Biological control of Spodoptera frugiperda (Nixon) (Lepidoptera: Noctuidae) in new invaded countries using insect pathogens

Background The fall armyworm (FAW), Spodoptera frugiperda (J.E. Nixon) (Lepidoptera: Noctuidae), is the major insect pest that infests cereal crops recently in African and Asian countries. The insect is polyphagous that attacks large numbers of host plants, especially maize and rice, causing considerable losses in their annual yield. The integrated pest management (IPM) of the insect depended mainly on insecticides and to some extent on biological control agents including insect pathogens (nematodes, fungi, bacteria and viruses). Results Different species of entomopathogens (nematodes, fungi, viruses and bacteria) infecting the insect could be isolated in such newly invaded countries. Laboratory and field experiments indicated that the insect was found to be susceptible to the isolated entomopathogens, and thus, they could be promising biocontrol agents against this insect. Conclusion This review article proved the susceptibility of S. frugiperda to the most of tested entomopathogens. However, more field studies have to be carried out in order to include such entomopathogens within integrated pest management programs against this insect pest


Background
The fall armyworm (FAW), Spodoptera frugiperda (Lepidoptera: Noctuidae), is the major insect pest that recently infests cereal crops in African and Asian countries.The insect is polyphagous that attacks large numbers of plants, especially maize and rice, causing 11.6-32-47% losses in maize yield of the total production/year.Tendeng et al. (2019) reported that the total life cycle of FAW averaged 25 days (22-28 days) at 25 °C.The female can deposit 1500-2000 eggs during its life span which ranges 7-10 days at 28 °C and 60% R.H. (Kumar et al. 2022).On rearing the insect on maize and Okra, the egg duration averaged 2.5 days while the larval duration averaged 13-15 days and the longevity of adults averaged 10.5 and 11.5 days in males and females, respectively (Kumar et al. 2022).Spodoptera frugiperda was detected starting from 2016 in West and Central Africa: Rwanda, Senegal, Sudan, Egypt, and over 44 African countries (Abbas 2023).As well, the pest has been recorded in Asia; India, China, Korea, Japan, Vietnam, Philippines, Sri Lanka, Syria, Jordan and Israel since 2016 (Pehlivan and Atakan 2022).The insect successfully invaded Europe (Germany, the Netherlands, Turkey) as well as Australia (Abbas 2023).Tendeng et al. (2019) claimed that FAW has long-distance migration ability that can covers 100 km per night, whereas Goergen et al. (2016) reported that in the Americas, adult moths of FAW could travel hundreds of kilometers per night on prevailing winds from their endemic zone to the worm regions.This ability might be one of the factors that facilitates its invasion of different parts of the world.
The IPM of FAW has been carried out worldwide mainly by using chemical insecticides, but because the pest has developed resistance to different groups of insecticides, biological control agents were found to have a great attention.Abbas et al. (2023) recorded 66 parasitoid and predator species of S. frugiperda in African countries as well as 35 species in Asian countries since its invasion starting from 2016.
The entomopathogenic nematodes (EPNs) in the two families Steinernematidae and Heterorhabditidae have been recorded as successful biocontrol agents against insect pests worldwide.
The entomopathogenic bacterium, Bacillus thuringiensis (Bt), is the major biocontrol agent infecting insect pests.It produces one or more crystalline proteinaceous inclusions (Cry toxins) adjacent to the endospore, which have been found to be toxic to invertebrates, mainly insects.Bt crops are plants genetically engineered (modified) to contain the toxins (Cry toxins) of Bt to be resistant to certain insect pests (Abbas 2018).
The entomopathogenic fungi (EPF) are associated with insects in almost all orders, and are among the most common pathogens that cause diseases in insects naturally.
However, very few commercial products of fungi proved to be successful insecticides.
The entomopathogenic viruses include more than 1100 species in 16 families (Adams 1991) and among these, baculoviruses are more promising and have been used as biopesticide with fair success (Inceogluetal 2006).FAW nucleopolyhedrovirus (SfMNPV), (a baculovirus), has been registered and used commercially in several African and Asian countries for FAW management (Firake et al. 2020;Hussain et al. 2021;Wennmann et al. 2021).
The present article deals with survey as well as laboratory and field evaluation of insect pathogens as biocontrol agents against the FAW in the new invaded countries in Africa, Asia and Australia.

Nematodes (Entomopathogenic Nematodes, EPNs)
In Nigeria, Ottun et al. (2021) found that at the concentration of 250 IJs/5 larvae of FAW, Heterorhabditis sp.caused 60% mortality in the treated larvae at 3 days posttreatment.In Egypt, Mohamed and Shairra (2023), evaluated S. carpocapsae and H. indica against 2nd-6th larval instars and found that S. carpocapsae caused 100% mortality in all larval instars at all the tested concentrations (150-2400 IJs).H. indica caused 100% mortality in 2nd and 3rd larval instars after 96 h, but 5th and 6th larval  Bateman et al. (2018), in Kenya, reported that S. carpocapsae and S. feltiae were registered as biological control agents of FAW in Cameron.Fallet et al. (2022) in Rwanda could isolate S. carpocapsae from soil samples.The nematode caused 100% mortality in FAW larvae when formulated in a carboxymethyl cellulose gel at a concentration of 3000 IJs/maize plant in the laboratory.However, it caused 72% mortality when applied in water.Danso et al. (2021), in Ghana, estimated the efficacy of unidentified Steinernematid and Heterorhabditid EPNs compared to the insecticide Emamectin benzoate in a greenhouse planted with maize (in pots).The results showed that both EPNs were as effective as Emamectin benzoate in reducing the numbers of FAW larvae as well as the infestation in maize cobs compared to the control.
The bacterium Bacillus thuringiensis (Bt) and Bt toxins Botha et al. (2019), in South Africa, evaluated the insecticidal efficiency of Bt maize against larvae of FAW fed on leaves of Bt maize expressing Cry1Ab (Bt1) and Cry1A.105+ Cry2b2 (Bt2).The results showed moderate survival (4-35%) on Bt1 maize and very high mortality (99%) on Bt2.The authors attributed the moderate survival on Bt1 to low dose of Cry1Ab in Bt1 or to that the insects arrived to the continent might have carried alleles with resistance to the Bt1.Fungi (entomopathogenic fungi, EPF) Akutse et al. (2019), in Kenya, tested 20 isolates of EPF at 1 × 10 8 conidia/ml for their efficacy against eggs and 2nd instar larvae of FAW as well as the neonate larvae that emerged from the treated eggs.Only B. bassiana (ICIPE676) isolate caused mortality of 30% to 2nd instar larvae.Metarhizium anisopliae (ICIPE 40) caused 83% mortality in the treated eggs.In addition, M. anisopliae (ICIPE 41) and (ICIPE 7) caused 96.5 and 93.7% mortality rates in the neonate larvae.Later, Akutse et al. (2020) tested 22 isolates (16 M. anisopliae and 6 B. bassiana) against FAW moths.All the 22 isolates were pathogenic to the moths, but the mortality varied significantly among the isolates 7 days post-treatment.Beauveria bassiana (ICIPE 621) and M. anisopliae (ICIPE 7) were superior among all the other isolates by causing 100% mortality of the moths with the lowest LT 50 values of 3.6 and 3.9 days, respectively.Sisay (2018), in Ethiopia, found that Beauveria sp. and Metarhizium sp. were highly effective against FAW larvae inducing 100% and 80% mortality rates, respectively, 5 days post-treatment.
Viruses (entomopathogenic Viruses: Baculoviruses) Bateman et al. (2018) stated that the commercial product of the cotton leafworm, Spodoptera littoralis nucleopolyhedrovirus (SlNPV), which targeted S. littoralis was found to be effective against FAW in Cameron and was being registered there.Further, Bateman et al. (2021) reported that commercial isolates of S. frugiperda NPV (SfMNPV) have been registered and successfully employed to control the FAW in some African and Asian countries (Kenya, Zambia, Bangladesh and Sri Lanka).In Nigeria, a new S. frugiperda multiblenucleopolyhedrovirus (SfMNPV-KA1) could be isolated from larvae of S. frugiperda (Wennmann et al. 2021).

Nematodes (Entomopathogenic nematodes, EPNs)
In Korea, laboratory studies were conducted by Acharya et al. (2020) to evaluate the efficiency of EPNs against 6th instar larvae and 5-day-old pupae at a concentration of 600 IJs/larva and 600 IJs/10 pupae.In larvae, 100% mortality was achieved by Heterorhabditis indica and Steinernema carpocapsae, whereas S. arenarium, S. longicaudum and H. bacteriophora caused 97, 93 and 53% mortality rates, respectively.The emergence rates of adults from treated pupae were 33% (S. carpocapsae), 37% (S. longicaudum), 40% (H.indica), 57% (S. arenarium) and 60% (H.bacteriophora).Similarly, Lalramliana et al. (2021), in India, treated 3rd and 5th larval instars and 1-day-old pupae of S. frugiperda by four EPNs at five concentrations for larvae (10-800 IJs/larva) and four concentrations for pupae (200-1600 IJs/pupa).The tested nematodes were H. indica, H. baujardi, S. sangi and S. surkhetense.The results revealed that rates of mortality in larvae ranged 43-100% in the 3rd instar larvae, 25-100% in the 5th instar larvae and 37-69% in the pupae.Wattanachaiyingcharoen et al. (2021), in Thailand, evaluated the efficiency of two indigenous H. indica and S. siamkayai against FAW larvae in laboratory and greenhouse.In laboratory, 2nd and 5th larval instars were treated at six concentrations (50-300 IJs/larva), whereas in the greenhouse, maize plants in pots, infested with 2nd instar larvae, were sprayed with 100 ml of each of the two species at 20,000 and 50,000 IJs/ml one day after releasing the larvae on the plants.Rates of mortality in 2nd instar  2022a) compared the virulence of an indigenous strain of Heterorhabditis sp. to a commercial S. feltiae against 3rd and 6th larval instars and 5-dayold pupae of FAW in laboratory.The tested concentrations of the nematodes were 100 IJs/larva and 1200 IJs/5 pupae.Rates of mortality in 3rd instar larvae were 84 and 100%, whereas in the 6th instar larvae, they were 56 and 93%, by Heterorhabditis sp. and S. feltiae, respectively.Rate of adult emergence from treated pupae was 26.7% by both nematodes compared to 93.3% in the control.Chen et al. (2023) evaluated the infectivity of H. bacteriophora (HbSD) against S. frugiperda under laboratory, greenhouse and field conditions.In laboratory assays, the nematode was highly virulent against the larvae.In greenhouse assays, spraying aqueous formulation of the nematode showed good performance in killing larvae on maize leaves.Patil et al. (2022), in India, reported that H. indica and S. carpocapsae caused 100% mortality in third-and fourth-larval instars of S. frugiperda, and 85% and 72% in pupae, respectively.Treatment of two maize fields, naturally infested with the insect by the nematodes at 2.5 × 10 8 IJs/ha, showed that H. indica significantly reduced the number of larvae and leaf damage.The efficacy of S. carpocapsae and H. indica applied twice against FAW in sweet corn field was tested by Ratnakala et al. (2023) in India.The results showed that S. carpocapsae reduced, significantly, the larval population and leaf damage in the treated crop.Shinde et al. (2022), in India, evaluated two native strains of H. indica (HI-MN and HI-CL) against larval instars of FAW.The LC 50s were 21.6 and 48.9 IJs (3rd instar larvae) and 25.5 and 52.4 IJs (4th instar larvae) for HI-MN and HI-CL, respectively.In China, treatment of 2nd instar larvae of S. frugiperda with S. carpocapsae and Also in China, Li-Mei et al. (2021), in field experiments, found that the larval density and percentages of damaged leaves and plants were significantly lower in Bt plants than in conventional ones.Li et al. (2019) evaluated five Bt toxins against neonate larvae of FAW using artificial diet assays.The results showed that the LC 50 values were 50.3, 161.3, 207.8, 603.7 and over 800 ng/cm 2 of diet for Cry1Ab, Cry1Ac, Cry1F, Cry2Ab and Vip3A, respectively.Zhao et al. (2023), in laboratory bioassays, found that Bt corn (DBN3601T) differed significantly in causing mortality in neonate larvae of fall armyworm during 3-year field trials compared to conventional corn.Zhang and Wu (2019) stated that the mortality of FAW larvae fed on leaves of Chinese Bt-Cry1Ab maize was less than 65%, whereas the mortality in larvae fed on Chinese Bt-(Cry1Ab + Vip3A) leaves ranged 53-100%.Similarly, Liang et al. (2021) reported that the mortality of FAW larvae that fed on different tissues of Chinese Bt-Cry1Ab maize DBN9936 ranged from 34 to 100%.
In India, an experiment was conducted to evaluate the efficiency of three products of fungi, a product of a baculovirus, a product of Bt and the insecticide, azadirachtin against S. frugiperda in a maize field during 2019/2020.Four sprays were applied at 12-day intervals.The results revealed that with fourth spray, Bt was the most effective (Wayal et al. 2021).
Endophytic Bt Karshanal and Kalia (2023), in India, reported that Bt could be established as endophyte in maize plants using different inoculation methods; seed treatment (ST), soil dranching (SD), foliar application (FA) and combination of all (ST + SD + FA).Feeding FAW larvae on the leaves of such plants caused 50% mortality by ST + SD + FA followed by 40% by ST.
Yan-li et al. ( 2022) could isolate eight fungal strains from diseased larvae of S. frugiperda in China.They tested the pathogenicity of M. rileyi against eggs, 1st-4th larval instars and pupae at 1 × 10 7 conidia/ml.Rates of mortality were 88.7, 72.5 and 56.7% in the 1st, 2nd and 3rd larval instars, respectively.Mortality was 56.6% in the eggs and 68.8% in the hatched neonate larvae after 3 days and reached 80.6% after 6 days.However, low mortality was found in the 4th instar larvae (5%) and pupae (14.8%) and did not differ from the control.Xu et al. (2020) compared the virulence of 3 B. bassiana strains isolated from soil in China (bbbj, bbzj and bbhn) against 3rd instar larvae of FAW at different concentrations.The strain bbbj had the highest virulence with LC 50 of 3.37 × 10 5 spores/ ml followed by the strain bbzj (with slightly less effect than bbbj) and then the strain bbhn.The strain bbhn, however, at the highest concentration (1 × 10 8 spores/ml) caused a mortality less than 50% in the treated 3rd instar larvae in 7 days.Visalakshi et al. (2020) found that spraying 3rd instar larvae of FAW with M. rileyi in laboratory at 2 × 10 8 spores/ml caused 90% mortality in treated larvae within 7 days.Ramanujam et al. (2020), in India, evaluated 10 indigenous strains of B. bassiana, M. anisopliae and M. rileyi, in India, against larvae of FAW in laboratory.Among the 10 isolates, M. anisopliae (strain Ma 35) caused 68% mortality, followed by B. bassiana (strain 1) with 64% and B. bassiana (strain 2) with 57% mortality.The other seven strains caused 11-29% mortality.In addition, field experiments in maize plots during 2019 showed that M. anisopliae (strain Ma 35) caused 70% reduction of infestation and 44% increase in the yield.The respective values for B. bassiana (strain 1) were 76% and 55%.Also, Ramanujam et al. (2021) reported that a survey of S. frugiperda in maize fields in Karnataka, India, in 2018 revealed that about 20-30% of the collected 4th and 5th larval instars were found to be infected with the fungus, B. felina.In China, Gao et al. (2022) stated that the LC 50s of B. bassiana (strain PfPb) for 1st to the 6th larval instars of S. frugiperda were found to be 7.7 × 10 5 , 5.5 × 10 6 , 2.2 × 10 7 , 3.1 × 10 8 , 9.6 × 10 8 and 2.5 × 10 11 spores/ml, respectively, 7 days post-treatment.Firake and Behera (2020) reported that M. reliye and S. frugioerda NPV (SfMNPV) were observed to be the dominant mortality factors of FAW larvae in corn fields in various locations across Northeast India throughout the season and were responsible for more than 50% mortality of the larvae.Ginting et al. (2020), in Indonesia, reported that N. rileyi was found, naturally, infecting 5.3-79% of S. frugiperda larvae collected from corn fields in five villages.
In Australia, Apirajkamol et al. (2022) tested six Beauveria isolates and five Metarhizium isolates against 3rd and 6th larval instars, pupae and moths of FAW.Two Beauveria isolates exhibited the highest mortality 7 days post-treatment at 3 × 10 3 conidia/ml.The isolate B-0571 caused 83, 61, 17 and 94% mortality in 3rd and 6th larval instars, pupae and moths, respectively.The respective values for the isolate B-1311 were 74, 72, 19 and 98%.Sari et al. (2022), in Indonesia, reported that 2-week old seedlings of maize already inoculated with the endophytic fungi, B. bassiana and M. anisopliaea were used to feed the neonate larvae of S. frigiperda for 6 h and were then fed on healthy non-inoculated leaves until pupation.The results showed that the fungal-colonized maize increased the developmental periods of larvae and pupae, significantly.However, % adult emergence, adults' longevity and number of eggs deposited/female were significantly decreased than the control.In addition, Gustianingtyas et al. (2021), in Indonesia, could obtain eight isolates of the endophytic fungi from the corn roots in Indonesia belonging to Aspergillus sp., Beauveria sp., Chaetomium sp. and Curvularia sp. and were found to have insecticidal activity against 2nd instar larvae of S. frugiperda at 1 × 10 6 spores/ml.The two most pathogenic isolates belong to Beauveria sp.(isolates JgCrJr and JgSPK) with larval mortality of 29.33 and 26.67%, respectively.Also, Herlinda et al.(2022), in Indonesia, stated that out of 20 isolates from endophytic fungi inoculated in corn, four isolates of B. bassiana, one isolate of M. anisopliae and one isolate of Curvularia lunata were found to be more pathogenic to S. frugiperda larvae.

Entomopathogenic viruses
Firake and Behera (2020) reported that Metarhizium reliye and S. frugioerda NPV (SfMNPV) were observed to be the dominant mortality factors of FAW larvae in various locations across Northeast India throughout the season and were responsible for more than 50% mortality of the larvae.Raghunandan et al. (2019) could isolate a nucleopolyhedrovirus from naturally infected larvae of FAW in India.A suspension of 10 8 occlusion bodies (OBs)/ml caused considerable mortality in treated larvae as well as malformation in the formed pupae and adults.Onkarappa et al. (2023), in India, studied the sublethal effects of the virus SfNPV on the biological parameters of FAW at five concentrations ranged from 3 × 10 4 to 1.18 × 10 7 OBs/100ul/late 2nd instar larvae.The results showed that the percentage of pupal mortality ranged from 40 to 72% at varying doses.Longevity of males and females was reduced compared to the control and the egg production of females ranged 150-338/ female at the different concentrations compared to 474 in the control.Sivakumar et al. ( 2020) noticed natural infection of FAW larvae by nucleopolyhedrovirus (NPV) in 2018 in three districts in India.The laboratory bioassay revealed that 1st, 2nd and 3rd larval instars were equally susceptible to infection (LC 50 3.71-5.02OBs/mm 2 of diet).Firake et al. (2020) could isolate a nucleopolyhedrovirus from collected 687 FAW larvae (in the 2nd and 4th larval instars) from corn fields in India.It was found that 23.7% of the larvae could not reach the adult stage due to infection with SfNPV.Similarly, Lei et al. (2020), in China, could isolate SfMNPV from FAW larvae and analyzed its molecular and biological characteristics.Kenis et al. (2022) reported that S. frugiperda NPV (SfMNPV) was registered in Bangladesh, Kenya and Cameroon to be used against FAW.Kasi et al. (2022), in India, evaluated the combination of the insecticide, Spinosad and each of the nematodes, S. feltiae and H. bacteriophora against the larvae of FAW in laboratory.The study showed that S. feltiae caused 60% mortality at a concentration of 2268 IJs/ larva.When combined with Spinosad at a concentration of 400 ppm, the mortality reached 90%.Similarly, H. bacteriophora caused 65% mortality alone and 95% when combined with Spinosad.In contrast, Spinosad alone caused 27.5% mortality in the treated larvae.

Conclusion
This review article proved the susceptibility of S. frugiperda to the four tested entomopathogens (nematodes, fungi, viruses and bacteria).However, more field studies have to be carried out in order to include such entomopathogens within integrated pest management programs against this insect.

Table 1
Virulence of Entomopathogens against Spodoptera frugiperda larvae in African countries

Table 2
Abbas et al. (2022)pathogens against Spodoptera frugiperda larvae in Asian countries 5% by H. indica and S. siamkayai, respectively.The respective values in the 5th instar larvae were 45 and 32.5% at 300 IJs/larva.In the greenhouse, rates of mortality 10 days post-treatment at 20,000 IJs/pot were 37.9% by H. indica and 28.75% by S. siamkayai.The respective values at 50.000 IJs/pot were 57.8 and 44.7%.In China,Abbas et al. (2022)reported that 280 IJs of Steinernema sp. could kill 100% of 3rd instar larvae of S. frugiperda, whereas 400 IJs of H. indica killed only 75%.Also,Wang et al. (