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Brevibacterium from Austrian hard cheese harbor a putative histamine catabolism pathway and a plasmid for adaptation to the cheese environment.

ABSTRACT: The genus Brevibacterium harbors many members important for cheese ripening. We performed real-time quantitative PCR (qPCR) to determine the abundance of Brevibacterium on rinds of Vorarlberger Bergkäse, an Austrian artisanal washed-rind hard cheese, over 160 days of ripening. Our results show that Brevibacterium are abundant on Vorarlberger Bergkäse rinds throughout the ripening time. To elucidate the impact of Brevibacterium on cheese production, we analysed the genomes of three cheese rind isolates, L261, S111, and S22. L261 belongs to Brevibacterium aurantiacum, whereas S111 and S22 represent novel species within the genus Brevibacterium based on 16S rRNA gene similarity and average nucleotide identity. Our comparative genomic analysis showed that important cheese ripening enzymes are conserved among the genus Brevibacterium. Strain S22 harbors a 22 kb circular plasmid which encodes putative iron and hydroxymethylpyrimidine/thiamine transporters. Histamine formation in fermented foods can cause histamine intoxication. We revealed the presence of a putative metabolic pathway for histamine degradation. Growth experiments showed that the three Brevibacterium strains can utilize histamine as the sole carbon source. The capability to utilize histamine, possibly encoded by the putative histamine degradation pathway, highlights the importance of Brevibacterium as key cheese ripening cultures beyond their contribution to cheese flavor production.

SUBMITTER: Anast JM

PROVIDER: S-EPMC6467879 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundBrevibacterium strains are widely used for the manufacturing of surface-ripened cheeses, contributing to the breakdown of lipids and proteins and producing volatile sulfur compounds and red-orange pigments. The objective of the present study was to perform comparative genomic analyses in order to better understand the mechanisms involved in their ability to grow on the cheese surface and the differences between the strains.ResultsThe genomes of 23 Brevibacterium strains, including twelve strains isolated from cheeses, were compared for their gene repertoire involved in salt tolerance, iron acquisition, bacteriocin production and the ability to use the energy compounds present in cheeses. All or almost all the genomes encode the enzymes involved in ethanol, acetate, lactate, 4-aminobutyrate and glycerol catabolism, and in the synthesis of the osmoprotectants ectoine, glycine-betaine and trehalose. Most of the genomes contain two contiguous genes encoding extracellular proteases, one of which was previously characterized for its activity on caseins. Genes encoding a secreted triacylglycerol lipase or involved in the catabolism of galactose and D-galactonate or in the synthesis of a hydroxamate-type siderophore are present in part of the genomes. Numerous Fe3+/siderophore ABC transport components are present, part of them resulting from horizontal gene transfers. Two cheese-associated strains have also acquired catecholate-type siderophore biosynthesis gene clusters by horizontal gene transfer. Predicted bacteriocin biosynthesis genes are present in most of the strains, and one of the corresponding gene clusters is located in a probable conjugative transposon that was only found in cheese-associated strains.ConclusionsBrevibacterium strains show differences in their gene repertoire potentially involved in the ability to grow on the cheese surface. Part of these differences can be explained by different phylogenetic positions or by horizontal gene transfer events. Some of the distinguishing features concern biotic interactions with other strains such as the secretion of proteases and triacylglycerol lipases, and competition for iron or bacteriocin production. In the future, it would be interesting to take the properties deduced from genomic analyses into account in order to improve the screening and selection of Brevibacterium strains, and their association with other ripening culture components.

Project description:Lactococcus garvieae is a lactic acid bacterium found in raw-milk dairy products as well as a range of aquatic and terrestrial environments. The plasmids in L. garvieae have received little attention compared to those of dairy Lactococcus lactis, in which the genes carried by these extrachromosomal elements are considered of adaptive value. The present work reports the sequencing and analysis of the plasmid complement of L. garvieae IPLA 31405, a strain isolated from a traditional, Spanish, starter-free cheese made from raw-milk. It consists of pLG9 and pLG42, of 9,124 and 42,240 nucleotides, respectively. Based on sequence and structural homology in the putative origin of replication (ori) region, pLG9 and pLG42 are predicted to replicate via a theta mechanism. Real-time, quantitative PCR showed the number of copies per chromosome equivalent of pLG9 and pLG42 to be around two and five, respectively. Sequence analysis identified eight complete open reading frames (orfs) in pLG9 and 36 in pLG42; these were organized into functional modules or cassettes containing different numbers of genes. These modules were flanked by complete or interrupted insertion sequence (IS)-like elements. Among the modules of pLG42 was a gene cluster encoding specific components of a phosphoenolpyruvate-phosphotransferase (PEP-PTS) system, including a phospho-β-galacosidase. The cluster showed a complete nucleotide identity respect to that in plasmids of L. lactis. Loss of pLG42 showed this to be involved in lactose assimilation. In the same plasmid, an operon encoding a type I restriction/modification (R/M) system was also identified. The specificity of this R/M system might be broadened by different R/M specificity subunits detected in pLG9 and in the bacterial chromosome. However, challenges of L. garvieae IPLA 31405 against L. lactis phages proved that the R/M system was not involved in phage resistance. Together, these results support the hypothesis that, as in L. lactis, pLG42 contribute towards the adaptation of L. garvieae to the dairy environment.

Project description:In this study, we combined metabolic reconstruction, growth assays, metabolome and transcriptome analyses to obtain a global view of the sulfur metabolic network and of the response to sulfur availability in Brevibacterium aurantiacum. In agreement with the growth of B. aurantiacum in the presence of sulfate and cystine, the metabolic reconstruction showed the presence of a sulfate assimilation pathway and of thiolation pathways that produce cysteine (cysE and cysK) or homocysteine (metX and metY) from sulfide, of at least one gene of the transsulfuration pathway (aecD) and of genes encoding three MetE-type methionine synthases. We also compared the expression profiles of B. aurantiacum ATCC9175 during sulfur starvation and in the presence of sulfate, cystine or methionine plus cystine. In sulfur starvation, 690 genes including 21 genes involved in sulfur metabolism and 29 genes encoding amino acids and peptide transporters were differentially expressed. We also investigated changes in pools of sulfur-containing metabolites and in expression profiles after growth in the presence of sulfate, cystine or methionine plus cystine. The expression of genes involved in sulfate assimilation and cysteine synthesis was repressed in presence of cysteine, while the expression of metX, metY, metE1, metE2 and BL613 encoding a probable cystathionine-γ-synthase decreased in the presence of methionine. We identified three ABC transporters: two stronger transcribed during cysteine limitation and one during methionine depletion. Finally, the expression of genes encoding a methionine γ-lyase, BL929, and a methionine transporter (metPS) was induced in the presence of methionine, in conjunction with a significant increase of volatile sulfur compounds production. Refer to individual Series. This SuperSeries is composed of the following subset Series: GSE25418: BA-Methionine plus Cystine vs Cystine GSE25419: BA-Sulfate vs Cystine GSE25420: BA-Methionine plus Cystine vs Sulfate GSE25421: BA-Sulfate vs Sulfate starvation

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Brevibacterium from Austrian hard cheese harbor a putative histamine catabolism pathway and a plasmid for adaptation to the cheese environment.

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