Background Dengue fever is the most rapidly spreading mosquito-borne viral disease over the past 50 years, with a 30-fold increase in global incidence. Dengue vector control is a key component for the dengue control strategy, since no absolutely effective vaccine or drug is available yet. However, the rapid rise and spread of mosquito insecticide resistance have become major threats to the efficiency of insecticide-based vector control activities. Thus, innovative vector control tools are badly needed. This study aims to confirm the antivirus effectiveness of ivermectin on dengue virus type 2 (DENV-2) in Aedes albopictus (Skuse, 1894), then to explore its potential use in the combating to the dengue epidemics.
Methods Aedes albopictus were first infected with DENV-2 in human whole blood, and at the fourth day after infectious blood feeding, they were divided into eight groups. Seven of them were held for six days with access to 0, 2, 4, 8, 16, 32 and 64 ng/ml ivermectin, respectively, and the last one was set as a historical control group, which was stored at -80°C until being detected at the same time with the other groups. Each mosquito was detected using real-time fluorescent RT-PCR kit. DENV-2 RNA concentration (copies/ml) and infection rate in each group were compared.
Ivermectin can directly or indirectly inhibit DENV-2 multiplication in Aedes albopictus. Moreover, the actual concentration for application in zooprophylaxis needs to be confirmed in the further field trials.
Efficacy of ivermectin on DENV-2 infection rate in Aedes albopictus
The average of infection rates in the seven groups treated with 0, 2, 4, 8, 16, 32 and 64 ng/ml ivermectin from 4 to 10 days post ingesting infectious blood were 84.62%, 85.29%, 82.54%, 74.24%, 63.33%, 54.29% and 42.62%, respectively, And the average of infection rates in historical control group was 81.67% (Table 1). Compared with the parallel control group or historical control group, infection rates in the mosquitoes fed with 2, 4, 8 ng/ml ivermectin were not significantly lowered; while infection rates in the mosquitoes fed with 16, 32, 64 ng/ml ivermectin were much lower (Table 1), with infection rate being reduced by as much as 49.63% (Fig 1). The regression equation between infection rates (Y2) and log2 values of ivermectin concentration (X2) was obtained as Y2 = 91.41–7.21*X2 with R2 = 0.89. (Table 1, Fig 2). What might confuse us here was that infection rate (85.29%) in mosquitoes fed with 2ng/ml ivermectin was seem to be higher than that in the historical control group (81.67%) or parallel control group (84.62%), but this differences were meaningless for being without statistical significance. In this part of experiment, antivirus effectiveness on DENV-2 in Aedes albopictus was observed in the ivermectin treatment groups at certain concentration, and the more ivermectin mosquito ingested, the lower the infection rate was.
Compared to mosquitoes fed with 0ng/ml ivermectin, the reduction degree of infection rate for each treatment group.
The calculation formula is as follows: (infection rate in mosquitoes fed with 0ng/ml ivermectin-infection rate in each treatment group)/ infection rate in mosquitoes fed with 0ng/ml ivermectin*100%.
The correlation regression between infection rates and log2 values of ivermectin concentrations.
Efficacy of ivermectin on DENV-2 loads in Aedes albopictus
Related parameters indicating the DENV-2 loads in mosquitoes, including Max, median, P75, P25, Min and Q in each group were presented in Table 2. Compared with mosquitoes fed with 0 ng/ml ivermectin, mosquitoes fed with 2, 4, 8 ng/ml ivermectin carried the same level of DENV-2 RNA concentrations (copies/ml), and mosquitoes fed with 16, 32, 64 ng/ml ivermectin exhibited much lower DENV-2 RNA concentrations (copies/ml) (Table 2), with Max, median, P75 and P25 of DENV-2 RNA concentrations (copies/ml) being reduced by up to 85.89%, 99.99%, 99.99% and 84.06%, respectively (Fig 3). On the other hand, compared with mosquitoes in historical control group, DENV-2 had well developed inside mosquitoes fed with 0, 2, 4, or 8 ng/ml ivermectin showing significantly higher DENV-2 RNA concentrations (copies/ml), and was effectively inhibited in mosquitoes fed with 16, 32, or 64 ng/ml ivermectin showing the same level of DENV-2 RNA concentrations (copies/ml). The evidences confirmed the observation of antivirus effectiveness that virus loads in Aedes albopictus were statistically reduced by treatment of ivermectin when concentration of ivermectin was more than 16ng/ml. (Table 2)
Compare to mosquitoes fed with 0ng/ml ivermectin, the reduction degree of maximum, median, P75, P25 of DENV-2 RNA concentration for each treatment group.
The calculation formula is as follows: (parameter (e.g. median) in mosquitoes fed with 0ng/ml ivermectin-parameter in each treatment group)/ parameter in mosquitoes fed with 0ng/ml ivermectin*100%.
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