C. pipiens blood feeding rhythm exhibits a circadian rhythm that have the ability to persist under absent of environmental cues, consistent with endogenous control. Variations observed in the blood feeding rhythm is accompanied by changes in CYCLE (CYC) expression and olfactory sensitivity, supporting a role for CYC in pathways related to feeding propensity control. In our study, we explore the endogenous circadian control of blood feeding and the effects of mistimed feeding. The blood-feeding rhythm of C. pipiens is governed by an endogenous circadian clock, wherein the variations observed in blood feeding propensity is accompanied by coordinated changes in CYCLE expression and the olfactory sensitivity. We observed a light-induced masking effect that can elevate feeding propensity in subjective daytime in the absence of light, suggesting acute involvement of light in feeding regulation. Under extended starvation induced molecular stress, olfactory sensitivity was reduced along with variations in CYC expression, that is consistent with an olfactory shutdown under extreme metabolic stress. The proposed mechanism placed CYC as a molecular mediator to shut down the olfactory system in mosquitoes in response to extreme metabolic stress. We confirmed through CYCLE knockdown and behavioral assays that the loss of function of CYCLE can alter the feeding propensity in mosquitoes. We also found that mistimed feeding severely compromises mosquito reproductive health and notably evokes compensatory mechanisms. This study provides evidence for endogenous regulation in the blood-feeding behavior of C. pipiens, from host-seeking through olfaction to reproductive fitness. The observed changes in blood feeding rhythm are associate with the light conditions provided, and the underlying rhythm is governed through an internal clock, including core clock component CYCLE and accompanied by the rhythmic changes in small neuropeptide F expression. Through our study, we provide insights into complex genetics involved in chronobiology that control mosquito biting behaviour and reveal a novel target in chronobiology-based vector control strategies aimed at reducing mosquito-human contact.
Sexual dimorphism in Aedes albopictus is typically strict, with blood-feeding restricted to females. Here, we report a rare field-collected intersex Ae. albopictus captured by human-landing catch in Guangzhou, China. Morphological examination revealed a structurally intact piercing-sucking proboscis, intermediate antennal whorled setae, and male-type claspers, although maxillary palp asymmetry and other cephalic traits could not be conclusively assigned to female morphology due to tissue condition. Blood was visible in the abdomen at the time of capture; however, molecular identification of the blood meal was unsuccessful, likely due to limited blood volume and tissue partitioning during DNA extraction. DNA sequencing of the male-determining gene Nix confirmed a genotypic male individual. These observations indicate region-specific disruption of sexual differentiation and suggest that genotypic males may occasionally exhibit female-like blood-feeding under rare circumstances. This study highlights the developmental plasticity of sexually dimorphic traits in mosquitoes and provides insights into the potential limits of rigid sex-specific feeding behavior.
The controlled and continuous production of mosquitoes is critical for conducting basic and operational research. The blood-feeding source is crucial, as it determines the amount of protein mosquitoes can obtain, which is essential for mosquito female fecundity and fertility. The objective of this study was to evaluate the fecundity and fertility of Aedes aegypti females carrying Wolbachia when fed on bovine and lamb blood sources. Females were fed for two consecutive days using bovine or lamb blood, respectively. Thirty fully engorged females per blood source (bovine or lamb) were selected and placed individually for oviposition. The eggs were weighed and measured to provide morphometric data, then counted and hatched. The average number of eggs laid by females fed bovine blood was significantly higher than those laid by females fed lamb blood; however, no significant differences were observed in average hatching percentages. Bovine blood may offer some advantages, potentially associated with a higher level of nutrients for mosquito mass rearing and release strategies due to its positive impact on egg production. However, lamb blood remains a viable alternative. La producción controlada y continua de mosquitos es fundamental para realizar investigación básica y operativa. La fuente de sangre utilizada para la alimentación es clave, ya que determina la cantidad de proteína que pueden adquirir los mosquitos, lo cual es esencial para la fecundidad y fertilidad de las hembras. El objetivo de este estudio fue evaluar la fecundidad y la fertilidad de hembras de Aedes aegypti portadoras de Wolbachia alimentadas con sangre bovina y de cordero. Las hembras fueron alimentadas durante dos días consecutivos con su respectiva fuente de sangre (bovina o de cordero). Se seleccionaron 30 hembras completamente ingurgitadas por tratamiento y se colocaron individualmente para la oviposición. Los huevos se pesaron y se midieron para obtener datos morfométricos; posteriormente se contaron y se evaluó la eclosión. El número promedio de huevos puestos por hembras alimentadas con sangre bovina fue significativamente mayor que el de hembras alimentadas con sangre de cordero; sin embargo, no se observaron diferencias significativas en los porcentajes promedio de eclosión. La sangre bovina puede ofrecer ventajas probablemente asociadas con un mayor nivel de nutrientes para estrategias de cría masiva y liberación, debido a su impacto positivo en la producción de huevos. No obstante, la sangre de cordero sigue siendo una alternativa viable.
Dermacentor andersoni, the Rocky Mountain wood tick, is an important vector for pathogens impacting human and animal health, including bovine anaplasmosis, Colorado tick fever, and Rocky Mountain spotted fever. A better understanding of the biology of this tick is needed for developing disease prevention and vector control strategies. A reference genome was assembled for D. andersoni using high-fidelity (HiFi) long-read PacBio sequences and Hi-C contact mapping, yielding a contiguous assembly in which most contigs matched one of 11 chromosomes. Genome annotation by the NCBI eukaryotic genome annotation pipeline revealed high gene content completeness, yielding a genome completeness score of 94.0% using the Arachnida ortholog dataset. Following genome sequencing, we identified specific genes involved in blood feeding across a range of tissue types and life stages for D. andersoni. To accomplish this, RNA-seq analysis was used to investigate differential gene expression across most organs in adult, nymphal, and larval D. andersoni before and after feeding. Based on this analysis, we identified several gene groups that are involved in blood feeding. Furthermore, we establish sex- and developmental-stage-specific transcriptional profiles. Collectively, this study advances knowledge of D. andersoni biology and enables the development of strategies to limit the spread of diseases transmitted by this tick.
Reference strains of Aedes aegypti, reared over decades under laboratory conditions, are commonly used in research due to their consistency and ease of handling. While their use is relevant in terms of reproducibility between labs, complementary work on field-collected mosquitoes and their progeny is essential to capture biological and behavioral variations of natural populations. However, experimental set-ups optimized on reference strains are not always successful with field-derived mosquitoes; their lack of attraction to experimental blood meals is a recurrent issue. In this study, we evaluated methods to improve blood-feeding rate of field-derived mosquitoes from Cayenne (French Guiana), using the New Orleans reference strain as a control. We tested the impact of several blood-feeding systems for mosquitoes kept in a cage or in individual vials and adjusted starvation parameters. Individual mosquitoes offered a blood meal in lids of microtubes had the highest blood-feeding rate. For field-derived mosquitoes, starvation, or alternatively, provision of a 3% salt solution instead of the sugar solution, also consistently improved blood-feeding rate, with a minimal impact on survival. Our results may be helpful for establishment of new colonies, including in resource-limited settings, studies on fitness of field-derived mosquitoes and on experiments requiring individual level monitoring.
Spatial emanators disrupt mosquito behaviour by inducing movement away from chemical stimuli and interfering with host detection and feeding. These tools were recently endorsed by the World Health Organization (WHO) for malaria control, based largely on clinical evidence from East Africa. Mosquito Shield™ is a passive, transfluthrin-based emanator designed to provide month-long protection in enclosed or semi-enclosed spaces. This study evaluated its entomological efficacy under experimental hut conditions in Benin, West Africa to generate evidence in support of WHO prequalification. An experimental hut trial was conducted against wild free-flying pyrethroid-resistant Anopheles gambiae s.l. at the Covè field station in southern Benin over two 32-day product life cycles of Mosquito Shield™. Sixteen West African–style experimental huts were assigned to Mosquito Shield™ or a placebo control. Efficacy was measured using human landing catches (HLC) and mosquito aspirations following standard hut testing methods. Primary endpoints included reductions in the numbers of mosquitoes caught through HLC (protective efficacy) and in the numbers of blood-fed mosquitoes caught through aspiration (personal protection). Secondary endpoints included deterrence, exophily, mortality, and blood-feeding inhibition. WHO susceptibility bioassays confirmed high levels of resistance to pyrethroids, including transfluthrin, in the local Anopheles gambiae s.l. population during the trial. A total of 5,682 An. gambiae s.l. and 6,158 Mansonia africana were collected through HLCs, and 1,436 An. gambiae s.l. by mosquito aspirations. Mosquito Shield™ significantly reduced mosquito landing, providing 43.0% protective efficacy (95% CI: 24.0–57.0; p < 0.001) against An. gambiae s.l. and 38.0% protective efficacy (95% CI: 12.0–57.0; p = 0.008) against Mansonia africana. Mosquito aspiration data showed 48.5% deterrence, 29.9% blood-feeding inhibition, comparable exiting rates between treatment and control huts (47.2% vs. 46.7%; p = 0.962) and 64% personal protection (95% CI: 21.9–81.8; p < 0.001) against An. gambiae s.l. No mosquito mortality was recorded in the control huts. In contrast, Mosquito Shield™ induced mortality of Anopheles gambiae s.l. reaching 49.0% in HLC collections and 22.7% in aspiration collections. Mosquito Shield™ also induced > 96% mortality in Mansonia africana, demonstrating both lethal and behavioural effects against both vector species. Overall, Mosquito Shield™ significantly reduced mosquito entry, landing, blood-feeding, and survival of pyrethroid-resistant An. gambiae s.l. under semi-field experimental hut conditions in West Africa, with additional effects against Mansonia africana. These results support its WHO prequalification and highlight its potential as a complementary vector control tool to strengthen malaria prevention and provide additional benefits for integrated control of other vector-borne diseases.
Although exposure of Anopheles mosquitoes to insecticide-treated vector control tools is not always lethal, the sublethal effects of these insecticides on their life-history traits remain poorly explored. Therefore, this study examined the impact of sublethal exposure to broflanilide, chlorfenapyr and deltamethrin on longevity; blood meal acceptance; viable egg production; and the proportion of females with viable eggs in insecticide-resistant An. gambiae s.l. populations from Tiassalé, Côte d'Ivoire. The sublethal effects of broflanilide, chlorfenapyr and deltamethrin were assessed on 3-5-day-old adult female An. gambie s.l. from the field collected Tiassalé strain (F0), using the insecticide-susceptible Kisumu strain as reference, following standardized World Health Organization (WHO) bottle bioassay procedures. For each insecticide, the lethal dose causing 20% mortality (LD20) was determined through dose-response analysis and then subsequently applied in treated bottles. Post-exposure, longevity, blood-feeding success, egg viability and the proportion of ovipositing females were recorded. Comparisons were made between the control and LD20 treatment groups. Sublethal exposures produced distinct effects depending on both the mosquito strain and the insecticide type. Broflanilide significantly reduced longevity in the Tiassalé population (P < 0.0001), whereas chlorfenapyr and deltamethrin had stronger effects on the Kisumu strain. Deltamethrin exposure notably impaired blood-feeding behaviour, reducing feeding success by 25.5% in Kisumu and 19.5% in Tiassalé. Reproductive parameters were also affected: both chlorfenapyr and broflanilide reduced viable egg production and the proportion of females producing viable eggs in both strains. In contrast, deltamethrin showed a more limited impact on reproduction. In the Tiassalé strain, the proportion of females producing viable eggs dropped to 41.5% following chlorfenapyr exposure, compared with 65.7% in unexposed individuals. These findings demonstrate that sublethal doses of insecticides can adversely affect reproductive success and blood-feeding behaviour in both susceptible and resistant Anopheles mosquitoes, potentially reducing their vectorial capacity. This study underscores the need to systematically account for sublethal effects when evaluating insecticides in vector control programs, particularly in settings where resistance is widespread among malaria vectors.
Granulocytic anaplasmosis is a zoonotic disease that affects various domestic mammals (dogs, horses, and, more rarely, cats). In ruminants, it is better known as tick-borne fever (TBF) and is responsible for significant economic losses on European livestock farms, mainly due to a drop in milk production, abortions, and immunosuppression, which can lead to secondary infections. The disease is caused by the strictly intracellular bacterium Anaplasma phagocytophilum, whose biological vectors are ticks of the genus Ixodes. Other blood-feeding arthropods may be involved in transmitting this bacterium, notably Stomoxys calcitrans, a major ectoparasite of livestock that is implicated in transmitting other pathogens, including bacteria of the genus Anaplasma. This study aimed to evaluate the potential of S. calcitrans to act as a mechanical vector of A. phagocytophilum under laboratory conditions. Two experimental models were employed: one mimicking immediate transmission, and the other delayed transmission. In both models, A. phagocytophilum DNA and RNA were detected in S. calcitrans for the first time, but no traces of the bacterium's DNA or RNA were found in the glass feeder's blood. Further research is needed to confirm these findings through field studies investigating the presence of the bacterium in flies under natural conditions. This study also describes two original infection models of stable flies designed to reproduce their ex vivo blood-feeding, promoting alternative experimental approaches in accordance with animal welfare regulations and 4R principles. Stomoxys calcitrans comme vecteur mécanique potentiel d'Anaplasma phagocytophilum : évaluation à l'aide de modèles d'alimentation ex vivo originaux. L’anaplasmose granulocytaire est une maladie zoonotique pouvant toucher divers mammifères domestiques (chiens, chevaux et plus rarement le chat). Chez les ruminants, cette maladie plus connue sous le nom de Tick-Borne Fever (TBF), est responsable d’importantes pertes économiques dans les élevages européens en raison notamment de chutes de production laitière, d’avortements et d’immunodépressions pouvant conduire à des infections secondaires. Cette maladie est causée par Anaplasma phagocytophilum, bactérie intracellulaire stricte dont les vecteurs biologiques sont les tiques du genre Ixodes. D’autres arthropodes hématophages pourraient être impliqués dans la transmission de cette bactérie, notamment Stomoxys calcitrans, ectoparasite important des bovins incriminé dans la transmission d’agents pathogènes, dont des bactéries du genre Anaplasma. L’objectif de cette étude a été d’évaluer le rôle de S. calcitrans en tant que vecteur mécanique potentiel d’A. phagocytophilum en conditions de laboratoire. Deux modèles expérimentaux ont été utilisés, l’un mimant une transmission immédiate et l’autre mimant une transmission retardée. Dans ces deux modèles, la présence d’ADN et d’ARN d’A. phagocytophilum a été détectée pour la première fois dans S. calcitrans, cependant aucune trace d’ADN ou d’ARN de la bactérie n’a été détectée dans le sang du receveur. Il serait intéressant de confirmer ces données par des études de terrain en recherchant la présence de la bactérie dans les mouches en conditions naturelles. Cette étude présente également deux modèles originaux d’infection des stomoxes permettant de reproduire leur prise de repas sanguins ex vivo, et promeut ainsi des approches expérimentales alternatives respectueuses de l’éthique animale, conformément à la règle des 4R.
The genus Bartonella comprises facultative intraerythrocytic bacteria capable of causing long-lasting bacteremia in their natural hosts, with zoonotic potential across multiple species. Rodents serve as important reservoirs for a broad diversity of Bartonella spp., with blood-feeding arthropods, particularly fleas, mediating transmission. Despite their frequent association with humans, the role of edible dormice (Glis glis) and their fleas (Ceratophyllus sciurorum) in Bartonella ecology remains poorly understood. We combined long-term ecological and epidemiological data with gut and body microbiome analyses of C. sciurorum to investigate the prevalence, diversity, host specificity, and transmission of Bartonella across flea life stages. The study was conducted over 6 years in a natural dormouse population. Bartonella detection and characterization were performed using multilocus PCR targeting gltA, rpoB, ftsZ, and ITS loci, bacterial cultivation on selective media, and long-read nanopore sequencing. Flea gut microbiomes were assessed in pooled and individual samples to determine the impact of Bartonella infection on microbial community structure. Transmission across flea life stages was evaluated by analyzing larvae, newly emerged adult fleas, and adults directly collected from dormice. We observed a consistently high prevalence of Bartonella in both dormice and their fleas. Four species were identified: B. gliris and B. grahamii subsp. shimonis dominated, each represented by multiple genotypes, whereas B. washoensis and B. bilalgolemii were each detected in a single flea. Moreover, mixed infections of B. gliris and B. grahamii subsp. shimonis were frequent in both dormice and their fleas. Detection of Bartonella DNA in flea larvae and newly emerged adults indicates possible transstadial perpetuation. Flea gut microbiomes were highly variable but consistently dominated by Bartonella in infected fleas. In contrast, uninfected fleas exhibited more diverse communities, often enriched with Staphylococcus and other environmental or host-associated taxa. We, therefore, suggest that flea populations can maintain Bartonella over extended periods, even in the absence of continuous contact with vertebrate hosts. The dormouse-flea-Bartonella system represents a valuable natural model for studying flea-borne zoonoses. The high prevalence and persistent bacteremia in dormice of different ages indicate their role as suitable reservoirs for at least two Bartonella species. Flea populations are capable of sustaining Bartonella over long periods, and detections in immature life stages suggest continuity of infection across flea generations. Therefore, C. sciurorum appears to play an important role in maintaining B. grahamii subsp. shimonis and B. gliris in nature. Infection is consistently associated with pronounced restructuring of the flea gut microbiome, highlighting the ecological and microbial dimensions of vector-borne pathogen transmission. These findings underscore the importance of considering vector-associated microbial communities when evaluating disease dynamics and zoonotic risk in natural host-vector systems.
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, Ethiopia. Flight monitoring assays revealed that An. stephensi exhibited pronounced activity at dusk, beginning between 18:00 and 19:00 with scotophase onset, and little to no activity during photophase. Blood-feeding propensity, defined as the proportion of mosquitoes taking a blood meal when offered, peaked during 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, 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.
Culicoides biting midges (Diptera: Ceratopogonidae) remain economically important dipteran pests of livestock and other animals worldwide. They are known for their blood-feeding nuisance behaviours, but also serve as biological vectors of several animal pathogens, including bluetongue virus, epizootic hemorrhagic disease virus and vesicular stomatitis virus. Efforts to limit biting midge access to target hosts such as cattle and sheep have relied predominantly on insecticide use. However, research regarding biting midge insecticide efficacy largely has been focused on animal and barrier treatments using formulated commercial products, with little effort to identify local endemic species level susceptibility to specific active ingredients. This study was conducted to determine the susceptibility of biting midges under laboratory and field conditions, as well as to determine the potential for insecticide resistance development in these pests. Under laboratory conditions, the susceptibility of a laboratory-reared biting midge colony (Culicoides sonorensis Wirth and Jones (Diptera: Ceratopogonidae)) was significantly less than wild Culicoides spp. Similarly, wild Culicoides spp. also were significantly more susceptible than wild mosquitoes when exposed to an ultra-low volume permethrin application under field conditions. Additionally, a long-established C. sonorensis colony subjected to permethrin selection under laboratory conditions suggested that insecticide resistance development in this pest could occur in field populations if pressured. This is the first account of the potential for insecticide resistance development in Culicoides. The implications of Culicoides spp. insecticide resistance and the lack thereof in assessed field populations are discussed.
Blood-feeding stable flies, Stomoxys calcitrans (L.), are reportedly attracted to conspecific feces. We investigated whether stable flies are attracted to fly feces-derived bacteria, and whether attraction varies with bacterial species and growth substrate, and the sex and reproductive status of foraging flies. We isolated seven bacterial species from fecal deposits of stable flies. In laboratory two-choice bioassays with paired adhesive-coated traps baited with sterile agar or bacteria-inoculated agar, the seven bacterial species in combination repelled flies. However, the bacterial congeners Serratia marcescens and Serratia surfactantfaciens attracted flies when tested singly. Analyses of the headspace volatiles of S. marcescens and S. surfactantfaciens by gas chromatography-mass spectrometry revealed near-identical odor profiles, which may explain their comparable attractiveness to flies. Flies were more attracted to S. marcescens grown on trypticase soy agar than to S. marcescens grown on bovine blood agar, likely due to substrate-dependent odor profiles. Feces-dwelling bacteria appear to function in the context of foraging rather than oviposition given that S. marcescens attracted both female and male stable flies, and both gravid and non-gravid females. That agar plates inoculated with all seven bacterial species failed to attract foraging flies could mean that the microbial community on agar did not sufficiently resemble that in fly feces, perhaps due to the limitations of culture-dependent isolation methods. Alternatively, additional feces-derived odors may also be necessary for fly attraction. Further investigation is warranted to identify the semiochemicals, and possibly other cues, produced by S. marcescens that make this bacterium distinctively attractive to stable flies.
Mosquitoes use their sense of smell in a variety of behaviors, such as searching for nectar, host-seeking, and finding oviposition sites. These behaviors, many of which are sexually dimorphic, require modulation in response to the internal state and the external environment of the animal. For example, a female mosquito is attracted to human odors, but after blood-feeding, its attraction to human hosts reduces and the attraction to oviposition sites eventually increases. Such changes in the internal state and their effects on the sensory systems are implemented in the insect brain by an intricate choreography involving many neuromodulators in a sex-specific manner. With a focus on the mosquito olfactory system, we review the expression profiles and the functional roles of monoamine neuromodulators (dopamine, serotonin, octopamine and tyramine) and some of the major neuropeptides, including FMRFamide, SIFamide, short neuropeptide F, allatostatin A, myoinhibitory peptide, allatotropin, and tachykinin-related peptide.
European deer keds (Diptera: Hippoboscidae: Lipoptena cervi (Linnaeus, 1758)) are blood-feeding ectoparasitic flies that usually feed on deer and other cervids but sometimes bite humans. Introduced into North America in the late 1800s, they have since become widespread in the Northeast. European deer keds were historically reported at the regional or state level, but recent work has begun to clarify their range by compiling county-level occurrence data. We expand upon this trend by using multiple methods to determine where deer keds are relatively more and less abundant in Pennsylvania. Based on our results, European deer keds are most abundant in northern Pennsylvania, present but less abundant in central and western Pennsylvania, and absent or nearly so from southeastern counties. Understanding where deer keds occur will inform public education and allow the public to better protect themselves from deer ked bites.
The gut microbiota has co-evolved with its host and plays a vital role in maintaining physiological homeostasis and health. Understanding the composition of microbial communities in leech guts may reveal important insights into their ecological adaptations and feeding strategies. This study aimed to compare the gut microbiota of hematophagous (blood-feeding) and non-hematophagous leeches, to identify microbial signatures associated with dietary divergence and niche specialization. Gut contents were collected from representative species of hematophagous and non-hematophagous leeches. The microbial community composition was analyzed via 16 S rRNA gene sequencing. A total of 751 microbial species were identified, encompassing 535 genera, 332 families, 203 orders, 86 classes, and 39 phyla. At the phylum level, Proteobacteria and Firmicutes were significantly more abundant in hematophagous leeches than in non-hematophagous leeches, whereas Bacteroidetes predominated in non-hematophagous counterparts. At the genus level, hematophagous leeches exhibited higher abundances of Elstera, unclassified genus of Rhodospirillaceae, Aeromonas, unclassified genus of Rhodospirillales, unclassified genus of Peptostreptococcales-Tissierellales, and unclassified genus of Oscillospirales. In contrast, unclassified genus of Comamonadaceae, Nubsella, Cetobacterium, Mucispirillum, unclassified genus of Peptostreptococcaceae, and Bacteroides were enriched in non-hematophagous leeches. Functional prediction analysis revealed significant differences in five key metabolic pathways between the two groups: lipid transport and metabolism, amino acid transport and metabolism, nucleotide transport and metabolism, translation and ribosomal structure biogenesis, and coenzyme transport and metabolism. Distinct taxonomic and functional profiles characterize the gut microbiota of hematophagous and non-hematophagous leeches. These results provide microbiological evidence for dietary specialization in leeches and offer a basis for guiding domestication and artificial breeding strategies.
Sialic acid commonly decorates the surface of cells and secreted proteins of Eukaryotes. It plays roles in cell signalling and adhesion, affects immune reactions and alternative complement pathways, and neurogenesis. While sialic acid is a saccharide present in high abundance in vertebrates, its minor representation was reported in arthropods and among medically necessary parasites. Ixodes ricinus tick is a significant vector of pathogens causing severe infections in humans and livestock. It has been known mainly through indirect evidence that sialylated glycoproteins are present in tick tissues. As a blood-feeding parasite, the host's blood, full of sialylated molecules, could be the source of such glycoproteins within the tick's body. To detect solely the tick sialylated glycoproteins, we used an azide-modified precursor of sialic acid to be metabolically incorporated by the tick cell line IRE/CTVM20 originating in this tick or by ticks if the corresponding biosynthetic pathway is present and active. Our I. ricinus genome screen revealed two sialyltransferase genes, which were expressed in all ticks' life stages, and we confirmed sialyltransferase activity in tick cells. We combined in vitro feeding along with Click chemistry to track the presence of sialylated glycoproteins from fed female ticks through their eggs to the larvae. Similarly, sialylated glycoproteins were produced by the IRE/CTVM20 cell line. Thus, we confirmed the ability of ticks to produce their own sialylated glycoproteins in addition to those originating from the host blood.
Climate change is driving the rapid range expansion of Aedes aegypti into temperate regions, presenting novel seasonal cues that can affect seasonal plasticity. Seasonal plasticity in Ae. aegypti is well studied in eggs but remains understudied in adults. Here, we investigated the effects of photoperiod and temperature on thermal plasticity, measured by the ability to cold acclimate, and other dormancy-related traits in female Ae. aegypti adults. Cold acclimation was done under a short-day (SD) and long-day (LD) photoperiod. LD photoperiod was introduced at three life stages: parental generation, egg state and adult acclimation, to determine if timing of LD photoperiod introduction affected thermal plasticity. LD photoperiod had a significant influence on both cold tolerance metrics assessed, chill coma onset temperature and survival after a cold stress. Introduction of LD photoperiod reduced thermal plasticity, with the magnitude of change in plasticity depending on when LD was introduced. At warm temperatures, LD photoperiod modified the rhythmic profile of one of the circadian clock genes analysed in this study, timeless, but not period. In contrast, cold acclimation abolished the cyclical expression of both clock genes. Cold acclimation under both photoperiods supressed blood-feeding behaviour, which resumed upon warming, but cold acclimation did not change relative transcript abundance of dormancy-implicated genes. This study presents the first evidence of an adult photosensitive seasonal phenotype in Ae. aegypti, with improved cold tolerance and reproductive quiescence. These findings contribute to a deeper understanding of how environmental cues may facilitate the continued expansion of this important disease vector into temperate environments.
Fleas are holometabolous, blood-feeding ectoparasites capable of transmitting diverse pathogens of significant veterinary and public health concerns. Their occurrence and abundance within a given habitat depend on environmental factors and the availability of suitable hosts. In Pakistan, research on flea fauna and their associated pathogens has been neglected. To date, no reports have documented the molecular characterization of Ctenocephalides felis felis and their associated bacteria in Pakistan. In the present study, three hundred and eighty morphologically identified C. felis felis specimens-comprising 184 collected from free roaming (stray) dogs (n = 69) and 196 from cats (n = 86), were subjected to DNA extraction followed by amplification of the cytochrome oxidase subunit 1 gene (cox1) and citrate synthase gene (gltA) for flea identity confirmation and screening for the presence of associated bacteria, respectively. Amplicons of appropriate base pair sizes were sequenced and submitted to BLASTn and subsequently subjected to phylogenetic analyses. The obtained cox1 sequence from the morphologically identified C. felis felis in this study showed 100% identity and phylogenetically clustered with C. felis felis sequences from India, Australia, Thailand, China, and Laos in GenBank. Similarly, gltA sequences showed 100% identity with the Wolbachia endosymbiont of C. felis reported from the United Kingdom. This study provides the first genetic characterization of C. felis felis infesting dogs and cats, and their associated Wolbachia endosymbiont in Khyber Pakhtunkhwa (KP), Pakistan. These findings provide baseline molecular data and highlight the need for systematic surveillance and management measures to mitigate any potential veterinary and public health threats.
In various dipteran taxa, selection of oviposition sites by gravid female flies is informed by the presence of adult conspecific flies, their eggs, or their larval offspring. Working with blood-feeding stable flies, Stomoxys calcitrans (Diptera: Muscidae), we tested three hypotheses: (1) stable flies preferentially oviposit on sites offering conspecific eggs with egg-derived airborne semiochemicals (message-bearing chemicals), (2) egg-borne bacteria elicit oviposition, and (3) egg- or egg bacteria-derived semiochemicals attract flies. Two-choice laboratory experiments, testing for fly attraction and oviposition, revealed that female flies were attracted to, and preferentially oviposited on, sites with deposits of conspecific eggs. Semiochemicals emitted from bacteria isolated from stable fly eggs, and grown on agar, attracted flies and prompted oviposition. Of the eight bacterial isolates, Serratia marcescens and Sporosarcina sp. were most attractive to gravid female flies. Gas chromatographic-mass spectrometric analysis of S. marcescens headspace volatile extracts identified 6 volatiles, and gas meters recorded elevated levels of carbon dioxide and ammonia in the headspace of S. marcescens. These two gases in binary combination, liberated from aqueous sodium bicarbonate and ammonium hydroxide, attracted flies and prompted oviposition. The attraction and oviposition-inducing effects of these gases were not enhanced by the 6-volatile blend. Our data suggest that gravid stable flies engage in conspecific cueing to locate and select suitable sites for oviposition and development of larval offspring. Unraveling all semiochemicals that mediate stable fly attraction and oviposition site selection may improve stable fly control tactics.
Spotted lanternflies and mosquitoes are among the most concerning invasive insects, which have introduced a myriad of public health, social, economic, and agricultural problems. In addition to their characteristic blood-feeding behavior, mosquitoes exhibit frequent sugar-feeding activity, targeting a diverse range of sugar sources. Spotted lanternflies are ravenous sap-feeders and, as a result, produce large aggregates of honeydew, a sugary excreta. Here, we used spotted lanternfly honeydew-specific DNA barcoding to determine that both invasive and native mosquito species feed on spotted lanternfly honeydew in the field. The implications and applications of these findings for invasive interspecies interactions and disease vector management are discussed.