Irritant and repellent behaviors of sterile male Aedes aegypti (L.) (Diptera: Culicidae) mosquitoes are crucial in the development of disease control strategies applying sterile insect technique.

The mosquito Aedes aegypti, known to transmit important arboviral diseases, including dengue, chikungunya, Zika and yellow fever. Given the importance of this disease vector, a number of control programs have been proposed involving the use of the sterile insect technique (SIT). However, the success of this technique hinges on having a good understanding of the biology and behavior of the male mosquito. Behavioral responses of Ae. aegypti male populations developed for SIT technology were tested under laboratory conditions against chemical and natural irritants and repellents using an excito-repellency (ER) chamber. The results showed that there were no significant behavioral escape responses in any of the radiation-sterilized male Ae. aegypti test populations when exposed to citronella, DEET, transfluthrin, and deltamethrin, suggesting that SIT did not suppress the expected irritancy and repellency (avoidance) behaviors. The type of information reported in the current study is vital in defining the effects of SIT on vector behavior and understanding how such behavior may influence the success of SIT technology with regard to other vector control interventions.

Spatial repellency and attractancy responses of some chemical lures against Aedes albopictus (Diptera: Culicidae) and Anopheles minimus (Diptera: Culicidae) using the high-throughput screening system.

We evaluated the behavioral responses of Aedes albopictus and Anopheles minimus to 3 isovaleric acid and lactic acid-based chemical lure blends and 2 individual alcohols, using Spatial Repellency Assay in a high-throughput screening system (HITSS). Five doses of 0.0002, 0.001, 0.0025, 0.005, and 0.01 g were tested per lure. A BG-lure was used as a reference standard. After 10-min exposure, the number of mosquitoes moving toward or away from the treated chamber was calculated. The results showed that all lures were repellent against Ae. albopictus except for Lure-4 (4% w/v isovaleric acid + 2% w/v lactic acid + 0.0025% w/v myristic acid + 2.5% w/v ammonium hydroxide) which showed a nonsignificant attractancy at the lowest dose. Significantly high spatial repellency was observed at the highest dose of all the tested lures including BG-lure. Lure-2 (isoamyl alcohol) was significantly repellent at all the tested doses. Against An. minimus, Lure-5 (0.02% w/v isovaleric acid + 2% w/v lactic acid) showed significant spatial repellency while Lure-4 was significantly attractant, at all the tested doses. All lures, except Lure-4, showed strong spatial repellency at high doses and attractancy or weak spatial repellency at the lowest dose of 0.0002 g. In summary, our study demonstrated that spatial repellency and attractancy of the tested lures were influenced by both the dose tested and the mosquito species. Lure-2 and Lure-4 are potential spatial repellents and attractants, respectively, for malaria and dengue vectors. However, further studies are necessary to confirm these results at a semifield and open field level.

Enhanced Excito-Repellency of Binary Mixtures of Plant-Based Mosquito Repellents Against Culex quinquefasciatus Say (Diptera: Culicidae), a Night Biting Mosquito Species.

Efficacies of essential oils (EOs) of Vetiveria zizanioides (L.) Nash. (Poales: Poaceae) (VZ EO), Cananga odorata (Lam) Hook. F. & Thomson (Magnoliales: Annonaceae) (CO EO), and crude extract (CE) of Andrographis paniculata (Burm.F.) Wall ex. Nees (Lamiales: Acanthaceae) (AP CE), against laboratory (lab) and field strains of Culex quinquefasciatus Say were investigated. Irritant and repellent activities of individual and binary mixtures of plant extracts were compared with N,N-diethyl-m-toluamide (DEET) using an excito-repellency system. The irritant activity (direct tarsal contact), the mean percent escape response of VZ EO (91.67%, 83.33%), and CO EO (80%, 88.33%) were not significantly different compared with DEET (88.33%, 95%) against lab and field strains, respectively. Similarly, irritant responses in combinations (1:1 and 1:2, v:v) of either VZ EO or CO EO with AP CE were not significantly different from DEET against both strains (P > 0.001). The repellent activity (no tarsal contact), the mean percent escape response of VZ EO (68.33%), CO EO (61.67%), and VZ EO+AP CE (1:1, v:v) (81.67%) against lab strain and CO EO (85%) against field strain were not significantly different from that of DEET (P > 0.001). Interestingly, the greatest contact irritancy of VZ EO+AP CE (1:1, v:v) (96.67%) (P = 0.0026) and a stronger repellency response of CO EO (85%) (P = 0.0055) produced significantly different patterns of escape response compared with DEET against both lab and field strains, respectively. The EOs of VZ EO and CO EO or their mixture with AP CE showed potential as plant-based active ingredients for mosquito repellents. In addition, the major chemical constituents of VZ EO were ß-vetivone (6.4%), khusimol (2.96%), and α-vetivone (2.94%) by gas chromatograpy-mass spectrometry.

Insecticidal and Behavioral Avoidance Responses of Anopheles minimus and Culex quinquefasciatus (Diptera: Culicidae) to Three Synthetic Repellents.

Escape responses, knockdown (KD), and toxicity of laboratory strains of Anopheles minimus Theobald and Culex quinquefasciatus Say to three synthetic mosquito repellents, DEET (N, N-diethyl-3-methylbenzamide), IR3535, or picaridin, at 5% v/v concentrations, were evaluated using repellent-treated papers in standard WHO tube assays and an excito-repellency (ER) test chamber system. The tube assays recorded knockdown effects of each repellent immediately after 30-min exposure and the final morality following a 24-h holding period. DEET showed 100% KD at 30 min and complete toxicity at 24 h against both species. Both actions were either minimal or absent for IR3535 and picaridin, respectively. Culex quinquefasciatus showed significantly greater escape with DEET compared with the other compounds in both contact irritancy (excitation) and noncontact spatial repellency trials. Anopheles minimus showed much more pronounced irritancy and repellency flight escape to IR3535 than picaridin. DEET was the most active irritant and repellent compound against Cx. quinquefasciatus. When adjusting contact test responses based on paired noncontact repellency assays, DEET and IR3535 showed much stronger spatial repellent properties than irritancy with An. minimus. Picaridin performed poorly as an irritant or repellent against both species. We conclude that DEET, followed by IR3535, act as strong spatial repellents at 5% concentration. DEET also performs as a strong toxicant. Our findings show that different mosquitoes can respond contrastingly to repellents, thus the importance to test a wider range of species and populations to assess the full range of chemical action.

Behavioral responses of Anopheles species (Culicidae: Diptera) with varying surface exposure to pyrethroid-treated netting in an excito-repellency test system.

Indoor Residual Spray (IRS) with insecticides has been a procedure used for decades to protect humans from biting mosquitoes and potential vectors of pathogens. The objective of this study was to determine the responses of three wild-caught species of malaria vectors exposed to pyrethroids of three different surface coverage percents using an excito-repellency test box. Each species was exposed to three insecticide-treated surfaces at varying exposure levels (full coverage, 50%, and 25% of the maximum allowable by the test system) to a single standard field dose of either lambda-cyhalothrin or alpha-cypermethrin. Larger numbers of mosquitoes escaped the treated chambers in the direct contact test compared to the spatial repellent chambers in all three different treated surface exposures. No significant differences in the percent of escaped mosquitoes were detected in the 50% and full coverage surface coverage exposures, whereas the 25% coverage produced significantly lower avoidance responses for both compounds. This study found that varying levels of surface exposure with synthetic pyrethroids can impact the behavioral avoidance responses of Anopheles; however, it may also be possible to reduce the amount of coverage to achieve similar avoidance actions. This information may assist policy makers in designing more cost effective strategies involving residual insecticides to control mosquito vectors.

Excito-repellency of essential oils against an Aedes aegypti (L.) field population in Thailand.

An investigation of the behavioral responses of Aedes aegypti (= Stegomyia aegypti) to various concentrations of essential oils (2.5, 5, and 10%) extracted from hairy basil (Ocimum americanum Linn), ginger (Zingiber officinale Roscoe), lemongrass (Cymbopogon citratus Stapf), citronella grass (Cymbopogon nardus Rendle), and plai (Zingiber cassumunar Roxb) were performed using an excito-repellency test chamber. Results showed that Ae. aegypti exhibited varying levels of escape response in both the contact and noncontact chambers in response to different essential oils. The magnitude of the behaviors changed in a dose-response fashion depending on the percent volume to volume concentration of oil used. A 2.5% concentration of hairy basil oil produced a significantly greater escape response compared to the other extracts at the same concentration (P< 0.05). Oils of ginger, lemongrass, and citronella produced stronger irritant and repellent responses at the median 5% concentration compared to the lowest and highest concentrations. There was marked suppression of escape for both contact and noncontact tests using 10% concentrations of hairy basil, lemongrass, and citronella, with high knockdown for all three oils after 30 min. Hairy basil and lemongrass had the highest insecticidal activity to Ae. aegypti, with LC50 values of 6.3 and 6.7 percent, respectively. We conclude that the essential oils from native plants tested, and likely many other extracts found in plants, have inherent repellent and irritant qualities that should to be screened and optimized for their behavior-modifying properties against Ae. aegypti and other biting arthropods of public health and pest importance.

Irritant and repellent responses of Anopheles harrisoni and Anopheles minimus upon exposure to bifenthrin or deltamethrin using an excito-repellency system and a live host.

Feeding responses of Anopheles harrisoni and An. minimus were evaluated following exposure to 2 pyrethroid insecticides, bifenthrin or deltamethrin, using an excito-repellency test system in the presence and absence of live host cues. The results demonstrated that contact irritancy was the primary action of bifenthrin or deltamethrin in both mosquito species. There was no noncontact repellency effect elicited by either insecticide. Anopheles minimus showed rapid escape response with high mortality rates following direct contact with deltamethrin in the absence of a host and delayed escape responses when a host was present. Similarly, exposure of An. minimus to bifenthrin also elicited a delayed escape response in the presence of a host but with lower mortality rates. In experiments using An. harrisoni, the presence or absence of a host had no significant effect on behavioral responses to either insecticide (P > 0.05). We conclude that deltamethrin elicited stronger irritant chemical effects than bifenthrin but that behavioral responses in vector populations are dampened in the presence of an available host. This information is useful for estimating probability of pathogen transmission when using irritant chemicals in proximity to a blood-meal source.

Host feeding responses of Aedes aegypti (L.) exposed to deltamethrin.

Escape responses of mated and unmated nulliparous Aedes aegypti mosquitoes were compared using three different concentrations of deltamethrin in the presence or absence of a live animal host using an excito-repellency (ER) test system. Both insecticide contact (excitation) and non-contact (repellency) test configurations were compared. For contact trials, mated mosquitoes showed similar escape movements among the three concentrations when host stimuli were absent. Significant differences in responses were seen between the lower concentrations of (LC(50) and LC(75) ) deltamethrin with and without hosts present (P<0.05). Presence or absence of host stimuli produced no significant differences in escape response for unmated females when exposed to the highest concentration (LC(90) ) of deltamethrin. Our findings indicate that as deltamethrin concentrations decrease to sublethal levels, mating status and host cues play a more significant role in escape behavior. Therefore, insemination can influence the outcome of feeding success and flight movement of nulliparous female Ae. aegypti in contact with deltamethrin and in the presence of live host stimuli. The ER assay system serves as a useful tool for observing excitation and repellency responses of Ae. aegypti to insecticides in the presence or absence of other environmental and biological cues that can affect mosquito behavior.

Frequency of pyrethroid resistance in Aedes aegypti and Aedes albopictus (Diptera: Culicidae) in Thailand.

Thirty-two Aedes aegypti populations collected throughout Thailand and five populations of Aedes albopictus from southern Thailand were subjected to standard WHO contact bioassays to assess susceptibility to three commonly used synthetic pyrethroids: permethrin, deltamethrin, and lambda-cyhalothrin. A wide degree of physiological response to permethrin was detected in Ae. aegypti, ranging from 56.5% survival (Lampang, northern Thailand) to only 4% (Kalasin in northeastern and Phuket in southern Thailand). All 32 populations of Ae. aegypti were found to have evidence of incipient resistance (62.5%) or levels of survival deemed resistant (37.5%) to permethrin. Four populations of Ae. albopictus were found with incipient resistance (97 - 80% mortality) and one with resistance (< 80%) to permethrin. The majority of Ae. aegypti populations (68.7%) was susceptible (> 98% mortality) to deltamethrin, with incipient resistance (observed 97-82% mortality) in other localities. In contrast, all populations of Ae. aegypti were completely susceptible (100% mortality) to the recommended operational dosage of lambda-cyhalothrin. All five populations of Ae. albopictus were found completely susceptible to both deltamethrin and lambda-cyhalothrin. Evidence of defined incipient or resistance to synthetic pyrethroids mandates appropriate response and countermeasures to mitigate further development and spread of resistance. In light of these findings, we conclude that routine and comprehensive susceptibility monitoring of dengue mosquito vectors to synthetic pyrethroids should be a required component of resistance management policies and disease control activities.