Effects of mosquito age and batch size on knockdown and mortality of laboratory-reared Anopheles dirus, Anopheles minimus, and wild-caught Anopheles harrisoni (Diptera: Culicidae) exposed to transfluthrin using WHO tube and CDC bottle bioassays.

WHO tube and CDC bottle bioassays are currently available for insecticide resistance monitoring and malaria transmission research. Multiple parameters including mosquito density, age, and nutritional status may affect the readout in these bioassays' tests. This study aims to assess the effects of experimental factors on knockdown and mortality measurements in dominant malaria vectors in Thailand following exposure to sublethal and lethal doses of transfluthrin. The effects of (i) 3 different mosquito batch sizes (5, 10, and 20 individuals) and (ii) 2 age groups (3-5 and 20-23 days old) on outcomes measured using the WHO tube (14.7 µg/cm2) and CDC bottle bioassay discriminating concentration (0.006 µg/cm2) against 2 laboratory strains: Anopheles dirus Peyton & Harrison and Anopheles minimus Theobald (species A) and wild-caught Anopheles harrisoni Harbach & Manguin (species C). Our results showed higher knockdown at 1-h exposure using WHO tube and CDC bottle bioassays containing 20 individuals compared to batches containing 10 and 5 individuals. Older mosquitoes showed greater susceptibility than younger test population, especially for An. mininus. Our study supports WHO recommendations for using 3- to 5-day-old mosquitoes. It also validates Praulin et al. (2022) proposal to divide the cohort into smaller batches with more test replicates when it is not practicable to test 25 mosquitoes per replicate.

Inference for entomological semi-field experiments: Fitting a mathematical model assessing personal and community protection of vector-control interventions.

The effectiveness of vector-control tools is often assessed by experiments as a reduction in mosquito landings using human landing catches (HLCs). However, HLCs alone only quantify a single characteristic and therefore do not provide information on the overall impacts of the intervention product. Using data from a recent semi-field study which used time-stratified HLCs, aspiration of non-landing mosquitoes, and blood feeding, we suggest a Bayesian inference approach for fitting such data to a stochastic model. This model considers both personal protection, through a reduction in biting, and community protection, from mosquito mortality and disarming (prolonged inhibition of blood feeding). Parameter estimates are then used to predict the reduction of vectorial capacity induced by etofenpox-treated clothing, picaridin topical repellents, transfluthrin spatial repellents and metofluthrin spatial repellents, as well as combined interventions for Plasmodium falciparum malaria in Anopleles minimus. Overall, all interventions had both personal and community effects, preventing biting and killing or disarming mosquitoes. This led to large estimated reductions in the vectorial capacity, with substantial impact even at low coverage. As the interventions aged, fewer mosquitoes were killed; however the impact of some interventions changed from killing to disarming mosquitoes. Overall, this inference method allows for additional modes of action, rather than just reduction in biting, to be parameterised and highlights the tools assessed as promising malaria interventions.

Distribution of mosquitoes (Diptera: Culicidae) in Thailand: a dataset.

Mosquitoes play a crucial role as primary vectors for various infectious diseases in Thailand. Therefore, accurate distribution information is vital for effectively combating and better controlling mosquito-borne diseases. Here, we present a curated dataset of the mosquito distribution in Thailand comprising 12,278 records of at least 117 mosquito species (Diptera: Culicidae). The main genera included in the dataset are Aedes, Anopheles, Armigeres, Culex, and Mansonia. From 2007 to 2023, data were collected through routine mosquito surveillance and research projects from 1,725 locations across 66 (out of 77) Thai provinces. The majority of the data were extracted from a Thai database of the Thailand Malaria Elimination Program. To facilitate broader access to mosquito-related data and support further exploration of the Thai mosquito fauna, the data were translated into English. Our dataset has been published in the Global Biodiversity Information Facility, making it available for researchers worldwide.

A semi-field evaluation in Thailand of the use of human landing catches (HLC) versus human-baited double net trap (HDN) for assessing the impact of a volatile pyrethroid spatial repellent and pyrethroid-treated clothing on Anopheles minimus landing.

BACKGROUND: The mosquito landing rate measured by human landing catches (HLC) is the conventional endpoint used to evaluate the impact of vector control interventions on human-vector exposure. Non-exposure based alternatives to the HLC are desirable to minimize the risk of accidental mosquito bites. One such alternative is the human-baited double net trap (HDN), but the estimated personal protection of interventions using the HDN has not been compared to the efficacy estimated using HLC. This semi-field study in Sai Yok District, Kanchanaburi Province, Thailand, evaluates the performance of the HLC and the HDN for estimating the effect on Anopheles minimus landing rates of two intervention types characterized by contrasting modes of action, a volatile pyrethroid spatial repellent (VSPR) and insecticide-treated clothing (ITC). METHODS: Two experiments to evaluate the protective efficacy of (1) a VPSR and (2) ITC, were performed. A block randomized cross-over design over 32 nights was carried out with both the HLC or HDN. Eight replicates per combination of collection method and intervention or control arm were conducted. For each replicate, 100 An. minimus were released and were collected for 6 h. The odds ratio (OR) of the released An. minimus mosquitoes landing in the intervention compared to the control arm was estimated using logistic regression, including collection method, treatment, and experimental day as fixed effects. RESULTS: For the VPSR, the protective efficacy was similar for the two methods: 99.3%, 95% CI (99.5-99.0) when measured by HLC, and 100% (100, Inf) when measured by HDN where no mosquitoes were caught (interaction test p = 0.99). For the ITC, the protective efficacy was 70% (60-77%) measured by HLC but there was no evidence of protection when measured by HDN [4% increase (15-27%)] (interaction test p < 0.001). CONCLUSIONS: Interactions between mosquitoes, bite prevention tools and the sampling method may impact the estimated intervention protective efficacy. Consequently, the sampling method must be considered when evaluating these interventions. The HDN is a valid alternative trapping method (relative to the HLC) for evaluating the impact of bite prevention methods that affect mosquito behaviour at a distance (e.g. VPSR), but not for interventions that operate through tarsal contact (e.g., ITC).

The effect of transfluthrin-treated jute and cotton emanator vests on human landing and fecundity of Anopheles minimus in Thailand.

Complementary approaches to tackle outdoor and early evening biting mosquitoes are urgently required. Transfluthrin (TFT) is a volatile pyrethroid capable of altering mosquito feeding behavior. This study investigated the efficacy of TFT-treated jute (TFT-J) and cotton (TFT-C) fabrics on human landing activity, feeding and fecundity of Anopheles minimus in Thailand. Jute and cotton fabrics each measuring 1024 cm2 were impregnated with 7.34 × 10-4 g/cm2 TFT (20%, w/v), and evaluated in a semi-field screen house system. Two collectors, wearing an untreated control or TFT-treated vests, conducted human-landing collections of released 100 laboratory-reared adult females of An. minimus from 18:00-00:00 h for 16 consecutive nights. Recaptured mosquitoes were given a blood meal for 30 min. with a membrane feeding system for assessment of blood feeding and fecundity. TFT-J, relative to control, significantly reduced human landings (Odds Ratio (OR) =0.27 (95% Confidence Interval (CI) [0.10-0.74], p = 0.011)), however no significant reduction was observed for TFT-C (OR=0.67 [95% CI 0.24-1.82], p = 0.43). Blood feeding was significantly lower among mosquitoes exposed to TFT-J (12.45% [95% CI, 2.04-22.85], p = 0.029) and TFT-C (13% [95% CI, 0.99-26.84], p = 0.016) relative to control. Impregnated fabrics had no effect on the mean number of egg oviposition. However, egg hatchability was reduced in TFT-J (49.5% [95% CI, 21.74-77.26], p = 0.029) and TFT-C (40.2% [95% CI, 17.21-63.19], p = 0.008) relative to control. TFT-J significantly reduced the landing, blood feeding, and fertility of An. minimus. Further studies are needed to evaluate different treatment methods on fabrics and their incorporation in integrated mosquito management.

Semi-field evaluation of human landing catches versus human double net trap for estimating human biting rate of Anopheles minimus and Anopheles harrisoni in Thailand.

Background: Whilst the human landing catch (HLC) technique is considered the 'gold standard' for estimating human-biting rates, it is labor-intensive and fraught with potential risk of exposure to infectious mosquito bites. This study evaluated the feasibility and performance of an alternative method, the human double net trap (HDNT) relative to HLC for monitoring host-seeking malaria vectors of the Anopheles minimus complex in a semi-field system (SFS). Methods: HDNT and HLC were positioned in two rooms, 30 m apart at both ends of the SFS. Two human volunteers were rotated between both traps and collected released mosquitoes (n = 100) from 6:00 pm till 6:00 am. Differences in Anopheles mosquito densities among the trapping methods were compared using a generalized linear model based on a negative binomial distribution. Results: There were 82.80% (2,136/2,580) of recaptures of wild-caught and 94.50% (2,835/3,000) of laboratory-reared mosquitoes that were molecularly identified as An. harrisoni and An. minimus, respectively. Mean density of An. harrisoni was significantly lower in HNDT (15.50 per night, 95% CI [12.48-18.52]) relative to HLC (25.32 per night (95% CI [22.28-28.36]), p < 0.001). Similarly, the mean density of a laboratory strain of An. minimus recaptured in HDNT was significantly lower (37.87 per night, 95% CI [34.62-41.11]) relative to HLC (56.40 per night, 95% CI [55.37-57.43]), p < 0.001. Relative sampling efficiency analysis showed that HLC was the more efficient trap in collecting the An. minimus complex in the SFS. Conclusion: HDNT caught proportionately fewer An. minimus complex than HLC. HDNT was not sensitive nor significantly correlated with HLC, suggesting that it is not an alternative method to HLC.

Comparing Light-Emitting-Diodes Light Traps for Catching Anopheles Mosquitoes in a Forest Setting, Western Thailand.

Light traps are a common method for attracting and collecting arthropods, including disease vectors such as mosquitoes. Various types of traps have been used to monitor mosquitoes in a forest in Western Thailand. In this study, four Light Emitting Diodes (LED) light sources (UV, blue, green, and red) and two fluorescent lights (white and UV) were used to trap nocturnal adult mosquitoes. These traps were used with light alone and not any additional attractant. The experiment was conducted from 18:00 to 06:00 h. on six consecutive nights, every two months, across dry, wet, and cold seasons. All specimens were first identified by morphological features and subsequently confirmed by using PCR. We collected a total of 873 specimens of 31 species in four genera, Anopheles, Aedes, Culex, and Armigeres. Anopheles harrisoni was the predominant species, followed by Aedes albopictus, Culex brevipalpis, Culex nitropunctatus, and Armigeres (Leicesteria) longipalpis. UV fluorescent light was the most effective light source for capturing forest mosquitoes, followed by UV LED, blue LED, green LED, white fluorescent, and red LED. The optimal times for collection were from 21:00 to 03:00 h in the dry season. Our results demonstrate that appropriate sampling times and light sources should be selected for optimal efficiency in vector surveillance programs.

Transmitted Light as Attractant with Mechanical Traps for Collecting Nocturnal Mosquitoes in Urban Bangkok, Thailand.

Mosquito surveillance is the cornerstone for determining abundance, species diversity, pathogen infection rates, and temporal and spatial distribution of different life stages in an area. Various methods are available for assessing adult mosquito populations, including mechanical trap devices using different forms of attractant cues (chemical and visual) to lure mosquitoes to the trap. So-called "light traps" use various electromagnetic wavelengths to produce a variety of visible spectral colors to attract adult mosquitoes. However, this type of trapping technology has not been widely used in Thailand. This study compared the efficacy of 4 light-emitting diodes (LEDs) (blue, green, yellow, and red) and 2 fluorescent (ultraviolet [UV] and white) lights for collecting mosquitoes in urban Bangkok. Using a Latin square experimental design, 6 light traps equipped with different lights were rotated between 6 trap site locations within the Kasetsart University (KU) campus. Each location received 6 replicate collections (6 consecutive trap-nights represented 1 replicate) over 36 collection nights for a total of 216 trap-nights. Traps were operated simultaneously (1800 to 0600 h), with captured mosquitoes removed at 3-h intervals. In total, 2,387 mosquitoes consisting of 11 species across 5 genera (Aedes, Anopheles, Armigeres, Culex, and Mansonia) were captured. Collectively, Culex species represented the predominant group sampled (2,252; 94.4%). The UV light source captured 1,544 (64.7%) of the total mosquitoes collected, followed by white 389 (16.3%), with the 4 LED sources collecting between 6.8% (blue) and 1.9% (yellow). Traps equipped with UV light were clearly the most effective for capturing nocturnally active mosquito species on the KU campus.

Susceptibility of Aedes aegypti and Aedes albopictus (Diptera: Culicidae) to Temephos in Thailand and Surrounding Countries.

Aedes-borne virus disease control relies on insecticides to interrupt transmission. Temephos remains a key chemical for control of immature stage Aedes in Thailand and much of Southeast Asia. However, repeated use of insecticides may result in selection for resistance in vector populations, thus compromising operational intervention. Herein, the phenotypic response to temephos by Aedes aegypti (L.) and Aedes albopictus (Skuse) collected in Thailand and surrounding countries is presented. Data from 345 collection sites are included: 283 from literature review (244 sites with Ae. aegypti, 21 with Ae. albopictus, and 18 having both species sampled), plus 62 locations with Ae. aegypti in Thailand conducted between 2014 and 2018. Susceptibility assays followed WHO guidelines using the recommended discriminating dose of temephos (0.012 mg/liter) against late third to early fourth instar Ae. aegypti. Findings revealed 34 locations with susceptible Ae. aegypti, 13 with suspected resistance, and 15 indicating resistance. Published data between 1999 and 2019 in Thailand found Ae. aegypti resistant in 73 of 206 collection sites, whereas 3 locations from 11 sampled with low-level resistant in Ae. albopictus. From surrounding countries conducting temephos assays (Cambodia, Lao PDR, Myanmar, Malaysia, and Singapore), resistance is present in Ae. aegypti and Ae. albopictus from 27 of 56 and 19 of 28 locations, respectively. Routine insecticide susceptibility monitoring should be an operational requirement in vector control programs. Given the wide distribution and apparent increase in temephos-resistance, alternative larvicidal compounds must be considered if chemical control is to remain a viable vector control strategy.

Excito-repellency Activity of Andrographis paniculata (Lamiales: Acanthaceae) Against Colonized Mosquitoes.

Excito-repellency activity of plant extracts have been increasingly studied as mosquito repellents. In this study, the crude extract of Andrographis paniculata was evaluated for its noncontact repellency, contact excitation (irritancy + repellency), and knockdown/toxicity response against five colonized mosquitoes; Aedes aegypti (L.), Aedes albopictus (Skuse), Anopheles dirus Peyton & Harrison, Anopheles epiroticus Linton & Harbach, and Culex quinquefasciatus Say (Diptera: Culicidae) using an excito-repellency assay system under laboratory-controlled conditions. The escape responses were observed at four different concentrations (0.5-5.0% w/v) with A. paniculata showing strong spatial repellency against Ae. albopictus (96.7% escape) and Ae. aegypti (71.7% escape) at the 2.5% and 0.5% concentrations, respectively. At 0.5% and 5.0% concentrations, the greatest repellency was seen for An. dirus (48.2% escape) and Cx. quinquefasciatus (59.7% escape), respectively. Comparatively, low repellency action was observed against An. epiroticus (1.6-15.0% escape). Escape in contact assays (before adjustment) was generally less pronounced compared to noncontact spatial repellency, with Ae. albopictus showing highest percent escape (71.4% escape) in the contact assay at 1.0% concentration. After adjusting for spatial repellency, escape due to contact irritancy alone was either not present or an insignificant contribution to the overall avoidance response for all species. No knockdown or mortality at 24-h postexposure was observed in any trials. These findings indicate that the A. paniculata crude extract is more active against day-biting mosquitoes; however, this may be a reflection of the time of testing. This study demonstrates compelling evidence that A. paniculata extract performs primarily as a spatial repellent. Further investigations exploring the use A. paniculata as a potential active ingredient in repellent products are needed.