Biosafety in microbiology laboratory

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Biosafety in Microbiology laboratoryBIOSAFETY : Preventing lab-acquired infections

Bacteria Viruses Fungi Human blood, unfixed tissue Human cell lines Recombinant DNA

Why is Biosafety Important? Laboratories recognize hazards of processing infectious agents Guidelines developed to protect workers in microbiological and medical

labs through engineering controls, management policies, work practices

Standard Microbiological Practices

NOT permitted in laboratories:

Eating Drinking Smoking Handling contact lenses Pipetting by mouth Storing food and drink

Amjad Khan Afridi 21/07/ 2016

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Biosafety Levels

Precautions so people researching or trying to identify organisms do not become infected

While handling or testing clinical specimens, workers could accidentally infect themselves or coworkers

Labs must adhere to very specific safety regulations to work with organisms that pose a threat to human health

Laboratories divided on basis of nature of microbes

Labs divided into 4 biosafety levels; protective practices increase with each

Biosafety Level 1 labs : work with least dangerous agents, require fewest precautions

Biosafety Level 4 labs : have strictest methods because dealing with agents that are most dangerous to human health

Barriers : Primary barriers• Primary barriers: physical barriers or personal protective

equipment between lab worker and pathogen• Gloves, masks, special breathing apparatuses


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Barriers Secondary barriers:

• Secondary barriers: structural aspects of the laboratory that make working environment safer against infection• Sinks for hand washing, special containment areas, special air

ventilation patterns

Universal Precautions:

• Include hand hygiene, gloves, gown, masks, eye protection, face shields, safe injection practices

• Require that all equipment or contaminated items are handled to prevent transmission of infectious agents

• Special circumstances may require additional precautions

• Protective clothing, special site decontamination


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Levels of Containment


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BL1 - microorganisms that don’t consistently cause disease in healthy adults– E. coli K12, S. cerevisiae, polyomavirus – Basic laboratory – Standard Microbiological Practices

Biosafety Level 1 (BSL-1)

Agents not known to cause disease in healthy adults

– Some organisms may cause disease in immunocompromised individuals

Agents include Bacillus subtilis, Naegleria gruberi, infectious canine hepatitis virus, non-pathogenic E. coli species

Standard practices required:

– frequent handwashing

– door that can be kept closed when working;– limits on access to the lab space when working;

– no smoking, eating, drinking, storage of food in laboratory;


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– care to minimize splashes and actions that may create aerosols (tiny droplets);

– decontamination of work surfaces after every use after any spills;

Standard practices (continued):

– decontamination of laboratory wastes;

– use of mechanical pipettes only (no mouth pipetting);

– "sharps" precautions, including special containers for disposing of needles and other sharp objects;

– maintenance of insect/rodent control program;

– use of personal protective equipment (lab coats, latex gloves, eye protection or face shields)

– Open bench top sink for hand washing

Biosafety Level 2 (BSL-2)

Agents associated with human disease

– Generally required for any human-derived blood, bodily fluids, tissues in which infectious agent may be unknown

– Agents include measles virus, Salmonella species, pathogenic Toxoplasma, Clostridium botulinum, hepatitis B virus

Levels of Containment BSL-2 : microorganisms of moderate potential hazard, transmitted by contact, ingestion, puncture

– Salmonella, herpesvirus, human blood

– Basic laboratory – Standard Practices PLUS


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Primary hazards:

– accidental needle sticks– exposure to eyes and nose (mucous membranes)

– ingestion of infectious materials Agents do not cause lethal infections, are not transmissible via

airborne route

– (do not cause infection if tiny droplets become airborne and are inhaled, which might occur if the material were spattered)

Agents are pathogens for which immunization or antibiotic treatment is available

Extreme care should be taken with contaminated needles and sharp lab instruments

Risk Group 2Pathogenic for humans Unlikely a serious hazard Treatment and preventive measures available Limited risk of spread of infection


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Standard practices include BSL-1 plus:

– policies to restrict access to lab;– biohazard warning signs posted outside lab;

– surveillance of laboratory personnel with appropriate immunizations offered;

– biosafety manual with definitions of needed waste decontamination or medical surveillance policies;

– supervisory staff who have experience working with infectious agents and specific training for laboratory personnel in handling these agents

Primary barriers: biosafety cabinets or other approved containment devices

Personal protective equipment: lab coats, gloves, face protection as needed

Protective clothing removed when personnel leave laboratory area

Cabinets thoroughly decontaminated daily and monitored for radiation for personal protection

Secondary barriers: BSL-1 barriers plus autoclave for glassware

Levels of Containment :


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BSL-3 - microorganisms that cause serious disease, transmitted by inhalation

– M. tuberculosis, yellow fever virus, hantavirus, Y. pestis (plague)

– Containment lab: double door entry; directional airflow; all work in biosafety cabinet

Biosafety Level- 3 (BSL-3):Agents with potential for respiratory transmission, may cause serious and potentially lethal infection

– May be studied at BSL-2 for diagnosis Agents include Mycobacterium tuberculosis, St. Louis encephalitis virus, Francisella tularensis, Coxiella burnetii

TB diagnostics and laboratory strengtheningCare of patients with tuberculosis starts with a quality assured diagnosis, obtained by growing and identifying Mycobacterium tuberculosis from clinical specimens and conducting DST of the organism to confirm or exclude resistance. Uptake of TB diagnostic technologies requires appropriate laboratory infrastructure and adequate policy reform at country level to enable their effective use in TB screening and diagnostic algorithms

• Laboratory infrastructure, appropriate biosafety measures and maintenance Equipment validation and maintenance Specimen transport and referral mechanisms Management of laboratory commodities and supplies Laboratory information and data


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management systems Laboratory quality management system are a priority.

Mdr – Tb , xmdr-tb and biosafety

• With growing incidences of MDR-TB and XMDR-TB it is highly essential all Microbiology laboratories must install Grade 3 Biosafety cabinets to prevent exposure to Infection. It necessary precaution's are not taken a fraction of Medical and Technical personal will be infected with grave consequences.

Risk Group 3Pathogenic, cause serious disease Effective treatment and preventive measures usually available Little person-to-person spread.

Biosafety Level -3 (BSL-3)


– strictly controlled access to the lab;

– specific training for lab personnel in handling potentially lethal agents;


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– decontaminating all waste;

– changing contaminated protective lab clothing, decontaminating lab clothing before laundering;

– institutional policies regarding specimen collection and storage from workers to establish exposure.

Primary barriers:

– Similar to BSL-2 personal protective equipment

– Respiratory equipment if risk of infection through inhalation

Secondary barriers:

– All BSL-2 barriers

– Corridors separated from direct access to lab

– Access through self-closing double doors

– Air handling systems to ensure negative air flow (air flows into the lab)

– Air pumped into lab not re-circulated in building .

Levels of Containment

BSL-4 - microorganisms that cause lethal disease, with no known treatment or vaccine

Ebola virus, Marburg virus

Maximum containment lab; positive pressure ventilated suits (moon suits)

Risk Group 4Amjad Khan Afridi 21/07/ 2016

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– Lethal, pathogenic agent Readily transmittable– direct, indirect Effective treatment and preventive measures not usually available

Biosafety Level- 4 (BSL-4)• Dangerous and exotic agents with high risk of life-threatening disease, aerosol-

transmitted

• Related agents with unknown risk of transmission

• Agents (all viruses) include Marburg virus, Ebola virus, viruses that cause Congo-Crimean hemorrhagic fever, Lassa fever

• Primary hazards:

respiratory exposure to infectious aerosols

mucous membrane exposure to infectious droplets

accidental sticks with needles or other sharp objects contaminated with infectious material

• For example

In late 1960s, 25 laboratory-acquired Marburg infections, including 5 deaths

Workers studying infected monkeys from Uganda

First documented naturally-occurring human case occurred in 1975

Personnel must receive specialized training in handling extremely dangerous infectious agents, containment equipment and functions


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Access to lab is restricted: immunocompromised persons are never allowed to enter the lab


strictly controlled access to the laboratory;

changing clothing before entering and exiting lab (showering upon exiting recommended);

decontaminating all material exiting facility

Primary barriers:

Biosafety cabinets used at other biosafety levels

Full-body, air-supplied, positive pressure personnel suit

Secondary barriers:

All physical barriers at BSL-3

isolated zone or a separate building;

dedicated supply and exhaust, vacuum, decontamination systems;

a recommended absence of windows (or sealed and resistant to breakage)

Laboratory Locations


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BSL-1: high schools, community colleges, municipal drinking water treatment facilities

BSL-2: local health departments, universities, state laboratories, private laboratories (hospitals, health care systems), industrial laboratories (clinical diagnostic companies)

BSL-3: state health departments, universities, private companies, industry, federal government (NIH, CDC)

BSL-4: only 15 facilities in the US

– 9 federal (CDC, NIH), 4 university (Georgia State University, University of Texas Medical Branch), 1 state, 1 private

– Renovations underway at several labs, new facilities proposed at additional sites

Standard Microbiological Practices

NEVER

– recap, bend, or break needles

– discard needles or sharps into biological waste bags

– discard needles into regular trash

Biosafety is everyone's concern

Laboratorians have long recognized hazards of processing infectious agents Biosafety guidelines developed to protect workers in microbiological and medical labs

through a combination of safeguards including engineering controls, management policies and work practices.

Issue described differences between biosafety levels Help you understand process labs may have to undertake to identify microorganism, why

every lab cannot test for every organism .


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Demonstrated by Dr. Hayat Khan


Health & Medicine

Biosafety in microbiology laboratory