Colitis is a significant factor that poses a threat to animal health and welfare, and its association with the gut microbiota has been extensively studied. However, there is limited information available on the changes in gut microbial composition in cats with colitis. Here, we investigated the gut microbial composition and variability of cats during colitis. Our findings indicated that Firmicutes, Actinobacteriota, and Proteobacteria were abundant in all the samples. However, there were significant changes in the gut microbial diversity and composition. Analysis of microbial diversity revealed a significant decrease in gut microbial alpha diversity in cats with colitis, along with notable changes in the gut microbial structure. Bacterial taxonomic analysis demonstrated an obvious increase in the relative abundance of 1 phylum (Firmicutes) and 10 genera (Acidithiobacillus, Candidatus_Bacilloplasma, Shewanella, Ferrimicrobium, Sellimonas, unclassified_Butyricicoccaceae, V9D2013_group, unclassified_Euzebyaceae, UCG_002 and Providencia), while the relative abundance of 24 phyla and 475 genera decreased significantly during colitis. Remarkably, 11 phyla and 370 genera completely disappeared among the reduced taxa. Overall, this study demonstrates that colitis can lead to significant alterations in the diversity and composition of the gut microbiota in cats. Moreover, these findings contribute to our understanding of the underlying causes of colitis from the perspective of the gut microbiota and provide a theoretical basis for the development of prevention and treatment strategies.