WITHDRAWN: Temporal Disruption and Recovery of Gut Microbiota During Chemotherapy: A Metagenomic Perspective

Background Chemotherapy remains indispensable in oncology but frequently causes gastrointestinal toxicity through gut-microbiota disruption. The microbiome is now recognized as a determinant of immune regulation, metabolism, and therapeutic response. A cytidine analogues used in several solid tumours can exacerbate intestinal dysbiosis, whereas dietaries flavonoid with antioxidant and anti-inflammatory properties has been proposed to restore microbial balance. Understanding these reciprocal interactions is vital for improving cancer-therapy tolerance and outcomes. To investigate the temporal alterations of gut-microbiota composition induced by gemcitabine and to assess whether quercetin modulates or mitigates chemotherapy-associated dysbiosis. Controlled animal experimentation conducted under approved ethical standards. Thirty-six female Wistar rats were randomly assigned to six groups (Control, Gemcitabine (Gem), Quercetin 25 (Q25), Quercetin 100 (Q100), G+Q25, G+Q100). Treatments lasted 7 days. Faecal samples were collected at baseline and on days 3 and 8. Full-length 16S rRNA gene sequencing (Oxford Nanopore GridION) provided taxonomic profiling from phylum to species level. Alpha-diversity indices (Shannon, Chao1, Simpson) and beta-diversity (weighted UniFrac, PCoA, UPGMA) were computed in QIIME2. Group differences were analysed using PERMANOVA and Fisher’s exact test for categorical variables (SPSS v26, p ≤0.05). Mortality and diarrhoea were recorded clinically. Results: Quercetin monotherapy was well tolerated, showing no diarrhoea or deaths. Gemcitabine caused severe gastrointestinal toxicity with diarrhoea by day 5 and 50 % mortality by day 7. Co-administration with quercetin failed to prevent toxicity, particularly in Gem+Q25, where early diarrhoea and sequential deaths occurred. Metagenomic analysis revealed marked reductions in microbial richness and evenness in all gemcitabine-containing groups (p≤0.05). Phylum-level data showed an elevated Firmicutes/Bacteroidetes ratio and enrichment of Proteobacteria signatures of dysbiosis. At the genus level, Lactobacillus depletion was accompanied by expansions of Enterococcus, Escherichia, and Fusobacterium. Species-level profiling identified a hierarchical ecological shift: loss of Lactobacillus johnsonii and Turicibacter sp. , overgrowth of Enterococcus hirae , Escherichia coli , and Fusobacterium nucleatum , and partial compensatory rise of Allobaculum spp. Quercetin alone preserved control-like communities but, when combined with gemcitabine, did not fully counteract dysbiosis. Beta-diversity (PCoA, UPGMA) confirmed distinct clustering of gemcitabine-treated samples from control and quercetin groups, reflecting profound structural disruption of microbial communities. Conclusion: Gemcitabine induces severe microbiota dysbiosis characterized by depletion of protective commensals and proliferation of opportunistic pathogens, correlating with diarrhoea and mortality. Quercetin monotherapy maintains microbial stability but offers very limited protection when combined with gemcitabine. These findings underscore the clinical need to monitor and preserve gut-microbiota integrity during chemotherapy. Incorporating microbiota-directed or high-dose antioxidant-based adjuncts may enhance treatment tolerance and reduce gastrointestinal complications.