Therapeutic doses of ivermectin injected to local Burkinabé Metis cattle rendered blood meals toxic to sympatric An. coluzii females and reduced both survivorship and fecundity of the mosquitoes feeding on treated animals for up to 28 days. For the 2 weeks following the treatment, mean survival time of mosquitoes that fed on treated cattle was two to 3.5 days (corresponding to a time to 100 % mortality of 3–5.5 days), meaning that the great majority would die before being able to resume a new gonotrophic cycle by biting a host, achieve sporogony, and eventually transmit malaria parasites. Mosquitocidal effect of ivermectin was not complete the third and fourth week after treatment and a proportion of mosquitoes was able to survive. Yet, 100 % mortality was achieved in 17 and 19 days, a timeframe just long enough to become infectious and potentially transmit the parasite only once (considering that the first bloodmeal was infectious and considering a sporogony lasting 12 days in average). Using the same scenario, control mosquitoes would survive for 27 days in average, and would be infectious through at least three gonotrophic cycles. Moreover, the fecundity of mosquitoes fed at 21 and 28 DAI was significantly reduced by 33 and 20 %, respectively. Mosquitoes weren’t allowed to lay their eggs nor the hatching rate and larvae survival followed, which represent a limitation of this study, probably leading to an under-estimation of the ivermectin treatments effects on mosquito’s fitness. Considering that a mosquito becomes infectious on average 12 days after gametocyte ingestion [39], corrected mortalities [40], i.e., 100 × (% dead mosquitoes fed on treated cattle − % of dead mosquitoes fed on control cattle)/(100 − % dead mosquitoes fed on control cattle) were 75 and 45 % at 21 and 28 days post injection, respectively. For up to 1 month, more than half of the mosquitoes would die before being able to transmit malaria parasites if they were blood fed on treated cattle before or the same day as the infectious blood meal. Moreover, a second ivermectin blood meal at sub-lethal concentrations further increased the mortality, so much that cumulative mosquito mortality was 100 % by day 12 after the second meal.
As previously reported [5, 14, 15, 41], the present study confirms that ivermectin reduces both the life span and fecundity of important and dominant malaria vectors of sub-Saharan Africa feeding on ivermectin-treated hosts. The mosquitocidal effect vanishes at different rates between calves, suggesting a fair variability in the kinetic and dynamic processes of ivermectin distribution, metabolism or clearance, which may impact on the compound availability in peripheral blood vessels. In a recent study, a greater availability of ivermectin was reported in female human volunteers, which has been associated with the greater body mass indices of female by comparison to male participants [15]. Although ivermectin is the less lipophilic of the macrolactones used as antiparasitic compounds, it nevertheless concentrates particularly in adipose tissues, where the limited vascularization and slow turnover rate of fat prolong the residence of the drug in the peripheral blood [32] and, therefore, its availability to vector mosquitoes. The “slow-release reservoir effect” [15] of body fat might also explain why the mosquitocidal effect of ivermectin could last more than a month in this study while in others, this effect disappeared more quickly and is incomplete even shortly after ivermectin administration [5, 15]. Subcutaneous injection distributes a much greater proportion of the ivermectin into lipid reservoirs than oral route and increases its residence time [42]. Moreover, ivermectin maximum concentration (Cmax) is much lower when the drug is administrated by oral route [43], which might further explain the more sustained mosquitocidal effect presented here, but also the greater, complete toxicity of the blood meals taken by An. coluzzii on treated cattle, for up to 2 weeks after ivermectin subcutaneous injection. Interestingly, at the sub-lethal doses imbibed by mosquitoes at 31 and 37 days, the deleterious effects of ivermectin on survival diminished after a second blood meal 4 days later, which obviously contained ivermectin at less concentration than the first [10]. A second blood meal that does not contain or contains less ivermectin than the first has been also shown to mitigate the effect of the drug on fecundity and hatch rate in Aedes aegypti, Aedes albopictus and Culex quinquefasciatus [44] and on survival in An. gambiae s.s [10]. These cumulative effects must be further examined, considering malaria vectors proclivities for frequent blood feeding in the field [45].
As opposed to in vitro studies [16], the present study failed to demonstrate any transmission blocking properties of sub-lethal concentrations of ivermectin when the ivermectin blood meal was ingested 3 days before an infectious blood meal, but yet at a time post injection where the drug concentration remains toxic enough to impact mosquito survival and fecundity. However, this is in line with recent in situ field studies where ivermectin does not impact gametocyte infectivity [15]. Since only a single DAI (28 days) has been tested on infection prevalence and intensity at the oocyst stage, the present study cannot rule out the possibility that ivermectin has sporontocidal effects. More experiments are needed using different sub-lethal doses, and have to be more adequately designed to study the impact of ivermectin when imbibed with a blood meal at different times after or before the infectious blood meal. However, the present results demonstrate that the stress and corresponding fitness costs induced by sub-lethal doses of the drug did not positively impact infection output, which could have had harmful and counterproductive consequences in terms of transmission, jeopardizing further use of ivermectin to control vector mosquito populations.
The integrative control measure adjunction to existing tools offered by ivermectin has a potential for the management of insecticide resistance since the mode of action of the drug and insecticides currently used for vector control are different. However, despite crucial importance, only a few studies have addressed the question of potential cross-resistance to ivermectin in insecticide-resistant vector mosquitoes. Deus et al. [46] found an increased tolerance to ivermectin imbibed in a blood meal in different Ae. aegypti strains resistant to pyrethroid insecticides. For An. gambiae s.l., ivermectin decreases mosquito life span of Anopheles in different areas of Burkina Faso [14, 15] where the frequency of insecticide target-site mutations, including knock-down resistance (kdr) and insensitive acetylcholinesterase (Ace-1R) alleles, has been regularly monitored and where detoxifying enzymes also contribute to the diversity of resistant phenotypes observed in the field [47, 48]. Thanks to the presence of one out of three mosquitoes carrying the mutated kdr allele in the An. coluzzii colony used here, this study suggests that no cross-resistance to ivermectin exists in kdr carriers, at least at the plasmatic concentrations where mosquitocidal effect is complete. However, proper phenotypic characterization of mutated kdr carriers using bioassays would have been needed to actually check the adequacy between the genotype and resistance phenotype. Hence, more studies are definitely needed to decipher this question, knowing the great diversity and complexity of physiological mechanisms allowing wild An. gambiae s.l. populations to resist most, if not all, of the insecticide classes used to date as vector control tools [31, 47].
Ivermectin is of capital importance for the control of many parasitic diseases in animals and humans and resistance appearance in endo- or ectoparasitic fauna classically targeted by ivermectin treatments would represent a public health disaster. Ivermectin resistance was reported in small ruminants and cattle nematodes after frequent host treatment [43–45, 49]. Hence, if the “One-Health” approach was to be implemented as an alternative method for the control of malaria vectors, a careful monitoring of potential resistance appearance must be undertaken. Researches must also be prompted to apprehend the risk of an emerging resistance to ivermectin in Anopheles field populations. Indeed, with a much longer mosquitocidal and anti-fecundity effect in cattle serum, longer insecticidal pressure from mosquitocidal cattle blood could select for ivermectin resistance in Anopheles. Recent attempts have been made to better understand the IVM-mosquitoes interactions, where canonical detoxification mechanisms seem to be only marginally involved in the mosquito’s response to ivermectin ingestion, whereas non-canonical pathways are highlighted, notably those involving Nieman–Pick type C-2 family genes [50]. Moreover, the recent discovery of ivermectin sensitive and insensitive glutamate-gated chloride channels generated through alternative splicing, questions this mechanism as the potential target of selection for ivermectin resistance in the field [17]. These findings are important in the sense that they clearly emphasize the complexity of IVM-mosquitoes interactions, which need to be unravelled to better evaluate the risks of emergence of ivermectin resistance in the mosquito populations targeted by ivermectin treatments. Similarly, treating cattle might select for Anopheles species/populations with altered behavior toward increased anthropophagy. Hence, the proposed approach would stand only if integrative measures are taken where treatments to humans and cattle and their potential consequences are considered concomitantly. Ivermectin resistance in human or animal targeted parasites and in Anopheles populations are dark shadows in the board of “One-Health” MDAs, and facing these caveats even before they become an issue is the only way to leave this promising approach a reality.
Although subcutaneous administration of ivermectin generates lower faeces concentrations of the product when compared to oral or poor-on formulations [32], non-targeted coprophagic fauna could especially be at risk if MDAs to livestock were implemented [51]. Dung pats are widely used for agricultural purposes in rural Burkina Faso, as everywhere in sub-Saharan rural Africa. Coprophagic fauna accelerate the degradation of dung pats and maintain soil productivity by enhancing the activity of the micro-organisms therein that participate in the mineralization of animal waste. Even sub-lethal doses of ivermectin induce an acute toxicity, altering the sensory and locomotor capacities of dung beetles, and preventing their basic biological activities, ultimately leading to their premature death [52]. However, knowing that sensitivity to ivermectin may vary among species of the same taxa [53], further studies are needed to properly assess ivermectin sensitivity of the coprophagic fauna present in the areas targeted for the “One-health approach”. Such studies are needed so the health benefits to humans and animals of integrated MDAs will not be hampered by potentially high economic losses, which might mitigate the acceptation of this approach by communities.
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