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Early-onset sepsis remains a common and serious problem for neonates, especially preterm infants. Group B streptococcus (GBS) is the most common etiologic agent, while Escherichia coli is the most common cause of mortality. Current efforts toward maternal intrapartum antimicrobial prophylaxis have significantly reduced the rates of GBS disease but have been associated with increased rates of Gram-negative infections, especially among very-low-birth-weight infants [1,2]. The diagnosis of neonatal sepsis is based on a combination of clinical presentation, the use of nonspecific markers, including C-reactive protein and procalcitonin (where available), blood cultures and the use of molecular methods, including PCR. Cytokines, including interleukin 6 (IL-6), interleukin 8 (IL-8), gamma interferon (IFN- ) and tumor necrosis factor alpha (TNF-) and cell surface antigens, including Soluble Intercellular Adhesion Molecule (sICAM) and CD64, are also being increasingly examined for use as nonspecific screening measures for neonatal sepsis [3]. Viruses, in particular enteroviruses, parechoviruses and Herpes Simplex Virus (HSV), should be considered in the differential diagnosis [4]. Empirical treatment should be based on local patterns of antimicrobial resistance but typically consists of the use of ampicillin and gentamicin or ampicillin and cefotaxime if meningitis is suspected, until the etiologic agent has been identified. Current research is focused primarily on development of vaccines against GBS.

It encompasses various systemic infections of the newborn such as septicemia, meningitis, pneumonia, pyogenic arthritis, osteomyelitis and urinary tract infections. Sepsis is the commonest cause of neonatal mortality; it is responsible for about 30-50% of the total neonatal deaths in developing countries. It is estimated that up to 20% of neonates develop sepsis and approximately 1% die of sepsis related causes. Sepsis related mortality is largely preventable with rational antimicrobial the rapid and aggressive supportive care [5].

Sola et al. showed that the likelihood of developing thrombocytopenia increases with the degree of prematurity [6]. Roberts et al., calculated that low-birth weight infants were at a 2.52-fold increased risk for thrombocytopenia [7]. Thrombocytopenia develops in 18-35% of neonates admitted to intensive care units. In addition, the rate and severity of thrombocytopenia in neonates of pregnancy induced hypertensive mothers vary. Chakravorty et al. demonstrated in those pregnancies complicated by preeclampsia that the usual course of thrombocytopenia includes diagnosis within the first 2-3 days and resolution by 7-10 days of life in most cases [8].

Platelet production or thrombopoiesis, is a complex process that results in the production of thrombopoietin as the thrombopoietic stimulus leading to the generation and proliferation of megakaryocyte progenitors.

Platelet Transfusions (PT) in Neonatal Thrombocytopenia (NT) are commonly administered to reduce the risk of bleeding. However, there are few evidence-based guidelines to inform clinicians’ decision-making processes. Developmental differences in hemostasis and differences in underlying disease processes make it difficult to apply PT practices from other patient populations to neonates [9]. Specifically, it is important to identify neonates at risk of bleeding who would benefit from PT and to determine whether PT either abrogate or exacerbate common neonatal complications such as sepsis, chronic lung disease, Necrotizing Enterocolitis (NEC) and retinopathy of prematurity. Among 972 Very- Low-Birth Weight (VLBW) infants from a multicenter retrospective cohort study, Sparger et al. reported that 231 (24%) had received a total of 1002 PT. A large proportion of PT were given to VLBW infants with platelet counts greater than 50,000/μL. Additionally, they found that the severity of illness influenced transfusion decisions. However, the severity of NT did not correlate with the risk of Intraventricular Hemorrhage (IVH) and in contrast, PT did not reduce the risk of IVH [10].

In a large multi-center study, Wiedmmeier et al. defined neonatal thrombocytopenia as a platelet count less than 150,000 cells/mm³ [11]. Neonatal thrombocytopenia has been categorized into two groups depending on the time of onset: early onset, which is within 72 h of life and the late onset, after 72 h of life [12,13]. The degree of severity of thrombocytopenia can be further subcategorized according to platelet count in affected individuals: Mild thrombocytopenia-platelet count 100000 to 150000 cells/mm³, moderate thrombocytopenia-platelet count 50000 to 99000 cells/mm³, severe thrombocytopenia-platelet count <50000 cells/mm³ [11].

The classification of sepsis based on time of appearance as EOS or LOS is important as it helps in determining the most probable organism and mode of transmission and guide for empiric treatment. Thrombocytopenia is one of the early but non-specific indicators of neonatal sepsis. It can be caused by bacterial, viral, fungal and parasitic infections and other non-infectious causes. Bleeding is a major complication of thrombocytopenia but I is generally limited to infants with count <30000 cells/mm³. Studies have shown that approximately 50% cases of culture proven sepsis have thrombocytopenia virtually any organism capable of causing sepsis can induce thrombocytopenia. Changes in other platelet indices, like MPV (Mean Platelet Volume) and PDW (Platelet Distribution Width) have also been examined. Increased Platelet Volume (IPV) indicates an increased proportion of young platelets in the circulation [14]. The present study aimed to know the incidence of thrombocytopenia and to evaluate the feasibility of thrombocytopenia as a screening tool for neonatal sepsis in a risky neonate.

Aims and objectives

The present study aimed to know the incidence of thrombocytopenia in neonatal sepsis and to evaluate the feasibility of neonatal thrombocytopenia as a screening tool for neonatal sepsis.

This hospital based prospective observational cross sectional study was conducted in Neonatal Intensive Care Unit in Universal College of Medical Sciences, a tertiary care hospital over a period of 3 months, from May 2019 to July 2019. Inclusion criteria were all newborn admitted to UCMS-TH NICU with screening positive sepsis or clinically suspected sepsis and neonates presence of 2 or more risk factor positive were considered as suspected sepsis were included. Other neonates were excluded from study according to exclusion criteria. A total number of 205 neonates suspected sepsis under the age of 28 days admitted in NICU, were studied and included in our study.

Investigations done-sepsis screening

Absolute Neutrophil Count (ANC)<1800 cells/mm³ Immature to Total Neutrophil (I/T)

RATIO>0.2(immature neutrophils/ANC), highly sensitive of NNS

Total Leukocyte Count (TLC)<5000 cells/mm³ or >20,000 cells/ mm³ CRP>6 mg/dL

MICRO ESR (μ-ESR)>15 mm/1^st h, specific but moderate sensitivity. Platelets<150000 cells/mm³.

Blood culture

NOTE: (≥ 2) positive screening parameters (TLC, ANC, I/Ratio, CRP, u-ESR), platelets counts. Taken as sepsis screen positive (Sn 93%, PPV 39%, >2 parameters-NPV 99%) and that neonate is with sepsis.

Exclusion criteria

MOTHER with History s/o ITP, SLE/ other autoimmune disorders, on medication during pregnancy (Sulfonamides, Quinine/Quinidine) (Thiazides, Tolbutamide, Vancomycin, Hydralazine and Heparin) Neonate with history s/o bleeding disorder in family, trisomies, Turner /Noonan’s syndromes, TAR syndrome. Conditions were associated with sequestration of platelets. (Kasabach–merritt syndrome with gianed haemangiomas, renal vein thrombosis, polycythemia, CCHD, placental vascular thrombi- PIH/preeclampsia/eclampsia). Severe Rh-HDN (marked erythropoiesis is in bone marrow → Neutropenia and thrombocytopenia) massive bleed from causes like birth trauma, accidental slipping of cord clamp causing hemodynamic disturbance/ exchange transfusion (dilutional NNT). Sick neonate with RVT, CHD, Congenital leukemia. Neonate who had received IV antibiotics for ≥ 48 h prior to our study.

Ethical clearance

The approval of Institutional Review Committee of Universal College of Medical Sciences, Bhairahawa, Nepal was taken before the initiation of experiment. Registration No. UCMS/IRC/118/19. All the protocols and experiments were conducted in compliance with the ethical principles and guidelines

Statistical analysis

Data analysis was done using Statistical Package for Social Sciences (SPSS) software version 14.0. The level of significance for tests was set at p<0.05. Since our variables is categorical variables and to find the association between two categorical variables we have to use Chisquare test for all category.

A total of 205 neonates with clinical sepsis were included for the study and were evaluated accordingly.

During the study period, a total 205 newborn with clinical sepsis were admitted. There were 128 (62.4%) Hindu, 53(25.9%) Muslim and others were 24 (11.7%). 148 (72.2%) male and 57 (27.8%) female neonate with male to female ratio of 2.5:1. However 108 (52.7%) were delivered by normal vaginal delivery and 97 (47.3%) were delivered by caesarean section. maternal risk factor for sepsis were 61 (29.8%) and without risk factor for sepsis 144 (70.2%). PV leakage were present in 22 (10.7%) cases. 39 (19%) neonate were born low birth weight and 166 (81%) were normal weight (>2.5 kg). 38 (18.5%) were preterm, 159 (77.6%) were term baby and 8 (3.9%) cases were post-term baby (Table 1).

Table 1: Demographic data of neonatal sepsis (n=205).

Table 2 showed out of 205 suspected neonatal sepsis cases shows 81 (39.5%) thrombocytopenia, in which mild is 50 (24.4%), moderate is 25 (12.2%), severe 6 (2.9%) (Table 2).

Table 2: Severity of neonatal sepsis.

The total 205 cases were evaluated for sepsis, the blood culture was positive in 51 (24.9%) cases. In which coagulase negative Staphylococci (CONS) 22 (43.3%) and Kleibsella species 12 (23.52%) were the most common Gram positive and Gram negative organisms (Table 3).

Table 3: Distribution of isolated organism.

Blood culture and sensitivity platelets count category showed that: Out of 205 case, 51 (24.87%) cases of thrombocytopenia were present in culture positive cases in which: 1 (2.0%) was severe, 12 (23.5%) were moderate, 12 (23.5%) were mild and 26 (51.0%) were normal type and remaining 154 (75.12%) cases were present in sterile cases out of which 5 (3.2%) were severe, 13 (8.4%) were moderate, 38 (24.7%) were mild and 98 (63.6%) were normal type. The chi square value is 8.403 and its p-value is 0.038 and it is statically significant (Table 4).

Table 4: Blood culture and sensitivity* platelets count category cross tabulation.

Thrombocytopenia in mode of delivery showed that, out of 205 cases, 108 (52.68%) were seen in normal vaginal delivery among which 5 (4.6%) were severe, 6 (5.6%) were moderate, 27 (25.0%) were mild and 70 (64.8%) were normal and remaining 97(47.31%) cases seen were in lower segment cesarean section among which 1(1.0%) was severe, 19 (19.6%) were moderate, 23 (23.7%) were mild and 54 (55.7%) were normal type. The chi square value is 11.814 and p value is 0.008 and it is statistically significant (Table 5).

Table 5: Thrombocytopenia in mode of delivery.

Thrombocytopenia in weight for age showed that out of 205 cases, 31(15.12%) were showed in SGA case of which:- 1(3.2%) severe, 4 (12.9%) moderate and 12 (38.7%) mild type, remaining 171 (83.41%) were seen in AGA cases out of which: 5 (2.9%) severe, 21 (12.3%) moderate and 35 (20.5%) mild type and other 3 (1.46%) cases were showed in LGA and all of them were mild type. The chi square value is 13.567 and p value is 0.035 and it is statistically significant (Table 6).

Table 6: Thrombocytopenia in weight for age.

Thrombocytopenia in C-Reactive Protein showed out of 205 cases 113 (55.12%) were seen in positive among which 6 (5.3%) were severe, 12 (15%) were moderate, 28 (24.8%) were mild type and 62 (54.9%) were normal type and remaining 92 (44.87%) were seen in CRP negative among which 8 (8.7%) were moderate, 22 (23.9%) were mild and 62 (67.4%) were normal type. The chi square value is 10.198 and its P value is 0.017 and it is statistically significant (Table 7).

Table 7: Thrombocytopenia in C-reactive protein.

The aim of the study was to evaluate the causes for NT, the duration of NT and the indications and effects of PT by means of a retrospective cohort study. we conducted this study for 3 months periods at UCMS NICU Unit it was shown that occurrence of thrombocytopenia in the newborn treated at NICU was 39.5%, which proves that frequency of thrombocytopenia in our region is still high among the newborn hospitalized at the NICU. A similar study carried out in Indonesia showed that thrombocytopenia in the children hospitalized at NICU was 12% [15]. Gram positive bacteria coagulase negative was isolated as the most frequent cause of thrombocytopenia in the new born in our study (43.13%) and gram negative bacteria i.e. Kleibseilla was found to be second common cause of thrombocytopenia in the newborn (23.52%). Riedler et al. found that an 80% incidence of thrombocytopenia in gram negative septicemia and 65% of incidence in gram positive septicemia [14].

From all the findings from this study, we can conclude our discussion that was in blood culture and sensitivity test, of thrombocytopenia was found approx. 24.87% cases in blood culture positive and approx. 75.12% in sterile cases. Similarly thrombocytopenia in normal vaginal delivery was found to be approx. 52.68% and about 47.13% in lower segment cesarean section showing that result was high in former one. Also thrombocytopenia was found maximum in appropriate for gestational age (AGA) i.e. 83.41% followed by SGA (15.12%) and then in LGA (1.46%). Thrombocytopenia was found more in CRP Positive cases approx. 55.12% followed by CRP Negative cases which was approx. 54.9%. Guida et al. had reported that 54% septic Very Low Birth Weight (VLBW) neonates developed thrombocytopenia [16]. Khair et al. studied ‘Role of Hematologic Scoring System in Early Diagnosis of Neonatal Septicemia they found that platelet count <1,00,000 cells/ mm³ had a sensitivity of 60%, specificity 82%, PPV 31% and NPV 94% [17]. A cross sectional analytical study on CRP and hematological parameters in NNS, in military hospital, Rawalpindi 4, over 7 months. It included 100 clinically septic and 100 normal neonates and observed that NNT has 64.3% sensitivity in detecting NNS. Culture, although not specific. NNT is almost equal in normal (49.38%) and low birth weight (50.62%) neonates. NNT can be used as screening tool in NNS as it is easy and cost effective [18]. It requires further large scales studies and meta-analysis to validate useful in developing countries like India. In the present study, blood culture positivity was observed in 51 (24.9%) neonates. Thrombocytopenia was found in 81 septic neonates (39.5%). These findings indicate that low platelet count is important finding in bacterial septicemia. Further it was also observed that thrombocytopenia was noted in majority of cases in which blood culture was negative. Therefore, it was observed from the study that platelet count is an important indicator of septicemia and not related with blood culture, although not specific.

It was observed that the study of platelet count is an important indicator of septicemia and not related with blood culture and sensitivity. In the present study was found that NNT (<1,50,000 cells/ mm³ ) can be used to screen neonate with sepsis (NNS) Especially in at risk neonates which is cost effective and available in almost all hospitals. Apart from that attention should be paid on the early detection of thrombocytopenia in the newborn hospitalized at the NICU because development of severe infection and at the same time high mortality in the risky group of the newborn can be prevented. Thrombocytopenia in neonatal sepsis increases the risk of mortality nearly four-fold, with another six-fold increase in mortality in case of gram negative sepsis.

Authors would like to thank the department of Pediatric and Neonatal intensive Care and also thankful to Laboratory staff Unit of Universal College of Medical Sciences Bhairahawa, Nepal.