Single Nucleotide Polymorphisms (SNPs) in the binding regions (Repeats 1–11) of fibronectin binding protein B (FnBPB) in the derivation cohort (A) and the external validation cohort (B).

<p>Red boxes indicate SNPs that were only present in isolates from the prosthetic joint infection group (PJI), blue boxes indicate SNPs that were only present in the uninfected (PJU) group, and purple boxes indicate SNPs that were present in both PJI and PJU isolates.</p

Single Nucleotide Polymorphisms (SNPs) in fibronectin binding protein A (<i>fnbA)</i> in the derivation cohort, external validation cohort, and late <i>S</i>. <i>aureus</i> bacteremia (SAB) group.

<p>*When false discovery rate control is applied, this raw p-value no longer maintains statistical significance (p = 0.22).</p><p>In the derivation cohort, no SNPs occurred with greater frequency in the prosthetic joint infection group (PJI) relative to the uninfected prosthetic joint group (PJU). In the external validation cohort, one SNP (S839N) was significantly associated with the PJU group, though when the two cohorts were combined the S839N association did not reach statistical significance (p = 0.22). Late SAB was defined as bacteremia occurring >1 year after placement or manipulation of prostheses, and here contains data from both the derivation and external validation cohorts. In the late SAB group, no SNPs occurred with greater frequency in PJI or PJU.</p

Demographic and clinical characteristics of patients in the derivation cohort with <i>S</i>. <i>aureus</i> bacteremia and infected or uninfected prostheses.

<p>*Late infection is defined as bloodstream infection occurring >1 year after the prostheses was implanted or surgically manipulated.</p><p>Demographic and clinical characteristics of patients in the derivation cohort with <i>S</i>. <i>aureus</i> bacteremia and infected or uninfected prostheses.</p

Comparison of the biofilm-forming capacity of <i>S</i>. <i>aureus</i> isolates from prosthetic joint infection (PJI) and uninfected prosthetic joint (PJU) groups.

<p>Values were calculated as percentage capacity to form biofilms relative to <i>S</i>. <i>aureus</i> control strain UAMS-1. Box ends represent the 25^th and 75^th percentiles, and whisker ends represent the minimum and maximum. There was no difference in biofilm-forming capacity between isolates in the PJI and PJU groups.</p

Single Nucleotide Polymorphisms (SNPs) in fibronectin binding protein B (<i>fnbB)</i> in <i>fnbB-</i>containing isolates.

<p>*When false discovery rate control is applied, this raw p-value no longer maintains statistical significance (p = 1.00).</p><p>No SNP was associated with the prosthetic joint infected (PJI) or uninfected (PJU) isolates in the derivation cohort, external validation cohort, or late <i>S</i>. <i>aureus</i> bacteremia (SAB) group. Late SAB was defined as SAB occurring >1 year after placement or manipulation of prostheses.</p

Comparison of the fibronectin binding capacity of <i>S</i>. <i>aureus</i> isolates from prosthetic joint infection (PJI) and uninfected prosthetic joint (PJU) groups.

<p>Values were calculated as percentage capacity to bind fibronectin relative to <i>S</i>. <i>aureus</i> control strain 8325–4. Box ends represent the 25^th and 75^th percentiles, and whisker ends represent the minimum and maximum. There was no difference in fibronectin binding capacity between isolates in the PJI and PJU groups.</p

Candidate genes on murine chromosome 8 are associated with susceptibility to <i>Staphylococcus aureus</i> infection in mice and are involved with <i>Staphylococcus aureus</i> septicemia in humans

<div><p>We previously showed that chromosome 8 of A/J mice was associated with susceptibility to <i>S</i>. <i>aureus</i> infection. However, the specific genes responsible for this susceptibility are unknown. Chromosome substitution strain 8 (CSS8) mice, which have chromosome 8 from A/J but an otherwise C57BL/6J genome, were used to identify the genetic determinants of susceptibility to <i>S</i>. <i>aureus</i> on chromosome 8. Quantitative trait loci (QTL) mapping of <i>S</i>. <i>aureus</i>-infected N2 backcross mice (F1 [C8A] × C57BL/6J) identified a locus 83180780–88103009 (GRCm38/mm10) on A/J chromosome 8 that was linked to <i>S</i>. <i>aureus</i> susceptibility. All genes on the QTL (n~ 102) were further analyzed by three different strategies: 1) different expression in susceptible (A/J) and resistant (C57BL/6J) mice only in response to <i>S</i>. <i>aureus</i>, 2) consistently different expression in both uninfected and infected states between the two strains, and 3) damaging non-synonymous SNPs in either strain. Eleven candidate genes from the QTL region were significantly differently expressed in patients with <i>S</i>. <i>aureus</i> infection vs healthy human subjects. Four of these 11 genes also exhibited significantly different expression in <i>S</i>. <i>aureus</i>-challenged human neutrophils: <i>Ier2</i>, <i>Crif1</i>, <i>Cd97</i> and <i>Lyl1</i>. CD97 ligand binding was evaluated within peritoneal neutrophils from A/J and C57BL/6J. CD97 from A/J had stronger CD55 but weaker integrin α5β1 ligand binding as compared with C57BL/6J. Because CD55/CD97 binding regulates immune cell activation and cytokine production, and integrin α5β1 is a membrane receptor for fibronectin, which is also bound by <i>S</i>. <i>aureus</i>, strain-specific differences could contribute to susceptibility to <i>S</i>. <i>aureus</i>. Down-regulation of <i>Crif1</i> with siRNA was associated with increased host cell apoptosis among both naïve and <i>S</i>. <i>aureus</i>-infected bone marrow-derived macrophages. Specific genes in A/J chromosome 8, including <i>Cd97</i> and <i>Crif1</i>, may play important roles in host defense against <i>S</i>. <i>aureus</i>.</p></div

Increased apoptosis is associated with lower <i>Crif1</i> expression in A/J and CSS8 macrophages.

<p>Apoptosis rates in Figs 8A and 8B cannot be compared because transfection experiments, which intrinsically elicit apoptosis, were performed in (B) but not (A). <b>(A)</b> Bone-marrow derived macrophages from A/J and CSS8 demonstrate higher apoptosis level as compared with C57BL/6J in both uninfected (25.8% and 18.7% vs 12.6%; p <0.05) and <i>S</i>. <i>aureus</i>-challenged conditions (23.3% for A/J, 16.7% for CSS8 and 10.2% for C57BL/6J; p<0.05) (n = 5 in each group). <b>(B)</b> Knockdown of <i>Crif1</i> by siRNA enhances apoptosis in both uninfected and <i>S</i>. <i>aureus</i>-challenged conditions. Knockdown of <i>Crif1</i> in BMDMs from C57BL/6J mice enhances apoptosis in uninfected status as compared with scramble siRNA (38.7% vs 17.3%; p<0.05). <i>S</i>. <i>aureus</i> stimulation exhibits similar patterns of apoptosis (40.6% vs 32.3%; p<0.05) (n = 5 in each group). Mice were 8-week old males.</p

<i>Crif1</i> expression is upregulated in <i>S</i>. <i>aureus</i>-infected A/J and C57BL/6J mice by qPCR.

<p>Consistent expression pattern of <i>Crif1</i>in A/J and C57BL/6J mouse (n = 5 in each group). <i>Crif1</i> showed consistent uninfected vs infected expression patterns between mouse and humans (Figs <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0179033#pone.0179033.g004" target="_blank">4</a> and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0179033#pone.0179033.g005" target="_blank">5</a>) At 3 hours and 6 hours post <i>S</i>. <i>aureus</i> challenge, <i>Crif1</i> is upregulated 6.7 fold (p<0.05) and 1.8 fold (p = 0.4) respectively in A/J, and 4.4 fold (p<0.05) and 2.5 fold (p = 0.14) in C57BL/6J. The normalization was conducted within each strain for comparing different time points (e.g. C57BL/6J time points were normalized to C57BL/6J pre-infection time point and A/J time points were normalized to A/J pre-infection time point.) All mice were 8-week old males.</p

Human orthologues of 11 candidate genes were significantly differentially expressed between patients with blood stream infection (BSI) due to <i>S</i>. <i>aureus</i> (S. A.), <i>E</i>. <i>coli</i> (E.C.) and healthy subjects (Control).

<p>Human orthologues of 11 candidate genes (<i>Crif1</i>, <i>Farsa</i>, <i>Inpp4b</i>, <i>Tnpo2</i>, <i>Cd97</i>, <i>Hook2</i>, <i>Ier2</i>, <i>Lyl1</i>, <i>Mylk3</i>, <i>Rnaseh2a</i> and <i>Tbc1d9</i>) were significantly differentially expressed between patients with <i>S</i>. <i>aureus</i> BSI and healthy subjects by microarray. Human blood RNA from patients with <i>S</i>. <i>aureus</i> BSI (n = 32) and healthy subjects with no infection (n = 44) were extracted and analyzed and applied to microarray. The expression of <i>Cd97</i> (1.17 fold; p<0.05), <i>Crif1</i> (1.87 fold; p<0.0001), <i>Hook2</i> (1.30 fold; p = 0.0001) were significantly higher in <i>S</i>. <i>aureus</i> BSI patients as compared with healthy controls. By contrast, the expression of <i>Farsa</i> (0.66fold; p<0.0001), <i>Ier2</i> (0.93 fold; p<0.05), <i>Inpp4b</i> (0.54fold; p<0.0001), <i>Lyl1</i> (0.72 fold; p<0.001), <i>Rnaseh2a</i> (0.81 fold; p = 0.001), <i>Tbc1d9</i> (0.51 fold; p<0.0001) and <i>Tnpo2</i> (0.84fold; p<0.0001), were significantly lower in <i>S</i>. <i>aureus</i> BSI patients. All of the 11 genes except <i>Cd97</i> showed similar expression changes in <i>Escherichia coli</i> BSI (n = 19) patients.</p