Mechanisms of resistance in Brinjal Shoot and fruit broer, Leucinodes orbonalis (Guenee) (Lepidoptera:Crambidae) to diamide insecticides
By: Anu Thomas.
Contributor(s): Smitha, M S(Guide).
Material type:
Item type | Current location | Collection | Call number | Status | Date due | Barcode |
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KAU Central Library, Thrissur Theses | Thesis | 632.6 ANU/ME Ph.D (Browse shelf) | Not For Loan | 176238 |
Ph.D
The development of insecticide resistance among insect pests is a major concern
in pest management. Generating data on the baseline susceptibility of field populations
to insecticides facilitates to track the resistance development in insects. This helps in
designing suitable insecticide resistance management (IRM) strategies; thereby, to
delay the development of resistance and extend the useful life of an insecticide.
Understanding the mechanisms of resistance, the possibilities of developing cross- and
multiple-resistance, and the fitness costs involved in resistance are also crucial to
maintain the sustainability of an insecticide. In this context, the present investigation
entitled “Mechanisms of resistance in brinjal shoot and fruit borer, Leucinodes
orbonalis (Guenee) (Lepidoptera: Crambidae) to diamide insecticides” was undertaken
during 2019-2024 with the objectives to study the baseline susceptibility of brinjal
shoot and fruit borer to diamide insecticides, to assess the possible mechanisms of
resistance development, to investigate the potential of developing cross and multiple
resistance in diamide-resistant populations and to analyze the fitness costs associated
with resistance.
Field populations of brinjal shoot and fruit borer were collected from brinjal
fields of Kerala, Kullarayanpalayam (Palakkad) and Anchal (Kollam); Tamil Nadu,
Devarayapuram (Coimbatore) and Trichy (Tiruchirappalli); and Karnataka,
Heggadadevankote (Mysore) and Doddaballapur (Banglore Rural). A laboratorysusceptible
population, maintained without any insecticide exposure since 2012, was
procured from NBAIR, Bangalore. The collected populations were assigned unique
accession codes such as PKD, KLM, CMB, TRY, MYS, BAN, and Lo-S, respectively,
and maintained separately in the laboratory. Information on insecticide usage pattern in
brinjal cultivation obtained from farmers at collecting sites revealed the usage of
insecticides of different modes of action and application above recommended dosages.
An intensive application of various insecticdes including diamides was practiced in
Bangalore Rural, Tiruchirappalli, Coimbatore, and Palakkad. In Kollam, on the other
hand, a need-based application in recommended doses was followed. In Mysore, the
farmers did not use any diamide insecticide; instead, they relied on organophosphates
and neonicotinoids applied at frequent intervals.
The laboratory bioassay of field populations of L. orbonalis was performed
against flubendiamide (Fame 39.35% SC) and chlorantraniliprole (Coragen18.5% SC)
to determine the median lethal concentration (LC50). The resistance ratio (RR) was
assessed by comparing the LC50 values with that of the Lo-S population. All field
populations of L. orbonalis except MYS displayed a significant shift in LC50 value
compared to the Lo-S population and were considered resistant to flubendiamide and
chlorantraniliprole based on the hypothesis of equality. Among the resistant
populations, the BAN and TRY were resistant homozygotes according to the hypothesis
of parallelism. The LC50 value of field populations ranged from 0.83 ppm in MYS to
544.07 ppm in TRY for flubendiamide compared to 0.50 ppm recorded with the Lo-S
population. The TRY population showed the highest resistance ratio (RR) of 1079.5-
fold to flubendiamide, followed by the BAN (845.85-fold), CMB (532.56-fold), and
PKD (218.84-fold) populations. However, the KLM population showed comparatively
less RR with 47.59-fold resistance, while the MYS population was considered
susceptible to flubendiamide. The LC50 value for chlorantraniliprole ranged from 0.169
ppm in MYS to 116.80 ppm in BAN in contrast to 0.119 ppm recorded with the Lo-S
population. Concerning chlorantraniliprole, the highest RR of 979.22-fold was
recorded with the BAN population followed by 788.18-fold in TRY, 480.30-fold in the
CMB, 244.75-fold in PKD, and 18.22-fold in KLM populations. The lowest RR of
1.41-fold was observed with MYS population and was considered susceptible
to chlorantraniliprole based on the hypothesis of equality.
The activity of detoxifying enzymes in the field populations of L. orbonalis was
quantified and compared with the Lo-S population to examine the role of detoxifying
enzymes in diamide resistance. The field populations, BAN, PKD, CMB, TRY, MYS,
and KLM showed 1.66-, 1.47-, 1.24-, 1.21-, 1.21-, and 1.17-fold increase in
carboxylesterase (CarE) activity, respectively. Significantly higher titers of
cytochrome P450 (Cyt P450) activity were displayed by all the field populations with
a 5.54- fold increase in the TRY population, followed by BAN (5.38-fold), CMB (5.26-
fold), PKD (5.13-fold), MYS (4.56-fold), and KLM (3.12-fold). The relative activity
of glutathione S-transferase (GST) was >4-fold in PKD, CMB, TRY, and BAN
populations displaying 4.31, 4.33, 4.34, and 4.44-fold variations, respectively,
compared to the Lo-S population. Meanwhile, KLM and MYS populations exhibited
only 1.5-fold and 1.25-fold increases, respectively.
To assess the potential for developing cross-resistance, a laboratory bioassay
was carried out against cyantraniliprole, a less commonly used diamide insecticide. The
field populations displaying resistance to flubendiamide and chlorantraniliprole also
exhibited resistance to cyantraniliprole, even though the population had no previous
exposure to the chemical. However, the populations that were not previously exposed
to emamectin benzoate and spinosad did not show resistance to these insecticides, and
thereby the possibility of developing multiple resistance in diamide-resistant
populations was rejected.
Fitness costs associated with diamide resistance were ascertained by comparing
the life table parameters of different field populations of L. orbonalis with the Lo-S
population. In the study, diamide-resistant populations, TRY, BAN, PKD, and CMB
displayed significantly longer larval and pupal duration, and less oviposition period and
female adult longevity than the Lo-S population. Male adult longevity in the TRY and
BAN populations was also notably shortened compared to the Lo-S population. The
survival of egg, larva, and pupa as well as fecundity of the diamide-resistant populations
except KLM were significantly less compared to the Lo-S population. The BAN and
TRY populations exhibited a relative fitness of 0.62, suggesting a higher fitness cost
associated with diamide resistance. The CMB population displayed a relative fitness of
0.68 followed by PKD (0.74) and the lowest was in KLM (0.82). These results indicated
a survival disadvantage with the resistant populations compared to the Lo-S
population.
In investigating the molecular basis of resistance in L. orbonalis, a partial
sequence of the ryanodine receptor (RyR) gene involved in diamide resistance was
analyzed. Thymine (T) was found to be substituted with cytosine (C) in the sequences
of the BAN and TRY samples that were tested, but the Lo-S sample did not exhibit this
substitution, which is consistent with the reference sequence (Acc. No.
PQWD01009585.1) of L. orbonalis. Alignment of the protein sequences revealed a
non-synonymous amino acid alteration from isoleucine to methionine (I to M) in the
RyR of BAN and TRY, which was not observed in the Lo-S gene sequence, indicating
the crucial role of point mutation for higher diamide resistance in TRY and BAN
populations.
The study revealed a shift in susceptibility status and the development of higher
folds of resistance to diamide insecticides among the field populations of L. orbonalis.
The possible role of detoxifying enzymes and target site mutations in the development
of diamide resistance was also established. The study on cross- and multiple-resistance
provided valuable information for the selection of insecticides in IRM programs. The
involvement of fitness costs in the diamide-resistant populations indicated a negative
impact of resistance on pest fitness and the chance of resistance reversion in the absence
of insecticides.
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