Marco Sandri, Paola Zuccolotto
Sara Zenoni, Marianna Fasoli
Giovanni Battista Tornielli
The Molecular Phenology Scale (MPhS) is a tool aimed to map the ontogenetic development of the fruit.
The statistical pipeline developed to define the MPhS is an unsupervised learning procedure yielding an innovative combination of semiparametric, smoothing and dimensionality reduction tools. From a statistical perspective, the most noticeable features of the proposed method are that (1) time information is exploited by considering only the ordering of timepoints and not their distance and (2) the procedure is based on the extraction of Principal Components that have revealed exceptionally able to summarize different characteristics of data (berry variety, vintage, stage). From issue (1) follows that the MPhS measurement unit is not time, but an ideal step of the berry development process, which can take a longer or a shorter time, according to a multiplicity of different factors. From issue (2) follows that only genes directly involved with the berry development process can be selected, depurating from those impacted by the berry variety and the vintage effects.
The MPhS scale paves the way for the development of tools that aspire at predicting the phenological stage of the fruit in various climate conditions, such as models that can account for temperature and other environment clues. The quality of these tools will benefit from the combination of various modeling techniques (molecular, metabolite, physical, visual levels), providing that great coordination and knowledge-transfer between modelers, biologists and growers will be established.
So far, MPhS has been developed for grape berry, but it could be potentially extended and successfully applied to any other fruit species, provided that the available data, which are intended to be used to develop the scale, have the following characteristics:
MPhS R PackageGiovanni Battista Tornielli, Marco Sandri, Marianna Fasoli, Alessandra Amato, Mario Pezzotti, Paola Zuccolotto, Sara Zenoni (2023), A molecular phenology scale of grape berry development, Horticulture Research, 10, 5.
Fruit growth and development consist of a continuous succession of physical, biochemical, and physiological changes driven by a genetic program that dynamically responds to environmental cues. Establishing recognizable stages over the whole fruit lifetime represents a fundamental requirement for research and fruit crop cultivation. This is especially relevant in perennial crops like grapevine (Vitis vinifera L.) to scale the development of its fruit across genotypes and growing conditions. In this work, molecular-based information from several grape berry transcriptomic datasets was exploited to build a molecular phenology scale (MPhS) and to map the ontogenic development of the fruit. The proposed statistical pipeline consisted of an unsupervised learning procedure yielding an innovative combination of semiparametric, smoothing, and dimensionality reduction tools. The transcriptomic distance between fruit samples was precisely quantified by means of the MPhS that also enabled to highlight the complex dynamics of the transcriptional program over berry development through the calculation of the rate of variation of MPhS stages by time. The MPhS allowed the alignment of time-series fruit samples proving to be a complementary method for mapping the progression of grape berry development with higher detail compared to classic time- or phenotype-based approaches.
MPhS R Package: A molecular phenology scale of grape berry development
With the MPhS package, the user can map their transcriptomic dataset onto the Molecular Phenology Scale (MPhS). User instructions in the paper Scaling the grape berry development stage by molecular phenology: Application details and caveats (see “Other contributions”)
Giovanni Battista Tornielli, Ron Shmuleviz, Alessandra Amato, Marco Sandri, Paola Zuccolotto, Mario Pezzotti, Nick Dokoozlian, Marianna Fasoli, Sara Zenoni (2025), Scaling the grape berry development stage by molecular phenology: Application details and caveats, Plants People Planet.
Societal Impact Statement
Global climate change is adversely affecting grape quality by accelerating developmental processes. We developed an advanced method to define grape berry phenological stages based on gene expression information and made it accessible for widespread research. By precisely identifying these growth stages we can improve our understanding of how environmental conditions and agricultural practices influence grape quality. This has significant implications for developing mitigation strategies in viticulture and emphasizes the need for adaptive policies that can sustain grape production in the face of the changing climate.
Summary
Determining the developmental stage of grape berries is essential to understand the effect of environmental and/or cultivation factors. Our work aims to illustrate thoroughly the application of the recently established Molecular Phenology Scale (MPhS) to any custom berry transcriptomic dataset to map the ontogenic development of the fruit with high precision.
We detail the code components and instructions to run the MPhS package in R for berry transcriptomic datasets. We describe the output of the application of this tool on berry samples from plants subjected to various cultivation and environmental factors (e.g. cluster thinning, defoliation, water limitation, and varying temperature regimes). We illustrate the procedure for unveiling molecular responses uniquely related to the tested factor by comparing differentially expressed genes upon alignment of fruit samples by MPhS stage.
We demonstrated that the MPhS application allows defining the shifts of fruit development driven by various agronomic and environmental factors. Moreover, by performing statistical analysis on grape samples aligned according to the MPhS, we clearly highlighted some modulation of secondary metabolism specifically triggered by crop load manipulation, beyond the anticipation or delay of developmental progression.
The application of the MPhS to align time-series samples has proven to be an advanced method to define the developmental stage of grape berries. This is particularly important when the effect of cultivation/environmental factors is studied through gene expression analysis. The method is now fully available to all users for their own research scopes and applications.
Alessandra Amato, Giovanni Battista Tornielli, Ron Shmuleviz, Marco Sandri, Paola Zuccolotto, Mario Pezzotti, Marianna Fasoli, Sara Zenoni, A molecular phenology scale of grape berry development, 67th Congress of the Italian Society of Agricultural Genetics, 11 – 13 September 2024, Bologna, Italy.
The Molecular Phenology Scale (MPhS) was applied to to a highly comprehensive RNA-sequencing dataset and allowed the alignment of time series fruit samples, proving to be a complementary method for mapping the progression of grape berry development with greater classification detail compared to classic time- or phenotype-based approaches.
Giovanni Battista Tornielli, Marco Sandri, Marianna Fasoli, Nick Dokoozlian, Mario Pezzotti, Paola Zuccolotto, Sara Zenoni, A novel berry phenological scale based on gene expression, 11th International Symposium on Grapevine Physiology & Biotechnology, 31 October – 5 November 2021, Stellenbosch, South Africa.
Phenology scale systems widely adopted by viticulturists define stages of the annual development of the vine based on the visual description of well recognizable traits related to organ growth and morphology, including grape features from fruit set to maturity. However, although some stages can be easily described (e.g. fruit set, veraison), defining a comparable developmental stage for grapes of the same cultivar when grown in different conditions or for grapes of different cultivars can be challenging, in particular after the onset of ripening. By analysing transcriptomic data collected over berry development, it was shown that the variations of a portion of the transcriptome exhibited conserved dynamics across cultivars and growing condition of grapevines, and thus may be used to describe the developmental stage of berry development. In this work, we used the transcriptomic data generated from grape berries weekly sampled from Cabernet Sauvignon and Pinot noir vines grown in the same location over three consecutive vintages, focusing on conserved annual dynamics rather than on the biological significance of the expression program inferred by gene function. By interpolating the transcriptomic samples dispersed in a 3D space of a PCA we built a 30-stage Transcriptional Phenology Scale (TPhS) precisely defining the progression of development from berry formation to full ripening. The performance of the scale was assessed projecting onto the TPhS both RNA-seq and microarray transcriptomic samples from the same dataset used to elaborate the scale, and from several other public datasets. The results allowed to align samples on the new phenological scale and to highlight differences related to variables like the grape variety, the cultivation site, the vintage, or the applied treatment such as cluster thinning, defoliation, water limitation and temperature regimes. In some cases, the phenological re-scaling of sample collections from previous studies provided valuable hints to re-interpret the experimental results. Overall, we show that the transcriptomic information can be accessed to precisely define a transcriptional phenology scale that can be used to map the ontogenetic development of the fruit with high precision and to align the stage of berry development of different grapes.
