Microorganisms can be identified by analyzing their biochemical reactions, such as carbon utilization and other metabolic tests. In smaller or educational laboratories, these assays are performed using a limited number of test tubes to determine the genus and species of a bacterial isolate based on its metabolic capabilities.

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Because many bacterial species appear morphologically identical, biochemical tests are crucial for identifying chemicals unique to specific species. The biochemical characteristics of a bacterium provide essential traits for its identification and classification. Specific advanced methods for this analysis include MALDI-TOF analysis, Fatty acid methyl ester (FAME) analysis, Phospholipid-derived fatty acids (PLFA) analysis, and proteomic analysis.

Biochemical Identification of Microorganisms

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Microbiology covers the scope and sequence requirements for a single-semester microbiology course for non-majors. The book presents the core concepts of microbiology with a focus on applications for careers in allied health. The pedagogical features of the text make the material interesting and accessible while maintaining the career-application focus and scientific rigor inherent in the subject matter. Microbiology’s art program enhances students’ understanding of concepts through clear and effective illustrations, diagrams, and photographs.

Microbiology is produced through a collaborative publishing agreement between OpenStax and the American Society for Microbiology Press. The book aligns with the curriculum guidelines of the American Society for Microbiology.

OpenStax Microbiology Textbook

Some microorganisms store specific chemical compounds as intracellular granules in their cytoplasm. Identifying the presence, absence, or content of these storage granules can serve as a diagnostic phenotypic biochemical characteristic for classifying and identifying microorganisms.

Microbial Identification Using Storage Granules

Microbial Identification via Metabolic and Physiological Assays

The Biolog system is a modern microbial identification method that simultaneously performs a panel of biochemical reactions analyzed by software. It identifies cells manually or using semi- or fully automated instruments by testing their ability to metabolize all major classes of biochemicals and assessing physiological properties like pH and chemical sensitivity.

Biolog Identification System

MALDI-TOF mass spectrometry is an automated method used for microbial identification by measuring a specimen's unique mass spectrum. During this process, a microorganism is mixed with a specialized matrix reagent on a disposable plate and irradiated with a high-intensity pulsed ultraviolet laser. This laser irradiation ejects gaseous ions from the microbe's chemical constituents, which are accelerated through a mass spectrometer at velocities determined by their mass-to-charge ratio ($$m/z$$). Reaching the detector at different times, these gaseous ions generate a mass spectrum plot of detector signal versus $$m/z$$ that is uniquely related to the organism's biochemical composition. Finally, comparing this mass spectrum to a library of reference spectra from known microorganisms allows for the identification of the unknown microbe.

MALDI-TOF Mass Spectrometry for Microbial Identification

Fatty acid methyl ester (FAME) analysis is a biochemical technique used to identify microbes by analyzing their membrane lipid composition. In this analysis, fatty acids are extracted from the cell membranes of microorganisms and chemically altered to produce volatile methyl esters. These methyl esters are then analyzed using gas chromatography (GC) to generate a chromatogram representing the microbe's specific fatty acid profile. By comparing the resulting chromatogram with a database of reference chromatograms from known bacterial isolates, the unknown microorganism can be identified.

Fatty Acid Methyl Ester (FAME) Analysis

Proteomic analysis is a microbial identification method based on analyzing the proteins produced by a pathogen under specific growth conditions within the human body. In this process, proteins from the pathogen are separated by high-pressure liquid chromatography (HPLC), and the collected fractions are then digested into smaller peptide fragments. These peptides are identified by mass spectrometry and compared with database profiles of known microorganisms to identify the pathogen.

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