Immobilization of β-glucosidase from different sources on nylon tube

Cellulase from four different fungi and β-glucosidase from almonds were immobilized on the inner surface of nylon tubing. The highest values of β-glucosidase activity retention on the support were obtained when P. funiculosum and N. crassa were used as the enzyme source. A comparative study of the thermal stability referring to β-glucosidase activity was developed using free and immobilized enzymes. The most stable β-glucosidases (from P. funiculosum and A. niger) did not show an appreciable change in its thermal stability after immobilization. An important increase in thermal stability was observed when less stable β-glucosidases (from T. reesei, N. crassa and almonds) were immobilized.     

Fumigant activity of essential oils and their components from Eucalyptus codonocarpa and E. dives against Tetranychus urticae (Acari: Tetranychidae) at three temperatures

Essential oils derived from eighteen species of the Myrtaceae family native to Australia, and major constituents of two oils selected from these oils were tested for their fumigant activity against adult females and eggs of Tetranychus urticae Koch (Acari: Tetranychidae) at 5, 15 and 25°C. Essential oils of Eucalyptus codonocarpa and Eucalyptus dives showed the highest fumigant activity against female adults at 10 μl/l at 15 and 25°C. Among major constituents of the two essential oils, piperitone was the most effective against female adults, followed by terpinene‐4‐ol at 10 μl/l at all three temperatures. The two essential oils and these two constituents lowered egg hatchability at 10 μl/l at 25°C. Our results suggest that piperitone should be further investigated as a potential fumigant against T. urticae.     

Insecticidal activity of Citrus aurantium peel extract against Bactrocera oleae and Ceratitis capitata adults (Diptera: Tephritidae)

Citrus aurantium (L.) peel extracts in petroleum ether were evaluated for toxicity against olive fruit fly, Bactrocera oleae (Gmelin) and medfly, Ceratitis capitata (Wiedemann) adults. Bactrocera oleae flies were more susceptible to the extract than C. capitata in contact and residual bioassays. Fumigation bioassay had no effect. Both sexes of B. oleae were equally susceptible in both types of bioassays. However, males of C. capitata were more susceptible than the conspecific females. LD₅₀ values (concentration causing 50% mortality) after 96 h for the males and females of B. oleae were 44.8 and 40.1 μg/insect in contact bioassay through topical application. Whereas, LD₅₀ values for the males and females of C. capitata were 38.8 and 67.8 μg/insect respectively. LC₅₀ values after 96 h for the males and females of B. oleae were 18.8 and 17.8 μg/cm² in Petri dish residual bioassay. Whereas, LC₅₀ values for the males and females of C. capitata were 70.6 and 147.1 μg/cm² respectively.
Fractionation of the extract on a silica gel column with three different polarity solvents resulted in three fractions with only the intermediate polarity solvent fraction having substantial insecticidal activity. Toxicity of the mixtures of active and inactive fractions was equal to the original extract. Our results indicate that C. aurantium has potential for controlling insect pests.     

Lethal activity of beauvericin,a Beauveria bassiana mycotoxin,against the two‐spotted spider mites,Tetranychus urticae Koch

The entomopathogenic fungi Beauveria bassiana (Balsamo) Vuillemin is known to produce a broad range of secondary metabolites. Beauvericin, a cyclic hexadepsipeptide, is the best known mycotoxin produced by B. bassiana; however, reports discussing the insecticidal activity of beauvericin per se are limited. In this study, we assessed the lethal activity of beauvericin against Tetranychus urticae Koch (Acarina: Tetranychidae). In addition, screening for suitable application of the mycotoxin against T. urticae on greenhouse strawberries is discussed. Beauvericin was able to control successfully T. urticae where concentrations of 10, 100 and 1,000 µg/g recorded mortalities of 84%, 100% and 100%, respectively, against motile stages. Furthermore, beauvericin inhibited egg hatching up to 83.3%, 69.3% and 53.3%, respectively, using the same concentrations under laboratory conditions. Under greenhouse conditions, the efficacy recorded was 52.6%, 85.7%, 72.4% and 72.4% at 1, 3, 7 and 10 days post‐inoculation, respectively. Beauvericin was efficacious under greenhouse conditions since the application increased strawberry yields while showing no phytotoxicity and ecotoxicological risk. Resistance to beauvericin was not detected initially at the unselected strain of T. urticae. Yet, the laboratory selection of populations of T. urticae exposed to beauvericin resulted in relatively resistant T. urticae strain that displayed no cross‐resistance to cyflumetofen and bifenazate. The acaricidal activity of beauvericin documented in this study would increase the efficacy of integrated pest management strategies.     

Natural occurrence of soil-borne entomopathogenic fungi in the Moroccan Endemic forest of <Emphasis Type="Italic">Argania spinosa</Emphasis> and their pathogenicity to <Emphasis Type="Italic">Ceratitis capitata</Emphasis>

The occurrence and abundance of entomopathogenic fungi were analysed in 203 soil samples of the Moroccan endemic forests of Argania spinosa, the world main refuge of the Mediterranean fruit fly, Ceratitis capitata. Using the Galleria baiting method and selective media, entomopathogenic fungi were isolated from 186 of the 203 (91.62%) soil samples, with only three species found: Beauveria bassiana (Balsamo) Vuillemin, Metarhizium anisopliae (Metschnikoff) Sorokin and Paecilomyces lilacinus (Thom.) Samson. B. bassiana was the most widespread entomopathogenic fungi (90.64%) in the Argan forest whereas M. anisopliae was less common (15.27%) and P. lilacinus was very rare (1.48%). This is the first report of natural occurrence of M. anisopliae and P. lilacinus in Morocco. Furthermore, 118 Moroccan B. bassiana isolates were studied for their pathogenicity to C. capitata and thermotolerance. Most of these autochtonous B. bassiana isolates were virulent (86.44%) to Medfly pupae and tolerant (55.08%) to temperature stress at 45°C for 2 h. Only 60.17% of Moroccan B. bassiana isolates might be considered as highly entomopathogenic and will serve as a source of potential biological control agents to C. capitata. The percentage of thermotolerant and pathogenic B. bassiana to C. capitata were shown to decrease significantly at winter time characterized by low temperatures and absence of any noticeable medfly in the Argan forest. The occurrence, thermotolerance and virulence of B. bassiana isolates to C. capitata seemed to be related to the sampling periods and location. Our data are discussed with respect to fungal ecology and biocontrol potential of B. bassiana isolates in relation to their habitat.     

Susceptibility of Ceratitis capitata Wiedemann (Diptera: Tephritidae) to Entomopathogenic Fungi and Their Extracts

The effectiveness of seven strains of entomopathogenic fungi against Ceratitis capitata adults was evaluated in the laboratory. Adults were susceptible to five of seven aqueous suspensions of conidia. Metarhizium anisopliae and strain CG-260 of Paecilomyces fumosoroseus were the most pathogenic fungi, with 10-day LD₅₀ values of 5.1 and 6.1 × 10³ conidia/fly, respectively, when applied topically. Sublethal effects on fecundity and fertility of the fungal-exposed females were also studied. The most effective fungus in reducing fecundity was P. fumosoroseus CECT 2705, with reductions on the order of 65% at 1 × 10⁶ conidia/fly. M. anisopliae and Aspergillus ochraceus also showed significant reductions of fecundity (40–50% for most of the assayed concentrations). Fertility was moderately affected by the fungi. M. anisopliae at 1 × 10⁶ conidia/fly was the most effective fungus, showing egg eclosion reduction of over 50% compared with the control. In addition, culture broth dichloromethane extracts from the entomopathogenic fungi were tested for insecticide activity against C. capitata, including effects on fecundity and fertility. The extract from M. anisopliae was the most toxic, resulting in about 90% mortality at a concentration of 25 mg/g of diet; under these conditions, fecundity and fertility of treated females were reduced by 94 and 53%, respectively, compared with untreated controls.     

Integrated pest management of two-spotted mite Tetranychus urticae on greenhouse roses using petroleum spray oil and the predatory mite Phytoseiulus persimilis

From 1995 to 1999, four experiments were conducted on greenhouse roses to assess the effectiveness of the nC24 petroleum spray oil (PSO), D-C-Tron Plus, against two-spotted mite, Tetranychus urticae Koch (Acarina: Tetranychidae), and to determine how the oil could be most efficiently and effectively used in combination with the predatory mite Phytoseiulus persimilis Athias-Henriot (Acarina: Phytoseiidae) in an integrated pest management program. The results showed that 0.5% PSO applied fortnightly to roses gave excellent protection from T. urticae infestation when the mite population was not already established. However, PSO applied after roses were infested with T. urticae above the economic threshold only stabilised populations without reducing them below that threshold. Populations of P. persimilis in the upper and lower canopies were unchanged after two sprays of PSO at 7-day intervals, and application of PSO to the upper canopy was as effective in controlling T. urticae in the presence of P. persimilis as spraying the entire plant. Combining PSO with P. persimilis gave better control of T. urticae than using P. persimilis alone. The most cost-effective use of PSO in the presence of P. persimilis is, therefore, to apply spray only to the upper canopy. This will not affect control of powdery mildew with PSO. Comparison of a control program for T. urticae based on the monitored use of synthetic miticides with that based on calendar application of PSO revealed that both gave equally effective control. The benefits of combining PSO and P. persimilis in an integrated pest management program for T. urticae on roses over a program based on synthetic fungicides are discussed.     

Abiotic factors affecting Diachasmimorpha longicaudata (Hymenoptera: Braconidae) activity as a natural enemy of Ceratitis capitata (Diptera: Tephritidae) under semi‐natural conditions in the Mediterranean region

The larval–pupal endoparasitoid Diachasmimorpha longicaudata (Hymenoptera: Braconidae) is currently the most commonly employed biological control agent against Tephritid fruit flies in the Americas. However, this parasitoid remains largely ignored and is not used in many regions, including the Mediterranean Basin. In this study, the potential of D. longicaudata as a biocontrol agent against the Mediterranean fruit fly (medfly) Ceratitis capitata (Diptera: Tephritidae) was addressed in an area of eastern Spain (the Valencian community). The parasitic activity of parasitoids and the effects of climatic conditions were evaluated throughout a 1‐year period in field‐cage experiments in which parasitoids were confined with apples artificially infested with medfly larvae. The following parameters were calculated and related statistically to several environmental conditions: the parasitism rate, the induced mortality and progeny sex ratio. The results show that D. longicaudata is able to parasitize medfly larvae throughout the year under semi‐natural conditions. Important fluctuations in the parasitism rate (from almost zero to 42%) and the induced mortality (from 6% to 80%) were partially influenced by climatic conditions. The parasitism rate increased with mean temperature and decreased with mean relative humidity, while the induced mortality decreased with minimum relative humidity. The optimal climatic conditions for the activity of the parasitoid were a mean temperature of 16–24°C combined with a relative humidity of 45%–60%. Overall, these results suggest that reduction in the medfly population due to D. longicaudata activity is feasible and provide information about the optimal time period for parasitoid release in the field. In conclusion, D. longicaudata has a significant potential to control C. capitata in the Mediterranean region.     

Life history of the predatory mite <Emphasis Type="Italic">Phytoseiulus fragariae</Emphasis> on <Emphasis Type="Italic">Tetranychus evansi</Emphasis> and <Emphasis Type="Italic">Tetranychus urticae</Emphasis> (Acari: Phytoseiidae,Tetranychidae) at five temperatures

Tetranychus evansi Baker and Pritchard and Tetranychus urticae Koch (Acari: Tetranychidae) are important pests of Solanaceae in many countries. Several studies have demonstrated that T. urticae is an acceptable prey to many predatory mites, although the suitability of this prey depends on the host plant. T. evansi, has been shown to be an unfavorable prey to most predatory mites that have been tested against it. The predator Phytoseiulus fragariae Denmark and Schicha (Acari: Phytoseiidae) has been found in association with the two species in Brazil. The objective of this work was to compare biological parameters of P. fragariae on T. evansi and on T. urticae as prey. The study was conducted under laboratory conditions at 10, 15, 20, 25 and 30°C. At all temperatures, survivorship was lower on T. evansi than on T. urticae. No predator reached adulthood at 10°C on the former species; even on the latter species, only about 36% of the predators reached adulthood at 10°C. For both prey, in general, duration of each life stage was shorter, total fecundity was lower and intrinsic rate of population increase (r _m ) was higher with increasing temperatures. The slower rate of development of P. fragariae on T. evansi resulted in a slightly higher thermal requirement (103.9 degree-days) on that prey than on T. urticae (97.1 degree-days). The values of net reproduction rate (R ₀), intrinsic rate of increase (r _m ) and finite rate of increase (λ) were significantly higher on T. urticae, indicating faster population increase of the predator on this prey species. The highest value of r _m of the predator was 0.154 and 0.337 female per female per day on T. evansi and on T. urticae, respectively. The results suggested that P. fragariae cannot be considered a good predator of T. evansi.     

Seed-borne fungi of five cruciferous vegetables and relative efficacy of aqueous seed extracts against some associated fungi

A total of 121 seed samples of cabbage, cauliflower, cress, radish and turnip collected from five localities in Upper Egypt were assayed for their fungal flora. The highest count of fungi was recorded on cabbage seeds (75%), whereas the lowest count was observed on turnip seeds (33%). Thirty-five fungal species and two varieties belonging to 16 genera were identified. The broadest number of species (22 species + 1 variety) were isolated from cress seeds. However, only five species,viz. Aspergillus flavus, A. niger, Penicillium chrysogenum, P. funiculosum andRhizopus stolonifer, were found to be associated with seeds of the five plants. The relative efficacy of aqueous seed extracts against these five fungal species was tested. The antifungal drug Trosyd (tioconazole) was taken as a standard inhibitor. The aqueous extract of cabbage seeds inhibited three fungal species, cress extract two and cauliflower and turnip one species; radish seed extract was ineffective.     

Preliminary evaluation of <Emphasis Type="Italic">Cladosporium cladosporioides</Emphasis> (Fresen.) de Vries in laboratory conditions,as a potential candidate for biocontrol of <Emphasis Type="Italic">Tetranychus urticae</Emphasis> Koch

A survey for natural entomopathogenic fungi of two spotted spider mite (Tetranychus urticae) adults was made in Erzurum, Turkey, during the period 2006. Tetranychus urticae (65.8%) infected with a strain of the fungus Cladosporium cladosporioides were found. Thirteen isolates of C. cladosporioides were assessed against T. urticae, in a single dose (8 × 10⁶ conidia ml ⁻¹), laboratory bioassay on bean leaflets. The total mortality percentage caused by C. cladosporioides isolates varied from 50.95 to 74.76% and LT₅₀ values ranged from 2.34 to 3.90 days. The results revealed that isolates of C. cladosporioides were effective against two spotted spider mite. This is the first record of natural infection of T. urticae by C. cladosporioides.     

Cellulases from Penicillium funiculosum: production, properties and application to cellulose hydrolysis

The objective of this work is to investigate the utilization of two abundant agricultural residues in Brazil for the production and application of cellulolytic enzymes. Different materials obtained after pretreatment of sugarcane bagasse, as well as pure synthetic substrates, were considered for cellulase production by Penicillium funiculosum. The best results for FPase (354 U L^?1) and β-glucosidase (1,835 U L^?1) production were observed when sugarcane bagasse partially delignified cellulignin (PDC) was used. The crude extract obtained from PDC fermentation was then partially characterized. Optimal temperatures for cellulase action ranged from 52 to 58°C and pH values of around 4.9 contributed to maximum enzyme activity. At 37°C, the cellulases were highly stable, losing less than 15% of their initial activity after 23 h of incubation. There was no detection of proteases in the P. funiculosum extract, but other hydrolases, such as endoxylanases, were identified (147 U L^?1). Finally, when compared to commercial preparations, the cellulolytic complex from P. funiculosum showed more well-balanced amounts of β-glucosidase, endo- and exoglucanase, resulting in the desired performance in the presence of a lignocellulosic material. Cellulases from this filamentous fungus had a higher glucose production rate (470 mg L^?1 h^?1) when incubated with corn cob than with Celluclast^®, GC 220^® and Spezyme^® (312, 454 and 400 mg L^?1 h^?1, respectively).     

Phytochemical,acaricidal and biological activity of Dodonaea viscosa L. against the two spotted spider mite Tetranychus urticae Koch

Alcohol extract, saponins and flavones fractionated from Dodonaea viscosa leaves were tested for their acaricidal activity against the adult females of Tetranychus urticae Koch. It was found that crude alcohol extract was more potent than the isolates obtained. At LC₅₀ and LC₉₀ levels, the crude extract ranked the first. All the biotic factors are significantly affected. The fecundity was seriously reduced however, the hatchability of eggs was not affected. Two flavonoids were identified as Querectin 3–0-glucose 7-0-rhamnoside and the other as Querectin 3-0 arabinose 7-0 rhammoside from the ethyl acetate fraction of the alcohol extract.     

Saccharification of used paper with different cellulases

Various used paper materials have been exposed to the action of cellulases from Penicillium funiculosum, Trichoderma reesei, Trichoderma viride and Aspergillus niger. A 2 h incubation period showed cellulase from T. viride the most active except for office paper that was maximally degraded by A. niger cellulase. Cellulase mixtures increased saccharification while sequential treatment with cellulases from T. reesei and P. funiculosum increased biodegradation at values between 15% and 190%. The maximum increase of saccharification (190%) was obtained when T. reesei cellulase initiated the sequential treatment of newspaper relative to the sole action of P. funiculosum cellulase on this non-pretreated and pretreated material.     

Influence of Temperature on Virulence of Fungal Isolates of <Emphasis Type="Italic">Metarhizium anisopliae</Emphasis> and <Emphasis Type="Italic">Beauveria bassiana</Emphasis> to the Two-Spotted Spider Mite <Emphasis Type="Italic">Tetranychus urticae</Emphasis>

Twenty-three isolates of Metarhizium anisopliae (Metschnikoff) Sokorin and three isolates of Beauveria bassiana (Balsamo) Vuillemin (Ascomycota: Hypocreales: Clavicipitaceae) were assessed for their virulence against the two-spotted spider mite, Tetranychus urticae Koch (Acari: Tetranychidae). Based on the screening results, nine isolates of M. anisopliae and two isolates of B. bassiana were tested for their virulence against young adult (1- to 2-day-old) female T. urticae at constant temperatures of 20, 25, 30 and 35°C. At all temperatures tested, all the fungal isolates were pathogenic to T. urticae but mortality varied with isolates and temperatures. Fungal isolates were more virulent at 25, 30 and 35°C than at 20°C. The lethal time to 50% mortality (LT₅₀) and lethal time to 90% mortality (LT₉₀) values decreased with increased temperature. There were no significant differences in virulence between fungal isolates at 30 and 35°C; however, significant differences were observed at 20 and 25°C.     

Toxicity and ovicidal activity of different plant extracts on two-spotted spider mite,Tetranychus urticae Koch (Acari: Tetranychidae)

The acaricidial, persistent and ovicidal activities of four methanolic extracts of Anisosciadium orientale DC., Scaligeria meifolia Boiss., Trigonella elliptica Boiss. and Ptelea viscosa L. were assessed against the two-spotted spider mite, Tetranychus urticae Koch, in a bioassay under laboratory conditions. All the extracts exhibited significant adult female mite mortality as compared to control and S. meifolia extract showed highest toxicity based on the LC₅₀ when compared with the other extracts tested. In the persistence test, the mites were exposed to 1, 24, 48 and 72 h old residues of 50% lethal concentration of methanolic extracts. The results showed that the lethal concentration effects of plant extracts fade within two or three days. S. meifolia caused high ovicidal activity (45.84%) followed by A. orientale (41.40%), T. elliptica (40.11%) and P. viscosa (37.66%). The results suggest that the methanolic extracts of A. orientale, S. meifolia, T. elliptica and P. viscosa have the potential to be used in mite pest management.     

Investigation of native isolates of entomopathogenic fungi for the biological control of three citrus pests

The pathogenicity of 15 isolates of Beauveria bassiana (Balsamo) Vuillemin, five isolates of Metarhizium anisopliae (Metschnikoff) Sorokin and one isolate of M. flavoviride (Gams and Rozsypal) were tested under laboratory conditions against the subterranean life stages of the citrus pests, Ceratitis rosa Karsch, C. capitata Wiedemann (Diptera: Tephritidae) and Thaumatotibia leucotreta Meyrick. (Lepidoptera: Tortricidae). When these citrus pests were treated with a concentration of 1×10⁷ conidia mL^?1, fungal isolates had a significantly greater effect on the adults of C. rosa and C. capitata than they did on the puparia of these two fruit fly species. Further, C. rosa and C. capitata did not differ significantly in their response to entomopathogenic fungi when adult and pupal mycosis were considered. Depending on fungal isolate, the percentage of T. leucotreta adults which emerged from fungal treated sand ranged from 5 to 60% and the percentage of pupae with visible signs of mycosis ranged from 21 to 93%. The relative virulence of the four most promising fungal isolates, as well as the commercially available B. bassiana product, BroadBand® (Biological Control Products, South Africa), were compared against one another as log-probit regressions of mortality against T. leucotreta which exhibited a dose-dependent response. The estimated LC₅₀ values of the three most virulent B. bassiana isolates ranged from 6.8×10⁵ to 2.1×10⁶ conidia mL^?1, while those of the least pathogenic ranged from 1.6×10⁷ to 3.7×10⁷ conidia mL^?1.     

The ground beetle Pseudophonus rufipes revealed as predator of Ceratitis capitata in citrus orchards

The Mediterranean fruit fly Ceratitis capitata (Wiedemann) is a major citrus pest in Spain. Although medfly is being controlled chemically, alternative biorational strategies should be developed, like those based on conservation biological control of polyphagous predators. The ground beetle Pseudophonus rufipes (De Geer) is the most abundant carabid inhabiting the ground of citrus orchards in eastern Spain. However, little is known about its activity–density and predatory role against C. capitata in the citrus agroecosystem. Here, we report on its predatory potential on the medfly’s developmental stages that take place in the citrus ground. This carabid species preyed efficiently on C. capitata third instar larvae and especially on pupae but not on teneral adults. Moreover, predation under field conditions was evaluated by detecting C. capitata DNA remains using PCR-based gut-content analysis. Half-life DNA detectability of C. capitata was of 32.33 h. Pseudophonus rufipes specimens were field-collected after C. capitata sterilized pupae were deployed in a commercial citrus orchard. Thereafter, the carabids captured by pitfall traps were analyzed, being DNA-remains of C. capitata detected in 22.2% of them. Data reported here clearly suggest that P. rufipes could play an important role in regulating medfly populations in citrus orchards. This information is particularly useful when biological control conservation strategies are being considered to control this pest.     

Predation-related odours reduce oviposition in a herbivorous mite

When adult females of the herbivorous mite, Tetranychus urticae, were exposed to the predatory mite, Phytoseiulus persimilis, they laid fewer eggs than females that had not been exposed to P. persimilis when transferred onto a new leaf patch. However, when T. urticae females were exposed to either products of P. persimilis or artificially damaged conspecific eggs on a leaf patch, the number of T. urticae eggs on a new leaf patch did not differ significantly from the control. The reduced oviposition was neither due to the feeding activity on the leaf patch with P. persimilis nor to that on the new leaf patch. There was also no significant difference between the number of T. urticae eggs produced on a new leaf patch following exposure to the odours of a neighbouring leaf patch where there had previously been either P. persimilis or T. urticae adults. However, female T. urticae that had been exposed to odours from neighbouring leaf patches on which both T. urticae and P. persimilis had been placed produced significantly fewer eggs on a new leaf patch than those that had not been exposed to such odours. Neither odours from neighbouring intact leaf patches on which T. urticae eggs were preyed on by P. persimilis, nor odours from a neighbouring Parafilm patch on which T. urticae was preyed on by P. persimilis affected the oviposition of T. urticae. These data suggest that the presence of T. urticae, P. persimilis and a leaf patch are needed for the emission of odours to reduce oviposition in T. urticae.