Comparative genomics of the Staphylococcus intermedius group of animal pathogens

Abstract

The Staphylococcus intermedius group consists of three closely related coagulase-positive bacterial species including S. intermedius, Staphylococcus pseudintermedius, and Staphylococcus delphini. S. pseudintermedius is a major skin pathogen of dogs, which occasionally causes severe zoonotic infections of humans. S. delphini has been isolated from an array of different animals including horses, mink, and pigeons, whereas S. intermedius has been isolated only from pigeons to date. Here we provide a detailed analysis of the S. pseudintermedius whole genome sequence in comparison to high quality draft S. intermedius and S. delphini genomes, and to other sequenced staphylococcal species. The core genome of the SIG was highly conserved with average nucleotide identity (ANI) between the three species of 93.61%, which is very close to the threshold of species delineation (95% ANI), highlighting the close-relatedness of the SIG species. However, considerable variation was identified in the content of mobile genetic elements, cell wall-associated proteins, and iron and sugar transporters, reflecting the distinct ecological niches inhabited. Of note, S. pseudintermedius ED99 contained a clustered regularly interspaced short palindromic repeat locus of the Nmeni subtype and S. intermedius contained both Nmeni and Mtube subtypes. In contrast to S. intermedius and S. delphini and most other staphylococci examined to date, S. pseudintermedius contained at least nine predicted reverse transcriptase Group II introns. Furthermore, S. pseudintermedius ED99 encoded several transposons which were largely responsible for its multi-resistant phenotype. Overall, the study highlights extensive differences in accessory genome content between closely related staphylococcal species inhabiting distinct host niches, providing new avenues for research into pathogenesis and bacterial host-adaptation.

Keywords: Staphylococcus; animal; antibiotic resistance; genomics; host-adaptation; pathogenesis.

Figure 1

Genome atlas of S. pseudintermedius ED99 compared to other staphylococci. Orthologs depicted in other species are identified by reciprocal best hits using BLASTP. Major features of the S. pseudintermedius ED99 are indicated on the outermost circle, including MGE (blue), CWA proteins (black), and SpPI1 (maroon) with positions on the reference genome indicated in Mb. From the outer to inner circles, circle 2 and 3, represent S. pseudintermedius ED99 genes on plus and minus strands, respectively (colored according to their COG category); circle 4, tRNA (green), rRNA (brown); circle 5, S. delphini 8086 (dark blue); circle 6, S. intermedius NCTC11048 (blue); circle 7, S. aureus Mu50 (red); circle 8, S. epidermidis RP62A (orange); circle 9, S. haemolyticus JCSC1435 (yellow); circle 10, S. saprophyticus ATCC15305 (light green); circle 11, S. carnosus TM300 (dark green); circle 12, G + C content plot (black and gray); and circle 13, GC skew plot (purple and khaki).

Figure 2

Average nucleotide identity (ANI)-based Neighbor-Joining tree of 23 sequenced strains from eight staphylococcal species in addition to an outgroup Macrococcus caseolyticus. The evolutionary relatedness was examined by calculation of ANI, based on core genes only. Species and strains of animal origin are highlighted in blue (scale indicates the % difference ANI).

Figure 3

Comparison of SpPI1 with the major staphylococcal pathogenicity islands. The pathogenicity island SpPI1 from S. pseudintermedius ED99 is compared to SaPI2 from S. aureus RN3984, SaPI4 from S. aureus MRSA252, SaPI3 from S. aureus COL, SaPI1 from S. aureus RN4282, and SaPIbov1 from S. aureus RF122. The similarity between regions is indicated by a spectrum of blue to gray, from 100 to 82% similarity. Genes are colored according to their sequence and function, as described previously (Novick et al., 2010): int (integrase) and xis (excisionase) in yellow; transcription regulators in dark blue; replication genes including the primase gene, pri, and the replication initiator gene, rep in purple; encapsidation genes in green; terminase small subunit gene (terS) in light green; superantigen and other accessory genes in pink; pif (related to phage interference) in light blue; and genes encoding hypothetical proteins in orange. Figure produced with EasyFig (Sullivan et al., 2011).

Figure 4

Comparison of the CRISPR loci of S. pseudintermedius ED99 and S. intermedius NCTC 11048 with related Nmeni and Mtube types. The CRISPR locus subtype Nmeni of S. pseudintermedius ED99 and S. intermedius NCTC 11048 are compared to those of Streptococcus M1 GAS and Ruminococcus lactaris ATCC 29176. The CRISPR locus subtype Mtube of S. intermedius NCTC 11048 is compared to those of S. epidermidis RP62A and S. lugdunensis HKU09-01. The similarity between regions is indicated by a spectrum of blue to gray colors depicting a range of 100–30% similarities. Genes of the csn family are shown in purple, the cas family in green, and the csm family in light blue. Of note, the csm6 gene in S. lugdunensis HKU09-01 appears to be a pseudogene. Black boxes represent clusters of direct repeats regions with the number of spacers between the direct repeats indicated. The red box at the beginning of each Nmeni subtype CRISPR locus corresponds to a single copy of the repeats identified in the cluster of downstream direct repeats. The gray boxes indicate the approximate number of direct repeats at the locus in S. intermedius NCTC 11048 (due to the unfinished status of the genome). Figure produced with EasyFig (Sullivan et al., 2011).

Figure A1

Comparison of the genomic island containing the bi-component leukotoxin Luk-I and the L-ascorbate transport and utilization operon in S. pseudintermedius ED99 and S. delphini 8086. The similarity between regions is indicated by a spectrum of blue to gray, ranging from 100 to 82% similarity. Genes are colored according to their sequence and function as follow: phage-related genes in red; leukotoxin genes in pink; putative phosphoglucomutase in dark green; L-ascorbate transport and utilization genes in green; putative transcriptional regulator in gray; tRNA genes are represented by black boxes; and genes encoding hypothetical proteins in orange.