Next Article in Journal
Identification and Transfer of a New Powdery Mildew Resistance Gene PmCAHM from Landrace Changanhongmai into Common Wheat
Next Article in Special Issue
Natural Enemies of the Pear Psylla, Cacopsylla pyri (Hemiptera: Psyllidae), and the Possibilities for Its Biological Control: A Case Study Review in the Western Balkan Countries
Previous Article in Journal
Rhizosphere-Associated Microbiota Strengthen the Pathogenicity of Meloidogyne incognita on Arabidopsis thaliana
Previous Article in Special Issue
Electrolyzed Oxidizing Water in Controlling Pseudomonas syringae pv. tomato in Tomato Crops
 
 
Search for Articles:
Title / Keyword
Author / Affiliation / Email
Journal
Article Type
 
 
Advanced Search
 
Section
Special Issue
Volume
Issue
Number
Page
 
Journals
Agronomy
Volume 14
Issue 4
10.3390/agronomy14040665
agronomy-logo
Submit to this Journal Review for this Journal Propose a Special Issue
► Article Menu

Article Menu

share Share announcement Help format_quote Cite question_answer Discuss in SciProfiles thumb_up
...
Endorse textsms
...
Comment
first_page
settings
Order Article Reprints
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Article

Impact of Three Entomopathogenic Fungal Isolates on the Growth of Tomato Plants—Ectoapplication to Explore Their Effect on Tetranychus urticae

by
Waqas Wakil
1,2,*,
Maria C. Boukouvala
3,
Nickolas G. Kavallieratos
3,*,
Aqsa Naeem
1,
Muhammad Usman Ghazanfar
4 and
Saleh S. Alhewairini
5
1
Department of Entomology, University of Agriculture, Faisalabad 38040, Pakistan
2
Senckenberg German Entomological Institute, D-15374 Müncheberg, Germany
3
Laboratory of Agricultural Zoology and Entomology, Department of Crop Science, Agricultural University of Athens, 75 Iera Odos Str., 11855 Athens, Greece
4
Department of Plant Pathology, College of Agriculture, Sargodha University, Sargodha 40100, Pakistan
5
Department of Plant Protection, College of Agriculture and Food, Qassim University, P.O. Box 6622, Buraidah 51452, Qassim, Saudi Arabia
*
Authors to whom correspondence should be addressed.
Agronomy 2024, 14(4), 665; https://doi.org/10.3390/agronomy14040665
Submission received: 18 February 2024 / Revised: 11 March 2024 / Accepted: 19 March 2024 / Published: 25 March 2024
(This article belongs to the Special Issue Sustainable Strategies for the Control of Crop Diseases and Pests to Reduce Pesticides)
Versions Notes

Abstract

:
Tetranychus urticae is an important pest of tomato crops globally, affecting plant yield and growth. Beauveria bassiana and Metarhizium robertsii have the potential to control T. urticae. We investigated the influence of two B. bassiana (i.e., WG-12 and WG-19) isolates and one M. robertsii (WG-02) isolate when colonizing different plant organs (leaves, stems, and roots) and their influence on the growth of tomato plants, through foliar, root-dipping, and seed-soaking application techniques. We also examined the acaricidal activity of the three isolates against T. urticae (female adults), spraying tomato leaf discs with each isolate separately. After 28 days, WG-12 and WG-19 colonized 97 and 91% of the leaves after foliar inoculation, whereas WG-02 exhibited the lowest leaf colonization (76%). The height of the tomato plants, the root length, the number of leaves, and the weight of the biomass above and below the ground were enhanced significantly after inoculation with WG-02 vs. B. bassiana isolates and control. The complete mortality of T. urticae was caused by WG-12 and WG-02 after 10 days, whereas WG-19 killed 94% of the adults. For the effective management of T. urticae, we propose the application of the WG-02 isolate since it provides complete protection and promotes the growth of tomato plants.

1. Introduction

Tomato, Solanum lycopersicum L. (Solanales: Solanaceae), is a species of high economic importance that is widely cultivated worldwide since it adapts to various agro-climatic environments [1,2,3,4]. The global tomato production is approximately 180,000,000 tons [5]. This fruit is also ranked second after the potato crop in terms of production and consumption [6]. However, tomato is attacked by several mites belonging to the Tarsonemidae, Eriophyidae, and Tetranychidae families, resulting in serious losses [7,8,9,10]. Among the tetranychids, Tetranychus urticae Koch (Acari) is a destructive pest of economic importance to tomato crops globally [8,9]. Tetranychus urticae damages a wide spectrum of plants, including ornamentals, vegetables, medicinals, and orchards [11,12,13]. This polyphagous pest has been recorded to feed on >1100 plant species from >140 plant families [14,15]. It causes an increase in water stress and a decrease in plant transpiration and photosynthesis by consuming leaf cell substances (e.g., chlorophyll) [16]. The feeding activity of T. urticae negatively affects plant yield and growth, causes leaf discoloration, also known as “bronzing”, or, more rarely, plant death [17,18,19]. Tetranychus urticae is short-lived and can rapidly develop high population densities, requiring approximately 7 days from the egg to the adult stage under optimal conditions (i.e., 27 °C and 55–60% humidity) [15,20]. Females may oviposit up to 150 eggs during their lifetime [15,21]. In addition, T. urticae reproduces by arrhenotokous parthenogenesis, where females emerge from fertilized eggs, while males emerge from unfertilized eggs [15,22]. Consequently, through this type of reproduction, a single unmated female can establish a mixed population after oedipal copulation (a female mates with her male progeny), resulting in serious economic losses [23,24].
Chemical acaricides have been widely used globally for the management of T. urticae [19,25,26]. However, several studies over the years have revealed that this pest has developed resistance to organophosphates [27,28], carboxamides [29], carbazates [25], pyrazols [30,31,32], quinazolines [33], organosulfurs [34], and organometallics [32]. Totally, T. urticae is resistant to 96 active ingredients in 526 cases; therefore, it is considered one of the most resistant pests among the arthropods [35]. Several factors contribute to the rapid development of acaricide resistance in T. urticae, such as the ability to reproduce by arrhenotokous parthenogenesis, large number of offspring, and short life cycle [36,37].
Many predatory mites have been used in controlled growth environments, such as greenhouses, to effectively manage T. urticae, but this approach is not as common in field crops where pesticide applications are dominant [38,39,40]. Another biocontrol option is the use of entomopathogenic fungi (EPF) against T. urticae, which can contribute to the natural population regulation of this pest [41]. In a biological control strategy, EPFs could replace conventional acaricides [41] or act synergistically with other biocontrol agents, such as predatory mites [42,43,44]. In addition, EPFs are not detrimental to non-target organisms or the environment, and are safe for humans [37]. Fungi are usually more advantageous over other microbial biopesticides (e.g., viruses and nematodes) because of their broader host selectivity since one isolate can attack and control different arthropod pests simultaneously [45,46,47,48]. Upon contact, the fungal conidia are attached to the host and penetrate the cuticula by secreting cuticle-degrading enzymes [49,50]. Then, the hyphae colonize the host’s body, leading to its death [51,52].
Several EPF species of Ascomycota, especially of the genera Metarhizium and Beauveria, have the potential to control T. urticae [53,54,55]. These fungi can also form symbiotic associations with plants (endophytes), living in plant tissues without symptoms through colonization [56,57]. Furthermore, EPFs have the potential to endophytically colonize a variety of plant species, improving plant growth and enhancing yields, while being harmful to pests [55,58,59,60]. There is evidence that EPFs make plants more resistant to stress caused by biotic or abiotic factors, enhance plant nutrient uptake, and excite hormone production, thereby contributing to plant growth [59,61,62].
Taking into account the limited knowledge related to the impact of isolates of entomopathogenic fungi on T. urticae and tomatoes in Pakistan [63], the current work aims to investigate for the first time the impact of three isolates of Beauveria bassiana (Balsamo-Crivelli) Vuillemin (Hypocreales: Cordycipitaceae) (WG-12 and WG-19) and Metarhizium robertsii J.F. Bisch., Rehner & Humber (formerly known as Metarhizium anisopliae (Metchnikoff) Sorokin) (Hypocreales: Clavicipitaceae) (WG-02) on the colonization and growth of tomato plants. Furthermore, the acaricidal efficacy of these isolates were tested against T. urticae after ectoapplication.

2. Materials and Methods

2.1. Mite Culture

Tetranychus urticae used in the bioassays was initially collected from a tomato field of the University of Agriculture, Faisalabad, Pakistan. This population had been nurtured for five years on the tomato cultivar (variety Moneymaker) susceptible to mites in an environment-controlled climate chamber set at a cycle of 16: 08 h light: dark, at 27 °C and 65% relative humidity (RH) [64]. Adult males are conical dorsally, while females are oval dorsally, and smaller than males, approximately 0.3–0.5 mm in length [65].

2.2. Tomato Plants

Tomato seeds (variety Moneymaker) were purchased from the local market and were sown in a seedling tray filled with moss and watered daily. This is the most promising cultivar and is extensively cultivated by Pakistani tomato growers [66,67]. For a period of three weeks post-sowing in the nursery, the seedlings were transferred to 3 L plastic pots filled with Sphagnum Peat Moss and were maintained in an environment-controlled chamber with a 16: 08 h light: dark cycle (27 °C, 65% RH). The plants were irrigated after a 3-day interval and fertilizer (macro- and micro-nutrients) was applied at 15-day intervals. The plants showed uniform growth characteristics and were used in further bioassays.

2.3. Suspensions of Beauveria bassiana and Metarhizium robertsii Isolates

In the current study, we used two entomopathogenic B. bassiana isolates (WG-12, WG-19) and one M. robertsii isolate (WG-02) [68]. Maintenance of these isolates was carried out on PDA (potato dextrose agar) (BD-Difco, Franklin Lakes, NJ, USA) stored in a refrigerator set at 4 °C, at the Microbial Control Laboratory, University of Agriculture, Faisalabad, Pakistan. Activation of the isolate was carried out in Petri dishes on SDA (sabouraud dextrose agar) (BD-Difco, Franklin Lakes, NJ, USA). Parafilm (Sigma-Aldrich Chemie GmbH, Taufkirchen, Germany) was used to seal the dishes and they were incubated for 10 days in 16: 08 h light: dark cycle at 25 °C (ICP-260, Memmert GmbH, Schwabach, Germany). These dishes produced an abundant amount of conidia two weeks post-inoculation. The layers of the conidia exiting from the media were scraped off with the help of a sterilized scalpel into a 10 mL tube. A falcon tube with a volume of 50 mL was used to suspend a portion of harvested conidia, containing a sterilized solution (30 mL) with 0.05% Tween 80 (Merck, Kenilworth, NJ, USA). For homogenized mixture, the conidia suspension was vortexed using a laboratory magnetic mixer (IRMECO GmbH, Lütjensee, Germany) with 8 glass beads. Measurement of the desired concentration was performed under the microscope with a Neubauer-Improved hemocytometer (Marienfeld, Germany). To measure conidia viability, two 60 mm Petri dishes with SDAY (sabouraud dextrose agar with 1% yeast) were inoculated with 0.1 mL (1 × 106 conidia/mL) of the conidial suspension, then sealed using parafilm and stored for 18 h for incubation at 25 °C with a photoperiod of 14: 10 h light: dark. Two SDAY dishes per B. bassiania and M. robertsii isolates were utilized and two counts including 200 conidia were recorded for each dish. Sterile cover slips were placed on the dishes after incubation. Conidia germination was considered successful when the germ tube length was 2× the conidia diameter [69,70]. This measurement was performed at magnification 400× using a microscope Euromex BB.1152-PLi microscope (Euromex Microscopen bv, Arnhem, The Netherlands). Before initiation of the experiments, conidia viability was >92%.

2.4. Inoculation Methods of Tomatoes with Suspensions of Beauveria bassiana and Metarhizium robertsii Isolates

Surfaces of seeds of tomato were sterilized with ethanol (70%) for 60–120 s, and later with a sodium hypochlorite solution (1.5%) for 120–180 s. The seeds were washed thrice using sterile distilled H2O to remove excess sodium hypochlorite, and then superfluid water was removed by blotting on sterile paper towels. The final rinsed H2O was spread on an SDA dish to evaluate the efficacy of the surface sterilization process [71]. Before inoculation, the viability of each entomopathogenic isolate was determined through germination tests as in the previous section. Three different methods were used for each isolate for their endophytic establishment in tomato plants as follows [72]: (1) Seed inoculation method: The sterilized seeds were immersed in a suspension of 1 × 108 conidia/mL in a 100 mL glass bottle with the respective fungus for 16–24 h [59,73]. Seeds that were treated with 0.01% Triton X-100 acted as controls. Then, seeds were placed on a sterile kitchen towel to air dry for an interval of 20 min. The seeds treated with WG-12, WG-19, and WG-02, and control were sown separately in 0.5 L volume plastic pots (8 cm × 10 cm height: diameter). Pots contained three-times-sterilized autoclaved plant substrates comprising compost and vermiculite. The planting material was autoclaved at 121 °C for 30 min, and left for 20 h intervals within each sterilization [73,74]. Next, the material was allowed to cool for 72 h before being transferred to the pots. The latter were maintained in a climatic chamber with a 16: 08 h light: dark cycle (27 °C, 65% RH) using fluorescent bulbs (Philips, Karachi, Pakistan) [75]. The pots were irrigated on alternate days [55], and 3 mL Hoagland solution (Merck KGaA, Darmstadt, Germany) was applied to each pot per week. (2) Foliar application: Six- to eight-week-old plants, raised from the sterilized seeds sown in the pots as described previously, were sprayed with a suspension (1 × 108 conidia/mL, 0.5 mL per leaf) of each isolate [76] with the help of a hand-held plastic sprayer (Kissan Ghar, Sargodha, Pakistan). Plants that were treated with 0.01% Triton X-100 acted as controls. Plants were maintained in an incubator with 16: 08 h light: dark cycle (27 °C, 65% RH) [64]. (3) Root-dipping: To observe the colonization of EPFs by root-dipping method, sterilized seeds were sown in pots as described previously. Then, a 15-day-old plant was gently removed from the soil, and its roots were immersed in a 300 mL volume of suspension (1 × 108 conidia/mL), corresponding to WG-12, WG-19, or WG-02, for 1.5 h [77]. Plants that were treated with 0.01% Triton X-100 acted as controls. Afterward, the plant was put back in the pot and kept in an incubator with 16: 08 h light: dark cycle (27 °C, 65% RH) [64].

2.5. Evaluation of Endophytic Colonization of Beauveria bassiana and Metarhizium robertsii isolates in Tomato Plants

The plants were uprooted gently to avoid any breakage of roots and other tissues. Per plant, five pieces of randomly selected leaves of 10 mm2 each, stems of 10 mm, and roots were cut with a sterilized blade [72]. They were sterilized with 1% NaClO for 300 s, then washed thrice with distilled H2O, and dried on sterile tissue paper [78]. The final rinsed H2O was spread on SDA media to check the efficiency of the disinfection process of each fungal isolate [79]. No contamination was observed in any of these dishes after sterilization of leaves, stems, and roots. After the sterilization process, with the help of a scalpel, the edges of the samples were trimmed and cut into pieces of 5 mm2 for leaves, 5 mm for stems, and 5 mm for roots [72]. These inoculated plant samples were placed in dishes containing the respective Beauveria or Metarhizium selective media as described by Rivas-Franco et al. [80]. Different dishes were used per group of leaves, stems, and roots per plant. The tissues were pressed in the selective media carefully to develop contact with the media [72], and the dishes were sealed with parafilm. Dishes were incubated for two weeks in an incubator at 26 °C, 16: 08 h light: dark cycle, and 84% RH [72]. Fungal growth was observed after 7, 14, 21, and 28 days of inoculation at the interface of the plated tissues, which were re-isolated and identified relying on conidia/conidiophores structures through microscope (BB.1152-PLi, Euromex Microscopen bv, Arnhem, The Netherlands) and comparing them with the original culture of each fungal isolate [72]. Different dishes with leaves, stems, and roots were prepared per exposure to evaluate the fungal growth originating from the same plant. The experiment was repeated independently using nine plants per inoculation method, including a total of 81 plants (i.e., 9 plants × 3 isolates × 3 inoculation methods). Data were presented as % colonization: number of tissues of plants exhibiting fungal outgrowth per total number of plant tissues) × 100 [72].

2.6. Influence of Endophytic Beauveria bassiana and Metarhizium robertsii Isolates on Plant Growth

The effect of the endophytic colonization of the isolates WG-12, WG-19, and WG-02 was evaluated on different plant growth parameters, by sowing new tomato seeds as described in the Section 2.4. The plants were inoculated with 1 × 108 conidia/mL of each fungal isolate separately by seed-soaking, foliar, and root-dipping methods. Per plant, the following growth physical parameters were evaluated 26 days post-inoculation: plant height (cm), i.e., distance between the tip of stem and base of the plant, using a steel ruler; stem diameter (mm), i.e., up to 30 mm from the ground, using Vernier caliper; number of leaves per plant; root length (cm), i.e., axial root, using a steel ruler; root dry weight (mg), using Shimadzu (Japan) balance; and above ground biomass (mg), i.e., leaves and whole stem. Above- and below-ground parts of the tomato were first dried inside an oven at 70 °C, and then weighed separately [72]. The same parameters for control plants were also recorded. The experiment was repeated independently using nine plants per inoculation method, including a total of 108 plants, i.e., 9 plants × 4 (isolates and control) × 3 inoculation methods.

2.7. Direct Action of Beauveria bassiana and Metarhizium robertsii Isolates against Tetranychus urticae under Laboratory Conditions

The bioassay was conducted in three Petri dishes (90 mm diameter) (subreplications) containing, each, a 3 cm diameter tomato leaf disc [81]. Each disc was put with the adaxial surface on water-saturated cotton to keep the leaf moist/fresh [82]. Twenty 1-day-old female adults [64] were placed separately in each disc [81]. Then, discs with the mites were sprayed with 1 mL [81] of the suspension (1 × 108 conidia/mL) of each isolate, using an airbrush (Master Multi-purpose Airbrush, USA). Control leaves were sprayed with 1 mL of H2O [37]. After spraying, leaves were left to air-dry for about 300 sec [82]. Afterward, dishes were covered with a PVC membrane (polyvinyl chloride), with fine holes for aeration [83], and stored in an incubator at 27 °C and 65% RH with a photoperiod of 16: 08 h light: dark. The mortality data were collected after 5, 7, and 10 days. Different leaves with mites were prepared per exposure. Observations were carried out under a stereomicroscope (Leica Wild M3B, Heerbrugg, Switzerland). Mites were considered dead if they could not move their appendages when touched with a camel hair brush [82]. The dead mites were placed on new plates containing a wet filter paper and stored in the same conditions for 3 days to observe any fungal growth to confirm their mortality caused by EPF [37,84]. The experiment was repeated four times using new plant leaves (i.e., 3 subreplications × 4 isolates and control × 4 replications).

2.8. Statistical Analysis

The data for the mortality of T. urticae were computed with Abbott’s formula [85]. All the data were transformed using the formula log (x + 1) to normalize variance prior to statistical analysis [86,87]. Regarding the EPF colonization experiment, the main effects were fungal isolate, inoculation method, different parts of the plant, and interval. Colonization was the response variable. The colonization data underwent a four-way analysis of variance (ANOVA). Data of plant growth were separately analyzed for plant height, stem diameter, root length, root dry weight, above-ground biomass, and number of leaves. The response variable was the plant growth of each part of the plant. The main effects were EPF isolate and inoculation method. The data for different plant growth parameters (i.e., plant height, stem diameter, root length, root dry weight, above-ground biomass, and number of leaves) were analyzed using two-way ANOVA. Regarding mortality data, exposure interval and inoculation methods were the main effects. Mortality was the response variable. Mortality data were subjected to two-way ANOVA. In all cases, the main effects and their interactions are considered in the analyses. Mortalities observed in the controls were <5%. Means were distinguished using the Tukey test (HSD) [88] at a level of significance of 5%. All analyses were carried out with the Minitab statistical package [89].

3. Results

3.1. Colonization by Endophytic Entomopathogenic Fungi Beauveria bassiana and Metarhizium robertsii Isolates in Tomato Plants

All the main effects and their associated interactions were significant, apart from isolate × plant part × interval, and isolate × methods × plant part × interval (Table S1). Among the inoculation methods, significantly different colonization rates were recorded throughout the experiment for WG-12. The colonization rate of the WG-12 isolate in the different parts of the tomato remained low, regardless of the inoculation method, reaching 46.66% in the leaves by the foliar method (Table 1). After 14 days, colonization by the foliar method was moderate in leaves (68.14%), whereas colonization of stems (47.40%) and roots (37.77%) was lower. This trend was observed after 21 days, with a higher colonization in leaves (83.70%) than stems (62.96%) and roots (49.63%) with the foliar method. Twenty-eight days later, leaf colonization reached 97.03% with the foliar inoculation method. However, stem and root colonization ranged between 47.40 and 76.29% with the three inoculation methods.
During the experimental period, there were significant differences among inoculation methods in WG-19 colonization rates (Table 1). The percentage of WG-19 colonized leaves, stems, and roots of tomato plants was low, not exceeding 40.74% regardless of the inoculation method, after 7 days. The percentage of colonization further increased at 14 days post-inoculation, with the highest percentage recorded for leaves (57.77%) with foliar inoculation, while the lowest percentage was reported for roots (17.77%) by seed-soaking inoculation. Similar results were obtained after 21 days, where leaves after foliar inoculation (74.81%) showed the highest colonization percentage, while colonization on tomato stems by the same inoculation method was lower (56.29%). However, in the rest of the plant part/inoculation method, colonization remained below 46.66%. At 28 days, an increase in colonization was observed in all cases, but, again, the highest percentages were found in leaves (91.11%) and stems (72.59%) after inoculation by the foliar method.
Significant differences were recorded in the colonization percentages of WG-02 to the different tomato parts when comparing inoculation methods at all observed intervals (Table 1). The percentage of colonization of the WG-02 isolate in the leaves, stems, and roots of tomato plants was very low and ranged from 5.18 to 24.44%, after 7 days. Fourteen days later, there was a slight increase in the colonization rate for all cases, reaching 42.96%. Twenty-one days after inoculation, the WG-02 isolate exhibited 58.51% colonization on leaves by the foliar method. However, colonization ranged from 16.29 (root by seed-soaking method) to 49.63% (stem by foliar method). After 28 days post-inoculation, the highest colonization percentages were reported in leaves (76.29%) and stems (67.40%) using the foliar method, while the proportion of colonized WG-02 isolate in the remaining cases did not exceed 41.48%.

3.2. Effect of Inoculation with Beauveria bassiana and Metarhizium robertsii Isolates on Tomato Plant Growth

All main effects/interactions were significant for the growth of the plant parts (Table S2). Inoculation with isolate WG-02 significantly improved the height of tomato plants compared to inoculation with isolates WG-12 or WG-19, or non-inoculated plants (control) (Table 2). In the case of isolate WG-02, significant differences were also observed among the three inoculation methods; i.e., the root-dipping method enhanced plant height more (27.02 cm) than seed-soaking (24.34 cm) and the foliation method (20.11 cm).
No significant differences in stem diameter were found by the foliar method between the fungal isolates and the control (Table 2). However, inoculation with WG-12 and WG-19 isolates with the root-dipping methods significantly increased stem diameter (i.e., 4.48 and 3.90 cm for WG-12 and WG-19, respectively) in comparison to inoculation by the foliar method (i.e., 3.61 and 3.17 cm for WG-12 and WG-19, respectively). Regarding the WG-02 isolate, inoculation with the root-dipping method resulted in a significantly wider stem (5.31 cm) than the other two methods (i.e., 4.26 and 3.75 cm for seed-soaking and foliar and methods, respectively).
The root was significantly longer with WG-02 isolate regardless of the inoculation method, when comparing to WG-12 and WG-19 isolates, and non-inoculated plants (Table 2). Inoculation with the three fungal isolates with the root-dipping method significantly enhanced root length (16.51, 18.27, and 19.37 cm for WG-19, WG-12, and WG-02 isolates, respectively) in contrast to the seed-soaking (14.38, 15.66, and 17.43 cm for WG-19, WG-12, and WG-02 isolates, respectively) and foliar method (12.88, 13.72, and 16.12 cm for WG-19, WG-12, and WG-02 isolates, respectively).
The root dry weight was significantly higher with the three inoculation methods using WG-02, in comparison to plants inoculated with WG-19 and WG-12, and non-inoculated plants (Table 2). Significant differences were noted among the inoculation methods for the three fungal isolates. The root dry weight was the highest with the root-dipping method (25.56, 29.16, and 38.36 mg for WG-19, WG-12, and WG-02 isolates, respectively), followed by the seed-soaking (21.42, 26.28, and 34.62 mg for WG-19, WG-12, and WG-02 isolates, respectively) and foliar method (17.62, 23.52, and 31.08 mg for WG-19, WG-12, and WG-02 isolates, respectively).
Inoculation with the three fungal isolates by the root-dipping method significantly increased the above-ground biomass weight (i.e., 187.47, 210.88, and 276.47 mg for WG-19, WG-12, and WG-02 isolates, respectively), compared to the seed-soaking (176.96, 198.23, and 223.39 mg for WG-19, WG-12, and WG-02 isolates, respectively) and foliar method (151.17, 165.80, and 194.27 mg for WG-19, WG-12, and WG-02, respectively) (Table 2). Regardless of the inoculation method, significant differences were noted among fungal isolates and controls in the above-ground biomass weight (i.e., WG-02 > WG-12 > WG-19 > control).
Concerning the number of leaves, the inoculated plants showed a significantly higher number of leaves than the control plants, regardless of the method used, except for the WG-19 isolate when inoculated by the foliar method, where the number of leaves did not vary significantly with the number of non-inoculated plants (i.e., 19.72 and 18.92 leaves/plant, for WG-19- and un-inoculated plants, respectively) (Table 2). The performance of WG-02 using the three inoculation methods resulted in a significantly higher number of leaves (24.26, 28.01, and 35.27 leaves/plant for the foliar, seed-soaking, and root-dipping methods, respectively), followed by WG-12 (21.17, 25.06, and 32.11 leaves/plant for the foliar, seed-soaking, and root-dipping methods, respectively), and WG-19 (19.72, 21.66, and 26.65 leaves/plant for the foliar, seed-soaking, and root-dipping methods, respectively). Inoculation by root-dipping method with the three isolates significantly enhanced leaf production in contrast to the seed-soaking and foliar methods.

3.3. Mortality Caused by Beauveria bassiana and Metarhizium robertsii Isolates on Tetranychus urticae in the Laboratory

All of the main effects/interactions (i.e., interval DF = 2, 107, F = 2264.5, p < 0.01, EPF isolate DF = 2, 107, F = 252.2, p < 0.01, and interval × EPF isolate DF = 4, 107, F = 50.6, p < 0.01) were significant. Significant differences were observed in adult mortality caused by the three fungal isolates after 5 days, with WG-02 (57.91%) showing moderate adult mortality, whereas mortality caused by WG-12 (45.41%) and WG-19 (29.58%) were low (Table 3). Mortality increased significantly after 2 days in all cases, with high mortality caused by WG-02 (81.25%), followed by WG-12 (68.75%) and WG-19 (63.33%). Complete mortality was caused by WG-12 and WG-02, while, in WG-19, adult mortality reached 94.58%, after 10 days.

4. Discussion

Entomopathogenic fungi have been artificially inoculated and successfully established as endophytes in several crops of economic importance, including tomato plants [73,77,90,91]. The results of our study demonstrated that the isolates tested were capable of endophytically colonizing the leaves, stems, and roots of tomato plants using three different inoculation methods. However, the percentage colonization of tomato tissues depended on the inoculation method and fungal isolate. It has been previously documented that these factors can affect the successful establishment of EPFs as endophytes [57,92,93]. In the current investigation, the three fungal isolates exhibited the highest percentage of leaf colonization by foliar inoculation as opposed to the root-dipping and seed-soaking methods. Allegrucci et al. [94] reported that foliar spray inoculation had a superior effect on the colonization percentage of an Argentine B. bassiana strain (LPSC 1067) on tomato leaves, compared to seed immersion and root-dipping methods. A significantly higher colonization of M. robertsii (80.0%) and B. bassiana (72.5%) on maize leaves was achieved by foliar spray than the seed inoculation (50 and 60% for B. bassiana and M. robertsii, respectively) [95]. Different inoculation methods (i.e., root/soil drenching, leaf soaking, and stem injection) failed to establish the colonization of the three B. bassiana isolates LPP139, LEF140, and LEF141 on tomato plants, whereas only LPP139 was able to successfully colonize all parts of the plant when inoculating water-stressed seedlings [96].
Interestingly, in each fungal strain, we noticed significant differences in the colonization percent among the different parts of tomato plants, regardless of the inoculation method used. The highest abundance of fungi was found in leaves, followed by stems and roots. Isolates WG-12 and WG-19 showed higher foliar colonization when compared to the WG-02 isolate, especially by the foliar method. Russo et al. [97] reported variation in the colonization performances of five B. bassiana and two Metarhizium isolates on the organs of soybean plants using different inoculation techniques. The leaf aspersion method benefited the establishment of fungi on leaves rather than the stems and roots, while B. bassiana strains showed a greater colonization rate over Metarhizium strains. The factors that caused an increase in the fungal colonization of leaves compared to the other organs are not yet clear. This could be due to variations in physiological conditions among the plant parts [97], an issue that merits further investigation.
The endophytic presence of EPFs in host plants has been reported to be beneficial, not only by protecting the plant from its enemies (e.g., pathogens or arthropods), but also by promoting plant development and yield [97,98,99,100]. Our results confirmed the promotion of plant growth, since, in most combinations (inoculation method/isolate), the three fungal isolates significantly enhanced the growth of tomato plants above and below the ground, increased the number of leaves per plant, the root dry weight, and the weight of the biomass above the ground, in comparison to the control plants. Several studies have shown the positive effect of different strains of both fungal species on the development of a wide range of host plants, including tomatoes [73,78,97,101,102,103]. Even though the mechanisms of plant growth enhancement by EPFs have not been clarified yet, some hypotheses attempted to clarify this issue. For instance, Jaber and Ownley [91] attributed the growth enhancement in colonized plants to the production of bioactive substances by Beauveria spp. González-Pérez et al. [102] correlated the high level of chlorophyll content found in inoculated plants with M. robertsii strains to an increase in their biomass, and the resulting plants were more vigorous than non-inoculated ones. Again, the inoculation method and fungal isolate affected plant growth. Our findings indicate that the three tested isolates have different effects on the growth enhancement of tomato plants. Furthermore, our results revealed that the inoculation method significantly affected plant growth, regardless of the isolate used here. Root-dipping inoculation proved more beneficial for the development of the plant organs and the biomass, followed by seed-soaking and foliar-spraying. In addition, root-dipping inoculation with WG-02 significantly enhanced plant growth in comparison to the plants inoculated with B. bassiana isolates or control plants.
Concerning the acaricidal activity against T. urticae, all three isolates killed 100% of the exposed adult females after 10 days, except WG-19 which resulted in 94.58% mortality. Interestingly, WG-02 exhibited higher mortality after 5 days compared to B. bassiana strains. The differences in the virulence of the three isolates against T. urticae may be related to differences in the protease activity of EPF [49,104]. Elhakim et al. [52] observed higher protease activity in the M. robertsii isolate compared to the B. bassiana isolate, which was correlated with the elevated virulence of the former strain against T. urticae. Furthermore, in a recent study, Khamis Al-Zahrani et al. [37] found that the activity of protease in B. bassiana was the lowest vs. other fungi belonging to the genera Fusarium, Aspergillus, Scopulariopsis, and Penicillium after a screening performed on skimmed milk agar medium. All EPFs were obtained from Rhynchophorus ferrugineus (Olivier) (Coleoptera: Curculionidae) with the exception of one Fusarium isolate obtained from T. urticae.

5. Conclusions

Our results revealed a multifactorial activity of the WG-12, WG-19, and WG-02 isolates that can be utilized to protect tomato plants from T. urticae and enhance plant growth at the same time. The WG-02 isolate was by far the most effective isolate that provided complete protection in tomatoes from the mite and promoted plant development in several ways. The application method of the EPFs should be taken into consideration since it played a significant role in fungal colonization, plant growth, and protection against T. urticae. Taking into account the results of the current study, we recommend the foliar spraying of WG-02, since, from a practical point of view, it is the easiest method that can be used in the field [93]. As a next step, further research should be conducted to demonstrate that the fungal isolates of M. robertsii and B. bassiana of the present study can act as endophytes protecting tomato plants against T. urticae or other mite species in the greenhouse and in the field.

Supplementary Materials

The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/agronomy14040665/s1, Table S1. ANOVA parameters for main effects and associated interactions for colonization of Beauveria bassiana and Metarhizium robertsii isolates with three inoculation methods in tomato plants (Total DF = 971). Table S2. ANOVA parameters for main effects and associated interaction for plant growth (Total DF = 109).

Author Contributions

Conceptualization, W.W. and N.G.K.; methodology, W.W. and N.G.K.; software, W.W., A.N. and M.U.G.; validation, W.W., A.N., M.C.B., N.G.K. and S.S.A.; formal analysis, W.W., A.N., M.C.B., N.G.K. and S.S.A.; investigation, W.W., A.N. and M.U.G.; resources, W.W. and M.U.G.; data curation, W.W., A.N. and M.U.G.; writing—original draft preparation, M.C.B., W.W., N.G.K., M.U.G. and S.S.A.; writing—review and editing, W.W., M.C.B., N.G.K., A.N., M.U.G. and S.S.A.; visualization, W.W., A.N., M.C.B. and N.G.K.; supervision, W.W.; project administration, W.W.; funding acquisition, W.W. All authors have read and agreed to the published version of the manuscript.

Funding

This study was partly funded by Project 3244 of the Higher Education Commission, Islamabad, Pakistan.

Data Availability Statement

The data are contained within the article.

Conflicts of Interest

The authors declare no conflicts of interest.

References

  1. Gupta, V.J.; Chattoo, M.A.; Singh, L. Drip irrigation and fertigation technology for improved yield, quality, water and fertilizer use efficiency in hybrid tomato. J. Agric. Res. 2015, 2, 94–99. [Google Scholar]
  2. Gerszberg, A.; Hnatuszko-Konka, K.; Kowalczyk, T.; Kononowicz, A.K. Tomato (Solanum lycopersicum L.) in the service of biotechnology. Plant Cell Tissue Organ Cult. 2015, 120, 881–902. [Google Scholar] [CrossRef]
  3. Naz, N.; Khan, I.; Gul, B.; Ayub, G.; Jan, F.; Jang, N.; Shuaib, M. Response of tomato (Lycopersicon esculentum Mill.) growth to different phosphorous levels and sowing dates. Acta Ecol. Sin. 2019, 39, 30–35. [Google Scholar] [CrossRef]
  4. Tekkam, S.; Pidigam, S.; Adapa, K.K.; Natarajan, S.; Amarapalli, G. Evaluation of parents and hybrids for yield and yield contributing traits in tomato (Solanum lycopersicum L.) for per se performance. J. Pharmacogn. Phytochem. 2021, 10, 1770–1776. [Google Scholar]
  5. FAOSTAT (Food and Agriculture Organization of the United Nations). Crops. Available online: http://www.fao.org/faostat/en/#data/QC/visualize (accessed on 29 January 2024).
  6. Al-Shaal, A.; Murshed, R.A.; Albiski, F.A. Molecular characterization of some tomato genotypes using inter simple sequence repeats (ISSR) technique. J. Genet. Environ. Resour. Conserv. 2021, 9, 134–142. [Google Scholar]
  7. Navajas, M.; de Moraes, G.J.; Auger, P.; Migeon, A. Review of the invasion of Tetranychus evansi: Biology, colonization pathways, potential expansion and prospects for biological control. Exp. Appl. Acarol. 2013, 59, 43–65. [Google Scholar] [CrossRef]
  8. Leppla, N.C.; Johnson, M.W.; Merritt, J.L.; Zalom, F.G. Applications and trends in commercial biological control for arthropod pests of tomato. In Sustainable Management of Arthropod Pests of Tomato; Wakil, W., Brust, G.E., Perring, T.M., Eds.; Academic Press: London, UK, 2018; pp. 283–303. [Google Scholar]
  9. Assouguem, A.; Kara, M.; Mechchate, H.; Korkmaz, Y.B.; Benmessaoud, S.; Ramzi, A.; Abdullah, K.R.; Noman, O.M.; Farah, A.; Lazraq, A. Current situation of Tetranychus urticae (Acari: Tetranychidae) in Northern Africa: The sustainable control methods and priorities for future research. Sustainability 2022, 14, 2395. [Google Scholar] [CrossRef]
  10. Vervaet, L.; De Vis, R.; De Clercq, P.; van Leeuwen, T. Is the emerging mite pest Aculops lycopersici controllable? Global and genome-based insights in its biology and management. Pest Manag. Sci. 2021, 77, 2635–2644. [Google Scholar] [CrossRef] [PubMed]
  11. Naher, N.; Islam, T.; Haque, M.M.; Parween, S. Effects of native plants and IGRs on the development of Tetranychus urticae Koch (Acari: Tetranychidae). Univ. J. Zool. Rajshahi. Univ. 2006, 25, 19–22. [Google Scholar] [CrossRef]
  12. Vacante, V. The Handbook of Mites of Economic Plants: Identification, Bio-Ecology and Control; CABI International: Wallingford, UK, 2016. [Google Scholar]
  13. Migeon, A.; Nouguier, E.; Dorkeld, F. Spider mites web: A comprehensive database for the Tetranychidae. In Trends in Acarology; Sabelis, M.W., Bruin, J., Eds.; Springer: Dordrecht, The Netherlands, 2010; pp. 557–560. [Google Scholar]
  14. Rioja, C.; Zhurov, V.; Bruinsma, K.; Grbic, M.; Grcic, V. Plant-herbivore interactions: A case of an extreme gemeralist, the two-spotted spider mite Tetranychus urticae. Mol. Plant Microbe Interact 2017, 30, 935–945. [Google Scholar] [CrossRef] [PubMed]
  15. Namin, H.H.; Zhurov, V.; Spenler, J.; Grbic, M.; Grbic, V.; Scott, I.M. Resistance to pyridaben in Canadian greenhouse populations of two-spotted spider mites, Tetranychus urticae (Koch). Pestic. Biochem. Physiol. 2020, 170, 104677. [Google Scholar] [CrossRef]
  16. Bensoussan, N.; Santamaria, M.E.; Zhurov, V.; Diaz, I.; Grbic, M.; Grbic, V. Plant-herbivore interaction: Dissection of the cellular pattern of Tetranychus urticae feeding on the host plant. Front. Plant Sci. 2016, 7, 1105. [Google Scholar] [CrossRef]
  17. Tomczyk, A.; Kropczynska, D. Effects on the host plant. In Spider Mites, Their Biology, Natural Enemies and Control; Helle, W., Sabelis, M.W., Eds.; Elsevier: Amsterdam, The Netherlands, 1985; pp. 312–330. [Google Scholar]
  18. Park, Y.L.; Lee, J.H. Leaf cell and tissue damage of cucumber caused by two spotted spider mite (Acari: Tetranychidae). J. Econ. Entomol. 2002, 95, 952–957. [Google Scholar] [CrossRef]
  19. Attia, S.; Grissa, K.L.; Lognay, G.; Bitume, E.; Hance, T.; Mailleux, A.C. A review of the major biological approaches to control the worldwide pest Tetranychus urticae (Acari: Tetranychidae) with special reference to natural pesticides. J. Pest Sci. 2013, 86, 361–386. [Google Scholar] [CrossRef]
  20. Ghongade, D.S.; Sood, A.K. Diversity and seasonal abundance of tetranychid mites in polyhouses in Himachal Pradesh. Indian J. Entomol. 2018, 80, 1688–1695. [Google Scholar] [CrossRef]
  21. Shih, C.T.; Poe, S.L.; Cromroy, H.L. Biology, life table and intrinsic rate of increase of Tetranychus urticae. Ann. Entomol. Soc. Am. 1976, 69, 362–364. [Google Scholar] [CrossRef]
  22. Helle, W.; Sabelis, M.W. Spider Mites: Their Biology, Natural Enemies, and Control; World Crop Pests; Elsevier: Amsterdam, The Netherlands, 1985; Volume 1A. [Google Scholar]
  23. Toyoshima, S.; Amano, H. Comparison of development and reproduction in offspring produced by females of Phytoseiulus persimilis Athias-Henriot (Acari: Phytoseiidae) under two prey conditions. Appl. Entomol. Zool. 1999, 34, 285–292. [Google Scholar] [CrossRef]
  24. Tuan, S.J.; Lin, Y.H.; Yang, C.M.; Atlihan, R.; Saska, P.; Chi, H. Survival and reproductive strategies in two-spotted spider mites: Demographic analysis of arrhenotokous parthenogenesis of Tetranychus urticae (Acari: Tetranychidae). J. Econ. Entomol. 2016, 109, 502–509. [Google Scholar] [CrossRef]
  25. Van Leeuwen, T.; Van Pottelberge, S.; Tirry, L. Biochemical analysis of a chlorfenapyr-selected resistant strain of Tetranychus urticae Koch. Pest Manag. Sci. 2006, 62, 425–433. [Google Scholar] [CrossRef] [PubMed]
  26. Choi, Y.S.; Baek, S.; Kim, M.J. Effect of temperature on the development and survival of Feltiella acarisuga (Vallot) (Diptera: Cecidomyiidae) preying on Tetranychus urticae (Koch) (Acari: Tetranychidae). Insects 2021, 12, 508. [Google Scholar] [CrossRef] [PubMed]
  27. Edge, V.E.; James, D.G. Organotin resistance in Tetranychus urticae (Acari: Tetranychidae) in Australia. J. Econ. Entomol. 1986, 79, 1477–1483. [Google Scholar] [CrossRef]
  28. Sato, M.E.; Suplicy Filho, N.; de Souza Filho, M.F.; Takematsu, A.P. Resistência do ácaro rajado Tetranychus urticae (Koch, 1836) (Acari: Tetranychidae) a diversos acaricidas em morangueiro (Fragaria sp.) nos municípios de Atibaia-SP e Piedade-SP. Ecossistema 1994, 19, 40–46. [Google Scholar]
  29. Herron, G.; Rophail, J. Genetics of hexythiazox resistance in the two-spotted spider mite, Tetranychus urticae Koch. Exp. Appl. Acarol. 1993, 17, 423–431. [Google Scholar] [CrossRef]
  30. Sato, M.E.; Miyata, T.; da Silva, M.; Raga, A.; de Souza Filho, M.F. Selections for fenpyroximate resistance and susceptibility, and inheritance, cross-resistance and stability of fenpyroximate resistance in Tetranychus urticae Koch (Acari: Tetranychidae). Appl. Entomol. Zool. 2004, 39, 293–302. [Google Scholar] [CrossRef]
  31. Van Pottelberge, S.; Van Leeuwen, T.; Nauen, R.; Tirry, L. Resistance mechanisms to mitochondrial electron transport inhibitors in a field-collected strain of Tetranychus urticae Koch (Acari: Tetranychidae). Bull. Entomol. Res. 2009, 99, 23–31. [Google Scholar] [CrossRef] [PubMed]
  32. Simma, E.A.; Hailu, B.; Jonckheere, W.; Rogiers, C.; Duchateau, L.; Dermauw, W.; Van Leeuwen, T. Acaricide resistance status and identification of resistance mutations in populations of the two-spotted spider mite Tetranychus urticae from Ethiopia. Exp. Appl. Acarol. 2020, 82, 475–491. [Google Scholar] [CrossRef] [PubMed]
  33. Vassiliou, V.A.; Kitsis, P. Resistance in Tetranychus urticae (Acari: Tettranychidae) populations from Cyprus. J. Econ. Entomol. 2013, 106, 1848–1854. [Google Scholar] [CrossRef]
  34. Kumari, S.; Chauhan, U.; Kumari, A.; Nadda, G. Comparative toxicities of novel and conventional acaricides against different stages of Tetranychus urticae Koch (Acarina: Tetranychidae). J. Saudi Soc. Agric. Sci. 2017, 16, 191–196. [Google Scholar] [CrossRef]
  35. Adesanya, A.W.; Lavine, M.D.; Moural, T.W.; Lavine, L.C.; Zhu, F.; Walsh, D.B. Mechanisms and management of acaricide resistance for Tetranychus urticae in agroecosystems. J. Pest Sci. 2021, 94, 639–663. [Google Scholar] [CrossRef]
  36. Van Leeuwen, T.; Vontas, J.; Tsagkarakou, A.; Tirry, L. Acaricide resistance mechanism in the two-spotted spider mite Tetranychus urticae and other important Acari: A review. Insect Biochem. Mol. Biol. 2010, 40, 563–572. [Google Scholar] [CrossRef]
  37. Al-Zahrani, J.K.; Al-Abdalall, A.H.; Osman, M.A.; Aldakheel, L.A.; AlAhmady, N.F.; Aldakeel, S.A.; AbdulAzeez, S.; Borgio, J.F.; ElNaggar, M.A.; Alabdallah, N.M.; et al. Entomopathogenic fungi and their biological control of Tetranychus urticae: Two-spotted spider mites. J. King Saud Univ. Sci. 2023, 35, 102910. [Google Scholar] [CrossRef]
  38. Van Leeuwen, T.; Tirry, L.; Yamamoto, A.; Nauen, R.; Dermauw, W. The economic importance of acaricides in the control of phytophagous mites and an update on recent acaricide mode of action research. Pestic. Biochem. Physiol. 2015, 121, 12–21. [Google Scholar] [CrossRef]
  39. Golec, J.R.; Hoge, B.; Walgenbach, J.F. Effect of biopesticides on different Tetranychus urticae Koch (Acari: Tetranychidae) life stages. Crop Prot. 2020, 128, 105015. [Google Scholar] [CrossRef]
  40. Ndiaye, S.G.; Welty, C. Augmentation and conservation biological control of Tetranychus urticae on hops in Ohio. Biol. Control 2022, 173, 104980. [Google Scholar] [CrossRef]
  41. Tehri, K.; Gulati, R.; Geroh, M.; Dhankhar, S.K. Dry weather: A crucial constraint in the field efficacy of entomopathogenic fungus Beauveria bassiana against Tetranychus urticae Koch (Acari: Tetranychidae). J. Entomol. Zool. Stud. 2015, 3, 287–291. [Google Scholar]
  42. Dogan, Y.O.; Hazir, S.; Yildiz, A.; Butt, T.M.; Cakmak, I. Evaluation of entomopathogenic fungi for the control of Tetranychus urticae (Acari: Tetranychidae) and the effect of Metarhizium brunneum on the predatory mites (Acari: Phytoseiidae). Biol. Control 2017, 111, 6–12. [Google Scholar] [CrossRef]
  43. Zhang, X.N.; Guo, J.J.; Zou, X.; Jin, D.C. Pathogenic differences of the entomopathogenic fungus Isaria cateniannulata to the spider mite Tetranychus urticae (Trombidiformes: Tetranychidae) and its predator Euseius nicholsi (Mesostigmata: Phytoseiidae). Exp. Appl. Acarol. 2018, 75, 69–84. [Google Scholar] [CrossRef] [PubMed]
  44. Castillo-Ramírez, O.; Guzmán-Franco, A.W.; Santillán-Galicia, M.; Tamayo-Mejía, F. Interaction between predatory mites (Acari: Phytoseiidae) and entomopathogenic fungi in Tetranychus urticae populations. BioControl 2020, 65, 433–445. [Google Scholar] [CrossRef]
  45. Montalva, C.; Rocha, L.F.; Fernandes, É.K.; Luz, C.; Humber, R.A. Conidiobolus macrosporus (Entomophthorales), a mosquito pathogen in Central Brazil. J. Invertebr. Pathol. 2016, 139, 102–108. [Google Scholar] [CrossRef]
  46. Lee, S.J.; Lee, M.R.; Kim, S.; Kim, J.C.; Park, S.E.; Li, D.; Kim, J.S. Genomic analysis of the insect-killing fungus Beauveria bassiana JEF-007 as a biopesticide. Sci. Rep. 2018, 8, 12388. [Google Scholar] [CrossRef]
  47. Canassa, F.; Esteca, F.C.; Moral, R.A.; Meyling, N.V.; Klingen, I.; Delalibera, I. Root inoculation of strawberry with the entomopathogenic fungi Metarhizium robertsii and Beauveria bassiana reduces incidence of the two-spotted spider mite and selected insect pests and plant diseases in the field. J. Pest Sci. 2020, 93, 261–274. [Google Scholar] [CrossRef]
  48. Basnet, P.; Dhital, R.; Rakshit, A. Biopesticides: A genetics, genomics, and molecular biology perspective. In Advances in Bio-inoculant Science, Biopesticides; Rakshit, A., Meena, V.S., Abhilash, P.C., Sarma, B.K., Singh, H.B., Fraceto, L., Parihar, M., Singh, A.K., Eds.; Woodhead Publishing: Cambridge, UK, 2022; pp. 107–116. [Google Scholar]
  49. Revathi, N.; Ravikumar, G.; Kalaiselvi, M.; Gomathi, D.; Uma, C. Pathogenicity of three entomopathogenic fungi against Helicoverpa armigera. J. Plant Pathol. Microbiol. 2011, 2, 114. [Google Scholar] [CrossRef]
  50. Charnley, A.K. Fungal pathogens of insects: Cuticle degrading enzymes and toxins. Adv. Bot. Res. 2003, 40, 241–321. [Google Scholar]
  51. Mondal, S.; Baksi, S.; Koris, A.; Vatai, G. Journey of enzymes in entomopathogenic fungi. Pac. Sci. Rev. A Nat. Sci. Eng. 2016, 18, 85–99. [Google Scholar] [CrossRef]
  52. Elhakim, E.; Mohamed, O.; Elazouni, I. Virulence and proteolytic activity of entomopathogenic fungi against the two-spotted spider mite, Tetranychus urticae Koch (Acari: Tetranychidae). Egypt. J. Biol. Pest Control 2020, 30, 30. [Google Scholar] [CrossRef]
  53. Castro, T.; Eilenberg, J.; Delalibera, I. Exploring virulence of new and less studied species of Metarhizium spp. from Brazil for two-spotted spider mite control. Exp. Appl. Acarol. 2018, 74, 139–146. [Google Scholar] [CrossRef]
  54. Canassa, F.; Tall, S.; Moral, R.A.; Lara, I.A.R.; Delalibera, I., Jr.; Meyling, N.V. Effects of bean seed treatment by the entomopathogenic fungi Metarhizium robertsii and Beauveria bassiana on plant growth, spider mite populations and behavior of predatory mites. Biol. Control 2019, 132, 199–208. [Google Scholar] [CrossRef]
  55. Rasool, S.; Cárdenas, P.D.; Pattison, D.I.; Jensen, B.; Meyling, N.V. Isolate-specific effect of entomopathogenic endophytic fungi on population growth of two-spotted spider mite (Tetranychus urticae Koch) and levels of steroidal glycoalkaloids in tomato. J. Chem. Ecol. 2021, 47, 476–488. [Google Scholar] [CrossRef]
  56. Bamisile, B.; Dash, C.K.; Akutse, K.; Keppanan, R.; Afolabi, O.G.; Hussain, M.; Qasim, M.; Wang, L. Prospects of endophytic fungal entomopathogens as biocontrol and plant growth promoting agents: An insight on how artificial inoculation methods affect endophytic colonization of host plants. Microbiol. Res. 2018, 217, 34–50. [Google Scholar] [CrossRef]
  57. Vega, F.E. The use of fungal entomopathogens as endophytes in biological control: A review. Mycologia 2018, 110, 4–30. [Google Scholar] [CrossRef]
  58. Jaber, L.R.; Araj, S.-E. Interactions among endophytic fungal entomopathogens (Ascomycota: Hypocreales), the green peach aphid Myzus persicae Sulzer (Homoptera: Aphididae), and the aphid endoparasitoid Aphidius colemani Viereck (Hymenoptera: Braconidae). Boil. Control 2018, 116, 53–61. [Google Scholar] [CrossRef]
  59. Sánchez-Rodríguez, A.R.; Raya-Díaz, S.; Zamarreño, Á.M.; García-Mina, J.M.; del Campillo, M.C.; Quesada-Moraga, E. An endophytic Beauveria bassiana strain increases spike production in bread and durum wheat plants and effectively controls cotton leafworm (Spodoptera littoralis) larvae. Biol. Control 2018, 116, 90–102. [Google Scholar] [CrossRef]
  60. Tall, S.; Meyling, N.V. Probiotics for plants? Growth promotion by the entomopathogenic fungus Beauveria bassiana depends on nutrient availability. Microb. Ecol. 2018, 76, 1002–1008. [Google Scholar] [CrossRef]
  61. Ownley, B.H.; Gwinn, K.D.; Vega, F.E. Endophytic fungal entomopathogens with activity against plant pathogens: Ecology and evolution. BioControl 2010, 55, 113–128. [Google Scholar] [CrossRef]
  62. Sasan, R.K.; Bidochka, M.J. The insect-pathogenic fungus Metarhizium robertsii (Clavicipitaceae) is also an endophyte that stimulates plant root development. Am. J. Bot. 2012, 99, 101–107. [Google Scholar] [CrossRef] [PubMed]
  63. Amjad, M.; Bashir, M.H.; Afzal, M.; Sabri, M.A.; Javed, N. Synergistic effect of some entomopathogenic fungi and synthetic pesticides, against two spotted spider mite, Tetranychus urticae Koch (Acari: Tetranychidae). Pakistan J. Zool. 2012, 44, 977–984. [Google Scholar]
  64. Keskin, N.; Kumral, N.A. Screening tomato varietal resistance against the two-spotted spider mite [Tetranychus urticae (Koch)]. Int. J. Acarol. 2015, 41, 300–309. [Google Scholar] [CrossRef]
  65. Sarwar, M. Mite (Acari Acarina) vectors involved in transmission of plant viruses. In Applied Plant Virology; Awasthi, L.P., Ed.; Academic Press: Cambridge, MA, USA, 2020; pp. 257–273. [Google Scholar]
  66. Chaudhry, Z.; Abbas, S.; Yasmin, A.; Rashid, H.; Ahmed, H.; Anjum, M.A. Tissue culture studies in tomato (Lycopersicon esculentum) var. Moneymaker. Pak. J. Bot. 2010, 42, 155–163. [Google Scholar]
  67. Ijaz, A.; Khan, I.; Zareen, S.; Khan, M.I.; Khan, R.; Haroon, M. Yield and yield attributes of tomato (Lycopersicon esculentum Mill) cultivars influenced by weed management techniques. Pak. J. Weed Sci. Res. 2017, 23, 431–438. [Google Scholar]
  68. Tahir, T.; Wakil, W.; Ali, A.; Sahi, S.T. Pathogenicity of Beauveria bassiana and Metarhizium anisopliae isolates against larvae of the polyphagous pest Helicoverpa armigera. Entomol. Gen. 2019, 38, 225–242. [Google Scholar] [CrossRef]
  69. Usman, M.; Gulzar, S.; Wakil, W.; Wu, S.; Piñero, J.C.; Leskey, T.C.; Nixon, L.J.; Oliveira-Hofman, C.; Toews, M.D.; Shapiro-Ilan, D. Virulence of entomopathogenic fungi to Rhagoletis pomonella (Diptera: Tephritidae) and interactions with entomopathogenic nematodes. J. Econ. Entomol. 2020, 113, 2627–2633. [Google Scholar] [CrossRef]
  70. Gulzar, S.; Wakil, W.; Shapiro-Ilan, D.I. Combined effect of entomopathogens against Thrips tabaci Lindeman (Thysanoptera: Thripidae): Laboratory, greenhouse and field trials. Insects 2021, 12, 456. [Google Scholar] [CrossRef]
  71. Parsa, S.; Ortiz, V.; Vega, F.E. Establishing fungal entomopathogens as endophytes: Towards endophytic biological control. J. Vis. Exp. 2013, 74, e50360. [Google Scholar]
  72. Zheng, Y.; Liu, Y.; Zhang, J.; Liu, X.; Ju, Z.; Shi, H.; Mendoza-Mendoza, A.; Zhou, W. Dual role of endophytic entomopathogenic fungi: Induce plant growth and control tomato leafminer Phthorimaea absoluta. Pest Manag. Sci. 2023, 79, 4557–4568. [Google Scholar] [CrossRef]
  73. Dash, C.K.; Bamisile, B.S.; Keppanan, R.; Qasim, M.; Lin, Y.; Islam, S.U.; Hussain, M.; Wang, L.; Ravindran, K. Endophytic entomopathogenic fungi enhance the growth of Phaseolus vulgaris L. (Fabaceae) and negatively affect the development and reproduction of Tetranychus urticae Koch (Acari: Tetranychidae). Microb. Pathog. 2018, 125, 385–392. [Google Scholar] [CrossRef]
  74. Sani, I.; Jamian, S.; Saad, N.; Abdullah, S.; Mohd Hata, E.; Jalinas, J.; Ismail, S.I. Inoculation and colonization of the entomopathogenic fungi, Isaria javanica and Purpureocillium lilacinum, in tomato plants, and their effect on seedling growth, mortality and adult emergence of Bemisia tabaci (Gennadius). PLoS ONE 2023, 18, e0285666. [Google Scholar] [CrossRef] [PubMed]
  75. Wakil, W.; Ghazanfar, M.U.; Usman, M.; Hunter, D.; Shi, W. Fungal-based biopesticide formulations to control nymphs and adults of the desert locust, Schistocerca gregaria Forskål (Orthoptera: Acrididae): A laboratory and field cage study. Agronomy 2022, 12, 1160. [Google Scholar] [CrossRef]
  76. Nishi, O.; Sushida, H.; Higashi, Y.; Iida, Y. Epiphytic and endophytic colonisation of tomato plants by the entomopathogenic fungus Beauveria bassiana strain GHA. Mycology 2021, 12, 39–47. [Google Scholar] [CrossRef]
  77. Qayyum, M.A.; Wakil, W.; Arif, M.J.; Sahi, S.T.; Dunlap, C.A. Infection of Helicoverpa armigera by endophytic Beauveria bassiana colonizing tomato plants. Biol. Control 2015, 90, 200–207. [Google Scholar] [CrossRef]
  78. Wei, Q.Y.; Li, Y.Y.; Xu, C.; Wu, Y.X.; Zhang, Y.R.; Liu, H. Endophytic colonization by Beauveria bassiana increases the resistance of tomatoes against Bemisia tabaci. Arthropod Plant Interact. 2020, 14, 289–300. [Google Scholar] [CrossRef]
  79. Muvea, A.M.; Meyhöfer, R.; Subramanian, S.; Poehling, H.M.; Ekesi, S.; Maniania, N.K. Colonization of onions by endophytic fungi and their impacts on the biology of Thrips tabaci. PLoS ONE 2014, 9, e108242. [Google Scholar] [CrossRef] [PubMed]
  80. Rivas-Franco, F.; Hampton, J.G.; Narciso, J.; Rostás, M.; Wessman, P.; Saville, D.J.; Jackson, T.A.; Glare, T.R. Effects of a maize root pest and fungal pathogen on entomopathogenic fungal rhizosphere colonization, endophytism and induction of plant hormones. Biol. Control 2020, 150, 104347. [Google Scholar] [CrossRef]
  81. Seiedy, M.; Moezipour, M. The entomopathogenic fungus Beauveria bassiana and its compatibility with Phytoseiulus persimilis (Acari: Phytoseiidae): Effects on Tetranychus urticae (Acari: Tetranychidae). Persian J. Acarol. 2017, 6, 329–338. [Google Scholar]
  82. Wu, S.; Sarkar, S.C.; Lv, J.; Xu, X.; Lei, Z. Poor infectivity of Beauveria bassiana to eggs and immatures causes the failure of suppression on Tetranychus urticae population. BioControl 2020, 65, 81–90. [Google Scholar] [CrossRef]
  83. Wu, S.Y.; Xie, H.C.; Li, M.Y.; Xu, X.N.; Lei, Z.R. Highly virulent Beauveria bassiana strains against the two-spotted spider mite, Tetranychus urticae, show no pathogenicity against five phytoseiid mite species. Exp. Appl. Acarol. 2016, 70, 421–435. [Google Scholar] [CrossRef] [PubMed]
  84. Sewify, G.; Mikhail, W.; Rizk, M.; Hassan, D. Using a biological control method for controlling red spider mite. Egypt Acad. J. Biol. Sci. 2015, 7, 115–126. [Google Scholar] [CrossRef]
  85. Abbott, W.S. A Method of Computing the Effectiveness of an Insecticide. J. Econ. Entomol. 1925, 18, 265–267. [Google Scholar] [CrossRef]
  86. Zar, J.H. Biostatistical Analysis; Pearson Education Limited: Essex, UK, 2014. [Google Scholar]
  87. Scheff, D.S.; Arthur, F.H. Fecundity of Tribolium castaneum and Tribolium confusum adults after exposure to deltamethrin packaging. J. Pest Sci. 2018, 91, 717–725. [Google Scholar] [CrossRef]
  88. Sokal, R.R.; Rohlf, F.J. Biometry; Freeman & Company: New York, NY, USA, 1995. [Google Scholar]
  89. Minitab, L.L.C. Getting Started with Minitab 18; Minitab Inc.: State College, PA, USA, 2017; p. 73. [Google Scholar]
  90. Russo, M.L.; Pelizza, S.A.; Cabello, M.N.; Stenglein, S.A.; Scorsetti, A.C. Endophytic colonisation of tobacco, corn, wheat and soybeans by the fungal entomopathogen Beauveria bassiana (Ascomycota, Hypocreales). Biocontrol Sci. Technol. 2014, 25, 475–480. [Google Scholar] [CrossRef]
  91. Jaber, L.R.; Ownley, B.H. Can we use entomopathogenic fungi as endophytes for dual biological control of insect pests and plant pathogens? Biol. Control 2018, 116, 36–45. [Google Scholar] [CrossRef]
  92. Akello, J.; Dubois, T.; Gold, C.S.; Coyne, D.; Nakavuma, J.; Paparu, P. Beauveria bassiana (Balsamo) Vuillemin as an endophyte in tissue culture banana (Musa spp.). J. Invertebr. Pathol. 2007, 96, 34–42. [Google Scholar] [CrossRef]
  93. Posada, F.; Aime, M.C.; Peterson, S.W.; Rehner, S.A.; Vega, F.E. Inoculation of coffee plants with the fungal entomopathogen Beauveria bassiana (Ascomycota: Hypocreales). Mycol. Res. 2007, 111, 748–757. [Google Scholar] [CrossRef] [PubMed]
  94. Allegrucci, N.; Velazquez, M.S.; Russo, M.L.; Pérez, M.E.; Scorsetti, A.C. Endophytic colonisation of tomato by the entomopathogenic fungus Beauveria bassiana: The use of different inoculation techniques and their effects on the tomato leafminer Tuta absoluta (Lepidoptera: Gelechiidae). J. Plant Prot. Res. 2017, 54, 205–211. [Google Scholar] [CrossRef]
  95. Altaf, N.; Ullah, M.I.; Afzal, M.; Arshad, M.; Ali, S.; Rizwan, M.; Al-Shuraym, L.A.; Alhelaify, S.S.; Sayed, S. Endophytic colonization by Beauveria bassiana and Metarhizium anisopliae in maize plants affects the fitness of Spodoptera frugiperda (Lepidoptera: Noctuidae). Microorganisms 2023, 11, 1067. [Google Scholar] [CrossRef]
  96. Silva, A.C.L.; Silva, G.A.; Abib, P.H.N.; Carolino, A.T.; Samuels, R.I. Endophytic colonization of tomato plants by the entomopathogenic fungus Beauveria bassiana for controlling the South American tomato pinworm, Tuta absoluta. CABI Agric. Biosci. 2020, 1, 3. [Google Scholar]
  97. Russo, M.L.; Pelizza, S.; Vianna, M.; Allegrucci, N.; Cabello, M.N.; Toledo, A.V.; Mourelos, C.; Scorsetti, A.C. Effect of endophytic entomopathogenic fungi on soybean Glycine max (L.) Merr. growth and yield. J. King Saud Univ. Sci. 2018, 31, 728–736. [Google Scholar] [CrossRef]
  98. Brownbridge, M.; Reay, S.D.; Nelson, T.L.; Glare, T.R. Persistence of Beauveria bassiana (Ascomycota: Hypocreales) as an endophyte following inoculation of radiata pine seed and seedlings. Biol. Control 2012, 61, 194–200. [Google Scholar] [CrossRef]
  99. Lopez, D.C.; Sword, G.A. The endophytic fungal entomopathogens Beauveria bassiana and Purpureocillium lilacinum enhance the growth of cultivated cotton (Gossypium hirsutum) and negatively affect survival of the cotton bollworm (Helicoverpa zea). Biol. Control 2015, 89, 53–60. [Google Scholar] [CrossRef]
  100. Barelli, L.; Moonjely, S.; Behie, S.W.; Bidochka, M.J. Fungi with multifunctional lifestyles: Endophytic insect pathogenic fungi. Plant Mol. Biol. 2016, 90, 657–664. [Google Scholar] [CrossRef]
  101. Elena, G.J.; Beatriz, P.J.; Alejandro, P.; Lecuona, R. Metarhizium anisopliae (Metschnikoff) Sorokin promotes growth and has endophytic activity in tomato plants. Adv. Biol. Res. 2011, 5, 22–27. [Google Scholar]
  102. González-Pérez, E.; Ortega-Amaro, M.A.; Bautista, E.; Delgado-Sánchez, P.; Jiménez-Bremont, J.F. The entomopathogenic fungus Metarhizium anisopliae enhances Arabidopsis, tomato, and maize plant growth. Plant Physiol. Biochem. 2022, 176, 34–43. [Google Scholar] [CrossRef] [PubMed]
  103. Liu, Y.; Yang, Y.; Wang, B. Entomopathogenic fungi Beauveria bassiana and Metarhizium anisopliae play roles of maize (Zea mays) growth promoter. Sci. Rep. 2022, 12, 15706. [Google Scholar] [CrossRef] [PubMed]
  104. Zare, M.; Talaei-Hassanloui, R.; Fotouhifar, K. Relatedness of proteolytic potency and virulence in entomopathogenic fungus Beauveria bassiana isolates. J. Crop Prot. 2014, 3, 425–434. [Google Scholar]
Table 1. Colonization (mean% ± SE) of tomato plants with WG-12, WG-19 isolates of Beauveria bassiana, and WG-02 isolate of Metarhizium robertsii under three different inoculation methods (foliar, root-dipping, and seed-soaking). Per each plant part, within each column, means followed by the same lowercase letters are not significantly different (DF = 2, 26; Tukey HSD test at p = 0.05).
Table 1. Colonization (mean% ± SE) of tomato plants with WG-12, WG-19 isolates of Beauveria bassiana, and WG-02 isolate of Metarhizium robertsii under three different inoculation methods (foliar, root-dipping, and seed-soaking). Per each plant part, within each column, means followed by the same lowercase letters are not significantly different (DF = 2, 26; Tukey HSD test at p = 0.05).
WG-12
IntervalsMethodLeaf ColonizationStem ColonizationRoot Colonization
7 daysFoliar46.66 ± 1.92 a34.81 ± 1.85 a26.67 ± 1.92 a
Root-dipping25.18 ± 1.48 b21.48 ± 1.48 b32.59 ± 1.73 a
Seed-soaking28.14 ± 2.15 b37.03 ± 1.17 a18.51 ± 1.85 b
F38.530.414.8
p<0.01<0.01<0.01
14 daysFoliar68.14 ± 2.15 a47.40 ± 1.73 a37.77 ± 1.57 a
Root-dipping34.07 ± 1.73 b28.14 ± 2.15 b41.48 ± 1.48 a
Seed-soaking39.25 ± 2.06 b45.92 ± 2.06 a23.70 ± 1.95 b
F84.728.931.0
p<0.01<0.01<0.01
21 daysFoliar83.70 ± 1.95 a62.96 ± 1.17 a49.63 ± 2.25 a
Root-dipping44.44 ± 1.57 b35.55 ± 1.92 c46.66 ± 2.72 a
Seed-soaking46.66 ± 1.92 b54.07 ± 1.73 b37.03 ± 1.17 b
F146.072.59.4
p<0.01<0.01<0.01
28 daysFoliar97.03 ± 2.25 a76.29 ± 1.61 a55.55 ± 1.11 a
Root-dipping53.33 ± 1.92 b47.40 ± 1.73 c58.51 ± 1.48 a
Seed-soaking59.25 ± 2.06 b66.66 ± 1.57 b50.37 ± 1.17 b
F129.080.210.6
p<0.01<0.01<0.01
WG-19
7 daysFoliar40.74 ± 1.33 a31.58 ± 1.48 a22.96 ± 1.17 a
Root-dipping19.25 ± 1.33 c17.03 ± 1.17 b28.14 ± 3.09 a
Seed-soaking25.92 ± 2.06 b32.59 ± 1.73 a12.59 ± 0.74 b
F46.435.116.3
p<0.01<0.01<0.01
14 daysFoliar57.77 ± 1.57 a43.70 ± 1.17 a31.85 ± 1.85 a
Root-dipping30.37 ± 1.61 c22.96 ± 1.17 c34.07 ± 1.73 a
Seed-soaking36.29 ± 1.17 b39.25 ± 1.33 b17.77 ± 1.57 b
F96.879.026.3
p<0.01<0.01<0.01
21 daysFoliar74.81 ± 1.48 a56.29 ± 1.17 a42.96 ± 1.95 a
Root-dipping38.51 ± 1.48 b29.63 ± 1.17 c41.48 ± 1.48 a
Seed-soaking40.74 ± 1.73 b46.66 ± 1.57 b34.81 ± 1.48 b
F168.0105.06.9
p<0.01<0.01<0.01
28 daysFoliar91.11 ± 1.11 a72.59 ± 2.06 a47.40 ± 2.06 ab
Root-dipping48.14 ± 1.48 b40.74 ± 1.73 c52.59 ± 1.73 a
Seed-soaking52.59 ± 1.73 b59.25 ± 1.73 b43.70 ± 1.17 b
F260.074.66.9
p<0.01<0.01<0.01
WG-02
7 daysFoliar24.44 ± 1.57 a18.51 ± 1.48 a15.55 ± 1.11 a
Root-dipping10.37 ± 1.61 b13.33 ± 1.11 b19.25 ± 1.73 a
Seed-soaking7.40 ± 0.74 b11.85 ± 1.48 b5.18 ± 0.97 b
F44.26.530.6
p<0.01<0.01<0.01
14 daysFoliar42.96 ± 2.51 a26.66 ± 1.57 a19.25 ± 1.73 b
Root-dipping14.81 ± 0.97 b17.03 ± 1.17 b25.18 ± 1.85 a
Seed-soaking12.59 ± 1.73 b18.51 ± 1.48 b9.62 ± 1.17 c
F83.613.423.6
p<0.01<0.01<0.01
21 daysFoliar58.51 ± 1.85 a49.63 ± 1.61 a38.51 ± 1.48 a
Root-dipping27.40 ± 2.34 b31.11 ± 1.57 b33.33 ± 1.57 b
Seed-soaking21.48 ± 1.48 b26.66 ± 1.92 b16.29 ± 1.17 c
F107.050.767.2
p<0.01<0.01<0.01
28 daysFoliar76.29 ± 1.17 a67.40 ± 1.73 a41.48 ± 2.15 a
Root-dipping30.37 ± 1.95 b35.55 ± 1.57 b39.25 ± 1.73 a
Seed-soaking28.14 ± 1.48 b32.59 ± 1.73 b22.22 ± 1.57 b
F299.0131.032.8
p<0.01<0.01<0.01
Table 2. Plant growth parameters (plant height, stem diameter, root length, root dry weight, above-ground biomass, and number of leaves/plant) (mean ± SE) of tomato plants inoculated with three isolates of Beauveria bassiana (WG-12 and WG-19) and Metarhizium robertsii (WG-02) using three different methods (foliar, root-dipping, and seed-soaking). Within each row, means followed by the same lowercase letters are not significantly different (DF = 3, 35; Tukey HSD test at p = 0.05). Per response, within each column, means followed by the same uppercase letters are not significantly different (DF = 2, 26; Tukey HSD test at p = 0.05).
Table 2. Plant growth parameters (plant height, stem diameter, root length, root dry weight, above-ground biomass, and number of leaves/plant) (mean ± SE) of tomato plants inoculated with three isolates of Beauveria bassiana (WG-12 and WG-19) and Metarhizium robertsii (WG-02) using three different methods (foliar, root-dipping, and seed-soaking). Within each row, means followed by the same lowercase letters are not significantly different (DF = 3, 35; Tukey HSD test at p = 0.05). Per response, within each column, means followed by the same uppercase letters are not significantly different (DF = 2, 26; Tukey HSD test at p = 0.05).
Plant Growth Parameter
Plant Height (cm)
MethodWG-12WG-19WG-02ControlFp
Foliar16.02 ± 0.26 Bb15.84 ± 0.24 Bb20.11 ± 0.37 Ca14.81 ± 0.52 Bb40.1<0.01
Root-dipping22.10 ± 0.27 Ab19.78 ± 0.37 Ac27.02 ± 0.23 Aa16.45 ± 0.21 Ad245.0<0.01
Seed-soaking21.87 ± 0.31 Ab18.62 ± 0.36 Ac24.34 ± 0.33 Ba15.76 ± 0.22 ABd142.0<0.01
F146.036.6118.05.5
p<0.01<0.01<0.010.01
Stem diameter (mm)
Foliar3.61 ± 0.19 Ba3.17 ± 0.18 Ba3.75 ± 0.13 Ba3.68 ± 0.16 Aa2.40.08
Root-dipping4.48 ± 0.25 Aab3.90 ± 0.16 Ab5.31 ± 0.24 Aa2.55 ± 0.32 Bc20.6<0.01
Seed-soaking3.86 ± 0.15 ABab3.58 ± 0.18 ABab4.26 ± 0.27 Ba3.38 ± 0.20 ABb3.30.03
F4.74.212.16.0
p0.010.02<0.01<0.01
Root length (cm)
Foliar13.72 ± 0.52 Cb12.88 ± 0.42 Cb16.12 ± 0.26 Ca10.43 ± 0.31 ABc35.2<0.01
Root-dipping18.27 ± 0.28 Ab16.51 ± 0.24 Ac19.37 ± 0.30 Aa9.45 ± 0.24 Bd267.0<0.01
Seed-soaking15.66 ± 0.25 Bb14.38 ± 0.37 Bc17.43 ± 0.21 Ba11.27 ± 0.31 Ad77.1<0.01
F37.225.738.338.3
p<0.01<0.01<0.01<0.01
Root dry weight (mg)
Foliar23.52 ± 0.25 Cb17.62 ± 0.22 Cc31.08 ± 0.36 Ca15.63 ± 0.23 ABd642.0<0.01
Root-dipping29.16 ± 0.38 Ab25.56 ± 0.45 Ac38.36 ± 0.36 Aa16.51 ± 0.29 Ad571.0<0.01
Seed-soaking26.28 ± 0.26 Bb21.42 ± 0.34 Bc34.62 ± 0.30 Ba15.23 ± 0.30 Bd718.0<0.01
F86.2126.0113.05.3
p<0.01<0.01<0.010.01
Above-ground biomass (mg)
Foliar165.80 ± 2.20 Cb151.17 ± 2.48 Cc194.27 ± 1.59 Ca98.62 ± 1.27 Ad422.0<0.01
Root-dipping210.88 ± 3.45 Ab187.47 ± 2.70 Ac276.47 ± 1.12 Aa96.55 ± 1.66 Ad952.0<0.01
Seed-soaking198.23 ± 2.03 Bb176.96 ± 2.17 Bc223.39 ± 1.65 Ba97.51 ± 1.56 Ad843.0<0.01
F77.357.5796.00.5
p<0.01<0.01<0.010.63
Number of leaves/plant
Foliar21.17 ± 0.49 Cb19.72 ± 0.40 Cc24.26 ± 0.29 Ca18.92 ± 0.61 Ac25.5<0.01
Root-dipping32.11 ± 0.47 Ab26.65 ± 0.36 Ac35.27 ± 0.35 Aa20.21 ± 0.43 Ad261.0<0.01
Seed-soaking25.06 ± 0.36 Bb21.66 ± 0.44 Bc28.01 ± 0.41 Ba19.74 ± 0.39 Ad81.0<0.01
F153.077.8241.01.8
p<0.01<0.01<0.010.18
Table 3. Mortality (mean% ± SE) of Tetranychus urticae female adults treated with Beauveria bassiana (WG-12 and WG-19) and Metarhizium robertsii (WG-02) isolates. Within each row, means followed by the same uppercase letter do not differ significantly. Within each column, means with the same lowercase letter do not differ significantly (DF = 2, 35 in all cases, Tukey HSD test at p = 0.05).
Table 3. Mortality (mean% ± SE) of Tetranychus urticae female adults treated with Beauveria bassiana (WG-12 and WG-19) and Metarhizium robertsii (WG-02) isolates. Within each row, means followed by the same uppercase letter do not differ significantly. Within each column, means with the same lowercase letter do not differ significantly (DF = 2, 35 in all cases, Tukey HSD test at p = 0.05).
Exposure
Isolate5 Days7 Days10 DaysFp
WG-1245.41 ± 1.14 Cb68.75 ± 1.64 Bb100.00 ± 0.00 Aa561.0<0.01
WG-1929.58 ± 1.29 Cc63.33 ± 1.42 Bc94.58 ± 1.29 Ab587.0<0.01
WG-0257.91 ± 2.08 Ca81.25 ± 1.39 Ba100.00 ± 0.00 Aa212.0<0.01
F82.438.017.4--
p<0.01<0.01<0.01--
Disclaimer/Publisher’s Note: The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of MDPI and/or the editor(s). MDPI and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content.

Share and Cite

MDPI and ACS Style

Wakil, W.; Boukouvala, M.C.; Kavallieratos, N.G.; Naeem, A.; Ghazanfar, M.U.; Alhewairini, S.S. Impact of Three Entomopathogenic Fungal Isolates on the Growth of Tomato Plants—Ectoapplication to Explore Their Effect on Tetranychus urticae. Agronomy 2024, 14, 665. https://doi.org/10.3390/agronomy14040665

AMA Style

Wakil W, Boukouvala MC, Kavallieratos NG, Naeem A, Ghazanfar MU, Alhewairini SS. Impact of Three Entomopathogenic Fungal Isolates on the Growth of Tomato Plants—Ectoapplication to Explore Their Effect on Tetranychus urticae. Agronomy. 2024; 14(4):665. https://doi.org/10.3390/agronomy14040665

Chicago/Turabian Style

Wakil, Waqas, Maria C. Boukouvala, Nickolas G. Kavallieratos, Aqsa Naeem, Muhammad Usman Ghazanfar, and Saleh S. Alhewairini. 2024. "Impact of Three Entomopathogenic Fungal Isolates on the Growth of Tomato Plants—Ectoapplication to Explore Their Effect on Tetranychus urticae" Agronomy 14, no. 4: 665. https://doi.org/10.3390/agronomy14040665

Note that from the first issue of 2016, this journal uses article numbers instead of page numbers. See further details here.

Article Metrics

Back to TopTop