Abstract
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.
Highlights
Lactococcus garvieae is a lactic acid bacterium (LAB) that was first isolated from the udder of a cow with mastitis [1]
L. lactis strains from non-dairy sources contain plasmids with genes that allow complex polysaccharides to be utilised and different metals to be taken up [13, 16]; these properties are unimportant for bacteria growing in milk
The plasmid complement-pLG9 and pLG42- of L. garvieae IPLA 31405 was examined at the molecular level and the results deposited in the GenBank database
Summary
Lactococcus garvieae is a lactic acid bacterium (LAB) that was first isolated from the udder of a cow with mastitis [1]. Human L. garvieae infections are rare, but cases of opportunistic endocarditis and spondylodiscitis have been reported [4] This microorganism is found in many farmhouse dairy products manufactured from raw milk, from which it is occasionally retrieved as a majority component of their native microbiota [5,6,7,8]. Dairy and environmental strains of Lactococcus lactis can carry a complex extrachromosomal complement consisting of many plasmids [9, 10]; these are thought to provide genes of use in adaptation to new environments [11,12,13]. L. lactis strains from non-dairy (plant) sources contain plasmids with genes that allow complex polysaccharides to be utilised and different metals to be taken up [13, 16]; these properties are unimportant for bacteria growing in milk
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