A study was undertaken to investigate developing biofilms in a petrochemical wastewater treatment plant encompassing the architecture, microflora and the chemical nature of the matrix. Biofilms were developed on glass slides immersed in the activated sludge unit and analysed at known time intervals using a range of techniques. Initially, biofilms were investigated using conventional and emerging microscopic approaches to select a suitable technique. Scanning Confocal Laser Microscopy (SCLM) allowed visualisation of biofilms in situ with minimal background interference and non-destructive and optical sectioning which were amenable to quantitative computer-enhanced microscopy. SCLM was superior over Light microscopy and Scanning Electron Microscopy. This study demonstrated biofilm growth, presence of extracellular polymer substances (EPS) in early biofilms associated with cells and the development of porous nature of mature biofilms including channel-like structures. Overall new information has been obtained on developing biofilms in an Australian petrochemical wastewater treatment plant / Doctor of Philosophy (PhD) (Biological Sciences)