مجله دانش گیاه پزشکی ایران
سال چهل و سوم شماره 2 (پاییز و زمستان 1391)

Citrus blast disease caused by Pseudomonas spp is among the most injurious diseases of citrus. in the northern citrus growing provinces of Iran. To determine the species and pathovars of Pseudomonas strains associated with the disease in Mazandaran province and in the neighboring areas in Guilan and Golestan provinces and as well to assess their phenotypic heterogeneity and virulence, symptomatic samples were collected from the affected trees during 2009-2010. Among the strains isolated from the leaves and twigs, 115 strains appeared, in preliminary testing to be pathogenic to sour orange (Citrus aurantium) leaves. Based on biochemical properties, the isolates were grouped into 16 separate categories. Strains were positive in tests for hypersensitive reaction in tobacco, negative in production of oxidase but varied in the production of levan, arginine dihydrolase and rotting of potato tuber slices. Some isolates despite their ability to rot potato tuber slices, were not sufficiently similar to P. viridiflava to be called as such. Based on their phenotypic characteristics, strains of group XII to XVI (39% of total isolates) were identified as Pseudomonas syringae and those of group XI (3% of total isolates) as P. viridiflava. Further studies are needed to clarify the taxonomic status of the remaining strains which appeared distinct from Pss and P. viridiflava. The virulence of the strains was evaluated through inoculation of each strain onto sour orange, sweet orange (C. sinensis, Washington navel) and alemow (C. macrophylla) leaves. There were noticeable differences observed in the size of lesions produced by different strains, on the basis of which basis the isolates of either pathogen could be differentiated into weak, moderate, or highly virulent strains.

Association of satellite RNA with Cucumber Mosaic Virus (CMV) leads to either attenuation or enhancement of the viral symptoms. For detection of the satellite RNAs in Iranian isolates of CMV, plant samples showing viral symptoms were collected from different farms in the provinces of East- and West- Azerbayjan and Ardebil. These samples were inoculated onto the experimental host plants in greenhouse with the virus being detected using common serological methods. To study the possible effects of the satellite RNA on symptoms of CMV, biological purifications were performed using local lesion and propagation hosts. One pair of primers, specific to the coat protein coding region of the virus was employed to detect CMV. Based on the behavior of CMV isolates on the test plants, especially on N.tabacum cv. Samsun on which the satellite RNA causes chlorosis, N.rustica, and also on tomato, seven isolates were found and designated as a group of the virus variants that cause chlorosis on N.tabacum cv. Samsun. For detection of the satellite RNA, a PCR product of about 360bp was amplified using a pair of specific primers(CMV-Sat –F and Sat-R) from one of the isolates. Several inoculations on tobacco and on tomato plants showed the possible presence of the satellite RNA attenuated CMV symptoms on the host plant, the characteristic of non-necrogenic Satellite RNAs. This is the first report from Iran on the possible presence of satellite RNA in CMV isolates.

Sublethal effects of two juvenile hormone analogue, flufenoxuron and lufenuron, were evaluated on larval, pupal and adult stages of Habrobracon hebetor Say under laboratory conditions. Dose-response bioassays were carried out on the insect’s immature and adult stages through dipping and contact residue methods. In the sublethal effect tests, field rates (500 ppm) of the two insecticides were employed. In sublethal effect tests, mean longevity and fecundity showed no significant differences between flufenoxuron (27.8±1.4 d and 434.5±30.7 eggs) and lufenuron (27.3±2.2 d and 316.9±34.6 eggs) as compared with control (28.3±2.4 d and 390.7±52.9 eggs). However, net reproductive rate and intrinsic rate of increase indicated significant differences between flufenoxuron (282.5±19.9 and 0.286±0.006) and lufenuron (209.3±22.9 and 0.263±0.01) as compared with control (177.3±24 and 0.260±0.008). In summary it can be concluded that both flufenoxuron and lufenuron exerted little negative effects on H. hebetor. However, negative effect of Flufenoxuron on intrinsic rate of increase (rm) was recorded lower than that in lufenuron.

The mill moth, Ephestia kuehniella Zeller and Indian meal moth, Plodia interpunctella (Hubner) are two injuriously important pests of stored products worldwide. Harobracon hebetor (Say) has been used as a biological control agent of moth’s larva in peanut storehouses and as well in packaged cereal products. In the present study, the efficacy of H. hebetor in parasitizing larva of the mill moth and of the Indian meal moth was evaluated at different densities of parasitoid (1, 2 and 4 pairs) and host larvae (1, 2, 4, 8, 16 and 32) in a growth chamber set at 25±1 °C, 65±5 RH, and 14L:10D photoperiodism. When one pair of parasitoid released, the number of eggs laid was increased at the densities of 1, 2, 4 and 8 larvae of either host, respectively but, it decreased at the densities of 16 and 32 larvae of either host. Also, the number of emerged females and adults of wasp were increased at the densities of 1, 2, 4 and 8 larvae of both hosts, respectively but, did not significantly increased at the densities of 16 and 32 larvae of either host. When two pairs of parasitoid were released, the highest number of eggs laid was observed at the density of 16 larvae of the mill moth. Whereas, the number of eggs laid at the densities of 4, 8, 16 and 32 larvae of the Indian meal moth were significantly higher than those for the other larval densities. Also, the number of emerged females and adults of wasp increased at the densities of 1, 2, 4, 8 and 16 larvae of either host, respectively but, they did not significantly increase at the density of 32 larvae for either host. When four pairs of the parasitoid released, the number of eggs laid at the densities of 8 and 16 larvae of either host was found significantly higher than those for the other larval densities. Also, the number of emerged females and adults of wasp increased at the densities of 1, 2, 4, 8 and 16 larvae of either host, respectively but, did not significantly increase for the density of 32 larvae in either host. Based on the results obtained it can be concluded that at one pair of parasitoid treatment, a number of 8 larvae of either host, at the two and four pairs of parasitoid treatments, a number of either 16 or 32 larvae of either host form the most suitable combinations for a mass rearing of H. hebetor.

The effect of fenpyroximate on life table parameters of two-spotted spider mite Tetranychus urticae Koch was evaluated under laboratory conditions (25±20C, 60±5% RH and 16:8 L:D). Data were analyzed on the basis of age-stage, two-sex life table theory. The estimated value of LC50 for T. urticae was found to be 380 Ml/l. Durations of total preadult stage and preoviposition period in T. urticae treated with fenpyroximate (12.64 and 2.36 d, respectively) were significantly longer than those in control (11.56 and 1d, respectively). The lifetime fecundity of T. urticae treated with fenpyroximate was 1.35 (eggs/female) compared with control (52.27 eggs/female).The instric rate of increase (r), finite rate of increase (λ), net reproductive rate (R0), the mean generation time (T) and Gross Reproduction Rate (GRR) in T. urticae treated with fenpyroximate were -0.001 d-1, 0.998 d, 0.96 offspring, 14.25 d, 2.94 offspring, respectively. The study shows that the LC50 of fenpyroximate decreases the fecundity of two-spotted spider mite under laboratory conditions and makes the pest population growth almost face extinction. So the acaricide can be effectively used in two-spotted spider mite control.

To evaluate the effect of different concentrations of copper, manganese and nickel on the growth and sporulation of Trichoderma species of; T. harzianum, T.longibrachiatum and T. tomentosum, an experiment was carried out in vitro at 25oC±2 on soil extract agar medium of five replicates. The results revealed that among these species T. longibrachiatum had the highest growth rate (47.78mm) after 72 hours past. During this time nickel not only didn’t exhibit any inhibitory effect on T. longibrachiatum, but also increased the growth of the species (7%). Throughout the experiment, the inhibitory concentration of 50% (IC50) values for the copper and nickel and for the case of T. tomentosum was estimated at 32.22ppm after 96 hours past. The results showed that the sporulation of Trichoderma species on soil extract agar medium increased in the presence of heavy metals. Also in this study it was found that the absorption rate of heavy metals by Trichodema species depended not only on the fungal species but also on the metal type and its concentration, the highest absorption rate (48.67%) being observed at 1000ppm concentration of nickel on T. tomentosum.

Although host plant resistance is the healthiest, most reliable and cheapest way of pest control, to achieve a successful integrated pest management, pest-resistant plant varieties should be compatibity with the biological control agents. Throughout the present study, the performance of the predatory mite, Phytoseiulus persimilis، on four common bean cultivars (Naz, Khomein, Akhtar and G11867) and also, the emission extent of Herbivore Induced Plant Volatiles (HIPVs), (responsible for the attraction of predatory mite to plant), following the plants infestalion with two-spotted spider mite, Tetranychus urticae, were investigated. To this end the seeds of each genotype were planted in pots and grown under greenhouse conditions the plants when at two fully expanded leaf stage, were each infested with 50 T. urticae adult females on each of the plant's primary leaves. After 3 days past, the plants were tested for predatory preference, carryied out through Olfactometery. The test was performed for all possible pairs on the four cultivars. The level of herbivore induced plant volatile compounds was assessed in the spider mite infested plants of common bean cultivars through GC/MS. The results indicated that Khomein cultivar stood at the highest volatile compound emitter and could significantly attract more predatory mites than the cultivars Akhtar and G11867. Also, although there was not any significant difference observed between the levels of volatile compounds emission in of Naz and Akhtar varieties, Naz could attract significantly more predatory mites than Akhtar did. It is understood that both the quantity, and quality of the volatile compounds are responsible for the attraction of the predatory mite to the spider mite infested bean cultivars.

Mixed infection of plants with more than one virus results in different types of interactions. Interaction between two recently identified lettuce infecting viruses in Iran, Lettuce X potexvirus (LeVX) and Rubus Chlorotic Mottle sobemovirus (RuCMV), was studied on co-infected Chenopodium murale and Chenopodium quinoa plants. Plants were mechanically inoculated with either LeVX, RuCMV or both. Co-infected plants of C. quinoa developed much more severe symptoms than the singly infected ones, with enhanced RuCMV accumulation level, as determined by quantitative ELISA and Western blot assay, indicating that the interaction of RuCMV with LeVX is synergistic in C. quinoa. Co-infection of C. murale plants resulted in symptom severity while with no detectable increase in RuCMV accumulation level, indicating that RuCMV interaction with LeVX is not synergistic in this plant. Conversely, LeVX accumulation level in either of C. quinoa or C. murale dually infected plants was significantly reduced, suggesting that LeVX interacts with RuCMV antagonistically in co-infected plants of both Chenopodium species.

Elm leaf beetle, Xanthogaleruca luteola, is a serious pest (particularly at its larval stage) of elm trees in many areas. The existence of three important enzyms namely: α-amylase, α- glucosidase and β-glucosidase in the midgut of the insect at its larval stage was investigated. Optimum activity for α-, β-glucosidase and α-amylase was recorded at pHs 5.0, 6.0 and 6.0 against p-nitrophenyl-α-D-glucopyranoside, p-nitrophenyl-β-D-glucopyranoside and starch respectively. α and β-glucosidase exhibited more stability at pHs 4.0-6.0 than at either highly acidic or and alkaline pHs. These enzymes were respectively more stable at pH 5.0 and 6.0 with increased incubation time. α -amylase was stable at pHs 4.0-8.0. α and β-glucosidase and also α-amylase, when incubated at different temperatures were found to be maximally active at 45, 60 and 40°C, respectively. Alpha- glucosidase activity significantly decreased in the presence of Ca2+ and Mg2+. On the contrary, the ions K+ and Na+ did not significantly affect the enzyme’s activity. The ions K+, Mg2+ and Na+ did not significantly affect the enzyme activity. Zymogram analysis revealed the presence of one band each of amylase, α- and β-glucosidase activity in the insect’s larval midgut extract.

To identify the faunal parasitic nematodes of Mazandaran, Guilan and Ardebil provinces, roughly 30 soil samples were collected from the rhizosphere of forest trees during 2005-2009. Nematodes were extracted from soil through centrifugal flotation technique. The extracted nematodes were transferred to pure glycerin using common methods. Permanent slides were made and nematodes identified as based on their morphological and morphometric characteristic. Throughout the study, three species of genus Nagelus belonging to family Belonolaimidae were identified viz, N. hehxagrammus, N. neohexagrammus and N. obscuruss from which the second species is introduced as a new record within the nematode fauna of Iran. It could be separated from most closely related species namely N. grandis, N. affinis and N. hexagrammus by its possession of well developed cephalic framework, six annules on head, areolated lateral fields, length of stylet and number of annules on tail.

This experiment was carried out on lily cut flowers in greenhouse conditions and at the Agricultural Faculty of Guilan University. The statistical factorial experiment was performed in a completely randomized design of three replications. The main factors studied were cultivars Navona (Asiatic) and Fangio (LA) along with four levels of calcium sulfate (0, 2.5, 5 and 10 mM) in combination with three levels of calcium nitrate (0, 1.5 and 2 mM). The results indicated that the combined effects, conjugated of calcium nitrate and calcium sulfate led to plants resistance against grey mold, and also with an improvement in the post harvest life. The resulting means indicated that the indexes were different at a 5 percent level of significance. The highest petal resistance to Grey Mold was observed at 5 and 1.5 mM of calcium sulfate and calcium nitrate levels respectively. Also the lowest ethylene and highest phenolic compounds production were observed in the highest resistant flowers. The lowest grey mold resistance was observed in the case of 0 mM calcium nitrate and calcium sulfate (control), while ethylene production being the highest. The highest vase life was observed in 1.5 mM calcium nitrate level of treatment for both Navona and Fangio cultivars. All in all, the results indicated that, the best treatment for Botrytis resistance in Navona and Fangio varieties is 1.5 and 5 mM of calcium nitrate and calcium sulfate respectively. The results finally indicated that one of the methods of decreasing grey Mold injury and increasing vase life of lily cut flower is the proportionate use of calcium nitrate and calcium sulfate.

Rhizosphere soil samples were collected from olive orchards of Golestan, Guilan, Zanjan and Qom provinces. Fluorescent pseudomonad strains were isolated from the collected samples on King’s B medium and their phenotypic features characterized as based on the standard bacteriological methods. For all the tested strains oxidase reaction, fluorescent pigment, growth at 4˚C and arginine dihydrolase were positive while tobacco hypersensitivity reaction, levan production on 5% sucrose and pectolytic activity on potato slices were proved negative. Most of the tested strains reduced nitrate while liquidizing gelatin. Based upon the phenotypic features of the tested strains most were identified as P. fluorescns while a few identified as P. Putida. Nematicidal activity of the cultural filtrate of the Psudomonas strains was tested on fresh hatched juvenile of Meloidogyne javanica under laboratory conditions. They caused nematode mortality ranging from 13.33 to 100% while showing significant differences. Efficacy of the representative strains along with fenamiphos on nematode egg productions, juvenile and mature nematode activity on olive root were studied under green-house conditions. Significant differences were observed among the applied treatments. The highest and lowest nematode juvenile and mature formation on olive seedling roots were observed for control plants and plants treated with fenamiphos respectively. The most effective strains on the different nematode life stages were P. fluorescens strains CHAO as well as strain 208. The highest and lowest nematode egg production on olive seedling roots were observed for control plants and plants treated with strains CHAO vs. strain 99 respectively. Some strains proved to be more effective than fenamiphos in nematode egg production on olive seedlings roots. This is the first report of the biocontrol of olive root knot nematode using flourescent pseudomonads.

Seven antagonistic bacteria belonging to Pseudomonas fluorescens species were evaluated on the basis of inhibitory zone of Sclerotinia sclerotiorum fungal growth Finally, the most suitable strain (P4) which exerted the greatest effect on reducing the causal agent of Sclerotinia root rot was selected for further germinator trails. Nine formulations were prepared on the basis of talc powder from the strain P. fluorescens P4. Three culture mediums: M1, NBY and KB; along with three stickers: gum arabic, xunthan gum and carboxy methyl cellulose were used in these formulations. A survey of the effect of the formulations as regards the control of Sunflower Sclerotinia root rot was done on Progress variety of Sunflower and in germinator. Studies after six weeks past, indicated that, treated pots with free cell bacteria, with 100 percent healthy plants, treated pots with Bordofix fungicide containing 90 percent healthy plants, treated pots with NBY-CMC, KB-CMC, NBY-XG, NBY-AG and M1-CMC formulations with 87.5 percent healthy plants as well as treated pots with M1-AG formulation with 75 percent healthy plants, without any significance of difference as compared with healthy control plants, led to the most diminishing percentage being even more effective than Bordofix fungicide bearing a 62.5 percent of healthy plants. Furthermore, among powder formulations, the survival and maintenance of the population of bacteria in M1 culture medium with gum arabic sticker, at 4o C was more pronounced than those in the other formulations, during 150 days. Thus, the type of culture medium proved to have a significant impact on survival of the bacteria population.

Cotton whitefly, Bemisia tabaci Gennadius (Homoptera: Aleyrodidae) is one of the most seriously damaging insect pests of agricultural products worldwide. It is highly polyphagous, damaging a broad range of food as well as non-food crops through direct feeding, excretion of honeydew and transmission of over 100 plant viruses. In Iran, growers usually use insecticides as the main control agent of Bemisia tabaci which may lead to insecticide resistance in the pest. An investigation of the susceptibility of three populations of this pest, collected from Tabriz, Gorgan and Qom regions, to imidacloprid and amitraz was carried out. Bioassays were conducted using an adult leaf-dip method. The results revealed that Tabriz strain was more susceptible to imidacloprid than the other two strains. Resistance ratios for Gorgan and Qom strains relative to Tabriz strain were 2.6 and 2.68, respectively. Bioassay results with amitraz revealed that Tabriz strain was 1.44 and 1.47 fold less susceptible than Gorgan and Qom strains, respectively. Biochemical assays were conducted through on estimation of esterase activity using α- naphthyl acetate and β-naphthyl acetate substrates. The results of with β-naphthyl acetate substrate showed that esterase activity in Gorgan and Qom strains were respectively 1.37 and 0.85 fold of Tabriz strain. Esterase activity was also investigated using page method. Different esterase banding patterns in Gorgan and Qom strains were detected for comparison with Tabriz strain.

Pathogenicity of wheat stem rust was investigated during two cropping seasons of the years 2010 and 2011. towards this end, 47 isogenic lines of wheat stem rust along with 43 cultivars/lines of the crop were tested in field conditions (natural infection) and also in the greenhouse conditions. The results showed, similarity of pathogenicity for plants carring gene/s Sr7B, Sr8A, Sr8B, Sr9B, Sr9D, S9G, Sr11, Sr13, Sr16, Sr17, Sr25, Sr34 and SrDP2 as for two isolates. As for the in genetic variation of two wheat stem rust isolates (in greenhouse conditions), Kelardasht isolate did not show virulence for 15 isogenic lines carrying gene/s Sr5, Sr8A, Sr22, Sr24, Sr26+Sr9G, Sr27, Sr28, Sr29, Sr31, Sr32, Sr33, Sr35, Sr36, SrGT and SrWLD. From Dashte-Azadegan isolate of Khuzestan, virulence for gene Sr31 as a variant of Ug99 race was detected, while this isolate being avirulent on plants carrying gene/s Sr5, Sr22, Sr24, Sr26+Sr9G, Sr27 and SrGT. The results of evaluation of resistance for commercial wheat cultivars and elite lines indicated that, such new released cultivars as Morvareid, Sivand and Parsi were resistant with a range of reaction of 5R-50M to stem rust diseases as observed during the two years of the study.

Tomato ringspot virus (ToRSV) is a member of the genus Nepovirus, family Secoviridae, causing substantial economically declining in perennial crops, including stone fruits, worldwide. During the years 2008 and 2009 a total number of 414 symptomatic and asymptomatic leaf samples were randomly collected from different stone fruit trees including peach, apricot, plum and sour cherry from Fars and Golestan provinces. Applying DAS-ELISA method samples were tested for the presence of ToRSV, through specific polyclonal antibody. Serological tests revealed that, during the years 2008 and 2009, different stone fruits were infected with ToRSV, with the infection rates of 8.5% and 10.2% in gardens of Fars and in Golestan with respective infection rates of 30% and 23.1%. One hundred of different stone fruit leaf samples that had weakly reacted with ToRSV antibody in DAS-ELISA test, were tested again through Dot- immunobinding assay (DIBA), ToRSV infection being confirmed in 85 samples. For an evaluation the biological properties of the detected isolates, they were subjected to mechanical inoculation on herbaceous plants and the morphology of isometric viral particles observed through immunosorbent electron microscopy for apricot and peach samples. According to the host range study the Iranian isolates did not differe in biological properties from other isolates. Such Symptoms as necrotic ring spots and leaf mosaics were frequently related to the viral infection. Applying with RT-PCR, a DNA fragment of 499 bp, belonging to the coat protein coding sequence of the ToRSV genome, was amplified for 20 out of 30 peach, apricot, plum and sour cherry samples the samples were serologically tested as positive to ToRSV infection. Results proved that different stone fruits in some gardens in South and Northern parts of Iran are infected with ToRSV.

Septoria tritici blotch (STB), caused by Mycosphaerella graminicola (anamorph: Septoria tritici), is one of the most important diseases of wheat worldwide that has been intensified recently in some regions of Iran including Moghan Plain. In this study reaction of 23 bread and durum wheat cultivars to 10 isolates of the pathogen collected from Moghan region during 2007 and 2008, were evaluated under greenhouse condition. Analysis of variance (ANOVA) of pycnidial coverage of leaf areas revealed significant differences (P<0.01) among fungal isolates as well as wheat genotypes. Moreover, genotype× isolate interaction term in the ANOVA analysis was highly significant (P<0.01), indicating isolate-specific interaction between wheat cultivars and fungal isolates. Although about half of wheat cultivars were susceptible to all isolates, 40 isolate-specific resistances were identified that most of them are new records. Among the new sources of resistance, Dehdasht, Seimareh, Galanco, Saisson and Hirmand with specific resistance to nine, eight, six, five and three isolates, respectively, were the most resistant cultivars. Furthermore, the isolates exhibited different virulence patterns on wheat cultivars. Although results of this study indicated high virulence diversity of M. graminicola isolates from Moghan Plain and susceptibility of the most wheat genotypes cultivated in this region to STB, the new sources of resistance could be utilized in the wheat-breeding programs.