Serious bacterial infection (SBI) remains an important contributor to mortality and morbidity in children, particularly in resource-poor countries. Child deaths attributable to invasive Streptococcus pneumoniae
and Haemophilus influenzae
type b infections are estimated to equal those collectively attributable to HIV, tuberculosis and malaria
]. The prompt recognition of SBI in children is challenging clinically, especially so in the context of co-infection with malaria and HIV
], and is of the utmost importance in ensuring both effective, and rational antibiotic treatment. Diagnostic uncertainty in bacterial sepsis is compensated by the overuse of antibiotics in self-limiting infections, leading to antibiotic resistance.
Biomarkers may be used alone or in combination to allow classification of an individual to a unique group with defined characteristics. Biomarkers which reflect the host response to infection are useful adjuncts to good clinical acumen and improved molecular diagnostic techniques in confirming SBI. Advances in transcriptomics and proteomics have provided basis for biomarker panels unravelled from these technologies, that might be useful in differentiating sepsis-associated inflammation from non-infectious inflammation
]. An improved understanding of the mechanism of sepsis, and the ability to identify and quantify key mediators involved in its pathophysiology, offers the prospect of reliable diagnostic and prognostic tools. Transcription profiling has been used in other diseases to identify candidate biomarkers
Over 170 biomarkers have been related to sepsis, but relatively few have been evaluated as diagnostic markers. A comprehensive review on sepsis biomarkers, concluded that it is unlikely that a single ideal biomarker will ever be found, and that a combination of biomarkers may be more effective
]. Many of the markers so far investigated are costly and time-consuming to measure. Studies of panels of biomarkers have yielded encouraging results, but none of them have potential to be measured as an affordable point-of- care (POC) test.
The performance of biomarkers is traditionally assessed using receiver operator characteristic (ROC) curves. ROC curves evaluate the discrimination of a test (or combination of tests), but may be insensitive to the addition of a new biomarker, especially if a good biomarker is already included in the model. Net reclassification improvement (NRI) quantifies improvement in the model as a result of adding one or more new biomarkers. This approach has been used in cardiovascular disease
] and acute lung injury
] to improve the accuracy of risk prediction.
Resistin is an adipokine first related to insulin resistance in mice
]. In humans, resistin expression occurs predominantly in macrophages, monocytes and neutrophils
]. Circulating resistin levels peak 8-16 h after the administration of LPS in healthy subjects, mediated by pro-inflammatory cytokines and NF-κB
]. Resistin is significantly elevated in intensive care patients with sepsis
], and levels correlate with severity of disease
Neutrophil gelatinase-associated lipocalin (NGAL) – or lipocalin 2 –has a role in innate immunity through its ability to bind siderophores, required by many bacterial pathogens to scavenge iron from the host
]. Like resistin, it is highly upregulated by inflammatory stimuli via NF-κB
]. NGAL is more significantly elevated in sepsis than in the systemic inflammatory response syndrome (SIRS)
]. In a large study of adult patients with suspected sepsis presenting to an emergency department, the optimal 3-marker panel was NGAL, protein C, and interleukin-1 receptor antagonist. The area under the curve for the accuracy of the sepsis score derived from these three biomarkers was 0.80 for severe sepsis, 0.77 for septic shock, and 0.79 for death
Granulysin is an antimicrobial protein co-localised with perforin in the granules of cytotoxic T lymphocytes (CTLs) and natural killer (NK) cells. It has a direct cytotoxic effect on numerous extracellular pathogens, and is postulated to kill intracellular organisms such as Mycobacterium tuberculosis
in conjunction with perforin
]. Granulysin levels correlate inversely with disease activity in M. tuberculosis
and normalise with treatment
]. Granulysin is transiently elevated in acute viral infections, and has been suggested as a marker of host cellular immunity
We aimed to identify differentially regulated biomarker transcripts in children with SBI using microarray technology. Three previously identified biomarkers were validated by quantitative real-time PCR in an independent prospective cohort. In order to determine the diagnostic utility of the biomarkers to predict SBI, we measured the proteins of the differentially regulated biomarker genes in plasma samples, to determine if they could reliably predict SBI, or outcome from SBI.