Standard Operating Procedure (SOP) for Collecting Sewage for Bacteriophage Isolation
1. Purpose
1.1 To safely and effectively collect sewage samples for the isolation of
bacteriophages, ensuring minimal contamination and maximum recovery of viable
phages.
2. Scope
2.1 This SOP applies to laboratories involved in bacteriophage research or
environmental microbiology studies.
3. Required Materials
3.1 Personal Protective Equipment (PPE)
3.1.1 Lab coat
3.1.2 Nitrile or latex gloves
3.1.3 Safety goggles
3.1.4 Face mask or respirator (if working with aerosol-generating samples)
3.1.5 Waterproof boots or shoe covers (for fieldwork)
3.1.6 Disposable gown (optional for high-risk areas)
3.2 Instruments and Equipment
3.2.1 Sterile containers (50 mL and/or 500 mL, preferably polypropylene or
glass)
3.2.2 Autoclave-safe sample bottles with screw caps
3.2.3 Sampling pole (optional, for hard-to-reach areas)
3.2.4 Cooler box with ice packs (for sample transport)
3.2.5 Sterile syringes and filters (0.22 μm or 0.45 μm)
3.2.6 pH meter or pH strips
3.2.7 Thermometer
3.2.8 Forceps and sterile scoops (if needed)
3.3 Chemicals and Reagents
3.3.1 Phosphate-buffered saline (PBS) or sterile saline solution
3.3.2 Ethanol (70%) for surface sterilization
3.3.3 Sodium thiosulfate (optional, for neutralizing residual disinfectants)
3.3.4 Peptone water (optional, for enriching bacteriophages)
3.3.5 Nutrient broth or LB broth (for host bacteria cultivation)
3.3.6 Agar media (for plaque assays)
4. Procedure
4.1 Preparation
4.1.1 Assemble all required equipment and PPE.
4.1.2 Label sterile containers with relevant details (e.g., sample ID, date,
time, and location).
4.1.3 Calibrate pH meter and thermometer if needed.
4.2 Collection
4.2.1 Identify a suitable collection point (e.g., raw sewage, effluent
discharge, septic tanks).
4.2.2 Put on all required PPE before entering the sampling site.
4.2.3 Sterilize the sampling container or ensure it is pre-sterilized.
4.2.4 Collect the sample directly from the sewage source:
4.2.4.1 For liquid samples, submerge the container or use a sterile sampling
pole.
4.2.4.2 For solid or semi-solid samples, use a sterile scoop to transfer
material into the container.
4.2.5 Fill the container, leaving some headspace (10-20% of the container
volume) for aeration.
4.3 Post-Collection Handling
4.3.1 Secure the lid of the container to prevent leaks.
4.3.2 Place the sample in a cooler box with ice packs to maintain a temperature
of ~4°C.
4.3.3 Record environmental data such as temperature, pH, and any notable
observations.
4.4 Transport and Storage
4.4.1 Transport samples to the laboratory as soon as possible.
4.4.2 Store samples at 4°C if immediate processing is not possible (process
within 24 hours for best results).
4.5 Processing in the Laboratory
4.5.1 Use sterile syringes to filter the sample through a 0.22 μm or 0.45 μm
filter to remove bacteria and debris.
4.5.2 Store the filtered supernatant at 4°C until use in isolation experiments.
5. Safety Precautions
5.1 Avoid direct contact with sewage. Handle samples with care to minimize
risk of contamination and infection.
5.2 Dispose of all used PPE and waste materials as per biohazard regulations.
5.3 Decontaminate all equipment and surfaces with a suitable disinfectant after
use.
6. Documentation
6.1 Maintain detailed records of sample collection, including location,
date, time, environmental conditions, and any anomalies.
6.2 Record sample processing details in the laboratory logbook.
END OF THE DOCUMENT
You may like to read these links:
2. List of All SOPs and Documents for In-vitro Laboratory
3. List of All SOPs and Documents for the Animal House Facility
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