7.5 Using Biochemistry to Identify Microorganisms
3 min read•june 18, 2024
reveal microbes' unique metabolic fingerprints. By analyzing how bacteria process sugars, amino acids, and enzymes, scientists can identify specific species. These tests create distinct profiles, helping distinguish between different microorganisms in clinical and research settings.
Mass spectrometry and lipid analysis offer advanced identification methods. provides rapid, accurate bacterial identification by analyzing protein spectra. Lipid analysis techniques like and examine fatty acid compositions, offering insights into microbial communities and individual species.
Biochemical Identification of Microorganisms
Biochemical identification of microorganisms
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- Biochemical tests assess metabolic capabilities and enzymatic activities of microorganisms
- Tests include (, ), (, ), and (, )
- Results create a biochemical profile unique to each microbial species (E. coli, )
- These profiles reflect the specific of different microorganisms
- contain specific substrates to distinguish between different microorganisms
- allow growth of certain microbes while inhibiting others ( for Gram-negative bacteria)
- change color due to pH shifts or specific metabolic products ( for lactose fermentation)
- Automated identification systems, such as and , use miniaturized biochemical tests
- Reactions are compared to a database to identify the microorganism
- Systems provide rapid, standardized results for clinical (diagnosis of infections) and research settings (screening of environmental isolates)
- patterns can provide insights into and aid in identification
Mass spectrometry for bacterial identification
- (MALDI-TOF) mass spectrometry
- Bacterial proteins are ionized and separated based on their mass-to-charge ratio ()
- Generates a unique mass spectrum for each bacterial species (fingerprint)
- Spectra are compared to a reference database for identification (, )
- Advantages of MALDI-TOF over traditional methods
- Rapid identification (minutes vs. hours or days)
- High accuracy (>95%) and reproducibility
- Minimal sample preparation (direct colony analysis) and cost-effective
- Applications in clinical microbiology and research
- Identification of pathogens from clinical specimens (blood, urine, cerebrospinal fluid)
- Strain typing and epidemiological studies (outbreak investigations)
- Discovery of novel biomarkers for specific microorganisms (antibiotic resistance, virulence factors)
Lipid analysis methods in microbial identification
-
- Bacterial fatty acids are extracted, methylated, and analyzed by
- Generates a unique fatty acid profile for each microbial species ()
- Requires pure cultures and standardized growth conditions (temperature, medium)
- (PLFA) analysis
- Measures the composition of phospholipid fatty acids in microbial membranes
- Can be performed on environmental samples without cultivation (soil, water, sediments)
- Provides insights into microbial community structure and biomass (fungi, bacteria, actinomycetes)
- (LPS) analysis
- LPS is a major component of the outer membrane of Gram-negative bacteria
- Variations in LPS structure can be used for serotyping and strain differentiation (, )
- Methods include SDS-PAGE (silver staining), immunoassays (ELISA, )
- Comparison of lipid analysis methods
- FAME and PLFA provide species-level identification, while LPS is more suitable for strain typing
- PLFA is advantageous for environmental samples, while FAME requires pure cultures
- Lipid analysis complements other identification methods, such as DNA sequencing (
) and mass spectrometry (MALDI-TOF)[16S rRNA gene](https://www.fiveableKeyTerm:16S_rRNA_Gene)
Enzymatic and Metabolic Profiling
- are key to understanding microbial metabolism and identification
- Specific enzyme assays can be used to differentiate between closely related species
- Metabolic profiling helps in by revealing unique biochemical characteristics
- Analysis of metabolic end-products can provide insights into microbial physiology and ecology
Key Terms to Review (52)
16S rRNA Gene: The 16S rRNA gene is a highly conserved genetic marker used for the identification and classification of bacteria and archaea. It is a component of the 30S small subunit of the prokaryotic ribosome and is essential for protein synthesis.
Amino Acid Decarboxylation: Amino acid decarboxylation is a biochemical process in which an amino acid's carboxyl group (-COOH) is removed, resulting in the production of a corresponding amine and carbon dioxide. This reaction is an important mechanism used by microorganisms to identify and differentiate between various species.
API: API, or Application Programming Interface, is a set of protocols, tools, and routines that enable different software applications to communicate and interact with each other. It provides a standardized way for software components to exchange data and functionality, allowing developers to build upon existing systems and create more integrated and efficient software solutions.
Bacterial Taxonomy: Bacterial taxonomy is the science of classifying and naming bacteria based on their physical, genetic, and biochemical characteristics. It provides a structured system to organize and identify different bacterial species, which is crucial for understanding their diversity, evolution, and roles in various environments and processes.
Biochemical Identification: Biochemical identification is a laboratory technique used to identify microorganisms based on their unique metabolic and enzymatic profiles. It involves analyzing the specific biochemical reactions and characteristics exhibited by a microbial isolate to determine its identity and classify it within the taxonomic hierarchy.
Biochemical Tests: Biochemical tests are analytical procedures used in microbiology to identify and characterize microorganisms based on their metabolic and enzymatic properties. These tests provide valuable information about the unique biochemical profiles of different microbial species, allowing for their accurate identification and classification.
Bruker Biotyper: The Bruker Biotyper is a mass spectrometry-based system used for the rapid and accurate identification of microorganisms. It is a widely-used tool in the field of microbiology for the identification and classification of bacteria, yeasts, and fungi.
Carbohydrate Fermentation: Carbohydrate fermentation is a metabolic process in which microorganisms, such as bacteria and yeast, break down carbohydrates in the absence of oxygen to produce various end products, including alcohols and organic acids. This process is a crucial part of microbial identification and characterization.
Catalase: Catalase is an enzyme found in most living organisms that catalyzes the decomposition of hydrogen peroxide (H2O2) into water and oxygen. It is a crucial enzyme that helps protect cells from oxidative damage caused by reactive oxygen species.
Core Oligosaccharide: The core oligosaccharide is a critical component of the lipopolysaccharide (LPS) molecule found in the outer membrane of Gram-negative bacteria. It serves as a link between the lipid A and the O-antigen, providing structural integrity and playing a key role in the identification and classification of microorganisms.
Cystic fibrosis: Cystic fibrosis (CF) is a genetic disorder that affects the respiratory and digestive systems, caused by mutations in the CFTR gene. It leads to thick, sticky mucus production, which obstructs airways and promotes bacterial infections.
Differential Media: Differential media are types of microbiological growth media that are designed to distinguish between different types of microorganisms based on their unique metabolic or morphological characteristics. These media allow for the selective isolation and identification of specific bacteria or other microbes of interest.
Enzymatic Reactions: Enzymatic reactions are the chemical transformations that occur within living organisms, facilitated by specialized protein molecules called enzymes. These reactions are central to the process of metabolism, enabling the breakdown, synthesis, and modification of various biomolecules necessary for life.
Enzyme Production: Enzyme production is the biological process by which enzymes, which are complex proteins that act as catalysts to speed up chemical reactions, are synthesized within cells. This process is crucial for the proper functioning of various metabolic pathways and cellular activities in microorganisms.
FAME: FAME stands for Fatty Acid Methyl Esters, which are the products of a chemical process that converts fatty acids into methyl esters. This process is commonly used in the identification and characterization of microorganisms, as the FAME profile can serve as a unique biochemical fingerprint for different bacterial species.
FAME analysis: FAME analysis (Fatty Acid Methyl Ester analysis) is a technique used to identify microorganisms based on the unique fatty acid profiles of their cell membranes. The method involves converting fatty acids into methyl esters for analysis via gas chromatography.
Fatty Acid Methyl Ester: Fatty Acid Methyl Ester (FAME) is a chemical compound formed by the esterification of a fatty acid with methanol. It is commonly used as a biodiesel fuel and in the identification and characterization of microorganisms through biochemical analysis.
Fatty acid methyl ester (FAME) analysis: Fatty acid methyl ester (FAME) analysis is a method used to identify microorganisms by analyzing the unique patterns of fatty acids in their cellular membranes. This technique involves converting fatty acids into their methyl esters for easier gas chromatography.
Gas Chromatography: Gas chromatography is an analytical technique used to separate, identify, and quantify the individual components of a complex mixture. It is a powerful tool in the field of microbiology, enabling the identification and characterization of microbial metabolites and other biomolecules.
Gas chromatography (GC): Gas chromatography (GC) is a technique used to separate, identify, and quantify compounds in a mixture based on their volatility and interaction with the stationary phase. It is widely used in microbial biochemistry for analyzing metabolic products and identifying microorganisms.
Glucose: Glucose is a simple sugar (monosaccharide) that serves as the primary source of energy for the body's cells. It is a crucial component of carbohydrates and plays a central role in cellular metabolism and microbial identification.
Granules: Granules are small particles or droplets within cells that store substances such as nutrients, pigments, or secretory products. In microbiology, they can be found in both prokaryotic and eukaryotic cells.
High-pressure liquid chromatography (HPLC): High-pressure liquid chromatography (HPLC) is a technique in analytical chemistry used to separate, identify, and quantify each component in a mixture. It relies on pumps to pass a pressurized liquid solvent containing the sample mixture through a column filled with a solid adsorbent material.
Hot tub rash: Hot tub rash, also known as Pseudomonas dermatitis, is a skin infection caused by the bacterium Pseudomonas aeruginosa. It typically occurs after exposure to contaminated water in hot tubs or swimming pools.
Indicator Media: Indicator media are specialized culture media used in microbiology to identify and differentiate microorganisms based on their ability to produce certain metabolic byproducts or enzymes. These media contain indicators that change color or appearance in response to specific microbial activities, allowing for the rapid detection and characterization of different bacterial species.
Lactose: Lactose is a disaccharide sugar composed of glucose and galactose, found primarily in the milk of mammals. It is an important carbohydrate that plays a crucial role in the identification and classification of microorganisms.
Lancefield groups: Lancefield groups are a classification system for streptococci based on the carbohydrate composition of their cell wall antigens. Developed by Rebecca Lancefield, these groups help in identifying and differentiating pathogenic streptococci.
Lipopolysaccharide: Lipopolysaccharide (LPS) is a complex molecule found in the outer membrane of Gram-negative bacteria. It is a key component of the bacterial cell wall and plays a crucial role in various aspects of microbiology, immunology, and pathogenesis.
Lysine: Lysine is an essential amino acid that plays a crucial role in protein synthesis and various metabolic processes. As a building block of proteins, lysine is essential for maintaining the structural integrity of cells and tissues, and it is also involved in the production of hormones, enzymes, and other important biomolecules.
MacConkey Agar: MacConkey agar is a selective and differential medium used to isolate and differentiate Gram-negative, lactose-fermenting bacteria, particularly members of the Enterobacteriaceae family, from clinical and environmental samples. It is an important tool in the identification and characterization of microorganisms.
MALDI-TOF: MALDI-TOF (Matrix-Assisted Laser Desorption/Ionization-Time of Flight) is a mass spectrometry technique used for identifying microorganisms by analyzing their protein composition. It provides rapid and accurate microbial identification by comparing the protein spectra to a reference database.
Matrix-Assisted Laser Desorption/Ionization Time-of-Flight: Matrix-Assisted Laser Desorption/Ionization Time-of-Flight (MALDI-TOF) is an analytical technique used to identify and characterize biomolecules, particularly proteins and peptides, by ionizing and detecting them based on their mass-to-charge ratio. It is a powerful tool employed in the context of using biochemistry to identify microorganisms.
Metabolic Pathways: Metabolic pathways are a series of interconnected chemical reactions that occur within cells to sustain life. These pathways are responsible for the breakdown and synthesis of molecules, providing the energy and building blocks necessary for cellular processes and organismal function. Metabolic pathways are central to the topics of 7.5 Using Biochemistry to Identify Microorganisms and 8.4 Fermentation, as they underlie the biochemical mechanisms that allow microorganisms to be characterized and utilize different energy sources.
Microbial Physiology: Microbial physiology is the study of the biological processes and functions that enable microorganisms, such as bacteria, archaea, and fungi, to survive, grow, and reproduce. It encompasses the mechanisms by which microbes obtain and utilize energy, nutrients, and other resources from their environment to carry out essential life activities.
O-antigen: The O-antigen is a polysaccharide component of the outer membrane of Gram-negative bacteria that serves as an important structural and functional feature, particularly in the context of lipids and microbial identification.
Ornithine: Ornithine is a non-proteinogenic amino acid that plays a crucial role in the urea cycle, a metabolic pathway responsible for the removal of excess nitrogen from the body. It is an important intermediate in this cycle, facilitating the conversion of ammonia into urea, which can then be excreted.
Oxidase: Oxidase is an enzyme that catalyzes the transfer of electrons from a substrate to molecular oxygen, producing water as a byproduct. This enzyme is particularly important in the context of prokaryote habitats, relationships, and microbiomes, as well as in the biochemical identification of microorganisms.
Phenol Red: Phenol red is a pH indicator dye commonly used in microbiology to detect changes in the acidity or basicity of a solution. It is particularly useful in identifying microorganisms based on their ability to ferment or metabolize various carbohydrates, which can be observed through color changes in the medium containing phenol red.
Phospholipid-Derived Fatty Acid: Phospholipid-derived fatty acids are a class of lipid molecules that are essential components of cell membranes. They are formed by the breakdown or metabolism of phospholipids, which are the primary structural lipids in biological membranes. These fatty acids play a crucial role in the identification and characterization of microorganisms.
Phospholipid-derived fatty acids (PLFA) analysis: Phospholipid-derived fatty acids (PLFA) analysis is a technique used to profile microbial communities based on the unique fatty acid compositions of their cell membranes. It helps in identifying and quantifying different microorganisms present in an environmental sample.
PLFA: PLFA, or Phospholipid Fatty Acid analysis, is a technique used in microbiology to identify and characterize the microbial community structure within a given environmental sample. It relies on the unique phospholipid fatty acid profiles of different microorganisms to provide insights into the composition and diversity of the microbial population.
Poly-β-hydroxybutyrate (PHB): Poly-β-hydroxybutyrate (PHB) is a type of biopolymer produced by certain bacteria as a form of energy storage. It is a biodegradable plastic with properties similar to petrochemical-derived plastics.
Proteomic analysis: Proteomic analysis is the large-scale study of proteins, particularly their structures and functions. It is used to identify and quantify proteins in a given sample to understand biological processes at the molecular level.
Pseudomonas: Pseudomonas is a genus of Gram-negative, rod-shaped bacteria known for their metabolic diversity and ability to thrive in various environments. They are often opportunistic pathogens, particularly Pseudomonas aeruginosa, which can cause infections in humans.
Pseudomonas aeruginosa: Pseudomonas aeruginosa is a Gram-negative, opportunistic pathogen that can cause serious infections in humans. It is known for its resistance to antibiotics and ability to thrive in various environments.
S. aureus: S. aureus, or Staphylococcus aureus, is a Gram-positive bacterium known for its ability to cause a wide range of infections, from mild skin conditions to life-threatening diseases. This versatile pathogen is a significant concern in both healthcare and community settings due to its virulence factors and potential for antibiotic resistance.
S. aureus is a key topic in the context of Gram-positive bacteria, biochemical identification of microorganisms, and bacterial infections of the skin and eyes. Its unique characteristics and clinical significance make it an important subject of study for students of microbiology.
Selective Media: Selective media are specialized growth media designed to support the growth of specific microorganisms while inhibiting the growth of others. These media contain specific nutrients, inhibitors, or selective agents that allow only the target organisms to thrive, making them a crucial tool in the identification and isolation of microbes.
Sherlock Microbial Identification System: The Sherlock Microbial Identification System is a powerful tool used in microbiology to accurately identify and classify microorganisms based on their biochemical profiles. This system leverages advanced analytical techniques to provide a comprehensive analysis of a microbial sample, enabling researchers and clinicians to make informed decisions about microbial identification, diagnosis, and treatment.
Substrate Utilization: Substrate utilization refers to the ability of microorganisms to use various chemical compounds as energy sources or building blocks for cellular processes. It is a crucial aspect of microbial identification and classification, as different microbes exhibit distinct patterns of substrate utilization.
VITEK: VITEK is an automated system used in clinical microbiology laboratories to identify and perform antimicrobial susceptibility testing on bacterial isolates. It utilizes a series of miniaturized biochemical tests to rapidly and accurately determine the identity and antibiotic resistance profile of various microorganisms.
VITEK MS: VITEK MS is an automated system used in clinical microbiology laboratories to rapidly identify microorganisms based on their mass spectrometry profiles. It is a valuable tool for the identification of bacteria, yeasts, and other microbes, aiding in the diagnosis and treatment of infectious diseases.
Western Blot: The Western blot is an analytical technique used to detect and quantify specific proteins within a complex mixture of proteins extracted from cells or tissues. It is a powerful tool employed in various fields, including microbiology, immunology, and virology, to identify and characterize proteins of interest.