Characterization of Nomuraea rileyi strains using polymorphic DNA, virulence and enzyme activity

Abstract

The characterization of entomopathogenic microorganisms is important for the selection of more effective strains for use in integrated pest-control programs. Five Nomuraea rileyi strains (SA86101, GU87401, SR86151, CG128 and VA9101) were characterized using random amplified polymorphic DNA (RAPD) analysis, virulence studies and assessment of chitinolytic and proteolytic activity. RAPD analysis divided the strains into two groups with a similarity coefficient of 0,76%, group 1 consisting of strains SA86101, GU87401 and SR86151 and group 2 of strains CG128 and VA9101. The LT50 varied from 165h with strain VA9101 to 246h with strain GU87401. Chitinolytic and proteolytic activity of the fungi after 144h growth in minimal medium were tested using colloidal chitin as substrate. All strains exhibited enzyme activity, with strain VA9101 having the highest chitinase activity (0,0040 mumol/mL/min the 40ºC) and strain SA86101 the highest proteolytic activity. No relationship was found between RAPD analysis, virulence and chitinase or protease activity.<br>Caracterizaram-se cinco linhagens de N. rileyi por análise de RAPD, avaliação da virulência e atividade enzimática (quitinolítica e proteolítica). Pela técnica de RAPD foi possível distinguir, entre as cinco linhagens avaliadas, dois grupos com similaridade 0,76: grupo I (linhagens SA86101, GU87401, SR86151) e grupo II (linhagens VA9101 e CG128). O TL50 variou de 165 horas com a utilização da linhagem VA9101 a 246 horas com GU87401. Todas as linhagens apresentaram atividade quitinolítica e proteolítica, destacando-se quanto à produção de quitinases a linhagem VA9101 (com 0,0040 mimol/mL/min a 40ºC) e em relação à protease, a linhagem SA86101. Não foi possível evidenciar relações entre virulência, análise genética e atividade enzimática

Genotypic analysis of Nomuraea rileyi collected from various noctuid hosts

“…In order to better understand the pathogen-host relationships of this mycopathogen, various molecular methods have been used to delineate strains of N. rileyi. For example, DNA extracted from diVerent pools of N. rileyi isolates have been subjected to random ampliWed polymorphic DNA (RAPD) (Tigano and Aljanabi, 2000;Vargas et al, 2003), inter-simple sequence repeat (ISSR) analysis (Han et al, 2002) and ampliWed fragment length polymorphism (AFLP) (Boucias et al, 2000a,b). Vargas et al (2003) were unable to link the results of RAPD analysis conducted on a series of N. rileyi isolates to phenotypic traits (virulence, enzyme levels).…”

“…For example, DNA extracted from diVerent pools of N. rileyi isolates have been subjected to random ampliWed polymorphic DNA (RAPD) (Tigano and Aljanabi, 2000;Vargas et al, 2003), inter-simple sequence repeat (ISSR) analysis (Han et al, 2002) and ampliWed fragment length polymorphism (AFLP) (Boucias et al, 2000a,b). Vargas et al (2003) were unable to link the results of RAPD analysis conducted on a series of N. rileyi isolates to phenotypic traits (virulence, enzyme levels). Tigano and Aljanabi (2000) using RAPDs were able to clearly delineate N. rileyi into two phenetic groups that were associated with the insect host.…”

Molecular ecology of fungal entomopathogens: molecular genetic tools and their applications in population and fate studies

“…Resulting products are separated by gel electrophoresis, which provides RAPD-banding profiles that allow to analyze presence or absence of bands. This profiling technique has been widely employed to study intraspecific variation within species like Entomophaga grylli (Entomophthoromycotina: Entomophthorales) (Bidochka et al 1995), E. muscae (Jensen et al 2001), P. neoaphidis (Rohel et al 1997;Nielsen et al 2001;Tymon and Pell 2005), Zoophthora phytonomi (Entomophthoromycotina: Entomophthorales) (Hajek et al 1996), Z. radicans (Hodge et al 1995), B. bassiana (Maurer et al 1997;Fernandes et al 2006), B. brongniartii (Cravanzola et al 1997Piatti et al 1998), Hirsutella thompsonii (Ascomycota: Hypocreales) (Mozes-Koch et al 1995;Aghajanzadeh et al 2007), L. lecanii (Mor et al 1996), M. anisopliae (Fegan et al 1993;Leal et al 1994;Velásquez et al 2007), N. rileyi (Boucias et al 2000Vargas et al 2003), and Paecilomyces farinosus (=Isaria farinosus) (Ascomycota: Hypocreales) (Chew et al 1998). Studies have focused for instance on associations of fungal genotypes with specific hosts (Hodge et al 1995;Bridge et al 1997;Maurer et al 1997;Jensen et al 2001) or on correlations between RAPD profiles and geographical origin of a species (Leal et al 1994;Hajek et al 1996;Boucias et al 2000;Nielsen et al 2001).…”

Pathogenicity of Native Entomopathogenic Fungus Nomuraea rileyi Against Spodoptera litura

“…The full potential of entomopathogenic fungi has not been approached. However, previous studies indicate that the insect mycopathogen N. rileyi a dimorphic hypomycete causes fungal epizootics in population of several noctuid pests (14,28,31). Moreover, previous bioassays have shown that N. rileyi isolates from different geographical locations and different hosts vary in their virulence and specificity (3,12,30,34).…”