Snap beans for organic farming and evaluation of resistance to the common bacterial blight

Snap beans have been widely used in organic farming as a good income source and an alternative to diversify production, with increasing use in crop rotation. This work reports the evaluation of 25 bush-type snap beans accessions for their suitability to integrate a breeding program for organic farming, as well as for their resistance to the common bacterial blight (CBB). Agronomic performance was assessed in two field experiments (September-December, 2013; April-June, 2014), in complete blocks at random, while resistance to CBB was assessed in greenhouse, in a completely randomized trial. Plants were challenged with two isolates, one from Xanthomonas axonopodis pv. phaseoli and another from X. fuscans subsp. fuscans. Accessions UEL 402, UEL 405, UEL 407, UEL 408, UEL 412, UEL 417 and UEL 420 were highly productive in both seasons (averages of 10.3, 8.7, 9.5, 9.2, 8.9, 9.3 and 9.2 t/ha, respectively), and are promising for use both as cultivars by organic farmers in the region of Londrina and also as germplasm in breeding programs for developing cultivars adapted to the region. Although all accessions were moderately susceptible to CBB, accessions UEL 407, UEL 409, UEL 411, UEL 412, UEL 424 and UEL 431 presented the lowest values for the area under CBB progress curve for both isolates.


Received on June 8, 2016; accepted on February 21, 2017
expanding steadily in last decades in several countries (Willer & Kilcher, 2012).In Brazil, 10,505 farmers currently meet official organic farming requirements, a 56% increase when compared to 2013, when not more than 6.7 thousand farmers were accredited as organic producers (MAPA, 2015).
Vegetable production is pioneer in organic farming.Even though, it still faces difficulties as, for instance, lack of enough organic seeds to meet the certification process throughout the production chain.In addition, cultivars meant for organic farming should be developed according to agroecological principles, using germplasm adapted to local conditions.Very often, in comparative experiments between conventional and organic systems, cultivars with good performance in the first do not necessarily perform well in the second (Kirk et al., 2012, Spagnuolo et al., 2016).In organic farming, the ideal cultivar is expected to be rustic, resistant to pests and diseases and reasonably productive in the absence of soluble fertilizers (Vidal et al., 2007).
Among snap beans diseases, the common bacterial blight (CBB), caused by Xanthomonas axonopodis pv.phaseoli (Xap) or X. fuscans subsp.fuscans (Xff) (EFSA Panel on Plant Health, 2014), is considered the main bacterial disease affecting beans in Brazil (Trindade et al., 2012;Silva et al., 2013).Xap and Xff are present in all environments where P. vulgaris is produced, causing damage mainly under high temperature and humidity (Miklas et al., 2003).CBB symptoms are observed throughout plant aboveground part: leaves, stems, pods and seeds (Rava & Sartorato, 1994).At first, leaves show water-soaked lesions that evolve into necrotic spots, usually surrounded by a yellow halo.Adjacent lesions coalesce and, in severe infections, CBB causes leaf death and, consequently, plant defoliation.In pods, lesions are initially circular and soaked, becoming necrotic and reddish.Seeds present discoloration in the thread, spots on the tegument and wrinkling, although in some cases seed infection is symptomless (Bianchini et al., 2005).
CBB must be controlled by employing several management measures simultaneously, since bactericides alone are not efficient.Among recommended measures, the use on non-infested areas, healthy seeds, crop rotation, adequate irrigation systems and resistant cultivars are highlighted (Trindade et al., 2015).Nevertheless, the development of snap beans cultivars with resistance to CBB is a challenge to breeders.Breeding for CCB resistance in snap beans still waits for the identification of resistance sources in either snap beans or common beans, once high levels of resistance to Xanthomonas spp.have not been found in P. vulgaris yet.
The present work describes the screening of accessions of bush-type snap beans from the Germplasm Bank of the Universidade Estadual de Londrina (UEL) for their performance in organic farming and resistance to CBB.

MATERIAL AND METHODS
Twenty-five snap beans accessions from the germplasm bank of the Universidade Estadual de Londrina (UEL) were evaluated.Agronomic traits were assessed in two experiments carried out at UEL School Farm (23°20'23''S, 51°12'32''W, altitude 535 m), in an area kept under organic farming for five years.Experiment I was carried out from September to December 2013 and, experiment II, from April to June 2014.
CBB resistance was evaluated in greenhouse, at the Department of Agronomy (UEL), from December 2013 to February 2014.We used a completely randomized design and with ten replications.Accessions were grown in five-liter pots, filled with a 50%-soil/30%-manure/20%-sand substrate.We planted four seeds per pot, thinned after emergence to two plants per pot.
Inoculation was carried out 35 days after sowing with two bacterial isolates, Xanthomonas axonopodis pv.phaseoli (Xap), and X. fuscans subsp.fuscans (Xff), received from the Biological Institute of São Paulo and from the Agronomic Institute of Paraná, respectively.Isolates were cultured in DYGS medium (Rodrigues Neto et al., 1986) in Petri dishes (streaks made with smear loop) and kept in growth chamber, 28°C for 36 hours.After, bacterial colonies were suspended in 100 mL distilled water and concentration adjusted to 10 7 CFU/mL, using a spectrophotometer at 640 nm absorbance.For inoculation, we took a three-blade leaf per plant and inoculated each bacterial isolate in a separate leaflet.Inoculation consisted of two 2-cm cuts per leaflet made with scissors previously immersed in the two bacterial suspensions.
CBB severity was evaluated daily for 30 days, using the diagrammatic scale of Pastor-Corrales et al. (1981), where: 1= no symptoms; 2= 1 to 5% necrosis; 3= 6 to 25% necrosis; 4= 26 to 50% necrosis; 5= over 50% necrosis.Data were used to calculate the area under the disease progress curve (AUDPC).Data from field and greenhouse experiments were submitted to analysis of variance individually and, subsequently, once homogeneity of residual variances was detected, field experiments were submitted to joint analysis of variance.Means were grouped using the Scott-Knott test, at 5% significance, whenever significant differences were found.Data were analyzed with the aid of the R program (http://www.r-project.org)using the Agricolae and Scott-Knott packages.

RESULTS AND DISCUSSION
The analysis of variance carried out independently for each field experiment indicated the occurrence of significant differences among snap beans genotypes for number of commercial and noncommercial pods per plant and productivity in season I; and for mass of commercial pods and productivity in season II, pointing to a wide variability among snap beans accessions.Accuracy estimates had moderate (≥50<70), high (≥70<90) or very high (≥90) magnitude in all cases, confirming experimental precision (Cargnelutti Filho et al., 2009).Accuracy is a very robust indicator of experimental precision, since the experimental mean does not influence it (Resende & Duarte, 2007).
Cargnelutti Filho et al. (2009) consider selective accuracy more adequate to assess experimental precision in trials with beans than the coefficient of variation or the honest significant difference used on the Tukey test, as percentage of the mean.The F test for the joint analysis of variance carried out considering the two field experiments indicated significant differences for all traits, for both sources of variation, e.g., genotypes and environments (seasons).Genotype x environment interaction (GE) was significant for all traits as well, indicating a differential response from genotypes to the environmental variation.
Agronomic traits averages were higher for the experiment carried out from April to June 2014 (Season II)  (Kirk et al., 2012;Luby et al., 2013).Mason & Spaner (2006) stated that the increase of plant stress in organic farming can affect cultivar performance strongly enough to justify the development of breeding programs specific for organic farming.
No significant differences among accessions were found for NnCP by the Scott-Knott clustering test in season II.Nevertheless, in season I accessions UEL 402, UEL 416, UEL 431, UEL 436, UEL 447, UEL 1, UEL 2 and cultivar Alessa were grouped as having significantly highest NnCP, respectively 52.3, 54.0, 62.7, 65.7, 67.3, 60.0, 57 There are no breeding programs of bush-type snap beans specific for organic farming.In the present work, accessions UEL 402, UEL 405, UEL 407, UEL 408, UEL 412, UEL 417 and UEL 420 have shown to be promising both as cultivars for organic farming in the region and as advanced germplasm in breeding.
The analysis of variance of the area under the disease progress curve (AUDPC) revealed significant differences and wide variability among accessions when inoculated with the two Xanthomonas isolates (Table 2).Average AUDPC values were 82.09

Table 1 .
Number of commercial and non-commercial pods per plant, mass and yield of commercial pods of 25 bush-type snap beans.Londrina, UEL, 2013.
Pods longer than 10 cm with no mechanical or disease damages were considered commercial; Means followed by same capital letters in the line and small letters in the column do not differ significantly from each other by Scott-Knott test, 5%.

Table 2 .
Area under the disease progress curve (AUDPC) for the Common Bacterial Blight in 25 bush-type snap beans accessions inoculated with Xanthomonas axonopodis pv.phaseoli (Xap) and X. fuscans subsp.fuscans(Xff).Londrina, UEL, 2013.Means followed by same capital letters in the column do not differ significantly from each other by Scott-Knott test, 5%.