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Revista Chapingo serie ciencias forestales y del ambiente

versión On-line ISSN 2007-4018versión impresa ISSN 2007-3828

Rev. Chapingo ser. cienc. for. ambient vol.30 no.1 Chapingo ene./abr. 2024  Epub 03-Dic-2024

https://doi.org/10.5154/r.rchscfa.2022.10.079 

Scientific articles

Interaction and compatibility in reciprocal grafting with two varieties of Pinus pseudostrobus Lindl.

Rubén Barrera-Ramírez1 
http://orcid.org/0000-0002-0491-5721

J. Jesús Vargas-Hernández2 
http://orcid.org/0000-0001-7422-4953

Martín Gómez-Cárdenas1 
http://orcid.org/0000-0003-2765-957X

Eduardo J. Treviño-Garza3 
http://orcid.org/0000-0002-8921-857X

Alberto Pérez-Luna2  * 
http://orcid.org/0000-0002-6013-6028

1Instituto Nacional de Investigaciones Forestales Agrícolas y Pecuarias (INIFAP). Av. Latinoamericana, col. Revolución núm. 1101. C. P. 60150. Uruapan, Michoacán, México.

2Colegio de Postgraduados, Postgrado en Ciencias Forestales. km 36.5 carretera México-Texcoco. C. P. 56230. Montecillo, Texcoco, Estado de México, México.

3Universidad Autónoma de Nuevo León (UANL), Facultad de Ciencias Forestales. Carretera Nacional km 145. C. P. 67700. Linares, Nuevo León, México.


Abstract

Introduction:

Compatibility between the parts involved in grafting is one of the factors defining its success, growth, and productivity.

Objective:

The aim of this study is to determine the effect of the level of taxonomic affinity in the scion/rootstock combination on the survival and shoot growth of reciprocal grafts of Pinus pseudostrobus var. pseudostrobus and P. pseudostrobus var. oaxacana.

Materials and methods:

The effect of the following three factors was determined: (a) three affinity levels, (b) two varieties of P. pseudostrobus and (c) four scion/ rootstock genotypes, on survival and shoot growth (Sg). An ANOVA was performed to determine the effect of factors a, b and c and their interactions on survival and Sg, and a Log-Rank test was carried out for survival analysis.

Results and discussion:

Factor a determined that survival was higher in intervarietal grafts (53.3 %); factor b indicated that grafts with scions of var. pseudostrobus had higher survival (47.2 %); finally, factor c showed the existence of genotypes that increase success if used as scion and decrease if used as rootstock. Sg only showed statistical differences associated with the effect of factor c, being higher for genotype 62 of var. pseudostrobus (18.1 ± 0.63 cm), and in its interaction with factor (a).

Conclusions:

There is an adequate level of compatibility between the two varieties of P. pseudostrobus with significant effect of variety and scion genotype especially in intervarietal grafting (survival greater than 70 %).

Keywords: taxonomic affinity; terminal cleft grafting; homografts; intervarietal grafts; genotypes

Resumen

Introducción:

La compatibilidad entre las partes involucradas del injerto es uno de los factores que define su éxito, crecimiento y productividad.

Objetivos:

Determinar el efecto del nivel de afinidad taxonómica en la combinación púa/patrón sobre la supervivencia y crecimiento del brote de injertos recíprocos de Pinus pseudostrobus var. pseudostrobus y P. pseudostrobus var. oaxacana.

Materiales y métodos:

Se determinó el efecto de tres factores: (a) tres niveles de afinidad, (b) dos variedades de P. pseudostrobus y (c) cuatro genotipos de la púa/patrón, sobre la supervivencia y crecimiento del brote (Cb). Se realizó el análisis de varianza para determinar el efecto de los factores a, b y c y sus interacciones sobre la supervivencia y Cb, y una prueba de Log-Rank para el análisis de la supervivencia.

Resultados y discusión:

El factor a determinó que la supervivencia fue mayor en injertos intervarietales (53.3 %); el factor b indicó que los injertos con púas de la var. pseudostrobus presentan mayor supervivencia (47.2 %); finalmente, el factor c demostró que existen genotipos que aumentan el éxito si se utilizan como púa y disminuye si se utilizan como patrón. El Cb solo presentó diferencias estadísticas asociadas al efecto del factor c, siendo mayor en el genotipo 62 de var. pseudostrobus (18.1 ± 0.63 cm), y en su interacción con el factor (a).

Conclusiones:

Existe un nivel adecuado de compatibilidad entre las dos variedades de P. pseudostrobus con efecto significativo de la variedad y el genotipo de la púa sobre todo en los injertos intervarietales (supervivencia mayor de 70 %).

Palabras clave: afinidad taxonómica; injerto de fisura terminal; homoinjertos; injertos intervarietales; genotipos.

Highlights:

  • var. pseudostrobus genotypes had higher survival (47. 2 %) compared to var. oaxacana.

  • Some genotypes of var. pseudostrobus interact better as scion than as rootstock.

  • Survival was higher for intervarietal grafts (53.3 %).

  • Shoot growth was higher for genotype 62 of var. pseudostrobus (18.1 cm).

Introduction

Vegetative propagation allows full cloning of the desired superior genotypes (Darikova et al., 2011). Grafting has been used mainly for fruit tree propagation, while in conifers it has gradually increased, especially for establishing clonal seed orchards (Barrera-Ramírez et al., 2021; Muñoz et al., 2013; Pérez-Luna et al., 2020). The grafting technique is useful for propagating difficult-to-root species (Kita et al., 2018), accelerating seed production with scions from sexually mature individuals (Zobel & Talbert, 1988) and for preserving superior genotypes in clonal banks (Goldschmidt, 2014). Grafting success is higher when scion and rootstock present taxonomic and genetic affinity (Kita et al., 2018; López-Hinojosa et al., 2021).

Scion-rootstock compatibility is reflected by the degree of successful union between the two graft structures and their subsequent development as a single plant (Goldschmidt, 2014). Generally, graft compatibility and success are associated with the level of taxonomic affinity (Darikova et al., 2011). This varies from homografts (when scion and rootstock come from the same individual, with high compatibility), intraspecific grafts (scion and rootstock belonging to the same taxon) and interspecific grafts (scion and rootstock from different species of the same genus), to intergeneric grafts usually presenting incompatibility (Darikova et al., 2011; Goldschmidt, 2014).

Incompatibility between scion and rootstock, due to lack of cambium union, affects graft growth and survival (Castro-Garibay et al., 2017; Goldschmidt, 2014). However, initial cambium attachment alone does not guarantee long-term compatibility; incompatibility sometimes appears after several years (Castro-Garibay et al., 2017; Kita et al., 2018; López-Hinojosa et al., 2021). Although incompatibility is not a measurable quantitative trait several degrees can be differentiated, from a minor interference with a normal development of the graft, to the mortality of one or both structures (Goldschmidt, 2014). The most common failures occur during the choice of the genotypes involved in the graft, as failure is mainly due to differences in growth and development of the parts, where the scion grows faster than the rootstock or vice versa (Goldschmidt, 2014).

The information available on the relationship between taxonomic affinity and graft compatibility in pines is limited (Castro-Garibay et al., 2017), as most of the studies on this topic have been developed using fruit species (Kita et al., 2018). When interspecific grafting of conifers has been practiced (Pérez-Luna et al., 2020), the degree of taxonomic affinity of the parts has not been considered to evaluate the success of the grafts (Castro-Garibay et al., 2017).

Pinus pseudostrobus var. pseudostrobus Lindl. and P. pseudostrobus var. oaxacana Martínez have economic and ecological importance in Mexico, due to their wide distribution (case var. pseudostrobus) and common use in reforestation programs and commercial plantations (Barrera-Ramírez et al., 2020; Viveros-Viveros et al., 2006). In addition, these species have been incorporated in forest genetic improvement programs (Viveros-Viveros et al., 2006) and in vegetative propagation studies by grafting (Barrera-Ramírez et al., 2021; Muñoz et al., 2013) with varying degrees of success. Due to the genetic variation and differentiation within and among its populations, this species represents a good biological model to evaluate the relationship between the taxonomic affinity of the scion/ rootstock combination on grafting success. The identification of this relationship would increase the success of clonal propagation programs and seed production of higher genetic quality in seed orchards of this species.

The objective of the present study was to determine the effect of the level of taxonomic affinity in the scion/rootstock combination on the survival and shoot growth of grafts of P. pseudostrobus var. pseudostrobus and P. pseudostrobus var. oaxacana. The effects of scion/rootstock interaction were also evaluated using reciprocal grafting at three levels of taxonomic affinity. The research hypothesis indicates that at least one level of taxonomic affinity had a significant effect on survival and shoot growth of terminal cleft grafts in the two varieties of P. pseudostrobus.

Materials and Methods

Rootstock origin and preparation

The study was carried out at the forest nursery of the Agricultural and Forest Experimental Field, Uruapan, Michoacán of the Instituto Nacional de Investigaciones Forestales, Agrícolas y Pecuarias, located at 19° 24’ 25.35” N and 102° 3’ 4.16” W, at 1 610 m elevation, in Michoacán, Mexico. A shade net with 50 % light retention was used to reduce the temperature inside the nursery. The seed to produce the standard plant was collected from four superior trees of P. pseudostrobus var. oaxacana in different locations in Oaxaca; one superior tree per location was selected to reduce the error attributable to the effect of the parent. The same procedure was used for the variety pseudostrobus, but at locations in Michoacán. No evidence of natural hybridization was found at the collection sites, so the rootstocks produced were considered as representative trees of each variety and the plants as desired progenies (Table 1).

Table 1 Seed origin to produce rootstocks and scions of Pinus pseudostrobus var. oaxacana and P. pseudostrobus var. pseudostrobus

Variety Genotype Locality Latitude (N) Longitude (O) Altitude (m)
oaxacana 3 Santa Catarina Ixtepeji, Oaxaca 17° 10' 96° 33' 2 347
13 Santa María Jaltianguis, Oaxaca 17° 21' 96° 30' 2 367
38 Teococuilco de Marcos Pérez, Oaxaca 17° 18' 96° 41' 2 495
45 San Pedro Yolox, Oaxaca 17° 31' 96° 41' 2 536
pseudostrobus 6 Ciudad Hidalgo, Michoacán 19° 36' 100° 40' 2 376
15 El Rosario, Michoacán 19° 30' 100° 18' 2 680
62 Santa Ana Jeráhuaro, Michoacán 19° 48' 100° 37' 2 630
76 Nuevo San Juan Parangaricutiro, Michoacán 19° 27' 102° 12' 2 665

Germination was carried out under greenhouse conditions using plastic containers of 310 cm3 with 54 cavities containing substrate formed by a mixture of peat moss, vermiculite and perlite in a 2:1:1 ratio. After two months, the plant was placed in the nursery under 50 % light retention shade netting, a condition maintained until grafting. Subsequently, at 10 months, the plants were transplanted to black polyethylene bag, 400 caliber of 5.2 L in a substrate of 60 % loam (humic Andosol) and 40 % ground pine bark with Multicote® 15-7-15 + 2 MgO + micronutrients fertilizer, in the proportion of 3 kg∙m-3 of substrate. At the time of grafting, the rootstock of both varieties was 1.5 years old. The plants of the oaxacana and pseudostrobus varieties had average heights of 77.5 ± 8.5 cm and 88.5 ± 5.5 cm, respectively, and average diameter at the base of the stem of 11.0 ± 1.5 mm and 12.4 ± 2.2 mm, respectively.

Collecting the scions

To ensure the greatest genetic compatibility between the organs to be grafted (at least 50 % in homografts), the scions were collected from the same batches of rootstock, corresponding to the progenies of the genotypes selected for each variety; this practice allows obtaining high values of grafting (Loupit & Cookson, 2020). The scions corresponded to the elongation stage and met the growth characteristics described by Viveros-Viveros and Vargas-Hernández (2007). Scions were collected on the same day of grafting (March 10, 2019) by cutting the upper section at 35 ± 6.5 cm from the terminal apex toward the base of the plant, to perform the combinations at each level of scion/rootstock affinity. Scions were 12 ± 1.5 cm long and 10.5 ± 2.2 mm in diameter and were treated with Ridomil Gold® fungicide at a dose of 1 mL∙L-1 water to prevent fungal damage to the graft.

Grafting combinations by affinity level

Three levels of scion-rootstock affinity were considered in this test: 1) homografts (i.e., combination of scion and rootstock from progenies of the same parent tree) in the two varieties; 2) intravarietal grafts (combination of scion and rootstock from different progenies in the same variety) and 3) intervarietal grafts (combination of scion and rootstock from different varieties). The first affinity level included all eight possible combinations (four in each variety) with six repetitions each (48 grafts); the second level included 24 combinations (12 per variety, including reciprocal grafts) with five repetitions each (120 grafts), and the third level included 18 combinations (nine per variety) with five repetitions each (90 grafts) (Figure 1). The interaction ratio was 4:12:9 for levels one, two and three for each variety. A total of 258 grafts were made using the top cleft technique described by Muñoz et al. (2013) and Barrera-Ramírez et al. (2021), which consists of protecting the graft area with a microclimate created by a transparent polyethylene bag with water inside. Grafting was performed on the same day as scion collection.

Figure 1 Combinations at each level of scion-rootstock affinity of superior tree progeny of two Pinus pseudostrobus varieties; where 1 = homografts, 2 = intravarietal grafts and 3 = intervarietal grafts. 

Variables evaluated

Graft survival (GS) was evaluated using the criteria outlined by Muñoz et al. (2013) and Barrera-Ramírez et al. (2021) which include binary categories of 0 for unsprouting or decayed grafts, where functional cambial union was not achieved, and 1 for success grafting and with new needle growth response and shoot elongation. GS value was recorded from five to 90 days after grafting (dag). For grafting with GS = 1, shoot length was recorded from a reference point marked on the scion at 30 dag.

Statistical analysis

Three factors with different levels each were evaluated: a) three levels of scion-rootstock affinity, b) two varieties and c) four genotypes (origin) nested for each variety. A 3 x 2 x 4 factorial arrangement was used in a completely randomized experimental design with different number of repetitions per treatment; each graft represented an experimental unit.

The effect of the factors and their interactions in relation to survival values were determined with an analysis of variance (ANOVA) with the generalized linear mixed procedure (GLIMMIX) for data with binomial distribution (ɑ = 0.05); for shoot growth, the ANOVA was performed with the MIXED procedure for data with normal distribution. The data used in the analysis correspond to the evaluation at 90 dag. When significant differences were observed, a Tukey's mean comparison test was performed (P = 0.05). Survival dynamics were analyzed with the Log-Rank test, using curves constructed by the Kaplan-Meier method, using the function S (t) = P (T ≥ t); where, S (t) is the probability of a death occurring at a time T; P is the probability of survival at the time intervals during evaluation; T is the total survival time to the end of the evaluation, which must be greater than t, and t is the time at any time from the beginning of the evaluation (Kaplan & Meier, 1958).

The scion-rootstock interaction was analyzed with the results of the variety*affinity level interaction using a Chi2 test (α = 0.05) for individual samples, where the effect of each genotype on grafting success when used as scion or as rootstock was compared regarding the average survival values where it participated in one or the other category. All statistical analyses were performed with SAS 9.4 (SAS Institute, 2013).

Results and Discussion

Graft survival

At 35 dag, needle formation and shoot elongation were detected, indicators of P. pseudostrobus grafting success. At 90 dag, overall survival was 33 % which in comparison with other studies on the same species is a high value; for example, Barrera-Ramírez et al. (2021) reported 30 % survival at 90 dag with the same grafting technique, Muñoz et al. (2013) reported 80 % survival at 60 dag, but all died before 90 dag.

The age of the scions (1.5 years), which came from a batch of plants produced in a nursery, is not considered an impediment to complementing genetic improvement programs for the species, since P. pseudostrobus varieties, being fast-growing, begin to flower and reproduce between the ages of five and seven years (Patiño-Valera, 1973). In addition, grafting is a propagation technique that accelerates the flowering process of the species (Valdés et al., 2003).

The top cleft grafting technique has been widely used in species of the genus Pinus, because it is possible to obtain survival greater than 50 %, better shoot growth and precocity in the production of cones, from the first year of grafting (Gaspar et al., 2017; Kita et al., 2018).

Figure 2 shows the survival dynamics estimated with the Kaplan-Meier model. This test indicated significant differences (P < 0.01) between graft affinity levels. Survival of homografts and intravarietal grafts was less than 20 %, while in intervarietal grafts it was 32.8 %, with differences of up to 15.5 % between these and homografts.

Figure 2 Survival dynamics with the Kaplan-Meier model for grafting at three affinity levels with Pinus pseudostrobus var. oaxacana and var. pseudostrobus. Values with different letters indicate statistical differences between affinity levels according to the Tukey's test (P = 0.05). 

Survival was determined by pre- and post-grafting environmental conditions (early spring), since the excess relative humidity of the microclimate contributed to the presence of fungi, which, despite the application of fungicides, increased the percentage of mortality.

According to Table 2, the three individual factors and the interaction of affinity level with scion variety (a*b) had a significant effect (P < 0.05) on graft survival; shoot growth only showed statistical differences associated with the effect of genotype in the same variety (factor c) and its interaction with affinity level (a*c). The success in intervarietal grafting, with the top cleft technique and scions at the end of elongation, differs from that of studies with other pine species, as in grafts with scions of the hybrid of P. engelmannii Carr. x P. arizonica Engelm. on P. engelmannii rootstocks with 80 % survival (Pérez-Luna et al., 2020). It also differs with the results of Kita et al. (2018) with interspecific grafting of Larix gmelinii var. japonica on rootstocks of the hybrid L. gmelinii var. japonica × L. kaempferi with 93 % success. These authors conclude that the vigor and phenological stage of the scion (dormancy) significantly favored grafting. On the other hand, the use of young rootstocks and scions and the associated physiological characteristics, plus the grafting technique, result in greater success and growth of the grafts, because these factors favor a stronger, healthier, and more homogeneous vascular union between both structures, as pointed out by Milošević et al. (2015) and Kita et al. (2018).

Table 2 Analysis of variance of shoot survival and growth in reciprocal grafts of Pinus pseudostrobus var. oaxacana and P. pseudostrobus var. pseudostrobus. 

Factor Degrees of freedom Survival Shoot growth
Affinity level (a) 2 0.008* 0.228
Variety (b) 1 0.029* 0.834
Genotype in variety (c) 6 0.008* 0.019*
a*b interaction 2 0.003* 0.129
a*c interaction 10 0.204 0.0214*

*Significant at a P value ≤ 0.05.

Effect of affinity level

According to Table 3, Tukey's test (P = 0.05) established that the highest survival value was 53.3 ± 4.2 % in intervarietal grafts and the lowest in homografts with 22.8 ± 5.2 %, both at 90 dag. Therefore, intra- and intervarietal grafting can be performed with the two studied varieties of P. pseudostrobus. Shoot growth was not significantly affected (P > 0.05) by the scion/ rootstock affinity level, with an overall mean value of 15.0 ± 0.7 cm at 90 dag.

In most studies it has been pointed out that in grafts with higher taxonomic affinity (case of homografts), success is higher (Goldschmidt, 2014), but the results with P. pseudostrobus varieties indicated the opposite, as survival was higher (more than double) in intervarietal grafts.

Table 3 Survival and shoot growth in reciprocal grafts with three affinity levels in two varieties of Pinus pseudostrobus. 

Affinity level Observations (n) Survival (%) Shoot growth (cm)
Homografts 48 22.86 ± 5.2 b 15.74 ± 0.68 a
Intravarietal grafts 120 30.82 ± 4.2 ab 15.04 ± 0.38 a
Intervarietal grafts 90 53.28 ± 4.5 a 14.41 ± 0.41 a

Mean values ± standard error. Different letters indicate significant differences between affinity levels according to the Tukey's test (P = 0.05).

Some studies have reported that differences in age between scion donor trees or in vigor between scion and rootstock, regardless of affinity level, affect grafting success and development (López-Hinojosa et al., 2021). However, in this study, the scions and rootstocks used had the same age (1.5 years), therefore, differences in the success of intervarietal grafting cannot be attributed to the effect of this factor.

In homografts, inefficient tissue fusion between scion and rootstock could be due to external factors such as post-graft handling, climate, scion quality (Loewe-Muñoz et al., 2022) or due to early partial incompatibility (Gaspar et al., 2017; Goldschmidt, 2014). The low success of homografts can also be attributed to differences in diameter growth, as sometimes scion growth is usually more accelerated than in the rootstock (Pérez-Luna et al., 2019; Solorio-Barragán et al., 2021). The fact is that in these grafts an adequate coupling was not achieved. Goldschmidt (2014) and López-Hinojosa et al. (2021) mention that the reduction of cambial activity and nutrient transport cause low percentage of survival and lower shoot growth, so this study considered that the low success in homografts is mainly due to this factor, even without understanding the precise mechanism by which it occurs.

In contrast, intra- and intervarietal grafts apparently presented acceptable cambial activity and substance transport between scion and rootstock that was reflected in higher survival and adequate support of the two structures, as described by Milošević et al. (2015) and Loewe-Muñoz et al. (2022) for other pine species. Solorio-Barragán et al. (2021) note that the survival of interspecific grafts of Pinus rzedowskii Madrigal & Caball. Del. on rootstocks of P. pinceana, P. maximartinezii, P. ayacahuite var. veitchii and P. pseudostrobus var. pseudostrobus was higher (80 ± 0.3 %) when increasing phylogenetic affinity with rootstocks of P. pinceana than with rootstocks of P. pseudostrobus var. pseudostrobus (20 ± 0.9 %). Therefore, the success of intervarietal grafting on P. pseudostrobus indicates a close relationship between the two varieties.

On the other hand, scions from apical shoots on primary and secondary branches at the top of the crown generated high survival rates, as was the case in intervarietal grafting. Although the grafted scions were taken from nursery-produced plant, Gaspar et al. (2017) and López-Hinojosa et al. (2021) note that the origin of the scion, in terms of its position in the crown of the donor tree, significantly influences graft survival. This effect was also observed by Solorio-Barragán et al. (2021), who used P. rzedowskii scions from terminal buds in interspecific grafting.

Variety effect

Survival in grafts with scions of var. pseudostrobus (47.2 ± 5.0) was significantly higher (P = 0.05) than in grafts with scions of var. oaxacana (24.1 ± 5.2). One reason for the low percentage of survival in scions of this variety is due to poor scion-rootstock union, attributed to the physiological characteristics of the vegetative material, as well as scion provenance and origin and partial incompatibility at the three levels of affinity. In addition, the higher number of resiniferous canals in var. oaxacana (Martínez, 1943) could also have affected grafting success, as it has been documented that resin release in Pinus can cause mortality (Pérez-Luna et al., 2019), as well as the grafting season (autumn in this case) (Barrera-Ramírez et al., 2021). In the case of shoot, the effect of variety was similar (P > 0.05) with 15 cm of average growth.

The greater success of grafting with scions of var. pseudostrobus indicates good scion-rootstock compatibility. It is possible that grafting success with scions of var. oaxacana may increase if younger rootstocks are used; however, further studies are needed to support this assertion. In grafting with Araucaria angustifolia (Bertol.) Kuntze, it has been reported that variety, physiological stage of the scion, genotype, time and post-grafting conditions are determinants for high survival rates (>60 %; Gaspar et al., 2017); therefore, in P. pseudostrobus, the interaction of these factors also contributes to greater success and better shoot growth (Barrera-Ramírez et al., 2021; Muñoz et al., 2013). The same happened in intra- and intervarietal grafting of Prunus americana L., where Milošević et al. (2015) highlight the importance of proper scion choice of a variety that works in most crosses with rootstocks. Ahsan et al. (2019) obtained 80 % survival in intervarietal grafts of P. americana Mill ‘Hass’ x P. americana Mill ‘Velvick’ (scion/rootstock); however, success decreased in their reciprocals (-20 %). In the present study, buds from leading, young and healthy shoots were used, a factor that increased graft success, vigor and growth, as reported by Świerczyński et al. (2018) and Kita et al. (2018), especially in intervarietal grafts of var. pseudostrobus x var. oaxacana (scion and rootstock, respectively). Furthermore, phenological differences in scion and rootstock growth (Martinez-Ballesta et al., 2010) between the two varieties, together with genotype, significantly determined grafting success and growth (Koepke & Dhingra, 2013). This may be attributed to the fact that a young bud increases vegetative growth of the grafts (Martinez-Ballesta et al., 2010), as was the case in the present study, in addition to facilitating vascular connections between tissues in the scion/rootstock interaction.

Genotype in variety

Table 4 shows that survival was higher in var. pseudostrobus (40 a 62 %); however, not differences were found among genotypes. For var. oaxacana, grafting success with juvenile scions varied significantly among genotypes; the highest survival values were obtained with scions of genotype 3 (53.0 ± 7.8 %). In contrast, shoot growth showed significant differences only for var. pseudostrobus; grafts with scions of genotype 62 reached the best growth with 18.1 cm.

Table 4 Survival and shoot growth per genotype in the same variety in grafts made with scions of two varieties of Pinus pseudostrobus

Genotype (same variety) Observations (n) Survival (%) Shoot growth (cm)
var. oaxacana
3 36 53.0 ± 7.8 a 15.2 ± 0.55 a
13 36 11.0 ± 7.8 c 14.0 ± 0.63 a
38 21 19.0 ± 10.3 bc 14.5 ± 0.58 a
45 36 42.0 ± 7.8 ab 14.7 ± 0.60 a
var. pseudostrobus
6 36 40.0 ± 7.8 a 14.1 ± 0.63 b
15 36 44.0 ± 7.8 a 13.6 ± 0.61 b
62 36 42.0 ± 7.8 a 18.1 ± 0.63 a
76 21 62.0 ± 10.2 a 15.0 ± 0.73 ab

Mean values ± standard error. Different letters indicate significant differences between genotypes of the same variety according to the Tukey's test (P = 0.05).

Differences in survival and shoot growth of genotypes in P. pseudostrobus varieties indicate phenological and physiological differences in the stage of scion or rootstock development, as pointed out by Darikova et al. (2011) in their study with heteroplastic grafts of Pinus sibirica Du Tour and P. cembra L. grafted on P. sylvestris L. rootstocks.

Interaction effects

The interaction affinity level by genotype in the same variety (a*c) had significant effects (P = 0.05) on shoot growth. Graft growth differences among var. pseudostrobus genotypes were different from one affinity level to another; only genotype 62 had the best values in the three affinity levels with an average growth of 15.2 to 18.1 cm. For var. oaxacana, growth was similar (P > 0.05) among graft affinity levels with an average range of 15.4 to 17.3 cm.

The interaction of affinity level with scion variety indicated that reciprocal effects in scion-rootstock interaction are important in intervarietal grafting, since a higher percentage of survival was observed when grafting scions of var. pseudostrobus onto the rootstock of var. oaxacana (72 ± 1.2 %), than the other way around (33.9 ± 1.4 %), while in intravarietal grafts no significant differences were found (average success of 31 ± 2.1 %). Higher grafting success was also reported in homografts with scions and rootstocks of var. pseudostrobus (42 ± 0.14 %) (Figure 3).

Figure 3 Average survival (%) of grafts with different levels of affinity in the two varieties of Pinus pseudostrobus included in this experiment. 

The success of var. oaxacana seemed to remain the same when its scions were grafted on rootstocks of the same variety or on a rootstock of var. pseudostrobus. On the other hand, the success of var. pseudostrobus scions grafted on rootstocks of the same variety was good (42 %), although it was better when grafted on var. oaxacana rootstocks (72 %; Figure 3).

Differences in the survival of reciprocal intervarietal grafts may be associated with physiological differences in the scion/rootstock interaction, due to the contrast between the growth and diameter of scion and rootstock. The average diameter of scions and rootstocks was similar in grafts of var. oaxacana (homograft) (7.0 and 8.7 mm), as well as in intravarietal grafts with var. pseudostrobus (8.0 and 9.5 mm). However, the difference in diameter of intervarietal grafts of var. oaxacana on var. pseudostrobus rootstocks was greater (7.0 vs. 9.5 mm), which may have generated less contact between cambial zones and affected grafting. In contrast, reciprocal intervarietal grafts of var. pseudostrobus on var. oaxacana rootstocks had greater homogeneity in scion and rootstock diameter (8.0 and 8.7 mm), which favored greater contact between cambial tissue. The var. oaxacana had a greater number of resin channels (Martinez, 1943), compared to var. pseudostrobus, which probably affected the functional union of the scion and rootstock, since the excessive release of resin from the scion prevents contact with the cambium of the rootstock and reduces the possibility of grafting success, as mentioned by Pérez-Luna et al. (2019).

Despite the significant effect of the variety and affinity level interaction, indicating the importance of scion-rootstock interaction in intervarietal grafting, the Chi2 test did not detect significant differences (Table 5) in the scion or rootstock performance of the genotypes, except for genotype 15 in var. pseudostrobus, which was more successful when used as a scion (44.7 %) than as a rootstock (16.1 %). However, the small number of repetitions in each genotype in this experiment may have affected the ability of the statistical test to detect significant effects of reciprocal grafting at the genotype level. Therefore, it is necessary to identify the possible anatomical and physiological factors involved in the success or failure of these grafts, as well as the nature of scion-rootstock interaction in intervarietal and interspecific grafts, as compared to grafts between individuals with higher level of taxonomic and genetic affinity of P. pseudostrobus.

Table 5 Average percentage of success in genotypes of two Pinus pseudostrobus varieties when participating as scion/rootstock in intervarietal grafts. 

Variety Genotype Genotype function Chi2
Rootstock (%) Scion (%)
oaxacana 3 66.7 49.6 ns
13 25.7 11.4 ns
38 35.0 20.0 ns
45 40.0 42.9 ns
pseudostrobus 6 36.7 46.7 ns
15 16.1 44.7 <0.01
62 41.9 41.9 ns
76 45.8 52.8 ns

Significant differences at α = 0.05, ns = not significant; DF = 1 in X2 tables.

Conclusions

The survival of Pinus pseudostrobus grafts was good (>33 %) compared to that reported in other studies with the same species; however, further research is needed to identify the causes and pre- and post-grafting management factors that determine the success of this technique in this species and its varieties. The success and growth of grafts of both P. pseudostrobus varieties and their reciprocals are determined by the level of affinity, scion variety, scion genotype and scion-rootstock interaction at the variety level. Therefore, phylogenetic compatibility exists between the two varieties at the intervarietal level when scions of var. pseudostrobus are grafted onto rootstock of var. oaxacana (72 % survival), but not in their reciprocal grafts. This evidence indicates that the anatomical and physiological interaction between scion-rootstock is important in P. pseudostrobus, but more detailed studies are required for the identification of this interaction.

Acknowledgments

The authors thank the Consejo Nacional de Ciencia y Tecnología (CONACyT) for funding facilities to the first author and to project 277784 "Establishment of regional asexual seed orchards and progeny trials of Pinus pseudostrobus for genetic evaluation of parents" of the Fondo Sectorial CONACyT-CONAFOR.

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Received: October 29, 2022; Accepted: September 18, 2023

*Corresponding author: aperez@ujed.mx; tel.: +52 618 152 1186.

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