Unit A2 6: Microbiology — Applications of Microorganisms
Welcome to your study notes for Applications of Microorganisms! Microorganisms such as bacteria, fungi, and yeasts might be invisible to the naked eye, but they power enormous global industries. In this unit, we will explore how microbes are harnessed in medicine, food production, agriculture, and environmental clean-up, along with the essential practical and safety standards required for your A2 6 portfolio.
Don't worry if industrial microbiology feels overwhelming at first. We will break down every process step-by-step so you can master both the theoretical principles and the practical coursework skills.
---1. Industrial and Pharmaceutical Applications
A. Antibiotic Production & Fermenter Control
One of the most famous applications of biotechnology is the large-scale production of antibiotics, such as penicillin, produced by the filamentous fungus Penicillium chrysogenum (or Penicillium notatum).
Understanding Microbial Growth Kinetics:
• Primary Metabolites: Compounds essential for growth and reproduction (such as amino acids, proteins, or ethanol). These are produced during the trophophase (the exponential / log growth phase).
• Secondary Metabolites: Compounds not directly required for basic growth and survival, often synthesized as defense mechanisms. Antibiotics are secondary metabolites, produced during the idiophase (the stationary phase), when population growth slows down due to nutrient limitation.
Key Fermenter Environmental Controls:
Industrial fermenters are giant, highly controlled vessels designed to maximize microbial yield. They require strict regulation of several factors:
• Dissolved Oxygen: High levels of sterile air are continuously bubbled into the vessel because Penicillium undergoes aerobic respiration.
• Temperature & Cooling Jackets: Microorganisms release metabolic heat as they grow. Water-cooled jackets surrounding the vessel prevent temperatures from rising to levels that would denature microbial enzymes.
• pH Control: Automated probes monitor pH, adding sterile acids or alkalis to maintain optimal enzyme activity.
• Agitation (Paddles / Impellers): Keeps the microorganisms in continuous suspension, ensures even temperature and nutrient distribution, and improves oxygen dissolution.
• Sterile Nutrient Feeds: Nutrients such as glucose, lactose, and corn steep liquor are added in controlled amounts.
B. Recombinant Microorganisms & Biopharmaceuticals
Through genetic engineering, scientists insert human genes into microorganisms to turn them into living factories for therapeutic proteins. Common host cells include the bacterium Escherichia coli and the yeast Saccharomyces cerevisiae.
• Recombinant Human Insulin: Used to manage diabetes mellitus, replacing older animal-derived insulins.
• Human Growth Hormone (hGH): Used to treat growth disorders in children.
• Hepatitis B Surface Antigens: Produced in recombinant yeast to manufacture safe, effective vaccines without using live viral particles.
C. Enzyme Technology
Microbial enzymes are extracted, purified, and often immobilized (attached to inert support matrices) so they can be reused continuously without contaminating the final product:
• Lactase: Extracted from the yeast Kluyveromyces lactis; used to hydrolyse lactose into glucose and galactose for lactose-free milk.
• Proteases and Lipases: Extracted from Bacillus species; used in biological laundry detergents to break down protein and fat stains at lower wash temperatures.
• Glucose Isomerase: Converts glucose into sweeter fructose for high-fructose corn syrup production in the food and beverage industry.
Quick Review — Section 1 Takeaway: Primary metabolites are made during the active growth phase (trophophase); secondary metabolites like penicillin are synthesized during nutrient stress in the stationary phase (idiophase). Industrial fermenters must provide sterile aeration, continuous agitation, and water cooling to protect enzymes.
---2. Food and Agricultural Microbiology
A. Food Fermentations
Fermentation uses microbial anaerobic or microaerophilic pathways to produce desirable flavors, textures, and natural preservatives:
• Dairy Products (Yoghurt & Cheese): Lactic acid bacteria such as Lactobacillus bulgaricus and Streptococcus thermophilus ferment the sugar lactose into lactic acid. The resulting drop in pH causes the milk protein (casein) to denature and coagulate (curdle), thickening the product and inhibiting spoilage bacteria.
• Brewing and Baking: The yeast Saccharomyces cerevisiae carries out anaerobic fermentation of sugars to produce ethanol and carbon dioxide gas:
\(\text{C}_6\text{H}_{12}\text{O}_6 \rightarrow 2\text{C}_2\text{H}_5\text{OH} + 2\text{CO}_2\)
In baking, the released \(\text{CO}_2\) gas bubbles cause dough to rise, while the ethanol evaporates during baking. In brewing, the ethanol is collected as the active beverage component.
B. Single-Cell Protein (SCP)
Single-Cell Protein refers to edible protein biomass derived directly from microbial cultures.
• Mycoprotein (Quorn): Produced using the filamentous fungus Fusarium venenatum.
• Production Method: Cultured in a continuous fermenter using a carbohydrate feed (like glucose or starch) enriched with ammonia and minerals.
• Advantages: High growth rate, low land and water footprint compared to livestock, rich in dietary fiber, and low in saturated fats.
C. Agricultural Microorganisms
• Biofertilizers: Symbiotic bacteria such as Rhizobium live in root nodules of leguminous plants (e.g., peas, clover). They fix inert atmospheric nitrogen (\(\text{N}_2\)) into ammonium ions (\(\text{NH}_4^+\)), providing natural nitrogen nutrition and reducing the need for synthetic chemical fertilizers.
• Biopesticides: The soil bacterium Bacillus thuringiensis (Bt) produces crystalline protein endotoxins. When ingested by pest insect larvae, the alkaline conditions of the insect gut activate the toxin, causing gut perforation and pest death without harming non-target organisms.
Quick Review — Section 2 Takeaway: Lactic acid bacteria coagulate casein via acid production; S. cerevisiae ferments sugars into ethanol and \(\text{CO}_2\); Fusarium venenatum provides fungal SCP (mycoprotein); and Rhizobium fixes atmospheric \(\text{N}_2\) into \(\text{NH}_4^+\) in legume root nodules.
---3. Environmental & Waste Treatment Applications
A. Bioremediation
Bioremediation is the use of specialized microorganisms (bacteria and fungi) to remove, neutralize, or degrade environmental pollutants and toxic xenobiotics (man-made chemical compounds not naturally produced).
• Mechanism: Microbes metabolize toxic petroleum hydrocarbons and xenobiotics into harmless end products: carbon dioxide (\(\text{CO}_2\)), water (\(\text{H}_2\text{O}\)), and non-toxic biomass.
• Application: Cleaning up maritime crude oil spills, petrol leaks from storage tanks, and industrial soil contamination.
B. Wastewater and Sewage Treatment
Sewage treatment uses a combination of microbial processes to convert hazardous waste into clean effluent before releasing it into waterways:
• Aerobic Digestion (Activated Sludge / Trickling Filters): Aerobic bacteria decompose dissolved organic matter in the presence of oxygen. This drastically reduces the Biochemical Oxygen Demand (BOD) of the wastewater, preventing oxygen depletion in natural rivers.
• Anaerobic Digestion: In sealed anaerobic digester tanks, methanogenic archaea and bacteria break down concentrated solid sewage sludge, producing biogas (primarily methane, \(\text{CH}_4\)), which can be burned as a renewable fuel source for heat and electricity.
Quick Review — Section 3 Takeaway: Aerobic sewage treatment lowers Biochemical Oxygen Demand (BOD) to protect aquatic life, while anaerobic digestion by methanogens converts solid waste into renewable methane biogas.
---4. Core Practical & Quality Assurance Standards (A2 6 Portfolio)
For your Unit A2 6 portfolio, examiners expect rigorous scientific accuracy regarding aseptic technique, safety protocols, and cell quantification methods.
A. Aseptic Technique Standards
• Flaming: Inoculating loops and necks of glass culture bottles must be passed through a blue Bunsen flame before and after transfer.
• Sterile Airfield: Work within the upward convection current created by a lit Bunsen burner or inside a sterile laminar flow cabinet.
• Autoclaving Parameters: Growth media, glassware, and hazardous waste must be sterilized in an autoclave under specific physical conditions:
\(121^\circ\text{C}\) for 15 minutes at \(15\text{ psi}\) (\(103\text{ kPa}\) of pressure).
Why? Boiling water (\(100^\circ\text{C}\)) cannot destroy bacterial endospores; pressurized steam at \(121^\circ\text{C}\) ensures complete sterilization.
B. Safe Incubation Protocols
• Petri Dish Sealing: Agar plates must be taped with two or four small pieces of adhesive tape (cross-taped), never sealed completely airtight around the perimeter. Airtight sealing promotes anaerobic conditions, which favor the growth of dangerous human pathogenic bacteria.
• Incubation Temperature: In school and college laboratories, cultures must be incubated below \(25^\circ\text{C}\) (typically \(20\text{–}25^\circ\text{C}\)). Human pathogens are adapted to body temperature (\(37^\circ\text{C}\)); keeping the incubator below \(25^\circ\text{C}\) minimizes the risk of multiplying pathogenic strains.
C. Quantification Methods: Total vs. Viable Cell Counts
Students must be able to clearly distinguish between total counts and viable counts:
1. Total Cell Counts (Measures both living and dead cells):
• Haemocytometer: A specialized calibrated microscope grid used to directly count cells in a known volume under a microscope.
• Colorimetry / Spectrophotometry: Measures the optical density (turbidity/cloudiness) of a liquid broth culture. Higher cell biomass absorbs/scatters more light.
2. Viable Cell Counts (Measures living, reproducing cells only):
• Serial Dilution & Spread/Pour Plating: A concentrated microbial culture is diluted by factors of 10 in a series of sterile tubes. Measured volumes from each dilution are spread onto agar plates.
• Colony-Forming Units (CFUs): Each single living cell multiplies into a visible colony.
• Statistical Reliability Rule: When calculating original cell concentrations, select only plates displaying between 30 and 300 colonies. Plates with over 300 colonies suffer from overcrowding/overlapping (uncountable), while plates with fewer than 30 colonies introduce excessive statistical error.
Common Portfolio Pitfalls & Examiner Warnings
• Confusing Growth Phases: Do not claim that penicillin is harvested during exponential growth. Primary metabolites (e.g., ethanol) are made in the trophophase; secondary metabolites (e.g., penicillin) are made in the idiophase / stationary phase.
• Incorrect Incubation Explanations: Always explain why plates are incubated below \(25^\circ\text{C}\) (prevents the growth of human pathogens adapted to \(37^\circ\text{C}\)) and why they are not sealed airtight (prevents growth of harmful obligate anaerobes).
• Omitting Autoclave Conditions: State the exact values: \(121^\circ\text{C}\), 15 minutes, \(15\text{ psi}\) (\(103\text{ kPa}\)).
• Total vs. Viable Counts: Remember that optical density (colorimeter) measures total biomass (dead + alive), whereas spread plating (CFU count) measures viable cells only.
• Fermenter Oxygenation: Antibiotic production using Penicillium is an aerobic process requiring sparging with sterile air, unlike anaerobic ethanol fermentation by yeast.
Memory Aid for Aseptic Autoclaving: Remember "121 – 15 – 15" (\(121^\circ\text{C}\), 15 minutes, \(15\text{ psi}\)) to secure full marks on autoclave specification questions!