فصلنامه بیماریهای گیاهی
سال پنجاه و پنجم شماره 4 (زمستان 1398)

Recent improvements in instrumentation and data mining have made it possible to investigate the effects of various stressors on primary and secondary metabolisms. In this study, in order to study mechanisms of pathogenesis and identify the pathogenesis-related metabolic pathways, changes in metabolic profiles resulting from a compatible interaction between canola cultivar (Hyola 401) and Leptosphaeria maculans (anamorph Phoma lingam) (Phkv102 isolate) were investigated by GC-MS chromatography using the polar fraction of plant extract. Under completely controlled conditions, 14 days after planting, when cotyledons were completely unfolded, the plants were inoculated by 107 spores per ml suspensions and water (control) by drop-wound method, respectively. The tissue samples were taken 48 hours after inoculation and were shock frozen immediately. Metabolites were extracted using methanol as solvent, identified and quantified with GC-MS and subjected to statistical analyses. Results indicated that 70 metabolites showed significant difference over their controls at P≤0.05, which is caused by changes in 28 metabolic pathways. Infection of canola cultivar Hyola 401 to P. lingam resulted in change in pathways related to the host cell wall, phenolic compounds, alkaloids and terpenoids biosyntesis, energy generator, hormone biosynthesis, signal transduction.The abundance of a number of sugars was also decreased, suggesting the crucial role of these carbohydrates in pathogenesis in an incompatible interaction of  P. lingam and a susceptible cultivar.

Autophagy is a degradation process in eukaryotes through which damaged or unwanted intracellular components are degraded. This process is also involved in plant disease resistance, although its mechanisms are not precisely known. Autophagy is regulated by multiple autophagy-related proteins (ATGs).In this study, we investigated the possible impact of two Barley yellow striate mosaic virus (BYSMV) proteins, i.e., phosphoprotein (P) and ancillary protein 3 (P3), on four important genes involved in autophagy (ATG2, ATG6, ATG7, and AGO1) in N. benthamiana. P and P3 genes were cloned in pCAMBIA-1302 vector under the control of the 2 × 35S promoter and Hemagglutinin tag. Constructs of each gene were agroinfiltrated in the abaxial side of the N. benthamiana leaves. Five days after agroinfiltration, the expression level of these genes was measured using RT- qPCR. The results showed that expression of ATG2, ATG6 and ATG7 genes increased in all treatments (P, P3, P+P3).The level of ATG2 expression was 5.57, 15.6 and 5.6 fold, in P/GFP, P3/GFP and P/P3/GFP treatments, respectively, while a 1.5-fold reduction was obtained in expression of AGO1 in all treatments. These results implied that P and P3 proteins of BYSMV can modify the expression of autophagy related genes in N. benthamiana plant. These findings suggest the involvement of AGO1, ATG6, ATG7 and ATG2 in immune responses of N. benthamiana against BYSMV, which provide a better understanding of plant host defense mechanisms against virus infections and might be an opportunity to exploit a novel antivirus approach.

Grape powdery mildew (PM) caused by plant pathogenic fungus Erysiphe necator, is the most important disease on grapevine in world and Iran. In order to study the spread of this disease in vineyards of Sistan region, 30 vineyards in three county including Zabol, Zahak and Hamoon were selected during years 2017 and 2018. The vineyards were visited on a weekly basis to record PM incidence (I) and disease severity (S). Based on disease incidence, there were no significant differences between counties and vineyards in first, second and combined two years, and in combined two years Zabol and Zahak (57.6%, 57.2%) had the highest and lowest percent of disease incidence.Based on leaf and fruit disease severity, there were significant differences between counties and two years (P <0.001) and in combined two years Hamoon and Zahak (23.1%, 20.6%) had the highest and lowest percent of infection in leaves. Based on the fruit disease severity, Zabol and Zahak (25.4%, 21.8%) had the highest and lowest percent of infection in fruits respectively. Based on area under disease progress curves, there were no significant differences between years, counties and vineyards in disease incidence but in leaf and fruit disease severity, there were significant differences between years and counties and in combined two years Hamoon and Zabol (696.9 and 680.3) had the highest of leaf and fruit infections respectively.

Xanthomonas arboricola pv. pruni (Xap) is a causal agent of leaf, fruit spot, and branch bacterial canker in warm and humid conditions on stone fruit trees. Herein, the pathogenicity of 18 Xap strains from Khorasan-Razavi Province, Iran was evaluated in greenhouse trials on Santha-Rosa plum saplings. Some pathogenicity factors including pXap41, T3SS effectors, and virulence features such as motility, biofilm, xanthan and biosurfactant production, cell wall degrading enzymes and their possible correlation with pathogenicity rate were investigated. The studied strains were divided into two severe and weak pathogenicity groups based on percentage of necrotic lesions. All 18 strains besides ICMP7485, had pXap41, xopE3 and xopA effector genes. In some cases, there was a direct correlation between disease severity and factors involving in virulence. For example, two strains, ShL45 and NB28k, showed the highest percentage of necrosis spots (62.5%), produced high levels of biosurfactant and biofilm that indicated significant differences with other strains at 5% probability levels. It seems that xanthan has no significant effective role in Xap virulence. The rate of swarming in 92.3 % of strains belonging to the severe pathogenicity group was more than the swimming (P≤ 0.05). All strains were also capable of secreting different amounts of enzymes, including protease, cellulase, polygalactronase, and pectatelyase. However, the rate of secretion in all them was not entirely consistent with virulence. It is suggested that more Xap strains from different regions of the country, along with other X. arboricola pathovars, should be tested to get results that are more accurate.

Beet mosaic virus (BtMV), the only potyvirus infecting beets, is distributed worldwide in beet growing areas. Here we report the full-length genome sequence of a BtMV isolate from Iran (Ir-VRU), which has enabled us to better understanding the evolutionary history of the virus. The complete nucleotide sequence of Ir-VRU genome was determined using degenerate and specific primers and 5'-RACE and The complete genomes consisted of 9591 nucleotides, excluding the poly-A tails. The genome has a 165 nt 5´-UTR and a 168 nt 3´-UTR. The RNA of Ir-VRU potentially encodes a single polyprotein of 3086 amino acid residues and has a deduced genome organization typical for members of the genus Potyvirus. Study on five different BtMV isolates showed that Iranian isolate closelly related to German and Chinese isolates, and on the other hand, it showed more differences with American isolate. Phylogenetic analysis, along with other potyviruses, also presents the Ir-VRU as a member of the BCMV supergroup.

Potato virus Y, the type member of the genus Potyvirus, family Potyviridae, is one of the most destructive viruses of tobacco worldwide. To determine the relationships between incidence (I) and severity (S) of PVY disease in five cultivars in tobacco fields in Golestan province, several surveys were conducted during July to October 2017 at 15 days intervals in tobacco fields of Gorgan, Aliabad and Minoodasht. Two disease indicators (I and S) were evaluated in 50 plant samples for each farm. The presence of PVY was detected by double antibody sandwich-ELISA (DAS-ELISA) using PVY-specific polyclonal antibodies. The rate of PVY infection was different among tobacco cultivars and the maximum and minimum infection rates were belonged to PVH03 (33.6%) and Burley (6.2%) cultivars respectively. Based on the statistical analysis of data, there was a significant difference for disease indicators on different tobacco cultivars, as the maximum and minimum symptom severity mean value were evaluated in PVH03 (4.44) and Burley (1.82) cultivars, respectively. Also, Square Root model (Sqrt transformation of I and S) in 60 % of the fields (R2 = 82.32 %), and Allometric model (natural logarithm transformation of I and S) in 30 % of the fields (R2 = 89.90 %) showed the best fitness for the collected data, andwere the best model to describe I-S relationships in PVY disease in tobacco fields. Based on the regression slope of Sqrt transformed values of I and S, the cultivars were classified in three groups, including PVH03 (with the highest slope of 1.005), NC100 (with the moderate slope of 0.729) and K326, Burley and Basma (with thelowest slope of 0.6 – 0.66).

Medicinal plants have undoubtedly been considered by human beings since ancient times. Sambucus ebulus (Caprifoliaceae) is one of the best known medicinal herbs since ancient times and is native to Southern and Central Europe, Northwest Africa, and Southwest Asia (especially Northern Iran) (Westwood 1985; Jabbari et al. 2017). It plays an important role in traditional medicine in many countries from different regions of the world (Jabbari et al. 2017). During the summer of 2014, leaf spots have been observed on S. ebulus plants in Gilan and Mazandaran provinces, Iran. Symptoms on infected leaves appeared as brown necrotic spots that surrounded by distinct dark brown haloes and were circular to irregularly in shape. During the summer of 2014 and 2015, sampling was done in Gilan and Mazandaran provinces from symptomatic leaves of S. ebulus and the fungus was isolated on 2% water agar (WA 2%) and potato dextrose agar (PDA). Pure fungal cultures were obtained by picking up a single spore on PDA. For morphological identification, the isolates were cultured on Potato Carrot Agar medium (PCA) and incubated at 25 °C under cool-white fluorescent with 16 h dark, 8 h light photoperiod for 14 days (Simmons 2007). The seven-days-old colonies on PCA were dark olive to brown in color and 70 mm in diameter. Primary conidiophores were often simple, reached to 70 µm in length and produced numerous conidia in a simple long chain. The conidia were 8–12 × 20–45 µm in size, egg or club shaped and contained 3-5 transverse and 1-2 longitudinal septa (Fig. 1a-c).According to macro- and micro morphological characters, all recovered isolated were identified as Alternaria tenuissima which was later synonymized under the name of A. alternata by Woudenberg et al. (2015). PCR amplification of the nuclear ITS-rDNA and RNA polymerase II second largest subunit regions was performed using primers ITS1/ITS4 and RPB2–5F2/fRPB2–7cR respectively (White et al. 1990, Sung et al. 2007; Liu et al. 1999). The sequence of NG1 isolate was deposited into the GenBank with the accession numbers MK212914 for ITS-rDNA and MK262741 for rpb2 region and living cultures of this strain were deposited in the Agriculture Biotechnology Research Institute of Iran culture collection (ABRII 10119). Phylogenetic estimates were evaluated using the Maximum Parsimony Analyses in MEGA 6.0 (Tamura et al. 2013). The results of phylogenetic study showed, NG1 isolate was clustered in a well-supported clade (100%) including sec. Alternaria and related to Alternaria alternate isolates (Fig. 2).  Pathogenicity test was performed in greenhouse condition on the detached leaves taken from healthy S. ebulus. Ten healthy leaves per isolates were cleaned with sterile water and were inoculated with conidial suspension containing 1 × 106 conidia/mL using the spray method. The control leaves were inoculated with distilled water. Leaf spots similar to the original symptoms were observed on all inoculated leaves after 5 days since inoculation and the fungus re-isolated from leaf lesions (Fig. 1e-g). Also, no symptoms were seen in controls.