Alcides Moino

Compatibility of entomopathogenic fungi with neonicotinoid insecticides

The in vitro fungitoxic effect of the neonicotinoid insecticides acetamiprid (Saurus 200 SP), imidacloprid (Confidor 700 WDGr) and thiamethoxam (Actara 250 WG) in three concentrations (AR= average field recommendation; 0.7 AR and 1.3 AR) to the entomopathogenic fungi Beauveria bassiana, Metarhizium anisopliae and Paecilomyces sp. was studied. The effect of the insecticides on conidia germination, vegetative growth and conidiogenesis was compared. The insecticides had no effect on conidia germination, except under the highest concentration (1.3 AR) of acetamiprid, in which significant inhibition of M. anisopliae occurred. Vegetative growth of B. bassiana and M. anisopliae was significantly inhibited only by the three concentrations of acetamiprid treatment. Thiamethoxam treatment, in the AR and 1.3 AR concentrations, was significantly lower for B. bassiana vegetative growth. Paecilomyces sp. vegetative growth was higher than the control in the following treatments: 0.7 AR and AR of acetamiprid; all concentrations of imidacloprid; and in the two smallest concentrations, 0.7 AR and AR, of thiamethoxam. Conidia production was significantly smaller for Paecilomyces sp. and M. anisopliae, in the highest concentration (1.3 AR) of acetamiprid treatment and, for Paecilomyces sp., in the highest concentration (1.3 AR) of imidacloprid. Thiamethoxam significantly inhibited M. anisopliae conidia production, only in the smallest concentration (0.7 AR).Significant increase in the conidia production of Paecilomyces sp. fungus was observed in the following treatments: two concentrations, 0.7 AR and AR of acetamiprid; and in the two highest concentrations, AR and 1.3 AR, of thiamethoxam treatments. The use of insecticides, in the recommended formulations and other concentrations tested, in most cases, had no negative effect on conidia germination, conidia production and vegetative growth of B. bassiana, M. anisopliae and Paecilomyces sp. Consequently, these products, in the formulations and concentrations tested, may be used in IPM programs in which the entomopathogens are important pest regulators.

Effect of Biofertilizers and Neem Oil on the Entomopathogenic Fungi Beauveria bassiana (Bals.) Vuill. and Metarhizium anisopliae (Metsch.) Sorok

The in vitro fungitoxic effect of three biofertilizers, E.M.-4, Multibion O and Supermagro used in organic agriculture and the neem oil (Azadirachta indica A. Juss) on the entomopathogenic fungi Metarhizium anisopliae and Beauveria bassiana was studied. These products were mixed in a medium where the two fungi were inoculated, and germination, vegetative growth and conidiogenesis were assessed. The biofertilizers Supermagro and E.M.- 4 showed to be less toxic for the two fungi whereas MultibionO caused major inhibition on M. anisopliae, with reductions in germination (-37.74%), colony diameter (-30.26%) and conidiogenesis (-42.62%). Neem oil promoted a larger negative effect on B. bassiana, inhibiting germination (-45.27%), colony diameter (-36.62%) and conidiogenesis (-84.93%).

Efeito de imidacloprid e fipronil sobre Beauveria bassiana (Bals.) Vuill. e Metarhizium anisopliae (Metsch.) Sorok. e no comportamento de limpeza de Heterotermes tenuis (Hagen)

The fungitoxic effects of imidacloprid and fipronil on the entomopathogenic fungi Beauveria bassiana (Bals.) Vuill. and Metarhizium anisopliae (Metsch.) Sorok. were evaluated. The colony radial growth and conidial production were measured in the presence of the insecticides added to the culture medium. Imidacloprid was less toxic to the fungi than fipronil, and M. anisopliae was less affected by either insecticide than B. bassiana. The changes in the grooming behavior of Heterotermes tenuis (Hagen) under the action of the insecticides was also evaluated. Using scanning electron microscopy, conidia of B. bassiana and M. anisopliae were observed on the surface of the insect tegument at different times after inoculation. Imidacloprid at sublethal concentrations affected the grooming behavior of H. tenuis, while fipronil did not alter this behavior.

Compatibilidade de Beauveria bassiana com agrotóxicos visando o controle da cochonilha-da-raiz-do-cafeeiro Dysmicoccus texensis Tinsley (Hemiptera: Pseudococcidae)

Several chemical substances are used to control insects, diseases and weeds, however many of these products are toxic to mankind and the animals, besides reducing the potential of pest control by predators, parasitoids and pathogens. The integrated control using selective chemical pesticides and entomopathogenic fungi is a viable strategy, however some of these products can impact these microorganisms, reducing vegetative growth, viability and sporulation. The objectives of this work were to evaluate the effect of chemical pesticides used in the coffee crop on the entomopatogenic fungus Beauveria bassiana (isolate UEL 114), for the control of the cofee root mealybug. A fungal suspension of 1 x 107 viable conidia/ml was added to solutions of the products at the recommended concentrations. After 1h, conidia were inoculated onto PDA medium, and quantification of germinated conidia assessed after 20h. The vegetative growth and sporulation were appraised eight days after the fungus inoculation onto PDA medium containing the products at recommended concentrations and maintained at the temperature of 25 ± 1oC, 12h photofase and 70 ± 10% relative humidity. The mean diameter of the colonies was measured and conidial production quantified in a Neubauer chamber. Azafenidyne, Quintozene, Symazine + Ametryne, 2,4-D, Acetochlor and Oxyfluorfen affected the conidial germination. Thiamethoxan, Imidacloprid, Carbofuran and Pencycuron were compatible; whereas Glyphosate, Dimetilurea, Azafenidine, Quintozene, Symazine + Ametryne, 2,4-D, Acetochlor and Oxyfluorfen significantly impacted vegetative growth and sporulation of B. bassiana UEL 114.

External development of the entomopathogenic fungi Beauveria bassiana and Metarhizium anisopliae in the subterranean termite Heterotermes tenuis

The subterranean termite Heterotermes tenuis is one of the main pests of sugarcane and eucalyptus in Brazil, and the use of entomopathogenic fungi, alone or associated to chemicals, is an efficient and environmentally favorable method for its control. Studies related to the fungal development on these insects are important due to the effect of insect behavior on entomopathogens. The objective of this work was to describe the external development of Beauveria bassiana and Metarhizium anisopliae on H. tenuis using Scanning Electron Microscopy (SEM), determining the duration of the different phases of fungal infection. Two fixation techniques for preparing SEM samples were also evaluated. Worker specimens of H. tenuis were inoculated with a 1 x 109 conidia mL -1 suspension of the fungi and maintained at 25±1 o C and 70±10% relative humidity. Insects were collected from 0 to 144 hours after inoculation and prepared on SEM stubs for each of the two fixation techniques. The results obtained with the two techniques were compared and duration of the different phases of the infection process were estimated from SEM observations and compared for three fungal isolates. B. bassiana and M. anisopliae have similar development cycles on the termite, but some important differences exist. The penetration, colonization and conidiogenesis phases are relatively faster for M. anisopliae than for B. bassiana, which results in a faster rate of insect mortality. The fixation technique with OsO 4 vapor is suitable for preparation of insects to be used in SEM observation of the developmental stages

Heterorhabditis amazonensis n. sp. (Rhabditida: Heterorhabditidae) from Amazonas, Brazil

Summary – In a survey of entomopathogenic nematodes in Brazil, a nematode isolate of the genus Heterorhabditis was found. The nematode was collected from soil by the insect-baiting technique and maintained in the laboratory on last instar Galleria mellonella (L.) larvae. Morphological and molecular studies of the isolate showed that the nematode is a new species. Light and scanning electron microscopy, DNA characterisation and phylogeny were used for this description. Heterorhabditis amazonensis n. sp. is morphologically similar to H. baujardi, H. floridensis, H. mexicana and H. indica, and can be distinguished from these species mainly by male and female characters. Fifty percent of Heterorhabditis amazonensis n. sp. males have two pairs of bursal papillae in the terminal group; 25% with two papillae on one side and one papilla on the other side and 25% with one pair of papillae. Twenty percent of the population has a curved gubernaculum. The percentage of the gubernaculum to spicule length (GS%) is lower than that of H. mexicana (50 vs 56), and the length of the spicule relative to anal body diam. (SW%) is lower than that of H. mexicana (152 vs 167) and H. baujardi (152 vs 182). The female can be differentiated from related species by its unique vulva pattern. In a phylogenetic tree, the new species, H. floridensis, H. baujardi and H. mexicana form a monophyletic group, a sister group to H. indica. The new species has evolved three autapomorphic nucleotide character states, differing from its sister taxa H. mexicana at 22, and H. baujardi at 15, aligned positions.

Influence of soil temperature and moisture on the infectivity of entomopathogenic nematodes (Rhabditida: Heterorhabditidae, Steinernematidae) against larvae of Ceratitis capitata (Wiedemann) (Diptera: Tephritidae)

The Mediterranean fruit fly, Ceratitis capitata (Wiedemann), is considered one of the main pests that affect fruit production in the world. This insect spends part of its life cycle in the soil, making it a target for entomopathogenic nematodes. This work aimed at evaluating the influence of soil temperature and moisture on the infectivity of Heterorhabditis sp. RSC01 and Steinernema carpocapsae ALL to third-instars of C. capitata, and to compare the efficiency of these isolates at five different soil temperatures (19, 22, 25, 28, and 31°C) and three levels of relative soil moisture (100, 75, and 50% of field capacity). Ten C. capitata larvae were transferred to plastic jars (12 cm × 6 cm) containing 100 g soil, followed by the application of an aqueous suspension containing 125 infective juveniles (IJ)/cm². In the control treatment, 3 ml of distilled water was applied. Mortality evaluations were made five days later and were confirmed by observations of the characteristic symptoms and cadaver dissection. The infectivity was directly proportional to temperature increase, with maximum percent mortality of 86.7% and 80.0% for S. carpocapsae and Heterorhabditis sp., respectively, at 31°C. At 25°C, the highest mortality for both species was obtained at 75% of field capacity (96.7% and 26.7% for S. carpocapsae and Heterorhabditis sp., respectively).

Controle do cupim subterrâneo Heterotermes tenuis (Hagen) com iscas termitrap impregnadas com inseticidas e associadas ao fungo entomopatogênico Beauveria bassiana (Bals.) Vuill.

The control of the termite Heterotermes tenuis (Hagen) (Isoptera: Rhinotermitidae) was evaluated using the bait/trap Termitrap impregnated with insecticides in low concentrations and associated to the entomopathogenic fungus Beauveria bassiana (Bals.) Vuill. isolate 634 (from Solenopsis invicta F. Smith) in sugarcane (Saccharum officinarum L.). The experiment was conducted in Rio Claro-SP, in a sugarcane field highly infested with termites. The treatments consisted of: imidacloprid 0,01%; imidacloprid 0,01% + B. bassiana; WG 0,003%; WG 0,003% + B. bassiana; B. bassiana; and Untreated control. Each treatment was replicated five times. The insecticides were impregnated on baits by immersion in water, being their concentrations calculated according to the weigth of the bait, and the B. bassiana was applied as pure conidia (109 conidia/bait). The research was conducted during November 1996 through April 1997. The evaluations were made after 15, 30, 41, 63, 86 e 136 days by assigning indexes to populations levels. All treatments significantly reduced termite populations when compared to the control. However, it took longer for B. bassiana alone to reduced H. tenuis population. The treatments with imidacloprid and WG were the most efficient in the control of termites in sugarcane. The baits/traps did not repel the termites.

Movement of Heterorhabditis amazonensis and Steinernema arenarium in search of corn fall armyworm larvae in artificial conditions

Spodoptera frugiperda (Smith, 1797) (Lepidoptera: Noctuidae) is considered to be the main pest of maize crops in Brazil. Entomopathogenic nematodes (EPN) may be used to control this pest and exhibit different, unique abilities to search for their hosts. The movement of EPN in relation to S. frugiperda was evaluated. To test for horizontal movement, a styrofoam enclosure filled with sand was divided into segments, nematodes were placed at the entrance to the enclosure and a larva was placed at the end of each division. The same approach was used to evaluate vertical movement; however, PVC pipes were used in this case. In general, the mortality was inversely proportional to the initial distance between host and nematodes. In the vertical displacement test, both nematodes were able to kill the larvae up to a distance of 25 cm. Therefore, the infective juveniles of H. amazonensis and S. arenarium can search out, infect and kill larvae of S. frugiperda at distances of up to 60 cm and 25 cm of horizontal and vertical displacement, respectively.

ANTAGONISTIC EFFECT OF Trichoderma sp. ON THE DEVELOPMENT OF Beauveria bassiana (Bals.) Vuill. AND Metarhizium anisopliae (Metsch.) SOROK

The objective of this work was to evaluate the effect of Trichoderma sp. on the development of Beauveria bassiana and Metarhizium anisopliae. The fungus Trichoderma sp. was inoculated on PDA culture medium, 0, 48, 120 and 168 hours after inoculation of the same plates with either B. bassiana (isolate 634) or M. anisopliae (isolate E-9). The radial growth of fungal colonies was measured 48 and 120 hours after Trichoderma sp. inoculation. Trichoderma sp. affected the development of both entomopathogenic fungi when inoculated simultaneously or 48 hours later. B. bassiana and M. anisopliae had normal development when inoculated 168 hours before Trichoderma sp. The effect of a toxic extract from Trichoderma sp. on the entomopathogenic fungi was also evaluated. Aliquots of 0.1, 0.5, 1.0 and 5.0 ml of extract were added to 100 ml of PDA medium, used to grow the entomopathogens. The radial growth of B. bassiana and M. anisopliae colonies grown in the presence of the extract were measured and the number of conidia produced per colony was counted. With 5% of extract in the medium, mycelial growth and conidiogenesis of B. bassiana decreased. M. anisopliae growth was affected by culture medium containing 1% or more of the extract.