Evaluation of insecticide resistance in Aedes aegypti populations connected by roads and rivers: the case of Tocantins state in Brazil

BACKGROUND The longstanding application of insecticides for vector control without periodic monitoring of the population response to these chemicals can directly drive the selection of resistant populations of vector mosquitoes. Tocantins is the newest state of the Brazilian Federation. Despite a historically high number of dengue cases, studies and monitoring data concerning insecticide resistance in the state are lacking. OBJECTIVES To verify the resistance status of Aedes aegypti from 10 populations distributed throughout the state connected by rivers and roads. METHODS Between 50 and 150 ovitraps were installed in house gardens within each municipality. Collection points were established based on the importance of the towns and on geographic aspects. Dose response bioassays were performed in accordance with World Health Organization guidelines. Molecular assays were conducted to detect kdr mutations, which are related to pyrethroid resistance. FINDINGS Of the 3,200 ovitrap paddles analysed, 25.8% contained eggs, with a total of 55,687 eggs collected. With the exception of Caseara, all evaluated populations were considered to be resistant to temephos. The data showed different levels of resistance to deltamethrin among the samples. Caseara and Guaraí showed the lowest RR95 values. On average, the NaVR1 kdr allele was most frequent (40.3%), followed by NaVS (38.1%), and NaVR2 (21.6%). Palmas, the capital of the state, had the highest frequency of kdr alleles (87.5%). MAIN CONCLUSIONS With the exception of Palmas, the towns with the highest indexes (ovitrap positivity, number and density of eggs), as well with high levels of resistance and kdr alleles were located along the BR-153 road, indicating that the flow of people and cargo can contribute to the dispersion of the vector and potentially resistance. This study contributes substantially to knowledge regarding the insecticide resistance profile of Tocantins mosquito populations; the data generated via the study could facilitate the judicious use of insecticides by vector control programs.

First investigation of deltamethrin pyrethroid susceptibility and resistance status of Anopheles labranchiae (Falleroni, 1926), potential malaria vector in Tunisia

Objective:To evaluate the deltamethrin pyrethroid insecticides against Anopheles labranchiae,potential malaria vector in Tunisia.Methods:Six field populations of Anopheles labranchiae mosquitoes were collected from six localities in Northern and Central Tunisia between October and November 2016.Different bioassays were performed to estimate the level of resistance in each collected population.Two synergists were used to estimate the involvement of detoxification enzymes in insecticide resistance.Results:All studied strains were resistant and the RRs0 ranged from 12.5 in sample #1 to 72.5 in sample #6.Synergist tests using piperonyl butoxide indicated the involvement of monoxygenases enzymes in the recorded resistance.In contrast,the increase of deltamethrin mortality was not significant in presence of S,S,sributyl phosphorothioate (0.8 ＜ SR ＜ 1.2),suggesting no role of esterases (and/or GST) in the resistance phenotype.The correlation recorded between mortality due to DDT and the LC50 of deltamethrin insecticide indicated an insensitive sodium channel affected by Kdr mutation (Spearman rank correlation,r =-0.59,P ＜ 0.01).Conclusions:These results should be considered in the current mosquitoes control programs in Tunisia.The use of pesticides and insecticides by both agricultural and public health departments in Tunisia should be more rational to reduce the development of resistance in populations.Different insecticide applications should be implemented alternately.

First investigation of deltamethrin pyrethroid susceptibility and resistance status of Anopheles labranchiae (Falleroni, 1926), potential malaria vector in Tunisia

Objective To evaluate the deltamethrin pyrethroid insecticides against Anopheles labranchiae, potential malaria vector in Tunisia. Methods Six field populations of Anopheles labranchiae mosquitoes were collected from six localities in Northern and Central Tunisia between October and November 2016. Different bioassays were performed to estimate the level of resistance in each collected population. Two synergists were used to estimate the involvement of detoxification enzymes in insecticide resistance. Results All studied strains were resistant and the RR

Mecanismos de resistencia a la permetrina en dos poblaciones de Aedes aegypti del occidente de Venezuela / Mechanisms of Permethrin resistance in two populations of Aedes aegypti in westernVenezuela

Se determinó la resistencia a la permetrina en Aedes aegypti de los estados Trujillo y Zulia mediante bioensayos de botella. Los insectos derribados durante la hora de exposición fueron registrados y usados para calcular la Concentración Knock-down cincuenta (CK50) y los muertos a las 24 horas para la Concentración Letal cincuenta(CL50). La resistencia al derribo y post-recuperación fueron determinadas calculando del Factor de Resistencia FRCK50 y FRCL50, comparando los valores de CK50 y CL50de las poblaciones de Ae. aegypti de campo con los de la cepa susceptible New Orleans obtenidos mediante análisis de regresión log-probit. Mecanismos metabólicos y no metabólicos asociados a la resistencia, fueron evaluados midiendo los niveles de las enzimas alfa-esterasas, beta-esterasas, oxidasas de función múltiple y glutatión-S-transferasas mediante la técnica de microplacas y determinando la frecuencia alélica I1016 por PCR alelo específico. Ambas poblaciones mostraron baja resistencia al derribo (FRCK50 < 5) y moderada resistencia post-recuperación (FRCL50 entre 5 y 10). Sobre-expresión de alfa-esterasas fue observada en la población Loma Linda la cual se correlacionó significativamente con la CL50. En la población Pampanito la frecuencia del alelo I1016 fue de 0,1 y en Loma Linda de 0,17, observándose homocigotos mutantes solo en esta última población. Se evidencia la presencia de mecanismos metabólicos y no metabólicos asociados a la resistencia al derribo y post-recuperación a la permetrina en las poblaciones bajo estudio, lo cual debe ser considerado antes de la aplicación de piretroides para el control de Ae. aegypti en la zona de estudio.

Permethrin resistance was determined in Aedes aegypti populations from Trujillo and Zulia states using the bottle bioassay method. Insect knock-down rates during 1h of exposure were recorded and used to calculate the 50% knock-down concentration (KC50) and the mortality after 24 h (LC50). Knock-down and post-recovery resistance were determined by calculating the resistance factors, FRKC50 and FRLC50 . This was done by comparing the KC50 and LC50 values (obtained by regression analysis log-probit) of the field populations with a susceptible New Orleans strain. Metabolic and non-metabolic mechanisms associated with resistance were assessed by measuring the levels of the following enzymes: alpha-esterases, beta-esterases, mixed function oxidases (MFOs) and glutathione-S-transferases (GSTs) using the microplate technique. We also determined the alellic frequency of I1016 by allele specific PCR. Both populations showed a low knock-down resistance (FRKC50 < 5) and moderate post-recovery resistance (FRLC50 between 5 and 10). Overexpression of alpha-esterases was observed in the Loma Linda population and was significantly correlated with the LC50 . The frequency of the I1016 allele was 0.1 for the Pampanito population and 0.17 for the Loma Linda population, and in the latter we also observed homozygous mutants. The existence of metabolic and non-metabolic mechanisms associated with knockdown resistance and post-recovery to permethrin in the populations studied was demonstrated. This should be taken into account before introducing these insecticides to control populations of Ae. aegypti in the region.

Multiple insecticide resistances and kdr mutation of Culex pipiens pallens in north-central Anhui Province / 中国血吸虫病防治杂志

Objective To understand the status of resistance to multiple insecticides and the frequencies of kdr mutations in Culex pipiens pallens from north?central Anhui Province. Methods From July to September,2014,the C. pipiens pallens mosquito larvae were collected in Huaibei,Bengbu and Chuzhou cities of the north?central Anhui Province and reared to adults. The female adult mosquitoes at 3-5 days post emergence were tested for susceptibility to the four insecticides,namely 0.05%deltamethrin,5%malathion,0.1%bendiocarb and 4%DDT,by using the standard WHO resistance tube bioassay. The detec?tion of the point mutations of the kdr gene at codon 1014 was conducted by PCR and DNA sequencing in the deltamethrin?resis?tant and?susceptible mosquitoes. Results High levels of resistance to all the four insecticides were found in all the three tested populations,although mosquito mortality varied among populations and test insecticides. Among the test insecticides,DDT showed lowest mortality with no significant difference(F=1.027,P>0.05)in all test populations,whereas significantly differ?ent mortalities were observed among populations for the remained three insecticides tested(deltamethrin,malathion,and ben?diocarb)(F = 23.823,33.955,128.841;all P < 0.01). Two types of non?synonymous kdr mutation at codon position 1014 (L1014F and L1014S)were observed. A positive correlation between L1014F mutation frequencies and deltamethrin resistance levels were detected in the three mosquito populations(r2=0.718,P<0.01). Conclusions The observed high levels of resis?tance to multiple?insecticides coupled with the occurrence of medium to high kdr frequencies in populations of C. pipiens pallens could profoundly affect the mosquito vector control programme in China. The local health departments need to strengthen vector dynamic monitoring and implement rational resistance management strategies.

Evidence of increasing L1014F kdr mutation frequency in Anopheles gambiae s.l. pyrethroid resistant following a nationwide distribution of LLINs by the Beninese National Malaria Control Programme

Objective:To determine the susceptibility status to pyrethroid in Anopheles gambiae s.l. (An. gambiae), the distribution of kdr“Leu-Phe”mutation in malaria vectors in Benin and to compare the current frequency of kdr“Leu-Phe”mutation to the previous frequency after long-lasting insecticide treated nets implementation. Methods: Larvae and pupae of An. gambiae s.l. mosquitoes were collected from the breeding sites in Littoral, Zou, Borgou and Alibori provinces. CDC susceptibility tests were conducted on unfed females mosquitoes aged 2-5 d old. An. gambiae mosquitoes were identified to species using PCR techniques. Molecular assays were also carried out to identify kdr mutations in individual mosquitoes. Results: The results showed that An. gambiae Malanville and Suru-lere populations were resistant to deltamethrin. Regarding An. gambiae Parakou and Bohicon populations, they were resistant to permethrin. PCR revealed 100%of mosquitoes tested were An. gambiae s.s. The L1014F kdr mutation was found in An. gambiae s.s. Malanville and Parakou at various allelic frequencies. The increase of kdr allelic frequency was positively correlated with CDC bioassays data. Conclusions: Pyrethroid resistance is widespread in malaria vector in Benin and kdr mutation is the main resistance mechanism involved. More attention may be paid for the future success of malaria control programmes based on LLINs with pyrethroids in the country.

Evidence of increasing L1014F kdr mutation frequency in Anopheles gambiae s.l. pyrethroid resistant following a nationwide distribution of LLINs by the Beninese National Malaria Control Programme

<p><b>OBJECTIVE</b>To determine the susceptibility status to pyrethroid in Anopheles gambiae s.l. (An. gambiae), the distribution of kdr "Leu-Phe" mutation in malaria vectors in Benin and to compare the current frequency of kdr "Leu-Phe" mutation to the previous frequency after long-lasting insecticide treated nets implementation.</p><p><b>METHODS</b>Larvae and pupae of An. gambiae s.l. mosquitoes were collected from the breeding sites in Littoral, Zou, Borgou and Alibori provinces. CDC susceptibility tests were conducted on unfed females mosquitoes aged 2-5 d old. An. gambiae mosquitoes were identified to species using PCR techniques. Molecular assays were also carried out to identify kdr mutations in individual mosquitoes.</p><p><b>RESULTS</b>The results showed that An. gambiae Malanville and Suru-lere populations were resistant to deltamethrin. Regarding An. gambiae Parakou and Bohicon populations, they were resistant to permethrin. PCR revealed 100% of mosquitoes tested were An. gambiae s.s. The L1014F kdr mutation was found in An. gambiae s.s. Malanville and Parakou at various allelic frequencies. The increase of kdr allelic frequency was positively correlated with CDC bioassays data.</p><p><b>CONCLUSIONS</b>: Pyrethroid resistance is widespread in malaria vector in Benin and kdr mutation is the main resistance mechanism involved. More attention may be paid for the future success of malaria control programmes based on LLINs with pyrethroids in the country.</p>