Genetic Diversity Studies and Fusarium Wilt Screening in Chickpea (Cicer arietinum L.) Germplasm

Chickpeas are a nutrient dense food, serving as an important source of protein, dietary fibre, minerals and folate for a large portion of the global population. It is self-pollinated diploid (2n = 2x = 16) crop having a genome size of 740 Mbp (Varshney et al., 2013), and is cultivated in about 57 countries under diverse environmental locations. India tops in production and consumption of chickpeas in the world, accounting for 66 per cent of total world’s production. It is cultivated in an area of 9.69 Mha with a production of 11.91 MT and productivity of 1142 kg/ha (Anonymous, 2021). To attain self-sufficiency by 2050, the total pulse production in India needs to reach 39 MT (Vision 2050; IIPR) and amongst all pulses, chickpea production alone needs to reach about 16–17.5 MT from a limited area of 9.69 Mha with a mean productivity of 1500–1700 kg/ha. We can achieve the production goal by exploring the genetic distance present among genotype using D2 statistics for effective choice of parents in the hybridization (Saha et al., 2018).
               
The genetic base of chickpea has narrowed substantially during the domestication process (Thudi et al., 2016). Therefore, increasing the genetic divergence of chickpea has been a major goal for breeders. The assessment of genetic diversity of chickpea germplasm can provide valuable information to the breeder for parental selection strategies in plant breeding programs. Chickpea wilt (Fisarium oxysporum f. sp. Ciceri) is a very important disease and the pathogen in association with other soil-borne pathogens like root rots and foot rot also causes extensive damage to chickpea. It causes around 10% yield loss in India but under certain conditions and specific locations, the losses may go up to 60%. Thus, the present study was commenced to estimate the extent of genetic diversity present and field screening for fusarium wilt among 280 chickpea germplasm.

Two hundred and eighty chickpea germplasms including two checks (JG-11 and SA-1) were evaluated during Rabi 2020-21 at Zonal Agricultural Research Station (ZARS), Kalaburagi. Sowing was carried out by hand dibbling method and approximately 40 seeds were sown per genotype. Sowing was done on 6^th October, 2021 for germplasm characterization and the field experiment was carried out in Augmented Block Design (ABD) with two checks in each block. Each entry was planted in one row of 4-meter length having a spacing of 30 cm between the rows and 10 cm between the plants respectively. The data was noted for five polygenic traits viz., days to 50 per cent flowering, plant height (cm), days to maturity, 100 seed weight (grams) and seed yield/plant (grams) (Plate 1). The data were analysed with the technique of D² statistics (Mahalanobis 1936), as advocated by Rao (1952). The software package INDOSTAT version 8.5 was used to analyse the statistical data. Experimental layout for screening Fusarium wilt was laid out on National Wilt Sick Plot maintained at Agricultural Research Station, Kalaburagi [Latitude (N) 170 35’ and Longitude (E) 760 81’]. The sick plot is maintained since 1985-86 to till date. It is one of the national sick plot for screening AICRP and local genotypes. The wilt sick plot was maintained by adding chopped plants of Fusarium oxysporum f. sp. ciceris infested chickpea plants every year.
 

               
All the genotypes were sown in single row along with wilt susceptible (JG-62) and resistant check varieties (WR-315) during the rabi 2019 season. A row length of 4 meters each was maintained with a spacing of 30 cm and 10 cm between the rows and plants respectively (Plate 2). The observations on per cent disease incidence was recorded at 30, 60, 90 days after sowing by counting the number of diseased and dead plants (due to Fusarium wilt) among the total number of plants present per genotype and per cent disease incidence was estimated. Disease rating was categorized as per the standard wilt scoring format of All India Coordinated Research Project (AICRP) on chickpea, IIPR, Kanpur.
 

Analysis of variance (ANOVA) is an important tool to determine the variability present among the genotypes. The analysis (ANOVA) revealed the presence of considerable variability among 280 genotypes including checks (JG-11 and SA-1) for all the five polygenic traits (Table 1). The results show ample scope incorporating the promising genotypes in the breeding programmes aimed improvement in seed yield and its component characters.
 

 
Group constellations by tocher method
 
On the basis of D² values, two-hundred and eighty genotypes including two checks (JG-11 and SA-1) were clustered into 17 clusters based on Tocher’s method (Rao, 1952). Out of 17 clusters, Cluster II has the highest number of genotypes (89) followed by cluster I with 83 genotypes, cluster III with 31 genotypes, cluster VIII with 29 genotypes, cluster VI with 24 genotypes, cluster VII with 15 genotypes (Table 2 and Fig 1). Whereas, the remaining clusters IV, V, IX, X, XI, XII, XIII, XIV, XV, XVI, XVII are mono-genotypic containing one genotype each, which shows that the genotypes included in these clusters are more divergent than the genotypes belonging to other clusters. The results are in accordance with the results obtained by Gediya et al., (2018), Bohare et al., (2020). The solitary clusters are obtained due to the fact that they exhibit some distinctive characters which make them diverse from the other clusters.
 

 

 
Relative contribution of each character towards genetic divergence
 
The analysi revealed that, days to maturity had contributed maximum towards divergence (29.92%) followed by 100 seed-weight (28.02%), plant height (22.45%), days to 50 per cent flowering (18.67%). Whereas,least contribution was made by seed yield/plant (0.57%) towards the genetic divergence (Fig 2).
 

 
Intra-cluster and inter-cluster distances among the chickpea germplasm
 
Intra-cluster and inter-cluster distances were calculated using D² values. Out of 17 clusters, intra cluster distances were lower than inter-cluster distances indicating that genotypes included in different clusters are more diverse compared to the genotypes included in the same clusters. The mean intra-cluster D² values were ranged from 0 to 425.66. Cluster VII logged highest intra-cluster distance (425.66), followed by cluster VIII (401.40), indicating that wide genetic divergence was existing among the genotypes within these clusters. So, more emphasis will be given to the genotypes included in these clusters while selecting parents for future crop improvement. The clusters viz., cluster VI (260.71), cluster III (221.62), cluster II (176.78), cluster I (148.70) had moderate intra cluster distance. Whereas, no intra cluster distance was observed in nine clusters IV, V, IX, X, XI, XII, XIII, XIV, XV, XVI, XVII as they are mono-genotypic with one genotype per cluster (Table 3).
 

 
Cluster mean values of seventeen clusters
 
Cluster mean value ranged from 31 days (cluster XIV) to 83 days (cluster XVI) for the trait days to 50 per cent flowering. Cluster XIV genotypes exhibited early flowering habit with average number of days taken to flowering were 31, indicating that the Cluster XIV composed of early flowering genotypes.  Cluster mean for the trait days to maturity ranged from 90 (cluster XI) to 130 days (cluster XVI). The genotypes of Cluster XI demonstrated a unique early maturing behaviour, taking only 90 days to reach maturity. In terms of plant height, genotypes from cluster IX displayed the greatest cluster mean value (66 cm), indicating that they were taller than genotypes from other clusters. Cluster VI had the lowest cluster mean value (34.03 cm) indicating that the genotypes included in this cluster are dwarf. Cluster mean value for 100 seed-weight was ranged from 8 g (cluster XI) to 56.8 g (cluster XVII), the genotypes of the cluster XVII showed the highest 100 seed-weight followed by cluster III (31.7 g) which shows that the genotypes of this cluster have bold seed characters which are preferred by the consumers. The genotype MNK-1 (56.80 g) showed the highest 100 seed-weight and the genotype ICC-14077 (4.8 g) has the lowest 100 seed-weight. Cluster mean for seed yield/plant values ranged from 2.38 g (cluster XII) to 8.13 g (cluster XVII), with genotypes from cluster XVII had the highest seed yield/plant (8.13 g) suggesting that the genotypes of this cluster are preferred for the seed yield improvement of chickpea (Table 4).
 

 
Fusarium wilt screaning in chickpea (Cicer arietinum L.) germplasm
 
Among the two hundred and eighty-two chickpea genotypes along with two checks screened against wilt, there was a significant variation between genotypes for their disease reaction. Out of 280 germplasm screened, 30 showed resistant (10.71%), 83 moderately resistant (29.64%), 44 moderately susceptible (15.71%), 31 susceptible (11.07%) and 92 highly susceptible reaction to fusarium wilt. The per cent disease incidence (PDI) ranged from 0.44 (ICCV 16116) to 49.49 (ICCV-171101) among germplasm with resistance check WR-315 and susceptible check JG-62 recording the wilt incidence of 4.25 and 86.42 per cent respectively. The lowest PDI was observed for the line ICCV 16116 (0.44) followed by ICC 9942 (1.77), ICC 4657 (3.48) and ICC 9872 (5.50) and can be re-validate and used in fusarium wilt resistance breeding program (Table 5). Similar study was conducted by Ayyub et al., (2003) who reported high level of resistance to Fusarium wilt in chickpea germplasm originating from different sources. Bakhsh et al., (2007) reported 3 genotypes with disease incidence 0, 6.7 and 8.3 per cent as resistant and 4 with disease incidence of 18.2 to 20 per cent as tolerant Kumar et al., (2013) reported that out of 100 genotypes, 44 showed resistant reaction, 11 MR, 22 tolerant, 15 moderately susceptible and 8 susceptible to Fusarium wilt. Similar results were also reported by Shah et al., (2015) and Kumar et al., (2015), Ayana et al., (2019). Laxuman et al., (2022) used various wilt resistant chickpea genotypes and breeding lines for evaluating yield response in different environments. They observed higher average yields of KCD-11 and ICCV 191106 with wilt resistance.

In this current divergence study, the relative contribution of each character towards genetic divergence will provide effective data to assist the plant breeders in selecting superior genotypes from available germplasm collection to use them as parents in the future crop improvement programme. The maximum intra cluster distance (425.66) was observed in the cluster VII, followed by cluster VIII (401.40), indicating the existence of wide genetic diversity among the genotypes within these cluster. The maximum inter cluster distance was observed between cluster XIV and cluster XVI (4352.70), followed by cluster X and cluster XIV (3578.44), cluster XVI and cluster XVII (3383.30), cluster XIV and cluster XV (3063.34), cluster VI and cluster XVI (3050.80), revealing that genotypes included in these clusters are genetically diverse and is vital for future hybridization programme. The field screening for Fusarium wilt of two hundred and eighty chickpea genotypes in the wilt sick plot yielded thirty resistant genotypes, of which ICCV 16116 (0.44) followed by ICC 9942 (1.77), ICC 4657 (3.48), ICC 9872 (5.50) showed significantly higher level of resistance for wilt as they produced narrow range of per cent disease incidence (PDI) value. The characters which contributed more to genetic divergence may be given more weightage in future plant breeding programmes to improve the yield through selection. The Fusarium wilt resistant genotypes which were screened using sick plot can be directly released as variety or choice of parents for hybridization programme.

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