Temephos resistance in two forms of Aedes aegypti and its significance for the resistance mechanism.

Aedes aegypti, at the larval stage, has been subjected to the temephos selection in laboratory. The level of temephos resistance was detected in a microplate by biochemical assay using WHO bioassay technique. The major enzyme-based resistance mechanisms involved in temephos resistance include elevated nonspecific esterase, oxidase and insensitive acetylcholinesterase. After 19 generations of temephos selection, the selected group showed resistance ratios of 4.64 and 16.92, when compared with a non-selected group and the WHO susceptible strain, respectively. The two seperated forms, type form and the pale form of Ae. aegypti showed low levels of resistance to temephos after 19 generations of selection, with resistance ratios of 4.82 and 4.07 for the type form and the pale form, respectively; when compared with the non-selected strain, 17.58 and 14.84, when compared with the WHO susceptible strain. This showed that the type form could develop higher level resistance than the pale form. The esterase inhibitor (S,S,S-tributyl phosphorotrithioate, DEF) or synergist implicated detoxifying esterase in all the temephos selected groups and the presence of elevated esterase were confirmed by biochemical assay. There were significant differences in elevated esterase activity between the temephos selected groups and the non-selected group. However no significant difference between the type form and the pale form was found. Besides the elevated esterase, there was no change in monooxygenase activity and no evidence of insensitive acetylcholinesterease for all temephos selected groups. These results suggest that temephos resistance could be developed in Ae. aegypti under selection pressure and that the main mechanism is based only on esterase detoxification.

Potential development of temephos resistance in Aedes aegypti related to its mechanism and susceptibility to dengue virus.

The addition of temephos to water containers as a larvicide against Aedes aegypti was commonly used as a part of DHF control programs. The widespread, or long-term, application of insecticides can lead to the development of mosquito resistance to the insecticides through selection pressure. This presents a problem for disease control. Therefore, this study was conducted in the laboratory to observe the potential development of resistance to temephos and the mechanism involved in Ae. aegypti, and to study the significance for dengue infection. The larvae were selected in consecutive generations. The level of resistance to temephos was detected by WHO assay technique. After 19 generations of selection, a low level of resistance was found. The resistance ratio at LC50 was 4.64 when compared with the non-selected group. The assay for major enzyme-based resistance mechanisms was done in a microtiter plate to detect elevated non-specific esterases, monooxygenase, and insensitive acetylcholinesterase in the temephos-selected and non-selected groups. It revealed a significant increase in esterase activity when compared with the non-selected group. There was no elevation of monooxygenase or insensitive acetylcholinesterase activities. However, when an esterase inhibitor (S, S, S-tributyl phosphorotrithioate, or DEF) was added to temephos and the susceptibility in the selected group was studied, the resistance ratio was reduced from 16.92 to 3.57 when compared with a standard susceptible strain (Bora Bora). This indicates that the esterases play an important role in temephos resistance. Dengue-2 virus susceptibility was studied by oral feeding to females of the temephos-selected (S19) and the non-selected groups. The dissemination rates, when the titer of virus in the blood meal was 7.30 MID50/ml, were 11.11% and 9.38% for the selected and non-selected groups, respectively. When the titer of virus in the blood meal was 8.15 MID50/ml, the dissemination rates increased to 24.24% and 33.33%, respectively. A statistical difference in viral susceptibility was not found between the two groups. This suggested that the low level of temephos resistance might not affect oral susceptibility. However, this needs further study.