Introduction to Microbial and Ecological Practicals

Welcome! In this chapter, we explore how biologists study life at two very different scales: the microscopic world of bacteria and the broad landscapes of ecosystems. These Core Practicals are essential because they teach you the "gold standard" of lab safety (aseptic technique) and how to gather reliable data from the unpredictable natural world. Whether you are aiming for a career in medicine or conservation, these skills are the foundation of biological research and are frequently examined in Paper 3.

Part 1: Microbiology (Core Practicals 12 & 13)

Microbiology is the study of organisms too small to see with the naked eye. Because bacteria are everywhere, we must use aseptic techniques to ensure we only grow the specific microbes we want to study and to keep ourselves safe.

Aseptic Technique: The "Golden Rules"

Aseptic technique refers to a set of procedures used to prevent contamination. When you are culturing microbes, you must:

  • Flame the equipment: Pass metal inoculating loops through a Bunsen burner flame until they glow red to kill any stray bacteria.
  • Bottle neck flaming: Quickly pass the neck of a glass bottle through the flame before and after opening to create upward air currents that prevent dust from falling in.
  • Lid lifting: Only open the lid of a Petri dish slightly (at an angle) to prevent microbes from the air landing on the agar.
  • Disinfect surfaces: Clean your workspace with 70% ethanol or disinfectant before and after the practical.

Culturing Bacteria

Bacteria are grown in culture media, which provide the nutrients (like glucose and nitrogen) they need.
1. Broth: A liquid medium used for growing large numbers of bacteria quickly.
2. Agar: A solid jelly-like medium.
3. Selective Media: Special agar that contains specific nutrients or antibiotics to allow only certain species to grow while inhibiting others.

Core Practical 12: Measuring Bacterial Growth

We can measure how fast a population of bacteria grows in a liquid broth using two main methods:

  1. Turbidity (Optical Methods): As bacteria multiply, the broth becomes "cloudy" (turbid). We use a colorimeter to measure how much light is absorbed or scattered. Higher turbidity = more bacteria.
  2. Dilution Plating (Cell Counts): If you try to count bacteria in a standard culture, there are too many to see individually. We perform a serial dilution (e.g., taking \(1\text{ ml}\) of culture and adding it to \(9\text{ ml}\) of sterile water, repeating this several times). We then plate the dilutions. We count the colonies on the plate that has between 30 and 300. Since each colony grew from one original cell, we can calculate the original population size.
    Formula: \( \text{Total viable count} = \text{number of colonies} \times \text{dilution factor} \)

The Bacterial Growth Curve

When bacteria grow in a closed system (like a flask), they follow a predictable pattern:

  • Lag Phase: Bacteria are adapting to their new environment; they are synthesising enzymes, not yet dividing quickly.
  • Log (Exponential) Phase: Rapid cell division. The population doubles at a constant rate.
  • Stationary Phase: The rate of cell death equals the rate of cell division. Nutrients are running out, and waste products (like toxins) are building up.
  • Death Phase: Cell death exceeds cell division due to a lack of food and toxic environment.

Math Alert: You may be asked to calculate the exponential growth rate constant (\(k\)). This involves using the change in the logarithm of the cell number over time: \( k = \frac{\log_{10} N_t - \log_{10} N_0}{0.301 \times t} \), where \(N_t\) is the final number and \(N_0\) is the initial number.

Core Practical 13: Streak Plating

This technique is used to isolate a single species from a mixed culture. You "streak" the bacteria across an agar plate in a specific pattern. Each time you move to a new section, you flame the loop. This dilutes the bacteria so much that, by the final sector, individual cells are deposited far apart, growing into pure colonies.

Quick Review: Aseptic technique prevents contamination. Dilution plating measures viable (living) cells, while turbidity measures total (living and dead) cells.

Part 2: Ecological Sampling (Core Practicals 15 & 16)

Ecology is about understanding where organisms live and why. Since we can’t count every single organism in an ecosystem, we use sampling.

Sampling Methods

  • Random Sampling: Used when the area is fairly uniform. You use a random number generator to create coordinates for your quadrats. This avoids bias (like picking the "prettiest" spot).
  • Systematic Sampling: Used when there is a change in the environment (a gradient), such as moving from a sunny field into a dark forest. We use a transect (a line or tape measure) and sample at regular intervals.

Tools for Sampling

  1. Frame Quadrats: A square frame (usually \(0.25\text{ m}^2\)). Used to count individual species or estimate percentage cover.
  2. Point Quadrats: A frame with pins. Each time a pin touches a plant, it is recorded. This is very objective for calculating percentage cover.
  3. Transects:
    - Line Transect: Record what touches the line.
    - Belt Transect: Place quadrats at intervals along the line to get more detailed data.

Core Practical 15: Estimating Population Size

When counting organisms, we can use different measures depending on the species:

  • Individual Counts: Counting every single plant (easy for large, distinct plants like dandelions).
  • Percentage Cover: Estimating what percentage of the quadrat area is covered by a species (best for spreading plants like moss or grass).
  • ACFOR Scale: A qualitative scale (Abundant, Common, Frequent, Occasional, Rare). It is quick but subjective (one person's "Common" might be another's "Frequent").

Core Practical 16: Abiotic Factors and Distribution

An abiotic factor is a non-living part of the environment, such as light intensity, soil pH, or temperature. This practical looks at how these factors change the distribution (where they are) or morphology (the physical shape/size) of a species.

Example: Measuring the leaf length of nettles in the shade versus nettles in full sunlight. You would use a transect and measure the abiotic factor (light) at the same spot you sample the plant.

Statistical Analysis in Ecology

Once you have your data, you need to prove it is significant:

  • Student’s t-test: Use this when you want to compare the means of two different groups (e.g., the average height of plants in Area A vs. Area B).
  • Spearman’s Rank Correlation Coefficient: Use this to see if there is a correlation between two variables (e.g., Does light intensity increase as plant height increases?).

Key Takeaway: Choose random sampling to avoid bias, but use systematic sampling along a transect if you are investigating an environmental gradient.

Common Mistakes to Avoid

  • Confusing Accuracy and Precision: In microbiology, counting 100 colonies is more accurate than counting 2 (too small for a sample) or 1000 (too many to count accurately, overlapping).
  • Forgetting Units: Always include units in your math, whether it's \( \text{cells ml}^{-1} \) for bacteria or \( \text{lux} \) for light intensity.
  • Aseptic Safety: Never seal a Petri dish completely with tape. This creates anaerobic conditions, which can encourage the growth of dangerous pathogenic bacteria. Use two small pieces of tape to allow oxygen in.

Final Quick Summary

Microbiology: Focuses on aseptic technique, growth phases (lag, log, stationary, death), and measuring growth via turbidity or dilution plating.
Ecology: Focuses on choosing the right sampling method (random vs. systematic), using quadrats/transects, and applying stats (t-test for means, Spearman’s Rank for correlation) to understand the impact of abiotic factors.