Sensitive chemoluminescence-based immunological quantification of bacteria in soil extracts with monoclonal antibodies

Abstract

Azospirillum is a soil bacterium with plant growth promoting potential. We use strain specific monoclonal antibodies (mabs) against Azospirillum brasilense Sp7 to monitor this bacterium in the rhizosphere. Different kinds of indirect immunoassays (IA) were compared, to find the most sensitive way of detection and counting microorganisms in soil suspensions without using antibiotic resistant markers or culture techniques. With the standard enzyme-linked immunosorbent assay (ELISA)-teehnique it was possible to detect bacteria down to a density of 104ml −1 soil extract. The IA with the fluorescence marker fluorescein was not suitable for the quantification of bacteria in soil, because the background fluorescence of soil components interfered. With phycoerythrin the background fluorescence was reduced, but the overall sensitivity was not better than 10 3 bacteria ml −1' soil extract. An easy to perform, fast and sensitive method was the IA based on chemolumineseence (CIA). With this method it was possible to quantify A. brasilense Sp7 specifically down to a density of 100 bacteria ml −1 soil extract. In contrast to the ELISA method the CIA allowed to detect and quantify A. brasilense Sp7 in small root tip segments of the wheat rhizosphere.