Pseudomonas aeruginosa is a common Gram-negative bacterium frequently encountered in clinical and environmental settings. As an opportunistic pathogen, its identification is important in health care and water quality testing. Initial identification relies heavily on culturing the organism on solid media, which reveals characteristic visual and olfactory clues. Analyzing the growth pattern, colony structure, and metabolic byproducts provides the first step in differentiating this species from other microbes, guiding microbiologists toward a presumptive diagnosis.
General Colony Appearance
When P. aeruginosa is grown on non-selective laboratory media, such as Blood Agar or Nutrient Agar, its colonies exhibit a distinct macroscopic morphology. After 24 to 48 hours of incubation at 37°C, the colonies are typically large, opaque, and flat. Colonies can be smooth, but they often develop a rough or corrugated texture. Strains isolated from respiratory infections may appear mucoid due to the production of a polysaccharide capsule.
The edges of the colonies are frequently irregular, and some strains display localized swarming motility across the agar surface. A highly recognizable feature, particularly on Blood Agar, is the appearance of a greenish or silver-grey metallic sheen that reflects light. This iridescent quality is sometimes attributed to the production of an oily metabolite on the colony surface. Furthermore, when grown on Blood Agar, nearly all strains exhibit beta-hemolysis, appearing as a distinct, clear zone surrounding the colony where red blood cells have been lysed.
Distinctive Pigmentation and Aroma
A defining characteristic of P. aeruginosa on agar is its ability to produce diffusible, water-soluble pigments that color the surrounding medium. The most common is Pyocyanin, a blue compound synthesized by up to 95% of strains. Another prominent pigment is Pyoverdine, which is yellow-green and fluorescent under ultraviolet light. When Pyocyanin and Pyoverdine are produced together, they combine to create the striking bright green color characteristic of the organism.
Pyoverdine functions as a siderophore, a molecule used by the bacteria to scavenge and transport iron ions into the cell. The production of these pigments is often linked to the organism’s virulence; some strains may also produce the less common pigments Pyorubrin (red) or Pyomelanin (brown). Beyond the visual cues, a culture frequently emits a distinctive, pleasant odor. This characteristic smell is often described as “grape-like” or “fruity,” caused by the metabolic production of the volatile compound 2-aminoacetophenone.
Identification on Selective and Differential Media
While standard agar provides initial clues, laboratory protocols utilize selective and differential media to isolate and confirm P. aeruginosa. The most widely used selective medium is Cetrimide Agar, which contains the quaternary ammonium compound cetrimide. Cetrimide inhibits the growth of most competing bacterial species, allowing P. aeruginosa to grow predominantly. The medium also enhances the production of the fluorescent Pyoverdine pigment.
Observing the plate under short-wave ultraviolet light confirms the presence of fluorescent Pyoverdine with a bright green glow. Another common identification medium is MacConkey Agar, which tests for lactose fermentation. Since P. aeruginosa is a non-lactose fermenter, it produces colorless or clear colonies, distinguishing it from the red or pink colonies of lactose-fermenting bacteria. The growth at higher temperatures, such as 42°C, is often tested in conjunction with selective media as an additional confirmation step.