Research Article, J Genet Disor Genet Rep Vol: 6 Issue: 2
Pirimiphos-methyl Resistance in Culex pipiens (Diptera: Culicidae) in Southern Tunisia
*Corresponding Author : Tabbabi A, Laboratory of Genetics
Faculty of Medicine of Monastir, Monastir University, Monastir- 5019, Tunisia
Tel: +216 97 085 424
E-mail: [email protected]
Received: April 17, 2017 Accepted: May 02, 2017 Published: May 08, 2017
Citation: Tabbabi A, Daaboub J, Laamari A, Ben Cheikh R, Ben Cheikh H (2017) Pirimiphos-methyl Resistance in Culex pipiens (Diptera: Culicidae) in Southern Tunisia. J Genet Disor Genet Rep 6:2.doi: 10.4172/2327-5790.1000155
Pirimiphos-methyl is an organophosphorus insecticide having a fast action with less toxicity for humans and environment. The aim of this study was to investigate the pirimiphos-methyl resistance of three populations of Culex pipiens in southern Tunisia. Three Culex pipiens samples were collected in 3 localities (South east, south west, and extreme south) of southern Tunisia between March 2002 and October 2005. The susceptibility of the third population (extreme south) of Culex pipiens to pirimiphos-methyl was not considered due to control-level mortality. Both other studied samples were resistant to pirimiphos methyl and to propoxur. A significantly correlation was observed between the mortality due to propoxur and the LC50 of pirimiphos methyl (P<0.01) showed an insensitive acetylcholinesterase (AChE). The characterization of Esterase phenotypes confirmed the previous studies which showed the major implication of esterases in the resistance to organophosphorus insecticides. Our results can improve current control and mosquito surveillance strategies.
Keywords: Culex pipiens; Primiphos methyl; Resistance; Insensitive AChE; Esterases; Southern Tunisia
Culex pipiens is showing locally invasive or interfering with everyday life, tourism and economic activities [1-3]. Many control methods have been tested. Many of these actions have had unintended negative or collateral consequences, and none has shown lasting results. This mosquito is favored by the fact that it is rapidly reproduced in large numbers and is highly adaptive. In particular, it has demonstrated the ability to adapt to many insecticides, which are sometimes only temporarily or partially effective [4-9], while affecting non-target species that were precisely predators of the mosquito.
The majority of synthetic insecticides are compounds that inactivate a vital target of the nervous system, resulting in death of the insect. Unfortunately, the populations of treated insects evolve very rapidly towards resistance [10,11]. In order to combat resistance, the use of insecticidal molecules directed against another target may prove effective for a more or less short time depending on the frequency of the occurrence of new adaptive mutations.
After the massive use of DDT (organochlorine), resistance was very quickly selected in the 1960s. Two other families of insecticides were then used: organophosphates (OP) and carbamates (CX). These molecules inhibit AChE, the enzyme responsible for the hydrolysis of acetylcholine in cholinergic synapses. This inhibition prolongs the nerve impulse, which leads rapidly to the death of the insect by tetany. In the mosquito Culex pipiens, two main mechanisms of resistance to these insecticides have been demonstrated: The first corresponds to an increased detoxification of OPs by carboxylesterases that trap or degrade them . In the case of resistance, these esterases are overproduced by a process of amplification or overexpression of the genes that encode them [13,14]. The second mechanism corresponds to a modification of the target, AChE, which reduces its affinity for insecticides and leads to its insensitivity [15,16].
Pirimiphos-methyl is an organophosphorus insecticide used in a wide range of vectors control and is being considered by WHO for addition to potable water in containers as a mosquito larvicide treatment. Recently, results have shown that pyrimiphos-methyl alone has little effect on larvae of Aedes aegypti whereas its efficacy on adults is particularly good . The aim of this study is to investigate the pirimiphos-methyl resistance of three populations of Culex pipiens in southern Tunisia.
Materials and Methods
Three Culex pipiens samples were collected at preimaginal stages from breeding sites in 3 localities of southern Tunisia between March 2002 and October 2005 (Table 1 and Figure 1). Comparisons of different bioassays were made with a reference strain called S-Lab . SA2 and SA5 are two resistant strains homozygous for Ester2 and Ester5, respectively . They were used for correct identification of detected esterases.
Bioassays and data analysis
Assays were realized according to Raymond et al. , using ethanol solutions of pirimiphos methyl (99% [AI]), brought from laboratory Dr Ehrenstorfer, Germany, and propoxur (99.9% [AI], Bayer AG, Leverkusen , Germany). The effect on OPs resistance of 2 synergists, the DEF (98% [AI], Chem Service, England), and the Pb (94% [AI], Laboratory Dr Ehrenstorfer, Germany) , was studied. Data were analysed by probit analysis  using a BASIC program .
Results and Discussion
S-Lab was the only strain that has linear dose-mortality response. With the exception of susceptibility to propoxur, the susceptibility of the third population (sample # 3) of Culex pipiens to pirimiphosmethyl was not considered due to control-level mortality. Both other studied samples were resistant to pirimiphos methyl. This result can be explained by the frequent control of these areas against this mosquito (frequent monitoring by the pirimiphos methyl, Table 1) because of the alerted by inhabitants about the mosquito bites. The RR50 was 6.9 in sample # 1 and 27 in sample # 2 (Table 2).
|Code||Locality||Breeding site||Date of collection||Mosquito control(used insecticides)||Agricultural pest control|
|1||Tozeur||Ditch||Oct. 2005||Frequent (C, Pm, F, P, D)||None|
|2||Gabes||Drain||June 2005||Frequent (C, Pm, P, D)||None|
|3||Borj El Khadra||Water pond||Mar 2002||Occasional (P)||None|
C : Chlorpyrifos ; T : Temephos ; Pm : Pirimiphos methyl ; F : Fenitrithion ; P : Permethrin ; D : Deltamethrin
Table 1: Geographic origin of Tunisian populations, breeding site characteristics, and insecticide control.
|Population||Pirimiphosmethyl||Pirimiphosmethyl +DEF||Pirimiphosmethyl +Pb|
|LC50 in µg/l
|3-Borj El Khadra||-||-||-||-||-||-||-||-||-||-||-||-||-|
(a): 95% CI *The log dose-probit mortality response is parallel to that of S-Lab **Parallelism test positive but without probability; RR50: resistance ratio at LC50 (RR50=LC50 of the population considered / LC50 of Slab) SR50: synergism ratio (LC50 observed in absence of synergist / LC50 observed in presence of synergist) RR and SR considered significant (P<0.05) if their 95%CI did not include the value 1 RSR: relative synergism ratio (RR for insecticide alone / RR for insecticide plus synergist)
Table 2: Pirimiphosmethyl resistance characteristics of Tunisian Culex pipiens in presence and absence of synergists DEF and Pb.
The tolerance to pirimiphos methyl in S-Lab and the two field samples decreased significantly with the addition of the DEF (SR50>1, p<0.05) (Table 2). No synergistic action was recorded by the DEF in samples # 1 and 2. Therefore, esterases (and/or GST) had no effect on the pirimiphos methyl resistance in both samples. These results confirm those found by Tabbabi et al.  which showed the noninvolvement of esterases (and/or GST) in the resistance of Culex pipiens strain subjected to temephos (organophosphorus) selection pressure. The tolerance to pirimiphos-methyl decreased significantly with the addition of the Pb in S-Lab (SR50=7.24, p<0.05) and in both samples, and the SR was not significantly higher than that recorded in S-Lab (Table 2). Therefore, the CYP450 had no effect on the pirimiphos methyl resistance in both populations. Other previous studies on Culex pipiens from Tunisia , Culex quinquefasciatus from Venezuela  and USA , and Musca domestica  showed the minor role of oxydases in resistance to chlorpyrifos insecticide (organophosphorus).
Mortality caused by propoxur was 1% in samples # 2, 46% in samples # 1 and 100% in sample # 3. A significantly correlation was observed between the mortality due to propoxur and the LC50 of pirimiphos methyl (P<0.01) showed an insensitive AChE. These results are in agreement with previous studies [9,28,29] which showed the major role of insensitive AChE in resistance to OPs insecticides. It was not possible to carry out toxicological tests on population # 3 because of control-level mortality but as it was even possible to keep samples and to study its AChE and esterases activities. The latter were very sensitive to propoxur. This can be explained by the absence of control actions in this region (occasionally by the permethrin).
With the exception of the sample # 3, all the studied samples showed the presence of one or more esterases in their electrophoretic profiles. The frequency of 0.03 and 0.56 were those of A4-B4 (and / or A5-B5) esterases found in samples # 1 and 2, respectively. The frequency of 0.03 was those of The B12 esterases observed in samples # 2. The frequency of C1 esterases was 0.28 in sample # 1. The A2B2 esterases were observed just in sample # 2 with a frequency of 0.11. Our results confirms the previous studies which showed the major implication of esterases in the resistance to OPs insecticides [4,25,27,30-36].
This work was kindly supported by the Ministry of Higher Education and Scientific Research of Tunisia by funds allocated to the Research Unit (Génétique 02/UR/08-03) and by DHMPE of the Minister of Public Health of Tunisia. We are very grateful to S. Ouanes, for technical assistance, A. Ben Haj Ayed and I. Mkada for help in bioassays, S. Saïdi, Tunisian hygienist technicians for help in mosquito collecting, and M. Nedhif and M. Rebhi for their kind interest and help.
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