BIOSAFETY TRAINING 2017
The “Laboratory Biosafety Training Course” aims at training laboratory professionals in biosafety and biorisk management according to the highest international standards, promoting the knowledge and the dissemination of these standards in Italy and abroad, with particular attention to developing countries.
The presence of about 10 (maximum) participants from all over the world is envisaged. Although candidates will have homogeneous bio-medical background, the variety of presented topics addresses the needs and interests of different professionals in the field of bio-risk management.
Good command of English is required.
Please download the registration form here.
Laboratory Biorisk Management
Biorisk is defined as the combination of the probability of occurrence of harm and the severity of that harm where the source of harm is a biological agent or toxin. Biorisk management is a management system to manage the biorisk in an organisation. Any organisation that contains facilities where work is carried out with potentially hazardous biological materials or genetically modified organisms (GMO) should implement a biorisk management structure and policy.
A successful biorisk management system relies on commitment by the top management and a focus on continual improvement. Biorisk management starts with a risk assessment based on planned projects with biological agents. At the same time, the organisation must obviously follow any applicable legislation.
Many of the elements of a biorisk management system are also implied by general biosafety measures. However, the management system involves a higher level system to ensure that the organisation as a whole (top management, middle management, and all other involved employees) is functioning in a coordinated manner. It also implies monitoring, corrective actions, and reviewing.
An effective management system approach should be built on the concept of continual improvement through a cycle of planning, implementing, reviewing and improving the processes and actions that an organization undertakes to meet goals. This is known as the PDCA (Plan-Do-Check-Act) principle.
Recently, as part of a biorisk management system, the competence of biosafety professionals is increasingly seen as a crucial success factor. The International Federation of Biosafety Associations (IFBA) has published a handbook “Ensuring Quality Biorisk Management Through Certification of Professionals”.
 CEN Workshop Agreement CWA 15793 (2011) “Laboratory biorisk management”.
 CEN Workshop Agreement CWA 16393 (2012) ” Laboratory biorisk management – Guidelines for the implementation”.
According to a very general definition, biosafety is the prevention of large-scale loss of biological integrity, focusing both on ecology and human health. Related fields are ecology, agriculture, medicine, chemistry, exobiology, and synthetic biology. Several international treaties or conventions cover one or more of these topics. In this lecture the term biosafety will be limited to laboratory aspects. This is relevant for any field where potentially hazardous biological materials or genetically modified organisms (GMO) are used in a laboratory facility.
The main focus in such a setting is on containment and protection of workers. Containment is required to prevent escape from the laboratory and spread into the environment of potentially hazardous biological materials (mainly: pathogens). A pathogen can be any infectious agent (virus, bacterium, fungus, parasite, prion), mainly human pathogens, but also animal and plant pathogens which can have devastating effects on agriculture, economy, or ecology. In addition, non-infectious agents can include biological toxins, GMO’s, or as yet not clearly defined synthetic biological entities.
The basic approach since many years has been to define four risk levels, referred to as risk groups 1, 2, 3, and 4, with risk group 4 being the highest risk level. For each level, specific risk control measures are described, including physical measures (construction, barriers, ventilation, PPE), organisational measures (roles, functions, responsibilities, training), administration (biosafety manual, procedures, licences, database), and monitoring measures (validation, inspection).
Several guidelines or biosafety manuals that describe these control measures can be found on internet, including from the WHO, CDC (US), Canadian government, and the EU,. In addition, several countries have national legislation or guidelines. This lecture will not focus on these specific documents, but on the general principles of biosafety and its implementation.
Biosafety knowledge exchange is provided by several national and international biosafety associations, such as EBSA and ABSA.
 Wikipedia (https://en.wikipedia.org/wiki/Biosafety)
 EU directive 2000/54/EC on protection of workers from risks related to exposure to biological agents
 EU directive 2009/41/EC on the contained use of genetically modified micro-organisms
Laboratory Biosecurity, Dual Use of Biological Materials and Ethical Issues
Gijsbert Van Willigen
Laboratory biosafety describes containment principles, technologies, and practices to protect people from biological agents, and prevent accidental release of biological agents. In addition to biosafety, laboratory biosecurity measures aim to prevent theft and intentional or malicious use of biological agents. Thus, both biosafety and biosecurity should be an integral part of program management of organizations handling dangerous pathogens, in order to prevent potential dual-use research, undesired spread, theft, malicious use, and bioterrorism.
A well implemented laboratory biosecurity program should be implemented. Dual-use biological agents can be used in bioterrorism. Dual-use good are materials that have a civilian and a military application. For these dual-use organisms a stringent biosecurity regime should be in place. Besides the wild type agent themselves, biological agents can also be modified in the lab to become more dangerous than the wild-type organism itself. This is called dual-use research of concern (DURC). One type of DURC research id gain-of-function research of GOF.
Besides the risks of dual-use agents and DURC/GOF also ethical questions can arise from the application of GOF.
The lecture will focus on:
– what should be part of a proper laboratory biosecurity program and who are stakeholders in the program;
– what are dual-use materials;
– what are dual-use biological agents are and the basis why agents are classified as dual-use. The criteria include besides the pathogenicity also other factor such a how can they be cultured and dispersed;
– types of research will be presented as examples of DURC/GOF and published examples will be discussed;
– ethical questions.
Key Reference Reading:
Simon Whitby et al. Preventing Biological Threats: What You Can Do, (Bradford: University of Bradford, 2016).
Tatyana Novossiolova, Biological Security Education Handbook: The Power of Team-Based Learning, (Bradford: University of Bradford, 2016).
 Both books are freely available at: http://www.bradford.ac.uk/social-sciences/peacestudies/research/publications-and-projects/guide-to-biological-security-issues/
Most developed countries have legislation for work with biohazardous materials at a national level, including authorities and inspection bodies. All EU member states fall under the European biosafety legislation. The biosafety rules that apply in the US and Canada are often considered in the EU as valuable resources for comparison. There is extensive overlap between the various guidelines.
There is a basic distinction between legislation for work with naturally occurring hazardous biological agents (BA) and genetically modified organisms (GMO). Although risk assessment, risk levels, and safety measures are often comparable between work with either BA or GMO, there can be very relevant distinctions. At the EU level, there are specific guidelines for BA and for GMO. Specific legislation exists for transport of hazardous biological materials, e.g. the Dangerous Goods Regulations of IATA. Additional legislation might exist for publication of knowledge, dual use, import and export, use in environment (especially for GMO), and use in agriculture or food.
Legislation and guidelines can change over time. This can be caused by new knowledge of biohazard, evolving insight, or newly discovered pathogens. The planned eradication of poliovirus is causing the WHO to issue plans for stricter safety rules for work with poliovirus.
Accreditation is the process in which certification of competency, authority, or credibility is presented. If compliancy is concluded by an independent third party, an official written statement (certificate) is issued to confirm this. Certification is a more loose term that refers to the confirmation of certain characteristics of an object, person, or organisation. Usually this is accomplished by some form of external review or audit. Organisations that issue certificates are often in turn accredited by official accreditation bodies, which are established in many countries with the primary purpose of ensuring that conformity assessment bodies are subject to oversight by an authoritative body. Certification does not state an absolute level of competency, but establishes compliancy of something with a set of requirements. Compliancy is not meaningful if the requirements are not clearly described. In many cases certification is based on internationally established standards such as ISO.
 EU directive 2000/54/EC on protection of workers from risks related to exposure to biological agents
 EU directive 2009/41/EC on the contained use of genetically modified micro-organisms
 International Accreditation Forum (IAF), http://www.iaf.nu
Primary Barriers: PPE & Laboratory equipment
Personal protective equipment (PPE) consists of a wide range of equipment and materials that are worn by a person in order to protect him/her against biohazardous materials. Examples are gloves, boots, coveralls, eye and face protection, and breathing protection. The proper use of PPE is essential for biosafety. The user must be trained sufficiently. Training with live agents is very effective for building confidence. Training must include the correct donning (putting on) and doffing (taking off) of PPE. Decontamination of a person still wearing PPE is often a standard part of work before doffing. Usually a sequence of steps is chosen to exit from the hot zone via the decontamination area to the safe zone. Doffing of PPE in the correct order is part of such a sequence.
There is no standard PPE, not even for a specific biosafety level. PPE must always be adapted to the specific work conditions and surrounding, and especially to the particular infectious organism being handled. Knowledge of the usual routes of infection of an organism helps in selecting the appropriate PPE components.
PPE should not inhibit the work to be carried out. PPE should also not pose a high burden upon the worker, as this might encourage a worker to not consistently using it. This relates to weight and general comfort, but also to heat and moisture building up inside a coverall. Breathing should be easy even under moderately stressful work conditions.
Biosafety equipment is any type of equipment that is used for containing biohazardous materials inside the biosafety area, for preventing the spread of such materials, or for protecting workers against contact with such materials.
Perhaps the most central piece of equipment is the biosafety cabinet. Besides protecting the worker and preventing spread of materials in the laboratory (or outside), they also protect the biological materials from becoming contaminated with biological materials from the environment or even from within the work space inside the cabinet (cross contamination). Several types of biosafety cabinets exist, with different properties and adapted to different kinds of work or risk level. The proper use of biosafety cabinets is crucial to their performance and workers need to be trained for doing so. The placement of biosafety cabinets in a room and connection to the ventilation system are also important factors to address.
Secondary Barriers: Facility design and construction
Gijsbert van Willigen
The principles of biohazard control centre on the concept of containment. The purpose of containment is to eliminate or reduce exposure of the environment and the outside environment from potentially dangerously pathogens. Using containment infectious agents can be handled in laboratory in a safe way.
There are three major elements of containment:
- Laboratory practices and techniques, i.e. Good Laboratory Practices
- Safety equipment, i.e. primary containment
- Facility design and construction i.e. secondary containment
Facility design and construction contributes to the laboratory workers protection, provides a barrier to protect humans and animals outside the laboratory and provides protection from infectious agents if the primary barrier has failed or collapsed.
The strength of secondary containment depends on the risks that are posed by the biological agent. Secondary containment of a BSL1 or BSL2 laboratory can consist of a separation of the lab from public areas, hand washing facilities and the availability of an autoclave in the building.
When the risk of the agent increases or the transmission route is via aerosols multiple layers of secondary containment measures may become necessary to prevent escape of the infectious agent from the laboratory. These extra layers of secondary containment include the use of specialized Heating, Ventilation and Air Conditioning systems (HVAC systems) to ensure “negative differential pressure” or “inward directional airflow “, treatment of exhaust air using HEPA filtration to remove infectious agents from the exhaust air, the use of airlocks for entrance, controlled access, building management systems and even security systems to prevent theft of the infectious agents from the facility.
In the presentation the concepts of secondary containment together with the challenges from an engineering and users point of view will be discussed. The BSL3+ facility of the Leiden University Medical Center will be used as an example where biosafety and biosecurity are well integrated into one facility.
Good Laboratory Practices
Good laboratory practices are all those methods and procedures that in combination with other measures (e.g. physical barriers or equipment) ensure the safe handling of hazardous biomaterials. In several countries the most often used description is Good Microbiological Practice (GMP), not to be confused with Good Manufacturing Practice. Other terms used include Standard Microbiological Practice (SMP) or Safe Microbiological Practice (also SMP).
Any biosafety procedure or biosafety equipment can be rendered useless if the user makes mistakes or does not use the equipment properly. The correct use of equipment and proper execution of methods by the microbiological worker is at the core of biosafety. A proper level of education and sufficient training are the most essential starting points for GMP.
Many international guidelines include paragraphs on GMP, such as from the WHO, CDC, and others. International biosafety associations such as ABSA and EBSA also publish guidelines for GMP.
GMP involves aspects such as training, peer control, procedures, SOP’s, and other elements. Several organisations offer training courses for GMP. Many use videos and photos to illustrate essential elements of GMP. GMP is not a replacement for a microbiological education, it is merely an extension, relying on a solid microbiological education as basis.
Disinfection, Sterilization and Decontamination
Gijsbert van Willigen
For inactivation of biohazardous material, including biohazardous waste, a large number of methods are available ranging from chemical inactivation using chemical disinfectants to various types of a physical process, such as heat for inactivation. All methods aim at rendering the biohazardous material harmless for humans, animals and the environment. In the presentation a large numbers of methods for disinfection, sterilization and decontamination will be discussed that can be used in a laboratory setting together with their advantages and disadvantages for use.
The method of choice for inactivation of biohazardous materials greatly depends on the composition, volume and risk level of the biological material, but also if the material that is being decontaminated is reusable or disposable. Also economic factors can be considered when choosing an inactivation method. After a method is chosen the inactivation process must be validated.
For a proper disinfection, sterilization and decontamination, validation and verification of the processes should be in place. For this several methods can be used to demonstrate the effectiveness of the process depending on the method used. This ranges from biological tests to the use of biological indicators when physical processes are used for inactivation and measuring physical parameters of the process. In the presentation pros and cons of these methods will be discussed.
Biohazardous Waste Management
Gijsbert Van Willigen
Solid (including sharps) or liquid biohazardous waste is unavoidably produced in facilities where laboratory experiments, experiments with animals are performed during the treatment of patients. For save handling of this waste it should be collected in such a way that it does not pose any risk for the people who handle or transport this waste inside or outside of the institute. Special bags, over packs and containers should be used for safe collection of biohazardous waste. They should be leak and puncture proof and can be used for various types of decontamination. Preferably they cannot be reopened. For biohazardous waste national and international legislation is in place for dealing with biohazardous waste.
The presentation will focus on the collection, storage and transport of biohazardous waste. Also some of the commonly made mistakes will be shown. Waste treatment i.e. inactivation will be the topic of the lecture on disinfection, sterilization and decontamination and will only be briefly touched in this lecture.
Practical exercises with fluorescent markers and UV lights
Gijsbert Van Willigen, Martien Broekhuijsen
Purposes of the exercises:
1. To see if hand washing was done thoroughly.
2. To demonstrate how easy a contamination occurs.
3. To see if surface cleaning was done thoroughly, or see effect of different cleaning methods.
Further details were provided during the exercises.
The security of work place is one of the most important aspects of the working activity. The risk assessment for the scientific laboratory, chemical and biological, is linked with the presence of hazardous factors present in the daily lab activities. Among the different elements to be considered in a risk assessment, there are the agents, the equipment, the problems linked with the space limitations, organizational/management aspects and sometimes the lack of information, education and training of the staff. The presence of these factors in a laboratory setting could cause laboratory accidents, such as break of tubes in a centrifuge, projection of liquid in eyes, spills, accidental injection of a contaminated solution, aerosol of liquid solution that could generate the spread of agents into the laboratory environment thereby causing the infection of workers or the contamination of equipment and materials, etc. Accordingly, a relevant problem in the context of the laboratory accidents is all infection acquired through laboratory or laboratory-related activities regardless whether they are symptomatic or asymptomatic in nature. This infections are defined as Laboratory Acquired Infection (LAI) or laboratory-associated infections. Among the LAI, the bloodborne Pathogens are important especially for HBV, HCV and HIV infections.
A prevention plan is mandatory to minimize the risk of accidents. Every laboratory should perform a risk assessment in order to evaluate which are the risks that could be encountered inside the laboratory. Risk assessment must take in account every material, every procedures, workers, environment and the different protective equipment. In the context of laboratory accidents, personnel training plays a key role: the workers are directly exposed to risk in case of emergency and they are the first operators involved in emergency maneuvers.
Laboratory Acquired Infections
Laboratory-acquired infections due to a wide variety of bacteria, viruses, fungi, and parasites have been described. Although the precise risk of infection after an exposure remains poorly defined, surveys of laboratory-acquired infections suggest that Brucella species, Shigella species, Salmonella species, Mycobacterium tuberculosis, and Neisseria meningitidis are the most common causes. Infections due to the bloodborne pathogens (hepatitis B virus, hepatitis C virus, and human immunodeficiency virus) remain the most common reported viral infections, whereas the dimorphic fungi are responsible for the greatest number of fungal infections. Because of the increasing attention on the role of the laboratory in bioterrorism preparation, I discuss the risk of laboratory-acquired infection with uncommon agents, such as Francisella tularensis and Bacillus anthracis. Physicians who care for a sick laboratory worker need to consider the likelihood of an occupationally acquired infection while advising exposed laboratory workers about postexposure prophylaxis. In addition, physicians should be aware of the importance of alerting the laboratory if infection with a high-risk agent is suspected.
Introduction to Occupational Medicine
The aims of occupational medicine are the identification of health risks in the workplace, health surveillance of workers, diagnosis and treatment of occupational diseases, and in collaboration with professionals with other expertise (e.g. engineers, chemists, industrial hygienists, psychologists) participation to training programs and activities of risk management measures. Over the year, a shift has been observed in the attitude towards health and safety in the workplace. From the XVIII century Bernardino Ramazzini’s first textbook on occupational diseases to the modern legislation, the active involvement of the workers has been increased. In fact, in the XXI century workers are required to actively participate in the management of health and safety in the workplace. This has two consequences: on one hand workers are entitled to provide suggestions and participate in the planning of the activities related to their health and safety through their representatives; on the other hand workers also became responsible and liable of their behaviour with respect to their and their co-workers’ health and safety.
Accidents (injuries) in the workplace, and occupational or work-related diseases are taken care of by specific insurance schemes, that may differ between countries, even within the European Union. According to the Organisation of Economic Co-operation and Development (OECD) “An occupational injury is any personal injury, disease or death resulting from an occupational accident; an occupational injury is therefore distinct from an occupational disease, which is a disease contracted as a result of an exposure over a period of time to risk factors arising from work activity”. Instead, the term occupational disease “is linked to the identification of a specific cause-effect relationship between a harmful agent and the affected human organism. However, it is not easy – and considerably more difficult than in the case of accidents – to prove that a disease is occupationally conditioned, i.e. caused by conditions at, not outside work”. Because of the difficulty in proving a disease to be occupational in origin, most countries have produced lists of prescribed occupational diseases. These are generally limited to those diseases where a strong cause-effect relationship has been proven. However, with the number of categories ranging from 50 to 90, national lists vary in terms of those diseases recognised as occupational. Recommended lists developed by the International Labour Organisation and the European Communities seem to have led only to limited degree of harmonisation. In addition, in certain countries, including Italy, the list is not prescriptive, and any worker can claim a disease as occupational or work-related provided that she/he is able to prove the causal relationship. Each country has different legislative approaches related to registration, notification, and compensation for occupational injuries or diseases and their health consequences.
The risks related to exposure to biological agents at work are subjects to an EU directive (2000/54/EC of 18 September 2000). This Directive has general provisions that have been implemented by local legislation in EU member Countries. General provisions of the directive include the definitions and the assessment of risks. Employers’ obligations including replacement, reduction of risks, information to authority, provision of adequate hygiene and individual protection devices, information and training of workers, compilation of a list exposed workers, active participation of workers, and notification to the competent authority. In addition, health surveillance, measures for the workplace, classification of biological agents based on their characteristics. The class of the biological agent will determine the different provisions that should be taken to protect the health and safety of the workers, and the consequent administrative, technical, preventive and sanitary measures.
BSL3: Agents & Specificities
Microorganisms are internationally classified by a risk group (1,2,3,4).
Different microorganisms (viruses and bacteria) are reported as high priority agents that pose a threat to national security. They can be easily disseminated or transmitted person-to-person, cause high mortality, with potential for major public health impact, might cause panic and social disruption and require special public health preparedness.
For their diagnosis different methods are used: microscopy, molecular tests, antigens and antibodies.
BSL 3 agents pose a risk for operators and communities. For this reason, their manipulation requires dedicated measures, in order to minimize the occurrence of laboratory acquired infections and the spread among the population. Different national and international institutions (i.e.: WHO, CDC) provide indications to properly work with these agents.
BSL 3 facilities can serve both research and diagnostic purposes; obviously, differences related to these aims exist, mainly concerning the use of animal models.
BSL 3 laboratories are enclosed environments: air and fluids systems are dedicated and not shared with other areas; moreover, only authorized personnel can enter. Such a condition can be obtained with an engineering planning before the construction building phase. Regarding the work in the laboratory, specific rules indicate which standards that must be followed as regards: Personal Protective Equipment, safety cabinets, decontamination agents, work procedures. Also emergency situations must be considered: they need particular measures, not adopted in daily routine activity.
In the Laboratory of Clinical Microbiology, Virology and Bioemergencies (CLIMVIB) at ASST Fatebenefratelli Sacco, a BSL 3 facility is present: its use is strictly related to diagnostic purpose, without any animal specificity. The examples presented refer to clinical experience and depict a real-life situation: Biosafety rules and indications are adapted to CLIMVIB needs and they could differ from other institutions. Naturally, each institution must meet Biosafety principles, in order to prevent personnel infections, epidemic and/or pandemic and related panic in the communities.
BSL4: Agents & Specificities
Romualdo Grande, Davide Mileto
The facilities of biosafety level 4 (BSL4) are designed in order to manage biological agents belonging to risk group 4 and patients infected by such kind of agents.
The main feature of this facility is related with the need to avoid the infection of workers but also with the requirement to prevent the spread of the level 4 biological agents into the environment.
Different international guidelines (WHO or CDC guidelines) describe all the specificities and the correct standards of behavior to be adopted for the BSL4 facilities.
Among them surely the need to have always almost two workers inside of the laboratory and an external support team outside of the laboratory. Communication between the two different teams, internal and external of the laboratory, is mandatory in order to monitor all the different steps and procedures and to act in case of emergency. Laboratory protective clothing must be of the type with solid-front or wrap-around gowns, scrub suits, coveralls, head covering and, where appropriate, shoe covers or dedicated shoes. All the personal protective equipment must be decontaminated before being disposed of. The complexity of the structure makes it mandatory to organize periodic training also to cope with accidents or illnesses.
Different technical features are needed for the BSL4 laboratory.
A negative pressure system inside the laboratory guarantees the primary containment. This condition is also provided thanks to the system of interlocked door necessary to enter and exit from the laboratory. The biosafety cabinet of level III are used inside the laboratory whereas supplies and materials are introduced through a double-door autoclave or fumigation chamber. A HEPA filter system cleans incoming and outgoing air flows including the air flow need to work with the positive pressure suit. All effluents from the suit area, decontamination chamber, decontamination shower, or class III biological safety cabinet must be decontaminated before final discharge.Emergency power and dedicated power supply lines must be provided.
HOW TO REACH L. SACCO UNIVERSITY HOSPITAL
Via G.B. Grassi 74
Stop Milano Certosa (2 km distant to L. Sacco University Hospital)
-Suburban line S5 (Varese – Treviglio )
-Suburban line S6 (Novara – Treviglio)
-Line 12, 19 (end of line in front of L.Sacco University Hospital)
The course will take place at the Laboratory of Clinical Microbiology, Virology and Bioemergencies, of L. Sacco University Hospital, headed by Prof. Maria Rita Gismondo. The laboratory includes BSL2, BSL3 and BSL4 facilities fully equipped not only for diagnostic purposes, but also for training and hands-on educational modules.