A field bioassay for assessing ivermectin bio-efficacy in wild malaria vectors.

BACKGROUND: Ivermectin (IVM) mass drug administration is a candidate complementary malaria vector control tool. Ingestion of blood from IVM treated hosts results in reduced survival in mosquitoes. Estimating bio-efficacy of IVM on wild-caught mosquitoes requires they ingest the drug in a blood meal either through a membrane or direct feeding on a treated host. The latter, has ethical implications, and the former results in low feeding rates. Therefore, there is a need to develop a safe and effective method for IVM bio-efficacy monitoring in wild mosquitoes. METHODS: Insectary-reared Anopheles gambiae s.s. were exposed to four IVM doses: 85, 64, 43, 21 ng/ml, and control group (0 ng/ml) in three different solutions: (i) blood, (ii) 10% glucose, (iii) four ratios (1:1, 1:2, 1:4, 1:8) of blood in 10% glucose, and fed through filter paper. Wild-caught An. gambiae s.l. were exposed to 85, 43 and 21 ng/ml IVM in blood and 1:4 ratio of blood-10% glucose mixture. Survival was monitored for 28 days and a pool of mosquitoes from each cohort sacrificed immediately after feeding and weighed to determine mean weight of each meal type. RESULTS: When administered in glucose solution, mosquitocidal effect of IVM was not comparable to the observed effects when similar concentrations were administered in blood. Equal concentrations of IVM administered in blood resulted in pronounced reductions in mosquito survival compared to glucose solution only. However, by adding small amounts of blood to glucose solution, mosquito mortality rates increased resulting in similar effects to what was observed during blood feeding. CONCLUSION: Bio-efficacy of ivermectin is strongly dependent on mode of drug delivery to the mosquito and likely influenced by digestive processes. The assay developed in this study is a good candidate for field-based bio-efficacy monitoring: wild mosquitoes readily feed on the solution, the assay can be standardized using pre-selected concentrations and by not involving treated blood hosts (human or animal) variation in individual pharmacokinetic profiles as well as ethical issues are bypassed. Meal volumes did not explain the difference in the lethality of IVM across the different meal types necessitating further research on the underlying mechanisms.

Expression of metallothionein and alpha-tubulin in heavy metal-tolerant Anopheles gambiae sensu stricto (Diptera: Culicidae).

Anopheles mosquitoes have been shown to adapt to heavy metals in their natural habitats. In this study we explored the possibility of using Anopheles gambiae sensu stricto as bio-reporters for environmental heavy metal pollution through expressions of their metal-responsive metallothionein and alpha-tubulin genes. The study was undertaken with third instar larvae after selection by cadmium, copper, or lead at LC(30) through five successive generations. Expression levels were determined in the 5th generation by semi-quantitative RT-PCR on the experimental and control populations. The data were analyzed using one-way ANOVA. The highest metallothionein (F(3,11)=4.574, P=0.038) and alpha-tubulin (F(3,11)=12.961, P=0.002) responses were observed in cadmium-tolerant treatments. There was significantly higher expression of metallothionein in cadmium or copper treatments relative to the control (P=0.012), and in cadmium than in lead treatments (P=0.044). Expressions of alpha-tubulin were significantly higher in cadmium than in control treatments (P=0.008). These results demonstrate the capacity of An. gambiae s.s. to develop tolerance to increased levels of heavy metal challenge. The results also confirm the potential of heavy metal-responsive genes in mosquitoes as possible bio-indicators of heavy metal environmental pollution. How the tolerance and expressions relate to An. gambiae s.s. fitness and vectorial capacity in the environment remains to be elucidated.