title: Unbiased Characterization of Anopheles Mosquito Blood Meals by Targeted High-Throughput Sequencing.
abstract: Understanding mosquito host choice is important for assessing vector competence or identifying disease reservoirs. Unfortunately, the availability of an unbiased method for comprehensively evaluating the composition of insect blood meals is very limited, as most current molecular assays only test for the presence of a few pre-selected species. These approaches also have limited ability to identify the presence of multiple mammalian hosts in a single blood meal. Here, we describe a novel high-throughput sequencing method that enables analysis of 96 mosquitoes simultaneously and provides a comprehensive and quantitative perspective on the composition of each blood meal. We validated in silico that universal primers targeting the mammalian mitochondrial 16S ribosomal RNA genes (16S rRNA) should amplify more than 95% of the mammalian 16S rRNA sequences present in the NCBI nucleotide database. We applied this method to 442 female Anopheles punctulatus s. l. mosquitoes collected in Papua New Guinea (PNG). While human (52.9%), dog (15.8%) and pig (29.2%) were the most common hosts identified in our study, we also detected DNA from mice, one marsupial species and two bat species. Our analyses also revealed that 16.3% of the mosquitoes fed on more than one host. Analysis of the human mitochondrial hypervariable region I in 102 human blood meals showed that 5 (4.9%) of the mosquitoes unambiguously fed on more than one person. Overall, analysis of PNG mosquitoes illustrates the potential of this approach to identify unsuspected hosts and characterize mixed blood meals, and shows how this approach can be adapted to evaluate inter-individual variations among human blood meals. Furthermore, this approach can be applied to any disease-transmitting arthropod and can be easily customized to investigate non-mammalian host sources.
authors.affiliation: Genomic Medicine Institute, Cleveland Clinic, Cleveland, Ohio, United States of America.|Department of Biology, Case Western Reserve University, Cleveland, Ohio, United States of America.|Center for Global Health and Diseases, Case Western Reserve University, Cleveland, Ohio, United States of America.
authors.forename: Kyle
authors.initials: K
authors.lastname: Logue
authors.affiliation: Department of Microbiology and Molecular Genetics, Michigan State University, East Lansing, Michigan, United States of America.|Papua New Guinea Institute of Medical Research, Madang, Papua New Guinea.
authors.forename: John Bosco
authors.initials: JB
authors.lastname: Keven
authors.affiliation: Genomic Medicine Institute, Cleveland Clinic, Cleveland, Ohio, United States of America.
authors.forename: Matthew V
authors.initials: MV
authors.lastname: Cannon
authors.affiliation: Liverpool School of Tropical Medicine and Hygiene, Liverpool, United Kingdom.
authors.forename: Lisa
authors.initials: L
authors.lastname: Reimer
authors.affiliation: Papua New Guinea Institute of Medical Research, Goroka, Papua New Guinea.
authors.forename: Peter
authors.initials: P
authors.lastname: Siba
authors.affiliation: Department of Microbiology and Molecular Genetics, Michigan State University, East Lansing, Michigan, United States of America.
authors.forename: Edward D
authors.initials: ED
authors.lastname: Walker
authors.affiliation: Department of Biology, Case Western Reserve University, Cleveland, Ohio, United States of America.|Center for Global Health and Diseases, Case Western Reserve University, Cleveland, Ohio, United States of America.
authors.forename: Peter A
authors.initials: PA
authors.lastname: Zimmerman
authors.affiliation: Genomic Medicine Institute, Cleveland Clinic, Cleveland, Ohio, United States of America.
authors.forename: David
authors.initials: D
authors.lastname: Serre
mesh_terms: D000818:Animals; D000852:Anopheles / Q000502:physiology*; D016000:Cluster Analysis; D004275:DNA, Ribosomal / Q000737:chemistry / Q000235:genetics; D004772:Entomology / Q000379:methods*; D005247:Feeding Behavior; D005260:Female; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans; D062407:Meals; D008969:Molecular Sequence Data; D010219:Papua New Guinea; D010802:Phylogeny; D012336:RNA, Ribosomal, 16S / Q000235:genetics*; D017422:Sequence Analysis, DNA
chemical_list: D004275:DNA, Ribosomal; D012336:RNA, Ribosomal, 16S