HOST PREFERENCE, BIOLOGY, AND POTENTIAL OF FLEA BEETLE

(ALTICA SP.) AS BIOLOGICAL CONTROL AGENT

FOR LUDWIGIA SPECIES

VAL C. GARCIA

A Master’s Thesis Outline Submitted to the Faculty of the Department of Crop Protection,
College of Agriculture, Central Luzon State University,
Science City of Muñoz, Nueva Ecija, Philippines,
in Partial Fulfillment of the Requirements
for the Degree

MASTER OF SCIENCE IN CROP PROTECTION

(Entomology)

NOVEMBER 2023
 ACCEPTANCE SHEET

This undergraduate thesis entitled “HOST PREFERENCE, BIOLOGY, AND

POTENTIAL OF FLEA BEETLE (ALTICA SP.) AS BIOLOGICAL CONTROL
AGENT FOR LUDWIGIA SPECIES” prepared and submitted by VAL C. GARCIA,
in partial fulfillment of the requirements for the degree MASTER OF SCIENCE IN
CROP PROTECTION (ENTOMOLOGY), is hereby accepted:

ELAIDA R. FIELGALAN, Ph.D. FERNAN T. FIEGALAN, Ph.D.

Member, Advisory Committee Member, Advisory Committee

_____________________ _____________________
Date Signed Date Signed

MARITA S. LABE, Ph.D.

Chair, Advisory Committee

_____________________
Date Signed

Accepted as partial fulfillment of the requirements for the degree of MASTER OF

SCIENCE IN CROP PROTECTION (ENTOMOLOGY):

MARIBETH L. PARUGRUG, Ph.D.

Acting Head, Department of Crop Protection

_____________________
Date Signed

ALONA T. BADUA, Ph.D.

Acting Dean, College of Agriculture

_____________________
Date Signed
 INTRODUCTION

Rice (Oryza sativa) is the primary dietary source for over 50 percent of the

population. Globally, rice is cultivated on approximately 158 million hectares, yielding

over 700 million kilograms in 2015 (USDA, 2016). Asia accounts for more than 90

percent of global rice production and consumption, with China, India, Indonesia,

Thailand, Bangladesh, Myanmar, Vietnam, Laos, Cambodia, Japan, Korea, and the

Philippines among the leading rice-producing nations.

In the Philippines, rice is an important staple diet. It provides more than 45 percent

of the daily calorie requirements for Filipinos, whose annual rice consumption averages

125.26 kilograms (BAS, 2022). Taking into account the country's population in 2021, this

equals 13.8 million kilograms of rice. As the global population continues to increase and

agricultural land becomes more limited, it is estimated that the world will need to

produce an additional 115 million tons of rice by 2035 to meet the increasing demand

worldwide.

The Philippines' food security has been determined by the volume of grain stored in

its granaries. However, Willocquet et al. (2004) estimate that between 120 and 200

million tons of grains are lost due to pests in rice fields in tropical Asia. Globally,

pathogens, weeds, and invertebrates cause significant crop losses, thereby restricting the

accomplishment of global food security and poverty reduction. In terms of food security,

crop losses due to pests may equal the amount of food sufficient to feed more than one

billion people (Birch et al., 2011). Sustainable rice production requires effective pest

management (Savary et al., 2006).

3

For centuries, insecticides have been used throughout the world to control insects.

Insecticides are used most extensively in agriculture, horticulture, and forestry (Gupta et

al., 2019). Chemical pesticides have contributed significantly to the battle against pests

and diseases over the years. Due to human health and environmental concerns, extensive

damage to the environment, pest resurgence, pest resistance to insecticide, and lethal

effects on non-target organisms, their use resulted in the development of insecticide

resistance, the use-cancellation and de-registration of some insecticides (Abudulai et al.,

2001). Pest control has been essential for crop productivity and pest toxin protection

since agriculture began. Food production intensified due to the invention and widespread

use of highly effective chemical pesticides (fungicides, herbicides, and insecticides).

Pesticide resistance in insect populations hindered this progress. Modern pesticides are

more specialized to decrease environmental impacts, but this makes it simpler for target

pests to acquire resistance.

Weeds are a significant issue in many ecosystems, including broadacre cropping,

rangelands, water bodies, natural systems, and recreation places. They are unwanted

plants that, under certain conditions, can have detrimental effects, such as contributing to

yield loss and yield quality reduction in an agricultural context by competing with the

crop for resources such as light, water, and nutrients (Vilà et al., 2004). By competing

with the crop and affecting yield quality, weeds can reduce production more than pests

and diseases. This rivalry, together with the cost of weed management measures like

tillage and herbicides, can cost billions of dollars (Soltani, 2017).

4

Uncontrolled, weeds could reduce global yields of main crops by as much as 34%

(Oerke, 2006). Overuse of herbicides can result in environmental pollution (Relyea,

2005), non-target damage to crops, loss of natural vegetation and soil biodiversity (Rose

et al., 2016), and negative impacts on farmworker and public health (Mamane et al.,

2015; Myers et al., 2016). Unfortunately, the excessive reliance on chemical control has

also resulted in a number of environmental and agricultural issues. Additionally, repeated

use of herbicides may result in the development of herbicide resistance. In this view, the

use of biocontrol agents for controlling weeds is essential.

Biocontrol agents are broadly defined as any living organism that can inhibit the

growth of weeds, although specialist agents such as arthropods (insects, mites) and plant

pathogens are more appropriate and extensively used for targeted weed control

(Schwarzländer, 2018). Typically, biological weed control is conducted using either a

classical or an augmentative approach. The classical approach, which has been employed

most frequently, consists of managing a problematic weed in a particular region by

introducing natural enemies from the weed's native range (Culliney, 2005).

Today, the use of herbivorous bio-control agents to control weeds without

interfering with the cropping system is the most effective ecologically friendly method,

not the exception. For example, a number of coleopteran chrysomelid insects have been

discovered in association with aquatic weeds in areas of their native range; these insects

can function as weed regulators (Roy, 2016). In addition, numerous researchers from

eastern and southeast Asian nations have been on the lookout for suitable biological
5

control agents for common marshy herbaceous vegetation in rice fields (Mukherjee and

Barik, 2013).

Statement of the Problem

In India, the flea beetle Altica cyanea (Weber) (Coleoptera:Chrysomelidae) feeds

on Ludwigia adscendens (Nayek and Banerjee, 1987) so the interest in using this beetle

as a biological control agent for weed in rice paddies. However, little is known of the life

history and host specificity of Altica sp. as a bio-control agent against the Ludwigia

species under Philippine condition. It is well known that host-preference by herbivorous

insects depends on chemical cue including nutritional requirements of insects. Therefore,

studies on its life history, and host specificity to fill these gaps are in order. The potential

of Altica cyanea as a biological control agent against the three Ludwigia species and to

determine if Altica can effectively suppress the growth of Ludwigia need to be explored.

Significance of the Study

The genus Ludwigia (Onagraceae) is dispersed throughout the world, and 82

species are on record (Wagner, 2007). These weeds are currently problematic in rice-

fields as they compete with rice for resources and nourishment. Control strategies in

current use against these weeds are largely on chemical herbicides but intensive use

creates an ecological imbalance through destruction of non-target beneficial insects, and

accumulation of toxic residues in the environment. Further, repeated use of herbicides

6

may result in the development of resistance against herbicides. In this perspective, it is

pre-requisite to use biological agents to control weeds.

Today, the control of weeds through herbivorous bio-control agents without

interfering the cropping system is a most effective eco-friendly approach. Altica sp. are

considered as biocontrol agent of Ludwigia species, the gregarious larvae of Altica

cyanea consume leaves and stems of L. adscendens for 3 weeks to complete three instars,

and the adults subsequently feed for a further 7-8 weeks on this plant (Nayek and

Banerjee, 1987).

To date no record is available on the life history and host-preference of Altica sp.

on Ludwigia octovalvis, L. hyssopifolia and L. perennis under Philippine conditions.

Therefore, studies on the life history and the host-preference of the flea beetle, Altica sp.

to determine its potential value as a biological control agent against the three Ludwigia

species (L. octovalvis, L. hyssopifolia and L. perennis) for their ecologically sustainable

management in near future are needed.

Objectives of the Study

The general objective of the study is to determine the life history, host range and

feeding preference of Altica sp. on Ludwigia species and to evaluate the impact of Altica

on the growth and development of Ludwigia. Specifically, it is aimed (a) to understand

the life cycle, reproductive strategies, growth and survival mechanism of Altica sp. (b) to

evaluate the host-preference of Altica on different species of Ludwigia, (c) and to

determine the biological control potential of Altica against Ludwigia.

7

Scope and Limitations

The first study will concentrate on life history of Altica, an experiment to help

understand the life cycle, reproductive strategies, growth and survival mechanism. The

second study will focus on host range and feeding preference of Altica on Ludwigia

species. The host range of Altica and experiments on the feeding behavior or preference

of Altica on the different species of Ludwigia will be observed.

Time and Place of the Study

The study will be conducted at the Philippine-Sino Center for Agricultural

Technology Central Luzon State University, Science City of Muñoz, Nueva Ecija from

November, 2023 to February, 2024.

8

REVIEW OF RELATED LITERATURE

Identification, Origin and Geographic Distribution of Ludwigia

Ludwigia Linn. (Onagraceae) is a serious weed of rice in many countries, including

India, Malaysia, and the Philippines (Holm et al., 1997; Moody, 1989). Ludwigia species

are considered invasive throughout the world and are a menace to indigenous plant

species. This weed has been observed throughout the rice growing season in direct-

seeded rice (DSR) and transplanted rice fields in Asia (Tomita et al., 2003), especially

when the plant stand is not uniform. In DSR fields, Ludwigia has been reported to occur

in seven countries in dry-seeded rice and six countries in wet-seeded rice (Rao et al.,

2007). The widespread introduction of direct seeding in Asia may increase its abundance

in rice crops. In addition to occurring in the crop fields, Ludwigia is also known to

frequent shallow ditches, pools, and river edges. A single plant of Ludwigia grown in rice

may yield up to 75,000 seeds, with 16,000 seeds per gram (Pancho, 1964). The nearly

cosmopolitan wetland genus Ludwigia, with 83 species currently placed in 23 sections, is

the only member of the subfamily Ludwigioideae of Onagraceae (Wagner et al., 2007;
9

Pesamosca and Boldrini, 2015). Ludwigia species are annual or perennial herbs or shrubs,

with some annual species. Most species (66) occur from temperate Midwestern and

eastern Northern America through subtropical South America, the remainder in the

warmer parts of the Old World (a few species reaching temperature Eurasia), and a few in

both.

Ecological Impacts of Ludwigia

Ludwigia species are considered invasive throughout the world and are a menace to

indigenous plant species (Moody, 1989; Holm et al., 1997; Tomita et al., 2003; Chauhan

et al., 2011). Four species, Ludwigia adscendens (L.) Hara, Ludwigia hyssopifolia (G.

Don) Exell, Ludwigia octovalvis (Jacq.) Raven, and Ludwigia perennis (L.) Raven, are

abundant in rice fields of Philippines, and these weeds are a major impediment to rice

production as they compete with rice for resources in fields (Mitra et al., 2019). L.

adscendens, has stems that creep on the ground, stems are also rooting at the nodes with

their white spongy pneumatophores arising in clusters. L. hyssopifolia has stems are erect

and branched, stems are angled, and woody at the base, while L. perennis stems are also

erect and branched and stems are irregularly rigged and sometime reddish (Donayre et

al., 2018) and L. octovalvis is generally a robust and well branched recurrent weed

(Barua, 2010).
10

The rapid biomas production of these species has a negative impact on ecological

processes in aquatic ecosystems, displacing desirable native fauna and vegetation

(Grewell et al., 2016). The plants form dense masses on the surface of the water, which

impede navigation and hinder recreational activities, irrigation, drainage, and agricultural

production (Thouvenot et al., 2013).

Management Approaches of Ludwigia

To date, the management of Ludwigia species has relied solely on physical,

thermal, and chemical techniques. These options frequently provide short-term control

that necessitates repeated annual interventions, have not been suitable due to ongoing

cost, and may be limited by regulatory constraints (Grewell et al., 2016). Biological

control is a sustainable alternative to these conventional methods because it can be used

alone or as a component of integrated pest management programs. Unlike other methods

of weed control, biological control may provide persistent suppression of weeds at the

landscape level if exotic natural enemies are carefully selected and established upon

introduction.

Classical biological control, the deliberate introduction of host-specific natural

adversaries from the native range of an exotic pest, is an alternative to conventional

control methods. Unlike conventional control methods, biological control has the

potential to suppress vegetation populations at the landscape level and, if successful, is a

long-term, sustainable option (Van Driesche et al., 2010). Biological control is frequently

a component of multifaceted integrated management programs for invasive vegetation

11

because it is frequently used alone or in conjunction with conventional methods

(Holtkamp, 2004; Paynter and Flanagan, 2004).

Natural Enemies

In an effort to provide an alternative to conventional control methods, recent

research has focused on the potential biological control agents of Ludwigia species. The

interest in biological control of exotic Ludwigia species is not, however, new. Cordo and

Deloach (1982) collected one leaf beetle and four species of weevils from L. pepliodes in

Argentina, according to published accounts: Lysathia flavipes (Coleoptera:

Chrysomelidae), two Tyloderma species (Coleoptera: Curculionidae), Sudauleutes bosqi

(Coleoptera: Curculionidae; formerly Auleutes bosqi [Colonnelli, 2004]), and one

Onychylis nigrirostris (Coleoptera: Curculionidae). All adult stages of species eat foliage.

The larvae of Tyloderma spp. and O. nigrirostris are stemborers, whereas the larvae of S.

bosqi feed on the developing meristem of leaves.

Hernández and Cabrera Walsh (2014) gathered insect herbivores from the plants

L. peploides subsp. montevidensis, L. hexapetala, and L. grandiflora. They discovered 19

insect species eating on L. hexapetala, belonging to six feeding guilds. Liothrips ludwigi

(Thysanoptera: Phlaeothripidae), six stem-boring beetle species (Merocnemus binotatus),

five Tyloderma spp. (Coleoptera: Curculionidae), and one fruit-feeding weevil

(Tyloderma nigromaculatum) are among the promising biological control agents.

Liothrips ludwigi is a cell-content feeder that consumes and reproduces in plant apical
12

buds. In young leaves, adults and larvae share the same refuge-feeding sites. Additionally,

the thrips species feeds on L. grandiflora and L. peploides subsp. montevidensis.

Biological Control of Weeds

In agricultural contexts, chemical, mechanical, and physical control methods have

been the mainstay for weed management (Kelton and Price, 2009). However,

conventional control methods are not economically viable over an extended period of

time or for the management of weeds in remote areas or on enormous tracts of public

land with low agricultural value (Culliney, 2005). Continuing globalization and

intensified international trade will increase the naturalization of plant species outside

their native range, leading to the problem and which requires alternative

weeds management strategies.

Biocontrol of weeds with insects intends to control the target weed by introducing

host-specific insects that contribute to the plant's extinction in its native country. Classical

weed biocontrol, a management program that offers safe and effective options for the

control of invasive plant species (Hinz et al., 2020), entails the introduction and

establishment of host-specific coevolved herbivores from the native range of the invasive

plant, with the goal of facilitating the permanent suppression of the plant. There are two

major hazards associated with biocontrol weeds. First, that the invasive insects pose a

risk to non-target flora. Second, that the biocontrol agent will not control the target plant

effectively and will be a poor investment of resources.

13

The use of biological control agents that attack the leaves, branches, and flowers of

weeds is viewed as a promising method of weed management. As a conventional method

of biological control, natural enemies from the native range are introduced to the new

invasion site. This method is regarded as having several advantages, including effective

control of the weed population by impeding its ability to establish and disseminate to

other areas, as well as a lower control cost than herbicide application (Day et al., 2020).

Biological Control of Weeds with Insects

Biological weed control was initially used in India when Dactylopius ceylonicus

(Hemiptera: Dactylopiidae) was introduced to Opuntia monacantha for dye production

(Rao et al., 1971). The insect decimated plant populations and was subsequently

dispersed across the country and into Sri Lanka for the control of O. monacantha.

However, the first official program to control a weed with biological control agents began

in 1902, when 23 insect species were shipped from Mexico to Hawaii to control Lantana

camara (Swezey, 1923). At least 90 countries have introduced weed biological control

agents intentionally (Schwarzländer et al., 2018).

Within the Pacific region, 17 of the 22 countries or territories have implemented

biological weed control, frequently employing agents from other countries, particularly

Australia, resulting in numerous successes (Mbati and Neuenschwand, 2005). In ten of

the thirteen countries where the psyllid has become established, Heteropsylla spinulosa

(Hemiptera: Psyllidae) is reported to control Mimosa diplotricha (Winston et al., 2014).

Calligrapha pantherine (Coleoptera: Chrysomelidae) is controlling Sida acuta in all four

14

Pacific island nations where it has been intentionally introduced (Day and Blue, 2016;

Day and Winston, 2016). In the Pacific region, biological control agents appear to have a

significant impact on Chromolena odorata (five countries), Eichhornia crassipes (four

countries), and Clidemia hirta (three countries) as well (Day et al., 2020).

A comprehensive analysis of the status of biological weed control in Africa and

Asia is not well documented. However, of all the Asian nations, India has been the most

proactive, releasing 20 agents against 10 plant species. Some of the more notable

successes in the region are the control of Salvinia molesta by Cyrtobagous salviniae

(Coleoptera: Erirhinidae) in Sri Lanka (Doeleman, 1989), Alternanthera philoxeroides by

Agasicles hygrophila (Coleoptera: Chrysomelidae) in China (Day, 2018) and Opuntia

spp. by Cactoblastis cactorum (Lepidoptera: Pyralidae) and Dactylopius opuntiae

(Hemiptera: Dactylopiidae) in India (Winston et al., 2014).

The most effective biological control programs have been applied to Eichhornia

crassipes, Pistia stratiotes, and Salvinia molinosa. In many of the 20 countries where the

two weevils Neochetina bruchi (Coleoptera: Erirhinidae) and N. eichhorniae have

become established, Eichhornia crassipes has been controlled or partially controlled

(Beshir and Bennett, 1984; Ajuonu et al., 2003; Coetzee et al., 2009; Winston et al.,

2014;). Pistia stratiotes has been controlled by Neohydronomus affinis (Coleoptera:

Curculionidae) in eight countries (Mbati and Neuenschwander, 2005; Chikwenhere,

1994; Ajuonu and Neuenschwander, 2003) while S. molesta has been controlled by C.

salviniae in five countries (Mbati and Neuenschwander, 2005; Chikwenhere and

Keswani, 1997; ‌Muniappan et al., 2009).

15

The Genus Altica

Altica (Fabricius 1775) is a large genus (approximately 250 species) with a

worldwide distribution; at least 50 species have been identified in the Palearctic region,

and 14 are found in Kazakhstan and Central Asia (Lopatin, 1984). Some species of Altica

cause damage to wood and shrub plantations. All species of Altica share a similar

morphology. In general, their bodies are large or moderate in size, oval or ovate in shape,

dorsally moderately enlarged, pubescent, and metallic green, blue, or violet in color.

Large tubercles are typically delineated by furrows on the frons, and tubercles are more

rarely on top by a weak transverse depression (Lopatin, 1984).

Alticinae is the largest subfamily of the family Chrysomelidae, with over 8,000

species distributed globally, including 1,000 species in the Palaearctic subregion. They

are commonly referred to as flea beetles due to their typically small size (0.5 mm in

length) and their ability to leap; their bodies are small or moderate in size, ovoid,

elongated oval, or cylindrical, and extremely variable in color; many are shiny and

metallic. They have prognathous to hypognathous heads, antennae that are typically 11-

segmented, but occasionally 10- or 9-segmented, filiform, or gradually broaden toward

the apex. The head extends into the pronotum up to the posterior optic region, which is

sometimes completely concealed by it, and the majority of the frons has frontal tubercles

separated from the upper portion of the frons by narrow furrows. The strongly dilated

metafemora contain an endoskeletal leaping organ (Aslan et al., 1999).

16

Altica spp. as Bio-Control Agents

A number of coleopteran chrysomelid insects have been found in association with

aquatic weeds in areas of the native range and can serve the purpose of regulating weeds.

The search for suitable biological control agents for marshy herbaceous weeds common

in rice fields has been conducted by many researchers from the oriental and southeast

Asian countries (Nayek and Banerjee, 1987).

Altica species (Coleoptera: Chrysomelidae) are considered as biocontrol agent of

Ludwigia species, as Altica cyanea feeds on L. adscendes, L. octovalvis and L. parviflora

(Nayek and Banerjee, 1987; Mitra et al., 2019), while A. litigata, consumes L. hexapetala

(Carruthers et al., 2011). Altica foeveicollis is highly effective in Thailand for the natural

control of L. adscendens and other Ludwigia species (Napompeth and Hirose, 1992). In

China, A. cyanea feeds on L. prostrata and Rotala indica (Xiao-Shui, 1990). Larvae and

adult of A. cyanea were recorded from rice fields of tropical and subtropical regions

where Luwigia species are growing, and A. cyanea is perfectly harmless on rice (Nayek

and Banerjee, 1987; Xiao-Shui, 1990; Azad et al., 2015). Both larvae and adult of A.

cyanea are abundant on Ludwigia weeds from October to April in West Bengal, India,

while adults are observed in under shed undisturbed marshy areas and close navigation

channels in summer. Several states in India, as well as Bangladesh, Thailand, Vietnam,

Pakistan, China, Japan, and Malaysia, have reported the presence of the beetle, indicating

that it has a wide niche (Alam and Karim, 1983; Dubey, 1981; Nayek and Banerjee,

1987; Xiao-Shui, 1990; Azad et al., 2015). Adult A. cyanea feeds for 7-8 weeks on L.
17

adscendens, while its larvae gregariously consume leaves and stems of these plants for

three weeks to complete three instar. The insect also feeds on Ludwigia prostrata,

Ludwigia hyssopifolia, Trapa natan, and Rotala indica (Roy, 2018). This insect also feeds

on other widespread weeds, including Ammania rotundifolia (Lefroy, 1978), Trapa

natans, and T. bisponsis (Nayek and Banerjee, 1987). Both larvae and adults of A. cyanea

are abundant in rice fields of tropical and subtropical regions where Ludwigia species are

present, but no damage to rice has been documented (Nayek and Banerjee, 1987; Xiao-

Shui, 1990; Lemmes and Bunyapraphatsara, 2003; Azad et al., 2015). The black larvae of

A. cyanea feed on the plant's leaves and stems. The stem portions are not completely

submerged and remain partially exposed for feeding, and the insect breeds freely in L.

adscendens during the rains and until December (Nayek and Banerjee, 1987).

MATERIALS AND METHODS

Conceptual Framework

Figure 1 demonstrates the conceptual framework for this study. The study will

concentrate on the life history, and host preference of Altica sp. as a bio-control agent

against the selected three Ludwigia species (L. octovalvis, L. hyssopifolia, and L.

perennis), and the impact of Altica on Ludwigia's growth and development. The results

will be useful in a better understanding of Altica's life cycle, reproductive strategies,

growth, and survival. The other host of Altica will be noted and if it prefers specific
18

species of Ludwigia. The potential value of Altica sp. as a biological control agent against

the three Ludwigia species will be determined to attain an ecologically sustainable

management program in the near future.

Evaluation of
Altica spp.

Impact of
Life Altica Host
History on Ludwigia Preference

Potential bio-
control of
Ludwigia species

Figure 1. Conceptual framework for the study

Life History Study

Establishment of Test Plants

Ludwigia will be planted in pots and maintained in caged conditions. These will be

used for the establishment of test insects and for life history studies. The plants will be

kept in the screenhouse until it is ready to be used. About 2-3 weeks old Ludwigia plants

will be used to study the life history of Altica sp.

Establishment of Test Insect

19

To start the mass rearing, a mixture of male and female adults Altica sp. will be

collected from Ludwigia species growing in rice fields near Philippine-Sino Center for

Agricultural Technology (PhilSCAT). The collected insects will be allowed to oviposit

on potted Ludwigia plants enclosed in a 20cm (height) x 15cm (diameter) fine mesh

nylon cage.

The deposited eggs will be allowed to develop and hatch, and the emerging insects

will be placed in a cage plant and maintained in such a way that the proper development

will be ensured, while the resulting colonies will not result in the death of the plants.

The established colonies will serve as insect sources for life history and host

preference studies.

Life History Set-up

Adult male and female Altica sp. will be carefully collected from the stock culture

using an insect aspirator and confined for mating and oviposition on detached twigs of

Ludwigia with fully expanded leaves kept in plastic transparent containers lined with

moist paper towel.

20

Adults will be removed from plastic container immediately after laying eggs. The

eggs will be regularly observed, once in the morning and once in the afternoon, for

hatching. Incubation period of the eggs will be noted.

Once the eggs hatched, the larvae will be individually transferred by carefully

letting individual larva crawl in a young, fresh leaf of Ludwigia hyssopifolia. The base of

the leaf will be wrapped in moistened cotton ball. The set-up will be maintained in plastic

petri dish. The leaves will be replaced when necessary.

The larva will be observed in the morning and in the afternoon for molting. Pupae

will be left undisturbed until the insects emerge as adults.

The adults will be maintained for fecundity and longevity data. Mating and

oviposition behavior will be observed and recorded.

The experiment will be conducted under laboratory condition with fluctuating

temperature of 25-30OC, relative humidity of 60% and natural lighting. Temperature will

be monitored using ordinary thermometer. Observation of the different stages will be

undertaken with the aid of a hand lens, or when necessary, under a dissecting microscope.

Description of Life Stages

Representative specimens of each of the stages of the insect will be collected and

described. These will be photographed accordingly before preservation in 70% ethyl

alcohol.
21

Host Preference Study

Test Plant

The propagation of Ludwigia hyssopifolia, L. octovalvis, L. perennis, and eggplant

(Solanum melongena) will be carried out using seed propagation in order to assure

uniformity in the age of the resulting plants. While the propagation of rice (Oryza sativa)

and corn (Zea mays) will be done by the method of direct-seeding.

After the test plant has successfully established roots, it will be transferred to pots

(15 cm diameter x 18 cm high). The plants will be subjected to natural day lengths and

temperatures inside the screenhouse.

Plants will get irrigation when required, while refraining from the application of

pesticides or fertilizers in order to prevent any potential impact on the biology or

behavior of Altica sp.

Adult Free-Choice Test

Host-preference tests will be conducted with potted plants. Each pot will be filled

with soils and contained 2 plants of each test plants (three species of Ludwigia, rice, corn,

and eggplant). The test plants will have at least 2-3 fully grown leaves. The potted test
22

plants will be arranged in circular pattern with the foliage of the plant touching to

facilitate movement of the insect between plants. All of the potted test plants will be

surrounded by a nylon-net cage, which will be secured to the borders of the pots.

Before the test, starved (4-6 hours) 20 newly emerged paired adults will be

introduced to each cage and will left undisturbed and subsequently removed after 24

hours. Thereafter, plants will be examined for feeding damage following the rating scale

by Dixon et al. (2023) (Table 1). The set-up will be replicated three times and will be

repeated thrice.

Adult No-Choice Test

Cages, pots and experimental conditions will be similar to those of the free-choice

test but the test plants will be caged individually.

Twenty starved (4-6 hours) adult will be released in each caged potted plant. Plants

will be examined after 24 hours for feeding damage. Treatments will be replicated 3

times and the test will be repeated 3 times.

Table 1. Rate used in assessing the feeding damage of Altica sp. (Dixon et al., 2023)

Scale Description (Visual estimation of damage )

23

1 evidence of exploratory feeding

3 minimal feeding

5 minimal feeding damage,

evidenced by minimal feeding scars and holes

7 extensive feeding damage

Data Analysis

All the data on host specificity as well as the data on growth duration (i.e. larval

and post larval duration and adult longevity) survivability and fecundity of Altica sp.

reared on the three host-plants will be subjected to one way analysis of variance

(ANOVA). Means associated with all the data for each variable will be separated using

Tukey’s Test when significant values will be obtained.

LITERATURE CITED
24

(BAS) Bureau of Agricultural Statistics. 2008. Department of Agriculture of the Republic

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