THE AMB2 LOCUS FROM SERRATIA-ENTOMOPHILA CONFERS ANTI-FEEDING EFFECT ON LARVAE OF COSTELYTRA-ZEALANDICA (COLEOPTERA, SCARABAEIDAE)

Serratia entomophila (Se) causes amber disease in the soil-dwelling pe st, Costelytra zealandica (Cz). The disease presents two main signs: a nti-feeding effect (AFE) and development of amber coloration (AC). To identify the genetic loci involved in pathogenicity, non-pathogenic (P ath(-)) Se mutants were created by transposon (TnphoA) mutagenesis [Up adhyaya et al., J. Bacteriol. 174 (1992) 1020-1028]. The mutant UC24 l ost the ability to produce amber disease signs and it was shown to con tain a single TnphoA insertion. The TnphoA insertion site was mapped i n a 5.3-kb DNA fragment, which was named amb2 locus. Cosmids containin g amb2 fully restored AFE and partially restored AC in UC24. Escherich ia coli (Ec) HB101 bearing the amb2 locus was able to cause AFE in a m ultiple-dose bioassay. SDS-PAGE analysis of the amb2 gene products pro duced in minicells showed the synthesis of two proteins of 16 and 19.5 kDa, named AnfA and AnfB. The genes encoding these proteins were mapp ed by deletion analysis. Pathogenicity tests with insect larvae fed wi th bacteria carrying the anfA and anfB gene regions separately showed that both regions are essential for AFE. It is proposed that the AnfA and AnfB proteins are virulence factors (toxin-like proteins) causing AFE in Cz.