Metagenomic and metabolic analysis of the rumen in cattle submitted to heat stress conditions

Abstract

Heat stress is a physiological condition resulting from an animal’s inability to dissipate heat to maintain its homeothermy. Changes in bovine metabolism may induce alterations in the ruminal microbiota and fermentative process, affecting its productivity. The objective of this project was to study prokaryote metagenomes and the metabolic profile in the rumen of beef cattle subjected to comfort and heat stress conditions. Six pure Zebu cattle, cannulated in the rumen, were used in the experiment. The animals were confined in bioclimatic chambers equipped with ventilators, exhausters, air conditioners, infrared lamps and humidity controllers. Six treatments were tested in a latin square design 6 × 6 with a 2 × 2 + 2 factorial arrangement, containing two factors (temperature and diet energy concentration) and two additional treatments, in order to obtain means of comparison of the temperature effect on the proposed variables without consumption level interference. The two proposed temperatures represent thermoneutral and heat stress conditions (34º C from 6 am to 6 pm and 24ºC from 6 am to 6 pm). Total DNA was extracted from each collected sample using a phenol-chloroform protocol. The total DNA was sequenced by the next -generation sequencing platform MiSeq Sequencing System (Illumina), and the diversity index of each sample was determined. The sequences taxonomic classification was determined using Geneious 10.2.3 bioinformatics software. Acetic, propionic, butyric, valeric, isobutyric and isovaleric acids were quantified by high performance liquid chromatography. Gas chromatography-mass spectrometry was used to characterise the compound profile present in the ruminal environment. Sequences were classified into 80 different genera, belonging to 10 phyla. In all treatments, Firmicutes and Bacteroidetes represented more than 70% of all prokaryotes identified. The phyla Verrucomicrobia and Spirochaetes and the genera Flavonifractor, Treponema and Ruminococcus presented a reduction (P<0.03) in their population when the cattle were submitted to high temperatures. The genus Saccharibacteria (incertae sedis), belonging to a phylum of uncultivated bacteria (Candidatus Saccharibacteria), presented higher (P=0.05) population in animals under thermal stress conditions. The concentration of isobutyric acid was lower (P<0.01) in the rumen of bovine under heat stress conditions. A total of 45 different compounds were identified in the ruminal samples. There was no standard compound profile related to heat stress. The amine dihydro-5-pentyl-2 (3H)-f uranone was identified in 100% of the samples following heat stress treatment. The heat stress caused alterations in the ruminal microbiota composition and also in the rumen fermentative process.