IRANIAN JOURNAL OF PLANT PATHOLOGY
Year:2013 | Volume:49 | Issue:2 (194)|Papers Count:21

After studying of the plant parasitic nematodes, collected from different plants and regions of Iran, seven species belong to Paratylenchus, including P. arculatus, P. colinus, P. conicephalus, P. neoprojectus, P. similis, P. straeleni and P. veruculatus were identified. Among them, P. colinus, P. neoprojectus and P. veruculatus are new records for Iranian nematode fauna that full description of them and also that of P. straeleni were given. These species were collected and identified from different hosts and localities as follows: P. colinus from the rhizosphere of apple trees in Divandareh, Kurdistan province, P. neoprojectus from both shade-trees in Tabriz and apricot seedlings in Sanandej, and P. veruculatus from sugarcane fields of Khuzestan province. Furthermore, a dichotomous key for identification of Iranian species of this genus was presented.

To identify pathotypes, phenotypes and genetic diversity of Iranian wheat leaf (brown) rust (Puccinia triticina) isolates, eleven samples were collected from different geographical regions. Pathotype and phenotype of each isolate was identified and the intergenic spacer of rDNA (IGS1) wassequenced. One PCR product (870 bp) was recovered from all isolates. Multiple sequence alignments showed 0.0 to1.2 % difference indicating that IGS1 sequences were highly conserved among all analyzed isolates. The unrooted phylogenetic tree of sequences was drawn based on neighbor joining (NJ) procedures. Based on NJ tree, all sequences were grouped into six different clades. The results showed no direct relationship between P. triticina pathotypes and rDNA IGS1 sequence differences.

The rice brown spot disease caused by Bipolaris oryzae is distributed in most of rice cultivated regions and some time causes economic recession. Plants are under attack by diverse pests and pathogens. They have evolved different mechanisms to survive against invaders. Similar to the other plants, rice employs structural and chemical barriers to defense against pathogens. The pathogenesis- related proteins have crucial role in chemical defense. In this survey, the expression pattern of several resistance genes (Thionin, Defensing, Allen oxide synthase, Proxidase) were considered in two rice cultivars under treatment of B. oryzae fungus, using Quantitative Real Time PCR. The Tarom and Khazar cultivars have been selected as susceptible and resistance cultivars, respectively. Results showed considerable increase in expression rate of all genes in both genotypes after inoculation than that of control plant in every hours. Data analysis using T-student test program also showed that there are significant differences between Tarom and Khazar cultivars with the expression of mentioned genes. The findings proves that Thionin, Defensin, Allen oxide synthase and Proxidase genes play a prominent role in rice resistance to B. oryzae. This result can be an introduction to use of mentioned genes for overexpression and generate resistant rice to destructive brown spot disease in breeding program.

There are many reports that Fusarium species causing wilt (F. oxysporum f. sp. ciceri) and root rot (F. solani) are among the main causes of yield loss in chickpea production. Since chemical treatment against these fungi are not so effective, it seems that application of non-toxic chemicals to induce defense mechanism in plant host is desirable. Induction of host resistance in chickpea against these diseases using salicylic acid (SA) and chitosan were investigated. Foliage spray of solutions in 0, 200 and 400 ppm concentration were applied. Plants were grown in growth chamber; at 24 oC and 16: 8 (L: D) photoperiod for forty days. Application of 400 ppm of SA had a significant effect on reduction of wilt symptoms, but other applied concentrations of SA had no significant effects on root rot symptoms. Application of 400 ppm concentration of chitosan had a significant effect on wilt symptoms reduction, whereas application of 200 ppm of chitosan had partial effect on reduction of root rot symptoms. An in vitro experiment carried out, application of different SA & chitosan concentrations had a direct effect on mycelium growth of both fungal species on PDA. The level of free SA variations in plant tissues treated by 400 ppm concentration of SA was detected by using HPLC method in different post inoculation intervals. The results indicated that the free SA levels decreased after 168 h post inoculation, and wilt disease symptoms appeared gradually afterwards. Chitinase and b-1, 4 glucanase enzymes activity and total phenol content in chickpea leaf tissues were evaluated as induced resistance indicators (among others) at 0, 48, 96, and 168 h after inoculation. Increased levels of enzymes activities were observed in 200 ppm of SA however, no significant effects on root rot disease control were obtained in this treatment. The results would suggest that the application of this elicitor (SA) could have partial effects on chickpea wilt symptoms reduction. Maximum levels of indicators activities were observed in 400 ppm of chitosan however, no significant effects on root rot disease control were obtained in this treatment. The results would support the idea that application of this elicitor (chitosan) can play an important role in inducing systemic resistance in chickpea plants against Fusarial wilt disease.

Fire blight is important disease of pome fruit trees. The role of mitochondrial electron transport chain in interaction of host-pathogen has been verified. The purpose of this research was to study the role of chloroplast electron transport chain in this interaction and to determine the eventual site of pathogen effecter protein (s) influence. For this, effects of several chloroplasts electron transport chain inhibitors including Methylviologen, Glutaraldehyde and Diuron, with various sites of inhibition was evaluated on symptoms progress. All tests were performed and compared on in vitro shootlets of apples, MM-111 (tolerant) and MM-106 (susceptible) and pears, Harrow Sweet (tolerant) and Spadona (susceptible). For distinctness of necrosis causes by inhibitory effects inhibitors from those from pathogen, in a series of primary experiments, the effects of inhibitors concentrations were evaluated on shootlets in absence of pathogen and provided at least 168 h without necrosis of inhibitors. Subsequently, effects of inhibitors were evaluated on necrosis of pathogen after shootlets inoculation. The results showed that chloroplasts electron transport chain inhibitors retarded symptom progress in all tested cultivars. The results confirm previous results observed by uracil inhibition of chloroplasts and represent more evidence for role of this chain in host/E. amylovora interaction. In addition, according to impeding site of inhibitors, NADP reduction complex is the most probable site of effecter proteins interaction in host chloroplasts.

Antagonistic effects of four isolates of Paecilomyces lilacinus and an isolate of Isaria farinosa on root-knot nematode Meloidogyne javanica were examined under in vitro and greenhouse conditions. Root-knot nematode as well as fungal isolates were identified using morphological and species specific primers. Infection ability of different fungal isolates on the eggs of M. javanica and effects of fungal culture filtrates on egg hatching and mortality of M. javanica juveniles (J2) were tested in vitro. Results indicated that nematode eggs parasitization percent by various fungal isolates and J2 mortality as well as egg hatching inhibition of the culture filtrate of these fungi have variable effects on target nematode. P. lilacinus (isolate P3) showed more efficiency than the other isolates. In greenhouse experiment, effects of P. lilacinus and I. farinosa on tomato plant growth factors and nematode population were tested. Fungal isolates were propagated on sterilized wheat seed and mixed (0.5% w/w) with autoclaved soil. Four leaf seedlings of tomatos were transplanted into each pot and inoculated by4000 eggs and juveniles (J2) after ten days. Pots were kept in greenhouse for two months. Results showed that P. lilacinus isolate P3, P1, P4, and P2 and I. farinosa had 65%, 44%, 42%, 29% and 23% nematode conrol, respectively, indicating a good potential of some isolates in reducing nematode population.

Cucurbit yellow stunting disorder virus (CYSDV), a member of the genus Crinivirus (family Closteroviridae), causes considerable losses in cucurbits. Using of the resistant cultivars is a potential option for control of the disease induced by CYSDV. In this research twenty five accessions of Cucumis melo, including local melons, collected from different regions in Iran, were evaluated under controlled conditions for their reaction to Iranian CYSDV isolate, (Bushehr). Inoculations were made at first true leaf-stage with Bemisia tabaci. Assays included mean of disease severity index 6 weeks after inoculation, time of symptoms development (infection percentages 20 days after inoculation) and virus concentration in plant (mean ELISA value 8 weeks after inoculation). Only two accessions Chorook zard and Til zard showed delay in symptom development, low disease severity index and low concentration of the virus. They were considered resistant to the virus and tolerant to the disease. The accessions Zard khareji, Zard moshabbak dorosht, Zard talaee and Sefid because of low concentration of the virus in ELISA and Sirjan and Kashefi because of delay in symptoms development were resistant to virus. Yazdi, Zard talaee, Sirjan, Zard moattar and Sefid had low disease severity index and rated tolerant. Other accessions were highly sensitive to the virus.

In order to identify the nematode fauna of the family Qudsianematidae Jairajpuri, 1969, more than 150 soil samples were collected from different locations of Chaharmahal va Bakhtiari province during 2009 - 2011. The nematodes were extracted from soil samples by centrifugal flotation method, killed by adding hot 4% formaldehyde solution (De Grisse 1969) and transferred into pure glycerin. Permanent slides were made and morphological characters were studied using light microscopy. Drawings and morphometrics were done using a drawing tube attached to the microscope. In this study, seven species belonging to the family Qudsianematidae, namely Eudorylaimus sabulophilus, E. bombilectus, Crassolabium cylindricum, Discolaimus agricolus, D. bicorticus, Ecumenicus monohystera and Eudorylaimus subdigitalis were identified from which the first four species are new reports for Iran’s nematode fauna.

During the last four years (2009- 2012) several reports of forest tree decline were received, complaining about serious damage and death of many trees. The disease has been spread throughout forests of Zagross mountains which extend from north west to south west of Iran covering approximately 4, 000, 000 ha. Also the disease has been spread throughout forests of Alborz mountains in the north of Iran. The infected tree species included Q. brantii, Q. castaneifolia and Zelkova carpinifolia. Q. brantii Lindl. (Persian oak) has dominated in all parts of Zagros mountains from north to south, especially on the southern side of mountain in Ilam, Lorestan, Kohgilouyeh va Boyer-Ahmad, Fars and Kermanshah provinces. Zelkova carpinifolia (Pall.) Dippel, and Quercus castaneifolia C.A.Mey are native to the Caucasus and Alborz mountains in northern parts of Iran. The decline began with browning of the leaves, viscous liquid exudatation on the branches and trunks resulting in a brown-black discoloration of bark and woody tissues. In the winter of the next year fungal growth induces a typical charcoal-black surface on diseased branches and trunks. The perithecia of B. mediterranea were observed in a black carbonaceous layer on the stem surface erupting from the declined trees and the ascospores were visible under light microscope. Perithecia were obovoid, containing, amyloid asci, with dark brown ellipsoid ascospores, with straight germ slits along the spore-length. On the basis of these morphological characteristics, the fungus was identified as Biscogniauxia mediterranea. Pathogenicity tests were conducted using an isolate of B. mediterranea on sixmonth- old Q. brantii seedlings using a mycelial plug of B. mediterranea colonized potato-dextrose agar and the symptoms were observed after two months and the same fungus was re-isolated. Based on previous studies infections occur in healthy living trees as endophyte and then become invasive under water stress conditions in most reports. B. mediterranea has been reported to be aggressive on drought stressed hosts. During the last ten years climate changes has occurred in Zagros forests and resulting drought stress extending desert areas followed by occurrence of the fine dust phenomenon has reduced the photosynthesis of the forest trees and making them more vulnerable to the disease. In spite of tolerance of Persian oak species to the range of temperatures from -31°C until+45, the incidence of charcoal disease has been increased dramatically on Q. brantii forests. On the basis of our current knowledge, Q. brantii represents a new host of B. mediterranea and this is the first report of Persian oak charcoal disease outbreaks throughout Zagros mountain forests of Lorestan, Ilam, Fars, Kohgiloye va Boyer – Ahmad and on Zelkova carpinfolia in Daland forests of Gorgan area.

Three species of Tuberculina viz. T. persicina, T. sbrozzii and T. maxima are newly reported from Iran. Information about distribution and host range of species is provided. Each species is re-described and illustrated by photomicrographs. A key for reported taxa is also provided.

In summer of 2009, a disease with symptoms such as stem and trunk cankers were seen in apple Golden Delicious cultivars trees in west Azerbaijan orchards. The disease was in progress due to environmental stresses such as drought and frostbite. Bark and vascular system of the tree was darkened at the site of infection and cankers were scaling on main trunk and branches. To isolation of the fungi, infected tissues were surface sterilized for 3 min in 70% ethanol and were washed 3 times with sterile distilled water. Pieces were taken from sterilized tissues and were transferred to PDA, and then were kept at 25 oC, in the dark. The growing fungi were purified by hyphal tip method. To induce the formation of asexual fruiting bodies, the method of using pine needles were used. Inoculated Petri plates were kept for 4 weeks at 25 oC under nearultraviolet light (near-UV) with 12: 12h photoperiod to produce the pycnidia (Pavlic et al.2008). Different macroscopic and microscopic characters such as color and growth characteristics of colony, color, shape and dimensions of conidium, conidiogenous cell and pycnidium were studied. Also, the ITS region of one selected isolate was amplified and sequenced. Based on morphological and ITS sequence analysis, Diplodia malorum was identified. Descriptive characters of the species are as: Colony at first white, after two weeks turned to olive buff to olive greenish and finally after three weeks became dark. Conidiomata pycnidial, 600×500 mm in diameter, solitry or aggregated, spherical to oblong, dark brown to black, immersed, partially erumpent when mature. Conidiogenous cells hyaline, smooth, cylindrical, swollen at the base, 7 - 16×2.5 - 5 mm, proliferating at the same level to produce periclinal thickening, or proliferating percurrently, giving rise to 2-3 annelations. Conidia aseptate, thick-walled, smooth outer surface and verruculose inner surface, oblong to cylindrical with broadly rounded ends, hyaline, becoming dark brown and 1-septate after release from the pycnidium (rarely two septum), 17.5-34×11-16 mm. The results of Blast search showed high similarity (100 percent) of the studied isolate with the isolates belong to Diplodia malorum deposited in Gene bank (Phillips et al.2012). The pathogenicity tests were carried out on potted 2 years old seedlings and fruits of apple Golden cultivar based on completely randomized design. After eight weeks post inoculation, symptoms of bark darkening and canker were seen in inoculated areas. In fruits, six days after inoculation the brown rot symptoms were seen in the inoculated area which was progressed into the fruit flesh. In the controls, no symptoms were developed. Reisolation and identification of the inoculated fungi were done from the newly infected tissues and fulfilled Koch's postulates. To our knowledge, this is the first report of isolation and pathogenicity confirmation of Diplodia malorum from apples in Iran.

The stem and bulb nematode, Ditylenchus dipsaci (Kuhn, 1875) Filipjev, 1936 is one the most destructive plant-parasitic nematode which is widely distributed in temperate regions of the world. It is a serious pest on a wide range of crops including lucerne, red clover, sugar beet, maize, onion, garlic, strawberry and many other agricultural and horticultural plants, especially flower bulbs such as narcissus, hyacinth and tulip. Ditylenchus dipsaci has been reported from alfalfa, garlic and onion from Iran. Occurrence of D. dipsaci on strawberry, which was first reported in 1913 from Britain, has not already been observed in Iran. During a survey of the strawberry fields in Bahnamir and Babol cities of Mazandaran province, symptoms of disease including dwarf, leaf shrinkage and deformities, reduced petiole length and surface of lamina, reduced fruit size and reduced fruit setting in severe infections were observed. The nematodes in the soil and aerial parts of the infected strawberry plants were extracted by tray method, fixed and transferred to the anhydrous glycerin. After preparing permanent slides, the nematode was studied by light microscopy. On the basis of morphological and the following morphometric characters, the population extracted from infected strawberry, identified as Ditylenchus dipsaci: Females (n=10): L=1169±122 (1011-1411) mm, a=44.1±2.9 (40.7-51.0), b=6.3±0.8 (5.0-7.2), c=14.9±1.6 (13.2-18.6), c'=5.1±0.4 (4.6-5.7), stylet=11.1±0.7 (10.2-11.8) mm, V=78.9±2 (74.5-81.3). Males (n=5): L=1183±175 (947-1430) mm, a=50.2±4 (46.2.-50.8), b=6.4±0.8 (5.8-7.6), c=14.8±1.2 (13.5-16.2), c'=4.8±0.5 (4.2-5.5), stylet=11.4±0.2 (11.1-11.6) mm, spicules=26±1 (24.8-27) mm, gubernaculum=7.7±0.3 (7.3-8.0) mm.Initial investigation indicated that approximately 90% and 40% of 20 and 10 strawberry fields in Bahnamir and Babol cities, respectively, showed the symptoms of disease. The occurrence of disease in different fields was estimated between 20-70%, with respect to the age of cultivation. The average number of nematodes in infected shoots obtained from five plant samples per each field was 147 nematodes per gram of plant tissue.

Safflower (Carthamus tinctorius L.) of the family Asteraceae is an annual broadleaf oilseed crop with 35-40 % oil. It was originated from southern Asia and is known to have been cultivated in China, India, Iran and Egypt. In Iran the safflower cultivation area has increased over 10000 ha in 2008. Some pathogenic fungi such as Pythium ultimum, Phytophthora drechsleri, Puccinia carthami, and Fusarium oxysporum f. sp. carthami affect safflower crop in the country. During summer of 2011 safflower plants showing yellowing and wilting were collected from fields in Bajgah, Shiraz (Fars province). Segments of root and crown were surface sterilized using 0.5% sodium hypochlorite, plated on acidified PDA and incubated at 25 °C. The growing colonies were purified using single spore method. The colonies appeared yellow pink on PDA. Carnation leaf agar was used for macroconidium formation. Macroconidia were curved with five septa; Chlamydospores were round and verrucose; no microconidia were produced. The colonies on acidified PDA at room temperature produced light yellow aerial mycelium and appeared dark pink when looked upon at the back of Petri plates.. Based on morphological features and available sources such as Nelson et al. (1983), Burgess et al. (1994) and Leslie and Summerell (2006), the fungus was identified as Fusarium compactum. This fungus has been previously reported from Iran on wheat by Zare and Ershad (1997). To study the pathogenicity of the fungus to safflower, an inoculum was prepared according to Westerlund et al. (1974). The fungus was grown on millet seeds. The colonized seeds were placed around crown and roots of 14-day old and 60- day old safflower plants and grown under greenhouse conditions. After four weeks, the inoculated plants developed leaf yellowing and root and crown necrosis. The fungus was reisolated from infected plants and identified to fulfill the Koch's postulates.

Brown stripe of rice disease was first reported from rice nursery in Mazandaran province. The agent was identified as Acidovorax avenae subsp. avenae (Aaa) (Rahimian.1986, Iran Agric. Res.5: 63-71).Assessment of the possible genetic diversity of Aaa isolates in eastern Mazandaran was attempted in the present study. On the basis of biochemical and physiological characteristics, the isolates were identified, phenotypically, as Aaa. All isolates were oxidase and catalase positive and hydrolysed starch, casein and Tween 80. They did not assimilate sucrose or salicin but were capable of using sorbitol and monnitol as carbon sources (Rahimian.1986, Iran Agricultural Res.5: 63-71). Genomic DNA of isolates was extracted and subjected to rep-PCR using REP and BOX primers, under the previously described conditions with only minor modifications (Versalovic et al.1991, Nucleic Acids Res.19: 6823-6831). Dendrograms were constructed using data sets obtained from amplified fragments, using the Jaccard coefficient, the UPGMA algorithm and NTSYS-PC program. Comparison of the genomic fingerprints demonstrated that, at 20%, 50%, 80% similarity levels isolates formed 7, 16, 23 clusters with BOX primer, respectively and 5, 13, 19 clusters with REP primer at the stated levels of similarity, respectively. The clusters produced in REP-PCR were highly congruent with those of BOX-PCR. There has been no report on the presence of such any high level of genetic diversity in this bacterial subspecies elsewhere.