Insecticide resistance profiles of Anopheles gambiae s.l. in Togo and genetic mechanisms involved, during 3-year survey: Is there any need for resistance management?

This work is licensed under a Creative Commons Attribution 4.0 International License.Background Malaria, one of the world’s greatest public health challenges, is an endemic disease with stable transmission in Togo. Combating malaria requires an effective vector control. This study provides temporal data on insecticide resistance status in the major malaria vector Anopheles gambiae sensu lato (s.l.) from Togo. Methods Two to 5 days old females of An. gambiae s.l., originating from three localities (Baguida, Kovié, Kolokopé) were subjected to insecticide-impregnated papers during 3 years (2012, 2013, 2016) as follows: organochlorides (4% DDT), pyrethroids (0.05% deltamethrin, 0.75% permethrin, 0.05% lambdacyhalothrin), carbamates (0.4% bendiocarb and 0.1% propoxur), and organophosphates (5% malathion, 0.4% chlorpyrifos methyl, 1% fenitrothion) following the WHO standard protocol. Dead and surviving mosquitoes were stored separately in Eppendorf tubes containing silica gel for DNA extraction, species identification, and kdr and ace-1 genotyping. Results Knockdown times (KDT50 and KDT95) were high in An. gambiae s.l. The lowest KDTs were recorded at Baguida in 2013 for deltamethrin (KDT50 = 24.7, CI [22.4–27.12] and KDT95 = 90.78, CI [76.35–113.49]). No KDTs were recorded for DDT and in some instances for permethrin. In general, An. gambiae s.l. was resistant to most of the four classes of insecticides during the survey periods regardless of locality and year, except to chlorpyrifos methyl. In some instances, mosquitoes were fully susceptible to fenitrothion (Kolokopé: 100% and Kovié: 98.05%, CI [95.82–100.26]) and malathion (100% at both Kolokopé and Kovié) in 2013, and malathion only (Kolokopé; 100%) in 2016. Anopheles coluzzii, An. gambiae and Anopheles arabiensis were the three sibling species identified at the three localities with some hybrids at Baguida (2013), and Kovié (2012 and 2016), respectively. Anopheles gambiae was relatively dominant (61.6%). The kdr 1014F allele frequency was > 0.9 in most of the cases, except at Kolokopé (f (1014F) = 0.63, CI [0.55–0.71]) in 2013. The kdr 1014S allele frequency was below 0.02. The highest ace-1 frequencies were identified in An. gambiae at Baguida (2012: 0.52, CI [0.34–0.69] and 2013: 0.66, CI [0.46–0.86]). Conclusion The resistance status is worrying in Togo and should be considered in future malaria vector resistance management programmes by decision-makers.Organization for Women in Science for the Developing Worl

Efficacy of two PBO long lasting insecticidal nets against natural populations of Anopheles gambiae s.l. in experimental huts, Kolokopé, Togo.

LLINs containing an insecticide plus the synergist, piperonyl butoxide (PBO) have been designed for increased efficacy against pyrethroid-resistant malaria vectors. In this study, two LLINs with PBO, PermaNet® 3.0 and Olyset® Plus, and a pyrethroid-only LLIN, Yorkool®, were evaluated in experimental huts against a free-flying, wild population of Anopheles gambiae s.l. in Kolokopé, a cotton cultivated area of Togo. WHO susceptibility tube tests and subsequent molecular assays determine the An. gambiae s.l. populations to be resistant to pyrethroids and DDT with both target site kdr and metabolic resistance mechanisms involved in the resistance observed. Anopheles gambiae s.s. and An. coluzzi were present in sympatry though the kdr (L1014F) mutation was observed at a higher frequency in An. gambiae s.s. The experimental hut results showed that both PermaNet® 3.0 and Olyset® Plus nets induced similar levels of deterrence, exophily, and reduced blood feeding rate against wild An. gambiae s.l. in contrast to the pyrethroid only LLIN, Yorkool®. The proportion of wild An. gambiae s.l. killed by unwashed PermaNet® 3.0 was significantly higher than unwashed Olyset® Plus (corrected mortality 80.5% compared to 66.6%). Similar blood feeding inhibition rates were observed for unwashed PermaNet® 3.0 and Olyset® Plus; however, PermaNet® 3.0 washed 20 times demonstrated significantly higher blood feeding inhibition rate than Olyset® Plus washed 20 times (91.1% compared with 85.6% respectively). Yorkool® performed the worst for all the parameters evaluated. In an area of pyrethroid resistance of An. gambiae s.l involving kdr target site and metabolic resistance mechanisms, LLINs with PBO can provide additional protection in terms of reduction in blood feeding and increase in mosquito mortality compared to a pyrethroid-only net, and should be considered in malaria vector control strategies

Characterization of the <i>An</i>. <i>gambiae s</i>.<i>l</i>. mosquito populations from Kolokopé.

<p>Characterization of the <i>An</i>. <i>gambiae s</i>.<i>l</i>. mosquito populations from Kolokopé.</p

WHO cone bioassay against susceptible <i>An</i>. <i>gambiae s</i>.<i>s</i>. Kisumu of nets before and after washing, and after the hut trial.

<p>WHO cone bioassay against susceptible <i>An</i>. <i>gambiae s</i>.<i>s</i>. Kisumu of nets before and after washing, and after the hut trial.</p

Active ingredient and synergist contents of each net sample before and after the experimental hut trial.

<p>Active ingredient and synergist contents of each net sample before and after the experimental hut trial.</p

Summary of trial results obtained for free flying <i>An</i>. <i>gambiae s</i>.<i>l</i>. in experimental huts (98 nights) in Kolokopé, Togo.

<p>Summary of trial results obtained for free flying <i>An</i>. <i>gambiae s</i>.<i>l</i>. in experimental huts (98 nights) in Kolokopé, Togo.</p

WHO cone bioassay against wild resistant <i>An</i>. <i>gambiae s</i>.<i>l</i>. Kolokopé of all treatment arms before and after the hut trial.

<p>WHO cone bioassay against wild resistant <i>An</i>. <i>gambiae s</i>.<i>l</i>. Kolokopé of all treatment arms before and after the hut trial.</p

Insecticide resistance intensity and efficacy of synergists with pyrethroids in Anopheles gambiae (Diptera: Culicidae) from Southern Togo

Abstract Background This study was designed to provide insecticide resistance data for decision-making in terms of resistance management plans in Togo. Methods The susceptibility status of Anopheles gambiae sensu lato (s.l.) to insecticides used in public health was assessed using the WHO tube test protocol. Pyrethroid resistance intensity bioassays were performed following the CDC bottle test protocol. The activity of detoxification enzymes was tested using the synergists piperonyl butoxide, S.S.S-tributlyphosphorotrithioate and ethacrinic acid. Species-specific identification of An. gambiae s.l. and kdr mutation genotyping were performed using PCR techniques. Results Local populations of An. gambiae s.l. showed full susceptibility to pirimiphos methyl at Lomé, Kovié, Anié, and Kpèlè Toutou. At Baguida, mortality was 90%, indicating possible resistance to pirimiphos methyl. Resistance was recorded to DDT, bendiocarb, and propoxur at all sites. A high intensity of pyrethroid resistance was recorded and the detoxification enzymes contributing to resistance were oxidases, esterases, and glutathione-s-transferases based on the synergist tests. Anopheles gambiae sensu stricto (s.s.) and Anopheles coluzzii were the main species identified. High kdr L1014F and low kdr L1014S allele frequencies were detected at all localities. Conclusion This study suggests the need to reinforce current insecticide-based malaria control interventions (IRS and LLINs) with complementary tools