Genetic diversity of parasitoids and entomopathogenic nematodes of Spodoptera frugiperda Smith, 1797 (Lepidoptera: Noctuidae) in Senegal

J. Biodiv. & Environ. Sci. Babou et al. RESEARCH PAPER OPEN ACCESS Genetic diversity of parasitoids and entomopathogenic nematodes of Spodoptera frugiperda Smith, 1797 (Lepidoptera: Noctuidae) in Senegal Farma Fall Babou*1, Toffène Diome1, Mama Racky Ndiaye2, Mbacké Sembene1 1Entomology and Acarology Laboratory, Team Genetic for Population Management, Department of Animal Biology, Faculty of Science and Technology, Cheikh Anta Diop University, Dakar, Senegal 2Department of Agronomy and Plant Production, University of Sine Saloum El-Hâdj Ibrahima NIASS, Kaolack, Senegal Key words: Spodoptera frugiperda, Genetic diversity, Mermithidae, Parasitoids, Senegal DOI: https://dx.doi.org/10.12692/jbes/27.5.69-84 [ Published: November 09, 2025 ] ABSTRACT The pest Spodoptera frugiperda, when it invades Africa, is associated with a parasitic fauna, justifying the need to characterize the genetic diversity of its natural enemies in Senegal. Thus the rDNA of the large subunit, 28S was amplified. The study revealed the existence of 12 species of parasitoids obtained and 3 species of entomopathogenic nematodes. Parasitoids are divided into two orders (Hymenoptera and Diptera) and five families (Ichneumonidae, Braconidae, Eulophidae, Encyrtidae and Tachinidae). These parasitoids are Sinophorus exartemae, Chelonus sp., Parapanteles sp., Dolichogenidea sp. 1, Dolichogenidea sp. 2, Meteorus sp., Meteorus orocrambivorus, Euplectrus phthorimaeae, Prochiloneurus testaceus, Ooencyrtus masii, Drino sp. and Senometopia cariniforceps. Nematodes are members of the family Mermithidae and correspond to Hexamermis sp., Mermithidae sp. 1 and Mermithidae sp. 2. All these species attack the larval stage except O. masii which is oophagous. This study also reports new associations between S. frugiperda and the parasitoids S. exartemae, M. orocrambivorus, E. phthorimaeae, P. testaceus, O. masii and S. cariniforceps. To our knowledge, these six species, in addition to Drino sp., have just been known in the entomological fauna of Senegal. Therefore, it is urgent to protect these auxiliaries from the abusive and inappropriate use of chemical insecticides for sustainable management. *Corresponding Author: Farma Fall Babou  [email protected] * https://orcid.org/0009-0009-6704-6701 Co-authors: Toffène Diome: https://orcid.org/0000-0002-6810-4450 Mama Racky Ndiaye: https://orcid.org/0000-0002-0133-9030 Journal of Biodiversity and Environmental Sciences | JBES ISSN: 2220-6663 (Print); 2222-3045 (Online) Website: https://www.innspub.net E-mail contact: [email protected] Vol. 27, Issue: 5, p. 69-84, 2025 J. Biodiv. & Environ. Sci. Vol. 27, Issue: 5, p. 69-84, 2025 70 Babou et al. Journal of Biodiversity and Environmental Sciences | JBES Website: https://www.innspub.net INTRODUCTION Noctuidae is a family of insects belonging to the order Lepidoptera. The pests of this family are among the most harmful in the world. They can cause economic losses that can be estimated at more than 4500 million US dollars per year (Sisay et al., 2019 ; Riaz et al., 2021 ; Rahmawati and Rahardjo, 2024). Most of the adults in this family have a high migratory capacity, high fertility and a very high reproduction rate (Riaz and Ly., 2021; Altaf et al., 2022; Chen et al., 2022; El-Refaie et al., 2024). In addition, Noctuidae larvae are highly polyphagous and voracious, possessing a wide and diverse host range (Babou et al., 2022; Rahmawati and Rahardjo, 2024). Fall armyworm (FAW), Spodoptera frugiperda Smith, 1797 is a perfect illustration of this. It causes economic damage especially to maize, its preferred host plant (Sisay et al., 2019; Zhou et al., 2021). This species has been invasive in Africa since 2016 (Nuambote-Yobila et al., 2022; Nuambote-Yobila, 2023) and Asia from 2018 (Ge et al., 2021). Due to the crop losses caused, especially in sub-Saharan Africa, chemical pesticides have been used (Nuambote-Yobila et al., 2025). However, this method of control is proving to be difficult due to resistance to many classes of insecticides, linked among other things to their overuse but also to the insects' adaptation and detoxification mechanism (Boaventura et al., 2020; Guo et al., 2022; Moustafa et al., 2024; Van den Berg and du Plessis, 2022). With dificulty of management of the pest, it is essential to develop ecologically respectful and sustainable means of control (Arias et al., 2019; Mansour et al., 2023; Babou et al., 2025). For this reason, beneficial insects are at the forefront, in an invasion context where native natural enemies can be registered as potential pathogens to S. frugiperda (Abang et al., 2021; Ganou et al., 2024). Its enemies can be parasitoids or entomopathogenic nematodes (Sun et al., 2020; Kumar et al., 2024). However, identifying these auxiliaries accurately, reliably and quickly can be difficult in some situations (Chen et al., 2021; Ivezić et al., 2021). This is explained on the one hand by the large number of entomophagous present in the agrosystems of the tropical and subtropical zone; on the other hand, it may be linked to a lack of specialists in morphological identification (Delvare, 1991; Chen et al., 2021). This suggests that there could be many more species than are usually recognized. Hence, the use of genetics as an approach to identify the natural enemies of S. frugiperda (Sagar et al., 2022). In this respect, this activity aims to genetically characterize the parasitoids and entomopathogenic nematodes of S. frugiperda in Senegal. MATERIALS AND METHODS Sampling Spodoptera frugiperda caterpillars found on various plants in the Groundnut Basin (‘Bassin arachidier’), Casamance and the Niayes area (‘Zone des Niayes’) were collected. Egg masses were also collected. The samples were sent to the Entomology and Acarology Laboratory of the Cheikh Anta Diop University in Dakar. They were kept at room temperature. Then the caterpillars were fed with 60g of fresh leaves, replaced every two days. Monitoring was daily and was done until the emergence of adult S. frugiperda, parasitoids or entomopathogenic nematodes. The natural enemies emerging from the caterpillar were dipped in 96% alcohol for genetic identification. All individuals are recorded in Table 1 below. Molecular analysis of entomopathogenic parasitoids and nematodes Extraction, polymerase chain reaction and DNA sequencing Total genomic Deoxyribonucleic Acid (DNA) from entomopathogenic nematodes and parasitoids was extracted using the Zymo Research Tissue Kit, according to the manufacturer's instructions, with some modifications. The extraction was carried out on the entire body of the individuals, with the exception of those belonging to the family Tachinidae where the abdomen was removed. A number of 28 samples were extracted for parasitoids and 21 for nematodes. They were chopped into thin pieces or ground according to the stiffness of the specimens' cuticle. The quality of the extracts obtained was approximately assessed by electrophoretic migration using a DNA Ladder size marker, 100 base pairs. J. Biodiv. & Environ. Sci. Vol. 27, Issue: 5, p. 69-84, 2025 71 Babou et al. Journal of Biodiversity and Environmental Sciences | JBES Website: https://www.innspub.net Table 1. Characteristics of the samples to be identified Orders Families Morphotypes Number of individuals Hymenoptera Ichneumonidae Charops sp. 1 Braconidae Chelonus sp. 2 Meteorus sp. 4 Microgastrinae 1 3 Microgastrinae 2 2 Undetermined Braconidae 2 Eulophidae Eulophidae 2 Encyrtidae Encyrtidae 1 1 Encyrtidae 2 1 Indeterminate Hymenoptera Indeterminate 2 Diptera Tachinidae Tachinidae 1 6 Tachinidae 2 2 Mermithida Mermithidae Mermithidae 21 The 28S ribosomal DNA (28S rDNA) was then amplified. The 28sF3633 primer pair: 5'- TACCGTGAGGGAAAGTTGAAA-3' and 28sR4076: 5'- AGACTCCTTGGTCCGTGTGTTT-3' were used to amplify this gene, specifically in the D2 domain (28SD2). The study of Schmidt et al. (2006) even reveals that at the level of the 28S gene, the D2 region is the most suitable for inferencing relationships, especially for taxonomic purposes. The conditions of the thermal cycler in amplification were as follows: an initial denaturation at 94°C for 3 min; 35 cycles of denaturation at 92°C for 30 s, hybridization at 52°C (45s) characterized by the attachment of the primers and elongation of the complementary strands at 72°C (1 min). Finally, a single period of final elongation at 72°C for 10 min was performed. The sequencing has been outsourced. Genetic analyses The resulting sequences were thoroughly checked, corrected and aligned using the Clustal-W algorithm (Thompson et al., 1997) with BioEdit 7.7.1 software (Hall, 1999). The sequences obtained in this study were compared to authenticated sequences available on the GenBank database based on the E-score through the BLAST (Basic Local Alignment Search Tool) program in NCBI (National Center for Biotechnological Information). This made it possible to verify their membership of the different taxonomic groups. The species considered to be identical to the sequences compared were those with the highest percentage of identity and percent coverage and the lowest E value. Phylogenetic reconstructions were performed using the Bayesian inference (with 1,000,000 generations and 4 Markov chains) and the maximum parsimony method (Fitch 1971) using Mr. Bayes version 3.2 and MEGA version 10.2 software (Kumar et al., 2016) and the GTR (General Time Reversible) model with a gamma distribution. Branch robustness was assessed by bootstrapping with 1,000 replications for maximum parsimony. To the sequences obtained from the present study, others have been added from the Genbank database and which are associated with their accession number. The reconstructions were rooted with a sequence of Helicoverpa armigera Hübner, 1808 (Lepidoptera: Noctuidae) for parasitoids and Daubaylia sp. (Nematoda: Daubayliidae) for nematodes. RESULTS Sequences obtained Of the forty-nine (49) samples sequenced, including twenty-eight (28) parasitoids and twenty-one (21) entomopathogenic nematodes, seven (7) were eliminated with regard to chromatograms, due to poor adhesion of the primers during sequencing. Of the seven, two are nematode samples and five are insect samples, including the 2 undetermined Braconidae, 1 of the two individuals of Eulophidae and two of the 6 Tachinidae 1. The rest of the analyses were carried out on 23 parasitoid sequences and 19 nematode sequences. J. Biodiv. & Environ. Sci. Vol. 27, Issue: 5, p. 69-84, 2025 72 Babou et al. Journal of Biodiversity and Environmental Sciences | JBES Website: https://www.innspub.net Identification and phylogenetic reconstruction of parasitoids Table 2 shows that twelve (12) insect species parasitized the pest Spodoptera frugiperda, based on the Blast performed. The percentages of identities are recorded in the table below. Two orders of insects are confirmed. These are the Hymenoptera with four families (Braconidae, Ichneumonidae, Eulophidae and Encyrthidae) and which make up most of the species (10) which are Sinophorus exartemae, Chelonus sp., Parapanteles sp., Dolichogenidea sp. 1, Dolichogenidea sp. 2, Meteorus sp., Meteorus orocrambivorus, Euplectrus phthorimaeae, Prochiloneurus testaceus, and Ooencyrtus masii. The second order concerns the Diptera consisting of a single family (Tachinidae) and two (2) species: Drino sp. and Senometopia cariniforceps. Table 2. List of S. frugiperda parasitoid species identified from 28S rDNA Orders/Families Morphotypes identified Species identified Accession No. Identity (%) E-value Hymenoptera: Ichneumonidae Charops sp. Sinophorus exartemae EF406253.1 94,61 0,0 Braconidae Chelonus sp. Chelonus sp. 2 AF029124.1 96,45-97,09 0,0 Hymenoptera indeterminate Chelonus sp. 2 AF029124.1 95,32-96,45 0,0 Microgastrinae 1 Parapanteles sp. MN645200.1 98,71-98,31 0,0 Microgastrinae 2 Dolichogenidea sp. 1 PV468782.1 94,57 0,0 Microgastrinae 2 Dolichogenidea sp. 2 MK568812.1 95,37 2.10-160 Meteorus sp. Meteorus orocrambivorus KJ591259.1 98,02-98,23 0,0 Meteorus sp. Meteorus sp. LC632021.1 97,67-96,88 0,0 Eulophidae Eulophidae Euplectrus phthorimaeae MW586890.1 96,82 0,0 Encyrtidae Encyrtidae 1 Prochiloneurus testaceus MT783605.1 88,53 5.10-151 Encyrtidae 2 Ooencyrtus masii PQ790242.1 88,48 3.10-148 Diptera: Tachinidae Tachinidae 1 Drino sp. OR082818.1 82,47-96,90 2.10-86-0,0 Tachinidae 2 Senometopia cariniforceps AB466064.1 96,93 -95,55 9.10-169-0,0 Fig. 1. Phylogenetic tree of parasitoids, inferred using the Bayesian approach The species names associated with accession numbers are taken from the Genbank database. Bootstrap values are indicated in front of the nodes. The analysis of the tree obtained by the Bayesian approach (Fig. 1) reveals the existence of 2 clades: The first clade is made up of individuals of the family Braconidae and Tachinidae. Tachinidae individuals have a strong genetic affinity for each other. Specimens of Senometopia cariniforceps separated from those of Drino sp. with a bootstrap value of 96% and 84% depending on the individual. However, the individuals sampled in Senegal are phylogenetically closer. As for the Braconidae, the sequences of Chelonus sp. and that of Chelonus insularis are genetically very close and are grouped together in a well-supported cluster (87%). The subclade made up only of Microgastrinae 1 (Parapanteles) and Microgastrinae 2 (Dolichogenidea) individuals reveals that they belong to the same taxonomic group (81%). Individuals of Dolichogenidea sp. 1 and those of Dolichogenidea sp. 2 share a well-supported subclade (81%). This shows the similarity between these sequences. This same remark is noted for the second subclade formed by Parapanteles sp. (95%). The two species Meteorus orocrambivorus and Meteorus sp. J. Biodiv. & Environ. Sci. Vol. 27, Issue: 5, p. 69-84, 2025 73 Babou et al. Journal of Biodiversity and Environmental Sciences | JBES Website: https://www.innspub.net obtained, have a genetic affinity of 100%. There is also an individual of M. orocrambivorus which formed a cluster with the two individuals of Meteorus sp. (53%). The second clade groups the individuals of Ichneumonidae, Eulophidae and Encyrtidae in one clade but in different subclades supported by strong bootstrap (99% and 100%). Individuals of the family Ichneumonidae, Sinophorus exartemae and Charops sp. (EF364443.1) share the same node for a maximum bootstrap of 100%. However, a stronger genetic affinity (100%) is noted between the species of Eulophidae (Prochiloneurus testaceus and Ooencyrtus masii) and Encyrtidae (Euplectrus phthorimaeae). Fig. 2. phylogenetic tree of parasitoids inferred by the maximum parsimony method The red diamonds indicate the species sampled and the others come from the Genbank database. The bootstrap values are shown below the node. Unlike the Bayesian method, the maximum of parsimony (Fig. 2) forms 3 clades, one of which includes individuals of Ichneumonidae, the second individuals of the family Braconidae and the third is formed by the species Eulophidae and Encyrtidae but in different subclades. Just like the first method, the branches are well supported with a slight difference in the bootstrap values. Identification and phylogenetic reconstruction of entomopathogenic nematodes Sequence analysis indicates the existence of 3 species of Mermithidae nematodes: Hexamermis sp. (LC844941.1) (4 individuals), Mermithidae sp. (PQ811687.1) (12 individuals) and Mermithidae sp. (LC788416.1) (3 individuals). The latter two are named in the study Mermithidae sp.1 and Mermithidae sp. 2 respectively. The percentages of identity are shown in Table 3. Fig. 3. Phylogenetic tree of entomopathogenic nematodes, inferred using the Bayesian approach The species names associated with accession numbers are taken from the Genbank database. Bootstrap values are indicated in front of the nodes. Fig. 4. phylogenetic tree of entomopathogenic J. Biodiv. & Environ. Sci. Vol. 27, Issue: 5, p. 69-84, 2025 74 Babou et al. Journal of Biodiversity and Environmental Sciences | JBES Website: https://www.innspub.net nematodes inferred by the maximum method of parsimony The red diamonds indicate the species sampled and the others come from the Genbank database. The bootstrap values are shown below the node. The trees of the individuals of bayesian inference (Fig. 3) and the maximum parsimony (Fig. 4), show the existence of 4 very distinct clades supported by high bootstrap values, between 67 and 100%. The Mermithidae species collected in this study form a single clade with a bootstrap of 76% and 100% depending on the methods (Fig. 3 and Fig. 4). In this clade, individuals of Mermithidae sp. 1 group into two subclades while individuals of Mermithidae sp. 2 and Hexamermis sp. are not resolved in this clade, according to maximum parsimony. The Bayesian method shows a grouping of individuals of Hexamermis sp. and Mermithidae sp. 2, in addition to two Mermithidae sp. 1. The species Hexamermis sp. (LC844941.1), which has more similarity to some sequences of the study and the species Hexamermis agrotis (KC784667.1) shares a clade that seems to have a strong affinity with a bootstrap of 82% and 93%. The species Mermithidae sp. 2 (LC788416.1) identified forms a subclade (Fig. 3) (73%) or a clade (Fig. 4) (100%) with another Mermithidae sp. (LC788415.1). However, Mermithidae sp.1 (PQ811687.1) does not form a clade with individuals of the genus Hexamermis, nor with other Mermithidae species, so does not seem to be resolving for both types of reconstructions performed. The species Hexamermis popilliae (3 sequences) and Hexamermis sp. (OR614376.1) constitute a clade backed by a bootstrap of up to 100%. Table 3. List of identified Spodoptera frugiperda entomopathogenic nematode species Morphotypes Species identified Accession No. Identity (%) E value Hexamermis sp. LC844941.1 89,29-89,32 3.10-148 Mermithidae Mermithidae sp. 1 PQ811687.1 89,15-89,61 6.10-150-1.10-146 Mermithidae sp. 2 LC788416.1 88.71-89.17 4.10-147-8. 10-144 DISCUSSION The present study, which aimed to genetically characterize the entomopathogenic parasitoids and nematodes of S. frugiperda in Senegal, revealed the presence of 12 species of insects and 3 species of entomopathogenic nematodes. This pest first reported in Africa by (Sisay et al., 2018) and in Senegal by Brévault et al. (2018) has been described with its community of parasitic species (Ruiz-Nájera et al., 2013; Sisay et al., 2018; Gani et al., 2023). In America, more than 130 species, parasitoids of S. frugiperda are listed by (Molina-Ochoa et al., 2003; Murúa et al., 2006) and fewer than five species of entomopathogenic nematodes in the Americas and Asia (Ruiz-Nájera et al., 2013; Sun et al., 2020). The first reports on its natural enemies in Africa reveal a dozen parasitoids (Sisay et al., 2018; Kenis et al., 2019; Chipabika et al., 2023) and Mermithidae nematodes (Gani et al. 2023). This is in line with the results of this study where parasitoids and nematodes parasitized the FAW. Of the 12 parasitoid species revealed by molecular identification, eleven (11) emerged from the larval stage versus one from the embryonic stage of S. frugiperda. These species belong to the orders Diptera and Hymenoptera. Diptera have long parasitized Lepidoptera, a pest of maize in Africa (Chinwada et al., 2014 ; Faithpraise et al., 2015). According to the study conducted by Molina-Ochoa et al. (2003) in America, more than fifty species of Tachinidae are in association with S. frugiperda. Several studies conducted on the African continent show that Tachinidae can cause parasitism rates of up to more than 15%. The parasitoid Tachinidae of S. frugiperda including species of the genus Drino, are well distributed across the African continent (Sisay et al., 2018; Durocher-Granger et al., 2021; Chipabika et al., 2023) particularly in West Africa (in Benin and Ghana) (Agboyi et al., 2020; Blakely et al., 2022). This is in line with the results of this study. A species, Drino sp. is observed in Burkina J. Biodiv. & Environ. Sci. Vol. 27, Issue: 5, p. 69-84, 2025 75 Babou et al. Journal of Biodiversity and Environmental Sciences | JBES Website: https://www.innspub.net Faso (Ahissou et al., 2021; Gani et al., 2023) and Zambia (Chipabika et al., 2023). Indeed, it predominates among the Tachinidae parasitoids of S. frugiperda, and has a wide distribution in these two countries (Chipabika et al., 2023; Gani et al., 2023). This confirms the results of this study, due to its presence in the Groundnut Basin and Casamance. This species is often cited in the literature, which may be the same or different species and may not yet be described. Concerning Senometopia cariniforceps Chao and Liang, 2002 (Carcelia cariniforceps), she is a solitary parasitoid. According to Pernek et al. (2016), the species of Senometopia can be generalist parasitoids. S. cariniforceps is first encountered in China (O'Hara et al., 2009). It should be noted that it is not yet reported in Africa, except in this study. However, two species of Senometopia were collected in Cameroon from Helicoverpa armigera by Di Giovanni et al. (2019). Also Senometopia illota Curran, 1927 is identified as a new association with S. frugiperda in Indonesia by Lestari et al. (2024). This same observation is noted by Shima and Tachi (2023) Come in S. cariniforceps and Catocala columbina Leech, 1900 in Japan. In fact, in parasitoids, It is common to meet associations with new guests (Efil and Kara, 2004). This is in line with the results of this research. S. cariniforceps has never been registered as a parasitoid of S. frugiperda and probably a new association. The parasitoids of S. frugiperda are mainly Hymenoptera with the family Braconidae dominating (Molina-Ochoa et al., 2003). This is in line with the results of this study where six species of Braconidae are recorded against two species of Encyrtidae, one species of Ichneumonidae and one species of Eulophidae. Species of the genus Chelonus are often reported as parasitoids of S. frugiperda in the world (Gupta et al., 2020; Sagar et al., 2022). They are reported in parasitic East and West Africa S. frugiperda or species of the same genus (Agboyi et al., 2020; Ahissou et al., 2021; Durocher-Granger et al., 2021; Koffi et al., 2022; Shen et al., 2023; Ganou et al., 2024). The results of the study are in line with those of Tendeng et al. (2019) in Senegal and those of Chipabika et al. (2023) in Zambia, which recorded the presence of a species of Chelonus not identified morphologically and genetically respectively. Our study indicated, based on the phylogenetic tree, that Chelonus sp. is 87% or 99% related to Chelonus insularis Cress, 1865, a promising biological control agent against S. frugiperda (Rezende et al., 1994). The results obtained indicate three species of the subfamily Microgastrinae: Parapantels sp. Dolichogenidea sp. 1 and Dolichogenidea sp. 2. Species of the genus Parapantels have a wide host range and are distributed in many countries. Some of its species have been recorded on Spodoptera cosmioides Walk, 1858 and Spodoptera eridania Stoll, 1782 (Whitfield et al., 2018; Freitas et al., 2019). A species named Parapantels sp. was newly associated with S. frugiperda Zambia (DurocherGranger et al., 2021). This is in line with the results of the study. A strong affinity was observed between the genus Parapanteles and Dolichogenidea, either 81% or 100% depending on the approach. In Senegal, three species of Dolichogenidea are listed by Crash et al. (2021). However, molecular identification does not show resemblance to these three species. The study of Crash et al. (2021) confirms that there are no keys to distinguish African species from Dolichogenidea, which can justify the existence of both Dolichogenidea sp. Indeterminate. Species in this genus have a very restricted host range and often attack Tuta absoluta Meyrick, 1917 (tomato pest) (Agboka et al., 2022; Aigbedion-Atalor et al., 2022) explaining his first report on S. frugiperda in a context where market gardening crops that shelter their main hosts are not very present in an area such as the groundnut Basin, especially during rainy periods when field crops are promoted. Concerning Meteorus orocrambivorus and Meteorus sp. species of the same genus are known as parasitoids of Spodoptera and among which S. frugiperda (Molina-Ochoa et al., 2001; Freitas et al., 2019). Species of the genus Meteorus have increased the list of native parasitoids of the pest in the invaded J. Biodiv. & Environ. Sci. Vol. 27, Issue: 5, p. 69-84, 2025 76 Babou et al. Journal of Biodiversity and Environmental Sciences | JBES Website: https://www.innspub.net areas. Mano et al. (2023) and Dickson et al. (2022) were able to show in Burkina Faso and Senegal respectively, the presence of the parasitoid Meteorus laphygmarum Brues, 1926 on moths. It is also a potential regulator of S. frugiperda in America, highlighting the ability to Meteorus to parasitize the FAW. This corroborates the results of this study. M. Orocrambivorus is not yet reported on S. frugiperda and in Senegal as well, according to the research carried out. This confirms the continuous adaptation of Meteorus to the FAW. The species Sinophorus exartemae Uchida, 1928 (Limnerium exartemae) is a family of Ichneumonidae. Studies have shown that species of this genus are distributed in central and northern America, where they are endemic, and in part of Asia and Europe (Sanborne, 1983; Sheng and Sun, 2014). In the Afrotropical zone, 3 species of Sinophorus are known among the 114 (Sheng and Sun, 2014). Studies have shown that this genus often parasitizes the larvae of Noctuidae, a pest of maize in the same way as S. frugiperda (Sertkaya and Bayram, 2005; Fortuna et al., 2023). S. exartemae is an endoparasitoid, solitary. However, it is can be documented and never seems to be reported in Senegal and even less as a parasitoid of S. frugiperda. Even if Charops sp., a species with which it is phylogenetically closely related (100%), is recorded on the FAW in Zambia (Durocher-Granger et al., 2021), in Mozambique (Caniço et al., 2020) and Charops ater Tanzania, Kenya (Sisay et al., 2018) and Niger (Moussa et al., 2023). The results of the genetic analysis were able to reveal the presence of Euplectrus phthorimaeae Ferrière, 1941. Studies show that wasps Euplectrus are parasitoids of Lepidoptera including Noctuidae, distributed in Africa (Burkina Faso) (Mano et al., 2023) (Euplectrus sp.) and in other continents (Hansson et al., 2015; Žikić et al., 2017). The species Euplectrus laphygmae Ferrière, 1941 is listed in Zambia as a new association with S. frugiperda (Durocher-Granger et al., 2021; Lestari et al. 2024). This is in line with the results of this study because indeed, Euplectrus phthorimaeae also is not yet registered on S. frugiperda. Referring to the study of Nagoshi and Meagher (2008), the females of Euplectrus prefer to lay their eggs on the corn strain of S. frugiperda. While individuals from Euplectrus phthorimaeae collected are sampled from maize and sorghum. Two preferred host plants of the maize strain according to Pashley (1986). This proves the possibility of new associations between Euplectrus at S. frugiperda. Moreover, a species Euplectrus frugiperda was recently discovered in Australia on S. frugiperda for the first time by Fagan‐Jeffries et al. (2024), hence its name. The sample of Encyrtidae 1 analyzed, corresponds to at Prochiloneurus testaceus Agarwal, 1965 (Achrysopophagus testaceus). Gender Prochiloneurus Silvestri, 1915 is cosmopolitan (Hayat et al., 2008). These species are known to be hyperparasitoids of species of the same family as it (Encyrtidae), Pteromalidae, Coccinellidae, among others (Noyes and Hayat, 1984; Pillai et al., 2009). In West Africa Prochiloneurus insolitus Alam, 1961 is recognized as the most abundant native hyperparasitoid on Pseudococcidae (Goergen and Neuenschwander, 1992). The author Godfray (1994) In his book, he points out that if the primary host is not parasitized, the hyperparasitoid can lay eggs in it and the larvae will develop as primary parasitoids. The authors Shivakumara et al. (2022) also claim to have found six species of hyperparasitoids Prochiloneurus sp. as primary parasitoids of two species of mealybugs. In the case of P. testaceus, it is difficult to say whether the parasitoid directly attacked the non-parasitized FAW or not, since it has never been recorded on the latter. In this study of Encyrtidae species in the same way as P. testaceus are sampled in the Groundnut Basin, in maize cultivation and at the same time. So the presence of these species could attract P. testaceus on S. frugiperda. Also, since the caterpillar has infested maize more than other pests since its invasion in Africa, its abundance could attract the native primary and secondary parasitoids that were once dependent on other J. Biodiv. & Environ. Sci. Vol. 27, Issue: 5, p. 69-84, 2025 77 Babou et al. Journal of Biodiversity and Environmental Sciences | JBES Website: https://www.innspub.net pests. However, its status as a hyperparasitoid or primary parasitoid of S. frugiperda, remains undetermined. Molecular analysis reveals that individuals of morphotype 2 of Encyrtidae are found to be Ooencyrtus masii Mercet, 1921. Species in this genus are parasitoids of a wide host range (Laraichi et al., 1979). The parasitized species can be of different orders, including the Lepidoptera (Laraichi et al., 1979; Žikić et al., 2017; Polaszek et al., 2023; Acheampong et al., 2024). This is consistent with the results of this study, where the species O. masii is found among the parasitoids of S. frugiperda. However, according to the research results obtained, this oophagous parasitoid has never been recorded on S. frugiperda. But insects, in certain conditions, can adapt according to the availability of the hosts in the wild (Abrams and Kawecki, 1999). This species was already considered to be solely dependent on the eggs of Malacosoma neustria Linnaeus, 1758 (Laraichi et al., 1979). The authors Fusu and Andreadis (2023) emphasize thatOoencyrtus seems to be able to adapt very quickly to invasive pests. This is in line with our results. To our knowledge, this study is the first recording ofO masii in Senegal. Proving Laraichi's words et al. (1979) on the restricted or wide distribution ofOoencyrtus. Molecular identification of Mermithidae nematodes indicated the existence of 3 species. These are Hexamermis sp., of Mermithidae sp.1 and Mermithidae sp. 2. This family of nematodes often attacks species of Lepidoptera that may belong to the family Noctuidae and specifically to the genus Spodoptera (Villemant et al., 2015; Babu et al., 2019). The species S. frugiperda since its invasion of Asia and Africa, Mermithidae have been associated with it. Species Ovomermis sinensis and Hexamermis cf. albicans are reported in China and India respectively between 2019 and 2020 (Firake and Behere, 2020; Sun et al., 2020). In two studies carried out in Burkina Faso, species or a species of Mermithidae, because it is undetermined, parasitized the caterpillar with very high rates (Ahissou et al., 2021; Gani et al., 2023). In Senegal, an undetermined species of the genus Hexamermis was collected from larvae of S. frugiperda by Tendeng et al. (2019). This is related to the results of this study, even if in the previous one only a morphological identification was applied. In addition Babu et al. (2019) have also found a species Hexamermis sp., parasitizing Spodoptera litura Fabricius, 1775 et S. exigua. This reinforces the idea that, Hexamermis can parasitize S. frugiperda. For the other two species whose molecular identification was limited to the family, one is similar to a species from Japan (Mermithidae sp. 2) and the other from Italy. The study of Kakui and Shimada (2022), carried out two years earlier in Japan and on the same host species (S. calcitrans), assigns nematodes to the genus Ovomermis, Hexamermis and Amphimermis thanks to the associated molecular data, the analysis of phylogenetic trees. This would also mean that Mermithidae sp. 2 would certainly not come out of these three genres. Since for the amplified region, there are already sequences of Hexamermis in the database while there is no one for the genera Ovomermis and Amphimermis. It can then be assumed that Mermithidae sp. 2 can correspond to Ovomermis or Amphimermis. It should be noted that individuals from Mermithidae sp. 2, due to the presence of a caudal appendage in post-parasitic juveniles and the presence of the stylet, it may be suspected that it is the genus Ovomermis although it has not yet been reported in Senegal. Moreover, in Senegal, apart from the morphological study of Tendeng et al. (2019) and that of Faye et al. (2024) that identified the family, only the genetic study of Kobylinski et al. (2012) was carried out on mosquito parasites and where an indeterminate Mermithidae was identified. The authors Kakui and Shimada (2022) confirm by adding that this is certainly linked to the unavailability of data in the databases. This can therefore minimize the number of sequences with which the sequences of a study could be compared. It may also be explained by the fact that the species is a new to the genus Hexamermis. 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