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(=Nomuraea) Rileyi (Farlow) Samson from Spodoptera Exigua (Hübner) Cross Infects Fall Armyworm, Spodoptera Frugiperda (J.E

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(=Nomuraea) Rileyi (Farlow) Samson from Spodoptera Exigua (Hübner) Cross Infects Fall Armyworm, Spodoptera Frugiperda (J.E Philippine Journal of Science 150 (1): 193-199, February 2021 ISSN 0031 - 7683 Date Received: 06 Apr 2020 Metarhizium (=Nomuraea) rileyi (Farlow) Samson from Spodoptera exigua (Hübner) Cross Infects Fall Armyworm, Spodoptera frugiperda (J.E. Smith) (Lepidoptera: Noctuidae) Larvae Melissa P. Montecalvo* and Marcela M. Navasero National Crop Protection Center, College of Agriculture and Food Science University of the Philippines Los Baños 4031 College, Laguna, Philippines Mycobiocontrol is a promising management strategy in mitigating the fall armyworm Spodoptera frugiperda (J.E. Smith) infestation in the Philippines. An isolate of Metarhizium (=Nomuraea) rileyi (Farlow) Samson from onion or beet armyworm, S. exigua, which induced high mortality to this pest, was assessed against different larval instars of S. frugiperda. Surface-sterilized corn leaves were treated with different conidial concentrations and fed to S. frugiperda larvae. Cross infection of this entomopathogenic fungus to S. frugiperda was confirmed with a fungal infection that was initiated at 1–2 d post-treatment depending on the age of the larvae. Larval mortality significantly increased at 4–5 d post-treatment. Up to 100% larval mortality was recorded at 7 d post-treatment. Early larval instars (1st–3rd) were more susceptible than late larval instars (4th–6th). Higher conidial concentrations caused a higher and faster rate of larval mortality than lower conidial concentrations. The inflicted mycoses due to M. rileyi resulted 5 8 –1 in a slightly lower lethal dose (LD50) (1.44 x 10 to 9.36 x 10 conidia ∙ mL ) and shorter mean time to death (4.51–8.89 d). Mummification of the cadaver confirmed fungal infection with white fungal growth that later changed to green during sporulation. Our findings suggest the infectivity of M. rileyi and can be explored as a biological control agent against S. frugiperda. Keywords: biological control, entomopathogenic fungus, fall armyworm, Metarhizium rileyi INTRODUCTION Damages in different growth stages of corn were observed in incidence areas and may cause significant yield losses. In the Philippines, armyworms are major and invasive Armyworm larvae feed on leaf tissues, which can result in lepidopterous insect pests of crops. The black armyworm, severe defoliation (Business Queensland 2020). Spodoptera exempta (Walker), infested corn, rice, and sugarcane crops in Southern Tagalog and Bataan provinces Armyworms are effectively controlled by entomopathogens in 2010 (Navasero et al. 2011). The onion or beet such as nucleopolyhedroviruses (Montecalvo and armyworm, S. exigua, caused an estimated crop loss of Navasero 2019), fungi (Grijalba et al. 2018; Montecalvo PHP 1.6 B from February–April 2016 in onion production and Navasero 2020), bacteria (Moar et al. 1995), and areas – including Nueva Ecija, Pangasinan, and Ilocos Sur protozoa and nematodes (Business Queensland 2020). (Navasero et al. 2017). In 2019, the invasive species of fall Among the entomopathogens, entomopathogenic fungi are armyworm, S. frugiperda, was reported in 17 municipalities promising biological control agents against Spodoptera in 10 provinces in the Philippines (Navasero et al. 2019). species. Mycobiocontrol utilizes fungi to lower the insect density to reduce the disease-producing activity and crop *Corresponding Author: [email protected] 193 Philippine Journal of Science Montecalvo and Navasero: Metarhizium (=Nomuraea) Vol. 150 No. 1, February 2021 rileyi from Spodoptera exigua damage (Sandhu et al. 2012). Over 750 fungal species can Bioassay using various conidial suspensions. Young corn provoke fungal infections in insect populations. leaves (5-cm length) were surface sterilized, following Asi et al. (2013). Conidial concentrations were sprayed to both Metarhizium species are known entomopathogenic fungi sides of the corn leaves using a mist sprayer. Tween 80 that are widely studied because of their global distribution, (0.1%) was applied to corn leaves for the control set-up. broad host range, safety, environmental friendliness, and Four treated corn leaves were introduced to S. frugiperda ease of mass production (Aw and Hue 2017). Reported larva/e in a Petri plate with moistened cotton. For early species of Metarhizium infecting lepidopterous pests (1st–3rd) larval instars, lower conidial concentrations (1 x include M. anisopliae against S. exigua (Han et al. 2014) 103 to 1 x 108 conidia ∙ mL–1) were used. Higher conidial and S. litura (Petlamul and Prasertsan 2012), and M. concentrations (1 x 105 to 1 x 1010 conidia ∙ mL–1) were rileyi against S. frugiperda (Grijalba et al. 2018) and S. assayed for late (4th–6th) larval instars. Ten (10) 1st and litura (Rajan and Muthukrishnan 2009). Infected insects 2nd larval instars were initially cultured per plate and die due to mechanical damage resulting from tissue single cultured upon reaching 3rd larval instar. One larva invasion, depletion of nutrient resources and toxicosis, and per plate was cultured for 3rd–6th larval instars. The Petri production of toxin in the insect body (Sandhu et al. 2012). plates were sealed with Parafilm. Fresh corn leaves that In our previous research, M. rileyi was successfully were surface-sterilized were introduced in the succeeding isolated from the cadavers of S. exigua and effectively days post-treatment. Each treatment was replicated thrice caused the lethal infection to this pest in a laboratory with 10 larvae per replicate. bioassay (Montecalvo and Navasero 2020). High lethal Mortality was observed daily. Mycosis of cadavers was infection to S. exigua resulted in a low pupation rate and confirmed in blotter set-up. Cadavers were dipped in 1% adult emergence. M. rileyi is a dimorphic hyphomycete sodium hypochlorite for 1min and washed twice in sterile that causes epizootic death in insect species belonging to distilled water for 1 min. The cadavers were dried in sterile Lepidoptera and Coleoptera (Sandhu et al. 2012). filter paper and placed in a microscope slide inside a Petri Considering the virulence of this entomopathogenic fungus plate lined with moistened filter paper. against Spodoptera species, this study aimed to assess the Percentage mortality was corrected using the equation: potential of M. rileyi as a new tool to be integrated into M (%) = [(t – c) / (100 – c)] × 100, where: M = corrected pest management programs against S. frugiperda. We mortality; c = percentage mortality in controls; t = conducted this bioassay to elucidate the pathogenicity of percentage mortality in treatments (Abbott 1925). this isolate to the different larval instars of this insect pest Mean time to death was calculated using the formula: and to determine the lethal dose (LD) and time as important mean time to death (d) = [(x1y1)+(x2y2)+(xnyn)] / total parameters in determining its biocontrol efficacy. mortality, where: x = number of larvae died on a given day; y = number of days of which the observation was made considering the time when the trial was initiated (El-Hawary and Abd El-Salam 2009). MATERIALS AND METHODS Laboratory rearing of S. frugiperda. Advanced generation Statistical Design and Analysis (F9) larvae of S. frugiperda were obtained from the The experimental set-up was arranged in CRD. The data existing culture at the Biocontrol Laboratory of NCPC, was analyzed following the general linear procedure of which was originally collected from Gonzaga, Cagayan, Statistical Tool for Agricultural Research. Treatment Philippines. Neonates were fed with fresh leaves of a means were compared by means of the analysis of variance native variety of corn (Lagkitan) until the desired stages using Tukey's honestly significant difference test. LD of larvae. Bioassays were conducted when the appropriate values were calculated using PriProbit ver 1.63. age/instar of test insects was reached. Preparation of conidial suspensions. M. rileyi isolate from S. exigua was revived and subcultured in PDA for at least 14 d. This entomopathogenic fungus was isolated from RESULTS mummified larvae of S. exigua collected from the infested The entomopathogenic fungus M. rileyi isolated from S. field of bunch onion from San Jose City, Nueva Ecija. exigua caused the lethal infection to S. frugiperda larvae Conidia were harvested from the cultures and suspended (Figure 1). Various larval instars of S. frugiperda have in 0.1% Tween 80 solution. Conidia in the suspension a different reaction to M. rileyi infection. Early larval were counted using a haemacytometer. Various conidial instars were more susceptible than late larval instars. concentrations (1 x 103 to 1 x 1010 conidia ∙ mL–1) were Fungal infection was initiated at 1–2 d after treatment prepared by diluting the stock in 0.1% Tween 80 solution. 194 Philippine Journal of Science Montecalvo and Navasero: Metarhizium (=Nomuraea) Vol. 150 No. 1, February 2021 rileyi from Spodoptera exigua Figure 1. Cumulative mortality of Spodoptera frugiperda larvae treated with different conidial concentrations of Metarhizium (=Nomuraea) rileyi. depending on the age of the larvae. Larval mortality significantly increased at 4–5 d post-treatment. Higher conidial concentrations caused higher and faster larval mortality than lower conidial concentrations. Conidial concentrations of 1 x 107 and 1 x 108 conidia ∙ mL–1 killed more than 50% of 1st and 2nd larval instars at 6 and 5 d post-treatment, respectively. The conidial concentration of 1 x 1010 conidia ∙ mL–1 caused 50% mortality to 4th and 5th larval instars at 4 d post-treatment. However, mortality in 3rd and 5th larval instars peaked at 7 d post-treatment. Lower conidial concentrations (1 x 103 to 1 x 106 conidia Figure 2. Mean mortality of early larval instars of Spodoptera ∙ mL–1) caused slower larval mortality. frugiperda infected with Metarhizium (=Nomuraea) rileyi at 7 d post-treatment. Zero mortality in Control. Fungal infection in early and late larval instars at 7 d post-treatment is presented in Figures 2 and 3. M. rileyi caused significantly different mortalities against different larval instars. The virulence of M. rileyi was dependent on conidial concentrations and age of larval instars. Concentrations of 1 x 106 to 1 x 108 conidia ∙ mL–1 caused 50–100% mortality in 1st and 2nd larval instars.
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