In vitro Inoculation of Fresh or Frozen Rumen Fluid Distinguishes Contrasting Microbial Communities and Fermentation Induced by Increasing Forage to Concentrate Ratio.

Zhi Yuan Ma,Si Yu Yi,Min Wang,Ju Wang Zhou,Zhi Liang Tan

doi:10.3389/fnut.2021.772645

Abstract

In vitro rumen batch culture is a technology to simulate rumen fermentation by inoculating microorganisms from rumen fluids. Although inocula (INO) are commonly derived from fresh rumen fluids, frozen rumen fluids are also employed for the advantages of storing, transporting, and preserving rumen microorganisms. The effects of frozen INO on microbial fermentation and community may be interfered with by substrate type, which has not been reported. This study was designed to test whether rumen fluid treatments (i.e., fresh and frozen) could interact with incubated substrates. A complete block design with fractional arrangement treatment was used to investigate the effects of INO (fresh or frozen rumen fluids) and concentrate-to-forage ratios (C/F, 1:4 or 1:1) on rumen fermentation and microbial community. The effects of increasing C/F were typical, including increased dry matter (DM) degradation and total volatile fatty acids (VFA) concentration (P < 0.001), and decreased acetate to propionate ratio (P = 0.01) and bacterial diversity of richness and evenness (P ≤ 0.005) with especially higher fermentative bacteria such as genus Rikenellaceae_RC, F082, Prevotella, Bacteroidales_BS11, Muribaculaceaege, and Christensenellaceae_R-7 (P ≤ 0.04). Although frozen INO decreased (P < 0.001) DM degradation and altered rumen fermentation with lower (P ≤ 0.01) acetate to propionate ratio and molar proportion of butyrate than fresh INO, typical effects of C/F were independent of INO, as indicated by insignificant INO × C/F interaction on substrate degradation, VFA profile and bacterial community (P ≥ 0.20). In summary, the effect of C/F on fermentation and bacterial diversity is not interfered with by INO type, and frozen INO can be used to distinguish the effect of starch content.

Highlights

In vitro rumen batch culture is a technology to stimulate rumen fermentation
Elevated concentrateto-forage ratio (C/F) increased dry matter (DM) degradation and gas production (P ≤ 0.001; Table 1), which agrees with many previous studies [2, 12, 28]
INO × C/F interaction was not observed for feed degradation expressed as DM degradation and gas production per g DM degraded, indicating independent effects of INO and C/F

Summary

INTRODUCTION

In vitro rumen batch culture is a technology to stimulate rumen fermentation. The advantage of this technique is to reduce the cost of animal and related animal welfare issues by testing multiple samples in batch [1]. Strong correlations of DM degradation have been observed between fresh and frozen INO [10], evidence indicates that frozen INO can alter the fermentation pattern with decreased VFA production and shift of fermentation to favor propionate production [11] Such changed rumen fermentation may interact with incubated types of substrates, an effect that has not been investigated. The contrasting substrates used were the increased concentrateto-forage ratio (C/F), which is characterized by increased DM degradation, shifted fermentation through propionate production, and changed microbial community [2, 12, 13] We hypothesized that both fresh and frozen INO showed a similar pattern concerning the effects of increasing dietary C/F on gas production, fermentation, and bacterial communities. Interaction between INO and C/F (INO × C/F) was analyzed

MATERIALS AND METHODS

RESULTS AND DISCUSSION

ETHICS STATEMENT