Two strategies to unravel gene expression responses of hostmicrobe interactions in cod (Gadus morhua) larvae

The commensal bacteria in the intestine play essential roles in the development and functionality of the host. To unravel the host^microbe interactions in Atlantic cod (Gadus morhua L.) larvae, we used two molecular approaches: (1) suppression subtractive hybridizationpolymerase chain reaction (SSH-PCR) to identify host gene responses and (2) expression analysis of selected genes reported to be di¡erentially expressed in gnotobiotic zebra¢sh in a previous study to determine whether these host responses are also conserved in cod. Suppression subtractive hybridization-PCR identi¢ed 156 transcripts putatively regulated by the presence of bacteria. However, out of 22 selected transcripts, only four were signi¢cantly di¡erentially expressed when quanti¢ed using quantitative (real-time) PCR. Expression analysis of selected genes from zebra¢sh revealed possible conservation of host responses for three out of eight genes analysed. For most of the genes quanti¢ed, the gene expression pattern varied between two biological replicates. This may re£ect differences in the bacterial composition in the rearing bottles, and denaturing gradient gel electrophoresis analysis con¢rmed signi¢cant di¡erences between the two replicates with regard to bacterial diversity. The varying e¡ects on gene expression caused by di¡erences in the microbial composition show the necessity of further studies where axenic cod larvae are compared with larvae raised in de¢ned and controlled (gnotobiotic) environments.