Genomic analysis of a Raoultella ornithinolytica strain causing prosthetic joint infection in an immunocompetent patient

Abstract

We sequenced the genome of Raoultella ornithinolytica strain Marseille-P1025 that caused a rare case of prosthetic joint infection in a 67-year-old immunocompetent male. The 6.7-Mb genome exhibited a genomic island (RoGI) that was unique among R. ornithinolytica strains. RoGI was likely acquired by lateral gene transfer from a member of the Pectobacterium genus and coded for a type IVa secretion system found in other pathogenic bacteria and that may have conferred strain Marseille-P1025 an increased virulence. Strain Marseille-P1025 was also able to infect, multiply within, and kill Acanthamoaeba castellanii amoebae.

Figure 1

Pan-genome analysis of R. ornithinolytica whole-genome sequences. A maximum likelihood tree was constructed from the accessory genome elements (left). The presence (blue) and absence (white) of accessory genome elements is presented on the right.

Figure 2

Comparison of sequences of the scaffold 21 from R. ornithinolytica strain Marseille-P1025 with those of R. ornithinolytica strains NBRC 105727 (A), 2-156-04_S1_C1 (B) and 2-156-04_S1_C2 (C). Figure 2D shows an alignment of all four compared genomes. Common and specific genes are displayed in orange and red, respectively.

Figure 3

Analysis of genomic recombinations in the R. ornithinolytica species based on the alignment of 13 genomes including 12 genomes mapped against that of strain Marseille-P1025, using ClonalFrameML. Recombination events are shown by dark blue horizontal bars. For a given branch, light blue sites mean no substitution. Any other color from white to red indicates a substitution. White indicates non-homoplasic substitutions and the increasing level of redness indicates the increasing degree of homoplasy. The arrow shows recombination events in scaffold 21 where the RoGI genomic island is located.

Figure 4

Transmission electron microscopy of R. ornithinolytica strain Marseille-P1025 using a Morgagni 268D transmission electron microscope (Philips) at an operating voltage of 60 kV. The scale bar represents 2 µm.

Figure 5

Co-culture of R. ornithinolytica and A. castellanii amoebae. (A) Rate multiplication of Raoultella ornithinolytica strains P2310 and Marseille-P1025 within A. castellanii in PAS at 32 °C. (B) Percentage of live A. castellanii infected with R. ornithinolytica strains P2310 and Marseille-P1025. Each bar represents the mean of triplicate wells, and the standard errors are represented by error bars. *P < 0.05.

Figure 6

Optical microscopy observation of A. castellanii trophozoites infected with R. ornithinolytica strain Marseille-P1025 and stained with the Gimenez staining. The presence of R. ornithinolytica was monitored for 3 days: (A) day 0, co-culture after 5 hours of incubation; (B) day1, after 24 hours of incubation; (C) day 2, after 48 hours of incubation; (D) day 3, after 72 hours of incubation.