Microbiology Study Guide: Microbial Growth, Control, and Genetics

Microbial Growth and Measurement

Measuring Microbial Growth

Accurate measurement of microbial growth is essential in microbiology for monitoring cultures, diagnosing infections, and assessing the effectiveness of antimicrobial treatments. Several methods are commonly used:

• Direct Microscopic Count: Microbes are counted directly under a microscope using a counting chamber (e.g., hemocytometer). This method is rapid but cannot distinguish between live and dead cells.

• Viable Plate Count: A sample is diluted and spread on an agar plate. After incubation, colonies are counted, and the number of viable cells is calculated. This method only counts living cells capable of forming colonies.

• Turbidity Measurement: The cloudiness (turbidity) of a liquid culture is measured using a spectrophotometer. Increased turbidity indicates higher cell density, but this method does not differentiate live from dead cells.

• Other Methods: Additional techniques include membrane filtration, most probable number (MPN) method, and measuring metabolic activity (e.g., CO2 production).

Example: Turbid solutions in IV fluids may indicate microbial contamination, which is detected by increased turbidity.

Clinical Application: Turbidity in IV Solutions

IV solutions should be clear and free of visible particles. Turbidity (cloudiness) suggests contamination, often by microbes or particulates.

• Problem: Turbid IV solution may contain microbial contamination, posing a risk of infection if administered.

• Correct Action: The nurse's decision to not use the turbid solution prevents potential harm to the patient.

Clinical Application: Handling of Urine Specimens

Proper handling of clinical specimens is crucial for accurate diagnosis.

• Diagnosis of UTI: Urinary tract infections are diagnosed when urine contains ≥100,000 microbial cells/ml.

• Improper Handling: If a urine specimen is left at room temperature, bacteria may multiply, artificially increasing the count and potentially leading to a false diagnosis.

• Example: A specimen with 1,000 cells/ml left for 4 hours may exceed the diagnostic threshold due to bacterial growth during storage.

Control of Microbial Growth

Sterilization vs. Disinfection

Controlling microbial growth is essential in healthcare, laboratory, and food settings. Two key processes are:

• Sterilization: The complete destruction or removal of all forms of microbial life, including spores. Examples:

  • Autoclaving surgical instruments before use in surgery.

  • Flaming inoculation loops in microbiology labs.

• Disinfection: The elimination of most pathogenic microorganisms (except bacterial spores) on inanimate objects. Examples:

  • Using bleach to clean hospital floors.

  • Applying alcohol to disinfect thermometers.

Factors Affecting Disinfectant Choice and Use

Several factors must be considered when selecting and using disinfectants:

• Nature of the Microbe: Some microbes are more resistant than others (e.g., spores, mycobacteria).

• Concentration and Contact Time: Higher concentrations and longer exposure increase effectiveness.

• Presence of Organic Matter: Blood, pus, or feces can inhibit disinfectant action.

• Surface Type: Porous vs. non-porous surfaces may affect efficacy.

• Toxicity and Safety: Disinfectants should be safe for users and the environment.

Microbial Resistance to Disinfectants

Certain microbes are harder to kill due to their unique structures:

• Mycobacterium: Has a waxy, lipid-rich cell wall (mycolic acids) that resists penetration by disinfectants.

• Bacillus and Clostridium: Form endospores, which are highly resistant to heat, chemicals, and desiccation.

Definitions and Examples of Key Terms

Term	Definition	Example/Application
Pasteurization	Heat treatment that kills pathogens and reduces spoilage organisms without sterilizing.	Milk is pasteurized to kill Mycobacterium tuberculosis.
Disinfection	Elimination of most pathogenic microbes (not spores) on inanimate objects.	Bleach used on hospital surfaces.
Antisepsis	Destruction of pathogens on living tissue.	Alcohol swab before injection.
Sanitization	Reduction of microbial population to safe levels.	Washing dishes in a restaurant.
Sterilization	Complete destruction/removal of all microbes, including spores.	Autoclaving surgical tools.
Degerming	Removal of microbes from a limited area.	Handwashing before surgery.

Microbial Genetics and Biotechnology

Structure and Functions of DNA and RNA

Genetic material in cells is composed of DNA and RNA, each with distinct structures and functions.

• DNA (Deoxyribonucleic Acid): Double-stranded helix, stores genetic information, composed of nucleotides (adenine, thymine, cytosine, guanine).

• RNA (Ribonucleic Acid): Single-stranded, involved in protein synthesis, nucleotides include adenine, uracil, cytosine, guanine.

• Functions:

  • DNA: Long-term storage of genetic information.

  • RNA: Transmits genetic information (mRNA), brings amino acids (tRNA), forms ribosomes (rRNA).

Key Genetic Terms

• mRNA (Messenger RNA): Carries genetic code from DNA to ribosomes for protein synthesis.

• tRNA (Transfer RNA): Brings amino acids to the ribosome during translation.

• Gene: Segment of DNA that encodes a functional product (usually a protein).

• 5' to 3': Directionality of nucleic acid synthesis; nucleotides are added to the 3' end.

• Transcription: Process of copying DNA into RNA.

• Translation: Process of synthesizing proteins from mRNA template.

• Codon: Three-nucleotide sequence in mRNA that specifies an amino acid.

• Anticodon: Three-nucleotide sequence in tRNA complementary to mRNA codon.

Genes and Their Products

• Genes are made of: DNA molecules.

• Genes code for: Proteins (enzymes, structural proteins, etc.).

Non-Protein Molecule Synthesis in Microbes

Although genes encode proteins, microbes synthesize non-protein molecules (e.g., phospholipids) through enzyme-mediated pathways. Enzymes, coded by genes, catalyze the biosynthesis of these molecules.

• Example: Phospholipid synthesis involves enzymes that assemble fatty acids and glycerol into membrane lipids.

Transcription and Translation Locations

Process	Prokaryotes	Eukaryotes
Transcription	Cytoplasm	Nucleus
Translation	Cytoplasm (often simultaneous with transcription)	Cytoplasm (after mRNA export from nucleus)

Transcription and Translation Example

Consider the following DNA strand:

• DNA Template Strand: 3'-TAC GGA TGC-5'

• Transcription (mRNA): 5'-AUG CCU ACG-3'

• Translation (Amino Acids): Methionine (AUG), Proline (CCU), Threonine (ACG)

Diagram: (Textual description)

• DNA is transcribed to mRNA by RNA polymerase.

• mRNA codons are read by ribosomes; tRNA brings corresponding amino acids.

• Polypeptide chain is formed according to the mRNA sequence.

Formula:

$\text{DNA} \xrightarrow{\text{Transcription}} \text{mRNA} \xrightarrow{\text{Translation}} \text{Protein}$

Example: For the DNA sequence above, the resulting polypeptide is Met-Pro-Thr.

Additional info: Some explanations and examples have been expanded for clarity and completeness.