidae and Ranidae families), 15 species of reptiles (Agamidae,
Gekkonidae, Scincidae, Varanidae, Colubridae, Elapidae, Phytonidae, and Typhloidae), and
11 species of bats (families Pteropodidae, Emballonuridae, Rhinolophidae, Hipposideridae,
and Miniopteridae).
Limited access, resources and funding to explore biodiversity in Tanimbar Islands has
led to limitations in uncovering species diversity and its potential. One of the new species
findings of Vitessa segereri (Lepidoptera, Pyralidae) from P. Yamdena was described by
Buchsbaum et al. (2014) who explained that the species of this genus had never been known
before in Tanimbar Islands. Further findings by Narakusumo et al. (2019), which has
described seven new species of beetle insects long snouted Trigonopterus spp. (Coleoptera,
Curculionidae) from P. Yamdena, P. Larat and P. Selaru. The discovery of this beetle species
also adds information about the role of islands in Maluku including Tanimbar Islands as
stepping stones for the spread of this group from Papua which is widespread in Sulawesi and
the Greater Sunda. The most recent description by Weijola and Kraus (2023) found a new
species of monitor lizard, Varanus tanimbar, thus adding to the record of endemic fauna in
Tanimbar Islands, namely six species of reptiles. The discovery of other new species of flora
and fauna is still very much open, especially if it can reach islands with a high level of
17
difficulty so that it can contribute to revealing biodiversity and its potential for science and
society.
II METHODS
3.1
Time and Place of Research
Ecological data collection was conducted in September-December 2015, March-April
2017, April-May 2018, December 2018, and November 2020 on P. Yamdena, P. Larat, P.
Selaru, and P. Vaimar in the Tanimbar Islands, while sampling of natural populations for
genetic studies was conducted in November 2020 on P. Yamdena, P. Larat, and P. Selaru
(Figure 6). Confiscated bird samples were obtained from law enforcement activities of the
East Java Natural Resources Conservation Center in Surabaya and Jember in July 2019. Data
on the illegal trade of C. goffiniana between 2009 and 2020, as well as its utilization by zoos
and captive breeders were collected from 2018-2020. Data were obtained from WCS-IP,
BKSDA, and various media reports. Data on ex-situ conservation efforts to conserve this bird
were obtained from the Indonesian Zoo Association (PKBSI), while captive breeding data
were obtained through inspections of several captive breeders in Bali, East Java, Central Java
and West Java. Data analysis and laboratory activities were conducted at the Biology
Research Center-Indonesian Institute of Sciences (now the National Research and Innovation
Agency) Cibinong, Bogor.
3.2
Tools and Materials
Ecological data collection used several tools including binoculars, cameras, Global
Position System (GPS), Wallacea regional bird guidebook, stationery, data sheets, meters,
calico bags and 6m, 9m and 12m mist nets. Landsat 8 OLI image data for the study area was
downloaded from the United States Geological Survey (USGS) website
(http://earthexplorer.usgs.gov/). This image has a spatial resolution of 30 m covering the
entire Tanimbar Islands region consisting of two scenes, namely path/row: 106/065 and
path/row: 106/066. Most of the Tanimbar Islands land area is recorded in path/row 106/65,
while path/row 106/66 records less land and ocean area (Figure 5.1). In 2019, 22 Landsat
images captured the Tanimbar Islands. The images used were taken on November 14, 2019 at
01:21:57 and 01:22:21 (Figure 7).
Genetic data collection used the following equipment: sampling tube, micropipette,
18
spin down centrifuge, centrifuge, Polymerase Chain Reaction (PCR) machine Applied
Biosystems Thermal Cycler (Type 2700), vortex, incubator, magnetic stirrer, rotary shaker,
aspirator, electrophoresis apparatus, freezer, UV Transilluminator, Bioanalyze, and various
other laboratory support tools. Genetic research used 49 samples of blood or feathers of C.
goffiniana birds from various locations (Table 1). Four samples were used for mitochondrial
genome (mitogenome) analysis and 45 samples were used for genetic diversity analysis with
three marker genes, namely COI, Cyt B, and ND2. Natural population samples were obtained
based on SATS-DN No. S.54/K.19/ SATS-DN/11/2020 from Tanimbar Kep. namely P.
Yamdena, P. Selaru, P. Larat, and confiscated birds obtained from law enforcement by the
East Java Natural Resources Conservation Center based on SATS-DN No.
344/K.2/BIDTEK.1/KSA/7/2019.
Laboratory activities require various wear materials such as various sizes of pipette
tips, collection tubes, 0.2 ml thin-walled PCR tubes, Eppendorf DNA LoBind microcentrifuge
tubes 1.5ml, Safe-Lock Tube 1.5 ml, rubber gloves, tissues, and several other materials.
Laboratory materials used to obtain genetic diversity data include QIAGEN Dneasy Blood
and Tissue Kit (Buffer AL, Buffer ATL, Buffer AW1, Buffer AW2, Buffer AE, proteinase K),
ethanol (96-100%), Dneasy mini spin column, distilled water, 1% and 2% agarose gel,
Fluoroscense, TAE, loading dye, marker, oligo primer; MgCl2, dNTP; PCR buffer, Taq-
polymerase, Isopropanol, Tris (1 M), RNase-free, NaCl (5 M) RNase-free, TE buffer pH 8.0.
Materials used for the structure and composition of the mitochondrial genome require various
materials including NEBNext® Ultra™ II DNA Library Prep Kit for Illumina® and Agilent
2100 Bioanalyzer.
3.3
Work Procedure
3.3.1 Ecological characters of C. goffiniana
Data collection on bird communities in Tanimbar Islands and their interactions with C.
goffiniana was carried out using the roaming method (Siegel 2009) to make bird observations
and capture using mist nets. Observations were conducted at 06.00-10.00 WIT and 15.00-
18.00 WIT. The use of mist nets was only conducted in P. Yamdena, precisely in Lorulun
Village in 2015 and 2018, and Tutukembong in 2015. This method was used to obtain data on
birds that are shy and have limited movement, making it difficult to observe directly.
Observations recorded species encounters, habitat use, breeding period and some interaction
behaviors between C. goffiniana and other bird communities in Tanimbar Islands.
19
Preliminary information on nest sites was obtained from interviews with local communities,
followed by observations, identification and measurements of trees. Indicators used to
determine which bird species are entering the breeding period are by observing breeding
behavior, including making nests, finding eggs or chicks in the nest. Interactions between C.
goffiniana and other bird communities were observed in foraging areas and potential nesting
trees on P. Yamdena and P. Selaru.
Data processing for land cover classification is carried out through several stages,
namely: image pre-processing, image classification and accuracy testing. The pre-processing
stage involved changing the format of GeoTiff data from each band to image format (.img).
The next stage is layerstacking so that multispectral analysis can be carried out, merging
(mosaic) two or more spatially separated photos and images.
The class classification processing on the Tanimbar Islands image uses 315 sample area
data (training area). This data was obtained from Google Map and direct object recognition
on P. Yamdena, P. Selaru and P. Larat according to the classification to be used, such as
forest, wetland, mangrove, dry land and open land agriculture, mixed plantation, and built-up
land. The forest cover class uses sample areas from direct checks of both primary and
secondary forests. Built-up land uses training data in the form of residential areas, airports,
ports, and other infrastructure. The determination of dryland agriculture and open land uses
sample areas of shrubs, grasslands, agricultural fields with annual crops, and land without
vegetation. The mixed plantation classification identifies land planted with plantation crops,
such as coconut, teak and banana. Mangrove land cover consists of mangrove vegetation
around rivers and beaches.
3.3.2 Genetic Characterization of C. goffiniana
Genetic character data were obtained by analyzing the mitochondrial genome and its
genetic diversity. DNA extraction of bird samples was carried out using Qiagen DNeasy
Blood & Tissue Kits. The principles of DNA extraction include cell membrane destruction,
DNA separation, and DNA precipitation. The main stages of DNA extraction using the kit
consist of cell lysis, DNA binding, DNA washing, and elution using reagents/buffers
provided by the Extraction Kit manufacturer. The work stages follow the Extraction Kit
protocol used, which is as follows: 5 mg of blood sample that has been preserved in 95%
alcohol is taken from the sample tube. The remaining alcohol in the sample is absorbed using
tissue paper and put in a 1.5 ml ependorf tube to be vacuumed to dry and then pulverized by
20
grinding to accelerate the process of cell membrane lysis. Next, Buffer ATL was added as
much as 200 μl and Proteinase K 20 μl and vortexed for 15 seconds, then incubated at 56o C
(1-2 hours) while vortexing every 20 minutes. The next step was that 200 μl AL buffer was
added, then vortexed for 15 seconds, absolute ethanol was added as much as 200 μl, then
vortexed 15 seconds. The whole mixture was pipetted and put into a 2 ml Dnaesy mini spin
column tube, then centrifuged at 800 rpm for 1 minute. Discard the liquid and collection tube
and transfer the DNeasy mini spin column to a new collection tube. Add 500 μl Buffer AW 1,
then centrifuged 800 rpm for 1 minute. Discard the liquid and place the DNeasy mini spin
column back into a 2 ml collection tube. Add 500 μl Buffer AW 2, then centrifuged 14,000
rpm for 3 minutes. The liquid and collection tube were discarded, and the DNeasy mini spin
column was transferred to a new 1.5 ml tube. Next, 100 μl of Buffer AE (Ellution buffer) was
added to wash the DNA on the mini spin column matrix. Incubate for 1 minute, then
centrifuge 800 rpm for 1 minute and obtain total DNA.
The next stage after obtaining total DNA is to amplify the mitochondrial DNA
(mtDNA) gene with the Polymerase Chain Reaction (PCR) method on a PTC-100 machineTM
Programmable Thermal Controller. In this study, we will use several Mitochondrial DNA
(mtDNA) marker primers, namely the Cytochrome C Oxidase Subunit I (COI) gene,
Cytochrome-b (cyt-b), and NADH dehydrogenase subunit-2 (ND2) (Table 2).
PCR amplification refers to Hebert et al. (2004), Astuti (2011), and Tavares et al.
(2006). The PCR reaction volume made was 25 µl with a composition of 1 µg DNA sample;
25 pmol/µl primers; 1.5 mM MgCl2 ; 0.1 mM dNTP; 10 µl 10xPCR buffer and 2 units of
Taq-polymerase. PCR conditions for mtDNA amplification were as follows predenaturation
94o C for 5 minutes, then denaturation 94o C for 40 seconds, primer attachment at 50o C for
40 seconds, elongation at 72o C for 1 minute, post extension 72o C for 7 minutes and storage
20o C for 5 minutes. The amplification cycle was performed 36 times. PCR results were
checked by electrophoresis on 2% agarose gel. Visualization is done using ethidium bromide
staining and the help of ultraviolet light read using Gel-DOC tool (UVItec). The PCR results
obtained were then carried out the DNA sequencing process using the ®3130 Genetic
Analyzer machine (Applied Biosystems, USA).
Mitochondrial genome (mitogenome) analysis begins with sample preparation and
sequencing. The sample preparation stage is to make a DNA library by using 50 μl of DNA
as the starting material for DNA library construction with the NEBNext® Ultra™ II DNA
Library Prep Kit for Illumina®. The stage begins with Fragmentation, which is the process of
fragmenting DNA into one reaction to produce a DNA library that is consistent and has a
21
uniform fragment size. The steps in fragmentation are the results of DNA extraction
quantified with Qubit® using the dsDNA HS Assay Kit (Thermo Fisher Scientific) and
diluted to produce a DNA concentration of 1 ng/µL. A total of 50 μL of diluted DNA was
loaded into a Covaris microTUBE-15 AFA Beads Screw-cap (PN 520145). DNA was then
cut on Covaris model M220 Focused-ultrasonicator with XTU Holder (PN 500414) with a
length of 500 bp in duplicate for each concentration.
The fragmentation results are used for the End prep stage by taking 50 μl of
Fragmented DNA into a tube and mixing it with 7 μl (green) NEBNext Ultra II End Prep
Reaction Buffer, and 3 μl (green) NEBNext Ultra II End Prep Enzyme Mix so that it becomes
60 μl. Use a 100 μl or 200 μl pipette to mix the solution until it is well mixed. Place the tube
in a thermal cycler that has been set to a temperature ≥ 75°C, and run the program as follows
30 minutes @ 20°C, 30 minutes @ 65°C, and hold at 4°C.
The next stage is Adaptor Ligation, starting with checking the results of the previous
stage, if the input result is ≤ 100 ng then dilution with NEBNext Adaptor for Illumina in 10
mM Tris-HCl, pH 7.5- is required.
8.0 with 10 mM NaCl. Next, make the following reaction mixture: 60 μl of End Prep Reaction
Mixture, 2.5 μl (red) NEBNext Adaptor for Illumina**, 30 μl (red) NEBNext Ultra II
Ligation Master Mix*, 1 μl (red) NEBNext Ligation Enhancer for a total of 93.5 μl reaction.
Mix all the solutions until they are well mixed using a pipette and spin quickly to lower the
solution that sticks to the wall of the tube. Then incubate at 20°C for 15 minutes in a thermal
cycler, then add 3 μl of USER® Enzyme (red / blue) solution into the reaction solution. Mix
well and incubate at 37°C for 15 minutes. Furthermore, the sample can be stored overnight at
-
20°C.
The next stage Size Selection or Cleanup of Adapter-ligated DNA is to select the size of
the ligated DNA adapter, starting with checking the results of the previous stage must have a
volume of> 50 ng. The initial volume used is 96.5 μl with 150 bp insert size selection so that
the Final Library Size Distribution (insert + adapter + primers) is around 270 bp. Add 50 μl
1st Bead Addition and 25 μl 2nd Bead Addition and vortex NEBNext Sample Purification
Beads. Add 40 μl (~0.4X) bead into the 96.5 μl ligation reaction and mix well by pipetting up
and down at least 10 times. Vortexing for 3-5 seconds at high speed can also be used. If
centrifuging the sample after mixing, be sure to stop the centrifugation before the granules
begin to settle. Incubate the sample for at least 5 minutes at room temperature. Place the tube
on a suitable magnetic stand to separate the bead from the supernatant. If necessary, rotate the
22
sample quickly to collect liquid from the sides of the tube or plate orifice before placing on
the magnetic stand. After 5 minutes (or when the solution is clear), carefully transfer the
supernatant containing the DNA to a new tube (caution: do not discard the supernatant).
Discard the bead that contains large unwanted fragments. Add 20 μl (0.2X) resuspended
NEBNext Sample Purification Beads to the supernatant and mix at least 10 times. Take care
to remove all liquid from the tip during the last stir.
The next step is to incubate the sample for at least 5 minutes at room temperature. Place
the tube on a suitable magnetic stand to separate the bead from the supernatant. If necessary,
rotate the sample quickly to collect liquid from the sides of the tube before placing on the
magnetic stand. After 5 minutes (or when the solution is clear), carefully remove and discard
the supernatant containing unwanted DNA. Carefully remove the bead containing the desired
target DNA (Caution: do not discard the bead). Add 200 μl of 80% ethanol to the tube while
it is in the magnetic stand. Incubate at room temperature for 30 seconds, then carefully
remove and discard the supernatant. Be careful of the bead containing the target DNA.
Repeat the above steps once more for a total of two washes. Be sure to remove all visible
liquid after the second wash. If necessary, briefly rotate the tube, put it back on the magnet
and remove the remaining ethanol with a p10 size pipette tip. Dry the bead for up to 5
minutes while the tube is on the magnetic stand with the cap open. Caution: the bead should
not be too dry as this may result in lower DNA target recovery. Elute the sample when the
bead is still dark brown in color and appears shiny when all visible liquid has evaporated. If
the bead turns light brown and starts to crack, the bead is too dry. Remove the tube from the
magnetic holder. Elute the target DNA from the bead into 17 μl of 10 mM Tris-HCl or 0.1X
TE. Mix well with a vortex mixer or by pipetting up and down 10 times. Incubate for at least
2 minutes at room temperature, if necessary, rotate the sample rapidly to collect liquid from
the sides of the tube. Place the tube on the magnetic stand After 5 minutes (or when the
solution is clear), transfer 15 μl to a PCR tube
(amplification). Samples can be stored at -20°C.
The next step is PCR Enrichment of Adaptor-ligated DNA. Make a reaction mixture
into a new tube with a composition of 15 μl Adaptor Ligated DNA Fragments, 25 μl (blue)
NEBNext Ultra II Q5 Master Mix, 5 μl (blue) Index Primer/i7 Primer, and 5 μl (blue)
Universal PCR Primer/i5 Primer for a total volume of 50 μl. Mix well by pipetting up and
down 10 times and rotating quickly to collect the reaction that sticks to the wall of the tube.
Place the tube on a thermal cycler and perform PCR amplification using the following PCR
conditions predenaturation at 98°C for 30 seconds for 1 cycle, denaturation at 98°C for 10
23
seconds for 15 cycles, annealing/extension at 65°C for 75 seconds, final extension at 65°C for
5 minutes for 1 cycle, and post extention at 4°C.
Next, the purification process is carried out using magnetic bead, by adding 45 μl
(0.9X) of resuspended bead into the PCR reaction. Mix well by pipetting up and down at
least 10 times. Be careful to remove all liquid from the tip during the last stir. Vortexing for
3-5 seconds at high speed can also be used. If centrifuging the sample after mixing, be sure to
stop the centrifugation before the granules begin to settle. Incubate the sample on a table for
at least 5 minutes at room temperature. Place the tube on a suitable magnetic stand to separate
the bead from the supernatant. If necessary, rotate the sample quickly to collect liquid from
the sides of the tube before placing on the magnetic stand. After 5 minutes (or when the
solution is clear), carefully remove and discard the supernatant. Be careful of the bead
containing the target DNA (Caution: do not discard the bead). Add 200 μl of freshly prepared
80% ethanol to the tube while it is in the magnetic stand. Incubate at room temperature for 30
seconds, then carefully remove and discard the supernatant. Repeat the step once for a total of
two washes. Be sure to remove all visible liquid after the second wash. If necessary, briefly
rotate the tube/plate, reattach it to the magnet and remove the remaining ethanol with a p10
pipette tip. Dry the bead for 5 minutes while the tube is on the magnetic stand with the cover
open. Remove the tube from the magnetic stand. Elute the target DNA from the bead by
adding 33 μl of 0.1X TE. Mix well by pipetting up and down 10 times, incubate for at least 2
minutes at room temperature. Place the tube back on the magnetic stand, after 5 minutes (or
when the solution is clear), transfer 30 μl to a new PCR tube and store at -20°C. Then, the
DNA library results were measured for quality and concentration using the Agilent 2100
Bioanalyzer, then the samples can be stored at -20°C. Four samples of DNA library results
were then sent using Novogene sequencing services, China for sequencing with Illumina
Hiseq Xten for 2 x 150 bp.
3.3.3 Conservation Implementation
Legal trade data of C. goffiniana from 1981-2018 were obtained from the CITES Trade
Database (UNEP-WCMC 2019) and downloaded on May 15, 2020. The category used to
obtain the data was the year range (1981-2018), exporting country (Indonesia), importing
country (all countries), source, destination, and form traded. CITES classifies these types into
four: live, whole body, feather, and specimen. In addition, the source of the birds is divided
into six codes: captive-bred (C), wild (W), undetermined (U), captive-born (F), pre-
convention (O), and confiscated (I). The traded birds had seven purposes: zoo-keeping,
24
personal, commercial, scientific, circus, captive-bred, and unspecified.
Data Analysis
Ecological Character Analysis:
Descriptive analysis was used to determine the species status and associations between
C. goffiniana and other species. Species status determines the species category in terms of
protection, endangerment (IUCN Red List), endemicity, migration and trade (CITES
Appendix). The protection status of the species is categorized based on the appendix of the
species list in Permen LHK No. P.106/MENLHK/SETJEN/KUM.1/12/2018 concerning the
Second Amendment to the Regulation of the Minister of Environment and Forestry No.
P.20/MENLHK/SETJEN/KUM.1/6/2018 concerning Protected Plant and Animal Species.
Species threat status is categorized based on the IUCN Red List of Threatened Species, which
consists of nine (9) categories, namely Critically Endangered, Endangered, Vulnerable, Near
Threatened, Not Evaluated, Data Deficient, Extinct, Extinct in the Wild, and Least Concern.
The CITES Appendix species categories follow the checklist of CITES species
(www.cites.org) which categorizes threats into three Appendices. Appendices I are species
that are threatened with extinction if trade is not stopped. Appendix II are all species that are
not immediately threatened with extinction, but will become so if trade continues. Appendix
III are all species that are not threatened with extinction, but all CITES member countries
may only trade with export licenses that are in accordance with the certificate of origin.
Endemicity status refers to a restricted distribution with a range of less than 50,000 km2
(Stattersfield et al. 1998) where birds are either endemic or endemic to the Tanimbar Islands.
Migratory status refers to birds that travel long distances from the northern or southern
hemisphere in winter to Indonesia but continue to breed in their home range.
Supervised classification is used for digital image classification analysis that provides
guidance to the computer in the classification process with the Erdas Imagine 2014 program.
To create, display, edit and layout maps using ArcGIS
10.3. After clustering the object class, it is evaluated with separability and accuracy. The
separability evaluation is used to show the separation of classes. The accuracy evaluation is
u se d t o see the level of accuracy that occurs in the classification of the sample area so that
the percentage of accuracy can be known. This evaluation tests the visual accuracy of the
guided classification. Accuracy mapping is done by creating a contingency matrix or error
matrix. The accuracy calculated is user's accuracy and producer's accuracy. Class evaluation
25
uses separability and contingency values, while classification accuracy is assessed based on
overall accuracy and kappa statistics. The habitat classification used was divided into five
types: forest, open land, swamp, mangrove and coastal. Forest type was not distinguished in
detail as primary and secondary monsoon forest. Open land habitats consist of settlements,
grasslands, farmlands, and open land. Swamp consists of all water bodies such as swamps,
ponds and rivers. Mangroves are specific vegetation located near rivers and coasts. Coastal
habitats consist of swamps, beaches and the sea.
3.4.2 Genetic Character Analysis
The sequenced nucleotide sequence was edited to remove unclear peaks at the end of
the sequence using Codon Code Aligner and then performed alligment using the MEGA X
program (Kumar et al. 2018). Sequence data analysis for the analysis of nucleotide base
variation in C. goffiniana populations was carried out with the MEGA program.
X and DnaSP version 6 (Rozas et al. 2017) to analyze haplotype number and haplotype
diversity. This analysis also used genetic data from several other cockatoo species such as C.
moluccensis as an outgroup for comparison with C. goffiniana.
Analysis of the mitochondrial genome begins with quality checking of the sequence
data using FastQC [http://www.bioinformatics.babraham.ac.uk/ projects/fastqc/]. Sequence
data generated from Illumina was then de-novo assembled using Novoplasty to produce
circular mitogenome data. This data was then validated using Geneious Prime by aligning
with the reference genome of C. goffiniana (Kim et al. 2021). To determine the protein
structure of genes in protein coding and ribosomal RNA, MITOS web server was used (Bernt
et al. 2013).
Phylogeny tree reconstruction was performed using the Maximum Likelihood (ML)
method with the Generalized Time-Riversible substitution model (GTR Model) built with
FastTree 2.1.11 (Price et al. 2010) in Geneious Prime 2023.0.4 software (Geneious Prime
2023). The outgroups used in the phylogeny tree reconstruction were C. pastinator, C.
galerita, and C. moluccensis.
3.4.3 Conservation Implementation Analysis
An assessment of C. goffiniana trade was conducted using descriptive analysis.
Furthermore, the data were tested for normality using the Kolmogorov-Smirnov Test, while
the Independent Samples T-Test was used to determine the difference in the number of C.
goffiniana traded before and after the bird was included in the list of protected birds and
26
Appendix I. Univariate ANOVA analysis was used to determine the effect of protection and
CITES listing on the number of legally traded birds. Statistical tests were conducted using
IBM SPSS Statistic 22 (IBM Corp. Released 2013).
4.1.1 Habitat of C. goffiniana and Avifauna in Tanimbar Islands
Tanimbar Islands, as the distribution area of C. goffiniana, is also a habitat for various
bird species. The results showed that the distribution of C. goffiniana was confirmed in P.
Yamdena, P. Selaru, P. Larat, P. Sera, and P. Molu. The distribution in P. Sera and P. Molu
has never been reported before. This study also recorded 92 bird species on P. Yamdena, P.
Selaru, P. Larat, and P. Vaimar (Tanimbar Islands). A total of 12 species have not been
reported in previous publications, most of which belong to shorebird and waterbird groups.
Mapping of habitat types used by birds in Tanimbar Islands was carried out with a land
cover classification analysis that resulted in six categories: forest, wetland, developed land,
mixed plantation, dry and open land agriculture, and mangrove (Figures 9, 10). Land cover
classification in the form of forest in Tanimbar Islands dominates in P. Yamdena, P. Waliaru,
P. Selu, P. Molu, and P. Maru. While the land cover on P. Selaru,
P. Larat, and P. Sera are dominated by dryland agriculture and open land, mixed plantation
land is dominant in P. Wotab and P. Laibobar. Built-up land is located mainly along the coast
and scattered on the larger islands (Yamdena, Larat, and Selaru).
Mangrove forests dominate watersheds, estuaries, and coastal areas. Dryland
agriculture and open land consist of shrubs, grasslands, farmland/gardens, and other vacant
land. Wetlands are shown in blue, while vegetation is shown in bright green. The brightness
of the green color indicates the density of the vegetation. High-density forests will appear
dark green compared to low-density or mixed forests. Furthermore, the forest class has the
lowest user accuracy (97.22%), while wetlands, settlements, dryland and openland
agriculture, and mangrove forests can be well separated with high user and producer accuracy
(100%).
Forest vegetation has a similar morphology to the plantation class due to the difficulty
of identifying object boundaries and lack of training data from direct observation. Built-up
land, dry land agriculture and open land based on visual interpretation cannot be clearly
classified, although the classification evaluation results show that the lowest separation value
occurs between the built-up land class and dry land agriculture and open land which has a
27
divergence transformation value of 480.151, meaning that the two classes are not separated.
In contrast, the other classes have a separation value between 1800-2000 because they can be
distinguished well.
Based on contingency, the highest level of fit for the training data was the forest class
(99.64%), while the lowest was mixed plantation (29.39%). Nevertheless, the overall
classification accuracy and kappa of 99.60% and 99.54% provide high accuracy. By
Therefore, the land cover classification resulting from the processing of Landsat image data
dated November 14, 2019 has high accuracy and the pixels are well classified. However,
forest cover is not further detailed, such as primary forest, secondary forest, and production
forest.
The analysis of land cover classification in Tanimbar Islands shows that forest is the
most extensive class among other classes. As the largest island in Tanimbar Islands, this
island has the largest area in almost all classes except dry land farming and open land located
on P. Selaru with an area of about 21,097.53 ha. The forest area in P. Yamdena is around
55,052.01 ha, which is the largest percentage (43.7%) of the total forest area in Tanimbar
Islands. In general, wetlands in Tanimbar Islands are very small, with P. Larat and P. Selaru
having only 147.6 ha and 144.36 ha respectively. The total area of each land cover
classification in Tanimbar and area on the three main islands of P. Yamdena, P. Larat and P.
Selaru are presented in Table 4.
The bird diversity of Tanimbar Islands is influenced by the diversity of habitat types,
namely forests, dryland and open land agriculture, wetlands, mangroves, mixed plantations,
and developed land. The avifauna of Tanimbar Islands is dominated by the order
Passeriformes and the families Ardeidae, Columbidae, Accipitridae. The number of bird
species based on the use of habitat types varies greatly (Figure 11).
4.2 Genetic Characteristics of C. goffiniana
4.2.1 Structure and Composition of the C. goffiniana Mitochondrial Genome
Characterization of the complete mitogenome (mitocondrial genome) sequence of C.
goffiniana from Tanimbar Islands obtained a circular genome with a total sequence length of
19,083 bp, its structure consists of 13 protein (protein coding genes), two ribosomal RNA
genes, 24 transfer RNA coding genes, and two control regions (Figure 25).
28
The difference between the two populations was caused by mutations in the samples
from the zoo in Seoul, Korea. The mutations that occurred were transitions, transversions,
deletions and insertions. The zoo samples had deletions in the 16,806th and 16,807th
nucleotide sequences with the loss of the cytosine base (C), while the 17,261th and 17,854th
sequences had deletions of the Guanine base (G). The 17,676th nucleotide sequence in the
zoo sample also had a Guanine (G) base insertion mutation.
4.2.2 Genetic diversity of C. goffiniana
Analysis of 49 samples with three marker genes (COI, Cyt-B and ND2) resulted in a
total nucleotide length of 2476 bp. The haplotype network formed three main clusters, the
largest cluster consisting of 14 haplotypes derived from 9 confiscated samples, seven samples
of the natural population of P. Yamdena, three samples of P. Larat and three samples from P.
Selaru. The second cluster was formed by eight haplotypes derived from 12 confiscated
samples, two samples from P. Yamdena and one sample from P. Larat. The third cluster was
formed by four haplotypes derived from four samples from P. Yamdena, one sample from P.
Larat, and P. Selaru, as well as two confiscated samples, while the four samples from P.
Yamdena analyzed by the mitogenome formed a separate haplotype (Figure 27). The analysis
showed high genetic diversity in C. goffiniana with variation at 44 sites with a Haplotype
diversity (Hd) value of 0.9464 and Nucleotide diversity (π)= 0.0017, forming 27 haplotypes
(Appendix 3).
The three clusters formed indicate that the population on P. Yamdena is the main
haplotype of the population that has spread to other islands, namely P. Larat, and P. Selaru. A
total of three samples from P. Selaru joined P. Yamdena to form the largest haplotype (H_6)
consisting of eight samples. The second largest haplotype (H_2) consisted of six samples
formed from P. Yamdena and confiscated samples, while the third largest haplotype (H_20)
consisted of five samples from P. Yamdena and P. Larat. This haplotype diversity was
formed due to variation caused by mutations, most of which were transitional mutations,
while transversion mutations only occurred at 16 sites (Appendix 3). Separate analysis with
COI, Cyt-B and ND2 resulted in a different number of variations and haplotype diversity
from the combined analysis of the three genes (Figure 28). Genetic diversity analysis with the
COI marker gene in C. goffiniana produced 681 bp, with a total of seven haplotypes and
nucleotide variation of six sites and Hd: 0.3002 (Table 11).
Nucleotide variations that occur due to mutations both transitions and transversions,
29
with COI analysis showing nucleotide variations at all six sites are transitional mutations
where there is a replacement of similar bases on two pyrimidine base sites (Cytosine-Timin)
and four purine base sites (Adenine-Guanine). This indicates that the mitochondrial COI gene
is relatively stable and low intra-specific variability so that it is not easily changed compared
to other genes. Based on the number of haplotypes and the number of nucleotide variations,
the COI marker gene for the analysis of C. goffiniana showed the least diversity compared to
other genes Cyt B and ND2.
Analysis of genetic diversity with the Cyt-B marker gene in C. goffiniana produced 746
bp, with a total of 13 haplotypes and nucleotide variation of 20 sites and Hd: 0.7577 (Table
12). Genetic variations that occur are 14 sites occur transitional mutations and six sites occur
transversions. The use of Cyt B markers to determine genetic variation at the species level is
more visible than COI, as evidenced by the number of haplotypes and mutated nucleotide
sites more than COI.
Genetic diversity with mitochondrial analysis of the ND2 gene has a nucleotide length
of 1049 bp showing a greater number of haplotypes than using the COI and Cyt-B genes,
namely 14 haplotypes, but Haplotype diversity and nucleotide variation are relatively lower
than COI, Hd: 0.7568 at 18 sites (Table 13). However, the use of Cyt-B and ND2 genes did
not show a very high difference, only one haplotype and two nucleotide sites. A total of ten
sites had transition mutations (Adenine-Guanine) and (Cytosine-Timin) and eight sites had
transversion mutations (Guanine-Cytosine) and (Adenine-Cytosine).
4.3 Conservation Implementation of C. goffiniana
4.3.1 Permanfaaatan C. goffiniana for Trade
The total number of legally traded C. goffiniana reported to CITES from 1981-2018
was 151,681. There are differences in numbers reported by exporters and importers as some
trade transactions are only reported by the exporting or importing country. Export trade after
the establishment of protection in 1990 and inclusion in Appendix I of CITES in 1992
declined sharply to around 10,240 birds, and since 2004, there has been no legal export of
this bird by Indonesia (Figure 30 and Table 15).
Trade dynamics were strongly influenced by the designation of C. goffiniana as a
protected bird in Indonesia. The number traded during 1981-1990 was 141,441 birds (about
14,144 individuals per year). Due to these strict regulations and the lack of breeding success
for this species, since 1993 there have been no large exports of C. goffiniana. CITES
30
recorded two birds in 1998-1999, one exported to Paraguay, and one to the United States.
Information on the destination and source of most of the birds traded is not specifically
mentioned in the CITES trade database, despite the fact that the numbers are very large
(Table 16), while the United States is the largest importing country (Table 17). The
unspecified destination of trade data is most likely in the commercial category. Similarly, data
on the source is unknown and it is possible that the figures are of wild origin.
Therefore, from year to year, the most common trade designation for C. goffiniana was
for purposes not specifically described in the document (58.6%), followed by commercial
(41.4%), private (0.002%), and zoo (0.001%). Birds traded were wild-caught (6.4%),
followed by captive-bred (0.032%) and confiscated (0.011%). A total of 141,949 individual
records (93.6%) did not include information on their source. These records were mainly those
that listed the exporting country (Indonesia) as the source. Therefore, it is likely that these
birds originated from the wild as breeders in Indonesia do not yet have the capacity to export
large quantities.
Commercial purposes and not specified in the documents account for the largest
amount of C. goffiniana international trade data. However, both are possible for the same
purpose of being used as a pet. Data clearly describing personal, zoo collection and breeding
purposes totaled only 25 individuals. The data for personal use of three individuals may also
be used as pets. The CITES Trade Database records a breeding trade of 20 individuals for
South African importing countries in 2003. The United States and Germany imported the
largest numbers of C. goffiniana from Indonesia between 1981 and 2018, totaling 118,356
and 8354, respectively. The destination countries of C. goffiniana exports from Indonesia in
1981-2018 were 34 countries (one country was not mentioned).
It is therefore possible that these numbers were used for breeding, although the original
documents reported commercial purposes. Only eight countries imported more than 1000
birds and the United States was the largest importer with a total of
118,356 birds or about 78% of the total birds exported by Indonesia. Cockatoos are the most
popular birds to be kept in the United States. This study shows that the source of the traded
birds is mostly from direct collection from the wild and
unknown source, as only 0.032% were reported to be of captive-bred origin.
4.3.2 Ecological Conservation of C. goffiniana
The Indonesian government encourages various parties to conserve C. goffiniana
through in-situ conservation in captivity and conservation institutions such as safari parks,
zoos, and nature parks (Table 18). Based on data from PKBSI, 10 out of 51 member
conservation organizations only have 46 birds of this species.
Furthermore, only six breeders are known to have attempted to breed this bird in
captivity, so the numbers available for breeding are very small, as there are only 56 males, 36
females and 4 individuals of unknown sex in captivity. This is not enough to produce a
second generation that can supply the local and international legal trade markets.
4.3.3 Threats to C. goffiniana from Illegal Activities
Illegal trade and capture from the wild continues to occur despite the species being
protected and included in Appendix I of CITES. This study through media monitoring,
NGOs, and BKSDA seizure tabulations found that 237 C. goffiniana were illegally traded
between 2009 and 2020 (Table 19). Data on the distribution of confiscated illegal possession
of C. goffiniana are mostly in Java, with a small proportion in Sulawesi and Maluku. From
2009-2020, 237 C. goffiniana were illegally traded and confiscated from various sources
(Figure 31). Meanwhile, based on data recorded in CITES, during the same years, there were
no legal exports of this bird from Indonesia.
Ecological Characteristics of C. goffiniana
5.1.1 C. goffiniana Habitat and Avifauna in Tanimbar Islands
The Tanimbar Islands is the confirmed distribution of the endemic bird C. goffiniana on
P. Yamdena, P. Selaru, P . Larat, P. Sera, and P. Molu. The distribution on P. Sera and P.
Molu has not been previously reported by Cahyadin et al. (1994) and Roselaar and Michels
(2004) who only recorded C. goffiniana as having a natural distribution in P. Yamdena, P.
Larat, and P. Selaru. These distribution records are also still limited to a few islands, so
several other islands have not been accessed to obtain data on the existence of C. goffiniana
distribution. This limited distribution data is due to limited access t o small islands, so
further research is needed to determine the distribution of this bird on other islands.
32
Classification of land cover in the Tanimbar Islands that became the distribution area
C. goffiniana produced six habitat classes: forest, wetland, developed land, plantation, open
land and mangrove. This classification is simple, but represents all habitat types that provide
resources for birds in Tanimbar Islands, making it easier and more implementable to develop
bird conservation management plans. Various bird species including C. goffiniana, which
have a wide range of ranges, use a variety of habitat types, and often visually the boundaries
between land cover classes cannot be clearly distinguished. This is in contrast to the
classification made by the Ministry of Forestry in 2001 (Jaya 2014), which divides in detail
into 23 land cover categories. Among them, forest land cover is further divided into seven
classes, namely primary lowland dryland forest, secondary lowland dryland forest, swamp
forest, production forest, and so on. The same is true for open land cover which is detailed
into classes of shrubs, savannah, agricultural land, paddy fields, ponds, airports, and
transmigration land. Based on its utilization, there are four types of forest in Tanimbar
Islands, namely protected forest, limited production forest, convertible production forest, and
nature reserve forest (West Southeast Maluku District Government 2012), while based on
canopy cover it is divided into semi-seasonal forest (characterized by stratified canopy and
young trees) and seasonal forest with discontinuous canopy with tall trees over 30 m
(Cahyadin et al. 1994).
The diversity of habitat types in Kep. Tanimbar is one of the factors supporting the
richness of biodiversity, one of which is avifauna. Based on the results of this study and data
from Bishop and Brickle (1998), the total avifauna recorded from P. Yamdena, P. Larat, P.
Selaru, P. Lutu, P. Molu, and P. Kirimoen, and P. Vaimar in Tanimbar Islands totaled 142
species consisting of 16 orders and 54 families (Appendix 1). Limited resources and
accessibility are challenges in revealing species data and discovering new bird species, so it
has not been able to record the entire avifauna in the Tanimbar Islands. Observations made on
P. Yamdena in this study were also very limited in the east and represents the few forests that
dominate the island, while in the west only a few locations were observed, namely Makatian
and Batu Putih villages, which have mangrove forests and coastal areas.
The land cover classifications used by C. goffiniana are forest, open land, and
plantations which are also habitats for other avifauna, the number of bird species recorded in
each habitat type is 65 species, 55 species, and 22 species. These three habitat types were
originally forests, where people cleared land for gardening, but during the dry season they
will be abandoned and become open areas in the form of shrubs. Rosantika (2006) explained
that the degradation rate was 9,473 ha/year in P. Yamdena between 1998-2008 caused by
forest clearing for conversion to agricultural land, fields, and open land. The shifting
agriculture system led to the expansion of land that provided food sources such as corn and
beans for all bird species including C. goffiniana. Some agricultural land extends into the
forest, reducing the main habitat in the forest for foraging, nesting and roosting.
These three land cover classes provide various resources needed by C. goffiniana and
its avifauna for survival and breeding such as food sources, nests, reproductive expression,
social behavior, and others. The total forest area in Tanimbar Islands, reaching 240,661 ha or
about 54% of the total area of Tanimbar Islands, is the main habitat for C. goffiniana and its
avifauna.
C. goffiniana and other avifauna. This habitat provides these key resources and cannot be
replaced by other habitat types, even if these resources are found in open fields or plantations.
Loss of forest habitat will disrupt the survival of C. goffiniana and other birds. Diverse
vegetation types provide a variety of food sources, ensuring year-round availability. Birds are
able to breed well and safely when they can choose tree species, materials, placement and
other nest criteria. Open land and plantation habitats are highly vulnerable to disturbance and
threats to the survival of C. goffiniana. Food sources in both habitat types are highly
dependent on the season and are mainly food crops grown by the community. It is also
vulnerable to hunting and trapping, which can reduce its population in the wild, and the
limited availability of resources for nest building and construction can lead to suboptimal
breeding. Therefore, forest habitat must be maintained both in quality and quantity.
The C. goffiniana bird does not use built-up land, mangrove, and wetland classes with a
total area of about 10% of the entire Tanimbar Islands. In contrast to Jepson et al. (2001) who
reported that
C. goffiniana uses these habitats. These different results may be due to differences in the
location of the study with the previous one that used the transect walking method from the
eastern to the western part of P. Yamdena. In this section, it is possible that the mangrove
habitat is adjacent to the forest in the wildlife reserve so that it is possible to find C.
goffiniana activity in the mangrove forest.
These birds did not show any direct interactions at both space and time scales with
avifauna using these three land cover classes. The number of avifauna using wetlands was 51
species, mangroves 19 species, and six species using built-up land. The total area of wetlands
and mangroves of about 36,892.88 ha (8.32 % of the total area of Tanimbar Islands)
consisting of water, river, swamp and coastal areas was not used by C. goffiniana due to the
incompatibility of resources available in both habitat types. There are no records of potential
34
food sources or nesting trees in the class. Food sources available in these habitats include
fish, molluscs, crustaceans, and other aquatic animals that have never been recorded as being
eaten by C. goffiniana. In addition, the main vegetation is the Rhizophoraceae tribe, which
has no potential as food and becomes a nesting tree, which is an attraction for C. goffiniana to
use these two types. The same is true for the built-up land cover class, which mainly consists
of settlements, ports, airports, and other infrastructure that do not provide the needs of this
bird.
Based on its biogeographical location and geologic history, Tanimbar Islands is one of
the important centers of bird endemicity in the Wallacea region. The study recorded 26
Indonesian endemic species with limited distribution in the Wallacea region, 10 endemic
species with distribution only in Tanimbar Islands, and 15 subspecies endemics. The
Tanimbar Kep. endemic species is a species whose distribution is only in Indonesia (endemic
to Indonesia) but has a distribution in several other islands with different character variations
from one another. At the beginning of the discovery of the species, descriptions were made
based on differences in morphological characteristics and limited island distribution but still
within one species. Species in Tanimbar Islands can also be found in Nusa Tenggara, Maluku
or Papua as different species. A total of three species share the same distribution in the Lesser
Sundas, four species share the same distribution in Maluku, and six species share the same
distribution i n t he Lesser Sundas and Maluku. Some species even share the same
distribution in Papua and Australia. Birdlife International has recognized Tanimbar Islands as
a center of bird endemism. The archipelago is included in the Banda Sea Islands, which are
designated as Endemic Bird Area EBA 165 (Bishop and Brickle 1998).
The endemicity of species and subspecies in Tanimbar Islands shows the similarity of
characters from the Lesser Sundas (Nusa Tenggara) and Maluku, so despite the different
geological history with Nusa Tenggara, especially Solor, Alor, Wetar and Banda which are
part of the inner Banda arc, Coates and Bishop (1997) included birds in Tanimbar Islands as
part of the Nusa Tenggara subregion even though administratively it is part of Maluku
Province. This is due to the similarities in climate, topography, geography and
geomorphology between the Tanimbar Islands and the Lesser Sundas, which are drier, arid
and have low vegetation density. Different groupings were made by White and Bruce (1986),
Eaton et al. (2016) that Tanimbar and Kai Islands are part of the South Maluku subregion.
This is also consistent with the geologic history that Tanimbar Islands originated as part of
the The outer Banda arc, together with Timor, Kai, Buru and Seram, share a common
geology with the southern Moluccas.
The development of science and technology with molecular approaches has obtained a
variety of new data and findings to evaluate the naming and taxonomic status of several
species and their subspecies. Therefore, this study also updated the species list using the new
name system. Myzomela sanguinolenta annabellae has had its species name changed to
Myzomela boiei (Banda Myzomela) due to the separation of names for different distributions.
Myzomela sanguinolenta (Scarlet Myzomela) is used for Myzomela species whose
distribution is in eastern Australia (Higgins et al. 2020). A name change due to distribution
separation also occurred for Pachycephala pectoralis fuscoflava (Golden Whistler) which has
been changed to Pachycephala macrorhyncha fuscoflava (Yellow-throated Whistler), because
Pachycephala pectoralis is used for species with a distribution in eastern Australia, while
Pachycephala macrorhyncha is used for species with a distribution in Indonesia (Boles et al.
2020). Ficedula dumetoria riedeli (Rufous-chested Flycatcher) has changed its status from a
subspecies to a separate species, Ficedula riedeli (Tanimbar Flycatcher), making it endemic
to Tanimbar Islands (del Hoyo et al. 2020). Morphological differences are evident in the
white eyebrows of F. riedeli, which are not present in F. dumetoria.
The genus name change occurred for Monarcha mundus (Black-bibbed Monarch)
which changed to Symposiachrus mundus which has a distribution in Tanimbar Islands
(Clement 2020). These two genera were previously in the same genus in the Monarchidae
family, but were eventually separated into different genus. The Symposiachrus genus consists
of 21 species that are generally dominated by gray-black and white plumage colors, while
Monarcha consists of eight species that are generally gray-black and brown. A name change
also occurred for C. goffini to C. goffiniana after redescription. The identification of the
previous specimen RMNH 87994 from Rotterdam Zoo as C. goffini or Lophochroa goffini or
a synonym of C. tanimberensis was a misidentification, after re-identification the specimen is
C. ducorpsii which has a distribution in Solomon. Therefore, using specimens RMNH 90750
and RMNH 90751 from the Tanimbar Islands, they are described as
C. goffiniana (Roselaar and Michel 2004).
The geological history of the Tanimbar Islands, which formed about one million years
ago and remained unconnected to the Sahul shelf during the Pleistocene (Charlton et al.
1991; Voris 2000), makes the Tanimbar Islands an interesting biogeographic region for
research other than birds. Tanimbar Kep served as a stepping stone for several groups of taxa
in their dispersal from widespread Papua to Sulawesi and the Greater Sunda. Jønsson et al.
(2010) used the Tanimbar Islands as a geological calibration point for phylogenetic analysis
of songbirds in the split between Pachycephala macrorhynca and Pachycephala orioloides at
A
36
0.8 ± 0.2 million years ago. Several decades have seen the discovery of other endemic species
such as Lepidoptera insects (Buchsbaum et al. 2014), beetle insects, and beetles long-snouted
Coleoptera (Narakusumo et al. 2019), and Weijola and Kraus (2023) who discovered a new
species of Varanus tanimbar so that there are six endemic reptile species in Tanimbar
Islands.
Tanimbar Islands is also part of the East Asian / Australian flyway migration route from
the northern hemisphere and Australia. The presence of wetland habitat types is very limited
in Tanimbar Islands. The presence of migratory birds indicates that Tanimbar Islands still has
a suitable habitat type as a stopover or migration destination. Bird migration behavior from
cold to tropical regions is another interesting topic for research, such as migration paths,
home ranges, mechanisms, periods, and various other things. Migratory birds will cross
national borders so that countries that are part of the route will coordinate and cooperate to
preserve migratory birds in the face of population threats. Migratory birds in Tanimbar
Islands are also threatened by habitat change and infrastructure development. Galbraith et al.
(2014) generally identified several main threats to the existence of migratory birds, namely
land use change, illegal hunting, disease, pollutants, and infrastructure development that
causes habitat loss and human disturbance.
The high diversity of bird species has various important roles for the ecosystem in the
kep. Tanimbar. Birds as part of the food chain also play an ecological role in seed dispersal,
pollination, pest control and maintaining ecosystem balance. Based on the type of food, birds
can be grouped into frugivores, insectivores, granivores, nectarivores, omnivores, fishivores
and predators. Seed and fruit-eating birds will play a role in seed dispersal in maintaining
forest vegetation regeneration, insectivorous birds will maintain insect populations and
prevent the development of pest populations that can cause losses to farmers. Predators will
maintain the population of snakes and rats that can also potentially disturb humans. The high
species diversity and endemicity of birds found only in Tanimbar Islands also provide
interesting ecotourism potential and can be developed for sustainable use. Therefore, the
preservation of bird populations and their existence in natural habitats must be of concern to
all parties so that they do not become extinct. The extinction of species, especially endemic
birds such as C. goffiniana, will lead to the extinction of species on this earth because its
natural population distribution is only limited to Tanimbar Islands.
5.1.2 Interactions of C. goffiniana with Avifauna Communities in Tanimbar Islands Forest and
open land are the most common habitat types.
used by birds in carrying out their various life cycles. In addition to these two land cover
classes, C. goffiniana also uses plantation habitat types. The similarity in the use of habitat
types, food types, nest types, breeding periods will lead to interactions of sharing and
competing for resources needed by these various birds.C. goffiniana birds are generalists and
opportunists in obtaining food sources, so that a diversity of sources is obtained high feed.
The 27 food species found in this study are fewer than the results of Mioduszewska et al.
(2018) and O'Hara et al. (2018) who identified 32 species of food for C. goffiniana and 35
potential food species. These forage species are found in forest and open land, especially in
agricultural land. A total of 17 food species in the forest habitat type vary in availability
depending on the season, including throughout the year, or during the dry season. These food
species are not available in large quantities in one tree, so that these types of trees are usually
only visited by a small number of C. goffiniana. In contrast to the 10 food species in open
fields, which are generally food crops grown by the community in large quantities and
simultaneously such as corn, red beans, green beans, long beans, and peanuts. This makes
them a favorite food source and a target for birds to visit during the harvest season. Birds will
visit these food sources in large groups.
This is not the case with other food species in open fields such as coconut, papaya,
cassava, banana and cashew nuts. In these species, variations occur in irregular distribution,
not simultaneous planting, different harvest ages, and not abundant numbers. These
conditions meant that C. goffiniana did not arrive in large numbers, and often only one was
found. An interesting behavior encountered when foraging in this open field is extractive
feeding. Birds take food that is in the soil or eat fruits or seeds that are covered by a thick
skin or layer of fruit such as cassava, peanuts, papaya seeds, coconut or nuts (Huber and
O'Hara 2016).
This study observed the interaction in the utilization of food objects between C.
goffiniana and E. roratus in sharing banana fruit food sources during the dry season with
limited quantities and food sources in the dry season.
P. Yamdena. Both species share the same banana fruit bunches at different times. Limited
food resources during the dry season also cause birds to search for food resources alone, in
pairs or small groups. The two parrot species also share similar breeding and nesting patterns,
so competition for nest trees was observed.
Canarium spp. trees are the most common tree species used by C. goffiniana to build
nests. The same tree species are also used as nest trees by C. alba in P. Halmahera (Rosyadi
et al. 2019). The selected tree has a larger diameter and height than the surrounding trees.
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Research by O'Hara et al. (2018) and Mioduszewska et al. (2018) reported that the breeding
period is estimated to start between June and early July or between December and February
(Jepson et al. 2001). Observations of C. alba in P. Halmahera and P. Bacan (North Maluku)
recorded its breeding season starting in mid-October (Rosyadi et al. 2019). Limited
availability of nest cavities may lead to competition for nests, especially when two or more
species have overlapping breeding periods (Rosyadi et al. 2019).
To maintain the trees that have been controlled, C. Goffiniana will make loud noises
and try to chase away or attack other species that are approaching the tree, especially E.
roratus. E. roratus also does the same thing when it finds a tree to nest in. These parrots also
use tree holes for their nests. However, there is a slight difference in the position of the nest
hole between the two species. C. goffiniana uses a tree hole on the former branch break so
that the door of the nest hole will look slightly prominent, while E. roratus uses a hole
located on the main trunk of the nest tree. Large trees are generally located in forests, so the
availability of potential nest trees that meet the criteria can still be maintained. Although C.
goffiniana is well adapted to using nests that do not meet the criteria, these conditions are
prone to predator threats or human disturbance to take the chicks. Large and tall nest trees are
not available on P. Selaru, forcing C. goffiniana to adapt to using small and short nest trees.
Mioduszewska et al. (2018) found several animals that threaten C. goffiniana as adults,
juveniles, chicks or eggs, including eagles, jungle cats, monitor lizards and snakes.
Different interactions occurred between C. goffiniana and A. crassa and E. reticulata,
which shared the use of the same nest tree to build their respective nests. This interaction
occurs in walnut trees in the forest habitat of P. Yamdena. The three bird species have
different patterns and placement of nests on different sides. The type of nest made by A.
crassa is different from the other two species, which use cavities or holes in the tree. This
species makes nests from dry twigs and other materials that are shaped like a bowl between
tree branches in a position between the nests of E. reticulata and C. goffiniana. However, at
the top of the same tree, C. goffiniana also did not repel D. concinna, which used the tree for
roosting or foraging. It is possible that A. crassa, E. reticulata, and D. concinna are not
considered threats or competitors to C. goffiniana.
The availability of tree resources for roosting is also an object of sharing and
competition between C. goffiniana and other communities. Competition for roosting trees
occurred between C. goffiniana and Corvus orru in milkwood (Alstonia scholaris) trees in the
forest habitat of P. Selaru. Similar to the shooing behavior and loud shouts to E. roratus, this
behavior was applied to C. orru that flew over and approached trees that had been used as
roosts by C. goffiniana. These parrots were treated the same as in the walnut trees of P.
Yamdena with D. concinna, which did not engage in expulsion when this species roosted or
foraged in these A. scholaris trees.
Nest tree conservation is an effort that can be done to support the C. goffiniana
conservation program, namely population recovery and increase. The threat of biodiversity
loss occurs mainly due to changes in land use, the introduction of invasive foreign species,
damage to ecosystems and habitats, and excessive exploitation of biological resources,
including flora and fauna (Widjaja et al. 2014).
5.2 Genetic Characteristics of C. goffiniana
The structure and composition of the mitogenome of C. goffiniana from P. Yamdena
are similar to the results of Kim et al. (2021) who analyzed C. goffiniana, C. galerita, and C.
alba obtained from Seoul Zoo Korea. The structure of mitochondrial DNA in C. goffiniana
and Cacatuidae is different from other aves such as chickens, Picidae, Cuculidae,
Passeriformes and Falconiformes. The gene sequence in the mitochondrial genome of
vertebrates (other than chicken) is ND5, ND6, tRNAglu , Cyt-b, tRNA , tRNAThrPro and D-
loop, while the chicken mitochondrial genome has the gene sequence ND5, Cyt-b, tRNAThr ,
tRNAPro , ND6, tRNAglu and D-loop. The gene arrangement in Picidae, Cuculidae,
Passeriformes and Falconiformes becomes ND5, Cyt-b, tRNAThr , D-loop, tRNAPro , ND6
and tRNAglu (Mindell et al. 1998). The mitochondrial gene structure is relatively more
different in humans, which is a difference of two genes encoding tRNA, as in Anderson et al.
(1981) which explains that the gene structure in the human mitochondrial genome consists of
two parts, (a) the coding region (coding region) consists of 37 genes, namely 13 genes
encoding proteins, two genes encoding rRNA and 22 genes encoding tRNA, (b) non coding
region (non coding region) is the control region (control region) which plays a role in the
process of transcription and replication of the mitochondrial genome Mitochondrial DNA is a
double thread DNA which is generally circular. Mitochondrial DNA consists of heavy strand
DNA and light strand DNA. The heavy strand consists of two rRNA encoding genes (16 S
and 12 S), 12 protein encoding genes (NADH dehydrogenase subunits 1, 2, 3, 4, 5, and 4L;
Cytochrome Oxidase I, II, and III; Cytchrome-b, ATPase 6 and ATPase 8) and 14 tRNA
encoding genes. The animal mtDNA light thread consists of one protein-coding gene (NADH
dehydrogenase subunit 6 or ND6) and eight tRNA-coding genes namely tRNA Glutamic
Acid, Proline, Serine, Tyrosine, Cystein, Asparagines, Alanine and Glutamine.
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Differences in the structure of the mitochondrial genome in aves including the
Cacatuidae family and with other vertebrates are thought to be due to changes due to the
addition and subtraction of genes during mtDNA replication, resulting in a new arrangement
of genes Quinn and Wilson (1993), one of the differences in the sequence of protein encoding
genes, especially ND5 and D-loop (Kvist (2000). Mitochondrial structure correlates with
environmental variations (Cheviron and Brumfield 2012; Ribeiro et al. 2011), differences in
mitochondrial haplotypes occur due to different adaptations to environmental conditions
(Pavlova et al. 2013).Environmental variations that occur are caused by changes in the
landscape. Landscape is a heterogeneous area formed by the interaction of various types of
ecosystems. Landscape change occurs due to disturbances in the structure and function of the
landscape. One of the changes that occur includes deforestation which causes problems in
conservation, namely habitat loss, fragmentation and species isolation (Prasetyo 2017).
Genetic diversity is a characteristic of individuals and populations that describes the
variation of alleles and genotypes in the genome and affects population characteristics such
as potential extinction risk and evolution. A decrease in genetic variation can reduce the
ability of a population to adapt to environmental changes, thereby reducing the ability of
individuals for long-term survival (Arif and Khan 2009). This is often the case in very small
populations that have the potential to increase the likelihood of extinction Inbreeding
ultimately leads to a decline in the superior quality of a population (Crnokrak and Roff 1999).
Inbreeding is known to decrease genetic diversity and reduce reproduction and survival rates,
leading to an increased risk of extinction. Wildlife populations with lower genetic diversity
are at greater risk of extinction (Saccheri et al. 1998). This is not currently the case for C.
goffiniana populations in Tanimbar Islands. The analysis showed high genetic diversity with
a Haplotype diversity (Hd) of 0.9464 and low nucleotide diversity (π)= 0.0017. There was
variation at 44 sites, forming 27 haplotypes.
5.3 Conservation Implementation of C. goffiniana
Birds have become one of the favorite pets because of their beautiful colors,
vocalizations, and intelligence, including Psittaciformes (parrots), Phoenicopteriformes
(flamingos), Accipitriformes (eagles), Passeriformes (songbirds) (Gilardi, 2006).
Psittaciformes are the most popular pet birds on all continents due to their colorful plumage,
capacity to learn and vocalize human voices, and tolerance of living outside their habitat
(White et al. 2011; VKM et al. 2020).
The stipulation of these two statuses has the consequence of stricter regulation and
licensing by the government for its utilization. The designation of C. goffiniana in the CITES
Appendix I category also affects exports made by Indonesia, because CITES makes and
applies strict provisions for plants and wildlife included in the Appendix I list in utilization
for international trade. Article II paragraph 1 of the CITES provisions prohibits international
trade in specimens of this type unless the purpose of import is not commercial, for example
for scientific research. In article VII, exceptions for trade can be granted, namely that an
import permit and export permit have been obtained and the species is captive-bred (CITES
2018).
Based on this data, it can be seen in two perspectives. First, this number is a potential
utilization market that can be managed properly to bring economic benefits to Indonesia.
Trade is conducted to meet market demand to utilize wildlife as food sources, high-value
collections, show attractions, medicines, and pets (Nijman 2010; Harris et al. 2015). The
international illegal wildlife trade generates billions of dollars annually (Wilson-Wilde 2010).
As well as the potential of the global bird trade, Indonesia has at least 300 traded bird species,
22% of households have pet birds, and the bird trade and associated businesses contribute
nearly $78.8 million to the national economy annually (Jepson and Ladle 2009, Jepson et al.
2011, Shepherd et al. 2004). Therefore, it is necessary to manage C. goffiniana properly in
order to be able to provide market needs in a way that is in accordance with applicable legal
provisions. In accordance with applicable regulations such as PP 8 of 1999 and its derivative
regulations, the utilization of protected species is the result of captive breeding of the second
generation and so on. The second point of view is that this trade is a threat to the
sustainability of populations in the wild. The amount traded at that time was only a small
amount from captive breeding (0.032%), so it is possible that most of it came from direct
collection from the wild. The wildlife trade has caused population declines of several species
in Asia, South America and Africa (Alves et al. 2013; van Balen et al. 2013; Shepherd et al.
2013; Martin 2018), and contributed to the spread of alien species outside their natural
habitats (García-Díaz et al. 2017).
CONCLUSIONS :
Tanimbar Islands as a strategic area in the Wallacea region is also one of the centers of
bird endemicity and high avifauna diversity and recorded as many as 142 species. C.
goffiniana is one of the endemic birds in Tanimbar Islands that is opportunistic and generalist
so that it has a very varied list of food sources due to the limited availability of food sources
that depend on the season in forest habitats and agricultural land. C. goffiniana has a
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distribution in Tanimbar Islands, especially P. Yamdena, P. Larat. P. Selaru, P. Sera, and P.
Molu, however observations at other island locations still allow new distribution records that
were previously unreported. C. goffiniana using forest, open land and plantation habitat types
will interact with other bird communities by sharing and competing, especially for food and
nests.
The most commonly used nest tree is the large and tall walnut tree, birds have the
ability to adapt to using small trees in open land such as the limited resources available on P.
Selaru,. This adaptability is an indicator that genetic diversity is still high. Birds will have a
high ability t o adapt to environmental changes and the ability to survive in the long term and
prevent extinction. There was no relationship between the influence of habitat, especially
land cover used by C. goffiniana, on its genetic diversity, so the population did not show
separation or grouping based on the islands in Tanimbar Islands.
The high genetic diversity indicates that the populations of P. Yamdena, P. Selaru, and
P. Larat were formed randomly by mixed breeding between various population origins. This
occurs because the separation distance between P. Yamdena and P. Larat and P. Selaru is not
too far and is still within their range. Genetic analysis can be used to identify species and
trace the origin of confiscated C. goffiniana birds from law enforcement activities, as well as
estimating the bird's range. The range of C. goffinaina using the genetic analysis approach is
estimated to be around 154.51 km. Various activities that can threaten the sustainability of C.
goffianiana still occur in Tanimbar Islands such as hunting, illegal trade, forest conversion,
and illegal logging which causes the loss of potential trees for nests.
