BIOHAZARD SAFETY & WASTE MANAGEMENT IN THE LABORATORY — Key Terms

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• Objectives of today: a) Identify biological hazards in biomedical, pharmaceutical, material, biotechnology, and environmental labs. b) Understand risk groups and factors for biosafety levels. c) Recognize required infrastructures for safe handling at each biosafety level (BSL). d) Describe SOPs and GDP for handling, storage, and disposal of samples and waste. e) Apply SOPs and GDP to safely identify, handle, store, and dispose of biological samples and waste.

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• Key concepts: Biological hazards (biohazards), Biological Agent Risk Groups, and classification of Biosafety Levels (BSLs). These guide containment and safety practices in labs.

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• Biohazards are biological substances that threaten health, animals, or the environment.
• Sources: microorganisms (bacteria, viruses, fungi, parasites), biological toxins, human/plant/animal tissues.
• Hazards can cause infections, allergies, or toxicity.
• Transmission routes include direct contact, air, water, and contaminated surfaces; exposures require specific safety measures.

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• Types of biohazardous samples:
  • Diagnostic and research samples: Blood, Urine, Tissue, Stool, etc.; cell lines, viruses, protein extracts.
  • Investigations occur in: Research laboratories (pharmaceutical/biomedical/biological sciences) and Diagnostic/Medical laboratories, among others.

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• Sample collection and testing contexts:
  • Clean-environment sectors test for microbial contamination.
  • Swab samples (surfaces) and food samples are common.
  • Investigations occur in various agencies (environmental, biomedical, biological sciences).

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• Additional sample categories:
  • Ecological monitoring includes samples from Plants, Marine Life, Fungi, Algae, etc.

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• Security breach vs safety breach vs emergencies:
  • Security breach: deliberate loss/theft/misuse; unauthorized access; potential release.
  • Safety breach: accidental loss of containment; exposure; lab-acquired infections/allergies.
  • Emergencies: medical events, fires, natural disasters, power/ HVAC failures.
• Purpose of biosafety and biosecurity: protect people, environment, and materials from misuse or accidental release.

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• Case highlight: Accidental contamination of SARS in Singapore illustrates cross-contamination and the need for proper BSL-3 containment features.
• Key takeaway: Inappropriate lab practices can lead to infections; BSL-3 requires heightened safety standards and engineering controls.

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• Principles of biosafety:
  • Objective: Containment to reduce/eliminate exposure to potentially harmful biological agents.
  • Step 1: Determine the biological agent's Risk Group.
  • Step 2: Determine the suitable containment to handle the sample.

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• Factors determining risk group (WHO guidelines):
  • Availability of preventive measures (vaccines) and effective treatment (antibiotics).
  • Capability to infect and cause disease.
  • Virulence (severity) and communicability (transmissibility).

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• Risk Group classifications (summary):
  • RG1: Individual risk Nil/Low; Community risk Nil/Low; Pathogenicity: unlikely to cause disease.
  • RG2: Individual risk Moderate; Community risk Low; serious infections possible but treatable; spread limited.
  • RG3: Individual risk High; Community risk Low; serious disease; not ordinarily transmitted; treatments exist.
  • RG4: Individual risk High; Community risk High; serious disease; highly transmissible; no vaccines or treatments.

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• National/regional risk grouping considerations:
  • Pathogenicity, mode of transmission/host range, and local preventive/treatment options.
  • Influenced by local immunity, population density/mobility, vectors, and environmental hygiene.
  • Prophylaxis, antisera, antimicrobials, antivirals may inform risk designation.
• Note: Risk group guidance is a starting point; it must be validated by a local risk assessment.

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• Biosafety Levels: representation of containment requirements from low to high risk.
  • BSL-1: Basic level; minimal hazard; standard practices; no special barriers; sink for handwashing; open bench allowed.
  • BSL-2: Moderate hazards; work on open bench with precautions; BSC for aerosols; restricted access; waste decontamination; PPE.
  • BSL-3: Indigenous or exotic agents with respiratory transmission; enhanced containment; directional airflow; BSC; separated area.
  • BSL-4: Dangerous/exotic agents with high risk and no vaccines; full containment; Class III BSC or full-body suits with supplied air; separate building.

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• Important distinction: Biological Agent Risk Group ≠ Biosafety Level.
• Higher volume/concentration of a RG-3 agent may require BSL-4 containment.
• Conduct a formal risk assessment to determine the appropriate biosafety level.

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• Activity hint: Map biosafety considerations to different applied-science scopes (Biomedical, Pharmaceutical, Biotechnology, Environmental/M marine science) and suggest suitable BSL levels.

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• Risk assessment (RA) basics:
  • RA is done by those most familiar with the work.
  • If information is insufficient, default to precautionary measures: Basic containment (BSL-2) + standard precautions.
  • Key RA review factors: organisms used, equipment, procedures, containment and facilities, and any animal models used.
  • RA is not a one-off; it should be routinely reviewed.

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• Factors to include in microbial risk assessment (RA):
  • Pathogenicity and infectious dose;
  • Natural route of infection;
  • Alternative infection routes due to lab manipulations;
  • Stability of the agent in the environment;
  • Presence of a suitable host;
  • Genetic manipulations that may extend host range;
  • Local prophylaxis/treatment availability;
  • Data from animal studies/clinical reports;
  • Agent concentration and volume to be manipulated.

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• Break time.

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• Basic containment overview: Biosafety Levels (BSL-1 to BSL-4) and core concepts of containment, safety practices, equipment, and facility design.

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• BSL-1: Suitable for well-characterized agents not known to cause disease in healthy humans; basic practices; standard PPE; open bench allowed; no special barriers; hazard symbol posted when infectious agents present.

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• BSL-1 facilities and practices:
  • PPE used for potential splashes; handwashing; doors for access control; lab designed for easy cleaning; impervious bench/chairs; sinks; screens on exterior windows.
  • Biosafety Cabinets (BSCs) generally not required at BSL-1.

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• BSL-2 overview:
  • Builds on BSL-1; handles moderate hazards; work with pathogenic agents; access restricted; BSC used for procedures with aerosols; general lab practices extended with biosafety protocol.

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• BSL-2 special practices and PPE:
  • Biohazard warning signs on doors;
  • All procedures with potential aerosols inside a BSC;
  • Routine decontamination of equipment and spills;
  • Use durable leak-proof containers for handling/in storage;
  • Immediate incident reporting; medical surveillance; immunization where applicable;
  • Only trained personnel may work with BSL-2 agents.

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• BSL-2 PPE specifics:
  • Eye protection when handling hazardous materials outside BSC;
  • Lab coats, gloves (change when contaminated); no reuse; strict handwashing;
  • Additional eye/face protection when needed; protective equipment for working with animals.

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• BSL-2 lab setup example:
  • HEPA-filtered air from Class II BSC; air recirculation per manufacturer guidance;
  • Showers/eye-wash stations mandatory;
  • BSC placement to minimize airflow disruption;
  • Self-closing locked doors; rules on waste decontamination (autoclave or chemical disinfection).

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• Quick BSL snapshots:
  • BSL-1: GMT (Good Microbiological Techniques); safe practice; non-pathogenic examples (e.g., E. coli K-12, Bacillus subtilis);
  • BSL-2: GMT plus protective clothing; biohazard signs; examples include S. aureus, Hepatitis B virus, Salmonella, Toxoplasma, HIV, human-derived materials.

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• Comparison: BSL-1 vs BSL-2 lab practices and barriers:
  • BSL-1: Standard practices; minimal barriers; open bench; basic PPE as needed.
  • BSL-2: Limited access; decontamination of waste/clothing; use of PPE; primary barriers include BSCs for procedures with aerosols; autoclave capability.

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• BSL-3 and BSL-4 scope:
  • BSL-3: Indigenous or exotic microbes with respiratory transmission; higher containment; enhanced PPE and practices; negative airflow; separate building often required for large scales.
  • BSL-4: Exotic/dangerous microbes with high aerosol risk; no vaccines/treatments; two lab models exist (cabinet lab with Class III BSC or suit lab with full-body suits); strict entry/exit and decontamination.

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• BSL-3/BSL-4 design and entry specifics:
  • Doors: biosafety level-specific entry controls; medical surveillance and immunizations may be required; manipulatio ns inside BSCs; no open bench work; serum sampling programs for at-risk personnel.

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• PPE for BSL-4 and higher:
  • Class III BSCs or positive-pressure suits; regular changes of clothing; head/foot protection; eye/face protection; PPE required at all times for BSL-4; sometimes powered air-purifying respirators (PAPR).

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• BSL-4 facility design highlights:
  • Isolation by airlock; negative pressure; HEPA-filtered, non-recirculated exhaust; dedicated systems; waste decontamination; double-door autoclaves; pass-through decontamination.
  • Facilities may be in a separate building or isolated zone.

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• BSL-4 lab models and practices:
  • Cabinet Lab (Class III BSC) or Suit Lab (full-body suit);
  • Two-person rule; complete clothing change before/after entry; emergency procedures and communication plans.
  • Adhere to BSL-3 code of practice with added containment for BSL-4.

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• Reference materials and examples of worldwide BSL facilities; (links and visuals cited in slide content).

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• Secondary barriers and facility design (BSL-4 focus):
  • Primary containment and secondary barriers; airlocks; autoclave at entry; containment drains; negative pressure; dedicated power and waste decontamination lines.
  • Specimens, materials, and animals enter via airlocks; double-door pass-through autoclave; all effluents decontaminated before discharge.

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• BSL-3 vs BSL-4 recap (practices, barriers, facilities):
  • BSL-3: Limited access, negative airflow, animal/agent handling inside containment; primary barriers include BSCs and PPE; separate building often required for higher containment.
  • BSL-4: Maximum containment; explicit isolation features; full-body suits or Class III BSC; separate building; dedicated systems; highest safety standards.

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• Summary comparison (continued):
  • Practices rise with level: BSL-1 (GMT) → BSL-2 (restricted access, waste decon) → BSL-3 (entry control, shower on exit, decontaminated exit) → BSL-4 (airlock, suit/ Class III containment).
  • Primary barriers adapt accordingly (PPE/BSCs) and secondary barriers (facility design) scale with risk.

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• Summary table (conceptual):
  • BSL-1: Low hazard; standard practices; no primary barriers beyond basic PPE; facilities include basic sinks and doors.
  • BSL-2: Moderate hazards; BSCs for some procedures; controlled access; waste decontamination; autoclave available.
  • BSL-3: Serious/high hazard; enhanced barriers; directional airflow; entry/exit controls; separate containment; autoclave and decontamination systems.
  • BSL-4: Extreme hazard; Class III BSC or suit-based containment; separate building; complete decontamination and dedicated systems.

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• Activity 2: Apply knowledge to real-life scenarios; explore risk-control measures using local resources; consider different geographic contexts.

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• Break time.

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• Title: BIOHAZARD SAMPLE MANAGEMENT overview.

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• 3PS framework (Official Closed/Non-Sensitive):
  • Protect User
  • Protect Sample
  • Protect Environment

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• Protect User: treat all samples as potentially infectious; apply Hazard Identification and Risk Assessment (HIRA) to select safeguards.
• Protect Sample: maintain sample integrity; prevent spills/breakage; control transit conditions; prevent contamination.
• Protect Environment: ensure proper containment at all stages; use appropriate containment equipment (e.g., biosafety cabinet or fume hood) during processing; safeguard proper disposal per HIRA.

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• Testing and Storage process:
  • Use correct sample type; proper labeling, containers, and safety precautions.
  • Record-keeping; transport from off-site if needed.
  • Pre-examination storage; testing; post-testing storage.
  • Requires Good Documentation Practices (GDP) and Good Laboratory Practices (GLP).

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• Do’s for GDP documentation:
  • Entries in indelible ink, clear handwriting.
  • If blank, mark with a cancellation or N.A.; sign with date.
  • Initial/cix date corrections; record reasons for corrections when appropriate.

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• Don’ts for GDP documentation:
  • Do not leave columns blank; use N.A. or nil.
  • Do not overwrite entries; cross out and preserve original text; write corrected entry nearby.
  • No pencil entries; no backdating; do not use correction tape.

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• GDP exercise prompts:
  • Identify good and bad GDP practices; propose correct practices for the bad GDP items.

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• GLP framework and its components:
  • Organization & Employee, SOPs, GDP, GLP; ensures traceability and regulatory compliance; supports planning, execution, documentation, reporting, and archiving.

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• Biohazard waste management principles:
  • Segregate and contain potentially infectious materials.
  • Use rigid, puncture-resistant, leak-proof containers labeled with biohazard symbols; fill to about 80 hundred? capacity to avoid puncture; ensure proper containment for sharps, liquids, and solids.
  • Liquid waste: treat with disinfectant (e.g., bleach) before disposal; Sharps: seal lid at 80\ ext{%} full; Solids: autoclave before licensed disposal; All waste managed by designated waste company; do not discharge liquids into sinks.

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• What you have learned (summary):
  1) Sources of biological hazards in labs.
  2) Factors in biological risk-group classification.
  3) Infrastructures of four biosafety levels for safe handling.
  4) SOPs and GDP for handling, storage, and waste management.
  5) Application of SOPs and GDP for safe sample handling and disposal.