A Study in Neonatal Sepsis Using Polymerase Chain Reaction-Based DNA Sequencing to Detect the Aetiological Agent in the Peripheral Blood of the Neonates with Sepsis

Blood culture, the gold standard for diagnosis of neonatal sepsis has low sensitivity and is time consuming. Therefore, objective is development and application of Polymerase chain reaction (PCR)-based DNA sequencing to detect and identify bacterium causing neonatal sepsis in culture negative specimens. Sixty three peripheral blood and 8 cerebrospinal fluid (CSF) referred from tertiary level Neonatal care Unit of the multi-specialty hospital from January 2012 to August 2013 were included. Specific Gram positive and Gram negative PCR followed by 16SrRNA region DNA sequencing was carried out. Based on DNA homology with Genbank sequences bacterial species were identified. Clinical details of these neonates including risk factors for early and late onset sepsis, reports of septic screen, blood culture and short term outcome till discharge were collected by retrospective chart review. PCR based DNA sequencing revealed 56 bacterial genomes comprising of 55 single bacterial species and 1 mixed bacterial species. Among them, 47 (84%) were Gram negative, 5 (8.9%) were Gram positive and 4 (7.1%) were uncultured bacterium. Escherichia coli (n =16; 27.5%) was the predominant bacterium identified. This study revealed newer emerging bacterial pathogensEscherichia fergusonii, Cronobacter dublinensis, Enterobacter hormachei, Janibacter species, Kluyvera georgiana, Streptococcus pseudopneumoniae as etiological agents of neonatal sepsis. To conclude, PCR based DNA sequencing is a rapid and specific molecular biological test to identify the causative agent of neonatal sepsis. Our study revealed novel bacterial pathogens as aetiological agents of neonatal sepsis. K e y w o r d s