Development of a Chip based Detection System for Bacterial Sepsis

Sepsis is a serious medical condition caused by an overwhelming response of a host upon a pathogen invasion. Till date, blood culture based identification of causative pathogen is considered as the gold standard method. But this process has long sample to result times and proven to be less sensitive for slow growing and fastidious organisms. To address these, a molecular assay involving a novel asymmetric Polymerase Chain Reaction (PCR) based amplification method called Asymmetric Exponential and Linear Amplification (AELA) is developed in this work. The AELA-PCR is coupled with a Hierarchical-Microarray for differentiation and identification of bacterial species.

The merit of AELA-PCR lies in its efficiency of generating single stranded amplicons which are know to have higher binding efficiency to microarray spots. The hierarchical microarray is fabricated on unmodified polystyrene substrate using a photo-crosslinking copolymer. This is also designed on the principle of taxonomical bacterial identification where a respective bacteria can be detected and differentiated at different taxonomical levels. The assay with AELA-PCR and Hierarchical-Microarray showed an LOD of 10cfu/ml for 40 sepsis causing pathogens tested representing 22 bacterial genera. The initial clinical validation studies of the assay showed 20% higher specificity and sensitivity levels compared to the gold standard blood culture based identification. This makes it a promising candidate for Sepsis diagnosis.