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Pathophysiological Mechanisms of Staphylococcus Non-aureus Bone and Joint Infection: Interspecies Homogeneity and Specific Behavior of S. pseudintermedius.

Maali Y, Martins-Simões P, Valour F, Bouvard D, Rasigade JP, Bes M, Haenni M, Ferry T, Laurent F, Trouillet-Assant S - Front Microbiol (2016)

Bottom Line: Compared to S. aureus, S. pseudintermedius was the only species that significantly adhered to human fibronectin.These results suggest the involvement of β1 integrin in the invasion process, although this mechanism was previously restricted to S. aureus.In summary, our results suggest that internalization into NPPCs is not a classical pathophysiologic mechanism of SNA BJIs.

View Article: PubMed Central - PubMed

Affiliation: Centre International de Recherche en Infectiologie, INSERM U1111, CNRS UMR5308, Université de Lyon 1, ENS de Lyon, Team "Pathogenesis of staphylococcal infections"Lyon, France; Department of Clinical Microbiology, Northern Hospital Group, Hospices Civils de LyonLyon, France.

ABSTRACT
Implicated in more than 60% of bone and joint infections (BJIs), Staphylococci have a particular tropism for osteoarticular tissue and lead to difficult-to-treat clinical infections. To date, Staphylococcus aureus internalization in non-professional phagocytic cells (NPPCs) is a well-explored virulence mechanism involved in BJI chronicity. Conversely, the pathophysiological pathways associated with Staphylococcus non-aureus (SNA) BJIs have scarcely been studied despite their high prevalence. In this study, 15 reference strains from 15 different SNA species were compared in terms of (i) adhesion to human fibronectin based on adhesion microplate assays and (ii) internalization ability, intracellular persistence and cytotoxicity based on an in vitro infection model using human osteoblasts. Compared to S. aureus, S. pseudintermedius was the only species that significantly adhered to human fibronectin. This species was also associated with high (even superior to S. aureus) internalization ability, intracellular persistence and cytotoxicity. These findings were confirmed using a panel of 17 different S. pseudintermedius isolates. Additionally, S. pseudintermedius internalization by osteoblasts was completely abolished in β1 integrin-deficient murine osteoblasts. These results suggest the involvement of β1 integrin in the invasion process, although this mechanism was previously restricted to S. aureus. In summary, our results suggest that internalization into NPPCs is not a classical pathophysiologic mechanism of SNA BJIs. S. pseudintermedius appears to be an exception, and its ability to invade and subsequently induce cytotoxicity in NPPCs could explain its severe and necrotic forms of infection, notably in dogs, which exhibit a high prevalence of S. pseudintermedius infection.

No MeSH data available.


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Fibronectin adhesion and internalization of 17 S. pseudintermedius clinical isolates into MG63 osteoblasts.(A) Adhesion of the isolates to human fibronectin was evaluated by measuring the absorbance at 620 nm normalized to the reference strain S. aureus. Plots and means were derived from three experiments performed in triplicate. The difference in the capacity of the SNA strains to adhere to fibronectin compared to DU5883 Δfnb S. aureus was assessed using the Mann–Whitney one-tailed test with an α risk of 0.05 (∗∗p < 0.01, ∗∗∗p < 0.001; OD: optical density). (B) Capacity of internalization of S. pseudintermedius clinical isolates into MG63 osteoblasts. Bars represent means ± standard deviation derived from two experiments performed in triplicate, and the results are expressed as percentages relative to the initial inoculum († = 0.01 ± 0.01%).
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Figure 2: Fibronectin adhesion and internalization of 17 S. pseudintermedius clinical isolates into MG63 osteoblasts.(A) Adhesion of the isolates to human fibronectin was evaluated by measuring the absorbance at 620 nm normalized to the reference strain S. aureus. Plots and means were derived from three experiments performed in triplicate. The difference in the capacity of the SNA strains to adhere to fibronectin compared to DU5883 Δfnb S. aureus was assessed using the Mann–Whitney one-tailed test with an α risk of 0.05 (∗∗p < 0.01, ∗∗∗p < 0.001; OD: optical density). (B) Capacity of internalization of S. pseudintermedius clinical isolates into MG63 osteoblasts. Bars represent means ± standard deviation derived from two experiments performed in triplicate, and the results are expressed as percentages relative to the initial inoculum († = 0.01 ± 0.01%).

Mentions: The original results observed for the reference strain S. pseudintermedius led us to investigate whether it’s particularly high abilities to adhere to fibronectin and internalize into osteoblasts was species-specific and consequently observed for other S. pseudintermedius isolates. Sixteen out of the 17 tested clinical S. pseudintermedius isolates adhered significantly more frequently than DU5883 Δfnb S. aureus (24.87 ± 2.90%; p < 0.01). The adhesion rate of S. pseudintermedius isolates varied from 28.54 ± 1.7% to 109.74 ± 2.28% compared to S. aureus 8325-4, which was used as a reference (100%; Figure 2A). In addition, 14 out of the 15 gentamicin-susceptible isolates exhibited higher internalization and persistence rates than DU5883 Δfnb S. aureus based on the gentamicin protection assay (p < 0.01 – Figure 2B and Supplementary Figure S1). Concerning infection-induced cytotoxicity, 14 out of 15 isolates harbored a comparable phenotype to the reference strain, and cells infected with these isolates exhibited a significantly higher cytotoxicity rate (p < 0.001) than uninfected cells (100%; Supplementary Figure S1).


Pathophysiological Mechanisms of Staphylococcus Non-aureus Bone and Joint Infection: Interspecies Homogeneity and Specific Behavior of S. pseudintermedius.

Maali Y, Martins-Simões P, Valour F, Bouvard D, Rasigade JP, Bes M, Haenni M, Ferry T, Laurent F, Trouillet-Assant S - Front Microbiol (2016)

Fibronectin adhesion and internalization of 17 S. pseudintermedius clinical isolates into MG63 osteoblasts.(A) Adhesion of the isolates to human fibronectin was evaluated by measuring the absorbance at 620 nm normalized to the reference strain S. aureus. Plots and means were derived from three experiments performed in triplicate. The difference in the capacity of the SNA strains to adhere to fibronectin compared to DU5883 Δfnb S. aureus was assessed using the Mann–Whitney one-tailed test with an α risk of 0.05 (∗∗p < 0.01, ∗∗∗p < 0.001; OD: optical density). (B) Capacity of internalization of S. pseudintermedius clinical isolates into MG63 osteoblasts. Bars represent means ± standard deviation derived from two experiments performed in triplicate, and the results are expressed as percentages relative to the initial inoculum († = 0.01 ± 0.01%).
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4940379&req=5

Figure 2: Fibronectin adhesion and internalization of 17 S. pseudintermedius clinical isolates into MG63 osteoblasts.(A) Adhesion of the isolates to human fibronectin was evaluated by measuring the absorbance at 620 nm normalized to the reference strain S. aureus. Plots and means were derived from three experiments performed in triplicate. The difference in the capacity of the SNA strains to adhere to fibronectin compared to DU5883 Δfnb S. aureus was assessed using the Mann–Whitney one-tailed test with an α risk of 0.05 (∗∗p < 0.01, ∗∗∗p < 0.001; OD: optical density). (B) Capacity of internalization of S. pseudintermedius clinical isolates into MG63 osteoblasts. Bars represent means ± standard deviation derived from two experiments performed in triplicate, and the results are expressed as percentages relative to the initial inoculum († = 0.01 ± 0.01%).
Mentions: The original results observed for the reference strain S. pseudintermedius led us to investigate whether it’s particularly high abilities to adhere to fibronectin and internalize into osteoblasts was species-specific and consequently observed for other S. pseudintermedius isolates. Sixteen out of the 17 tested clinical S. pseudintermedius isolates adhered significantly more frequently than DU5883 Δfnb S. aureus (24.87 ± 2.90%; p < 0.01). The adhesion rate of S. pseudintermedius isolates varied from 28.54 ± 1.7% to 109.74 ± 2.28% compared to S. aureus 8325-4, which was used as a reference (100%; Figure 2A). In addition, 14 out of the 15 gentamicin-susceptible isolates exhibited higher internalization and persistence rates than DU5883 Δfnb S. aureus based on the gentamicin protection assay (p < 0.01 – Figure 2B and Supplementary Figure S1). Concerning infection-induced cytotoxicity, 14 out of 15 isolates harbored a comparable phenotype to the reference strain, and cells infected with these isolates exhibited a significantly higher cytotoxicity rate (p < 0.001) than uninfected cells (100%; Supplementary Figure S1).

Bottom Line: Compared to S. aureus, S. pseudintermedius was the only species that significantly adhered to human fibronectin.These results suggest the involvement of β1 integrin in the invasion process, although this mechanism was previously restricted to S. aureus.In summary, our results suggest that internalization into NPPCs is not a classical pathophysiologic mechanism of SNA BJIs.

View Article: PubMed Central - PubMed

Affiliation: Centre International de Recherche en Infectiologie, INSERM U1111, CNRS UMR5308, Université de Lyon 1, ENS de Lyon, Team "Pathogenesis of staphylococcal infections"Lyon, France; Department of Clinical Microbiology, Northern Hospital Group, Hospices Civils de LyonLyon, France.

ABSTRACT
Implicated in more than 60% of bone and joint infections (BJIs), Staphylococci have a particular tropism for osteoarticular tissue and lead to difficult-to-treat clinical infections. To date, Staphylococcus aureus internalization in non-professional phagocytic cells (NPPCs) is a well-explored virulence mechanism involved in BJI chronicity. Conversely, the pathophysiological pathways associated with Staphylococcus non-aureus (SNA) BJIs have scarcely been studied despite their high prevalence. In this study, 15 reference strains from 15 different SNA species were compared in terms of (i) adhesion to human fibronectin based on adhesion microplate assays and (ii) internalization ability, intracellular persistence and cytotoxicity based on an in vitro infection model using human osteoblasts. Compared to S. aureus, S. pseudintermedius was the only species that significantly adhered to human fibronectin. This species was also associated with high (even superior to S. aureus) internalization ability, intracellular persistence and cytotoxicity. These findings were confirmed using a panel of 17 different S. pseudintermedius isolates. Additionally, S. pseudintermedius internalization by osteoblasts was completely abolished in β1 integrin-deficient murine osteoblasts. These results suggest the involvement of β1 integrin in the invasion process, although this mechanism was previously restricted to S. aureus. In summary, our results suggest that internalization into NPPCs is not a classical pathophysiologic mechanism of SNA BJIs. S. pseudintermedius appears to be an exception, and its ability to invade and subsequently induce cytotoxicity in NPPCs could explain its severe and necrotic forms of infection, notably in dogs, which exhibit a high prevalence of S. pseudintermedius infection.

No MeSH data available.


Related in: MedlinePlus