LIFE CYCLE AND OVIPOSITIONAL PREFERENCE OF Bactrocera carambolae

FEEDING ON STARFRUITS

Nur Amirah binti Abdul Latif

Bachelor of Science with Honours

(Animal Resource Science and Management) 2013

Faculty of Resources Science and Technology

Declaration

I hereby declare that the thesis is based on my original work except for the citation

which has been duly acknowledgement. I also declare it has not been previously or

concurrently submitted for any degree for any other degree at UNIMAS or any other

institution of higher learning.

______________________

Nur Amirah Abdul Latif

Animal Resource Science and Management Programme

Department of Zoology

Faculty of Resource Science and Technology

Universiti Malaysia Sarawak

i

ACKNOWLEDGEMENTS

Praise be to Allah S.W.T Most Gracious, Most Beneficent

My deepest appreciation to both of my supervisor Prof. Dr. Sulaiman bin

Hanapi and Miss Ratnawati binti Hazali for their concern, continuous advice and

encouragement throughout the final year project. My gratitude also to my co-

supervisor and his team, Mr. Hj. Sulaiman bin Zulkifli which is an officer from

MARDI Serdang, Selangor, Mr. Hassan and Mr. Khalid for being patience and gave

valuable guidance based on their many years of experienced on fruit flies.

I would like to record my sincere thanks to Dr. Mohamad Roff bin Mohamad

Nor and to all staffs of MARDI (Malaysian Agriculture Research and Development

Institute), and FRST (Faculty of Resource Science and Technology) UNIMAS for

assist me and turn this research into valuable experience.

My utmost appreciation to my beloved family and friends for their

understanding, patience and support throughout the entire project.

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TABLE OF CONTENTS

ACKNOWLEDGEMENTS i

TABLE OF CONTENTS ii

LIST OF TABLES iv

LIST OF FIGURES V

LIST OF ABBREVIATIONS vi

LIST OF APPENDICES vii

ABSTRACT viii

CHAPTER 1 INTRODUCTION

1.1 Background of Study 1

1.2 Objectives 5

CHAPTER 2 LITERATURE REVIEWS

2.1 Family Oxalidaceae. 6

2.2 Maturity Index of Starfruit. 8

2.3 Taxonomic Classification of Bactrocera carambolae. 10

2.4 Bactrocera carambolae Distribution. 10

2.5 The Characteristics and Life Cycle of Bactrocera. 11

2.5.1 Ovipositor of Female Bactrocera carambolae. 12

2.6 Sampling of Fruit Flies. 12

2.7 Hatchability Percentage. 12

iii

CHAPTER 3 MATERIALS AND METHODS

3.1 Study Site. 13

3.2 Sampling of Fruit Flies. 13

3.3 Fruit Flies Identification. 14

3.4 Laboratory Hygiene. 15

3.5 Fruit Fly Rearing. 15

3.6 Life Cycle Experiment. 15

3.6.1 Egg Collection. 16

3.6.2 Hatchability Percentage (HBT%). 16

3.6.3 Inoculation Technique. 16

3.6.4 Larvae Collection. 17

3.6.5 Puparia Collection. 17

3.6.6 Adults Collection. 17

3.6.7 Preserving Insects. 18

3.7 Ovipositional Preference Experiment. 18

CHAPTER 4 RESULTS

4.1 Life Cycle of Bactrocera carambolae Feeding on Starfruits. 20

4.2 Susceptibility of Different Maturity Index of Starfruit to B.

carambolae Infestation.

25

CHAPTER 5 DISCUSSIONS

5.1 Life Cycle of Bactrocera carambolae Feeding on Starfruits. 27

5.2 Susceptibility of Different Maturity Index of Starfruit to B.

carambolae Infestation.

35

CHAPTER 6 CONCLUSION AND RECOMMENDATIONS 39

REFERENCES 41

APPENDICES 48

iv

LIST OF TABLES

Table 2.1 The colour indicator for seven maturity stages of starfruits.

8

Table 4.1 The hatchability percentage (%) of B.

carambolae species.

20

Table 4.2 Life cycle of B. carambolae on it host, carambola fruits in laboratory condition.

21

Table 4.3 Number of percentage (%) survival during life cycle of B. carambolae (Drew & Hancock).

23

Table 4.4 Ovipositional preference of B.

carambolae species on different maturity stages of starfruit.

25

v

LIST OF FIGURES

Figure 2.1 The life cycle of starfruit. Adapted from Sapii and Muda (2011).

7

Figure 2.2 Maturity index of starfruit. Adapted from Sapii and Muda (2011).

9

Figure 4.1 The life cycle of B. carambolae species observed during this study.

22

Figure 4.2 Instar stages of B. carambolae. A: First instar, B: Mouth hook features of first instar, C: Second instar, D: Mouth hook features of second instar, E: Third instar, F: Mouth hook features of third instar.

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Figure 4.3 The number of mean infestation on 10 fruits for each maturity stage in relation with three oviposition site.

26

vi

LIST OF ABBREVIATIONS

mm Millimetre

cm Centimetre

HBT% Hatchability percentage

g Gram

ml Millilitre

h Hours

˚C Degree Celsius

ha Hectare

NaCIO Sodium hypochlorite

TKPM “Taman Kekal Pengeluaran Makanan”

µl Microlitre

% Percentage

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LIST OF APPENDICES

Appendix 1

Samples collection.

48

Appendix 2

Steps involved in life study.

49

Appendix 3

Life cycle of B. carambolae.

51

Appendix 4

Data collected for the study of B. carambolae life cycle.

60

Appendix 5

Ovipositional preference of B. carambolae on starfruits.

64

Appendix 6

Collected data for the study of ovipositional preferences of B. carambolae species on different maturity index of starfruit.

68

Appendix 7

Preliminary study of B. papayae development using ripe Chok Anan mangoes.

70

Appendix 8

Data collected for the study of B. papayae life cycle.

74

viii

Life Cycle and Ovipositional Preference of Bactrocera carambolae

Feeding on Starfruits.

Nur Amirah binti Abdul Latif

Animal Resource Science and Management Programme Faculty of Resource Science and Technology

Universiti Malaysia Sarawak

ABSTRACT

Malaysia is the main starfruit exporter for Hong Kong, Europe and Singapore market. Hence, starfruit contributes financially to Malaysian economics. But due to the presence of its enemies, Bactrocera

carambolae, this species have created havoc in horticulture industries which lead to financial loss. This study focuses more on life history of B. carambolae that are need to be determine before any management control takes place. Generally, life cycle of fruit fly indicated six development stages: mating, oviposition, egg, larvae, pupae and adults while the larval have three development stages (1st, 2nd and 3rd instar). Techniques used in life cycle study involve: fruit fly sampling, fruit fly rearing, egg collection, inoculation, hatchability percentage, disinfestation, pupa collection and the emergence of adults. Eggs of B. carambolae took about 2 days to develop into larva. Larval stage took 6 days to complete and develop into pupa. Then, teneral adults were emerge (26 days) from pupa and will undergo 2-3 weeks of pre-oviposition period before they were fully matured. Regarding experiment 2, investigations were carried out to identify susceptibility of different maturity stage of starfruit to adult females of B. carambolae. B. carambolae species did not infest Index 1 of starfruit while Index 2 was slightly attacked. Fruits from Index 3, Index 4 and Index 5 showed significantly higher infestations than starfruit of Index 2. Among ripe starfruits, Index 5 was more attractive to B. carambolae than fruits that from Index 6 and Index 7. As starfruits ripens, it become more susceptible to B. carambolae attacked.

Keywords: Starfruits, life cycle, Bactrocera carambolae, ovipositional preference.

ABSTRAK

Malaysia ialah pengeksport utama belimbing untuk pasaran Hong Kong, Eropah dan Singapura.

Oleh itu, buah belimbing menyumbang dari segi kewangan kepada ekonomi Malaysia. Tetapi

disebabkan kehadiran serangga perosak, Bactrocera carambolae, spesis ini telah mewujudkan huru-

hara dalam industri hortikultur yang menjurus kepada kerugian kewangan. Kajian ini lebih

memfokuskan kepada kitaran hidup B. carambolae yang perlu di ambil kira sebelum mana-mana

kawalan pengurusan mengambil tempat. Pada umumnya, kitaran hidup lalat buah mengandungi

enam peringkat: mengawan, pengoviposian, telur, larva, pupa dan lalat dewasa di mana larva

mempunyai tiga peringkat tumbesaran (1, 2, dan 3 peringkat ulat). Teknik-teknik yang digunakan

dalam kajian kitaran hidup lalat melibatkan: pensampelan lalat buah, pembiakan lalat buah,

pungutan telur, penginokulatan, peratusan menetas, mengeluarkan ulat, koleksi kepompong dan

kemunculan lalat dewasa. Telur B. carambolae mengambil masa sekitar 2 hari untuk bertukar

menjadi larva. Peringkat larva mengambil masa 6 hari sebelum menjadi kepompong. Kemudian,

lalat-lalat dewasa yang kurang matang muncul dari kepompong dan akan menjalani 2-3 minggu

tempoh pra pengoviposian sebelum mereka matang sepenuhnya. Mengenai eksperimen ke-2, siasatan

dijalankan bagi mengenalpasti kerentanan peringkat kematangan berbeza buah belimbing untuk

betina dewasa B. carambolae. B. carambolae spesis tidak mengerumuni Skala 1 belimbing manakala

Skala 2 sedikit diserang. Buah dari Skala 3, Skala 4 dan Skala 5 menunjukkan serangan lalat betina

dewasa lebih tinggi daripada buah belimbing Skala 2. Antara buah belimbing yang masak, Skala 5

lebih menarik kepada B. carambolae daripada buah yang daripada Skala 6 dan Skala 7. Apabila

buah belimbing masak, ia menjadi lebih rentan kepada B. carambolae yang menyerang.

Kata kunci: Buah belimbing, kitaran hidup, Bactrocera carambolae, keutamaan pengoviposian.

1

CHAPTER 1

INTRODUCTION

1.1 Background of Study

The family Tephritidae or commonly known as ‘true’ fruit flies by White and

Elson-Harris (1992), are classified in the order of Diptera. White and Elson-Harris

(1992) stated that, family Tephritidae consists of approximately 4 000 species

worldwide. Endemically, 75 described species of tropical fruit flies were found in

Southeast Asia including the small number of international significance polyphagous

pests which are Bactrocera carambolae, B. papayae, B. philippinensis and B.

dorsalis sensu stricto (Clarke et al., 2005).

According to Chua (1998), approximately 100 species of B. dorsalis complex

were found in Malaysia and only half of them have been recorded. The species

include the carambola fruit fly, B. carambolae which had been widely infesting more

than 151 kinds of fruits and vegetables including starfruit, mango, cashew, lemon,

grapefruit, guava, mandarin, tomato, jackfruit, orange, avocado, sugar palm,

breadfruit and sapodilla (Vijayasegaran, 1984). Therefore, it is included as major

2

pest in agriculture throughout Asia and the Pacific which financial loss is significant

(Chua et al., 2010).

Fruit flies create havoc in horticulture industries because they cause

tremendous economic loss to fruit and vegetable growers all over the world. The

hosts that are preferable to fruit flies larval include a broad array of fruits, vegetables,

flower heads and seeds (Kapoor, 1993). Due to their polyphagous nature, these

tropical species tend to increase their host range, thereby creating problems in their

control.

Nishida (1963) and Meats (1981), believed that temperature has a bearing on

their emergence number. Whereas Drew et al. (1983), relates the variety of available

food with the higher emergence number of fruit flies in cultivated or wild plants.

Based on their observation, adult fruit flies feed on glandular secretions of plants,

plant sap exudates, pollen, honey dew secretions of homopteran insects, nectar,

decaying fruit, bird droppings, leaf and fruit surface bacteria. Recently, bacteria are

proven to be a well-known diet for adult fruit flies and these bacteria provide food

for fruit fly larvae which will then cause serious damage to the fruit and the fruit

could not be eaten (Drew et al., 1983; Courtice & Drew, 1984; Lauzon, 2003; Behar

et al., 2009; Ansari et al., 2012). Due to this reason, the fruits are not marketable to

large and profitable country, for example, United State of America (USA) and Japan

(Ariff et al., 2011).

This pest can ruin all the fruits in the area if not controlled and the damage

will reach 100%. Fruit with puncture mark will fall faster and become rotten. Bagged

3

fruit can also be damaged by the sting of fruit flies. The sting can penetrate baggage

especially when the wrapping paper is wet and attached to the surface of the fruit

(Rahman et al., 1992). The larvae hatch from the eggs and dig into the fruit and

making it unsuitable to be eaten. Other physical methods include the use of netted

structure which is less convenience for large orchards because it is hard to maintain

(Appendix 1). In addition, the use of pesticides in the spray coverage is not

encouraged as it will kill the honey bees and other beneficial organisms. An effective

way to control fruit fly is by doing it biologically and, therefore, the understanding

on life history of the species is important.

This study was conducted using tropical popular fruit that is the starfruit from

the family Oxalidaceae. Strafruits were already being included in the National

Agriculture Policy 3 (NAP3) among the other 13 major fruits (Ariff et al., 2011).

Malaysia is the main exporter in starfruit production and demand for tropical fruit are

increasing either in Malaysia or abroad (Ariff et al., 2011). The fruit not only

famous in the domestic market, in fact, it has been exported to other countries such

as Hong Kong, Singapore, European countries and Middle East countries. There is

no accurate data on the production and supply of starfruit but based on population

growth and per capita consumption of 0.69 kg/year, the estimated demand and

supply of fresh starfruit for the country is increasing (Ariff et al., 2011).

The specific statistics on fresh starfruit is hardly found because the fruit is

included in the group of other tropical fruit in statistical reports before 2010 (Ariff et

al., 2011). However, demand in the major markets of Netherlands, Hong Kong and

Singapore have increases. At present, due to high demand in starfruit production, a

4

total of 50 to 100 tons of fresh starfruits were exported to other countries every

month by air (Ariff et al., 2011).

According to Kapoor (1993), although fruit flies are commonly thought of as

pests, some species are valuable agent for the biological control of weeds. Most

species that have been used or tested for biological control belongs to the subfamily

Tephritinae and attack plants of the family Asteraceae (Kapoor, 1993). Chaetorellia,

Urophora and Terellia are some tephritid genera that used as biological control

agents (Griffiths, 1999). In Western North America, Urophora stylata has reduced

seed production in Cirsium vulgare (Savi) Tenore, and U. affinis and U.

quadrifasciata have reduced seed production in Centaurea diffusa Lam. and C.

maculosa Lamarck (White & Clement, 1987).

Several intensive studies have been conducted on the taxonomic and genetic

variation of the adults and larvae in Malaysia since 1986 (Elson-Harris, 1988; Ooi,

1988; Vijayasegaran & Mohd, 1991). However, the studies that focus on growth and

development of B. carambolae on Averrhoa carambola L. are still lacking and there

is no previous study based on their ovipositional preference on carambola fruit. It is

important to understand the growth and development aspects of the insects in

predicting its development, emergence, distribution and abundance in the field.

5

1.2 Objectives

The aims of this study are:

1) To study the development stages of Bactrocera carambolae using starfruits

as a hosts.

2) To identify the most susceptible maturity stage and oviposition site of

starfruits that is prone to B. carambolae attack.

6

CHAPTER 2

LITERATURE REVIEWS

2.1 Family Oxalidaceae

Starfruit or scientifically known as Averrhoae carambolae L. from the family

Oxalidaceae, derived from clusters of islands in Southeast Asia. It is the most

preferable hosts of B. carambolae rather than Syzygium species (Wee & Tan, 2000).

Starfruit trees are not seasonal and harvest can be done four to six times per year

(Sapii & Muda, 2011). Starfruit can be categorized into three varieties which are B2,

B10, and B17. In this experiment, B10 variety was used and observed for the study

of life cycle and ovipositional preference of B. carambolae species. B10 variety was

chosen because it was the most popular clones and being cultivated on a large scale

(Rahman et al., 1992). B10 varieties were originally come from Serdang Baru,

Selangor and it was registered in 1968 with weight range from 180 g to 250 g per

fruit (Sapii & Muda, 2011). Sometimes, their size can be more than 15 cm long. In

addition, when it is ripe, they appeared to be orange or reddish in color. Their flesh is

juicy and sweet, with brittle texture and slightly fibrous with aromatic smell. Today,

B10 variety is well-known for its high yield production.

7

Production of

young shoots

Fruits 10-30 mm

long

Fruit less

than 10

mm long

Fruit more than

60 mm long

Fruits 30-60 mm

long

Fertilization

Flower bud

development

Flower bud

expansion

Beginning of

flower bud

The maturity

state of main

branches

60-90 days

2-3 days

7-10 days

1 week

2 weeks

1 week

2 weeks

2 weeks

Figure 2.1: The life cycle of starfruit. Adapted from Sapii and Muda (2011).

8

2.2 Maturity Index of Starfruit

Sapii and Muda (2011) mentioned that, the colour of carambola fruit was

formed by certain maturity level that can be used as an index of maturity (Figure

2.2). Skin colours of carambola fruit were classified into seven categories as

indicated in Table 2.1:

Table 2.1: The colour indicator for seven maturity stages of starfruits.

Stages Fruit coloration

Index 1: Overall in dark green color

Index 2: Fruit is light green and shiny

Index 3: Fruit much more greener than yellow

Index 4: 50% of the fruit is green and another 50% is yellow

Index 5: Fruit much more yellowish than green

Index 6: Overall in yellow color

Index 7: Whole fruit is yellow-orange

9

Figure 2.2: Maturity index of starfruit. Adapted from Sapii and Muda (2011).

10

2.3 Taxonomic Classification of Bactrocera carambolae

B. carambolae was classified as new species in 1994 by Drew and Hancock

(Drew & Hancock, 1994). Carambolae or carambola came from a Latin word which

gives a meaning of its major host, starfruit (Averrhoa carambola). The taxonomic of

B. carambolae were classified by Drew and Hancock (1994) as below:

2.4 Bactrocera carambolae Distribution

In worldwide, the family Tephritidae or known as ‘true’ fruit flies, under the

order Diptera consists of around 4 000 species (White & Elson-Harris, 1992).

According to Clarke et al. (2005), the species of B. dorsalis complex from the family

Tephritidae contains 75 described species and they are largely endemic to Southeast

Asia. In Malaysia, as stated by Chua et al. (2010), there are approximately 100

Bactrocera species of which only half of them had been recorded and one of the

species is B. carambolae (Chua, 1998; Chua et al., 2010). In all scientific research

stated that B. carambolae are the most critical pest in agricultural production,

especially fruits (Vijayasegaran & Mohd, 1991; White & Elson-Harris, 1992;

Yesmin & Clyde, 2012). B. carambolae species had been recorded infesting 151

Kingdom: Animalia

Phylum: Arthropoda

Class: Insecta

Order: Diptera

Family: Tephritidae

Genus: Bactrocera

Species: carambolae

Common name: Carambola fruit fly

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kinds of fruits and vegetables and are widely distributed in Malaysia, Singapore,

Indonesia, Thailand and Brunei.

2.5 The Characteristics and Life Cycle of Bactrocera

According to Yesmin and Clyde (2012), B. carambolae, B. papayae and B.

dorsalis species are considered as sibling species. Hence, they conclude that these

species shared the same growth and development. As described by White and Elson-

Harris (1992), B. carambolae species have clear wings, dark T-shaped pattern and

rectangular dark bands on its abdomen and spot on female’s foreleg. This species

differs from B. papayae by having broad medial longitudinal black band on

abdominal terga lll-V, a broader coastal band apically, and shorter male aculeus and

female ovipositor (White & Elson-Harris, 1992). They undergo four development

stages which are egg, larva, pupa and adult (Yesmin & Clyde, 2012). Based on

previous study conducted by Mohd Noor et al. (2011), the adult B. papayae laid eggs

in one to two days to produce larvae that feed on their respective host causing

premature fruit fall. The larva grows in size by three larval stages: first instar, second

instar, and third instar. Based on previous research conducted on B. papayae, these

three stages being differentiated based on their body size by measuring the length

and width of the larvae (White & Elson-Harris, 1992; Pena et al., 1998; Mohd Noor

et al., 2011). When fully grown, the larva escapes from the fruit, burrows a few

centimeters into the soil and its skin thickens and hardens to form a puparium, in

which the larva will soon transforms itself into the adult. After 10 to 14 days, the

adult fly emerges from the puparium and digs its way out from the soil (Mohd Noor

et al., 2011).

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2.5.1 Ovipositor of Female Bactrocera carambolae

There are no records on female adult B. carambolae ovipositor but as

described by White and Elson-Harris (1992) on its sibling species, female adult B.

papayae, this species has long needle-like ovipositor which is an egg-laying tube at

the tip of abdomen to puncturing the fruit and pushes bacteria from the skin into the

flesh. These bacteria cause fruit decay, which results as the source of food for the

larvae (Drew et al., 1983).

2.6 Sampling of Fruit Flies

Steiner’s trap is used to monitor the population of Bactrocera species in the

field or orchard (White & Elson-Harris, 1992). Mostly it is baited with insecticides

and lure (methyl eugenol) in ratio of 1:3 (Clarke et al., 2001). Most people tend to

modified it according to their purpose. The advantage is, in a short period of time,

Steiner’s trap are capable to catch a large number of flies since it has large open area

at each end which helps in wider distribution of attractant vapour (Kapoor, 1993).

Next, plastic McPhail traps is used to sample an adult fruit flies and it is baited with

food to attract the adult fruit fly (Marques da Silva, 2008).

2.7 Hatchability Percentage (HBT%)

Hatchability percentage or HBT% referred to percentage of survival of the

eggs. According to Ekesi and Mohamed (2011), hatchability is done to give an

estimation number of individual that will hatch from the eggs in the given

temperature and humidity before the actual experiment starts. Usually HBT is done

for fruit fly rearing or for the study of the life cycle.

13

CHAPTER 3

MATERIALS AND METHODS

3.1 Study Site

For the study of life cycle of Bactrocera carambolae, the experiment were

conducted under laboratory at Horticulture laboratory 4, MARDI while the sampling

for B. carambolae species where conducted at 3 ha of ‘Ladang Tanaman Belimbing

(LTB)’ at MARDI, Serdang, Selangor. In addition, fruits were collected at 150 acres

of ‘Taman Kekal Pengeluaran Makanan (TKPM)’ at Lanchang, Pahang. MARDI

was chosen because B. carambolae species need to bring for further identification

because this species have almost the same characteristics with its siblings, B.

papayae and B. dorsalis (Smith et al., 2003).

3.2 Sampling of Fruit Flies

Monitoring for the abundance of B. carambolae species in the field was

carried out using modified Steiner’s trap (Appendix 1) containing a lure, usually 7.5

ml of methyl eugenol with 2.5 ml of 0.1% malathion. Whereas, the sampling of adult

fruit flies were carried out by using plastic McPhail’s trap containing 10% of starfruit

juice and Steiner’s trap by using cotton wick containing lure. The specimen were

14

collected and the lure being replaced every three days. The traps were installed under

the branch of the tree and the locations were randomly picked. Marques da Silva

(2008) described that the number of traps per unit area can vary according to project

objectives. In this project, five traps were put in 1 ha area and the traps are placed

equidistant with each other. The samples were collected at ‘Ladang Tanaman

Belimbing (LTP)’, MARDI Serdang, Selangor under the supervision of Mr. Hj.

Sulaiman Zulkifli, MARDI officer.

3.3 Fruit Flies Identification

Not only B. carambolae species infesting carambola fruit but also B.

papayae, and B. dorsalis, its sibling species (Smith et al., 2003). They are

polyphagous pests which preferred the same host. Thus, the probability for them to

be caught in the traps is high. Prabhakar et al. (2012), presented the actual

photographs of North Western Himalaya of India fruit flies collected during 2009

until 2010 which included 13 species under two genera of fruit flies namely

Bactrocera and Dacus of subfamily Dacinae. White and Elson-Harris (1992)

provided useful information for the identification key of fruit flies with additional

information regarding on fruit fly larvae. After the first stage of identification, the

specimens were brought to Entomology Laboratory at MARDI for further

identification.