Blood Meal Analysis Research Articles

Host associations of biting midges (Diptera: Ceratopogonidae: Culicoides) at deer farms in Florida, USA.

Documenting the host use of vector species is important for understanding the transmission dynamics of vector-borne pathogens. Biting midges (Diptera: Ceratopogonidae: Culicoides) are vectors of epizootic hemorrhagic disease virus (EHDV) and bluetongue virus (BTV) worldwide. However, relative to mosquitoes and many other vector groups, host associations of this group are poorly documented. In this study, we used PCR-based bloodmeal analysis to determine species-level host associations of 3,603 blood-engorged specimens belonging to 18 Culicoides species at 8 deer farms in Florida, USA. We used a binomial mixed model with a Bayesian framework to compare the effect of host composition on the feeding patterns of Culicoides spp. and employed the Morisita-Horn Index to investigate the similarity of host use between farms for Culicoides stellifer and Culicoides insignis. Results show that the estimated probability of Culicoides spp. feeding upon white-tailed deer depends on the availability of cattle or exotic game and demonstrates differences in host-feeding selection among species. Culicoides insignis had high host similarity across farms suggesting that its host-use patterns are somewhat conserved. Culicoides stellifer had lower host similarity across farms suggesting that it is a more opportunistic feeder. White-tailed deer are fed upon by many Culicoides species on deer farms in Florida, and while most Culicoides species feed on white-tailed deer, the ratio of white-tailed deer bloodmeals to other bloodmeals is likely influenced by host availability. Culicoides spp. taking a majority of their bloodmeals from farmed white-tailed deer should be assessed for their vector competence for EHDV and BTV.

The first detection of avian haemosporidia from Culicoides biting midges in Japan, with notes on potential vector species and the transmission cycle.

Biting midges (Ceratopogonidae) are capable of transmitting a variety of pathogens including viruses, trypanosomes and haemosporidia. The majority of Haemoproteus parasites are transmitted by biting midges predominantly of the genus Culicoides and are known to cause significant physical and reproductive impacts on both wild and domestic birds. In Japan, Haemoproteus had been detected from various avian hosts, but not from arthropod vectors. In this study, we investigated the prevalence of avian haemosporidia at an educational forest in central Japan in attempt to reveal possible vector species of Haemoproteus, which would help to better understand the transmission cycle of Haemoproteus within Japan and to develop preventative measures for captive and domestic birds. Biting midges were caught using UV light traps from 2016 to 2018. The collected samples were morphologically identified, and haemosporidian parasites were detected using PCR-based methods. The detected lineages were phylogenetically analyzed and compared with lineages previously detected from birds. Bloodmeal analyses were also carried out for part of the blood-fed individuals. Six Haemoproteus lineages were detected from 17 of 1042 female Culicoides (1.63%), including three species (C. sigaensis, C. arakawae, and C. pictimargo) in which Haemoproteus was detected for the first time. All detected lineages were placed in the subgenus Parahaemoproteus clade and were previously detected from crows of central Japan, strongly suggesting that parasites of these genetic lineages are transmitted between Culicoides and crows. Two Plasmodium lineages were also detected but are thought to be transmitted between Culex mosquitoes and birds of the educational forest based on previous detections. No amplifications were seen in bloodmeal analysis, possibly due to insufficient amount of blood, denaturation via digestion, or insufficient detectability of the used protocol. Haemoproteus DNA was detected from Culicoides for the first time in Japan, suggesting that transmission is possible within the country. These findings highlight the necessity to investigate Culicoides populations and Haemoproteus infections dynamics in Japan. However, vector competence could not be confirmed in this study and further studies are anticipated.

Differences in Longevity and Temperature-Driven Extrinsic Incubation Period Correlate with Varying Dengue Risk in the Arizona-Sonora Desert Region.

Dengue transmission is determined by a complex set of interactions between the environment, Aedes aegypti mosquitoes, dengue viruses, and humans. Emergence in new geographic areas can be unpredictable, with some regions having established mosquito populations for decades without locally acquired transmission. Key factors such as mosquito longevity, temperature-driven extrinsic incubation period (EIP), and vector-human contact can strongly influence the potential for disease transmission. To assess how these factors interact at the edge of the geographical range of dengue virus transmission, we conducted mosquito sampling in multiple urban areas located throughout the Arizona-Sonora desert region during the summer rainy seasons from 2013 to 2015. Mosquito population age structure, reflecting mosquito survivorship, was measured using a combination of parity analysis and relative gene expression of an age-related gene, SCP-1. Bloodmeal analysis was conducted on field collected blood-fed mosquitoes. Site-specific temperature was used to estimate the EIP, and this predicted EIP combined with mosquito age were combined to estimate the abundance of "potential" vectors (i.e., mosquitoes old enough to survive the EIP). Comparisons were made across cities by month and year. The dengue endemic cities Hermosillo and Ciudad Obregon, both in the state of Sonora, Mexico, had higher abundance of potential vectors than non-endemic Nogales, Sonora, Mexico. Interestingly, Tucson, Arizona consistently had a higher estimated abundance of potential vectors than dengue endemic regions of Sonora, Mexico. There were no observed city-level differences in species composition of blood meals. Combined, these data offer insights into the critical factors required for dengue transmission at the ecological edge of the mosquito's range. However, further research is needed to integrate an understanding of how social and additional environmental factors constrain and enhance dengue transmission in emerging regions.

Collection of triatomines from sylvatic habitats by a Trypanosoma cruzi-infected scent detection dog in Texas, USA.

Triatomine insects, vectors of the etiologic agent of Chagas disease (Trypanosoma cruzi), are challenging to locate in sylvatic habitats. Collection techniques used in the United States often rely on methods to intercept seasonally dispersing adults or on community scientists' encounters. Neither method is suited for detecting nest habitats likely to harbor triatomines, which is important for vector surveillance and control. Furthermore, manual inspection of suspected harborages is difficult and unlikely to reveal novel locations and host associations. Similar to a team that used a trained dog to detect sylvatic triatomines in Paraguay, we worked with a trained scent detection dog to detect triatomines in sylvatic locations across Texas. Ziza, a 3-year-old German Shorthaired Pointer previously naturally infected with T. cruzi, was trained to detect triatomines. Over the course of 6 weeks in the fall of 2017, the dog and her handler searched at 17 sites across Texas. The dog detected 60 triatomines at 6 sites; an additional 50 triatomines were contemporaneously collected at 1 of these sites and 2 additional sites without the assistance of the dog. Approximately 0.98 triatomines per hour were found when only humans were conducting searches; when working with the dog, approximately 1.71 triatomines per hour were found. In total, 3 adults and 107 nymphs of four species (Triatoma gerstaeckeri, Triatoma protracta, Triatoma sanguisuga, and Triatoma indictiva) were collected. PCR testing of a subset revealed T. cruzi infection, including DTUs TcI and TcIV, in 27% of nymphs (n = 103) and 66% of adults (n = 3). Bloodmeal analysis of a subset of triatomines (n = 5) revealed feeding on Virginia opossum (Didelphis virginiana), Southern plains woodrat (Neotoma micropus), and eastern cottontail (Sylvilagus floridanus). A trained scent detection dog enhanced triatomine detections in sylvatic habitats. This approach is effective at detecting nidicolous triatomines. Control of sylvatic sources of triatomines is challenging, but this new knowledge of specific sylvatic habitats and key hosts may reveal opportunities for novel vector control methods to block the transmission of T. cruzi to humans and domestic animals.

Blood meal survey reveals insights into mosquito-borne diseases on the island of Santiago, Cape Verde

IntroductionThe transmission of pathogens by blood-sucking insects to humans and other animals depends on vector-host interactions. As yet unexplored in Cape Verde, mosquito feeding behavior plays a crucial role in pathogen transmission. Herein, we aim to explore, through blood meal analysis, the relationship between mosquito species and common hosts in Santiago Island, Cape Verde.MethodsEngorged female mosquitoes were collected through mechanical aspiration from May 2016 to December 2017 in three municipalities of Santiago Island (Praia, Santa Cruz, and Santa Catarina). Blood-feeding behavior in each municipality was assessed through blood meal analysis using an enzyme-linked immunosorbent assay (ELISA).ResultsWe were able to determine that single-host blood meals were common in Aedes aegypti, Anopheles arabiensis, and Culex pipiens sensu lato (s.l.). In general, the mosquitoes preferred to feed on humans, dogs, and chickens, and on multiple hosts, mainly two hosts. The human blood index (HBI) was highest (i.e., 1.00) in Ae. aegypti, with the lowest value (0.40) observed in An. arabiensis. It was observed that, among single-host blood meals, the likelihood of Cx. pipiens s.l. feeding on humans was significantly high, whereas the likelihood of An. arabiensis feeding on humans was significantly low (log-odds ratio (LOR) = 0.85 and –2.44, respectively). In addition, a high likelihood of Ae. aegypti feeding on humans was observed, but this was not statistically significant (LOR = 0.85).DiscussionOverall, our findings demonstrate a lack of feeding preference in Culex pipiens s.l. compared with Ae. aegypti and An. arabiensis. These results provide insights into possible parasite transmission and pathogen spillover/spillback, which threaten human/animal health and the economy in Cape Verde.

Nanopore adaptive sampling for targeted mitochondrial genome sequencing and bloodmeal identification in hematophagous insects

BackgroundBlood-feeding insects are important vectors for an array of zoonotic pathogens. While previous efforts toward generating molecular resources have largely focused on major vectors of global medical and veterinary importance, molecular data across a large number of hematophagous insect taxa remain limited. Advancements in long-read sequencing technologies and associated bioinformatic pipelines provide new opportunities for targeted sequencing of insect mitochondrial (mt) genomes. For engorged hematophagous insects, such technologies can be leveraged for both insect mitogenome genome assembly and identification of vertebrate blood-meal sources.MethodsWe used nanopore adaptive sampling (NAS) to sequence genomic DNA from four species of field-collected, blood-engorged mosquitoes (Aedes and Culex spp.) and one deer fly (Chrysops sp.). NAS was used for bioinformatical enrichment of mtDNA reads of hematophagous insects and potential vertebrate blood-meal hosts using publically available mt genomes as references. We also performed an experimental control to compare results of traditional non-NAS nanopore sequencing to the mt genome enrichment by the NAS method.ResultsComplete mitogenomes were assembled and annotated for all five species sequenced with NAS: Aedes trivittatus, Aedes vexans, Culex restuans, Culex territans and the deer fly, Chrysops niger. In comparison to data generated during our non-NAS control experiment, NAS yielded a substantially higher proportion of reference-mapped mtDNA reads, greatly streamlining downstream mitogenome assembly and annotation. The NAS-assembled mitogenomes ranged in length from 15,582 to 16,045 bp, contained between 78.1% and 79.0% A + T content and shared the anticipated arrangement of 13 protein-coding genes, two ribosomal RNAs, and 22 transfer RNAs. Maximum likelihood phylogenies were generated to further characterize each insect species. Additionally, vertebrate blood-meal analysis was successful in three samples sequenced, with mtDNA-based phylogenetic analyses revealing that blood-meal sources for Chrysops niger, Culex restuans and Aedes trivittatus were human, house sparrow (Passer domesticus) and eastern cottontail rabbit (Sylvilagus floridanus), respectively.ConclusionsOur findings show that NAS has dual utility to simultaneously molecularly identify hematophagous insects and their blood-meal hosts. Moreover, our data indicate NAS can facilitate a wide array of mitogenomic systematic studies through novel ‘phylogenetic capture’ methods. We conclude that the NAS approach has great potential for broadly improving genomic resources used to identify blood-feeding insects, answer phylogenetic questions and elucidate complex pathways for the transmission of vector-borne pathogens.Graphical

A meta-transcriptomic study of mosquito virome and blood feeding patterns at the human-animal-environment interface in Guangdong Province, China

Mosquitoes are a formidable reservoir of viruses and important vectors of zoonotic pathogens. Blood-fed mosquitoes have been utilized to determine host infection status, overcoming the difficulties associated with sampling from human and animal populations. Comprehensive surveillance of potential pathogens at the interface of humans, animals, and the environment is currently an accredited method to provide an early warning of emerging or re-emerging infectious diseases and to proactively respond to them. Herein we performed comprehensive sampling of mosquitoes from seven habitats (residential areas, hospital, airplane, harbor, zoo, domestic sheds, and forest park) across five cities in Guangdong Province, China. Our aim was to characterize the viral communities and blood feeding patterns at the human-animal-environment interface and analyze the potential risk of cross-species transmission using meta-transcriptomic sequencing. 1898 female adult mosquitoes were collected, including 1062 Aedes and 836 Culex mosquitoes, of which approximately 12% (n = 226) were satiated with blood. Consequently, 101 putative viruses were identified, which included DNA and RNA viruses, and positive-stranded RNA viruses (+ssRNA) were the most abundant. According to viral diversity analysis, the composition of the viral structure was highly dependent on host species, and Culex mosquitoes showed richer viral diversity than Aedes mosquitoes. Although the virome of mosquitoes from different sampling habitats showed an overlap of 39.6%, multiple viruses were specific to certain habitats, particularly at the human-animal interface. Blood meal analysis found four mammals and one bird bloodmeal source, including humans, dogs, cats, poultry, and rats. Further, the blood feeding patterns of mosquitoes were found to be habitat dependent, and mosquitoes at the human-animal interface and from forests had a wider choice of hosts, including humans, domesticated animals, and wildlife, which in turn considerably increases the risk of spillover of potential zoonotic pathogens. To summarize, we are the first to investigate the virome of mosquitoes from multiple interfaces based on the One Health concept. The characteristics of viral community and blood feeding patterns of mosquitoes at the human-animal-environment interface were determined. Our findings should support surveillance activities to identify known and potential pathogens that are pathogenic to vertebrates.

Effects of storage conditions and digestion time on DNA amplification of biting midge (Culicoides) blood meals

BackgroundMolecular analysis of blood meals is increasingly used to identify the hosts of biting insects such as midges and mosquitoes. Successful host identification depends on the availability of sufficient host DNA template for PCR amplification, making it important to understand how amplification success changes under different storage conditions and with different durations of blood meal digestion within the insect gut before being placed into the storage medium.MethodWe characterised and compared the digestion profile of two species of Culicoides over a 96-h period using a novel set of general vertebrate primers targeting the 16S rRNA gene. A set number of individuals from each species were killed over 13 time points post-blood feeding and preserved in 95% ethanol. Samples were stored either at ambient room temperature or in a − 20 °C freezer to examine the effect of storage condition on the PCR amplification success of host DNA.ResultsWe found that amplification success across the 96-h sampling period post-feeding was reduced from 96 to 6% and 96% to 14% for Culicoides nubeculosus and Culicoides sonorensis, respectively. We found no effect of storage condition on PCR amplification success, and storage in 95% ethanol was sufficient to maintain high rates of amplifiable host DNA for at least 9 months, even at room temperature.ConclusionsThese findings highlight the limited time frame during which an individual may contain amplifiable host DNA and demonstrate the importance of timely sample capture and processing post-blood feeding. Moreover, storage in 95% ethanol alone is sufficient to limit host DNA degradation. These results are relevant to the design of studies investigating the biting behaviour and disease transmission potential of Culicoides and other biting Diptera.Graphical

Aedes albopictus host odor preference does not drive observed variation in feeding patterns across field populations

Laboratory and field-based studies of the invasive mosquito Aedes albopictus demonstrate its competency to transmit over twenty different pathogens linked to a broad range of vertebrate hosts. The vectorial capacity of Ae. albopictus to transmit these pathogens remains unclear, partly due to knowledge gaps regarding its feeding behavior. Blood meal analyses from field-captured specimens have shown vastly different feeding patterns, with a wide range of anthropophagy (human feeding) and host diversity. To address this knowledge gap, we asked whether differences in innate host preference may drive observed variation in Ae. albopictus feeding patterns in nature. Low generation colonies (F2–F4) were established with field-collected mosquitoes from three populations with high reported anthropophagy (Thailand, Cameroon, and Florida, USA) and three populations in the United States with low reported anthropophagy (New York, Maryland, and Virginia). The preference of these Ae. albopictus colonies for human versus non-human animal odor was assessed in a dual-port olfactometer along with control Ae. aegypti colonies already known to show divergent behavior in this assay. All Ae. albopictus colonies were less likely (p < 0.05) to choose the human-baited port than the anthropophilic Ae. aegypti control, instead behaving similarly to zoophilic Ae. aegypti. Our results suggest that variation in reported Ae. albopictus feeding patterns are not driven by differences in innate host preference, but may result from differences in host availability. This work is the first to compare Ae. albopictus and Ae. aegypti host preference directly and provides insight into differential vectorial capacity and human feeding risk.

Predicting potential transmission risk of Everglades virus in Florida using mosquito blood meal identifications.

The overlap between arbovirus host, arthropod vectors, and pathogen distributions in environmentally suitable habitats represents a nidus where risk for pathogen transmission may occur. Everglades virus (EVEV), subtype II Venezuelan equine encephalitis virus (VEEV), is endemic to southern Florida where it is transmitted by the endemic vector Culex cedecei between muroid rodent hosts. We developed an ecological niche model (ENM) to predict areas in Florida suitable for EVEV transmission based upon georeferenced vector-host interactions from PCR-based blood meal analysis from blood-engorged female Cx. cedecei females. Thirteen environmental variables were used for model calibration, including bioclimatic variables derived from Daymet 1 km daily temperature and precipitation values, and land use and land cover data representing percent land cover derived within a 2.5 km buffer from 2019 National Land Cover Database (NLCD) program. Maximum temperature of the warmest month, minimum temperature of the coldest month, and precipitation of the driest month contributed 31.6%, 28.5% and 19.9% to ENM performance. The land cover types contributing the greatest to the model performance were percent landcover of emergent herbaceous and woody wetlands which contributed 5.2% and 4.3% to model performance, respectively. Results of the model output showed high suitability for Cx. cedecei feeding on rodents throughout the southwestern portion of the state and pockets of high suitability along the northern east coast of Florida, while areas with low suitability included the Miami-Dade metropolitan area and most of northern Florida and the Panhandle. Comparing predicted distributions of Cx. cedecei feeding upon rodent hosts in the present study to historical human cases of EVEV disease, as well as antibodies in wildlife show substantial overlap with areas predicted moderate to highly suitable for these vector/host associations. As such, the findings of this study likely predict the most accurate distribution of the nidus of EVEV to date, indicating that this method allows for better inference of potential transmission areas than models which only consider the vector or vertebrate host species individually. A similar approach using host blood meals of other arboviruses can be used to predict potential areas of virus transmission for other vector-borne diseases.

First Evidence of Co-Circulation of Emerging Leishmania martiniquensis, Leishmania orientalis, and Crithidia sp. in Culicoides Biting Midges (Diptera: Ceratopogonidae), the Putative Vectors for Autochthonous Transmission in Southern Thailand.

Since 1996, autochthonous cases of emerging leishmaniasis caused by Leishmania (Mundinia) martiniquensis and Leishmania (Mundinia) orientalis have been more frequently reported, especially in the northern and southern parts of Thailand. However, the accurate identification of their natural vectors and reservoirs remains unconfirmed. Previous studies have suggested that these emerging parasites might be transmitted by other non-phlebotomine vectors. Herein, we speculated that Culicoides biting midges might act as the competent vectors responsible for autochthonous leishmaniasis in southern Thailand. In this research, 187 non-engorged, parous and gravid Culicoides females and 47 blood-engorged ones were trapped from the residences of two recently diagnosed visceral leishmaniasis patients in Sadao District and the unaffected site in Rattaphum District, Songkhla Province, southern Thailand. Species diversity and abundance of biting midges varied among the trapping sites. Using ITS1-PCR and BLASTn analysis, L. martiniquensis was predominantly detected in several Culicoides species, including C. peregrinus, C. oxystoma, C. mahasarakhamense, and C. huffi from the vicinity of patients' houses; and in C. fordae and C. fulvus from the unaffected site. L.&amp;nbsp;orientalis was also co-circulated in C. peregrinus and C. oxystoma caught near the second patient's house. Additionally, Crithidia sp. were also detected using SSU rRNA-PCR across Culicoides spp. Host blood meal analysis of eight different Culicoides species from the unaffected site also revealed that all trapped Culicoides had fed on cows and goats, indicating the possible role of these mammalian species as reservoir hosts. Essentially, this study is the first entomological investigation, revealing the co-circulation of emerging trypanosomatids among several species of Culicoides biting midges and strongly supporting the potential role of this insect group as the main vectors responsible for the epidemiology of autochthonous leishmaniasis in southern Thailand.

Molecular Identification of Host Blood Meals and Detection of Blood Parasites in Culicoides Latreille (Diptera: Ceratopogonidae) Collected from Phatthalung Province, Southern Thailand.

Simple SummaryBiting midges, which feed on a variety of hosts, transmit a wide range of harmful human and animal viruses and parasites. According to recent studies in Thailand, biting midges may act as potential vectors for leishmaniasis and trypanosomiasis. The purpose of this study is to look for Leishmania and Trypanosoma DNA in biting midges obtained in the field in a leishmaniasis-endemic area in Phatthalung province, Southern Thailand. In addition, we analysed engorged midges for host blood DNA and screened the collected midges for avian haemosporidian parasites. According to our findings, biting midges have a diversified feeding habit and may be able to transmit various pathogens, including trypanosomatid and avian haemosporidian parasites.Five hundred and fifty-nine female biting midges were collected, and seventeen species in six subgenera (Avaritia, Haemophoructus, Hoffmania, Meijerehelea, Remmia, and Trithecoides) and two groups (Clavipalpis and Shortti) were identified. The dominant Culicoides species was C. peregrinus (30.94%), followed by C. subgenus Trithecoides. From blood meal analysis of engorged biting midges, they were found to feed on cows, dogs, pigs, and avians. The majority of blood preferences of biting midges (68%; 49/72) displayed a mixed pattern of host blood DNA (cow and avian). The overall non-engorged biting midge field infectivity rate was 1.44 % (7/487). We detected Leucocytozoon sp. in three Culicoides specimens, one from each species: C. fulvus, C. oxystoma, and C. subgenus Trithecoides. Crithidia sp. was found in two C. peregrinus specimens, and Trypanosoma sp. and P. juxtanucleare were separately found in two C. guttifer. More consideration should be paid to the capacity of biting midges to transmit pathogens such as avian haemosporidian and trypanosomatid parasites. To demonstrate that these biting midges are natural vectors of trypanosomatid parasites, additional research must be conducted with a greater number of biting midges in other endemic regions.

Leishmania (Sauroleishmania) tarentolae isolation and sympatric occurrence with Leishmania (Leishmania) infantum in geckoes, dogs and sand flies

The trypanosomatid protist Leishmania tarentolae is a saurian-associated parasite vectored by the Sergentomyia minuta sand fly. This study aimed to confirm the circulation of L. infantum and L. tarentolae in sand flies, reptiles and dogs and to isolate new strains of these protists. Reptilian and sheltered dog blood samples were collected, and sand flies were captured. Samples were tested for Leishmania spp. using duplex real-time PCR (dqPCR) and real-time PCR (qPCR); the origin of blood meal was identified in engorged sand flies by conventional PCR. The reptilian blood and intestinal content of sand fly females were cultured. Dog sera were tested by IFAT using both Leishmania species. Four Tarentola mauritanica geckoes were molecularly positive for L. infantum or L. tarentolae, with no co-infections; moreover, amastigote-like forms of L. infantum were observed in the bone marrow. 24/294 sand flies scored positive for Leishmania spp. by dqPCR, 21 S. minuta and two Phlebotomus perniciosus were positive for L. tarentolae, while only a single Ph. perniciosus was positive for L. infantum. Blood meal analysis confirmed reptile and dog in S. minuta, dog and human in Ph. perniciosus and dog in Phlebotomus neglectus. Two axenic strains of L. tarentolae were obtained. Twelve of 19 dogs scored positive for L. infantum and L. tarentolae by IFAT and three of them also for L. infantum by dqPCR, and six by qPCR. These data confirm the sympatric circulation of L. infantum and L. tarentolae in geckoes, sand flies, and dogs, and suggest that geckoes may be infected with L. infantum.

Host preference of bluetongue virus vectors, Culicoides species associated with livestock in West Bengal, India: Potential relevance on bluetongue epidemiology

Determination of host choice of Culicoides species (Diptera: Ceratopogonidae), the vectors of bluetongue virus (BTV), is pivotal to ascertain the role of each species in the transmission of pathogens, pest management and enumeration of disease prediction models. Host preference of livestock associated Culicoides midges was investigated in West Bengal, India with four replicates of a 3 × 3 Latin square design during August and September 2021. Adult Culicoides were mouth aspirated from three BTV hosts viz., cattle, sheep and goats. Mouth aspirating was validated by the sweep net collections. The host-baited collections recorded seven Culicoides species; with the highest landing rate on cattle (n = 5,667; 92.9%) followed by sheep (n = 365; 6.0%) and goat (n = 67; 1.1%). Based on the Jacob's selectivity index, all midge species, except for Culicoides fulvus Sen & Das Gupta, encountered, preferred cattle over other mammalian hosts. Culicoides oxystoma Kieffer, the subgenus Trithecoides Wirth & Hubert and Culicoides actoni Smith, predominated on the ventral region (belly/flank) of the cattle. However, Culicoides peregrinus Kieffer and C. actoni were observed to be prevalent in the leg region of sheep. A significantly higher percentage of female (99.9%) with only 0.3% of male were trapped in aspiration based animal baited collections. On the other hand sweep net and light trap catch comprises of 50.7%, 89.7% female and 49.2%, 10.2% male respectively. Surprisingly, DNA based blood meal analysis revealed human blood from the midges trapped in UV-LED light traps. Supplying the first evidence that Culicoides similis Carter, Ingram & Macfie, C. fulvus and Culicoides palpifer Das Gupta & Ghosh, feed on humans.

The resting behavior of malaria vectors in different ecological zones of Ghana and its implications for vector control

BackgroundIn sub-Saharan Africa there is widespread use of long-lasting insecticidal nets and indoor residual spraying to help control the densities of malaria vectors and decrease the incidence of malaria. This study was carried out to investigate the resting behavior, host preference and infection with Plasmodium falciparum of malaria vectors in Ghana in the context of the increasing insecticide resistance of malaria vectors in sub-Saharan Africa.MethodsIndoor and outdoor resting anopheline mosquitoes were sampled during the dry and rainy seasons in five sites in three ecological zones [Sahel savannah (Kpalsogo, Pagaza, Libga); coastal savannah (Anyakpor); and forest (Konongo)]. Polymerase chain reaction-based molecular diagnostics were used to determine speciation, genotypes for knockdown resistance mutations (L1014S and L1014F) and the G119S ace1 mutation, specific host blood meal origins and sporozoite infection in the field-collected mosquitoes.ResultsAnopheles gambiae sensu lato (s.l.) predominated (89.95%, n = 1718), followed by Anopheles rufipes (8.48%, n = 162) and Anopheles funestus s.l. (1.57%, n = 30). Sibling species of the Anopheles gambiae s.l. revealed Anopheles coluzzii accounted for 63% (95% confidence interval = 57.10–68.91) and 27% (95% confidence interval = 21.66–32.55) was Anopheles gambiae s. s.. The mean resting density of An. gambiae s.l. was higher outdoors (79.63%; 1368/1718) than indoors (20.37%; 350/1718) (Wilcoxon rank sum test, Z = − 4.815, P < 0.0001). The kdr west L1014F and the ace1 mutation frequencies were higher in indoor resting An. coluzzii and An. gambiae in the Sahel savannah sites than in the forest and coastal savannah sites. Overall, the blood meal analyses revealed that a larger proportion of the malaria vectors preferred feeding on humans (70.2%) than on animals (29.8%) in all of the sites. Sporozoites were only detected in indoor resting An. coluzzii from the Sahel savannah (5.0%) and forest (2.5%) zones.ConclusionsThis study reports high outdoor resting densities of An. gambiae and An. coluzzii with high kdr west mutation frequencies, and the presence of malaria vectors indoors despite the use of long-lasting insecticidal nets and indoor residual spraying. Continuous monitoring of changes in the resting behavior of mosquitoes and the implementation of complementary malaria control interventions that target outdoor resting Anopheles mosquitoes are necessary in Ghana.Graphical abstract

Species Diversity, Habitat Distribution, and Blood Meal Analysis of Haematophagous Dipterans Collected by CDC-UV Light Traps in the Dominican Republic.

Haematophagous insects cause major economic losses by both direct damage and the transmission of pathogens. However, the biting Diptera species in the Caribbean region have been poorly documented. During 2021, CDC downdraft suction traps with UV light were employed to assess both the species occurrence and blood meal sources across three different habitats in the Dominican Republic. Eighteen species of mosquitoes (n = 274), six species of Culicoides (n = 803), two black fly species (n = 2), and one species of muscid fly (n = 25) were identified at species-level by morphology and/or molecular phylogenetic approaches based on the mitochondrial cytochrome c oxidase subunit 1 (COI). Engorged mosquito (n = 5) and Culicoides (n = 28) females showed host preferences derived exclusively from mammals (cows and pigs), except Culex species containing the blood of chickens. Our study provides new records of the Diptera Dominican catalogue (Culex salinarius for the Greater Antilles, Culicoides jamaicensis for Hispaniola, and Culicoides haitiensis and Culicoides borinqueni for the Dominican Republic), the first available COI DNA sequences of different Diptera in the GenBank, some pictures of diagnostic features of closely related specimens, spatial distribution across the habitats studied, and new insights on their feeding preferences in the Caribbean region.

Culicoides species community composition and feeding preferences in two aquatic ecosystems in northern Spain

BackgroundAquatic ecosystems provide breeding sites for blood-sucking insects such as Culicoides biting midges (Diptera: Ceratopogonidae), but factors affecting their distribution and host choice are poorly understood. A study was undertaken at two nature reserves in northern Spain to examine the abundance, species composition, population dynamics and feeding patterns of biting midges between 2018 and 2019.MethodsCulicoides were captured by light suction traps baited with CO2 and by sweep netting vegetation. Blood meals and species identification of blood-fed specimens were determined using cytochrome c oxidase I subunit (COI) DNA barcoding. Multivariate generalized linear models were used to evaluate the associations between the abundance of Culicoides, the species richness and other parameters.ResultsThe 4973 identified specimens comprised 28 species of Culicoides. These included two species reported for the first time in northern Spain, thus raising to 54 the number of Culicoides species described in the region. Specimens of all 28 species and 99.6% of the total specimens collected were caught in suction traps, while sweep netting vegetation revealed just 11 species and 0.4% of the total specimens. Midge abundance peaked in June/early July, with five species comprising > 80% of the captures: Culicoides alazanicus (24.9%), Culicoides griseidorsum (20.3%), Culicoides poperinghensis (16.2%), Culicoides kibunensis (10.7%) and Culicoides clastrieri (9.6%). DNA barcode analysis of blood meals from eight Culicoides species revealed that they fed on 17 vertebrate species (3 mammals and 14 birds). Species in the subgenus Avaritia were primarily ornithophilic, except for C. griseidorsum and C. poperinghensis. Host DNA from blood meals was successfully amplified from 75% of blood-fed females. A pictorial blood meal digestion scale is provided to accurately assess the blood-fed status of female Culicoides.ConclusionsThe large number of different blood meal sources identified in the midges captured in this study signals the likely importance of wild birds and mammals (e.g. red deer and wild boar) as reservoir/amplifying hosts for pathogens. Available hosts are more exposed to being bitten by biting midge populations in aquatic ecosystems in late spring and early summer.

Blood meal analysis of Culicoides species associated with livestock in West Bengal, India

Knowledge gaps exist on the feeding pattern and host range of bluetongue virus vectors, Culicoides species, associated with livestock in India. Adult midges were trapped with ultraviolet light traps at 13 household farms adjacent to human biotope. Host DNA was isolated from individual females (n=101; blood engorged-82, gravid-4 and parous-15) and subjected to PCR amplification targeting CytB and 16S rRNA gene fragments followed by sequencing of amplified DNA samples. However, DNA sequences from only 71 individuals (70.3%) comprising of 10 Culicoides species were obtained. Blood meal analysis revealed at least 10 species that fed on five mammalian hosts including humans, but surprisingly none tested positive for birds. Results revealed that Culicoides innoxius tested positive for four not previously recognized species indicating a potential role as a vector species. Likewise, Culicoides shortti and Culicoides hegneri preferred goat and cattle respectively as hosts, whereas Culicoides palpifer preferred cattle along with buffalo as hosts, which is being reported for the first time. This is the first document on DNA-based blood meal identification and feeding preference of Culicoides midges associated with livestock in India.

Blood feeding habits of mosquitoes: hardly a bite in South America.

Mosquito blood feeding plays a key role in epidemiology. Despite its importance and large number of studies worldwide, less attention has been paid in South America. We summarized some general concepts and methodological issues related to the study of mosquito blood feeding habits, and compiled and analyzed all published information regarding the subject in the continent until 2020. Available literature comprised 152 scientific studies, that pursued different approaches: human landing catches (102 studies), baited trap (19), and blood meal analyses of collected specimens (38). Among the latter, 23 used serological and 15 molecular techniques. Species most frequently studied were those incriminated in malaria transmission, whereas relevant vectors such as Aedes aegypti, Ae. albopictus, and Haemagogus janthinomys were surprisingly neglected. Brazil was the leading country both in number of works and species studied. For over 70% of the species and three out of 13 South American countries there is no single information on mosquito blood feeding habits. Data from baited traps included 143 mosquito species, 83.9% of which were attracted to humans, either exclusively (10.5%) or in combination with other vertebrates (73.4%). Host blood identification of field collected specimens provided data on 102 mosquito species, and 60.8% of these fed on humans (55.9% combined with other vertebrates). Only 17 of the 73 species assessed by both methods yielded similar feeding patterns. Finally, supplementary tables are provided in a comprehensive summary of all information available and information gaps are highlighted for future research in the continent.Supplementary InformationThe online version contains supplementary material available at 10.1007/s00436-022-07537-0.

Blood Meal Analysis and Molecular Detection of Leishmania DNA in Wild-Caught Sand Flies in Leishmaniasis Endemic Areas of Turkey and Northern Cyprus.

Phlebotomine sand flies (Diptera: Psychodidae) are known as the vector of diseases such as leishmaniasis, bartonellosis and viral diseases. The aim of this study is to detect the host feeding pattern of sand flies in the endemic areas for leishmaniasis in Turkey (Antalya, Kayseri) and Northern Cyprus (TRNC) as well as the presence of Leishmania DNA in the specimens. One-hundred seventy-six blood-fed sand fly specimens were examined for blood meal analysis. A SYBR Green-PCR assay was performed with specific forward primers for each host and a universal reverse primer. Primers of human and goat were used together in multiplex PCR while goat and cow were studied separately. ITS-1 qPCR assay was also performed on both blood-fed and non-blood-fed females to detect Leishmania parasites. Blood sources could be detected in 69 out of 176 blood-fed sand fly specimens. The results of blood meal analysis showed that specimens were fed mostly on cows (22.2%) followed by humans (5.7%), goats (2.8%) and dogs (0.6%). Multiple feeding patterns were also detected as human + cow (3.4%), cow + goat (2.8%) and human + goat (1.7%). Five of the blood-fed specimens were Leishmania spp. positive: P. major s.l. (n = 1), P. tobbi (n = 2) were L. tropica positive from Antalya, P. simici was positive for L. infantum from Kayseri and P. papatasi (n = 1) was positive for L. major from Cyprus. Leishmania infection rates were determined as 3.79%, 1.69% and 2.63% among the blood-fed sand fly specimens in Antalya, Kayseri and TRNC, respectively. The SYBR-Green-based multiplex PCR assay is a cost-effective and promising tool for blood meal identification of wild-caught sand flies as well as other blood-sucking arthropods. Feeding patterns of important vector species detected in the present study show the high risk in these endemic areas. As a next step, to identify the blood source in a shorter time and to make the test more sensitive, development of this assay to probe-based and multiplex PCR will be also planned.