فصلنامه گیاه پزشکی
سال چهل و پنجم شماره 4 (زمستان 1401)

The Mediterranean flour moth, Ephestia kuehniella Zeller (Lepidoptera: Pyralidae), is one of the world’s most significant pests of stored products. E. kuehniella eggs and larvae are widely used in rearing parasitoids and predators for biological control programs. The purpose of this study was to look at the effects of various cereal hosts and cultivars, including wheat (Meraj, Paya and Aftab cultivars), barley (Oxin, Faraz and Fardan cultivars), rice (Hashemi cultivar), maize (704 Hybrid), sorghum (Spidfit cultivar), rye (Danko cultivar) and millet (Morvaridi cultivar) on life history variables, biological characteristics and population growth parameters of E. kuehniella. In addition, biochemical (protein and starch contents) and physical (humidity and hardiness index) characteristics of the cereal hosts were evaluated to understand any possible correlation between important demographic parameters and biochemical and biological features of the cereal hosts.

The E. kuehniella larvae were reared in a growth chamber at 26 ± 2°C, 60 ± 5% RH, and a photoperiod of 14:10 (L:D) hours on each cereal host. The duration of each stage, developmental period, and survival rate were observed and recorded daily. TWOSEX-MS Chart was used to analyze the life table parameters of E. kuehniella. The biochemical metabolites of cereals were measured using Bradford’s (1976) and Bernfeld’s (1955) methods.

Our findings showed that various kinds of cereal significantly impact the life history and demographical parameters of E. kuehniella. The millet host had the longest development time (66.57 days), while the rice and maize hosts had the shortest on Paya and Aftab cultivars. Total fecundity (mean number of eggs laid during the reproductive period) of E. kuehniella varied significantly depending on the hosts tested. The lowest fecundity values were observed on the millet host (49.10 ± 2.92 eggs), while the highest was on the maize host (136.18 ± 7.36 eggs). The maize host had the highest intrinsic rate of increase (0.067 day-1), and the millet host had the lowest (0.033 day-1) host. Significant differences in physical characteristics and biochemical metabolites were observed between the various hosts in this study. Furthermore, significant positive or negative associations were discovered between different hosts’ life history variables and humidity percentage, hardiness index, protein, and starch contents.

According to the findings, sorghum (Spidfit cultivar) and millet (Morvaridi cultivar) were the least suitable (relatively resistant) hosts for E. kuehniella growth. In contrast, wheat (Meraj cultivar) a

Brevicoryne brassicae is one of the most serious cabbage pests in the world. This pest feeds on the leaves, mostly on the host plant's middle leaves. Despite having many natural enemies, aphids are not entirely controlled by natural enemies due to their rapid reproduction. Applying chemical insecticides is one of the essential methods used to control aphids; using one pesticide to control several pests can reduce the costs and frequency of spraying. In this study, the lethal and sub-lethal effects (LC25) of matrine insecticide with a contact and stomach function on the cabbage aphid, were evaluated under laboratory conditions.

The dipping method was used in bioassay and life table studies against adults of aphid insects. The LC50 value of matrine insecticide for adults of cabbage aphid was 83.33 μl/ml after 24 hours, and LC25 concentration (30.11 ml/ml) was used to estimate the sub-lethal effects of matrine on the biological parameters of the cabbage aphid. Experiments were performed on the acephala variety of cabbage. One common method of investigating insecticides' non-lethal effects is demographic toxicology, which uses life table and growth population parameters to evaluate toxicity.

According to the results, the female lifespan/longevity and fertility affected by sub-lethal concentration were significantly reduced compared to the control (distilled water and citowett). The mean oviposition period of adult aphids decreased from 8.92 days in control to 3.79 days in LC25 concentration of the insecticide. The present study's data also showed that the sublethal concentration also affected the offspring of treated cabbage aphids. Accordingly, the sublethal concentration of matrine reduced the net reproductive rate (R0) from 23.96 nymphs per female in each generation in the control treatment to 3.26 nymphs in LC25. The intrinsic rate of increase (r) in sublethal concentration was recorded as 0.36 (day-1) and in control as 0.1 (day-1). Other parameters, such as finite rate of increase (l) and gross reproductive rate (GRR), were also significantly lower than the control. The mean generation time (T) of treatment affected by LC25 concentration of matrine increased compared to the control treatment.

Results of the current research in laboratory conditions showed that this insecticide has significant lethal and sublethal effects on the cabbage aphid. Therefore, matrine, as an herbal insecticide, can be considered in the management programs of cabbage aphids.

Citrus canker disease caused by Xanthomonas citri subsp. citri and bacterial leaf spots caused by X. gardneri and X. perforans are critical bacterial diseases. The control of these pathogens is a serious challenge due to the problems caused by the restrictions on the use of poisons and antibiotics, and the issue of toxic resistance. Therefore, plant essential oils can be a suitable alternative to control plant pathogens. In the present study, the effect of Lippia citriodora, Mentha piperita, M. aquatica, M. saulavens, M. spicata and M. pulegium essential oils against X. citri subsp. citri, X. gardneri and X. perforans have been evaluated.

After plant essential oils extraction by clevenger, their constituent compounds were identified by GC-MS. Then, the antibacterial effect of essential oils was surveyed using the disk diffusion method. Also, the Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC), the combined effect of plant essential oils, and the effect of M. pulegium essential oil against X. gardneri were investigated using Transmission electron microscopy.

The results of GC-MS essential oils analysis showed that a total of 28, 28, 30, 36, 39, and 42 compounds were identified in M. saulavens, M. piperita, M. pulegium, M. aquatica, M. spicata, and L. citriodora essential oil compounds, respectively. The highest and lowest amount of inhibitory zone was related to oregano species on the bacteria X. citri subsp. citri with an amount of 24 ± 2.08 mm and apple mint essential oil on X. gardneri with an amount of 5.66 ± 0.66 mm, respectively. MIC and MBC for the examined essential oils against the pathogenic bacteria were between 1-5 and 2-6 μg/ml, respectively. Also, the synergistic effect was observed between different essential oils such as M. piperita- L. citriodora, M. pulegium- M. spicata, M. pulegium- M. aquatic, M. aquatic- M. piperita, M. aquatic- M. pulegium, M. pulegium- M. saulavens on pathogenic bacteria. Transmission electron microscope photos related to the effect of M. pulegium essential oil against X. gardneri bacteria showed complete cell destruction, cell wall damage, swollen nuclear, area and changes in the cytoplasm density of bacterial cells.

Essential oils are regarded safe substances when considering the control limitations of bacterial infections, bacterial resistance to toxins, and environmental issues associated with the use of toxins. Also, due to the different compositions of essential oils and the observation of synergistic properties between them in this study, the risk of developing resistance to them decreases, and their antibacterial effect increases.

Loose smut caused by Ustilago nuda is one of the barley’s critical seed-borne diseases, causing crop yield to decrease. Using a healthy seed disinfected with chemical fungicides is the simplest and most cost-effective way to prevent the damage caused by loose smut disease of barley. The purpose of this study was to investigate the health status of barley fields concerning Loose smut disease and assess the effect of seed infection. It also sought to examine the effectiveness of seed disinfection with Iprodione-carbendazim (Rovral-TS®) and Carboxin thiram (Vitavax-thiram®) fungicides on traits related to germination, disease incidence, and antioxidant defense system responses.

The fields of Isfahan, Markazi, Zanjan, Tehran, Alborz, West Azerbaijan, Hamedan, Razavi Khorasan, Qazvin, Qom, Semnan, Fars, South Khorasan, North Khorasan, Sistan and Baluchestan, Kohgiluyeh and Boyer-Ahmad, Khuzestan, and Ilam provinces of Iran were sampled according to the International Rules for Seed Testing to investigate the health status of barley seeds. Detecting the seed infected with the agent of the Loose smut disease was carried out using the embryo test. Then, the activity level of antioxidant enzymes (superoxide dismutase, catalase, and polyphenol oxidase enzymes), the oxidative damage index malondialdehyde, and the content level of phenolic compounds and total protein were investigated.

In the range of 1.1% to 0.05%, approximately 13% of the barley seed samples collected from various fields were infected with loose smut disease. Infected barley seeds can significantly reduce vigor indices without affecting germination percentage. The results showed that seed disinfection with chemical fungicides improved the dry and fresh weight, average root length, seedling weight vigor index, coefficient of germination velocity, and mean times germination. Chemical fungicides’ effectiveness in reducing disease incidence varied depending on the type of fungicide and the infection level of the seed. Compared to Carboxin thiram fungicide, seed disinfection with Iprodione-carbendazim fungicide significantly improved some quality characteristics in seed germination and reduced disease incidence. The findings of the biochemical mechanisms investigating revealed that the activity level of antioxidant enzymes (superoxide dismutase, catalase, and polyphenol oxidase enzymes) and the content of phenolic compounds in spikes obtained from infected and non-infected seeds were higher than the control. In addition, seed disinfection with chemical fungicides increased total protein content while decreasing the membrane peroxidation index compared to the control.

The current study found that chemical fungicides seed disinfection increased the levels of defensive enzymes and the content of phenolic and protein compounds, which may help to limit infection and improve the level of defense-related antioxidant enzymes in barley. These findings shed new light on the effect of seed infection levels and seed disinfestations on the stimulation of the antioxidant defense system in barley basal resistance, which can be used to manage loose smut disease effectively.

Legumes are the second most significant plant source of human protein-rich diets after cereals. Additionally, these plants improve soil structure through biological nitrogen fixation. Plant pathogens are one of the most important global constraints on chickpea production. Plant-parasitic nematodes are key damaging agents to crop production. Broad beans, soybeans, lentils, beans, mung beans, and chickpeas are hosts for P. ritteri, and under greenhouse conditions, this nematode reduces the majority of plant growth indices. Various methods are employed to control plant-parasitic nematodes, most of which rely on chemical control. This study was conducted under laboratory and greenhouse conditions to examine the effects of refined wood vinegar, unrefined wood vinegar, chloro carboxylic acid, and sodium dioctyl sulfosuccinate on P. ritteri.

In this study, 66 soil and plant root samples were collected from chickpea fields in Lorestan province's counties. P. ritteri nematodes were identified after extraction using conventional techniques. The number of P. ritteri nematodes per kilogram of soil in an infected field soil was determined, and this soil was subsequently used in laboratory and greenhouse experiments. The effect of four compounds, including refined wood vinegar, unrefined wood vinegar, chloro carboxylic acid, and sodium dioctyl sulfosuccinate, on P. ritteri was investigated under laboratory and greenhouse conditions, as was the mortality of P. ritteri in chickpea. The data were analyzed using SPSS22 software, and tests were conducted using randomized complete block designs.

A comparison of the mean mortality of P. ritteri under laboratory conditions revealed that the highest mortality was associated with a 1% chloro carboxylic acid treatment for 24 h (100%), and the lowest mortality was attributed to a 0.5% refined wood vinegar treatment for 12 hours (25%). Under greenhouse conditions, the chloro carboxylic acid treatment resulted in the highest mortality, 1% (94%), while the unrefined wood vinegar treatment had the lowest mortality, 0.5% (49%). In addition, the mortality rate demonstrated a significant difference at P≤0.05 in two frames of 12 and 24 h for most compounds (except for sodium dioctyl sulfosuccinate 0.5%) and at P≤0.01 in two levels of 1% and 0. 5% for all compounds.

All investigated compounds induced nematode mortality, with chloro carboxylic acid proving to be the most effective for nematode mortality and unrefined wood vinegar the least effective. Consequently, these compounds can be used as pre-planting to control these nematodes.

Ultrasonic waves are mechanical waves that can adversely affect insects' biological and reproductive characteristics. On the other hand, low temperatures are regarded as a non-chemical and residue-free method for controlling stored-product insects because they affect the growth, development, population parameters, survival, reproduction, and various physiologic aspects of insects. In this research, due to the delayed effects of low temperatures, we examined the influence of pre-exposure of different developmental stages of the Mediterranean flour moth, Ephestia kuehniella Zeller, to ultrasonic waves on the lethal and sub-lethal effects of 4 °C temperature against emerged adults. The study's main goal was to mitigate the delayed effects of low temperatures.
 

All experiments were conducted at the Entomological Research Laboratory within the Department of Plant Protection at the Gorgan University of Agricultural Sciences and Natural Resources. The insect utilized for the research was reared on wheat flour at room conditions. Ultrasonic waves (20-60 kHz) were applied to eggs, first- and fifth instar larvae, and pupae. In addition, the following developmental stages were continuously exposed to ultrasonic waves: 4 days: egg pre-exposure; 9 days: eggs and first-instar larvae; 14 days: eggs to second-instar larvae; 19 days: eggs to third-instar larvae; 24 days: eggs to fourth-instar larvae; 33 days: eggs to fifth-instar larvae; and 40 days: eggs, five larval instars, and pupae. After pre-exposure treatments, emerged adults were exposed to 4 °C. Furthermore, LT50 and LT95 values were estimated for adults. Moreover, the sublethal effects of low temperature on male and female longevity, the fecundity of newly-emerged adults, and the percentage of egg hatches were investigated.

According to the results, pre-treatment of Mediterranean flour moth eggs with ultrasonic waves for 3 and 4 days significantly reduced the LT50 and LT95 of the exposed eggs when exposed to low temperature. In addition, pre-treatment of first and fifth instar larvae to ultrasonic waves did not reduce LTs of low temperature on emerged adults, whereas pre-treatment of pupae for eight days and consecutive pre-treatment of different developmental stages for 24, 33, and 40 days reduced LTs of low temperature significantly against emerged adults. In this study, although low temperature ultimately caused the death of adult insects, it increased the adults' longevity from 8-10 days up to 28-46 days. Furthermore, no significant effect of pre-treatment of eggs and first instar larvae with ultrasonic waves was observed on the longevity, fecundity and percentage of egg hatch of emerged adults. Moreover, pre-treatment of fifth instar larvae and pupae had a noticeable effect on adult longevity and fecundity, but did not affect egg hatchability. In all cases, pre-treatment of newly emerged adults with low temperature had a major effect on these traits. According to the findings, the longevity of male and female adults exposed to low temperature was significantly shorter than that of the control group following their release from these conditions. The results demonstrated that low temperature reduced pre-treated adult insects' fecundity and egg- hatching rate.

The current study's findings amply demonstrated that pre-exposure of some developmental stages (for example, larvae and pupae) of the Mediterranean flour moth could reduce LTs of low temperatures against emerged adults while also increasing the negative effects of these temperatures on their longevity, fecundity, and percentage egg hatch. This observation can be used to control stored-product insect pests.

The date is one of the most nutrient-dense fruit which is low in fat and protein but high in sucrose. Date syrup is rich in macro and micro nutrients. The possibility of replacing the date syrup with sugar in Apis mellifera was investigated. For this purpose, the stimulatory feeding effects on the biological and biochemical characteristics were studied, under natural conditions, in Karbala, Iraq. The average of brood rearing area in the sugar syrup (50%) grew over time (during February and March 2019) from 147 to 272.7 inches2; however, in the other treatments ("date syrup" and "date syrup+ sugar syrup"), there was no increase in brood rearing area. Totally, there was no significant difference in the areas of capped honey storage and pollen storage in honeybee colonies fed various diets. Concerning biochemical features of honey, the highest concentration of diverse chemicals was associated with sugar, while the lowest concentration was associated with date syrup. Furthermore, the honeybees fed with date syrup had an average of 51.64 percent protein, whereas those treated with sucrose syrup had a percentage of 47.5. The highest fat reserve belonged to the honeybees treated with a "sugar syrup + date syrup" with an average of 9.1 %, in comparison with sugar treatment (6 %). The present study revealed that sugar syrup and date syrup treatments resulted in the highest and the lowest average percentage of carbohydrates (43.9% and 37.72%), respectively.

Betasatellites, single-stranded circular DNAs, are multifunctional agents associated with monopartite begomoviruses (family Geminiviridae), that act as symptoms determinant. Begomoviruses are replicated by species-specific interactions between the viral replication-associated protein (Rep) and iteron motifs at the upstream of the origin of replication (ori). In contrast, promiscuous replication of betasatellites could be supported by different geminiviruses. In this study, the interaction of Cotton leaf curl Multan virus (CLCuMuV, genus Begomovirus) and Beet curly top virus (BCTV, genus Curtovirus) encoded Rep proteins with the iteron-like sequence of betasatellite, 5́-GAGGACC-3́, was investigated using in silico approaches. Nucleotide sequences of two Rep-encoding genes were obtained from the GenBank database, NCBI. Physicochemical characteristics of Rep proteins and their secondary and tertiary structures were predicted using the SOMPA tool and I-TASSER servers, respectively. The binding affinity of the best-predicted models of both proteins toward betasatellite iteron-like sequence was assessed using Docking simulations. The results represented reliable tertiary structures and showed structural similarity for Rep of different analyzed geminiviruses. Cluster analysis of HADDOCK revealed more total binding energy for CLCuMuV Rep toward the iteron-like sequence than BCTV complex. These in silico results confirmed the more trans-replication activity for relative geminiviruses in replication of betasatellite genomes. They emphasized the role of iteron-like sequences in interactions with the Rep of helper geminiviruses. Furthermore, targeting of identified activate sites within Rep protein structures to interact with betasatellite genomes could be considered as a control measure for begomovirus/betasatellite complexes.

Rice blast disease caused by Pyricularia oryzae is one of the most important rice diseases worldwide. Given the growing demand for non-toxic and chemical-free products, identification of plant growth-promoting microorganisms that can assist in mitigating challenges to plant growth is very important in sustainable rice cultivation. This study aimed to investigate the effect of two plant growth-promoting rhizobacteria (PGPR) isolates, Alcaligenes faecalis strain O1R4 and Bacillus idriensis strain MR2, on the induction of resistance-related enzymes such as peroxidase and catalase and on the severity of rice blast in Hashemi and Hassan Sarai cultivars.

The roots of the seedlings (cv. Hashemi and Hassan Sarai) were inoculated with a suspension of bacterial isolates (106-107 CFU/ml) for one hour and planted into pots. The plants were then sprayed in the four-leaf stage with an aqueous spore suspension (2×105 spore/ml) until run-off. The inoculated plants were transferred to the greenhouse at 28 °C and 90% relative humidity. Catalase and peroxidase activity in rice leaf extracts from different treatments were assayed one, three, five, and seven days after inoculation with blast fungus using a spectrophotometer. The activity was expressed in U µg–1 protein. The experiment was done in a completely randomized design with eight treatments and three replications for each cultivar. Also, the blast severity was assessed in the leaves of inoculated plants with six replications. The data were analyzed by SAS software (version 9.1) and Tukey's test at 0.05 and 0.01 probability levels. 

According to the analyses of variance, there was a significant influence of bacterial inoculation on the activity of peroxidase enzyme on the third, fifth, and seventh days. On the other hand, the activity of catalase enzyme increased on the first day to the seventh day at the 1% probability level. Mean comparison revealed in the treatments consisting of A. faecalis + P. oryzae, the levels of peroxidase and catalase enzymes were respectively elevated by 97% and 43% in Hashemi cultivar and by 174% and 125% in Hassan-Sarai cultivar compared to the positive control. Also, in the presence of B. idriensis + P. oryzae, the amount of peroxidase and catalase enzymes were respectively heightened in cv. Hashemi by 69% and 33% and in cv. Hassan-Sarai by 195% and 121% compared to the positive control. In the simultaneous use of two bacterial isolates and P. oryzae, the amount of catalase enzyme increased in Hashemi and Hassan-Sarai cultivars by 73% and 166% and the amount of peroxidase enzyme reached 174% and 319% in Hashemi and Hassan-Sarai cultivars, respectively compared to the positive control. Finally, the results showed that the blast severity was reduced using A. faecalis strain O1R4 and B. idriensis strain MR2 by 37% and 20% in the cultivars respectively.

In the present study, peroxidase activity in all treatments consisting of blast fungus was approximately at the same level within 24 hours post-inoculation. Then, the peroxidase level increased and maintained until the seventh day. Further, the catalase activity increased in the presence of the blast fungus and PGPRs. The level of peroxidase activity increased one to three days after inoculation. In comparison, the level of catalase increased within 24 hours after inoculation. The simultaneous use of two bacterial isolates along with pathogenic fungus was more effective than usage of each isolate separately. In general, the results of this research were consistent with previous studies that indicate the bacterial isolates can reduce the blast severity by inducing systemic resistance in rice.

 Orosanga japonica (Hemiptera: Ricaniidae) is an invasive and polyphagous pest in the Palearctic region collected and reported from the northern part of Iran on citrus orchards in 2010. Nymphs and adults of the pest cause economic and critical damage to different plants by direct feeding and heavy deposits of honeydew on leaves. Despite various management practices, successful control of this pest has been difficult due to its rapid development and reproduction rate. Beauveria bassiana, an entomopathogenic fungus (EPF), has been extensively researched for its ability to manage a variety of insect pests and appears to be a potential alternative to conventional insecticides.

 In the present study, in the course of a forest survey in the distribution areas of O. japonica in Mazandaran province, larvae were infected with some native isolates of fungal mycelia were observed. After morphological and molecular identification, two fungal-infected specimens were confirmed as B. bassiana. Fungal collections were characterized by amplifying and sequencing the ITS region of rDNA using primers ITS1 and ITS4. The susceptibility of adults of O. japonica to two EPFs isolates was evaluated using the leaf dipping method and the direct spraying method under laboratory conditions. Data were analyzed using Analysis of Variance (SAS, 2002) with insect mortality as the response variable. Means were separated at alpha=0.01 using Duncan´s test. The LC50 and LC90 values of EPFs and the mean lethal times of LT50 were estimated by Probit analysis using SPSS Statistics 17.0

Two different fungal isolates presented white to cream colonies with irregular edges and powdery appearance, reproductive structures, and conidia with typical morphology, size, and color of B. bassiana species. The mean lethal concentration values (LC50) of isolate 1 and isolate 2 to were 4.00 × 106 and 3.3 × 106 conidia ml-1 (leaf dipping method) and 5.4 × 106 and 4.7× 106 conidia ml-1 (direct spraying method) on the adults of O. japonica respectively in the laboratory tests after 96 h post treatments, at 25°C and 60% relative humidity. The similarities of the ITS sequences of both two EPFs were 99-100% to those of B. bassiana in the GenBank database.

The natural pathogens of pests have long been known to play a significant role in the population dynamics of many important forest pests and provide assistance to manage pests and develop tools for management. Fungal diversity in the moist and temperate Hyrcanian forests can vary widely due to diverse ecological conditions. Generally, these findings could help establish an efficient management strategy for O. japonica as an invasive pest and the development of mycoinsecticide from an indigenous fungus. Future research could investigate the effect of other native EPF isolates and developing techniques in fungal formulation against O. japonica.