Type-IV pili spectroscopic markers: applications in the quantification of piliation levels in Moraxella bovis cells by a FT-IR ANN-based model

Abstract

Type-IV pili are cell surface organelles found in a wide variety of Gram-negative bacteria. They have traditionally been detected by electron microscopy and ELISA techniques. However, these methodologies are not appropriate for the rapid discrimination and quantification of piliated and nonpiliated cells in industrial or field conditions. Here, the analysis of FT-IR spectra of piliated, nonpiliated and sheared Moraxella bovis cells, together with purified pili suspensions spectra, allowed the identification of 3 IR regions associated to spectroscopic markers of Type-IV pili: 1750-1600, 1450-1350 and 1280-950 cm(-1). Such IR-specific markers were found for piliated cells grown in different culture systems (liquid or solid media), independently of the strain or pili serotype. They were also sensitive to pili expression levels. Therefore, on the bases of these specific spectral features, an FT-IR ANN-based model was developed to classify piliation levels in 5 distinct groups. An overall classification rate of almost 90% demonstrates the strong potential of the ANN system developed to monitor M. bovis cultures in vaccine production.