World-wide increases in Vibrio-associated diseases have been reported in aquaculture and humans in co-occurrence with increased sea surface temperatures. Twelve species of Vibrio are known to cause disease in humans, but three species dominate the number of human infections world-wide: Vibrio cholerae, Vibrio parahaemolyticus, and Vibrio vulnificus. Fluorescent protein (FP)-labelled bacteria have been used to make great progress through in situ studies of bacterial behavior in mixed cultures or within host tissues. Currently, FP-labelling methods specific for Vibrio species are still limited by time-consuming counterselection measures that require the use of modified media and temperatures below the optimal growth temperature of many Vibrio species. Within this study, we used a previously reported R6K-based suicide delivery vector and two newly constructed transposon variants to develop a tailored protocol for FP-labelling V. cholerae, V. parahaemolyticus, and V. vulnificus environmental isolates within two days of counterselection against the donor Escherichiacoli. This herein presented protocol worked universally across all tested strains (30) with a conjugation efficiency of at least two transconjugants per 10,000 recipients.

中文翻译：

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开发一种通用且有效的方法来快速选择稳定的荧光蛋白标记病原性弧菌

据报道，在水产养殖和人类中，与弧菌相关的疾病在全球范围内均有增加，同时海面温度升高。已知有十二种弧菌会引起人类疾病，但在全世界范围内，三种感染占主导地位：霍乱弧菌，副溶血性弧菌和创伤弧菌。荧光蛋白（FP）标记的细菌已通过混合培养或宿主组织内细菌行为的原位研究取得了巨大进展。当前，特定于弧菌的FP标签方法该种仍然受到费时的反选择措施的限制，这些措施需要使用改良的培养基和低于许多弧菌物种最佳生长温度的温度。在这项研究中，我们使用了先前报道的基于R6K的自杀传递载体和两个新构建的转座子变体，以针对针对FP标记的霍乱弧菌，副溶血弧菌和V. vulnificus环境分离株开发了量身定制的方案，在两天内反选针对捐助大肠埃希氏大肠杆菌。本文提出的方案可在所有测试菌株（30）上通用，其结合效率为每10,000个接收者至少两种转导结合体。