PLATELET-TO-LYMPHOCYTE RATIO IN EARLY ONSET NEONATAL

SEPSIS

Abstract
Objectives
Neonatal sepsis is a leading cause of infant morbidity and mortality worldwide. Although blood
culture is the gold-standard of sepsis diagnosis, test sensitivity and diagnostic speed offer
challenges in accurate and timely diagnosis and treatment. Platelet-to-lymphocyte ratio (PLR) is
a widely available, effective, simple, and affordable marker that can be performed to predict
early onset neonatal sepsis (EONS) without specific qualification. The goal of this study is aimed
to find the association between PLR and EONS.
Methods
All inpatient neonates in the study hospital who were treated suspected to have EONS were
included in the study. Subjects were then categorized into sepsis and non-sepsis groups based on
clinical findings, laboratory results, and blood culture. The independent variable was PLR and
the dependent variable was sepsis occurrence. Bivariate analysis and point-biserial correlation
coefficient were used to identify any significant association between PLR and EONS.
Results
Out of 193 enrolled subjects, 135 (69%) had confirmed sepsis with positive blood cultures, and
55.6% were males. The blood cultures were dominated by Klebsiella pneumoniae (42.2%).
There was a higher PLR in the sepsis group (61.90 ± 18.30) in comparison to the non-sepsis
group (53.72 ± 20.37) subjects. There was a positive association between PLR and EONS (P
<0.001) with point-biserial correlation coefficient of 0.491 (P <0.001).
Conclusion
This study found that there is a significant positive association between PLR and the incidence of
EONS. PLR can be used as a reliable, inexpensive, easily calculated marker in predicting EONS.
Authors suggest using cut-off point of 45.6 as the value to predict EONS.
Keywords: early-onset neonatal sepsis, PLR, sepsis predictor, cut-off
Corresponding author: Rocky Wilar, Department of Child Health, Universitas Sam Ratulangi/
Prof Dr. R. D. Kandou Hospital Manado. Jalan Raya Tanawangko No. 56, Manado, 95115,
North Sulawesi, Indonesia. Phone/fax: (0431) 821652/(0431)859091.

Introduction
Neonatal sepsis is one of the leading causes of morbidity and mortality globally, accounting for
over 500,000 neonatal deaths per year.1,2,3 In 2009, the World Health Organization (WHO)
reported 3.3 million neonatal deaths out of a total of 6 million infant deaths.7 According to the
2012 Indonesian Health Demographic Survey, the infant mortality rate in Indonesia is 32 deaths
per 1.000 live births.4,5 The United Nations International Children’s Emergency Fund (UNICEF)
reports a neonatal mortality rate of 14 deaths per 1000 live births in Indonesia.6
Early diagnosis of neonatal sepsis is critical to managing the disease and improving the patient’s
prognosis. A late diagnosis is linked to an increased mortality risk of the neonate and worse
prognosis.4 Blood culture has been the gold-standard of sepsis diagnosis, but test sensitivity is a
concern, and processing speeds can be very slow. This can lead to a delay in treatment resulting
in worse clinical outcomes or even death. However, if the culture shows a false positive,
overtreatment will increase antibiotic use and hospitalization days, decreasing cost-effectiveness.
These problems indicate a need for other diagnostic approaches to increase the timeliness and
accuracy of sepsis diagnosis.8
Platelet-to-lymphocyte ratio (PLR) is a marker with a test that is widely available, effective,
simple, affordable, and can be performed without specific qualification.9-11 Several studies have
found that increased PLR is associated with an increased morbidity and mortality in patients with
severe sepsis and septic shock.12-14 This study has been conducted to assess the association
between PLR and Early Onset Neonatal Sepsis (EONS).
Methods
This is an analytic observational study with cross-sectional design. The study took place in the
NICU of Neonatology Division, Department of Paediatrics, Sam Ratulangi University/R.D.
Kandou Hospital, Manado, North Sulawesi, Indonesia from May to September 2019. The study
was conducted under the approval of Research Ethics Committee of R.D. Kandou Hospital
Manado with the number of 077/EC-KEPK/VI/2019.
Study Participants
The study population were all inpatient neonates treated initially for suspected EONS at NICU,
Department of Paediatrics, Sam Ratulangi University/R.D. Kandou Hospital, Manado, North
Sulawesi, Indonesia. Suspicion of EONS was indicated by the presence of two major or one
major and two minor risk factor criteria for neonatal sepsis within the first 72 hours after birth.
Major criteria include: prolonged rupture of membranes >18 hours, maternal intrapartum fever
>38oC, chorioamnionitis, sustained foetal heart rate >160 bpm, and foul-smelling amniotic fluid.
Minor criteria include prolonged rupture of membranes >12 hours, maternal intrapartum fever
>37.5oC, low APGAR scores, very low birth weight <1,500 grams, untreated maternal
leukorrhea, and untreated maternal urinary tract infection.4,15
Inclusion criteria for this study were all neonates with suspected EONS that were born vaginally
or operatively. Neonates with congenital postoperative defects, birth trauma, or congenital heart
disease were excluded from the study. By using a correlative sample size formula, researchers
calculated that the minimum sample size is 92 subjects. Sampling method was done by
consecutive sampling.

Definitions and Outcome Measures

Neonatal sepsis is defined by having three categories of criteria:
1. The presence of clinical abnormalities in 4 of 6 systems (respiratory, cardiovascular,
metabolic, neurologic, gastrointestinal, or hematologic system)15-17
2. Blood test indicating two of the following hematologic profiles (Hb <15 g/dL, white
blood cell >25,000/mm3 or <5,000/mm3, platelet <100,000/mm3, CRP >6 mg/dL, or
immature-to-total (I/T) ratio >0.2)18 AND
3. Positive blood culture results.
Patients who did not fulfil these criteria were categorized into the non-sepsis group.
All neonates with suspected EONS based on the major and minor criteria listed above were
enrolled into the study. After blood culture results were available, subjects were categorized into
the sepsis or non-sepsis group. The main outcome of this study is observing the correlation
between the occurrence of culture-proven sepsis and the independent variable (PLR). PLR was
calculated by dividing absolute platelet count with absolute lymphocyte count.
Statistical Analyses
The occurrence of sepsis is expressed as a binominal variable into sepsis and non-sepsis. PLR is
a numeric variable expressed as Mean ± Standard Deviation (SD). Bivariate analysis was used to
identify any significant association between PLR and EONS. For all analyses, P <0.05 is
considered as statistically significant. The point-biserial correlation coefficient was also
calculated to find the strength and direction of the association between those variables.
All statistical analyses were done using IBM SPSS version 25.0 (IBM Corp. IBM SPSS Statistics
for Windows, Armonk, NY).

Results
This study was conducted between May and September 2019. A total of 193 neonates with
suspected EONS were included. Within this study population, 105 were males (54.4%) and 88
were females (45.6%). After blood culture results were available, 135 neonates were confirmed
to have sepsis (69%) while the other 58 were categorized as non-sepsis (31%). In the sepsis
group, 75 were males (55.6%); while in the non-sepsis group, there were 30 males (51.7%).
Within the sepsis group, the most common microorganisms found in blood cultures were
Klebsiella pneumoniae (42.2%), Candida albicans (10.4%), Serratia marcescens (6.7%),
Staphylococcus aureus (6.7%), Enterobacter aerogenes (5.9%), and Escherichia coli (4.4%).
Distribution of these microorganisms can be seen on Table 1.
 Table 1. Microorganisms found in blood culture results

Percentage
Microorganism N
(%)

Acinetobacter baumannii 5 3.7

Acinetobacter junii 2 1.5

Candida albicans 14 10.4

Candida pelliculosa 2 1.5

Candida tropicalis 1 0.7

Elizabethkingia meningoseptica 1 0.7

Enterobacter aerogenes 8 5.9

Enterobacter cloacae complex 1 0.7

Enterococcus faecium 1 0.7

Escherichia coli 6 4.4

Klebsiella pneumoniae 57 42.2

Listeria monocytogenes 1 0.7

Micrococcus luteus 1 0.7

Methicillin-resistant staphylococcus
1 0.7
aureus

Pseudomonas aeruginosa 2 1.5

Salmonella spp 1 0.7

Serratia marcescens 9 6.7

Staphylococcus aureus 9 6.7

Staphylococcus epidermidis 7 5.2

Staphylococcus haemolyticus 4 3.0

Staphylococcus hominis 1 0.7

Staphylococcus saprophyticus 1 0.7

Total 135 100

 Table 2. Distribution and statistical analyses of PLR among groups of study population

Category N Minimum Maximum Mean ± SD P-value

Sepsis 135 21.25 92.30 62.88 ± 12.88

< 0.001
Non-sepsis 58 18.68 87.50 43.19 ± 15.19

SD = Standard Deviation

The PLR among subjects with sepsis (62.88 ± 12.80) was higher than in the non-sepsis group
(39.19 ± 15.19) (Table 2). By using logistic regression, it was found that there was a very
significant association between PLR and the incidence of EONS (P<0.001). By observing the
tendency of the curve (Figure 1), it can be concluded that PLR was positively associated with
EONS. Point-biserial correlation coefficient was rpb = 0.626 with P-value <0.001. This result
means that there is a significant positive correlation between PLR and the incidence of EONS
(Figure 2).
Probability of Sepsis

PLR

Figure 1. Scatterplot of the Association Between PLR and Sepsis Probability

 PLR

Figure 2. Box-Whisker plot distribution of PLR by EONS category

Discussion
In this study, sepsis was found slightly more in males than females. This finding is consistent
with previous research conducted by Juniatiningsih, et.al. in Jakarta, Indonesia, which reported
that male neonates suffered from sepsis more than females.19 This result is also consistent with
research done by Putra20 and Kardana21 in Indonesia.
A majority the positive cultures revealed bacteria as the infectious pathogen, whereas 12.5% of
patients were infected by fungi (Candida albicans, Candida pelliculosa, and Candida tropicalis).
The most common microorganism found was Klebsiella pneumoniae (42.2%), consistent with
the results from research done by Patel, et.al.23 and Kayange et.al.24. Blood culture remains the
“gold-standard” in diagnosing neonatal sepsis, but a negative result cannot rule out the presence
of sepsis. A positive culture merely indicates the presence of bacteremia or contamination. Blood
culture is sometimes difficult to use as a diagnostic tool because of its long waiting period (3 to 5
days).17,25 Therefore, the authors sought to find another accurate, fast alternative to diagnose
EONS.
Platelets and lymphocytes are important components in the immune system that provides the first
line of defence against infection. In sepsis, activated platelets secrete proteins like cytokine,
chemokine, coagulation mediator, and antimicrobial peptides. Activated platelets also combine
with neutrophils to increase the secretion of neutrophil extracellular trap, which will induce more
coagulation.26,27 As infection progresses, antigen-presenting cells and lymphocytes interact and
lymphocytes move from the vascularization into the infection site, which presents as
lymphocytopenia. In sepsis patients, apoptosis of the lymphocyte is also found. 28,29 Between the
movement of lymphocytes to the infection area and apoptosis of lymphocytes, sepsis patients
present with lymphocytopenia, which is the basis for using PLR in predicting the diagnosis of
sepsis in adults.30 Studies about PLR usage in predicting neonatal sepsis is still rare.
This study finds a significant positive association between PLR and EONS. A prior study by
Can, et.al. found that subjects with EONS have significantly higher neutrophil count, axillary
temperature, neutrophil-to-lymphocyte ratio, PLR, CRP, and procalcitonin levels.31 Another
study by Arcagok and Karabulut found that neonates with confirmed and suspected EONS have
significantly higher PLR compared to the control group of healthy newborns.32 This Arcagok
study stated that PLR has an area under the curve (AUC) of 0.89 to 0.93. The cut-off points for
using PLR to predict suspected EONS was 39.5 (sensitivity 88.9%, specificity 94.7%) and 57.7
for confirmed cases of EONS (sensitivity 91.3%; specificity 97.6%). Similarly, our study found
the predictive cut-off value in patients who are suspected to have EONS was a PLR of 45.6
(sensitivity 89.6%; specificity 79.3%; AUC 0.879; P < 0.001).

Research Limitations
There were several limitations that can be taken into consideration for further study. One
limitation is that this study did not separate term and preterm neonates to determine separate cut-
offs for the two groups. Further research is recommended to determine cut-off values for
different groups to better diagnose EONS in neonates.

Conclusion
There is a strong positive association between PLR and the incidence of EONS. PLR can be used
as a reliable, easily calculated marker for diagnosing EONS. Authors suggest using cut-off point
of 45.6 as the value to predict EONS in patients with suspected EONS.

References
1. Modell B, Berry RJ, Boyle CA, Christianson A, Darlison M. Global, regional, and national
causes of child mortality. Lancet. 2012;380:1251-61. DOI:10.1016/S0140-6736(12)60560-
1
2. Mathur NB. Neonatal Mortality and Infections: Global Perspective. In: Mathur NB, editor.
Neonatal Sepsis. New Delhi: Elsevier; 2010. p.8-9.
3. Black RE, Cousens S, Johnson HL, Lawn JE, Rudan I, Basani DG, et al. Global, Regional
and National Causes of Child Mortality in 2008: A Systematic Analysis.
Lancet.2010;375:1969-87. DOI: 10.1016/S0140-6736(10)60549-1.
4. Aminullah A. Sepsis in Neonate. In: Kosim MS, Yunanto A, Dewi R, Sarosa GI, Usman
A, editors. Neonatology Textbook, 1st Ed. Jakarta: IDAI, 2012. p.170-87.
5. Indonesian National Demographic and Health Survey 2012. Available from:
http://kesga.kemkes.go.id/images/pedoman/SDKI%202012-Indonesia.pdf
6. UNICEF. Maternal and Newborn Health Disparities in Indonesia. 2015. Accessed March
20th, 2020.
7. World Health Organization. Neonatal and Infant Mortality: Global Health Observatory
(GHO). 2009. Available from: https://www.who.int/data/gho
8. McGowan KL, Foster JA, Coffin SE. Outpatient pediatric blood cultures time to positivity.
Pediatrics 2000;106:251-5. DOI: 10.1542/peds.106.2.251.
9. Balta S, Ozturk C. The platelet-lymphocyte ratio: A simple, inexpensive, and rapid
prognostic marker for cardiovascular events. Platelets 2015;26:680–1.
DOI: 10.3109/09537104.2014.979340.
10. Yazar FM, Bakacak M, Emre A, Urfalıoglu A, Serin S, Cengiz E, et al. Predictive role of
neutrophil-to-lymphocyte and platelet-to-lymphocyte ratios for diagnosis of acute
appendicitis during pregnancy. Kaohsiung J Med Sci 2015;31:591–6.
DOI: 10.1016/j.kjms.2015.10.005.
11. Yavuzcan A, Cağ lar M, Ustün Y, Dilbaz S, Ozdemir I, Yildiz E, et al. Mean platelet
volume, neutrophil-lymphocyte ratio and platelet-lymphocyte ratio in severe
preeclampsia. Ginekol Pol 2014;85:197–203. PMID: 24783431
12. Biyikli E, Kaymaz AE, Kavalci C. Effect of platelet–lymphocyte ratio and lactate levels
obtained on mortality with sepsis and septic shock. Am J Emerg Med 2018;36:647-50.
DOI: 10.1016/j.ajem.2017.12.010.
13. Yildiz A, Yigit A, Benli A.R. The prognostic role of platelet to lymphocyte ratio and mean
platelet volume in critically ill patients. Eur Rev Med Pharmacol Sci 2018;22: 2246-52.
DOI: 10.26355/eurrev_201804_14811.
14. Kartal O, Kartal AT. Value of neutrophil to lymphocyte and platelet to lymphocyte ratios
in pneumonia. Bratisl Lek Listy 2017; 118: 513-6. DOI: 10.4149/BLL_2017_099.
15. Tripathi S, Malik GK. Neonatal sepsis: past, present and future; a review article. Int J Med
2010; 5: 45-54. DOI: 10.4314/ijmu.v5i2.56163.
16. Polin RA. Committee on fetus and newborn. Management of neonates with suspected or
proven early onset bacterial sepsis. Pediatrics 2012;129:1005-16. DOI: 10.1542/peds.2012-
0541.
17. Shah BA, Padbury OF. Neonatal sepsis: an old problem with new insights. Virulence 2014;
5: 170-8. DOI: 10.4161/viru.26906.
18. Haque KN. Definitions of bloodstream infection in the newborn. Pediatr Crit Care Med
2005; 6: 45-9. DOI: 10.1097/01.PCC.0000161946.73305.0A.
19. Juniatiningsih A, Aminullah A. Firmansyah A. Microorganism profile of neonatal sepsis in
Child Health Department of Ciptomangunkusumo Hospital Jakarta. Sari Pediatri 2008; 10:
60-5. DOI: 10.14238/sp10.1.2008.60-5.
20. Putra PJ. Incidences and factors related to neonatal sepsis in Sanglah Hospital, Denpasar.
Sari Pediatri 2012; 14: 205-10. DOI: 10.14238/sp14.3.2012.205-10.
21. Kardana IM. Incidence and factors associated with mortality of neonatal sepsis. Paediatr
Indones 2011; 51: 144-8. DOI: 10.14238/pi51.3.2011.144-8.
22. Lawn JW, Katende KW, Cousens SN. Estimating the causes 4 million neonatal deaths in
the year 2000. Int J Epid 2006; 35: 706-18. DOI: 10.1093/ije/dyl043.
23. Patel D, Nimbalkar A, Sethi A, Kungwani A, Nimbalkar S. Blood culture isolates in
neonatal sepsis and their sensitivity in Anand District of India. Indian J Pediatr 2014; 1: 1-
6. DOI: 10.1007/s12098-013-1314-2.
24. Kayange N, Kamugisha E, Mwizamholya DL, Jeremiah S, Mshana SE. Predictors of
positive blood culture and deaths among neonates with suspected neonatal sepsis in a
tertiary hospital, Mwanza- Tanzania. BMC Pediatr 2010; 10: 1-9. DOI: 10.1186/1471-
2431-10-39.
25. Goldstein B, Giroir B, Randolph A. International consensus conference on pediatric sepsis.
Definitions for sepsis and organ dysfunction in pediatrics. Pediatr Crit Care Med 2005; 6:
2-8. DOI: 10.1097/01.PCC.0000149131.72248.E6.
26. Sacha F., Cornelis V, Poll T. The role of platelets in sepsis. Thromb Haemost
2014;112:666–77. DOI: 10.1160/TH14-02-0126.
27. Greco E, Lupia E, Bosco O, Vizio B and Montrucchio G. Platelets and multi-organ failure
in sepsis. Int J Mol Sci. 2017; 18: 2200. DOI: 10.3390/ijms18102200.
28. Shubin NJ, Monaghan SF, Heffernan DS, Chung CS, Ayala A. B and T lymphocyte
attenuator expression on CD4+ T-cells associates with sepsis and subsequent infections in
ICU patients. Crit Care 2013; 17: 1-33. DOI: 10.1186/cc13131.
29. Hotchkiss RS, Monneret G, Payen D. Sepsis induced immunosuppression: from cellular
dysfunction to immunotherapy. Nat Rev Immunol 2013; 13: 862-72. DOI:
10.1038/nri3552.
30. Kutlucan L, dkk. The predictive effect of initial complete blood count of intensive care unit
patients on mortality, length of hospitalization, and nosocomial infections. Eur Rev Med
Pharmacol Sci 2016; 20: 1467-73. PMID: 27160116.
31. Can E, Hamilcıkan S, Can C. The value of neutrophil to lymphocyte ratio and platelet to
lymphocyte ratio for detecting early-onset neonatal sepsis. J Pediatr Hematol Oncol 2018;
40: e229-32. DOI: 10.1097/MPH.0000000000001059.
32. Arcagok BC, Karabulut B. Platelet to lymphocyte ratio in neonates: A predictor of early
onset neonatal sepsis. Mediterr J Hematol Infec Dis. 2019; 11: e2019055.
DOI: 10.4084/MJHID.2019.055.