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Title: Evaluating RNAlater® as a preservative for using near-infrared spectroscopy to predict Anopheles gambiae age and species

Author
item SIKULU, MAGGY - Queensland Institute Of Medical Research
item DOWELL, KAYLA - Volunteer
item HUGO, LEON - Queensland Institute Of Medical Research
item WIRTZ, ROBERT - Centers For Disease Control And Prevention (CDC) - United States
item MICHEL, KRISTIN - Kansas State University
item PEIRIS, KAMARANGA H - Kansas State University
item MOORE, SARAH - Ifakara Health Institute
item KILLEEN, GERRY - Ifakara Health Institute
item Dowell, Floyd

Submitted to: Malaria Journal
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/8/2011
Publication Date: 7/8/2011
Citation: Sikulu, M., Dowell, K.M., Hugo, L.E., Wirtz, R.A., Michel, K., Peiris, K.S., Moore, S., Killeen, G.F., Dowell, F.E. 2011. Evaluating RNAlater® as a preservative for using near-infrared spectroscopy to predict Anopheles gambiae age and species. Malaria Journal. 10:186.

Interpretive Summary: Determining mosquito age and species is important to determine the effectiveness of vector control programs. Near-infrared spectroscopy (NIRS) has previously been applied to determine the age and species of freshly anesthetized mosquitoes that transmit malaria. However, this has only been achieved on freshly-collected specimens, and future applications will require samples to be preserved between field collections and scanning by NIRS. In this study, a sample preservation method (storage in RNAlater®) was evaluated for mosquito age and species identification by NIRS. The average accuracy obtained for predicting the age of young (<7 days) or old (greater than or equal to 7 days) of all fresh and all preserved mosquito samples was 83% and 90%, respectively. For species identification, accuracies were 82% for fresh against 80% for preserved. Thus, RNAlater® can be used to preserve mosquitoes for subsequent scanning and analysis by NIRS to determine their age and species with minimal costs and with accuracy similar to that achieved from fresh insects.

Technical Abstract: Mosquito age and species identification is a crucial determinant of the efficacy of vector control programs. Near-infrared spectroscopy (NIRS) has previously been applied successfully to rapidly, non-destructively, and simultaneously determine the age and species of freshly anesthetized African malaria vectors from the Anopheles gambiae s.l. species complex: An. gambiae s. s. and Anopheles arabiensis. However, this has only been achieved on freshly-collected specimens and future applications will require samples to be preserved between field collections and scanning by NIRS. In this study, a sample preservation method (RNAlater®) was evaluated for mosquito age and species identification by NIRS against scans of fresh samples. Two strains of An. gambiae s.s. (CDC and G3) and two strains of An. arabiensis (Dongola, KGB) were reared in the laboratory while the third strain of An. arabiensis (Ifakara) was reared in a semi-field system. All mosquitoes were scanned when fresh and rescanned after preservation in RNAlater® for several weeks. Age and species identification were determined using a cross-validation. The mean accuracy obtained for predicting the age of young (<7 days) or old (greater than or equal to 7 days) of all fresh (n=633) and all preserved (n=691) mosquito samples was 83% and 90%, respectively. For species identification, accuracies were 82% for fresh against 80% for RNAlater® preserved. For both analyses, preserving mosquitoes in RNAlater® was associated with a highly significant reduction in the likelihood of a misclassification of mosquitoes as young or old using NIRS. Important to note is that the cost for preserving mosquito specimens with RNAlater® ranges from 3-13 cents per insect depending on the size of the tube used and the number of specimens pooled in one tube. RNAlater® can be used to preserve mosquitoes for subsequent scanning and analysis by NIRS to determine their age and species with minimal costs and with accuracy similar to that achieved from fresh insects. Cold storage availability allows samples to be stored longer than a week after field collection. Further study to develop robust calibrations applicable to other strains from diverse ecological settings is recommended.