Insect pests pose significant threats to global agroecosystems due to their increasing resistance to chemical pesticides and their detrimental impacts on biodiversity (Bamisile et al. 2021). Conventional pest control methods, reliant on synthetic chemicals, raise environmental and health concerns, necessitating the development of sustainable alternatives. Among these, entomopathogenic fungi have emerged as promising biocontrol agents, capable of infecting and killing arthropod pests while minimizing ecological disruption (Chandler et al. 2011; Tiwari & Tripathi 2014).

Entomopathogenic fungi, such as Beauveria bassiana (Bals.-Criv.) Vuill. and Metarhizium spp., naturally regulate insect populations by colonizing their cuticles, leading to disease and death (Świergiel et al. 2016; Wang & Wang 2017). These fungi also exhibit dual roles – suppressing plant pathogens and parasitic nematodes – enhancing their utility in integrated pest management (IPM) (Sharma et al. 2021). Advances in genomic studies have further elucidated their host adaptation mechanisms, enabling targeted biocontrol strategies (Wang & Wang 2017).

Recent innovations include myco-nanopesticides, such as silver nanoparticles derived from fungal metabolites, which demonstrate high efficacy against pests (Bihal et al. 2023). Additionally, microbial biopesticides based on Bacillus thuringiensis Berliner and Metarhizium anisopliae (Metschn.) Sorokīn are widely used in agriculture (Lacey et al. 2015; Kumar et al. 2019). Field studies highlight the success of fungal agents in reducing pest populations, such as Cydia pomonella (codling moth) and Caliroa cerasi (cherry slug sawfly), key pests in orchards (Gürlek et al. 2018; Mesquita et al. 2023).

In Uzbekistan, apple and cherry orchards face severe infestations from C. pomonella and C. cerasi, yet research on native entomopathogenic fungi remains limited. Preliminary studies identified Aspergillus, Fusarium, and Alternaria spp. in these pests (Akhmedova et al. 2024), suggesting untapped potential for biocontrol.

This study aims to isolate and characterize entomopathogenic fungi from naturally deceased C. pomonella and C. cerasi in Uzbek orchards, evaluate their diversity and prevalence, with a focus on genera like Metarhizium and Beauveria, and assess their potential as biocontrol agents to support sustainable pest management. By leveraging native fungal strains, this research contributes to eco-friendly pest control strategies, reducing reliance on synthetic pesticides while enhancing orchard productivity.

Modified entomological, microbiological, and mycological approaches were employed to investigate microbial associations in orchard-dwelling insect pests. The study focused on apple and cherry orchards in Uzbekistan, with sampling conducted during May–June in 2023 and 2024.

Insect specimens were manually collected from tree trunks, branches, and foliage following a systematic inspection of the study area. Captured specimens were placed in sterile Petri dishes and transported to the Laboratory of Entomophage Ecology and Biomethod Theoretical Foundations at the Institute of Zoology, Academy of Sciences of the Republic of Uzbekistan, for further analysis. Target pest species included Cydia pomonella (Linnaeus, 1758) and Caliroa cerasi (Linnaeus, 1758).

Species identification was performed using taxonomic keys (Kuchurova & Maksakova 2003), while pest prevalence and distribution were assessed following established methodologies (Zhelokhovtsev 1988; Gilligan et al. 2018; Rakhmonova 2018).

Insect specimens were surface-sterilized in 70% ethanol (2–3 min), rinsed in 0.7% saline solution, and homogenized via agitation. Serial dilutions were prepared, and microbiological culturing was conducted in triplicate under aseptic conditions using a FH 1500x laminar flow hood.

Aliquots were plated on solid media and incubated in a DX 210 precision incubator at 26 ± 0.5 °C for 5–6 days. Emerging fungal colonies were subcultured onto Czapek-Dox agar (CZA) and potato dextrose agar (PDA) for purification and morphological characterization.

Fungal isolates were examined using a BS203 binocular microscope (400× magnification) equipped with a MICDC MOS 5mp USB camera. Taxonomic identification was based on macro- and micromorphological traits (e.g., hyphal structure, sporulation patterns, conidiophore morphology) following standard mycological references (Litvinov 1967; Pidoplichko & Milko 1971; Gorlenko 1976; Bilay 1982).

From naturally infected cadavers of Cydia pomonella and Caliroa cerasi collected across Uzbek orchards, we isolated 38 fungal strains representing 6 families, 8 genera, and 15 species. The isolates included members of the Mucoromycetes class (order Mucorales, family Mucoraceae, genus Mucor Fresen., 1850), with the most numerous families being Mucoraceae, Aspergillaceae, Cladosporiaceae, Pleosporaceae, Nectriaceae, and Clavicipitaceae. The dominant genera isolated were Aspergillus P. Micheli, 1729, Penicillium Link, 1809, Alternaria Nees ex Wallroth, 1816, Cladosporium Link, 1816, Fusarium Link, 1809, and Metarhizium Sorokīn.

Comparative analysis revealed differences in fungal communities between the two insect hosts. From C. cerasi we isolated 11 fungal species, while C. pomonella harbored 12 species. Both insect species shared several common fungal species (Table 1). Two potentially entomopathogenic species were identified: Fusarium solani (Mart.) Sacc. and Metarhizium anisopliae (Metschn.) Sorokīn. The fungal isolates showed characteristic morphological features when cultured on different media, with distinct colony morphologies and microscopic structures that aided in their identification (Table 1). The distribution patterns of these fungi varied between the two insect hosts, suggesting potential host-specific associations in their natural mycobiota.