Diagnostic Utility of Cord Blood Serum Procalcitonin in Early Onset Neonatal Sepsis in Comparision to Sepsis Screening Parameters

Abstract

Neonatal sepsis is an important cause of neonatal deaths globally. Diagnosis of neonatal sepsis is established based on microbiological tests of sepsis screen and clinical status. Mid phase markers of inflammation like CRP & Serum Procalcitonin are considered useful and sensitive for diagnosis. Most of the studies evaluating serum PCT as a diagnostic marker for neonatal septicemia have been carried out in peripheral venous blood with smaller sample sizes with inclusion of neonates without considering perinatal sepsis score. This hospital based, prospective study compares the diagnostic utility of cord blood Procalcitonin (PCT) with venous blood PCT; alone and as part of sepsis screening parameters currently in use in perinatal sepsis score positive neonates. Statistical analysis for cord blood Serum Procalcitonin (PCT) for detecting blood culture positive patients showed that PCT has a sensitivity of 44.4%, a high specificity of 86.4%, a low PPV of 33.3% and a high NPV of 91.1%. Overall diagnostic accuracy is 80.9%, indicating that cord blood PCT is a good test for identifying these patients (p < 0.05). However, venous blood PCT failed to demonstrate similar results.

Conclusions- Umbilical blood sampling protects the neonates from pain of venipunctures. Cord blood PCT estimations have statistically significant correlation with blood culture and other sepsis screen parameters and better sensitivity and specificity than venous blood PCT. This early serological biomarker is valuable for the diagnostic armamentarium of neonatal septicemia for early diagnosis and management while awaiting blood culture reports and helps in reducing separation of probable sepsis neonates from mother, thus contributing in developmental supportive care.

Introduction

Neonatal sepsis causes significant morbidity and 30%-50% of total neonatal deaths in developing countries. South Asia accounts for 3.5 million cases per year of the 6.9 million global neonatal sepsis burdens [1]. India, with its 1.5 billion population, has a large proportion of this disease burden. For diagnosis of clinical sepsis C-reactive protein (CRP), Total leukocyte count (TLC), ANC, immature/total neutrophil ratio (I:T ratio), platelet count and microbiological test combined with clinical status of the patient are parameters being used currently [2]. Screening of serological markers like mid phase marker (Procalcitonin/PCT) and late phase marker (CRP) is considered useful and sensitive indicators for diagnosis. The half-life (25-30 hours) and serum/plasma stability of PCT make it an easy parameter for routine use as compared to other inflammatory cytokines [3]. Most of the studies on role of serum PCT as a diagnostic marker for neonatal septicemia have been carried out in peripheral venous blood with smaller sample sizes.

         Blood sample is drawn by painful venipuncture (repeated multiple times if unsuccessful/inadequate) from the neonates for culture and hematology. Adequate umbilical cord blood can be obtained in the labor or delivery room by noninvasive and nontraumatic process, babies would experience less pain and rapid institution of antibiotic therapy is possible [4]. We planned the present study aimed to evaluate the diagnostic efficacy of serum Procalcitonin (PCT) alone and in combination with other diagnostic modalities for detection of early onset neonatal sepsis in cord blood and to compare the diagnostic utility of Procalcitonin, with neonatal sepsis screening parameters in venous blood; with an appropriate sample size as per statistical calculation, to obtain statistically relevant results. 

Materials And Methods

This hospital based, prospective study was conducted at the Department of Pediatrics, S.M.S medical college, Jaipur and attached group of hospitals providing obstetric services; over a period from April 2020 to 2021 after getting approval from institutional ethics committee.

Sample size (247 subjects) was calculated at 95% confidence interval to verify an expected 80.43% sensitivity of PCT for diagnosis of culture positive sepsis at absolute allowable error of 9%; assuming Prevalence of culture positive sepsis to be 30.4% as reported by previous studies.

Newborns delivered with positive perinatal sepsis score (Suspected sepsis), were enrolled after applying inclusion and exclusion criteria with prior informed consent. Neonates with Major congenital anomaly, Meconium aspiration syndrome, respiratory distress syndrome (RDS), Gestational diabetes mellitus, Pneumothorax, Intracranial hemorrhage and negative consent were excluded. 

Perinatal sepsis score (PSS) was determined by maternal history and the presence of risk factors like Low birth weight (< 2500 grams) or prematurity, Febrile illness in the mother with evidence of bacterial infection within two weeks prior to delivery, Foul smelling liquor, Rupture of membrane>24hours, Single unclean or >3 sterile vaginal examination(s) during labor, Prolonged labor (duration of 1st and 2nd stage of labor ≥24 hours), Perinatal asphyxia (Apgar score < 4 at 1 minute). Presence of two risk factors or more was subjected to sepsis screen. Presence of foul-smelling liquor or three of the above-mentioned risk factors warrant initiation of antibiotics [5].

In Neonates with suspected sepsis cord blood serum PCT, Sepsis screen and blood culture were sent at the time of birth.   They were observed for 72 hours, for signs and symptoms, suggestive of sepsis. Based on clinical history and examination subjects were divided in two groups: Group 1 (cord blood) and Group 2 (venous blood). Group 1, based on cord blood investigations was subdivided into three groups: Group a -Sepsis screen negative, Blood culture negative (No sepsis), Group b -Sepsis screen positive, Blood culture negative (Probable sepsis) and Group c -Sepsis screen positive, Blood culture positive (Proven sepsis).

In Group 2: in subjects with signs, and symptoms suggestive of sepsis, venous blood sample (2^nd sample) was withdrawn within 72 hours and sent for serum PCT, sepsis screen and blood culture. Based on 2^nd sample Group 2 was subdivided into three groups: Group a-Sepsis screen negative, Blood culture negative (Clinical sepsis), Group b-Sepsis screen positive, Blood culture negative (Probable sepsis) and Group c-Sepsis screen positive, Blood culture positive (Proven sepsis).

Umbilical cord blood sampling- Under strict aseptic precautions, umbilical cord blood samples were taken from 247 newborns (Group 1) with maternal high-risk factors for early onset of sepsis or positive perinatal sepsis score; venous blood samples were taken from 40 newborns (Group 2) with clinical signs & symptoms suggestive of sepsis within 72 hours. Umbilical cord blood was collected at birth, after resuscitation (if needed). Post-delivery, the umbilical cord was clamped on both the placental and the umbilical end (approximately 2-4 cm distance from the infant side) and was cut between each pair of clamps. The placental end (approximately 5 cm) was wiped with 70% isopropyl alcohol and with a 22-gauge needle and syringe 6-7 ml blood was collected. 2 ml of cord blood sample was collected from the umbilical cord just after the delivery of the baby in one plain uncoated glass tube for PCT, CRP (quantitative), 2-3 ml of cord blood EDTA glass tube for CBC WITH DLC, I/T RATIO, Micro ESR. Needle from syringe was replaced with a new sterile needle and 2-3 ml of cord blood injected into aerobic blood culture bottle after wiping the top of blood culture bottle with 70% isopropyl alcohol, followed by povidone-iodine, and followed again by 70% isopropyl alcohol. 5-6 ml of Venous blood sample was collected from the peripheral vein. 

STATISTICAL ANALYSIS: Categorical / Nominal variables were expressed as number and percentage and were analyzed using Chi square test. ROC curve was drawn to determine the area under the curve (with 95% confidence interval) for various parameters for diagnosis of sepsis. Sensitivity, specificity, Positive predictive value, negative predictive value, and diagnostic accuracy were calculated by using 2x2 contingency table. A p value <0.05 was taken as statistically significant. All statistical analysis was done using SPSS trial version 20.

Results

Baseline characteristics of study participants are depicted in Table:1.

Table: 2 summarizes Serum Procalcitonin (PCT) levels in relation to sepsis screen in Group 1 and Group 2. 154 subjects (62.34%) were PSS equal to two and 93 subjects (37.66) were PSS more than two. In Group 1, Out of 247; sepsis screen was positive (≥2) in 52.23% subjects and negative (<2) in 47.77% subjects. In group 2, 17 subjects (42.5%) were sepsis screen negative and 23 subjects (57.5%) were sepsis screen positive.

Serum Procalcitonin (PCT) levels in Group 1 and group 2 newborns according to sepsis screen and blood culture are depicted in Table 2 &3.

Serum Procalcitonin (PCT) levels in Group 1 newborns according to sepsis screen and blood culture- For detecting probable sepsis newborns PCT had a test sensitivity 47.1%, a higher specificity 86.4% and low PPV of 33.3%. overall diagnostic accuracy was found to be 81.5% indicating the PCT is a good test for diagnosing EOS (p value< 0.002). In probable sepsis serum PCT estimation has a high NPV (91.9%), this is the peak of high specificity of this test for excluding neonatal sepsis. On comparing Group-a, and Group c the PPV was 69.2% and NPV was 57.3% which only give that estimation of serum PCT values have reasonably good sensitivity (Sn) and specificity (Sp). For detecting probable sepsis (SS positive, blood culture negative) newborns, PCT has a sensitivity of 47.1%, indicating that it can only correctly identify 47.1% of these patients. A high specificity of 86.4% indicates that PCT is good for excluding these cases. A low PPV of 33.3% indicates that if PCT is raised, then only 33.3% patients will have SS positive, blood culture negative. A high NPV indicates that if PCT is low then 91.9% patients will not have SS positive, blood culture negative. Overall diagnostic accuracy is 81.5%, indicating that PCT is a good test for identifying these patients (p <0.05). 

Serum Procalcitonin (PCT) levels in Group 2 newborns according to sepsis screen and blood culture- In group 2; The sensitivity (Sn), specificity (Sp) and positive predictive value (PPV) and negative predictive value (NPV) and diagnostic accuracy (DA) and p value of Group b were respectively 61.5%, 47.1%, and 47.1%, 61.5% and 53.3%, p value>0.334, statistically not significant between Group a (clinical sepsis) and Group b (Probable sepsis) for PCT. Out of 10 subjects in Group c, 40% subjects had Serum PCT<0.5 ng/mL and 60% subjects had values ≥0.5 ng/mL. The sensitivity (Sn), specificity (Sp) and positive predictive value (PPV) and negative predictive value (NPV) and diagnostic accuracy (DA) and p value were respectively 60%, 47.06% and 40%,66.7% and 51.8%, p value>0.721 of Group c, statistically not significant between Group a (clinical sepsis) and Group c (Proven sepsis) for PCT.

Relationship of Serum Procalcitonin (PCT) levels in Group 1 and 2 newborns with blood culture are depicted in Table 3.  

Serum Procalcitonin (PCT) levels in Group 1 newborns according blood culture- Serum PCT equal or more than 0.5 ng/mL was found in 31.15% cases of blood culture positive subjects. The sensitivity, specificity and PPV and NPV and diagnostic accuracy (DA), p value respectively was 44.4%,86.4% and 33.3%,91.1% and 80.09%, p value <0.005(s), statistically significant for PCT in cord blood. For detecting blood culture positive patients, PCT has a sensitivity of 44.4%, indicating that it can only correctly identify 44.4% of these patients. A high specificity of 86.4% indicates that PCT is good for excluding these cases. A low PPV of 33.3% indicates that if PCT is raised, then only 33.3% patients will have blood culture positive. A high NPV indicates that if PCT is low then 91.9% patients will not have blood culture positive. Overall diagnostic accuracy is 80.9%, indicating that PCT is a good test for identifying these patients in cord blood (p <0.05). 

Serum Procalcitonin (PCT) levels in Group 2 newborns according to blood culture- However, venous blood (Group 2) PCT failed to demonstrate similar results. in Group 2(n=40); Blood culture was positive in 25% cases. Serum PCT levels <0.5 ng/mL in 4 (40%) subjects and ≥0.5 ng/mL in 6 (60%) subjects out of 10 blood culture positive subjects. Serum PCT levels < 0.5 ng/mL in 13 (43.44%) and ≥0.5 ng/mL in 17 (56.66%) subjects, out of 30 blood culture negative subjects. (Table 4). Venous blood serum PCT levels were increased in 60% of blood culture positive newborns and 56.66 % in blood culture negative subjects. P value= 0.853 is not significant. Diagnostic utility of venous blood serum PCT showing sensitivity, specificity, PPV, NPV and Diagnostic accuracy with blood culture taken as a gold standard were 60%,43.3%,26.1%,76.5% and 47.5%. P value is not significant for PCT in venous blood; therefore, it cannot replace the blood culture as gold standard test.

Comparison of Receiver operating characteristic curve (ROC) of PCT, CRP, TLC, ANC, m-ESR and Platelets for diagnosis of early onset of sepsis in Group 1 newborns (Figure:1): On Receiver operating characteristic curve (ROC) analysis, cord blood serum PCT, TLC, ANC and platelet showed statistically significant area under curve (AUC) value ranging from 0.587 to 0.624 (95% CI). P value is highly significant for PCT and TLC, significant in ANC and platelet.

Comparison of Receiver operating characteristic curve (ROC) of PCT, CRP, TLC, ANC, m-ESR and Platelets for diagnosis of early onset of sepsis in Group 2 newborns (Figure:2): On ROC curve analysis, venous blood m-ESR showed statistically significant area under curve (AUC)value 0.686 significant (95%CI) and p value is significant (0.046) for m-ESR. Serum PCT, CRP, TLC and Platelets did not show significant area under curve (AUC >0.5). 

On ROC curve analysis, area under curve (AUC) value were found to be significant for PCT, TLC, ANC and platelet counts (AUC>0.5 values between 0.587 to 0.624 - 95% confidence interval); PCT and TLC had the highest diagnostic value (p value<0.001 and p < 0.004) and ANC & Platelets also had significant diagnostic values (p<0.021 and p<0.018).   speaking about their diagnostic value for EOS in cord blood samples, while in venous blood sample only micro-ESR had shown significant area under curve value more than 0.5 (0.686 -95% CI and significant p value 0.046) while no other parameters had shown significant area under curve (AUC<0.5). (Figure-1,2)

In summary, slightly higher percentage positive cut off value of PCT (29.04%) was observed in newborns with strongly positive perinatal sepsis screen compared with positive perinatal sepsis score that is 21.43%, the difference was statistically not significant. Newborns with sepsis screen positive status had shown a significantly higher percentage (34.11%) having PCT value equal or more 0.5ng/mL, while only 13.55% of sepsis screen negative newborns had shown positive cutoff value. Our results show a significant role of PCT in diagnosing early onset of sepsis (EOS) in cord blood samples, but PCT estimation failed to demonstrate similar diagnostic value in newborns of Group 2 (p value=0.859). For detecting probable sepsis newborns had a PCT test sensitivity 47.1%, a higher specificity 86.4% and low PPV of 33.3%. overall diagnostic accuracy was found to be 81.5% indicating the PCT is a good test for diagnosing EOS (p value< 0.002). However, PCT estimation failed to demonstrate similar results in venous blood samples taken from neonatal sepsis. Cord blood PCT was increased in 32.15% blood culture positive newborns and 17.77 % in blood culture negative subjects and P value (p<0.005) is highly significant. Diagnostic utility of Cord blood serum PCT showing sensitivity, specificity, PPV, NPV and Diagnostic accuracy with blood culture taken as a gold standard were 44.4%,86.4%,33.3%,91.1% and 80.9%. However, venous blood (Group 2) PCT failed to demonstrate similar results. On ROC curve analysis, area under curve (AUC) value were found to be significant for PCT, TLC, ANC and platelet counts (AUC>0.5) speaking about their diagnostic value for EOS in cord blood samples, while in venous blood sample only micro-ESR had shown significant area under curve value more than 0.5 while no other parameters had shown significant area under curve (AUC<0.5). 

Discussion

Male newborns are more (144: 58.29%). As per UN (World Population Prospectus 2019), the sex ratio of the total population in India is 108.18 males per 100 females [6]. High male predominance was observed in some studies [7,8,9]; other studies report high female sex predominance [10,11].

        Caesarean births are very high (46.96%) than WHO, UNICEF and UNFPA guidelines of desired proportion of Caesarean births (5 to 15%) [12].  Based on DLHS-3 (District Level Household and Facility Survey-3) data, the overall caesarean section rate in India is 9.2% and in Rajasthan 4.2%.[13]; this hospital being a tertiary care center, catering complicated deliveries referred from small peripheral institutions and factors   like decreasing acceptance for fetal risk (e.g., routine Caesarean for breech presentation), decreasing tolerance for perineal trauma (Caesarean instead of forceps delivery), over-estimation of risk with labor after prior Caesarean (decreased Vaginal Birth after Caesarean rates, vaginal twin delivery) are the reasons. Similar results were observed by Abdel Hakeem et al and other studies [7,10,14]. 

            Preterm subjects (87 ;35.22%) are more than WHO normal ranges.  According to WHO, every year about 15 million babies are born prematurely around the world and the rate of preterm birth ranges from 5% to 18% of babies born. In India, the rate of preterm birth is 12.96%. Similar results were found in other studies [9,10,12, 15,]. The proportion of LBW (24.70%) is low, as compared to incidence of LBW of Rajasthan (32%). Similar results were observed in other studies [12,15]. The incidence of LBW is estimated to be around 15% worldwide with a range of 3.3-38% in developing countries. The incidence is very high in India (23%) [16]. 87.45% subjects are AGA, 20 (8.09%) subjects are LGA, and 11 (4.46%) subjects are SGA. The 4.46% incidence of SGA in present study is within usual range as most of the south Asian countries have 5.3% to 41.5%SGA births [17]. No other study has mentioned such classification based on Birth weight in relation to gestational maturity & perinatal sepsis score (PSS) and religion-based distribution, so the results cannot be compared.

             Serum procalcitonin levels (PCT) in Group 1 subjects in relation to sepsis screen and blood culture in Olivia Oria de et al reported that in ‘no sepsis’ neonates’ serum PCT was positive in 4.77% while 100% probable sepsis & proven sepsis neonates were PCT positive [18]. Similar results noted in Noemie Huetz et al study [19]. 

         Serum PCT was also increased in the 17.77% case of blood culture negative subjects. Raised PCT in blood culture negative cases may be due to physiological rise of PCT within 96 hours of birth or due to non - infective cause. Serum PCT in Group 2 subjects in relation to blood culture was also reported by other workers [8,20]. 67.85% subjects with positive blood culture had low PCT (<0.5ng/mL). This may be because of antibiotics administration during antepartum and intrapartum period, which may affect PCT concentration in umbilical cord blood; even postnatal administration of antibiotics is known to decrease PCT concentration [21].

          In our study higher results of positive sepsis screen are due to higher proportion of preterm subjects (35.22%), higher percentage of high-risk perinatal sepsis score (PSS>2) subjects (37.66%), and various maternal factors like poor nutrition, anemia, low maternal education, and low socioeconomic status. Other studies [7,19] report low positive sepsis screen, but these studies were done in developed countries having better socioeconomic status of families.

             In Group 2, out of 40 subjects, 17 (42.5%) subjects were having sepsis screen negative (<2) and 23 (57.5%) subjects were having sepsis screen positive (≥2). Similar results observed by other studies [9,10,15,28]. Slightly lower results were observed in some studies [11,12,22].

           Olivia Oria de et al study reported no sepsis, probable sepsis and proven sepsis in 92.64%,5.88% and 1.48% neonates; and Noemie Huetz et al study reorted 98.74%,1.23%,0.03% respectively [18,19]. The higher number of no sepsis neonates, lower number of probable and proven sepsis neonates in these studies reflects overall higher quality of health services, higher maternal education, good health education and high socioeconomic status.

            In Group 2 ;42.5% in Group a (clinical sepsis) subjects suggests need to confirm sepsis and start precautionary antibiotics. In Group b (probable sepsis) 32.5% and in Group c (proven sepsis) in 25%; is due to enrolled neonates having positive PSS and in this group neonates with signs & symptoms suggestive of sepsis were included. Naher BS et al study reports clinical sepsis probable sepsis and proven sepsis in 38%, 22% and 20%; & Seema Shah et al study reports as 21%, 16.5% and 11.5% respectively [8,23,24]. We included neonates with positive PSS, while they included all the neonates.

          The higher value of PCT positive newborns in positive PSS subjects in our study is most probably due to high number of preterm and low birth weight. Olivia Oria de et al study reported that serum PCT levels were negative in 120 (88.23%) & positive in 16 (11.77%) subjects, out of 136 subjects [18]

           Relationship in Serum procalcitonin levels (PCT) and sepsis screen reported by Noemie Huetz et al as 57 subjects had serum PCT levels ≥0.6 ng/ml with positive sepsis screen in 16(28.07%) and negative in 41(71.93%) & serum PCT levels < 0.6 ng/ml in 3023 subjects, with positive sepsis screen in 23(0.76%) and negative in 3000 (99.24 %), other studies also depicted similar results [7,19,25,26,29]. Variations in the results of studies by different workers are due to respective cut-off values. In our study, we have taken sepsis negative subjects as control and compared the PCT values (cut -off value ≥0.5 ng/mL) with sepsis screen positive subjects.  

Similar results of Serum PCT in group 2 (venous blood) subjects in relation to sepsis screen observed by Haniya jafar et al in their study, serum PCT value<0.5 ng/ml in 44(36.66%) and serum PCT value > 0.5 ng/ml in 76(63.34%) subjects out of 120 sepsis screen positive subjects; while Om Shankar et al found serum PCT value positive in 77(96.25%) subjects (very high %) and negative in 3(3.75%) subjects, out of 80 positive sepsis screen subjects.  [20,27]. 

        In present study the percentage of serum PCT positive is much higher (probable sepsis 47.05%, proven sepsis 32.15%) again this can be explained by inclusion of high-risk newborns showing positive perinatal sepsis score. Serum procalcitonin levels (PCT) in Group 1 subjects in relation to blood culture revealed a lower sensitivity (44.4%) and PPV (33.3%) in our study but a very good specificity (86.4%) and NPV (91.1%) was seen in our study so this test can serve as good mean to exclude neonatal sepsis in early neonatal sepsis in newborns. 

Conclusions

PCT is an early marker of neonatal sepsis with good diagnostic value, high NPV and high specificity (Sp); which make PCT a test of utility to exclude neonatal sepsis. PCT estimation in cord blood has shown a better specificity and sensitivity in comparison to venous blood collected on day 1 to 3. PCT test has shown better sensitivity and specificity in newborns having more perinatal risk factors and higher number of sepsis screen parameter positivity in their serum. PCT qualifies to be an important early serological biomarker in the diagnostic armamentarium of neonatal septicemia. Therefore, it should become an essential component of all the sepsis screen tests in all institutions. For specific analysis and to detect the sensitivity, specificity, PPV, NPV and diagnostic accuracy it is recommended that multicentric, multinational studies are to be conducted with uniform reagent kits and similar cut-off values, so that its actual status can be arrived at.

Declarations

Funding: None of the authors has taken any financial support from anyone 
Conflicts of interest/Competing interests: None
Availability of data and material: Yes
Code availability: Not applicable
Authors' contributions: CL drafted the study design, collected data, contributed in statistical analysis and inference of results. RS supervised the entire work, drafted the study design, collected data, contributed in statistical analysis and inference of results. DRB did critical analysis of all data, counselled the parents and hypothesized the study, drafted the study design, collected data, contributed in statistical analysis and inference of results. NS drafted the conclusions, reviewed the literature, drafted the study design, collected data, contributed in statistical analysis and inference of results   
Ethics approval: This study was approved by clinical trial screening committee and ethics committee of SMS Medical College, Jaipur.                 
Consent to participate: Consent of parents was taken.
Consent for publication:  not applicable     
 

Abbreviations

ANC-Absolute Neutrophil Count

AUC -Area under curve

BC-Blood Culture

CBC-Complete Blood Count

CRP-C-reactive protein

EOS- early onset sepsis

I/T Neutrophil ratio- immature to total neutrophil ratio

LOS-late onset sepsis

micro ESR-erythrocyte sedimentation ratio

NPV-Negative Predictive Value

PPV-Positive Predictive Value

PSS-Perinatal sepsis score

PCT- serum procalcitonin

ROC-Receiver operating characteristic curve

Sn-Sensitivity

Sp-Specificity

SS-sepsis screen

TLC -Total leukocyte count

UPT- Up-converting phosphor technology

References

1. Seale AC, Blencowe H, Manu AA, Nair H, Bahl R, Qazi SA, et al. Estimates of possible severe bacterial infection in neonates in sub-Saharan Africa, south Asia, and Latin America for 2012: a systematic review and meta-analysis. Lancet Infect Dis. 2014; 14:731–741. [PubMed: 24974250] 
2. Fleischmann-Struzek C, Goldfarb DM, Schlattmann P, Schlapbach LJ, Reinhart K, Kissoon N. The global burden of paediatric and neonatal sepsis: a systematic review. Lancet Respir Med. 2018; 6 (3):223–230. https://doi.org/10.1016/S2213-2600(18)30063-8 PMID: 29508706.
3. Liu L, Oza S, Hogan D, Chu Y, Perin J, Zhu J, Lawn JE, Cousens S, Mathers C, Black RE. Global, regional, and national causes of under-5 mortality in 2000–15: an updated systematic analysis the Sustainable Development Goals. The Lancet.2016;388 (10063):3027-35.
4. Hansen A Potential substitution of cord blood for infant blood in the neonatal sepsis   evaluation. BiolNeonate.2005;88(1): 128.Epub2005Feb10. 
5. Singh M, Narang A, Bhakoo ON. Predictive perinatal score in the diagnosis of neonatal sepsis. J Trop Pediatr. 1994 Dec;40(6):365-8. 
6. World Population Prospects - Population Division - United Nations Inbox, https://population.un.org/wpp/Download/Standard/Population/
7. Kordek A, Hałasa M, Podraza W. Early detection of an early onset infection in the neonate based on measurements of procalcitonin and C-reactive protein concentrations in cord blood. Clin Chem Lab Med 2008; 46:1143-1148.
8. Seema Shah, Anil Kumar Goel, Renu Garg, Mamta Padhy, Amit Gupta, Procalcitonin and C-reactive protein in early diagnosis of neonatal sepsis, Indian Journal of Medical Specialities, Volume 5, Issue 2, 2014, Pages 102-106, ISSN 0976-2884, https://doi.org/10.1016/j.injms.2015.08.002.(https://www.sciencedirect.com/science/article/pii/S0976288415000703)
9. Neeraj Kumar, R. Dayal, D. Agrawal, P. Kumar, R. Bhatia, A. Goyal, D. Verma. “Is Procalcitonin a better Diagnostic Marker than CRP in Neonatal Sepsis?” Journal of Evolution of Medical and Dental Sciences 2014; Vol. 3, Issue 12, March 24; Page: 3140-3147, DOI: 10.14260/jemds/2014/2257
10. Predictive values for procalcitonin in the diagnosis of neonatal sepsis Abdel Hakeem Abdel Mohsen, Bothina Ahmed Kamel Electron Physician. 2015 Aug; 7(4): 1190–1195. Published online 2015 Aug 10. doi: 10.14661/2015.1190-1195 PMCID: PMC4578539
11. Eisa O. El-Amin Abdalla, Fathya A.M. Salih, Haytham F. Salih, Osman E. Elamin, Mutaz A. Gamaleldin & Babikir M. Mustafa | Tsai-Ching Hsu (Reviewing Editor) (2017) Procalcitonin in the diagnosis of early-onset neonatal infection in resource-limited settings, Cogent Medicine, 4:1,      
     DOI: 10.1080/2331205X.2017.1283085
12. UNICEF,WHO,UNFPA,New York: United Nations Children’s Fund, 1997,1.
13. International Institute for Population Sciences (IIPS), 2010. District Level Household and Facility Survey (DLHS-3), 2007-08: India. Mumbai: IIPS.
14. Umran RMR,Hashim JM, Jameel H. Significance of Serum Procalcitonin Level in the Early Diagnosis of Neonatal Sepsis. Iranian Journal of Neonatology. 2020 Mar: 11(3). DOI: 10.22038/ijn.2020.42296.1702
15. Diagnostic value of procalcitonin in neonatal sepsis | Nigerian Journal of Paediatrics AO Arowosegbe et al, Vol. 43 No. 1 (2016) DOI: 10.4314/njp.v43i1.3
16. Preterm birth | National Health Portal Of India, http://www.nhp.gov.in/disease/reproductive-system/female-gynaecological-diseases-/preterm-birth.
17. Koyanagi A, Zhang J, Dagvadorj A, Hirayama F, Shibuya K, Souza JP, et al. Macrosomia in 23 developing countries: an analysis of a multicountry, facility-based, cross-sectional survey. Lancet. 2013;381(9865):476–83.
18. Oria de Rueda Salguero O, Beceiro Mosquera J, Barrionuevo González M, Ripalda Crespo MJ, Olivas López de Soria C. Procalcitonina en sangre de cordón en la valoración del riesgo de sepsis neonatal precoz [Cord blood procalcitonin in the assessment of early-onset neonatal sepsis]. An Pediatr (Barc). 2017 Aug;87(2):87-94. Spanish. doi: 10.1016/j.anpedi.2016.09.003. Epub 2016 Oct 14. PMID: 27751752.
19. Potential Impact of Umbilical-Cord-Blood Procalcitonin-Based Algorithm on Antibiotics Exposure in Neonates with Suspected Early-Onset Sepsis. Front Pediatr. 2020; 8: 127. Published online 2020 Apr 17. doi: 10.3389/fped.2020.00127 PMCID: PMC7181674 PMID: 32363168 Noémie Huetz, Elise Launay, Géraldine Gascoin, Bertrand Leboucher, Christophe Savagner, Jean B. Muller, Sophie Denizot,Cécile Boscher,Jocelyne Caillon, Damien Masson, and Christèle Gras Le Guen.
20. Jafar H, Agarwal J, Kalyan RK, Radera S, Verma S, Kumar M, Tripathi S. Role of serum procalcitonin as a marker for diagnosing neonatal sepsis. Int J Contemp Pediatr 2019;6:1991-7.
21. Stocker M, Hop WC, van Rossum AM. Neonatal Procalcitonin Intervention Study (NeoPInS): Effect of Procalcitonin-guided decision making on duration of antibiotic therapy in suspected neonatal early-onset sepsis: A multi-centre randomized superiority and non-inferiority Intervention Study. BMC pediatr. 2010;10(1):89.
22. Liu S, Hou Y, Cui H. Clinical values of the early detection of serum procalcitonin, C-reactive protein and white blood cells for neonates with infectious diseases. Pak J Med Sci. 2016 Nov-Dec;32(6):1326-1329. doi: 10.12669/pjms.326.11395. PMID: 28083019; PMCID: PMC5216275. 
23. Park IH, Lee SH, Yu ST, Oh YK. Serum procalcitonin as a diagnostic marker of neonatal sepsis. Korean J Pediatr. 2014;57(10):451-456. doi:10.3345/kjp.2014.57.10.451
24. Naher BS, Mannan MA, Noor K, Shahiddullah M. Role of serum procalcitonin and C-reactive protein in the diagnosis of neonatal sepsis. Bangladesh Med Res Counc Bull. 2011 Aug;37(2):40-6. doi: 10.3329/bmrcb. v37i2.8432. PMID: 21877603.
25. Joram N, Muller JB, Denizot S, Orsonneau JL, Caillon J, RozéJ C, et al. Umbilical cord blood procalcitonin level in early neonatal infections: a 4-year university hospital cohort study. Eur J Clin Microbiol Infect Dis 2011; 30:1005-1013.
26. Lencot S, Cabaret B, Sauvage G, Laurans C, Launay E, Orsonneau JL, Caillon J, Boscher C, Roze JC, Gras-Le Guen C. A new procalcitonin cord-based algorithm in early-onset neonatal infection: for a change of paradigm. Eur J Clin Microbiol Infect Dis. 2014 Jul;33(7):1229-38. doi: 10.1007/s10096-014-2065-3. Epub 2014 Feb 11. PMID: 24515097. 
27.  Akter, J., Ahammad, F., & Begum, T. (2018). Procalcitonin – A Reliable Marker for Diagnosis of Neonatal Sepsis in Compare to CRP. Bangladesh Journal of Child Health, 42(1), 19-25. https://doi.org/10.3329/bjch.v42i1.37046
28. Chaurasiya OS, Ahmad T, Chhabra K, Nath D, Shobhne HJ: Procalcitonin Level in Neonatal Sepsis. PJSR ;2017:10(1):29-32. 
29. Joram N, Boscher C, Denizot S, Loubersac V, Winer N, Roze JC, et al. Umbilical cord blood procalcitonin and C reactive protein concentrations as markers for early diagnosis of very early onset neonatal infection. Arch Dis Child Fetal Neonatal Ed 2006;91: F65-F66.

Tables

Table 2: Serum Procalcitonin (PCT) levels in relation to sepsis screen in newborns of Group 1&2 

*SS-sepsis screen

Table 3: Relationship of Serum Procalcitonin (PCT) levels in Group 1 and group 2 newborns with sepsis screen and blood culture 

Table 4: Serum Procalcitonin (PCT) levels in Group 1 and 2 newborns according to blood culture