Sclerotinia stem rot is a major disease of oilseed Brassica, with up to 80% incidence of plants affected in the worst affected crops in the Punjab and Haryana states of Northern India (Kang and Chahal, 2000). In Haryana, 5 to 20% of plants affected by this disease are common, varying with crop growth stage and region (personal communication). Estimated yield losses from Sclerotinia stem rot vary throughout the world, with 13% losses in North Dakota and Minnesota (Lamey and Bradley, 2002);  20% (Fitt et al., 1992) to 50% (Pope et al., 1989) in the UK; 0.4-1.5 tonne/ha losses in Australia (Kirkegaard et al., 2006) and 70% in China (Deng and Tang,  2006).  Sclerotia  of  Sclerotinia  sclerotiorum   can germinate either myceliogenically or carpogenically, lead to stem base infection and aerial infections, respectively. Being a ubiquitous necrotrophic pathogen with many different hosts, Sclerotinia stem rot is difficult to manage. Primary methods of management rely upon use of non-host crops, fungicide application and manipulation of management practices, but all have been proved unreliable and frequently of little if any economic benefit. The variation among genotypes for Sclerotinia stem rot under different environmental conditions was observed by Li et al. (2006). Genetic resistance to Sclerotinia stem rot offers the best long term prospect for making oilseed Brassica crops more  profitable in regions prone to this disease. For this reason, using a field stem inoculation technique, we evaluated oilseed Brassica napus and Brassica juncea genotypes from India, China and Australia for resistance to S. sclerotiorum in the field. Inpresent investigation, 92 genotypes of oilseed Brassica were evaluated over three environments that is, three crop seasons (2009, 2010 and 2011) to identify stable genotypes with Sclerotinia stem rot resistance.

Seed of accessions of B. napus and B. juncea was obtained from Australia, China and India through an Australian Centre of International Agricultural Research (ACIAR) collaborative program between these 3 countries. The B. napus and B. juncea genotypes tested are enlisted in Table 2. Ninety two genotypes of B. napus and B. juncea were tested in the field at the Chaudhary Charan Singh Haryana Agricultural University (CCSHAU), Hisar, India. The germplasms were hand sown on 23^rd October, 2009, 27^th October, 2010, and 25^th Oct 2011 in plots with 5 x 2 m. Row to row and plant to plant spacing was maintained 45 and 15 cm, respectively. All recommended agronomic practices were followed, including 100 kg N/ha, 30 kg P₂O₅ /ha fertilizer application at sowing time, two irrigations at 45 and 90 days after sowing in sandy loam soil having pH 8.0 and electrical conductivity 3.5 mMhos/cm. Each genotype was sown in a complete randomised block design (RBD) with three replicates and used for further study. The single isolate of S. sclerotiorum (‘CCS HAU-Hisar’) used in this study was isolated from sclerotia collected from diseased B. juncea plants stem pith in the previous crop season. Sclerotia were surface sterilized using 0.1% mercuric chloride solution in distilled water, cut in two pieces with sterilized Gilllete platinum blade and plated cut side face down on potato dextrose agar medium in 90 mm Petri® plates under aspetic conditions in laminar flow. Thirty plants of each genotype within each replication were picked at random and inoculated at the flowering stage (when 50% of plants in the row had at least one opened flower). This was stage GS 61 BBCH on the scale of Lancashire et al. (1991) and equivalent to stage GS 41-42 on the scale of Harper and Berkenkamp (1975). Stem inoculation was carried out according to the method of Buchwaldt et al. (2005) in all crop seasons. A single 5 mm diameter agar plug disc cut from the S. sclerotiorum colony edge of 3 to 4 days old culture growing on a glucose rich medium (Glucose 20 g, NaOH 1 g, NH₄ NO₃ 2 g, MgSO₄ 7 H₂O 0.1 g, Malic acid 3 g, KH₂ PO₄ 1 g and Agar 30 g in 1 L of distilled H₂O) was used to inoculate each plant. The agar plug was placed mycelium side up on a small piece of Parafilm® (about 5 cm long). The plug was then placed on to the stem at the first internode above the middle internode of each stem (with the mycelium in contact with stem) by wrapping the Parafilm® strip around the stem to secure the plug onto the stem. A wet cotton wool swab was also wrapped around the stem just above the top of the Parafilm® strip to maintain high humidity during the infection period. The observation on four characters viz. plant age at inoculation (days) time, stem diameter (mm), stem lesion length (mm) and wilted/dead plant (%) were recorded. The statistical analysis for stability was carried out according to the method of Eberhart and Russell (1966).

The pooled analysis of variance (Table 1) revealed that mean sum of squares due to Genotype (G) × Environment (E) interaction tested against pooled error and was found significant for wilted plants (%). It indicated that genotypes and environments not independent in causing variation but also have involvement of G×E interaction in the expression of wilted plant (%). Non significant G×E interaction observed for plant age at inoculation, stem diameter and stem lesion length indicate that these characters are least influenced by the environments. The absence of differential response of the genotypes for plant age at inoculation, stem diameter and stem lesion length in the present investigation indicates the stable expression of these characters.

 

 

Highly significant environmental (linear) variance for all the characters suggested that variation among the environments was linear. A linear environmental variance signifies unit changes in environmental conditions. The G×E (linear) variance was non significant for plant age at inoculation, stem diameter and stem lesion length implying thereby, differential performance of genotypes under diverse environments with nearly uniform reaction norms.

 

On the other hand, non significant pooled deviations for all the characters suggested that performance of different genotypes non-fluctuated significantly from their respective linear path of response to environments (Table 1). In other words, the predictable environments formed the major portion of G×E interactions. Moreover, by observing the individual varietal fluctuation from linearity, it becomes clear that only a very few genotypes fluctuated significantly from linearity. The environmental index (Ij) for all the environments and for all the characters was estimated. A critical analysis revealed that E₃ that is, environmental condition prevailed during 25^th Oct 2011 sown genotype expressed high environmental index for the characters viz. plant age at inoculation, stem diameter, and wilted plant (%) and E₂ that is, environmental condition prevailed during 27^th Oct 2010  sown genotype for stem lesion length (Table 2).

 

 

On the contrary, E₂ exhibited lowest value for plant age at inoculation time, and E₁ that is, environmental condition prevailed during 23^rd 2009 sown genotype exhibited lowest value for stem diameter and stem lesion length (Table 2).

 

Plant age at inoculation (days)

 

On mean basis, 48 genotypes were early in plant age at inoculation (days) time and 44 genotypes were older in plant age. Out of 92 genotypes, only 8 genotypes expressed below average (b_i<1) response, 27 genotypes expressed above average (b_i>1) response and remaining 57 genotypes exhibited average response value of regression coefficient (Table 2). A consideration of the stability parameters together, 57 genotypes (30 below mean and 27 above mean) were average in response (b_i = 1) and good in stability (deviation from regression that is, S²_di=0). 

 

Stem diameter (mm)

 

An examination of individual stability parameter for stem diameter (Table 2) revealed that as many as 43 genotypes had above average mean performance and 49 genotypes had below average mean performance (Table 2). Further, all the genotypes were found to be stable (S²_di=0). Majority of genotypes were having average response for stem diameter (b_i = 1). Only 15 genotypes exhibited below average response (b_i<1) and only one genotype that is, Rivette exhibited above average (b_i>1) response.

 

Stem lesion length (mm)

 

On the basis of mean stem lesion length (mm) it was observed that 42 genotypes exhibited below mean, 6 genotypes average mean and 50 genotypes above mean for stem lesion length. Further, all the genotypes exhibited non significant S²_di value indicating the absence of non linear component of G×E interaction. Out of 92 genotypes,  only  38  genotypes  exhibited   totally absence of G×E interaction having b_i = 1 and S²_di=0, 17 genotypes (b_i > 1) and 55 genotypes (bi <1) exhibited the presence of linear component of G×E interaction (Table 2).

Considering the three stability parameters, simultaneously (high resistance/ small stem lesion, b_i = 1 and S²_di = 0) RH 13, Ringot, Brassica juncea 1, Brassica juncea 2, Brassica juncea 3 and RQ 011 were highly resistant and stable for stem lesion length over the environments or three crop seasons (Table 2). Moreover, Ag outback was highly resistant and suitable for conducive environment (small lesion length, b_i <1, S²_di = 0) for disease development against CCSHAU-Hisar isolate.

 

Wilted/ dead plants

 

An examination of data on wilted /dead plant reflected that 38, 13 and 41 genotypes were below mean, at average mean and above mean, respectively. In majority of genotypes linear components of G×E interaction was noticed, except 31 genotypes which showed  the  absence of linear component of G×E interaction. However, the majority of genotypes (81) exhibited the non significant for S²_di value means absence of non-linear component of G×E interaction (Table 2). Out of top resistant genotypes against Sclerotinia stem rot, only Ag spectrum was found suitable for general environment conditions (highly resistant, b_i = 1, S²_di = 0) and six other (JM018, Ag outback, Monty,  B. juncea 1, B. juncea 2 and B. juncea 3), were suitable for  conducive environment (highly resistant, b_i<1, S²_di = 0) of disease development (Table 2). In contrast to this, JM 3 was highly susceptible to Sclerotinia stem rot; but it was also stable susceptible (S²_di = 0 and b_i = 1).

 

On the basis of environmental index, it was found that E₃ was most conducive environment for disease development. The estimates of three stability parameters namely X, b_i and S²_di (Table 2) revealed that the non significant value of S²_di indicating thereby the totally absence of nonlinear component of G×E interaction in all the genotypes.

 

However, only linear component of G×E interaction was noticed which is expressed by significant value of regression coefficient (b_i = 1) only in 35 genotypes for this characters. Rivette exhibited above average (b_i>1) response showing their adaptability to favourable environmental conditions of disease development.

 

Out of 92 genotypes, only 16 genotypes were found to have b_i significant values indicating thereby the presence of linear component of G×E interaction in these genotypes only. Moreover, S²_di value for all the genotypes were found non significant indicating totally absence of non-linear component of G×E interaction for stem diameter. On stability parameters basis (high resistance/ small stem lesion, b_i = 1 and S²_di = 0) RH 13, Ringot, B. juncea 1, B. juncea 2, B. juncea 3 and RQ 011 were highly resistant and stable for small stem lesion length over the environments. Ag spectrum was found suitable for general environment conditions (small lesion length, b_i = 1, S²_di = 0) for disease development. Six genotypes that is, JM018, Ag outback, Monty, B. juncea 1, B. juncea 2 and B. juncea 3, were stable and suitable for conducive environment of disease development (highly resistant, b_i<1, S²_di = 0). In contrary to it, JM 3 was highly susceptible to Sclerotinia stem rot, but it was also stable susceptible (S²_di = 0 and b_i = 1).          

 

The genotypes showing resistance against Sclerotinia stem rot as indicated by lesser number of wilted / dead plant were also exhibited short stem lesion length, wider stem diameter and older plant age for inoculation. Similar observations have also been reported by Li et al. (2006) that there is significant positive linear relationship between plant death and stem lesion length. Hence, the identified stable resistance genotypes could be utilized directly as cultivar after evaluation over time and space if found suitable. Moreover, these should be incorporated in resistance breeding programme to enhance the genetic level of resistance in future cultivars against recalcitrant necrotroph.

 

The authors have not declared any conflict of interest.