Introduction

Most agricultural production systems worldwide employ chemical insecticides to control pests. The global pesticide consumption in 2019 was approximately 4.19 million metric tons ^{(Pathak et al. 2022)}. In Mexico, the average annual use of pesticides from 2010 to 2014 was 7.87 kg/ha, ranking third worldwide and first in Latin America ^{(Zhang 2018)}, this has had significant consequences for the environment, public health, and the development of insect resistance ^{(Herrera-Moreno et al. 2018}, ^{Li et al. 2023)}. Biological control has been developed as an environmentally friendly alternative to chemical pest control methods. It involves the introduction or enhancement of natural enemies of the pest, these can include insects, mites, nematodes, fungi, bacteria, and viruses. Specifically referring to microorganisms to enhance crop growth, increase yield, and protect crops from pathogens and pests; the global agricultural microbial market is valued at USD 6.4 billion in 2022 since the consumer preference for organic food products, gradual phase-out of crucial ingredients, residue levels in food, pest resurgence, and resistance, and increasing need for agricultural sustainability ^{(Markets & Markets 2023)}.

Currently the use of Agricultural Fungi-based Insecticides (AFI) is part of the biological control strategies in integrated pest management and agroecological pest management. Unlike chemical insecticides, AFI only reduces insect populations to a level that does not cause economic losses; their toxicity is categorized as reduced or null in humans and other organisms, and due to their co-evolutionary process, they have the advantage of behaving as endophytic microorganisms that function as growth promoters and also protect plant hosts from pathogens and herbivores ^{(Behie et al. 2015}, ^{Butt et al. 2016}, ^{Vega 2018)}. AFI have increased in popularity in the last two decades with a record of 110 commercial products based on Entomopathogenic Fungi (EF), of which 40 % have Beauveria bassiana and 39 % Metarhizium anisopliae sensu lato ^{(Faria an& Wright 2007}, ^{Mascarin & Jaronski 2016)}.

Since B. bassiana interaction with silkworms was observed by Agostino Bassi in 1835, and M. anisopliae studied by Eli Metchnikoff in 1878, the EF have been studied to develop microbial pest control agents ^{(Lacey et al. 2015)}. These discoveries have promoted the study, use, and marketing of these and other EF worldwide in the 21th century.

The effectiveness of EF in agricultural pest control is attributed to their host range and sophisticated infection mechanisms. The infection process begins when susceptible insects come into contact with fungal conidia or blastospores, which then adhere to the insect cuticle. This triggers the expression of various hydrolytic enzymes, including proteases, chitinases, and lipases, as well as other factors that facilitate fungal germination and growth across the host’s surface, leading to the penetration of the cuticular layers. Upon breaching the cuticle, the fungus enters the hemocoel, where host immune responses such as melanization, sclerotization, phagocytosis, nodulation, and encapsulation take place ^{(Ortiz-Urquiza et al. 2013)}. Within the hemocoel, blastospores proliferate, invading other tissues, absorbing nutrients, and producing secondary metabolites. Ultimately, the fungus forms sporulation structures that emerge from the insect cadaver, thus continuing its life cycle ^{(Butt et al. 2016)}.

Among the most commonly and effectively used biopesticides are the AFI whose active ingredients are entomopathogenic fungi (EF) and are used mainly as microbial insecticides against agricultural pests, as well as insect vectors of human diseases ^{(Murillo-Alonso et al. 2019}, ^{Ortiz and Sansinenea 2023)}.

In Mexico, the Centro Nacional de Referencia de Control Biológico (CNRCB) maintains the largest collection of EF strains, which are crucial for developing biological pest control strategies, primarily in the agricultural sector. These strains encompass a variety of genera, including Metarhizium, Beauveria, Cordyceps, Hirsutella, Simplicillium, Aschersonia, Purpureocillium, Entomophthora, Akanthomyces, and Gibellula ^{(Montesinos-Matías et al. 2020)}.

Agricultural Fungi-Based Insecticides (AFI)

The development of a bioinsecticide is a complex process that requires extensive research and development over several years. The process begins with seeking infected insects or the use of insect baits to obtain entomopathogenic fungi from the soil. These fungi undergo isolation and morphological and molecular characterization. Additionally, the optimal growth conditions are evaluated, including culture media, fermentation type, thermotolerance tests, UV light tolerance, chemical tolerance, tests for synergism and antagonism, and water stress. Also, the biological effectiveness of the bioinsecticide is assessed under both laboratory and field conditions, focusing on metrics such as mean lethal concentration, mean lethal dose, and mean lethal time. Efficient mass production methods are developed, followed by the creation of a suitable formulation. This includes testing the formulation’s tolerance to ingredients, quality parameters, hydrophobicity, packaging, and labeling. Further assessments ensure persistence in the field and determine the shelf life, considering both the physical and biological stability of the formulation. Before commercialization, the bioinsecticide must undergo evaluation by regulatory agencies. In Mexico, this involves obtaining authorization and registration from the Comisión Federal para la Protección contra Riesgos Sanitarios (COFEPRIS) in coordination with the Secretaría de Medio Ambiente y Recursos Naturales (SEMARNAT) and the Secretaría de Agricultura y Desarrollo Rural (SADER).

In Mexico, the study, production, and application of EF in the field began in 1990 in some research centers and mainly at the CNRCB ^{(Zelaya-Molina et al. 2022}, ^{Rodríguez del Bosque et al. 2015)}. One of the first cases of the development of microbial pest control agents with EF in Mexico was the control of the locust (Schistocerca piceifrons) with M. acridum, where the CNRCB had an essential role in laboratory and field studies and the diffusion of the technology ^{(Hernández-Velázquez and Arredondo-Bernal 2003)}. In the mid-90s, the first companies that produced and marketed AFI were consolidated; one of these companies was “Agrobiológico del Noroeste, S.A. de C.V.’’ (Agrobionsa). At the end of the 90s, twenty institutions and private companies producing and marketing beneficial organisms for insect control in Mexico had at least one EF species ^{(Tamez-Guerra et al. 2001)}. By 2001, there were 11 products based on EF, of which five had B. bassiana as active ingredient, three had M. anisopliae, and another three had Paecilomyces fumosoroseus. These fungi were produced by four companies and two laboratories from the Mexican government. In 2010, a total of 28 plants producing biopesticides were documented (17 companies and 11 laboratories of propagation centers and/or units), which mainly produced B. bassiana, followed by M. anisopliae, P. fumosoroseus, Verticillium lecanni, M. anisopliae var. acridum and Paecilomyces sp. ^{(García de León & Mier 2010)}.

According to the “Directorio de Laboratorios Reproductores y Comercializadores de Agentes de Control Biológico DGSV-CNRCB, y DGSV-CNRF-Departamento de Control Biológico (DCB) ^{(SENASICA 2020}, ²⁰²²⁾” and the “Registro Sanitario de Plaguicidas, Nutrientes Vegetales de la Comisión Federal para la Protección contra Riesgos Sanitarios ^{(COFEPRIS 2022)}”, 76 products with EF as an active ingredient are produced and marketed (Tables 1-4). Of these, 53 contain as an active ingredient a single EF species, 23 products of B. bassiana, 16 M. anisopliae, 7 I. fumosorosea, 4 A. lecanii, 2 M. acridum, and only 1 product that contains I. javanica. The remaining products contain two or more EF strains and even a mixture with entomopathogenic bacteria (Table 4). At least 56 other products containing EF were not documented and are not described in the tables because the information provided was not accurate or the label lacked an adequate description, such as the amount of the conidial concentration, the type of formulation, and target pests. Products containing B. bassiana, according to the description on the labels and technical sheets, are used to control arthropods belonging to Coleoptera, Hemiptera, Lepidoptera, Orthoptera, Thysanoptera, and Trombidiformes (Table 1). Products that contain M. anisopliae as an active ingredient are used to control Coleoptera, Hemiptera, Dermaptera, Diptera, Hymenoptera, Lepidoptera, Orthoptera, Thysanoptera and Trombidiformes (Table 2). In contrast, products containing I. fumosorosea aim to control insects of the order Hemiptera (Table 3) and A. lecanii controls insects of the orders Homoptera and Hemiptera (Table 3). Sixty-three percent of companies and institutions dedicated to the production and sale of this type of AFI indicate conidia or spores as the active ingredient, while the rest only indicate that the products contain Colony Forming Units (CFU). The formulations of AFI are diverse, and according to the classification of formulations provided by the Food and Agriculture Organization of the United Nations-World Health Organization ^{(FAO-OMS 2017)} the 61.8 % of the documented products have a wettable powder (WP) formulation and contain a variable concentration of CFU or conidia in presentations of 100, 200, 240, 250, 300, 400, 500 and 1000 g. The rest of the formulations are diverse, with 5.2 % in the form of emulsifiable concentrate (EC), 14.5 % soluble concentrate (SL), 9.2 % suspension concentrate (SC) and 9.2 % granules (GR).

The formulations of AFI marketed in Mexico differ significantly in the concentration of the active ingredient or the infective units (conidia/gram, conidia/liter, spores/gram, spores/liter, CFU/gram, and CFU/liter). The taxonomic identity of the marketed AFI usually needs to be more precise due to the complexity of fungal nomenclature, which is constantly revised and changing. For example, according to current taxonomic and phylogenetic reviews, Nomurea rileyi belongs to the genus Metarhizium; Verticillium lecanii changed to Lecanicillium lecanii and now to Akanthomyces lecanii; Paecilomyces fumosorosea changed to Isaria fumosorosea and now to Cordyceps fumosorosea (Table 5).

Use of AFI in phytosanitary campaigns and programs

Phytosanitary campaigns in Mexico aim to detect, control, and prevent the spread of pests and pathogens from Mexican vegetable production. Epidemiological surveillance, chemical and biological control, and legal activities are carried out to improve the crop’s competitiveness ^{(SENASICA 2023)}.

Among the phytosanitary campaigns that have used AFI are those carried out with I. javanica and M. anisopliae to control the “Asian citrus psyllid” (Diaphorina citri), which is a hemipteran that causes damage to citrus trees, mainly from the Rutaceae family that represents 589,683 ha in 2017 ^{(SENASICA 2019a)}. From 2012 to 2015, 34,644 ha of citrus trees were treated with the strains CHE-CNRCB 303 and 307 of I. javanica, and strain 224 of M. anisopliae in the states of Colima, Hidalgo, Jalisco, Nayarit, Oaxaca, San Luis Potosí, Tamaulipas and Veracruz ^{(Sánchez-González et al. 2015)}. During 2018, applications of M. anisopliae were carried out on 3,884 ha in the state of Hidalgo in March, in addition to applications of I. javanica in June on 770, 2,500 and 16,000 ha, for the states of Campeche, Quintana Roo and Yucatán respectively. It is considered that with the applications of these AFI, a reduction in the psyllid population between 60 to 81.8% has been achieved ^{(SENASICA 2018)}.

Another relevant pest is the American locust (Schistocerca spp.), which is considered among the most harmful pests in the world. Their different species devastate thousands of hectares cultivated worldwide, affecting cereals, fruit trees, legumes, and grasses. The biological control of S. piceifrons has been carried out since 2009 using applications of M. acridum in the states of Campeche, Chiapas, Hidalgo, Oaxaca, San Luis Potosí, Tabasco, Tamaulipas, Veracruz, and Yucatán. From 2009 to 2016, applications of the fungus were carried out on 11,102 ha which means an annual average of 1,586 ha treated per year. In 2017 and 2018, 886 and 1,090 ha were treated, respectively ^{(SENASICA-DGSV 2016a)}. Furthermore, the control of grasshoppers of the species Brachystola magna, B. mexicana, Melanoplus differentialis, and Sphenarium purpurascens has been treated with M. acridum in the states of Guanajuato and Tlaxcala with applications on 4,000 and 8,000 ha, respectively since 2011. From 2012 to 2016, applications with M. acridum were carried out on 54,256 ha in the states of Chihuahua, Guanajuato, and Tlaxcala ^{(SENASICA-DGSV 2016b)}. Finally, a campaign from 2009 to 2015 with I. javanica strain CHE-CNRCB 305 to control the brown citrus aphid Toxoptera citricida, which is a severe threat to citrus farming, was implemented on 12,336 ha ^{(Barrera 2020}; ^{SENASICA 2019b)}.

Conclusion

AFI research, production, and application in the field have increased in Mexico since the 1990s. AFI has gained prestige as functional products that control insect pests of fruit, vegetables and cereal crops. Currently, there are 76 commercial products described in this work and 56 products that were not considered in the tables. The most common fungi in AFI are B. bassiana and M. anisopliae, which are mostly formulated in wettable powders. This review represents a database that can be updated periodically, providing the scientific community and Mexican agriculture with a source of information on AFI.