فصلنامه گیاه پزشکی
سال چهل و سوم شماره 3 (پاییز 1399)

 Bemisia tabaci (Genn.)(Hem: Aleyrodidae)is considered as one significant pest in the agricultural products in Iran. In this research, the effects of ZnO NPs, kaolin, B. bassiana (EUTP105 Isolate) were assessed alone or in combination to Neemarin® compared to thiacloprid insecticide under field conditions on eggs and nymphs of B. tabaci and pupae of its parasitoid Eretmocerus mundus.

This research was handled in the research field of the scientific staff of the Cotton Research Center of East Iran, Kashmar, in a randomized complete block design with three replications in August 2019. The experimental plots were separately 6 m2 and 1.5 m. The recommended concentration of Neemarin® (15 mg L-1 AZA), thiacloprid (0.3 mL L-1), Kaolin (30 g L-1), a concentration of 20 mg L-1 of ZnO NPs, a concentration of 1×1010 conidia mL-1 of B. bassiana, half- concentration of Neemarin® (7.5 mg L-1 AZA) + 10 mg L-1 of ZnO NPs, half- concentration of Neemarin® (7.5 mg L-1 AZA) + 15 g L-1 of Kaolin and half- concentration of Neemarin® (7.5 mg L-1 AZA) + a concentration of 1×105 conidia mL-1 of B. bassiana were prepared which sprayed on upper and lower surfaces of cotton plants using a knapsack sprayer, equivalent to an application volume of 500 l ha-1. The control treatment was sprayed by water. The mortality of B. tabaci eggs and nymphs and pupal stage of E. mundus were calculated using Henderson and Tilton equation . To this purpose, five plants were randomly selected in each plot and three leaves from each one (top, middle, and bottom section of plant canopy) were excised and transferred to a laboratory. Two pieces of 1 cm2 from each leaf were chosen so that the units included into main vein. Total live eggs and nymphs of pest as well pupae of parasitoid in each sample unit were counted using a binocular. Samplings were handled one day before and 3, 7 and 14 days after spraying.

In 3 DAT (days after treatment), the most effective treatments on eggs and nymphs of pest were Kaolin + Neemarin® and thiacloprid, respectively. While, the least effective treatments were B. bassiana. In 7 DAT, Kaolin + Neemarin® and ZnO NPs exerted the highest and lowest mortality, respectively, on the eggs of pest, Neemarin® and B. bassiana exerted the highest and lowest mortality, respectively to the nymph. In 14 DAT, the highest efficacy on eggs and nymphs was observed in Kaolin + Neemarin® and Neemarin®, respectively, while the lowest efficacy both on eggs and nymphs belonged to B. bassiana. The highest and lowest mortality of the pupae of parasitoid in 3 DAT belonged to thiacloprid and B. bassiana, respectively. In 7 DAT, Neemarin® and ZnO exerted the highest and lowest mortality on the parasitoid pupae, respectively.  In 14 DAT, Kaolin exerted the highest mortality on the pupae of parasitoid that was followed by Kaolin + Neemarin®, Neemarin®, and thiacloprid. The maximum yield and benefit/cost ratio were observed in Kaolin + Neemarin®, followed by Kaolin treatments. 

Considering the results, ZnO NPs and Kaolin alone, or in combination to Neemarin® can be the suitable candidates as an alternative in the IPM programs of B. tabaci on the cotton field.

Date palm Phoenix dactylifera L. is one of the income sources of cultural and economic importance in several countries. Leaf wilt disease is one of the important diseases observed in palm gardens in Khuzestan province. Disease symptoms are observed as asymmetrical leaves drying. Fusarium proliferatum is a common pathogen infecting numerous crops, including banana, carnation, cycads, date palm, garlic, mango, maize, onion, rice, and sesame plants.

To identify the causal agent(s), infected samples were collected from Abadan, Khorramshahr, and Shadegan cities and transferred to the laboratory. The samples were then disinfected and cultured on potato dextrose agar (PDA) plates. Fungi were purified by the single-spore technique. For molecular identification, the colonies were inoculated in potato-dextrose broth (PDB) and the grown mycelium mass was used for DNA extraction. Polymerase chain reaction was done with reaction initiators of Tef171F and Tef1997R to replicate a part of the tef1 region.

BLAST search showed that the sequence of the studied isolate was 99.55% similar to both the confirmed isolate and the reference of F. proliferatum strain TOR-55. Phylogenetic analysis of the isolate with sequences related to type strains, using the maximum likelihood, confirmed that they belonged to this species. Pathogenicity test was followed by re-isolation from the inoculated plants.

These results indicated that all four concentrations were able to infect trees and cause disease symptoms and pathogenicity. However, disease severity was higher at 5×105 and 5×106 spores per ml than the other concentrations of spores used here. The second two concentrations of spores were more effective in drying of leaflets and leaves.

Potato bacterial soft rot is considered one of the most common bacterial diseases of Potato.Soft rot of potatoes was caused by a range of bacteria worldwide, such as Pectobacterium carotovorum subspecies carotovorum, Pectobacterium atrosepticum, and Dickeya species.The bacteria mainly attack the fleshy storage organs of their hosts (tubers, corms, bulbs, and rhizomes), they also affect succulent buds, stems, and petiole tissues. Controlling the disease is not always so effective, sanitary practices in production, storing, and processing which can be done to slow the spread of disease and protect yields. Little information is available on the effect of temperature on pathogen interaction with plants and the induction of PR genes. In the present study, an attempt was made to establish a relationship between the accumulation of PR proteins and the development of resistance obtained by infecting Potato tuberculosis cells with bacterial pathogens at different temperatures to  effectively control potato bacterial soft rot disease.

Pectobacterium carotovorum, strains A2 and A14, Pectobacterium atrosepticum, strains A21 and A36, and Dickeya-dadantii ENA49 were used. Potato cultivars Scarab, Vetraz, and Odyssay, were used for the bacterial infection. The experiment was factorial with three replications based on a completely randomized design. The macerated tissue weight was measured in mg per disc. In other experiment, the relative value of mRNA copies of studying genes was determined in potato tuber cells of two semi resistant and susceptible cultivars which infected with bacterial strains of Pectobacterium carotovorum 2A, Pectobacterium atrosepticum 36A, and Dickeya dadantii ENA49 which were incubated under temperatures of 18, 28 and 33 ° C.

Odyssay cultivar of Potato as a susceptible cultivar and Scarb cultivar as a semi-resistant cultivar to Potato pathogens under different temperatures (18°С, 28°С and 33°С) were identified. When potatoes are infected with pectolytic bacteria, the genes PR-3, PR-5t and   PR-10 were expressed. In tubers and leaves of resistant potato cultivars, the level of PR-5t gene expression is significantly higher than that of susceptible cultivars, when infected with bacteria and without infection. There is a correlation among expression levels of PR-5t gene in the tissues of potato tubers with their resistance to bacterial soft rot.

Several pathogens usually attack plants. Different defence pathways in plants have evolved in reaction to the pathogens. These defence mechanisms can be stimulated and activated by some microorganisms or chemicals. Temperature can be considered a virulence factor. It has a significant effect on the pathogenicity of pectolytic bacteria Pc, Pa, and Dd. Plant resistance to pathogens is directly related to pathogenesis expression related genes and defence response genes. When potatoes are infected with pectolytic bacteria, the induction of pathogenesis related genes and Potato defence response genes occurs which is in consistent to other researchers' findings on plant resistance mechanisms to pathogens through the induction of defence response genes. The experimental results showed that when potatoes are infected with pectolytic bacteria, the induction of several PR genes occurs, especially PR-3, PR-5t, and PR-10. The induction of the PR-10 gene is stimulated by the infection of Pc and Pa at low temperature (18 ° C).

Chymotrypsin-like enzymes are involved in many physiological processes of insects including digestion, development, survival, and immunity. The enzyme cleaves the peptide bonds on C-terminal of the aromatic amino acids in proteins and release active peptides as well as amino acids required for growth, reproduction, and development of the insect. Due to the importance of food proteins on insect survival and development, more fundamental entomologists are researching the structures, functions, mechanisms, and interactions of the involved digestive enzymes. Discovering biochemical and structural properties of insect proteases can help develop rational control strategy based on specific protease inhibitors. Accordingly, the present study focuses on protein structural analysis, protein sequence alignment, phylogenetic analysis as well as conserved motif assessment from various insect species using different bioinformatics tools. The multiple sequence alignment revealed different conserved stretches of amino acids along with highly conserved catalytic sites (His, Asp, Ser) and ten conserved motifs were also discovered by MEME and MAST tools. The phylogenetic data suggest that the insect chymotrypsins might share a common ancestor. The three-dimensional structures of chymotrypsins were generated by I-TASSER, and the 3D model was further verified using PROCHECK, ERRAT, and Verify-3D. The protein – protein interactions network constructed by STRING 11 provided ten enriched pathways in Aedes aegypti (L.) chymotrypsin. Totally, the present work can provide new insights for designing alternative pesticides based on specific digestive enzyme inhibitors.

Chickpea is an important grain legume in Asia, and makes a significant contribution to the food and nutrition security of the people. Fusarium wilt is an important disease of chickpea, causing significant yield loss. This malady is the most important and destructive soil-borne disease of chickpea in Iran. Excessive use of chemical fungicides to control this soil-born pathogen can have adverse effects on human health and environment, and can lead to fungal resistance to fungicides. Increasing induced resistance using resistance inducers as bio-fertilizers can be considered as an alternative method to plant disease control. The use of nitroxine a bio-fertilizer containing Azosprillium and Azotobacter species as well as mycorrhizal fungi to induce systemic resistance mechanisms has been demonstrated in different plants under biotic stress. The aim of this study was to investigate the effect of bio-fertilizers to induce acquired resistance in chickpea plant under pathogen stress as compared to non-infected plants.

The change of some antioxidant enzymes and gene expression analysis was examined to evaluate the effect of biofertilizers on increasing resistance in chickpea plants infected with Fusarium oxysporum fsp. ciceri. A factorial experiment based on complete randomized block design with three replications was conducted under greenhouse conditions in 2014. The eight treatments included: 1) Plant control without infection (sh), 2) Fusarium oxysporum (Foc), 3) Mycorrhiza (Gi), 4) Nitroxin (N), 5) Mycorrhiza+Nitroxin+Infection (GI+N+F), 6) Mycorrhiza+Nitroxin (GI+N), 6) Mycorrhiza + Fusarium oxysporum (Gi+F) and 8) Nitroxin+Fusarium (N+F). Mycorrhiza, Fusarium oxysporum and Nitroxin+Fusarium (M+F+N) were used in this study. Catalase, Proxidase, and Polyphenoloxidase activity was assessed by spectrophotometer with corresponding wavelengths. Total RNA was isolated using an extraction kit, according to the manufacturer’s protocol. Real-time RT-PCR was performed in a thermocycler using the following program: 5 min at 94 °C, followed by 35 cycles of 30sec at 95°C, 30 min at 59°C and 30 min at 72 °C, with final extension for 10min at 72°C. All acquired data were analyzed using SAS software version 9 and mean values were compared using Duncan's multiple range test. The changes of transcript level were measured via a comparative technique. Actin genes were used as internal reference.

Data analysis revealed that the highest rate of total phenol and protein (50/292 mg/mL) in treated plants was related to nitroxin and mycorrhiza treatments, respectively. The greatest change in all three enzymes was observed in mycorrhiza treatment 72h after plant infection. Mean comparison of gene expression data at different time intervals showed a high expression level 72h after plant infection in mycorrhizal application. Our results indicated that the maximum effect of bio-fertilizer application occurred 72h following plant infection with the pathogen. Application of both biofertilizers without Fusarium infection did not show any significant change in the expression level of the two tested gene.

Biofertilizer agents can improve plant growth through several different mechanisms such as protecting the plant under stressful conditions and defense against plant pathogen which can lead to reduced disease and death. The increase of total phenol and protein as the precursor of plant defense mechanism through apply biofertilizers in infected plants indicated the efficiency of these compounds to reduce disease. Antioxidant enzymes as a biomarker were used to evaluate the effect of exogenic compounds on increasing plant resistance against biotic stress. As with a similar work, we found a correlation between biofertilizer application and enzyme activity. Other parameters such as phenotypic characters and yield components can be used as a marker to investigate the effect of biofertilizers in these domains. Analysis of variance of the data related to gene expression confirmed the efficiency of biofertilizers on inducing resistance. Based on our results, we recommended use of biofertilizers in a single or mixture form to improve growth conditions in stressed plants especially under biotic stress. To make the bio-fertilizer more efficient in the field, a coordinated work by different science domains as bacteriology, chemistry, genetics, and agronomy as well as farmers as bio-fertilizer consumers seems necessary. This coordination could facilitate the adaptation of fungi and bacterium-based bio-fertilizers to different agriculture systems.

Phenacoccus solenopsis Tinsley is a global pest which recently causing serious damage to various crops and ornamental plants such as hibiscus shrubs in southwestern Iran. The field biology of this pest was investigated in Dezful during 2015-2016 season. Results indicated that in Dezful, P. solenopsis passes 11 generations per year. The first generation begins from early March to early April, and the 11th generation ends in the mid-March. The mealybug activities did not stop during the winter. The 8th generation was the shortest one which lasted 28.71±1.39 days from the late September to the mid-October. The longest generation time was also the 2nd one (47.47±0.79 days), from early April to early May. The reproduction parthenogenetically occurred, and females produced eggs and crawlers. The highest fecundity was163.11±3.11 offspring per female with the highest ovipositional period (4.41±0.14days) in the 4th generation in summer. The lowest fecundity (1.67±0.12offspring) was in the 11th generation in winter. The female longevity was significantly different among various generations, from 13.79±2.03 days in 11thgeneration to 26.12±0.70 days in 2nd generation. Males lived in a short time (1 to 2 days). The present study confirms that P. solenopsis has a rapid development, high fecundity with a pattern of crawlers’ production, and the ability to adapt to the unconditional warm and dry climate of Dezful. Unfortunately, due to lack of management, and a wide range of host plants, it could be a serious threat for farmlands, greenhouses, and urban areas in Khuzestan province.