Neonatal sepsis refers to an infection involving the bloodstream in new-born infants less than 28 days old. It remains a leading cause of morbidity and mortality among neonates, especially in middle and lower-income countries [1] Sepsis is an ongoing public health problem worldwide and a major cause of morbidity and mortality. Late Onset Sepsis is usually hospital or community acquired such as contact from healthcare workers or caregivers. Septicaemia was the most common clinical category with incidence of 23/1000 live births while the incidence of meningitis was reported to be 3/1000 live births [2]. Late onset septicaemia (LOS) usually presents after 72 h of age [3,4]. Any new-born with bacterial sepsis is also at risk of meningitis. However, the incidence of meningitis in neonatal sepsis has varied from 0.3% to 3% in various studies, but Late Onset Sepsis has been reported to be fairly associated with meningitis; with percentage ranging from 3% to 30%.[3,4,5]. Due to the nonspecific neonatal presentation for sepsis and the high risk of mortality and morbidity without treatment, many asymptomatic neonates undergo a sepsis workup if risk factors are present and/or clinically indicated. Although approximately 7% to 13% of all neonates are worked up for sepsis, only 3% to 8% have positive cultures [2]. The most frequently encountered pathogens in the late-onset group, the organisms were coagulase-negative staphylococci and Enterobacteriaceae, including E. coli, Klebsiella pneumoniae, and Acinetobacter baumannii but infection with Pseudomonas aeruginosa was associated with the highest morality rate (55.0%) [6]. In cases of Late Onset Sepsis, Lumbar Puncture should be done in all infants prior to starting antibiotics. Lumbar Puncture could be postponed in a critically sick neonate but should be performed once the clinical condition stabilizes [7]. Neonatal sepsis remains a significant contributor to morbidity and mortality in neonates. Prematurity and delayed treatment are commonly associated with adverse outcomes. VLBW infants have been found to have a higher risk of chronic lung disease, and extremely low birth weight (ELBW) infants are at a greater risk of neurodevelopmental risks, such as hearing and visual deficits, cerebral palsy, and impaired psychomotor and mental development [8]. On the other hand, the unnecessary overuse of antibiotics can increase the chances of severe candidiasis and multi-drug resistant organisms. There is marked variability in the prevalence of meningitis in neonatal sepsis among different studies done previously. So, the rationale behind present study was to timely detect the cases of meningitis among neonates having sepsis, so that prompt treatment could be started. This study also helped us to know the frequency of neonatal meningitis in late onset sepsis in developing country like ours and also show the importance of doing lumbar puncture in every patient admitted with neonatal sepsis. Furthermore microorganisms thus detected during study will be help in framing antimicrobial policy in neonatal intensive care unit (NICU) of this hospital.

Neonates with features of sepsis and positive CRP were subjected to Lumber puncture (included cerebrospinal fluid (CSF) cytology, biochemistry and culture), complete blood count, blood sugar, serum bilirubin, renal function test, blood culture and chest X Ray. Meningitis was labelled in neonate whose CSF findings satisfied criteria of WBC>10 /Glucose <25 /Protein > 170 among neonates with blood culture proven sepsis in preterm babies & WBC > 8/ Glucose < 20/ Protein > 120 for term babies. Other investigations such as liver function tests, renal function test, Prothrombin Time Index (PTI), arterial blood gas analysis, carried out as and when required. The neonates were followed-up, to determine their final outcome. Also, antibiotic resistance and sensitivity patterns were assessed to formulate hospital antimicrobial drug policy. The data were analysed with the help of computer software SPSS (authorized version) 23.0 for Windows. Chi-square test was used to ascertain statistical significance among proportions. Prevalence along with 95% confidence limits was calculated to express the magnitude disease under study. Odd's ratio with 95% confidence limits using logistic regression model was estimated vis a vis the presence or absence of meningitis in late onset neonatal sepsis. A P value of equal to or <0.05 was considered statistically significant.

A total of 1,565 neonates were admitted during study period with various clinical indications. Out of these 225 neonates presented with Late Onset Sepsis were eligible for study after fulfilling exclusion criteria. The Demographic profile of neonates with late onset sepsis is depicted in Table1. Out of total 225 neonates meningitis was reported in 26% (n =58) neonates on blood culture proven sepsis. 37% (n= 35) in preterm and 17% (n=17) in term babies. The proportion of meningitis in low birth weight reported to be 64.8% which was highly significant with p value of 0.00. Similarly, Prevalence reported more in males (47%) than females (21%) which was statistically highly significant. Also, significant association observed with mothers’ education & mothers’ occupation with P value <0.05. Furthermore, 13% subjects died of meningitis and around 87% with meningitis recovered from disease with p value 0.01. No significant association was observed with socio economic status, place of residence (Urban/ Rural), mothers’ parity, and gestational age of neonate with p value > 0.05 Table 2. The most common presenting feature among neonates in late onset sepsis with or without meningitis was respiratory distress (53%) followed by lethargy (43%), Refusal to feed (40%). Other presenting features observed among neonates were fever (17%), irritability (15%), convulsions (8%), apnea (6%), shock (2%), abdominal distention (2%), hypothermia (2%), vomiting (2%), loose stools (2%) respectively Table 3. None of the neonates with late onset sepsis had history of bleeding from any site, umbilical lesions and bulging fontanelle. Thrombocytopenia was present in 19%, skin mottling was present in 5% study subjects and X ray report was abnormal in 8% neonates. 4% films showed Hyaline membranous changes and respiratory distress syndrome in (4%) subjects. Blood culture was positive in 27% subjects with high positivity rate in preterm (60%) than term (40%) Table 4. On CSF cytological findings, meningitis reported to be around 34% more in term (37%) than preterm neonates (31%). The most common organism isolated in blood culture was Staphylococcus aureus (26%) followed by Aceinitobacter (15%), Klebshiella (13%), Enterobacter (12%). Cerebrospinal fluid was turbid in 3%, red in 6% and clear in 91% study subjects in present research. However, all CSF culture were sterile. 32% of samples showed antibiotic resistance. Maximum antibiotic resistance was observed against penicillin (17%) followed by erythromycin (6%), clindamycin (4%), gentamicin (2%), Vancomycin (2%). Duration of stay observed to be within 14 days in 72% study subjects & 28% had duration of stay in NICU more than 14 days. After being discharged from NICU 47% had hospital stay within 14 days & 53% had hospital stay more than 14 days. Mortality reported to be statistically significant between meningitis and no meningitis group. On independent logistic regression model to ascertain independent association of various risk factor with meningitis is shown in Table 5. Table 1: Demographic profile of neonates with Late Onset Sepsis Variable Number (%) Age at time of admission 72 hours 175 (78) 4-14 days 30 (13) 15-28 days 20 (9) Socioeconomic status Upper 18 (8) Upper Middle 57 (25.3) Middle 59 (26.2) Lower middle 55 (24.4) Lower 36 (16) Gender & Gestational Age Male 115 (51) Term 65 (56.5) Preterm 50 (43.5) Female 110 (49) Term 66 (60) Preterm 44 (40) Birth weight <2.5 kg 91 (40) >2.5 kg 134 (60) Table 2: Association of neonatal meningitis with various risk factors Variable Meningitis (CSF findings) Test of Significance Yes (n=77) No (n=148) P value Mothers’ occupation Employed 18 (47.3) 20 (52.6) .04 Home maker 59 (31.5) 128 (68.4) Residence Urban 18 (41.8) 25 (58.1) .160 Rural 59 (32.4) 123 (67.5) Parity ≥2 67 (36.2) 118 (63.7) .202 <2 10 (25) 30 (75) Gender Male 54 (46.9) 61(53.1) .00 Female 23 (20.9) 87 (79.09) Gestational Age Term 48 (36.6) 83 (63.3) .224 Preterm 29 (30.8) 65 (69.14) Birth weight Low 59 (56.7) 45 (43.2) .00 Normal 18 (14.87) 103 (85.12) Outcome of disease Recovery 63 (30.43) 144(69.56) .00 Death 14 (77.77) 4 (22.22) Blood culture Positive 6 (10.34) 52 (89.65) .00 Negative 71 (42.51) 96 (57.48) Table 3: Presenting features among neonates with late onset sepsis with or without meningitis Presenting Features N (%) Respiratory Distress 119 (53) Lethargy 97 (43) Refusal to feed 90 (40) Fever 38 (17) Irritability 34 (15) Convulsions 18 (8) Apnoea 14 (6) Shock 5 (2) Abdominal Distension 5 (2) Hypothermia 5 (2) Vomiting 5 (2) Loose Stools 5 (2) Bleeding from any site 0 (0) Table 4: Prevalence of Meningitis in Late Onset Sepsis as per blood culture Gestational Age Blood Culture Positive Blood Culture Negative Total Term 23 (17) 108 (83) 131 (100) Preterm 35 (37) 59 (63) 94 (100) Total 58 (25.7) 167 (74.3) 225 (100) Table 5: Independent association of risk factors in neonatal meningitis Variable aOR 95% CI P value Mother’s Occupation Home maker 1.68 [.779-3.570] .188 Employed Reference Gender Male .796 [.410-1.546] .501 Female Reference Birth weight Low .294 [.163-.528] .000 Normal Reference

During the study period, there were 225 study subjects. All were evaluated for sepsis with Lumbar puncture followed by blood culture. In the present study, overall prevalence of meningitis in late onset sepsis was reported to be 25.7% on blood culture where as on CSF cytology (lumbar puncture) meningitis was reported among 34% neonates. Blood culture found to be positive in 17% among term neonates & 37% among preterm neonates. 78% newborn presented at 72hours from birth, 13% presented with 4 to 14 days and rest presented more than 14 days to 28 days. 40% neonates were low birth weight. In contrast to the previous studies, this study reported that respiratory distress, lethargy, refusal to feed common clinical symptoms among neonates with late onset sepsis. Other presenting features observed among neonates with late onset sepsis is fever (17%), irritability (15%), convulsions (8%), apnea (6%), shock (2%), abdominal distention (2%), hypothermia (2%), vomiting (2%), loose stools (2%) respectively. Similar observation was reported by [18,19, 9]. The study revealed prevalence of neonatal meningitis in late onset sepsis was 25.7% which was almost similar to other Indian observational studies [11, 25]. However, Bhagat et al. [9] (16%) and Mehta [10] (17%) reported lower prevalence in Indian settings. Schwersenski et al.[23] and Hristeva et al.[24] reported the prevalence of meningitis in LOS as 3.5% and 6.8% respectively, but the former excluded neonates more than 72 h and less than 7 days from their study and the latter delayed the lumbar puncture procedure in all the babies who had respiratory distress. Similar observational studies from Kenya,[12] Brazil,[13] and Asia [14] reported the prevalence of meningitis in neonates with late onset sepsis were 17.9%, 17%, and 17.2%, respectively. The minor difference in the prevalence can be attributed to the various epidemiological and geographical factors in community-acquired infection in late-onset variety of sepsis. We cross tabulated the results i.e. frequency of meningitis with effect modifier i.e. age, gender, parity, birth weight, outcome of disease. Frequency of meningitis was higher i.e. 89.3% in infants between 1-15 days, as compared to older infants (>15 days) i.e. 10.7%. However, the results were not statistically significant (p= 0.235). Similarly, in a descriptive study by Khurshid A et al, Majority of the neonates with sepsis i.e. 52.1% were between 1-7 days of their life. 47% and 21% were males and females respectively and this difference was statistically significant. Our study showed male predominance similar to other observational studies. Kaul et al. reported male predominance in neonates who had meningitis [11]. However, similar observational studies like Jiang JH et al [15], Gheibi S et al [16] & Umate [17] et al reported no gender predominance. For pre-terms, the same data extrapolated wasn’t statistically significant. This difference in the pattern of observance can be explained on the basis of better available neonatal intensive care set up and early initiation of the drug therapy based on the available CSF cytology reports which takes only 3 hours from sample collection to reporting in contrast to blood culture report that takes 3 days. Around 40% neonates had low birth weight. Prevalence of meningitis in Late Onset Sepsis in LBW neonates was 64.8% in our study. On applying logistic regression model, birth weight alone was observed to be statistical significant. Being born with normal birth weight babies had protective (71%) factor against neonatal meningitis compared to babies born with low birth weight. Similarly, in a descriptive study by Khurshid A et al, 39.3% neonates had low birth weight (1.5-2.5 kg), however mean weight was 2.55 + 0.39 kg (range 1.8- 3.6 kg). This high incidence of meningitis in LBW and especially preterm has also been reported by Kaul et al, [11] Bhagat R et al [9] & Jiang J et al [15] observed that late onset sepsis was significantly more common in low birth weight and preterm neonates. However, in present study no statistical interaction was observed between birth weight & gestational age. The mean WBC count of the infants with late onset sepsis was 29.78±9.32 in our study. Majority of infants i.e. 60.05% had less than 75% neutrophils in our study. More than one -third of infants with late onset sepsis i.e. 38% had Positive C-reactive protein in our study. Blood culture was positive in 26% cases of late onset sepsis in our study. However, 7.8% of blood culture positive cases had meningitis and 92% of blood culture negative cases had meningitis. Bhagat R et al [20] reported 12.7% positive blood culture in septic neonates. However, Bhagat et al.[9], Mehta [10] and Umate et al [17] reported 42.6% ,49.6% and 53.8% positive blood culture in neonates with meningitis, respectively. The commonest microorganism isolated from culture was gram positive Staphylococcus aureus (26%) followed by Aceinitobacter (15%), Klebshiella (13%), Enterobacter (12%) & MRSA (7%). Moreno et al, [21] Karthikeyan et al., [19] and Sundaram et al [22] also reported S.aureus as the commonest gram positive isolate followed by MRSA (7%) & Staphylococcus epidermidis (7%). Of the gram negative isolates in our study the commonest were Aceinitobacter comprising 15%, Klebshiella (13%) ,Enterobacter 12.% , E coli (7%) and Pseudomonas (4%). The overall mortality in neonatal meningitis was 8% which was comparatively less as reported by other studies such as Mehta [10] (20.6%), Tiskumara et al. [14] (20%), Kaul et al.,[11] (26.1%) and Bhagat et al.[9] (17.6%). Out of the neonates having meningitis, 13% expired in the present study which is similar to the observations made by Tisukumara et al who observed that meningitis was associated with a mortality of 20% in his study. This low mortality in our study can be attributed to continuous efforts to improve intensive care facilities and prompt treatment based on CSF cytology report.

An attempt was made to establish concordance between blood and Cerebrospinal Fluid analysis including cytology & cultures which may affect management strategies for sepsis and meningitis. The present study showed importance of doing lumbar puncture in every patient admitted with neonatal sepsis so that prompt treatment could be started in minimal time based on CSF cytology reports. The study concluded that prevalence of neonatal meningitis in late onset meningitis was 26% on blood culture & 34% on CSF cytology. Of various risk factors, only birth weight was reported to be most significant risk factor associated with meningitis in late onset sepsis. Maximum antibiotic resistance was observed against penicillin followed by erythromycin, clindamycin, gentamicin, Vancomycin. Microorganisms thus detected during study helped in framing antimicrobial policy in neonatal intensive care unit (NICU) of this hospital. Recommendations Routine CSF analysis should be practiced in all neonates with neonatal sepsis regardless of their postnatal age and clinical features enabling prompt treatment, better prognosis & reduced mortality among neonates with meningitis in late onset sepsis.

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