The aquatic plant Wolffia globosa and the probiotic Limosilactobacillus reuteri KUB-AC5 have been proposed as candidate synbiotic components targeting gut microbiota modulation. In this study, we investigated the effects of W. globosa supplementation, alone or in combination with L. reuteri KUB-AC5, on the gut microbiota and health status in healthy adult dogs. For a 28-day feeding trial, 24 healthy dogs were randomly assigned to three dietary treatment groups: control, prebiotic (PRE; 3.5% W. globosa powder), and synbiotic (SYN, 3.5% W. globosa powder combined with L. reuteri KUB-AC5 at 1 × 10⁸ CFU/day). No significant differences were observed in body weight, body condition score, fecal consistency, feed intake, or blood parameters among groups (p > 0.05). Although overall gut microbial diversity remained unchanged, linear discriminant analysis effect size analysis revealed the selective enrichment of specific bacterial taxa. The PRE group exhibited higher relative abundances of Parabacteroides merdae, Romboutsia lituseburensis, Subdoligranulum variabile, and Phocaeicola sartorii, whereas the SYN group showed increased levels of Escherichia coli and Terrisporobacter glycolicus and reduced levels of Anaerobiospirillum succiniciproducens and Campylobacter upsaliensis (p < 0.05). In conclusion, W. globosa, alone or combined with L. reuteri KUB-AC5, was safely used to modulate gut bacterial composition as a potential functional ingredient.