During 2006-2007, many surveys are done from rapeseed fields in Mazandaran and Golestan Provinces (N. Iran) and the sexual form of Blackleg fungus as Leptosphaeria maculans (Desmaz.) Ces. & De Not. [Anamorph=Phoma lingam (Tode: Fr.) Desmaz.] was diagnosed on rapeseed debris both winter (R & C variety) and spring (HYOLA 401 hybrid and PF variety) type oilseed rape among OCT-NOV months. Ascocarps were globose, black, 300-500 mm diameter (Fig. 3 B) and observed on stem branches (Fig. 3 A). Asci were 80-120 ´ 15-22 mm, cylindrical to clavate, bitunicate wall and with eight spores.

Leptosphaeria maculans causes blackleg disease on Brassica oilseedcrops, which globally is one of the great threats for oilseed production. In order to determine virulence types of this fungi, several isolates were collected from canola fields in Mazandaran and Golestan provinces, Totally 12 isolates were selected and tested by three standard (definitional) cultivars including Westar, Quinta and Glacier in which two virulence types including PGT and PG2 were identified. In order to screen some existence local and some new canola genotypes, 24 genotypes of B. rapa, B. napus, B. juncea and B. nigra species were tested against PGT and PG2 isolates. As a result, among tested genotypes, Option and Champlain cultivars were resistance but PF and Adriana were moderate resistance to PG2 virulence type. In addition, Option and Champlain cultivars were moderate resistance to PGT virulence type of the disease.

Blackleg caused by Leptosphaeria maculans is one of the important diseases widely affecting canola production worldwide. Effectiveness of three plant defence related genes transferred from pea (Pisum sativum) to canola was tested against blackleg in two experiments. In the first experiment, three transgenes including PR10.1, Chitinase, and DRR206 were transferred from transgenic Wester variety into four commercial cultivars (Apollo, Sentry, OAC Triton, and MillenniUM 03) via backcrossing to examine the effects of different genetic backgrounds on disease response. In the second experiment, to study the effect of gene pyramiding on level of disease resistance, three transgenic Wester lines were crossed. Cotyledon inoculation was performed for indoor disease screening tests of blackleg. DRR206 in Sentry and Chitinase×DRR206 appeared to be the best combinations conferring disease resistance. Overall, DRR206 had the highest impact on disease, probably due to delayed infection development. Promising lines of this research could be utilized in breeding programs and cultivar release projects after field trials.

Blackleg disease of rapeseed, which is caused by Leptosphaeria maculans (Phoma lingam), is considered as one of the destructive diseases of crucifera family and has resulted in total yield loss in some locations. It appears as an endemic disease in many areas of the world, especially in rapeseed fields. In this study, the response of six winter cultivars of rapeseed to the two isolates of the fungus (Aliabad and Bandar Torkman) was evaluated. Plants were inoculated in different growth stages: cotyledon, and stages, 2.3-2.4, 2.1, and 3.2, using spore suspension under 23±1°C on the basis of established procedures for a successful infection. Inoculated plants were kept under plastic covers for 48 hours in order to provide a 100% relative humidity. After inoculation period, disease severity was assessed in cotyledon stage by the method of William and Delwich, in stage 3.2 by McNabb and Rimmer, and in stage 5 on the basis of Newman and Bailey, and also Mc Nabb and Rimmer. It was revealed that responses of Okapi and SLM046 to disease were homogenous in all stages. Okapi was recognized more susceptible and SLM046 more resistant than other cultivars under study. Talaych, Regent × Cobra, Fomax and Ceres which showed different reaction during different stages of inoculation, indicated the heterogenicity of response to blackleg disease in these cultivars.          

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.

Introduction: Canola is one of the important oil crops in Iran and blackleg disease caused by Leptosphaeriamaculansis an economically important disease of rapeseed especially in the northern provinces of Iran. At the beginning of the season in early autumn, L. maculans areinitiatesby air-borne ascospores released from infected stubbles of previous crops. These ascospores germinate and produce leaf lesions. The fungus then grows systemically from the leaf lesions to stem where cankers are produced which can result in major yield loss.Blacklegiscaused by a complex of at least two species of Leptosphaeria: L. maculans and L. biglobosa. Of these two species which L. maculans wasis much more prevalent and virulent than L. biglobosa and was reported from almost all oilseed rape growing regions of the world.A number ofmethods has been developed to differentiate isolates of these two groupsthrough morphological, physiological, genetic and molecular markers. Among these virulence markers of group pathogenicity are most widely used for characterizing the pathogen population. The purpose of this research work was to investigate isolates identification based on colony morphology and molecular methods. Moreover, for management strategies to be effectiveit is important to recognize the population structure based on pathogenicity groups and an understanding of possible shift in fungus population in thenorth of Iran.Materials and Methods: Infected plants of oilseed rape were collected from the northeast Iranduring 2013-2016. Diseased plant organs with clearly defined symptoms of the disease were used for the isolation of the fungi. All isolates were analyzed using a combination of morphological and physiological. In order to confirm the identification ofL. maculans from oilseed rape isolates, DNA was prepared using the standard method described previously. The ITS region of genome of all isolates was amplified using polymerase chain reaction with specific primers pair LmF, lmR. Some isolates were sequenced with ITS1 and ITS2 along with the 5.8S rRNAregion and then sequence data were compared with that of availableL. maculans isolates in Gene Bank.Aggressive isolates ofL. maculans collected in widely separated geographic regions were further divided into pathogenicity groups based on virulence of three differentBrassica napus cultivars.Results and Discussion: Morphological and physiological and molecular characterizations of 72 isolates were performed. Isolates grewslowly, the pycnidia of the fungus were black, globose to subglobose in shape and conidia were single-celled, hyaline. The most of isolates formed yellow and pigment formation was scored in six groups in PDB at 18°C in the dark on the shaker. The PCR detection showed that all isolates were amplified by L. maculans-specific primer pair and a 334-bp PCR product was reliably amplified from L. maculans. The sequence analysis of the ITS region revealed that the sequences had 99.7% sequence similarity with the ITS sequences of knownL. maculans isolates in GenBank by NCBI Blast search. This isolates were registered with accession number KX649997 and KX792142 in gene bank and this confirmed that the pathogen exists in Brassica napussamples. The role of morphological identification in virulence is complex and the production of pigment and mycelial growth is not always correlated with virulence. Therefore, based on amplification with type specific primers the amplified product fragment specific for group A was found in all isolates. It is important to know which pathotype is present, because the pathotypes ofL. maculans differ in the amount of damage they cause. Based on the reaction of isolates on differential cultivars all four group pathogenicity PG-2, PG-3, PG-4 and PGT were observed in aggressive isolates. The most isolates were classified to be a pathogenicity PG-4 virulent on three differentcultivars and for the first time wasidentified in Iran.Conclusions: In order to determine whether isolates belonging to the canker L. maculans are present in the north of Iran, initial species identification based on colony morphology was confirmed using molecular methods.Asthe pathogen causes considerable losses, thefast identification and pathotype determination are important foragriculture and successful management of blackleg disease. Our results showed that an understanding of possible shift in fungus populations of PG2 to PG4 will be of value in developing strategies for successful management of blackleg disease.

In the course of a research project during 2008-09, diseases of mulberry shoots in Izadshahr (Mazandaran), Astaneh Ashrafieh (Guilan) were investigated. The results showed that phloeospora leaf-spot caused by Phloeospora maculans on Morus species is a widespread disease in the above mentioned cities. Also, diatrype die-back on white mulberry, caused by Diatrype stigma, is a rather important disease in Astaneh Ashrafieh. A full account of the symptoms of the diseases and also details of their causal agents are presented here. Malberry is a new host for Cytospora cincta and Diatrype stigma in Iran. Specimens of the two diseases are preserved under IRAN 14695F and IRAN 14692F in the Herbarium of Iran (IRAN) at the junior author's address.