# Buccal Swab Samples from Japanese Brown Cattle Fed with Limonite Reveal Altered Rumen Microbiome

## TL;DR

Feeding limonite to pregnant Japanese Brown cattle in Japan changes their rumen microbiome, reducing core bacteria for roughage digestion and increasing non-core bacteria.

## Contribution

This study reveals how limonite feeding alters the rumen microbiome in pregnant Japanese Brown cattle.

## Key findings

- Limonite feeding reduced core-rumen bacteria involved in roughage digestion.
- Non-core-rumen bacteria increased in the limonite-fed group.
- Beta-diversity analysis showed significant differences between limonite and control groups.

## Abstract

Farmers in the Mount Aso area, a southern district of Japan, raise the Kumamoto strain of Japanese Brown (Japanese brown) cattle on its grassland and often supplement their feed with Aso limonite, a mineral derived from deposited volcanic ash and magma, particularly for pregnant animals. However, the mechanisms associated with limonite’s beneficial effects have not been characterized. In this analysis, groups of Japanese brown pregnant cattle were either fed with limonite or without (control), and buccal swab samples were collected every 30 days for 90 days. DNA extracted from buccal swab samples was then subjected to a 16S rRNA gene sequence analysis, identifying core-rumen, non-core-rumen, as well as oral, bacteria. Limonite feeding reduced the proportion of core-rumen bacteria focused primarily on roughage digestion, instead promoting the non-core-rumen microbiome.

The areas of the Mount Aso grasslands in Kumamoto, Japan, are the primary location for the breeding of the Kumamoto strain of Japanese Brown cattle (JBRK). Although Aso limonite, deposited by volcanic ash and magma, has been commonly fed to pregnant JBRK in this area, the mechanisms of its salutary effects on pregnant JBRK have not yet been elucidated. Approximately 100 days before the expected day of calf delivery, seven JBRK (four supplemented with limonite and three controls without limonite) were assigned to this study, from which a buccal swab was collected at the highest rumination every 30 days for 90 days. DNA extracted from these swabs was then analyzed using a 16S rRNA gene amplicon sequence analysis. Statistically significant differences between the two groups were discovered through beta-diversity analysis, though results from alpha-diversity analysis were inconclusive. The microbiota identified were classified into six clusters, and three of the main clusters were core-rumen bacteria, primarily cellulose digestion in cluster 1, oral bacteria in cluster 2, and non-core-rumen bacteria in cluster 3. In the limonite group, core-rumen bacteria decreased while non-core-rumen bacteria increased, suggesting that limonite feeding alters rumen microbiota, particularly activation of non-core-rumen microbiota.