The effect of silkworms ( Bombyx mori ) chitosan on rumen fermentation, methanogenesis, and microbial population in vitro

Ruminant enteric methane (CH ) is one of the largest sources of greenhouse gases that contribute to global warming. To minimize environmental harm caused by ruminants' CH production, natural substances can be used to suppress it. Chitosan from crustacean sources had been known to obstruct CH generation in the rumen. About 18% of silkworm pupae is chitin, but little is known about the impact of silkworm pupae chitosan on rumen methanogenesis. This study investigated the efficacy of the silkworm chitosan extraction method and its impact on rumen fermentation, methanogenesis, and microbial growth . This study employed a randomized complete block design featuring five treatments and four batches for rumen incubation as the blocking factor. In this study, five treatments were implemented: Control (CO) (basal diet with no added chitosan), basal diet with 6% chitosan from the Chinese Silkworm strain 804 (CHI804), basal diet with 6% chitosan from the PS 01 Hybrid Silkworm strain (CHIPS01), basal diet with 6% chitosan from the Hybrid F1 Japanese 102 × Chinese 202 races (CHIJC02), and basal diet with 6% commercial shrimp shell chitosan as the positive control (CHICOMM). The experiments assessed digestibility, pH, total gas generation, CH production, ammonia nitrogen (NH -N), and short-chain fatty acid levels, along with microbial population. Data were analyzed using a general linear model followed by Duncan's test when applicable. A significant effect on dry matter digestibility (DMD), total gas production, CH , NH -N, and rumen microbial populations (Methanogens, , , , , , spp., and spp.) was observed (p < 0.05). The extracted chitosan (CHIJC02) used in this study exhibited a similar quality to that of commercial chitosan (CHICOMM). CHI804 treatment could reduce gas production, NH -N production, and population significantly (p < 0.05), while CHIJC02 could reduce CH production, methanogen population, acetate (C ) production, and increase propionate (C ) production significantly (p < 0.05). CHIJC02 and CHICOMM treatments could also increase the population of , , and spp. significantly (p < 0.05). Chitosan addition significantly (p < 0.05) reduced DMD but did not impact organic matter digestibility or pH. The extracted chitosan mimics commercial chitosan in physico-chemical properties. Chitosan derived from Japanese and Chinese F1 hybrid silkworm strains demonstrated superior capacity for inhibiting CH generation compared to commercial chitosan. The quality and effect