Biofilms of sessile <i>Pseudomonas syringae</i> cells formed on top of plant host’s leaves or fruits allow surviving harsh environmental conditions (desiccation) and improve their resistance to antibacterial treatments of crops. Better understanding of these biofilms can help minimize their effect on harvests. In the present study, infrared attenuated total reflection (IR-ATR) spectroscopy with optical and confocal laser scanning microscopy have been applied to analyze Pseudomonas syringae pathovar morsprunorum biofilm development in real-time, for the first time. The biofilm development was observed within the spectral window 4000-800 cm^-1 at constant flow conditions during 72 h. The kinetics of representative integrated band areas (nucleic acids with polysaccharides at 1141-1006 cm^-1, amino acid side chains with free fatty acids at 1420-1380 cm^-1, proteins at 1580-1490 cm^-1, and lipids with proteins at 2935-2915 cm^-1) were analyzed with regards to the observed biofilm structure and following P. syringae biofilm developmental stages were attributed: The inoculation phase, washing of weakly attached bacteria closely followed by recolonization of vacated surface, the restructuration phase, and finally the maturation phase.