A two-step metagenomics approach for prey identification from the blood meals of common vampire bats (Desmodus rotundus)

ABSTRACTThe feeding behaviour of the sanguivorous common vampire bat (Desmodus rotundus) facilitates the transmission of pathogens that can impact both human and animal health. To formulate effective strategies in controlling the spread of diseases, there is a need to obtain information on which animals they feed on. One DNA-based approach, shotgun sequencing, can be used to obtain such information. Even though it is costly, shotgun sequencing can be used to simultaneously retrieve prey and vampire bat mitochondrial DNA for population studies within one round of sequencing. However, due to the challenges of analysing shotgun sequenced metagenomic data such as false negatives/positives and typically low proportion of reads mapped to diet items, shotgun sequencing has not been used for the identification of prey from common vampire bat blood meals. To overcome these challenges and generate longer mitochondrial contigs which could be useful for prey population studies, we shotgun sequenced common vampire bat blood meal samples (n=8) and utilised a two-step metagenomic approach based on combining existing bioinformatic workflows (alignment and de novo mtDNA assembly) to identify prey. Further, we validated our results to detections made through metabarcoding. We accurately identified the common vampire bats’ prey in seven out of eight samples without any false positives. We also generated prey mitochondrial contig lengths between 138bp to 3231bp (mean=985bp, SD=981bp). As we develop more computationally efficient bioinformatics pipelines and reduce sequencing costs, we can expect an uptake in metagenomics dietary studies in the near future.

Survey of Mississippi Mosquito Blood Meals for Vertebrate Host Identification

ABSTRACT Exploring particular mosquito and vertebrate relationships provide insight to potential transmission of several agents of disease. In the current study, the relationship between white-tailed deer (Odocoileus virginianus) and mosquitoes was explored by identifying blood meals within mosquitoes captured throughout Mississippi between June and September of 2013 and 2017. We captured 72 bloodfed mosquitoes between 2 collection years, with a majority of specimens identified as Culex erraticus or Psorophora mathesoni. Seventy-nine percent (26/33) of blood meals in Cx. erraticus originated from the white-tailed deer. These findings implicate mosquitoes may primarily be feeding on white-tailed deer in rural areas of Mississippi.

The connection between Trypanosoma cruzi transmission cycles by Triatoma brasiliensis brasiliensis: A threat to human health in an area susceptible to desertification in the Seridó, Rio Grande do Norte, Brazil

An outbreak of Chagas disease, possibly involving its vector Triatoma brasiliensis brasiliensis, was identified in the state of Rio Grande do Norte (RN). Given the historical significance of this vector in public health, the study aimed to evaluate its role in the transmission dynamics of the protozoan Trypanosoma cruzi in an area undergoing desertification in the Seridó region, RN, Brazil. We captured triatomines in sylvatic and anthropic ecotopes. Natural vector infection was determined using parasitological and molecular methods and we identified discrete typing units (DTUs) of T. cruzi by analyzing the COII gene of mtDNA, 24Sα rDNA, and mini-exon gene. Their blood meals sources were identified by amplification and sequencing of the mtDNA cytochrome b gene. A total of 952 T. b. brasiliensis were captured in peridomestic (69.9%) and sylvatic ecotopes (30.4%). A wide range of natural infection rates were observed in peridomestic (36.0% - 71.1%) and sylvatic populations (28.6% - 100.0%). We observed the circulation of TcI and TcII DTUs with a predominance of Tcl in sylvatic and peridomestic environments. Kerodon rupestris, rocky cavy (13/39), Homo sapiens, human (8/39), and Bos taurus, ox (6/39) were the most frequently detected blood meals sources. Thus, Triatoma b. brasiliensis is invading and colonizing the human dwellings. Furthermore, high levels of natural infection, coupled with the detection of TcI and TcII DTUs, and also the detection of K. rupestris and H. sapiens as blood meals sources of infected T. b. brasiliensis indicate a risk of T. cruzi transmission to human populations in areas undergoing desertification.

Plasmodium metabolite HMBPP stimulates feeding of main mosquito vectors on blood and artificial toxic sources

Recent data show that parasites manipulate the physiology of mosquitoes and human hosts to increase the probability of transmission. Here, we investigate phagostimulant activity of Plasmodium-metabolite, (E)-4-hydroxy-3-methyl-but-2-enyl pyrophosphate (HMBPP), in the primary vectors of multiple human diseases, Anopheles coluzzii, An. arabiensis, An. gambiae s.s., Aedes aegypti, and Culex pipiens/Culex torrentium complex species. The addition of 10 µM HMBPP to blood meals significantly increased feeding in all the species investigated. Moreover, HMBPP also exhibited a phagostimulant property in plant-based-artificial-feeding-solution made of beetroot juice adjusted to neutral pH similar to that of blood. The addition of AlbuMAXTM as a lipid/protein source significantly improved the feeding rate of An. gambiae s.l. females providing optimised plant-based-artificial-feeding-solution for delivery toxins to control vector populations. Among natural and synthetic toxins tested, only fipronil sulfone did not reduce feeding. Overall, the toxic-plant-based-artificial-feeding-solution showed potential as an effector in environmentally friendly vector-control strategies.

Biological Parameters of Two Triatoma protracta Subspecies (Hemiptera: Reduviidae)

In recent years, concerns about Chagas disease in the United States have increased. Triatomine bug (Hemiptera: Reduviidae) populations are the vectors of the parasite Trypanosoma cruzi Chagas (Trypanosomatida: Trypanosomatidae), which causes Chagas disease, although the route of transmission is considered inefficient in United States. However, more studies on triatomine feeding and defecation behavior are needed. In this study, six related biological parameters from two populations of Triatoma protracta protracta (Uhler) and T. p. woodi (Uhler) from Mexican locations near the U.S. border were evaluated. The four population life cycles were less than 6 mo (161–171 d), with 9–10 blood meals needed to molt. Mortality rates were similar (31–38%) among the four populations. Triatoma p. woodi from Hidalgo, Coahuila was the most aggressive one. Feeding times were over 10 min, increasing with instar in all populations. Defecation behaviors varied among populations. High percentages of male and female fourth- and fifth-instar nymphs of T. p. protracta from Imuris and both populations of T. p. woodi defecated immediately after or <1 min of feeding. Lower percentages were observed for T. p. protracta from Jacumé. Because most parameters were similar among the four populations, independent of their subspecies and their geographic origin, we considered that T. p. protracta and T. p. woodi are efficient vectors of T. cruzi. In contrast, defecation patterns were noticeably different among some of the four triatomine populations studied. Our results highlight the importance of studying the biological parameters of local triatomine populations. They also contribute to increasing the knowledge of North American triatomine behavior and defecation patterns.

Seasonal Prevalence and Host Preference of Some Medically Important Aedes Species of Doon Valley, India

In the recent past, several outbreaks of mosquito-borne diseases in Dehradun city make it necessary for a comprehensive understanding of man-vector contacts, disease transmission and determination of appropriate resting sites for adult mosquitoes. The study was conducted within the periphery of Dehradun city in Doon valley, Uttarakhand state, India consisting of eight (08) locations namely Sahaspur, Premnagar, Raipur, Karanpur, Sahastradhara, Dudhli, Doiwala and Rani Pokhri with the objective to analyze and assess the seasonal prevalence and host choice of the mosquitoes with special reference to Aedes species. Among the eight locations of the current study, Raipur registered the highest mosquito abundance (17%) trailed by Sahaspur (16%) and Karanpur (15%). A low abundance of mosquitoes was registered conjointly from Doiwala and Rani Pokhri areas (9% from each area). In these areas, the profile plots of the 2-way ANOVA displayed the highest variation for the data collected from human habitations followed by bovine sheds, whereas the lowest recorded variation was from the poultry. With regards to seasonal prevalence, the highest mosquito abundance was registered during the monsoon followed by the post-monsoon. In provender analysis, a total of 1912 blood meals were collected for antibody test among which 1851 were found positive, with the highest reaction with human antisera (57.75%), followed by bovine antisera (21.34%) and pigs antisera (4.97%). Ae. aegyptihad the highest Anthropophilic Index (A.I.) with 68.78%, followed by Ae. albopictus (64.89%) and Ae. vittatus (55.55%). However, Human Blood Predominance (H.B.P.) and Pure Human Blood Pervasiveness (P.H.B.P.) were found highest for Ae. albopictus with values 0.43 and 0.44 respectively. The results also show the seasonal prevalence and feeding habits of these vectors. This study would be helpful in formulating season-wise strategies for the control of Aedes vector mosquitoes in different eco-epidemiological situations.

The Extrinsic Incubation Period of Zika Virus in Florida Mosquitoes Aedes aegypti and Ae. albopictus

The Asian genotype of Zika virus (ZIKV) emerged in Brazil in 2015 and subsequently spread throughout the Americas. In July 2016, Florida experienced its first locally acquired ZIKV infection in the continental U.S. Concerns about health risks from ZIKV infection have increased the need to investigate the interactions between potential mosquito vectors and ZIKV. The time it takes for an arbovirus to propagate within a mosquito, and become transmissible, is the extrinsic incubation period (EIP). The EIP for potential mosquito vectors in Florida is unknown. To address this gap in the understanding of ZIKV epidemiology, Florida Aedes aegypti (L.) and Ae. albopictus (Skuse) were orally exposed to ZIKV infected blood meals and fully engorged mosquitoes were held at a constant temperature of 28 °C through the duration of the experiment. Saliva expectorates were collected from cohorts of mosquitoes and tested for the presence of ZIKV at three-day intervals over a period of 24 days to allow for an evaluation of the EIP of the emergent Asian lineage of ZIKV. High rates of infected bodies in Ae. albopictus (75–94%) and Ae. aegypti (68–86%) were observed throughout the incubation period, which did not differ by species. Higher rates of disseminated infection were observed later during the incubation period but did not differ between species. We calculated the 50% EIP to be shorter in Ae. albopictus than Ae. aegypti (16.2 and 18.2 days post infection, respectively). The competence for ZIKV observed in both species may contribute to high rates of ZIKV transmission in Florida populations.

Nonrandom Selection and Multiple Blood Feeding of Human Hosts by Anopheles Vectors: Implications for Malaria Transmission in Papua New Guinea

Nonrandom selection and multiple blood feeding of human hosts by Anopheles mosquitoes may exacerbate malaria transmission. Both patterns of blood feeding and their relationship to malaria epidemiology were investigated in Anopheles vectors in Papua New Guinea (PNG). Blood samples from humans and mosquito blood meals were collected in villages and human genetic profiles (“fingerprints”) were analyzed by genotyping 23 microsatellites and a sex-specific marker. Frequency of blood meals acquired from different humans, identified by unique genetic profiles, was fitted to Poisson and negative binomial distributions to test for nonrandom patterns of host selection. Blood meals with more than one genetic profiles were classified as mosquitoes that fed on multiple humans. The age of a person bitten by a mosquito was determined by matching the blood-meal genetic profile to the villagers’ genetic profiles. Malaria infection in humans was determined by PCR test of blood samples. The results show nonrandom distribution of blood feeding among humans, with biased selection toward males and individuals aged 15–30 years. Prevalence of Plasmodium falciparum infection was higher in this age group, suggesting males in this age range could be super-spreaders of malaria parasites. The proportion of mosquitoes that fed on multiple humans ranged from 6% to 13% among villages. The patterns of host utilization observed here can amplify transmission and contribute to the persistence of malaria in PNG despite efforts to suppress it with insecticidal bed nets. Excessive feeding on males aged 15–30 years underscores the importance of targeted interventions focusing on this demographic group.