Determining the minimum postmortem interval (minPMI) from an entomological perspective relies mainly on development data recorded for various species of flies collected from a crime scene or suspicious death. This study focused on the larval and pupal development of Lucilia sericata (Meigen), with an emphasis on the changes of the external morphology of the puparium and its pupal content throughout the duration of metamorphosis. Colonies of L. sericata were reared on 3 types of swine tissue (skeletal muscle, liver tissue, and heart tissue) at 2 different temperature regimes; 24 ± 1 °C and 30 ± 1 °C. The overall developmental time, larval width and length, and inner and outer pupal morphology changes were observed and recorded. The results show that: (i) temperature significantly influenced overall development time, as well as changes in larval width and length, but this effect was not dependent on tissue type; (ii) larval development duration was longest on heart tissue, and shortest on skeletal muscle for both temperatures; and (iii) pupation was longest for larvae reared on skeletal muscle at 24 ± 1 °C, and on liver tissue at 30 ± 1 °C, while those larvae reared on liver tissue at 24 ± 1 °C and heart tissue at 30 ± 1 °C had the shortest pupation period. A seven-character checklist plus 4 landmark stages were developed comprising the external morphology of the puparium and pupal content changes of L. sericata. In conclusion, the study provides larval and pupal development timetables, as well as checklists and photo guides for pupal character development that may be useful for future postmortem determinations.

Numerous factors influence insect colonization and development on decomposing remains and can influence postmortem interval (PMI) estimations when insects are used in investigations of suspicious deaths. This study aimed to determine whether insect repellent influences insect colonization of decomposing remains. Pig feet were divided into 4 groups: a control not sprayed with repellent, 40% DEET repellent, 98% DEET repellent, and lemon eucalyptus repellent. The feet were placed in containers, and insect visitation was observed over the course of 3 days, then insects were collected at the end of the third day. This was repeated twice a month from April through November 2022. The analysis focused on the order Diptera. Pairwise comparison determined that there was a significant delay in visitation time for remains with 98% DEET compared to remains with no repellent and 40% DEET. There were significantly fewer adult Diptera collected and more remains with no larvae or eggs present among remains treated with 98% DEET. It was concluded that low concentrations of DEET and lemon eucalyptus repellent did not delay colonization, but that high concentrations of DEET delayed visitation, reduced the number of adult flies collected from the remains, and reduced the likelihood of collecting eggs or larvae from the remains. This contributes to the understanding of factors that can influence insect colonization and development that allow for more accurate PMI estimations by suggesting that insect repellent does not need to be accounted for in PMI estimation, but a high concentration of DEET can deter insects in the short term.

Many disciplines are utilized within the field of veterinary forensic sciences, including forensic entomology. Understanding the initial colonization period by flies of forensic importance can contribute to estimating the minimum postmortem interval. There is limited data regarding the time of colonization of animals with fur, and the interpretation of this data is difficult due to the variation in animal models used. The purpose of this study was to examine the initial insect colonization of cats (Felis catus), with light and dark fur. Twelve domestic short-haired cats were placed in cages 15.2 m apart in a grassy field in West Lafayette, IN, United States. Weather data (temperature, precipitation, sun/cloud exposure, humidity), insect activity, time to oviposition, and decomposition changes were documented. Eggs from initial oviposition events were collected and reared to identify the primary colonizing species. Although the time of first oviposition event was not different between the treatments, fur color did affect fly colonization, and cats with dark fur had more oviposition events than cats with light fur (t = 2.639, df = 4, P = 0.029). Three species of Lucilia (Diptera: Calliphoridae) colonized the cats on the initial day of placement. Further studies in cats should include the decompositional studies to understand the unique characteristics that occur during each stage of decomposition, which could aid in developing a scoring system for animals with fur. Additional studies could include analyzing how fur length would affect colonization.

Proper fixing and long-term preservation of entomological evidence are essential in collections and research and crucial in applied fields such as forensic entomology. Incorrectly stored samples may lose important morphological features over time, rendering molecular analyses exceedingly difficult. The most effective method for preserving soft samples such as larvae is fluid preservation. It uses a combination of a wide range of fixatives and storage fluids. However, very little comparative work has been done to determine the effects of long-term storage on sample quality in terms of color, shape, and DNA stability. Moreover, the current golden standard in forensic entomology has been tailored for age estimation of larvae of Diptera, which differ from larvae of Coleoptera in morphology and subsequently in applied methods. We compared the effects of combinations of 6 commonly used fixatives and 6 commonly used storage fluids on midsized larvae of the forensically important beetle, Necrodes littoralis (Linnaeus, 1758), in terms of color, shape, and suitability for DNA analyses over a 2-yr period. We were looking for combinations that can preserve specimens in a satisfactory state, can be used on a regular basis, do not require advanced protection or skills of the personnel, and are not toxic or too harmful to the environment. We found not only several methods that scored significantly better in the tested parameters compared with the golden standard but also several common methods that should be avoided. The effects of agents on each tested category are discussed in detail.

The genetic structure of forensically important blow fly (Brauer & Bergenstamm) (Diptera: Calliphoridae) populations has remained elusive despite high relatedness within wild-caught samples. This research aimed to determine if the implementation of a high-resolution spatiotemporal sampling design would reveal latent genetic structure among blow fly populations and to elucidate any environmental impacts on observed patterns of genetic structure. Adult females of the black blow fly, Phormia regina (Meigen) (Diptera: Calliphoridae), were collected from 9 urban parks in Indiana, USA over 3 yr and genotyped at 6 polymorphic microsatellite loci. The data analysis involved 3 clustering methods: principal coordinate analysis (PCoA), discriminant analysis of principal components (DAPC), and STRUCTURE. While the PCoA did not uncover any discernible clustering patterns, the DAPC and STRUCTURE analyses yielded significant results, with 9 and 4 genetic clusters, respectively. Visualization of the STRUCTURE bar plot revealed N = 11 temporal demarcations indicating barriers to gene flow. An analysis of molecular variance of these STRUCTURE-inferred populations supported strong temporally driven genetic differentiation (F_ST = 0.048, F'_ST = 0.664) relative to geographic differentiation (F_ST = 0.009, F'_ST = 0.241). Integrated Nested Laplace Approximation and Boosted Regression Tree analyses revealed that collection timepoint and 4 main abiotic factors (temperature, humidity, precipitation, and wind speed) were associated with the genetic subdivisions observed for P. regina. A complex interplay between environmental conditions, the unique reproductive strategies of the blow fly, and the extensive dispersal abilities of these organisms likely drives the strong genetic structure of P. regina in the Midwestern US.

The field of veterinary entomology is dominated by research concerning insects and arthropods that negatively impact the health of domestic animals. A curated selection of peer-reviewed research which was highlighted at the 2022 Joint Meeting between the Entomological Society of America, the Entomological Society of Canada, and the Entomological Society of British Columbia, which prioritized exploring entomology through the lens of art, science, and culture are summarized. Articles are categorized into (i) biting and non-biting flies, (ii) ectoparasites, and (iii) review articles, with the overall goal of showcasing innovative methodologies, addressing overlooked questions in veterinary entomology, and highlighting comprehensive reviews. While not exhaustive, the selected studies represent a myriad of arthropods, methodologies, and perspectives, to inspire future scientists with diverse research avenues and emphasize the continual evolution and importance of entomological studies in today's world.

In efforts to intensify malaria control through vector control and hasten the progress towards elimination, the impact of control interventions needs to be evaluated. This requires sampling vector population using appropriate trapping methods. The aim of this article is to critically review methods of sampling malaria vectors and their reliability in estimating entomological indicators of malaria transmission in Africa. The standard methods are human landing catch (HLC), pyrethrum spray catch, and pit shelter for sampling host-seeking, indoor resting, and outdoor resting malaria vectors, respectively. However, these methods also have drawbacks such as exposure of collectors to infective mosquito bites, sampling bias, and feasibility issue. Centers for Disease Control and Prevention (CDC) light traps placed beside human-occupied bed nets have been used as an alternative to the HLC for sampling host-seeking malaria vectors. Efforts have been made to evaluate the CDC light traps against HLC to generate a conversion factor in order to use them as a proxy estimator of human biting rate and entomological inoculation rates in Africa. However, a reproducible conversion factor was not found, indicating that the trapping efficiency of the CDC light traps varies between different geographical locations. Several other alternative traps have also been developed and evaluated in different settings but most of them require further standardization. Among these, human-baited double net trap/CDC light trap combination and mosquito electrocuting trap have the potential to replace the HLC for routine malaria vector surveillance. Further research is needed to optimize the alternative sampling methods and/or develop new surveillance tools based on vector behavior.

Insecticide resistance is a great challenge facing mosquito operational control agencies across the United States, where few active ingredients with unique modes of action are available for use, increasing resistance pressure and further hampering resistance management strategies. Emergence and expansion of insecticide resistance in mosquitoes can be detected by resistance monitoring programs; however, there are gaps in our knowledge regarding the link between resistance bioassay results and operational control outcomes. Here, we review both public health and agricultural studies on pesticide resistance bioassays and control outcomes. A discussion on the main gaps in our knowledge of insecticide resistance and a review of resistance management practices is also presented. We conclude with research questions that can advance our understanding of resistance monitoring and control.

Vector-borne disease incidence and burden are on the rise. Weather events and climate patterns are known to influence vector populations and disease distribution and incidence. Changes in weather trends and climatic factors can shift seasonal vector activity and host behavior, thus altering pathogen distribution and introducing diseases to new geographic regions. With the upward trend in global temperature, changes in the incidence and distribution of disease vectors possibly linked to climate change have been documented. Forecasting and modeling efforts are valuable for incorporating climate into predicting changes in vector and vector-borne disease distribution. These predictions serve to optimize disease outbreak preparedness and response. The purpose of this scoping review was to describe the use of climate data in vector-borne disease prediction in North America between 2000 and 2022. The most investigated diseases were West Nile virus infection, Lyme disease, and dengue. The uneven geographical distribution of publications could suggest regional differences in the availability of surveillance data required for vector-borne disease predictions and forecasts across the United States, Canada, and Mexico. Studies incorporated environmental data from ground-based sources, satellite data, previously existing data, and field-collected data. While environmental data such as meteorological and topographic factors were well-represented, further research is warranted to ascertain if relationships with less common variables, such as oceanographic characteristics and drought, hold among various vector populations and throughout wider geographical areas. This review provides a catalogue of recently used climatic data that can inform future assessments of the value of such data in vector-borne disease models.

To accurately model the food webs, we need to acquire precise data on food ecology of the interacting species. This allows better understanding of the trophic interactions and for the necrophagous species this information could be used in medico-legal investigations. For this reason, we recently proposed standardized laboratory methodology to assess the foraging strategies based on parallel testing of 2 food items (meat, dead larvae) (Jakubec et al. 2021). The original methodology had 2 shortcomings. It was not suited for testing living larvae, which could prove predatory behavior of the species. The methodology was also based on parallel experimental design, where the food items are tested together, which could underestimate the maximum consumption of the tested subject for some items. To test if these concerns are valid, we improved original methodology allowing testing living larvae as well as a new sequential experimental setup, where consumption of each item is tested individually in a random order, thus theoretically giving an unbiased maximum consumption estimate. These methodologies were tested head-to-head on 3 forensically relevant species from the genus Thanatophilus (Thanatophilus micans (Fabricius 1794)(Fabricius 1794), Thanatophilus rugosus (Linnaeus, 1758), and Thanatophilus sinuatus(Fabricius, 1775)). The experiments have confirmed that all 3 species are almost strictly necrophagous, although they were capable of predation, despite the presence of preferred food (meat). The comparison also showed that the sequential design has indeed improved capability to quantify the maximal consumption of the given food item. Thus, we suggest following this methodology in future studies.

The growth and associated traits of Aedes aegypti (L.) mosquitoes may adapt and evolve in response to the costs associated with body size in relation to latitudinal variation. We analyzed the life-history traits and energy reserves of field-collected mosquitoes from Taiwan, Thailand, and Indonesia along a latitudinal range spanning from 23°N to 6°S. A U-shaped relationship between body size and latitude was observed. Our study demonstrated the role of latitudinal temperature variations in determining the body size patterns of Ae. aegypti. Notably, the body size of the northern populations (from Taiwan) was significantly larger than those of the tropical populations from Thailand and Indonesia. Models have demonstrated that regional precipitation levels may contribute to body trait variations in certain high-latitude populations in Thailand. However, Indonesian populations have high development rates and large body sizes, indicating the involvement of other physiological traits in determining mosquito body size. The reproductive output of the adult females in this study was positively correlated with body size, but our measure of longevity did not covary significantly with the body size. By contrast, the reproductive output of mosquito-tested populations was in inverse proportion to longevity. Additionally, the mean teneral glycogen levels in the Indonesian and Thai populations were 2.5 times higher than those in theTaiwanese populations. The Indonesian and Thai populations had low mean generation and doubling times, resulting in a high intrinsic rate of increase compared with that of the Taiwanese populations, despite the Taiwanese populations having the highest net reproduction rate.

Graphical Abstract

Following several days of blood feeding by larval and nymphal ixodid (hard) ticks, the salivary glands degenerate and are completely replaced in the next life stage. Yet, what happens during the molt of immature argasid (soft) ticks after their rapid and small bloodmeal has remained a mystery. Multiple studies of nymphal Ornithodoros hermsi Wheeler (Acari: Argasidae) ticks infected with the relapsing fever spirochete Borrelia hermsii suggested the salivary glands in these ticks may not disintegrate after feeding. Therefore, cohorts of second-stage O. hermsi nymphs were fed and examined daily after the bloodmeal by fresh dissections and weekly by histological cross-sections of the entire tick. The composition of the salivary glands was typical for argasid ticks in having agranular (Type I) and granular (Type II) acini, the latter being surrounded by a myo-epithelial sheath. In all 197 ticks examined from 1 to 63 days after feeding, morphologically intact salivary glands were present. During apolysis, 5 ticks had extralimital clusters of granular acini adhering to otherwise intact glands. Our observations demonstrate that the salivary glands of nymphal O. hermsi do not disintegrate after feeding and new acini are produced during the molt for incorporation into the existing glands. Cumulatively, these findings suggest a fundamental difference in the transstadial development of argasid and ixodid ticks.

There is growing interest in insecticide resistance in the mosquito, Aedes albopictus (Skuse), as its potential for spreading diseases is increasing as urbanization and control efforts intensify. Here we review the presence and diversity of mutations in the voltage-sensitive sodium channel (Vssc) gene associated with pyrethroid resistance and report on additional surveys of these mutations in new populations with an analysis of their spread. The known diversity of these mutations has increased in recent years including the identification of 26 non-synonymous mutations, although phenotypic data associating mutations with resistance remain limited. We provide data on mutations in several new locations including those inTimor Leste, Indonesia, and Vanuatu. We use population genomic data from ddRAD analyses of target populations with the 1534C mutation to identify single nucleotide polymorphisms (SNPs) associated with the mutant to test for clustering of SNPs based on the presence of the 1534C mutation rather than population origin. Our findings suggest spread of resistance alleles via genetic invasion, which is further supported by patterns from a genome-wide principal components analysis. These data point to movement of resistance alleles across wide areas with likely impacts on local control options.

In temperate regions of the United States, female Anopheles mosquitoes respond to low temperatures and short photoperiods by entering an overwintering dormancy or diapause. Diapause in Anopheles results in reduced frequency of blood-feeding and reproductive arrest, indicating a period when pathogen transmission by these mosquitoes is unlikely. However, it is unclear precisely how late into the fall and how early in the spring these mosquitoes are biting, reproducing, and potentially transmitting pathogens. This is further complicated by the lack of clear markers of diapause in Anopheles (e.g., changes in egg follicle length). Our goal was to characterize the seasonal reproductive activity of female Anopheles in central Ohio, United States and evaluate egg follicle length as an indicator of Anopheles diapause. We used traditional mosquito traps and aspirators to collect Anopheles from urban woodlots and culverts, respectively, from late September 2021 through mid-May 2022 in central Ohio. By measuring their egg follicle length, reproductive status, and blood-feeding status, we found that egg follicle length is not a reliable indicator of Anopheles diapause. We also found that a small proportion of An. punctipennis (Say), An. perplexens (Ludlow), and An. quadrimaculatus (Say) continued to bite and reproduce into early November 2021 and that females of these species terminated reproductive dormancy and began biting by mid-March 2022. This period of reproductive activity extends beyond current mosquito surveillance and control in Ohio. Our findings suggest that within temperate regions of North America, Anopheles have the capacity to transmit pathogens throughout the spring, summer, and fall.

Water mites (Hydrachnidia) commonly parasitize mosquitoes; however, the nature of these parasitic interactions remains poorly understood. We sampled mites collected from mosquitoes taken from CDC light traps placed in a botanical garden in Northern Florida from April to November 2022. Mites were found almost exclusively parasitizing the mosquitoes, Anopheles crucians sensu lato (Wiedemann, 1828), Anopheles quadrimaculatus sensu lato (Say, 1824), and Culex erraticus (Dyar and Knab, 1906). All sampled mites were of the genus Arrenurus. Further identification proved to be impossible given the available resources. Seasonality of the mites corresponded with the seasonality of their hosts, with the highest numbers being recorded in May and September. Nomenclature for mite attachment sites on mosquitoes was developed and provided. Mites most commonly attached to the second abdominal segments of all sampled mosquitoes with varied positions around the segment depending on mosquito species. We found significance for the relationship between the abdominal segment mites attached to and what position on the segment mites would take for Cx. erraticus, which indicates a preference of attaching directly underneath the second and fourth abdominal segments. Such a relationship was not found for either Anopheles species.

A spatiotemporal investigation of hematophagous fly prevalence was conducted over a 1-year period on 12 beef cattle farms located in major livestock areas of Bangkok,Thailand, using Vavoua traps. The survey revealed 5,018 hematophagous flies belonging to Muscidae and Tabanidae, with the 3 dominant species identified as Stomoxys calcitrans (Linnaeus) (2,354; 46.91%), Musca crassirostris Stein (1,528; 30.45%), and Haematobia exigua de Meijere (922; 18.37%). The abundance of S. calcitrans per trap per week was significantly higher during the rainy season (45.64 ± 14.10), followed by the cold and dry seasons (6.39 ± 2.16 and 3.04 ± 1.27, respectively). The relative abundance of S. calcitrans reached the highest apparent density per trap per day (ADT) index of 9.83 in August 2022 during the rainy season. Subsequently, there was a rapid decline, and the ADT index dropped to nearly zero in December 2022 during the cold season. This low abundance continued through the dry months from March to May 2023. The higher rainfall and relative humidity could significantly contribute to the high relative abundance of S. calcitrans. In contrast, M. crassirostris and H. exigua showed population fluctuations that were not significantly associated with seasonal changes and weather conditions. Remote sensing data and spatial regression analyses using ordinary least squares regression showed the high spatial density of S. calcitrans in the north direction of the Khlong Sam Wa district during the rainy season; it shifted toward the south in the cold and dry seasons, corresponding with rainfall.

Volatile pyrethroids are effective in reducing mosquito populations and repelling vectors away from hosts. However, many gaps in knowledge exist for the sublethal impacts of volatile pyrethroids on mosquitoes. To that end, transfluthrin exposures were conducted on a field strain of Aedes albopictus (Skuse) held as a laboratory colony. Dose–response analysis was conducted on both sexes at either 1–4 days old or 5–10 days old. Resultant concentration data were used to evaluate the LC₂₀ and LC₅₀ values in various mate pairings of treatments and controls in which either the male or female was from a selectively treated group and mated with a counterpart that was treated independently. Blood feeding proportion, delayed mortality after a 24-h recovery period, egg collection totals, and F₁ larval survival were determined following transfluthrin treatment in the F₀, but outcomes were not significant. In contrast, sterility was predicated on male treatment, with treated females resulting in higher overall egg viability. Treated males in the mating pair resulted in significantly lower egg viability and accelerated larval hatch in the F₁. Additionally, the presence of sperm in female spermathecae was significantly diminished in test groups containing treated male mosquitoes. Male sublethal effects may be a critical determinant of a mixed population's reproductive success.

Forest management practices designed to meet varied landowner objectives affect wildlife habitat and may interrupt the life-cycle stages of disease vectors, including the black-legged tick, Ixodes scapularis Say (Acari: Ixodidae). Ixodes scapularis transmits multiple pathogens including Borrelia burgdorferi, the causative agent of Lyme disease, which is the most common tick-borne disease in the United States. There is evidence that a range of active forest management practices (e.g., invasive plant removal, prescribed burning) can alter tick densities and pathogen transmission. However, few studies have investigated relationships between forest stand structural variables commonly manipulated by timber harvesting and tick ecology. Foresters may harvest timber to create certain forest structural conditions like the mean number of trees, or basal area, per hectare. This study used a spatially replicated experiment in a blocked design to compare forest stands with a range of overstory structures and document variations in the midstory, understory, and forest floor, as well as microclimate conditions within tick off-host habitat. Greater numbers of trees or basal area per hectare correlated with greater canopy closure but less understory cover, stabilized microclimate temperature, higher microclimate humidity, and greater I. scapularis nymph densities. A random forest model identified understory forest structure as the strongest predictor of nymph densities. There was no relationship between the number of trees or basal area per hectare and daily deer (Odocoileus virginianus Zimmermann) activity or nymphal infection prevalence. These findings provide a deeper understanding of tick-habitat associations within a forest stand and have the potential to inform forest management decisions.

Insecticide resistance in mosquito populations has long been recognized as a significant global public health challenge, motivating the development of new control chemistries. ReMoa Tri is a novel triple-action space spray that employs a different mode of action than traditional adult mosquito control formulations. It combines 3 components: fenpropathrin, a mixed-type I/II pyrethroid; abamectin, a macrocyclic lactone; and C8910, a patented fatty acid chain. As an ultra-low volume adulticide, ReMoa Tri has the potential to target mosquito species that are resistant to pyrethroid and organophosphate-based control materials. To determine whether ReMoa Tri effectively targets resistant mosquito species in Florida's Collier County, United States, we conducted ground-based field cage trials using field-caught pyrethroid-resistant Culex quinquefasciatus (Say) and Aedes aegypti (L.), of which the latter also displayed developing resistance to organophosphates. Trials were also conducted against the same mosquito populations with Merus 3.0, a pyrethrin-based adulticide used by the Collier Mosquito Control District. ReMoa Tri was effective against Collier's pyrethroid-resistant Cx. quinquefasciatus, resulting in more than 95% mortality in semifield cage trials by 24 h postapplication. Similarly, ReMoa Tri applications against Collier's pyrethroid-resistant Ae. aegypti resulted in 72%–89% mortality at 24 h postapplication and 74%–97% mortality at 48 h postapplication. This study represents the first field data on this novel space spray, and its findings shed light on the performance of ReMoa Tri against local mosquito populations that have developed resistance to currently available adulticides.

Long-lasting insecticide nets (LLINs) are the recommended tools against mosquito-borne diseases. However, their physical integrity and bioefficacy in the field could be affected by several factors. This study evaluated the physical integrity and bioefficacy of nets used in Makenene since 2016. Cross-sectional field surveys were carried out after 6 y. A questionnaire was first administered to the heads of households, and then the physical integrity of the LLINs was determined by calculating the proportional hole index (pHI). WHO cone bioassays were conducted to determine the bioefficacy of LLINs currently being used against wild strains of Anopheles gambiae s.l., Culex pipiens s.l., and laboratory-reared pyrethroid-susceptible strain of Anopheles coluzzii (Ngousso). Of the 167 LLINs examined in households, 39.5% were fairly good, 26.4% were acceptable, and 34.1% were damaged. The most torn faces of the nets were the sides used for entering and exiting. None of the 30 LLINs used for WHO cone bioassays was still effective against An. gambiae s.l. and Cx. pipiens s.l. while up to 85.7% of these LLINs were at least effective against the susceptible strain after 24 h, with a significant difference observed when comparing the mortality rates between wild and laboratory-susceptible strain of Anopheles (P-value < 0.01). Anopheles gambiae s.l. were all (100%) identified as An. gambiae s.s. by PCR. The LLINs distributed in Makenene since the 2016 campaign are only effective on susceptible strain and should be replaced for a better control of residual malaria transmission and the nuisance by Culex mosquitoes in the locality.

The emergence of insecticide resistance in mosquitoes necessitates the exploration and validation of sustainable biological strategies for controlling mosquitoes in their natural habitats. We assessed the predatory effect of Utricularia aurea Lour (Lamiales: Lentibulariaceae), an aquatic carnivorous plant found in the Indian subcontinent, Japan, and Australia, on 4 instars of Anopheles stephensi Liston, Culex quinquefasciatus Say, and Aedes aegypti Linn (Diptera: Culicidae), in the laboratory and field settings. In the laboratory setting, predation of larvae by U. aurea was highest during the first hour when it predated 45%, 61%, and 58% of first instars of An. stephensi, Cx. quinquefasciatus, and, Ae. aegypti, respectively, and, within 12 h, U. aurea preyed upon ∼95% of the first, second, and third instars of the 3 mosquito species, ∼80% of the fourth instars of An. stephensi and Ae. aegypti, and ∼60% of fourth instars of Cx. quinquefasciatus. The predatory effect of U. aurea varied with mosquito species and instar. Broadly, predation risk declined with the increase of the instar size. In the field setting, at the end of 16 days, U. aurea predated 76% and 71% of the immature An. stephensi and Ae. aegypti, respectively. Our findings suggest U. aurea can be utilized as a potential biocontrol agent for controlling mosquito larvae in natural habitats; however, the current claim warrants additional investigations in a variety of natural habitats.

Culex panocossa, Dyar and Knab, an important enzootic vector of Venezuelan equine encephalitis virus subtype ID in Central and South America, was found to have invaded and become established in southern Florida in 2016. No information is currently available regarding the ecology of this invasive mosquito in the United States. Here, we use PCR-based blood meal analysis to investigate vertebrate host associations of Cx. panocossa from Florida to provide information necessary for determining the potential importance of this mosquito for arbovirus transmission in the United States. Culex panocossa fed mainly upon birds (49.5%) but took a substantial fraction of blood meals from mammals (33.3%) and reptiles (17.1%). By feeding upon amplifying hosts of Everglades virus (hispid cotton rat) and eastern equine encephalitis virus (wading birds) and humans, Cx. panocossa could act as a bridge vector for these pathogenic Alphaviruses in Florida, potentially resulting in increased human disease.

Spiroplasma (Mycoplasmatales: Spiroplasmataceae) is one of the most widely distributed symbionts of arthropods. Spiroplasma species can infect their hosts via vertical or horizontal transmission. However, the mode of transmission of Spiroplasma between different arthropod taxa has not been elucidated. In this study, we investigated the potential for the transmission of Spiroplasma to non-native arthropod species, using 2 Spiroplasma spp. isolated from ticks, namely Spiroplasma ixodetis and Spiroplasma mirum, and 3 species of mosquito laboratory colonies, namely Aedes albopictus, Aedes aegypti, and Culex pipiens pallens (Diptera: Culicidae).After feeding the adult mosquitoes with Spiroplasma-containing artificial meals, they were kept at 25 °C for 10 days. Homogenates prepared from Spiroplasma-fed mosquitoes were used to re-isolate Spiroplasma using the in vitro culture method. Nine weeks after culture initiation, the presence of Spiroplasma was tested using the polymerase chain reaction (PCR).The results revealed that only S. ixodetis was detected from all 3 species of mosquitoes and re-isolated from 2 of them.The differences in the infection ability of different Spirolasma species could be attributed to several factors, including environmental effects. Nevertheless, this is the first experimental demonstration of Spiroplasma transmission among different arthropod taxa. Further studies are needed to elucidate the evolutionary mechanism that supports the survival of Spiroplasma in nature.

Graphical Abstract

Arthropod-derived cell lines serve as crucial tools for studying arthropod-borne viruses (arboviruses). However, it has recently come to light that certain cell lines harbor persistent infections of arthropod-specific viruses, which do not cause any apparent cytopathic effects. Moreover, some of these persistent viral infections either inhibit or promote the growth of arboviruses. Therefore, it is of utmost importance to identify the presence of such persistent viruses and understand their impact on arboviral infections. In this study, we conducted a comprehensive virome analysis of several arthropod-derived cell lines, including mosquito-derived NIID-CTR, Ar-3, MSQ43, NIAS-AeAl-2, CCL-126 cells, and tick-derived IDE8 cells, along with flesh fly-derived NIH-Sape-4 cells. The aim was to determine if these cells were infected with persistent viruses. The results revealed the presence of 15 persistent viruses in NIID-CTR, Ar-3, MSQ43, NIAS-AeAl-2, and IDE8 cells. Among these, 11 were already known arthropod-specific viruses, while the remaining 4 were novel viruses belonging to Orthophasmavirus, Rhabdoviridae, Totiviridae, and Bunyavirales. In contrast, CCL-126 and NIH-Sape-4 cells appeared to be free of viral infections. This study provides valuable insights into the diversity and latency of arthropod-specific viruses within arthropod-derived cell lines. Further investigations are required to explore persistent viral infections in other arthropod-derived cell cultures and their effects on arbovirus replication. Understanding these factors will enhance the accuracy and reliability of experimental data obtained using these cell lines.

Biting midges in the genus Culicoides Latreille (Diptera: Ceratopogonidae) are known to transmit many pathogens of veterinary and medical concern. Although much work has been done globally and in certain regions of North America, Culicoides spp. research in rural Appalachia is limited. To begin characterizing the distribution and community structure of Culicoides spp. in Appalachia, we surveyed 2 distinct sites in the Ridge and Valley ecoregion of northeastern Tennessee, USA, from April 2021–September 2021. Culicoides spp. were sampled using 2 methods: Centers for Disease Control ultraviolet LED light traps and potential larval habitat substrate collection (coupled with water chemistry values). Site 1 was dominated by natural features, and Site 2 was a beef cattle operation. During 96 trap nights, a total of 1,568 Culicoides were collected, representing 24 species. Site 1 yielded the highest diversity, with 24 species, while Site 2 yielded 12 species. Overall, the most abundant species in light traps were C. stellifer Coquillett (44%), C. bergi Cochrane (18%), C. haematopotus Malloch (12%), and C. debilipalpis Lutz (11%). From substrate sampling, 8 species were identified. Culicoides haematopotus was the most abundant and was collected during each sampling period. Water chemistry values taken at the time of substrate collection were not significantly related to which Culicoides spp. emerged from a given substrate. Our results indicate a diverse community of Culicoides spp. in our study area, however, further work is needed to identify Culicoides species composition across a variety of landscapes in Appalachia and inform research on vector presence and associated vector disease dynamics.

Three Asian longhorned ticks (Haemaphysalis longicornis) were collected on Staten Island, Richmond County, New York, in 2014–2015 as part of a tick-borne disease surveillance program conducted by the New York City Department of Health and Mental Hygiene and the Defense Centers of Public Health – Aberdeen Tick-Borne Disease Laboratory. These records mark the earliest known occurrence of H. longicornis in New York State outside of quarantine areas, predating previously reported detections by several years. Robust populations of H. longicornis were collected in subsequent years at the Staten Island site where these few ticks were found, demonstrating that small infestations have the potential to proliferate quickly. Haemaphysalis longicornis is a 3-host ixodid tick native to eastern Asia but now established in the United States, as well as Australasia and several Pacific islands. Although H. longicornis has not yet been associated with human disease transmission in the United States, it warrants attention as a potential vector, as it is demonstrated to harbor various pathogens of medical and veterinary interest across its native and introduced range.

A total of 2,504 ticks of 5 species (Ixodes scapularis, Dermacentor variabilis, Amblyomma americanum, Haemaphysalis leporispalustris, and H. longicornis) were collected over 2 yr (2014–2015) in New York City parks. Specimens were collected via tick-dragging, identified to species, and tested for pathogens of human diseases. The causative agents of 5 human diseases (Lyme borreliosis, ehrlichiosis, babesiosis, anaplasmosis, and Rocky Mountain spotted fever) were detected in a subset of samples. Results of this surveillance effort further illustrate that risk of tick-borne disease is considerable even in parks located adjacent to densely populated areas.

The Pacific Coast tick (Dermacentor occidentalis Marx, 1892) is a frequently encountered and commonly reported human-biting tick species that has been recorded from most of California and parts of southwestern Oregon, southcentral Washington, and northwestern Mexico. Although previous investigators have surveyed populations of D. occidentalis for the presence of Rickettsia species across several regions of California, populations of this tick have not been surveyed heretofore for rickettsiae from Baja California, Oregon, or Washington. We evaluated 1,367 host-seeking, D. occidentalis adults collected from 2015 to 2022 by flagging vegetation at multiple sites in Baja California, Mexico, and Oregon and Washington, United States, using genus- and species-specific assays for spotted fever group rickettsiae. DNA of Rickettsia 364D, R. bellii, and R. tillamookensis was not detected in specimens from these regions. DNA of R. rhipicephali was detected in D. occidentalis specimens obtained from Ensenada Municipality in Baja California and southwestern Oregon, but not from Washington. All ompA sequences of R. rhipichephali that were amplified from individual ticks in southwestern Oregon were represented by a single genotype. DNA of the Ixodes pacificus rickettsial endosymbiont was amplified from specimens collected in southwestern Oregon and Klickitat County, Washington; to the best of our knowledge, this Rickettsia species has never been identified in D. occidentalis. Collectively, these data are consistent with a relatively recent introduction of Pacific Coast ticks in the northernmost extension of its recognized range.

Mosquito-borne diseases can pose significant burdens. In many countries, they pose a risk to national economies and the well-being of humans and animals. To mitigate this, mosquito surveillance is crucial to assess the real and potential transmission of mosquito-borne diseases. Between 2020 and 2023, mosquito larvae were collected from both indoor and outdoor breeding sites in urban and rural areas of 4 municipalities of Santiago and Boavista Islands in Cabo Verde. Mosquitoes were identified morphologically and by polymerase chain reaction-based techniques that targeted the mitochondrial cytochrome C oxidase subunit I sequence. During this period, 6,825 breeding sites were assessed, and of 8,094 mosquito specimens screened, 194 specimens of Culex thalassius were identified for the first time in the country in 4 municipalities of Santiago and Boavista Islands. This new finding highlights the importance of including entomological surveillance in health systems. Although this species has only been detected on a few islands, it is important to continuously monitor it to determine its distribution, spread/dispersal, density, and potential involvement in pathogen transmission.

The hard tick, Ixodes keiransi Beati, Nava, Venzal, & Guglielmone, formerly the North American lineage of Ixodes affinis Neumann, is expanding its range northward along the US East Coast. In July 2023, we collected I. keiransi adult female and nymph in a single sampling event, suggesting its range now includes southern New Jersey. In this area, I. keiransi is sympatric with northern populations of Ixodes scapularis Say (Acari: Ixodidae), the primary vector of Lyme disease. Given its status as an enzootic vector of spirochaetes in the Borrelia burgdorferi sensu lato complex, proper differentiation of these 2 species will be critical for accurate estimates of entomological risk. Targeted surveillance should be implemented to monitor further I. keiransi expansion and to elucidate the phenology and enzootic role of this and other understudied Ixodes spp. in the northeastern United States.

The New Jersey Light Trap has been among the earliest trap models used for mosquito surveillance in the United States. This trap was modernized in the 1950s to the miniature CDC light trap, with the addition of CO₂ following soon after. The incandescent light has the tendency to attract nontarget insects, as well as losing a substantial portion of their energy as heat. Few studies have delineated whether heat or light in isolation make a difference in field collections using the former traps within the United States. Our study focused on isolating heat and light variables by using incandescent bulbs, light emitting diode (LED) bulbs, and electric heating patches affixed to a base model CO₂ trap as designed at the Salt Lake City Mosquito Abatement District. Sites were selected in the urban and suburban foothills and canyons of the Wasatch Mountain front, industrial areas near the Salt Lake City International Airport, and rural wetlands in the marshes outlying the Great Salt Lake. Five traps were replicated within each sector during the summer and fall summer seasons. Collections were composed of Aedes dorsalis (Meigen), Culex pipiens L., Culex tarsalis Coquillett, and Culiseta inornata (Williston). Composition changes were a result of seasonal, rather than spatial, shifts. The results showed that LED light traps depressed collections of key species. Otherwise, there were negligible differences in collections among incandescent, heat film, and base model traps. In the Intermountain West, the miniature CDC trap is reliable enough to make programmatic decisions even if light usage varies by district.

Malaria vector surveillance tools often incorporate features of hosts that are attractive to blood-seeking females. The recently developed host decoy trap (HDT) combines visual, thermal, and olfactory stimuli associated with human hosts and has shown great efficacy in terms of collecting malaria vectors. Synthetic odors and yeast-produced carbon dioxide (CO₂) could prove useful by mimicking the human odors currently used in HDTs and provide standardized and easy-to-use olfactory attractants. The objective of this study was to test the attractiveness of various olfactory attractant cues in HDTs to capture malaria vectors. We compared 4 different odor treatments in outdoor field settings in southern Benin and western Burkina Faso: the standard HDT using a human, HDT with yeast-produced CO₂, HDT with an artificial odor blend, and HDT with yeast-produced CO₂ plus artificial odor blend. In both experimental sites, the standard HDT that incorporated a real human produced the greatest catch of Anopheles gambiae s.l (Diptera: Culicidae). The alternatives tested were still effective at collecting target vector species, although the most effective included CO₂, either alone (Benin) or in combination with synthetic odor (Burkina Faso). The trap using synthetic human odor alone caught the fewest An. gambiae s.l. compared to the other baited traps. Both Anopheles coluzzii and Anopheles gambiae were caught by each trap, with a predominance of An. coluzzii. Synthetic baits could, therefore, represent a more standardized and easier-to-deploy approach than using real human odor baits for a robust vector monitoring strategy.

Graphical Abstract

Herein we present the first-known case report of a structural infestation by the bird flea (Dasypsyllus gallinulae perpinnatus) (Dale 1878) in the United States. In March of 2023, the San Mateo County Mosquito and Vector Control District was contacted by a resident in South San Francisco, CA who reported the presence of fleas inside their condominium. The resident had 2 dogs who were on oral flea medication and only 1 inhabitant out of 4 reported receiving flea bites. The front walkway, backyard, and garage were flagged and a small passerine nest was removed from a fire alarm bell in the front walkway. A total of 31 fleas (13 males, 18 females) were collected by flagging from the front entryway underneath the nest. One female flea was collected from the backyard, and 20 (9 males, 11 females) were collected by the resident from inside the house. A total of 387 fleas (163 males, 224 females) were collected from the nest of a small passerine. All life stages (egg, larvae, pupae, and adult) were observed within the nest. Additional parasites in the nest included bird lice and larval western black-legged ticks (Ixodes pacificus, Cooley and Kohls 1943). Research should be conducted into whether bird fleas are capable of transmitting avian pathogens to humans and this information should be communicated to healthcare providers as part of a One Health approach. Additionally, resources to aid in species-level flea identification should be made accessible to pest control operators as this will aid the development of targeted treatments as part of an integrated pest management plan.

Arboviruses can be difficult to detect in the field due to relatively low prevalence in mosquito populations. The discovery that infected mosquitoes can release viruses in both their saliva and excreta gave rise to low-cost methods for the detection of arboviruses during entomological surveillance. We implemented both saliva and excreta-based entomological surveillance during the emergence of Zika virus (ZIKV) in French Guiana in 2016 by trapping mosquitoes around households of symptomatic cases with confirmed ZIKV infection. ZIKV was detected in mosquito excreta and not in mosquito saliva in 1 trap collection out of 85 (1.2%). One female Ae. aegypti L. (Diptera: Culicidae) was found with a ZIKV systemic infection in the corresponding trap. The lag time between symptom onset in a ZIKV-infected individual living near the trap site and ZIKV detection in this mosquito was 1 wk. These results highlight the potential of detection in excreta from trapped mosquitoes as a sensitive and cost-effective method to non invasively detect arbovirus circulation.

Graphical Abstract