Collection of Chicken GI Content Samples

The chicken trial performed in this study was approved by the Institutional Animal Care and Use Committee at The University of Tennessee, Knoxville, with protocol number 1387-0219. Briefly, 15 newly hatched broiler chicks (Hubbard Efficiency Plus) were fed with regular feed for 6 weeks. At 2, 4, or 6 weeks of age, five chickens were randomly selected and euthanized. The pH in gizzard and small intestine (the junction site between the duodenum and jejunum) was measured on site using a pH spear tester (Thermo Fisher, MA, USA). Subsequently, the fresh GI contents (5 ml per sample) were collected using clean stainless steel laboratory scoops, transferred into 15 ml sterile plastic tubes, stored on ice, and immediately subjected to downstream work as detailed below.