Denature Gradient Gel Electrophoresis Microbiota Profiling

We initially analyzed microbiota community profiles using DGGE (Denature Gradient Gel Electrophoresis). A 457-bp fragment from the V6 to V8 region of the bacterial 16S rDNA gene was amplified with primers U968-GC and L1401. DGGE was performed in an 8% (wt/vol) polyacrylamide gel with a denaturing gradient ranging from 40 to 50% at the top and bottom of the gel, respectively (100% denaturing conditions were defined as 7 M urea and 40% formamide). After electrophoresis (16 h, 65 V, 60°C), the gel was stained with SYBER Green (Novex, San Diego, CA, United States). DGGE analysis was performed only for F.O.P. bacteriophage therapy, ampicillin therapy, and PBS therapy groups for Days 0, 1, 5, and 10. We compared the degree of microbiota distortion between treatment groups by comparing banding patterns indicative of microbiota diversity within each group and evaluating levels of band conservation. Distortion for both F.O.P. therapy and ampicillin therapy were compared against the PBS control group. We did not evaluate microbiome profiles for the EcoShield PX^TM bacteriophage therapy or the diluted F.O.P. 1:10 bacteriophage therapy.