Abstract
INTRODUCTION:
In the developing countries, neonatal sepsis is the most common complication in neonatal period. It is as a systemic inflammatory response because of infection. Laboratory indicators, do not have satisfactory sensitivity. Thus, early identification of sepsis is still needed. Because PTX3 may be a faster acute-phase protein that is not liver-dependent, it is probable that it is superior to traditional biomarkers for mirroring rapid inflammatory courses.
METHODS:
A prospective case control study design was used to determine the sensitivity of pentraxin 3 in the diagnosis of neonatal sepsis to allow early diagnostic tool. This study was carried out on neonatal ICU unit in Suez Canal University Hospital and the studied population were divided into two groups, including patients diagnosed with neonatal sepsis, based on clinical, laboratory and positive blood culture results, and control group
RESULTS:
The study found that there was statistically significant differences between both groups in serum CRP values in diseased and control group (Mean = 49.3±37.4 mg/L, 26.8±17.2 mg/L, p < 0.05), and pentraxin values in diseased and control group (Mean = 5.2±3.7 mg/L, 2.3±0.78 mg/L, p < 0.0001). In addition, there were statistically significant correlations between pentraxin and serum CRP concentrations (p < 0.05) in diseased group. ROC curve showed that serum CRP demonstrated good diagnostic accuracy in predicting neonatal sepsis AUC = 0.875 with sensitivity of 100% and specificity of 92.3%.
CONCLUSION:
Serum PTX3 may be a promising acute-phase protein for interpretation of affected newborns with symptoms and signs of sepsis.
Introduction
There is notable heterogeneity among studies concerning the definition of neonatal sepsis [1]. The existence of a positive blood culture traditionally is the “benchmark” for it [1]. This assumption needs at least two finding: 1) the neonate would not have been assessed for sepsis (have a blood culture drawn) in the nonappearance of clinical signs of a systemic inflammatory response syndrome (SIRS), 2) the isolated bacteria did not signify contamination (type I error). [1]. As the clinical presentation of sepsis are non-specific and associated elevated mortality rates, early diagnosis and starting management before blood culture results is critical [2].
Laboratory tests do not have enough early predic-tive value [3]. Therefore, markers to identify early onset sepsis are needed [3]. The pentraxin units are comprised of proteins with cyclic multimeric structu-res. The acute-phase protein CRP goes to the short members, whereas pentraxin 3 (PTX3) belongs to the long counterparts. PTX3 is secreted from macro-phages and endothelial cells in which the protein is produced and stockpiled. During an acute inflammatory response PTX3 increased its concentrations to nearly 100 fold within couple hours, as seen in other mediators like interleukin-1 (IL-1), tumor necrosis factor-alpha (TNF-α), and toll-receptor-mediated signals, PTX. PTX3 synthesis is produced close to the site of inflammation and is not dependent on cytokine production or liver [4, 5].
As PTX3 is not liver-dependent, it is probable superior to traditional biomarkers for mirroring rapid inflammatory courses and may be a faster acting ac-ute-phase protein. Elevated serum concentration of PTX3 predicts multiple organ failure and infections [6, 7]. Therefore, the present study aimed to assess the applicability of pentraxin3 as early diagnostic markers in neonatal sepsis.
Methods
The aim of this study was to evaluate concentration of serum PTX3 l in neonatal sepsis at time of diagnosis and compare serum PTX3 level and serum CRP in a case-control study in eligible full term and late preterm neonates admitted in the NICU of Suez Canal University between first of December 2018 and 1st of September 2019. The subjects were divided into two groups; patients group (Group 1) including full term and late preterm neonates with neonatal sepsis and (Group 2) including age and sex matched healthy full and late preterm neonates.
Study groups
Participants were divided into two groups. Group 1 included neonates diagnosed with neonatal sepsis s based on clinical, laboratory and positive blood culture results. The clinical findings included the presence of three or more of the following: temperature instability, respiratory alterations, cardiovascular alterations, neurologic alterations, gastrointestinal alterations. CRP values > 6 mg/L were considered to be positive. Preterm neonates and neonates with suspected metabolic disease were excluded from the study population. Group 2 included neonates with no symptoms, signs, or risk factors for infections who had been hospitalized for physiological jaundice in the neonatal unit, and all neonates and with negative CRP level for neonates in control group was negative (CRP < 6 mg/L).
Sample size
Sample size was calculated according to the fol-lowing equation [8]: Zα/2 2 (SD)2/d2 where: Zα/2 = 1.96 (The critical value that divides the central 95% of the Z distribution from the 5% in the tail). SD: Standard deviation of Mean plasma PTX3 concentration of newborns = 7.83μg/L [9]. D: absolute error or precision = 2μg/L [9]. According to above equation the sample size will be 65 neonates after adding 20% dropout. Convenience sample was used; group 1 consists of 35 patients suffering from sepsis, group 2 consists of 35 normal serving as control group.
Data collected
Neonates included in the study were subjected to the following: Complete history, laboratory investigations namely complete blood count and C-reactive protein. Serum C-reactive protein was determined by standard nephlometric method. Blood culture: One to two ml of blood was added to blood culture media (Biphasic) and incubated at 37°C for 5–7 days. Bottles with positive results were sub cultured on blood agar (Himedia, India) and EMB media. The isolated microbes were identified by standard bacteriological methods. Measurement of pentraxin 3 levels: Plasma was separated from EDTA-anticoagulated blood samples by centrifuging in 2700 g for 10 min and stored at –80°C until analyzed. PTX3 was determined by enzyme linked immunosorbent assay (ELISA) kits (Adipo Bioscience®, CA, USA). The concentrations were interpolated from standard curves expressed in ng/mL.
Statistical analysis
Results were analyzed using SPSS (ver. 22.0; IBM, Chicago, IL, USA). Quantitative data were displayed in the form of mean±standard deviation (SD). Qualitative data were demonstrated through figures of frequency and percentage. Charts of different types were used to illustrate data and relations where app-ropriate and p < 0.05 was accepted as indicating statistical significance. The following tests were done: t-test was used when comparing between two means. Man Whitney U-test of significance was used when comparing between two means (Non-parametric). Chi-square (X2) test (Fisher exact) of significance was used in order to compare proportions between two qualitative parameters. Spearman rank correlation between significance relations. ROC (receiver operating characteristic) curve was used to detect cutoff value of Pentraxin 3 (PTX3) to diagnose neonatal sepsis. A probability value of p-value <0.05 was considered statistically significant.
Consent
Written informed consent was obtained from parents of all participants with full explanation of hazards and benefits of the procedure and ethical app-roval is obtained from ethical committee, medicine, Suez Canal University, Egypt.
Results
There were 70 neonates enrolled in this study, divided equally in to two groups, each has 35 neonates as designated by the study protocol after excluding 25 neonates.
The demographic data of the study population and comparison between the two study groups revealed statistically significant difference between two gro-ups in gestational age, weight and mode of delivery as shown in Table 1.
Characteristics of the studied neonates
Characteristics of the studied neonates
*Statistically significant at p < 0.05 1Chi-square test used.
The distribution of all 70 neonates from two groups according to age as showed below in group 1, the neo-nates more than 7 days was bigger (85.7 %) than less than 7 days (14.3%) while in group 2, the two groups were nearly equal. Group 1 neonates were more than female (51.4 %) while in-group 2 female neonates were more than male (51.4%). In-group 1 preterm neonates were bigger than term neonates were with (54.3%) while in-group 2 full term neonates were bigger (80 %).
Neonates weighed less than 2.5 kg (57.2%) were bigger than in sepsis while in-group 2 neonates wei-ghted more than 2.5 kg (80%) were bigger. In-group 1 the percentage of neonates who were delivered by normal vaginal delivery was (54.3 %) in opposite the percent of cesarean section was (91.4 %) in healthy neonates.
At time of presentation of the diseased neonates, 19 (54.3%) of them presented with respiratory distress only, while 10 (28.8. %) presented with refusal of feeding, 3 (8.4%) presented with pneumonia, 2 (5.7%) presented with neonatal jaundice and 1 presented with hematemesis. While in-group 1, 28(80%) presented with jaundice and six (20%) presented with respiratory distress with statistically significant difference between both groups. 17 (48.6%) diseased neonates were put on oxygen support as following; six (17.2%) of them were put on nasal cannula and 11 (31.4%) on mechanical ventilator.
All diseased neonates had positive blood culture. Most cases of neonatal sepsis were Staphylococci 55% were the most frequent isolated pathogens, followed by E. coli 20%, Klebsiella pneumonia 20% and Serratia marcescens 5%.
Table 2 shows statistically significant differences between both groups in serum CRP values in group 1 and 2 respectively (Mean = 49.3±37.4 mg/L, 26.8±17.2 mg/L, p < 0.05) and pentraxin values in diseased and control group (Mean = 5.2±3.7 mg/L, 2.3±0.78 mg/L, p < 0.05), and here were statistically significant correlations between pentraxin and serum CRP concentrations (p < 0.05). Table 3
Relation between CRP and Pentraxin in both groups
*Statistically significant at p < 0.05 1Man Whitney U-test used.
Correlation between pentraxin and Serum CRP in diseased group
*Statistically significant at p value < 0.05.
Receiver-operator characteristics (ROC) curve for pentraxin in diseased group.
In this study, ROC curve was run and showed that pentraxin demonstrated good diagnostic accuracy in predicting neonatal sepsis [AUC = 0.875 (95% CI 0.868 –1.000)] with sensitivity of 100% and specificity of 94.3%. Fig. (1)
The problem of neonatal sepsis is still a common cause of NICU mortality with a higher incidence in developing country than in advanced countries [10].
Globally, infections account for two-thirds of the annual deaths in children who are younger than 5 years. Still, the neonatal period is considered the highest lifetime risk of serious infections, [11]. In developing countries like Egypt, the rates of sepsis exceed 25 % in neonatal intensive care units as found by a study done by Medhet et al. with increased mortality and morbidity [12].
The delay in diagnosis and management
In the present study, the difference in mean serum CRP concentrations between septic neonates and controls were statistically significant. CRP values must be interpreted in the context of an infant’s clinical condition and not used alone to guide clinical decision-making [13]. Hisamuddin et al. [10] study have showed good CRP validity in the prediction of sepsis with low sensitivity and specificity of 58.33% and 56.52% respectively.
The similar results were observed by Mather NJ et al. [14] that showing a sensitivity rise from 22% to 61% with increasing time after admission. Wagle and Colleagues [15] studied the role of CRP in sepsis in preterm neonates and documented that the sensitivity/specificity of CRP on Day 1 was 62% and 87.7% increasing up to 70.2 and 97% on Day 2 of admission.
Even though CRP is one of the most commonly used diagnostic tool used to diagnoses neonatal sepsis however, in the interpretation of an increased serum CRP concentration during the first days of life, especially in preterm is questionable [16].
Pentraxins are endogenous substances that elaborated by macrophages and the complement system. These complex biological responses to tissue injury are part of the innate defense system [17]. Pentraxin 3 has been used as a reliable biomarker in the evaluation of cardiovascular disease and pulmonary hypertension in previous pediatric studies [4].
C-reactive protein and serum amyloid P component are the prototype of the short pentraxin family, which are acute phase proteins produced in the liver in response to inflammatory signal. However, their specificity [4] and poor prognostic ability for mortality [18] limit their diagnostic value in sepsis.
For instance, several studies have reported high level of serum CRP in burns patients with or without septic complications [19, 20].PTX-3 is the prototype of the long pentraxin family, whose members were first identified in the 1990s as cytokine inducible genes or molecule expression in specific tissues [21]. PTX-3 was then under the spotlight of investigation and has been established as an essential pattern recognition receptor at the crossroad between innate immunity, inflammation and matrix deposition [22].
The usefulness of PTX-3 in assessing the sepsis severity has also been established by various studies, the correlations is especially strong in critically ill patient, for example in an ICU setting. Different studies demonstrate a positive correlation between PTX-3 plasma level and severity of inflammatory conditions, including fungal infection, autoimmune disease tuberculosis [22–24].
In this study, we have found that the difference in pentraxin values between septic neonates and controls was statistically significant, and ROC curve showed that pentraxin demonstrated good diagnostic accuracy in predicting neonatal sepsis with significant sensitivity and specificity.
In a study by Hamed et al. [25] revealed that valuable and consistent discrimination of sepsis and septic shock was achieved by measurements of PTX-3 on day one, the marker being higher in the sepsis cohort.
In this study, there were statistically significant correlations between pentraxin and serum CRP concentrations in-group 1 with nonspecific clinical presentation including temperature instability, bradycardia and apnea, and hypotension in critically ill neonates with sepsis similar to adrenal insufficiency presentation [26–29].
To conclude, PTX3 represent a helpful tool in diagnosis of sepsis in the neonate. Still we need larger studies to prove our conclusion.
Conflict of interest
None
Consent for publication
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Ethics
The study obtained ethical approval and procedures were in accordance with the guidelines of by Suez Canal University ethics committee. Written informed consent was obtained from all parents after full explanation of the benefits & hazards of the procedures that will be carried out for each patient before getting them involved in the study.