**6. Sepsis biomarkers**

#### **6.1. Introduction**

The current gold standard for the diagnosis of infections of the circulatory stream is blood culture, unfortunately its important limitation is the time necessary to complete the survey that goes from 1 to 5 or more days in the case of suspected sepsis caused by yeasts. The ideal situation would be to analyze the patient's on-site blood and provide all the necessary information to immediately target a targeted antibiotic therapy, and then the test should provide data for therapy evaluation by measuring the clearance of pathogenic nucleic acids in the blood in a certain period of time.

The ideal molecular test does not exist but these characteristics will have to be set as a longterm objective in the design of new molecular techniques for the diagnosis of sepsis.

A short-term objective is to analyze blood in parallel to culture methods and identify pathogens responsible for infection including non-cultivable organisms, and also to detect some of the determinants of drug resistance [30]. The molecular techniques currently available on the market can be distinguished in those based on the principle of amplification of the bacterial and/or fungal genome and those that exploit the principle of hybridization. Blood culture coupled with molecular investigations involves some disadvantages inherent to the traditional method such as delay in response with its possible alteration due to an unsuitable collection and the inability to provide the nutrients necessary for the development of demanding microorganisms. In the market, there are several kits in molecular biology that identify the pathogen from positive blood culture samples.

In the medical field, a circulating biochemical marker is defined as a demonstrable, therefore measurable, substance in the blood whose variation in concentration constitutes the signal of the presence of a pathology [32].

Regarding the role of biomarkers in the management of sepsis, over time various molecules were analyzed that, taken singly or combined in a panel, could be able to allow rapid diagnosis and prognosis, and that in the near future could also be able to guide the therapy itself to improve the clinical response and management of these patients [33]. Among the various biomarkers, some are used daily in clinical practice. Among these great importance has always been attributed to the role of lactates. They are considered useful by the guidelines because they have a prognostic value, being related to disease severity especially for values ≥ 2 mmol/L. However, according to other authors, the additive prognostic role of lactates to qSOFA would not be significant [34].

A limitation of blood culture is the lack of sensitivity in the search for particular bacteria that are often responsible for community-acquired pneumonia, such as *Mycoplasma pneumoniae*, *Legionella pneumophila* and *Chlamydia pneumoniae* and other germs that are difficult to grow or even non-cultivable such as *Coxiella burnetii*, *Francisella tularensis*, *Bartonella* spp., *Rickettsia*

The value of blood culture as a diagnostic test for bacteraemia and sepsis is limited, in fact, it emerges that in 50% of cases, the blood culture is negative even if the diagnosis of sepsis is certain and also the first results are provided after 48 h to conclude analysis with the identifi-

The current gold standard for the diagnosis of infections of the circulatory stream is blood culture, unfortunately its important limitation is the time necessary to complete the survey that goes from 1 to 5 or more days in the case of suspected sepsis caused by yeasts. The ideal situation would be to analyze the patient's on-site blood and provide all the necessary information to immediately target a targeted antibiotic therapy, and then the test should provide data for therapy evaluation by measuring the clearance of pathogenic nucleic acids

The ideal molecular test does not exist but these characteristics will have to be set as a long-

A short-term objective is to analyze blood in parallel to culture methods and identify pathogens responsible for infection including non-cultivable organisms, and also to detect some of the determinants of drug resistance [30]. The molecular techniques currently available on the market can be distinguished in those based on the principle of amplification of the bacterial and/or fungal genome and those that exploit the principle of hybridization. Blood culture coupled with molecular investigations involves some disadvantages inherent to the traditional method such as delay in response with its possible alteration due to an unsuitable collection and the inability to provide the nutrients necessary for the development of demanding microorganisms. In the market, there are several kits in molecular biology that identify the pathogen from positive blood culture samples. In the medical field, a circulating biochemical marker is defined as a demonstrable, therefore measurable, substance in the blood whose variation in concentration constitutes the signal of

Regarding the role of biomarkers in the management of sepsis, over time various molecules were analyzed that, taken singly or combined in a panel, could be able to allow rapid diagnosis and prognosis, and that in the near future could also be able to guide the therapy itself to improve the clinical response and management of these patients [33]. Among the various biomarkers, some are used daily in clinical practice. Among these great importance has always been attributed to the role of lactates. They are considered useful by the guidelines because they have a prognostic value, being related to disease severity especially for values

term objective in the design of new molecular techniques for the diagnosis of sepsis.

cation and sensitivity to the pathogen's antibiotics after 5 days or more [1, 30, 31].

spp. and *Nocardia* spp. [28, 29].

64 Biomarker - Indicator of Abnormal Physiological Process

**6. Sepsis biomarkers**

in the blood in a certain period of time.

the presence of a pathology [32].

**6.1. Introduction**

CRP and PCT are routinely used in patients with suspected first aid infection. CRP, released as an acute phase protein in inflammatory states, has a high specificity but low sensitivity and is therefore used for its negative predictive value. PCT, on the other hand, has been shown to have diagnostic and prognostic value in the management of sepsis. It is released in the course of bacterial infections, being therefore aids in the differential diagnosis, and is also included within the guidelines as a valid aid in guiding antibiotic therapy, representing a "mirror" of the therapeutic response to infection, and for its high prognostic power.

More recently, however, we have focused on the role of adrenomedullin (ADM). It is a peptide that several studies have shown to increase in septic patients [35]. The secretion mechanism depends to a large extent on stimulation by the lipopolysaccharide of bacteria, and its plasma levels derive mainly from secretion by endothelial cells [36]. ADM plays a fundamental role in hyperdynamic response during the early stages of sepsis and its main function is vasodilator [37]. After demonstrating the prognostic role of its precursor MR-proADM [38], thanks to a novel instrument based on a chemiluminescent sandwich detection (in which specific monoclonal antibodies are used against the C-terminal part of the peptide), today, it is possible directly measure the biologically active molecule (Bio-ADM or ADM). Thus, in a study published in 2014, our group showed that ADM values are significantly higher in patients with severe sepsis and septic shock than those with sepsis [39]; in particular, ADM, considered one of the most potent endogenous vasodilators, was found to be closely related to low mean arterial pressure (PAM) values and to the need to use vasopressors, thus detecting septic shock markers. Furthermore, higher values have been correlated with greater probability of death; and finally, the detection of multi-stroke ADM has proved to be fundamental in identifying patients with a worse prognosis [39].

From what has emerged, considering that sepsis determines endothelial dysregulation, excessive vasodilatation and consecutively collapse of arterial pressure and micro-vascular homeostasis and that ADM has as its main function vasodilator and therefore regulator of vascular tone, new studies currently in progress and still in the experimental phase, are directed to the development of a murine anti-ADM monoclonal antibody (HAM1101) that can become a therapy in septic shock, according to the hypothesis that it can improve the hemodynamics and perfusion of the organs and, consequently, also reduce the incidence of acute renal failure. To date, a humanized antibody has been selected, the HAM8101, which will be the subject of phase III experimental studies in the near future [39, 40].

Establishing diagnosis and therapy is very important but to hinder their definition is the difficulty in differentiating sepsis from non-infectious stimuli in SIRS situations, especially in those critically ill patients who may have developed SIRS for other causes such as pancreatitis, trauma, burns, etc. Therefore, the detection of an accurate biomarker in sepsis is decisive in critical situations with the ability to exclude or confirm an acute bacterial infection, to evaluate the systemic inflammatory response to infection and the host response to the established therapy [41].

An ideal biological marker must have different characteristics:

• useful in early diagnosis, provide information for a definitive diagnosis or help to identify a probable diagnosis;

• provide information regarding the prognosis including the patient in subpopulations whose outcome is better/worse than the population in question;

**6.2. White blood cell count (WBC)**

predictor for bacteremia [56].

0.77 [56–58].

**6.3. Serum lactate**

≥4 mmol/L alone [61, 62].

vasopressor which caused further tissue hypoxia [62].

White blood cell is a part of innate immune response by localizing infection. Otherwise in systemic sepsis process, there is profound leucocyte activation. Systemic sepsis also leading to organ damage and organ dysfunction attenuated by inhibition of leukocyte-endothelial

Biomarkers Utility for Sepsis Patients Management http://dx.doi.org/10.5772/intechopen.76107 67

The WBC results were considered abnormal if both the total number of neutrophils and the immature/total neutrophil ratio were abnormal simultaneously [53]. The sensitivity and specificity WBC for sepsis diagnosis in the literature vary widely, since there are significant differences in definitions used to count total neutrophils and sub-fractions; with sensitivity varying from 17 to 100% and specificity from 31 to 100%. A study by Caldas et al., reported

In 2001, Zahorec introduced the use of neutrophil and lymphocyte count ratio (NLCR) as one of infection marker [55]. There is also correlation between NLCR and disease severity, also

The sepsis criteria recently changed from Sepsis-2 to Sepsis-3 highlighting on life-threatening organ damage caused by dysregulated host response to infection. This also affecting the use of biomarker in the diagnosis of sepsis, based on Ljungstrom et al., the AUC of NLCR to predict positive culture was 0.71, similar to previous study by Loonen showed AUC of 0.73 and

Lactate is an important source of energy, particularly during starvation. Lactate also contributes to acidic environment by converting to lactic acid. Lactate value of 1400–1500 mmol/L per day resulted from anaerobic glycolysis activity as the reduction of pyruvate, moreover in tissue hypoxia [59]. Excretion of lactate mostly occurs in liver (60%) followed by kidney (30%) and other organs [60]. Shock status, such as cardiogenic or septic shock, is an important source of lactate production. The mortality rate was 46.1% for patients with both hypotension and lactate ≥4 mmol/L, 36.7% for septic patients with hypotension alone and 30% for patients with lactate

According to the new definition, septic shock can be diagnosed under two circumstances. The first one is persistent hypotension and the second one is increase of lactate serum level for more than 2 mmol/L, with additional note that lactate cut off were changed from 4 to 2 mmol/L. Therefore, increase of lactate serum level for more than 2 mmol/L can be recognized as a sign of septic shock. This condition greatly affects by hypotension and the use of

Ljungstrom et al. showed lactate correlate well with Sepsis-2 criteria in diagnosis of sepsis and septic shock, with specificity, accuracy and DOR were 97, 92 and 56.3%, respectively, by using cutoff 3.5 mmol/L. The downside of lactate is that even lactate level alone is widely used for diagnosis in early sepsis, the elevated level is not considered specific for sepsis diagnosis, hence it is proven to be more valuable in predicting mortality. The suggestion is to use combination of several biomarkers including lactate in diagnosing septic and septic shock [56].

interactions, systemic leukocyte activation and disseminated leukocyte adhesion [52].

that combination of WBC and CRP had sensitivity better than WBC alone [54].


The use of biomarkers must always be integrated with the information deriving from a careful medical history collection, from a complete clinical objective examination, from the results of laboratory analyzes and diagnostic methods required in different cases. In fact, only the set of all these elements make it possible to reduce the number of evaluation errors deriving from a hasty decision, often dictated by superficiality or by incorrect knowledge. At present, there is no biomarker that ensures 100% correlation with a single pathology. In fact, we speak of "highly significant values" for a given pathological condition. Other potential uses of biomarkers include their prognostic role and, consequently, the ability to drive antibiotic therapy and to evaluate the response to the therapy itself, identifying those patients who most likely will face complications and organ dysfunction [32].

In the literature, there are many biomarkers already evaluated, such biomarkers for chemokine, cellular, receptor, hemostasis and vascular and others [42].

The available markers for the diagnosis of sepsis are numerous. There are leukocyte count, C-reactive protein (CRP), procalcitonin (PCT), endotoxin, cytokine, IL-1 receptor, complement factors, endothelin-1, ICAM-1 and VCAM-1, fosofolipase A2, PGE2, lactoferrin, neopterin, elastase, different interleukins (ILS), adrenomedullin (ADM) and proADM, atrial natriuretic peptide (ANP) and proANP, pro-vasopressin (copeptin), interferon-γ (IFN-γ), triggering receptor expressed on myeloid cells 1 (TREM-1) and resistin [41–44]. In several recently published studies, the most relevant biomarkers used, as they have a high diagnostic and prognostic capacity are CRP [45], PCT [46], ADM [47], copeptin [48], natriuretic peptide (MR-proANP) [49], presepsin (or CD 14 ligand) [50] and suPAR [51].

Unfortunately, in the sepsis, an ideal marker has not yet been identified and those normally used (fever, leukocytosis, CRP, PCT, etc.) often have little sensitivity and specificity, consequently they have limited use in patient management. Their dosage should be used and evaluated in the context of the clinical situation in which the patient is located and therefore it is essential to determine the appropriateness of the individual markers' request in collaboration with the clinician to exploit their potential to the full diagnostics, therapeutic monitoring and prognosis.

This section will discuss about white blood cell count (WBC), serum lactate, C-reactive protein (CRP), procalcitonin (PCT), presepsin and bioadrenomedullin as biomarker of sepsis.
