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Unlabelled: Abstract Introduction: Minimally invasive surgery in children with long-segment intestinal aganglionosis aims to reduce the number of abdominal wall incisions. Conventional laparoscopic and laparoendoscopic single-site (LESS) surgeries fulfill this goal. In children, natural orifice translumenal endoscopic surgery (NOTES™; American Society for Gastrointestinal Endoscopy [Oak Brook, IL] and Society for American Gastrointestinal and Endoscopic Surgeons [Los Angeles, CA]) has been limited because of fear of access site complications. We present a novel technique of totally transanal LESS pull-through colectomy (TLPC), avoiding abdominal wall incision, which combines LESS technology and the NOTES approach., Subjects and Methods: Two boys and one girl (2.5 months, 6 months, and 5 years of age, respectively) with sigmoid and transverse colon aganglionosis underwent surgery. The TLPC procedure consisted of an endorectal technique with submucosal dissection starting 1 cm orally from the dentate line to above the peritoneal reflection, where the rectal muscle was divided circumferentially. After ligation of the rectal mucosa, the proximal bowel was replaced into the abdominal cavity, and a TriPort(®) (Olympus Surgical Technologies Europe, Hamburg, Germany) was introduced transanally. Mesenterial resection of the aganglionic bowel was accomplished via transanal LESS until the normoganglionic colon segment was reached and pulled down to the site of anastomosis. After removal of the port, a conventional pull-through procedure was performed., Results: All children displayed normal bowel movements and were complication-free during the follow-up period of up to 7 months., Conclusions: TLPC combines the minimally invasive LESS surgery with the scarless concept of NOTES and allows resection of long-segment aganglionosis without abdominal incision. TLPC is a safe, effective, and feasible surgical procedure in children with long-segment intestinal aganglionosis.