Incidental prophylactic surgeries are performed in certain situations. Incidental prophylactic appendectomies were common practice within opened bariatric surgeries. The gut microbiota has emerged as an important actor within the homeostasis of the host. A new hypothesis has been formulated about the appendix function in relation to gut microbiota. Our objective was to study the gut microbiota profiles of patients that had suffered from an incidental prophylactic appendectomy during their bariatric surgeries, while comparing them to patients whose appendixes had remained intact. A case-control observational prospective study of 40 patients who underwent bariatric surgery, with or without an incidental prophylactic appendectomy, during 2004–2008 with an evaluation of their gut microbiota populations at the end of 2016 was conducted by sequencing the 16 S rRNA gene by Next Generation Sequencing of patients’ stools and appendix tissues. Patients with their appendix removed showed lower levels of richness and diversity of their gut microbiota populations. Odoribacter, Bilophila, Butyricimonas, and Faecalibacterium levels were increased in the Intact group, while Lachnobacterium suffered an expansion in the group without the appendix. Moreover, a linear regression model introduced the concept that Butyricimonas and Odoribacter may be implicated in insulin regulation. Thus, gut microbiota should be considered in the decisions of practical surgery, regarding the appendix as a mediator of homeostasis in the host. Butyricimonas and Odoribacter require further investigation as key bacteria implicated in insulin regulation.