PS7-26. Increased Dietary DL-Methionine Supplementation Enhanced Ileal Bacterial Diversity in Diquat-Challenged Growing Pigs.

Abstract Dietary methionine (Met) possesses antioxidant properties and may help maintain intestinal microbiota homeostasis, thereby supporting gut health under oxidative stress conditions. In a previous study, we reported that an intraperitoneal diquat injection significantly altered the relative abundance of certain beneficial bacterial species in the ileum of growing pigs. The present study aimed to evaluate the effects of increased dietary Met supplementation on the ileal bacterial profile of pigs challenged with diquat-induced oxidative stress. Twenty barrows (initial body weight: 21.5 ± 2.50 kg) were randomly assigned to two dietary treatment groups. Group 1 received a corn-soybean meal-based diet formulated to meet or exceed the pigs’ nutrient and energy requirements as recommended by NRC (2012). The standardized ileal digestible (SID) Met + Cys was 0.62% while the SID Lys was 1.0% in this basal diet. Group 2 received the same basal diet but supplemented with 0.16% additional DL-methionine to provide 25% more SID Met + Cys compared to Group 1. At the end of 3 weeks of dietary treatment (as Phase 1), all pigs were intraperitoneally injected with 10 mg of diquat (dissolved in 10 mL saline) per kilogram of body weight. Following the injection challenge, pigs remained on their respective diets for one more week (as Phase 2). At the end of Phase 2, all pigs were euthanized, and ileal digesta samples were collected for microbiota analysis. Full-length 16S rRNA gene amplicon sequencing was performed using the Oxford Nanopore MinION long-read sequencing platform. Generated FASTQ files were analyzed using the Emu database for taxonomic annotation and relative abundance. Alpha and beta diversity indices, including species richness and diversity Shannon index and Bray-Curtis distance, were calculated using the R vegan package. Linear discriminant analysis effect size (LEfSe) was used to identify the taxa differentially enriched between the two groups. Results showed that ileal alpha diversity, as measured by the Shannon index, was greater in Group 2 than in Group 1 (1.91 ± 0.17 vs. 1.24 ± 0.17; P = 0.04). Furthermore, LEfSe analysis revealed that the relative abundances of several bacterial species, including Clostridium butyricum, Clostridium disporicum, Clostridium bornimense, Clostridium sacchariperbutylacetonicum, Clostridium saccharobutylicum, and Paeniclostridium sordellii, were higher in the ileum of the pigs in Group 2 compared with those in Group 1. Clostridium butyricum, in particular, is known as a beneficial butyrate producer species. In conclusion, increased DL-methionine supplementation increased the ileal bacterial alpha diversity and enhanced the relative abundance of some specific bacterial taxa in pigs exposed to diquat-induced oxidative stress, suggesting a potential role for supplemental Met in maintaining intestinal microbial homeostasis and gut health during oxidative stress.