96 R. B. Veegala and S. Vemuri
pest in many countries, namely India, Korea,
Japan, China, Malaysia, Sri Lanka, and Vietnam
(Dale 1994 ). The over-use of broad-spectrum
insecticides, such as methyl parathion, monocro-
tophos, and endosulfan has been cited as a major
cause of outbreaks of C. medinalis because ex-
cessive spraying of insecticide disrupts natural
control (Dale 1994 ). Insecticides still remain the
major control tactics against leaf folder. Carbo-
furan and fenthion (Chandramohan and Jayaraj
1976 ), bendiocarb, acephate and carbosulfan,
quinolphos, monocrotophos, and phosphamidon
(Raju et al. 1990 ) and fenvalerate (Ramaraju and
Natarajan 1997 ) were the common insecticides
used against rice leaf folder.
Genetically modified rice lines expressing
B. thuringiensis insecticidal crystal proteins
have been produced that are highly tolerant to
lepidopteran pests. In China, three GM rice lines
transformed with cry1Ac/cry1Ab genes(GM
Minghui 63), cry1Ac/CpTI genes (GM Minghui
86) and cry1Ab genes(GM Kemingdao) effective
against Scirpophaga incertulus, C. medinalis,
and Chilo suppressalis have been tested both at
field and laboratory levels and are on the verge
of commercialization (Tu et al. 2000 ; Ye et al.
2003 ; Han et al. 2007 ). Though economic and
environmental benefits of GM crops are well
established, a matter of concern is the possibil-
ity of the target insect pest developing resistance
to B. thuringiensis insecticidal toxins. Though a
couple of resistance mechanisms have been re-
ported for conferring resistance to B. thuringien-
sis viz. reduced binding of crystal toxins to the
brush border membrane vesicles (BBMVs) of
midgut epithelium and alteration in the midgut
proteases that cleave the protoxin to active toxin.
A new resistance mechanism to B. thuringiensis
Cry toxins is identified and is associated with
increased activity of midgut carboxylesterase ac-
tivity (Gunning et al. 2005 ). The involvement of
glutathione-S-transferase, carboxylesterase, and
microsomal mono-oxygenase in insecticide re-
sistance has been reported in insecticide-resistant
strains of many insect species. Carboxylesterases
(CES, EC 3.1.1.1) are members of a superfamily
of serine hydrolases that hydrolyze ester, amide,
and carbamate bonds. Several different carbo-
xylesterase (CarE) genes exist with evidence of
multiple gene duplication in insects. Esterases
hydrolyze ester bonds from various substrates
with a carboxylic ester. Esterases are frequently
implicated in the resistance of insects to organo-
phosphorus, carbamates, pyrethroids, neonicoti-
noids, and to many other new classes of insec-
ticides through gene amplification, upregulation,
coding sequence mutations, or a combination of
these mechanisms (Li et al. 2007 ).Material and Methods
Two field populations of C. medinalis were col-
lected from the Directorate of Rice Research
(DRR), Rajendranagar and ICRISAT, Patancheru.
C. medinalis adults were collected from rice fields
during the boot leaf stage in rabi 2011. The col-
lected adults were released into pots containing
TN-1 plants for egg-laying and were covered with
a muslin cloth for aeration, 20 % honey solution
was also provided for feeding. Ten pairs of C. me-
dinalis adults were released into each TN-1 pot.
C. medinalis populations from different locations
were reared separately and after larval hatching
the third instar larva was used for bioassay.
The leaf-dip bioassay method was used.
Three–four long tender leaves from TN-1 were
used. The leaves were first washed with distilled
water and were then dipped in monocrotophos so-
lution and thoroughly air-dried for about 10 min,
different concentrations of the insecticide were
prepared, and bioassays were carried out first at
tenfold variation. Based on 20–80 % mortality,
concentrations were prepared in a narrow range of
fivefold for further bioassays. Six concentrations
were tested with ten third instar larvae per treat-
ment and replicated thrice. Larvae were allowed
to feed on insecticide treated leaves for 24 h and
mortality was recorded for 24 h after treatment.
Control treatments with larval mortality more
than 20 % were discarded and bioassays were
repeated. Statistical analysis for calculating the
LC 50 values for the bioassay was estimated using
maximum likelihood program MLP 3.01 (Ross
1987 ). The corrected percentage mortality was
calculated using Abbott’s formula (Abbott 1925 ).