aphicidal assessment of entomopathogenic fungi by the statistical analysis of corrected mortality in mustard aphids

Optimized Bioassay for Entomopathogenic Fungi Against Mustard Aphid

The mustard aphid (L. erysimi) is a notorious pest that infests a variety of cruciferous crops, causing significant economic losses1. While several systematic insecticides have been recommended to combat aphid infestations, the frequent use of these insecticides raises concerns about pesticide resistance2,3. Therefore, in terms of eco-friendly pest management, entomopathogenic fungi (EPF) could serve as a suitable alternative control strategy. EPF is an insect pathogen with the ability to infect hosts by penetrating their cuticles, making it a potent agent for controlling aphids and other plant-sucking insects4. Furthermore, EPF has proven to be a feasible and sustainable pest management technique, offering benefits such as plant pathogen antagonism and plant growth promotion5.
EPF can be obtained through insect-soil baiting or isolated from insect cadavers in the field6,7. However, before further use of fungal isolates, pathogenicity screening is necessary. Several studies have been conducted on the effectiveness of EPF against aphids, which are significant crop pests that can cause severe damage8,9. Mustard aphids, among various species of aphids, have been tested for susceptibility to several strains of Beauveria spp., Metarhizium spp., Lecanicillium spp., Paecilomyces spp., and even Alternaria, which is primarily known as a saprophytic and plant pathogenic fungus but has shown some lethal effects against mustard aphids10,11,12.
To evaluate the effectiveness of EPF against aphids under laboratory conditions, bioassays can be divided into two main parts: the inoculation chamber and fungal inoculation. The current protocol describes the construction of an inoculation chamber, where aphids can be maintained using various methods such as an excised leaf with a petiole wrapped in moist cotton, an excised leaf disc with a Petri dish lined with damped filter paper, direct maintenance on pot plants, or an excised leaf disc embedded in water agar within a Petri dish or container10,11,13. Common methods for fungal inoculation include conidia spraying, aphid immersion into a conidia suspension, leaf dipping into a conidia suspension, and plant endophyte inoculation11,14,15,16. While various inoculation methods exist, the bioassays should simulate field application conditions. For example, in the case of the leaf dipped method12,17, the efficiency of EPF can be evaluated, but since the aphids infest the fungus-loaded leaves, the dorsal side of the aphid, which is a preferential penetration site, does not usually get exposed to the fungus.
To evaluate the aphidicidal effect of EPF under laboratory conditions, this protocol suggests using the detached-leaf method described by Yokomi and Gottwald18 with some modifications, followed by conidia inoculation using a micro-sprayer. This method maintains approximately 100% humidity in the bioassay chamber for at least seven days without requiring additional replenishment of water18,19. Additionally, confining aphids to one surface ensures their exposure to conidia spraying and facilitates observations20. However, aphids may become stuck in the exposed agar surface while moving within the inoculation chamber. Furthermore, recording data in the Petri dish experiment with mustard aphids, which are parthenogenetic insects, can be challenging due to their rapid development and reproduction. It is difficult to distinguish between inoculated adults and their progeny without removal. The details of how to proceed with this step are seldom mentioned, and some inconsistent factors, such as leaf consumption area, need to be optimized.
This protocol demonstrates a stable system for screening the virulence of EPF isolates against mustard aphids, enabling the selection of potential isolates against various aphid species from an extensive EPF library. Field-collected aphids can be identified, and a sufficient laboratory population of mustard aphids can be established to evaluate the aphidicidal effect of various fungal isolates using an easy and feasible methodology with consistent outcomes. Aphids have developed multiple evolutionary mechanisms in response to intense and repeated anthropogenic pressures in agroecosystems, posing challenges to food security9. Therefore, this described method could be extended to evaluate potential EPF isolates against various aphid species.

Author Spotlight: Eco-Friendly Pest Management Using Entomopathogenic Fungi Against Mustard Aphids 

Assessment of Aphidicidal Effect of Entomopathogenic Fungi against Parthenogenetic Insect, Mustard Aphid, Lipaphis erysimi (Kalt.)

Watch this Scientific Journal Video about Aphicidal Assessment of Entomopathogenic Fungi by the Statistical Analysis of Corrected Mortality in Mustard Aphids at JoVE.com

겨자 진딧물의 수정된 사망률에 대한 통계적 분석에 의한 곤충병원성 곰팡이의 살균 평가

먼저 SPSS 소프트웨어 플랫폼을 실행한다. 치료 및 수정된 사망률이라는 변수를 만듭니다. 그런 다음 동일한 농도로 동일한 시점에서 접종된 다른 분리물에 대해 계산된 결과를 입력합니다.분석, 평균 및 비율 비교로 이동한 다음 독립 T-검정 분석을 위해 SPSS에서 독립 표본 T 검정으로 이동합니다. 다음으로, 그룹화 변수 상자에 처리를 입력하고 검정 변수 상자에 사망률을 수정합니다. 서로 다른 곰팡이 분리에 따라 그룹을 분류하고 OK를 눌러 분석을 시작합니다.치사 시간 중앙값과 치사 농도 중앙값을 계산하려면 총계, 반응 및 기간 또는 집중으로 정의된 변수를 만듭니다. 그런 다음 기록된 결과를 스프레드시트에 입력합니다. 마지막으로 분석(Analyze), 회귀(Regression), 프로빗(Probit)을 차례로 선택하여 치사 시간 중앙값과 치사 농도 중앙값을 계산합니다.밀리리터당 7개의 분생포자를 10개에 접종한 결과, 3 및 4 DPI에서 메타히지움 균주 197과 동충하초 균주 213에 대해 유의하게 다른 보정 사망률이 나타났다. 동충하초 균주 213은 다른 치료법에 비해 현저히 짧은 치사 기간을 보였다. 또한 메타히지움 213 균주에 비해 더 낮은 치사량 값을 보였다.

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JoVE Journal

Biology

Aphicidal Assessment of Entomopathogenic Fungi by the Statistical Analysis of Corrected Mortality in Mustard Aphids

70 조회수. 먼저 SPSS 소프트웨어 플랫폼을 실행한다. 치료 및 수정된 사망률이라는 변수를 만듭니다. 그런 다음 동일한 농도로 동일한 시점에서 접종된 다른 분리물에 대해 계산된 결과를 입력합니다.분석, 평균 및 비율 비교로 이동한 다음 독립 T-검정 분석을 위해 SPSS에서 독립 표본 T 검정으로 이동합니다. 다음으로, 그룹화 변수 상자에 처리를 입력하고 검정 변수 상자에 사망률?...

비디오: 겨자 진딧물의 수정된 사망률에 대한 통계적 분석에 의한 곤충병원성 곰팡이의 살균 평가

The mustard aphid (L. erysimi) is a pest that infests various cruciferous crops and transmits plant viruses. To achieve eco-friendly pest management, entomopathogenic fungi (EPF) are potential microbial control agents for controlling this pest. Therefore, virulence screening of EPF isolates under Petri dish conditions is necessary before field application. However, the mustard aphid is a parthenogenetic insect, making it difficult to record data during Petri dish experiments. A modified system for detached-leaf bioassays was developed to address this issue, using a micro-sprayer to inoculate conidia onto aphids and prevent drowning by facilitating air-drying after spore suspension. The system maintained high relative humidity throughout the observation period, and the leaf disc remained fresh for over ten days, allowing parthenogenetic reproduction of the aphids. To prevent offspring buildup, a process of daily removal using a painting brush was implemented. This protocol demonstrates a stable system for evaluating the virulence of EPF isolates against mustard aphids or other aphids, enabling the selection of potential isolates for aphid control.

This protocol presents an optimized detached-leaf bioassay system for evaluating the effectiveness of entomopathogenic fungi (EPF) against the mustard aphid (Lipaphis erysimi (Kalt.)), a parthenogenetic insect. The method outlines the data collection process during Petri dish experiments, enabling researchers to consistently measure the virulence of EPF against mustard aphids and other parthenogenetic insects.

The mustard aphid (L. erysimi) is a pest that infests various cruciferous crops and transmits plant viruses. To achieve eco-friendly pest management, ...