Biochemical Tests for Identifying Citrobacter freundii

Identifying bacterial species is essential in clinical diagnostics, environmental studies, and food safety. Citrobacter freundii, a Gram-negative bacterium, can be an opportunistic pathogen responsible for various infections. Accurate identification of this organism ensures appropriate treatment and management strategies.

Biochemical tests are fundamental tools used to differentiate C. freundii from other closely related bacteria. These tests focus on specific metabolic activities unique to the organism, allowing researchers and clinicians to identify C. freundii with precision.

Enzyme Activity Assays

Enzyme activity assays provide insights into the organism’s metabolic capabilities. These assays detect specific enzymes characteristic of C. freundii. One such enzyme is β-galactosidase, detected using the ONPG test, which results in a yellow color change. The ability to produce β-galactosidase distinguishes C. freundii from other Enterobacteriaceae.

Lysine decarboxylase is another enzyme of interest, assessed through the lysine iron agar test. This assay evaluates the bacterium’s ability to decarboxylate lysine, leading to an alkaline reaction. The resulting color change in the medium aids in differentiating C. freundii from other bacteria lacking this enzymatic function.

Carbohydrate Fermentation

The ability to ferment carbohydrates aids in identifying Citrobacter freundii. This metabolic feature involves the breakdown of sugars, resulting in acidic byproducts. For C. freundii, glucose, lactose, and mannitol fermentation are particularly informative. The organism’s ability to ferment glucose with gas production is a distinguishing trait, often observed in a Durham tube setup.

Lactose fermentation is another important aspect. Unlike some other Enterobacteriaceae, C. freundii can typically ferment lactose, leading to acid production and a color change in lactose-containing media. This feature is frequently evaluated using MacConkey agar, where lactose fermentation results in pink colonies.

Mannitol fermentation, though less emphasized, can also provide additional clues. C. freundii’s capacity to ferment mannitol supports its identification, especially when combined with other biochemical tests.

Amino Acid Decarboxylation

Amino acid decarboxylation provides valuable insights into the biochemical identity of Citrobacter freundii. This process involves the removal of a carboxyl group from amino acids, resulting in the formation of amines and carbon dioxide. The metabolic versatility of C. freundii is highlighted by its ability to decarboxylate multiple amino acids, including ornithine and arginine.

The decarboxylation of ornithine leads to the production of putrescine, contributing to an alkaline shift in the culture medium. This reaction can be observed using ornithine decarboxylase broth, where a color change signifies the presence of the enzyme. Similarly, the decarboxylation of arginine results in the formation of agmatine, further demonstrating the organism’s enzymatic capabilities.

Hydrogen Sulfide Production

Hydrogen sulfide (H₂S) production is a biochemical trait that plays a role in identifying Citrobacter freundii. This process occurs when the organism reduces sulfur-containing compounds to produce H₂S gas. The presence of H₂S is typically detected using media that contain iron salts, such as triple sugar iron (TSI) agar. When C. freundii produces H₂S, it reacts with the iron salts to form iron sulfide, a black precipitate.

The ability to produce H₂S is common among several members of the Enterobacteriaceae family. However, when combined with other biochemical tests, it becomes a valuable piece of the puzzle in distinguishing C. freundii from other closely related bacteria.

Urease Activity

Urease activity is an important biochemical feature that can help in the identification of Citrobacter freundii. This enzyme catalyzes the hydrolysis of urea into ammonia and carbon dioxide, leading to an increase in the pH of the medium. The resultant alkaline environment can be detected using a pH indicator present in the urease test medium, typically phenol red.

While urease activity is common among several bacteria, its presence in combination with other biochemical markers can provide a more comprehensive profile of C. freundii. The urease test is often conducted alongside other tests, providing a holistic view of the organism’s metabolic capabilities.

Citrate Utilization

Citrate utilization is another metabolic feature that aids in the identification of Citrobacter freundii. This process involves the organism’s ability to use citrate as its sole carbon source, assessed using Simmons’ citrate agar. The medium contains bromothymol blue, a pH indicator that shifts from green to blue as the medium becomes alkaline due to citrate metabolism.

The ability to metabolize citrate provides insights into the metabolic flexibility of C. freundii, reflecting its adaptability to various environmental niches. This trait can be particularly informative when differentiating C. freundii from other Enterobacteriaceae that may not possess this capability. In conjunction with other biochemical tests, citrate utilization helps create a detailed metabolic profile of the bacterium.