MSc

Spider mites of the family Tetranychidae are considered as one of the
most serious sucking pests of vegetable crops worldwide. Intensive use of
conventional acaricides had lead to the development of resistance in many
mite species around the globe. In view of this, several novel acaricides with
unique chemical structure and mode of action were introduced and
commercialized for mite management.
In Kerala, mite management in vegetable crops solely depends on the
use of novel acaricides. Of late, several farmers have raised concern over
their poor efficacy against mite pests. Spider mites have the ability to develop
resistance quickly on continuous exposure to a pesticide. In this context, the
present study was undertaken to investigate the status, biochemical and
molecular bases of acaricide resistance in Tetranychus truncatus Ehara, the
predominant species of spider mite infesting vegetable crops of Kerala.
Purposive surveys were conducted in the vegetable fields of
Vellanikkara, Thrissur and spider mites were collected and reared in the
laboratory by assigning accession numbers. Three accessions/strains viz.,
VkOk1 (okra), VkAm3 (amaranthus) and VkPm3 (pumpkin) which were
identified as T. truncatus were used for the study.
Susceptibility of the three field strains to three commonly used
acaricides, viz., spiromesifen, fenazaquin and diafenthiuron was evaluated in
the laboratory following leaf dip bioassay in comparison with a laboratory
maintained susceptible strain (SS). Bioassay study revealed that the strain
VkOk1 recorded highest LC50 value and has developed 8, 13 and 10 fold
resistance to spiromesifen, fenazaquin and diafenthiuron, respectively. This
was followed by VkAm3 which showed 7.0, 5.53 and 1.67 fold resistance,
while VkPm3 recorded 1.35, 1.13 and 1.03 fold resistance.
The activity of the detoxifying enzymes viz., cytochrome P450 and
carboxylesterase, was significantly higher in VkOk1strain followed by
VkAm3. The strains VkOk1, VkAm3 and VkPm3 showed 2.69, 1.24 and
1.09 fold enhanced activity of Cytochrome P450, respectively compared to
SS, while carboxylesterase in VkOk1 and VkAm3 showed an increased
activity by 2.59 and 1.18 fold. However, the strain VkPm3 recorded a
decrease in activity of carboxyleasterase by 0.78 fold compared to the
susceptible strain.
DNA isolated from the two resistant strains (VkOk1 and VkAm3) and
the susceptible strain (SS) was amplified with the help of gene specific
primers for cytochrome P450 and carboxylesterase. The results of PCR for
cytochrome P450 gene showed that there was no amplification in the case of
SS, whereas there were distinct markers in the resistant strains, okra and
amaranthus at 1300 bp size. However, PCR amplification showed distinct
markers for carboxyl esterase in all the three strains. The strains VkOk1 and
SS showed markers at 1500 bp and 1300bp size respectively, while VkAm3
strain showed both the markers.
The sequence homology search by BLASTn analysis showed that the
sequences of cytochrome P450 of T. truncatus has similarities with
cytochrome P450 sequences from different species of spider mites with an
identity match ranging from 81 to 97 per cent, while carboxylesterase
sequences showed similarity with two mRNA sequences of carboxylesterase
of T. urticae. Further, the translated sequences of cytochrome P450 and
carboxylesterase aminoacids from T. truncatus when analysed by BLAST P
showed similarity with the amino acid sequences from other spider mites.
The sequences of carboxylesterase genes from the resistant and susceptible
strains did not align together, showing that there are two different
caboxylesterase genes controlling the resistance to acaricides in T. truncatus.
The study recorded resistance in T. truncatus to three novel acaricide
molecules, for the first time from India. This is the first report of cytochrome
P450 and carboxylesterase genes conferring resistance to acaricides in T.
truncatus. The study also developed standard markers for discriminating the
resistant and susceptible population in T. truncatus.