The feasibility of using autofluorescence to detect lignin deposition pattern during defense response in apple roots to Pythium ultimum infection

Authors: Zhu, Yanmin

Submitted to: Horticulturae
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 11/15/2022
Publication Date: 11/17/2022
Citation: Zhu, Y. 2022. The feasibility of using autofluorescence to detect lignin deposition pattern during defense response in apple roots to Pythium ultimum infection. Horticulturae. 8(11). Article 1085. https://doi.org/10.3390/horticulturae8111085.
DOI: https://doi.org/10.3390/horticulturae8111085

Interpretive Summary: In the post-genomics era, detailed and accurate phenotypic evaluation remains a major operational bottleneck for genetic studies of target traits, comparing to relatively accessible genomic data. To elucidate the molecular mechanisms underpinning apple root resistance to infection by Pythium ultimum, comprehensive transcriptome analysis has provided a wide-angled and high-resolution picture of the genome-wide transcriptional networks regulating the defense activation. On the other hand, evaluating cellular responses and anatomical changes remains challenging in young roots of apple as a woody species of Rosacea family. For example, our genomics data have strongly suggested that laccase-directed lignin formation is likely a crucial component of apple root resistance trait. Yet, little is known about the phenotypic features of lignin deposition or cell wall modification during defense activation to P. ultimum infection. Taking advantage of the autofluorescence from lignin in plant tissues and the specific lignin staining by Wiesner reagent, the feasibility of using microscopy to detect lignin deposition patterns in apple root tissue was investigated in this study. The preliminary results indicated that a combination of brightfield and fluorescence images may provide the opportunity to reveal the location and degree of lignification in sectioned roots. The current procedure represents a proof-of-concept of using fluorescence microscopy to examine cellular features relevant to apple root resistance traits. With further improvements, the standardized method should be able to distinguish features of cell wall lignification between rootstock genotypes, under various treatments and at different infection timepoints. The capability of detecting subtle phenotypic variation, such as lignin deposition patterns, is critically needed for bridging the gap between genomics data and macro-level plant resistance phenotypes. Therefore, the result from this study is crucial for further identifying the resistance mechanisms in apple root.

Technical Abstract: The molecular mechanisms underpinning apple root resistance to infection by Pythium ultimum have not been carefully investigated until recently. A comprehensive transcriptome analysis has generated a panoramic and high-resolution picture of the genome-wide transcriptional networks regulating the defense activation in apple roots. Among the most notable transcriptome changes are the upregulation of phenylpropanoid and flavonoid biosynthesis, transportation of secondary metabolites, and laccase-directed lignin formation. Key candidate genes were identified based on their differential expression at both transcriptional and post-transcriptional levels. On the other hand, assessing cellular responses and anatomical changes in young roots of apple as a woody species of Rosacea family remains challenging. Little is known about the phenotypic features of lignin deposition, cell wall modification, and accumulation of other phenolic compounds in apple roots during defense activation to P. ultimum infection. Taking advantage of the autofluorescence of lignin and the specific lignin staining by Wiesner reagent, the feasibility of using microscopy to detect lignin deposition in young apple roots was investigated. The preliminary results indicated that a combination of brightfield and fluorescence images provide the opportunity to reveal the location and degree of lignification in sectioned roots. The procedure represents a proof-of-concept of using fluorescence microscopy to examine cellular features relevant to apple root resistance traits. With further improvements to the procedure, the standardized method should be able to differentiate features of cell wall lignification between rootstock genotypes, under various treatments and at different infection timepoints. The ability to inspect subtle phenotypic variation, such as lignin deposition patterns, is critically needed for bridging the gap between genomic data and macro-level plant resistance phenotypes and further identifying the mechanisms of resistance in apple root.