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SELECTION of STRAINS of Beauveria Bassiana and Metarhizium Anisopliae (ASCOMYCOTA: HYPOCREALES) for ENDOPHYTIC COLONIZATION in COCONUT SEEDLINGS

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SELECTION of STRAINS of Beauveria Bassiana and Metarhizium Anisopliae (ASCOMYCOTA: HYPOCREALES) for ENDOPHYTIC COLONIZATION in COCONUT SEEDLINGS Chilean J. Agric. Anim.Gaviria Sci., et ex al. Agro-Ciencia Strains of B. bassiana(2020) 36(1): y M.3-13. anisopliae in coconut sedlings 3 ISSN 0719-3882 print ISSN 0719-3890 online SELECTION OF STRAINS OF Beauveria bassiana AND Metarhizium anisopliae (ASCOMYCOTA: HYPOCREALES) FOR ENDOPHYTIC COLONIZATION IN COCONUT SEEDLINGS Jackeline Gaviria 1*, Pedro Pablo Parra 2, Alonso Gonzales 3 1 Corporación Colombiana de Investigación Agropecuaria (AGROSAVIA), Diagonal a la intersección de la Carrera 36A con Calle 23, Palmira, Colombia. 2 Tropical Research & Education Center, University of Florida, Homestead, FL 33031-3314, USA. 3 AGO Consulting, Cali, Colombia. * Corresponding author E-mail: [email protected] ABSTRACT Beauveria bassiana and Metarhizium anisopliae are considered virulent pathogens of the coconut weevil Rhynchophorus palmarum (Linnaeus). The objective of this study was to determine the ability of B. bassiana (Beauveriplant SBb36) and M. anisopliae (JGVM1) to establish an endophytic relationship with coconut Cocos nucifera (Linnaeus) seedlings. Strains were selected based on the mortality of adults of R. palmarum exposed to these fungi. Three methods of inoculation were used to inoculate the seedlings obtained through seed germination: foliar spray, stem injection and drench to the roots. Immersion of seedlings in a conidial suspension was used to inoculate seedlings obtained from tissue culture. Colonization was determined through the re-isolation of the fungi four weeks after inoculation. Beauveriplant SBb36 and JGVM1 colonized endophytically 100% of the seedlings obtained through tissue culture and 91.6% of seedlings obtained from germinated seeds. For plants inoculated by immersion with B. bassiana, the colonization rate in petioles (43%) was higher than in leaves and roots, 14 and 17%, respectively. Metarhizium anisopliae did not show significant differences in the colonization rate in leaves, petioles and roots. Plants inoculated by foliar spraying showed that colonization was higher in leaves > petioles > stems > roots. Colonization in stem injection was higher in stems than in the roots. Soil drench inoculation was higher in roots > stems. To our knowledge, this is the first report describing the endophytic relationship that B. bassiana and M. anisopliae establish with coconut seedlings obtained through seed germination and tissue culture when targeted inoculations are performed. Key words: Biological control, entomopathogens, endophyte, coconut palm, seeds, tissue culture. RESUMEN Beauveria bassiana y Metarhizium anisopliae son considerados patógenos virulentos contra el picudo Rhynchophorus palmarum. El objetivo de este estudio fue determinar la habilidad de la cepa comercial Beauveriplant SBb36 de B. bassiana y la cepa JGVM1 de M. anisopliae, para establecer una relación endofítica con plantas de coco Cocos nucifera (Linnaeus) obtenidas a partir de semilla sexual y de cultivo de tejidos. Se evaluaron diferentes métodos de inoculación: aspersión foliar, inyección en tallo y aplicación de suspensión conidial en las raíces, fueron usados para inocular plantas obtenidas a partir de semilla sexual, mientras que el método de inmersión fue usado para inocular plantas obtenidas a partir de cultivo de tejidos. El porcentaje de colonización en diferentes tejidos de las plantas se determinó a través del re-aislamiento de los hongos cuatro semanas después de la inoculación. Las cepas Beauveria bassiana (Beauveriplant SBb36) y Metarhizium anisopliae (JGVM1) colonizaron endofíticamente el 100% de las plantas de coco obtenidas a partir de cultivo Received: 02 October 2019 Accepted: 09 December 2019 4 Chilean J. Agric. Anim. Sci., ex Agro-Ciencia (2020) 36(1):3-13. de tejidos y el 91.6% de las plantas de coco obtenidas por semilla sexual. En el presente estudio, se reporta por primera vez que B. bassiana and M. anisopliae establecen una relación de endofitismo con plántulas de coco obtenidas a partir de la germinación de semilla sexual y cultivo de tejidos a través de inoculaciones dirigidas. Palabras clave: Control biológico, entomopatógenos, endófitos, palma de coco, semillas, cultivo de tejidos. INTRODUCTION highly impractical because of the height of palms (Aldana et al., 2011), development of resistance The coconut palm Cocos nucifera (Linnaeus) against pesticides, environmental pollution (Arecaceae) is an industrial crop from which and high costs to farmers. An alternative to several and highly valuated products with reduce the damage caused by larval feeding important nutritional composition (Manivannan and transmission of the nematode is the et al., 2018), such as oil, milk, cream, fibers, biological control of larval and adult stages of activated carbon, and coconut water, are the insect by using entomopathogenic fungus obtained. It is also an important source of food like B. bassiana (Bals.) Vuill and M. anisopliae and income for small-scale farmers around (Metsch) Sorokin (Ascomycota: Hypocreales). the world. In Colombia, coconut is especially Despite entomopathogenic fungi are used as important for low-income Afro-Colombian biopesticides, this alternative requires recurrent communities along the Pacific coast. It is grown applications and their efficacy depend on under very special conditions (influenced by the environmental conditions, such as temperature, tide) where no other agricultural crop is able humidity, radiation, among other factors to grow, becoming almost the only alternative (O’Callaghan et al., 2001). Other biocontrol for these communities. Despite its importance, alternatives that do not have these restrictions are the coconut production in the Pacific region has ideal in plantations that require labor-intensive decreased dramatically due to the American management practices, such as mass trapping palm weevil (APW) Rhynchophorus palmarum and pesticide applications. (Linnaeus). This insect is the most important The use of entomopathogenic fungi as coconut pest in many countries of Latin America endophytes may play an important role in plant and the Caribbean (Löhr et al., 2015). The APW protection against pest and diseases (Jaber and attacks not only coconut palms but also a large Ownley, 2017). They could be the most suitable number of cultivated and wild palms. alternative to combat larval stages found inside In Colombia R. palmarum is widely distributed plants (Azevedo et al., 2000). Fungal species like along different coconut and oil palm growing B. bassiana and M. anisopliae are often used in the zones (Aldana et al., 2011). It is the main vector biological control of pests in agricultural crops of Bursaphelenchus cocophilus (Cobb) (Aldana et (Vega et al., 2008; Jaronski, 2010). al., 2011), the nematode associated with the red The fungus B. bassiana is the best studied ring disease (RRD), which is lethal to coconut endophytic fungal entomopathogen (Ownley and other palm species. In addition, APW larval et al., 2008; Jaber, 2015; Vidal and Jaber, 2015; stages cause direct damage to palms by feeding Greenfield et al., 2016). This fungus has been at the base of petioles, inflorescences and stem. inoculated in corn (Zea mays) to control Ostrinia The larval feeding causes palm weakening and nubilalis (Hübner) (Wagner and Lewis, 2000), in sometimes palms die because of severe larval date palm to control red date scale Phoenicococcus damage on meristematic tissue (Oehlschlager et marlatti (Cockerell) (Gómez-Vidal et al., 2006), al., 2002; Lohr et al., 2015). in coffee to control the coffee berry borer Management strategies against the APW Hypothenemus hampei (Ferrari) (Posada et al., have shown limited success, mainly because this 2007), and in cocoa to control the cocoa pod insect develops most of its life cycle inside the borer Conopomorpha cramerella (Snellen) (Posada plant tissues (Aldana et al., 2011). This behavior and Vega, 2005). In adittion, B. bassiana has been makes detection and eradication difficult to inoculated in banana to control the banana weevil achieve. Cultural practices such as mass trapping Cosmopolites sordidus (Germar), showing a damage (Oehlschlager et al., 2002) and eradication of Red reduction of 50% caused during immature stages Ring Diseased palms (Aldana et al., 2011) are of banana plants (Musa spp.) obtained from tissue currently the most commonly used management culture (Akello, 2008). strategies in Colombia. However, these practices Another fungus used as endophyte is M. are costly and labor intensive. The use of anisopliae; this fungus has the ability to colonize chemical pesticides for the APW management is rhizomes and roots of plants, such as cabbage Gaviria et al. Strains of B. bassiana y M. anisopliae in coconut sedlings 5 (Brassica oleracea), fir (Abies), tomato (Solanum commercial sources and reactivated on adults of lycopersicum), corn (Zea mays) (Hu and Leger, R. palmarum (Akello et al., 2009). The reactivation 2002; Bruck, 2005; Garcia et al., 2011), broad bean was carried by diluting commercial strains in a (Aphis fabae) (Akello and Sikora, 2012), and cassava 0.1% Tween-80 solution to obtain a concentration (Manihot esculenta) (Greenfield et al., 2016). When of 1x108 conidia mL-1. Five adult females of R. M. anisopliae was found in corn as an endophyte, palmarum were immersed into the suspension for it promoted plant development, increased seed five minutes and then placed individually inside germination rate and protected the plant against a 90 mm sterile Petri plate with sterile moist Coleopteran pests (Kabaluk et al., 2007; Garcia et filter
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