• 1.

    World Health Organization, 2015. Global Technical Strategy for Malaria 2016–2030. Geneva, Switzerland: WHO, 135.

  • 2.

    Service M, 2012. Medical Entomology for Students, Vol. 90. Cambridge, England: Cambridge University Press.

  • 3.

    Lounibos LP, 2002. Invasions by insect vectors of human disease. Annu Rev Entomol 47: 233266.

  • 4.

    Coetzee M, Craig M, Sueur D, 2000. Distribution of African malaria mosquitoes belonging to the Anopheles gambiae complex. Parasitol Today 16: 19982001.

    • Search Google Scholar
    • Export Citation
  • 5.

    Dhimal M, Ahrens B, Kuch U, 2014. Species composition, seasonal occurrence, habitat preference and altitudinal distribution of malaria and other disease vectors in eastern Nepal. Parasit Vectors 7: 540.

    • Search Google Scholar
    • Export Citation
  • 6.

    Logue K, Keven JB, Cannon MV, Reimer L, Siba P, Walker ED, Zimmerman PA, Serre D, 2016. Unbiased characterization of Anopheles mosquito blood meals by targeted high-throughput sequencing. PLoS Negl Trop Dis 10: 118.

    • Search Google Scholar
    • Export Citation
  • 7.

    Das S, Muleba M, Stevenson JC, Pringle JC, Norris DE, 2017. Beyond the entomological inoculation rate: characterizing multiple blood feeding behavior and Plasmodium falciparum multiplicity of infection in Anopheles mosquitoes in northern Zambia. Parasit Vectors 10: 45.

    • Search Google Scholar
    • Export Citation
  • 8.

    Githeko AK et al. 1996. Some observations on the biting behavior of Anopheles gambiae s.s., Anopheles arabiensis, and Anopheles funestus and their implications for malaria control. Exp Parasitol 82: 306315.

    • Search Google Scholar
    • Export Citation
  • 9.

    Keven JB, Reimer L, Katusele M, Koimbu G, Vinit R, Vincent N, Thomsen E, Foran DR, Zimmerman PA, Walker ED, 2017. Plasticity of host selection by malaria vectors of Papua New Guinea. Parasit Vectors 10: 95.

    • Search Google Scholar
    • Export Citation
  • 10.

    Hemingway J et al. 2016. Averting a malaria disaster: will insecticide resistance derail malaria control? Lancet 387: 17851788.

  • 11.

    Coleman M, Hemingway J, Gleave KA, Wiebe A, Gething PW, Moyes CL, 2017. Developing global maps of insecticide resistance risk to improve vector control. Malar J 16: 86.

    • Search Google Scholar
    • Export Citation
  • 12.

    Ranson H, Lissenden N, 2016. Insecticide resistance in African Anopheles mosquitoes: a worsening situation that needs urgent action to maintain malaria control. Trends Parasitol 32: 187196.

    • Search Google Scholar
    • Export Citation
  • 13.

    Cooke MK et al. 2015. ‘A bite before bed’: exposure to malaria vectors outside the times of net use in the highlands of western Kenya. Malar J 14: 259.

    • Search Google Scholar
    • Export Citation
  • 14.

    Lindsay SW, Emerson PM, Charlwood JD, 2002. Reducing malaria by mosquito-proofing houses. Trends Parasitol 18: 510514.

  • 15.

    Glunt KD, Abílio AP, Bassat Q, Bulo H, Gilbert AE, Huijben S, Manaca MN, Macete E, Alonso P, Paaijmans KP, 2015. Long-lasting insecticidal nets no longer effectively kill the highly resistant Anopheles funestus of southern Mozambique. Malar J 14: 17.

    • Search Google Scholar
    • Export Citation
  • 16.

    Saul A, 2003. Zooprophylaxis or zoopotentiation: the outcome of introducing animals on vector transmission is highly dependent on the mosquito mortality while searching. Malar J 2: 118.

    • Search Google Scholar
    • Export Citation
  • 17.

    Burkot TR, Russell TL, Reimer LJ, Bugoro H, Beebe NW, Cooper RD, Sukawati S, Collins FH, Lobo NF, 2013. Barrier screens: a method to sample blood-fed and host-seeking exophilic mosquitoes. Malar J 12: 19.

    • Search Google Scholar
    • Export Citation
  • 18.

    Grjébine A, 1966. Faune de Madagascar: Insectes Diptères Culicidae Anophelinae. XXII. Paris, France: Centre National de la Recherche Scientifique, Office de la Recherche Scientifique et Technique Outre-Mer. Available at: https://books.google.mg/books?id=TKsEDQEACAAJ.

    • Search Google Scholar
    • Export Citation
  • 19.

    Gillies MT, De Meillon B, 1968. The Anophelinae of Africa South of the Sahara: Ethiopian Zoogeographical Region. Johannesburg, South Africa: South African Institute for Medical Research. Available at: https://books.google.com/books?id=GfF6QgAACAAJ.

    • Search Google Scholar
    • Export Citation
  • 20.

    Gillies MT, Coetzee M, 1987. A Supplement to the Anophelinae of Africa South of the Sahara (Afrotropical Region). Johannesburg, South Africa: South African Institute for Medical Research. Available at: https://books.google.com/books?id=LchhAAAACAAJ.

    • Search Google Scholar
    • Export Citation
  • 21.

    Huber JT, 1998. The importance of voucher specimens, with practical guidelines for preserving specimens of the major invertebrate phyla for identification. J Nat Hist 32: 367385.

    • Search Google Scholar
    • Export Citation
  • 22.

    Koekemoer LL, Kamau L, Hunt RH, Coetzee M, 2002. A cocktail polymerase chain reaction assay to identify members of the Anopheles funestus (Diptera: Culicidae) group. Am J Trop Med Hyg 66: 804811.

    • Search Google Scholar
    • Export Citation
  • 23.

    Choi KS, Coetzee M, Koekemoer LL, 2010. Simultaneous identification of the Anopheles funestus group and Anopheles longipalpis type C by PCR-RFLP. Malar J 9: 316.

    • Search Google Scholar
    • Export Citation
  • 24.

    Joshi D, Park MH, Saeung A, Choochote W, Min GS, 2010. Multiplex assay to identify Korean vectors of malaria. Mol Ecol Resour 10: 748750.

  • 25.

    Bass C, Williamson MS, Field LM, 2008. Development of a multiplex real-time PCR assay for identification of members of the Anopheles gambiae species complex. Acta Trop 107: 5053.

    • Search Google Scholar
    • Export Citation
  • 26.

    Vezenegho SB, Bass C, Puinean M, Williamson MS, Field LM, Coetzee M, Koekemoer LL, 2009. Development of multiplex real-time PCR assays for identification of members of the Anopheles funestus species group. Malar J 8: 19.

    • Search Google Scholar
    • Export Citation
  • 27.

    Van De Vossenberg BT, Ibáñez-Justicia A, Metz-Verschure E, Van Veen EJ, Bruil-Dieters ML, Scholte EJ, 2015. Real-time PCR tests in Dutch exotic mosquito surveys; implementation of Aedes aegypti and Aedes albopictus identification tests, and the development of tests for the identification of Aedes atropalpus and Aedes japonicus japonicus (Diptera: Culicidae). J Med Entomol 52: 336350.

    • Search Google Scholar
    • Export Citation
  • 28.

    Henry-Halldin CN et al. 2011. High throughput multiplex assay for species identification of Papua New Guinea malaria vectors: members of the Anopheles punctulatus (Diptera: Culicidae) species group. Am J Trop Med Hyg 84: 166173.

    • Search Google Scholar
    • Export Citation
  • 29.

    Henry-Halldin CN et al. 2012. Multiplex assay for species identification and monitoring of insecticide resistance in Anopheles punctulatus group populations of Papua New Guinea. Am J Trop Med Hyg 86: 140151.

    • Search Google Scholar
    • Export Citation
  • 30.

    Kent RJ, 2009. Molecular methods for arthropod bloodmeal identification and applications to ecological and vector-borne disease studies. Mol Ecol Resour 9: 418.

    • Search Google Scholar
    • Export Citation
  • 31.

    McNamara DT, Kasehagen LJ, Grimberg BT, Cole-Tobian J, Collins WE, Zimmerman PA, 2006. Diagnosing infection levels of four human malaria parasite species by a polymerase chain reaction/ligase detection reaction fluorescent microsphere-based assay. Am J Trop Med Hyg 74: 413421.

    • Search Google Scholar
    • Export Citation
  • 32.

    Hillis DM, Dixon MT, 1991. Ribosomal DNA: molecular evolution and phylogenetic inference. Q Rev Biol 66: 411453.

  • 33.

    Beebe NW, Saul A, 1995. Discrimination of all members of the Anopheles punctulatus complex by polymerase chain reaction-restriction fragment length polymorphism analysis. Am J Trop Med Hyg 53: 478481.

    • Search Google Scholar
    • Export Citation
  • 34.

    Beebe NW, Whelan PI, Van Den Hurk AF, Ritchie SA, Corcoran S, Cooper RD, 2007. A polymerase chain reaction-based diagnostic to identify larvae and eggs of container mosquito species from the Australian region. J Med Entomol 44: 376380.

    • Search Google Scholar
    • Export Citation
  • 35.

    Garros C, Koekemoer LL, Kamau L, Awolola TS, Van Bortel W, Coetzee M, Coosemans M, Manguin S, 2004. Restriction fragment length polymorphism method for the identification of major African and Asian malaria vectors within the Anopheles funestus and An. minimus groups. Am J Trop Med Hyg 70: 260265.

    • Search Google Scholar
    • Export Citation
  • 36.

    Weeraratne TC, Surendran SN, Reimer LJ, Wondji CS, Perera MDB, Walton C, Karunaratne SHPP, 2017. Molecular characterization of Anopheline (Diptera: Culicidae) mosquitoes from eight geographical locations of Sri Lanka. Malar J 16: 234.

    • Search Google Scholar
    • Export Citation
  • 37.

    Toma T, Miyagi I, Crabtree MB, Miller BR, 2000. Identification of Culex vishnui subgroup (Diptera: Culicidae) mosquitoes from the Ryukyu Archipelago, Japan: development of a species-diagnostic polymerase chain reaction assay based on sequence variation in ribosomal DNA spacers. J Med Entomol 37: 554558.

    • Search Google Scholar
    • Export Citation
  • 38.

    Batovska J, Blacket MJ, Brown K, Lynch SE, 2016. Molecular identification of mosquitoes (Diptera: Culicidae) in southeastern Australia. Ecol Evol 6: 30013011.

    • Search Google Scholar
    • Export Citation
  • 39.

    Li J, Wirtz RA, McConkey GA, Sattabongkot J, Waters AP, Rogers MJ, McCutchan TF, 1995. Plasmodium: genus-conserved primers for species identification and quantitation. Exp Parasitol 81: 182190.

    • Search Google Scholar
    • Export Citation
  • 40.

    Howes RE et al. 2018. Risk factors for malaria in central Madagascar: insights from a cross-sectional population survey. Am J Trop Med Hyg 99: 9951002.

    • Search Google Scholar
    • Export Citation
  • 41.

    R Core Team, 2017. R: A Language and Environment for Statistical Computing. Vienna, Austria: R Foundation for Statistical Computing.

  • 42.

    Wickham H, 2017. Tidyverse: Easily Install and Load the “Tidyverse”. Vienna, Austria: R Foundation for Statistical Computing.

  • 43.

    McNamara DT, Kasehagen LJ, Grimberg BT, Cole-Tobian J, Collins WE, Zimmerman PA, 2005. Diagnosing infection levels of four human malaria parasite species by a PCR/LDR fluorescent microsphere-based assay. Am J Trop Med Hyg 73: 2021.

    • Search Google Scholar
    • Export Citation
  • 44.

    Gaffigan T, Pecor J, 1997. Collecting, Rearing, Mounting and Shipping Mosquitoes. Walter Reed Biosystematics Unit, Suitland, MD, 18.

  • 45.

    Reeves LE, Holderman CJ, Gillett-kaufman JL, Kawahara AY, Kaufman PE, 2016. Maintenance of host DNA integrity in field-preserved mosquito (Diptera: Culicidae) blood meals for identification by DNA barcoding. Parasit Vectors 9: 503.

    • Search Google Scholar
    • Export Citation
  • 46.

    Müller P, Pflüger V, Wittwer M, Ziegler D, Chandre F, Simard F, Lengeler C, 2013. Identification of cryptic Anopheles mosquito species by molecular protein profiling. PLoS One 8: e57486.

    • Search Google Scholar
    • Export Citation
  • 47.

    Schaffner F, Kaufmann C, Pflüger V, Mathis A, 2014. Rapid protein profiling facilitates surveillance of invasive mosquito species. Parasit Vectors 7: 142.

    • Search Google Scholar
    • Export Citation
  • 48.

    Niare S, Almeras L, Tandina F, Yssouf A, Bacar A, Toilibou A, Doumbo O, Raoult D, Parola P, 2017. MALDI-TOF MS identification of Anopheles gambiae giles blood meal crushed on Whatman filter papers. PLoS One 12: e0183238.

    • Search Google Scholar
    • Export Citation
  • 49.

    Laroche M, Almeras L, Pecchi E, Bechah Y, Raoult D, Viola A, Parola P, 2017. MALDI-TOF MS as an innovative tool for detection of Plasmodium parasites in Anopheles mosquitoes. Malar J 16: 5.

    • Search Google Scholar
    • Export Citation
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

 

 

 

A Novel Assay for Simultaneous Assessment of Mammalian Host Blood, Mosquito Species, and Plasmodium spp. in the Medically Important Anopheles Mosquitoes of Madagascar

View More View Less
  • 1 The Center for Global Health and Diseases, Case Western Reserve University, Cleveland, Ohio;
  • | 2 Department of Biology, Case Western Reserve University, Cleveland, Ohio;
  • | 3 Direction de Lutte contre le Paludisme/National Malaria Control Program Madagascar, Antananarivo, Madagascar;
  • | 4 Department of Microbiology and Molecular Genetics, Michigan State University, East Lansing, Michigan;
  • | 5 Faculty of Medicine and Faculty of Sciences, University of Antananarivo, Antananarivo, Madagascar
Restricted access

Anopheles mosquitoes vary in habitat preference, feeding pattern, and susceptibility to various measures of vector control. Consequently, it is important that we identify reservoirs of disease, identify vectors, and characterize feeding patterns to effectively implement targeted control measures. Using 467 anopheline mosquito abdomen squashes captured in Madagascar, we designed a novel ligase detection reaction and fluorescent microsphere assay, dubbed Bloodmeal Detection Assay for Regional Transmission (BLOODART), to query the bloodmeal content, identify five Anopheles mosquito species, and detect Plasmodium infection. Validation of mammalian bloodspots was achieved by preparation and analysis of known hosts (singular and mixed), sensitivity to degradation and storage method were assessed through mosquito feeding experiments, and quantification was explored by altering ratios of two mammal hosts. BLOODART identifications were validated by comparison with mosquito samples identified by sequenced portions of the internal transcribed spacer 2. BLOODART identification of control mammal bloodspots was 100% concordant for singular and mixed mammalian blood. BLOODART was able to detect hosts up to 42 hours after digestion when mosquito samples were stored in ethanol. A mammalian host was identified in every field-collected, blood-fed female Anopheles mosquito by BLOODART. The predominant mosquito host was cow (n = 451), followed by pig (n = 26) and human (n = 25). Mixed species bloodmeals were commonly observed (n = 33). A BLOODART molecular identification was successful for 318/467 mosquitoes, with an overall concordance of 60% with all field-captured, morphologically identified Anopheles specimens. BLOODART enables characterization of large samples and simultaneous pathogen detection to monitor and incriminate disease vectors in Madagascar.

Author Notes

Address correspondence to Peter A. Zimmerman, The Center for Global Health and Diseases, Case Western Reserve University, Biomedical Research Building 4th Floor, 10900 Euclid Ave. LC: 4983, Cleveland, OH 44106. E-mail: paz@case.edu

Financial support: Stipend support for R. E. T. was provided by the U.S. Navy Health Services Collegiate Program. Additional support was provided to P. A. Z. through the CWRU School of Medicine.

Ethics approval and consent to participate: These epidemiological surveys are routinely performed by the Madagascar NMCP and are consistent with protocols approved by the Madagascar Ministry of Health (No. 099-MSANP/CE). In addition, community and household approvals were obtained following fokontany-based meetings before initiating all study activities.

Authors’ addresses: Riley E. Tedrow and Peter A. Zimmerman, The Center for Global Health and Diseases, Case Western Reserve University, Cleveland, OH, E-mails: ret31@case.edu and paz@case.edu. Jocelyn Ratovonjato and Arsene Ratsimbasoa, Direction de Luttecontre le Paludisme/National Malaria Control Program Madagascar, Ministry of Health, Antananarivo, Madagascar, E-mails: njatovo_joc@yahoo.fr and aratsimbasoa@gmail.com. Edward D. Walker, Department of Microbiology and Molecular Genetics, Michigan State University, East Lansing, MI, E-mail: walker@msu.edu.

Save