**7. Performance of CRP in diagnosis of neonatal sepsis can further be enhanced by combining it with early sensitive markers**

An important limitation of CRP is the low sensitivity during the early phases of sepsis. By then values are often still normal, though the consequences of the bacterial invasion are

already apparent and a delay of the initiation of antibiotic therapy may be associated with an adverse outcome. CRP takes ten to twelve hours to significantly change after the onset of infection. [6] Earlier in the inflammatory cascade activated macrophages release proinflammatory cytokines (IL-1, IL-6, TNF-α) and growth factors (IL-3, CSFs) inducing the hepatic synthesis of acute-phase-reactants and the activation of neutrophils. The increase of cytokines therefore precedes the changes of CRP. Of the many mediators studied, much attention has been focused on IL6, IL8, and TNF-α.

The Role of C-Reactive Protein in the Diagnosis of Neonatal Sepsis 53

Sensitivity is low during the early phase of infection. The performance of serial

improves diagnostic accuracy.

whom infection has resolved.

compared to term newborns.

antibiotics in these infants.

applied to clinical practice.

identifying infected cases.

**Table 3.** CRP facts.

later periods. [62]

CRP values in otherwise healthy newborns.

may better reflect neonatal physiology.

determinations 24 to 48 hours after the onset of symptoms is recommended, as it clearly

CRP is particularly useful for ruling out an infection and for monitoring the response to treatment and guiding the duration of the antibiotic therapy. Two consecutive values <10 mg/l determined more than 24 hours apart identify infants unlikely to be infected or in

CRP values undergo a physiological 3-day-rise after birth and non-infectious confounders such as meconium aspiration syndrome and perinatal maternal risk conditions may elevate

Preterm neonates have lower baseline CRP values and a lower CRP response to infection in

Data on non infectious CRP elevations in otherwise healthy newborns are inconsistent and does currently not allow drawing recommendations on the continuation or withdrawal of

Up to date the most used cut-off value is 10 mg/l irrespective of the gestational and postnatal age of the neonate. Cut-off values adapted to the gestational and postnatal age

In order to compensate for the diagnostic weakness during the early phases of infection initial CRP determination should be combined with determination of early and sensitive markers. Suitable markers include but are not limited to PCT, IL6, and IL8. Many further parameters may provide similar good results but are not yet sufficiently examined to be

At the moment none of the described current diagnostic markers are sensitive and specific enough to influence the judgment to withhold antimicrobial treatment independent of the clinical findings. Efforts were done to improve diagnostic accuracy by combining multiple markers in order to further enhance the diagnostic accuracy of these mediators in

CRP has been investigated in combination with a variety of "new" infection markers including cytokines, surface markers, and other acute-phase-reactants with promising results. Especially the combination with an early sensitive marker such as PCT, IL6, IL8, CD11b, and CD64 increases the sensitivity to values between 90% and 100% in most studies.

Especially in the early neonatal period, many physiological and metabolic processes are in change and differ from every later moment in life. These changes affect several laboratory parameters as well and many reference values and serum kinetics substantially differ to

**8. Do special subpopulations need special CRP reference values?** 

IL-6 increases rapidly after the bacterial invasion and was demonstrated to have a high sensitivity during the early stages of sepsis (80%-100%) even when determined from umbilical cord blood (87%-100%). [29, 53) However, a short half life caused by plasma protein binding, hepatic clearance, and inactivation results in a rapid normalization of serum levels and a decrease of sensitivity during the later course of the disease, even though the infection persists. IL-8 and TNF-α have very similar characteristics and kinetic properties to IL-6. Both are pro-inflammatory cytokines predominantly produced by activated phagocytes in response to systemic infection and inflammation. [53] While studies report on a reliable diagnostic accuracy of IL-8 with a sensitivity of 69%-100%, the usefulness of TNF-α as a diagnostic marker has not been found to be as good as either IL-6 or IL-8. [53, 54]

Similar to CRP procalcitonin is another important acute-phase reactant produced by monocytes and hepatocytes. It has the advantage of increasing more rapidly after contact to bacterial endotoxin with levels rising after four hours and peaking at six to eight hours [55] In a recent meta-analysis the sensitivity and specificity in the diagnosis of early onset sepsis were 76% (range 68–82%) and 76% (60–87%). [56] Though the sensitivity during the early stages of sepsis may be superior to CRP, the significant rapid variations of basal levels after birth, the increase after non-infectious conditions such as asphyxia, maternal pre-eclampsia, and intracranial hemorrhage,[57] and the need for several different cut-off values with changing neonatal age, have limited its diffusion as an early marker in comparison to CRP.

Specific leukocyte cell surface antigens are known to be expressed in substantial quantities after inflammatory cells are activated by bacteria or their cellular products. [62] From the amount of surface markers studied neutrophil CD11b and CD64 appear most promising for diagnosis of neonatal sepsis. CD11b expression increases considerably within a few minutes after the inflammatory cells come into contact with bacteria and endotoxins. [58, 59] The sensitivity and specificity of CD11b for diagnosing early onset neonatal sepsis are 86–100% and 100% respectively. [7, 53] CD64 has a sensitivity ranging between 81% and 96% and a NPV between 89% and 97%. [60] Though promising, estimation of cell surface markers is limited by the need for sophisticated equipment and the need to process blood samples rapidly before neutrophils die from apoptosis or the surface antigens are down regulated. [61]

Despite the favorable claims by many studies, many of these diagnostic markers fail to meet the stringent demands required for clinical practice. High costs, limited availability of specimens at the appropriate time, and complexity of the assay methods all limit the clinical applicability. More importantly, the relatively small sample size in most studies, the lack of clear reference values for many diagnostic markers still prohibit the use of most of these parameters in clinical practice.

Sensitivity is low during the early phase of infection. The performance of serial determinations 24 to 48 hours after the onset of symptoms is recommended, as it clearly improves diagnostic accuracy.

CRP is particularly useful for ruling out an infection and for monitoring the response to treatment and guiding the duration of the antibiotic therapy. Two consecutive values <10 mg/l determined more than 24 hours apart identify infants unlikely to be infected or in whom infection has resolved.

CRP values undergo a physiological 3-day-rise after birth and non-infectious confounders such as meconium aspiration syndrome and perinatal maternal risk conditions may elevate CRP values in otherwise healthy newborns.

Preterm neonates have lower baseline CRP values and a lower CRP response to infection in compared to term newborns.

Data on non infectious CRP elevations in otherwise healthy newborns are inconsistent and does currently not allow drawing recommendations on the continuation or withdrawal of antibiotics in these infants.

Up to date the most used cut-off value is 10 mg/l irrespective of the gestational and postnatal age of the neonate. Cut-off values adapted to the gestational and postnatal age may better reflect neonatal physiology.

In order to compensate for the diagnostic weakness during the early phases of infection initial CRP determination should be combined with determination of early and sensitive markers. Suitable markers include but are not limited to PCT, IL6, and IL8. Many further parameters may provide similar good results but are not yet sufficiently examined to be applied to clinical practice.

**Table 3.** CRP facts.

52 Neonatal Bacterial Infection

attention has been focused on IL6, IL8, and TNF-α.

parameters in clinical practice.

already apparent and a delay of the initiation of antibiotic therapy may be associated with an adverse outcome. CRP takes ten to twelve hours to significantly change after the onset of infection. [6] Earlier in the inflammatory cascade activated macrophages release proinflammatory cytokines (IL-1, IL-6, TNF-α) and growth factors (IL-3, CSFs) inducing the hepatic synthesis of acute-phase-reactants and the activation of neutrophils. The increase of cytokines therefore precedes the changes of CRP. Of the many mediators studied, much

IL-6 increases rapidly after the bacterial invasion and was demonstrated to have a high sensitivity during the early stages of sepsis (80%-100%) even when determined from umbilical cord blood (87%-100%). [29, 53) However, a short half life caused by plasma protein binding, hepatic clearance, and inactivation results in a rapid normalization of serum levels and a decrease of sensitivity during the later course of the disease, even though the infection persists. IL-8 and TNF-α have very similar characteristics and kinetic properties to IL-6. Both are pro-inflammatory cytokines predominantly produced by activated phagocytes in response to systemic infection and inflammation. [53] While studies report on a reliable diagnostic accuracy of IL-8 with a sensitivity of 69%-100%, the usefulness of TNF-α as a

Similar to CRP procalcitonin is another important acute-phase reactant produced by monocytes and hepatocytes. It has the advantage of increasing more rapidly after contact to bacterial endotoxin with levels rising after four hours and peaking at six to eight hours [55] In a recent meta-analysis the sensitivity and specificity in the diagnosis of early onset sepsis were 76% (range 68–82%) and 76% (60–87%). [56] Though the sensitivity during the early stages of sepsis may be superior to CRP, the significant rapid variations of basal levels after birth, the increase after non-infectious conditions such as asphyxia, maternal pre-eclampsia, and intracranial hemorrhage,[57] and the need for several different cut-off values with changing neonatal age, have limited its diffusion as an early marker in comparison to CRP. Specific leukocyte cell surface antigens are known to be expressed in substantial quantities after inflammatory cells are activated by bacteria or their cellular products. [62] From the amount of surface markers studied neutrophil CD11b and CD64 appear most promising for diagnosis of neonatal sepsis. CD11b expression increases considerably within a few minutes after the inflammatory cells come into contact with bacteria and endotoxins. [58, 59] The sensitivity and specificity of CD11b for diagnosing early onset neonatal sepsis are 86–100% and 100% respectively. [7, 53] CD64 has a sensitivity ranging between 81% and 96% and a NPV between 89% and 97%. [60] Though promising, estimation of cell surface markers is limited by the need for sophisticated equipment and the need to process blood samples rapidly before neutrophils die from apoptosis or the surface antigens are down regulated. [61] Despite the favorable claims by many studies, many of these diagnostic markers fail to meet the stringent demands required for clinical practice. High costs, limited availability of specimens at the appropriate time, and complexity of the assay methods all limit the clinical applicability. More importantly, the relatively small sample size in most studies, the lack of clear reference values for many diagnostic markers still prohibit the use of most of these

diagnostic marker has not been found to be as good as either IL-6 or IL-8. [53, 54]

At the moment none of the described current diagnostic markers are sensitive and specific enough to influence the judgment to withhold antimicrobial treatment independent of the clinical findings. Efforts were done to improve diagnostic accuracy by combining multiple markers in order to further enhance the diagnostic accuracy of these mediators in identifying infected cases.

CRP has been investigated in combination with a variety of "new" infection markers including cytokines, surface markers, and other acute-phase-reactants with promising results. Especially the combination with an early sensitive marker such as PCT, IL6, IL8, CD11b, and CD64 increases the sensitivity to values between 90% and 100% in most studies.
