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Utility of Sepsis Screen in the Early Diagnosis of Neonatal Sepsis |
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Chandrashekhar P Bhale, Ap urva Vasant Kale, Sachin S Kale, Meera Mahajan, Smi ta SMulay 1. Professor, Department of Pathology, M. G. M. Medical College and Hospital, Aurangabad, India. 2. Resident, Department of Pathology, M. G. M. Medical College and Hospital, Aurangabad, India. 3. Associate Professor, Department of Pathology, M. G. M. Medical College and Hospital, Aurangabad, India. 4. Assistant Professor, Department of Pathology, M. G. M. Medical College and Hospital, Aurangabad, India. 5. Professor and Head, Department of Pathology, M. G. M. Medical College and Hospital, Aurangabad, India. |
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Correspondence Address : Dr. Apurva Vasant Kale, C/O Dr. V. M. Kale, R. H. No K-20, Tirupati Supreme Enclave, Jalan Nagar, Near Railway Station, Aurangabad, India. E-mail: Apurvakale33@gmail.com |
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| ABSTRACT |  | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
: Introduction: Neonatal sepsis could be defined as a clinical entity because of generalized bacterial infection within 28 days of life and showing a positive blood culture. It is probably responsible for 30- 50% of the total neonatal deaths each year. Timely diagnosis of neonatal sepsis is critical because in neonates the illness can progress more rapidly than adults. The blood culture report takes at least 72 hours. Therefore, a simple test with quick availability of results can be helpful to reduce neonatal morbidity and mortality. Aim: To evaluate the utility of sepsis screen in early diagnosis of neonatal septicemia and to study various hematological parameters, changes in peripheral blood smear, evaluate the performance of microerythrocyte sedimentation rate, serum C-reactive protein and serum direct bilirubin in neonates with clinical suspicion of sepsis. Materials and Methods: The present, study was done in our institute from October 2013 to October 2015. CBC was done on 191 clinically suspected cases of neonatal sepsis along with Micro ESR, Serum CRP and direct bilirubin. Differential leukocyte count, absolute neutrophil count, immature neutrophils: total neutrophils ratio was done from Field stained peripheral smears. Blood culture was done in Microbiology Department. Exclusion criteria were neonates with major congenital anomalies and those who have already received antibiotics. Statistical Analysis: Statistical analysis was done using SPSS software, version 20th and unpaired ‘t’- test. Result: Out of 191 cases studied, 91 were culture positive. CRP (84.6%) and immature : total neutrophils ratio(75.8%) showed highest sensitivity, Whereas absolute neutrophil count(99.0%) along with serum direct bilirubin (93.0%) and corrected total leucocyte count (93.0%)showed highest specificity. Positive predictive value was highest for absolute neutrophil count (97.5%) and CRP (84.8%). Conclusion: Serum CRP is the most sensitive marker of sepsis. Use of peripheral smear study, serum direct bilirubin and micro ESR together with CRP can be used effectively as a sepsis screen for early diagnosis of neonatal sepsis. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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| Keywords : Bilirubin, C - reactive protein, Leukocyte Count, Newborn | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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| INTRODUCTION |
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Neonatal sepsis could be defined as a clinical entity because of generalized bacterial infection within 28 days of life and showing a positive blood culture (1). It is probably responsible for 30-50% of the total neonatal deaths each year (1). Neonatal sepsis can be early or late onset. Onset of symptoms within 72 hrs of life is early onset, this period can be extended up to one week. Onset of symptoms later than that is late onset sepsis. Infections are more common in low birth weight and preterm babies (2). In neonates the illness can progress more rapidly than in adults, therefore early diagnosis is of utmost importance (2). Positive blood culture is a gold standard for diagnosis, but it is time consuming (requires 72 hours, atleast 24 hours in case of BacT- ALERT®) and demands a well equipped laboratory (3). Many investigators have evaluated various inflammatory markers. But these are sophisticated and impractical for developing countries [4-7]. A good diagnostic test should have high sensitivity and specificity and should be cost effective with early availability of results [4,5]. The parameters used are absolute neutrophil count, total leukocyte count, immature: mature neutrophil ratio, micro-erythrocyte sedimentation rate, C-reactive protein and serum direct bilirubin. They together can be used as sepsis screen. Presence of two or more abnormal parameters in case of strong clinical suspicion is considered as positive sepsis screen. The results can be obtained much earlier than blood culture and antibiotic therapy can be instituted early. This can be helpful to reduce neonatal mortality and morbidity (8). b#bClassification of Neonatal Sepsis:b?b Early onset sepsis (within first 72 hours of life) and late onset sepsis (more than 72 hours) (8). b#bAimb?b The aim of the present study was to study the various hematological parameters including various changes seen in the peripheral smears of neonates clinically suspicious of sepsis and to evaluate the performance of micro-erythrocyte sedimentation rate and C-reactive protein in neonatal sepsis. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| Material and Methods |
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The present study was a cross-sectional study carried out in MGM’s Medical College, Aurangabad for the duration of 2 years during period from October 2013 to October 2015 (prospective study). The study comprised of 191 neonates clinically suspected of neonatal sepsis. Ethical committee approval was obtained prior to the study. All the neonates (age 0 to 28 days) admitted to NICU, who presented with signs and symptoms of neonatal sepsis were included in the study and those who had received the antibiotics prior to admission and neonates with major congenital anomalies were excluded from the study. Sepsis screen included following tests: Absolute Neutrophil Count (ANC), C-reactive protein (CRP), immature: total neutrophil ratio (I:T ratio), serum direct bilirubin, micro erythrocyte sedimentation rate (microESR) and corrected total leukocyte count (Corrected TLC). Blood culture report was considered as a gold standard. About 3-4 ml of blood was drawn using all aseptic precautions, out of which 1 ml of the blood sample was inoculated aseptically into a yellow BacT/ALERT pediatric blood culture bottle, 1 ml of the blood, was allowed to clot in a plane vacutainer to collect serum for the estimation of C-reactive protein and direct bilirubin. The remaining 2 ml of blood was collected in a vacutainer containing the anticoagulant EDTA (2-2.5 mg/ml) for smear preparation and estimation of the Total WBC count, Absolute neutrophil count, I: T ratio. Total leukocyte count and absolute neutrophil count were noted from Advia 2120i automated cell counter. A drop of EDTA blood was taken on a clean dry slide and a thin tongue-shaped smear was made, air dried and stained with Field’s stain. The differential count and I: T ratio was calculated as per standard hematological methods. CRP was estimated using Siemens Dimension® clinical chemistry system for quantitative determination of C-reactive protein in serum. Micro -ESR was estimated with capillary blood obtained by heel prick, collected in a standard 75 mm microhematocrit tube with internal diameter of 1.1 mm. The air was not allowed to interrupt the column of blood to avoid false normal result and one end of the tube was sealed with 2-3 mm plasticin. The 45 degrees Set Square (rule) was used to draw a vertical line on the wall and the capillary tube was placed along that line. The rule was used to measure the distance from the highest point of the plasma column to the meniscus of the packed red cell column (9). Neonatal Sepsis screen was considered positive if any two criteria of the following were present (8). • Absolute Neutrophil Count of =1800/cumm • CRP =1 mg/dL. • I/T ratio =0.2 • Micro-ESR =15mm at the end of 1st hour • Serum direct bilirubin = 2mg/dL • Total Leucocyte Count (TLC) of =5000/cumm b#bStatistical analysisb?b All the study parameters were entered in the excel sheet and were analyzed using SPSS software, version 20. Sensitivity, specificity, positive predictive value and negative predictive value of septic screen was compared with culture outcome (gold standard). Unpaired t test was used to find p-value (p< 0.05 was considered significant). | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| Results |
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Total 191 cases were studied in the present study. None of the babies were excluded from the study. Out of 191 cases, 91 (47.47%) cases were blood culture positive, remaining 100 (52.3%) were reported as sterile. The distribution of cases among culture positive group was as shown in [Table/ Fig-1]. The percentage of male babies was more as compared to female in culture proven septicemia cases. The percentage of septicemia among low birth weight babies was considerably more than in normal birth weight babies. Preterm babies were more affected than the term babies. Early onset sepsis episodes were more than late onset. The babies born outside were affected more than the babies born inside NICU [Table/ Fig-2]. C-reactive protein showed highest sensitivity and negative predictive value. Absolute neutrophil count showed highest specificity and positive predictive value (Table/Fig 3). All the parameters showed a relation with blood culture report which was statistically very significant (p<0.001). The sensitivity, specificity, positive predictive value and negative predictive value were high for sepsis screen. The sensitivity of two or more abnormal parameters was more than any other single parameter used for screening (Table/Fig 3). The commonest organisms were Klebsiella pneumonia and Staphylococcus aureus. Gram positive organisms were found more commonly than gram negative (Table/Fig 4). | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| Discussion |
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Definitive diagnosis rests on a positive blood culture, to identify the pathogen and determine its antibiotic susceptibility pattern, but for better survival and outcome, simple and rapid diagnostics tests are required as adjuncts to the blood culture for early and effective initiation of treatment to the septicemia neonates. The ratio of culture positive neonatal septicemia cases were higher among males than the females in the present study, showing a ratio of 1.46 : 1. The male preponderance in neonatal septicemia may be linked to the X- linked immune-regulatory gene factor resulting in the host’s susceptibility to infections in males (4) [Table/ Fig-5]. Maximum culture positive cases were seen in neonates of age =72 hours as compared to neonates aged more than 72 hours. The higher proportion of early onset sepsis cases may be due to the immature immunological responses of the neonates in the first week of life, making them more susceptible to infections in this period [Table/ Fig-5]. In present study, the percentage of culture positive cases in low birth weight neonates was considerably higher than in normal birth weight neonates. According to Barbara Stoll et al., (14) 1991, the rate of infection is inversely proportional to the birth weight, and low IgG levels due to impaired cellular immunity in the very low birth weight neonates contributes to the increased susceptibility to infections in these neonates (14) [Table/ Fig-6]. The sepsis was more common in preterm neonates than in term babies. Preterm babies are more susceptible to infections due to inherent deficiencies of both humoral and cellular defense mechanisms. According to Barbara J. Stoll et al., (14) the incidence of septicemia increased with the decreased gestational age of the neonates (14) [Table/ Fig-6]. In present study, the percentage of culture positive cases in neonates born inside our institute (40.68%) was lesser than the neonates born outside (50.9%). The predominant organism found in blood culture was Klebsiella pneumoniae, followed by Staphylococcus aureus. In this study, gram positive organisms (62.6%) were more commonly found in blood culture as compared to gram negative organisms (37.4%) [4-6,10,11]. Cut off value of absolute neutrophil count=1800/µl was taken as diagnostic criterion for sepsis screen. Absolute neutrophil count in the sepsis screen showed low sensitivity (42.9%) and high specificity (99.0%). The positive predictive value was 97.5% and negative predictive value was 65.6% Absolute neutrophil count showed highest specificity and positive predictive value among all the other parameters of sepsis screen [Table/ Fig-7]. C-reactive protein =1mg/dl was considered as positive result for sepsis screen. In present study, CRP had a high sensitivity (84.62%), specificity (78.0%), positive predictive value (77.78%) and negative predictive value (84.78%) CRP proved to be the most sensitive of all the markers of sepsis. The highest negative predictive value was seen with CRP (Table/Fig 7). In present study, immature to total neutrophils ratio=0.2 was diagnostic criterion for sepsis screen which had good sensitivity (75.82%), specificity (66.35%), positive predictive value (65.2%) and negative predictive value (74.71%) (Table/Fig 8). Micro ESR=15 mm at the end of 1 hour was considered as positive for sepsis screen. The results in the present study showed specificity (87.0%) more than sensitivity (69.23%) and positive predictive value (82.89%) more than negative predictive value (75.65%). Micro ESR was a good predictor of neonatal sepsis but had lower sensitivity but higher specificity than C-reactive protein (Table/Fig 8). Serum direct bilirubin level =2 mg/dl was taken as positive diagnostic criterion for sepsis screen. The sensitivity was 57.14%, specificity was 93.0%, negative predictive value was 88.14% and negative predictive value was 79.45%.Therefore serum direct bilirubin proved to be a reliable marker of neonatal sepsis. Though the sensitivity was not high but specificity, positive predictive value and negative predictive value as well were high. Corrected total leukocyte count =5000/µl was taken as positive for sepsis screen. The specificity was 93.0% whereas positive predictive value and negative predictive value were 83.33% and 62.42% respectively. Leucopenia had low sensitivity, but high specificity (Table/Fig 9). Two or more abnormal parameters had a high accuracy in predicting neonatal sepsis. The results in the present study were in accordance with Gerdes et al., (23), and Jadhav et al., (7). The sensitivity of two or more abnormal parameters was 93.4%, specificity was 77.00%, positive predictive value was 78.7% and negative predictive value was 92.77% (Table/Fig 10). The sepsis screen should be considered as a positive septic screen, If two parameters are abnormal and antibiotic therapy can be started. If there is strong clinical suspicion and sepsis screen is negative, in 12 hours the screen can be repeated. If the screen is negative even after that, then sepsis may not be present. b#bLimitationsb?b The results obtained from sepsis screen cannot establish or rule out neonatal sepsis completely. The final diagnosis is obtained by culture and sensitivity only. The sepsis screen cannot replace blood culture. False positive cases may receive unnecessary antibiotic therapy. Obtaining blood sample in neonates is difficult. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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