Medical and Veterinary Entomology

Aedes koreicus is a mosquito (Diptera: Culicidae) from Northeast Asia with a rapidly expanding presence outside its original native range. Over the years the species has been discovered in several new countries, either spreading after first introduction or remaining localised to limited areas. Notably, recent studies have demonstrated the ability of the species to transmit zoonotic parasites and viruses both in the field and in laboratory settings. Combined with its invasive potential, the possible role of Ae. koreicus in pathogen transmission highlights the public health risks of its invasion. In this study, we used a recently established population from Italy to investigate aspects of biology that influence reproductive success in Ae. koreicus: autogeny, mating behaviour, mating disruption by the sympatric invasive species Aedes albopictus, and the presence of the endosymbiont Wolbachia pipientis. Our laboratory population did not exhibit autogenic behaviour and required a blood meal to complete its ovarian cycle. When we exposed Ae. koreicus females to males of Ae. albopictus, we observed repeated attempts at insemination and an aggressive, disruptive mating behaviour initiated by males. Despite this, no sperm was identified in Ae. koreicus spermathecae. Wolbachia was not detected in this Ae. koreicus population and therefore had no effect on Ae. koreicus reproduction.

Reference strains of Aedes aegypti, reared over numerous generations under laboratory conditions, are selected to fit laboratory conditions and commonly used in research due to their consistency and ease of handling. However, working exclusively with such strains may not fully reflect the traits of natural populations. While working on such strains is relevant in terms of reproducibility between labs, it is also important to perform some work on field-collected mosquitoes and their progeny to capture more representative biological and behavioural variation. In this study, we used the New Orleans reference strain as a control to adjust and evaluate methods to manipulate the F1 progeny of field-collected mosquitoes from Cayenne (French Guiana). To improve blood feeding rate, we tested the impact of several blood feeding systems for mosquitoes kept in a cage or in individual vials and adjusted starvation time. To monitor fertilization, we assessed if dissection buffer affects burst of spermatheca during dissection, whether mosquitoes were collected alive of shortly after death. The results described here may be helpful for studies on mosquito fitness, particularly on field-derived mosquitoes or on experiments requiring individual level monitoring. Author summaryAedes aegypti is the major vector of dengue virus, causing hundreds of millions of cases per year. To fight against disease transmission, mosquitoes are studied in the laboratory, notably using colonies of mosquitoes kept over decades in the laboratory. These reference strains have been selected to fit laboratory conditions and may thus not fully reflect the traits of natural populations. While working on such strains is relevant in a sake of consistency between labs and ease of handling, it is also important to work on field-collected mosquitoes and their progeny to capture more representative biological and behavioural variation. In this study, we adjusted and evaluated methods to manipulate the F1 progeny of field-collected mosquitoes from Cayenne (French Guiana), using the New Orleans reference strain as a control. To improve blood feeding rate, we tested the impact of several blood feeding systems for mosquitoes kept in a cage or in individual vials and adjusted starvation time. To monitor fertilization, we assessed if dissection buffer affects burst of spermatheca during dissection, whether mosquitoes were collected alive of shortly after death. The results described here may be helpful for studies on mosquito fitness, particularly on field-derived mosquitoes or on experiments requiring individual level monitoring.

Identifying the current geographic range of disease vectors is a critical first step towards determining effective mechanisms for controlling and potentially eradicating them. This is particularly true given that historical vector ranges may expand due to changing climates and human activity. The Aedes subgenus Stegomyia contains over 100 species, and among them, Ae. aegypti and Ae. albopictus mosquitoes represent the largest concern for public health, spreading dengue, chikungunya, and Zika viruses. While Ae. aegypti has been observed in the country of Zambia for decades, Ae. albopictus has not. In 2015 we sampled four urban and two rural areas in Zambia for Aedes species. Using DNA barcoding, we confirmed the presence of immature and adult Ae. albopictus at two rural sites: Siavonga and Livingstone. These genotypes seem most closely related to specimens previously collected in Mozambique based on CO1 sequence from mtDNA. We resampled Siavonga and Livingstone sites in 2019, again observing immature and adult Ae. albopictus at both sites. Relative Ae. albopictus frequencies were similar between sites, with the exception of immature life stages, which were higher in Siavonga than in Livingstone in 2019. While Ae. albopictus frequencies did not vary through time in Livingstone, both immature and adult frequencies increased through time in Siavonga. This report serves to document the presence of Ae. albopictus in Zambia, which will contribute to the process of determining the potential public health implications of this disease vector in Central Africa.

Dengue cases in Bangladesh have surged in recent years. The existing insecticide-based control program, implemented in parts of the country, is challenged by issues of insufficient household coverage and high levels of insecticide resistance in the primary dengue virus (DENV) vector, Aedes aegypti. A more sustainable, effective alternative could be the implementation of a Wolbachia-mediated disease management strategy. Infecting mosquitoes with Wolbachia can change their reproductive compatibility and their ability to transmit DENV. These new phenotypes can be exploited to suppress or replace wild-type Ae. aegypti populations. Such strategies require the development of well-characterised Wolbachia-infected strains with biological characteristics that are comparable with local mosquitoes. We created and characterised a Wolbachia-infected Ae. aegypti strain with a Dhaka wild-type genetic background, and compared its reproductive compatibility, maternal inheritance, fitness, and virus-blocking ability to the parental strains (Dhaka wild-type and wAlbB2-F4). The new Ae. aegypti strain wAlbB2-Dhaka demonstrated complete cytoplasmic incompatibility with the wild-type strain and complete maternal transmission, retaining levels of pyrethroid resistance of the Dhaka wild-type (70% survival to 10 times the dose of permethrin expected to kill susceptible mosquitoes). No significant fitness costs were detected during laboratory comparisons of fecundity, fertility, survival, mating competitiveness, or desiccation tolerance. Compared to the wild-type strain, wAlbB2-Dhaka mosquitoes had a significantly reduced number of DENV genome copies in the bodies (44.4%, p = 0.0034); two-fold reduction in dissemination to legs and wings (47.6%, p < 0.0001); and >13-fold reduction of DENV in saliva expectorates (proxy of transmission potential) (92.7%, p < 0.0001) 14 days after ingesting dengue-infected blood. Our work indicates that the wAlbB2-Dhaka strain could be used for Ae. aegypti suppression or replacement strategies for dengue management in Bangladesh. Author summaryBangladesh is currently grappling with a series of severe dengue outbreaks, resulting in more than 300 thousand infections, 1700 fatalities, and a significant influx of patients requiring hospitalisation in 2023. These outbreaks coincide with the emergence of high levels of insecticide resistance in the Ae. aegypti population, which is contributing to the failure of conventional, insecticidal control strategies. In search of more sustainable, effective dengue control strategies, approaches using the endosymbiotic bacteria Wolbachia that render mosquitoes resistant to arbovirus infection are being trialled in a number of countries. We transinfected Dhaka wild-type Ae. aegypti with the Wolbachia strain wAlbB2 by backcrossing it with a Wolbachia wAlbB2-infected Ae. aegypti strain and assessed the new strains suitability for field release. We compared the fitness and virus-blocking ability of the backcrossed strain with its parental strains. We demonstrated that the new strain, wAlbB2-Dhaka, is as fit as the Dhaka wild-type strain and blocked [~]92% of dengue virus transmission. This strain has great potential for Wolbachia-mediated dengue management strategies in Bangladesh.

The mosquito microbiome significantly influences vector competence, including in Aedes albopictus, a globally invasive ectoparasite. Describing the microbiome and Wolbachia strains of Ae. albopictus from different regions can guide area-specific control strategies. Mosquito samples from Spain and Sao Tome were analyzed using 16S rRNA gene sequencing and metagenomic sequencing. Wolbachia infection was genotyped, and statistical analysis compared infection patterns by sex and population. Results revealed a dominance of dual Wolbachia infections in the microbiome of both populations of Ae. albopictus, especially among females. The two populations presented a shared microbiome, although 5 and 9 other genera were only present in Spain and Sao Tome populations, respectively. Genera like Pelomonas and Nevskia were identified for the first time in Aedes mosquitoes. This study is the first to describe the Ae. albopictus bacteriome in Spain and Sao Tome, offering insights for the development of targeted mosquito control strategies.

Anopheles stephensi is rapidly expanding across Africa, posing new challenges for malaria control. Its biting time patterns, however, remain poorly characterized, raising uncertainty about the effectiveness of bed nets against this invasive vector. To address this gap, we investigated diel biting activity, feeding propensity, and flight behavior using complementary behavioral assays on females reared from wild-caught larvae in Hawassa City, southern Ethiopia. Flight monitoring assays revealed that An. stephensi exhibited pronounced activity at dusk, beginning between 18:00 and 19:00 with the onset of scotophase, and little to no activity during the photophase. Blood-feeding propensity, defined as the proportion of mosquitoes taking a blood meal when offered, peaked during the early scotophase (18:00-22:00) at 33.3-51.7%, but was markedly reduced during daylight hours (0-16.7%). Human landing catches in large-cage enclosures confirmed this early evening activity: 83.5% of total landings occurred between 18:00 and 22:00 with a sharp peak at 18:00-19:00, corresponding to a mean biting rate of 17.8 bites per person per hour. These findings demonstrate that invasive An. stephensi primarily seek hosts and bite during the early evening, a time when people are often unprotected by bed nets. This behavior reduces the protective impact of conventional net-based interventions and underscores the need for African National Malaria Control Programs to deploy complementary measures such as spatial repellents and larval source management to mitigate early-evening transmission. Moreover, this study highlights the utility of integrated behavioral assays for estimating biting time, offering approaches that can be extended to other vector species across Africa.

We report the first records of Aedes euedes Howard, Dyar, and Knab, and Coquillettidia perturbans (Walker) from Canadas Yukon Territory, and the first record of Ae. decticus Howard, Dyar, and Knab from British Columbia. We also report range extensions in northern BC for the western treehole mosquito, Aedes sierrensis (Ludlow), the common house mosquito, Culex pipiens L., and the cool weather mosquito Culiseta incidens (Thomson).

BackgroundThe invasive mosquito Aedes koreicus has established populations in several European countries during the past decade, raising increasing public health concerns due to its potential role as a vector of pathogens. While species identification is primarily based on morphological characters, Ae. koreicus exhibits distinct morphological variants originating from mainland Korea and Jeju Island, which complicates surveillance and may lead to misidentification, particularly in regions where closely related species co-occur. To date, the genetic basis and population-level relevance of these morphological forms in Europe remain poorly understood. MethodsWe investigated the co-occurrence of two morphological forms of Ae. koreicus in Hungary, representing the first confirmed European location where both forms were detected sympatrically and even within the same breeding sites. Adult mosquitoes were morphologically characterised using diagnostic traits, and individuals representing both morphotypes were subjected to comprehensive genetic analyses. We sequenced multiple mitochondrial markers (COX1, COX2, COX3, ATP6, ND1, ND3) and the nuclear ITS2 region using Oxford Nanopore long-read sequencing. Phylogenetic reconstructions and haplotype network analyses were applied to assess genetic differentiation between morphotypes and to compare them with Aedes japonicus as a closely related reference species. ResultsAcross all analysed mitochondrial and nuclear markers, no genetic differentiation was detected between specimens identified as the "mainland" or "Jeju-do" morphological forms of Ae. koreicus. Phylogenetic and haplotype network analyses consistently grouped individuals independently of their morphotype, indicating a shared genetic background at the population level. In contrast, Ae. japonicus formed a clearly distinct genetic lineage, confirming the robustness of the applied markers for interspecific discrimination. ConclusionsOur results demonstrate that the observed morphological variability in Ae. koreicus populations are not underpinned by detectable genetic differentiation using commonly applied mitochondrial and genomic markers. These findings highlight the limitations of using morphology alone to infer population origin or structure and emphasise the need for heightened awareness of intraspecific variability in routine surveillance. Accurate morphological identification remains critical, particularly in citizen science-based monitoring programmes and AI-based automatization platforms, such as Mosquito Alert, to avoid confusion with morphologically similar invasive species. Further studies integrating genomic and ecological approaches are required to elucidate the mechanisms underlying morphological variability in this emerging vector species.

The wMel and wAlbB strains of the bacterial endosymbiont Wolbachia are being introgressed into Aedes aegypti populations as a biocontrol method to reduce the transmission of medically important arboviruses. Successful introgression of Wolbachia relies on both persistence of Wolbachia throughout the host life cycle and a high fidelity of maternal transmission of Wolbachia between generations. wMel has been introgressed into field populations in 14 countries to date. Monitoring of field sites has shown that wMel prevalence can fluctuate substantially over time, prompting concerns this could lead to reduced efficacy of the biocontrol method. To explore the fidelity of wMel persistence and transmission, we developed molecular methods to measure the prevalence of Ae. aegypti negative for Wolbachia infection but carrying the "founder" mitochondrial haplotype of the single female first transinfected. As all released wMel-infected mosquitoes and any subsequent offspring will carry this founder mitochondrial haplotype, any mosquitoes with this mitochondrial haplotype and without wMel indicate that wMel was lost from this lineage at some point. We observed loss of wMel ranging from 0 to 20.4% measured at various time intervals after wMel-infected mosquito releases in five different countries. Despite some field sites showing Wolbachia loss, overall Wolbachia prevalence was sustained during the time periods studied. We then employed laboratory studies to explore factors that could contribute to the loss of wMel. Surprisingly, near-perfect maternal transmission was measured across laboratory conditions of early blood feeding, starvation, and salinity. Collectively, these findings underscore that although wMel transmission can be imperfect it does not necessarily undermine population-level establishment, providing encouragement that the intervention will be robust in most dengue-endemic environments.

Tropilaelaps spp. are a parasitic mite that feed and reproduce within honey bee brood (Apis spp.) and cause significant damage and mortality to Apis mellifera colonies. T. mercedesae is found outside the range of its natural host A. dorsata and was recently detected in Europe for the first time in 2024. It is widely believed that Tropilaelaps spp. are unable to survive without brood. However, studies have demonstrated that T. mercedesae can survive during broodless periods while parasitising A. dorsata and without brood in overwintering colonies of A. mellifera in temperate climates. This study sought to examine the survival of T. mercedesae on a range of matrices and found that mites could survive for more than 96 hours on live adult A. mellifera and more than 144 hours and 168 hours on decomposing A. mellifera pupae and adults respectively. These findings could indicate one possible mechanism which allows T. mercedesae to survive without brood. These findings also have implications towards better understanding possible transmission routes for Tropilaelaps. Previously bee sales in the form of queens and packages and used beekeeping equipment were considered a relatively low risk for transmitting Tropilaelaps spp. due to the absence of live brood. This study demonstrates Tropilaelaps ability to survive in these scenarios and the increased potential for it to spread globally.

Mating often changes behavior and physiology of female insects. In many species, these changes have been attributed to receipt of seminal fluid molecules (SFMs). SFMs influence phenotypes including feeding, egg production, and response to male courtship and insemination attempts. These same phenotypes are potential targets for management of insect pests. Aedes aegypti mosquitoes are the primary vector of several pathogens including dengue, Zika, and chikungunya. Within an hour after an initial insemination, female Ae. aegypti are generally refractory to subsequent inseminations, a response attributed to SFMs. However, the specific molecules involved in inducing long-term insemination refractoriness have not been identified. In a previous study, we identified adipokinetic hormone (AKH) precursor protein as an SFM in Ae. albopictus. AKH is a well-studied insect neuropeptide that impacts phenotypes including those related to metabolism, locomotion, and reproduction. In this study, we investigated whether AKH is an SFM in Ae. aegypti and whether it impacts female re-insemination patterns. We first established that AKH is produced in the male reproductive tract and transferred to females during mating, and is, therefore, an SFM. We then created an AKH-null line which allowed us to demonstrate that seminal fluid AKH contributes to long-term insemination refractoriness of females. Together, these results have established a novel expression pattern for AKH and identified AKH as a contributor to Ae. aegypti insemination refractoriness, laying the groundwork for understanding the evolution and mode of action of novel seminal fluid proteins as well as for investigating novel pathways or approaches for mosquito control.

The mosquito midgut is an important site for bloodmeal regulation while also acting as a primary site for pathogen exposure within the mosquito. Recent studies show that exposure to dehydrating conditions alters mosquito bloodfeeding behaviors as well as post-feeding regulation, likely altering how pathogens interact with the mosquito. Unfortunately, few studies have explored the underlying dynamics between dehydration and bloodmeal utilization, and the overall impact on disease transmission dynamics remains veiled. In this study, we find that dehydration-based feeding in the yellow fever mosquito, Aedes aegypti, prompts alterations to midgut gene expression, as well as subsequent physiological factors involving water control and post-bloodfeeding (pbf) regulation. Altered expression of ion transporter genes in the midgut of dehydrated mosquitoes and rapid reequilibration of hemolymph osmolality after a bloodmeal indicate an ability to expedite fluid and ion processing. These alterations ultimately indicate that female A. aegypti employ mechanisms to ameliorate the detriments of dehydration by imbibing a bloodmeal, providing an effective avenue for rehydration. Continued research into bloodmeal utilization and the resulting effects on arthropod-borne transmission dynamics becomes increasingly important as drought prevalence is increased by climate change.

Mating is a vital behavior for mosquito reproduction, yet it remains poorly understood under captive conditions. We examined the copulation dynamics of two key malaria vectors, Anopheles funestus, and Anopheles arabiensis, in controlled laboratory settings in Tanzania. We observed how variations in mosquito age and artificial lighting influence mating success for these two mosquito species within cages under controlled conditions. We conducted observations in 24-hour cycles, monitoring copulation events and insemination in females. We used generalized linear mixed models (GLMMs) for statistical analyses to assess how environmental conditions influence mating behavior. We found that Anopheles arabiensis exhibited rapid copulation, with 32.4% of individuals mating by Day 3 post-emergence, while An. funestus showed delayed activity, reaching a similar mating rate by Day 8. The introduction of artificial red light significantly accelerated copulation in An. funestus but did not affect An. arabiensis. Dissection confirmed successful sperm transfer and mating plug delivery in over 92% of copulating pairs for both species. Mating occurred primarily at night, with distinct peaks at 22:00 for An. arabiensis and 23:00 for An. funestus. In conclusion, our findings reveal species-specific differences in reproductive behavior, which could improve the colonization of An. funestus, a species historically challenging to rear in captivity. These insights may also inform the development of new vector control technologies, such as sterile insect techniques and genetic-based approaches, that exploit mosquito mating behaviors.

Aedes aegypti and Aedes albopictus are competent vectors of arboviruses such as dengue and chikungunya viruses which co-exist in some areas including La Reunion island. A kind of reproductive interference called satyrization has been described in sympatric species where a dominant species tends to control the spread of the other species. Here, we investigated satyrization in laboratory experiments to provide insights on the potential impact on Ae. aegypti of a control campaign including a sterile insect technique component against Ae. albopictus. Different mating crosses were used to test sympatric, conspecific-interspecific and allopatric effects of irradiated and non-irradiated male Ae. albopictus on female Ae. aegypti, including in a situation of skewed male ratio. Our results suggest that there was only a low level of satyrization between sympatric populations of Ae. aegypti and Ae. albopictus colonised from La Reunion island. A male Ae. albopictus to female Ae. aegypti ratio of 3:1 did not increase the level of satyrization. Female Ae. aegypti previously mated to male Ae. albopictus were not prevented from being inseminated by conspecific male Ae. aegypti. A satyrization effect was not seen between allopatric Ae. albopictus and Ae. aegypti strains from La Reunion Island either. The tested Ae. aegypti strain from La Reunion island has therefore developed full resistance to satyrization and so releasing sterile male Ae. albopictus may not suppress Ae. aegypti populations if an overflooding of irradiated male Ae. albopictus leads to similar results. The management strategy of two competent species in a sympatric area is discussed.

Mosquitoes display highly variable oviposition strategies and behavior. By using a high-speed camera, we detailly documented for the first time the egg-throwing strategy of the sylvatic yellow fever vector Sabethes albiprivus Theobald in laboratory. An oviposition trap made with sapucaia nut for field collection of tree-hole mosquitoes and obtaining Sa. albiprivus eggs either in the field or in the laboratory colony is described.

Temperature is a major factor that impacts tick populations by limiting geographic range of different species. Little is known about the thermal characteristics of these pests outside of a few studies on survival related to thermal tolerance. In this study, thermal tolerance limits, thermal preference, impact of temperature on metabolic rate, and temperature-activity dynamics were examined in larvae for six species of ixodid ticks. Tolerance of low temperatures ranged from -15 to -24{degrees}C with Dermacentor andersoni surviving at the lowest temperatures. High temperature survival ranged from 41 to 47 {degrees}C, with Rhipicephalus sanguineus having the highest upper lethal limit. Ixodes scapularis showed the lowest survival at both low and high temperatures. Thermal preference temperatures were tested from 0-41{degrees}C. D. variabilis exhibited a significant distribution of individuals in the lower temperatures, while the majority of other species gathered around 20-30{degrees}C. Activity was measured from 10-60{degrees}C, and the highest activity was observed in most species was near 30{degrees}C. Metabolic rate was the highest for most species around 40{degrees}C. Both activity and metabolic rate dropped dramatically at temperatures below 10{degrees}C and above 50{degrees}C. In summary, tick species vary greatly in their thermal characteristics, and our results will be critical to predict distribution of these ectoparasites with changing climates.

BackgroundThe control of vector-borne diseases is increasingly challenged by widespread insecticide resistance. Therefore, innovative vector control strategies with alternative modes of action are urgently needed. Ivermectin (IVM), an endectocide, has demonstrated toxicity to mosquito species such as Anopheles and Aedes when they feed on treated humans or livestock. In this study, we conducted a laboratory experiment to assess the effect of IVM, at concentrations equivalent to human plasma levels following mass drug administration (MDA), on the survival and fecundity of Anopheles coluzzii and Aedes aegypti in Burkina Faso. MethodsTwo laboratory experiments were conducted using 3-5-day old wild-derived female An. coluzzii and Aedes aegypti. Each experiment included four replicates per IVM concentration and was performed on separate dates. Mosquitoes were fed via membrane feeding on rabbit blood treated with five concentrations of IVM (C=112 ng/ml, C2=29 ng/ml, C3=15 ng/ml, C4=6.5 ng/ml, C5=2.5ng/ml), corresponding to the mean human plasma levels at 2, 4, 7, 14, and 28 days post-MDA with IVM at a dose of 300 g/kg. A negative control (C6=0.0 ng/ml) was also included. Mosquito mortalities were recorded daily for 7 days. Fecundity was measured by counting both laid eggs and developed eggs (via ovary dissection). ResultsIVM significantly reduced the survival of An. coluzzii compared to the control group (p<0.001), with the risk of death increasing from 4.2-fold at the lowest concentration (2.5 ng/ml) to 64.2-fold at the highest (112 ng/ml). In contrast, IVM had no significant effect on Aedes aegypti (p>0.05). Additionally, in An. coluzzii, IVM significantly reduced both egg laying and egg development (p<0.0001 and p<0.001, respectively), whereas no significant impact on fecundity was observed in Ae. aegypti (all p>0.80). ConclusionIvermectin concentrations typically achieved in human plasma during mass drug administration campaigns were sufficient to significantly reduce both survival and fecundity of wild type An. coluzzii, but had no measurable effect on recently colonized Ae. aegypti. These findings highlight the species-specific efficacy of ivermectin and support its potential role in integrated vector control strategies targeting malaria vectors in Africa.

Aedes (Aedimorphus) vexans (Meigen, 1830) is a globally distributed mosquito species commonly associated with temporary flooded habitats, including those in the South Pacific region. While its presence has been documented in several Pacific archipelagos, its occurrence on Tahiti island had remained unconfirmed. A retrospective examination of historical mosquito collections held at the Institut de Recherche pour le Developpement (IRD, France) and the Institut Louis Malarde (ILM, Tahiti) revealed evidence of Ae. vexans on Tahiti dating back as early as 1993. This historical detection was corroborated by targeted field surveys conducted between 2023 and 2025, which confirmed the species presence at multiple sites, particularly in the southern part of the island. To support these observations, COI gene sequences from newly collected specimens were analyzed alongside reference Ae. vexans sequences from GenBank. The Tahitian specimens fall unambiguously within the Pacific clade of Ae. vexans and show no genetic divergence that would support their assignment to Ae. nocturnus or Ae. nipponii. Similarly, sequences published under these names in GenBank do not form distinct lineages within Ae. vexans. This study provides the first confirmed record of Ae. vexans on Tahiti, based on both morphological identification and molecular evidence. The findings underscore the need to integrate molecular tools into entomological monitoring to improve species detection and prevent misidentifications in island environments where vector diversity is often underestimated.

Tropilaelaps mercedesae is a parasitic mite species that negatively affects the health of the Apis mellifera colonies. Recent reports show that it is spreading westwards, through Central Asia into Europe. Several field and laboratory methods have been proposed to detect Tropilaelaps spp. in A. mellifera colonies; however, most of them are either laborious, costly or ineffective. Here we introduce a novel method for detecting T. mercedesae based on the mites characteristic biology (reduced feeding as bee pupae mature, brief dispersal phase on adult bees and agility) and the use of wax strips. Sealed worker brood cells at the development stage of white to purple-eyed pupae are swiftly decapped with wax strips to observe and count surfacing adult mites. Our study shows over 90 % detection efficacy and brood survival, and ease of application. Therefore, we recommend the novel "Rapid brood decapping" method as a reliable tool for detecting and monitoring T. mercedesae infestation.

Zika is a vector-borne disease caused by an arbovirus (ZIKV) and overwhelmingly transmitted by Ae. aegypti. This disease is linked to adverse fetal outcomes, mostly microcephaly in newborns, and other clinical aspects such as acute febrile illness and neurologic complications, for example, Guillain-Barre syndrome. One of the actual most promising strategies to mitigate arbovirus transmission involves releasing Ae. aegypti mosquitoes carrying the maternally inherited endosymbiont bacteria Wolbachia pipientis. The presence of Wolbachia is associated with a reduced susceptibility to arboviruses and a fitness cost in mosquito life-history traits as fecundity and fertility. However, the mechanisms by which Wolbachia influences metabolic pathways leading to differences in egg production remains poorly known. To investigate the impact of co-infections on the reproductive tract of the mosquito, we applied an isobaric labeling-based quantitative proteomic strategy to investigate the influence of Wolbachia wMel and ZIKV infection in Ae. aegypti ovaries. To the best of our knowledge, this is the most complete proteome of Ae. aegypti ovaries reported so far, with a total of 3,913 proteins identified, also,were able to quantify a total of 1,044 Wolbachia proteins in complex sample tissue of Ae. aegypti ovary.. Furthermore, we discuss proteins and pathways altered in Ae. aegypti during ZIKV infections, Wolbachia infections, co-infection Wolbachia/ZIKV, and compared with no infection, focusing on immune and reproductive aspects of Ae. aegypti. The modified aspects were mostly related to the immune priming enhancement by Wolbachia presence and the modulation of the Juvenile Hormone pathway caused by both microorganisms infection. HighlightsO_LIProteome changes in Ae. aegypti, Wolbachia, and ZIKV interactions C_LIO_LIA great diversity of Wolbachia proteins were quantified in Ae. aegypti ovary C_LIO_LIJuvenile Hormone pathway is modulated by both infections C_LIO_LIWolbachia enhances Ae. aegypti immune priming mechanism C_LIO_LIZIKV unsettles host immune response by reducing antimicrobial peptides production C_LIO_LICoinfection triggers oxidative stress and a lack of vitellogenin precursors C_LI