1. Current mark–release–recapture methodologies are limited in their ability to address complex problems in vector biology, such as studying multiple groups overlapping in space and time. Additionally, limited mark retention, reduced post-marking survival and the large effort in marking, collection and recapture all complicate effective insect tracking.
2. We have developed and evaluated a marking method using a fluorescent dye (SmartWater^®) combined with synthetic DNA tags to informatively and efficiently mark adult mosquitoes using an airbrush pump and nebulizer. Using a handheld UV flashlight, the fluorescent marking enabled quick and simple initial detection of recaptures in a field-ready and non-destructive approach that when combined with an extraction-free PCR on individual mosquito legs provides potentially unlimited marking information.
3. This marking, first tested in the laboratory with Anopheles gambiae s.l. mosquitoes, did not affect survival (median ages 24–28 days, p-adj > 0.25), oviposition (median eggs/female of 28.8, 32.5, 33.3 for water, green, red dyes, respectively, p-adj > 0.44) or Plasmodium competence (mean oocysts 5.56–10.6, p-adj > 0.95). DNA and fluorescence had 100% retention up to 3 weeks (longest time point tested) with high intensity, indicating marks would persist longer.
4. We describe a novel, simple, no/low-impact and long-lasting marking method that allows separation of multiple insect subpopulations by combining unlimited length and sequence variation in the synthetic DNA tags. This method can be readily deployed in the field for marking multiple groups of mosquitoes or other insects.

中文翻译：

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一种新颖的荧光和 DNA 组合，用于对蚊子进行多功能、长期标记

1. 目前的标记-释放-重新捕获方法在解决载体生物学中的复杂问题（例如研究在空间和时间上重叠的多个群体）方面的能力有限。此外，有限的标记保留、标记后存活率的降低以及标记、收集和重新捕获方面的巨大努力都使有效的昆虫追踪变得复杂。
2. 我们开发并评估了一种标记方法，该方法使用荧光染料 (SmartWater ^® ) 与合成 DNA 标签相结合，使用喷枪泵和喷雾器对成年蚊子进行信息丰富且高效的标记。使用手持式紫外线手电筒，荧光标记能够以现场准备的非破坏性方法快速、简单地对重新捕获进行初步检测，当与单个蚊子腿上的免提取 PCR 相结合时，可以提供潜在的无限标记信息。
3. 该标记首次在实验室用冈比亚按蚊进行测试，不影响存活（中位年龄 24-28 天，p -adj > 0.25）、产卵（水、绿色、中位卵数/雌性为 28.8、32.5、33.3）红色染料，分别，p -adj > 0.44）或疟原虫能力（平均卵囊 5.56–10.6，p -adj > 0.95）。DNA 和荧光以高强度 100% 保留长达 3 周（测试的最长时间点），表明标记将持续更长时间。
4. 我们描述了一种新颖、简单、无/低影响且持久的标记方法，该方法允许通过结合合成 DNA 标签中的无限长度和序列变化来分离多个昆虫亚群。这种方法可以很容易地在现场部署，用于标记多组蚊子或其他昆虫。