Insecticide susceptibility status of Anopheles gambiae mosquitoes and the effect of pre-exposure to a piperonyl butoxide (PBO) synergist on resistance to deltamethrin in northern Namibia.

BACKGROUND: Pyrethroid-based indoor residual spraying (IRS) and long-lasting insecticidal nets (LLINs) have been employed as key vector control measures against malaria in Namibia. However, pyrethroid resistance in Anopheles mosquitoes may compromise the efficacy of these interventions. To address this challenge, the World Health Organization (WHO) recommends the use of piperonyl butoxide (PBO) LLINs in areas where pyrethroid resistance is confirmed to be mediated by mixed function oxidase (MFO). METHODS: This study assessed the susceptibility of Anopheles gambiae sensu lato (s.l.) mosquitoes to WHO tube bioassays with 4% DDT and 0.05% deltamethrin insecticides. Additionally, the study explored the effect of piperonyl butoxide (PBO) synergist by sequentially exposing mosquitoes to deltamethrin (0.05%) alone, PBO (4%) + deltamethrin (0.05%), and PBO alone. The Anopheles mosquitoes were further identified morphologically and molecularly. RESULTS: The findings revealed that An. gambiae sensu stricto (s.s.) (62%) was more prevalent than Anopheles arabiensis (38%). The WHO tube bioassays confirmed resistance to deltamethrin 0.05% in the Oshikoto, Kunene, and Kavango West regions, with mortality rates of 79, 86, and 67%, respectively. In contrast, An. arabiensis displayed resistance to deltamethrin 0.05% in Oshikoto (82% mortality) and reduced susceptibility in Kavango West (96% mortality). Notably, there was reduced susceptibility to DDT 4% in both An. gambiae s.s. and An. arabiensis from the Kavango West region. Subsequently, a subsample from PBO synergist assays in 2020 demonstrated a high proportion of An. arabiensis in Oshana (84.4%) and Oshikoto (73.6%), and 0.42% of Anopheles quadriannulatus in Oshana. Non-amplifiers were also present (15.2% in Oshana; 26.4% in Oshikoto). Deltamethrin resistance with less than 95% mortality, was consistently observed in An. gambiae s.l. populations across all sites in both 2020 and 2021. Following pre-exposure to the PBO synergist, susceptibility to deltamethrin was fully restored with 100.0% mortality at all sites in 2020 and 2021. CONCLUSIONS: Pyrethroid resistance has been identified in An. gambiae s.s. and An. arabiensis in the Kavango West, Kunene, and Oshikoto regions, indicating potential challenges for pyrethroid-based IRS and LLINs. Consequently, the data highlights the promise of pyrethroid-PBO LLINs in addressing resistance issues in the region.

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.

Baseline characterization of entomological drivers of malaria transmission in Namibia: a targeted operational entomological surveillance strategy.

BACKGROUND: Namibia's focus on the elimination of malaria requires an evidence-based strategy directed at understanding and targeting the entomological drivers of malaria transmission. In 2018 and 2019, the Namibia National Vector-borne Diseases Control Program (NVDCP) implemented baseline entomological surveillance based on a question-based approach outlined in the Entomological Surveillance Planning Tool (ESPT). In the present study, we report on the findings of the ESPT-based NVDCP on baseline vector species composition and bionomic traits in malaria endemic regions in northern Namibia, which has the aim of generating an evidence base for programmatic decision-making. METHODS: Nine representative sentinel sites were included in the 2018 entomological surveillance program (Kunene, Omusati, Oshana, Ohangwena, Oshikoto, Otjozondjupa, Kavango West, Kavango East and Zambezi); the number was reduced to four sites in 2019 due to limited funding (Ohangwena, Kavango West, Kavango East, and Zambezi). In the 2018 baseline collections, multiple sampling methods (human landing catches, pyrethroid spray catches, U.S. Centers for Disease Control and Prevention light traps [CDC-LTs], resting boxes [RBs] and larval sampling) were utilized to evaluate indoor/outdoor human biting rates, resting behaviors and insecticide resistance (IR). CDC-LTs and RBs were not used in 2019 due to low and non-representative sampling efficacies. RESULTS: Overall, molecular evidence demonstrated the presence of three primary mosquito vectors, namely Anopheles arabiensis, rediscovered Anopheles gambiae sensu stricto and Anopheles funestus sensu stricto, alongside Anopheles squamosus and members of the Anopheles coustani complex. Vectors were found to bite throughout the night (1800 hours 0600 hours) both indoors and outdoors, with An. arabiensis having the highest biting rates outdoors. Low numbers of indoor resting Anopheles point to possible low indoor residual spraying (IRS) efficacy-with An. arabiensis found to be the major vector species resting indoors. The IR tests demonstrated varying country-wide resistance levels to the insecticide deltamethrin, with the resistance levels confirmed to have increased in 2019, evidence that impacts national programmatic decision-making. Vectors demonstrated susceptibility to the insecticides dichlorodiphenyltrichloroethane, bendiocarb and Actellic 300CS in 2018, with mosquitoes from only one site (Kavango West) demonstrating possible resistance to DDT. Targeted and question-based entomological surveillance enabled a rapid and focused evidence base to be built, showing where and when humans were being bitten and providing entomological data on long-lasting insecticidal nets, IRS efficacy and insecticide resistance, which the Ministry of Health and Social Services-Namibia can use to further build a monitoring and evaluation framework for understanding the drivers of transmission. CONCLUSION: Identification and characterization of species-specific bionomic traits allows for an understanding of where and when vector human contact may occur as well as the potential impact of interventions. Low indoor resting rates as well as the presence of insecticide resistance (and the increase in its frequency) point to the need for mosquito-behavior-directed and appropriate interventions as well as the requirement for a resistance mitigation strategy. The ESPT-based question- and minimal essential indicator-based operational research strategy provides programs with directed and focused data for facilitating decision-making while requiring limited funding and capacity.

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).

Entomological Surveillance Planning Tool (ESPT)-generated actionable evidence on human and vector behaviours optimizes present interventions and reduces exposure to Anopheles vectors in two communities of Guna Yala, Panamá.

BACKGROUND: Although most of Panamá is free from malaria, localized foci of transmission persist, including in the Guna Yala region. Government-led entomological surveillance using an Entomological Surveillance Planning Tool (ESPT) sought to answer programmatically relevant questions on local entomological drivers of transmission and gaps in protection to guide local vector control decision-making. METHODS: The ESPT was used to design a sampling plan to answer priority programmatic questions about the appropriateness of Long Lasting Insecticidal Nets (LLINs) and spaces and times where humans remain exposed to Anopheles bites (gaps in protection) in the communities of Permé and Puerto Obaldía, Guna Yala. Adult Anopheles were sampled at three time points via human landing catches (HLCs) during the rainy and dry seasons (2018/2019). Human behaviour observations (HBOs) were conducted alongside HLCs to examine intervention use, indoor versus outdoor activity, and sleeping patterns. HLC and HBO data were integrated to evaluate HBO-adjusted human biting rate (HBR). RESULTS: A total of 7,431 adult Anopheles were collected across both sites. Of the 450 specimens molecularly confirmed to species-level, 75.5% (n = 340) were confirmed as Anopheles Nyssorhynchus albimanus, followed by Anopheles (Ny.) aquasalis. Anopheles host seeking activity was demonstrated to be primarily exophagic throughout all sampling periods and in both communities. When adjusted with HBOs, exposure to mosquito bites was predominantly indoors and overnight in Permé (Nov, Mar), compared to predominantly outdoors in Puerto Obaldía (Nov, Mar, Jul). Differences in site-specific human-vector exposure profiles were due to contrasting cultural and lifestyle practices between Permé and Puerto Obaldía (possibly partly influenced by the absence of electricity in Permé), and lower LLIN use in Permé. This evidence supported a previously planned LLIN campaign alongside a social behaviour change communication (SBCC) strategy in the Guna Yala Comarca (Jul 2019), which increased LLIN use. In turn, this led to a reduction of indoor exposure to mosquito bites, and a shift to predominant outdoor exposure to mosquito bites. CONCLUSION: ESPT-based question-driven planning and the integration of HBOs, intervention, and HLC data generated evidence towards answering the programmatic questions. This evidence enabled the characterization of site-specific human-vector exposure profiles, and the quantification of remaining gaps in protection. These data also provide important insights into remaining gaps in protection that must be addressed to further reduce human exposure to mosquito bites at these sites.

Human and vector behaviors determine exposure to Anopheles in Namibia.

BACKGROUND: Although the Republic of Namibia has significantly reduced malaria transmission, regular outbreaks and persistent transmission impede progress towards elimination. Towards an understanding of the protective efficacy, as well as gaps in protection, associated with long-lasting insecticidal nets (LLINs), human and Anopheles behaviors were evaluated in parallel in three malaria endemic regions, Kavango East, Ohangwena and Zambezi, using the Entomological Surveillance Planning Tool to answer the question: where and when are humans being exposed to bites of Anopheles mosquitoes? METHODS: Surveillance activities were conducted during the malaria transmission season in March 2018 for eight consecutive nights. Four sentinel structures per site were selected, and human landing catches and human behavior observations were consented to for a total of 32 collection nights per site. The selected structures were representative of local constructions (with respect to building materials and size) and were at least 100 m from each other. For each house where human landing catches were undertaken, a two-person team collected mosquitoes from 1800 to 0600 hours. RESULTS: Surveillance revealed the presence of the primary vectors Anopheles arabiensis, Anopheles gambiae sensu stricto (s.s.) and Anopheles funestus s.s., along with secondary vectors (Anopheles coustani sensu lato and Anopheles squamosus), with both indoor and outdoor biting behaviors based on the site. Site-specific human behaviors considerably increased human exposure to vector biting. The interaction between local human behaviors (spatial and temporal presence alongside LLIN use) and vector behaviors (spatial and temporal host seeking), and also species composition, dictated where and when exposure to infectious bites occurred, and showed that exposure was primarily indoors in Kavango East (78.6%) and outdoors in Ohangwena (66.7%) and Zambezi (81.4%). Human behavior-adjusted exposure was significantly different from raw vector biting rate. CONCLUSIONS: Increased LLIN use may significantly increase protection and reduce exposure to malaria, but may not be enough to eliminate the disease, as gaps in protection will remain both indoors (when people are awake and not using LLINs) and outdoors. Alternative interventions are required to address these exposure gaps. Focused and question-based operational entomological surveillance together with human behavioral observations may considerably improve our understanding of transmission dynamics as well as intervention efficacy and gaps in protection.

Anopheles drivers of persisting malaria transmission in Guna Yala, Panamá: an operational investigation.

BACKGROUND: Though most of Panamá is free from malaria, localized foci of transmission persist, including in the Guna Yala region. Government-led entomological surveillance using an entomological surveillance planning tool (ESPT) sought to answer programmatically-relevant questions that would enhance the understanding of both local entomological drivers of transmission and gaps in protection that result in persisting malaria transmission to guide local vector control decision-making. METHODS: The ESPT was used to design a sampling plan centered around the collection of minimum essential indicators to investigate the relevance of LLINs and IRS in the communities of Permé and Puerto Obaldía, Guna Yala, as well as to pinpoint any remaining spaces and times where humans are exposed to Anopheles bites (gaps in protection). Adult Anopheles were collected at three time points via human landing catches (HLCs), CDC Light Traps (LT), and pyrethrum spray catches (PSCs) during the rainy and dry seasons. Mosquitoes were identified to species via molecular methods. Insecticide susceptibility testing of the main vector species to fenitrothion was conducted. RESULTS: In total, 7537 adult Anopheles were collected from both sites. Of the 493 specimens molecularly confirmed to species, two thirds (n = 340) were identified as Nyssorhynchus albimanus, followed by Anopheles aquasalis. Overall Anopheles human biting rates (HBRs) were higher outdoors than indoors, and were higher in Permé than in Puerto Obaldía: nightly outdoor HBR ranged from 2.71 bites per person per night (bpn) (Puerto Obaldía), to 221.00 bpn (Permé), whereas indoor nightly HBR ranged from 0.70 bpn (Puerto Obaldía) to 81.90 bpn (Permé). Generally, peak biting occurred during the early evening. The CDC LT trap yields were significantly lower than that of HLCs and this collection method was dropped after the first collection. Pyrethrum spray catches resulted in only three indoor resting Anopheles collected. Insecticide resistance (IR) of Ny. albimanus to fenitrothion was confirmed, with only 65.5% mortality at the diagnostic time. CONCLUSION: The early evening exophagic behaviour of Anopheles vectors, the absence of indoor resting behaviours, and the presence of resistance to the primary intervention insecticide demonstrate limitations of the current malaria strategy, including indoor residual spraying (IRS) and long-lasting insecticidal nets (LLINs), and point to both gaps in protection and to the drivers of persisting malaria transmission in Guna Yala. These findings highlight the need for continued and directed entomological surveillance, based on programmatic questions, that generates entomological evidence to inform an adaptive malaria elimination strategy.

Rhamphomyia Meigen of the Canadian Arctic Archipelago, Greenland and Iceland (Diptera: Empididae).

Rhamphomyia of the Canadian Arctic Archipelago, Greenland and Iceland, comprising 23 species, including five new species, are revised: R. (Ctenempis) albopilosa Coquillett, R. (Dasyrhamphomyia) erinacioides Malloch, R. (Dasyrhamphomyia) hovgaardii Holmgren, R. (Dasyrhamphomyia) leptidiformis Frey, R. (Dasyrhamphomyia) nigrita Zetterstedt, R. (Eorhamphomyia) shewelli Sinclair, Vajda, Saigusa Shamshev sp. nov., R. (Pararhamphomyia) diversipennis Becker, R. (Pararhamphomyia) filicauda Henriksen Lundbeck, R. (Pararhamphomyia) frigida Sinclair, Vajda, Saigusa Shamshev sp. nov., R. (Pararhamphomyia) helleni Frey, R. (Pararhamphomyia) hilariformis Frey, R. (Pararhamphomyia) hoeli Frey, R. (Pararhamphomyia) kjellmanii Holmgren, R. (Pararhamphomyia) lymaniana Sinclair, Vajda, Saigusa Shamshev sp. nov., R. (Pararhamphomyia) omissinervis Becker, R. (Pararhamphomyia) petervajdai Sinclair, Vajda, Saigusa Shamshev sp. nov., R. (Pararhamphomyia) septentrionalis Sinclair, Vajda, Saigusa Shamshev sp. nov., R. (Pararhamphomyia) simplex Zetterstedt, R. (Pararhamphomyia) ursinella Melander, R. herschelli Malloch, R. hirtula Zetterstedt, R. laevigata Loew, R. setosa Coquillett. The following six new synonyms are proposed: R. calvimontis Cockerell, 1916 and R. wuorentausi Frey, 1922 = R. albopilosa Coquillett, 1900; R. fridolini Frey, 1950 = R. laevigata Loew, 1861; R. hirticula Collin, 1937 = R. setosa Coquillett, 1895; R. uralensis Becker, 1915 = R. kjellmanii Holmgren, 1880; R. zaitzevi Becker, 1915 = R. hovgaardii Holmgren, 1880. Lectotypes are designated for the following species: R. diversipennis Becker, R. filicauda Henriksen Lundbeck, R. helleni Frey, R. herschelli Malloch, R. hirticula Collin, R. hoeli Frey, R. leptidiformis Frey, R. omissinervis Becker, R. setosa Coquillett, R. uralensis Becker, R. wuorentausi Frey, R. zaitzevi Becker. A neotype is designated for R. laevigata Loew. Keys to male and female species of Rhamphomyia and distribution maps of this region are provided. DNA barcode data are presented for 16 species of arctic Rhamphomyia.

New Record of Wyeomyia mitchellii (Diptera: Culicidae) on Guam, United States.

Wyeomyia (Wyeomyia) mitchellii (Theobald) (Diptera: Culicidae) was recovered for the first time on Guam, United States of America, in 2017. Larval specimens were collected from water-filled axils of bromeliads during a larval survey carried out in a residential neighborhood of the Chalan Pago/Ordot area. Native to the New World, Wy. mitchellii has likely made its way to the Pacific Islands through the possibly illegal import of ornamental bromeliads. While this mosquito does not represent a significant threat to public health, this finding highlights the vulnerability of the Pacific Islands to the introduction of exotic species, including mosquito species that may increase public health risks.

Assessing the Risk Factors Associated with Malaria in the Highlands of Ethiopia: What Do We Need to Know?

Malaria has been Ethiopia's predominant communicable disease for decades. Following the catastrophic malaria outbreak in 2003⁻2004, the Federal Ministry of Health (FMoH) took drastic public health actions to lower the burden of malaria. The FMoH achieved significant declines in malaria mortality and incidence, and recently declared its objective to achieve malaria elimination in low malaria transmission areas of Ethiopia by 2020. However, while the overall malaria prevalence has decreased, unpredictable outbreaks increasingly occur irregularly in regions previously considered "malaria-free". Such outbreaks have disastrous consequences on populations of these regions as they have no immunity against malaria. The Amhara Region accounts for 31% of Ethiopia's malaria burden and is targeted for malaria elimination by the FMoH. Amhara's epidemiological surveillance system faces many challenges to detect in a timely manner the unpredictable and irregular malaria outbreaks that occur in areas of otherwise low transmission. Despite the evidence of a shift in malaria transmission patterns, Amhara's malaria control interventions remain constrained to areas that are historically known to have stable malaria transmission. This paper discusses the influence of temperature and precipitation variability, entomological parameters, and human population mobility on malaria transmission patterns across the Amhara Region, and in particular, in areas of unstable transmission. We argue that malaria epidemiological surveillance systems can be improved by accounting for population movements in addition to environmental and entomological factors. However, to date, no study has statistically analyzed the interplay of population dynamics on environmental and entomological drivers of malaria transmission.