1 Towards environmental detection of Chagas disease vectors and pathogen 2 3 Grace Gysin 1 , Plutarco Urbano 2,3 , Luke Brandner-Garrod 1 , Shahida Begum 1 , Mojca Kristan 1 , 4 Thomas Walker 1 , Carolina Hernández 3,4 , Juan David Ramírez 2,5 , Louisa A. Messenger 1 5 6 1 Department of Disease Control, Faculty of Infectious and Tropical Diseases, London School 7 of Hygiene and Tropical Medicine, London, United Kingdom 8 9 2 Centro de Investigaciones en Microbiología y Biotecnología-UR (CIMBIUR), Facultad de 10 Ciencias Naturales, Universidad del Rosario, Bogotá, Colombia 11 12 3 Grupo de Investigaciones Biológicas de la Orinoquia, Universidad Internacional del Trópico 13 Americano (Unitrópico), Yopal, Colombia 14 15 4 Centro de Tecnología en Salud (CETESA), Innovaseq SAS, Bogotá, Colombia 16 17 5 Microbiology Division, Department of Pathology, Molecular and Cell-Based Medicine, 18 Icahn School of Medicine at Mount Sinai, New York, NY, USA 19 20 Abstract 21 22 Background 23 Accurate surveillance of triatomine household infestation is crucial for Chagas disease vector 24 control. However, no gold standard detection method with high levels of sensitivity or 25 specificity is currently available. Several intrinsic features of triatomine bug behaviour and 26 the lifecycle of Trypanosoma (T.) cruzi lead to deposition of environmental DNA (eDNA) in 27 infested houses. This study evaluated the use of FTA cards and cotton-tipped swabs as low- 28 technology, cost-effective tools for simultaneous detection of T. cruzi and vector eDNA in 29 the laboratory and field. 30 31 Methods/Principal Findings 32 This study had two components: (1) laboratory evaluation and optimisation of QIAcard® 33 FTA® classic cards to detect Rhodnius (R.) prolixus eDNA by altering five different 34 environmental variables (darkness, triatomine number, temperature, feeding status and 35 degradation at ambient temperature); (2) detection of R. prolixus and T. cruzi eDNA from 36 cotton-tipped house wall swabs from an endemic region in Casanare Department, Colombia. 37 eDNA was extracted from all specimens and amplified using a multiplex TaqMan qPCR 38 assay targeting the R. prolixus 12S rRNA gene and T. cruzi satellite DNA region. R. prolixus 39 eDNA from five 3 rd /4 th instar nymphs was successfully amplified from FTA cards after as 40 little as 15 minutes of contact time under standard insectary conditions. Factors significantly 41 increasing eDNA detection from FTA cards were increasing temperature from 21 o C to 27- 42 32 o C, triatomine bug density from 1-25 bugs and recent blood-feeding. eDNA was detectable 43 from FTA cards stored at room temperature for at least two weeks. In cotton-tipped swabs 44 from the field, the sensitivity and specificity of R. prolixus eDNA detection was 60.6% 45 (n=20/33) and 100% (n=33/33), respectively. T. cruzi eDNA was amplified from 93.9% 46 (n=31/33) of infested houses. 47 48 Conclusions/Significance 49 FTA cards are a highly sensitive tool for entomological surveillance of R. prolixus and 50 exhibit little variability under different environmental conditions. Additionally, cotton-tipped . CC-BY 4.0 International license It is made available under a perpetuity. is the author/funder, who has granted medRxiv a license to display the preprint in (which was not certified by peer review) preprint The copyright holder for this this version posted December 27, 2021. ; https://doi.org/10.1101/2021.12.24.21268369 doi: medRxiv preprint NOTE: This preprint reports new research that has not been certified by peer review and should not be used to guide clinical practice.
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