Rapid detection of West Nile and Dengue viruses from mosquito saliva by loop-mediated isothermal amplification and displaced probes

被引:0
|
作者
Kim, Dongmin [1 ]
DeBriere, Terry J. [2 ]
Eastmond, Bradley H. [1 ]
Alomar, Abdullah A. [1 ]
Yaren, Ozlem [3 ,4 ]
McCarter, Jacquelyn [3 ,4 ]
Bradley, Kevin M. [4 ]
Benner, Steven A. [3 ,4 ]
Alto, Barry W. [1 ]
Burkett-Cadena, Nathan D. [1 ]
机构
[1] Univ Florida, Florida Med Entomol Lab, Vero Beach, FL 32962 USA
[2] TrakitNow Inc, Columbia, SC USA
[3] Firebird Biomol Sci LLC, Alachua, FL 32615 USA
[4] Fdn Appl Mol Evolut, Alachua, FL 32615 USA
来源
PLOS ONE | 2024年 / 19卷 / 02期
关键词
EQUINE ENCEPHALITIS-VIRUS; ARBOVIRUS SURVEILLANCE; VECTOR COMPETENCE; AEDES-AEGYPTI; DIPTERA; LAMP; CULICIDAE; SUSCEPTIBILITY; IDENTIFICATION; TEMPERATURE;
D O I
10.1371/journal.pone.0298805
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
Arthropod-borne viruses are major causes of human and animal disease, especially in endemic low- and middle-income countries. Mosquito-borne pathogen surveillance is essential for risk assessment and vector control responses. Sentinel chicken serosurveillance (antibody testing) and mosquito pool screening (by RT-qPCR or virus isolation) are currently used to monitor arbovirus transmission, however substantial time lags of seroconversion and/or laborious mosquito identification and RNA extraction steps sacrifice their early warning value. As a consequence, timely vector control responses are compromised. Here, we report on development of a rapid arbovirus detection system whereby adding sucrose to reagents of loop-mediated isothermal amplification with displaced probes (DP-LAMP) elicits infectious mosquitoes to feed directly upon the reagent mix and expectorate viruses into the reagents during feeding. We demonstrate that RNA from pathogenic arboviruses (West Nile and Dengue viruses) transmitted in the infectious mosquito saliva was detectable rapidly (within 45 minutes) without RNA extraction. Sucrose stabilized viral RNA at field temperatures for at least 48 hours, important for transition of this system to practical use. After thermal treatment, the DP-LAMP could be reliably visualized by a simple optical image sensor to distinguish between positive and negative samples based on fluorescence intensity. Field application of this technology could fundamentally change conventional arbovirus surveillance methods by eliminating laborious RNA extraction steps, permitting arbovirus monitoring from additional sites, and substantially reducing time needed to detect circulating pathogens.
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页数:15
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