Validation of the new Sepsis-3 definitions: proposal for improvement in early risk identification.

OBJECTIVES: Sepsis-3 definitions generated controversies regarding their general applicability. The Sepsis-3 Task Force outlined the need for validation with emphasis on the quick Sequential Organ Failure Assessment (qSOFA) score. This was done in a prospective cohort from a different healthcare setting. METHODS: Patients with infections and at least two signs of systemic inflammatory response syndrome (SIRS) were analysed. Sepsis was defined as total SOFA ≥2 outside the intensive care unit (ICU) or as an increase of ICU admission SOFA ≥2. The primary endpoints were the sensitivity of qSOFA outside the ICU and sepsis definition both outside and within the ICU to predict mortality. RESULTS: In all, 3346 infections outside the ICU and 1058 infections in the ICU were analysed. Outside the ICU, respective mortality with ≥2 SIRS and qSOFA ≥2 was 25.3% and 41.2% (p <0.0001); the sensitivities of qSOFA and of sepsis definition to predict death were 60.8% and 87.2%, respectively. This was 95.9% for sepsis definition in the ICU. The sensitivity of qSOFA and of ≥3 SIRS criteria for organ dysfunction outside the ICU was 48.7% and 72.5%, respectively (p <0.0001). Misclassification outside the ICU with the 1991 and Sepsis-3 definitions into stages of lower severity was 21.4% and 3.7%, respectively (p <0.0001) and 14.9% and 3.7%, respectively, in the ICU (p <0.0001). Adding arterial pH ≤7.30 to qSOFA increased sensitivity for prediction of death to 67.5% (p 0.004). CONCLUSIONS: Our analysis positively validated the use of SOFA score to predict unfavourable outcome and to limit misclassification into lower severity. However, qSOFA score had inadequate sensitivity for early risk assessment.

Change of annexin binding of monocytes as an expression of cellular response to Candida albicans: down-regulation in severe sepsis.

To study the differences of monocyte activation by albicans and non-albicans species of Candida and its change in sepsis, peripheral blood mononuclear cells were isolated from 17 healthy volunteers and 26 patients with severe sepsis/shock, and incubated in the absence/presence of heat-killed (HK) isolates of four different Candida species and purified ß-D-glucan from C.albicans. Experiments were repeated in the presence and absence of inhibitors of intracellular activation pathways. Expression of annexin V on cells membranes of monocytes and lymphocytes, cytoplasmic activity of caspase-3, and DNA fragmentation of monocytes were studied. Membrane expression of annexin V on viable monocytes of healthy volunteers decreased significantly after incubation with C.albicans but not with non-albicans species. The decrease was dose-dependent from the Candida inoculum and by the concentration of ß-D-glucan. A relationship with inhibition of apoptosis was found as the activity of caspase-3 activity, and the level of DNA fragmentation were also decreased. Incubation in the absence/presence of inhibitors showed that the decrease by annexin V expression resulted by activation of the dectin-1 pathway and Raf-1 by ß-D glucan. The decrease of annexin V(+)/PI(-) expression was not shown on monocytes of patients with severe sepsis/shock, where no effect of inhibitors was found. Decrease of annexin V binding on monocytes can be viewed as a selective response to C.albicans partly effected through activation of dectin-1. This response is down-regulated after a septic insult.

Impact of haplotypes of TNF in the natural course of infective endocarditis.

Based on previous findings for the role of single nucleotide polymorphisms (SNPs) of TNF for the predisposition for bloodstream infections, this study investigates the role of these SNPs at the promoter positions -376, -308, -238 in infective endocarditis (IE). In a case-control study, 83 patients with IE and 83 controls were enrolled. Blood genotyping for the presence of G or A alleles of the three SNPs was carried out using restriction fragment length polymorphisms. Haplotypes were calculated. Patients were mostly infected by Staphylococcus aureus (32.5%) and by species of enterococci (14.3%) and streptococci (14.3%). Carriage of the minor frequency A alleles at -238 of the promoter region of TNF was greater than in controls (8.4% versus 1.2%, p 0.003). The presence of any of the three GGA/GAA/AGA haplotypes was more frequent in patients with IE (OR 8.22, 95CI% 1.8-37.4, p 0.001). After multivariate logistic regression analysis, it was found that the only factor related to fatal outcome was carriage of the wild-type GGG haplotype (OR, 3.29, 95CI%, 1.05-10.29, p 0.04). GGA, AGA and GAA haplotypes were more frequent in patients with IE than in controls, suggesting a predisposition for IE and a potential protective role against fatal outcome, as the wild-type GGG haplotype was independently related with death.

Indication for a role of regulatory T cells for the advent of influenza A (H1N1)-related pneumonia.

Regulatory T cells (T(regs) ) have an anti-inflammatory role. A former study in a limited number of patients found that absolute counts of T(regs) increase when infection by the new influenza H1N1 virus is complicated with pneumonia. These results generate the question if H1N1-related pneumonia is associated with a state of hypo-inflammation. A total of 135 patients were enrolled with blood sampling within less than 24 h from diagnosis; 23 with flu-like syndrome; 69 with uncomplicated H1N1-infection; seven with bacterial pneumonia; and 36 with H1N1-related pneumonia. T(regs) and CD14/HLA-DR co-expression were estimated by flow cytometry; concentrations of tumour necrosis factor-alpha (TNF-α), of interleukin (IL)-6 and of soluble triggering receptor expressed on myeloid cells-1 (sTREM-1) by an enzyme immunoassay; those of procalcitonin (PCT) by immuno-time-resolved amplified cryptate technology assay. Expression of human leucocyte antigen D-related (HLA-DR) on monocytes was similar between groups; absolute T(reg) counts were greater among patients with H1N1-related pneumonia than flu-like syndrome or H1N1-uncomplicated infection. Serum TNF-α of patients with bacterial pneumonia was greater than those of other groups, but IL-10 was similar between groups. Serum PCT was greater among patients with H1N1-related pneumonia and sTREM-1 among those with H1N1-related pneumonia. Regression analysis revealed that the most important factors related with the advent of pneumonia were the existence of underlying illnesses (P = 0·006) and of T(regs) equal to or above 16 mm(3) (P = 0·013). It is concluded that the advent of H1N1-related pneumonia is related to an early increase of the absolute T(reg) counts. This increase is probably not part of a hypo-inflammatory state of the host.

In vitro release of fusidic acid and teicoplanin from cancellous bone allografts.

The characteristics of cancellous bone allografts as carriers of fusidic acid and teicoplanin are described. Particles of cancellous bone were compressed into a wiremesh cylinder; five replicas were impregnated for one hour into fusidic acid; and another five for one hour into teicoplanin. Elution was estimated daily. Concentrations of fusidic acid and teicoplanin were determined by a microbiological assay. Both antibiotics were eluted at very high concentrations within the first days. Allografts impregnated in fusidic acid provided concentrations above 20 microg/ml for 20 days. Eluted teicoplanin after day 4 was below 10 microg/ml. It is concluded that cancellous bone allografts may allow adequate in vitro elution of fusidic acid but not of teicoplanin. The latter results support their application in experimental models of osteomyelitis.

Efficacy of teicoplanin, administered in two different regimens, in the treatment of experimental endocarditis due to Enterococcus faecalis.

Using a rabbit model of endocarditis, we studied the efficacy of teicoplanin against a strain of Enterococcus faecalis resistant to ampicillin. Rabbits were randomly assigned to receive no antibiotics, teicoplanin 12 or 18 mg/kg of body weight every 12h, for 9 days. The effect of treatment on bacterial counts of vegetations and survival of the animals was evaluated at the end of treatment and 10 days thereafter. The two treatment regimens of teicoplanin produced peak serum levels 18.51+/-1.84 and 34.66+/-4.19 microg/ml, and trough levels above 10 x MIC of teicoplanin for the infecting organism. Both regimens resulted in significant bacterial reduction in the vegetations as compared to the control group (p<0.001). The drug prevented relapse of the infection 10 days after discontinuation of treatment. By increasing the teicoplanin dosage no additional therapeutic benefit was observed in terms of bacterial killing, sterilization of the vegetations, and survival of the animals, although the higher doses gave numerically superior results. These findings may have meaning for the optimum use of teicoplanin in the treatment of enterococcal endocarditis.

The effect of the type of intraperitoneally implanted prosthetic mesh on the systemic inflammatory response.

BACKGROUND: The purpose of this study was to determine any differences in the systemic inflammatory response after the intraperitoneal implantation of three different types of polypropylene mesh. METHODS: Thirty-two male New Zealand rabbits underwent a 6-cm midline incision and opening of the peritoneal cavity. The animals were randomly divided into four groups. In groups A, B, and C, there was an intraperitoneal placement of polypropylene mesh, titanium-coated polypropylene mesh, and composite polypropylene/e-PTFE mesh, respectively. Group D received a sham operation. Blood was sampled preoperatively and at 6, 24, 48, and 168 h postoperatively to measure white blood cell count (WBC), tumor necrosis factor-alpha (TNF-alpha), and malondialdehyde (MDA). RESULTS: Statistically significant elevations of WBC, TNF-alpha and MDA were observed in all four groups at 6, 24, and 48 h postoperatively (P<0.05). There were no statistically significant differences in WBC, TNF-alpha, and MDA between groups A, B, and C at any time interval. However, a statistically significant elevation of WBC (P<0.05) and TNF-alpha (P<0.05) was observed between each of the groups with mesh implantation and group D at 24 h postoperatively. CONCLUSION: Intraperitoneal mesh implantation induces mild systemic inflammatory response regardless of the type of implanted mesh.

Early apoptosis of monocytes contributes to the pathogenesis of systemic inflammatory response and of bacterial translocation in an experimental model of multiple trauma.

The objective of this study was to investigate the occurrence of apoptosis of monocytes in an experimental model of multiple trauma and its probable correlation to bacterial translocation. Thirty-two rabbits were applied in three groups: A, controls; B, myotomy of the right femur; and C, myotomy and fracture of the right femur. Blood was sampled for the estimation of endotoxins [lipopolysaccharide (LPS)], tumour necrosis factor (TNF)-alpha, malondialdehyde (MDA) and isolation of peripheral blood mononuclear cells (PBMCs). PBMCs, derived after centrifugation over Ficoll, were incubated in flasks and apoptosis of non-adherent lymphocytes and adherent monocytes was estimated after staining for Annexin-V and flow cytometry. TNF-alpha of supernatants of cultured monocytes was also determined. Tissue segments were cultured after death. Median survival of groups A, B and C was > 14, > 14 and 9.00 days, respectively. Apoptosis of lymphocytes in group C was higher than group A at 2, 4 and 48 h and of monocytes in group C higher than group A at 2 and 4 hours. LPS in group C was higher than group A at 2, 4 and 48 h. Apoptosis of lymphocytes and monocytes was correlated positively with serum TNF-alpha and negatively with TNF-alpha of monocyte supernatants. Cultures of organ segments of group A were sterile. Pseudomonas aeruginosa was isolated from liver, lung and spleen in five animals in group B (45.45%) and in six in group C (54.54%). Early apoptosis of blood monocytes supervened after multiple trauma; the phenomenon was accompanied by apoptosis of blood lymphocytes and subsequent bacterial translocation.