Intestinal bacteria composition and prebiotics may play a role in the management of type 2 diabetes mellitus (T2DM). The objective of this systematic review was to evaluate the effect of prebiotics and substances with prebiotic properties on the metabolic and inflammatory biomarkers of individuals with T2DM compared with placebo. A literature search to identify articles published up to March 31, 2018, was conducted utilizing PubMed, Science Direct, and Cochrane Central Register of Controlled Trials. Individuals at aged 18 years or older with T2DM from randomized controlled trials investigating prebiotics or substances with prebiotic properties were included. Metabolic and inflammatory biomarkers associated with T2DM were the primary outcome measures. Twenty-seven publications were analyzed. All but seven of these publications reported a beneficial effect on metabolic and/or inflammatory biomarkers. Interventions included mostly women, lasted 4 days to 12 weeks, and diabetes duration ranged from 6 months to 11 years. Nineteen publications reported improvements in glycemia, 15 in cardiovascular markers, nine in body weight, and nine in inflammatory markers. Benefits from resistant starch, resistant dextrin, and oligofructose-enriched inulin were most frequent. A smaller number of studies utilizing other substances with prebiotic properties also yielded improvements. Based on these results, there is fair evidence that prebiotics and substances with prebiotic properties may improve metabolic and inflammatory biomarkers related to T2DM in women aged 18 years at least. Interventions with resistant starch, resistant dextrin, and oligofructose-enriched inulin exhibited the strongest evidence for improvements due to the quantity of publications and quality grades. Other prebiotics and substances with prebiotic properties show promise but the number of studies is few. Additional studies that are longer in duration, include both sexes, and include other prebiotics or substances with prebiotic properties are needed. Copyright © 2020 Academy of Nutrition and Dietetics. Published by Elsevier Inc. All rights reserved.