Dual Biocontrol Potential of the Entomopathogenic Fungus, Isaria javanica, for Both Aphids and Plant Fungal Pathogens

Dual biocontrol of both insects and plant pathogens has been reported for certain fungal entomopathogens, including Beauveria bassiana and Lecanicillum spp. In this study, we demonstrate, for the first time, the dual biocontrol potential of two fungal isolates identified by morphological and phylogenetic analyses as Isaria javanica. Both these isolates caused mortality in the greater wax moth, and hence can be considered entomopathogens. Spores of the isolates were also pathogenic to nymphs of the green peach aphid (Myzus persicae), with an LC₅₀ value of 10⁷ spores/mL 4 days after inoculation and an LT₅₀ of 4.2 days with a dose of 10⁸ spores/mL. In vitro antifungal assays also demonstrated a strong inhibitory effect on the growth of two fungi that are pathogenic to peppers, Colletotrichum gloeosporioides and Phytophthora capsici. These results indicate that I. javanica isolates could be used as novel biocontrol agents for the simultaneous control of aphids and fungal diseases, such as anthracnose and Phytophthora blight, in an integrated pest management framework for red pepper.

Cymbopogon citratus (Poaceae) essential oil on Frankliniella schultzei (Thysanoptera: Thripidae) and Myzus persicae (Hemiptera: Aphididae) / Óleo essencial de Cymbopogon citratus (Poaceae) Sobre Frankliniella schultzei (Thysanoptera: Thripidae) e Myzus persicae (Hemiptera: Aphididae)

Phytovirus vectors Frankliniella schultzei (Trybom, 1920) (Thysanoptera: Thripidae) and Myzus persicae (Sulzer, 1776) (Hemiptera: Aphididae) feed on crops of great economic importance brings large economic losses worldwide to cultivated species such as tomato and cotton. F. schultzei transmits Tospovirus, Groundnut ring spot virus (GRSV) and Tomato spotted wilt virus (TSWV) to tomato plants and M. persicae transmits Potato virus Y (PVY), Tomato yellow top virus (ToYTV) and Tomato bottom yellow leaf virus (TBYLV) to tomato crops. Chemical constituents of essential oils have been increasingly studied because they present a wide range of biological activities. The aim of this work was to characterize Cymbopogon citratus essential oil and evaluate its potential insecticide activity against F. schultzei and M. persicae. The essential oil was obtained from fresh leaves by hydrodistillation using a Clevenger apparatus. Its yield (1.04%) was determined relative to the dry mass of the plant. Qualitative analysis was performed by gas chromatography coupled to mass spectrometry and chemical constituent content was determined by gas chromatography with a flame ionization detector. Nine compounds were identified, with geranial (49.98%) and neral (37.78%) being the major components. The insects were sprayed with C. citratus essential oil at different concentrations using a Potter tower. The LC50 values for M. persicae and F. schultzei were 0.28% and 1.49%, respectively. Essential oil from C. citratus is a promising natural alternative for developing pesticides to manage M. persicae.

Os vetores de fitovírus Frankliniella schultzei (Trybom, 1920) (Thysanoptera: Thripidae) e Myzus persicae (Sulzer, 1776) (Hemiptera: Aphididae) se alimentam de culturas de grande importância econômica, trazendo grandes perdas econômicas em todo o mundo para as espécies cultivadas, como tomate e algodão. F. schultzei transmite Tospovirus, Groundnut ring spot virus (GRSV) e Tomato spotted wilt virus (TSWV) em tomateiro and M. persicae transmite Potato virus Y (PVY), Tomato yellow top virus (ToYTV) e Tomato bottom yellow leaf virus (TBYLV) ao tomateiro. Os constituintes químicos dos óleos essenciais têm sido cada vez mais estudados, pois apresentam uma ampla gama de atividades biológicas. O objetivo deste trabalho foi caracterizar o óleo essencial de Cymbopogon citratus e avaliar o seu potencial inseticida sobre F. schultzei e M. persicae. O óleo essencial foi obtido a partir de folhas frescas por hidrodestilação utilizando um aparelho Clevenger. O seu rendimento (1,04%) foi determinado em relação à massa seca da planta. A análise qualitativa foi realizada por cromatografia gasosa acoplada a espectrometria de massa e o teor dos constituintes químicos foi determinado por cromatografia gasosa com detector de ionização de chama. Nove compostos foram identificados, com geranial (49,98%) e neral (37,78%), sendo os componentes principais. Os insetos foram pulverizados com óleo essencial de C. citratus em diferentes concentrações utilizando uma torre de Potter. Os valores de CL50 de M. persicae e F. schultzei foram de 0,28% e 1,49%, respectivamente. Óleo essencial de C. citratus é uma alternativa natural promissor para o desenvolvimento de inseticidas para o manejo de M. persicae.

Biological Control of Aphid Using Fungal Culture and Culture Filtrates of Beauveria bassiana

Aphids are one of the most destructive pests in crop production such as pepper, cucumber, and eggplants. The importance of entomopathogenic fungi as alternative pest control agents is increasing. Conidia of entomopathogenic fungi are influenced by environmental conditions, such as temperature and relative humidity, and cause slow and fluctuating mortality. These factors have prevented wider application and use of biocontrol agents. For investigation of means of mitigation of such problems, we conducted bioassays with 47 fungal culture filtrates in order to evaluate the potential of secondary metabolites produced by entomopathogenic fungi for use in aphid control. Among 47 culture filtrates cultured potato dextrose broth, filtrate of Beauveria bassiana Bb08 showed the highest mortality (78%) against green peach aphid three days after treatments. Filtrate of Bb08 cultured in Adamek's medium showed higher toxicity as 100% to third instar nymphs of the aphid compared with seven other filtrates cultured in different broths amended with colloidal chitin or oil. The culture filtrates and fungal cultures from media amended with colloidal chitin or oil had lower control efficacies than filtrates without these additives in three different media. These results indicate that the fungal culture fluid or culture filtrate of B. bassiana Bb08 cultured in Adamek's medium has potential for development as a mycopesticide for aphid control.

HrpNEa-induced deterrent effect on phloem feeding of the green peach aphid Myzus persicae requires AtGSL5 and AtMYB44 genes in Arabidopsis thaliana.

Myzus persicae, a generalist phloem-feeding insect, develops with induced expression of the AtMYB44 gene. Special GLUCAN SYNTHESIS-LIKE (GSL) genes and β-1,3-glucan callose play an important role in plant defence responses to attacks by phloem-feeding insects. Here we report that AtGLS5 and AtMYB44 are both required for HrpNEainduced repression of M. persicae feeding from the phloem of Arabidopsis leaves. In 24 h successive surveys on large-scale aphid populations, the proportion of feeding aphids was much smaller in HrpNEa-treated plants than in control plants, and aphids preferred to feed from the 37 tested atgsl mutants rather than the wild-type plant. The atgsl mutants were generated previously by mutagenesis in 12 identified AtGSL genes (AtGSL1 through AtGSL12); in the 24 h survey, both atgsl5 and atgsl6 tolerated aphid feeding, and atgsl5 was the most tolerant. Consistently, atgsl5 was also most inhibitive to the deterrent effect of HrpNEa on the phloem-feeding activity of aphids as monitored by the electrical penetration graph technique. These results suggested an important role of the AtGSL5 gene in the effect of HrpNEa. In response to HrpNEa, AtGSL5 expression and callose deposition were induced in the wild-type plant but not in atgsl5. In response to HrpNEa, moreover, the AtMYB44 gene known to be required for repression of aphid reproduction on the plant was also required for repression of the phloem-feeding activity. Small amounts of the AtGSL5 transcript and callose deposition were detected in the atmyb44 mutant, as in atgsl5. Both mutants performed similarly in tolerating the phloem-feeding activity and impairing the deterrent effect of HrpNEa, suggesting that AtGSL5 and AtMYB44 both contributed to the effect.

Eficiência de Chrysoperla externa (Hagen, 1861)(Neuroptera: Chrysopidae) no controle de Myzus persicae (Sulzer, 1776) (Hemiptera: Aphididae) em pimentão (Capsicum annum L.) / Efficiency of Chrysoperla externa (Hagen, 1861) (Neuroptera: Chrysopidae) in the Myzus persicae (Sulzer, 1776) (Hemiptera: Aphididae) population reduction in sweet pepper (Capsicum annum L.)

Avaliou-se a eficiência de larvas de primeiro ínstar de Chrysoperla externa no controle Myzus persicae em pimentão, em experimentos realizados em câmara e sala climatizadas a 25 ± 1ºC, UR de 70 ± 10 por cento e fotofase de 12 horas. Larvas de C. externa recém-eclodidas foram liberadas em discos foliares, contendo 5; 10 ou 20 ninfas do afídeo, acondicionados em placas de Petri. Em um segundo experimento, larvas do predador foram liberadas nas proporções 1:5, 1:10 e 1:20, em plantas inoculadas com 60; 100 e 140 ninfas do pulgão. O potencial predatório de C. externa nos discos foliares foi influenciado pela densidade inicial do pulgão. A eficiência do predador nas densidades de 5; 10 e 20 pulgões foi de 100 por cento; 96,7 por cento e 79,3 por cento, respectivamente. A eficiência das larvas em eliminar as populações do pulgão nas plantas de pimentão variou em função do tempo. Na proporção 1:5, a eliminação das ninfas ocorreu entre um e dois dias após a liberação do predador, enquanto nas proporções 1:10 e 1:20 o período de quatro dias não foi suficiente para que os pulgões fossem eliminados. As larvas de C. externa quando liberadas nas três proporções promoveram reduções na população de M. persicae, se comparada à testemunha. No entanto, nas proporções 1:5 e 1:10 o predador se mostrou mais eficiente.

The efficiency of Chrysoperla externa first instar larvae in the control of Myzus persicae in sweet pepper was evaluated in experiments conducted in climatic chamber and room at 25 ± 1ºC, 70 ± 10 percent RH and 12h photophase. C. externa newly emerged larvae were released in sweet pepper foliar discs containing 5; 10 or 20 aphid nymphs, putted in Petri dishes. In a second experiment, predator larvae were released in 1:5, 1:10 and 1:20 proportions in sweet pepper plants inoculated with 60; 100 and 140 aphids nymphs. The C. externa predatory potential in the foliar discs was affected by aphid initial density. The predator efficiency in 5; 10 and 20 aphid densities was 100 percent; 96,7 percent and 79,3 percent, respectively. The larvae efficiency in eliminating aphids populations in sweet pepper plants range as a result of the time. In 1:5 proportion the nymphs elimination occurred between one and two days after the predator release, while in 1:10 and 1:20 proportions four days were not sufficient to the aphids elimination. C. externa larvae released in the three proportions promoted reductions in M. persicae population when compared to the control. The proportions 1:5 and 1:10 the predator was more efficient.

Relação entre a atividade de esterases e a resistência de Myzus persicae (Sulz. ) (Sternorrhyncha:Aphididae) a pirimicarbe / Relationship between esterases activity and the resistance of Myzus persicae (Sulz. ) (Sternorrhyncha: Aphididae) to pirimicarb

O objetivo deste estudo foi avaliar a relação entre a atividade total das esterases e a resistência de indivíduos de M. persicae a inseticidas em populações coletadas em plantas de batata, em Contenda e Piraí do Sul, PR e Ibicoara, BA. No bioensaio, os afídeos foram colocados em recipientes de plástico transparente com o fundo coberto por uma tela de malha fina, em cinco repetições de 10 indivíduos para cada concentração. O conjunto foi mergulhado em concentrações ( por cento) de pirimicarbe a 0,0031; 0,0062; 0,0125; 0,025; 0,05 e 0,1 por cento, durante 10 segundos e depois secos em papel filtro. Após uma hora em câmaras a aproximadamente 25°C, os afídeos foram examinados sob microscópio estereoscópico, anotando-se o número de insetos mortos. A atividade de carboxilesterase foi medida através de ensaio colorimétrico in vitro. De acordo com os níveis de absorbância, os afídeos foram classificados em S; S/R1 (S ou R1); R1; R1/R2 (R1 ou R2); R2 e R3 (suscetível; suscetível/ levemente resistente; levemente resistente; levemente resistente/ altamente resistente; altamente resistente e extremamente resistente, respectivamente). Foi efetuada uma correlação simples entre os resultados das CL50 obtidas no bioensaio e a freqüência desses indivíduos. Encontrou-se uma correlação positiva entre a CL50 e R2 , mas negativa entre a CL50 e S e S/R1. O bioensaio, apoiado pelo teste colorimétrico, mostrou-se como uma ferramenta capaz de estimar com precisão tanto uma população com alta freqüência de indivíduos suscetíveis quanto outra com elevada proporção de indivíduos resistentes e altamente resistentes.

The objective of this research was to evaluate the relationship between the total activity of carboxylesterase and the resistance of M. persicae populations to pirimicarb. The aphids were collected on potato plants in Contenda and Piraí do Sul, PR and Ibicoara, BA, Brazil. For the bioassay, five replicates for each concentration were prepared with 10 aphids placed in plastic vials with a fine fabric mesh on the bottom, and dipped into pirimicarb concentrations ( percent) at 0.0031; 0.0062; 0.0125; 0.025; 0.05, and 0.1 percent, during 10 seconds, then dried on filter paper. After 1h inside a 25°C chamber, the aphids were examined under a stereomicroscope, recording the number of dead insects. The carboxylesterase activity was assessed by in vitro colorimetric technique. A simple correlation was established between the results of the LC50 bioassay and the frequency of M. persicae resistant variants. According to the absorbance level, the aphids were classified as variants S; S/R1 (S or R1); R1; R1/R2 (R1 or R2); R2 and R3, respectively susceptible; partially resistant, resistant and highly resistant. It was found a significant positive correlation between the LC50 and the frequency of R2 forms and a significant negative correlation between LC50 and the frequency of S and S/R1 forms. The colorimetric data supported the bioassay results, indicating that the bioassay was precise enough to determine either populations with high frequency of susceptible individuals or population with high proportion of resistant and highly resistant individuals.

Exigências térmicas e tabelas de vida de fertilidade de Myzus persicae (Sulzer) (Hemiptera: Aphididae) em laboratório / Thermal requirements and age-specific life tables of Myzus persicae (Sulzer) (Hemiptera: Aphididae) in laboratory

This study aimed to determine the thermal requirements of the green peach aphid, Myzus persicae (Sulzer), to calculate age-specific life tables, and to predict the occurrence of adults inthe field by a degree-day model. The aphids were reared, individualized, in leaf cages set on kale, Brassica oleracea L. var. acephala DC., kept at 15, 20, 23, 25 and 30°C, in laboratory. The lowestdevelopmental thermal threshold (Tb) was 2.2°C and the thermal constant (K) was 165.6 day-degree. The degree-day model predicted the occurrence of adults 0-1 day before the time they were first observed in the field. The age-specific life table parameters, on a degree-day time scale, indicated that the temperatures 23°C and 25°C provided the best thermal conditions for the population growth of M. persicae. At these temperatures the highest intrinsic rate of natural increase (rm = 0.012) was observed as well as the lowest mean generation time (T = 303.8 degree-day and T = 272 degree-day, respectively) and population doubling time (TD = 57.8 degree-day).

Este estudo visou determinar as exigências térmicas do pulgão Myzus persicae (Sulzer), calcular tabelas de vida de fertilidade, e prever a ocorrência de adultos no campo, por meio de modelo de graus-dia. Os pulgões foram mantidos individualmente em gaiolas fixadas em folhas de couve, Brassica oleracea L. var. acephala DC, em laboratório, às temperaturas de 15, 20, 23, 25 e 30ºC. O limite térmicoinferior de desenvolvimento (Tb) e a constante térmica (K) foram 2,2ºC e 165,6 graus-dia, respectivamente.O modelo de graus-dia previu a ocorrência de adultos de M. persicae para 0 a1 dia antes da data em queeles foram observados no campo. Os parâmetros das tabelas de vida estimados na escala de tempo emgraus-dia evidenciaram que as temperaturas de 23°C e 25°C foram as que proporcionaram as melhores condições térmicas para o crescimento populacional de M. persicae. Nessas temperaturas observou-se o maior valor da capacidade inata de aumentar em número (rm = 0,012), o menor valor da duração média da geração (T = 303,8 graus-dia e 272 graus-dia, respectivamente) e o menor tempo necessário para a população duplicar em número de indivíduos (TD = 57,8 graus-dia).

Plasticidade morfológica, reprodutiva e assimetria flutuante de Myzus persicae (Sulzer) (Hemiptera: Aphididae) sob diferentes temperaturas / Morphological and reproductive plasticity and fluctuating asymmetry of Myzus persicae (Sulzer) (Hemiptera: Aphididae) under different temperatures

Quatro clones do pulgão Myzus persicae (Sulzer) foram utilizados para se avaliar a magnitude de suas respostas plásticas em sete condições de temperatura 15,0; 17,5; 20,0; 22,5; 25,0; 27,5 e 30,0°C. Um índice multivariado de tamanho foi obtido por ACP, a partir de cinco caracteres morfológicos. Quatro desses caracteres foram medidos bilateralmente para quantificação da Assimetria Flutuante (AF). A fecundidade foi estimada pelo número de ninfas presentes do trato reprodutivo das fêmeas. A variabilidade atribuída aos componentes ambientais e genéticos de variação foi estimada com o auxílio de um modelo de genética quantitativa. A maior parte da variabilidade fenotípica morfológica na população estudada ocorreu como resposta à temperatura (componente ambiental), mas componentes genéticos de variação também foram detectados. Acredita-se que o potencial plástico descrito seja adaptativo, contudo um ambiente a 30°C não permitiu a sobrevivência. Pequena variabilidade no comprimento do rostro com relação aos demais caracteres pode ter resultado de seleção estabilizadora, considerando seu valor adaptativo para a obtenção de alimento. Os pulgões apresentaram AF em alguns dos caracteres medidos. Entretanto, não foi detectada relação linear dos níveis de AF com valores de temperatura, nem houve um padrão geral de variação para este caráter. Os dados corroboram a hipótese de que adversidades do meio afetam mais efetivamente o tamanho dos organismos e que, portanto, este seria um indicativo de estresse mais eficiente do que a AF de um caráter propriamente dito.

Four clones of the aphid Myzus persicae (Sulzer) were used to assess the magnitude of their responses to seven different temperatures (15.0, 17.5, 20.0, 22.5, 25.0, 27.5 and 30.0°C). A multivariate index of size was obtained using five morphological characteristics simplified by PCA. Among these characteristics, four were measured bilaterally both sides in order to evaluate the fluctuating asymmetry (FA). Fertility was estimated according to the number of nymphs in the reproductive female apparatus. The variability attributed to the environmental and genetic components was estimated using a genetic quantitative model. A large part of the total variability found for size distribution was attributed to differences in temperature (environmental component), but genetic components of variation were also detected. The plastic potential was considered adaptive, since it increases clone survival in unstable environments and allows new habitat exploitation. However, 30°C temperature did not permit clone development and survival. Lower variability in one morphological character, the rostrum, in comparison with the other traits may be the result of a stabilizing selection, in relation to its adaptive value to obtain food. The aphids presented FA in some of the measured characteristics. However, no linear relation was detected for the levels of FA and temperature values and no general pattern of variability was found. The data corroborate the hypothesis that size is more effectively affected than FA by external influences, and consequently, is a much more efficient indicator of environmental stress.