Biosafety & Bioethics Principles & ideas

Gives brief idea about the laboratory safety and bioethical principles

1. BIOSAFETY AND BIOETHICS NEP Sem II Devlina Sengupta Department of Microbiology Kanchrapara College

2. Biosafety Biosafety is defined as prevention of large scale loss of Biological integrity with focuses both on ecology and human health It is a system of handling of toxic and dangerous biological and chemical substances Components of safety Safe handling, Storage and Disposal of Specimens, Chemicals, Instruments, Radioactive components and Fire and electrical safety Disasters management for natural calamities like Earthquakes, Hurricanes etc. Devlina’s Lectures 2

3. Technical Terms Biohazard: An agent of biological origin that has the capacity to produce deleterious effects on human That may be Microorganisms and/ or Toxins, allergens developed from them or Toxins and /or allergens developed from higher plants and animals (PBHA- potential Biological Hazardous Agents) Biosafety: The containment (Inhibitory) principles, technologies and practices that are implemented to prevent unintentional exposure to pathogens and toxins and their accidental release Biosecurity: Control of accidental or deliberate release of biohazardous materials Devlina’s Lectures 3

4. Containments (Inhibitions) The term Containment is used to describe the safe methods of managing infectious agents in laboratory environments where they are being handled and maintained To reduce the exposure of laboratory workers and other persons to hazardous environment Elements of Containments: Laboratory Practice and Techniques (GLP) Safety Equipment (Primary Barriers) Design facility (Secondary Barriers) Devlina’s Lectures 4

5. Types of Containments Physical Containments: Physical methods adopted in laboratory for preventing the escape of GEOs to the environment Ex. Air filters, Sterilization lights, Waste disposal and Protective handling Biological Containments: Biological Principles used to prevent the escape of GEOs and Microbes Ex. Conditions make the organism unable to survive outer environment Devlina’s Lectures 5

6. Devlina’s Lectures 6 Some unfortunate examples……. 1950- Typhoid fever 2003 -2014: Large outbreaks of SARS, Ebola, Dengue 1980 : Large outbreaks of Hepatitis, Tuberculosis 2019: Corona pandemic through out the world

7. Need of Biosafety To support the medical and scientific research on microbes and the human immune response to them To apply such research to the discovery and development of vaccines, drugs and diagnostic tests to designed to protect the general population To insure that the every has sufficient research facilities to carry out these activities Devlina’s Lectures 7

8. Scope of Biosafety Academic Research in the field of: Ecology Medicine Chemistry Agriculture Exobiology For providing advice on laboratory design, risk assessment of experiments involving infectious agents and in vivo and in vitro rDNA studies Devlina’s Lectures 8

9. Biosafety Issues Laboratory safety Blood born Pathogens (BBP) Recombinant DNA (rDNA) and production of genetically modified food and organisms Biological waste disposal Shipping of infectious substances and diagnostic specimen Devlina’s Lectures 9

10. Work practices: Good laboratory practices (GLP) First introduced in New Zealand in 1972 In 1992 organizations for Economic cooperation & Development (OECD) laid principles of GLP. DST established a national GLP compliance monitoring authority (NGCMA) in 2002 India is full time member for Mutual Acceptance of Data (MAD) in the OECD’s working group on GLP since 2011. Devlina’s Lectures 10

11. Why Good laboratory practices (GLP) It provides a framework within which the laboratory studies are planned, performed, monitored, recorded, reported and achieved. A primary requisite to ensure the safety of an individual and the environment. Principles of GLP apply to all non-clinical health and environmental safety studies (unless exempted by national legislation) Devlina’s Lectures 11

12. Significance of GLP: Quality & Integrity Data generated is used for risk assessment of hazards and risks to users, including environment & can be assessed for pharmaceuticals, agrochemicals, cosmetics, food & feed additives & contaminants, novel food & biocides. Devlina’s Lectures 12

13. Principles of GLP(by OECD) Test facility Organization & personnel Quality Assurance programme (QAP) Facilities Apparatus, materials & reagents Test systems Test & reference items Standard operating procedures(SOPs) Performance of the study Reporting of study results Storage & retention of records & materials Devlina’s Lectures 13

14. Areas of Risks Risk for health: Toxicity, Allergies, Pathogen drug resistance (Antibiotic resistance) Risk for Environment: Persistency of transgene elements and products, susceptibility to nontarget organisms and unpredicted expressions Risk in Agriculture: Resistance/ Tolerance of target organisms, production of weed or super weed, Alteration of nutritional value Risk of interaction with non target organisms: Genetic pollution, Horizontal gene transfer, Generation of new line viruses General concerns: Hight cost requirements, Loss of familiarities, Ethical issues Devlina’s Lectures 14

15. To: Lab Personals, Community Through: Various Specimens, Human blood, unfixed tissue, Human cell lines From: Fungi, Bacteria, Viruses, Parasites, Prions rDNA Devlina’s Lectures 15

16. Risk Groups RG 1: Agents not associated with diseases in healthy humans RG 2: Agents associated with rare human diseases RG 3: Agents associated with serious and lethal human diseases RG 4: Agents associated with serious and lethal human diseases for which preventive measures are not available Devlina’s Lectures 16

17. Biosafety Levels (BSL) It refers to classification system used to indicate the safety precautions required for known to be dangerous or lethal experiments It is a level of bio- containment precautions required to undertake while handling of hazardous biological agents in an enclosed facility through a combination of safeguards including Engineering control Management policies Standard work practice Devlina’s Lectures 17

18. BSL 1: Work with least dangerous agents and require fewest precautions BSL 2: Agents associated with human diseases BSL 3: Agents causing serious diseases transmitted by inhalation for which immunization and antibiotic treatment is available BSL 4: Agents causing serious diseases for which known treatment and immunization is not available Devlina’s Lectures 18

19. Barriers Primary Barriers: Physical barriers like personal protective equipment and breathing apparatus Secondary Barriers: Safe laboratory structural aspects, sinks for handwash, air exhausts and sterilization equipment and Staff immunization with TB, Rubella, Typhoid, Anthrax, Poliomylites and Diphtheria Devlina’s Lectures 19

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22. Biosafety Cabinets (BSC) Devlina’s Lectures 22

23. • Room air is drawn in through the front opening at a minimum velocity of 0.38 m/s, it passes over the work surface and is discharged from the cabinet through the exhaust duct • The front opening allows the operator’s arms to reach the work surface inside the cabinet while he or she observes the work surface through a glass window. •The air from the cabinet is exhausted through a HEPA filter: ( a) into the laboratory and then to the outside of the building through the building exhaust; ( b) to the outside through the building exhaust; or ( c) directly to the outside . •Does not produce reliable product protection but can be used in works w/ radionuclides & volatile toxic chemicals. Cheap due to its simple design. Biological safety cabinets: BSC Class I Devlina’s Lectures

24. • The Class II BSC was designed not only to provide personnel protection but also to protect work surface materials from contaminated room air . • Class II BSCs, of which there are four types (A1, A2, B1 and B2 ), differ from Class I BSCs by allowing only air from a HEPA-filtered (sterile) supply to flow over the work surface. • The Class II BSC can be used for working with infectious agents in Risk Groups 2 and 3. Can also be used w/ Risk Group 4 if positive pressure suits are used. • Class II A1 has an internal fan drawing room air into the cabinet through the front opening & into the front intake grill at an inflow velocity of 0.38 m/s. The supply air then passes through a supply HEPA filter before flowing downwards over the work surface. Exhaust is to room or thimble connection • Class II A2 air is vented to the outside with air intake velocity of 0.51m/s. Exhaust is to thimble connection • Class II B1 & B2 differ in air recirculation into the workspace and they exhaust into the hard duct •Used in pathology centres & research institutes. Biological safety cabinets: BSC Class II Devlina’s Lectures

25. • BSC Class III type provides the highest level of personnel protection and is used for Risk Group 4 agents . • All penetrations are sealed “gas tight”. • Supply air is HEPA-filtered and exhaust air passes through two HEPA filters. • Airflow is maintained by a dedicated exhaust system exterior to the cabinet, which keeps the cabinet interior under negative pressure • Access to the work surface is by means of heavy duty rubber gloves , which are attached to ports in the cabinet . • The Class III cabinet may be connected to a double-door autoclave used to decontaminate all materials entering or exiting the cabinet • suitable for work in BSL -3 & 4 laboratories Biological safety cabinets: BSC Class III Devlina’s Lectures

26. Risk group classification categorizes biological agents based on their potential for harm to humans and the environment. It’s a system used in laboratory settings and research to determine appropriate safety precautions and containment measures. Risk groups range from 1 (least hazardous) to 4 (most hazardous) Risk groups Devlina’s Lectures

27. Devlina’s Lectures 28 RG 1 Agents BSL 1 Practice Non- pathogenic E.coli Transgenic Plants Plasmids (Ti plasmid, pUC plasmid) Fungi Yeast Bench top work Daily Decontamination Manual pipetting Proper handwashing Red bag waste Safety glasses, gloves and lab coats Standard Microbiological practice Laboratory Facilities: Slip resistant, smooth, hard, & liquid tight floors, minimum 12 inches clearance above BSCs, eyewashes & safety showers

28. Devlina’s Lectures 30 RG 2 Agents BSL 2 Practice Human and Primate cells Herpes Simplex Virus Attenuated human immunodeficiency virus Hepatitis B virus Salmonella , Clostridium Patient specimens like blood, body fluids and infected tissues Limited access to lab Daily decontamination Mechanical pipetting Red bag waste Safety glasses, gloves and lab coats / PPE Laboratory Facilities: supply of generator for the support of essential equipments when necessary such as incubators, BSCs, freezers, animal ventilators etc , mechanical ventilation system that provide inward flow of air w/o recirculation of air, all other BSL1 facilities

29. Devlina’s Lectures 32 RG 3 Agents BSL 3 Practice Human immunodeficiency virus Mycobacterium tuberculosis Coxiella burnetii No public access Daily decontamination Foot activated handwashing sinks , micro incinerator No sharp containers Plastic disposable transfer loops Screw capped tubes & bottles Provision of active or passive immunization for workers Laboratory Facilities: Anterooms with self closing doors, Heating, ventilation & air-conditioning (HVAC), electronic surveillance & detection system, uninterruptible power systems, liquid effluent treatment system

30. Devlina’s Lectures 34 RG 4 Agents BSL 4 Practice Highly contagious viruses like Ebola, Herpes B, Lassa fever virus, Corona virus All facilities of BSL 3 w/ two person rule Maximum containment facilities (controlled delineated zone, double door autoclave, controlled air system by negative pressure, fumigation chamber Chemical decontamination showers Class III biosafety cabinets (cabinet room/suit laboratory is required) [refer lab biosafety manual by WHO at pg 26 for suit lab] Waste liquid collection and decontamination Containment drain, airlock entry ports for specimens, materials & animals must be provided

31. T he assignment of a biosafety level takes into consideration the organism (pathogenic agent) used, the facilities available, and the equipment practices and procedures required to conduct work safely in the laboratory . • Most common containments are listed accordingly for the four BSL levels Biosafety level requirements 35 Devlina’s Lectures

32. Aim of Biosafety guidelines Regulation of rDNA research with organisms that have no or limited adverse effects Minimizing the possibilities of occasional release of GEOs from the laboratory Banning the release of GEOs if they are supposed to be causing potential risk in the environment Devlina’s Lectures 36

33. Biosafety Guidelines Internationally WHO Laboratory Biosafety Manual has been in broad use at all levels of clinical & public health laboratories Many countries have established NBF (national biosafety framework) to ensure safe handling and use of GMOs. Refer Cartagena protocol on biosafety & IFBA (International Federation of Biosafety Association) to regulate the movement of genetically engineered organisms. Biosafety clearing house(BCH) has been set up by Cartagena protocol to facilitate exchange of scientific, technical, environmental & legal information on GMOs & LMOs. Devlina’s Lectures 37

34. Biosafety Guidelines Internationally Different organizations & database information on biosafety International plant protection convention (IPPC): Agreement that protects plants by assessing & managing risks of plant pest that are associated w/ GMO. Codex Alimentarius Commission : deals w/ health & food safety in case food is derived from genetic engineering. Provides guidelines for GM foods & lays emphasis on labelling GMOs. Convention on Biological Diversity: raises the concern of potential impact of biotechnology application on biodiversity & emphasizes & elaborates the needs & precautions to be taken in safe handling of biotechnology products. World Trade Organization (WTO): to ensure smooth & free flow of trade b/w nations Food & Agriculture Organization of the United Nations (FAO): Encourages the countries to take the advantage of advanced technologies in food & agro sector. Organization for Economic co-operation & Development (OECD): to promote policies & provide forums in which governments can work together to share experiences related to economic & social well being of people around the world. Devlina’s Lectures 38

35. International biosafety guidelines, like those established by the  World Health Organization (WHO)  and the  Biosafety Clearing-House , aim to ensure: • T he safe handling, transport, and use of biological materials, especially living modified organisms (LMOs).  • These guidelines cover a wide range of aspects, including risk assessment, containment measures, and emergency response procedures.  • They are crucial for protecting human health and the environment.  Biosafety Guidelines Internationally Devlina’s Lectures 39

36. Examples of International Biosafety Guidelines WHO Laboratory Biosafety Manual (LBM): This manual provides comprehensive guidance on biosafety in laboratories, covering various aspects from risk assessment to emergency response.  Cartagena Protocol on Biosafety: This international agreement governs the safe movement and use of living modified organisms (LMOs).  International Health Regulations (IHR): These regulations, adopted by the World Health Organization (WHO) and the  UNICEF , provide a framework for countries to address public health emergencies, including those related to biological agents.  Devlina’s Lectures 40

37. rDNA Biosafety Guidelines of India Policy proposed or Adopted by the Government to avoid the risks of GEOs on Environment and Public Health The Recombinant DNA Safety Guidelines proposed by DBT in 1983 which were amended in 1990 The Guidelines deals with a set of rules for hazardous organisms for: Production Use Import Export Storage Devlina’s Lectures 41

38. Committees for rDNA guidelines implantation in India Recombinant DNA advisory committee (RDAC) Institutional Biosafety Committee (IBSC): Controls research activities at institutional level Review Committee on Genetic Manipulation (RCGM): R eviews for situation for research on Hazardous organisms and grants permission to work and provide advice on IPR and Patents , it assists BIS (Bureau of Indian standards) to evolve standards of products coming from rDNA technology Genetic Engineering Approval Committee (GEAC): It works under the Department of Environment & Forests, approved the GEOs for large scale production and field use Devlina’s Lectures 42

39. Toxin gene cloning Cloning of genes for vaccine production Cloning of mosquito & tick DNA Cloning of antibiotic resistance gene Cloning of oncogenes Experiments with infectious animals and plant viruses Transgenesis experiments in animal cell culture Transfer of toxicity genes into plants Gene therapy for hereditary disease Permission of review & approval committee before commencement of following works Devlina’s Lectures 43

40. • Organized by DBT under DST, GOI •It provides regulatory control to the implementation committees • Provides a forum for public discussion to ensure that gene transfer research has the necessary public transparency . • Recombinant DNA Advisory Committee (RDAC) is mandated to review developments in Biotechnology at national and international levels and recommend suitable and appropriate safety regulations for India in recombinant research, use and applications from time to time. Recombinant DNA Advisory Committee ( RDAC) Devlina’s Lectures 44

41. • The Institutional Biosafety Committee ( IBSC ) is a standing committee and is responsible for reviewing all University research and teaching activities conducted by faculty, staff, students and/or visiting scientists on University property that involve the use of biological agents, but are not limited to, recombinant DNA, RNAi , pathogens, human materials and other potentially infectious material, as well as transgenic animals. • The IBSC provides recommendations to the intramural community in matters pertaining to the control of biohazards associated with the use of microbiological agents and their vectors . • It also represents the interests of the surrounding community with respect to public health and protection of the environment Institutional Biosafety Committee ( IBSC) Devlina’s Lectures 45

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43. The review committee on genetic manipulation (RCGM) established under the Department of Biotechnology, Ministry of Science and Technology monitor the safety of on-going research projects and activities (including small scale field trials, import, export etc ) involving genetically engineered organisms.  The  RCGM  shall include representatives of  •    Department of biotechnology (DBT) •    Indian council of medical research(ICMR) •    India council of agricultural research (ICAR) •    Council of scientific and industrial research (CSIR) •    Other experts in their individual capacity Review Committee on Genetic Manipulation (RCGM) Devlina’s Lectures 47

44. The committee's function is to determine the scope of laws and regulations for institutions involved in RDNA research and activities  •    review of current research involving harmful microorganisms  •    Visit the experimental site and make sure that the tests are carried out in accordance with the regulations.;  •    Provide advice to the customs authority on import of micro-organisms and products, G.M.O. Functions of Review Committee on Genetic Manipulation(RCGM) Devlina’s Lectures 48

45. GEAC plays a crucial role in balancing the benefits of biotechnology in agriculture with the need to protect human health and the environment. It has the following functions: • To appraise activities involving large scale use of hazardous microorganisms and recombinants in research and industrial production from the environmental angle. • To appraise proposals relating to release of genetically engineered organisms and products into the environment including experimental field trials. • The committee or any persons authorized by it has powers to take punitive action under the Environment Protection Act. Genetic Engineering Approval Committee ( GEAC) Devlina’s Lectures 49

46. The IAEC is responsible for ensuring ethical standards in animal research, reviewing protocols, and minimizing animal suffering during experiments (following Brambell’s five freedom) T hey encourage 3R to reduce the impact of research on animals- Replacement, Reduction Refinement. IAEC is responsible for ensuring ethical standards in animal research, reviewing protocols, and minimizing animal suffering during experiments . Institutional Animal Ethics Committee (IAEC) Prof. Roger Brambell reported this on livestock husbandry in 1965, approved by UK Farm animal welfare council (FAWC) Devlina’s Lectures 50

47. SUMMARY OF RECOMMENDED BIOSAFETY LEVELS FOR ANIMAL RESEARCH Devlina’s Lectures 51

48. Activities need to acquire permission and Approval Toxin gene cloning and transfer in plants Cloning of genes for Vaccine production Cloning of Antibiotic Resistance gene Cloning of Oncogenes Experiments with infectious animals Experiments with Plant Viruses Transgenesis experiments with animal cell cultures Gene therapy for hereditary diseases Devlina’s Lectures 52

49. Ethics and Morals Study of Ethical and Moral effects of new biological discoveries and biomedical advances The term Bioethics was first introduced by Van Rensselaer Potter in 70s for protection of Biosphere It was later used to refer to study of the ethical issues arising from health care, biological and medical sciences Devlina’s Lectures 53

50. Ethics Vs Moral It is concern with right or wrong conduct Guides to behaviour Making choice or Judgement Mor a ls are individual frame work for decision making Ethics are generalized conceptual frame work for decision making Devlina’s Lectures 54

51. Moral Considerations in Ethics Whether one ought to act to maximize the best outcomes Are we required only not to harm others or must we also act in ways that benefit them or make their lives better? What should be done when we think policies or law are unethical How could we access that all people have a fair or maybe an equal opportunity How and when should we share information about a medical treatment to best permit others Devlina’s Lectures 55

52. Bioethics Bioethics concerns itself with issues in healthcare, medicine, research, biotechnology, and the environment Devlina’s Lectures 56 Professionals working in the field of bioethics include philosophers, scientists, health administrators, lawyers, anthropologists and social workers Each field contributes important insights, resources and methodologies and efforts to think about or make changes to practices and policies that raise ethical concerns

53. Bioethics in Biotechnology Bioethics also needs to address emerging biotechnologies that affect basic biology and future humans These developments include: Gene Cloning Gene therapy Human genetic engineering Manipulation of basic Animal and Plant biology through altered DNA, RNA and proteins Devlina’s Lectures 57

54. However the use of this technology has raised some concerns, which include: Their ecological impact Their potential harm to human health Their negative impact due to development of resistance Their contribution to corporate monopoly So, there is a need to address life through Ethical point of views Devlina’s Lectures 58

55. Principles of Bioethics Autonomy: Right of self determination and ability to take decisions independently Beneficence (Kindness): To do well, to promote well being and work for the benefit of others Non maleficence: To do no harm or to avoid doing harm Justice: Treat every one alike and consider the equity One more….. Openness: Not to keep the information secret Role of International Bioethics Committee (IBC) of UNESCO Devlina’s Lectures 59

56. Scope of Bioethics Human genome projects Stem cell research Artificial reproductive technologies Pre- implementation diagnosis The synthesis of new life forms The possibilities of successful reproductive cloning A ll work related to human genome manipulation should be based on Nuremberg code. Devlina’s Lectures 60

57. Major Issues of Bioethics Eugenics: DNA manipulation for improvement of human race Human cloning: Scientific production of identical human cell, tissue and entire body Advancement of species and extinction of certain genetic diseases Lead to discrimination of races and possibilities of eradication of race Development of regenerative medicines, tissue and organ donation Lead to human farming and killing the clones for body parts Devlina’s Lectures 61

58. Stem cell research: Differentiation of stem cell into a range of specialized cell types Euthanasia: Purposefully ending a life Potential for disease curing Lead to destruction of life As a relief of unnecessary suffering Lead to end of blessed life Devlina’s Lectures 62

59. Devlina’s Lectures 63 Thank You…..