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Biochemical defense response and genetic basis of resistance against Ascochyta blight (Ascochyta rabiei (Pass.) Labr.) in chickpea (Cicer arietinum L.)
(Punjab Agricultural University, 2024) Abassy, Omer; Dr Upasana Rani
Chickpea (Cicer arietinum L.), the second-largest global pulse crop, is under threat from highly variable nature of Ascochyta rabiei, causing widespread disease damage under favourable environmental conditions. This disease, Ascochyta blight is a major concern worldwide, with insufficient resistant sources and frequent breakdowns due to rapid pathogen evolution. This study evaluated 238 kabuli chickpea genotypes for resistance to Ascochyta blight (AB) over the growing seasons from 2019-2022 at Punjab Agricultural University under artificial epiphytotic conditions, identifying 18 resistant lines (GLK 10-40, GLK 20054, GLK 20055, FLIP 10-298C-IFC-S2, FLIP 10298C-IFC-S1, CS-3-E-24, FLIP 09 256C-55, FLIP 07-314C-57, FLIP 10-243C, FLIP 09-194C, FLIP 08-104C, FLIP 04-219C, ICCV 55233, ICCV 55215, ICCV 55135, ICCV 55108 and ICCV 155141) belonging to different genetic backgrounds with disease scores of 1.0 to 3.0, whereas seventy-three (73) lines demonstrated a moderate level of disease resistance with a disease rating of 3.1-5.0. The activities of defense-related enzymes (PAL, TAL, PPO, POD) associated with phenol metabolism along with lignin and total phenol content were compared among six kabuli chickpea lines, viz., five resistant (GLK 10-40, GLK 20055, FLIP-09-194C, FLIP-04-219C, ICCV 55215) and one susceptible (GLK 17301) treated lines that exhibited differential responses to Ascochyta blight at tested time intervals of 48, 96, 144 and 240 hr post-inoculation respectively. The perusal of data showed increased activity of all enzymes (PAL, TAL, PPO, POD) till 96 hr after inoculation, whereas the non-enzymatic contents such as lignin and total phenols showed the maximum enzyme activity till 144 hr after inoculation. However, enzyme and non-enzymatic activity remained almost constant with no significant difference in all control/untreated resistant and susceptible line(s) at different time intervals. For understanding the genetic basis of resistance, the F2 population comprising of 162 plants derived from the cross, GLK 24096 (resistant) and L 556 (susceptible) was assessed under artificially epiphytotic conditions against local isolate of Ascochyta rabiei. Of 162 plants evaluated, 39 plants were grouped in resistant (R ) category (disease score ≤ 5.0 ) and 123 were classified as susceptible (S) with score of five and above. Consequently, the F2 plants exhibited a segregation ratio of 1R:3S, suggesting that a monogenic recessive gene governed resistance to Ascochyta blight. Of thirty four SSR markers associated with Quantitative Trait Loci (QTLs)/genes were used for confirmation of resistance in identified eighteen resistant and six susceptible lines, only 19 SSR markers depicted polymorphism. Notably, SSR markers, STMS11, TA194, TS12, TA146, and TA2 exhibiting significant polymorphism, indicated tight linkage to Ascochyta blight resistance. Phylogenetic analysis showcased distinct clustering based on resistance, offering insights into the genetic basis of resistance. These findings informed about targeted resistance breeding and sustainable disease management with defence-related enzymes/identified markers holding promise for developing superior, resistant and resilient chickpea varieties contributing to crop sustainability and productivity.
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Characterization of mycotoxic Fusarium species associated with ear rot of maize
(Punjab Agricultural University, 2024) Harinder Singh; Dr. Harleen Kaur
Ear rot samples of maize (Zea mays L.) were collected during Kharif and spring seasons from different districts of Punjab. In all, 48 Fusarium isolates were obtained and purified for further studies. These isolates were grouped into six major groups based on morphological characteristics viz., colony color, pigmentation, type of mycelium, pattern of growth, growth rate and sporulation. Two species of Fusarium viz., F. verticillioides and F. proliferatum were identified using ITS, TEF-1α, RPB2, VERT- 1/VERT-2 gene specific primers. The sequences of four representative isolates-Fus 15, Fus 28, Fus 44 and Fus 45 when submitted to BLAST showed 99-100 per cent similarity with F. verticillioides and Fus 48 isolate had shown 97.91 per cent similarity with F. proliferatum. Further, out of 47 isolates of F. verticillioides VERTIF-1/VERTIF-2 primers 40 isolates with fumonisin producing ability. FUM13F /13R primers identified 36 isolates of F. verticillioides with fumonisin producing ability. FUM1 primers revealed that out of 48 Fusarium isolates, 41 isolates possessed FUM1 gene. Genetic diversity analysis of Fusarium isolates using 20 SSR primers revealed high degree of polymorphism and clustered 48 isolates into two major groups and seven subgroups. The primer 3H07 amplified maximum number of loci (6) ranging from 100-1400 bp in size and had highest PIC value of 0.76. On comparing the clusters of morphological, molecular and pathological characterization defined correlation could not be obtained among morphological characteristics, SSR pattern and pathological behavior of Fusarium isolates. For standardization of inoculation technique, we used four techniques viz., needle inoculation into the cob, toothpick inoculation in the ear, needle inoculation into the silk and spray silk inoculation, out of which needle inoculation into the cob technique gave highest mean disease index of 26.44 and 36.77 per cent during spring and Kharif seasons, respectively. Higher disease severity of FER was obtained when cobs were inoculated at 7 days after silking in both spring (19.34 %) and Kharif (32.16%) seasons. On comparing the seasons, Kharif season showed comparatively more disease severity (25.25 %) than spring season (17.1 %). Using this standardized technique, forty-five advanced maize germplasm lines were screened artificially against FER during Kharif 2021, disease severity varied from 27.77 to 96.29 per cent being lowest (27.77 %) in LM13×Teosinte-1-2-1-1-1 (BC1F1) and highest (96.29 %) in LM14×Teosinte-1-1-1-1 (BC1F1) line. Out of 45 lines, none of lines showed resistance reaction to FER, however 13 lines were found moderately resistant. Pathological behavior of all Fusarium isolates was studied on four hybrids (PMH 1, PMH 2, PMH 10 and PMH 13) and two inbreds (LM 13 and EML 173) of maize during spring 2022. Based upon their virulence, heatmap was generated and all isolates were clustered into seven groups. Group I isolates comprising Fus 15 (Ludhiana), Fus 23 (Jalandhar) and Fus 48 (Sirmour) were highly aggressive giving maximum disease severity of 68.74 per cent. Among the hybrids, PMH 1 showed minimum mean disease severity (34.16%) giving moderately resistant reaction to all Fusarium isolates. Among the inbreds, LM 13 showed minimum mean disease severity of 41.16 per cent giving moderately resistant reaction. The fumonisin production potential of F. verticillioides isolate- Fus 15 and F. proliferatum isolate-Fus 48 was observed on PMH 1 and PMH 2 hybrids under artificial inoculation conditions. Concentration of Fumonisin B1 (FB1) produced by Fus 15 and Fus 48 isolates was comparatively higher in PMH 2 than PMH 1 hybrid. Maximum fumonisin production of both FB1 and FB2 in maize grains was estimated at 21 days after incubation.
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Resistance gene identification in Vigna species infected with Begomovirus
(Punjab Agricultural University, 2024) Kuppuraj J; Sirari, Asmita
The successful intra and inter-specific hybridization in Vigna revealed the dominant nature of yellow mosaic disease resistance with both monogenic and digenic gene action with dominant epistasis in Vigna mungo and digenic gene action with complementary epistasis in Vigna radiata. Further, differential expression of different classes of resistance genes in both compatible and incompatible Vigna – mungbean yellow mosaic India virus interactions was investigated. The qPCR primers were designed for amplifying plant resistant protein domain - specific regions (NB-ARC; SgS3; RdRp3) from genome of different Vigna species. Resistance protein domain specific sequences were shortlisted (with candidate genes, type species) and retrieved from NCBI database, followed by multiple sequence alignment through MEGA11 software on different Vigna species genome. A total of 37 qPCR primers with three domain - specific regions were designed using Primer3Web version 4.1.0 online software, were amplifying with specific amplicon size (90 to195 bp) and moreover without primer dimer formation. Most of the primers were amplified on genomic DNA of four resistant and two susceptible genotypes of different Vigna speices genome. Total RNA was isolated and cDNA panel was prepared from mock and MYMIV inoculated leaf tissues of both resistant and susceptible genotypes of different Vigna species at 1, 5, 10, 15, 20, 25 and 30 days post inoculations. SYBR green chemistry was performed in RT-qPCR to study differential gene expression in both compatible and incompatible interactions. All the three resistant protein domains i.e. NB-ARC, SgS3 and RDR were found to be involved in YMD resistance in Vigna radiata and Vigna umbellata with absence of NB-ARC functional, only in Vigna mungo. On the other hand, expression of mungbean yellow mosaic India virus specific coat protein gene AV1 was higher in susceptible Vigna radiata genotype (SML1082) as compared to Vigna mungo and vice-versa in case of replication gene AC1. Low level of expression of both the AV1 and AC1 genes was also observed in resistant genotypes of Vigna with first experimental evidence of Vigna umbellata as host of mungbean yellow mosaic India virus.
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Understanding sunflower - Macrophomina phaseolina interaction under water stress conditions
(Punjab Agricultural University, 2023) Navkiran Kaur; Sharma,Pankaj
A total of 35 isolates of Macrophomina phaseolina causing charcoal rot were collected from eleven kharif and rabi crops from different districts of Punjab and adjoining states. Macrophomina phaseolina isolated from sunflower field, Ludhiana was characterized on cultural and morphological and molecular basis. The isolates infecting sunflower showed highest sequence homology of ITS genomic region with M. phaseolina isolate from Indore, Madhya Pradesh NCBI accession no. MT127404. This isolate was further used to standardize the inoculation techniques under field and polyhouse conditions. Tooth pick inoculation technique was found most effective for large scale screening while sorghum grain inoculation method was proficient under polyhouse conditions. A set of 71 sunflower genotypes were evaluated using tooth pick inoculation technique against charcoal rot under irrigated and restricted irrigation conditions. Four genotypes viz; 75B, EC6078261, OPH137, and OPH 172 were found moderately resistant under irrigated conditions whereas only two genotypes 75B and EC6078261 showed moderately resistant reaction in restricted irrigated conditions. The present study was first attempt to investigate the water stress-Macrophomina phaseolina interaction on different components of physiological, plant growth and yield parameters along with disease incidence. A significant reduction in total chlorophyll content (54.78 %), relative water content (47.06 %), membrane stability (51.75 %), sugar (69.00 %), protein (13.90 %) and proline (56.66 %) content was observed with inoculum density of 4 g/kg soil at soil moisture levels of 40 % and 60 %. The inoculum density of 4 g/kg of dry soil resulted in maximum mortality during seedling stage and growth period. Thus, varied inoculum density and water stress conditions made the sunflower plants more prone to dismissive physiological alterations. It led to biomass reduction viz; root length (41.27 %), root width (76.74 %), root weight (85.37 %), shoot length (56.71 %) and shoot weight (79.06 %). Thus it resulted in per plant seed yield reduction (95.45 %) at 52.62 to 100 % disease severity at higher inoculum density (4g, 6g, and 8g/kg dry soil). The expression levels of the defense related genes LOX, ACCO1, PAL, SOD, and APX were also modified under varying degree of water stress and inoculum density. The strong correlation between low soil moisture and disease severity implies that the proper irrigation of the field can be suggested as cultural management practices for the farmers.
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Virulence pattern of leaf and neck blast isolates (Pyricularia oryzae Cavara) from basmati rice and identification of resistance donors
(Punjab Agricultural University, 2023) Anjali; Jain, Jyoti
The blast disease caused by Pyricularia oryzae was prevalent throughout the Punjab state with leaf blast incidence ranging from 9.47 to 54.26 percent and neck blast incidence ranging from 23.73 to 46.11 percent respectively. The highest disease incidence and disease severity were observed in the agro-ecological Zone-III comprising of Bathinda, Sangrur, Barnala and Muktsar districts on variety Pusa Basmati 1401 followed by Pusa Basmati 1509 and Pusa Basmati 1121. Leaf and neck blast isolates were found to be variable with respect to colony colour, elevation, margin, form, sectoring, zonation, mean radial growth (mm), spore size and extent of sporulation. Three leaf blast isolates (LB-3, LB-10 and LB-11) and five neck blast isolates (NB-51, NB-52, NB-65, NB-83 and NB-89) were categorizedas fast growers with Kr value of more than 8.1mm/24hr. All the tested isolates sporulated better on Oatmeal agar medium than on Paspalum grass medium. Genetic analysis of the leaf and neck blast isolates using SSR markers revealed high degree of polymorphism with PIC values ranging from 0.44 to 0.90 and 0.44 to 0.93 respectively discerning the isolates into two major clusters. Studies on virulence pattern and cross-infectivity exhibited substantial variability among leaf and neck blast isolates on a set of eight basmati varieties. Univariate and multivariate analysis of isolates originating from leaf or neck portions of basmati rice with respect to various components of aggressiveness viz., infection efficiency (IE), Incubation period (IP50), Latent period (LP), Lesion size (LS), Sporulation intensity (SI), Disease incidence (DI) and Disease severity (DS) demonstrated their cross-pathogenic behaviour. However, the neck blast isolates proved to be more aggressive on rice leaves than leaf blast isolates on rice necks. Among the different components of aggressiveness, IE, LS and SI were positively correlated with each other, whereas IP50 and LP50 showed a negative correlation with all the aggressiveness components. Out of 96 germplasm lines screened against leaf and neck blast phases separately under artificial inoculation conditions, none of the genotypes was found to be resistant to both phases. Only six genotypes namely, IR 67417-75-3-2-2-3, IR 72860-62-1-1-3, Acharamati, Basmati C122, Pusa Basmati 1637 and Tetep showed moderately resistant reaction to both leaf and neck blast phases with susceptibility index <3. Ten genotypes IR 67017-73-1-4, IR 67418-131-2-3-3-3, IR 68726-3-3-1-2, IR 68740-3-2, IR 70418-112-1-2, Basmati Surkh 161, JJ-92, Tulsi Phoola, UPR-3519-18-1-1 and IET 15392 exhibited moderately resistant reaction to leaf blast but were moderately susceptible to neck blast. Under natural epiphytotic conditions at hotspot location (RWRC Malan, H.P.), twelve genotypes namely IR 67017-73-1-4, IR 67417-75-3-2-2-3, IR 68726-3-3-1-2, IR 68740-3-2, IR 70418-112-1-2, IR 71740-7-3-1, Basmati C122, JJ-92, Tulsi Phoola, IET 15392, Pusa Basmati 1637 and Tetep showed resistant reaction to leaf blast and moderately resistant reaction to neck blast. The present study established that resistance to the leaf blast phase does not necessarily imply resistance to the neck blast, indicating a need to evaluate rice genotypes for both the leaf and neck blast phases in the ongoing blast breeding programs. Furthermore, the new insights gained into the biology of P. oryzae, its organ specificity, comparative aggressiveness, and the genetic sources of resistance will be useful in designing better management strategies.
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Detection, variability and translocation of Potato virus Y (PVY) infecting potato in Punjab and identification of host plant resistance
(Punjab Agricultural University, 2023) Belludi, Rakesh; Sharma, Abhishek
This study aimed to investigate the variability of Potato virus Y (PVY) infecting potato crop of Rabi season in Punjab, encompassing symptomatology, serology and molecular aspects. The study revealed prevalent symptoms, such as mild mosaic, chlorotic patches, interveinal chlorosis, puckering, veinal necrosis and stem necrosis, on potato plants caused by various potato viruses including PVY. ELISA tests confirmed the presence of six out of seven viruses in potato samples. Among them, PVX exhibited the highest incidence (78.08%), followed by PVYN (51.37%), PVM (20.55%), PVYO/C and PVS (13.01%). PLRV exhibited a lower incidence (2.74%) and no samples tested positive for PVA. Notable regional variations noticed, with Ludhiana showing high incidences of PVYN (78.43%), PVYO/C (31.37%), PVM (33.3%) and PVS (35.29%). Conversely, Kapurthala displayed a 100% PVX incidence, while Jalandhar had the highest PLRV (19.05%) incidence. In nucleic acid-based detection through RT-PCR, the primer NIb2F/3R effectively amplified the desired ~350bp amplicon in all PVYN and PVYO/C seropositive samples. Whereas, the qPVY NIb F2/CP R2 primer generated the ~1.5kb fragment in only 24 out of 67 samples, further confirmed via BLASTn sequence analysis as PVYO, PVYNTN and PVYN-Wi strains. Further virus translocation studies revealed a wide range of carryover rates, spanning from 40% to 92.46%, from infected mother plants to daughter tubers, exhibiting significant variation in viral titre across genotypes and among tubers. Spatial translocation within the tuber's root zone revealed that the bottommost tuber displayed a relatively lower PVY titre. Interestingly, freshly harvested tubers exhibited a higher viral titre at the heel end (stem end) compared to the rose end eyes in all examined tubers. After storage, a contrasting trend emerged in sprouted tubers, where the highest PVY titre was found in rose end eyes and the lowest in heel end eyes. In germplasm screening against resistance to PVY, one germplasm (KP -16-19-14) displayed high resistance (HR), while 16 exhibited resistant (R), 20 moderately resistant (MR), 22 moderately susceptible (MS), 10 susceptible (S) and two were highly susceptible (HS) to PVY. Following PVY inoculation, significant reduction was observed in chlorophyll fluorescence parameters, viz., Fv/Fm, Y(II), qP, and qL values in susceptible cultivars, accompanied by notable increases in NPQ, Y(NO) and qN values. Susceptible genotypes displayed yield losses ranging from 13.64 to 58.84% with viral titre ranging from 0.014 mc to 419.673 mc. Finally, a farmer-friendly and cost-effective vertical flow assay (VFA) was developed for rapid PVY detection in potato leaf samples. The sensitivity of the developed assay was equivalent to commercially utilized ELISA. Specificity test confirmed assays precision for prevalent PVY strains and its non-specificity to other potato viruses. Developed assay exhibited 95% accuracy compared to RT-PCR.
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 Kumar
The 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.
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Mapping of stripe rust resistance gene(s) in recombinant inbred lines of Triticum aestivum L.
(Punjab Agricultural University, Ludhiana, 2022) Waris, Mohammad; Jaspal Kaur
Stripe 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.
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Characterization of citrus greening pathogen(s) and its integrated management
(Punjab Agricultural University, 2023) Katoch, Sonali; Arora, Anita
Surveys conducted during 2019-21 to assess the distribution and intensity of greening disease in citrus orchards of Punjab revealed that its prevalence was high in the district Ludhiana (33.33%) followed by Hoshiarpur (32.60%), Fazilka (27.59%) and Faridkot (19.31%). The maximum mean disease incidence (16.60%) was observed in Hoshiarpur and the maximum mean disease index (11.98%) was observed in Ludhiana. Leaf mottling, yellowing, green islands and bi-coloured fruit symptoms of disease were observed on different cultivars of citrus. Multilocus sequence typing using 16S rDNA, CLIBASIA gene locus and tufB-secE-nusG-rplKAJL-rpoBC gene cluster was followed to identify and characterize the pathogen. Three primers (OA1/OI2c, CGB F/R and Las606/LSS) corresponding to bacterial 16S rDNA sequence were used on a total of 153 samples, and positive reactions were observed with all the three primer pairs. However, CGB F/R and Las606/LSS had higher detection rates (69.9%) than the OA1/OI2c primer (37.8%), indicating more robustness of the former. Similar results were observed with nursery samples of Punjab. The root samples from greening positive plants collected in the month of February were also found positive with five primer pairs (OA1/OI2c, CGB F/R, rplJ/K F/R, Las606/LSS and LapGP F/R) whereas, the root samples collected in May and September months showed no amplification. In addition, the CLIBASIA gene locus based primer pairs corresponding to sequences coding for hypothetical, heamolysin and type-I secretary system protein and one primer based on tandem repeats were used on 15 isolates of greening bacterium. All the samples showed positive reaction with five primer pairs. The disease was characterized by sequencing of 16S rDNA and CLIBASIA gene locus from five isolates with CGB F/R, three with Las606/LSS, two clones with OA1/OI2c and four with CLIBASIA gene locus which showed 99-100% similarity with reported isolates of Candidatus Liberibacter asiaticus (Ca. Las) from Asia. The phylogenetic analysis of these isolates showed 99-100% similarity with Ca. Las. The isolation of Ca. Las from leaves of PCR positive Kinnow plants was attempted on Liber A media, but it could not be cultured. The cultivars viz., Kinnow, Daisy, Musambi and Kagzi Lime were observed to be the best indicator plants for indexing. For integrated management of citrus greening, the treatment consisting of zinc sulphate + manganese sulphate + boric acid, tetracycline hydrochloride and 2,4-dichlorophenoxyacetic acid (2,4-D sodium salt) was proved significantly superior to other treatments in reducing the percent disease index (13.99) and providing 59.29 percent disease control with significant increase in yield (54.87 kg/tree) as compared to control (38.32 kg/tree).

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