Host immunity and the colon microbiota of mice infected with Citrobacter rodentium are beneficially modulated by lipid-soluble extract from late-cutting alfalfa in the early stages of infection.

K Fries-Craft,J M Anast,E A Bobeck,S Schmitz-Esser,Juan J Loor

doi:10.1371/journal.pone.0236106

Abstract

Alfalfa is a forage legume commonly associated with ruminant livestock production that may be a potential source of health-promoting phytochemicals. Anecdotal evidence from producers suggests that later cuttings of alfalfa may be more beneficial to non-ruminants; however, published literature varies greatly in measured outcomes, supplement form, and cutting. The objective of this study was to measure body weight, average daily feed intake, host immunity, and the colon microbiota composition in mice fed hay, aqueous, and chloroform extracts of early (1st) and late (5th) cutting alfalfa before and after challenge with Citrobacter rodentium. Prior to inoculation, alfalfa supplementation did not have a significant impact on body weight or feed intake, but 5th cutting alfalfa was shown to improve body weight at 5- and 6-days post-infection compared to 1st cutting alfalfa (P = 0.02 and 0.01). Combined with the observation that both chloroform extracts improved mouse body weight compared to control diets in later stages of C. rodentium infection led to detailed analyses of the immune system and colon microbiota in mice fed 1st and 5th cutting chloroform extracts. Immediately following inoculation, 5th cutting chloroform extracts significantly reduced the relative abundance of C. rodentium (P = 0.02) and did not display the early lymphocyte recruitment observed in 1st cutting extract. In later timepoints, both chloroform extracts maintained lower splenic B-cell and macrophage populations while increasing the relative abundance of potentially beneficially genera such as Turicibacter (P = 0.02). At 21dpi, only 5th cutting chloroform extracts increased the relative abundance of beneficial Akkermansia compared to the control diet (P = 0.02). These results suggest that lipid soluble compounds enriched in late-cutting alfalfa modulate pathogen colonization and early immune responses to Citrobacter rodentium, contributing to protective effects on body weight.

Highlights

Phytochemicals are a class of natural compounds that may improve livestock health through direct and indirect action on the host immune system and intestinal microbiota [1, 2]
At 21dpi, only 5th cutting chloroform extracts increased the relative abundance of beneficial Akkermansia compared to the control diet (P = 0.02). These results suggest that lipid soluble compounds enriched in late-cutting alfalfa modulate pathogen colonization and early immune responses to Citrobacter rodentium, contributing to protective effects on body weight
No body weight (BW) differences were observed during the feed enrichment period (Fig 1)

Summary

Introduction

Phytochemicals are a class of natural compounds that may improve livestock health through direct and indirect action on the host immune system and intestinal microbiota [1, 2]. One dietary phytochemical source is alfalfa, a documented source of bioactive compounds associated with a number of health benefits such as saponins [3, 4], phytoestrogens/flavonoids [5, 6], and non-cellulosic polysaccharides [7, 8]. Dietary alfalfa inclusion for nonruminant livestock is limited due to the high indigestible fiber content, reducing dietary energy density. Anecdotal evidence from swine producers suggests that feeding late-cutting alfalfa may confer greater health benefits to sows and piglets compared to earlier cuttings. The use of alfalfa extracts can possibly circumvent fiber-associated limitations while still conferring immunological benefits. Crude aqueous alfalfa extract and protein-rich concentrate increased lymphocyte proliferation while chloroform extracts improved mouse survival during LPS challenge and suppressed production of pro-inflammatory interleukin (IL)-6 in vitro [15,16,17,18]

Objectives

Methods

Results