Hubert Benjamin, Marchi Muriel, Ly Vu Joseph, Tranchant Camille, Tarkowski Łukasz P, Leprince Olivier, Buitink Julia
Univ Angers, Institut Agro, INRAE, IRHS, SFR QUASAV, F-49000, Angers, France.
INRAE, Université de Strasbourg, UMR SVQV, Colmar, France.
Plant Methods. 2024 Jan 29;20(1):16. doi: 10.1186/s13007-024-01144-z.
One of the levers towards alternative solutions to pesticides is to improve seed defenses against pathogens, but a better understanding is needed on the type and regulation of existing pathways during germination. Dormant seeds are able to defend themselves against microorganisms during cycles of rehydration and dehydration in the soil. During imbibition, seeds leak copious amounts of exudates. Here, we developed a nephelometry method to assay antimicrobial activity (AA) in tomato seed exudates as a proxy to assess level of defenses.
A protocol is described to determine the level of AA against the nonhost filamentous fungus Alternaria brassicicola in the exudates of tomato seeds and seedlings. The fungal and exudate concentrations can be adjusted to modulate the assay sensitivity, thereby providing a large window of AA detection. We established that AA in dormant seeds depends on the genotype. It ranged from very strong AA to complete absence of AA, even after prolonged imbibition. AA depends also on the stages of germination and seedling emergence. Exudates from germinated seeds and seedlings showed very strong AA, while those from dormant seeds exhibited less activity for the same imbibition time. The exudate AA did not impact the growth of a pathogenic fungus host of tomato, Alternaria alternata, illustrating the adaptation of this fungus to its host.
We demonstrate that our nephelometry method is a simple yet powerful bioassay to quantify AA in seed exudates. Different developmental stages from dormant seed to seedlings show different levels of AA in the exudate that vary between genotypes, highlighting a genetic diversity x developmental stage interaction in defense. These findings will be important to identify molecules in the exudates conferring antifungal properties and obtain a better understanding of the regulatory and biosynthetic pathways through the lifecycle of seeds, from dormant seeds until seedling emergence.
寻求农药替代解决方案的一个途径是增强种子对病原体的防御能力，但需要更深入了解种子萌发过程中现有防御途径的类型和调控机制。休眠种子在土壤中的吸水和脱水循环过程中能够抵御微生物。在吸水过程中，种子会大量渗漏渗出物。在此，我们开发了一种比浊法来测定番茄种子渗出物中的抗菌活性（AA），以此作为评估防御水平的指标。
本文描述了一种用于测定番茄种子和幼苗渗出物对非寄主丝状真菌链格孢抗菌活性水平的方法。可以调整真菌和渗出物的浓度来调节检测灵敏度，从而提供一个较大的抗菌活性检测范围。我们发现休眠种子中的抗菌活性取决于基因型。其抗菌活性范围从非常强到完全没有，即使经过长时间吸水也是如此。抗菌活性还取决于种子萌发和幼苗出土的阶段。在相同的吸水时间下，萌发种子和幼苗的渗出物显示出非常强的抗菌活性，而休眠种子的渗出物活性较低。渗出物的抗菌活性对番茄致病真菌链格孢的生长没有影响，这表明该真菌对其寄主具有适应性。
我们证明，我们的比浊法是一种简单而强大的生物测定方法，可用于量化种子渗出物中的抗菌活性。从休眠种子到幼苗的不同发育阶段，渗出物中的抗菌活性水平不同，且因基因型而异，这突出了防御过程中遗传多样性与发育阶段的相互作用。这些发现对于鉴定渗出物中具有抗真菌特性的分子以及更好地理解种子从休眠到幼苗出土整个生命周期中的调控和生物合成途径具有重要意义。
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