The diagnostic value of serum levels of C-reactive protein and procalcitonin in differentiation between active pulmonary TB and CAP

Introduction: C-reactive protein (CRP) and procalcitonin (PCT) levels are elevated in patients with community-acquired pneumonia (CAP), but PCT does not increase in patients with pulmonary tuberculosis (TB). Aim: To evaluate the diagnostic value of serum levels of CRP and PCT in differentiating between active pulmonary TB and CAP. Participants and methods: The present study was carried out on 90 individuals divided into the following groups: group I included 10 control participants, group II included 40 patients with active pulmonary TB, and group III included 40 patients with CAP. Serum levels of CRP and PCT were measured. Results: CRP was significantly increased in group III compared with groups I and II. PCT was significantly increased in group III compared with groups I and II; also, there was a significant increase in group II compared with group I. The cut-off value of CRP between group II and group III was more than 24 (mg/dl), with a sensitivity of 100%, a specificity of 70%, and that of PCT was more than 530 (pg/ml), with a sensitivity of 67% and a specificity of 97.5%. Conclusion: Measurements of CRP and PCT were complementary to each other to differentiate between pulmonary TB and CAP.


Introduction
Tuberculosis (TB) continues to be a major public health concern, with nine million incident cases and estimated 1.3 million deaths reported worldwide in 2007 [1]. Th e diagnosis of TB is made on the basis of clinical symptoms, chest radiograph, skin tuberculin test, and fi nally, the detection of the causative agent by direct microscopy of biological specimens and culture on solid and in liquid media. Smear examination and in-vitro culture of Mycobacterium tuberculosis bacilli has remained the gold standard [2].
Community-acquired pneumonia (CAP) is a major cause of hospital admission and the most important infectious cause of death [3]. A rapid diagnosis and appropriate antibiotic treatment are essential to reduce the morbidity and mortality caused by CAP.
In countries with a high TB burden, Mycobacterium tuberculosis is a frequent cause of CAP and the diff erential diagnosis of TB from common bacterial pneumonia is diffi cult [3].
C-reactive protein (CRP) is an acute-phase protein produced primarily in the liver and is stimulated by cytokine release [4]; CRP levels are elevated in patients with CAP compared with healthy control participants [5]. Procalcitonin (PCT) is an acutephase reactant protein and consists of 116 amino acids; it has been reported to be a sensitive marker of severe bacterial infection [2].
Recently, PCT has also been introduced as a promising alternative to CRP in guiding the antibiotic treatment of CAP and acute exacerbations of chronic obstructive pulmonary disease [1] on the basis of the ability of PCT to discriminate between patients with or without bacterial infection. In lower respiratory tract infections, measurement of serum PCT may enable physicians to diff erentiate between typical bacterial and nonbacterial causes of infl ammation using a cut-off value of 0.5 ng/ml [6][7][8][9]. In addition, PCT does not appear to be signifi cantly elevated in patients with pulmonary TB, making it an attractive potentially rapid diagnostic method for diff erentiating pulmonary TB from bacterial CAP [1]. Identifi cation of the etiology of CAP is a clinical diffi culty because single clinical, radiologic, or laboratory parameters have limited value in predicting the infectious organism [10], and no rapid test has been standardized for the diagnosis of 'atypical' or viral pathogens. As a result, broad-spectrum initial antibiotic therapy is usually empirically chosen [11,12]. PCT serum levels or other biological markers of bacterial infection might enable clinicians to choose targeted antibiotic Introduction C-reactive protein ( CRP) and procalcitonin ( PCT) levels are elevated in patients with communityacquired pneumonia (CAP), but PCT does not increase in patients with pulmonary tuberculosis (TB).
Aim To evaluate the diagnostic value of serum levels of CRP and PCT in differentiating between active pulmonary TB and CAP.

Participants and methods
The present study was carried out on 90 individuals divided into the following groups: group I included 10 control participants, group II included 40 patients with active pulmonary TB, and group III included 40 patients with CAP. Serum levels of CRP and PCT were measured.
Results CRP was signifi cantly increased in group III compared with groups I and II. PCT was signifi cantly increased in group III compared with groups I and II; also, there was a signifi cant increase in group II compared with group I. The cut-off value of CRP between group II and therapy in patients with CAP by diff erentiating between classic bacterial and atypical or viral etiology.
Th is work aims to evaluate the diagnostic value of serum levels of CRP and PCT in diff erentiating between active pulmonary TB and CAP.

Participants and methods
Th e present study was carried out on 90 participants; they were recruited from the chest department, Tanta University Hospital, and Abbassia chest hospital during the period from April 2012 till April 2013 and divided into three groups. Group I included 10 apparently healthy normal nonsmoker volunteers, six men and four women, mean age 37.1 ± 10.816 years. Group II included 40 patients with active pulmonary TB, 40 men and no women, mean age 38.1 ± 6.368 years. Group III included 40 patients with CAP caused by bacterial infection, 36 men and four women, mean age 49.425 ± 6.687 years. Informed consents were obtained from all the participants in this study. (d) Sputum Gram stain and culture showed that the causative organisms were bacteria. Sputum samples shoul d fulfi ll the following criteria to maintain a sensitivity of 62% and a specifi city of 85% to ensure that organisms present in the smear were not commensals: squamous epithelial cells (normally exfoliated from the oropharynx) less than 10 cells per lowpower microscopic fi eld, polymorphonuclear neutrophils 10-25 cells per low-power microscopic fi eld, and there should be at least 10 organisms per oil immersion fi eld [14].

Exclusion criteria
(1) Patients with lung infections other than pulmonary TB or CAP caused by bacterial infection.
(2) Bacterial CAP on top of viral infections of respiratory tract .Th is was diagnosed as pneumonia after fl u manifestation although many studies found that serum level of PCT is not increased or even low in viral infections [15,16] Th e following were carried out for all participants: Th orough assessment of history, full clinical examination, plain chest radiograph posteroanterior view, sputum examination fo r acid-fast bacilli by Ziehl-Neelsen stain three successive times for patients suspected of having tuberculous. In patients with CAP, sputum examination by gram stain and culture was performed to detect causative microorganisms; also, blood culture was required in severe cases. Fiv e milliliters of venous blood was withdrawn from each participant to measure the serum levels of CRP and PCT. Serum levels for CRP were measured using qualitative measurement of CRP (Biosystems S.A.Costa Brava 30, 08030 Barcelona-Spain on the basis of the latex agglutination method as follows: (1) All reagents, controls, and serum samples were brought to room temperature. (2) Th e CRP latex reagent was shaken gently before use. One drop of reagent was placed in the test circle. Using disposable pipettes, one drop of undiluted patient serum was placed on the same circle and both were mixed together with the paddle end of the pipette. (3) Positive and negative controls were obtained with each series of test serum in the same way as in step 2. (4) Th e slide was rotated back and forth for 2 min and the result was read under an indirect oblique light source [9].
Serum levels of PCT were measured using a commercially availabl e ELISA kit supplied by 'Raybiotech Inc.' (3607 Parkway Lane, Suite 100, Norcross GA 30092, USA) according to the manufacturer's instructions as follows: (1) All reagents and samples were brought to room temperature (18-25 °C) before use. (2) One hundred microliter of the standard and the sample were added to appropriate wells, covered, and incubated for 2.5 h at room temperature or overnight at 4°C with gentle shaking. (3) Th e solution was discarded and washed four times with 1×wash solution by fi lling each well with wash buff er (300 μl) using a multichannel pipette or autowasher. Complete removal of the liquid at each step was performed and was important to achieve good performance. After the last wash, any remaining wash buff er was removed by aspirating or decanting. Th e plate was then inverted and blotted on clean paper towels. (4) One hundred microliter of 1×prepared biotinylated antibody was added to each well and incubated for 1 h at room temperature with gentle shaking. (5) Th e solution was discarded and the wash was repeated as in step 3. (6) One hundred microliter of prepared Streptavidin solution was added to each well and incubated for 45 min at room temperature with gentle shaking. (7) Th e solution was then discarded and the wash was repeated as in step 3.

(8) One hundred microliter of TMB One-Step
Substrate Reagent was added to each well and incubated for 30 min at room temperature in the dark with gentle shaking. (9) Fifty microliter of stop solution was added to each well and was read at 450 nm [9].

Statistical analysis
Mean values, SD, range, sensitivity, and specifi city were determined in this study, and ANOVA test and Tukey's test were used.

Results
Th e participants were older and there was a signifi cant increase in serum CRP in group III compared with groups I and II, but there was no signifi cant diff erence between groups I and II; also, there was a signifi cant increase in serum PCT in group III compared with groups I and II, but there was signifi cant increase in group II compared with group I. Th ere was a signifi cant positive correlation between serum PCT and serum CRP in groups II and III. Th e cut-off point of serum CRP between group II and group III was more than 24 mg/dl with a sensitivity of 100% and a specifi city of 70%, but the cut-off point of serum PCT between both the groups was more than 530 pg/ml, with a sensitivity of 67% and a specifi city of 97.5% (Tables 1-5 and Figs. 1-4).

Discussion
Sahin and Yildiz [17] concluded that CRP is useful in the diff erential diagnosis of TB and pneumonia; the best serum CRP cut-off value was 9.4 mg/dl; also, a study by Yoon et al. [18] found that the mean values of serum CRP were signifi cantly higher in CAP patients compared with pulmonary tuberculous (PTB) patients, with a sensitivity of 67.5% and a specifi city of 85.1%. Th e CRP level is a nonspecifi c marker of acute-phase infl ammation because it is subjected to the infl uence of other factors such as the general characteristics of  Th is was in agreement with the study of Rasmussen et al. [1] as they found positive correlations between both PCT and CRP. Th e current study showed, that the cut-off value of serum PCT of more than 530 pg/ml Serum C-reactive protein (CRP) level in the three groups stud ied.

Fig. 1
Serum procalcitonin level in the three groups studied. CRP, C-reactive prot ein.

Fig. 2
Correlation between serum procalcitonin and serum C-reactive protein (CRP) in group II.

Fig. 3
Correlation between serum procalcitonin and serum C-reactive protein (CRP) in group III. had a sensitivity of 67% and a specifi city of 97.5% between group II and group III. Nyamande and Lalloo [20] concluded that PCT has a fairly high specifi city for diagnosing CAP because of common bacterial pathogens (88%) as well as PTB (82%); however, the sensitivity is relatively low. Th ey also concluded that PCT was more sensitive and specifi c than CRP in diff erentiating between both diseases. Ugajin et al. [19] concluded that serum PCT has better diagnostic validity than CRP for diff erentiation between PTB and CAP, with a higher sensitivity and specifi city. In this study, the low sensitivity and the high specifi city of PCT as well as the high sensitivity and the low specifi city of CRP in comparing between CAP and pulmonary TB prove that they are complementary tests for diff erentiation between both diseases. However, there are several limitations in this study; the sample size was not large, CAP bacterial causative pathogens were not clearly defi ned, and this work did not study the relation between PCT and CRP concentrations and the severity of CAP or the eff ect of antimicrobial therapy. Th erefore, further studies should be carried out taking into consideration the above-mentioned poi nts.