Microarray analysis of Acidovorax temperans

Abstract

Bacteria belonging to the genus Acidovorax have been shown to be a consistent member of the activated sludge microbial community. Two phenotypic variants of A. temperans CB2 isolated locally from activated sludge exhibit noteworthy characteristics, such as the ability to form biofilms and flocs, which are critical microbial processes underlying all modern wastewater treatment systems. Gene expression microarray technology is a functional genomics platform that enables the simultaneous interrogation of all expressed transcripts during normal cell ontogeny, or in response to specific environmental stimuli. Microarray technology offers the opportunity to investigate gene expression changes relevant to key processes in wastewater treatment, using A. temperans as a model organism. The aims of this research were to develop a full genome microarray platform for A. temperans CB2 and to use this microarray platform to investigate major differences in gene expression between the Hpos and Hneg phenotypic variants. An optimised gene expression microarray platform was established through the assessment of various experimental methods, such as RNA extraction, RNA amplification, microarray probe design, and quantitative PCR. Using the microarray platform, gene expression comparisons were obtained for planktonic broth cultures, static biofilms and bacterial colonies. Gene expression analyses have provided insights into the complex developmental processes involved in the transition from planktonic cells to stages of initial attachment, cell proliferation, biofilm maturation and nutrient limitation during the formation of A. temperans biofilms. Factors that have been identified in other bacterial systems such as type IV pili and activation of stress responses were also observed in A. temperans biofilms. In addition, several intriguing classes of genes, such as transcriptional regulators, a toxinantitoxin gene cassette, and nitrate metabolism were also found to be differentially expressed during the formation of A. temperans biofilm. The incorporation of microarray technology with other functional genomics techniques to investigate molecular mechanisms underlying the complex processes occurring in wastewater treatment will provide a scientific basis to improve the reliability of current wastewater treatment strategies and for the development of new treatment technologies.