SciELO - Scientific Electronic Library Online

vol.76 issue5Phenotyping for angular leaf spot severity and its implication in breeding common bean for resistanceHistopathological characterization of Coffea arabica cultivar IPR 106 resistance to Meloidogyne paranaensis author indexsubject indexarticles search
Home Pagealphabetic serial listing  

Services on Demand




Related links


Scientia Agricola

On-line version ISSN 1678-992X

Sci. agric. (Piracicaba, Braz.) vol.76 no.5 Piracicaba Sept./Oct. 2019  Epub May 20, 2019 

Genetics and Plant Breeding

Resistance to Pratylenchus brachyurus in Vitis species population through multivariate approaches and mixed models

Paulo Ricardo dos Santos1

Alexandre Pio Viana1  *

Vicente Martins Gomes2

Sandra da Costa Preisigke1

Odimar Ferreira de Almeida1

Eileen Azevedo Santos1

Daniele Lima Rodrigues1

Rosana Rodrigues1

Michael Andrew Walker3

1Universidade Estadual do Norte Fluminense Darcy Ribeiro/CCTA– Lab. de Melhoramento Genético Vegetal, Av. Alberto Lamego,2000 – 28013-602 – Campos dos Goytacazes, RJ – Brasil

2Instituto Federal Fluminense, R. Cambuci, km 5 – 28430-000 – Três Irmãos, RJ – Brasil

3University of California – Dept. of Viticulture & Enology, One Shields Av. – 95616-5270 – Davis, CA –USA


Genetically diverse interspecific hybrids of Vitis were selected for resistance to Pratylenchus brachyurus. Three segregating populations with 57 hybrid crosses were evaluated. The parents included Vitis romanetiiC166-043 × 07355-075, 06354-047 × Cereza and 06354-047 × Nocera, selected from the germplasm bank at the University of California, Davis, the United States. The experiment was arranged in a randomized block design, with three replications and three plants per plot. Root mass, nematodes per gram of root and reproduction factor were determined and used as quantitative variables; 16 multi categoric descriptors were also evaluated. The traits were analyzed using the Ward-Modified Location Model procedure (Ward-MLM) for the composition of genotype groups. Genetic parameters and prediction of genetic values by Restricted Maximum Likelihood / Best Linear Unbiased Prediction (REML / BLUP) were assessed. The Ward-MLM classification strategy supported the formation of three homogeneous groups. Group I comprised 13 hybrids; Group II, 26 hybrids; and Group III, 18 hybrids. Groups II and III contained hybrids resistant to P. brachyurus. High broad sense heritability values were found for root mass, reproduction factor and nematodes per gram of root, which provided genetic gain and allowed selection of resistant genotypes available for cloning, since the total genetic variance occurred due to the dominance effects. Of the 57 genotypes assessed, those with the lowest genotypic values for reproduction factor were selected as resistant, including: CH3.2, CH3.23, CH3.8, CH3.37, CH3.38, CH3.41, CH3.36, CH2.1, CH2.7, CH1.1, CH1.3 and CH1.2.

Keywords: Vitis genotypes; nematode; root; genetic variability


In the international scenario, the Brazilian viticulture occupied the 11thplace in grape production and the 13th in wine production in 2015, accounting for a total production of 1.6 million tons of grapes in 2016 (FAO, 2016). Therefore, viticulture in Brazil has huge social and economic importance for generating employment and income, considering both table grapes and wine production (IBGE, 2016).

However, nematodes seriously impair grapevine establishment by hindering development, decreasing vigor and greatly reducing the plant root system, thereby decreasing fruit yield and quality. It is estimated that nematode feeding reduces annual grape production by 20 to 25 % worldwide (Puerari et al., 2012). The specie Pratylenchus brachyurus stands out for its wide dissemination in regions of agriculture cultivation. It has been recognized as one of the worst soil-borne pests of grapevines, especially under high populations in the soil (Téliz et al., 2007; Ferris et al., 2012).

The increase in population levels and damage caused by P. brachyurus represent a serious problem for production systems, compromising tropical viticulture (Puerari et al., 2012). The application of nematicides as a control method is an expensive and inefficient practice. The use of resistant genotypes is the main strategy for nematodes control in the vine, as it does not increase production costs, nor does it harm the environment. Therefore, the selection of resistant hybrids is necessary to be used as rootstocks for good yield in viticulture (Walker et al., 1994; Zasada et al., 2012).

The inclusion of wild species in breeding programs is one of the key approaches to pest and disease resistance breeding, since these species may have resistance genes not found in cultivated plants. Therefore, knowing the genetic variability to select resistant clones in segregating populations is particularly important for breeders when the level of variation in breeding populations is not fully known or explored (Walker et al., 2010; Viana et al., 2011).

This study aimed to characterize populations of interspecific crosses of Vitis based on qualitative and quantitative descriptors and estimate genetic parameters capable of selecting interspecific hybrids resistant to P. brachyurus for grapevine breeding programs.

Materials and Methods

Populations assessed

Three segregating populations with 57 hybrids obtained from crosses involving Vitis spp. were investigated in relation to their resistance to P. brachyurus (Table 1).

Table 1 Parents and crosses used in the morphological characterization and evaluation for resistance to P. brachyurus at the University of California, Davis, USA. 

Parentsa Traits
07355-075 (V. vinifera) Advanced wine grape selection with resistance to Pierce's disease originating from V. arizonica
Vitis romanetii C166-043 (V. romanetii) Chinese species, DVIT2732, from the National Clonal Germplasm Repository - Davis
Nocera (V. vinifera) An Italian wine grape from Sicily
Cereza (V. vinifera) Large berried Italian table grape
06354-047 (V. vinifera/M. rotundifolia) Powdery mildew resistant wine grape selection with resistance from a V. vinifera × Muscadinia rotundifolia hybrid
Crosses Number of hybrids assessed
Vitis romanetii C166-043 × 07355-075 7 - CH1a
06354-047 ×Cereza 9 - CH2a
06354-047 ×Nocera 41 - CH3a
Total 57

aCH1, CH2 and CH3 = hybrid obtained from crossed 1, 2 and 3 respectively.

Experimental details and phenotyping

Cuttings of hybrids were rooted and then planted in 7 L pots containing a soil: sand mixture atratio2:1 (v/v) and kept in a greenhouse in the municipality of Campos dos Goytacazes, Rio de Janeiro, Brazil (21°44’ S, 41°19’ W, altitude 12 m), with average temperature and relative humidity of 31.3 °C and 83 %, respectively. Meteorological data were obtained from a thermo-hygrometer inside the greenhouse.

The experiment was arranged in a randomized block design, with three replications and three plants per plot of 57 interspecific Vitis hybrids, and a maize cultivar, ‘BR 106’, which was used as source of inoculum and as a susceptible control to verify the viability of nematode inoculum. Maize is very sensitive to P. brachyurus infestation and cultivar ‘BR 106’ was used in this work for its wide distribution in commercial crops throughout Brazil.

After four months (in Nov 2015), the plants were inoculated with a suspension of 800 adults and juveniles of P. brachyurus. The nematode suspension was calibrated to 400 nematodes mL−1 and placed into two evenly distributed holes around the grape and maize seedlings.

Maize was evaluated approximately 90 days after inoculation for nematode extraction. After that, each pot received a new plant to be evaluated in 180 days for grapes and a second 90 days for maize again. The root systems of grape and maize plants were collected for nematode extraction. Nematodes were extracted from the roots using the methodology proposed by Coolen and D’Herde (1972). The samples obtained were evaluated and all the specimens of P. brachyurusin each sample were counted under a stereoscopic microscope, using the Peters's counting slide.

From the evaluations, we determined the root fresh mass, nematodes per gram of root and the reproduction factor (RF = Pf / Pi, where FR, Pf and Pi refer to the reproduction factor, final population and initial population, respectively). The plants were classified as immune (FR = 0), resistant (0 <FR <1) and moderately resistant or susceptible (FR > 1), as described by Oostenbrink (1966). Fresh root mass was determined from the plants were carefully removed to the pots and the separate aerial part of the root system. The root systems were washed in running water, the roots were placed on absorbent paper until water excess was removed, then they were weighed and fresh root mass was obtained.

The interspecific hybrids were also characterized by morphological descriptors. Sixteen qualitative descriptors were used (vegetative stage) along with three quantitative characters related to P. brachyurus resistance, for 19 descriptors (Table 2). Ampelographic characters were described by using OIV descriptors (OIV, 2013) with small modifications. Sixteen parameters were observed and most of them were measured in different organs of the adult plant (Table 2). These characters were observed ten days after inoculations in plants of each plot (three plants), where five young and adult leaves were measured per plant and the statistic mode was calculated for each plant.

Table 2 Descriptors used for the characterization and assessment of a population from the interspecific cross in Vitis spp. UENF, Campos dos Goytacazes, Rio de Janeiro, Brazil, 2016 

Descriptors Description
Young branch: opening in the tip; 1 = closed; 3 = semi-open; 5 = fully open
Young branch: Anthocyanin pigmentation in the tip; 1 = absent or very weak; 3 = weak; 5 = medium; 7 = strong; 9 = very strong
Young leaf: color of the upper limb face; 1 = yellow-green; 2 = green with anthocyanic areas; 3 = clear red-copper; 4 = dark red-copper; 5 = wine-red
Adult leaf: limbsize; 1 = very small; 3 = small; 5 = medium; 7 = large; 9 = very large
Adult leaf: limbshape; 1 = cordiform; 2 = deltoid; 3 = pentagonal; 4 = orbicular; 5 = reniform
Adult leaf: number of lobes; 1 = none; 2 = three; 3 = five; 4 = seven; 5 = above seven
Adult leaf: depth of the upper lateral sinuses; 3 = shallow; 5 = medium; 7= deep
Adult leaf: arrangement of the lobes of the upper lateral sinuses; 1 = open; 2 = closed; 3 = slightly overlapped; 4 = very overlapped
Adult leaf: shape of the basis of the petiolar sinus; 1= concave (in “u”); 2 = straight (in “v”); 3 = convex
Adult leaf: arrangement of the lobes of the petiolar sinus; 1 = fully open; 2 = very open; 3 = half open; 4 = little open; 5 = closed; 6 = slightly overlapped; 7 = half overlapped; 8 = very overlapped; 9 = fully overlapped
Adult leaf: length of teeth; 3 = short; 5 = medium; 7 = long
Adult leaf: teeth length / width ratio; 1 = very small; 3 = small 5 = medium; 7 = large; 9 = very large
Adult leaf: teethshape 1 = both sides are concave; 2 = both sides are rectilinear; 3 = both sides are convex; 4 = one side is concave and the other is convex; 5 = combination of both sides are rectilinear with both sides are convex
Adult leaf: anthocyanin pigmentation of major veins on upper limb face 1 = absent or very weak; 3 = weak; 5 = medium; 7 = strong; 9 = very strong
Adult leaf: length of petiole in relation to central vein; 4 = shorter; 5 = equal; 6 = longer
Adult leaf: density of erect hairs on the petiole 1 = absent or very low; 3 = low; 5 = medium; 7 = high; 9 = very high
Root Mass (g) Total mass root system of plants 180 days after inoculation
Reproduction Factor Ratio between final population and initial population of nematodes
Nematodes per gram of root 10 g of roots were weighed and separated and chopped to determine nematode numbers

Ward-MLM multivariate analysis

The quantitative and qualitative variables were analyzed simultaneously, using the Ward-MLM procedure to compose genotype groups, using the procedure CLUSTER and IML in SAS program (Statistical Analysis System, version 9.2). The distance matrix was obtained by the Gower algorithm (Gower, 1971) using the Ward grouping method. The ideal number of groups was defined according to the pseudo-F and pseudo-t2 criteria, combined with the likelihood profile and the likelihood ratio test.

The Gower dissimilarity index was used because the set of variables in the study form a mixed group of qualitative and quantitative traits in which a single index of dissimilarity is generated, ranging from 0 to 1. Finally, a complete MLM analysis was carried out for the number of groups (g) defined, which described the classification results on a table presenting the groups formed, while a canonical analysis was used for the quantitative variables. To obtain the canonical variables, the CANDISC procedure in SAS program was used, which contained the canonical coordinates for observations (Crossa and Franco, 2004).

Estimates of genetic parameters and individual selection using the REML/BLUP procedure

To select the representative value of a clone, genotypic values for each quantitative trait were obtained using the REML (Restricted Maximum Likelihood) procedure and applied to the mixed linear model in its matrix form to evaluate related clones in randomized block designs with several plants per plot: Y = Xr + Za + Zd + Wp + e. Where: Y is the data vector; r= is the vector of repetition effects (assumed to be fixed) added to the general average; a= is the vector of the additive genetic effects (assumed to be random); d= is the vector of the dominance genetic effects (assumed to be random); p = is the vector of plot effects (assumed to be random); e = is the vector of errors or residuals (assumed to be random). Capital letters refer to incidence matrices for these effects.

The following genetic parameters were estimated: additive genetic variance ( σa2 ), dominance genetic variance ( σd2 ), environmental variance between plots ( σparc2 ), residual (environmental) variance ( σe2 ), individual phenotypic variance ( σf2 ), narrow sense individual heritability ( h^a2 ), individual heritability of dominance effects ( h^d2 ), broad sense individual heritability ( h^g2 ), coefficient of determination of plot effects ( cparc2 ) and overall average of the experiment.

The structures of averages and variances are given by:


where: V=ZAσa2Z'+ZDσd2Z'+WIσc2+Iσe2

Mixed Model Equations:




σd2 and ha2 : Genetic variance of dominance and broad sense heritability, respectively; D: matrix of genetic correlation of dominance between the individuals assessed.

The system provided predictions of the additive ( a^ ) and dominance ( d^ ) effects isolatedly. The total genotypic values, given by g^=a^+d^ , can be predicted directly from the equations of mixed models:


where: G=σa2+Dσd2

Iterative estimators of variance components by REML via EM algorithm:

σ^e2=[y'yb^'X'y=a^'Z'yd^'Z'yc^'W'y]/[Nr(x)]σ^c2=[c^'c^+σ^e2trC44]/s ;σ^a2=[a'A1a^'++σ^e2tr(A1C22)]/q;σ^d2=[d^'D1d^+σ^e2tr(D1C22)]/q.


Ward-MLM multivariate analysis

The Ward-MLM classification strategy, which simultaneously used quantitative traits related to resistance to P. brachyurus and qualitative, such as morphological and agronomic descriptors, defined the optimal number in three groups and was capable of discriminating 57 interspecific hybrids (Figure 1). The likelihood function was used to determine the ideal number of groups, with an increment value of 66.51 (Figure 1). According to Barbé et al. (2010), the analysis of the likelihood function can define accurate criteria for formation of groups, resulting in less subjective groups.

Figure 1 Logarithmic probability function (Log-likelihood) for the number of groups formed by Ward-Modified Location Model procedure (Ward-MLM) strategy in interspecific hybrids of Vitis spp. UENF, Campos dos Goytacazes, Rio de Janeiro, Brazil, 2016. 

The traits of young shoot tips showed that most hybrids presented opening of shoot tips, with predominance of hybrids presenting absent and weak anthocyanic pigmentation (70 %). Only group II contained four hybrids with strong anthocyanin pigmentation (Table 3).

Table 3 Traits and number of interspecific hybrids per group of multicategoric traits in each of the three groups formed by the Ward-MLM strategy. UENF, Campos dos Goytacazes, Rio de Janeiro, Brazil, 2016 

Descriptors Groups
G I (13) G II (26) G III (18)
Young branch: openness in the tip
Closed 6 3
Open 13 20 15
Young branch: anthocyanin pigmentation of the tip
Absent or weak 11 16 13
Medium 2 6 5
Strong 4
Young Leaf: color of the upper limb face
Yellow-green 1 6 6
Green with anthocyanicareas 10 19 11
Light copperred 1 1 1
Darkcopperred 1
Adult leaf: limbshape
Cordiform 3 2 2
Deltoid 8 21 13
Pentagonal 2 3 3
Adult leaf: number of lobes
None 4
Three 9 11 11
Five 14 7
Above seven 1
Adult leaf: depth of the upper lateral sinuses
Shallow 2 5 7
Medium 6 12 7
Deep 5 9 4
Adult leaf: arrangement of the upper lateral sinuses
Open 10 17 12
Closed 3 8 5
Very overlapped 1 1
Slightly overlapped
Adult leaf: shape of the basis of the petiolar sinus
Concave (in “u”) 5 7 1
Rectilinear (in “v”) 6 10 10
Convex 2 9 7
Adult leaf: arrangement of the lobes of the petiolar sinuses
Fully open 9 10 12
Half open 2 13 5
Slightly open 1 1 1
Closed 1 2
Adult leaf: tooth length
Short 9 13 14
Medium 3 10 3
Long 1 3 1
Adult leaf: tooth length/tooth width ratio
Short 2 6 3
Medium 10 17 14
Large 1 3 1
Adult leaf: size of the limb
Verysmall 1 4
Small 7 13 13
Medium 3 8 1
Large 2 2
Verylarge 1 2
Adult leaf: shape of the teeth
Both sides are concave 2 11 5
Both sides are rectilinear 3 12 9
Both sides are convex 8 1 4
One side is concave and the other is convex 2
Rectilinear sides with convex sides
Adult leaf: Anthocyanin pigmentation of the major veins in the upper limb
Absentorweak 7 20 15
Medium 5 3 2
Strong 1 3 1
Adult leaf: length of the petiole in relation to the central vein
Shorter 7 19 12
Equal 5 6 4
Longer 1 1 2
Adult leaf: Density of erect hairs on the petiole
Absent or low 8 17 16
Medium 4 4 2
High 1 5

Considering the traits evaluated in the adult leaves, hybrids of group I presented between zero and three lobes, whereas hybrids of groups II and III had predominantly between three and five lobes (91 %). Limb-shaped leaves were predominantly adeltoid (73 %) with upper lateral sinus ranging from shallow to deep. For the base shape of the petiole sinuses, most hybrids in each group presented leaves of the rectilinear type. Hybrids of groups I and II had leaflet size ranging from small to very large, while group III allocated hybrids with predominantly small leaf limbs (13 hybrids) (Table 3).

For the traits evaluated in adult leaves, hybrids had zero to five lobes, adeltoid-shaped leaves, shallow-to-deep superior lateral sinuses, petiolar sinuses ranging from concave, convex to rectiliniear, and limb sizes ranging from very small to very large (Table 3).

Regarding the variables related to resistance to P. brachyurus for the susceptible control, maize ‘BR 106’, the average values obtained for root mass, number of nematodes per gram of root and reproduction factor were 74.87, 86.58 and 29.40, respectively, confirming viability of the inoculum used in this work (Table 6).

Wide variation in the quantitative variables related to P. brachyurus resistance was detected among the hybrids evaluated (Table 4). Group I had 13 genotypes with the highest values for reproduction factor and nematodes per gram of root, but lower values for root mass. The minimum value for the reproduction factor of this group was 4.09 and the number of nematodes per gram of roots was 21.74. All hybrids were susceptible to high nematode reproduction rates and reproduction factors above 1.0. The susceptible hybrids had the highest values for this trait. One hybrid in this group was from cross V. romanetii C166-043 × 07355-075, two from cross 06354-047 × Cereza, and ten from cross 06354-047 × Nocera.

Table 4 Minimum and maximum values, average and standard deviation of quantitative traits for each of the three groups and coefficient of correlation of quantitative variables with the first two canonical variables (CAN). UENF, Campos dos Goytacazes Rio de Janeiro, Brazil, 2016 

Variables Parameters Groups CAN
G I (13) G II (26) G III (18) CAN 1 CAN 2
Root Mass Minimum 65.40 71.00 100.50 1.88 1.94
Maximum 97.00 133.20 134.50
Average 79.80 100.93 120.0
Standard Deviation 9.19 15.55 10.20
Reproduction Factor Minimum 4.09 0.71 0.35 -1.65 -6.55
Maximum 14.22 9.25 7.68
Average 8.34 5.04 2.69
Standard Deviation 3.38 2.60 2.11
Nematodes per gram of root Minimum 21.74 3.87 1.33 0.33 7.32
Maximum 72.00 33.64 22.80
Average 41.37 19.25 8.82
Standard Deviation 14.99 8.86 6.56

Most genotypes (26 hybrids) were clustered in Group II. Five were obtained from cross V. romanetii C166-043 × 07355-075; five, from cross 06354-047 × Cereza; and 16, from cross 06354-047 × Nocera. Some resistant genotypes had reproduction factors equal to 0.7. None of the hybrids was considered highly resistant to P. brachyurus, given that nematodes were found in root systems of these hybrids and their reproduction factors were higher than zero (Table 4).

Group III had 18 genotypes, five were resistant with the minimum reproduction factor of 0.35. Most hybrids in this group belong to 06354-047 × Nocera population (15 hybrids) and the other hybrids, from crosses 06354-047 × Cereza (two hybrids) and Vitis romanetii C166-043 × 07355-075 (one hybrid). Groups II and III were the most homogeneous, with an average reproduction factor value of 5.04 and 2.69, and low standard deviation estimate of 2.60 and 2.11, respectively. The most resistant genotypes with the highest estimates for root mass (100.93 and 120.0 g) and the smallest nematode populations (19.95 and 8.82), respectively, were found in these groups. Hybrids with the smallest number of nematodes per gram of root and, consequently, lower reproduction factor, also exhibited a more developed root system, a trait evaluated by root mass (Table 4).

The first two canonical variables (CAN) obtained through the Ward-MLM methodology explained 99 % of the total variation (Figure 2). According to Crossa and Franco (2004), if the first two canonical variables result in estimates above 80 % for the total variation, a satisfactory interpretation of variability of hybrids can be obtained, which was achieved in this study. This allowed an appropriate two-dimensional graphic presentation of relationships between groups and hybrids within the groups.

Figure 2 The first two canonical variables for the three groups formed by the Ward-Modified Location Model procedure (Ward-MLM). UENF, Campos dos Goytacazes, Rio de Janeiro, Brazil, 2016. 

Based on the first canonical variable, the traits that mostly contributed to the quantification of genetic divergence were root mass, with 1.88, and reproduction factor, −1.65 (Table 4). A separation between Groups I and III formed by the Ward-MLM procedure and approximation of Groups I and II and between Groups II and III was observed (Table 5 and Figure 2). Groups II and III include resistant hybrids with the smallest nematode populations in the roots.

Table 5 Distance between the groups formed by the Ward-MLM procedure (proposed by Franco et al., 1998). Data analyzed at UENF, Campos dos Goytacazes, Rio de Janeiro, Brazil, 2016 

Groups Group I Group II Group III
Group I 37.83 87.88
Group II 37.83 36.11
Group III 87.88 36.11

During the experiment, susceptible plants presented symptomatic reactions, such as necrotic lesions of various sizes and colors in the vine roots infested with P. brachyurus. In addition, several symptoms were detected on shoots of susceptible plants, including loss of vigor, varying plant size, yellowing and premature leaf drop, wilt during the hottest hours of the day, slow growth, necrotic leaves and plant death due to its inability to produce an adequate root system. Such symptomatic reactions are typical of the genus Pratylenchus in grapevines (Zheng et al., 2010; Zasada et al., 2012).

Estimates of genetic parameters and individual selection using REML / BLUP procedure

Estimates of coefficient of broad sense heritability that captured the total genotypic effects were higher than 74 % for root mass, reproduction factor and nematodes per gram of root, especially for the reproduction factor (84 %) having the highest value (Table 6). The high values for these traits show great potential for genetic progress in the selection of individuals resistant to P. brachyurus.

Table 6 Estimates of variance components and genetic parameters for variables: root mass (g), reproduction factor, and nematodes per gram of root. Data from 57 clones of Vitis spp. interspecific populations selected for resistance to nematode P. brachyurus. Data acquired and analyzed at UENF, Campos dos Goytacazes, Rio de Janeiro, Brazil, 2016 

Variancecomponents (Individual REML)* Traits
Root Mass Reproduction Factor Nematodes per root gram
σa2a 13.1238 0.5622 11.2973
σa2b 401.5690 24.7453 500.8805
σplotsc2 0.0955 0.0167 0.2839
σe2d 141.4035 4.4519 114.3441
σf2e 556.1918 29.7760 626.8058
h^a2f 0.0236 0.0188 0.0180
h^g2g 0.7456 ± 0.1495 0.8499 ± 0.1596 0.817 ± 0.1565
h^d2h 0.7220 0.8310 0.7991
Cplotsi2 0.002 0.006 0.005
Overall meanpopulation 104. 0917 6.1129 19. 5720
BR 106 mean 74.87 29.40 86.58

*REML = Restricted Maximum Likelihood / Best Linear Unbiased Prediction; aEstimates of additive genetic variance; bdominance genetic variance; cenvironmental variance between plots; dresidualvariance; eindividual phenotypic variance; findividual narrow-sense heritability; gindividual broad-sense heritability; hindividual heritability of the dominance effects; icoefficient of determination of the plot effects.

Estimates of genetic variances were primarily derived from dominance effects for root mass (401.57), reproduction factor (24.745) and nematodes per gram of root (500.881). Consequently, the individual heritability of dominance was increased, ranging from 0.722 to 0.831. Such values are close to estimates of broad sense heritabilities (Table 6). The narrow sense individual heritabilities were low for the three traits, ranging from 0.018 to 0.024, as evidenced by low estimates of additive variances, reaching values up to 13.124 for root mass (Table 6).

Considering phenotypic variances of root mass (556.192), reproduction factor (29.776) and nematodes per gram of roots (626.806), dominance variances were higher than residuals, ranging from 4,452 (reproduction factor) to 141,404 (root mass). These values corroborate that these conditions are favorable for selection and demonstrate that most parts of the phenotypes are attributed to genetic causes of dominance effects (Table 6).

Determination coefficients of plot effects ranged from 0.002 to 0.006 for all traits. The low values of these coefficients are attributed to very small environmental variations within the plots and indicate good experimental precision, corroborated by low estimates of environmental variance between the plots, ranging from 0.096 to 0.284 (Table 6).

A significant different was observed for genotypic values of hybrids for the reproduction factor of nematodes, which ranged from 0.63 to 18.48 (Table 7). The following hybrids stand out for the selection of resistant individuals based on their genotypic values (u + g): CH3.2, CH3.23, CH3.8, CH3.37, CH3.38, CH3.41 and CH3.36 from cross 06354-047 × Nocera; CH2.1 and CH2.7, from cross 06354-047 × Cereza; and CH1.1, CH1.3 and CH1.2 from cross 06354-047 × Nocera.

Table 7 Arrangement of 57 interspecific hybrids of Vitis spp. for resistance to nematode P. brachyurus: where (a) refers to predicted additive effects, (d) predicted dominance effects, (g) predicted genotypic effects, (u+g) genotypic average, genetic gain, and new average or genotypic values for trait reproduction factor in the selection of hybrids. Data acquired and analyzed at UENF, Campos dos Goytacazes, Rio de Janeiro, Brazil, 2016 

Order Hybrids a d g u + g Gain New Average
1 CH1.2 -0.0962 -5.3813 -5.4774 0.6355 0.0063 6.1193
2 CH3.36 -0.0700 -5.3257 -5.3956 0.7173 0.1043 6.2172
3 CH2.7 -0.0631 -5.1453 -5.2083 0.9046 0.2043 6.3172
4 CH3.38 -0.0663 -5.0817 -5.1479 0.965 0.3045 6.4174
5 CH1.3 -0.0905 -5.003 -5.0935 1.0194 0.4074 6.5203
6 CH3.37 -0.0642 -4.9428 -5.0069 1.106 0.5132 6.6261
7 CH1.1 -0.0891 -4.911 -5.0001 1.1129 0.6214 6.7343
8 CH3.8 -0.0633 -4.8855 -4.9488 1.1641 0.7338 6.8468
9 CH2.1 -0.0584 -4.8349 -4.8932 1.2197 0.8498 6.9627
10 CH3.23 -0.0605 -4.7036 -4.7641 1.3488 0.9694 7.0824
11 CH3.2 -0.0597 -4.6493 -4.7091 1.4039 1.0914 7.2044
12 CH3.41 -0.0493 -3.9642 -4.0136 2.0993 1.2175 7.3305
13 CH3.6 -0.0423 -3.499 -3.5413 2.5716 1.3338 7.4467
14 CH3.1 -0.0371 -3.1594 -3.1966 2.9164 1.4446 7.5575
15 CH3.9 -0.0358 -3.0694 -3.1052 3.0077 1.5525 7.6654
16 CH3.5 -0.0333 -2.9085 -2.9418 3.1711 1.6634 7.7763
17 CH3.13 -0.031 -2.7517 -2.7827 3.3302 1.7757 7.8887
18 CH3.16 -0.0259 -2.4205 -2.4464 3.6665 1.8897 8.0026
19 CH3.32 -0.0255 -2.3936 -2.4192 3.6937 2.0009 8.1138
20 CH3.3 -0.0236 -2.2642 -2.2878 3.8252 2.1172 8.2301
21 CH3.30 -0.0215 -2.1252 -2.1467 3.9663 2.2363 8.3492
22 CH2.2 -0.0166 -2.0788 -2.0954 4.0175 2.358 8.4709
23 CH3.24 -0.0119 -1.4905 -1.5024 4.6106 2.4852 8.5982
24 CH3.34 -0.0081 -1.2407 -1.2488 4.8641 2.6025 8.7155
25 CH3.10 -0.0066 -1.1472 -1.1539 4.9591 2.7192 8.8322
26 CH3.27 -0.0041 -0.9821 -0.9863 5.1267 2.8403 8.9532
27 CH3.25 -0.0031 -0.916 -0.9192 5.1938 2.9637 9.0766
28 CH3.12 0.0041 -0.4376 -0.4335 5.6794 3.0931 9.2061
29 CH3.7 0.0046 -0.4028 -0.3981 5.7148 3.2147 9.3277
30 CH3.28 0.0048 -0.3936 -0.3889 5.7241 3.3438 9.4567
31 CH3.35 0.0054 -0.3493 -0.3439 5.7691 3.482 9.5949
32 CH3.20 0.0055 -0.3467 -0.3412 5.7717 3.6292 9.7421
33 CH3.14 0.0143 0.2382 0.2526 6.3655 3.788 9.9009
34 CH2.9 0.0187 0.2505 0.2692 6.3821 3.9353 10.0482
35 CH2.6 0.0202 0.3503 0.3705 6.4834 4.0947 10.2076
36 CH2.5 0.0264 0.7609 0.7873 6.9003 4.264 10.3769
37 CH1.4 -0.0023 0.8154 0.8131 6.9261 4.4295 10.5424
38 CH3.18 0.0243 0.8952 0.9195 7.0324 4.6103 10.7233
39 CH3.31 0.0273 1.0956 1.1229 7.2359 4.8046 10.9175
40 CH3.4 0.0299 1.2661 1.296 7.409 5.0091 11.1221
41 CH3.22 0.0371 1.7385 1.7756 7.8885 5.2275 11.3405
42 CH1.5 0.0128 1.8131 1.8259 7.9388 5.4433 11.5562
43 CH1.3 0.0258 2.6683 2.6941 8.8071 5.6845 11.7974
44 CH3.15 0.0532 2.7998 2.853 8.9659 5.898 12.011
45 CH3.11 0.0542 2.8685 2.9227 9.0356 6.1323 12.2452
46 CH2.4 0.0599 2.9672 3.0271 9.14 6.3997 12.5127
47 CH3.26 0.0601 3.256 3.316 9.429 6.7064 12.8193
48 CH3.17 0.0765 4.3423 4.4188 10.5317 7.0454 13.1583
49 CH3.29 0.0779 4.4319 4.5098 10.6227 7.3372 13.4502
50 CH3.33 0.082 4.702 4.7839 10.8969 7.6907 13.8036
51 CH3.39 0.0863 4.9862 5.0725 11.1854 8.1059 14.2188
52 CH1.7 0.0636 5.1635 5.2271 11.34 8.6115 14.7244
53 CH2.3 0.1144 6.5668 6.6812 12.7941 9.2884 15.4013
54 CH2.8 0.1266 7.3685 7.4951 13.608 9.9401 16.0531
55 CH3.19 0.1489 9.117 9.2659 15.3788 10.7552 16.8681
56 CH3.40 0.1692 10.4602 10.6295 16.7424 11.4998 17.6127
57 CH3.21 0.1952 12.1749 12.3702 18.4831 12.3702 18.4831
BR 106


Based on the pseudo-F and pseudo-t2 criteria, the likelihood function defined that the ideal number of homogeneous groups is three. These groups were formed according to their similarities, which is mainly due to the greater increase in the likelihood function observed for group III, that is, 65.54 (Figure 1).

The likelihood function was used to define more accurate criteria for the formation of the three groups, which resulted in more objective groupings than the hierarchical grouping methods in which the number of groups is established in a more subjective manner (Gonçalves et al., 2008; Kurosawa et al., 2017).

For most hybrids allocated to each group formed, there was predominance of hybrids with leaves with weak anthocyanin pigmentation in the main veins of the upper limb. However, 35 % of the hybrids had pigmentation varying from medium to strong levels. Similarly, 29 % of these hybrids had anthocyanin pigmentation varying from medium to strong levels in the young branches (Table 3). In red grapes, high concentration of anthocyanins in leaves and branches are important for production of juice and red wine, because anthocyanins contribute to sensory attributes and mainly to color and flavor. However, quantity and composition of these anthocyanins differ according to species, variety, phenological stage and climate (Muñoz-Espada et al., 2004).

The knowledge of morphological traits assessed in this work (Table 2, Figure 2) is fundamental for plant breeding programs. These traits may not play a direct role in selection and development of new wine grapes; however, they do contribute to a better characterization of genetic diversity, exploration of genetic variability and subsequent conservation of species.

Vitis hybrids were separated into three groups according to their genotypic values that reactto P. brachyurus. Regarding the trait reproduction factor (RF): 46 genotypes were susceptible (RF ≥ 1); seven were moderately resistant (1.0 ≥ RF < 1.5); and four, resistant (RF ≤ 1). Eleven moderately resistant and resistant hybrids selected corresponded to 19 % of the total segregating populations (Table 7). Resistant hybrids derived from crosses that used 06354-047 as a parent, indicating that it may be the source of resistance to P. brachyurus.

Groups I and II had resistant hybrids; however, neither was immune to nematodes. The hybrids selected in our study have a dual application, that is, both for direct use as rootstocks or development of fruiting varieties.

Heritability is one of the most important genetic parameters, as it quantifies the fraction of the inheritable phenotypic variation available for selection. Nematodes per gram of root and root mass allow the best genetic progress of broad sense heritability for the traits of reproduction factor. In addition, since the results found that these traits are correlated, there is a good chance of success in advancing all three characters through selection of the reproduction factor alone (Santos et al., 2017; Santos et al., 2015;Kouassi et al., 2009). The sexual reproduction of these hybrids is not likely to provide significant genetic gains for these traits in grape breeding. However, genetic gains can be maximized (Kouassi et al., 2009; Piepho et al., 2009) by selecting hybrids for lower reproduction factor and nematodes per gram of root as well as by cloning the material selected. The number of nematodes per gram of fresh roots is a good parameter to assess nematode populations, since it correlates directly with losses caused by nematodes (Starr et al., 2002).

In this work, the conditions are favorable for selection since most phenotypes are attributed to genetic causes. The total phenotypic variances tended towards zero, due to the environmental effect between plots and determination coefficients of plot effects. Thus, it can be concluded that there is a small magnitude effect of plots and good selection efficiency regarding experimental precision for reproduction factor, root mass and nematode per gram of root (Santos et al., 2017).

For selection, using the average components predicted by BLUP, plant breeders should prioritize genotypic values, as these are necessary predicted true values (Piepho et al., 2009). Thus, selected hybrids should have average reproduction factors of up to 1.40 (Table 7).The use of reproduction factor for selection demonstrated that most hybrids are susceptible since they had reproduction factor values higher than 1.0. However, for several hybrids, the genetic values (u + g) for reproduction factor ranged from1.0 to2.0. In these cases, it is advisable to consider susceptibility as moderate resistance, since they are genotypes that allow nematode survival, without showing strong symptoms of their feeding. The same is applied to resistance found in hybrid CH3.38, whose genetic value was slightly less than 1.0 for reproduction factor.

On the other hand, several hybrids, with reproduction factor values above or very close to 2.0 should be considered susceptible. These individuals are efficient hosts and favor the establishment of P. brachyurus populations that are high enough to cause significant direct damage to grapevine growth and prevent their use in breeding programs.

Narrow sense heritability for root mass, nematodes per gram of root and reproduction factor were less than 2 % or almost null for these traits. This reveals that for grapevine breeding, selection for resistance to P. brachyurus by sexual reproduction in a population composed of these clones does not lead to genetic gains and is therefore impracticable. While in the broad sense, considering the additive and dominant variances, where heritabilities ( h^g2g ) reached values ranging from 74 to 84 %, meaning that selection based on these traits and cloning the selected material can be maximized the genetic gains. In other words, broad sense heritability relative to traits of root mass, nematodes per gram of root and reproduction factor had the greatest genetic advances associated to the cloning of selected genotypes for those traits. The individuals selected for resistance in this study had the lowest genotypic values, with predicted gains below 1 %. Hybrid CH1.2 had the lowest genotypic value for reproduction factor (u + g = 0.6355) and genetic gain close to zero (0.0063). This indicates that genotypes with lower values for reproduction factor can be cloned and advanced.

The use of resistant grapevines, along with other measures of nematode control, decreases P. brachyurus populations in the soil and increases commercial yield of vineyards. Resistant clones can now be evaluated for use as fruiting varieties or as commercial rootstocks (Walker etal., 1994).


The Ward-MLM classification strategy effectively discriminated among the variable genotypes and allowed the separation of 57 genotypes into three homogeneous groups. Groups II and III were the closest genetically and contained hybrids resistant to P. brachyurus. High estimates of dominance variance suggest that further genetic progress can be achieved through the cloning of resistant hybrids, more than their use as resistant parents. Based on the genotypic values for reproduction factor, the following resistant and moderately resistant clones were selected: CH3.2, CH3.23, CH3.8, CH3.37, CH3.38, CH3.41, CH3.36, CH2.1, CH2.7, CH1.1, CH1.3 and CH1.2. The hybrids with the lowest nematode populations per gram of root and lower breeding capacity for P. brachyurus resistance should be tested as rootstocks and evaluated for use in infested areas.


We are thankful to FAPERJ – Fundação de Amparo à Pesquisa do Estado do Rio de Janeiro for the granting and CAPES, of the doctoral scholarship and financial support provided to the first author, as well as to the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) for the accomplishment of the present work.


Barbé, T.C.; Amaral Júnior, A.T.; Gonçalves, L.S.A.; Rodrigues, R.; Scapim, C.A. 2010. Association between advanced generations and genealogy in inbred lines of snap bean by the Ward-Modifie Location Model. Euphytica 173: 337-343. [ Links ]

Coolen, W.A.; D’Herde, C.J. 1972. A method for the Quantitative Extraction of Nematodes from Plant Tissue. State Agricultural Research Centre, Ghent, Belgium. [ Links ]

Crossa, J.; Franco, J. 2004. Statistical methods for classifying genotypes. Euphytica 137: 19-37. [ Links ]

Ferris, H.; Zheng, L.; Walker, M.A. 2012. Resistance of grape rootstocks to plant-parasitic nematodes. Journal of Nematology 44: 377-386. [ Links ]

Food and Agriculture Organization [FAO]. 2016. FAOSTAT. FAO, Rome, Italy. Available at: [Accessed Mar 28, 2018] [ Links ]

Franco, J.; Crossa, J.; Villasenõr, J.; Taba, S. 1998. Classifying genetic resources by categorical and continuous variables. Crop Science 38: 1688-1696. [ Links ]

Gonçalves, L.S.; Rodrigues, R.; Amaral Júnior, A.T.; Karasawa, M.; Sudré, C.P. 2008. Comparison of multivariate statistical algorithms to cluster tomato heirloom accessions. Genetic and Molecular Research 7: 1289-1297. [ Links ]

Gower, J.C. 1971. A general coefficient of similarity and some of its properties. Biometrics 27: 857-874. [ Links ]

Brazilian Institute of Geography and Statistics [IBGE]. 2016. Agricultural Census. = Instituto Brasileiro de Geografia e Estatística. Censo Agropecuário. Available at: [Accessed Mar 28, 2018] (in Portuguese). [ Links ]

International Organisation of Vine and Wine [OIV]. 2013. International list of vine varieties and their synonyms. OIV, Paris, France. Available at: [Accessed 27 Mar, 2018] [ Links ]

Kouassi, A.B.; Durel, C.; Costa, F.; Tartarini, S.; Weg, E.V.; Evans, K; Fernandez-Fernandez, F.; Govan, C.; Boudichevskaja, A.; Dunemann, F.; Antofie, A.; Lateur, M.; Stankiewicz-Kosyl, M.; Soska, A.; Tomala, K.; Lewandowski, M.; Rutkovski, K.; Zurawicz, E.; Guerra, W.; Laurens, F. 2009. Estimation of genetic parameters and prediction of breeding values for apple fruit-quality traits using pedigreed plant material in Europe. Tree Genetics and Genomes 5: 659-672. [ Links ]

Kurosawa, R.N.; Amaral Junior, A.T.; Silva, F.H.; Santos, A.; Vivas, M.; Kamphorst, S.H.; Pena, G.F. 2017. Multivariate approach in popcorn genotypes using the Ward-MLM strategy: morpho-agronomic analysis and incidence of Fusarium spp. Genetic and Molecular Research 16: 1-12 [ Links ]

Muñoz-Espada, A.C.; Wood, K.V.; Bordelon, B.; Watkins, B.A. 2004. Anthocyanin quantification and radical scavening capacity of Concord, Norton, and Marechalfoch grapes and wines. Journal of Agricultural Food Chemistry 52: 6779-6786. [ Links ]

Oostenbrink, M. 1966. Major Characteristics of the Relation Between Nematodes and Plants. Beltsville, MD, USA. [ Links ]

Piepho, H.P.; Möhring, J.; Melchinger, A.E.; Büchse, A. 2009. BLUP for phenotypic selection in plant breeding and variety testing. Euphytica 161: 209-228. [ Links ]

Puerari, H.H.; Dias-Arieira, C.R.; Moura, M.F; Biela, F.; Chiamolera, F.M.; Cunha, T.P.L. 2012. Reaction of grape vine rootstocks to Pratylenchus brachyurus and Pratylenchus zeae. Tropical Plant Pathology 37: 220-222. [ Links ]

Santos, E.A.; Viana, A.P.; Freitas, J.C.O.; Rodrigues, D.L.; Tavares, R.F.; Paiva, C.L.; Souza, M.M. 2015. Genotype selection by REML/BLUP methodology in a segregating population from an interspecific Passiflora spp. crossing. Euphytica 204: 1-11. [ Links ]

Santos, P.R.; Preisigke, S.C.; Viana, A.P.; Cavalcante, N.R.; Bezerra, C.M.; Amaral Júnior, A.T. 2017. Associations between vegetative and production traits in guava tree full-sib progenies. Pesquisa Agropecuária Brasileira 52: 303-310. [ Links ]

Starr, J.L.; Cook, R.; Bridge, J. 2002. Plant resistence to parasitic nematodes. CABI, Wallingford, England. [ Links ]

Téliz, D.; Landa, B.B.; Rapoport, H.F.; Pérez Camacho, F.; Jiménez Díaz, R.M.; Castillo, P. 2007. Plant-parasitic nematodes infecting grapevine in southern Spain and susceptible reaction to root-knot nematodes of rootstocks reported as moderately resistant. Plant Disease 91: 1147-1154. [ Links ]

Viana, A.P.; Riaz, S.; Walker, M.A. 2011. Evaluating genetic diversity and optimizing parental selections in a segregating table-grape population. American Journal of Enology and Viticulture 62: 285-290. [ Links ]

Walker, M.A.; Ferris, H.; Eyre, M. 1994. Resistance in Vitis and Muscadiniaspecies to Meloidogyne incognita. Plant Disease 78: 1055-1058. [ Links ]

Walker, M.A.; Aguero, C.; Hwang, C.F.; Zheng, L.; Ferris, H.; Ng, D. 2010. Progress toward the breeding of rotundifolia-based rootstocks. American Journal of Enology and Viticulture 61: 4-31. [ Links ]

Zasada, I.A.; Riga, R.; Pinkerton, J.N.; Wilson, J.H.; Schreiner, R.P. 2012. Plant-parasitic nematodes associated with grapevines, Vitis vinifera, in Washington and Idaho. American Journal of Enology and Viticulture 63: 522-528. [ Links ]

Zheng, L.; Ferris, H; Walker, A.M. 2010. Interactions between nematodes and grape rootstocks. Journal of Nematology 42: 278-279. [ Links ]

Received: December 07, 2017; Accepted: May 18, 2018

*Corresponding author <>

Edited by: Leonardo Oliveira Medici

Authors’ Contributions

Conceptualization: Santos, P.R.; Viana, A.P.; Gomes, V.M.; Walker, M.A. Data acquisition: Santos, P.R.; Preisigke, S.C.; Almeida, O.F.; Rodrigues, D.L.; Data analysis: Santos, P.R.; Viana, A.P.; Design of Methodology: Santos, P.R.; Viana, A. P.; Gomes, V.M.; Rodrigues, R.; Software development: Santos, P.R.; Viana, A.P.; Writing and editing: Santos, P.R.; Viana, A.P.; Santos, E.A.; Rodrigues, R.; Walker, M. A.

Creative Commons License This is an Open Access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.