Identification of immunorelevant genes from greater wax moth (Galleria mellonella ) by a subtractive hybridization approach V. Seitz a, * , A. Clermont b , M. Wedde c , M. Hummel b , A. Vilcinskas c , K. Schlatterer d , L. Podsiadlowski e a Max-Planck Institute Molecular Genetics, Ihnestrasse 73, 14195 Berlin, Germany b Institute of Pathology, Free University, Hindenburgdamm 30, 12200 Berlin, Germany c Systematic Zoology and Evolutionary Biology, University of Potsdam, Villa Liegnitz, Lenne `str. 7a, 14417 Potsdam, Germany d Institute of Clinical Chemistry and Pathobiochemistry, Free University, Hindenburgdamm 30, 12200 Berlin, Germany e Institute of Zoology, Free University, Ko ¨nigin-Luise Str. 1–3, 14195 Berlin, Germany Received 17 July 2002; revised 12 September 2002; accepted 15 September 2002 Abstract In this study we have analyzed bacterial lipopolysaccharide (LPS) induced genes in hemocytes of the Lepidopteran species Galleria mellonella using subtractive hybridization, followed by suppressive PCR. We have found genes that show homologies to molecules, such as gloverin, peptidoglycan recognition proteins and transferrin known to be involved in immunomodulation after bacterial infection in other species. In addition, a few molecules previously not described in the innate immune reactions were detected, such as a RNA binding molecule and tyrosine hydroxylase. Furthermore, the full-length cDNA of a LPS-induced molecule with six toxin-2-like domains is described to be a promising candidate to further elucidate the relationship between toxin- and defensin-like domains in arthropod host defense. q 2002 Elsevier Science Ltd. All rights reserved. Keywords: Galleria mellonella; Lipopolysaccharide; Subtractive hybridization; Real-time PCR; Innate immunity; Gloverin; Peptidoglycan recognition protein; Toxin-2 1. Introduction Most immunological data for insects have been reported from Drosophila melanogaster [1]. In the pre-genome age immunomodulations were studied at the protein level in relatively large insects such as the Lepidopteran species Manduca sexta, Hyalophora cecropia, Bombyx mori and Galleria mellonella, as the larger body size facilitated the collection of haemolymph for protein purification [2–7].In G. mellonella various immunorelevant protein mol- ecules were described [6,8–11]. However, compared to Drosophila melanogaster only scarce molecular data are available. To fill this gap and to build a broader basis for understanding the evolution of the immune system, we have conducted a molecular approach for which no molecular pre-information is needed and which can also easily be applied to other 0145-305X/03/$ - see front matter q 2002 Elsevier Science Ltd. All rights reserved. PII: S0145-305X(02)00097-6 Developmental and Comparative Immunology 27 (2003) 207–215 www.elsevier.com/locate/devcompimm * Corresponding author. Tel.: þ49-30-8413-1267; fax: þ 49-30- 8413-1385. E-mail address: seitz_v@molgen.mpg.de (V. Seitz). Abbreviations: LPS, lipopolysaccharide; PCR, polymerase chain reaction; aa, amino acids; PGRP, peptidoglycan recognition protein.