A quantitative assessment of factors affecting the rooting of grapevine rootstocks (Vitis vinifera L.)

ABSTRACT. Grapevine is a fruit species of great agronomic interest since both fresh fruit and wine are highly marketable products. Therefore, there is growing interest in this crop among researchers in the pursuit of increased yields and better cultivation conditions. Asexual propagation is the most commonly used method for propagating grapevine seedlings, with cuttings used for rootstock formation and subsequent grafting of materials to form the canopy. The rootstock is responsible for support, water, and nutrient uptake, thus determining the vigor of the plant; therefore, it is essential to understand the suitable conditions in which roots can thrive and thereby enhance plant growth. The type of cutting, type of substrate, and application of exogenous auxins are among the factors that interfere with the rooting of grapevine rootstocks, and determining and implementing the best combination of these factors could provide for more vigorous plants. Reviewing high-quality published scientific research from diverse countries is a method of identifying data, grouping information, and obtaining reliable answers. Furthermore, it can allow for the detection and grouping of the main factors affecting the rooting of grapevine cuttings, thus enabling a clearer recommendation and better understanding of the issue.


Introduction
Grapevine (Vitis vinifera L.) is a species of the Vitaceae family, native to the Mediterranean region, Central Europe, Southwest Asia, Morocco, from northern Portugal to southern Germany, and eastern to northern Iran.The genus Vitis includes many species worldwide.Its diffusion and importance are highlighted by the diverse usage of grapes, including by the consumption of fresh and dried fruits (raisins), and the production of wines, juices, vinegar, ethanol, jams, fertilizers, and antioxidant compounds.In addition, the medicinal properties of the grape aid in the prevention of cancer, heart disease, allergies, and diabetes (Neto, Ricardino, Souza, Aguiar, & Marques, 2020).
To obtain quality fruit, attention to all stages of cultivation is paramount, and seedling production is considered crucial for the successful installation of a vineyard.The sexual propagation of grapevines results in large plant heterogeneity as well as late and uneven fruit sets (Fronza & Hamann, 2015).Therefore, the use of this technique is restricted to breeding programs (De Albuquerque & Choudhury, 1993;Villa et al., 2003).The rooting of cuttings is the most commonly used technique for producing grapevine seedlings (De Albuquerque & Choudhury, 1993), by using a small portion of the branch of a parent plant to produce a new plant by growing adventitious roots (Bettoni et al., 2014a).The cutting technique is widely used for the preservation of rootstocks (Fronza & Hamann, 2015) for subsequent grafting of a cultivar of interest and seedling production as the foundation of a vineyard (Regina, Souza, & Dias, 2012).
From a physiological and anatomical point of view, there is some complexity in root development by cuttings.Factors such as hormonal balance, cutting and substrate type (Nachtigal & Pereira, 2000), and hormone and plant growth regulators in the auxin group (Taiz & Zaiger, 2013;Vilarinho & Cândido, 2014) influence the dedifferentiation and redirection of totipotent cells for rooting.
The existence of qualitative and quantitative uncertainties of the aforementioned factors creates a demand for studies that present a clearer identification of the combinations that make the production of grapevine seedlings viable.Therefore, this study aimed to conduct a literature review on the main factors that affect the rooting of grapevine cuttings and to provide a clearer and more objective recommendation for more successful seedling production.
In this literature review, research results on the factors that interfere with the rooting of grapevine cuttings were raised and discussed.

Data collection methodology
We reviewed 55 scientific articles published in journals with different impact factors and data from the most diverse grape-producing countries and regions.We chose to search for a better geographic representation of the information collected in order to cover as many particularities of each location as possible, thus reducing the effect of environmental variability on our review and conclusions.The databases used for the research were Scielo, Google Scholar, Science Direct, and CAPES websites.
Table 1 shows the number of articles selected for each investigation period (years) and the countries in which each study was conducted.Twenty-five (25) of the 55 articles were from the last ten years, of which 14 were from the past five years.Thirty-four (34) were from Brazil, and 21 were from eight other countries.The independent variables of the materials were grouped and defined as auxin dose (mg L -1 ), immersion time (seconds or hour), cutting type, cutting length (cm), auxin, rootstock, substrate, and presence or absence of a leaf (Table 2).Rooting percentage was regarded as a dependent variable, and we considered the highest rooting percentage (%) of each article in this study (Table 3).A total of 926 rooting percentages were found, with the amount of data per paper varying from 1 (minimum) to 108 (maximum).Since large variability of the data was observed in most of the papers, the use of the average value could result in low rooting percentage values.In addition, using the highest value allowed us to obtain a combination of variables that result in better rooting of grapevine rootstock cuttings, which makes it possible to verify the trend of the results and simplify future decisions regarding new studies and/or practical applications that seek better results in the rooting of grapevine cuttings.Table 2 also shows the number of articles in which the results represent the levels of each variable.), indole-3-butyric acid (IBA), naphthalene acetic acid (NAA), and indole-3-acetic acid (IAA); leaf 0.3 (30% of a leaf), 0.5 (50% of a leaf), and 1 (100% of a leaf).
Table 3 shows the information required for the selected materials and relates them to our results.The variables were grouped in columns, and the main information was summarized and standardized to facilitate interpretation.Number of articles (n), indole-3-butyric acid (IBA), naphthalene acetic acid (NAA), and indole-3-acetic acid (IAA); leaf of 0.3 (30% of a leaf), 0.5% (50% of a leaf), and 1 (100% of the leaf).

Results and discussion
The results were grouped into four categories to facilitate data discussion.The groups were organized by the effect of the dose and immersion time, cutting type and length, auxin and rootstock, substrate type, and presence or absence of leaves.

Dose and time of immersion
Despite finding that the application of exogenous auxin improves histological characteristics through the formation of calluses and tissues and differentiation of vascular tissue (Mitra & Bose, 1954;Singh, 2018), it may be unnecessary in other circumstances for the grapevine crop.Our results showed that the highest rooting percentages (%) were obtained with the use of exogenous auxin (Figure 1A) at doses of 250, 500, 1500, and 3000 mg L -1 , all of which were close to 100%.However, even at a dose of 0 mg L -1 , the % values were above 80%.This may imply a recommendation other than the use of exogenous hormones to induce the root cuttings to reduce production costs.Several studies have reported little or no influence of plant growth regulators on the rooting of grapevine cuttings (Goode Jr. & Lane, 1983;Moretti & Borgo, 1985;Thomas & Schiefelbein, 2001;Keeley, Preece, & Taylor, 2003;Thomas & Schiefelbein, 2003;Villa et al., 2003;Keeley, Preece, Taylor, & Dami, 2004;Thomas & Schiefelbein, 2004;Botelho et al., 2005a;Tofanelli et al., 2014;Bartolini et al., 2017).However, our results indicate that there was a slight increase in the percentage of cuttings rooted in the application of exogenous auxins.Thus, its use could be justified when seeking greater rooting efficiency or when there is a limited amount of material available for propagation.

Cutting type and length
The data for cutting type showed the best results for rooting with hardwood cuttings (Figure 2A).In this cutting type, the average was 91.23%, while the averages for semi-hardwood and herbaceous cuttings were 76.72 and 84.00%, respectively.In general, rooting of hardwood grapevine cuttings is straight-forward (Biasi et al., 1997), and often 100% rooting can be obtained (Da Silva et al., 1986;Roberto et al., 2004c;Köse & Güleryüz, 2006;Sozim & Ayub, 2006;Amaral et al., 2009;Gökbayrak et al., 2010;Ahmed et al., 2017;Sabir & Sabir, 2018), compared to semi-hardwood (Bordin et al., 2005;Botelho et al., 2005b;Denega et al., 2009) and herbaceous cuttings (Leão, 2003;Villa et al., 2003;Nascimento et al., 2019) with percentageages below 85%.However, with the expansion of cultivation areas, the use of semi-hardwood (Alley, 1980;Egger, Moretti, & Borgo, 1985;Moretti & Borgo, 1985) and herbaceous cuttings (Winkler, Cook, Kliewerer, & Lider, 1974) are viable alternatives to respond to the demand for healthy and high-quality plant material.When there is a need to produce a large number of cuttings from a limited amount of available materials, shorter cuttings and a reduced number of buds can be used.Figure 2B shows higher values of with increasing cutting length.Cuttings lengths of 25-45 cm showed percentages above 90%.There are two possible reasons for these results.First, it is reasonable to relate the cutting length to the type; Figure 2B shows a direct relationship between the highest percentage and hardwood, which generally has longer cuttings.(Figure 5) (Leão, 2003;Monteguti et al., 2008;Daskalakis, Biniari, Bouza, & Stavrakaki, 2018;Jesus et al., 2018;Daskalakis et al., 2019).There is evidence that the degree of lignification of cuttings has a greater influence on the number of rooted cuttings compared to the length or number of buds per cutting (Leão, 2003).Second, there is likely a greater amount of nutritional reserves present and available for rooting in longer cuttings (Nicoloso, Cassol, & Fortunato, 2001).

Auxin and rootstock
The average value with the use of indole-3-butyric acid (IBA) as an exogenous inducer was 84.3% (Table 3).When combined, indole-3-acetic acid (IAA) and naphthalene acetic acid (NAA) represented only 6% of the selected studies (Table 2).Due to the small number of articles, it is not possible to generate conclusive information about the best auxin for inducing the rooting of grapevine cuttings.
The large number of studies which assessed IBA as a plant growth regulator suggests that this is the main synthetic hormone used for the rooting of grapevine cuttings (Machado et al., 2005).However, factors such as cultivar, time of year, and cutting type, reported 25 years ago (Fachinello, Hoffmann, Nachtigal, Kersten, & Fortes, 1995) still appear to cause variability in the results.
In view of the diverse range of rootstocks available for grapevine propagation, the main four (Table 2 and Figure 3) have a good rooting potential, particularly IAC 766 and Paulsen 1103 with close to 100% success.

Presence or absence of leaves and type of substrate
The presence of leaves proved to be a significant factor in the rooting of cuttings (Figure 4A).Despite the small number of articles in our database that compared rooting with and without leaves (Table 2), it is worth mentioning that the average was above 80% in the presence of leaves (regardless of the measurement unit) (Figure 4A).The presence of leaves in the rooting of herbaceous and semi-hardwood cuttings is essential for obtaining positive results with grapevine (Biasi et al., 1997), showing superior performance to that using hardwood cuttings (Zuffellato-Ribas & Rodrigues, 2001;Botelho et al., 2005b).Genetic materials that are generally challenging to root with hardwood cuttings (Goode Junior, Krewer, Lane, Daniell, & Couvillon, 1982) can provide good results with cuttings in the presence of leaves collected during the plant growth period (Goode Junior & Lane, 1983).IAC 766

Rootstock
This strong rooting of cuttings is justified by the fact that the leaf is an important auxin source (Hartman, Kester, Davies Junior, & Geneve, 2011), in addition to assisting in the production and transport of carbohydrates from the photosynthetic process to the base of the cutting (Pires & Biasi, 2003).However, the grapevine has large leaves, and a large leaf area is not preferable due to the increased risk of excessive dehydration, which can hinder rooting (Bordin et al., 2005).
The best results were 95, 92, and 89% obtained with nutrient solution, sand, and soil, respectively (Figure 4B).Based on the knowledge of the functionality of the substrate as a means for developing seedlings where structure, aeration, water retention, and absence of pathogens are essential, it is reasonable to conclude that the aforementioned substrates serve these functions well.Furthermore, sand and sandy soils are considered the most widely used substrates (De Albuquerque & Choudhury, 1993).However, despite the high average values, there is evidence of the formation of thick roots with little branching and brittleness when sand is used as a substrate (De Albuquerque & Choudhury, 1993;Hartmann et al., 2011).As a result, it is more advantageous to use soil because of its low cost, increased possibility of forming a healthy root system, and the likelihood of higher rooting percentages, as shown by our research data.However, it is not possible to generalize the use of soil as a substrate for grapevines.Specific recommendations have been made, such as vermiculite for herbaceous and hardwood cuttings (Gonçalves & Minami, 1994) and rice husks for the rooting of herbaceous cuttings of IAC 766 and IAC 572 rootstocks (Roberto et al., 2004a).

Recommendations for the grapevine
Figure 5 shows data on the relationship between the qualitative variable (cutting type) and the quantitative variables (length, rooting percentage, and immersion time).The Microsoft Excel "=AVERAGEIF (QUALITATIVE COLUMN, CRITERION, QUANTITATIVE COLUMN)" function allowed the grouping of this information.
It is possible to make recommendations based on the information presented in Figure 5, such as the fact that it is reasonable to consider applying higher doses of exogenous auxin to hardwood grapevine cuttings, in the absence of leaves at the time of cutting.To achieve a high rooting percentage (close to 100%) with the use of hardwood cuttings of approximately 25 cm (or 3 knots), we recommend using an auxin dose close to or above 1500 mg L -1 .However, this implies an increase in production costs with the purchase of exogenous auxin and increased consumption of plant material because of the larger size of the cuttings.The use of semi-hardwood and herbaceous cuttings for the propagation of grapevine rootstocks can be a viable alternative in situations where the availability of materials is limited, as the smaller size required for these cutting types maximizes the use of materials.Furthermore, the use of cuttings with leaves reduces the need for exogenous auxin application (Figures 4A and 5).Therefore, doses of 500 to 800 mg L -1 are sufficient to meet crop needs and obtain a good rooting percentage (close to 80%), thus reducing the costs for seedling production, and thereby allowing for increased net profit.

Conclusion
The use of hardwood cuttings for the propagation of grapevine rootstocks had the highest rooting percentage of all wood types evaluated, but with a presumed increase in the cost of seedling production due to the use of synthetic auxins.The presence of leaves in semi-hardwood and herbaceous cuttings increases seedling production efficiency and reduces the cost of purchasing plant growth regulators.For cultivars and/or varieties in which the amount of material available for propagation is limited, the use of semihardwood and herbaceous cuttings is recommended.

Table 1 .
Number of articles compiled over the past 40 years on the production of grapevine rootstock seedlings.

Table 2 .
Distribution of the number of scientific articles and the variables that influence the production of grapevine rootstock seedlings.