Fast and reliable assays to precisely define the nature of the infectious agents causing sepsis are eagerly anticipated. New molecular biology techniques are now available to define the presence of bacterial or fungal DNA within the bloodstream of sepsis patients. We have used a new technique (VYOO®) that allows the enrichment of microbial DNA before a multiplex polymerase chain reaction (PCR) for pathogen detection provided by SIRS-Lab (Jena, Germany). We analyzed 72 sepsis patients and 14 non-infectious systemic inflammatory response syndrome (SIRS) patients. Among the sepsis patients, 20 had a positive blood culture and 35 had a positive microbiology in other biological samples. Of these, 51.4% were positive using the VYOO® test. Among the sepsis patients with a negative microbiology and the non-infectious SIRS, 29.4% and 14.2% were positive with the VYOO® test, respectively. The concordance in bacterial identification between microbiology and the VYOO® test was 46.2%. This study demonstrates that these new technologies offer great hopes, but improvements are still needed.
Treatment of septic shock relies on appropriate antimicrobial therapy. Current culture based methods deliver final results after days, which may delay potentially lifesaving adjustments in antimicrobial therapy. This study was undertaken to compare PCR with blood culture results under routine conditions regarding 1. impact on antimicrobial therapy, and 2. time to result, in patients with presumed sepsis.
This was an observational study in a 50 beds ICU of a university hospital. In 245 patients with suspected sepsis, 311 concomitant blood cultures and blood for multiplex PCR (VYOO®) were obtained. 45 of 311 blood cultures (14.5%) and 94 of 311 PCRs (30.1%) were positive. However, blood culture or microbiological sampling from the presumed site of infection rarely confirmed PCR results and vice versa. Median time to positivity and interquartile range were 24.2 (18.0, 27.5) hours for the PCR and 68 (52.2, 88.5) hours for BC (p<0.01). PCR median time to result was dependent on technician availability (53.5 hours on Saturdays, 7.2 hours under optimal logistic conditions). PCR results showed good correlation with procalcitonin (p<0.001). In 34% of patients with positive PCRs antimicrobial therapy was considered inadequate according to assessment of clinical arbitrators including 5 patients with vancomycin-resistant enterococci (VRE), 3 cases with multiresistant staphylococci, and 4 patients with fungi.
The results of this observational study support the hypothesis that PCR results are available faster, are more frequently positive, and may result in earlier adjustment of antimicrobial therapy. However, shorter time to result can only be fully exploited when the laboratory is adequately staffed for a 24 hour/7 day service, or when point of care/automated assay systems become available.
Sepsis is a major health problem in newborns and children. Early detection of pathogens allows initiation of appropriate antimicrobial therapy that strongly correlates with positive outcomes. Multiplex PCR has the potential to rapidly identify bloodstream infections, compensating for the loss of blood culture sensitivity. In an Italian pediatric hospital, multiplex PCR (the LightCycler SeptiFast test) was compared to routine blood culture with 1,673 samples obtained from 803 children with suspected sepsis; clinical and laboratory information was used to determine the patient infection status. Excluding results attributable to contaminants, SeptiFast showed a sensitivity of 85.0% (95% confidence interval [CI] = 78.7 to 89.7%) and a specificity of 93.5% (95% CI = 92.1 to 94.7%) compared to blood culture. The rate of positive results was significantly higher with SeptiFast (14.6%) than blood culture (10.3%) (P < 0.0001), and the overall positivity rate was 16.1% when the results of both tests were combined. Staphylococcus aureus (11.6%), coagulase-negative staphylococci (CoNS) (29.6%), Pseudomonas aeruginosa (16.5%), and Klebsiella spp. (10.1%) were the most frequently detected. SeptiFast identified 97 additional isolates that blood culture failed to detect (24.7% P. aeruginosa, 23.7% CoNS, 14.4% Klebsiella spp., 14.4% Candida spp.). Among specimens taken from patients receiving antibiotic therapy, we also observed a significantly higher rate of positivity of SeptiFast than blood culture (14.1% versus 6.5%, respectively; P < 0.0001). On the contrary, contaminants were significantly more frequent among blood cultures than SeptiFast (n = 97 [5.8%] versus n = 26 [1.6%]), respectively; P < 0.0001). SeptiFast served as a highly valuable adjunct to conventional blood culture in children, adding diagnostic value and shortening the time to result (TTR) to 6 h.
Timely identification of pathogens is crucial to minimize mortality in patients with severe infections. Detection of bacterial and fungal pathogens in blood by nucleic acid amplification promises to yield results faster than blood cultures (BC). We analyzed the clinical impact of a commercially available multiplex PCR system in patients with suspected sepsis.
Blood samples from patients with presumed sepsis were cultured with the Bactec 9240™ system (Becton Dickinson, Heidelberg, Germany) and aliquots subjected to analysis with the LightCycler® SeptiFast® (SF) Test (Roche Diagnostics, Mannheim, Germany) at a tertiary care centre. For samples with PCR-detected pathogens, the actual impact on clinical management was determined by chart review. Furthermore a comparison between the time to a positive blood culture result and the SF result, based on a fictive assumption that it was done either on a once or twice daily basis, was made.
Of 101 blood samples from 77 patients, 63 (62%) yielded concordant negative results, 14 (13%) concordant positive and 9 (9%) were BC positive only. In 14 (13%) samples pathogens were detected by SF only, resulting in adjustment of antibiotic therapy in 5 patients (7,7% of patients). In 3 samples a treatment adjustment would have been made earlier resulting in a total of 8 adjustments in all 101 samples (8%).
The addition of multiplex PCR to conventional blood cultures had a relevant impact on clinical management for a subset of patients with presumed sepsis.
Patients identified with sepsis in the emergency department often are treated on the basis of the presumption of infection; however, various noninfectious conditions that require specific treatments have clinical presentations very similar to that of sepsis. Our aim was to describe the etiology of illness in patients identified and treated for severe sepsis in the emergency department.
We conducted a prospective observational study of patients treated with goal-directed resuscitation for severe sepsis in the emergency department. Inclusion criteria were suspected infection, 2 or more criteria for systemic inflammation, and evidence of hypoperfusion. Exclusion criteria were age of <18 years and the need for immediate surgery. Clinical data on eligible patients were prospectively collected for 2 years. Blinded observers used a priori definitions to determine the final cause of hospitalization.
In total, 211 patients were enrolled; 95 (45%) had positive culture results, and 116 (55%) had negative culture results. The overall mortality rate was 19%. Patients with positive culture results were more likely to have indwelling vascular lines (P = .03) be residents of nursing homes (P = .04), and have a shorter time to administration of antibiotics in the emergency department (83 vs 97 min; P = .03). Of patients with negative culture results, 44% had clinical infections, 8% had atypical infections, 32% had noninfectious mimics, and 16% had an illness of indeterminate etiology.
In this study, we found that >50% of patients identified and treated for severe sepsis in the emergency department had negative culture results. Of patients identified with a sepsis syndrome at presentation, 18% had a noninfectious diagnosis that mimicked sepsis, and the clinical characteristics of these patients were similar to those of patients with culture-positive sepsis.
Sepsis is among the top 10 causes of mortality in the United States. Rapid administration of antibiotics is one of the most important contributors to patient survival, yet only a limited number of methods exist for rapid identification of microbes cultivated from bloodstream infections, which can lead to sepsis. While traditional single-target molecular methods have been shown to greatly improve survival for septic patients by enabling rapid deescalation of broad-spectrum antibiotics, multiplex methods offer even greater possibilities. A novel multiplex method, PCR coupled to electrospray ionization mass spectrometry (PCR/ESI-MS), was used to identify the genus and species of microorganisms found to cause human bloodstream infections. DNA was directly extracted from 234 BacT-Alert blood culture bottles, and results were compared to those obtained by clinical reference standard methods. The study results demonstrated 98.7% and 96.6% concordance at the genus and species levels, respectively. Mixtures of microbes were identified in 29 blood culture bottles, including mixed species of the same genus, as well as mixtures containing Gram-positive and Gram-negative organisms, exemplifying the PCR/ESI-MS capability to identify multiple organisms simultaneously without the need for cultivation. This study demonstrates high analytical accuracy in comparison to routine subculture of blood culture bottles and phenotypic identification of microbes. Without foreknowledge of the microorganisms potentially present, the PCR/ESI-MS methods can deliver accurate results in as little as 5 to 6 h after a positive alarm from the automated blood culture system; however, current batch mode testing limits the method's clinical utility at this time.
Accurate and timely diagnosis of community-acquired bacterial infections in patients with systemic inflammation remains challenging both for clinician and laboratory. Combinations of markers, as opposed to single ones, may improve diagnosis and thereby survival. We therefore compared the diagnostic characteristics of novel and routinely used biomarkers of sepsis alone and in combination.
This prospective cohort study included patients with systemic inflammatory response syndrome who were suspected of having community-acquired infections. It was conducted in a medical emergency department and department of infectious diseases at a university hospital. A multiplex immunoassay measuring soluble urokinase-type plasminogen activator (suPAR) and soluble triggering receptor expressed on myeloid cells (sTREM)-1 and macrophage migration inhibitory factor (MIF) was used in parallel with standard measurements of C-reactive protein (CRP), procalcitonin (PCT), and neutrophils. Two composite markers were constructed – one including a linear combination of the three best performing markers and another including all six – and the area under the receiver operating characteristic curve (AUC) was used to compare their performance and those of the individual markers.
A total of 151 patients were eligible for analysis. Of these, 96 had bacterial infections. The AUCs for detection of a bacterial cause of inflammation were 0.50 (95% confidence interval [CI] 0.40 to 0.60) for suPAR, 0.61 (95% CI 0.52 to 0.71) for sTREM-1, 0.63 (95% CI 0.53 to 0.72) for MIF, 0.72 (95% CI 0.63 to 0.79) for PCT, 0.74 (95% CI 0.66 to 0.81) for neutrophil count, 0.81 (95% CI 0.73 to 0.86) for CRP, 0.84 (95% CI 0.71 to 0.91) for the composite three-marker test, and 0.88 (95% CI 0.81 to 0.92) for the composite six-marker test. The AUC of the six-marker test was significantly greater than that of the single markers.
Combining information from several markers improves diagnostic accuracy in detecting bacterial versus nonbacterial causes of inflammation. Measurements of suPAR, sTREM-1 and MIF had limited value as single markers, whereas PCT and CRP exhibited acceptable diagnostic characteristics.
Early infection diagnosis as the cause of a patient's systemic inflammatory syndrome is an important facet of sepsis care bundles aimed at saving lives. Microbiological culture provides the main route for infection diagnosis but by its nature cannot provide time-critical results that can impact on early management. Consequently, broad-spectrum and high-potency antibiotics are essential during the immediate management of suspected sepsis in critical care but are associated with the development of drug-resistant organisms and superinfections. Established molecular laboratory techniques based on polymerase chain reaction (PCR) technology can detect pathogen DNA rapidly and have been developed for translation into a clinical diagnostic setting. In the setting of sepsis in critical care, emerging commercial systems are now available for the analysis of whole blood within hours, with the presumed aim of adoption into the current care bundles. In this review, we consider the importance of early infection diagnosis in sepsis, how this is limited by culture approaches and how the emerging PCR methods are showing promise in early clinical observational studies. The strengths and weaknesses of culture and PCR pathogen detection in whole-blood samples will be highlighted and recommendations made for urgent appropriately powered diagnostic validation studies in advance of clinical effectiveness trials before these emerging PCR pathogen detection techniques can be considered for adoption in clinical practice.
Morbidity, mortality and social cost of sepsis are high. Previous studies have suggested that individual cytokines levels could be used as sepsis markers. Therefore, we assessed whether the multiplex technology could identify useful cytokine profiles in Emergency Department (ED) patients.
ED patients were included in a single tertiary-care center prospective study. Eligible patients were >18 years and met at least one of the following criteria: fever, suspected systemic infection, ≥2 systemic inflammatory response syndrome (SIRS) criteria, hypotension or shock. Multiplex cytokine measurements were performed on serum samples collected at inclusion. Associations between cytokine levels and sepsis were assessed using univariate and multivariate logistic regressions, principal component analysis (PCA) and agglomerative hierarchical clustering (AHC).
Among the 126 patients (71 men, 55 women; median age: 54 years [19–96 years]) included, 102 had SIRS (81%), 55 (44%) had severe sepsis and 10 (8%) had septic shock. Univariate analysis revealed weak associations between cytokine levels and sepsis. Multivariate analysis revealed independent association between sIL-2R (p = 0.01) and severe sepsis, as well as between sIL-2R (p = 0.04), IL-1β (p = 0.046), IL-8 (p = 0.02) and septic shock. However, neither PCA nor AHC distinguished profiles characteristic of sepsis.
Previous non-multiparametric studies might have reached inappropriate conclusions. Indeed, well-defined clinical conditions do not translate into particular cytokine profiles. Additional and larger trials are now required to validate the limited interest of expensive multiplex cytokine profiling for staging septic patients.
Sepsis is a serious medical condition that requires rapidly administered, appropriate antibiotic treatment. Conventional methods take three or more days for final pathogen identification and antimicrobial susceptibility testing. We organized a prospective observational multicenter study in three study sites to evaluate the diagnostic accuracy and potential clinical utility of the SeptiFast system, a multiplex pathogen detection system used in the clinical setting to support early diagnosis of bloodstream infections.
A total of 212 patients, suspected of having systemic inflammatory response syndrome (SIRS) caused by bacterial or fungal infection, were enrolled in the study. From these patients, 407 blood samples were taken and blood culture analysis was performed to identify pathogens. Whole blood was also collected for DNA Detection Kit analysis immediately after its collection for blood culture. The results of the DNA Detection Kit, blood culture and other culture tests were compared. The chosen antimicrobial treatment in patients whose samples tested positive in the DNA Detection Kit and/or blood culture analysis was examined to evaluate the effect of concomitant antibiotic exposure on the results of these analyses.
SeptiFast analysis gave a positive result for 55 samples, while 43 samples were positive in blood culture analysis. The DNA Detection Kit identified a pathogen in 11.3% (45/400) of the samples, compared to 8.0% (32/400) by blood culture analysis. Twenty-three pathogens were detected by SeptiFast only; conversely, this system missed five episodes of clinically significant bacteremia (Methicillin-resistant Staphylococcus aureus (MRSA), 2; Pseudomonas aeruginosa, 1; Klebsiella spp, 1; Enterococcus faecium, 1). The number of samples that tested positive was significantly increased by combining the result of the blood culture analysis with those of the DNA Detection Kit analysis (P = 0.01). Among antibiotic pre-treated patients (prevalence, 72%), SeptiFast analysis detected more bacteria/fungi, and was less influenced by antibiotic exposure, compared with blood culture analysis (P = 0.02).
This rapid multiplex pathogen detection system complemented traditional culture-based methods and offered some added diagnostic value for the timely detection of causative pathogens, particularly in antibiotic pre-treated patients. Adequately designed intervention studies are needed to prove its clinical effectiveness in improving appropriate antibiotic selection and patient outcomes.
Sepsis is a serious condition with a significant morbidity and mortality. New insight into the immunopathogenesis of sepsis could promote the development of new strategies for diagnosis and therapy. High mobility group box-1 protein (HMGB1) has been known for many years as a nuclear chromosomal protein. Its role as a pro-inflammatory cytokine in sepsis and rheumatoid arthritis has been described recently. The aim of our study was to evaluate HMGB1 as a molecular marker in patients with community-acquired infections.
Patients suspected of having infections/sepsis and admitted to a department of internal medicine were included in the study in a prospective manner. Demographic data, comorbidity, routine biochemistry, microbiological data, infection focus, severity score, and mortality on day 28 were recorded. Plasma and serum were sampled at the time of admission. HMGB1 levels were measured with a commercially available enzyme-linked immunosorbent assay (ELISA). Procalcitonin levels were measured with a TRACE (time-resolved amplified cryptate emission) assay. Lipopolysaccharide-binding protein and interleukin-6 were measured with a chemiluminiscent immunometric assay. Soluble haemoglobin scavenger receptor (sCD163) levels were measured with an in-house ELISA.
One hundred and ninety-four patients were included in the study. Levels of HMGB1 are presented as medians and interquartile ranges: healthy controls (0.77 ng/ml, 0.6 to 1.46), non-infected patients (1.54 ng/ml, 0.79 to 2.88), infected patients without systemic inflammatory response syndrome (2.41 ng/ml, 0.63 to 3.44), patients with sepsis (2.24 ng/ml, 1.30 to 3.75), and patients with severe sepsis (2.18 ng/ml, 0.91 to 3.85). In a receiver operator characteristic curve analysis discriminating between non-infected patients and all infected patients, the area under the curve for HMGB1 was 0.59 (P < 0.0001). HMGB1 correlated only weakly to levels of white blood cell count, neutrophils, C-reactive protein, interleukin-6, procalcitonin, and lipopolysaccharide-binding protein (P < 0.001). HMGB1 did not correlate to sCD163.
In a cohort of patients with suspected community-acquired infections and sepsis, HMGB1 levels were statistically significantly higher in patients compared to the healthy controls. There was no statistically significant difference between the infected and the non-infected patients. Levels of HMGB1 correlated only very weakly to other pro-inflammatory markers and did not correlate to the anti-inflammatory marker sCD163.
Bloodstream infections are associated with high mortality rates because of the probable manifestation of sepsis, severe sepsis and septic shock1. Therefore, rapid administration of adequate antibiotic therapy is of foremost importance in the treatment of bloodstream infections. The critical element in this process is timing, heavily dependent on the results of bacterial identification and antibiotic susceptibility testing. Both of these parameters are routinely obtained by culture-based testing, which is time-consuming and takes on average 24-48 hours2, 4. The aim of the study was to develop DNA-based assays for rapid identification of bloodstream infections, as well as rapid antimicrobial susceptibility testing. The first assay is a eubacterial 16S rDNA-based real-time PCR assay complemented with species- or genus-specific probes5. Using these probes, Gram-negative bacteria including Pseudomonas spp., Pseudomonas aeruginosa and Escherichia coli as well as Gram-positive bacteria including Staphylococcus spp., Staphylococcus aureus, Enterococcus spp., Streptococcus spp., and Streptococcus pneumoniae could be distinguished. Using this multiprobe assay, a first identification of the causative micro-organism was given after 2 h.
Secondly, we developed a semi-molecular assay for antibiotic susceptibility testing of S. aureus, Enterococcus spp. and (facultative) aerobe Gram-negative rods6. This assay was based on a study in which PCR was used to measure the growth of bacteria7. Bacteria harvested directly from blood cultures are incubated for 6 h with a selection of antibiotics, and following a Sybr Green-based real-time PCR assay determines inhibition of growth. The combination of these two methods could direct the choice of a suitable antibiotic therapy on the same day (Figure 1). In conclusion, molecular analysis of both identification and antibiotic susceptibility offers a faster alternative for pathogen detection and could improve the diagnosis of bloodstream infections.
Immunology; Issue 65; Infection; Medicine; Microbiology; Bacteria; real-time PCR; probes; pathogen detection; blood culture; 16S rDNA gene; antibiotic resistance; antibiotic susceptibility testing
The aim of the study was to test if the Mortality in Emergency Department Sepsis (MEDS) score accurately predicts death among emergency department (ED) patients with severe sepsis and septic shock.
This study was a preplanned secondary analysis of a before-and-after interventional study conducted at a large urban ED. Inclusion criteria were suspected infection, 2 or more criteria for systemic inflammation, and either systolic blood pressure of less than 90 mm Hg after a fluid bolus or lactate 4 mmol/L or higher. Exclusion criteria were: age of less than 18 years, no aggressive care desired, or need for immediate surgery. Clinical and outcomes data were prospectively collected on consecutive eligible patients for 1 year before and 1 year after implementing early goal-directed therapy (EGDT). The MEDS scores and probabilities of in-hospital death were calculated. The main outcome was in-hospital mortality. The area under the receiver operating characteristic curve was used to evaluate score performance.
One hundred forty-three patients, 79 pre-EGDT and 64 post-EGDT, were included. The mean age was 58 ± 17 years, and pneumonia was the source of infection in 37%. The in-hospital mortality rate was 23%. The area under the receiver operating characteristic curve for MEDS to predict mortality was 0.61 (95% confidence interval [CI], 0.50–0.72) overall, 0.69 (95% CI, 0.56–0.82) in pre-EGDT patients, and 0.53 (95% CI, 0.33–0.74) in post-EGDT patients.
The MEDS score performed with poor accuracy for predicting mortality in ED patients with sepsis. These results suggest the need for further validation of the MEDS score before widespread clinical use.
Sepsis is one of the leading causes of morbidity and mortality in hospitalized patients worldwide. Molecular technologies for rapid detection of microorganisms in patients with sepsis have only recently become available. LightCycler SeptiFast test Mgrade (Roche Diagnostics GmbH) is a multiplex PCR analysis able to detect DNA of the 25 most frequent pathogens in bloodstream infections. The time and labor saved while avoiding excessive laboratory manipulation is the rationale for selecting the automated MagNA Pure compact nucleic acid isolation kit-I (Roche Applied Science, GmbH) as an alternative to conventional SeptiFast extraction. For the purposes of this study, we evaluate extraction in order to demonstrate the feasibility of automation. Finally, a prospective observational study was done using 106 clinical samples obtained from 76 patients in our ICU. Both extraction methods were used in parallel to test the samples. When molecular detection test results using both manual and automated extraction were compared with the data from blood cultures obtained at the same time, the results show that SeptiFast with the alternative MagNA Pure compact extraction not only shortens the complete workflow to 3.57 hrs., but also increases sensitivity of the molecular assay for detecting infection as defined by positive blood culture confirmation.
The identification and treatment of critical illness is often initiated by emergency medical services (EMS) providers. We hypothesized that emergency department (ED) patients with severe sepsis who received EMS care had more rapid recognition and treatment compared with non-EMS patients.
Prospective observational study of ED patients with severe sepsis treated with early goal-directed therapy (EGDT). We included adults with suspected infection, evidence of systemic inflammation, and either hypotension after a fluid bolus or elevated lactate. Prehospital and ED clinical variables and outcomes data were collected. The primary outcome was time to initiation of antibiotics in the ED.
There were 311 patients with 160 (51.4%) transported by EMS. EMS transported patients had more organ failure (sequential organ failure assessment score 7.0 vs. 6.1, p =0.02), shorter time to first antibiotics (111 vs. 146 minutes, p=0.001) and, shorter time from triage to EGDT initiation (119 vs. 160 minutes, p=0.005), compared to non-EMS transported patients. Among EMS patients, if the EMS provider indicated a written impression of sepsis, there was a shorter time to antibiotics (70 vs. 122 minutes, p=0.003) and a shorter time to EGDT initiation (69 vs. 131 minutes, p=0.001), compared to those without an impression of sepsis.
In this prospective cohort, EMS provided initial care for half of severe sepsis patients requiring EGDT. Patients presented by EMS had more organ failure and a shorter time to both antibiotic and EGDT initiation in the ED.
Early structured resuscitation of severe sepsis has been suggested to improve short term mortality; however, no previous study has examined the long-term effect of this therapy. We sought to determine one year outcomes associated with implementation of early goal directed therapy (EGDT) in the emergency department (ED) care of sepsis.
We performed a longitudinal analysis of a prospective before and after study conducted at a large urban ED. Adult patients were enrolled if they had suspected infection, 2 or more systemic inflammatory response criteria, and either systolic blood pressure (SBP) <90 mmHg after a fluid bolus or lactate >4 mM. Exclusion criteria were: age <18 years, no aggressive care desired, or need for immediate surgery. Clinical and outcomes data were prospectively collected on consecutive eligible patients for 1 year before and 2 years after implementing EGDT. Patients in the pre-implementation phase received non-protocolized care at attending physician discretion. The primary outcome was mortality at one year.
285 subjects, 79 in the pre- and 206 in the post-implementation phases, were enrolled. Compared to pre-implementation, post-implementation subjects had a significantly lower ED SBP (72 vs. 85 mm Hg, P < 0.001) and higher sequential organ failure assessment score (7 vs. 5, P = 0.0004). The primary outcome of 1 year mortality was observed in 39/79 (49%) pre-implementation subjects and 77/206 (37%) post-implementation subjects (difference 12%; P = 0.04).
Implementation of EGDT for the treatment of ED patients with severe sepsis and septic shock was associated with significantly lower mortality at one year.
Bloodstream infections in neonates and infants are life-threatening emergencies. Identification of the common bacteria causing such infections and their susceptibility patterns will provide necessary information for timely intervention. This study is aimed at determining the susceptibilities of bacterial etiological agents to commonly-used antimicrobial agents for empirical treatment of suspected bacterial septicaemia in children.
This is a hospital based retrospective analysis of blood cultures from infants to children up to 14 years of age with preliminary diagnosis of sepsis and admitted to the Neonatal Intensive Care Unit (NICU) and Paediatric Wards of the Teaching Hospital Tamale from July 2011 to January 2012.
Out of 331 blood specimens cultured, the prevalence of confirmed bacterial sepsis was 25.9% (86/331). Point prevalence for confirmed cases from NICU was 44.4% (28/63) and 21.6% (58/268) from the Paediatric ward. Gram positive cocci (GPC) were the predominant isolates with Coagulase positive (32.2%) and Coagulase-negative (28.7%) Staphylococci accounting for 60.9% of the total isolates. Gram negative rods (GNR) comprised 39.1% of all isolates with Klebsiella, E.coli and Salmonella being the most common organisms isolated. Klebsiella was the most frequent GNR from the NICU and Salmonella typhi was predominantly isolated from the paediatric ward. Acinetobacter showed 100.0% susceptibility to Ceftriaxone and Cefotaxime but was resistant (100.0%) to Ampicillin, Tetracycline and Cotrimoxazole. Escherichia coli and Klebsiella were 80.0% and 91.0% susceptible to Ceftriaxone and Cefotaxime respectively. Klebsiella species showed 8.3% susceptibility to Tetracycline but was resistant to Ampicillin and Cotrimoxazole. Escherichia coli showed 40.0% susceptibility to Ampicillin, Chloramphenicol and Cotrimoxazole; 20.0% susceptibility to Tetracycline and 80.0% susceptible to Gentamicin and Cefuroxime. Coagulase negative Staphylococci was susceptible to Gentamicin (72.0%) but Coagulase positive Staphylococci showed intermediate sensitivity to Gentamicin (42.9%).
Coagulase Negative, Coagulase Positive Staphylococci, Salmonella and Klebsiella were the aetiological agents of bloodstream infection among children at TTH. While gram-positive and gram-negative bacteria showed low susceptibility to Ampicillin, Tetracycline and Cotrimoxazole, the GNR were susceptible to Gentamicin and third-generation cephalosporins.
Early diagnosis and rapid bacterial identification are of primary importance for outcome of septic patients. SeptiFast® (SF) real-time PCR assay is of potential utility in the etiological diagnosis of sepsis, but it cannot replace blood culture (BC) for routine use in clinical laboratory. Procalcitonin (PCT) is a marker of sepsis and can predict bacteremia in septic patients. The aim of the present study was to investigate whether PCT serum levels could predict SF results, and could help screening febrile patients in which a SF assay can improve the etiological diagnosis of sepsis.
From 1009 febrile patients with suspected sepsis, 1009 samples for BC, SF real-time PCR, and PCT determination were obtained simultaneously, and results were compared and statistically analysed. Receiver operating characteristic (ROC) curves were generated to determine the area under the curve and to identify which cut-off of PCT value produced the best sensitivity to detect SF results.
Mean PCT values of sera drawn simultaneously with samples SF positive (35.42±61.03 ng/ml) or BC positive (23.14±51.56 ng/ml) for a pathogen were statistically higher than those drawn simultaneously with SF negative (0.84±1.67 ng/ml) or BC negative (2.79±16.64 ng/ml) samples (p<0.0001). For SF, ROC analysis showed an area under the curve of 0.927 (95% confidence interval: 0.899–0.955, p<0.0001). The PCT cut-off value of 0.37 ng/ml showed a negative predictive value of 99%, reducing the number of SF assays of 53.9%, still identifying the 96.4% of the pathogens.
PCT can be used in febrile patients with suspected sepsis to predict SF positive or negative results. A cut-off value of 0.37 ng/ml can be considered for optimal sensitivity, so that, in the routine laboratory activity, SF assay should not be used for diagnosis of sepsis in an unselected patient population with a PCT value <0.37 ng/ml.
Procalcitonin (PCT) has emerged as a valuable marker of sepsis. The potential role of PCT in diagnosis and therapy monitoring of intravascular catheter-related bloodstream infections (CRBSI) in intensive care unit (ICU) is still unclear and was evaluated.
Forty-six patients were included in the study, provided they were free of infection upon admission and presented the first episode of suspected CRBSI during their ICU stay. Patients who had developed any other infection were excluded. PCT was measured daily during the ICU hospitalization. Primary endpoint was proven CRBSI. Therapy monitoring as according to infection control was also evaluated.
Among the 46 patients, 26 were diagnosed with CRBSI. Median PCT on the day of infection suspicion (D0) was 7.70 and 0.10 ng/ml for patients with and without proven CRBSI, respectively (p < 0.001). The area under the curve (AUC) for PCT was 0.990 (95% CI; 0.972 – 1.000), whereas a cut-off value of 0.70 ng/ml provided sensitivity and specificity of 92.3 and 100% respectively. In contrast, the AUC for white blood cells (WBC) was 0.539 (95% CI; 0.369 – 0.709), and for C-reactive protein (CRP), 0.603 (95% CI; 0.438 – 0.768). PCT was the best predictor of proven infection. Moreover, an increase >0.20 ng/ml of PCT between the D0 and any of the 4 preceding days was associated with a positive predictive value exceeding 96%. PCT concentrations from the D2 to D6 after suspected infection tended to decrease in controlled patients, whereas remained stable in non-controlled subjects. A PCT concentration exceeding 1.5 ng/ml during D3 was associated with lack of responsiveness to therapy (p = 0.028).
We suggest that PCT could be a helpful diagnostic and prognostic marker of CRBSI in critically ill patients. Both absolute values and variations should be considered.
Despite advances in modern medicine, sepsis remains a complex syndrome that has been associated with significant morbidity and mortality. Multiple organ failure associated with sepsis leads to high mortality and morbidity. About 28 – 50% deaths have been reported in patients with sepsis. The number of sepsis patients is increasing, with considerable burden on healthcare facilities. Various factors leading to a rise in the incidence of sepsis are (1) Improvement of diagnostic procedures (2) Increase in the number of immunocompromised patients taking treatment for various autoimmune disease, carcinomas, organ transplantation (3) Advances in intensive procedures (4) Nosocomial infections (5) Extensive use of antibiotics. With the better understanding of sepsis various modalities to modify pathophysiological response of septic patients have developed. Activated protein C and low-dose corticosteroid therapy have been tried in patients, with variable results.
Corticosteroid; human recombinant activated protein C (rhAPC); sepsis
Early diagnostic and prognostic stratification of patients with suspected infection is a difficult clinical challenge. We studied plasma pentraxin 3 (PTX3) upon admission to the emergency department in patients with suspected infection.
The study comprised 537 emergency room patients with suspected infection: 59 with no systemic inflammatory response syndrome (SIRS) and without bacterial infection (group 1), 67 with bacterial infection without SIRS (group 2), 54 with SIRS without bacterial infection (group 3), 308 with sepsis (SIRS and bacterial infection) without organ failure (group 4) and 49 with severe sepsis (group 5). Plasma PTX3 was measured on admission using a commercial solid-phase enzyme-linked immunosorbent assay (ELISA).
The median PTX3 levels in groups 1–5 were 2.6 ng/ml, 4.4 ng/ml, 5.0 ng/ml, 6.1 ng/ml and 16.7 ng/ml, respectively (p<0.001). The median PTX3 concentration was higher in severe sepsis patients compared to others (16.7 vs. 4.9 ng/ml, p<0.001) and in non-survivors (day 28 case fatality) compared to survivors (14.1 vs. 5.1 ng/ml, p<0.001). A high PTX3 level predicted the need for ICU stay (p<0.001) and hypotension (p<0.001). AUCROC in the prediction of severe sepsis was 0.73 (95% CI 0.66–0.81, p<0.001) and 0.69 in case fatality (95% CI 0.58–0.79, p<0.001). PTX3 at a cut-off level for 14.1 ng/ml (optimal cut-off value for severe sepsis) showed 63% sensitivity and 80% specificity. At a cut-off level 7.7 ng/ml (optimal cut-off value for case fatality) showed 70% sensitivity and 63% specificity in predicting case fatality on day 28.In multivariate models, high PTX3 remained an independent predictor of severe sepsis and case fatality after adjusting for potential confounders.
A high PTX3 level on hospital admission predicts severe sepsis and case fatality in patients with suspected infection.
The aim of this study was to determine the occurrence rate, demographics, clinical characteristics, and outcomes of patients with severe sepsis admitted to the emergency department.
A prospective study evaluating all patients admitted to the emergency department unit in a public hospital of tertiary complexity in a six-month period was conducted. During this period, the emergency team was trained to diagnose sepsis. Patients who met the diagnostic criteria for severe sepsis were followed until their discharge from the hospital.
A total of 5,332 patients were admitted to the emergency department, and 342 met the criteria for severe sepsis/septic shock. The median (interquartile range) age of patients was 74 (65–84) years, and 52.1% were male. The median APACHE II and SOFA scores at diagnosis were 19 (15–25) and 5 (3–7), respectively. The median number of dysfunctional organ systems per patient was 2 (1–3). The median hospital length of stay was 10 (4.7–17) days, and the hospital mortality rate was 64%. Only 31% of the patients were diagnosed by the emergency department team as septic. About 33.5% of the 342 severe sepsis patients admitted to the emergency department were referred to an ICU, with a median time delay of 24 (12–48) hours. Training improved diagnosis and decreased the time delay for septic patients in arriving at the ICU.
The occurrence rate of severe sepsis in the emergency department was 6.4%, and the rate of sepsis diagnosed by the emergency department team as well as the number of patients transferred to the ICU was very low. Educational campaigns are important to improve diagnosis and, hence, treatment of severe sepsis.
Sepsis; Emergency Department; Occurrence; Epidemiology; Mortality; Training
International guidelines concerning the management of patients with sepsis, septic shock and multiple organ failure make no reference to the nature of the infecting organism. Indeed, most clinical signs of sepsis are nonspecific. In contrast, in vitro data suggest that there are mechanistic differences between bacterial, viral and fungal sepsis, and imply that pathogenetic differences may exist between subclasses such as Gram-negative and Gram-positive bacteria. These differences are reflected in different cytokine profiles and mortality rates associated with Gram-positive and Gram-negative sepsis in humans. They also suggest that putative anti-mediator therapies may act differently according to the nature of an infecting organism. Data from some clinical trials conducted in severe sepsis support this hypothesis. It is likely that potential new therapies targeting, for example, Toll-like receptor pathways will require knowledge of the infecting organism. The advent of new technologies that accelerate the identification of infectious agents and their antimicrobial sensitivities may allow better tailored anti-mediator therapies and administration of antibiotics with narrow spectra and known efficacy.
Neonatal sepsis is difficult to diagnose and pathogens cannot be detected from blood cultures in many cases. Development of a rapid and accurate method for detecting pathogens is thus essential. The main purpose of this study was to identify etiological agents in clinically diagnosed neonatal sepsis using bacterial ribosomal RNA-targeted reverse transcription-quantitative PCR (BrRNA-RT-qPCR) and to conduct comparisons with the results of conventional blood culture. Since BrRNA-RT-qPCR targets bacterial ribosomal RNA, detection rates using this approach may exceed those using conventional PCR.
Subjects comprised 36 patients with 39 episodes of suspected neonatal sepsis who underwent BrRNA-RT-qPCR and conventional blood culture to diagnose sepsis. Blood samples were collected aseptically for BrRNA-RT-qPCR and blood culture at the time of initial sepsis evaluation by arterial puncture. BrRNA-RT-qPCR and blood culture were undertaken using identical blood samples, and BrRNA-RT-qPCR was performed using 12 primer sets.
Positive rate was significantly higher for BrRNA-RT-qPCR (15/39, 38.5%) than for blood culture (6/39, 15.4%; p = 0.0039). BrRNA-RT-qPCR was able to identify all pathogens detected by blood culture. Furthermore, this method detected pathogens from neonates with clinical sepsis in whom pathogens was not detected by culture methods.
This RT-PCR technique is useful for sensitive detection of pathogens causing neonatal sepsis, even in cases with negative results by blood culture.
Intensive care mortality of HIV-positive patients has progressively decreased. However, critically ill HIV-positive patients with sepsis present a worse prognosis. To better understand this condition, we propose a study comparing clinical, etiological and inflammatory data, and the hospital course of HIV-positive and HIV-negative patients with severe sepsis or septic shock.
A prospective observational study enrolling patients with severe sepsis or septic shock associated or not with HIV infection, and admitted to intensive care unit (ICU). Clinical, microbiological and inflammatory parameters were assessed, including C-reactive protein (CRP), procalcitonin (PCT), interleukin-6, interleukin-10 and TNF-α. Outcome measures were in-hospital and six-month mortality.
The study included 58 patients with severe sepsis/septic shock admitted to ICU, 36 HIV-positive and 22 HIV-negative. All HIV-positive patients met the criteria for AIDS (CDC/2008). The main foci of infection in HIV-positive patients were pulmonary and abdominal (p=0.001). Fungi and mycobacteria were identified in 44.4% and 16.7% of HIV-positive patients, respectively. In contrast, the main etiologies for sepsis in HIV-negative patients were Gram-negative bacilli (36.4%) and Gram-positive cocci (36.4%) (p=0.001). CRP and PCT admission concentrations were lower in HIV-positive patients (130 vs. 168 mg/dL p=0.005, and 1.19 vs. 4.06 ng/mL p=0.04, respectively), with a progressive decrease in surviving patients. Initial IL-10 concentrations were higher in HIV-positive patients (4.4 pg/mL vs. 1.0 pg/mL, p=0.005), with moderate accuracy for predicting death (area under receiver-operating characteristic curve =0.74). In-hospital and six-month mortality were higher in HIV-positive patients (55.6 vs. 27.3% p=0.03, and 58.3 vs. 27.3% p=0.02, respectively).
The course of sepsis was more severe in HIV-positive patients, with distinct clinical, etiological and inflammatory characteristics.
sepsis; HIV; AIDS; etiology; inflammatory markers; mortality