Attachment and germination of Entomophaga maimaiga conidia on host and non-host larval cuticle Ann E. Hajek * and Callie C. Eastburn Department of Entomology, Cornell University, Comstock Hall, Ithaca, NY 14853-0901, USA Received 15 January 2002; accepted 20 November 2002 Abstract The lepidopteran-specific fungal pathogen Entomophaga maimaiga is highly virulent against Lymantria dispar (gypsy moth) larvae, and other members of the family Lymantriidae. Numerous species in the subfamily Cuculliinae (Family Noctuidae) are not susceptible to E. maimaiga due to the inability of this fungus to penetrate the larval cuticle. Conidial attachment and germination were compared among five cuculliine species and L. dispar using bioassays and scanning electron microscopy. Although conidia were showered evenly across larvae during bioassays, on L. dispar conidia were most abundant on segments, where they adhered well to the cuticle and germinated at high percentages. Conidia on cuculliine cuticles were predominantly found in large, loose aggregations in intersegmental areas. Few conidia on cuculliine cuticle germinated and scanning electron microscopy revealed a thick film of mucous enveloping conidia. We hypothesize that the conidia on cuculliines become coated by this film and were only loosely attached to the larval cuticle. No such film was seen on L. dispar larvae where individual conidia appeared well attached. On L. dispar larvae many conidia also adhered to setae. To determine if hydrophobicity affected the ability of E. maimaiga conidia to attach and germinate on a substrate, a goniometer was used to determine relative hydrophobicity of larval cuticles. L. dispar cuticle was more hydrophobic than cuculliine cuticle, suggesting that a high level of hydrophobicity could be a required characteristic for hosts. Cuticles from four cuculliine species and L. dispar were sequentially extracted using hexane, chloroform, and methanol. Conidia were showered onto glass slides coated with the different extracts and germination was quantified. Methanol extracts of cuculliine cuticle consistently decreased germination, compared to all extracts of L. dispar cuticle. For all L. dispar extracts, the majority of conidia produced germ tubes, which is a normal prerequisite for cuticular penetration. For the cuculliines, conidia exposed to hexane and chloroform extracts produced secondary conidia as did all controls, but the conidia exposed to cuculliine methanol extracts that germinated produced germ tubes. These studies demonstrated that a range of factors act in concert to prevent E. maimaiga infection of the cuculliine species investigated. Ó 2003 Elsevier Science (USA). All rights reserved. Keywords: Entomopathogenic fungi; Entomophthorales; Germination; Adhesion; Cuticle hydrophobicity; Non-target effects; Biological control 1. Introduction Insect cuticle forms the first formidable barrier to pathogens. There are a number of discrete stages leading to breaching the cuticle, including spore attachment, spore germination, spore differentiation (with or without formation of appressoria) and then penetration of the cuticle by enzymatic and mechanical means (St. Leger, 1991). A diversity of factors have been identified as in- fluencing the activity of conidia at the cuticular level. Conidia of many species of entomopathogenic fungi are thought to initially attach nonspecifically (Boucias and Pendland, 1991). The hydrophilic conidia of Entom- ophthorales are covered by a mucilaginous coat that is released upon cuticular attachment and is thought to act as a glue (see Boucias and Pendland, 1991). Surface hydrophobicity has been associated with adhesion of plant pathogenic fungi (Kuo and Hoch, 1996; Terhune and Hoch, 1993). Conidial distribution on host cuticle can be region specific (e.g., Sosa-Gomez et al., 1997) and surface topography has been shown to influence growth of deuteromycetes after adhesion (Boucias and Pend- land, 1991). Chemical components of the cuticle can also affect conidial development after adhesion by either Journal of Invertebrate Pathology 82 (2003) 12–22 Journal of INVERTEBRATE PATHOLOGY www.elsevier.com/locate/yjipa * Corresponding author. Fax: +1-607-255-0939. E-mail address: aeh4@cornell.edu (A.E. Hajek). 0022-2011/03/$ - see front matter Ó 2003 Elsevier Science (USA). All rights reserved. doi:10.1016/S0022-2011(02)00198-2