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Malar J. 2019 Mar 22;18(1):95. doi: 10.1186/s12936-019-2723-0.

Application of matrix-assisted laser desorption/ionization mass spectrometry to identify species of Neotropical Anopheles vectors of malaria.

Author information

1
Centro de Biodiversidad y Descubrimiento de Drogas, Instituto de Investigaciones Científicas y Servicios de Alta Tecnología (INDICASAT AIP), City of Knowledge, Panama, 0843-01103, Republic of Panama.
2
Smithsonian Tropical Research Institute, Panama, Republic of Panama.
3
Programa Centroamericano de Maestría en Entomología, Universidad de Panamá, Panama, Republic of Panama.
4
College of Health Sciences, The University of Texas at El Paso, El Paso, TX, USA.
5
Grupo de Investigación en Biotecnología, Bioinformática y Biología de Sistemas, Centro de Producción e Investigaciones Agroindustriales, Universidad Tecnológica de Panamá, Panama, Republic of Panama.
6
Grupo de Investigación en Sistemas de Comunicaciones Digitales Avanzados, Facultad de Ingeniería Eléctrica, Universidad Tecnológica de Panamá, Panama, Republic of Panama.
7
ENSEIRB-MATMECA-Bordeaux INP, Talence, France.
8
Sam Noble Oklahoma Museum of Natural History and Department of Biology, University of Oklahoma, Norman, OK, USA.
9
Department of Biology, University of Massachusetts Boston, Boston, MA, USA.
10
Centro de Biodiversidad y Descubrimiento de Drogas, Instituto de Investigaciones Científicas y Servicios de Alta Tecnología (INDICASAT AIP), City of Knowledge, Panama, 0843-01103, Republic of Panama. rgittens@indicasat.org.pa.
11
Centro de Neurociencias, INDICASAT AIP, Panama, Republic of Panama. rgittens@indicasat.org.pa.

Abstract

BACKGROUND:

Malaria control in Panama is problematic due to the high diversity of morphologically similar Anopheles mosquito species, which makes identification of vectors of human Plasmodium challenging. Strategies by Panamanian health authorities to bring malaria under control targeting Anopheles vectors could be ineffective if they tackle a misidentified species.

METHODS:

A rapid mass spectrometry identification procedure was developed to accurately and timely sort out field-collected Neotropical Anopheles mosquitoes into vector and non-vector species. Matrix-assisted laser desorption/ionization (MALDI) mass spectra of highly-abundant proteins were generated from laboratory-reared mosquitoes using different extraction protocols, body parts, and sexes to minimize the amount of material from specimen vouchers needed and optimize the protocol for taxonomic identification. Subsequently, the mass spectra of field-collected Neotropical Anopheles mosquito species were classified using a combination of custom-made unsupervised (i.e., Principal component analysis-PCA) and supervised (i.e., Linear discriminant analysis-LDA) classification algorithms.

RESULTS:

Regardless of the protocol used or the mosquito species and sex, the legs contained the least intra-specific variability with enough well-preserved proteins to differentiate among distinct biological species, consistent with previous literature. After minimizing the amount of material needed from the voucher, one leg was enough to produce reliable spectra between specimens. Further, both PCA and LDA were able to classify up to 12 mosquito species, from different subgenera and seven geographically spread localities across Panama using mass spectra from one leg pair. LDA demonstrated high discriminatory power and consistency, with validation and cross-validation positive identification rates above 93% at the species level.

CONCLUSION:

The selected sample processing procedure can be used to identify field-collected Anopheles species, including vectors of Plasmodium, in a short period of time, with a minimal amount of tissue and without the need of an expert mosquito taxonomist. This strategy to analyse protein spectra overcomes the drawbacks of working without a reference library to classify unknown samples. Finally, this MALDI approach can aid ongoing malaria eradication efforts in Panama and other countries with large number of mosquito's species by improving vector surveillance in epidemic-prone sites such as indigenous Comarcas.

KEYWORDS:

Anopheles mosquito; MALDI; Malaria vector; Mass spectrometry; Panama; Taxonomic identification

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