Metagenomic analysis reveals the linkages between bacteria and the functional enzymes responsible for potential ammonia and biogenic amine production in alfalfa silage

Abstract

Aims:To clarify the molecular mechanisms underlying ammonia (NH 3 ) and biogenic amines (BAEs) formation in alfalfa silage, whole metagenomic sequencing analysis was performed to identify the linkages between functional bacteria and their responsible enzymes in alfalfa silage prepared with and without sucrose addition. Methods and Results:Genes encoding nitrite reductase (nirB) resulting in NH 3 formation were the most abundant and were mostly assigned to Enterobacter cloacae and Klebsiella oxytoca. Putrescine-related genes, classified mainly to encode ornithine decarboxylase (odcA), were predominantly carried by Escherichia coli, Ent. cloacae and Citrobacter sp. Escherichia coli and Kl. oxytoca were the important species responsible for cadaverine and tyramine formation. Ent. cloacae, E. coli, and Kl. oxytoca dominated the bacterial community in naturally fermented alfalfa silage, whilst sucrose-treated silages greatly inhibited the growth of these species by promoting the dominance of Lactobacillus plantarum, thus decreasing the concentrations of NH 3 , cadaverine, putrescine and tyramine.Conclusions: Enterobacteriaceae bacteria are mainly responsible for the NH 3 , putrescine, cadaverine and tyramine formations in alfalfa silage. Significance and Impact of the Study:Whole metagenomic sequencing analysis served as a useful tool to identify the linkages between functional bacteria and associated enzymes responsible for NH 3 and BAEs formation.

Effects of Cellulase and Lactiplantibacillus plantarum on the Fermentation Parameters, Nutrients, and Bacterial Community in Cassia alata Silage

“…In the early stages of silage, the dominant genus was Klebsiella , and the subdominant genera were Lactobacillus, Weissella , and Acetobacter . Li et al ( 2021 ) found that the genes involved in NH 3 formation were mainly classified as encoding nitrite reductase (nirB) and distributed mainly in Klebsiella . Klebsiella is an acid-intolerant facultative anaerobe that can grow well in anoxic environmental conditions using fermentation substrates (Stewart, 1988 ).…”

Effects of intrinsic tannins on proteolysis dynamics, protease activity, and metabolome during sainfoin ensiling

“…However, NH 4 + -N content in silage formed by deamination from FAA is mainly attributed to bacterial enzyme activity rather than plant enzyme activity ( McDonald et al, 1991 ). Metagenomic analysis revealed that NH 4 + -N contents were associated with nitrite reductase, with the gene encoding nitrite reductase mainly assigned to Enterobacter ( Li R.R. et al, 2021 ).…”

“…Enterococcus exhibit greater activity during the early fermentation stages of silage, with Leuconostocs, Pediococci , and Lactococci involved in the initiation of silage fermentation through lactic acid production; however, all the bacteria above are replaced gradually by Lactobacillus with the prolongation of ensiling time ( Cai and Benno, 1998 ). Additionally, Enterococcus could transform FAA to produce AN and bioamines through different metabolic pathways through corresponding deaminases and decarboxylases ( Oladosu et al, 2016 ); for example, lysine decarboxylase forms cadaverine, tyrosine decarboxylase forms tyramine, and arginine decarboxylase forms arginine ( Li R.R. et al, 2021 ).…”

The need for an integrated multi‐OMICs approach in microbiome science in the food system

“…By coupling RNA‐seq with metabolomics, it was also possible to see that perturbations during food processing modify the function of the microbiome. For example, process temperature during cheese ripening (De Filippis et al., 2016), fruit ripening (Li et al., 2021; Xu et al., 2019), plant‐based fermentation (Kim et al., 2021), and vinegar production (Wu et al., 2021) regulates the metabolic activity and modifies the gene expression of the microbiome with important changes in volatilome profile of the final products. Apart from observational studies showing how process parameters can modify the microbiome with important changes in the final sensorial characteristic of the products, the integration of RNA‐seq with metabolomics offers the possibility to study the mechanism of bacteria acid‐and‐ethanol tolerance directly connected with the quality characteristics and ammonia flavor formation in fermented fish (Zhao & Eun, 2020), the identification of microbes responsible for producing biogenic amines during soy sauce fermentation (Kim et al., 2021), or bacterial metabolism under high salinity‐induced osmotic stress (Chun et al., 2019).…”