Bacterial Identification Methods PDF

Importance of Bacterial Identification: Correctly identifying bacteria is crucial in clinical labs to help doctors choose the right treatment for patients. For example, in food poisoning, accurate identification of the bacteria helps doctors provide the right care. I...

Importance of Bacterial Identification: Correctly identifying bacteria is crucial in clinical labs to help doctors choose the right treatment for patients. For example, in food poisoning, accurate identification of the bacteria helps doctors provide the right care. In disease outbreaks, identifying the pathogen helps contain the spread of the epidemic. Methods for Identifying Microorganisms: There are different methods to identify bacteria, some based on physical (phenotypic) traits, and others on genetic (genotypic) traits. Common methods include: ○ Analyzing bacteria’s nutrition and metabolism to figure out its genus and species. ○ Checking for stored compounds (e.g., poly-β-hydroxybutyrate or PHB) inside the bacteria, which can help identify the species. Examples of Bacteria: Some Pseudomonas species, like P. aeruginosa (a human pathogen) and P. syringae (a plant pathogen), can be identified by their fluorescent pigments and whether or not they store PHB. Biochemical Testing: Traditional methods use test tubes for metabolic tests like carbon utilization, but modern systems (e.g., Biolog) use software to run multiple tests at once. Biolog identifies bacteria based on their biochemical reactions and other traits like pH and chemical sensitivity. Automated Mass Spectrometry (MALDI-TOF): Another method uses MALDI-TOF mass spectrometry to identify bacteria by analyzing their mass spectrum. The bacterial sample is hit with a laser, generating ions, which are analyzed by the mass spectrometer to create a unique mass spectrum. The spectrum is compared to a database to identify the microorganism. Microbes can be identified by their unique lipid profiles. Fatty acids in lipids differ by: Chain length Number and presence of double bonds Hydroxyl groups, branches, and rings To identify a microbe, the fatty acids in its membranes are analyzed. A common method for this is called fatty acid methyl ester (FAME) analysis. FAME analysis is used in clinical, public health, and food labs. In this method: Fatty acids are extracted from the microbe’s membrane. They are chemically changed into methyl esters. These esters are analyzed by gas chromatography (GC). The results are compared to a database to identify the microbe. PLFA analysis (Phospholipid-derived fatty acids): Membranes are made of phospholipids. Phospholipids can be broken down (saponified) to release fatty acids. These fatty acids are analyzed (FAME analysis) and compared with known profiles to identify microorganisms. Proteomic analysis (Protein-based identification): Bacteria can be identified based on the proteins they produce in specific conditions. The proteins are separated using high-pressure liquid chromatography (HPLC). These proteins are broken down into smaller pieces (peptides), identified by mass spectrometry, and compared with known microorganisms. Carbohydrate-based identification (Glycoproteins): Microorganisms can be identified by carbohydrates attached to proteins in their membrane or cell wall. Antibodies or proteins attach to specific carbohydrates, causing cells to clump together (serological tests). Examples include Lancefield group tests used to identify Streptococcus species.