Comparative transcriptome combined with morphophysiological analyses revealed the molecular mechanism underlying <i>Tetrahymena thermophila</i> predation-induced antiphage defense in <i>Aeromonas hydrophila</i>

Protozoan predation has been demonstrated to be a strong driving force for bacterial defence strategies in the environment. Our previous study demonstrated that <i>Aeromonas hydrophila</i> NJ-35, which evolved small-colony variants (SCVs), displayed various adaptive traits in response to <i>Tetrahymena thermophila</i> predation, such as enhanced phage resistance. However, the evolutionary mechanisms are largely unknown. In this study, we performed a genome- and transcriptome-wide analysis of the SCV1, representing one strain of the SCVs, for identification of the genes of mutation and altered expression underlying this phage resistance phenotype. Our study demonstrated that phage resistance caused by <i>T. thermophila</i> predation was due to the downregulation of a flagellar biosynthesis regulator, <i>flhF</i>, in SCV1. Interestingly, we confirmed that phage resistance in SCV1 was not straightforwardly attributable to the absence of flagella but to FlhF-mediated secretion of extracellular protein that hinders phage adsorption. This finding improves our understanding of the mechanisms by which <i>A. hydrophila</i> lowers the susceptibility to phage infection under predation pressure, and highlights an important contribution of bacterium–protozoan interactions in driving the adaptive evolution of pathogens in complex environments.原生动物捕食已被证实是驱动自然环境中细菌防御策略演化的强有力驱动因素。我们此前的研究表明，形成小菌落变异体（small-colony variants，SCVs）的嗜水气单胞菌（Aeromonas hydrophila）NJ-35菌株，在应对嗜热四膜虫（Tetrahymena thermophila）捕食时展现出多种适应性性状，例如增强的噬菌体抗性。然而，该过程背后的演化机制仍鲜为人知。本研究针对其中一株小菌落变异体SCV1开展了全基因组与全转录组分析，以期鉴定介导该噬菌体抗性表型的突变基因与差异表达基因。本研究证实，嗜热四膜虫捕食诱导的噬菌体抗性，源于SCV1中鞭毛生物合成调控因子flhF的下调表达。有趣的是，我们确认SCV1中的噬菌体抗性并非直接源于鞭毛缺失，而是通过FlhF介导的细胞外蛋白分泌阻碍噬菌体吸附所实现。这一发现加深了我们对嗜水气单胞菌在捕食压力下降低噬菌体感染易感性的机制的理解，并凸显了细菌-原生动物相互作用在驱动复杂环境中病原菌适应性进化方面的重要作用。