Comparative effects of temperature and thermoregulation on candidate strains of entomopathogenic fungi for Moroccan locust <Emphasis Type="Italic">Dociostaurus maroccanus</Emphasis> control

Strains IMI 330189 of Metarhizium acridum (Driver & Milner) J.F. Bisch., Rehner & Humber (Hypocreales: Clavicipitaceae) and EABb 90/2-Dm of Beauveria bassiana (Bals.-Criv.) Vuill. (Hypocreales: Cordycipitaceae) are a promising biocontrol tool of Dociostaurus maroccanus (Thunberg) (Orthoptera: Acrididae), although the effects of thermoregulation and Moroccan locust-fever on the infection process of these fungi remain unknown. In vitro experiments, measuring conidial germination and hyphal growth either at constant or at fluctuating temperatures simulating thermoregulatory conditions, indicated that strain IMI 330189 had greater fitness at temperatures above 27 °C and the strain EABb 90/2-Dm was better adapted to the temperature range of 10–25 °C. These effects were mirrored in vivo, where locust thermoregulation caused a marked reduction in the virulence of EABb 90/2-Dm in comparison to no thermoregulation conditions (average survival time: 6.10 vs. 15.83 days; mortality: 100% vs. 73.7%) but only a moderate reduction in the virulence of IMI 330189 (average survival time: 4.57 vs. 8.26 days; mortality: 100% vs. 100%). Thermal gradient experiments revealed that the strain IMI 330189 induced behavioral fever in D. maroccanus (preferred temperatures approximately 4 °C above the uninfected control), although it only led to a slight reduction in virulence. Strain EABb 90/2-Dm did not induce such a clear behavioral response. Under the temperature conditions of the main breeding areas of the Moroccan locust, strain IMI 330189 is likely a better candidate for use in biocontrol, although strain EABb 90/2-Dm could be also a good alternative in more temperate environments either as a stand-alone one or in mixed combinations of the two fungal strains, potentially providing more effective control over a broader range of temperatures.     

Insecticidal and antifeedant activities of proteins secreted by entomopathogenic fungi against Spodoptera littoralis (Lep., Noctuidae)

Abstract:  Entomopathogenic fungi are a poorly exploited source of insecticidal proteins, which may be used for the development of new natural insecticides and as alternative molecules for transgenic deployment. The crude soluble protein extracts in Adamek's liquid medium of 25 fungal isolates belonging to the fungal species Metarhizum anisopliae , Beauveria bassiana , Beauveria brongniartii and Scopulariopsis brevicaulis were screened by per os toxicity on Spodoptera littoralis larvae. Whilst extracts from two M. anisopliae and two B. bassiana isolates gave significant mortalities when applied either on alfalfa leaf discs or incorporated into artificial diet, the one from M. anisopliae 01/58-Su isolate was the only most toxic that showed promise for S. littoralis control. In leaf disc assays, this extract exhibited strong dose-related toxic and antifeedant activity against the larvae. At 10, 20 and 40  μ g protein/insect, the extract gave 61.3%, 96.6% and 100.0% mortality, respectively, and average survival time of 5.7, 4.3 and 3.1 days respectively. Not only the antifeeding index was dose-related, but it significantly increased over time in a dose-related manner. Longer exposure times led to a dose-related significant increase in larval mortality. The exposure times for 50% mortality were 91.3 h and 62.1 h for 20 and 40  μ g protein/insect respectively. The crude extract when exposed to higher temperature or protease treatment lost toxicity, indicating that toxicity was protein mediated. In addition, the liquid medium composition did not influence its insecticidal activity. The effects of the protein extract on midgut cells of second instar larvae of S. littoralis were investigated by using both light and electron microscopy. A progressive bleeding of the midgut epithelium into the gut lumen was observed along with lysis of the epithelium and deterioration of the microvilli.     

Insect-toxic secreted proteins and virulence of the entomopathogenic fungus Beauveria bassiana

Fungal virulence has been mostly associated with cuticle-degrading enzymes that can be regulated depending on nutrient conditions. However, few studies have related fungal virulence to insect-toxic secreted proteins. Here, we describe how the presence of secreted toxic proteins may be linked to conidial virulence, which can be affected by nutrient factors. In this study we evaluated: (1) the virulence of the conidia of four Beauveria bassiana strains (EABb 01/103-Su, EABb 01/12-Su, EABb 01/88-Su and EABb 01/110-Su) grown on three different media (malt extract agar (MA), Rice (Rice), Sabouraud dextrose agar (SDA) and harvested from the cadavers of fungal-infected Galleria mellonella larvae (CAD) and (2) the toxicity of the crude soluble protein extracts (CSPEs) obtained from Adamek’s liquid medium inoculated with these conidia. Conidial suspensions were obtained from the four media, assessed on G. mellonella larvae and used to produce CSPEs that were injected into healthy G. mellonella larvae. The larvae were also injected with conidia obtained from MA and CAD cultures to expose them to in vivo-secreted proteins. For all isolates, the CAD conidia were by far the most virulent, followed by conidia grown on SDA, Rice and MA. The injected CSPEs showed the same toxicity trends as the conidial suspensions. In addition, the outcomes of injection of the in vivo-secreted proteins showed that the toxic proteins secreted in vitro by the EABb 01/110-Su strain are not produced in vivo. However, the other strains produced toxic proteins both in vivo and in vitro, suggesting that these toxic proteins may be virulence factors involved in invertebrate pathogenesis.     

Reproduction of Symptoms of a Root Disease of Wheat by Toxic Metabolites Produced by Two Pythium Species and their Partial Characterization

Abstract Three Pythium isolates [ Pythium arrhenomanes isolates (320) & (323)], and Pythium irregulare produced in vitro metabolites that affect normal wheat seedling growth and development. P. arrhenomanes isolates (320 & 323) produced toxic metabolites that caused a general browning of root tissue of 2–3-day-old wheat seedlings, root stunting, inhibition of root hair formation, and reduced fresh and dry root weights. These symptoms were also seen in infected seedlings inthe field. Culture filtrates of P. irregulare only partly reproduced disease symptoms, as they inhibited root elongation, but did not cause any browning of root tissue or inhibition of root hair formation. P. irregulare filtrates also stunted shoot growth and stimulated root hair formation and elongation in a swollen area immediately behind the root tip. Autoclaving did not affect activity of P. irregulare culture filtrates, but did inactivate P. arrhenomanes culture filtrates, suggesting that they are different. Ultrafiltration separations of P. arrhenomanes (isolate 320) filtrates indicated that toxic metabolites were heterogeneous. One component was less than 1,000 mol.wt. and the other greater than 50,000 mol.wt., but less than 100,000 mol.wt.     

Effects of the timing of applications on the incompatibility of three fungicides and one isolate of the entomopathogenic fungus Beauveria bassiana (Balsamo) Vuillemin (Deuteromycotina)

Abstract: The effect of timing of application on the incompatibility of three fungicides (metalaxyl, mancozeb, copper oxide) and isolate MK2001 of the entomopathogenic fungus Beauveria bassiana (Balsamo) Vuillemin was assessed in vitro . Isolate MK 2001 at 1 × 10⁷ conidia/ml concentration was highly pathogenous to adults of Lygus lineolaris (Hemiptera) (84% mortality at day 4 post-treatment). The fungicides at the manufacturer recommended rate (1X) were strongly fungistatic and inhibited the fungal isolate radial growth on Sabouraud dextrose agar (SDA). Mancozeb, metalaxyl and copper oxide exhibited insecticidal activity of 24, 40 and 48% respectively, on L. lineolaris adults. It is clear that application of the fungal isolate MK2001 2–4 days before applying fungicides metalaxyl, mancozeb or copper oxide synergized the insecticidal effect of the isolate. On the contrary, application of metalaxyl, mancozeb, copper oxide 2–4 days before applying isolate MK2001 antagonized the insecticidal effect. The simultaneous use of each fungicide (metalaxyl, mancozeb or copper oxide) and the isolate gave lesser insect mortality. This study revealed that although some fungicides are incompatible for use alongside fungal isolates, the proper evaluation of their time of use could be beneficial in biological control or IPM programmes. Furthermore, the application of B. bassiana isolate MK2001 followed by fungicide application could synergize insecticidal activity.     

球形芽孢杆菌Ts—1毒蛋白的分离纯化

Bacillus sphaericus strain Ts-1 is highly insecticidal to larvae of the mosquito. It's insecticidal component is toxic proteins. The toxin was extracted from spore-crystal complexes by disruption in a Sonicator Cell Disruptor Model W-220F followed by treatment with 0.05 mol/L NaOH. Fraction recovered from chromatography of the spore-crystal complexes on column of Sephadex G-200 were assayed against mosquito larvae and the toxic fractions from gel chromatography were subjected to SDS-PAGE. The toxic proteins in B. sphaericus Ts-1 spore-crystal complex migrated in position corresponding to 42kD and 43kD. Bioassay of the two purified proteins prepared by PAGE indicated that they were all toxic to mosquito larvae. Toxic protein was further purified by DEAE-cellulose chromatography. The toxic protein with a molecular weight of 42kD was obtained.     

Laboratory evaluation of entomopathogenic fungi Beauveria bassiana and Metarhizium anisopliae against puparia and adults of Ceratitis capitata (Diptera: Tephritidae)

Laboratory experiments were done to measure the pathogenicity of 10 autochthonous isolates of Beauveria bassiana (Balsamo) Vuill. and of five Metarhizium anisopliae (Metsch.) Sorok. toward puparia and adults of the Mediterranean fruit fly, Ceratitis capitata (Wiedemann) (Diptera: Tephritidae). Although all isolates applied via inoculation of the fungal suspensions on the ventral surface of the abdomen were pathogenic to adults, with mortality rates ranging from 30 to 100% and average survival times (ASTs) from 6.5 to 8.6 d, when C. capitata puparia were immersed in the conidial suspensions, only B. bassiana Bb-1333 and EABb 01/103-Su and M. anisopliae EAMa 01/58-Su isolates caused >50% mortality of puparia. In a second series of bioassays conducted on five selected isolates, adults were sprayed with four 10-fold concentrations ranging from 1.0 x 10(5) to 1.0 x 10(8) colony-forming units (cfu)/ml. The median lethal concentrations (LC50) of the four most virulent isolates ranged from 4.9 x 10(5) to 2.0 x 10(6) cfu/ml with estimated time to kill 50% of the insects ranging from 4.6 to 5.3 d. The effect of a sublethal dose (ca. LD50) of either B. bassiana EABb 01/103-Su or M. anisopliae EAMa 01/58-Su isolate was studied by reciprocal crossing. Treatment with B. bassiana reduced fecundity and fertility at 6, 8, and 10 d after treatment, with fecundity and fertility reductions ranging from 20.0 to 71.2% and from 33.6 to 60.0%, respectively. These reductions occurred in pairing combinations of treated females with either treated or nontreated males. M. anisopliae was more effective in reducing fecundity and fertility at 6 d after treatment, with the reduction varying from 58.4 to 72.1% and from 28.6 to 45.9%, respectively. In addition, the first oviposition was significantly delayed for 1 d in females treated by either fungal species. The above-mentioned five selected isolates were assayed against C. capitata puparia treated as late third instars in sterilized soil at 25'C under three moisture conditions (-0.1, -0.01, and -0.0055 MPa). At -0.01 MPa, all isolates were low pathogenic to C. capitata puparia, whereas significant differences in the puparia mortality occurred between isolates at -0.1 and -0.0055 MPa. The highest pupal mortalities ranged from 52.5 to 70.0%, as a function of soil moisture and were caused by EAMa 01/58-Su and Bb-1333 isolates.     

Comparative analysis of the production of insecticidal and melanizing macromolecules by strains of Beauveria spp.: in vivo studies

Eleven strains of Beauveria bassiana, and a further five species of Beauveria sp., were tested by injection of 8x10(2) conidia into the haemocoel of the larvae of the lepidopteran Galleria mellonella with the aim of analysing their toxin producing activity in vivo. Although the virulent strains killed 100% of the insects at slightly different rates (4-6 days) there were significant differences in the pattern and intensity of host melanization caused by isolates. The majority of the isolates of Beauveria spp. induced a fast and intense melanization of the cuticle of the integument and of tracheal wall, which followed one of three patterns. Another small group of two B. bassiana strains, isolated from Ostrinia nubilalis, induced very weak or no melanization. Strains 618 and 101 of B. bassiana, were selected as models of "melanizing" and "non-melanizing" strains, respectively. Ultrastructural alterations of cells of hypodermal and tracheal epithelium and of haemocytes, assumed to be at least partially caused by fungal toxins, were revealed in larvae infected by both isolates. However, their effects on the fine structure of the hypodermis were different. Injection of sera obtained from haemolymph of insects infected with B. bassiana 618 showed that they have insecticidal, melanizing, and cytotoxic effects similar to those occurring during mycosis. Chromatographic studies and bioassays with fractions prepared from crude serum have allowed a partial identification of the toxic molecules secreted by the fungus in vivo. They are proteinaceous, as shown by protease treatments, thermolabile, negatively charged, and not glycosylated with alpha-d-mannose or alpha-d-glucose. If strain B. bassiana 618 produces melanizing macromolecules which are vivotoxins secreted during the mycosis, the mode of action of isolate 101 is different. Its capacity to kill the host depends on active mycelial development, and on the production of low molecular weight toxins.     

Simple and rapid screening of entomopathogenic fungi having radical-scavenging and anticancer activities

Entomopathogenic fungal metabolites exhibit a wide variety of insecticidal, antimicrobial, anticancer, antioxidant, and antiviral activities, and they have been suggested as potential candidates for the development of new bioactive agents. The rapid and simple screening of entomopathogenic fungi with such bioactivities is important for investigating bioactive substances. The radical-scavenging and anticancer activities of 28 species and 20 genera of 342 isolates of entomopathogenic fungi were evaluated using only culture filtrates. As a result, 23 isolates of Metarhizium anisopliae var. anisopliae, 1 isolate of Pochonia sp., and 1 isolate of Aspergillus sp. showed high radical-scavenging activity. Additionally, anticancer activity in HCT116 human colon cancer cells was also confirmed for selected culture filtrates. This simple screening method of entomopathogenic fungi using culture filtrates may be useful for the rapid detection of related substances with pharmacological activity from a large number of fungal samples.     

K24 T. gondii isolate is a hybrid and has the virulence of lineage I isolates

A permanently high virulence was found in tachyzoites of T. gondii K24 after serial passage in mice (90 passages during 324 days). Virulence tests revealed that a single tachyzoite of the 50th passage represented LD100 for mice. Analysis of genotype of K24 isolate was done by PCR/RFLP with ROP1/Ddel, SAG1/Ddel, 850/Rsal and IGS/Rsal and by RFLP/DNA with TGR1E sequence and Pstl enzyme. K24 isolate had an atypical genotype, with an association of type II (for ROP1, SAG1 genes and TGR1E sequence) and type I (for 850 gene) alleles, and a new pattern observed for IGS. All tested PCR/RFLP did not change through 2, 10, 20, 28, 40, 50, 60, 70, 81 and 90 tested passages. In RFLP/DNA with Pstl enzyme and TGR1E probe, K24 isolate produced a pattern with seven fragments of the size ranging from one to 23 kb and did not change through 7, 56, 70 and 83 tested passages. K24 T. gondii isolate is a hybrid and has the virulence of lineage I isolates.     

The potential for enhanced fungicide resistance in Beauveria bassiana through strain discovery and artificial selection

Our objectives were to determine the (1) natural variation in fungicide resistance among Beauveria bassiana strains, (2) potential to increase fungicide resistance in B. bassiana through artificial selection, and (3) stability of virulence in selected B. bassiana strains. Fungicides included dodine, fenbuconazole, and triphenyltin hydroxide, which are commonly used in pecan and other horticultural crops. Comparison of seven B. bassiana strains indicated some are substantially more resistant to fungicides than others; a commercial strain (GHA) was less resistant than all wild strains isolated from pecan orchards. Artificial selection resulted in enhanced fungicide resistance in the GHA strain but not in a mixed wild strain. Removal of selection pressure for three passages did not reduce the enhanced fungicide resistance. Sub-culturing with exposure to fungicides did not affect the GHA strain's virulence to pecan weevil, Curculio caryae, larvae, whereas fungicide exposure increased virulence in a mixed wild population of B. bassiana.     

球孢白僵菌胞外壳聚糖酶的纯化和性质*

球孢白僵菌Beauveria bassiana 1316-V1的培养上清液经硫酸铵分级沉淀,Sephadex G-75凝胶过滤,Chitosan-bead亲和层析,第二次Sephadex G-75凝胶过滤, 得到电泳纯的一种胞外壳聚糖酶,比活力达到45u/mg 。此酶的分子量为36 kD; 最适酶反应温度为60℃;最适pH为4.0;最适离子强度为 0.25mol/L NaCl; 37℃以下,pH 2.0~5.0之间稳定性好; Cu2+、Hg2+、Pb2+、Ni2+ 对该酶有强烈抑制作用;Ag+、Mn2+也有较强抑制作用;Fe2+有轻微激活作用。该壳聚糖酶是一种糖蛋白,含糖约为12.6%。酶的最适底物为脱乙酰度为90%的胶体壳聚糖;也能轻微水解CMC、DEAE-Cellulose和胶体几丁质;但不能水解片状的壳聚糖和几丁质。     

Production in vitro of toxic macromolecules by strains of Beauveria bassiana, and purification of a chitosanase-like protein secreted by a melanizing isolate

The production of macromolecular insecticidal toxins in Adamek's medium by two selected strains of Beauveria bassiana was investigated. Filtrates and dialysates of the melanizing strain 618 were toxic when injected into the lepidopteran insect Galleria mellonella. Separation by DEAE chromatography revealed that peaks eluted respectively with 100 and 200 mM NaCl (P 100 and P 200) had an insecticidal activity and induced cuticular blackening. A hydrophilic protein, Bclp, which causes the formation of brownish spots of the integument, was purified from P 200 by means of chromatographic and electrophoretic methods. Bclp exhibited clear sequence homologies with fungal chitosanases of Fusarium solani. It has a molecular mass of 28 kDa, a pHI of 4 and is thermolabile. Injection of Bclp causes the same cytoxic effects and alterations of the cuticule as those observed during mycosis, and may contribute to the virulence of strain 618. Comparatively, the most obvious characteristic of the weakly melanizing strain 101 is the lack of significant toxic activity of its P 200, which does not contain Bclp. However, this strain secretes other insecticidal molecules active on lepidopterans, presently unidentified.     

In Vitro Antibacterial Activity of Minocycline and Effect of Agar Medium Utilized in Its Susceptibility Testing

The in vitro activity of minocycline against 1,028 bacterial strains was determined in parallel in Mueller Hinton Agar and Trypticase Soy Agar. The broad antibacterial effect of minocycline against gram-positive cocci and gram-negative bacilli is confirmed. Minimal inhibitory concentrations for gram-positive bacteria in Mueller Hinton Agar were at least twofold less than in Trypticase Soy Agar. Minimal inhibitory concentrations for gram-negative bacilli in Mueller Hinton Agar were usually fourfold less than in Trypticase Soy Agar.     

Effects of virulence,sporulation, and temperature on Metarhizium anisopliae and Beauveria bassiana laboratory transmission in Coptotermes formosanus

Sporulation characteristics and virulence of Metarhizium anisopliae and Beauveria bassiana were examined in relation to laboratory transmission in Coptotermes formosanus. Fungal isolates significantly affected disease prevalence in termite populations. Sporulation of M. anisopliae played a more important role than virulence in producing epizootics within small groups of termites, but this was not the case for B. bassiana. Isolates characterized by quick sporulation (day 2 after death) did not exhibit better transmission in termites than those with high total sporulation (day 11 after death) in either fungal species. An isolate of M. anisopliae ranking highly in all three categories (virulence, quick sporulation, and total sporulation) produced better epizootics than an isolate that was inferior in all three characteristics. High temperatures (35 degrees C) significantly reduced fungal germination rates, leading to significant reduction of epizootics. M. anisopliae was better than B. bassiana in producing epizootics at 27 degrees C. Thus, fungal characteristics other than virulence should be considered for the seasonal colonization approach to termite microbial control.     

Preliminary characterization of glycogen phosphorylase activating hormone and adipokinetic hormone from Manduca sexta corpora cardiaca

ABSTRACT. An attempt was made to separate glycogen phosphorylase activating hormone (GPAH) and adipokinetic hormone (AKH) from the corpora cardiaca (CC) of the moth Manduca sexta (Lepidoptera: Sphingidae) by separating extracts of CC on various chromotographic media, but it was not possible to conclude whether GPAH and AKH are activities of one or of two different peptides. Both activities elute together from glass beads, from Sephadex G-25 and from Sephadex LH-20 columns. In the separation experiments with glass beads and G-25 the activities eluted as a single peak, but using LH-20 we found two peaks exhibiting both activities. The major peak eluted at 1.25 × V_t, which is very similar to locust AKH, while the smaller second peak eluted at O.74 × V _t. Cross injections of CC extracts from M. sexta into Locusta migratoria and CC extracts from L. migratoria into M. sexta suggest that GPAH and the AKH from M. sexta are not identical with the decapeptide AKH from locusts.     

Trypanosoma cruzi: infectivity modulation of a clone after passages through different hosts

Although Trypanosoma cruzi virulence can be modified through passages in vivo or long-term in vitro culture, the mechanisms involved are poorly understood. Here we report modifications in the infectivity of a T. cruzi clone after passages in different hosts without detectable changes in parasite genetic patterns. A clone was obtained from a T. cruzi IIe isolate and showed to be less virulent than the original isolate (p<0.05). This clone was enzymatically similar to the original isolate as shown by multilocus enzyme electrophoresis. Infection of this clone was compared by successive passages in mice and guinea pigs. The mouse-passaged subline became more virulent for both host species compared to the guinea pig-passaged subline (p<0.05). The clone line displayed similar random amplified polymorphic DNA patterns before and after passages in different hosts suggesting that alterations in virulence could be a result of a differential expression of virulence factors.     

Effect of temperature on virulence of Beauveria bassiana and Metarhizium anisopliae isolates to Tetranychus evansi

The virulence of three isolates of Beauveria bassiana (Bals.) Vuill. and 23 isolates of Metarhizium anisopliae (Metschnik.) Sorok. (Ascomycota: Hypocreales) against the tomato spider mite, Tetranychus evansi Baker and Pritchard (Acari: Tetranychidae), was assessed in the laboratory. The effect of temperature on germination, radial growth and virulence of selected isolates (two isolates of B. bassiana and nine of M. anisopliae) on T. evansi was also investigated in the laboratory. All the fungal isolates tested were pathogenic to the adult females of T. evansi, and there were significant differences in mortality between fungal isolates. The lethal time to 50% mortality (LT(50)) values ranged from 4.2 to 8.1 days and the LT(90) values from 5.6 to 15.1 days. Temperature had significant effects on germination, radial growth and virulence of the various isolates. The best fungal germination was observed at 25 and 30 degrees C, while for the fungal radial growth it was 30 degrees C. All the isolates germinated and grew at all temperatures, but germination and radial growth varied with isolate and temperature. The selected isolates were all virulent to T. evansi, but virulence varied also with isolate and temperature.     

In Vitro Stability of Nitrate Reductase from Wheat Leaves: III. Isolation and Partial Characterization of a Nitrate Reductase-inactivating Factor

A nitrate reductase (EC 1.6.6.1)-inactivating factor has been isolated from 8-day-old wheat leaves. The purification schedule involved ammonium sulfate precipitation, Sephadex G-100 filtration, DEAE-cellulose chromatography, and Sephadex G-150 filtration. No accurate assessment could be made as to the degree of purification relative to crude extract as the inactivating factor could not be detected in crude extract. However a 2,446-fold purification was achieved from the ammonium sulfate fraction to the pooled enzyme from the Sephadex G-150 step.     

Evaluation of Metarhizium anisopliae (Metsch) Sorok. to target larvae and adults of Capnodis tenebrionis (L.) (Coleoptera: Buprestidae) in soil and fiber band applications

The aim of this work has been to evaluate in the laboratory the potential of entomopathogenic fungi against adults and larvae of Capnodis tenebrionis (L.) (Coleoptera: Buprestidae) through fiber band application and a potted plant bioassay with soil application, respectively. Our previous findings revealed that Metarhizium anisopliae EAMa 01/58-Su isolate was the most virulent against neonate larvae of the buprestid. In the present work, M. anisopliae EAMa 01/58-Su isolate has been also shown to be highly virulent against adult beetles by immersion in a conidial suspension; thus it was selected to accomplish our objectives. When adult beetles were stimulated to climb 100 x 200 mm non-woven commercial fiber bands impregnated with conidia of M. anisopliae EAMa 01/58-Su isolate, total mortality rates varied from 85.7% to 100.0%; whereas no significant correlation was detected between the time needed to cross the band (mean value 648.7+/-22.4s) and the time of death, with mean average survival time ranging between 10.3 and 16.0 days, compared to 28 days of the controls. Potted seedlings (5-6 months old) of cherry plum (Prunus myrobalana Lois.), a commonly used apricot rootstock, were used to study the efficacy of soil treatment with M. anisopliae EAMa 01/58-Su isolate against neonate C. tenebrionis larvae. The soil inoculation with M. anisopliae EAMa 01/58-Su isolate had a significant effect on the mean number of dead larvae recovered from the roots, with mean mortality ranging from 83.3% to 91.6%; whereas no significant differences were detected between the three fungal doses. In all cases, dead larvae found within roots exhibited external signs of fungal growth. Hence, it may be possible to use M. anisopliae EAMa 01/58-Su isolate in a biocontrol strategy targeting both adults and larvae of C. tenebrionis.