Nighat Sarwar

Biochemical alterations in leaves of resistant and susceptible cotton genotypes infected systemically by cotton leaf curl Burewala virus

Leaf curl disease caused by Cotton Leaf Curl Burewala virus (CLCuBuV) has been recognized as serious threat to cotton in Indian subcontinent. However, information about cotton–CLCuBuV interaction is still limited. In this study, the level of phenolic compounds, total soluble proteins, and malondialdehyde (MDA) and the activities of phenylalanine ammonia-lyase (PAL), peroxidase (POX), catalase (CAT), proteases, superoxide dismutase (SOD), and polyphenol oxidase (PPO) were studied in leaves of two susceptible (CIM-496 & NIAB-111) and two resistant (Ravi and Co Tiep Khac) cotton genotypes. Disease symptoms were mild in the resistant genotypes but were severe in highly susceptible genotypes. The results showed that phenolic compounds, proteins, PAL, POX, CAT, proteases, SOD, PPO, and MDA play an active role in disease resistance against CLCuBuV. The amount of total phenols, proteases, MDA, and PPO was significantly higher in leaves of CLCuBuV-inoculated plants of both resistant genotypes as in non-inoculated plants, and decreased in CLCuBuV-inoculated plants of both susceptible genotypes over their healthy plants. POX, protein content, SOD, and PAL activities showed lower values in resistant genotypes, while they decreased significantly in susceptible genotypes as compared to the noninoculated plants except PAL, which showed non-significant decrease. CAT was found to be increased in both susceptible and resistant genotypes with maximum percent increase in resistant genotype Ravi, as compared to non-inoculated plants. The results showed significantly higher concentrations of total phenols and higher activity of protease, MDA, SOD, and PPO in resistant genotype Ravi after infection with CLCuBuV, suggesting that there is a correlation between constitutive induced levels of these enzymes and plant resistance that could be considered as biochemical markers for studying plant-virus compatible and incompatible interactions.

Field Evaluation of Mungbean Recombinant Inbred Lines against Mungbean Yellow Mosaic Disease Using New Disease Scale in Thailand

Studies were conducted to identify the sources of resistance in mungbean recombinant inbred lines (RILs) in Thailand against mungbean yellow mosaic disease (MYMD). 146 mungbean RILs in series were evaluated in a field including resistant parent NM-10-12-1 and susceptible parent KPS 2 during summer 2008 under high inoculum pressure. The RILs were subsequently scored for disease symptom severity ratings (DSSR) using a new scale. Observations regarding DSSR and % disease index (%DI) showed that the tested RILs responded differently to the disease. A large number of RILs (132) were found highly susceptible, 12 were susceptible, 3 were tolerant and one was resistant. Overall screening results showed that three RILs, viz. line no. 30, 100 and 101 had minimum DSSR and % disease index thus they are good source of resistance to MYMD in spite of high disease pressure and can therefore be used directly as varieties to manage the disease in Thailand.

CHINA ROSE (HIBISCUS ROSA-SINENSIS): A NEW NATURAL HOST OF COTTON LEAF CURL BUREWALA VIRUS IN PAKISTAN

China rose (Hibiscus rosa-sinensis) is a perennial ornamental plant grown throughout the tropics and subtropics. China rose plants with severe vein thickening/ greening, leaf curling, and enations on the lower leaf surface were found near cotton fields at the Nuclear Institute of Agriculture Biology (NIAB) in Faisalabad. The symptoms of disease exhibited on China rose plants were very much similar to Cotton leaf curl disease exhibited by cotton plants. DNA was extracted from ten naturally infected symptomatic China rose plants and subjected to PCR using begomovirus and betasatellite specific primers. The expected products of 2.8Kb and 1.4Kb for begomovirus and betasatellites were amplified, respectively. The begomovirus and its cognate betasatellite isolated from China rose were sequenced and submitted to the database with accession numbers HG003876 and HG003877, respectively. Owing to a 99% nucleotide sequence identity, this virus was designated as an isolate of Cotton leaf curl Burewala virus (CLCuBuV). Similarly the sequenced betasatellite showing 96% nucleotide sequence identity was designated as an isolate of Cotton leaf curl Multan betasatellite (CLCuMuB). The symptomatic induction of Cotton leaf curl disease (CLCuD) by indexing in LRA-5166, a CLCuBuV susceptible genotype of cotton that resists Cotton leaf curl Multan virus (CLCuMuV), further confirmed the presence of CLCuBuV in China rose. To our knowledge, this is the first report of CLCuBuV and its cognate betasatellite in infected China rose.

Evaluation of Bt-cotton Genotypes for Resistance to Cotton Leaf Curl Disease under High Inoculum Pressure in the Field and Using Graft Inoculation in Glasshouse.

Bt-cotton germplasm, consisting of 75 genotypes was evaluated against cotton leaf curl disease (CLCuD) under high inoculum pressure in the field and using graft inoculation in glasshouse by visual symptom scoring assessments. None of the tested genotype was found disease free under both evaluation tests. Under field conditions in 2011, 3 genotypes were found resistant, 4 moderately resistant, 3 tolerant, 2 moderately susceptible and one susceptible; in 2012, 3 genotypes were tolerant, 7 moderately susceptible, 5 susceptible and 38 highly susceptible; in 2013, one was moderately susceptible and 51 were highly susceptible with varying degree of percent disease index (PDI) and severity index (SI). However, through graft evaluation in glasshouse, none of the graft inoculated plant was symptomless. All tested genotypes showed disease symptoms with SI values ranging between 5.0 and 6.0, and latent period between 12 and 14 days. Of the 75 genotypes evaluated using graft inoculation, 11 were found susceptible with SI values of 5.0 to 5.4 while remaining 64 were highly susceptible with SI values of 5.5 to 6.0. Inoculated plants of all tested genotypes exhibited severe disease symptoms within 10 days after the appearance of initial symptoms. No reduction in SI value was observed until the end of the experiment i.e., 90 days after grafting. Information generated under the present study clearly demonstrates that no sources of resistance to CLCuD are available among the tested Bt-cotton genotypes. So, a breeding programme is needed to introgress the CLCuD-resistance from other resistant sources to agronomically suitable Bt-cotton genotypes.

PHYSIOLOGICAL RESPONSE OF COTTON LEAF CURL BUREWALA VIRUS-INFECTED PLANTS OF TOLERANT AND SUSCEPTIBLE GENOTYPES OF DIFFERENT GOSSYPIUM SPECIES

The effect of Cotton leaf curl Burewala virus (CLCuBuV) on cotton growth, yield, photosynthetic gas exchange, chlorophyll fluorescence and chlorophyll contents was studied in susceptible (CIM-496 and NIAB-111 from Gossypium hirsutum) and tolerant (Ravi from G. arboreum and Co Tiep Khac from G. herbaceum) cotton genotypes under glasshouse condition through artificial inoculation. Compared to the non-inoculated plants, the inoculated plants of susceptible genotypes showed stunted growth, reflected as significant decrease in plant height, internode length, number of leaves per plant, leaf area, number of bolls per plant, boll weight and yield per plant. Upon virus inoculation, transpiration rate decreased significantly in both susceptible genotypes while water use efficiency (WUE) was enhanced in CIM-496 compared to the non- inoculated plants. However, overall plant growth, net photosynthetic rate, leaf transpiration, stomatal conductance and WUE remained unaffected by CLCuBuV-infection in tolerant genotypes. Moreover, chlorophyll b and total chlorophyll content increased in one of the tolerant genotypes (Ravi) in response to CLCuBuV-infection while remaining unaffected in all other genotypes. Level of chlorophyll a and chlorophyll fluorescence (Fv/Fm) was not affected by CLCuBuV-infection in all genotypes. In conclusion, CLCuBuV infection adversely affected the overall plant growth and yield in susceptible genotypes while tolerant genotypes were able to maintain the growth and yield in the presence of infection. Reduced transpiration rate and increased WUE in susceptible genotypes and increased chlorophyll b and total chlorophyll content in tolerant genotypes on virus inoculation provided evidence for their significant involvement in CLCuBuV-tolerance mechanism in cotton.

Gibbago trianthemae causes Trianthema portulacastrum (horse purslane) blight in Pakistan

Trianthema portulacastrum (horse purslane) is among the most noxious summer annual weeds in Pakistan. During June 2010–2012, a severe outbreak of leaf and stem blight was observed for the first time on T. portulacastrum in Pakistan. Symptoms on leaves and stems were examined as round to oval straw colored spots with maroon margins. As the disease progressed, affected leaves became chlorotic and dried up causing severe defoliation and withering of stems. The pathogen was identified as Gibbago trianthemae on the basis of morphology and cultural characteristics. A pathogenicity test was performed and Koch’s postulates were fulfilled by re-isolation of the fungus G. trianthemae from diseased tissues of T. portulacastrum. This is the first report of G. trianthemae causing blight on T. portulacastrum in Pakistan.

Mungbean phyllody disease in Pakistan: symptomatology, transmission, varietal response and effects on yield characteristics

Mungbean plants showing typical symptoms of infection by a phytoplasma that causes phyllody disease (transformation of floral parts into green, leaf-like structures) were examined in field crops in both spring and summer seasons. The most common symptoms were floral virescence, phyllody, reduction in leaf size, extensive proliferation of branches and stunting of plants. Sometimes these symptoms were accompanied by longitudinal splitting of green pods followed by germination of green (physiologically mature) seeds, producing small plants. Phyllody disease pathogen was successfully transmitted from diseased to healthy mungbean plants using either grafting and the leafhopper Orosius albicinctus under laboratory conditions. Transmission via seeds and by insects that included leafhopper Amrasca devastans, pea aphid Acyrthosiphon pisum, beetles Brumoides suturalis, Hippodamia variegata, Menochilus sexmaculatus, and the whitefly Bemisia tabaci failed to produce disease under laboratory conditions. During the spring and summer seasons of 2009, a field trial of 162 exotic and indigenous mungbean genotypes obtained from 8 different geographical regions was conducted, aiming to shed light on the causes of susceptibility to phyllody disease. Sixty-one genotypes showed infection, ranging from 1.00% to 4.35%. Significant variation in all yield and morphological characteristics due to phyllody disease on mungbean plants depending on growth stages was also recorded. Comparison between healthy and diseased plants showed significant percentage reduction at all growth stages. Maximum reduction in all the tested charcateristics was recorded when plants were infected before flowering followed by infection at the flowering and pod-setting stages. The most severely affected plant variables at all infection stages were the number of seeds per plant, the number of normal seeds per plant and yield per plant. Severely infected plants produced no seeds but some partially infected plants produced very small, sickle-shaped, upright and leathery pods with degenerate seeds.

Effect of cucumber mosaic virus infection on morphology, yield and phenolic contents of tomato

Ten tomato genotypes were screened for their resistance against cucumber mosaic virus (CMV) and its vector Myzus persicae under natural infection in field, using aphids M. persicae under net-house and mechanical inoculation under greenhouse. Large differences were observed among genotypes for infection percentage (IP) and severity index (SI) among the testing methods used. All genotypes showing tolerance to CMV in the field or through aphid inoculation, however, become susceptible and highly susceptible after mechanical inoculation. All the test genotypes also showed susceptibility to the aphid M. persicae population. Plants inoculated with CMV showed substantial decrease in yield and yield-contributing parameters which varied with cultivars that probably depended upon its genetic make up. All the test genotypes exhibited 0.97–30.19% decrease in plant height, 11.47–52.65% decrease in root length, 46.56–95.56% decrease in fresh plant weight, 65.78–92.84% decrease in root fresh weight, 19.97–87.65% decrease in the dry weight of plants, 75.63–95.43% decrease in dry root weight, 69.51–95.65% reduction in the number of fruits and 89.04–99.89% decrease in yield per plants. After 15 days of inoculation, the quantitative analysis using double beam spectrophotometer showed an increase in total phenolics in CMV-inoculated plants as compared to un-inoculated plants among genotypes. Similarly the thin layer chromatography (TLC) on silica gel G indicated that the number of phenolic compounds was increased in most of the inoculated genotypes while in others they were either decreased or remained same.

EVALUATION OF TOMATO GENOTYPES FOR LATE BLIGHT RESISTANCE USING LOW TUNNEL ASSAY

Late blight (LB) caused by Phytophthora infestans is one of the major bottlenecks of low productivity of tomato worldwide including Pakistan. Transfer of resistance into elite lines is the principal way to combat this problem. However, lack of reliable methods to screen and evaluate plant resistance prior to breeding has been a limiting factor in the development of tomato varieties resistant to LB. In the present study, we screened 285 tomato genotypes of diverse genetic pool using a low tunnel assay for the first time to the best of our knowledge and confirmed that it is not an established method in use for the same purpose. Results over the multiyear (2013-2015) elucidated one resistant genotype LO6122 (Solanum arcanum) and 31 moderately resistant genotypes (one of S. arcanum and 30 of S. lycopersicum) to LB. Rest of the genotypes including mutants and differential plants having major LB resistant genes (Ph1, Ph2, Ph3 and Ph2 + Ph3) were ranged from susceptible to highly susceptible. Strong resistance conferred by LO6122 points out the presence of an alternate source of genes resistant to LB. Therefore, this line can possibly be used as a donor parent of resistance against virulent strains of P. infestans. This study also showed that low tunnel assay is practicable, efficient, simple, and cost effective in terms of space and pathogen inoculum required. It gives high output results to distinguish resistant and susceptible genotypes even when screening of large number of genotypes and breeding populations against LB is exercised.

Convolvulus arvensis, a new host for Alternaria solani causing early blight of Solanum lycopersicum in Pakistan

In March–April 2011, a severe outbreak of early blight disease caused by Alternaria solani was observed on Solanum lycopersicum (cultivated tomatoes) at the Nuclear Institute for Agriculture and Biology (NIAB), Faisalabad, Pakistan. A similar type of symptoms was also observed on Convolvulus arvensis (field bindweed, lehli). Based on disease symptoms, morphological characteristics and pathogenicity test, it has been confirmed that the A. solani could infect C. arvensis. To the best of our knowledge this is the first report of A. solani infecting C. arvensis in Pakistan.