The pea aphid Acyrthosiphon pisum has a worldwide distribution and causes serious losses for agricultural production. Drunken horse grass Achnatherum inebrians is a widely distributed perennial poisonous grass on the grasslands of Northern and Northwestern China. The present study focused on contact toxicity activity of aqueous extracts of endophyte-infected (E+) and endophyte-free (E−) A. inebrians in different growth periods of pea aphids, and the growth and development of two color morphs of F1 generation nymphs. Both of the color morphs had development durations in E+ treatments that tended to be longer at 1st, 2nd, 3rd, and 4th instars than E− and control (CK). The E+ treated aphids also showed decreased weights at maturity with over all lower mean relative growth rates (MRGR). Aphid survival of E+ treated aphids was lower than that of E− and CK at all growth periods. Seeding stage E+ extracts showed a greater propensity for negatively affecting aphids than did E+ extract at maturity and the yellowing stage. These results show that extracts from endophyte-containing plants may contain compounds that may be used to control insects.
Achnatherum inebrians, a perennial grass, is widely distributed in China. When infected by the endophyte Epichloë gansuensis, A. inebrians produces an abundance of alkaloids that enhance plant survival but are toxic to animals. Here we used in vitro fermentation to study the impact of endophyte-infected A. inebrians (E+) addition on rumen fermentation characteristics and on microbial community and diversity as assessed with amplicon sequencing technology. We examined E+ addition at five levels, E0, E25, E50, E75, and E100, corresponding to 0%, 25%, 50%, 75%, and 100% of the fermentation substrate, respectively. Both the fermentation characteristics and rumen microbial community structure differed significantly among treatments. E100 resulted in the highest values for pH, the Shannon index, Kiritimatiellaeota, and Lentisphaerae levels relative to the other treatments. In contrast, E25 was associated with higher levels of ammonia nitrogen, total volatile fatty acid, propionate, butyrate, isobutyrate, valerate, of the phyla Bacteroidetes and Firmicutes, and of the genus Prevotella_1, Succiniclasticum, Family_XIII_AD3011_group, Rikenellaceae_RC9_gut_group, Prevotellaceae_UCG-001, and Pyramidobacter as compared with other treatments. E50 resulted in the greatest values for the abundance-based coverage estimator (ACE) and the Chao1 index as compared with other treatments. E0 resulted in the greatest values for digestibility of dry matter, gas production, acetate, and Ruminobacter as compared with other treatments. This approach avoided animal toxicity experiments and confirmed that rumen fermentation characteristics and rumen microbiota were affected by E+ toxin. Therefore, E25 showed higher abundance in Prevotella_1, Prevotellaceae_ UCG-001, and Lachnospiraceae_XPB1014_group that implied they should play significant roles in E+ alkaloids degradation. And then, we can infer that rumen microorganisms should function as an antidote with respect to this poisoning reaction at moderate dietary percentages of E+.
Endophytic fungal infection is the major reason for intoxication of animals caused by drunken horse grass. Fortunately, it has been established that seed detoxification techniques and isolation of endophytic fungi infect non-endophytic fungi populations with the same genetic background as endophyte-infected Achnatherum inebrians. Moreover, sheep can use endophyte-free Achnatherum inebriants (EF) without obvious toxicity symptoms. The present study selected EF as a representative grass, consisting of five different replacement levels, EF0, EF25, EF50, EF75, and EF100, corresponding to 0%, 25%, 50%, 75%, and 100% of the fermentation substrate, respectively. Simultaneously, in vitro fermentation and the 16S rRNA amplicon sequencing method was used to explore the effect of EF on sheep ruminal fermentation and microbial diversity. The results revealed that EF100 had the highest values for pH, acetate: propionate, the Patescibacteria, Kiritimatiellaeota, and Synergistetes phylum levels, Ruminococcaceae, Prevotellaceae, and Saccharofermentans genus levels than the other treatments (p < 0.05). In contrast, EF25 was associated with higher levels of abundance-based coverage estimator (ACE), Chaol index of the phyla Synergistetes and Bacteroidetes, and of the genus Erysipelotrichaceae, Rikenellaceae, and Prevotella as compared with other treatments (p < 0.05). EF50 resulted in the greatest values for the genus Christensenellaceae and Lachnospiraceae as compared with other treatments (p < 0.05). EF75 resulted in the greatest values for the Shannon index as compared with other treatments (p < 0.05). EF0 resulted in the greatest values for gas production (GP), ammonia nitrogen (NH3-N), total volatile fatty acid (TVFA), acetate, propionate, butyrate, valerate, isobutyrate, isovalerate, and the phyla Firmicutes, Proteobacteria, and Spirochaetes, and the genus Succiniclasticum, Ruminobacter, Family_XIII and Treponema as compared with other treatments (p < 0.05). PICRUSt2 analysis indicated that most of the functional prediction pathways were involved in Carbohydrate metabolism and, Amino acid metabolism. Therefore, the recommended ratio of EF in sheep diet should range from 25% to 50%, and the maximum proportion should not exceed 75%.
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