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Bacteria that are Gram-negative are not stained dark blue or violet by Gram staining, in contrast to Gram-positive bacteria.

The difference lies in the cell wall of the two types; Gram-positive bacteria have a high amount of peptidoglycan in their cell wall which the stain interacts with, while Gram-negative bacteria have a cell wall made primarily of lipopolysaccharide. The Gram-negative cell wall is similar to a cytoplasmic membrane, typically only a few layers thick and generally much thinner than Gram-positive types.

Many species of Gram-negative bacteria are pathogenic. This pathogenic capability is usually associated with certain components of their cell walls, particularly the lipopolysaccharide (endotoxin) layer.

The proteobacteria are a major group of Gram-negative bacteria, including for instance Escherichia coli, Salmonella, and other Enterobacteriaceae, Pseudomonas, Moraxella, Helicobacter, Stenotrophomonas, Bdellovibrio, acetic acid bacteria, and a great many others. Other notable groups of Gram-negative bacteria include the cyanobacteria, spirochaetes, green sulfur and green non-sulfur bacteria.