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2014

Section VIII
Insect Bioinformatics

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NAL B OO

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Short Views on Insect Biochemistry and Molecular Biology

Vol. (1), 00 00, 2009
Vol. (2) 663 684, 2014

Invited Review
Invited Review

Chapter 29

Molecular expression and structure-function relationships

of apolipophorin III in insects with special reference
to innate immunity

Bharat Bhusan Patnaik1,3, Raman Chandrasekar 2, Yeon Soo Han1*

1

Department of Applied Biology, Division of Plant Biotechnology, Institute of Environmentally-Friendly

Agriculture (IEFA), College of Agriculture and Life Sciences, Chonnam National University,
Gwangju 500-757, Republic of Korea.
2
Department of Biochemistry and Molecular Biophysics, Kansas State University,
Manhattan 66506, KS, USA.
3
School of Biotechnology, Trident Academy of Creative Technology (TACT), Bhubaneswar 751013
Odisha, India.

Abstract

Insect immune response relies on the humoral and cellular mechanisms of innate immunity. The
key factors of the humoral immune mechanisms are the antimicrobial peptides that act in concert
against invading pathogens. One of the several such key factors is the apolipophorin III (apoLp-III)
gene that is constitutively expressed in the hemolymph and functions in immune sensing and
activation in insects against invading pathogenic microbes, apart from its role in lipid transport. We
have reported the functional role of apoLp-III in the epithelial cells of mosquito gut and established
its action as a negative regulator of Plasmodium development. Recently, we also studied the promoter
sequence of apoLp-III gene from a coleopteran beetle, Tenebrio molitor (TmapoLp-III) to understand
the functional regulation imposed by the putative transcription factors. In addition, the critical
function of TmapoLp-III in immune surveillance against an intracellular pathogen, Listeria
monocytogenes has been currently explored. Here we review the known functions of apoLp-III in the
hemolymph and also make an attempt towards understanding the intracellular role of Anopheles
gambiae apoLp-III (AgapoLp-III). A detailed analysis of AgapoLp-III sequence and structure has
been provided to dissect the evolution of the protein in insects that form a basis to authenticate
their multidimensional role.
Key words: Anopheles gambiae, apoLp-III, gene expression, molecular structure, innate immunity
.
*For Correspondence (email: hanys@jnu.ac.kr)

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Overview
1.
2.
3.
4.
5.
6.
7.
8.
9.
10.

Introduction
Insect innate immunity and apolipophorin-III
Apolipophorin-III expression and function in the hemolymph
Intracellular expression and function of apoLp-III
Sequence similarity and comparisons
Molecular structure of A. gambiae apoLp-III
Evolution of A. gambiae ApoLp-III
Functional significance and Conclusion
Acknowledgements
References

1. Introduction
Apolipophorin-III (apoLp-III) is a prototypical exchangeable insect homolog of
human apolipoprotein-E that functions in the transport of diacylglycerol (DAG) from
the fat body to the flight muscles in the adult life stage (1). ApoLp-III is synthesized in
a variety of tissues apart from the fat body cells and these include hemocytes, midgut
epithelial cells, ovaries and testes. The differential tissue-specific expression levels of
apoLp-III in insects suggest their additional functions in immune responses against
various pathogens. ApoLp-III has been abundantly characterized from insect
hemolymph and has been shown to mediate insect immune responses in various
species such as Galleria mellonella (wax moth) (2), Hyphantria cunea (fall webworm)
(3), Heliothis virescens (tobacco budworm) (4), Locusta migratoria (locust) (5),
Anopheles gambiae (mosquito) (6), Thitarodes pui (ghost moth) (7), and lately in
beetles Tribolium castaneum (8) and Tenebrio molitor (9).
It is well understood by now that insects possess a highly specialized innate
immune response mechanism to account for the lack of adaptive immunity. The innate
immunity in insects is regulated by the hemocyte-mediated cellular component and the
humoral component that includes the antimicrobial peptides and prophenoloxidase
(PO) cascade (10). ApoLp-III forms the humoral component with other factors such as
lysozyme, cecropins, attacins etc. and acts as a pattern recognition receptor in the initial
line of defense against microbial/parasitic infections. ApoLp-III has attracted more
attention than the other humoral factors due to their versatility in eliciting an immune
response against invading pathogens. ApoLp-III can act as a pattern recognition
protein, inducing antimicrobial activities against diverse groups of pathogens (11,12),
stimulate the PO cascade (13) and participate in cellular immune reactions such as
phagocytosis and encapsulation (14).
ApoLp-III has been extensively investigated with regards to the molecular details
of the binding interactions with cell wall components such as lipopolysaccharide (LPS),
peptidoglycan (PGN) and -1,3 glucan (15-17). Especially, in Galleria mellonella,
apoLp-III has been found to interact with the cell wall components that suggested its
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presence as a carrier molecule (14, 18). Binding of apoLp-III to wild-type and mutant
Candida albicans accelerated the removal of yeast cells from G. mellonella
hemolymph and enhanced hemocyte-mediated nodulation in yeast (19). Significantly,
the treatment of Metarhizium anisopliae conidia with apoLp-III prior to injection into
G. mellonella larvae decreased the mortality percentage (14 The most fascinating
model for the binding interaction relates to a conformational change of the protein
whereby the hydrophobic face of the protein interacts with the interior lipid region,
leading to the formation of a stable apoLp-III/lipid region complex (20, 21). The
reversible change in conformation of apoLp-III prevents lipids from coming into
contact with aqueous environment.
Many studies on insect apoLp-III have focused on sequence-structure
relationships and their evolutionary positions within the insect orders. The identities of
apoLp-III at the primary sequence level was found less striking within the orthologous
groups, although they fold to similar tertiary structures (6,22). The predicted structure
consists of five long amphipathic alpha-helices connected by short loops. Another
distinctive feature of apoLp-III helix bundle is helix 3 that is envisioned to act as the
best candidate amphipathic helix initiating lipid binding through one end of the bundle
(23).
The sequence diversity exemplified by apoLp-III in various insects, still folding
into a similar tertiary conformation leading to multi-dimensional roles has made it an
attractive target for efficient gene silencing. The structure-function relationships of
apoLp-IIIs have been well depicted in Manduca sexta, H cunea, A. gambiae and T. pui
model systems (6, 7, 24, 25). The sequence-specific knockdown of apoLp-III has been
demonstrated in several insects such as H. cunea, Plutella xylostella and T. castaneum
(8, 11, 26). Although gene silencing in H. cunea explored a novel function of apoLp-III
in the regulation of antioxidant enzymes such as manganese superoxide dismutase
(MnSOD), other reports emphasized their role in innate immunity mechanisms. We
have studied the effects of apoLp-III silencing in the mosquito A. gambiae and
established the participation of the protein in mosquito antiplasmodial responses (6).
We localized high apoLp-III levels in the cytoplasm of Plasmodium berghei invaded
cells and assigned a putative role as a cytoplasmic pathogen recognition receptor.
Recently, we have made an attempt to study the genetic regulation and expression of
apoLp-III gene isolated from the expressed sequence tag (EST) information of
T. molitor larvae with special reference to the sensing of an intracellular pathogen,
Listeria monocytogenes (9). The results indicated low sequence identity to other
orthologous proteins with similar structural folds as revealed in previous studies. Here,
we discuss the regulation of apoLp-III gene in insects by assembling and annotating the
apoLp-III sequences from the databases. This will be helpful in advancing further
research on the functional characterization of apoLp-III with special reference to insect
innate immunity.

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2. Insect innate immunity and apolipophorin-III

Insects are the most diversified organisms among the eukaryotes and can inhabit
various habitats, without constraining to latitudinal and altitudinal boundaries. This is
significant as they lack the adaptive immune response characteristic of vertebrates. The
extreme challenge of climatic regions as well as the variable pathogenic stresses has
evolved their innate immunity pathways. These pathways are sufficient to mount
protective responses to varying pathogens, including bacteria, fungi, and viruses. The
Toll pathway is necessary in mediating defense against Gram positive bacteria and
natural fungal infections, whereas the Imd pathway controls the Gram negative
infections (27). The antiviral response in insects is known to be regulated by the RNA
interference (RNAi), Toll, Imd and Jak-STAT pathways (28-31). Although these
pathways can work independently against the pathogens, there are evidences of an
integrated, versatile insect immune system capable of communication between
pathways and cells to give rise to an effective response (32).
In a pursuit to study the innate immune mechanisms in insects, different
approaches including advanced microscopy techniques and functional genomics using
(RNAi) have been utilized. Several genes involved in the recognition of pathogens and
the modulation of the extracellular and intracellular signaling pathways have been
elucidated in model insects. This chapter presents a comprehensive review of
apoLp-III in insects with special reference to innate immune mechanisms. Although
many reports have established the participation of apoLp-III in extracellular immune
responses, studies lately have focused on midgut immunity. We have made an attempt
to compile the previous reports on the constitutive and induced expression of apoLp-III
gene in different insects. Further, we gain insights into the role of apoLp-III in inciting
an immune response in the midgut epithelium of A. gambiae (G3 strain) mosquitoes in
response to P. berghei infection. This is followed up with the detailed analysis of the
structure-function relationships of insect apoLp-IIIs with special reference to the
A. gambiae apoLp-III (AgapoLp-III).

3. Apolipophorin-III expression and function in the hemolymph

Apolipophorin-III is an abundant hemolymph protein in larval and adult stages of
insects and is involved in several critical functions including eliciting an immune
response. It is synthesized not only by fat body cells but also in a variety of other cells,
including those of hemocytes, ovaries and testes (3). The gene has been known from
several insect orders such as Lepidoptera, Diptera, Orthoptera, Hymenoptera,
Hemiptera, Orthoptera and Coleoptera and has been studied for their role in activation
of immune mechanisms in the hemolymph. An account of the isolated and
characterized apoLp-III gene with their multidimensional roles have been depicted in
Fig. 1.
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Fig.1. The apoLp-III genes isolated and characterized from representative species of different insect
orders. # The apoLp-III gene from the coleopteran beetle has been recently characterized by our
group. The critical functions of the isolated apoLp-III have been highlighted with special reference to
innate immunity.

The level of hemolymph apoLp-III undergoes changes in different species of

insect depending on the pathogen/parasite. The variable hemagglutination
characteristics of apoLp-III, indicates its participation against infection by microbes
having different cell wall components (2, 33, 34). A study on the changes of apoLp-III
level in the hemolymph of G. mellonella larvae after bacterial infection is critical (35).
The authors found that the entomopathogenic strains of Pseudomonas aeruginosa and
Serratia marcescens increased the amount of apoLp-III in the hemolymph, whereas
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Bacillus thuringiensis showed an insignificant rise in apoLp-III level. In contrast,

apoLp-III protein increased in T. castaneum larvae exposed to spore-crystal mixtures
of B. thuringiensis (8). Further it was observed that the infection by entomopathogenic
bacterium, Xenorhabdus nematophila led to an increase in apoLp-III levels in the
larvae of P. xylostella (11). ApoLp-III was also highly expressed in the hemolymph of
Micrococcus luteus challenged herbivorous insect, Trichoplusia ni (36), although this
seems to be in contrast to the down-regulation of apoLp-III gene in Apis mellifera in
response to infection with the same pathogen (37).
ApoLp-III exists in the insect hemolymph in free as well as lipid-bound form.
They are able to interact with lipids variably, and these apoLp-III-lipid complexes can
act as the necessary signal to activate the immune system of insects. This can be
postulated as an early step of immune recognition by apoLp-III in hemolymph of
insects (20, 38). To explain the association of apoLp-III with lipid components, a
model showing the binding interaction of G. mellonella apoLp-III with LPS has been
known (21). A conformational change of the protein is found to be necessary for the
direct interaction of the hydrophobic protein interior with the lipid A region of LPS,
forming a stable apoLp-III/LPS complex. Although ionic interactions may be
important for the initial binding to the carbohydrate, the amphipathic nature of
-helices (particularly the non-polar face) may be considered the critical structural
feature required for LPS binding.
This two-pronged strategy of binding the lipids by apoLp-III resembles the
binding of antimicrobial peptides, which are expressed as potential therapeutic proteins
against LPS infections (39, 40). ApoLp-III has also been known to bind and detoxify
lipoteichoic acids (LTA). G. mellonella apoLp-III interacts with LTAs of Bacillus
subtilis, Enterococcus hirae and Streptococcus pyogenes (41). Apart from the same,
apoLp-III has been tested against bacteria lacking LTA in their cell wall (2). The
binding of apoLp-III to C. albicans accelerated the removal of the yeast cells from
G. mellonella hemolymph, enhancing hemocyte-mediated nodulation in yeast (19, 42).
Other studies have also described the interaction of apoLp-III with the cell surface of
selected fungi by binding to -1, 3 glucan (14, 42). The binding of insect apoLp-III to
microbial cell wall components establishes its role as a multiple-binding pathogen
recognition receptor (PRR) and a component of detoxification machinery. The role of
apoLp-III in LPS detoxification has been suggested. In Bombyx mori hemolymph, the
association of apoLp-III/LPS caused a significant depreciation in the biological
activity of LPS, reflected by a significant reduction in the induction of cecropin gene
(43). We have made a comprehensive review of apoLp-III gene expression and its
potential functions and presented in Table-1. This will be crucial to understand the
participation of apoLp-III in the signaling network between hemolymph, hemocytes
and fat body cells.
The role of apoLp-III as a signaling molecule in the hemolymph of insects is clear
as the lipid-associated apoLp-III was seen in the hemocytes of G. mellonella larvae
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(38). The protein increases phagocytosis by insect hemocytes (44) and stimulates
cellular encapsulation of the foreign material (41). ApoLp-III also inhibits the adhesion
of G. mellonella larval hemocytes by reducing protein kinase C (PKC) activity in the
lipid-enriched samples (45). Moreover, apoLp-III is known to be involved in
entrapping of pathogens in the net-structures formed by nucleic acids released from
ruptured hemocytes (46). Table-1 also shows that apoLp-III is constitutively expressed
in hemocytes of various insects such as P. xylostella, H. cunea and T. castaneum
enhancing the hemocyte nodulation behavior and preventing lethal pathogenicity. We
have also observed a greater constitutive expression of apoLp-III in the hemocytes of
T. molitor larvae (9).
The most striking function of apoLp-III in hemolymph of insects is the
potentiation of antimicrobial activity. The antibacterial activity induced through the
injection of native and recombinant apoLp-III from G. mellonella was similar to that
provoked by injecting bacteria (44, 47). In H. virescens, bacterial infection led to an
increase in apoLp-III protein levels in both larval and pupal hemolymph (4). In
T.molitor, E. coli challenge showed nearly 10-fold upregulation of apoLp-III
transcripts at 24 hrs followed by an eventual decline in the expression levels (9). Such
an intensification of hemolymph antimicrobial activity after apoLp-III injection would
be an outcome of the activation of antimicrobial peptides as also demonstrated in H.
cunea. The authors have also detected apoLp-III in H. cunea granulocytes and
postulated the local discharge of the protein in response to bacterial challenge (3).
Certain reports show that the antimicrobial activity of apoLp-III shows a synergistic
action with defense peptides. A study observed that the muramidase activity of G.
mellonella lysozyme considerably increased in the presence of apoLp-III. ApoLp-III
was also able to enhance the permeabilization activity of lysozyme towards E. coli cells
(12). Additionally, an increase in cecropin A anti-E. coli activity has been noticed in
the presence of apoLp-III (48).
Activation of the PO cascade has been reported to occur following the recognition
of cell wall components such as PGN and LPS from bacteria as well as -1, 3 glucans
from fungi. It also gets activated after tissue damage by mechanical wounding and
possibly by the enzymes released from pathogens (49). ApoLp-III acts as a strong
activator of PO pathway as well as the synthesis of antimicrobial peptides as it is a PRR
recognizing the cell wall components of microbes (48). The PO cascade was activated
by G. mellonella apoLp-III, although it was found to depress by induction with LTA
from B. subtilis (41). In T. castaneum, apoLp-III participates in immune response to B.
thuringiensis by activating the PO cascade. This was because the PO activity was
found to be reduced in apoLp-III silenced larvae than that of non-silenced intoxicated
larvae (8). An association of PRRs, lectins and regulatory proteins activating PO
including the lipophorins has been known (50). Lipophorins, including apoLp-III have
also been special because they are involved in clot formation in several insects such as
T. molitor, Locusta migratoria, Periplaneta americana and Leucophaea maderae (46,
51).
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Table 1. Comprehensive review of apoLp-III gene expression with their potential functions in insects. The GenBank
accession number for the nucleotide sequences has been provided. * represents partial sequence. up and down arrows
represent the upregulation and downregulation of the apoLp-III gene expression against microbial challenges.

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These studies have provided sufficient evidence regarding the central role of
apoLp-III in immune activation. Although this is true for extracellular immune
mechanisms, it would be important to divulge the function of apoLp-III in intracellular
signaling pathways, transcriptional regulation of antimicrobial peptides and other
effector genes in the nucleus. The larger question to be addressed here is whether
apoLp-III can sufficiently activate the serine protease cascade after binding to the lipid
moieties of pathogen associated molecular patterns (PAMPs). This would provide
some interesting insights, as an earlier study has reported the decline in the immune
responses of G. mellonella, including the PO and the cellular encapsulation reaction
after challenge with entomoparasite Steinernema feltiae. This attributes to the removal
of apoLp-III from the insect hemolymph after interaction with the parasite-cuticular
lipids, thus preventing the inducible response of the host (52). Such immunodepression
inflicted by the pathogen/parasite can result in greater host mortality percentage. In
future, such models of interference in antimicrobial peptide synthesis will be critical in
understanding the role of apoLp-III in the activation of the NFB family transcription
factors.

4. Intracellular expression and function of apoLp-III

The presence of apoLp-III protein in the granules of H. cunea prohemocytes,
plasmatocytes, granulocytes and spherulocytes has been known (3). Our earlier studies
with the dipteran model, A. gambiae (6) as well as a recent study in the coleopteran
beetle (9), showed greater expression of apoLp-III in the late pupal stages, the time
related to the programmed cell death. These are in agreement with a previous study
showing an increase in apoLp-III titers during programmed cell death (53). Towards
the understanding of the sensing and recognition processes implicated by apoLp-III
against intracellular microbes/parasites, we studied the plasmodial infection in A.
gambiae (G3 strain). This was significant as an earlier study was unable to establish the
role of apoLp-III in the intracellular immunity of A. gambiae (Yaounde strain)
mosquitoes infected with Plasmodium falciparum (54). We were able to localize
greater levels of apoLp-III protein in the cytoplasm of P. berghei invaded midgut cells
(6). This was crucial as we were able to address the local or gut-specific immune
response that has a direct application towards the perennial problems of pests and
diseases in the farmers field.
We established the negative regulation of P. berghei infection in the epithelial
cells of the midgut after silencing of apoLp-III gene. These results have shown the
participation of A.gambiae apolipophorin-III (AgapoLp-III) in enhancing an
antiplasmodial response in the cytoplasm of the host. This provides advancement on
the immune regulatory role of apoLp-III apart from its known role in lipid transport and
immune activation in the extracellular fluid. We have also observed the upregulation of
apoLp-III transcript from a coleopteran beetle T. molitor after infection with an
intracellular bacterium, Listeria monocytogenes (9).

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To understand the multidimensional roles of apoLp-III in insects, we have made

an analysis of the sequence and structure of AgapoLp-III and compared it with
apoLp-IIIs from orthologous groups of insects.

Fig.2 Alignment of the amino acid sequences of insect apoLp-IIIs. (A). Aligned amino acid sequences
for ApoLp-IIIs are from Anopheles gambiae (HQ112291), Cluex quinquefasciatus (XP_001849278),
Aedes aegpyti (XP_001659524), Hyphantria cunea (AAQ24031), Manduca sexta (P13276.1), Galleria
mellonella (CAA07363), Bombyx mori (AAQ7038), Trichplusia ni (ABV68867.1), Thitarodes pui
(ADM64569), Plutella xylostella (ADK78218). Five amphipathic -helical domains are indicated by
arrow line. The leucines that are putatively involved in the initial contact of the lipophorin surface are
indicated by a bold face. Conserved resides that are probably involved in breaks and /or turn between the
helices are designated with a star below the residues and non-conservative amino acid substitutions are
marked with : and . respectively. Cysteine residue (Cys12) involved in metal binding are marked in
red color. (B) Sequence homology in the insect Apolipophorin-III.

5. AgapoLp-III sequence similarity and comparisons

The amino acid sequence of AgapoLp-III was aligned with apoLp-III from ten
insects. The result of the amino acid alignment is illustrated in Fig. 2. The AgapoLp-III
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cDNA codes for a polypeptide of 193 amino acids, including a 23 amino acid putative
signal sequence for secretion. The predicted cleavage site between amino acids VRR
and DA was found as proposed [55]. The sequence alignment showed extensive
homology with all the insects and high degree of conservation in amino acids 21 -25
(VRRDA) that flanks the signal peptide cleavage site (marked as asterisk;
Fig. 2A). There are two short insertions in mosquito apoLp-IIIs at positions 3034
(ASEEP) and 6264 (GFQ) that are not present in Lepidopteran apoLp-IIIs. Fig. 2B
shows that the AgApoLp-III show a higher degree of sequence identity with other
mosquitoes such as, C. quinquefasciatus (66%), A. aegypti (58%), although a
substantial difference was found when the Orthopteran, L. migratoria was compared to
lepidopteran insects (17.8 - 20.6%).

6. Molecular structure of AgapoLp-III

The structural homology of AgapoLp-III was identified using a known 3D
structure (PDB-1EQ1 template of M. sexta structure) by PSI-BLAST. Structural
analysis reveals that AgApoLp-III molecules consist of five, long amphipathic alpha
helical bundles with short loops connecting the helix 1 (N-terminal helix), helix 2, 3
and 5 (C-terminal) to the center of helix 4 (Fig. 2A). In the predicted model of
AgapoLp-III, the hydrophobic faces oriented towards the center of the bundle, whereas
the hydrophilic faces interacted with the solvent. This molecular architecture resembles
to that of the four-helix, 22 kDa N-terminal domain of human apolipoprotein E. (56).
The predicted homology model demonstrates a five-helix bundle for lipid-free
AgapoLp-IIII (Fig. 3A) and is found similar to the X-ray crystallographic structure of
M. sexta and L. migratoria apoLp-III (57, 58). The modeling steps included are the
backbone superposition of the atom, loop modeling and orientation of the side chains
(59, 60). There are several ways to model the backbone (58, 59). The target backbone
(from N-terminus to C-terminus) was built by averaging the backbone atom position of
the template structures. Structural model was built based on the atomic coordinates in
the PDB data files and visualized in the appropriate representation.
The predicated model of AgapoLp-III was analyzed using energy minimization,
refinement and simulation program at PROCHECK using a full set of PDBsum
analyses (www.ebi.ac.uk/thornton-srv/software/PROCHECK/) (61, 62). This helped
to evaluate the geometry and stereo-chemical parameters (r0, and ) for the
qualitative analyses of predicted model. The resultant structure (1.8 2.0 ) was
visualized in several representations, such as line drawing, ribbon model (Fig. 3B),
cylindrical, (Fig. 3C),

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Fig.3 Deduced 3D structure of Anopheles gambiae (AgApoLp-III) predicted using Manduca sexta
apoLp-III model (PDB ID: 1EQ1) as a template. A) Structure superposition of AgApo3 (blue-cylinder
model) with template (silver-ribbon model); B) Ribbon model; C) Cylindrical model; D) Transparant
mesh surface ribbon model, E) sphere model; F) Transparent grid surface ribbon model; G) Stereo view of
molecular surface model based on type of residue. Color code: blue-charged amino acid; whitehydrophobic; red-negative charge.

ball and stick (Fig. 3D), transparent grid surface of ribbon model (Fig.3 E, F), space
filling model (showing secondary structure elements) and the molecular surface
(Fig. 3G), using PyMOL molecular graphics system, version 1.5.0.4 (www.pymol.
org/). Furthermore, Ramachandran plot was used to evaluate dihedral angles agreeing
with the values of allowed conformation for protein backbones. The plot diagram
showed that 84.8% of amino acid residues are in the most favored regions, 10.9%
residues in additional and generously allowed regions, and 7 residues (Ala25, Phe35,
Gln64, Asp100, Asn124, Val162 and Pro163) were found in disallowed regions (Fig.4A).
The comparable Ramachandran plot characteristics and G-factor score confirmed the
good quality of the predicted AgApoLp-III model. Based on the study on main-chain
and side-chain, we found the confirmation of the predicated model to be favorable; and
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stable with 99.8% accuracy. The AgApoLp-III model was deposited in Protein Model
Database -PMDB (http://www.caspur.it/ PMDB) bearing Model ID: PM0078769.

Fig.4 A) Ramachandran plot analysis of AgApoL-III model using Mol-Probity software.

B) Structure was highlighted 1-5 helix region and Leucine 32, 35 and 95 residues are possible
binding region for lipophorin surface. Thr 32 and Thr 44 residue for disulfide stability of helix
region. C) Graph sowed AgApoLp-III hydropahty and amphipathicity. Hydropathy (blue) and
amphipathcitiy (red) plot diagram. D) Surface electrostatic potential model (positive potential
in blue and negative in red).

The arrangement of amphipathic helices in AgapoLp-III is similar to those in

Lepidopteron groups (Spodoptera litura, M. sexta, G. mellonella, and B. mori and L.
migratoria). The protein shows structural and functional homology to other
Lepidopteran insects as well as to L. migratoria apoLp-III, but is non-glycosylated
sharing only 29% sequence identity (63). On the basis of numerous inter helical nuclear
overhauser enhancement contacts detected, it is postulated that hydrophobic
interactions between paired helical segments stabilize the bundle conformation (64, 65).
This effect may be counter balanced by several polar residues located in the proteins
hydrophobic interior, which may contribute to the overall low intrinsic stability of
lipid-free apoLp-III. The distinct structural feature of the apoLp-III helix bundle is
helix 3. This short helix connects helix 3 and helix 4, and orients almost perpendicular
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to the helix bundle (Fig. 4B). In terms of the proposed helical repositioning upon lipid
association (57), helix 3 may function in initiating the conformational opening of the
protein. The multiple sequence alignment and the hydrophobic plot (Fig. 4C & D)
reveals that despite helix 3 being short, its amino acid sequence is amphipathic,
containing both hydrophobic (Val, Leu) and hydrophilic residues (Lys, Gly). The
identification of the active binding sites in the predicated model of AgApoLp-III was
performed using the Q-site finder (www.modelling.leeds.ac.uk/qsitefinder/).
AgApoLp-III contains a conserved cysteine residue (Cys12) near the N-terminal,
which might serve as metal binding site.
ApLp-III protein has two conserved prolines and a glycine residue which are
involved in breaks or turns between -helices (66, 67). It is also envisioned that the
apoLp- III helix bundle will initiate lipid binding and contact with lipoprotein
surface-binding sites (68). These analyses speculate that a partially folded molten
globule-like state of apoLp-III may play an important physiological role in lipoprotein
metabolism. Based on the available information from this analysis, the conservation of
structurally and functionally important residues, and the experimental evidences
showing that M. sexta apoLp-III is functionally equivalent to L. migratoria apoLp-III
(69), we propose that insect apoLp-IIIs share a common structural motif of five
amphipathic helices with properties similar to the class A amphipathic helical domains
of vertebrate exchangeable apolipoproteins. In addition, precise amino acid identity
appears less important than the distribution of polar and nonpolar residues.

7. Evolution of AgapoLp-III
The evolutionary relationship of AgapoLp-III was studied by the alignment of the
thirty-five conserved N-terminal sequence motifs of the selected apoLp-III sequences.
The phylogenetic analysis was conducted by using the neighbor- joining (NJ),
maximum likelihood (ML) method (Fig. 5). An un-rooted tree shows the specifications
of relationships but does not order them according to their history. AgapoLp-III was
closely related within their group, but was distantly related to other insect groups such
as Orthoptera, Hemiptera, Hymenoptera, Coleoptera and Lepidoptera. Despite the fact
that the majority of non-identical residues represent conservative substitutions, the
degree of sequence identity appears rather low, compared with that of other
orthologous proteins from different groups. Observation of modular architecture and
phylogenetic analyses demonstrates that insect apoLp-III proteins are members of
lipophorin superfamily and are homologous to vertebrate apoE. Our data strongly
suggest that these sequences diverged after an ancient duplication event leading to the
separation of an apoLp-III paralogous from bacterial outer membrane lipophorin
(OML) sequences.

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Table 2. List of apoLp-III protein in insects and their accession number

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The fact that all the apoLp-III, so far characterized, have the same structure, tends to
make us believe that they have a similar function, and may have a common ancestor.
Through their homologous sequence they retained the capacity to bind and transport
lipids, a function that is shared within apoLp-III family members and thats probably
the ancestral function.

Fig.5 Phylogenetic analysis of the aligned apoLp-III protein sequences. The analyses were
conducted using the branch and bound parsimony algorithm and single un-rooted phylogram
three was identified. The length of the branch lines indicates the extent of divergence
according to the scale 2.0 bar. The Gene Bank accession numbers of the ApoLp-III protein
sequences are given in Table 2.

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8. Functional significance and Conclusion

The sequence and structure similarities found between AgapoLp-III and other
organisms suggest that they originate from a common ancestor, and that they possibly
share the same function of carrying hydrophobic ligands in a hydrophilic medium.
Further studies on apoLp-III lipid interaction, receptor binding and enzyme activation
will provide important clues about how this protein functions in plasma lipoprotein
metabolism. Most significantly, the intracellular role of apoLp-III is a rich area to be
exploited towards understanding of the innate defense mechanisms in insects. RNAi
mediated gene silencing will also add towards the elucidation of sequence-specific
function of apoLp-III in challenging areas of pest and disease control. We hypothesize
here that due to their greater expression in the late pupal stages, apoLp-III can be
significant in studies involving the pupal to adult metamorphosis and deficiencies in
the adult phenotype. In the near future, we can look forward to the development of
further methods and increased usage of all these techniques in the new era of functional
genomics, proteomics and bioinformatics. The wealth of structural and functional
knowledge will undoubtedly facilitate new avenues and advancements in the
functional characterization of apoLp-III and impending needs in immunity, pattern
recognition and programmed cell death.

9. Acknowledgements
This work was supported by the Next-Generation Bio Green 21 Program
(No. PJ008186) of the Rural Development Administration, Republic of Korea.

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____________________________________________________________________________________________
Article History: Received 15th June 2013; Revised 10th December 2013 and Accepted 5th January 2014; and
Published 30th October 2014.

Lepidopteran larvae and pupae

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Table Contents

SION
MIS

TERNA
IN
T

N AL B OO

IO

Page No.

Preface
Forward message
Contributors
Reviewers
Acknolwedgement

i
ii
iii
iv
v

Volume1
Section I: Insect Biochemical approaches

1. Introduction to Insect Molecular Biology.

Raman Chandrasekar, P.G., Brintha, Enoch Y.Park, Paolo Pelsoi, Fei Liu,
Marian Goldsmith, Anthony Ejiofor, B.R., Pittendrigh, Y.S., Han,
Fernando G. Noriega, Manickam Sugumaran, B.K., Tyagi, Zhong Zheng Gui,
Fang Zhu, Bharath Bhusan Patnaik, and P. Michailova

2.

Modulation of Botanicals on pests biochemistry.

57

Sahayaraj, K.

3.

Detoxication, stress and immune responses in insect antenna:

new insights from transcriptomics.

75

David Siaussat, Thomas Chertemps and Martine Maibeche

4.

Application of isotopically labeled compounds and tandem mass

spectrometry for studying metabolic pathways in mosquitoes.

99

Stacy Mazzalupo and PatriciaY.Scaraffia

5.

Field Response of Dendroctonus armandi Tsai & Li (Coleoptera:

Scolytinae) to Synthetic Semiochemicals in Shaanxi, China.

127

Shou-An Xie, Shu-Jie L.V., Hui-Chen, Raman Chandrasekar

xvii

Section II: Insect Growth

6. Insect Cuticular SclerotizationHardening Mechanisms and Enzymes.

149

Manickam Sugumaran

7. New Approaches to Study Juvenile Hormone Biosynthesis in Insects.

185

Crisalejandra Rivera-Perez, Marcela Nouzova and Fernando G. Noriega

8. The regulatory biosynthetic pathway of juvenile hormone.

217

Zhentao Sheng and Raman Chandrasekar

Section III:

Insect Immunity

9. The innate immune network in a hemimetabolous insect, the brown

planthopper, Nilaparvata lugens.

233

Yanyuan Bao, Raman Chandrasekar, Chuan-Xi Zhang

10. Immune Pathways in Anopheles gambiae.

253

Maria L. Simes and Raman Chandrasekar

11. Key biochemical markers in silkworms challenged with immuno-

271

elicitors and their association in genetic resistance for survival.

Somasundaram, P., Chandraskear, R., Kumar,K.A., and Manjula, A.

Section IV:

Insect Molecular Genetics

12. The recent progress of the W and Z chromosome studies of the

291

silkworm, Bombyx mori

Hiroaki Abe, Tsuguru Fujii and Raman Chandrasekar

13. Molecular characterization and DNA barcoding for identification of

317

agriculturally important insects.

Rakshit Ojha, Jalali, S.K., and Venkatesan, T.

14. Polytene chromosomes and their significance for Taxonomy,

331

Speciation and Genotoxicology

Paraskeva V. Michailova

15. Insect exuvium extracted DNA marker: a good complementary

molecular taxonomic characteristics with special reference
to mosquitoes.

355

Dhanenjeyan, K. J., Paramasivam, R., Thanmozhi, V., Chandrasekar,R., and Tyagi, B.K.
Index

363

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Volume2
Section V:

Molecular Biology of Insect Pheromones

16. Understanding the functions of sex-peptide receptors?

373

Orly Hanin, Ada Rafaeli

17. Current views on the function and evolution of olfactory receptors

385

in Lepidoptera.

Arthur de Fouchier, Nicolas Montagn, Olivier Mirabeau, Emmanuelle Jacquin-Joly

18. Molecular architecture, phylogeny and biogeography of pheromone

409

biosynthesis and reception genes / proteins in Lepidoptera.

Jian-Cheng Chang, P. Malini, R. Srinivasan

Section VI:

Insect Molecular Biology

19. Application of Nanoparticles in sustainable Agriculture :

429

Its Current Status.

Atanu Bhattacharyya , Raman Chandrasekar, Asit Kumar Chandra,

Timothy T. Epidi and Prakasham, R.S.

20. Mosquito Ribonucleotide Reductase: A Site for Control.

449

Daphne Q.-D. Pham, Victor H. Perez, Lissette Velasquez, Dharty Bhakta,

Erica L. Berzin, Guoli Zhou, and Joy. J. Winzerling.

21. Green protocol for synthesis of metal nanoparticles

to control insect pests.

473

Murugan, K., Chandrasekar, R., Panneerselvam, C., Naresh Kumar, A.,

Madhiyazhagan, P., Mahesh Kumar, P., Jiang-Shiou Hwang, Jiang Wei

22. Aquaporins in Blood-Feeding Arthropods.

497

Lisa L. Drake, Hitoshi Tsujimoto, Immo A. Hansen

23. Mimetic analogs of three insect neuropeptide classes

509

for pest management.

Ronald J. Nachman

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Section VII:

Insect Pest Management through

Biochemical and Molecular approaches

24. Induced resistance in plants against insect pests and

533

counter-adaptation by insect pests.

Abdul Rashid War and Hari C Sharma

25. Insect Chemical communication - an important component of

549

novel approaches to insect pest management.

Usha Rani, P.

26. Mosquito control using biological larvicides: Current Scenario.

575

Subbiah Poopathi, C. Mani and R. Chandrasekar

27. Application of RNAi toward insecticide resistance management.

595

Fang Zhu, Yingjun Cui, Douglas B. Walsh, Laura C. Lavine

Section VIII:

Insect Bioinformatics

28. Entomo-informatics: A prelude to the concepts in Bioinformatics.

621

Habeeb, S.K.M. and Raman Chandrasekar

29. Molecular expression and structure-function relationships of

633

apolipophorin III in insects with special reference to innate immunity.

Bharat Bhusan Patnaik, Raman Chandrasekar, Yeon Soo Han

30. Computer-aided pesticide design: A short view

685

Jitrayut Jitonnom

Index

709

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ISBN No. 978-1-63315-205-2 (USA)

First Edition: Volume 1, 2 October 2014

Total No. Pages: 398 + 372 = 770

Edited by Raman Chandrasekar

B.K. Tyagi
Zhong Zheng Gui
Gerald R. Reeck
Copyright Reserved
Published by International Book Mission, Academic Publisher, South India.

Printed in the K-State Union, Copy and Printing services,

Kansas State University, Manhattan 66506, KS, USA.
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Book Mission Project # 2: Initiated on June 2010; Completed on March 2014 and Published on Oct. 2014.

Volume 1 & 2, October 2014

Short Views on Insect Biochemistry

and Molecular Biology

PREFACE
Entomology as a science of inter-depended branches like biochemistry, molecular entomology, insect
biotechnology; has made rapid progress in its attributes in the light of modern discoveries. This also
implies that there is an urgent need to manage the available resources scientifically for the good of man.
In the past five decades, entomology in the world/country has taken giant steps ahead. Continued
research has evolved better pest management through molecular approaches. The aim of the Short
Views on Insect Biochemistry and Molecular Biology book is to integrate perspectives across
biochemistry and molecular biology, physiology, immunology, molecular evolution, genetics,
developmental biology and reproduction of insects. This century is proclaimed as the Era of
Biotechnology and its consists of all types of Mol-Bio applications, which is an essential component for
a through understanding of the Insect Biology. This volume 1 & 2 (8 section with 30 chapters)
establishes a thorough understanding of physiological and biochemical functions of proteins, genes in
insects life processes; the topics dealt with in the individual chapters include chemistry of the insect
cuticle, hormone and growth regulators; biochemical defenses of insects; the biochemistry of the toxic
and detoxification action; modern molecular genetics and evolution; inter- and intra-specific chemical
communication and behavior; insect pheromone and molecular architecture, phylogeny and chemical
control of insect by using insect pheromones biotechnology; insect modern biology and novel plant
chemical and microbial insecticides for insect control, followed by a discussion of the various
mechanisms of resistance (both behavioral and physiological) and resistance management; modern insect
pest management through biochemical and molecular approaches; Mimetic analogs of insect
neuropeptide for pest management; entomo-informatics and computer-aided pesticide designing. In short
this book provides comprehensive reviews of recent research from various geographic areas around the
world and contributing authors area recognized experts (leading entomologist/scientist) in their
respective filed of molecular entomology. We will miss this collaboration now it has ended, but will feel
rewarded if this book is appreciated by our team/colleagues and remarkable mile stone in entomology
field.
This book emphasizes upon the need for and relevance of studying molecular aspects of entomology in
Universities, Agricultural Universities and other centers of molecular research. To encompass this
knowledge and, particularly disseminate it to the scientific community free of cost, was the major
inspiring force behind the launch of Short Views on Insect Biochemistry and Molecular Biology.

Editors

Raman Chandrasekar
Brij Kishore Tyagi

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ShortViewson

InsectBiochemistryand
MolecularBiology
Editedby
Raman Chandrasekar, Ph.D.,
Kansas State University, USA.
B.K.Tyagi, Ph.D.,
Centre for Research in Medical Entomology (ICMR), India.
Zhong Zheng Gui, Ph.D.,
Jiangsu University of Science and Technology,
Sericultural Research Institute, Chinese Academy of
Agricultural Sciences, China.
Gerald R. Reeck, Ph.D.,
Kansas State University, USA.

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Contributing Authors

Dr. B.K.Tyagi

Prof.Fernando G. Noriega

Centre for Research in Medical Entomology,

4Sarojini Street, Chinna Chokkikulam,
Madurai 625002 (TN), India

Department of Biological Sciences

HLS 227, Florida International University
11200 SW 8th St, Miami, FL 33199, USA.

Prof. Gui Zhongzheng

Dr. Zhentao Sheng

Sericultural Research Institute,

Chinese Academy of Agricultural
Sciences, Zhenjiang, 212018,
Jiangsu, P. R. China.

Chicogo University, Chicogo, USA.

Prof. K. Sahayaraj

Prof.Yanyuan Bao
Institute of Insect Science,
Zhejiang University, China.

Dept. of Advanced Zoology and Biotechnology,

St. Xavier's College
Palayamkottai 627 002, Tamil Nadu, India.

Prof. Chuan-Xi Zhang,

Prof. David Siaussat

Dr. Maria L. Simes

Universit Pierre et Marie Curie (Paris 6/UPMC),

UMR 1272A Physiologie de l'Insecte:
Signalisation et Communication (PISC),
7 Quai Saint Bernard, Batiment A - 4me tage bureau 410, 75252 Paris Cedex 05, France.

Prof. PatriciaY.Scaraffia
Department of Tropical Medicine,
Tulane University, New Orleans,
LA 70112, USA.

Prof. Shou-An Xie

Institute of Insect Science,

Zhejiang University, China.

UEI Parasitologia Mdica,

Centro de Malria e Outras Doenas Tropicais,
Instituto de Higiene e Medicina Tropical,
Rua da Junqueira 96, 1300 Lisboa,
Portugal.

Dr. P. Somasundaram
Central Sericultural Germplasm Resources Centre,
P.B.No.44, Thally Road,
Hosur-635109,
Tamilnadu, India.

College of Forestry,
Northwest A & F University
Yangling, Shaanxi 712100, China

Dr. Hiroaki Abe

Dr. Raman Chandrasekar

Dr. S.K. Jalali

Department of Biochemistry and Molecular

Biophysics, Kanas State University,
Manhattan, 66506, KS, USA.

Prof. Gerald R. Reeck

Department of Biochem. and Molecular
Biophyscis, Kansas State University, KS, USA.

Prof. Manickam Sugumaran

Department of Biology
University of Massachusetts Boston
100 Morrissey Blvd,
Boston, MA 02125, USA.

Tokyo University of Agriculture and Technology,

Japan.

National Bureau of Agriculturally Important

Insects, ICAR, India.

Prof. Paraskeva V. Michailova

Institute of Biodiversity and
Ecosystem Research,
1 Tzar Osvoboditel boulv
Bulgarian Academy of Sciences
Sofia 1000, Bulgaria.

Prof. Ada Rafaeli

Associate Director for Academic Affairs &
International Cooperation
Agricultural Research Organization,
The Volcani Center, P. O. Box 6,
Bet Dagan 50250, Iseral.

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Prof. Emmanuelle Jacquin-Joly

Dr. Fei Liu

UMR PISC Physiologie de l'insecte

INRA, Route de Saint-Cyr
78026 Versailles cedex, France..

Department of Biological Science & Technol.,

Shaanxi Xueqian Normal University,
Shaanxi, China.

Dr. R. Srinivasan

Prof. Marian Goldsmith

Entomologist and Head of Entomology Group

AVRDC-The World Vegetable Center
60 Yi Ming Liao, Shanhua
Tainan 74151, Taiwan.

Biological Sciences Department,

University of Rhode Island,
Kingston, RI 02881, USA

Prof. Atanu Bhattacharyya

Prof. Anthony Ejiofor

Vidyasagar College for Women,

Post Graduate Department of Environmental
Science,
University of Kolkata, India.

Department of Biological Sciences,

College of Agriculture, Human & Natural
Sciences, Tennessee State University,
3500 John A Merritt Blvd., Nashville,
Tennessee 37209, USA.

Prof. Daphne Q.-D. Pham

Dr. Bharath Bhusan Patnaik

Dept of Biological Sciences,

University of Wisconsin-Parkside,
900 Wood Road, Kensoha,
WI 53144, USA.

School of Biotechnology,
Trident Academy of Creative Technology
(TACT), Bhubaneswar 751013 Odisha, India.

Prof. Jitrayut Jitonnom

Prof. B.R. Pittendrigh

School of Science
University of Phayao, Thailand.

Department of Entomology,
University of Illinois, Urbana-Champaign, IL,
61801, USA
.

Prof. K. Murugan

Dr. Subbiah Poopathi

Department of Zoology, School of Life Sciences,

Bharathiar University,
Coimbatore - 641 046, India.

Prof. Immo A. Hansen

Department of Biology,
New Mexico State University,
Las Cruces, NM, USA.

Dr. Ronald J. Nachman

USDA-ARS,
Food Animal Protection Research Laboratory,
USA.

Dr. Hari C Sharma

International Crops Research Institute for the
Semi-Arid Tropics (ICRISAT), Patancheru502324,
Andhra Pradesh, India.

Prof. Paolo Pelsoi

State Key Laboratory for Biology Plant Diseases
and Insect Pests, Institute of Plant Protection,
Chinease Academy of Agricultural Sciences,
Bejing, China.

Unit of Microbiology and Immunology,

Vector Control Research Centre
(Indian Council of Medical Research),
Medical complex, Indira Nagar,
Puducherry 60 5006, India.

Dr. P.Usha Rani

Biology and Biotechnology Division
Indian Institute of Chemical Technology
(CSIR)Taranaka,
Hyderabad - 500 007 (AP), India.

Dr. Fang Zhu

Irrigated Agriculture Research and Extension
Center, Dept.of Entomology,
Washington State University,
Prosser, WA, USA.

Prof. S.K.M. Habeeb

Department of Bioinformatics,
Faculty of Engineering & Technology,
SRM University, Kattankulathur,
Chennai 603203, Tamilnadu, India.

Prof. Yeon Soo Han

Division of Plant Biotechnology,
College of Agriculture & Life Science,
Chonnam National University,
Gwangju 500-757, South Korea

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Reviewer & External supportive members

Prof. Michael Riehle, Department of Entomology, University of Arizona, USA.

Dr. Dawn L.Geiser, College of Agriculture and Life Sciences, University of Arizona, USA.
Prof. Young Jung Kwon, School of Applied Biosci., Kyungpook National University, South Korea.
Dr. Kaliappandar Nellaiappan, CuriRx Inc. USA.
Prof. Patricia Y. Scaraffia, Department of Tropical Medicine, Tulane University, USA.
Prof. Richard Newcomb, Plant & Food Research, University of Auckland, New Zealand.
Dr. S. Krishnaswamy, School of Biotechnology, Madurai Kamaraj University, South India.
Dr. Mary-Anne Hartley, University of Lausanne, Switzerland.
Dr. Igor F. Zhimulev, Institute of Molecular and Cellular Biology, Novosibirsk, Russia.
Dr. S. Subramanin, Indian Agricultural Research Institute. India.
Prof. Gustavo F. Martins, Departament de Biologia Geral, Universidade Federal de Vicosa, Brazil.
Prof. Helena Janols, Infektionsklinien, Skanes Universitetsisjukhus, Sweden.
Prof. Donald R.Barnard, USDA, Agricultural Research Service, CMAVE, USA.
Dr. Keith White, Faculty of Life Science, University of Manchester, UK.
Prof. Marten J.Edwards, Biology Department, Muhlenberg College, USA.
Prof. E. Warchalowska-Sliwa, Polish Academy of Sciences, Poland.
Dr. K. Balakrishnan, Department of Immunology, Madurai Kamaraj University, India.
Dr. J.Joe Hull, USAD-ARS, Arid Land Agricultural Research Centre, USA.
Dr. Neil Audsley, The Food & Environment Research Agency, UK.
Dr. Raman Chandrasekar, Kansas State University, USA.
Dr. B.K. Tyagi, Centre for Research in Medical Entomology (ICMR), Madurai, TN, India.
Prof. Zhongzheng Gui, Sericulture Research Institute, Chinese Academy of Agricultural Sci., China.
Dr. Fang Zhu, Irrigated Agril. Research and Extension Center, Washington State University, USA.
Prof. K. Murugan, Department of Zoology, Bharathiar University, Coimbatore, India.
Dr. Xiao-Wei Wang, Institute of Insect Science, Zhejiang University, China.
Dr. Haijun Xu, Institute of Insect Science, Zhejiang University, China.
Dr. Alisha Anderson, CSIRO Ecosystem Sciences, Australia.
Prof. Eric D.Dodds, Department of Chemistry, University of Nebraska-Lincoln, USA.
Prof. P. Mosae Selvakumar, Department of Chemistry, Karnaya University, Coimbatore, India.
Prof. A.K.Dikshit, Indian Agriculture Research Institute, New Delhi.
Prof. K.R.S. Sambasiva Rao, Dept. of Biotech. & Zoology, Acharya Nagarjuna University, India
Dr. R. Rangeshwaran, National Bureau of Agriculturally Important Insects, Banglore, India.
Dr. V. Selvanarayanan, Faculty of Agriculture, Annamalai University, Tamil Nadu, India.
Prof. Fernando G. Noriega, Florida International University, Miami, USA.
Prof. Ada Rafaeli, Department of Food Quality and Safety, A.R.O., Israel.
Prof. Daphne Q.-D. Pham, Dept. of Biological Sciences, University of Wisconsin-Parkside, USA.
Prof. Emmanuelle Jacquin-Joly, INRA, UMR 1272 Physiologie de lInsecte, Versailles, France.
Prof. Manickam Sugumaran, University of Massachusetts Boston, USA.
Prof. Nannan Liu, Auburn University, USA.
Prof. Michihiro Kobyashi, Nagoya University, Japan.
Prof. Enoch Y.Park, Innovative Joint Research Center, Shizuoka University, Japan.
Prof. Luiz Paulo Moura ANDRIOLI, Universidade de So Paulo, SP - Brazil
Prof. SHIMADA Toru, The University of Tokyo, Japan.
Prof. Erjun Ling, Institute of Plant Physiology and Ecology, China.

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Acknowledgements
Writing and publishing a book requires the assistance of individuals who are
creative, talented, and hard-working. All of these qualities were present in the
individuals assembled to produce this book volume. I would like to express my
heartfelt gratitude to my former teacher Prof. Seo Sook Jae, (GSNU, South Korea),
Prof. Subba Reddy Palli (University of Kentucky, USA), and other external mentors
Prof. Marian R. Goldsmith (University of Rhode Island, USA), Prof. Enoch Y. Park
(Shizuoka University, Japan), Prof. M. Kobayashi (Nagoya University, Japan), Prof.
CHU Jang Hann (National University of Singapore, Singapore), Prof. Thomas W.
Sappington (USDA-ARS, USA), Prof. Fernando G. Noriega (Florida International
University, USA), Dr. Srinivasan Ramasamy, AVRDC, The World Vegetable
Center, Taiwan), Dr. H.C. Sharam (ICRISAT, India), who inspiration and
supported me at many ways for the commencement of this International Book
Mission Program. The book mission program was initiated on May 2010,
completed on March 2014 and published on October 2014. I have no words to
express my feeling for all those who provided valuable contributions from USA,
South Korea, Japan, China, India, Thailand, Taiwan, Bulgaria, France, Iseral, and
Portugal (Contributors name list, see page no. v) and made the completion of this
book possible. We express our appreciation to the following people (Reviewer
name list, see page no. vii) who reviewed various part of the manuscript as it was
being developed and improved quality of each chapter. I thank the ICMR, New
Delhi, and Chinese Academy of Agricultural, China, and Kansas State University for
support from several aspects. Many others (scientists and publishers) have also
allowed us to use their materials in the various chapters, their color image have then
been converted to gray color/BW. Iam especially indebted to International Book
Mission Organization, Academic Publishing Services for the production of book. I
thank my Co-Editors for their continuous vigilance over the book project and for
always giving advance notice of the editing and proofreading schedules. I thank also
my Brintha, P.G., (my wife), who in all possible way, encouragement helped
transform our original efforts into an acceptable final form. I apologize to those
whose work could not be cited owing to space considerations limitation. Further, I
wish to recognize the moral support extended by colleagues and friends. I hope that
this volume will inspire interest on the diverse aspects of insect biochemistry and
molecular biology in aspiring and established scientists.
Raman Chandrasekar

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A Note from the Publisher

Dear Readers,
This edition represents the first number of the Short Views on Insect
Biochemistry and Molecular Biology book series published by International
Book Mission. It serves to show the public how important entomology field in
expanding basic knowledge or in the development of new technologies nowadays,
in virtually all fields of knowledge. We called for piece of work falling into two
volumes (Basic and Advance aspects).
Far from being complete, the 30 chapters clearly structured and simply explained
experts contributions may provide an overview about current and prominent
advances in insect biochemistry and molecular biology which will help students and
researchers to broaden their knowledge and to gain an understanding of both the
challenges and the opportunities behind each approach.
We look forward to receiving new proposals for the new edition 2015 - 2017.
International Book Mission
Academic Publisher
Manager

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