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ThesisItem Restricted Epidemiology and Management of Post harvest diseases of Kinnow mandarin(Punjab Agricultural University, Ludhiana, 2021) Gurwinder Kaur; Raheja, SandeepSurveys were conducted periodically (twice in a month) from January to March during 2019 and 2020 to record the incidence and prevalence of major post-harvest rots of Kinnow mandarin and pathogens associated with it in South-western Punjab. Highest incidence of post-harvest rot of Kinnow mandarin 20.21 and 12.59 percent was observed in the month of March during the year 2019 and 2020, respectively. Penicillium digitatum and Penicillium italicum causing green and blue mould rots were found to be two most prevalent post-harvest pathogens with their prevalence of 30.88 and 26.44 percent, respectively during 2019 and 32.44 and 24.88 percent, respectively, during 2020. Temperature of 25˚C favoured maximum radial growth (84.67 mm) of both test pathogens also, maximum percent disease index of green (95.00 percent) and blue mould rot (90.00 percent) was recorded at 25˚C. The pH 5.5 and 6.0 supported maximum radial growth of P. italicum by 90.00 & 88.66 mm and of P. digitatum by 88.67 & 87.67 mm, respectively and were found to be at par with each other. Among different GRAS chemicals and botanicals evaluated against test pathogens, copper sulphate, boric acid, sodium bicarbonate and sodium carbonate at 100mM and above concentrations among chemicals and garlic extract at 10 per cent and above concentration among botanicals showed complete inhibition (100.0 percent) of both test pathogens under in vitro, whereas, under in vivo garlic extract at the concentration of 15 percent showed complete inhibition of green (100.0 percent) and of blue mold rot (92.22 percent). Boric acid at the concentration of 4 percent showed complete inhibition (100.0 percent) of both the test pathogens.ThesisItem Open Access DIVERSITY ANALYSIS OF Drechslera maydis, THE INCITANT OF MAYDIS LEAF BLIGHT OF MAIZE AND ITS MANAGEMENT(Punjab Agricultural University, Ludhiana, 2022) Bagaria, Pravin Kumar; Vineet KumarThe investigation entitled “Diversity analysis of Drechslera maydis, the incitant of maydis leaf blight of maize and its management” was carried out at PAU Ludhiana and ICAR-IIMR, Ludhiana during 2019-2021. The sixty-nine isolates of D. maydis were collected from Punjab and other states showed significant variation in terms of cultural, morphological, and pathological characteristics. The majority of isolates (31) were having greyish black growth of the mycelium on the PDA medium, while 10, 11 and 17 isolates were black, grey and whitish grey, respectively. Most of the isolates (37) showed rough appressed type growth pattern, whereas sectoring and irregular margins were present in 36 isolates. The twenty isolates recorded slow growth (6.0 mm/day). The isolates Dm_8, Dm_46, Dm_58, and Dm_69 showed minimum number septa (3-5 septa/conidium) whereas maximum number of septa (8-12) were observed in isolate Dm_22. Similarly, maximum and minimum spore count (×104/ml) were recorded in isolates, Dm_29 (58) and Dm_51 (11), respectively. Isolate Dm_25 (Ladhowal) recorded minimum incubation (2 days), maximum average lesion numbers (15.03), and highest PDI (94.07%) on susceptible genotype, CM 600, while Dm_26 (Gujarat), the least aggressive isolate, showed maximum incubation period of 4 days with minimum average lesions (11.73) and minimum PDI (61.85%). The isolates were clustered into four groups based on the PDI and representative isolates, viz., Dm_25, Dm_27, Dm_44, Dm_49, and Dm_61 were selected from these groups which were further tested on 10 maize genotypes. The isolate Dm_25 showed maximum average PDI (63.9%), number of lesions (11.1) and AUDPC (846.6), while Dm_27 exhibited minimum average PDI (50.7%), number of lesions (9.1) and AUDPC (668.8). Forty-eight unique polymorphic SSRs were obtained through in-silico analysis of D. maydis strains and filtered to 15 SSRs for further amplification and validation. Six markers (SSR1, SSR4, SSR5, SSR10, SSR13, and SSR15) exhibited high polymorphism with PIC values ranging from 0.81-0.92. Among the 69 isolates, maximum melanin production at 25ºC was recorded in the isolate Dm_25 (2.62 μg/g) while minimum production of melanin (0.37 μg/g) was recorded in Dm_62 isolate. The majority of isolates showed maximum melanin production at temperature range of 25-30°C. Out of 234 maize inbred lines screened under artificial epiphytotic conditions, thirteen genotypes (HKI 42050, V-373, IML66-1, HKI 484-5, LM 13, HKI 163, UMI 112, DQL- 2017, DQL- 2030, DQL- 2231, DQL- 2300-1-1, DQL-2294, and DQL-2105) were found resistant to MLB pathogen. The evaluation of SAR chemicals revealed that maximum disease control (36.72%) and highest yield (29.59 q/ha) were recorded in foliar application of BABA at 200 mg/L followed by ASM (28.66% and 28.34 q/ha) and SA (25.73% and 28.06 q/ha) at 200 mg/L, respectively, as compared to the inoculated control (24.62 q/ha). An overall trend of an upsurge in defensive enzymes (PAL, β-1,3 glucanase and chitinase), antioxidative enzymes (POX and PPO), total phenols and flavonoids was found in inoculated plants of Punjab Sweet Corn 1 treated with SAR chemicals up to 6 days after treatment. Thus, these resistant genotypes may serve as an important source in resistance breeding program against maydis leaf blight and SAR chemicals could be applied for disease management in maize.ThesisItem Restricted Mapping of stripe rust resistance gene(s) in recombinant inbred lines of Triticum aestivum L.(Punjab Agricultural University, Ludhiana, 2022) Waris, Mohammad; Jaspal KaurStripe rust caused by Puccinia striiformis f.sp. tritici (Pst) has been identified as the most devastating wheat rust disease and is considered one of the major constraints on wheat (Triticum aestivum L.). The pathogen's polycyclic and airborne nature causes the emergence of novel pathotypes, resulting in the breakdown of many resistance genes and considerable losses in grain yield and quality. To combat the emergence of novel pathotypes, breeders and plant pathologists are constantly on the lookout for stable and long-lasting sources of resistance. Thus, this research work was designed to find novel sources of resistance. A 441 germplasm entries were evaluated, first for their seedling response against the most prevalent pathotypes of Puccinia striiformis f.sp. tritici (238S119, 110S119, and 46S119). Based on infection-type data, out of the 441 lines, 30 lines were found highly resistant to all three pathotypes tested, and 27 lines were highly susceptible. The same entries were also tested for adult plant resistance against stripe rust under field conditions at Ludhiana and Gurdaspur during 2019-21. From the field response against stripe rust infection, i.e., based on final rust severity (FRS) and area under disease progress curve (AUDPC) for all three years at Ludhiana and Gurdaspur, the lines were categorized into highly resistant, resistant, moderately resistant, moderately susceptible, and susceptible. To see the performance of genotypes in multi-environments the data was subjected to GGE biplot analysis. Wheat genotypes were inconsistent in terms of stripe rust reaction at the tested locations. IC111939 (G1) was found to be the "ideal" genotype in both environments. Gene postulation in the 45 lines which shows both all-stage resistance as well as adult plant resistance was done by using 13 known YR geneassociated markers pertaining to 5 Yr genes (Yr5, Yr10, Yr15, Yr24, and Yr26). Yr5 presence was detected in sixteen lines with two linked markers, i.e., Xwmc175 and Xgwm120; Yr10 was detected in ten lines linked with the marker Xpsp3000; Yr15 was detected in fourteen lines with two linked markers, i.e., Xgwm413 and Xgwm273; Yr24/26 was detected in 15 lines with two linked markers, namely Xbarc181 and Xbarc187. For mapping of stripe rust resistance genes in RILs of bread whe at, four F5:6 RIL populations (IC530087 x PBW621, IC530078 x PBW621, IC553914 x PBW621, and IC529094 x PBW621) were tested with stripe rust pathotypes at the seedling stage (238S119, 46S119, and 110S119) as well as at the adult plant stage against a mixture of pathotypes. The same amounts of DNA from ten resistant (R) and ten susceptible (S) lines from each of the four populations were used to make the R and S bulks. A BSA-based 90K SNP array was performed, and 81,423 SNPs were reduced to 71,991 KASP SNPs. The polymorphic SNPs were then used for the mapping of stripe rust resistance genes in all the RILs. A total of 66 KASP markers were developed from the sequencing data. Out of 66 KASP markers, 35 were found to be polymorphic between the parents, and 19 were validated on RIL populations. Mapping using these 19 markers led to the identification of five major QTLs, qYr-pau1A_P5, qYr-pau-5B_P5At, qYr-pau-5A-p11, qYr-pau-6A, qYr-pau-2B_P7, and one minor QTL, qYrpau-P9-3b, on chromosomes 1A, 5B, 5A, 6A, 2B, and 3B, respectively. KASP markers associated with these genes can be directly utilized for marker-assisted breeding in wheat.ThesisItem Restricted Characterization of blast and bacterial blight resistance genes in basmati rice(Punjab Agricultural University, 2022) Vashisth, Tanvi; Jain, JyotiThe present study was conducted to evaluate MCF1 populations of basmati rice developed from complex crosses (designated as cross A and cross B). Phenotypic evaluation of cross A was done for resistance to neck blast (Pyricularia oryzae) using highly virulent isolate NB-7 under artificial inoculation conditions. Cross B was evaluated against neck blast (NB-7) as well as bacterial blight (Xanthomonas oryzae pv. oryzae) using two pathotypes PbXo-7 and PbXo-8 under artificial inoculation conditions. Among the MCF1 population comprising of 421 plants from cross A, 12 plants were immune, 6 were highly resistant, 30 were resistant and 59 were moderately resistant to neck blast isolate NB-7. A population of 103 plants from cross B showed immune response to neck blast in 9 plants, while 12 plants were resistant to pathotype PbXo-7 and 27 plants were resistant to PbXo-8. Both the crosses were also analyzed for known blast resistance genes (Pi54, Pi2, Pi9 and Pi68) and bacterial blight resistance genes (xa13 and Xa21). A total of 200 plants were tested in cross A, and 199 were found to have the Pi54 gene, while 155 had the Pi2 gene. Only 4 plants showed the presence of Pi9 gene in cross A. The combination of two blast resistance genes was found in 149 plants (Pi54 + Pi2) and in four plants there was a combination of three genes (Pi2 + Pi54 + Pi9). In the cross B, only two plants showed the presence of blast resistance gene (Pi9) and 40 plants showed the presence of Xa4 gene. Two plants were heterozygous for bacterial blight resistance gene (xa13) and four were heterozygous for Xa21. There were only two plants identified as having blast + BB resistance genes (Pi9+Xa4) and one plant showing two BB resistance genes (Xa21+Xa4). The assay of biochemical parameters indicated a significant increase in activity of peroxidase, PPO, PAL, catalase and hydrogen peroxide among resistant/moderately resistant genotypes as compared to the susceptible genotypes at 10 days after inoculation with P. oryzae. Out of 35 genotypes screened against leaf and neck blast phases under artificial inoculation conditions, none was found to be resistant to both the phases. Only two entries viz., Tetep and Pusa Basmati 1637 showed moderately resistant reaction to both neck and leaf blast with susceptibility index (Sx) value <3. Three genotype namely JJ92, UPR3519-18-1-1 and IET 15392 exhibited moderately resistant reaction to leaf blast but were moderately susceptible to neck blast under artificial inoculation conditions. Under natural epiphytotic conditions (hotspot location), five entries namely JJ92, UPR3519-18-1-1, IET 15392, Tetep and Pusa Basmati 1637 showed moderately resistant reaction to neck blast and resistant reaction to leaf blast. The current study has the potential to contribute towards strengthening ongoing blast and bacterial blight resistance breeding program by utilizing the identified donors harbouring single as well as multiple resistance genes to both the diseases.ThesisItem Embargo Factors affecting aggressiveness of Sclerotinia sclerotiorum (Lib.) de Bary inciting stem rot of rapeseedmustard(Punjab Agricultural University, Ludhiana, 2022) Chetan V. N.; Sandhu, P SThe stem rot caused by Sclerotinia sclerotiorum (Lib.) de Bary is one of the major diseases of rapeseed-mustard. In this study, factors affecting aggressiveness of the pathogen were evaluated using isolates collected from various rapeseed-mustard growing regions of India. These isolates were estimated for development of the disease, oxalic acid production, total cellulase and cutinase activity. These isolates were significantly different w.r.t. their aggressiveness which highly correlated with oxalic acid production and cutinase activity and moderately with total cellulase activity. Low and high aggressive isolates were sequenced to detect variation in genes Sscut and Ssoah1. No variation was seen in Sscut while Ssoah1 showed 7 synonymous SNPs. Further study was conducted to test the efficacy of alkali salts such as sodium carbonate (SC), potassium silicate (PS), sodium propionate (SP) and calcium propionate (CP) against Sclerotinia stem rot. Under in vitro conditions, the PDA media amended with SC (0.6-1 per cent), PS (0.8-1.2 per cent), CP and SP (1.6-1.8 per cent) showed complete mycelial growth inhibition when compared with un-amended PDA medium. In case of detached leaf assay, SC @ 1 per cent, PS @ 1.4 per cent, SP @ 1.8 per cent and CP @1.8 per cent were effective in inhibiting lesion development of pathogen when compared to untreated leaves. Effective treatments were tested under field conditions at two locationswhere different treatments were sprayed with different alkali salts 2 days before the inoculation. The pooled data of both the locations revealed that CP @ 1.8 per cent showed highest percent reduction in mean lesion length (63.35 per cent) followed by SP @1.8 per cent with 57.77 per cent reduction. The yield data showed negative correlation with lesion length which implies significant reduction in yield of control as compared to treatments.ThesisItem Embargo Web blight of mungbean (Vigna radiata L.) and its management(Punjab Agricultural University, Ludhiana, 2022) Raghveer Singh; Sharma, Krishan KumarWeb blight of mungbean, caused by Rhizoctonia solani, has a significant impact on grain yield and quality. The pathogen is soil-borne and has a broad host range. Field surveys were conducted in different districts of Punjab during the year 2019-20. The survey concluded that Ludhiana district had the lowest prevalence and severity of web blight, while Ballowal Saunkhri (SBS Nagar) had the most. Pathogen variability was investigated at morphological and molecular level. On the basis of morphological characterization viz. colour, mycelial abundance, shape and size of sclerotia, the pathogen was identified as Rhizoctonia solani. Of all the five isolates, BS (Ballowal Saunkhri) and NS (Nawanshahr) were found to be the fast-growing isolates with a 0.88 mm/hr growth rate followed by RO(Ropar) and L1(Ludhiana 1) and L2 (Ludhiana) 0.68 mm/hr. Among all the isolates, L1-L2, BS-L1, RO-L1, NS-L1, and NS-BS were vegetatively compatible, whereas BS-L2, NS-L2, BS-RO, RO-NS, RO-L2 were vegetatively incompatible. The isolates were characterized on the basis of ITS region sequence homology and confirmed as R. solani. The sequences have been registered with NCBI as accession numbers ON17668, ON176686, ON084814, ON084815 and ON084816. Further, Among the three inoculation techniques viz. soil inoculation, foliar spray and field screening at hotspot location, the foliar spray technique was found to be the most precise inoculation technique. Among the four biocontrol agents viz. recommended consortium (LSMR 1+RB3), a new consortium (Bacillus subtilis +RB1), Trichoderma viride, and Pseudomonas flourescens, the new consortium (Bacillus subtilis +RB1) was found most effective against web blight under both in vitro and in vivo conditions. Further, a set of 58 elite mungbean genotypes was screened for web blight resistance by field screening technique at hotspot location. Out of 58 genotypes, seven namely, SML 2061, SML 2056, SML 2064, SML 2055, SML 2073, SML 2065 and SML 2070 showed resistant reaction to web blight.ThesisItem Restricted Population structure and mating type distribution of Ascochyta rabiei from Northern India(Punjab Agricultural University, Ludhiana, 2022) Lovepreet Singh; Amarjit SinghA set of twenty-four isolates of Ascochyta rabiei causing Ascochyta blight (AB) of chickpea were collected from different chickpea growing areas of Northern India. The AB isolates were assessed for their virulence pattern on a set of varieties with varying levels of resistance viz; PBG 5, PBG 7, PBG 8 and L 555. Disease severity was observed as the lowest in case of AB 19 i.e. 2.83, indicating that it was the least virulent isolate on all four varieties. Likewise; the most virulent isolate was observed as AB 1 which showed the highest disease severity i.e. 7.9 on 1-9 rating scale. On the basis of virulence pattern, all the isolates were clustered into two main groups – Group A (14 isolates) and Group B (10 isolates). The mean disease severity of group A ranged from 4.75 to 7.83 whereas in case of group B, the disease severity ranged from 2.83 to 5.17. Genotyping of all 24 AB isolates with 27 SSR loci produced 119 alleles ranging from 2 to 8 in number and 138 to 490 bp in size with an average of 5.42 alleles per marker. The observed heterozygosity was zero in all the populations. This may be due to absence of sexual spores in India and presence of only haploid pycnidiospores of this fungus. Further, the percent variation between the populations was very small i.e. 0.56%. However, the genetic variation within the population was very high i.e. 99.93%. The unweighted neighbour-joining dendrogram grouped the 24 isolates of the three populations into three major clusters. Cluster I included isolates 1 and 6, cluster II consisted of 11 isolates 20, 4, 11, 24, 9, 14, 8, 21, 13, 15 and 18. Cluster III consisted of 11 isolates 2, 5, 12, 23, 3, 19, 7, 16, 17, 10 and 22. Principal coordinated analysis (PCoA) showed that overall, the AB isolates were grouped into four major groups. Clumpak (barplot) clustering of individuals into populations based on multi-locus genotyping grouped the isolates into 3 clusters with an admixture indicating no clear-cut geographical origin-based sub structuring of the population. The MAT1-2 primer amplified in all the isolates giving an amplicon of ~ 800bp whereas MAT1-1 did not amplify in any of the isolates. It indicated that only one mating type ie; MAT1-2 was present in all the AB isolates. For confirmatory study the 800 bp amplicon produced by MAT1-2 primer was sequenced and subjected to BLAST analysis which showed 81.05 to 98.29% homology with Didymella rabiei strain ATCC 76501MAT 1-2-1 (MAT 1-2-1) gene.ThesisItem Embargo Mapping of Karnal Bunt (Tilletia indica Mitra) resistance gene(s) in recombinant inbred lines of bread wheat(Punjab Agricultural University, Ludhiana, 2022) Bromand, FerdawsThe current study was conducted to evaluate a set of 972 NBPGR (National Bureau of Plant Genetics and Resources) accessions and three recombinant inbred lines (RILs) derived from Karnal bunt (KB) resistant donors viz., ALDAN, CMH77.308, and H567.71 in the background of susceptible parent WH542 at PAU, Ludhiana during crop seasons from 20192022. Out of the total 972 accessions evaluated for resistance against Tilletia indica; 417 lines were found resistant over the three years. These 417 lines included 302 highly resistant lines with zero percent KB infection and 115 lines having KB infection up to 5.0 percent. The phenotyping of three RIL populations of ALDAN „S‟ IAS 58 x WH 542, 208 lines screened for KB resistance reported 24 lines having complete resistance i.e., 0% KB infection and the rest of the 184 lines exhibited 0.1 to >20 percent infection. In RIL population CMH 77.308 x WH 542, out of 75 lines screened for KB resistance, 15 lines were reported to be completely resistant having 0.00 percent KB infection and the rest 60 lines showed 0.1 to >20 percent infection. Likewise, in the RIL population H567.71/3*PAR x WH542, a total number of 72 lines screened for KB resistance, 12 RILs were reported to be completely resistant i.e., 0% KB infection the rest 60 lines showed 0.1 to >20 percent infection. A further panel of 63 lines from two RIL populations i.e. ALDAN „S‟ IAS 58 x WH542 and H567.71/3*PAR x WH542 were subjected to genotyping using SNP markers. This panel consisted of 55 resistant and 8 highly susceptible lines from the two crosses. As a consequence, the present study has identified three major QTLs: qKB.pau.AW-4A.1, qKB.pau.AW-4A.2, and qKB.pau.AW1A from ALDAN „S‟ IAS 58 x WH542 RIL populations. Similarly, qKB.pau.HW-1A.1, qKB.pau.HW-1B, and qKB.pau.HW-1A.2 are the notable QTLs detected from the H567.71/3*PAR x WH542 RIL populations during 2019–2020 and 2020–21. The QTLs detected across these environments may be considered more stable and robust as well as may be utilized in the further breeding program for KB resistance.ThesisItem Embargo Characterization of Karnal bunt (Tilletia indica) resistant genotypes and disease dynamics under alternate cultivation practices of wheat(Punjab Agricultural University, Ludhiana, 2022) Jaspreet Kaur; Vineet KumarA set of 1305 genotypes was evaluated against Tilletia indica under artificial epiphytotic conditions during cropping seasons of 2020-21 and 2021-22. After screening for two years, 284 lines were found to be highly resistant reaction and 289 lines showed resistant reaction (0-5% KB infection). Out of 284 highly resistant lines, diversity analysis of 173 Triticum aestivum lines using 70 SSR markers concluded that majority of the lines were different from each other and have potential to be explored for identification of KB resistance loci. The effect of seed and soil borne inoculum under four different tillage methods viz. happy seeder, super seeder, zero tillage and conventional (control) was studied. Super seeder (infected seed in infected soil) showed highest teliospore count (22,580 per 250 g of soil) and lowest count (1602 per 250 g of soil) was observed with zero tillage (healthy seed in healthy soil). There was significant effect of tillage methods on teliospore count. However, no significant difference of sowing dates was observed on teliospore count. The effect of seed and soil borne inoculum on germination revealed that maximum (80.66%) germination percentage was observed when healthy seed were sown in healthy soil using super seeder while minimum germination (52.26%) was observed with infected seed sown in infected soil using happy seeder. The study needs to be undertaken further to confirm the results obtained in the present investigation.
