a conservation assessment of stingless bees (apidae: meliponini) in singapore

A Conservation Assessment of Stingless Bees (Apidae: Meliponini) in Singapore

Chui Shao Xiong

A thesis submitted to the Department of Biological Science National University of Singapore in partial fulfilment for the Degree of Bachelor of Science with Honours in Life Sciences

Cohort AY2014/2015 S1

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Acknowledgements This journey has involved detours, roadblocks, and a few dead ends, but like many of those who have made it before me, somehow we made to the finish line, a little worse for wear, but a little more knowledgeable, and hopefully a little wiser.

I have benefited in varying amounts from the following people, all of which have my thanks:

Asst. Prof. John S. Ascher, my supervisor, for the supervision that I greatly appreciate as a step into a wider world.

Ms. Samantha Lai from NParks, for processing and coordinating the approval of permit [NParks Permit number: NP/RP14-082], thereby allowing the study to be carried out.

Ms. Abigail Huang from the National Archives of Singapore, for her patience in facilitating the retrieval of the many historical maps that I requested for.

Ms. Heidi Lee from the Singapore Land Authority, for granting permission to use these historical maps as reference material for this study.

Mr. Randolph Quek Z. B., for the mutualism we shared from the intersection of separate studies on Fishtail Palms and the stingless bees that visit them.

To Rafi, Meng Hwee, Randolph, Wei Qiang, Ken Wei, Leonard, Weng Ngai, Joshua, Kelly, Eunice, Kiayi, and Sihao, because getting to the end of the final year required endurance for the first two, enjoyment of the third, and resolution for the last, all of which required healthy doses of friendship, which you all have provided in one way or another.

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Contents Acknowledgements ........................................................................................................................ i

Contents ........................................................................................................................................ ii

List of appendices .................................................................................................................... iii

Abstract ........................................................................................................................................ iv

1. Introduction .............................................................................................................................. 1

2. Methods .................................................................................................................................... 7

2.1 Present sampling of extant stingless bee species ............................................................... 7

2.2 Documentation of spatial land use changes ....................................................................... 9

2.3 Geometric morphometrics ................................................................................................ 10

3. Results & Discussion ............................................................................................................... 13 3.1 Sampling of extant species ................................................................................................ 13

3.2 Recorded species distribution ........................................................................................... 16

3.3 Implications of biology on conservation ........................................................................... 22

3.4 Morphometrics ................................................................................................................. 26

3.4.1 Traditional morphometrics ........................................................................................ 26

3.4.2 Geometric Morphometrics ........................................................................................ 29

4. Conclusion ............................................................................................................................... 37

4.1 Future directions ............................................................................................................... 37

4.2 Concluding remarks .......................................................................................................... 39

5. References .............................................................................................................................. 41

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List of appendices

Appendix I Historical maps of Singapore Appendix II Nests of stingless bees

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Abstract Stingless bees (Apidae: Meliponini) are important understorey pollinators of

lowland dipterocarp forests of Southeast Asia. The impact of continued regional forest

loss on these forest dependent, highly eusocial bees is not known. Singapore can be

used as a model to assess the impact of past deforestation on species loss, through

comparison of extant and historically recorded species. Of 11 species historically

recorded, 7 were recorded in Singapore during the present study, mirroring extinction

rates for vertebrates but not for other bees. Reduction of nest sites, as evidenced by the

paucity of larger trees in secondary forests, is seen as a potentially important driver of

species loss in Singapore. Species differed in their ability to adapt to the urban

environments. For example, among Tetragonula only T. laeviceps was commonly found

in urban parks, whereas Tetragonula geissleri and Tetragonula pagdeniformis were

restricted to larger forest fragments. Geometric morphometrics (quantitative analysis of

shape variation) of the forewing and hind leg of stingless bees provided an objective

method of shape comparisons between species taxa. It clearly separated all genera, but

not three morphologically similar species Tetragonula species (T. geissleri, T. laeviceps,

and T. pagdeniformis).

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1. Introduction Conservation assessment of stingless bees in Singapore

From the second half of the twentieth century, an alarming trend of sustained

deforestation rates in Southeast Asia of 0.67% per year in the 1990s and 0.59% per year

in the 2000s has resulted in forest loss of 32 Mha, or 6.5% of the regions land area in

two decades (Stibig et al., 2014). The main drivers of deforestation include conversion

to cash crops, rubber plantations, and increasingly oil palm plantations. There is urgent

need to understand how this deforestation will affect species diversity (particularly that

of invertebrates) (Lawton et al., 1998; Whitmore & Sayer, 1992), ecosystem services,

and the healthy functioning of an ecosystem. A baseline conservation assessment is

crucial to inform future monitoring and eventual habitat conservation.

Deforestation in Singapore is not a recent occurrence, as by the mid-1800s

lowland dipterocarp-dominated rain forests in Singapore had largely been cleared for

gambier and pepper plantations, leaving only small patches of original forest on a few

hill tops by 1854 (ODempsey, 2014; Wallace, 1869). Primary forest fragments now

comprise just 0.2% of total land area, and secondary forests an additional 4%. With

relatively mature secondary forests of mostly 50-80 years of age as the major extant

forest type within Singapore, understanding how animals make use of these forests as

compared to primary forest that are much more complex, both structurally and

floristically, will be key to establishing their conservation status (Chua et al., 2013;

Corlett, 1992). Singapore, as a model for the effects of tropical deforestation on species

diversity, can inform discussion of the future repercussions of continued forest loss in

the region and globally. Brook et al. (2003) estimated catastrophic extinctions following

deforestation in Singapore by comparing observed and inferred extinction rates across

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many taxa. Extending this approach to other invertebrate groups, such as pollinators,

will provide cross-taxa comparison and greater insights as to the effects of deforestation

on fauna.

The closed understorey of lowland dipterocarp forests is windless, so pollination

services must come from animals, with eusocial corbiculate bees (Apis, Meliponini) a

major contributor through their sheer abundance (Corlett, 2004). The effects of

deforestation on bees and the resulting effects on pollination cannot be fully appreciated

without a comparison of baseline inventories between historical and current species.

Bee species richness in the tropical forests of Southeast Asia is known to be

poor in comparison to the high plant species richness present (Roubik, 1990). Roubik

postulated that each highly eusocial species, supreme generalists, could occupy the role

of several solitary species in an ecological hierarchy of food resource usage. While

honey bees (Apis) have a low diversity (ca. 10 spp.) with a native paleotropics range

spanning Europe to their maximum diversity in Southeast Asia, stingless bees

(Meliponini) have a relatively high diversity of (ca. 500 spp.) distributed pantropically

(Ascher & Pickering, 2015; Charles D. Michener, 2007). However, while studies on

stingless bees are relatively lacking in Southeast Asia when compared to the relatively

prolific research from the Neotropics (Michener, 2007), research has been on the rise in

the region (Liow, 2001; Sakagami et al., 1990; Salim et al., 2012).

The most recent comprehensive general survey of bees in the forest understorey

of Singapore was conducted by Liow (2001) through baiting with a diluted honey

solution. Prior to this, patchy records of stingless bees (Cockerell, 1918; Sakagami,

1959, 1978; Smith, 1857) were probably netted at flowers, in rain forest dominated by

tropical dipterocarps, a habitat where scarce floral resources is the norm, punctuated

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only by infrequent (2-6 yrs) dipterocarp masting events (Sakai, 2002). Soh & Ngiam,

(2013) surveyed bees and wasps in parks within Singapore through netting at flowers,

but the majority of sites were too open and contained too low quality forests to sustain a

variety of stingless bees. As for solitary leaf-cutter bees (Megachile) in Singapore, of

which 10 new species records were added to the total of 21 species for Singapore

(Ascher et al., accepted), a comprehensive species inventory had yet to be consolidated

for the highly eusocial stingless bees as of 2013.

Application of the International Union for Conservation of Nature (IUCN) Red

List of Threatened Species, in the conservation assessment of stingless bees in

Singapore, is not presently possible as the IUCN has yet to evaluate Meliponini, and

had only just recently made a first review of European bee species (no Meliponini).

Fortunately, most, if not all Singapore species, even locally extirpated ones, are

widespread in Peninsular Malaysia as well as within the Southeast Asian region, and so

are not of a high concern globally.

Taxonomy

The genus Tetragonula (often treated as a subgenus of Trigona, sometimes as

part of subgenus Heterotrigona), a Southeast Asian group, includes Tetragonula

laeviceps sensu lato, a common species complex in the region. The neotype of T.

laeviceps was designated from a worker which originates from Singapore, after the

holotype was deemed to have been lost (Rasmussen & Michener, 2010). Notoriously

variable with overlapping morphological characters, Tetragonula species offer few

reliable characters for species identification, with species descriptions often based

primarily on a combination of size and subtle differences in hair pattern and colour. The

colour of various appendages varies within populations and is subject to interpretation

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(Sakagami, 1978), and variability of diagnostic characters has limited the utility of

taxonomic keys for species determination. In practice, study of comprehensive

reference collections including type material or specimens compared with types is often

necessary to consistently identify regional Meliponini, but these have not been available

in Singapore.

Advances in molecular systematics have enabled longer stretches of DNA to be

sequenced and analysed, potentially allowing for reliable species delimitation and

identifications and also increasingly accurate reconstructions of the molecular

phylogeny of stingless bees to be made (Rasmussen & Cameron, 2007). [However, not

all named forms are included in BOLD and other online databases, and many publicly

available sequences lack identifications or these are unreliable.]

Complementarity between varying lines of evidence, such as traditional

morphology, geometric morphometrics, and genetics, should inform decisions about

species delimitation and identification (Schlick-Steiner et al., 2010; Schwarzfeld &

Sperling, 2014). Exploring applications from the growing field of geometric

morphometrics, on the usage of shape variation of certain morphological characters for

species diagnosis, is one aim of the current study.

Traditional morphometrics, as practised by Sakagami (1978) in his measurement

of various parts of Tetragonula bees, involved the comparison of length measurements

of various characters as a means to diagnose species. While Sakagami found allometric

trends for some characters, the underlying flaw of the method is its inability to localise

morphological differences (Zelditch et al., 2012), which limits its utility for species

diagnosis.

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Geometric morphometrics offers an alternative interpretation of morphological

characters, focusing on quantification of shape rather than qualitative description as

found in traditional taxonomic works, or use of character lengths used as in traditional

morphometrics (Zelditch et al., 2012). There are two main approaches to geometric

morphometrics: 1) landmark-based method; 2) outline-method. The landmark-based

method compares a series of homologous characters between specimens to quantify

shape variation, whereas the outline method compares the outline of a morphological

character between specimens to quantify shape variation. Whereas the landmark-based

method has been used to quantify shape variation in the forewings of stingless bees in a

few studies (Combey et al., 2013; Vijayakumar & Jayaraj, 2013), there is potential for

the outline-based method to characterise shape variation for morphological characters

with few homologous points (Dujardin et al., 2014).

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Aims

This study investigates how deforestation has affected species diversity of a key

invertebrate group, stingless bees, in Singapore, within the context of potential loss of

invertebrate diversity and ecological interactions should present deforestation rates be

maintained in Southeast Asia. To this end, this study seeks to:

1) Determine stingless bee species loss in Singapore through comparison of extant

species collected during present sampling, and historically recorded species.

2) Examine land use changes in Singapore through historical maps, to correlate spatial

patterns of deforestation and recorded species distribution, in order to infer the

effects of deforestation on species loss.

3) Improve tools for species diagnosis through integrative taxonomy, particularly

among Tetragonula species present in Singapore:

a) explore the usage of geometric morphometrics, on selected characters (e.g.

forewing and hind leg), as an additional means to complement existing

species diagnosis.

b) examine success rates of species diagnosis between traditional

morphological-based taxonomy and geometric morphometrics.

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Table 1. Field collection sites within the Central Catchment Nature Reserves.

1MacRitchie Reservoir > Based on Yee & Tan (2010)

2. Methods 2.1 Present sampling of extant stingless

bee species

2.1.1 Sampling localities

The 8 sites selected (Fig. 1, Table 1) for

bait sampling were within the Central

Catchment Nature Reserves, the largest

continuous patch of remnant forest in

Singapore. Sites were selected from non-

convoluted, separate trails of at least 1km,

designated as official trails by the National

Parks Board (NParks), but sites 7 and 8

were contiguous. Bukit Timah Nature

Reserve (BTNR) was closed for restoration works during the course of this study.

no. Site Forest Type> Latitude Longitude 1 MCR1 (Lornie Trail) Mixed primary/secondary forest 1.34289 103.82710 2 MCR1 (MacRitchie Nature Trail) Mixed primary/secondary forest 1.35279 103.82545 3 MCR1 (Sime Track) Old secondary forest 1.35627 103.81422 4 MCR1 (Rifle Range Link) Old secondary forest 1.35292 103.80667 5 Nee Soon pipeline Freshwater swamp forest 1.38577 103.81120 6 Upper Peirce Reservoir (Pump

Station) Young secondary forest 1.37316 103.80425 7 Dairy Farm Nature Park Old secondary forest 1.36433 103.77625 8 Dairy Farm (Siap Rd) Old secondary forest 1.36065 103.76969

Figure 1. Field collection sites within the Central Catchment Nature Reserves. Refer to Table 1. for site details.

1 km

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2.1.2 Sampling method for stingless bees

Two main sampling methods are efficient for stingless bees: opportunistic

netting from flowers, and spray baiting with a sugar/honey solution on vegetation (Liow,

2001; Salim et al., 2012; Salmah et al., 1990). In particular, Liow found that the

spraying of a diluted honey solution on vegetation attracted a disproportionate number

of eusocial honey bees and stingless bees as compared to solitary bees, rendering this

method particularly suited for targeted sampling of stingless bees. Alternative methods

for sampling employed by Liow included funnel traps, bowl traps with a floating

platform, all of which have had comparatively low capture rates, while banana pulp

attracted no bees. Simple plastic bowl traps (filled with soapy water), which have been

used to great success to attract a variety of bees (non-Meliponini) in other more open

habitats such as meadows and arid areas, failed to attract bees in a trial run by Soh

(2014) (Ascher, pers. comm.).

2.1.3 Spray baiting of honey solution on vegetation

For sampling surveys of stingless bees, a modified sampling protocol using baits

was adapted from that of Liow (2001) and Salmah et al., (1990). Honey solution (1:4,

honey:water) with added salt (4 cm3/L), was used to attract stingless bees. Each site had

a 1 km transect consisting of 10 bait stations, spaced 100 m apart. The honey solution

was sprayed on vegetation in a 1 m radius of the sampling point to a height of 1.5 m at

each station along the transect.

A trial run compared sampling after pre-baiting the day before with sampling

without pre-baiting, and it was observed that both methods yielded stingless bees in

similar abundance at stations (pers. obs.). Sampling without pre-baiting was decided

upon due to the frequent, unpredictable inter-monsoonal rains that descended upon the

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island during the sampling period (September to December 2014), which often rendered

pre-baiting unviable. Sampling took place two hours after initial spraying of bait, and a

maximum of ten individuals of each stingless bee morphospecies were collected within

ten minutes from each station.

2.1.4 Netting from flowers

Another source of samples was opportunistic netting of stingless bees from

flowers found along the transect at each site, mainly from two plant species which

dominate patches of shrubland interspersed within the forest (Dillenia suffruticosa and

Melastoma malabathricum)(Corlett, 1997). In particular, 14 bee species were recorded

on M. malabathricum in Singapore (Chong, 2010). In a collaborative effort with R. Z.

B. Quek, stingless bees captured from the male inflorescence of fish tail palms (Caryota

mitis), the subject of his study (Quek, 2015), were also included in this study.

2.1.5 Specimens

Collected specimens were pinned and deposited in the Insect Diversity

laboratory (IDL) reference collection (National University of Singapore) for study.

Vouchers specimens will be deposited in the Lee Kong Chian Natural History Museum

(LKCNHM).

2.2 Documentation of spatial land use changes

To document land use changes in Singapore at varying points of its history,

historical topographic maps were obtained from the National Archives of Singapore

with approval granted by the Singapore Land Authority. Maps covering 1898, 1924, and

1937 were used (see Appendix I for details). A3-sized colour photocopies of the maps

were obtained, which were scanned at 600 dpi with a commercial scanner. The scanned

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maps were then geo-referenced in ArcGIS ver. 9.2 with existing coordinates on the

maps, as well as through manual alignment using landmarks. Shape-polygon layers

were traced for primary and secondary forests present in each map. These layers were

then compiled to reveal spatial changes of primary and secondary forest over time. A

present day map of parks/forests in Singapore was sourced from the Urban

Redevelopment Authority (URA) 2014 Master Plan Parks and Waterbodies Plan

(Singapore. Urban Redevelopment Authority, 2014).

2.3 Geometric morphometrics

2.3.1 Imaging of Stingless bee specimens

A Canon 6D digital camera, with an attached MP-E 65mm 1-5x f2.8 lens, was

used to capture images. The camera was mounted on a copy stand setup (Visionary

Digital Imaging Passport II system). Individual images were stacked to form a focused

composite image with great depth of field (Zerene Stacker Build T201412212230, 21

Dec 2014).

The right forewing and hind leg were removed from a subset of specimens and

prepared in the same manner before imaging using the aforementioned setup, after

which analytical methods differed.

2.3.2 Preparation for imaging

To standardize the plane orientation of forewing and hind leg with reference to

the camera axis, both forewings and hind legs were mounted on glass microscope slides,

held in place in glycerol gelatin (Kaisers glycerin jelly) and sandwiched between slide

and cover slip. The use of an electric cup warmer was useful to keep the glycerol gelatin

liquid during specimen manipulation.

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The hind legs of stingless bees required further preparation before mounting as

the corbicula (pollen basket) of the tibia was often coated with resin and pollen, which

if not cleaned off, made clear images of the hind tibia difficult. Soaking the legs in a

dishwashing liquid solution prior to sonication at 40kHz in a diluted glass cleaner

solution was sufficient to remove any debris (Harrison, 2012).

2.3.3 Geometric morphometric analyses

2.3.3.1 Landmark-based method

Landmarks were selected from homologous points on the forewing across all

species, usually situated at the junction of veins (Fig. 2). These landmarks were plotted

as an overlay on each wing image using tpsDig ver. 2.17 (Rohlf, 2013). The landmarks

were then aligned as a consensus shape using the Procrustes method in tpsRelw ver.

1.54 (Rohlf, 2014), from which principal component analysis (PCA) of wing shape

variation was obtained.

Figure 2. Landmark-based method. Forewing with 9 landmarks placed on homologous points.

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2.3.3.2 Outline-based method

Due to the hirsute nature of the hind legs of various species, trial automation of

hind leg shape tracing with Adobe Photoshop CS5 did not produce accurate tibia or

basitarsus shape. Instead, manual tracing of the hind leg was carried out with the same

software. The traces were put through the SHAPE ver. 1.3 (Iwata & Ukai, 2002) suite of

programmes for harmonics (closed outlines described with algorithms) generation (Fig.

3) of elliptical fourier descriptors, for use in elliptic fourier analysis of tibia and

basitarsus shape variation, from which principal component scores (PCS) were obtained.

The PCS were in turn used for PCA.

2.3.4 Data analyses

Canonical Variates Analysis (CVA) was performed on principal components generated

from PCA using PAST ver. 3.05 (Hammer et al., 2001).

K-means testing was performed on principal components generated from PCA using

CLUSTER ver. 3.0 (Eisen & de Hoon, 2002).

Figure 3. Outline-based method. Consecutive fitting of increasingly complex closed outlines, harmonics, is used to quantify shape.

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3. Results & Discussion 3.1 Sampling of extant species

The present study recorded 7 extant species of stingless bees in Singapore (Fig.

4), namely: Heterotrigona itama, Lepidotrigona terminata, Tetrigona apicalis,

Tetragonula fuscobalteata, Tetragonula geissleri, Tetragonula laeviceps, and

Tetragonula pagdeniformis. During the present study, Lepidotrigona terminata was first

found on a Nipa Palm (Nypa fruticans) inflorescence at Chek Jawa, Pulau Ubin in late

March 2015, too late for inclusion in morphometric analyses.

The localities of each stingless bee species, and the methods used to collect the

data, are presented in Figure 5. Honey-baiting and netting was conducted primarily

within the Central Catchment Nature Reserve (CCNR), with netting conducted at the

a b c

d e f

g 5mm

Figure 4. Workers of stingless bee species collected from present study. a: Heterotrigona itama; b: Lepidotrigona terminata; c: Tetrigona apicalis; d: Tetragonula fuscobalteata; e: Tetragonula geissleri; f: Tetragonula laeviceps; g: Tetragonula pagdeniformis.

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Figure 5. Sampling localities in Singapore for 2014-2015 survey. Central Catchment Nature Reserve in grey.

additional sites of Pulau Ubin, Kent Ridge Park, and the National University of

Singapore. Mangrove Insect Project (MIP) malaise trapping yielded few species, as did

most visual observations. Tetragonula laeviceps was present at most sites. One result is

the fewer number of species between CCNR and more urbanized areas. Two species

had single localities: Tetragonula fuscobalteata from SBG, and Lepidotrigona

terminata from Chek Jawa, Pulau Ubin.

The Singapore Botanic Gardens (SBG) was exceptional in having 4 species

despite their urban location, situated away (ca. 2 km) from CCNR, contrary to

expectations of few Meliponini from parks and gardens, especially those not adjacent to

CCNR. However, it does have a small fragment of remnant primary rainforest (ca. 6 ha),

which could account for the unexpected number of species. One probable reason for the

higher than expected number of species, is the profusion of flowering plants throughout

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the year in the botanic gardens, a rich food resource that could sustain multiple species.

Floral richness was such that while there was interference and exploitative competition

at individual flowers, species were not seen to engage in the exclusion of other species,

as in the case of niche preemption (pers. obs.) (Roubik, 1990).

Sampling was by no means exhaustive for the current study. Different sampling

techniques possibly yielded differing success rates of capturing species present at each

site. The sampling method used at the various sites in the study reflected a utilitarian

approach, where flowers were seen to be the best attractant for stingless bees; if flowers

were not present, baiting was the next best alternative.

Baiting with a diluted honey solution has been proven to be a viable method of

sampling for stingless bees in the forest understorey where flowering is scarce, as the

bees tended to recruit more foragers to the lucrative food source (Salim et al., 2012;

Salmah et al., 1990).

However, while this sampling technique is effective, it does not replace pollen

from flowers as an attractant, the second food resource stingless bees gather. For

instance, T. apicalis was found on Fishtail Palm (Caryota mitis) flowers, but not at baits

placed at the site. Palms in the forest provide a rich pollen source for stingless bees, in

particular the male inflorescences of Fishtail Palm (Caryota mitis) (R. Z. B. Quek,

2015), and should be considered as a good resource for the capture of stingless bees

should the chance arise.

Based on an examination of 61 sampling occurrences, (count of species

observed at each sampling event) the Chao1 estimator (Fig. 6) plateaus, at an estimated

species richness of 7 species for Singapore. Although the species, Lepidotrigona

terminata and Tetragonula fuscobalteata are represented by a singleton and doubleton

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respectively, Chao1 did not estimate higher species richness. One explanation would be

that the total sampling occurrence consisted of few singletons and doubletons, and it is

thus unlikely that additional undetected

species exist. The >5 species estimated

from the Central Catchment Nature

Reserve (CCNR) (Fig. 6) exceeds current

observed estimates, and suggests that

additional bait sampling is required to

conclude if undetected, extinct species

would be rediscovered from within it. In

stark contrast, estimates of undetected

species far exceeded observed estimates

for Megachile (Soh, 2014).

3.2 Recorded species distribution

Forest cover change maps (1898-2014) in Figure 7. Forests include both pristine

and degraded forests, which were not distinguished; Belukar is Malay for secondary

forest, likely to have been dominated by the tree species Adinandra belukar. Species

collected historically, with a specific locality in Singapore: A. R. Wallace 1854 (Bukit

Timah) Geniotrigona thoracica, Homotrigona fimbriata, Tetragonula laeviceps; C. F.

Baker 1911 (Bukit Timah) G. thoracica, Heterotrigona itama, Lophotrigona

canifrons, Tetragonula geissleri, Tetrigona apicalis; also (Changi) T. apicalis;

(Seletar) T. apicalis; (Singapore Botanic Gardens) G. thoracica, T. apicalis; H. C.

Abraham 1922 (Mandai) H. itama, T. geissleri; C. B. Kloss 1926 (Singapore Botanic

Gardens) T. apicalis; T. C. Maa 1958 (Nee Soon) T. laeviceps, G. thoracica;

Kawamichi 1974 (Bukit Timah) T. geissleri, T. laeviceps. Wallace records published

Figure 6. Sampling occurrence based rarefaction curve.

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by Smith (Smith, 1858), Baker and Abraham records by Schwarz (Schwarz, 1939), Maa

and Kawamichi records by Sakagami (Sakagami, 1978).

Historical records of stingless bee diversity available in the literature are patchy

at best, providing inadequate coverage even for the small island of Singapore. No

records exist of their diversity in an assumed pristine habitat before forest clearance in

the early 1800s for gambier plantations. By the time Wallace arrived in 1854, he had to

traverse bare open country to reach remnant patches of primary forest situated at hill

tops (Wallace, 1869). The famous entomologist Charles Fuller Baker (Welles, 1927)

collected from various forest patches scattered around Singapore in 1911, after the

establishment of crown forest reserves in 1889 following recommendations by Cantley

(1883), including the subsequently developed Changi Forest Reserve. Maa and

Kawamichi sampled within modern CCNR after its establishment in 1951 under the

Nature Reserves Ordinance, forerunner of the Park and Trees Act. Local specimens

from the Lee Kong Chian Natural History Museum (LKCNHM) were not examined due

to the closure of the collection during the course of the present study.

Collections made by various workers, probably through opportunistic netting of

bees from flowers, provide a snapshot of species present in each period. An overall

trend that can be observed from Figure 7 is the decline of stingless bee species present

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Figure 8. Similarity of stingless bee species within Sundaland. Srensen Similarity Index (.00-1.00) = (2 x number of shared species) / [(no. of species in area A) + (no. of species in area B)]. Figure adapted from Inger & Voris (2001).

over time, in particular of the larger-bodied non-Tetragonula species such as G.

thoracica, H. fimbriata and L. canifrons and recent general sampling in 2001 of bees by

Liow did not find these three species (Liow, 2001). However, to accurately chart the

decline of individual species, consistent, frequent sampling is required.

Examining the biogeography of the Sunda Shelf region, it is expected that there

is a high similarity of stingless bee species between Peninsular Malaysia (to which

Singapore is a part of biogeographically), Sumatra, and Borneo (Fig. 8).These three

land masses have a lower similarity with Java, and the least similarity with Sulawesi.

All the land masses, with the exception of Sulawesi, were connected with land bridges

during the last ice age during the Pleistocene (ca. 12000yrs ago), allowing for the spread

of shared species. Patterns of species similarity between these land masses are mirrored

in snakes and frogs (Inger & Voris, 2001).

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It is on the basis that Singapore is part of Peninsular Malaysia,

biogeographically, that Brook and colleagues inferred species loss in Singapore based

on what exists in the whole of Peninsular Malaysia (Brook et al., 2003). Adapting from

their calculations of inferred extinction for various fauna, observed (recorded) and

projected (inferred) species loss was calculated for both stingless bees and non-stingless

bees in Singapore (Fig. 9).

Three scenarios, depicting different

geographical assumptions of shared species,

were used for calculation of inferred species

loss. Scenario (1) follows the original

assumption that species present in Peninsular

Malaysia were once to be found in

Singapore. Scenario (2) takes into

consideration states in the lower half of

Peninsular Malaysia, while Scenario (3)

includes only Johor, the nearest Peninsular Malaysian state to Singapore. The inferred

extinction rate for Scenario (1) is not a good representation for the non-stingless bees as

the actual extinction rate is far lower than predicted (data provided by Ascher, J. S.).

Scenario (2) is the most realistic of the three, projecting half of historically recorded

stingless bee species (11 species) to have gone extinct, while only a few non-stingless

bee species are projected to have gone extinct in Singapore. This is expected as species

distributions need not be homogenous between Singapore and Peninsular Malaysia,

assumed by Brook and colleagues from the low species endemism in Singapore

attributed to the narrow Straits of Johor which separates both countries (Brook et al.,

2003). Scenario (3) is extreme, with the inferred extinction for both stingless bees and

Figure 9. Observed and projected stingless bee species diversity loss in Singapore. Mammals (MAM); Stingless bees (STB); Rest of the bees (BEE). Inferred species extinction based on checklists from: Peninsular Malaysia (1); Johor + Melaka + Negeri Sembilan + Pahang (2); Johor (3), arranged in descending distance from Singapore. Figure adapted from Brook et al. (2003.

21

non-stingless bees implausibly below that of observed extinctions, a result attributable

to the probable historical undersampling of Johor, relative to Singapore.

Even with the reasonable assumption that Singapore shared stingless bee species

with the lower half of Peninsular Malaysian states, as in Scenario (2), it is surprising

that Singapore has had a lower than expected number of historically recorded species.

This suggests that some factors may have had an effect on limiting species distribution

in Singapore. One factor may include a reduction of available nest sites in Singapore

through removal of large trees, a process well underway by the time A. R. Wallace

obtained the first local samples.

A representative distribution of stingless bee nests among living trees of varying

dbh (diameter at breast height ~1.3m) in Sabah, sourced from Eltz et al. (2003), with

additional inputs adapted from Chua et al. (2013), is shown in Figure 6. Eltz and

colleagues found that while stingless bee species did not have preferred tree species for

nest sites, 86.1% of nests were found in trees with dbh > 60cm. This seems to

corroborate the tree core decay study by Panzer (1976), which found about half of trees

larger than 60cm dbh tended to develop trunk hollowness, which translates to potential

nest sites for stingless bees.

The study by Chua et al. (2013), conducted on a 2 ha plot each of primary and

secondary forest in Singapore, found a lack of tree representation of dbh > 60cm in the

top five most common trees in the secondary forest plot (Fig. 10). While the sample of a

2 ha plot does not reflect the absolute absence of trees of dbh > 60 cm, it is a generality

reflective of the bulk of such forests in Singapore. It would mean secondary forests are

able to provide relatively few potential nest sites for stingless bees when compared with

primary forests.

22

One nest each of T. laeviceps and H. itama were found in the Singapore Botanic

Gardens during the course of the study (Appendix II). The T. laeviceps nest was found

in an unidentified tree of dbh 107 cm, while H. itama was found nesting in a Tembusu

tree (Fagraea fragrans) of dbh 147 cm (Fig. 10).

Figure 10. Distribution of stingless bee nests among varying sizes (dbh) of living trees in Sabah (sourced from Eltz et al., 2003). Diameters of trees housing one nest each of T. laeviceps and H. itama found locally in the Singapore Botanic Gardens overlayed on tree class sizes. Inverted axis gives a rough representation of tree size distributions of 2 ha primary and secondary forest plots in Singapore (adapted from Chua et al. 2013).

3.3 Implications of biology on conservation

3.3.1 Nest site requirements

With the nest site requirement of trunk hollows, the distribution of many

stingless bee species is limited by the availability of trees of sufficient diameter to

provide large hollows to house hives of the species in question. Moreover, the

correlation between trunk diameter, probability of hollowness, and trunk hollow volume,

is not homogenous among tree families, genera, or even species (Eltz et al., 2003).

23

Compounded with the high heterogeneity of tree species within local lowland

dipterocarp forests, it is difficult to reliably estimate of the availability of nest sites in

the remnant forests in Singapore, and through extrapolation, an estimate of the long

term viability of local populations.

3.3.2 Population replenishment

Natural replenishment of species stocks from Peninsular Malaysia, via crossing

the narrow Johor Straits that separates both countries (ca. 600 m) at its narrowest point,

may be expected for large, strong-flying, non-eusocial bees such as Xylocopa but is a

challenge for smaller, weaker-flying, and highly eusocial stingless bees (Michener,

1979), due to their unique nesting biology. During reproductive swarming, newly

emerged virgin queens (gyne) and a portion of older workers seek out a viable nest site.

The site has to be within flight distance from the existing hive as construction materials

(i.e. propolis) for the new nest has to be brought over, which necessitates constant

shuttling between both nests for an extended period. This limits the extent to which

stingless bees can establish a nest across water bodies broader than the maximum flight

distance of the species, which ranges from 600-2600 m depending on body size (Arajo

et al., 2004). A few nests, sealed from the elements, might possibly survive a water

crossing on floating vegetation, for example, but small founder would then be subjected

to inbreeding or extinction due the lack of viable population sizes (see next section).

3.3.3 Vertebrate-like population size

Only the queens of highly eusocial stingless bees (Meliponini) and honey bees

(Apini) are capable of breeding. However, there are differences in reproduction between

the two tribes. Honey bees and stingless bees have a single dominant reproductive

queen, thereby displaying relatively straightforward vertebrate-like population sizes

24

(Romiguier et al., 2014), although under some conditions nests contain a proportion of

egg-laying workers that, being unfertilised, produce only drones.

3.3.4 Nesting biology

One fundamental difference between stingless bees and honey bees, which could

explain differences in their distributional success in Singapore, is in their life strategy

(Chinh, 2004). Stingless bees take longer to develop from egg to adult than honey bees

(40 days vs 20 days), which allows the latter to respond more quickly to environmental

changes. Compared to honey bees, which have the option of moving en masse to

another site should conditions be unfavourable (Dyer & Seeley, 1994), stingless bees

accumulate relatively large stores of honey in permanent nests to tide them through lean

times.

Comparing fecundity, honey bees swarm up to 10 times yearly, while stingless

bees swarm on average once yearly (Chinh, 2004). In honey bees, the existing, old,

queen leaves the established nest with a substantial portion (ca. half) of the workers,

leaving the nest to be inherited by a new queen (Winston, 1987), but in stingless bees

each new gyne leaves with a far smaller proportion of workers (ca. 10-30% of workers)

and the old queen remains in the original nest (Inoue et al., 1984).

Collectively, these differences characterise honey bees as short-term strategists

that exploit seasonal resource abundance, typified by the correlation between the

migration of the giant honey bee (Apis dorsata) and masting events in dipterocarp

forests of Southeast Asia (Itioka et al., 2001), and stingless bees as long term strategists

that focus more on nest maintenance and resource accumulation. In more general terms,

honey bees are poised to exploit dynamic changes in the environment, while stingless

bees are poised to exploit stable environments (Chinh, 2004).

25

A key difference between the historical lowland rainforests which once

blanketed Singapore, and the remnant forests of the present, lies in the stability of the

environment. Forest fragments are more resource unstable as the effects of a reduction

in nest site availability from past forest loss, as well as food resources, are magnified

over a smaller area.

Singapore presents an interesting scenario in that it may be highly urbanised, but

areas outside the forest patches are heavily planted with an assortment of trees and

shrubs, unlike the sparse greenery which characterises other large cities. The managed

greenery provides year round food resource for bees, whereas flowers are scarce in the

forests in unfavourable seasons. It is a paradox that food resources in urban parks, even

that of Gardens by the Bay (built on reclaimed land), are more stable that what is

available in the forests, which are richer and comparatively more stable in terms of nest

site availability.

Both stingless bees and honey bees are able to exploit urban food resources, but

differ in the extent to which they are able to do so. While both are central place foragers,

returning to the same nest between foraging trips, stingless bee nests are restricted to

forests and as such they are not able to exploit food resources beyond their maximum

flight distance from the nest. Honey bees, with their adaptability to nest in urban

environments, are able to extend their collective foraging range beyond the confines of

forests and into non-forested areas (Ricketts, 2004).

26

3.4 Morphometrics

Species diagnosis using descriptive morphological character traits in taxonomy

functions well when traits are discrete, but ambiguity ensues when traits are continuous

and overlaps occur between species. Such is the case for the genus Tetragonula, where

reliable character traits are few and far between, are often difficult to consistently

observe from pinned specimens, and tend not to stand up to scrutiny (Sakagami, 1978).

Sakagami characterised colour of various parts of the bee for species diagnosis, but an

arbitrary system of six colour gradations ranging from the lightest colour hue (pale

fulvous) to the darkest (brownish black to black), with wide colour ranges for species

complexes such as T. laeviceps, with a tendency of colour range overlap between

closely related species, does not allow for confident species diagnosis.

3.4.1 Traditional morphometrics

Sakagami pointed out that the lack of reliable structural characters in

Tetragonula workers was the main difficulty in species diagnosis (Sakagami, 1978).

Consequently, species diagnosis had to depend on non-structural characters such as size.

Sakagami utilised traditional morphometrics to quantify size differences between

species for use in species diagnosis, which involves linear measurements of various

parts of the body (Fig. 11).

The permutations of the scatterplots of different measurements seem to be

highly correlated as evidenced by the constant slope; larger species will have a

proportionately larger body than smaller species. T. fuscobalteata is distinctly the

smallest species in all scatterplots, but size overlaps occur most frequently between T.

pagdeniformis and T. laeviceps, and to a lesser extent T. apicalis and T. itama.

.

27

While size ranges can be used for species diagnosis to an extent, overlapping

size ranges between some species limits the utility of traditional morphometrics as a

reliable diagnostic tool. Also, size conveys no information on geometric structure of the

species (Zelditch et al., 2012) and cannot be used to provide specific characters

(characters which distinguish one species from another).

To improve on traditional morphometrics, geometric morphometrics quantifies

shape variation, which allows for the objective comparison of shape (a specific

character) between species. The geometric morphometrics toolkit includes quantitative

shape comparison, as well as visual shape variation representation.

Figure 11. Body length (BL); head width (HW); wing length (WL1); length between M/Cu vein junction and base of 1st marginal cell (WL2); hind tibia length (HTL).

28

Figure 12. Relative warps of the wings of 6 species.

Relative wing warps show the direction of shape change between species and

the mean shape (T. laeviceps) through a visual representation of the distortion of a

virtual mesh on which landmarks are placed (Fig. 12). This method can be used to

complement quantitative analysis methods such as principal components analysis,

through an examination of which characters best explain quantitative shape variation

between species, as evidenced by localised distortion of landmarks. This can potentially

lead to the development of reliable character traits for use in species diagnosis (Zelditch

et al., 2012).

Visual inspection of the relative wing warps is instructive too. T. apicalis and T.

fuscobalteata have extremes of body sizes, with T. apicalis the largest and T.

fuscobalteata the smallest of extant species in Singapore. The two landmarks, circled in

red (Fig. 12), highlight opposing distortions of stigma shape between both species. A

29

strong negative distortion of the stigma shape for T. apicalis, and a slight positive

distortion for T. fuscobalteata, suggests that T. apicalis has a disproportionately smaller

stigma relative to T. fuscobalteata. This finding conforms to the allometric trend

suggested by Danforth (1989) that small Hymenoptera tend to have disproportionately

larger stigmas. Danforth postulated that the mass of the stigma aids in balancing and

stabilizing the forewing as a counterweight, which leads to increased flight efficiency.

However, relative stigma size is not a reliable character for species diagnosis, at least

within the Tetragonula in Singapore. The relative wing warps of three species with

overlapping size ranges, T. geissleri, T. laeviceps, and T. pagdeniformis, do not show

any differences in relative stigma size.

3.4.2 Geometric Morphometrics

Exploratory tools such as Principal Component Analysis (PCA) can be used to

summarise the data generated from geometric morphometrics. PCA is used to simplify

the dataset of related shape variables through the combination of likely correlated

variables that account for maximum variance in the data. Used on its own, PCA has

limited usefulness due to lack of critera to test the significance of the species clusters

produced.

However, the generated principal components can be cross validated with the

aforementioned relative wing warps (Fig. 12), to determine if the shape variance from

PCA can help explain biologically significant character differences among species.

3.4.2.1 Principal components analysis (PCA)

PCA was performed on the forewing, forewing veins, hind tibia, and hind

basitarsus, and the first and second principal components displayed in scatterplot graphs

(Fig. 13). The forewing (Fig. 13a), to which PCA is most widely applied within the

30

b a

c d

Figure 13. Comparison of principal component analysis scatterplots. a. forewing; b. forewing veins; c. hind tibia; d. hind basitarsus. Percentage variance explained by each component in brackets.

Hymenoptera (Aytekin et al., 2007; Pretorius, 2005; Villemant et al., 2007), has distinct

clusters for the non-Tetragonula species, H. itama and T. apicalis. T. fuscobalteata and

T. pagdeniformis clusters have some overlap, and T. geissleri and T. laeviceps have a

high overlap in their species clusters. This trend is repeated in a decreasing extent for

the wing veins and hind tibia, while no trend was observed for the hind basitarsus.

31

3.4.2.2 Canonical variates analysis

Canonical variates analysis (CVA) was then performed using the reduced

number of variables produced from PCA. A standardised number (10) of principal

components was used, representing an accumulative >95% variance of the data. CVA

was used to determine how much shape variance can be explained for by species. Like

PCA, CVA is an exploratory tool of shape variance, but CVA can be further tested,

such as through the calculation of the percentage of successful species assignment rates

via the jack-knife procedure. This procedure uses each specimen in turn as the test set

against the rest of the specimens (training set), to determine the percentage success rate

of species assignments (Table 3) over a predetermined number of runs (1000 runs).

CVA was performed on the forewing, forewing veins, hind tibia, hind basitarsus,

as well as on measurements made using traditional morphometrics. The first and second

canonical variates were used to plot scatterplot graphs (Fig. 14). As with the PCA

scatterplots, some general trends can be observed. For the forewing, the non-

Tetragonula species, H. itama and T. apicalis, have distinct clusters while Tetragonula

species clusters have high overlap. As hypothesised, CVA plot for forewing veins had

very high cluster overlap for all species, as the number of homologous landmarks used

(5 points) were few and tightly spaced, which probably led to the inability of the

method to detect small scale shape variation. For both the hind tibia and hind basitarsus,

T. fuscobalteata formed a cluster distinct from other Tetragonula species. This is as

opposed to the close morphological similarity noted by Sakagami between T.

fuscobalteata and T. laeviceps, discounting colour differences (Sakagami, 1978) .

The results of CVA for traditional morphometrics revealed that the first

canonical variate explained a very high percentage (94.17%) of variance in the data,

32

which can be viewed as traditional morphometrics having highly correlated variables.

Most of the variance of size can be explained by species. This supports the assertion

made earlier that traditional morphometrics is limited in its utility for species diagnosis

beyond highly correlated size measures.

3.4.2.3 K-means test

K-means testing was performed in order to test percentage success rate of

species assignments using PCA results. The K-means used in this study randomly

assigned each specimen to one of six a priori determined classes (K=6), to reflect the

number of species tested. The centroid (K-means) of each class was calculated,

followed by the distance of each specimen from the centroid. Each specimen was then

reassigned to the nearest centroid. The process was repeated until no specimens were

reassigned. As with the CVA jack-knife, jack-knifing was done for K-means (1000

runs). The percentage of successful species assignments (Table 3) was calculated from

the consensus of species assignments (Table 2).

A general trend that emerges from examination of K-means species assignments

(Table 2) is the frequent incorrect assignment of specimens belonging to T. geissleri, T.

laeviceps, and to a lesser extent T. pagdeniformis. T. fuscobalteata has the least

incorrect species assignments for Tetragonula, while non-Tetragonula H. itama and T.

apicalis had the least incorrect species assignments overall.

The results of the percentage of successful species assignments were compiled

for both CVA and K-means across both geometric morphometric and traditional

morphology (Table 3). Traditional morphology had the highest successful species

assignments for both CVA and K-means as expected, because of the high correlation

between size and species. Successful assignment rates for CVA were good for the

33

forewing, hind tibia and hind basitarsus, but poor for the wing veins. The same trend is

reflected with K-means assignment rates. K-means assignment rates were lower than

that for CVA as K-means tends to assign equally spaced centroids, and does not deal

well with clusters with high overlaps.

a

c

b

d

e

Figure 14. Comparison of canonical variate analysis scatterplots. a. forewing; b. wing veins; c. hind tibia; d. hind basitarsus. Percentage variance explained by each variate in brackets.

34

Table 2. K-means species assignments. Successful in bold, incorrect underlined.

Species H. itama T. apicalis

T. fuscobalteata

T. geissleri

T. laeviceps

T. pagdeniformis

H. itama 16 0 0 0 0 0 T. apicalis 0 16 0 0 0 0 T. fuscobalteata 0 0 11 0 0 1 T. geissleri 0 0 0 10 31 0 T. laeviceps 0 0 0 2 38 1 T. pagdeniformis 0 0 0 1 8 13


T. fuscobalteata

T. geissleri

T. laeviceps

T. pagdeniformis



T. fuscobalteata

T. geissleri

T. laeviceps

T. pagdeniformis



T. fuscobalteata

T. geissleri

T. laeviceps

T. pagdeniformis



T. fuscobalteata

T. geissleri

T. laeviceps

T. pagdeniformis


Fore

win

g W

ing

vein

s H

ind

tibia

H

ind

basi

tars

us

Trad

ition

al M

orph

omet

rics

35

Table 3. Comparison of successful species assignment rates

CVA jack-knife K-means

Geometric Morphometrics Forewing 88.51% 70.27%

Wing veins 30.41% 51.85% Hind tibia 85.61% 61.36% Hind basitarsus 87.31% 57.04%

Traditional Morphology 91.89% 76.35%

3.4.3 Geometric morphometrics in species diagnosis

Though successful species assignment rates are high for CVA (ca. 85%) across

most characters tested (Table 3), varying degrees of overlap among species clusters

exist within the taxonomically difficult group Tetragonula in PCA (Fig. 13) and CVA

(Fig. 14).

The utility of geometric morphometrics to complement existing morphological-

based species diagnosis tools remains unrealised in its potential within the current study.

As Zelditch and colleagues noted, the aim of using geometric morphometrics in the

quantification of shape variation is not merely to stop at PCA or CVA to show if species

clusters are significantly different, but to critically examine shape variables that

contribute to each component/variate, and relate that with biologically significant shape

variance, in order to discover character traits that can be used for species diagnosis

(Zelditch et al., 2012).

Based on the results, an assessment of the number of extant species cannot be

made with complete confidence through a quantitative shape analysis of workers alone,

for T. geissleri, T. laeviceps, and T. pagdeniformis, which also overlap in size and

colouration. A more comprehensive method of quantitative shape analysis is 3D

36

geometric morphometrics, which allows for complete shape visualisation. However,

miniaturisation of 3D scanning technology for small subjects such as insects is still in

its infancy (Centeno et al., 2011; Nguyen et al., 2014)

It is recommended that other lines of enquiry be included in species diagnosis

Availability of males and of nests, though more difficult to source than workers, would

permit the examination of male genitalia and the comparative study of nest structures

for these species.

37

4. Conclusion 4.1 Future directions

Future directions include the development of managed colonies of stingless bees

in Singapore for research and education investigation the role of stingless bees in

pollination within local forests, and development of sustainable meliponiculture

(stingless bee keeping) to produce honey in Singapore. One group (Pollen Nation)

currently carrying out the rehabilitation of stingless bee nests (of the small-bodied

adaptable T. laeviceps) from managed structures (i.e. lamp posts) to custom-built nest

sites, seeks to develop both meliponiculture and the establishment of nests in an urban

setting such as roof-top gardens where they could pollinate fruits and vegetables.

The present study provides baseline data for feasibility studies as to the eventual

broader establishment of stingless bee species such as H. itama and T. apicalis, with due

consideration of existing populations and re-establishment of Geniotrigona thoracica

[etc.] if follow-up surveys confirm these to be extirpated from Singapore. Before such

implementations by take place, the present study is necessary to document the present

state of Meliponini distributions in Singapore, before stingless bee rehabilitation and

meliponiculture alters current distributional extent, which may obscure natural patterns

and be undesirable if carried out wrongly.

To reverse the likely trend of declining nest site availability in forests, the

effectiveness of artificial nest sites in the form of hollow plastic containers can be

explored via trap-nest studies for stingless bees, to determine if methods used in the

Neotropics can be adapted for use in Singapore (Oliveira et al., 2013). The general nest

site preferences with regards to tree species for stingless bees, together with the ability

38

of species such as T. laeviceps to adaptively nest in artificial structures such as lamp

posts, is evidence that stingless bees can adapt to artificial nest sites of appropriate size.

Three historically recorded species (G. thoracica, H. fimbriata, and L. canifrons)

were not found during this study. Without more extensive sampling, it cannot be

concluded if they have gone locally extinct. If they had, it can only be speculated in

retrospect if loss of these species could have been prevented. Ethical considerations

remain as to whether reintroductions should be carried out, to bolster existing

populations, or reintroduced if these species are already locally extinct.

Most stingless bee species are neither as resilient nor adaptable as T. laeviceps in

urban Singapore. Forest-bound species such as T. geissleri and T. pagdeniformis may

not cope with artificial nests or an urban environment. To better estimate the availability

of nest sites of these forest dependent species in remnant forest fragments, tree species-

specific trunk hollow diameters should be measured, and correlated with stingless bee

occupancy. This will help to clarify whether the composition of dominant tree species in

secondary forests, which differs greatly from the dipterocarp dominated primary forests,

is able to sustain viable populations of diverse stingless bees in the long term.

Population genomics has been applied to bees in Singapore (Chua, 2015; Ng,

2015), and can also be used to examine genomic diversity of stingless bees at a local

level, as well as gene flow between colonies. This is to understand population dynamics

of the Singapore population within the context of a fragmented landscape, which will

inform on the long term survivability of forest dependent species in increasingly

deforested environments within the Southeast Asian region.

39

4.2 Concluding remarks

Of 11 stingless bee species recorded historically, 7 were found during the course

of the present study. Species differed in their ability to adapt to the urban environment.

T. laeviceps was the most adaptable, occurring in forests as well as urban parks, while

congeneric T. geissleri and T. pagdeniformis were restricted to the forests of the Central

Catchment Nature Reserve. These three species are morphologically similar, and while

the exploratory geometric morphometrics used in this study did not succeed in finding

any clear character trait differences between them, its potential remains to objectively

compare morphological characters between species, and develop reliable characters for

species delimitation and diagnosis.

Loss of stingless bee species from Singapore (or their decline to undetectable

levels) can be attributed to historical deforestation, which led to reduced nest site

availability in remnant forests. The nesting biology of stingless bees, in particular their

dependency on an old nest for resources necessary to establish a new nest, limits their

dispersal capability and causes them to be uniquely vulnerable to the effects of

deforestation. The compounded effects of the inability to recolonize, lack of immigrants

from major source areas, and inability to exchange genes between forest fragments, may

lead to population inbreeding and eventual extirpation.

While both solitary bees and highly eusocial honey bees are able to move

hibernate or disperse to avoid unfavourable environments and periods of resource dearth,

stingless bees are adapted to endure with stockpiled food resources. Stingless bee

species loss in Singapore, as a result of past deforestation, may be a microcosm of what

is to come if continued deforestation persists in the rest of Southeast Asia. These forest

understorey pollinators may be unique among bees in their vulnerability to

40

anthropogenic extinction, and the loss of their pollination services may have far-

reaching consequences.

41

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Appendices Appendix I Historical maps of Singapore

Year covered

Year published Title Prepared by Published by

1898 ? Map Of The Island Of Singapore And Its Dependencies Colonial Engineer Office

Surveyor General's Office

1924 1934 Johore and Singapore Geographical Section, General Staff

War Office

1937 1939 Johore and Singapore, 2nd edition Geographical Section, General Staff

War Office

Table I-1: Provenance of historical maps used

Appendix II Nests of Stingless bees

Figure II-1. Nest entrances of stingless bees from the Singapore Botanic Gardens. a: Heterotrigona itama. b: Tetragonula laeviceps as termitophiles in Nasutitermes sp. (Blattodea: Isoptera) nest.

a conservation assessment of stingless bees (apidae: meliponini) in singapore

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