Comparative genomic analysis of Staphylococcus lugdunensis shows a closed pan-genome and multiple barriers to horizontal gene transfer

Abstract

Background: Coagulase negative staphylococci (CoNS) are commensal bacteria on human skin. Staphylococcus lugdunensis is a unique CoNS which produces various virulence factors and may, like S. aureus, cause severe infections, particularly in hospital settings. Unlike other staphylococci, it remains highly susceptible to antimicrobials, and genome-based phylogenetic studies have evidenced a highly conserved genome that distinguishes it from all other staphylococci.

Results: We demonstrate that S. lugdunensis possesses a closed pan-genome with a very limited number of new genes, in contrast to other staphylococci that have an open pan-genome. Whole-genome nucleotide and amino acid identity levels are also higher than in other staphylococci. We identified numerous genetic barriers to horizontal gene transfer that might explain this result. The S. lugdunensis genome has multiple operons encoding for restriction-modification, CRISPR/Cas and toxin/antitoxin systems. We also identified a new PIN-like domain-associated protein that might belong to a larger operon, comprising a metalloprotease, that could function as a new toxin/antitoxin or detoxification system.

Conclusion: We show that S. lugdunensis has a unique genome profile within staphylococci, with a closed pan-genome and several systems to prevent horizontal gene transfer. Its virulence in clinical settings does not rely on its ability to acquire and exchange antibiotic resistance genes or other virulence factors as shown for other staphylococci.

Keywords: CRISPR; Comparative genomics; Core genome; Pan genome; Restriction-modification; Staphylococcus lugdunensis; Toxin/antitoxin.

Fig. 1

Pan-genome and core genome development plot projections for S. lugdunensis (Panel a and b), S. epidermidis (Panel c and d), and S. aureus (Panel e and f). All calculations were done using the EDGAR software platform. For pan-genome development plot extrapolation: the red curve shows the fitted exponential Heaps’ low function, and the blue and green curves indicate the upper and lower boundary of the 95% confidence interval. For Core genome development plot extrapolation: the red curve shows the fitted exponential decay function, and the blue and green curves indicate the upper and lower boundary of the 95% confidence interval

Fig. 2

Phylogenetic tree illustrating genetic relationships between strains. The phylogenetic tree was built based on the complete core genome of the analyzed strains. Alignments of the each individual core gene set were generated using MUSCLE and subsequently concatenated to one large supermatrix. Fasttree was used to infer a maximum likelihood tree from this core gene alignment. Shimodaira-Hasegawa support values. Calculations were made for the three species and show that they form three clearly separated clusters within the phylogenetic tree

Fig. 3

Structural genomic comparison of S. lugdunensis reference strain HKU0901 with the 14 other complete S. lugdunensis genomes. Synteny plots were produced by using EDGAR web server, showing the stop positions of orthologous gene pairs in different genomes. S. lugdunensis HKU09101 genome was compared with the 14 other genomes by splitting the analysis in two for better visual representation (panel a and b). Although some genomes have differing start positions, they all show a high degree of co-linearity, indicating a high level of relatedness and a low level of genomic rearrangement activity within the nalayzed set

Fig. 4

COG functional categories from the core genome of S. lugdunensis, S. aureus, and S. epidermidis strains. Gene lists were predicted using the EDGAR web server, and COG categories obtained by loading them into the WebMGA web server. COG categories are as follows: for cellular processes and signaling, d is cell cycle control, cell division, and chromosome partitioning; m is cell wall/membrane/envelope biogenesis; n is cell motility; o is post-translational modification, protein turnover, and chaperones; t is signal transduction mechanisms; u is intracellular trafficking, secretion, and vesicular transport; v is defense mechanisms; and z is cytoskeleton. For information storage and processing, b is chromatin structure and dynamics; j is translation, ribosomal structure, and biogenesis; k is transcription; and l is replication, recombination, and repair. For metabolism, c is energy production and conversion; e is amino acid transport and metabolism; f is nucleotide transport and metabolism; g is carbohydrate transport and metabolism; h is coenzyme transport and metabolism; i is lipid transport and metabolism; p is inorganic ion transport and metabolism; and q is secondary metabolite biosynthesis, transport, and catabolism. r is for general function prediction only, and s for unknown function

Fig. 5

DR sequences and spacers in type IIIA CRISPR/Cas systems from S. lugdunensis

Fig. 6

Genomic context of the PIN-like domain coding sequence of S. lugdunensis and other staphylococci