Effect of some feed additives on the gut microbiota of rainbow trout (Oncorhynchus mykiss)
Five fish were randomly collected from each tank (n=45) and processed individually for 16S rRNA gene-based profiling of fish gut bacteriomes. A five cm-long portion of the distal intestine ending with the rectum was excised aseptically, the intestine was washed with sterile saline to remove surface-associated contaminants and then digesta was gently removed by squeezing with a sterile scalpel. The intestinal material was collected into microcentrifuge tubes with 2 ml RNAlater solution and then stored at -20C until gDNA extraction. Intestine samples (approx. 200 mg wet weight) were thawed and homogenized with a tissue homogenizer and gDNA was extracted from the homogenates using PureLink Genomic DNA Mini Kit. The extracted gDNA was used as template and partial 16S rRNA-gene amplicons were obtained with universal bacteria primers. PCR-products were purified with AMPure XP beads and library preparation and indexing were performed with TG Nextera XT Index Kit v2 Set A and Nextera XT DNA Library Preparation Kit, respectively. Sequencing was performed, with the paired-end (PE) 2x150 bp option, on the Illumina Miseq platform.