Identification Of Blood Meal Sources Research Articles

Ecological interactions of sand flies, hosts, and Leishmania panamensis in an endemic area of cutaneous leishmaniasis in Colombia.

The transmission dynamics of leishmaniasis are complex. There is also a lack of information about the ecological relationships between the vector/host/parasite at a more local and specific level. The Andean region concentrates more than 50% of Colombia's cutaneous leishmaniasis (CL) cases. The study of the ecological interactions of sand flies through the identification of blood sources has provided information on the female's opportunistic behavior, feeding on various hosts. Therefore, this study aimed to determine sand flies' ecological interactions with Leishmania parasites and their blood sources in an endemic area of CL. A total of 4,621 sand flies were collected, comprising 20 species, in which the most abundant were Nyssomyia yuilli yuilli (55.4%), Psychodopygus ayrozai (14.5%) and Ps. panamensis (13.4%). Sequences of 12S gene fragment were analyzed using the BLASTn search tool. Blood-meal source identification was successfully performed for 47 sand flies, detecting seven vertebrate species, human and armadillo being the most frequent. Leishmania DNA was amplified in four female pools, constituted by Ny. yuilli yuilli and Ps. ayrozai, and the identification through RFLP detected Leishmania (Viannia) panamensis in the positive pools. The interactions between the sand fly species, local mammalian fauna and the Leishmania parasite in this active focus of CL, provide evidence of the potential role of two different species in the maintenance of the parasite transmission, important information for the understanding of the ecoepidemiology and transmission dynamics of the disease in Andean endemic areas. However its necessary further evaluations of the vector and host competence in the transmission and maintenance of Leishmania spp, in these complex and diverse areas.

Investigating potential sand fly vectors after the first reported outbreak of cutaneous leishmaniasis in Ghana

BackgroundLeishmaniasis is a parasitic disease caused by species of the genus Leishmania, which are transmitted through the bite of infected female sand flies. Since the first reported outbreak of cutaneous leishmaniasis in Ghana, in 1999, there has been limited published information on its vectors and reservoir hosts there. Previous studies have shown strong dominance of the sand fly genus Sergentomyia over the genus Phlebotomus in Ghana. Thus the aim of this study was to determine the possible sand fly vector species in Ghana, as well as their human-feeding behavior, from the time of the first reported outbreak of CL in the country.MethodsSand flies were collected from randomly selected houses in three communities. They were identified and used for blood meal source identification and the detection of Leishmania infection using molecular methods. ResultsA total of 1051 female sand flies were morphologically identified, of which Sergentomyia africana africana (29%) was the predominant species. Among the 275 female sand flies that had blood-fed, the identified blood meal sources included chicken (33.8%) and goat (12.4%); the percentage of human blood meals was 32%. Single-source and mixed-source blood meals were identified in Sergentomyia africana africana (11.6%), Sergentomyia ingrami (14.9%) and Sergentomyia simillima (20%), with S. simillima having the highest proportion of blood meals that included human blood (14.6%). Using molecular methods, unfed sand flies and identified human-feeding species were examined for the presence of Leishmania DNA. Pool screening analysis revealed three pools of S. ingrami positive for Leishmania major DNA, with an infection rate of 1.27% (95% confidence interval 2.467–3.647).ConclusionsThe findings suggest that some Sergentomyia species may be involved in the transmission of cutaneous leishmaniasis in Ghana. However, the role of S. ingrami as a vector of leishmaniasis in Ghana needs to be conclusively validated by isolating the parasite from this species and through experimental transmission studies.Graphical abstract

Blood meal source identification and RNA virome determination in Japanese encephalitis virus vectors collected in Ishikawa Prefecture, Japan, show distinct avian/mammalian host preference.

In Asia, Culex mosquitoes are of particular interest because of their role in maintaining endemic mosquito-borne viral diseases, including the Japanese encephalitis virus (JEV). Nonetheless, host-feeding preferences, along with naturally infecting RNA viruses in certain Culex species, remain understudied. In this study, selected blood-fed mosquitoes were processed for avian and mammalian blood meal source identification. Concurrently, cell culture propagation and high-throughput sequencing (HTS) approaches were used to determine the RNA virome of Culex mosquitoes collected in Ishikawa Prefecture, Japan. The identification of blood meal sources from wild-caught Culex spp. revealed that Culex (Culex) tritaeniorhynchus Giles, 1901, has a robust preference toward wild boar (62%, 26/42), followed by heron (21%, 9/42). The other two species, Culex (Oculeomyia) bitaeniorhynchus Giles, 1901, and Culex (Culex) orientalis Edwards, 1921, showed a distinct preference for avian species, including migratory birds. From the HTS results, 34 virus sequences were detected, four of which were newly identified virus sequences of unclassified Aspiviridae, Qinviridae, Iflaviridae, and Picornaviridae. The absence of observable cytopathic effects in mammalian cells and phylogenetic analysis suggested that all identified virus sequences were insect-specific. Further investigations involving other mosquito populations collected in different areas are warranted to explore previously unknown vertebrate hosts that may be linked to JEV dispersal in nature.

Utility of MALDI-TOF MS for determination of species identity and blood meal sources of primary malaria vectors on the Kenyan coast

Background: Protein analysis using matrix-assisted laser desorption/ionization time-of-flight mass-spectrometry (MALDI-TOF MS) represents a promising tool for entomological surveillance. In this study we tested the discriminative power of this tool for measuring species and blood meal source of main Afrotropical malaria vectors on the Kenyan coast. Methods: Mosquito collections were conducted along the coastal region of Kenya. MALDI-TOF MS spectra were obtained from each individual mosquito’s cephalothorax as well as the abdomens of blood-engorged mosquitoes. The same mosquitoes were also processed using gold standard tests: polymerase chain reaction (PCR) for species identification and enzyme linked immunosorbent assay (ELISA) for blood meal source identification. Results: Of the 2,332 mosquitoes subjected to MALDI-TOF MS, 85% (1,971/2,332) were considered for database creation and validation. There was an overall accuracy of 97.5% in the identification of members of the An. gambiae (An. gambiae, 100%; An. arabiensis, 91.9%; An. merus, 97.5%; and An. quadriannulatus, 90.2%) and An. funestus (An. funestus, 94.2%; An. rivulorum, 99.4%; and An. leesoni, 94.1%) complexes. Furthermore, MALDI-TOF MS also provided accurate (94.5% accuracy) identification of blood host sources across all mosquito species. Conclusions: This study provides further evidence of the discriminative power of MALDI-TOF MS to identify sibling species and blood meal source of Afrotropical malaria vectors, further supporting its utility in entomological surveillance. The low cost per sample (<0.2USD) and high throughput nature of the method represents a cost-effective alternative to molecular methods and could enable programs to increase the number of samples analysed and therefore improve the data generated from surveillance activities.

“Mi Casa, Tu Casa”: the coati nest as a hub of Trypanosoma cruzi transmission in the southern Pantanal biome revealed by molecular blood meal source identification in triatomines

BackgroundThe study of the ecology of Trypanosoma cruzi is challenging due to its extreme adaptive plasticity, resulting in the parasitism of hundreds of mammal species and dozens of triatomine species. The genetic analysis of blood meal sources (BMS) from the triatomine vector is an accurate and practical approach for gathering information on which wild mammal species participate in a local transmission network. South American coatis, Nasua nasua, act as important reservoir host species of T. cruzi in the Pantanal biome because of their high rate of infection and elevated parasitemia, with the main discrete typing unit (DTU) lineages (TcI and TcII). Moreover, the carnivore coati is the only mammal species to build high arboreal nests for breeding and resting that can be shared by various vertebrate and invertebrate species. Herein, we applied the sensitive and specific methodology of DNA barcoding and molecular cloning to study triatomines found in a coati nest to access the diversity of mammal species that explore this structure, and therefore, may be involved in the parasite transmission network.MethodsTwenty-three Triatoma sordida were collected in one coati’s nest in the subregion of Nhecolândia, Pantanal. The DNA isolated from the gut of insects was subjected to BMS detection by PCR using universal primers that flank variable regions of the cytochrome b (cytb) and 12S rDNA mitochondrial genes from vertebrates. The Trypanosoma spp. diagnosis and DTU genotyping were based on an 18S rDNA molecular marker and also using new cytb gene primers designed in this study. Phylogenetic analyses and chord diagrams were constructed to visualize BMS haplotypes, DTU lineages detected on vectors, and their interconnections.ResultsTwenty of 23 triatomines analyzed were PCR-positive (86.95%) showing lineages T. cruzi DTU TcI (n = 2), TcII (n = 6), and a predominance of TcI/TcII (n = 12) mixed infection. Intra-DTU diversity was observed mainly from different TcI haplotypes. Genetic analyses revealed that the southern anteater, Tamandua tetradactyla, was the unique species detected as the BMS of triatomines collected from the coati’s nest. At least three different individuals of T. tetradactyla served as BMS of 21/23 bugs studied, as indicated by the cytb and 12S rDNA haplotypes identified.ConclusionsThe identification of multiple BMS, and importantly, different individuals of the same species, was achieved by the methodology applied. The study demonstrated that the southern anteaters can occupy the South American coati’s nest, serving as the BMS of T. sordida specimens. Since anteaters have an individualist nonsocial behavior, the three individuals detected as BMS stayed at the coati’s nest at different times, which added a temporal character to BMS detection. The TcI and TcII infection, and significantly, a predominance of TcI/TcII mixed infection profile with different TcI and TcII haplotypes was observed, due to the discriminatory capacity of the methodology applied. Tamandua tetradactyla, a host which has been little studied, may have an important role in the T. cruzi transmission in that Pantanal subregion. The data from the present study indicate the sharing of coatis’ nests by other mammal species, expanding the possibilities for T. cruzi transmission in the canopy strata. We propose that coatis’ nests can act as the true hubs of the T. cruzi transmission web in Pantanal, instead of the coatis themselves, as previously suggested.Graphical

Species diversity, blood meal source and infection rate of malaria vectors in the village of Kodougou, Northwestern Burkina Faso

Background and Objectives: Anopheles gambiae members are the main malaria vectors in Burkina Faso seconded by Anopheles funestus. However, other anophelines species are found in particular environments such as forest and localities bordering the rivers. The aim of this study, was to assess the species diversity, the blood feeding preferences and the infection rate of all anophelines species found in Kodougou, a village situated near the Mouhoun River.Methods: Adult mosquitoes were collected from September to December 2018 using insecticide spraying catch in 160 randomly selected houses. DNA extracted from wing/legs, head/thorax, abdomens were used respectively for mosquitoes species molecular identification, infection detection and blood meal source identification from female anophelines. The entomological inoculation (EIR) rate was to estimate malaria transmission intensity in the study area.Results: A total of 1528 anophelines were collected consisting of 1392 (91.1%) Anopheles gambiae, 115 (7.5%) Anopheles nili, 8 (0.5%) Anopheles funestus, 13 (0.9%) Anopheles pharoensis. The most abundant specie was Anopheles coluzzii representing for 87% of total of Anopheles gambiae s.l. Blood meal source was mostly human host (88%), followed by cattle (11%) and pigs (1%). The overall EIR was 0.1 infective bite per human during the study period. The highest EIR with 0.08 infective bite per human (i.b/h) for Anopheles coluzzii. Likely, minor vectors Anopheles nili and Anopheles pharoensis presented an EIR of 0.02 i.b/h and 0.04 i.b/h respectively. Conclusion: This study shown that in addition to Anopheles gambiae, minor vectors like Anopheles nili and Anopheles pharoensis contribute to malaria transmission in the northern part of Burkina Faso. So, control measures should take into account these species for effective result.

Molecular detection of blood meal source up to three months since the last meal: Experimental starvation resistance in triatomines

The identification of Blood Meal Source (BMS) in hematophagous vectors contributes to a better understanding of the ecology of hemoparasite transmission. Those insects can endure long periods without feeding, waiting for a favorable setting. Although this represents an important behavior observed in those groups, such as triatomines, little is known about how time can affect BMS detection, especially considering extended periods. To comprehend to which extent this behavioral phenomenon can impact molecular detection, we submitted two groups of Rhodnius robustus to increasing periods of starvation under experimental conditions. It was possible to recover the BMS until the 12th week of the starvation process. Nymphs were more resistant to prolonged periods of starvation (up to more than 189 days) than adults (maximum of 137 days), with no significant difference between their weights after being fed. The study brought new insights to the understanding of Trypanosoma cruzi transmission by R. robustus in the nature, with a temporal perspective.

Identification of blood meal sources in species of genus Rhodnius in four different environments in the Brazilian amazon

Chagas disease is a zoonotic disease caused by the hemoflagellate Trypanosoma cruzi and transmitted primarily by triatomine vectors. Triatomines are hematophagous insects that feed on a variety of vertebrate hosts. The Chagas disease transmission cycle is closely related to the interactions between vectors, parasites, and vertebrate hosts. Knowledge of triatomine food sources is critical to understanding Chagas disease transmission dynamics. The aim of this study was to identify blood meal sources used by triatomines from different environments in the Brazilian Amazon. A total of 25 captures were conducted in four environments. Triatomine specimens were captured on palm trees and were identified by morphological and morphometric characters. Blood meal sources identification was conducted using a traditional PCR followed by Sanger sequencing of mtDNA cytb gene. Sequencing was successful in 167 specimens and a total of 21 blood meal sources were identified: two reptilians, six birds, and 13 mammals. Among these 21 species, three (Tamandua tetradactyla, Didelphis marsupialis and Rattus rattus) are considered reservoir of T. cruzi. Knowledge of the relationship between triatomines and possible reservoirs can help to elucidate the enzootic cycle of T. cruzi in the Amazon region and guide control strategies for Chagas disease transmission in that region.

Blood meal profile and positivity rate with malaria parasites among different malaria vectors in Sudan

BackgroundMalaria is a life-threatening public health problem globally with particularly heavy burden in the sub-Saharan Africa including Sudan. The understanding of feeding preference of malaria vectors on different hosts is a major challenge for hindering the transmission cycle of malaria. In this study, blood meals taken by blood-fed Anopheles mosquitoes collected from the field in malaria endemic areas of Sudan were analysed for source of blood meal and malaria parasite presence.MethodsAnopheles mosquitoes were collected from different regions in Sudan: Khartoum state, Sennar state, Northern state, and El Gedarif state between September 2020 and February 2021. Anopheles mosquitoes were collected using the standard pyrethrum spray catch and back-pack aspirator. Mosquito samples were sorted and morphologically identified to species level using international identification keys. Morphologically identified mosquito species were also confirmed using PCR. Genomic DNA was extracted from mosquitoes for molecular identification of blood meal source and parasite detection. The presence of Plasmodium species DNA in each mosquito sample was investigated using semi-nested PCR. Frequency of each blood meal source, Anopheles mosquito vector, and malaria parasite detected was calculated. Positivity rate of each fed female Anopheles mosquito was calculated for each species.ResultsA total of 2132 Anopheles mosquitoes were collected. 571 (26.8%) were males and 1561 (73.2%) were females classified based on their abdominal status into 1048 (67.1%) gravid, 274 (17.6%) fed, and 239 (15.3%) unfed females. Among the blood fed Anopheles mosquitoes, 263 (96.0%) were morphologically identified and confirmed using PCR to Anopheles arabiensis, 9 (3.3%) to Anopheles stephensi, and 2 (0.7%) to Anopheles rufipes. Of 274 blood-fed An. arabiensis, 68 (25.9%) fed on mixed blood meals from human and cattle, 8 (3.0%) fed on cattle and goat, and 13 (4.8%) fed on human, cattle and goat. For single blood meal sources, 70 (26.6%) fed on human, 95 (36.1%) fed on cattle, 8 (3.0%) fed on goat, and 1 (0.4%) fed on dog. While An. rufipes and An. stephensi fed on dog (2; 0.75%) and cattle (9; 3.3%), respectively. Plasmodium parasite detection in the blood meals showed that 25/274 (9.1%) An. arabiensis meals were positive for Plasmodium vivax and 19/274 (6.9%) An. arabiensis meals were positive for Plasmodium falciparum. The rate of positivity of An. arabiensis with any Plasmodium species was 16.7%. However, the positivity rate with P. falciparum only was 7.2%, while P. vivax was 9.5%. Both An. rufipes and An. stephensi were having positivity rates of 0.0% each.ConclusionsThis study which was mainly on blood-fed Anopheles mosquitoes showed a diversity in the type of diet from human, cattle, and goat. Anopheles mosquitoes especially An. arabiensis in Sudan, are opportunistic blood feeders and can feed broadly on both human and cattle. The application of blood meal identification is not only important in malaria vector epidemiological surveillance but also is very useful in areas where arthropods exhibit zoophilic feeding behaviour for mammals.

Detection of Plasmodium Sporozoites and Blood-Meal Source in a Population of Anopheles Coustani Senso Lato in Kakamega County, Western Kenya

Background Re-emerging of malaria vectors in the highlands of Western Kenya pose a challenge to malaria eradication efforts . Anopheles coustanis.l is a sub-Saharan mosquito species implicated in transmission of malaria in many parts of Africa as a secondary vector. It is zoo-anthropophilic species that has been assumed to be of negligible importance which may not be the case. This study therefore aimed at getting the malaria vectorial system of the study area, has tried to determine the relative abundance of the vectors and a new outdoor malaria vector. Methods A cross sectional study was carried out in April to June, 2020 in Eluche location, Mumias East sub-County, Kakamega County, Kenya. Pyrethrum spray collections (PSC) and Centers for Disease Control (CDC) and prevention light traps were used for sampling mosquitoes. Mosquitoes were collected both indoors; between 0700h and 1100h using PSC and outdoors between 1800h and 0700h using CDC light traps. All mosquitoes were identified morphologically and female Anopheles’ heads and thorax were analyzed using Polymerase Chain Reaction (PCR) for Plasmodium sporozoite detection and blood-meal source identification. Results A total of 376 female Anopheles mosquitoes were collected composed of: An. coustanis.l, 42.55%; An. funestus, 27.66%; An. maculipulpis, 25.00%; An. arabiensis, 4.26% and An. gambiaes.s, 0.53%. Malaria sporozoites were detected in only An. coustani (1.06%). Conclusion There is a possibility of Anopheles coustani mosquito involvement in malaria transmission in Mumias east, Kakamega County.

Identification of Blood Meal Sources in Triatomine Bugs in Four Diferent Environments in the Brazilian Amazon

Identification of Blood Meal Sources in Triatomine Bugs in Four Diferent Environments in the Brazilian Amazon

Genetic diversity of Anopheles stephensi in Ethiopia provides insight into patterns of spread

BackgroundThe recent detection of the South Asian malaria vector Anopheles stephensi in the Horn of Africa (HOA) raises concerns about the impact of this mosquito on malaria transmission in the region. Analysis of An. stephensi genetic diversity and population structure can provide insight into the history of the mosquito in the HOA to improve predictions of future spread. We investigated the genetic diversity of An. stephensi in eastern Ethiopia, where detection suggests a range expansion into this region, in order to understand the history of this invasive population.MethodsWe sequenced the cytochrome oxidase subunit I (COI) and cytochrome B gene (CytB) in 187 An. stephensi collected from 10 sites in Ethiopia in 2018. Population genetic, phylogenetic, and minimum spanning network analyses were conducted for Ethiopian sequences. Molecular identification of blood meal sources was also performed using universal vertebrate CytB sequencing.ResultsSix An. stephensi COI-CytB haplotypes were observed, with the highest number of haplotypes in the northeastern sites (Semera, Bati, and Gewana towns) relative to the southeastern sites (Kebridehar, Godey, and Degehabur) in eastern Ethiopia. We observed population differentiation, with the highest differentiation between the northeastern sites compared to central sites (Erer Gota, Dire Dawa, and Awash Sebat Kilo) and the southeastern sites. Phylogenetic and network analysis revealed that the HOA An. stephensi are more genetically similar to An. stephensi from southern Asia than from the Arabian Peninsula. Finally, molecular blood meal analysis revealed evidence of feeding on cows, goats, dogs, and humans, as well as evidence of multiple (mixed) blood meals.ConclusionWe show that An. stephensi is genetically diverse in Ethiopia and with evidence of geographical structure. Variation in the level of diversity supports the hypothesis for a more recent introduction of An. stephensi into southeastern Ethiopia relative to the northeastern region. We also find evidence that supports the hypothesis that HOA An. stephensi populations originate from South Asia rather than the Arabian Peninsula. The evidence of both zoophagic and anthropophagic feeding support the need for additional investigation into the potential for livestock movement to play a role in vector spread in this region.Graphical

Identification of Bloodmeals from Sand Flies (Diptera: Psychodidae) Collected in the Parque Nacional do Viruá, State of Roraima, Brazil.

The transmission of pathogens that cause leishmaniases occurs by the bite of female sand flies (Diptera: Psychodidae) in their vertebrate hosts, which makes the identification of their bloodmeal sources an important step for the control and epidemiology of these diseases. In Brazil, the state of Roraima has a great diversity of sand flies, vertebrate hosts, and protozoan Leishmania, but little is known about the host blood-feeding preferences of sand flies. Thus, we evaluated the bloodmeal sources of sand flies collected from their sylvatic habitats in Parque Nacional do Viruá, Roraima. Fieldwork was carried-out between 13th and 18th August 2019 using CDC light traps. Sand flies were slide-mounted and morphologically identified using the head and last segments of the abdomen. Engorged females had their DNA extracted, followed by amplification and sequencing of the cytochrome b (cytb) molecular marker for vertebrates. Sequences were analyzed and compared with those from GenBank using the BLASTn search tool, in addition to the reconstruction of a phylogenetic tree to demonstrate the clustering pattern of these sequences. A total of 1,209 sand flies were identified, comprising 20 species, in which the most abundant were Psychodopygus ayrozai (Barretto and Coutinho) (42.10%) and Psychodopygus chagasi (Costa Lima) (26.22%). Bloodmeal source identification was successfully performed for 34 sand flies, that confirm four vertebrate species, being the most abundant the armadillo Dasypus novemcinctus Linnaeus, 1758 (Cingulata: Dasypodidae).

Species diversity and blood meal sources of phlebotomine sand flies (Diptera: Psychodidae) from Los Tuxtlas, Veracruz, Mexico

Phlebotomine sand flies can transmit several species of the genus Leishmania, that cause leishmaniasis, a serious neglected tropical disease worldwide. Although Mexico is an endemic country for the disease, studies on the biology, ecology, and the identification of blood meal sources of phlebotomine sand flies in some states remain unexplored. For this reason, this study aimed to evaluate the species diversity of sand flies, and identify their blood meal sources in the Biosphere Reserve Los Tuxtlas, Veracruz, an area with a high prevalence of Leishmania infantum. The cumulative sampling effort of sand flies covered 300 trap-nights between March 2011 and May 2013. For estimating species diversity, we calculated the species richness (q = 0), the diversity of the species (q = 1) and the dominant species (q = 2). To identify the blood meal sources, we amplified and sequenced a fragment of ≈400 bp of the vertebrate Cytb gene. A total of 951 specimens belonging to 15 species were collected. Psathyromyia aclydifera and Psychodopygus panamensis were the most abundant species. We were able to identify seven terrestrial vertebrate species, among which human beings were the most common source of the blood meal. In this study, relevant information on the structure of sand fly populations and their blood meal sources was obtained, providing basic and important information about the interactions between sand flies, hosts and Leishmania species.

Modernizing the Toolkit for Arthropod Bloodmeal Identification

Simple SummaryThe ability to identify the source of vertebrate blood in mosquitoes, ticks, and other blood-feeding arthropod vectors greatly enhances our knowledge of how vector-borne pathogens are spread. The source of the bloodmeal is identified by analyzing the remnants of blood remaining in the arthropod at the time of capture, though this is often fraught with challenges. This review provides a roadmap and guide for those considering modern techniques for arthropod bloodmeal identification with a focus on progress made in the field over the past decade. We highlight genome regions that can be used to identify the vertebrate source of arthropod bloodmeals as well as technological advances made in other fields that have introduced innovative new ways to identify vertebrate meal source based on unique properties of the DNA sequence, protein signatures, or residual molecules present in the blood. Additionally, engineering progress in miniaturization has led to a number of field-deployable technologies that bring the laboratory directly to the arthropods at the site of collection. Although many of these advancements have helped to address the technical challenges of the past, the challenge of successfully analyzing degraded DNA in bloodmeals remains to be solved.Understanding vertebrate–vector interactions is vitally important for understanding the transmission dynamics of arthropod-vectored pathogens and depends on the ability to accurately identify the vertebrate source of blood-engorged arthropods in field collections using molecular methods. A decade ago, molecular techniques being applied to arthropod blood meal identification were thoroughly reviewed, but there have been significant advancements in the techniques and technologies available since that time. This review highlights the available diagnostic markers in mitochondrial and nuclear DNA and discusses their benefits and shortcomings for use in molecular identification assays. Advances in real-time PCR, high resolution melting analysis, digital PCR, next generation sequencing, microsphere assays, mass spectrometry, and stable isotope analysis each offer novel approaches and advantages to bloodmeal analysis that have gained traction in the field. New, field-forward technologies and platforms have also come into use that offer promising solutions for point-of-care and remote field deployment for rapid bloodmeal source identification. Some of the lessons learned over the last decade, particularly in the fields of DNA barcoding and sequence analysis, are discussed. Though many advancements have been made, technical challenges remain concerning the prevention of sample degradation both by the arthropod before the sample has been obtained and during storage. This review provides a roadmap and guide for those considering modern techniques for arthropod bloodmeal identification and reviews how advances in molecular technology over the past decade have been applied in this unique biomedical context.

Host feeding patterns of Nyssorhynchus darlingi (Diptera: Culicidae) in the Brazilian Amazon

Nyssorhynchus darlingi (Root) is the dominant malaria vector in the Brazilian Amazon River basin, with additional Anophelinae Grassi species involved in local and regional transmission. Mosquito blood-feeding behavior is an essential component to define the mosquito-human contact rate and shape the transmission cycle of vector-borne diseases. However, there is little information on the host preferences and blood-feeding behavior of Anophelinae vectors in rural Amazonian landscapes. The barrier screen sampling (BSS) method was employed to sample females from 34 peridomestic habitats in 27 rural communities from 11 municipalities in the Brazilian Amazon states of Acre, Amazonas, Pará and Rondônia, from August 2015 to November 2017. Nyssorhynchus darlingi comprised 97.94% of the females collected resting on barrier screens, and DNA sequence comparison detected 9 vertebrate hosts species. The HBI index ranged from 0.03–1.00. Results revealed the plasticity of Ny. darlingi in blood-feeding on a wide range of mainly mammalian hosts. In addition, the identification of blood meal sources using silica-dried females is appropriate for studies of human malaria vectors in remote locations.

Studies of host preferences of wild-caught Phlebotomus orientalis and Ph. papatasi vectors of leishmaniasis in Sudan.

IntroductionUnderstanding the feeding behavior and host choice of sand flies provides valuable information on vector-host relationships and elucidates the epidemiological patterns of leishmaniasis transmission. Blood meal analysis studies are essential for estimating the efficiency of pathogen transmission, assessing the relative human disease risk, and assist in identifying the other potential hosts of leishmaniasis. In Sudan and most of East Africa, there are large remaining gaps in knowledge regarding the feeding habits of phlebotomine vectors. The study aimed to identify the blood meal sources and, therefore, the host preferences of the principal vectors Phlebotomus orientalis and Ph. papatasi in leishmaniasis endemic areas of eastern and central Sudan.Materials and methodsSand flies were collected from two endemic villages in eastern and central Sudan using CDC light traps and sticky traps. The phlebotomine sand flies were morphologically and then molecularly identified. The source of blood meal of the engorged females was determined using a multiplex PCR methodology and specific primers of cytochrome b gene of mitochondrial DNA for human, goat, cow, and dog. The detection of the Leishmania parasite was done using PCR.ResultsThe total number of collected female phlebotomine sand flies was 180. Morphological identification revealed the abundance of Ph. orientalis 103 (57.2%), Ph. papatasi 42 (23.3%), Ph. bergeroti 31 (17.2%), Ph. rodhaini 2 (1.1%) and Ph. duboscqi 2 (1.1%) in the study sites. Out of the 180 collected, 31 (17%) were blood-fed flies. Three species were blood-fed and molecularly identified: Ph. papatasi (N = 7, 22.6%), Ph. bergeroti (N = 9, 26%), and Ph. orientalis (N = 15, 48.4%). Blood meal analysis revealed human DNA in two Ph. orientalis (6.4%), hence, the anthropophilic index was 13.3%.ConclusionsMultiplex PCR protocol described here allowed the identification of blood meal sources of many vertebrate species simultaneously. The results indicate that wild-caught Ph. orientalis are anthropophilic in the study areas. Further studies on larger blood-fed sample size are required to validate the potential applications of this technique in designing, monitoring and evaluating control programs, particularly in investigating the potential non-human hosts of leishmaniasis.

Prediction of malaria transmission drivers in Anopheles mosquitoes using artificial intelligence coupled to MALDI-TOF mass spectrometry

Vector control programmes are a strategic priority in the fight against malaria. However, vector control interventions require rigorous monitoring. Entomological tools for characterizing malaria transmission drivers are limited and are difficult to establish in the field. To predict Anopheles drivers of malaria transmission, such as mosquito age, blood feeding and Plasmodium infection, we evaluated artificial neural networks (ANNs) coupled to matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (MS) and analysed the impact on the proteome of laboratory-reared Anopheles stephensi mosquitoes. ANNs were sensitive to Anopheles proteome changes and specifically recognized spectral patterns associated with mosquito age (0–10 days, 11–20 days and 21–28 days), blood feeding and P. berghei infection, with best prediction accuracies of 73%, 89% and 78%, respectively. This study illustrates that MALDI-TOF MS coupled to ANNs can be used to predict entomological drivers of malaria transmission, providing potential new tools for vector control. Future studies must assess the field validity of this new approach in wild-caught adult Anopheles. A similar approach could be envisaged for the identification of blood meal source and the detection of insecticide resistance in Anopheles and to other arthropods and pathogens.

Molecular identification of blood meal sources in black flies (Diptera: Simuliidae) suspected as leucocytozoon vectors

Identification of the host blood meal of hematophagous insects can provide significant information regarding host preferences and the possibility of the transmission of disease agents. Currently, this knowledge is limited for black flies in the Oriental region. In this study, we used cytochrome b gene sequences for identification of blood meal sources of two black fly taxa, the Simulium asakoae complex and S. chumpornense Takaoka & Kuvangkadilok in Thailand. A total of 4,260 wild adult females were visually screened revealing 24 blood-engorged females from which cytochrome b was successfully amplified in 19 individuals. Comparisons in GenBank database revealed that all are closest to chicken (Gallus gallus) with sequence similarity of >98%. Therefore, these black fly species are feeding on chickens. We also molecularly investigated the hemosporidian blood protozoa genus Leucocytozoon in black flies and found 13 of 19 blood-engorged females positive for this protozoon. Sequence analysis revealed that this Leucocytozoon DNA could be assigned into two previously recognized groups, one with a Leucocytozoon reported from domestic chickens and black flies in Thailand, and another close to the L. schoutedeni. Our results indicate a high possibility that the S. asakoae complex and S. chumpornense are natural vectors of Leucocytozoon.

Identification of bloodmeal sources of triatomines captured in the Paraguayan Chaco region of South America by means of molecular biology analysis

ABSTRACTThe Paraguayan Chaco is an isolated environment with its own unique ecosystem. In this region, Chagas disease remains a health problem. Chagas disease is caused by the parasite Trypanosoma cruzi, and it is primarily transmitted by triatomines. In order to identify the blood meal sources of triatomines, specimens of the vector were collected in domestic and peridomestic areas and the PCR-RFLP method was implemented. Cytochrome b was amplified from the samples and later subjected to digestion with two restriction enzymes: Hae III and Xho I.It was possible to generate distinct restriction patterns on the amplified material to identify several blood meal sources for the vectors. We employed the blood from several species as positive controls: human, chicken, canine, feline, and armadillo blood. However, we identified only 3 sources for the blood meals of the insect vectors: human, chicken and canine blood. In total, 76 triatomines were captured. T. cruzi was not found in any of them. In 61% of the captured specimens, the blood meal sources for the vectors could be identified. In 30% of these cases, the presence of DNA from more than one vertebrate was detected in the same triatomine. The most common blood meal source found was chicken blood. The presence of human and chicken blood in triatomines captured in domestic and peridomestic areas strongly suggests that the parasite can freely move amongst both areas regardless of food availability. Free vector movement in these areas constitutes an epidemiological threat for the inhabitants of the community under study.