Validation of the methylene blue test for assessment of viability of protoscolices from hydatid cysts

Background: Methylene blue was used as a vital stain for assessment of viability of protoscolices from hydatid cysts taking advantage of the chemical nature of the dye as a redox indicator and the kinetically distinct molecular transfer systems of Echinococcus protoscolex for uptake of materials across the tegument. Aim: The present study attempts to validate the application of methylene blue staining for assessment of viability of protoscolices. Methods: To validate the criteria by which viability is assessed, control tests were performed using normal protoscolices and protoscolices previously treated with distilled water at 60 °C for 5 min. Performance of methylene blue was further studied at intervals over a period of 50 min after protoscolex exposure using 1% dye concentration. Results: Normal protoscolices were able to adsorb and reduce the dye and have therefore lost the blue color. Protoscolices previously treated with warm water on the other hand, being functionally dead, failed to reduce the adsorbed dye and permanently retained the blue color. Results also indicated that a clear distinction between dead and alive protoscolices can be made within one min. Reading of the test after 10 min would be misleading giving a false result. Conclusion: These findings suggest that viability of protoscolices can be assessed on the basis of acquisition and loss/retaining of the dye blue color. Increasing the concentration of methylene blue to 1% was noticed to be associated with remarkable enhancement of contractility, sucker movement and evagination. Such an excitatory action of the dye may be exploited in viability tests which adopt these criteria

Ação do fungo Beauveria bassiana, isolado 986, sobre o ciclo biológico do cascudinho Alphitobius diaperinus em laboratório Action of the fungus Beauveria bassiana, strain 986, over the biologic cycle of the Alphitobius diaperinus beetle in laboratory

O presente trabalho foi realizado para verificar a eficácia do fungo Beauveria bassiana em todas as etapas de desenvolvimento de Alphitobius diaperinus e assim saber em que fase ele é mais susceptível ao controle biológico. O fungo B. bassiana, isolado 986, foi testado em duas concentrações (3,4 X 10(6) e 3,4 X 10(8) conídios ml-1) mais um grupo controle. Foram realizadas duas leituras, uma sete e outra quatorze dias após a aplicação dos fungos. Na concentração 3,4 X 10(6), 54% dos ovos tratados estavam inférteis, ocorrendo 54% de mortalidade de larvas tratadas nos estádios I, II e III, 22,5% nos estádios IV, V e VI, 9,5% nos estádios VII e VIII e 24,5% de mortalidade das pupas. Na concentração 3,4 X 10(8), 66,8% dos ovos tratados estavam inviáveis, ocorrendo mortalidade de 56% das larvas tratadas nos estádios I, II e III, 34% nos estádios IV, V e VI, 24,5% nos estádios VII e VIII e 49,5% das pupas. Já no grupo controle, 13,3% dos ovos estavam inviáveis, ocorrendo mortalidade de 10% das larvas nos estádios I, II e III, 4% nos estádios IV, V e VI e 0% de mortalidade nos estádios VII, VIII e em pupas. Sobre o cascudinho adulto, o fungo não teve efeito nocivo em nenhum grupo analisado. Com base nesses resultados, observou-se que o fungo nas concentrações testadas apresentou efeito nocivo às fases do ciclo biológico do cascudinho, com exceção do inseto adulto.<br>The present research was carried out to verify the Beauveria bassiana fungus efficacy in all development phases of Alphitobius diaperinus and to know in which phase it is more susceptible for biologic control. The B. bassiana, 986-isolated fungus, was tested in two different concentrations (3.4 x 10(6) and 3.4 x 10(8) conidium ml-1) and a control group. Two readings were realized, one after 7 days of fungus application and the other after 14 days. At concentration of 3.4 x 10(6) conidium ml-1, 54% of treated eggs were infertile, with larva death of 54% at I, II and III phase, 22.5% at IV, V, and IV phase and 9.5% at VII and VIII phase. For pupa stage 24.5% of death was found. At concentration of 3.4 x 10(8) conidium ml-1, 66.8% of treated eggs were impracticable occurring treated larva death of 56% at I, II and III phase; 34% at IV, V and VI phase, 24.5% at VII and VIII phase and 49.5% at pupa phase. For control group, 13.3% of eggs were not viable with larva death of 10% at I, II and III phase, 4% at IV, V and VI phase and 0% at pupa, VII and VIII phase. About the adult lesser mealworms, the fungus did not present noxious effect at all analyzed group. In conclusion, these results showed that the fungus at tested concentrations presented noxious effect at all biologic cycle phases of lesser mealworms, except for the adult insect

Control of the Alphitobius diaperinus (Panzer) (Coleoptera: Tenebrionidae) with entomopathogenic fungi

The beetle Alphitobius diaperinus (Panzer), considered a worldwide pest in the poultry industry, is difficult to control and it is a vector for pathogens. The objective of this study was to evaluate the biological control of the lesser mealworm, by strains of fungi Beauveria bassiana, Cladosporium sp. and Trichoderma sp. Larvae and adults of the A. diaperinus were inoculated with suspensions of conidia in the concentration of 10(7) conídia.mL-1. The B. bassiana isolate caused higher insect mortality as compared to Cladosporium sp. and Trichoderma sp. isolates, with the larvae being more susceptible than adults. The entomopathogenicity of B. bassiana was further evaluated with 200 larvae and 200 adults of A. diaperinus inoculated with suspensions 10(6), 10(7), and 10(8) conidia.mL-1, and observed for ten days. Larvae mortality started at the fourth day at the lowest concentration, and the adult mortality was only observed on the sixth day at the concentration of 10(8) conidia.mL-1