Principles and Practices of Biosafety

Principles and Practices of Biosafety

Environmental Health and SafetyEnvironmental Health and Safety

San Diego State UniversitySan Diego State University

Abbreviations BSC – Biosafety Cabinet BSC – Biosafety Cabinet BSL – BBSL – Biosafety Leveliosafety Level ABSL – Animal BABSL – Animal Biosafety Leveliosafety Level BSO – Biosafety OfficerBSO – Biosafety Officer BUA – BUA – Biological Use AuthorizationBiological Use Authorization CA – CaliforniaCA – California CDC – Center for Disease Control and CDC – Center for Disease Control and

PreventionPrevention DGR – Dangerous Goods RegulationsDGR – Dangerous Goods Regulations DOT – Department of TransportationDOT – Department of Transportation EHS – Environmental Health and EHS – Environmental Health and

SafetySafety EPA – Environmental Protection EPA – Environmental Protection

AgencyAgency HMR – Hazardous Materials HMR – Hazardous Materials

RegulationsRegulations

IACUC – IACUC – Institutional Animal Care Institutional Animal Care and Use Committee and Use Committee

IATA – International Air Transport IATA – International Air Transport AssociationAssociation

IBC – Institutional Biosafety IBC – Institutional Biosafety CommitteeCommittee

IRB – Institutional Review BoardIRB – Institutional Review Board NIH – National Institute of HealthNIH – National Institute of Health OSHA – Occupational Health and OSHA – Occupational Health and

Safety Administration Safety Administration PI – Principal InvestigatorPI – Principal Investigator PP – Physical PlantPP – Physical Plant PPE – Personal Protective Equipment PPE – Personal Protective Equipment PS – Pubic SafetyPS – Pubic Safety SD – San DiegoSD – San Diego USPS – U.S. Postal ServiceUSPS – U.S. Postal Service

Introduction

The management of biological hazards through the The management of biological hazards through the proper application of engineered containment and proper application of engineered containment and administrative controls is referred to as administrative controls is referred to as biosafety biosafety or biohazard controlor biohazard control..

Biosafety or biohazard control is a team effort Biosafety or biohazard control is a team effort involving the PI, research lab personnel, BSO, involving the PI, research lab personnel, BSO, IBC, IRB, IACUC, EHS, PP and PS.IBC, IRB, IACUC, EHS, PP and PS.

Regulatory Requirements and Guidelines NIH Guidelines for Research Involving NIH Guidelines for Research Involving

Recombinant DNA Molecules (April 2002)Recombinant DNA Molecules (April 2002) CDC/NIH Biosafety in Microbiological and CDC/NIH Biosafety in Microbiological and

Biomedical Laboratories (May 1999)Biomedical Laboratories (May 1999) CAL/OSHA Bloodborne Pathogen StandardCAL/OSHA Bloodborne Pathogen Standard CA Medical Waste Management Act CA Medical Waste Management Act DOT Transportation of Hazardous DOT Transportation of Hazardous

MaterialsMaterials

SDSU Biosafety Requirements and Guidelines Policies established by the Institutional Biosafety Policies established by the Institutional Biosafety

Committee (IBC) that meet or exceed applicable Committee (IBC) that meet or exceed applicable guidelines and regulations for use of RG 2 and guidelines and regulations for use of RG 2 and RG 3 biohazardous materials or agents as well as RG 3 biohazardous materials or agents as well as non-exempt NIH Recombinant DNA research. non-exempt NIH Recombinant DNA research.

Policies established by Environmental Health and Policies established by Environmental Health and Safety (EHS) that meet or exceed applicable Safety (EHS) that meet or exceed applicable regulations and guidelines for minimizing regulations and guidelines for minimizing bloodborne pathogen exposure and disposal of bloodborne pathogen exposure and disposal of biohazardous wastes.biohazardous wastes.

Biohazardous Material Infectious microorganisms Infectious microorganisms

(bacteria, viruses, fungi, (bacteria, viruses, fungi, parasites, prions, parasites, prions, rickettsiae, etc.) affecting rickettsiae, etc.) affecting humans and animalshumans and animals

Diagnostic (clinical) Diagnostic (clinical) specimensspecimens

Recombinant DNA (viral Recombinant DNA (viral vectors, gene therapy, vectors, gene therapy, cloning)cloning)

Genetically Modified Genetically Modified Microorganisms Microorganisms (transgenic plants and (transgenic plants and animals)animals)

Human and non-human Human and non-human primate cells, cell culture primate cells, cell culture (primary and (primary and immortalized), tissues, immortalized), tissues, blood (whole blood or any blood (whole blood or any components) and body components) and body fluidsfluids

Animal or plant cells, cell Animal or plant cells, cell cultures, fluids, tissues or cultures, fluids, tissues or derived wastes which derived wastes which may contain pathogensmay contain pathogens

Animals known to be Animals known to be reservoirs of zoonotic reservoirs of zoonotic diseasedisease

Institutional Biosafety Committee and

Biological Use Authorization

Biological Use Authorization (BUA) Application BSL 1, ABSL 1 and NIH-Exempt BSL 1, ABSL 1 and NIH-Exempt

Reviewed and approved by BSO Reviewed and approved by BSO

BSL 2 or 3, ABSL 2 or 3 and NIH- Non-exemptBSL 2 or 3, ABSL 2 or 3 and NIH- Non-exempt Reviewed and approved by IBC Reviewed and approved by IBC

Currently, no facilities at SDSU meet the Currently, no facilities at SDSU meet the minimum criteria for BSL 4; thus this type of minimum criteria for BSL 4; thus this type of work is prohibited.work is prohibited.

Submission of New BUA

New infectious agents New infectious agents New cell lines New cell lines New vector system New vector system Enhanced replication or infectivity Enhanced replication or infectivity Expression of toxic products Expression of toxic products Partial genomes increased to more than two-thirds Partial genomes increased to more than two-thirds

of whole genome of whole genome New or altered procedures that pose increased risk New or altered procedures that pose increased risk

(e.g., aerosol or other type of exposure) (e.g., aerosol or other type of exposure) Work with non-human systems being changed Work with non-human systems being changed

into work with human system into work with human system

SDSU Biosafety Approval

Institutional Biosafety Committee (IBC)Institutional Biosafety Committee (IBC) Protocol ReviewProtocol Review Biological Use Authorization (BUA) Biological Use Authorization (BUA)

with respect to containment level with respect to containment level (biosafety level)(biosafety level)

TrainingTraining InspectionInspection

Principal Investigator Develops specific protocols to ensure the safe use Develops specific protocols to ensure the safe use

of biohazardous materials. of biohazardous materials. Submits a BUA application and obtains approval Submits a BUA application and obtains approval

from IBC prior to commencement of work. from IBC prior to commencement of work. Complies with specific biosafety protocols, Complies with specific biosafety protocols,

practices and procedures described in the practices and procedures described in the biohazard control manual. biohazard control manual.

Ensures that all laboratory staff are appropriately Ensures that all laboratory staff are appropriately trained on biosafety. trained on biosafety.

Reports any significant problems, violations of Reports any significant problems, violations of the policies, practices and procedures, or any the policies, practices and procedures, or any significant research-related accidents and/or significant research-related accidents and/or laboratory acquired infection to the BSO laboratory acquired infection to the BSO immediately.immediately.

Laboratory Staff

Comply with the specific biosafety Comply with the specific biosafety protocols, practices and procedures protocols, practices and procedures described in the biohazard control manual.described in the biohazard control manual.

Report to the PI or the lab manager all Report to the PI or the lab manager all problems, spills or violations in procedure problems, spills or violations in procedure immediately.immediately.

Risk Group and

Biosafety Level

Classification of Infective Agents by Risk Group

Pathogenicity Pathogenicity Infectious Dose Infectious Dose Mode of Transmission Mode of Transmission Host Range Host Range Availability of Effective Preventive Availability of Effective Preventive

Measures and TreatmentMeasures and Treatment

Classification of Infective Agents by Risk Group

Risk Group 1 Risk Group 2 Risk Group 3 Risk Group 4

Severity of Disease

unlikely to cause human or animal disease

can cause disease, unlikely to be serious, effective treatment and preventive measures available

can cause serious disease, does not ordinarily spread from one person to another, effective treatment and preventive measures usually available, exposure route: inhalation (often)

likely to cause serious or lethal disease, can be readily transmitted from one individual to another, effective treatment and preventive measures are not usually available, transmission: direct, indirect, inhalation

Host Range human (healthy adult) and animals

human (healthy adult) and animals



Individual Risk

low moderate (potential hazard)

high high

Community Risk

low low low high

Classification of Containment by Biosafety Levels Practices and Procedures Practices and Procedures Containment Equipment (Primary Barriers)Containment Equipment (Primary Barriers) Containment Facility (Secondary Barriers) Containment Facility (Secondary Barriers)

Note: CDC/NIH has 4 biosafety level classifications Note: CDC/NIH has 4 biosafety level classifications currently in use. Each level is appropriate for: currently in use. Each level is appropriate for: Operations Performed Operations Performed Routes of Transmission (ingestion, inoculation, Routes of Transmission (ingestion, inoculation,

inhalation, mucous membrane exposure)inhalation, mucous membrane exposure) Laboratory FunctionLaboratory Function

Risk Group and Biosafety Level Classifications Not appropriate to use risk group or Not appropriate to use risk group or

biosafety level when assessing toxins.biosafety level when assessing toxins. Can be found in the American Biological Can be found in the American Biological

Safety Association website:Safety Association website:

www.absa.org/resriskgroupwww.absa.org/resriskgroup Risk Groups and Biosafety Levels are not Risk Groups and Biosafety Levels are not

always the same! always the same!

Bloodborne Pathogen Standard

Introduction

California Code of Regulations, Title 8, Sec 5193California Code of Regulations, Title 8, Sec 5193 Applies to all employees who could “reasonably Applies to all employees who could “reasonably

anticipate” as a result of performing their job tasks anticipate” as a result of performing their job tasks contact with blood and other potentially infectious contact with blood and other potentially infectious material (OPIM) i.e., body fluids, unfixed tissues material (OPIM) i.e., body fluids, unfixed tissues or organs or organs

Limit occupational exposure to blood and OPIM Limit occupational exposure to blood and OPIM which could result in transmission of bloodborne which could result in transmission of bloodborne pathogens i.e., Hepatitis B virus (HBV), HIV pathogens i.e., Hepatitis B virus (HBV), HIV

Requires a written Exposure Control PlanRequires a written Exposure Control Plan

Elements of an Exposure Control Plan Exposure Determination Exposure Determination Method of Compliance Method of Compliance HIV, HBV and HCV Research Laboratories HIV, HBV and HCV Research Laboratories Hepatitis B Vaccination and Post-Exposure Hepatitis B Vaccination and Post-Exposure

Evaluation and Follow-Up Evaluation and Follow-Up Hazard Communication Hazard Communication Record Keeping Record Keeping Evaluation of Exposure Incidents Evaluation of Exposure Incidents Sharps Injury LogSharps Injury Log

Exposure DeterminationExposure Determination

Exposure evaluation based upon the job Exposure evaluation based upon the job description.description.

Exposure evaluation based upon reasonably Exposure evaluation based upon reasonably anticipated contact (skin, eye, mucous anticipated contact (skin, eye, mucous membrane, parenteral contact, etc.) with membrane, parenteral contact, etc.) with blood or other potentially infectious blood or other potentially infectious materials resulting from performing the materials resulting from performing the assigned tasks.assigned tasks.

Method of ComplianceMethod of Compliance Universal Precaution – treating Universal Precaution – treating all human blood and all human blood and

certain human body fluids as if infectious for bloodborne certain human body fluids as if infectious for bloodborne pathogens pathogens

Engineering and Work Practice Controls Engineering and Work Practice Controls Needleless system or needles/sharps with engineered sharps injury Needleless system or needles/sharps with engineered sharps injury

protection protection Needles and other sharps shall not be recapped, bent or broken Needles and other sharps shall not be recapped, bent or broken Needles and other sharps shall be disposed in rigid, puncture-Needles and other sharps shall be disposed in rigid, puncture-

proof, leak resistant and properly labeled sharps container proof, leak resistant and properly labeled sharps container Sharps container shall be closed immediately prior to removal or Sharps container shall be closed immediately prior to removal or

replacement to prevent spillage or protrusion of contents during replacement to prevent spillage or protrusion of contents during handling or transport handling or transport

Specimens of blood or OPIM shall be placed in a closable, Specimens of blood or OPIM shall be placed in a closable, leakproof, properly labeled red bag prior to handling, collection or leakproof, properly labeled red bag prior to handling, collection or transporttransport

Personal Protective Clothing and EquipmentPersonal Protective Clothing and Equipment

Hepatitis B Vaccination and Hepatitis B Vaccination and Post-Exposure Follow-UpsPost-Exposure Follow-Ups Hepatitis B vaccination shall be provided at Hepatitis B vaccination shall be provided at

no cost to employee who has a potential for no cost to employee who has a potential for becoming exposed to blood or OPIM. becoming exposed to blood or OPIM.

Post-exposure follow-ups shall be provided Post-exposure follow-ups shall be provided if an occupational exposure occurs. if an occupational exposure occurs.

Hazard CommunicationHazard Communication1.1. Signs and Labels Signs and Labels Signs shall be posted at the entrance to the work areas Signs shall be posted at the entrance to the work areas

which shall bear: which shall bear: Name of infectious agent Name of infectious agent International symbol for biohazard in fluorescent International symbol for biohazard in fluorescent

orange-red orange-red Special requirements for entering the area Special requirements for entering the area Name and telephone number of lab director or other Name and telephone number of lab director or other

responsible person responsible person Warning labels shall be affixed to containers of Warning labels shall be affixed to containers of

infectious wastes, refrigerators and freezers infectious wastes, refrigerators and freezers containing blood or OPIM, or other containers used containing blood or OPIM, or other containers used to store or transport blood or OPIM. to store or transport blood or OPIM.

Labels shall have the international symbol for biohazard Labels shall have the international symbol for biohazard in fluorescent orange-red in fluorescent orange-red

2.2. TrainingTraining

Pathogenic Microbiology

Bacterial Laboratory Acquired Infections 76% of exposures occurred in clinical labs; 76% of exposures occurred in clinical labs;

6% in vaccine manufacturing facilities; 8% 6% in vaccine manufacturing facilities; 8% in research labs. in research labs.

Exposure modes: 60% inhalation, other Exposure modes: 60% inhalation, other exposure modes included ingestion exposure modes included ingestion (intentional, poor technique such as mouth (intentional, poor technique such as mouth pipetting, smoking and eating in the lab) pipetting, smoking and eating in the lab) and secondary transmission.and secondary transmission.

Viral Laboratory Acquired Infections >70% associated with research labs; 32% of all >70% associated with research labs; 32% of all

viral LAIs associated with animals viral LAIs associated with animals 18% of total were Hantavirus; of these, 8% were 18% of total were Hantavirus; of these, 8% were

working with known infectious material or rodents working with known infectious material or rodents - others thought they were working with - others thought they were working with uninfected rodents uninfected rodents

16% were in clinical labs; rest were in production 16% were in clinical labs; rest were in production or field work or field work

Major exposure modes: inhalation, percutaneous Major exposure modes: inhalation, percutaneous (especially from animals)(especially from animals)

Rickettsial Laboratory Acquired Infections All rickettsial LAIs were associated with research All rickettsial LAIs were associated with research

laboratories laboratories 95% of overt infections were by 95% of overt infections were by Coxiella burnetiiCoxiella burnetii; ;

remainder were Murine typhus remainder were Murine typhus All Q fever cases by inhalation; all infected staff All Q fever cases by inhalation; all infected staff

worked with or were in close proximity to sheepworked with or were in close proximity to sheep Remaining cases were by percutaneous, Remaining cases were by percutaneous,

inhalation, mucous membrane or unknown inhalation, mucous membrane or unknown exposure modesexposure modes

Molecular Biology

It’s a Matter of Perspective

The investigators who submit IBC protocols The investigators who submit IBC protocols want to perform their experiments safely.want to perform their experiments safely.

However, their perception of the risks However, their perception of the risks involved will not necessarily be the same as involved will not necessarily be the same as that of a biosafety professional.that of a biosafety professional.

Risk Assessment

The following risk assessment will identify the The following risk assessment will identify the biological containment system to be used:biological containment system to be used: Properties of the donor organismProperties of the donor organism Nature of the DNA sequences that will be Nature of the DNA sequences that will be

transferredtransferred Properties of the recipient organismProperties of the recipient organism Properties of the environmentProperties of the environment

Biological Expression System

Most routine genetic engineering experiments Most routine genetic engineering experiments can be performed safely in can be performed safely in E. coliE. coli K12/pUC18 at BSL 1 provided the inserted K12/pUC18 at BSL 1 provided the inserted foreign DNA sequences do not require a foreign DNA sequences do not require a higher BSL.higher BSL.

Donor Organism and Cloned DNA

Insertion of well-characterized DNA sequences that Insertion of well-characterized DNA sequences that are unlikely to be involved in pathogenicity may are unlikely to be involved in pathogenicity may not require additional safety measures. not require additional safety measures.

In cases where these sequences are not characterized, In cases where these sequences are not characterized, a situation that is typically encountered when a a situation that is typically encountered when a library of genomic DNA of an organism is being library of genomic DNA of an organism is being established, a higher BSL will be required.established, a higher BSL will be required.

Cloning of genes coding for proteins that have Cloning of genes coding for proteins that have potential pharmacological activity such as toxins potential pharmacological activity such as toxins may therefore require higher BSL.may therefore require higher BSL.

Viral Vectors for Gene Transfer

Although viral vectors used in gene therapy or Although viral vectors used in gene therapy or gene transfer are replication-defective, they gene transfer are replication-defective, they should be handled at the same BSL as the should be handled at the same BSL as the parent viral vector from which they are parent viral vector from which they are derived since the virus stocks may be derived since the virus stocks may be contaminated with replication-competent contaminated with replication-competent viruses, which are generated by rare viruses, which are generated by rare spontaneous recombination events in the spontaneous recombination events in the complementing cell line.complementing cell line.

Transgenic and “Knock-Out” AnimalsAnimals carrying foreign genetic information Animals carrying foreign genetic information

(transgenic animals) should be handled in the (transgenic animals) should be handled in the containment level appropriate to the containment level appropriate to the characteristics of the products of the foreign characteristics of the products of the foreign genes. For each new line of transgenic animal, the genes. For each new line of transgenic animal, the routes by which the animals can be infected, the routes by which the animals can be infected, the inoculum size required for infection, and the inoculum size required for infection, and the extent of the virus shedding by the infected animal extent of the virus shedding by the infected animal must be determined.must be determined.

Animals with targeted deletions of specific genes Animals with targeted deletions of specific genes (“knock-out” animals) do not generally present (“knock-out” animals) do not generally present particular biological hazards.particular biological hazards.

Human and Other Primate Cells and Tissues

Human Source MaterialHuman Source Material

Blood and blood products Blood and blood products Vaginal secretions Vaginal secretions Semen Semen Amniotic fluid Amniotic fluid Unfixed tissues Unfixed tissues Cerebrospinal, synovial, Cerebrospinal, synovial,

pleural, pericardial and pleural, pericardial and peritoneal fluids peritoneal fluids

Cell culturesCell cultures

Saliva Saliva Urine Urine Tears Tears Sputum Sputum Feces Feces Vomit Vomit Other excretions and Other excretions and

secretions secretions

Second column not covered in Bloodborne Pathogen Standard, possibly not occupationally related.

Human Source MaterialHuman Source Material

May transmit infectious agents May transmit infectious agents Imperfect knowledge of infectious statusImperfect knowledge of infectious status

Incubation period (asymptomatic) Incubation period (asymptomatic) No test for every pathogenNo test for every pathogen

Most tissues and body fluidsMost tissues and body fluids Bloodborne Pathogens (HBV, HCV, HIV, Bloodborne Pathogens (HBV, HCV, HIV,

HTLV-1)HTLV-1) Pathogens causing Malaria, Syphilis, Pathogens causing Malaria, Syphilis,

Babesiosis, Brucellosis, Leptospirosis, Babesiosis, Brucellosis, Leptospirosis, Arboviral infections, Relapsing fever, Arboviral infections, Relapsing fever, Creutzfeldt-Jacob disease and viral Creutzfeldt-Jacob disease and viral hemorrhagic feverhemorrhagic fever

Cell Culture RisksCell Culture Risks Contaminating pathogenic agents Contaminating pathogenic agents

natural (often zoonotic) or inadvertent natural (often zoonotic) or inadvertent ~20 LAIs from primary cultures in last 30 years ~20 LAIs from primary cultures in last 30 years e.g., Herpes B (CHV-1), prions e.g., Herpes B (CHV-1), prions

Oncogenic potential Oncogenic potential could be oncogene or oncogenic agent could be oncogene or oncogenic agent e.g., HPV-18, MPMV genomes in HeLa cells e.g., HPV-18, MPMV genomes in HeLa cells

Unexpected (adventitious) agents Unexpected (adventitious) agents e.g., SIV, STLV, SV5 in primate cells, HHV-8 e.g., SIV, STLV, SV5 in primate cells, HHV-8

in BCBL-1 cells in BCBL-1 cells Hazardous chemicals added to culture medium Hazardous chemicals added to culture medium

Cell Culture under Bloodborne Cell Culture under Bloodborne Pathogen StandardPathogen Standard ABSA requested OSHA’s interpretation in 1994: ABSA requested OSHA’s interpretation in 1994:

Do human cell cultures fall under the Do human cell cultures fall under the Bloodborne Pathogen (BBP) Standard? Bloodborne Pathogen (BBP) Standard?

Response: Response: All primary human cell cultures (explants) and All primary human cell cultures (explants) and

subsequent subsequent in vitroin vitro passages fall under the passages fall under the BBP Standard BBP Standard

To be exempted from the BBP requirements, To be exempted from the BBP requirements, cell strains and lines must undergo testing and cell strains and lines must undergo testing and characterization (documented) for bloodborne characterization (documented) for bloodborne pathogens (not just HBV, HCV and HIV)pathogens (not just HBV, HCV and HIV)

Cell Culture SafetyCell Culture Safety

Extend Universal/Standard Precautions to all Extend Universal/Standard Precautions to all human and animal cell cultures human and animal cell cultures Consider working at BSL 2 (most work Consider working at BSL 2 (most work

there already to protect the cell cultures) there already to protect the cell cultures) Handle all cultures in a biosafety cabinet Handle all cultures in a biosafety cabinet If human origin and not demonstrated to If human origin and not demonstrated to

be free of human bloodborne pathogens, be free of human bloodborne pathogens, adhere to requirements of the BBP Standard adhere to requirements of the BBP Standard

Wear PPE appropriate to human source Wear PPE appropriate to human source materialmaterial

Human Source MaterialsHuman Source Materials May be regulated May be regulated Can be biohazardous Can be biohazardous Use Universal Precautions at Use Universal Precautions at

all times all times Visible blood means Visible blood means

increased risk increased risk Don’t consider “normal” Don’t consider “normal”

sourcesource

Human and Non-Human and Non-human Primate Cell human Primate Cell CulturesCultures

Treat human cultures as Treat human cultures as possible biohazards possible biohazards

Beware of non-human Beware of non-human primate cells primate cells

Beware of CNS, corneal, Beware of CNS, corneal, pituitary cells pituitary cells

Some cells may be OK at Some cells may be OK at BSL 1BSL 1

Summary

Animals and Allergens

Risk Assessment for Work with Research Animals Risks associated with the research agent used in Risks associated with the research agent used in

the animalthe animal chemical, physical, biologicalchemical, physical, biological

Risks associated with the species of animal usedRisks associated with the species of animal used zoonotic agentszoonotic agents

Risks associated with animal maintenanceRisks associated with animal maintenance ergonomic factors, bites, scratches, allergensergonomic factors, bites, scratches, allergens

Risks Associated with the Agent Used Chemical agentsChemical agents

carcinogens, mutagenscarcinogens, mutagens toxic chemicalstoxic chemicals anestheticsanesthetics

Physical agentsPhysical agents radiationradiation heatheat soundsound

Risks Associated with the Agent Used Potentially biohazardous agentsPotentially biohazardous agents

deliberate use of an infectious agent in deliberate use of an infectious agent in animals for research purposesanimals for research purposes

maintenance of infected animal for maintenance of infected animal for duration of experimentduration of experiment

sacrifice, necropsy and harvesting of sacrifice, necropsy and harvesting of

agent or infected tissueagent or infected tissue

Transmission of Biohazards During Work with Animals AirborneAirborne Release of infectious aerosols by animal by Release of infectious aerosols by animal by

sneezing, coughingsneezing, coughing Release during nasal infection or aerosol Release during nasal infection or aerosol

challengechallenge Aerosolization from bedding and excretaAerosolization from bedding and excreta During surgical proceduresDuring surgical procedures During birthing During birthing

Transmission of Biohazards During Work with Animals

Direct InoculationDirect Inoculation Needlesticks during injection/inoculation Needlesticks during injection/inoculation

processprocess Bites and scratches from infected animalBites and scratches from infected animal

Transmission of Biohazards During Work with AnimalsDirect exposure of mucous membranesDirect exposure of mucous membranes

(by splash or splatter)(by splash or splatter) During surgical proceduresDuring surgical procedures During injectionDuring injection During necropsyDuring necropsy

Transmission of Biohazards During Work with Animals

Indirect transmission and ingestionIndirect transmission and ingestion From contaminated hands or gloves to mouthFrom contaminated hands or gloves to mouth Facial contamination directly from animalFacial contamination directly from animal Transfer of parasites by animal handlingTransfer of parasites by animal handlingIndirect transmission with eye or mucousIndirect transmission with eye or mucousmembrane exposuremembrane exposure Dust from beddingDust from bedding Splash during cage washingSplash during cage washing ““Dirty” environmentDirty” environment

Risk Reduction: Containment of Infectious Agent Containment must include:Containment must include:

Primary containmentPrimary containment Enclosed filtered caging systemEnclosed filtered caging system Biosafety cabinetsBiosafety cabinets Safety equipmentSafety equipment PPEPPE

Secondary containmentSecondary containment The containment facilityThe containment facility

• Negative pressurizationNegative pressurization• Nonrecirculated air supplyNonrecirculated air supply• Ventilation must consider wellbeing of animal Ventilation must consider wellbeing of animal

Containment Caging Systems

No ContainmentNo Containment Open (standard) cageOpen (standard) cage

Some ContainmentSome Containment Filter top cageFilter top cage

(microisolator cage)(microisolator cage) Full ContainmentFull Containment

Fully enclosed in Fully enclosed in

ventilated rackventilated rack


Microisolator CageMicroisolator Cage works like a Petri dishworks like a Petri dish open gaps around lid edge allow limited air open gaps around lid edge allow limited air

exchangeexchange may lead to more labor intensive husbandry due may lead to more labor intensive husbandry due

to moisture and ammonia buildupto moisture and ammonia buildup


Individual cages sealed into rack with Individual cages sealed into rack with supplied air under negative pressuresupplied air under negative pressure

Both supply and exhaust usually HEPA Both supply and exhaust usually HEPA filteredfiltered

Ventilation must control humidity and Ventilation must control humidity and buildup of ammoniabuildup of ammonia


Can install cages in class III biosafety Can install cages in class III biosafety cabinetcabinet

Cages are completely contained with glove Cages are completely contained with glove port accessport access

Very motion-limitingVery motion-limiting Transfer in and out may be an issueTransfer in and out may be an issue


BioBubble (Ft. Collins, CO) makes soft-BioBubble (Ft. Collins, CO) makes soft-wall ventilated enclosureswall ventilated enclosures

Can be containment or barrier styleCan be containment or barrier style Large equipment can be surface-mounted in Large equipment can be surface-mounted in

wallwall

Special Animal Housing Situations

Barrier coloniesBarrier colonies Special breeds - Special breeds - often immunocompromised, often immunocompromised,

“fragile”, expensive (SCID-Hu, nude athymics)“fragile”, expensive (SCID-Hu, nude athymics) Transgenics - Transgenics - often even more fragile and often even more fragile and

expensive (knockouts, microinjected, combos)expensive (knockouts, microinjected, combos) Specific pathogen-free (SPFSpecific pathogen-free (SPF) - bred and raised ) - bred and raised

to be missing certain specific microorganismsto be missing certain specific microorganisms Isolation coloniesIsolation colonies

Extensive SPFsExtensive SPFs and and defined floradefined flora animals animals GnotobiotesGnotobiotes (an entirely different animal!) (an entirely different animal!)

Zoonoses

Zoonotic diseaseZoonotic disease: A disease of animals that : A disease of animals that can be transmitted under natural conditions can be transmitted under natural conditions and cause disease in humansand cause disease in humans Wild caught animals most hazardousWild caught animals most hazardous Random source animals (e.g., from a Random source animals (e.g., from a

pound) are also a riskpound) are also a risk Purpose bred animals pose least riskPurpose bred animals pose least risk

Some Animals and Their Zoonoses AnimalAnimal Macaque monkeysMacaque monkeys SheepSheep White mouseWhite mouse Dogs, cats, skunks, Dogs, cats, skunks,

raccoons, batsraccoons, bats Cattle, NHPCattle, NHP CatsCats Parrots, macawsParrots, macaws ChickensChickens

DiseaseDisease Herpes B virusHerpes B virus Q feverQ fever HantavirusHantavirus RabiesRabies TuberculosisTuberculosis ToxoplasmosisToxoplasmosis PsittacosisPsittacosis Avian influenzaAvian influenza

Rodent Zoonoses

Rat bite fever (Rat bite fever (Streptobacillus moniliformis, Streptobacillus moniliformis, Spirillum minusSpirillum minus) ) transmission: direct contact (bites)transmission: direct contact (bites)

Lymphocytic choriomeningitis (LCM, a virus)Lymphocytic choriomeningitis (LCM, a virus) transmission: inhalationtransmission: inhalation

Leptospirosis (Leptospirosis (LeptospiraLeptospira spp.) spp.) transmission: inhalationtransmission: inhalation

Others include ringworm (fungal), scabies (mites, Others include ringworm (fungal), scabies (mites, an ectoparasite) an ectoparasite)

Transmission of Zoonoses

Enteric route (fecal/oral) Enteric route (fecal/oral) Salmonella, Shigella, Campylobacter, Salmonella, Shigella, Campylobacter,

Giardia, Toxoplasma, Cryptosporidium,Giardia, Toxoplasma, Cryptosporidium,

EntamoebaEntamoeba, Hepatitis A, Hepatitis A Respiratory route Respiratory route

Q fever, Q fever, ChlamydiaChlamydia, Measles, Measles Skin contact Skin contact

Ringworm (Tinea), Measles, MonkeypoxRingworm (Tinea), Measles, Monkeypox

Control of Zoonoses

Get information on Get information on species and agentspecies and agent

Quarantine animals prior Quarantine animals prior to useto use

Use Engineering controlsUse Engineering controls facility construction and facility construction and

secondary barrierssecondary barriers

Consider the need for Consider the need for containment cagingcontainment caging

Use Administrative Use Administrative controlscontrols written SOPs and written SOPs and

manualsmanuals

Use PPEUse PPE additional protection for additional protection for

workerworker

Practice good facility and Practice good facility and personal hygienepersonal hygiene

Provide staff trainingProvide staff training

Laboratory Acquired Allergies (LAA) Significant occupational diseaseSignificant occupational disease Affects >30% of all personnel working with Affects >30% of all personnel working with

animalsanimals No minimum safe exposure levels to allergens No minimum safe exposure levels to allergens

have been establishedhave been established Animal allergens found in hair, dander, urine, Animal allergens found in hair, dander, urine,

saliva, serumsaliva, serum fel-d-fel-d-l cat allergen (in saliva and thus on skin) l cat allergen (in saliva and thus on skin)

is one of the strongest allergens known for is one of the strongest allergens known for humanshumans

Sources of Exposure to LAA

Hair and dander shed Hair and dander shed from animalfrom animal

Urine and feces dried Urine and feces dried in beddingin bedding

Particulates shed from Particulates shed from bedding materialbedding material

Animal salivaAnimal saliva

Routes of Exposure to LAA

Inhalation of airborne allergensInhalation of airborne allergens during cage changingduring cage changing during animal handlingduring animal handling

Skin or eye contactSkin or eye contact usually indirect by touching skin, eyesusually indirect by touching skin, eyes

Percutaneous exposurePercutaneous exposure animal bites (saliva)animal bites (saliva)

Risk Factors for Development of LAA Exposure to allergensExposure to allergens

durationduration frequencyfrequency intensityintensity

Previous allergic conditionsPrevious allergic conditions Other predisposing conditionsOther predisposing conditions

illnessillness ImmunocompromisedImmunocompromised petspets

LAA: Exposure Control

Engineering ControlsEngineering Controls enclosureenclosure dilution ventilationdilution ventilation

Administrative ControlsAdministrative Controls reduce time with animalsreduce time with animals reduce density of animalsreduce density of animals housekeeping practiceshousekeeping practices

Personal Protective EquipmentPersonal Protective Equipment respirators and clothingrespirators and clothing

Medical SurveillanceMedical Surveillance

Disinfection

Why Disinfect?Why Disinfect?

To reduce or eliminate exposure risk To reduce or eliminate exposure risk Biohazard waste disposal Biohazard waste disposal Spill cleanup Spill cleanup Routine surface decontamination Routine surface decontamination

To eliminate contamination risk To eliminate contamination risk Preparation of microbiological media and Preparation of microbiological media and

supplies supplies Preparation of work area for cleanliness-critical Preparation of work area for cleanliness-critical

taskstasks

Resistance to DisinfectantsResistance to Disinfectants

Prions Prions Bacterial spores Bacterial spores Coccidia Coccidia

((CryptosporidiumCryptosporidium) ) Mycobacterium Mycobacterium Nonlipid viruses (Hep A, Nonlipid viruses (Hep A,

Polio) Polio) Fungi Fungi Rickettsiae, Chlamydiae Rickettsiae, Chlamydiae Vegetative bacteria Vegetative bacteria Lipid-containing viruses Lipid-containing viruses

Classes of DisinfectantsClasses of Disinfectants

Chlorine Chlorine Iodine Iodine Alcohol Alcohol Phenolics Phenolics Quaternary Ammonium Quaternary Ammonium Glutaraldehyde Glutaraldehyde Hydrogen peroxideHydrogen peroxide

Factors Influencing EfficacyFactors Influencing Efficacy

Surface/TopographySurface/Topography – uneven, cracked or pitted – uneven, cracked or pitted surfaces especially wooden surfaces can hide surfaces especially wooden surfaces can hide microorganisms and are difficult to disinfect microorganisms and are difficult to disinfect

Temperature - Temperature - elevated temperatures may elevated temperatures may enhance germicidal action but also evaporation rate enhance germicidal action but also evaporation rate

Relative Humidity – Relative Humidity – many disinfectants have many disinfectants have optimal relative humidity range for maximum optimal relative humidity range for maximum effectiveness effectiveness

Water Hardness – Water Hardness – some disinfectants may be less some disinfectants may be less effective when diluted in hard watereffective when diluted in hard water

Organic Load Organic Load Blood, sputum, milk, bedding, feed, Blood, sputum, milk, bedding, feed,

manure manure Proteins physically protect and stabilize Proteins physically protect and stabilize

many microorganisms many microorganisms Adverse effect on action of many Adverse effect on action of many

disinfectants disinfectants

Concentration Concentration In most cases, the higher the In most cases, the higher the

concentration, the more rapid the kill concentration, the more rapid the kill Consider potential damage to surfaces or Consider potential damage to surfaces or

tissues tissues Reducing concentration to avoid Reducing concentration to avoid

damage will require additional contact damage will require additional contact time time

Ultimately, disinfectant will no longer Ultimately, disinfectant will no longer be active enough to be useful be active enough to be useful

Contact Time Contact Time Disinfectants should be effective with a Disinfectants should be effective with a

short contact time short contact time Manufacturer’s recommended contact Manufacturer’s recommended contact

time may be unrealistic under in-use time may be unrealistic under in-use condition condition

Contact time may depend on the method Contact time may depend on the method of applicationof application

For surface applications, loss by For surface applications, loss by evaporation may require frequent evaporation may require frequent applications to achieve contact timeapplications to achieve contact time

Some Other FactorsSome Other Factors Dirt, grease and oils – all can protect the organism Dirt, grease and oils – all can protect the organism

and will repel water based disinfectants and will repel water based disinfectants Types of microbes present – spores, vegetative Types of microbes present – spores, vegetative

cells, viruses cells, viruses Dried spills (from media, buffers) can protect Dried spills (from media, buffers) can protect

microorganisms from contact with the disinfectant microorganisms from contact with the disinfectant pH pH Age of the product/solution Age of the product/solution Method of application (spray vs. wipe)Method of application (spray vs. wipe) Rate of application Rate of application Storage conditionStorage condition

Medical Waste Disposal(Biohazardous and Sharps Wastes)

Who Regulates Medical Waste?Who Regulates Medical Waste?

Federal Federal EPA (40 CFR part 60.51c) EPA (40 CFR part 60.51c) DOT (49 CFR Part 173.134) DOT (49 CFR Part 173.134) OSHA (29 CFR Part 1920.1030b) OSHA (29 CFR Part 1920.1030b) USPS (39 CFR 111.1) USPS (39 CFR 111.1)

State CA Health and Safety Code State CA Health and Safety Code Local SD Code of Regulatory Ordinance Local SD Code of Regulatory Ordinance

Medical WasteMedical Waste

……biohazardous waste and/or sharps waste biohazardous waste and/or sharps waste that is produced or generated as a result of that is produced or generated as a result of diagnosis, treatment or immunization of diagnosis, treatment or immunization of human beings or animals; research human beings or animals; research pertaining thereto; production or testing of pertaining thereto; production or testing of biologicals or removal of regulated waste biologicals or removal of regulated waste from a trauma scene.from a trauma scene.

Forms of Medical WasteForms of Medical Waste Solid Solid

Labware (flasks, tubes, plates, bottle, vials) Labware (flasks, tubes, plates, bottle, vials) Pipettes (could also be sharps) Pipettes (could also be sharps) Lab waste (stocks, specimens, cultures, swabs, Lab waste (stocks, specimens, cultures, swabs,

vaccines) vaccines) Gloves, apparel, wipes Gloves, apparel, wipes

Liquid Liquid Aspirates, culture fluids, rinses, washes Aspirates, culture fluids, rinses, washes Sera, body fluids Sera, body fluids

Sharps Sharps Anything with a point or edge capable of Anything with a point or edge capable of

piercing or cuttingpiercing or cutting

Medical Waste Does Not Include:Medical Waste Does Not Include:

Waste generated in food processing Waste generated in food processing Urine, feces, saliva, sputum, nasal secretions, Urine, feces, saliva, sputum, nasal secretions,

sweat, tears, or vomitus, unless it contains fluid sweat, tears, or vomitus, unless it contains fluid blood blood

Medical solid wastes i.e., paper towels or empty Medical solid wastes i.e., paper towels or empty specimen containers that are not biohazardous, specimen containers that are not biohazardous, bandages/dressings containing dried blood bandages/dressings containing dried blood

Hazardous waste, radioactive waste, household Hazardous waste, radioactive waste, household waste waste

Waste generated on agricultural or livestock Waste generated on agricultural or livestock practices on a farm or ranchpractices on a farm or ranch

Biohazardous WasteBiohazardous Waste Laboratory Wastes Laboratory Wastes

Human or animal Human or animal specimen cultures specimen cultures

Cultures and stocks of Cultures and stocks of infectious agents infectious agents

Wastes from production of Wastes from production of biologicals, live and biologicals, live and attenuated vaccines, attenuated vaccines, culture dishes and devices culture dishes and devices

Pharmaceuticals Pharmaceuticals Human surgery specimens or Human surgery specimens or

tissues, including those fixed tissues, including those fixed in fixatives in fixatives

Animal parts, tissues, Animal parts, tissues, fluids, or carcasses fluids, or carcasses

Wastes containing Wastes containing recognizable fluid blood recognizable fluid blood or blood products, or blood products, containers with fluid containers with fluid blood, blood from animals blood, blood from animals

Wastes contaminated or Wastes contaminated or containing containing chemotherapeutic agents chemotherapeutic agents

Sharps WasteSharps Waste

… … means any device having acute rigid corners, means any device having acute rigid corners, edges, or protuberances capable of cutting or edges, or protuberances capable of cutting or piercing. piercing.

Needles, needles with syringes, contaminated Needles, needles with syringes, contaminated syringes, blades, needles with attached tubing syringes, blades, needles with attached tubing

Broken glass items i.e., Pasteur pipettes, blood Broken glass items i.e., Pasteur pipettes, blood vials contaminated with biohazardous waste vials contaminated with biohazardous waste

Mixed WasteMixed Waste

… … means mixtures of medical and nonmedical means mixtures of medical and nonmedical waste. Mixed waste is medical waste, except for waste. Mixed waste is medical waste, except for all of the following: all of the following:

““Mixed Waste Hierarchy”Mixed Waste Hierarchy”

Biohazard BagBiohazard Bag

……means a disposable red bag that is means a disposable red bag that is impervious to moisture and has a strength impervious to moisture and has a strength sufficient to preclude ripping, tearing, or sufficient to preclude ripping, tearing, or bursting under normal conditions of usage bursting under normal conditions of usage and handling of the waste-filled bag.and handling of the waste-filled bag.

Containment and StorageBiohazardous WasteBiohazardous Waste Must be segregated from other types of wastes Must be segregated from other types of wastes Must be contained in “biohazard bags” Must be contained in “biohazard bags”

Bags must be red.Bags must be red. Bags must be labeled either with the word Bags must be labeled either with the word

“Biohazardous” or with the biohazard symbol “Biohazardous” or with the biohazard symbol and the word “Biohazard”. and the word “Biohazard”.

Bags must also be labeled with the generator’s Bags must also be labeled with the generator’s name, address and phone number. name, address and phone number.

Bags must be securely tied to prevent leakage Bags must be securely tied to prevent leakage or expulsion of contents.or expulsion of contents.

Bags must be placed in a rigid container for Bags must be placed in a rigid container for storage, handling and transport. storage, handling and transport.

ContainersContainers Containers shall be leak resistant, have tight-fitting Containers shall be leak resistant, have tight-fitting

covers, kept clean and in good repair. covers, kept clean and in good repair. Containers may be of any color and shall be labeled Containers may be of any color and shall be labeled

with the word “Biohazardous” or with the biohazard with the word “Biohazardous” or with the biohazard symbol and the word “Biohazard” on the lid and on the symbol and the word “Biohazard” on the lid and on the sides so as to be visible on any lateral direction.sides so as to be visible on any lateral direction.

Reusable containers shall be washed and decontaminated Reusable containers shall be washed and decontaminated unless protected from contamination by disposable liners unless protected from contamination by disposable liners or bags. or bags.

Reusable containers shall be washed to remove visible soil Reusable containers shall be washed to remove visible soil and decontaminated by: and decontaminated by: Exposure to hot water (180°F) for 15 secs. Exposure to hot water (180°F) for 15 secs. Exposure to the following sanitizer for 3 mins. Exposure to the following sanitizer for 3 mins.

Hypochlorite soln. (500 ppm avail. Chlorine) Hypochlorite soln. (500 ppm avail. Chlorine) Phenolic soln. (500 ppm active agent) Phenolic soln. (500 ppm active agent) Iodoform soln. (100 ppm avail. Iodine) Iodoform soln. (100 ppm avail. Iodine) Quaternary ammonium soln. (400 ppm active agent) Quaternary ammonium soln. (400 ppm active agent)

Sharps ContainerSharps Container

……means a rigid puncture resistant container means a rigid puncture resistant container that, when sealed, is leak resistant and that, when sealed, is leak resistant and cannot be reopened without great difficulty.cannot be reopened without great difficulty.

Containment and StorageContainment and StorageSharps WasteSharps Waste Must be segregated from other types of wastes. Must be segregated from other types of wastes. Must be contained in “sharps container”. Must be contained in “sharps container”. Tightly close or tape closed the lid of a full sharps Tightly close or tape closed the lid of a full sharps

container ready for disposal. container ready for disposal. Store sharps container ready for disposal for not Store sharps container ready for disposal for not

more than 7 days. more than 7 days. Label sharps container with the word “Sharps Label sharps container with the word “Sharps

Waste” or the biohazard symbol and the word Waste” or the biohazard symbol and the word “Biohazard”. “Biohazard”.

Must also be labeled with the generator’s name, Must also be labeled with the generator’s name, address and phone number that is legible and address and phone number that is legible and easily visible on the outside of the container.easily visible on the outside of the container.

DisposalDisposal Take biohazard bag and sharps container to Take biohazard bag and sharps container to

designated Accumulation Sites: designated Accumulation Sites: Life Science, Room 14 Life Science, Room 14 Student Health Service, outside shed Student Health Service, outside shed Generators at other locations may call EHS at Generators at other locations may call EHS at

(619) 594-6778 for biohazard waste pick-ups (619) 594-6778 for biohazard waste pick-ups A biowaste vendor will collect biohazard bag and A biowaste vendor will collect biohazard bag and

sharps container for disposal on a weekly basis sharps container for disposal on a weekly basis from Life Science and Student Health Services. from Life Science and Student Health Services.

Biohazardous and sharps wastes will be Biohazardous and sharps wastes will be autoclaved while animal carcasses will be autoclaved while animal carcasses will be incinerated.incinerated.

Solid Medical Waste CollectionNot acceptable in CAMust be rigid, puncture-proof, leak-proof

Labels have to be affixed on all 4 sides of the container.

Sharps Waste Collection

Sharps containers <7 gal. should not be on the floor. Lids have to be difficult to open. Labels have to be affixed on all 4 sides of the container.

What’s Wrong with these Pictures?

Left: Sharps sticking out of Sharps Waste container.Right: Sharps Waste container past full line. No generator label.


Left: Bottle not labeled.Right: Cardboard box is not allowed for liquid waste. No labels. No lid.


Left and Right: Cardboard box is not an appropriate Sharps Waste container. No labels. No lids.


Left: Red bag should be inside the secondary container. Cardboard box is not an acceptable secondary container. Right: Bag must be red. Secondary container does not have to be red. No biohazard label. Red bag on floor ready for disposal must be transported to the accumulation site immediately.


Left: Do not fill red bags completely. Replace more often.Right: No biohazard label. Red bag on floor ready for disposal must be transported to the accumulation site immediately.


Left: Do not deface container. Incorrect label placed on container (need generator label).Right: Red bag must be transported in a secure secondary container to the accumulation site. Red bag must have biohazard label and generator label.


Left: Proper Sharps Waste container not used. No generator label.Right: Generator label should be on the outside of the red bag. Secondary container needs biohazard label on all visible sides including top. Use appropriately sized red bag for secondary container.


Left: Incorrect label placed on container (need generator label). Keep lid closed when not in use.Right: No lid. Use appropriately sized red bag for secondary container. Secondary container needs biohazard label on all visible sides including top.

Containment Equipment and Facilities

BiocontainmentBiocontainment

The principle of holding or being capable of The principle of holding or being capable of holding or including within a fixed limit or holding or including within a fixed limit or area area

Preventing the unintentional release of Preventing the unintentional release of biological agents through a combination of biological agents through a combination of laboratory practices, containment laboratory practices, containment equipment (primary barrier) and laboratory equipment (primary barrier) and laboratory facility design (secondary barrier)facility design (secondary barrier)

Primary Barrier

Primary barriers contain the agent at the Primary barriers contain the agent at the sourcesource

Equipment/Engineering ControlEquipment/Engineering Control Biological safety cabinet, fumehood, Biological safety cabinet, fumehood,

glove box, animal housing, centrifuge, glove box, animal housing, centrifuge, fermenterfermenter

Secondary Barrier

Secondary barrier is the structure Secondary barrier is the structure surrounding the primary barriersurrounding the primary barrier

Facility/Engineering ControlFacility/Engineering Control Rooms, buildingRooms, building

Types of FacilitiesTypes of Facilities Basic laboratoryBasic laboratory Containment laboratoryContainment laboratory

Primary Barriers - Equipment Personnel ProtectionPersonnel Protection

Any aerosol generated within the cabinet is Any aerosol generated within the cabinet is contained and kept away from the researchercontained and kept away from the researcher

Product ProtectionProduct Protection Air within the work space of the cabinet has Air within the work space of the cabinet has

been filtered so that is is virtually free of been filtered so that is is virtually free of airborne particles and organisms; thus airborne particles and organisms; thus protecting the work from outside contaminationprotecting the work from outside contamination

Environmental ProtectionEnvironmental Protection Aerosols generated within the unit are removed Aerosols generated within the unit are removed

from the air before the air is dischargedfrom the air before the air is discharged

Ventilation EquipmentClasses and Types

Chemical Fume Hood

100 fpm face velocity 100 fpm face velocity Offer only personnel protection Offer only personnel protection Always exhaust air to the outside Always exhaust air to the outside Do not offer protection to the product or the environment, Do not offer protection to the product or the environment,

as there is no filtration of intake and exhaust air as there is no filtration of intake and exhaust air (Sometimes air cleaning treatment is added to the exhaust.) (Sometimes air cleaning treatment is added to the exhaust.)

Do draw contaminants in the laboratory air directly over Do draw contaminants in the laboratory air directly over the product being worked on the product being worked on

Used for work with chemical hazards Used for work with chemical hazards

Any Comments?

Fumehood - keep hood clean, sash should be closed when hood is not in use, equipment should be 9” from sash

Clean Bench / Laminar Flow Hoods

Provide product protection only Provide product protection only Product protection is provided by creating a unidirectional Product protection is provided by creating a unidirectional

airflow generated through a HEPA filter airflow generated through a HEPA filter Discharge air goes directly into workroom Discharge air goes directly into workroom Applications Applications

– – Any application where the product is not hazardous but must Any application where the product is not hazardous but must be kept contaminant free be kept contaminant free – – Preparation of non-hazardous intravenous mixtures and media Preparation of non-hazardous intravenous mixtures and media – – Particulate free assembly of sterile equipment and electronic Particulate free assembly of sterile equipment and electronic devices devices

Eliminate Clean Bench in containment laboratory Eliminate Clean Bench in containment laboratory

Biological Safety Cabinets Designed to contain biological hazards Designed to contain biological hazards Inward airflow for personnel protection Inward airflow for personnel protection HEPA filtered exhaust air for environmental protection HEPA filtered exhaust air for environmental protection Supply air HEPA filter for product protection (except Supply air HEPA filter for product protection (except

Class I) Class I) Separated into Classes and Types Separated into Classes and Types

– – Class I Class I

– – Class II Class II

• • Type A1, A2 Type A1, A2

• • Type B1, B2 Type B1, B2

– – Class III Class III Microbiological studies, cell cultures, pharmaceutical Microbiological studies, cell cultures, pharmaceutical

research and procedures… research and procedures…

Class I Cabinet

75 fpm face velocity 75 fpm face velocity Provides personnel and environmental protection Provides personnel and environmental protection No product protection No product protection Requires an exhaust blower to pull the air through Requires an exhaust blower to pull the air through

- usually to the outdoors - usually to the outdoors Applications Applications

– – Housing centrifuges, fermenters Housing centrifuges, fermenters – – Cage dumping in an animal lab Cage dumping in an animal lab – – Aerating culturesAerating cultures

Class II Cabinets Ventilated cabinet Ventilated cabinet Provides personnel, product, and environmental Provides personnel, product, and environmental

protection protection Open front with inward airflow for personnel Open front with inward airflow for personnel

protection protection Downward HEPA filtered laminar airflow for Downward HEPA filtered laminar airflow for

product protection product protection HEPA filtered exhaust air for environmental HEPA filtered exhaust air for environmental

protection protection

Any Comments?

BSC - remove unnecessary objects, keep grill at front of cabinet unobstructed

Primary Barriers

PersonnelPersonnel ProductProduct EnvironmentEnvironment

Chemical FumehoodChemical Fumehood xx

Laminar FlowhoodLaminar Flowhood xx

Class I Biosafety CabinetClass I Biosafety Cabinet xx xx

Class II Biosafety CabinetClass II Biosafety Cabinet xx xx xx

Class III Biosafety CabinetClass III Biosafety Cabinet xx xx xx

IsolatorsIsolators xx xx xx

Types of Biosafety CabinetsNSF/ANSI Standard 49 – 2002

Type Face velocity (lfpm)

Airflow Pattern Radionuclides/ Toxic Chemicals

Bio-safety Level(s)

Product Protection

Class I 75 In at front; rear and top through HEPA filter

No 2, 3 No

Class II Type A1

7570% recirculated through HEPA; exhaust through HEPA

No 2, 3 Yes

Class II Type A2

10030% recirculated through HEPA; exhaust via HEPA and hard ducted

Yes (Low levels/volatility)

2, 3 Yes

Class II Type B1

100No recirculation; total exhaust via HEPA and hard ducted

Yes 2, 3 Yes

Class II Type B2

100Same as B1, but plena under negative pressure to room and exhaust air is ducted

Yes 2, 3 Yes

Class III NASupply air inlets and exhaust through 2 HEPA filters

Yes 3, 4 Yes

Biological Safety Cabinet Certification

First CertificationFirst Certification AnnuallyAnnually When movedWhen moved When filter is changedWhen filter is changed When repaired or modifiedWhen repaired or modified

Note: Certification is paid by the researcher, not EHS Note: Certification is paid by the researcher, not EHS

Other Primary Barriers- Engineering Control Gasketed blenders, homogenizersGasketed blenders, homogenizers Cotton plugs, filters for flasks in shakersCotton plugs, filters for flasks in shakers Filtered pipette tipsFiltered pipette tips HEPA and hydrophobic vacuum line filtersHEPA and hydrophobic vacuum line filters Plasticware substituted for glasswarePlasticware substituted for glassware Gas burners with shield, microincineratorGas burners with shield, microincinerator CentrifugesCentrifuges

Interlock, solid cover, safety buckets, O-ringsInterlock, solid cover, safety buckets, O-rings

Secondary Barrier- Facilities

Laboratory Biosafety Level 2Laboratory Biosafety Level 2

Lockable doors (a must for restricted agents)Lockable doors (a must for restricted agents) SinkSink Bench tops impervious and easily cleanedBench tops impervious and easily cleaned Biological safety cabinet (if applicable)Biological safety cabinet (if applicable) EyewashEyewash Inward airflow (desirable)Inward airflow (desirable)

Biosafety Practices and Procedures

Hierarchy of Controls

Administrative ControlAdministrative Control Engineering ControlEngineering Control Work PracticesWork Practices Personal Protective Clothing or Equipment Personal Protective Clothing or Equipment

Administrative Controls SubstitutionSubstitution Authorization/ApprovalAuthorization/Approval Written biosafety procedures required for the Written biosafety procedures required for the

experimental procedures and equipment including experimental procedures and equipment including inventory of biological agents or materialsinventory of biological agents or materials

Laboratory personnel biosafety trainingLaboratory personnel biosafety training Medical Surveillance (BSL 2 and above)Medical Surveillance (BSL 2 and above)

Health historyHealth history Medical screeningMedical screening ImmunizationImmunization Serum storageSerum storage Post-exposure prophylaxisPost-exposure prophylaxis

Engineering Controls

Biological safety cabinets, glove boxesBiological safety cabinets, glove boxes Animal containment caging systemsAnimal containment caging systems Safety equipment (filtered or sealed Safety equipment (filtered or sealed

equipment)equipment) Ventilation systemVentilation system Containment facilitiesContainment facilities

Personal Protective Clothing and Equipment

Provides barrier against skin, mucous Provides barrier against skin, mucous membrane or respiratory exposure to membrane or respiratory exposure to infectious agents during proceduresinfectious agents during procedures

Prevent spread of contaminationPrevent spread of contamination Does not eliminate the hazardDoes not eliminate the hazard Integrity wanes with use (i.e., change Integrity wanes with use (i.e., change

gloves frequently)gloves frequently)

BSL 1: Work Practices and Procedures ApplicationsApplications

Non-infectious agent and tissue culture, media Non-infectious agent and tissue culture, media preparationpreparation

Prevent Cross ContaminationPrevent Cross Contamination Keep cultures coveredKeep cultures covered Flame instruments and containersFlame instruments and containers Use sterile media and equipmentUse sterile media and equipment Keep hands or face away from culturesKeep hands or face away from cultures

Biosafety ProceduresBiosafety Procedures Work with agents may be conducted on open Work with agents may be conducted on open

benchbench Wash hands oftenWash hands often No mouth pipettingNo mouth pipetting No eating or drinking in labNo eating or drinking in lab Minimize aerosol generationMinimize aerosol generation Decontaminate work surfacesDecontaminate work surfaces Wear applicable PPEWear applicable PPE

BSL 1: Work Practices and BSL 1: Work Practices and ProceduresProcedures

BSL 2: Work Practices and Procedures Increasing emphasis on safety Increasing emphasis on safety

procedures and practicesprocedures and practices Increasing need for staff trainingIncreasing need for staff training Increasing need for competent Increasing need for competent

supervisionsupervision Biohazard sign posted at entry doorBiohazard sign posted at entry door Biohazard labels affixed on regulated Biohazard labels affixed on regulated

waste containerswaste containers Use of personal protective equipment Use of personal protective equipment

as a barrier to exposure: lab coat, as a barrier to exposure: lab coat, gloves, eye and face protectiongloves, eye and face protection

Some work on open bench allowedSome work on open bench allowed

BSL 2: Work Practices and Procedures Aerosol generating procedures Aerosol generating procedures

performed in a biosafety cabinet:performed in a biosafety cabinet: HomogenizingHomogenizing VortexingVortexing Vigorous mixingVigorous mixing Pipetting infectious liquidsPipetting infectious liquids SonicationSonication PouringPouring

If breach occurs:If breach occurs: Evacuate lab, post spill signEvacuate lab, post spill sign With appropriate PPE and With appropriate PPE and

disinfectant, decontaminate disinfectant, decontaminate centrifuge, buckets, other centrifuge, buckets, other items or areasitems or areas

Correct Use of Biosafety Cabinets Start UpStart Up

Turn off ultraviolet light (if so Turn off ultraviolet light (if so equipped) as soon as you enter the equipped) as soon as you enter the room. room.

Turn on all blowers and BSC Turn on all blowers and BSC illumination lights. illumination lights.

Allow five minutes of operation to Allow five minutes of operation to purge system; check flow alarm purge system; check flow alarm system audio and visual alarm system audio and visual alarm function (if so equipped).function (if so equipped).

Decontaminate readily accessible Decontaminate readily accessible interior surfaces and items with a interior surfaces and items with a disinfectant (appropriate for the disinfectant (appropriate for the agents or suspected agents present) agents or suspected agents present) before loading and wait at least 10 before loading and wait at least 10 minutes prior to start of work.minutes prior to start of work.

Correct Use of Biosafety Cabinets During UseDuring Use

Load supplies prior to Load supplies prior to work. work.

Do not overload cabinet. Do not overload cabinet. Separate clean and dirty Separate clean and dirty

side.side. Work in center of work Work in center of work

area. area. Do not block front or rear Do not block front or rear

grills.grills. Minimize disruption of Minimize disruption of

airflow (turbulence).airflow (turbulence). Clean up spill promptly.Clean up spill promptly. Discard waste within the Discard waste within the

cabinet.cabinet.

Correct Use of Biosafety Cabinets Shut DownShut Down

Decontaminate and remove all Decontaminate and remove all items from interior work area. items from interior work area.

Decontaminate readily Decontaminate readily accessible interior surfaces with accessible interior surfaces with a disinfectant appropriate for a disinfectant appropriate for the agents or suspected agents the agents or suspected agents present.present.

Turn on ultraviolet light (if so Turn on ultraviolet light (if so equipped). equipped).

Allow five minutes of Allow five minutes of operation to purge system. operation to purge system. Then wait at least 10 Then wait at least 10 minutes.minutes.

Turn off BSC blower.Turn off BSC blower.

Correct Use of Biosafety Cabinets Moving/InstallationMoving/Installation

BSCs must be decontaminated prior to moving. In order to BSCs must be decontaminated prior to moving. In order to ensure filter integrity, the equipment must be recertified after the ensure filter integrity, the equipment must be recertified after the BSC is installed at its final new location. Arrangements need to BSC is installed at its final new location. Arrangements need to be made well in advance in order for contractors to meet your be made well in advance in order for contractors to meet your schedule. The PI is responsible for contacting the contractor or to schedule. The PI is responsible for contacting the contractor or to schedule this work.schedule this work.

DecontaminationDecontaminationDecontamination is usually performed by certified professionals. Decontamination is usually performed by certified professionals.

CertificationCertificationAll BSCs that are used for handling biohazardous materials must All BSCs that are used for handling biohazardous materials must be recertified annually. SDSU has contracted with a specific be recertified annually. SDSU has contracted with a specific contractor to provide a consistent level of certification and contractor to provide a consistent level of certification and maintenance service. Contact EHS at (619) 594-6778 to obtain maintenance service. Contact EHS at (619) 594-6778 to obtain contractor information.contractor information.

DO NOT use Bunsen burners or open flamesDO NOT use Bunsen burners or open flames Fire hazardFire hazard Can damage HEPA filterCan damage HEPA filter Interferes with proper air flowInterferes with proper air flow Microincinerator preferredMicroincinerator preferred Burner with pilot light not a good alternative Burner with pilot light not a good alternative Open flames react with disinfectants (flammables)Open flames react with disinfectants (flammables)

Correct Use of Biosafety Cabinets – Open Flames

Use sealed tubes, rotors, and safety cups/buckets that are sealed with Use sealed tubes, rotors, and safety cups/buckets that are sealed with O-ringsO-rings

Inspect tubes, O-rings and rotors for wear, and buckets for cracks, Inspect tubes, O-rings and rotors for wear, and buckets for cracks, chips, erosion, etc.chips, erosion, etc.

Do not use aluminum foil to cap centrifuge tubesDo not use aluminum foil to cap centrifuge tubes Clean and maintain gaskets and O-ringsClean and maintain gaskets and O-rings Change O-rings if compromisedChange O-rings if compromised Load/unload centrifuge tubes, rotors and accessories in BSCLoad/unload centrifuge tubes, rotors and accessories in BSC Small, low speed centrifuges may be placed in a BSC; however, high Small, low speed centrifuges may be placed in a BSC; however, high

speed centrifuges pose additional hazards speed centrifuges pose additional hazards Do not overfill tubesDo not overfill tubes Balance buckets, tubes and rotors properly before centrifugationBalance buckets, tubes and rotors properly before centrifugation Wait 5 minutes (or 30 mins. for high speed centrifuge) after each run Wait 5 minutes (or 30 mins. for high speed centrifuge) after each run

before openingbefore opening Do not decant or pour off supernatant. Use a vacuum system with Do not decant or pour off supernatant. Use a vacuum system with

appropriate in-line reservoirs and filtersappropriate in-line reservoirs and filters

Safe Use of Centrifuge

Avoid use of glass blender jars, unless covered Avoid use of glass blender jars, unless covered with polypropylene jarwith polypropylene jar

Place disinfectant-moistened towel over the top of Place disinfectant-moistened towel over the top of the blender during usethe blender during use

Before opening the blender jar, allow the unit to Before opening the blender jar, allow the unit to rest at least 1 minute for aerosols to settle and then rest at least 1 minute for aerosols to settle and then open in a BSCopen in a BSC

Decontaminate promptly after useDecontaminate promptly after use

Safe Use of Blenders

Use careful pipetting practicesUse careful pipetting practices Avoid drops onto hard surfacesAvoid drops onto hard surfaces Wipe up spills promptly with appropriate Wipe up spills promptly with appropriate

disinfectantdisinfectant For ejection of liquid from micropipetteFor ejection of liquid from micropipette

No blowoutNo blowout No pressure ejectionNo pressure ejection Use wall contactUse wall contact

Use capped tubes when mixing, blending, or Use capped tubes when mixing, blending, or vortexingvortexing

Pour liquids carefullyPour liquids carefully Avoid bubblesAvoid bubbles

Minimizing Aerosols

Careful Pipetting Techniques Never blow out last drop in Never blow out last drop in

pipettepipette Use pipetting aids with filtersUse pipetting aids with filters Never mix by suction and Never mix by suction and

expulsion (mix by sonication)expulsion (mix by sonication) Discharge liquid down side Discharge liquid down side

of container, using tip-to-wall of container, using tip-to-wall contactcontact

Deliver as close as possible Deliver as close as possible to contentsto contents

Work over plastic-backed Work over plastic-backed absorbent matting (ensure it absorbent matting (ensure it doesn’t slide forward or doesn’t slide forward or backward blocking air grill)backward blocking air grill)

Use Extreme Care with Sharps

Use sharps if only absolutely Use sharps if only absolutely required as part of a processrequired as part of a process

Percutaneous exposure riskPercutaneous exposure risk Employ safe work practicesEmploy safe work practices Utilize safe sharp devicesUtilize safe sharp devices

Aerosol exposure riskAerosol exposure risk Use biosafety cabinet for Use biosafety cabinet for

removal of air from needleremoval of air from needle Use mechanical methods for Use mechanical methods for

needle removalneedle removal Never bend, recap or manipulate Never bend, recap or manipulate

sharps by handsharps by hand Keep hands away from needleKeep hands away from needle

Vacuum System Protection

In-line filter and disinfectant in collection and In-line filter and disinfectant in collection and overflow flasksoverflow flasks

Signs and Labels

Biohazard Signs

Completely filled out biohazard sign must be Completely filled out biohazard sign must be displayed at entrances to laboratories where displayed at entrances to laboratories where biohazards are present and tissue culture roomsbiohazards are present and tissue culture rooms

Features fluorescent orange-red sign with lettering Features fluorescent orange-red sign with lettering “biohazard” and the international biohazard symbol “biohazard” and the international biohazard symbol in contrasting color, name of biohazardous agent or in contrasting color, name of biohazardous agent or material, special entry requirement and PI and BSO material, special entry requirement and PI and BSO contact numbercontact number

Available free of charge from EHS, Available free of charge from EHS, call (619) 594-2865call (619) 594-2865

Biohazard Label

Must be attached to containers of biohazards or Must be attached to containers of biohazards or biohazardous waste containers.biohazardous waste containers.

Features fluorescent orange-red label with lettering Features fluorescent orange-red label with lettering “biohazard” and the international biohazard symbol in “biohazard” and the international biohazard symbol in contrasting color.contrasting color.

Red bags and sharps containers must also have the Red bags and sharps containers must also have the generator address labelgenerator address label affixed prior to use: affixed prior to use:

San Diego State UniversitySan Diego State University5500 Campanile Drive5500 Campanile DriveSan Diego, CA 92182San Diego, CA 92182(619) 594-6778(619) 594-6778

Laboratory Moves

It is important that the laboratory is safe:It is important that the laboratory is safe: For custodians to cleanFor custodians to clean For contractors to work inFor contractors to work in For the next group of laboratory personnel For the next group of laboratory personnel

to occupyto occupy

Disposal and Decontamination of Biohazardous WasteIn preparation for moving, observe the following In preparation for moving, observe the following

guidelines:guidelines: Label ALL biohazardous waste red bags and Label ALL biohazardous waste red bags and

sharps containersharps container Dispose of all biohazardous waste red bags and Dispose of all biohazardous waste red bags and

sharps containers at approved accumulation sitessharps containers at approved accumulation sites EHS will not pick-up biohazardous waste except EHS will not pick-up biohazardous waste except

in unusual situationsin unusual situations Chemically disinfect biohazardous wasteChemically disinfect biohazardous waste Autoclave liquid biohazardous waste and dispose Autoclave liquid biohazardous waste and dispose

of down the drainof down the drain

Moving the Biosafety Cabinet

Disinfect all BSC work surfaces prior to Disinfect all BSC work surfaces prior to moving the BSC to a new facility. BSCs moving the BSC to a new facility. BSCs used for work with pathogenic organisms used for work with pathogenic organisms may require paraformaldehyde may require paraformaldehyde decontamination before being moved.decontamination before being moved.

Each BSC must be recertified for correct Each BSC must be recertified for correct airflow and filter integrity after it has been airflow and filter integrity after it has been moved and placed in its final location.moved and placed in its final location.

Moving or Disposing of Refrigeration Units

Clear all materials stored inside the Clear all materials stored inside the refrigeration units.refrigeration units.

Disinfect all refrigeration units prior to Disinfect all refrigeration units prior to removal or disposal.removal or disposal.

Obtain clearance notification from EHS Obtain clearance notification from EHS prior to removal or disposal of refrigeration prior to removal or disposal of refrigeration units.units.

Shipment and Transportation

To Whom are the Shipping and Transportation Regulations Applicable?

Under IATA, DGR apply to anyone who handles, Under IATA, DGR apply to anyone who handles, offers for transport, transports dangerous goods or offers for transport, transports dangerous goods or causes dangerous goods to be transported.causes dangerous goods to be transported.

Under DOT, HMR apply to each person who Under DOT, HMR apply to each person who performs, or causes to be performed, functions performs, or causes to be performed, functions related to the transportation of hazardous materials related to the transportation of hazardous materials such as determination of, and compliance with, such as determination of, and compliance with, basic conditions for offering; filling packages; basic conditions for offering; filling packages; marking and labeling packages; preparing marking and labeling packages; preparing shipping papers; handling, loading, securing and shipping papers; handling, loading, securing and segregating packages within a transport vehicle, segregating packages within a transport vehicle, freight or cargo hold; and transporting hazardous freight or cargo hold; and transporting hazardous materials.materials.

Emergency Response to Biological Incidents

Response to Biological Spills in Response to Biological Spills in the Laboratorythe Laboratory

(Intentional or Accidental)(Intentional or Accidental)

Exposure Management

For splash to eyes, mucous membranes, or For splash to eyes, mucous membranes, or broken area of the skinbroken area of the skin

Irrigate eyes with clean water, saline or Irrigate eyes with clean water, saline or sterile irrigantssterile irrigants

Flush splashes to mouth, nose, and broken Flush splashes to mouth, nose, and broken area of skin with waterarea of skin with water

Exposure Management

For needlesticks or cuts with human blood, fluids, For needlesticks or cuts with human blood, fluids, infectious agents or antibiotic resistant organisminfectious agents or antibiotic resistant organism

Flush needlesticks and cuts with soap and waterFlush needlesticks and cuts with soap and water Get medical evaluation ASAPGet medical evaluation ASAP Inform PI, BSO and health professional (required Inform PI, BSO and health professional (required

mandatory reporting of incident)mandatory reporting of incident) Public Health Service has recommendations for Public Health Service has recommendations for

post-exposure follow-uppost-exposure follow-up

Spill Clean-Up

You can clean-up a biological spill if:You can clean-up a biological spill if: You are aware of the hazards and clean-up You are aware of the hazards and clean-up

procedures (training required)procedures (training required) There is no potential for personal or There is no potential for personal or

environmental damageenvironmental damage The appropriate spill clean-up equipment is The appropriate spill clean-up equipment is

availableavailable One or two people can clean-up the spill One or two people can clean-up the spill

thoroughly in less than an hourthoroughly in less than an hourNote: Spill incident still needs to be reported to Note: Spill incident still needs to be reported to

BSO/EHS. If spill is in gallons or liters, call BSO/EHS. If spill is in gallons or liters, call BSO/EHS.BSO/EHS.

Biological Spill Clean-Up Kit- Basic Nitrile gloves (double gloving), splash Nitrile gloves (double gloving), splash

goggles, shoe coversgoggles, shoe covers Small disposable broom with dustpan, tongs Small disposable broom with dustpan, tongs

or forceps (for picking up sharps)or forceps (for picking up sharps) Paper towels or other absorbent in the lab Paper towels or other absorbent in the lab Sharps container and/or biohazard waste Sharps container and/or biohazard waste

bagsbags Disinfectant agent suitable for the agents in Disinfectant agent suitable for the agents in

the labthe lab

Spill Clean-Up for BSL 1-2If there is a spill inside the biosafety cabinet:If there is a spill inside the biosafety cabinet: Keep the BSC running during spill and clean-up to contain aerosol.Keep the BSC running during spill and clean-up to contain aerosol. Place absorbent paper on spill and soak with disinfectant.Place absorbent paper on spill and soak with disinfectant. Allow 20 minutes of contact time. Wipe up spill, working from the edges to Allow 20 minutes of contact time. Wipe up spill, working from the edges to

the center. Clean spill areas with fresh paper towels soaked in disinfectant.the center. Clean spill areas with fresh paper towels soaked in disinfectant. Disinfect the BSC interior and any other equipment in the BSC with Disinfect the BSC interior and any other equipment in the BSC with

disinfectant.disinfectant. Discard contaminated disposable materials using appropriate biohazardous Discard contaminated disposable materials using appropriate biohazardous

waste disposal procedures.waste disposal procedures. Place contaminated reusable items in biohazard bags or autoclavable pans Place contaminated reusable items in biohazard bags or autoclavable pans

before autoclaving.before autoclaving. Run BSC 10 minutes after clean-up before resuming work or turning BSC Run BSC 10 minutes after clean-up before resuming work or turning BSC

off.off.

Note: If you are working in a BSC and the power went off in the room or the Note: If you are working in a BSC and the power went off in the room or the BSC fan stops blowing, IMMEDIATELY LEAVE THE ROOM.BSC fan stops blowing, IMMEDIATELY LEAVE THE ROOM.

Spill Clean-Up for BSL 1-2If the spill is in the laboratory but outside the biosafety cabinet:If the spill is in the laboratory but outside the biosafety cabinet: Call the BSO if the material is RG 2 or greater.Call the BSO if the material is RG 2 or greater. Clear area of all personnel. Wait at least 30 minutes for aerosol to settle Clear area of all personnel. Wait at least 30 minutes for aerosol to settle

before entering spill area.before entering spill area. Remove any contaminated clothing and place in biohazard bag to be Remove any contaminated clothing and place in biohazard bag to be

autoclaved.autoclaved. Put on disposable gown, safety glasses and gloves.Put on disposable gown, safety glasses and gloves. Initiate cleanup with disinfectant as follows:Initiate cleanup with disinfectant as follows:

Place dry paper towels on spill then layer a second set of disinfectant Place dry paper towels on spill then layer a second set of disinfectant soaked paper towels over the spill.soaked paper towels over the spill.

Encircle the spill with additional disinfectant being careful to minimize Encircle the spill with additional disinfectant being careful to minimize aerosolization while assuring adequate contact. aerosolization while assuring adequate contact.

Allow at least a minimum of 20 minutes contact time to ensure germicidal Allow at least a minimum of 20 minutes contact time to ensure germicidal action of disinfectant. Wipe up spill, working from the edges to the center. action of disinfectant. Wipe up spill, working from the edges to the center. Clean spill areas with fresh paper towels soaked in disinfectant.Clean spill areas with fresh paper towels soaked in disinfectant.

Decontaminate all items within the spill area.Decontaminate all items within the spill area. Discard contaminated disposable materials using appropriate biohazardous Discard contaminated disposable materials using appropriate biohazardous

waste disposal procedures.waste disposal procedures.

Spill Clean-Up for BSL 1-2

If the spill is outside the laboratory, in transit:If the spill is outside the laboratory, in transit: To prevent a spill, transport labeled biohazardous To prevent a spill, transport labeled biohazardous

material in an unbreakable, well-sealed primary material in an unbreakable, well-sealed primary container placed inside of a second unbreakable, container placed inside of a second unbreakable, lidded container (cooler, plastic pan or pail).lidded container (cooler, plastic pan or pail).

Should a spill occur in a public area, do not attempt Should a spill occur in a public area, do not attempt to clean it up without appropriate PPE.to clean it up without appropriate PPE.

Secure the area, keeping all people clear of the Secure the area, keeping all people clear of the spill.spill.

Call the BSO to assist in the clean-up.Call the BSO to assist in the clean-up. Stand by during spill response and cleanup activity Stand by during spill response and cleanup activity

to provide information and assistance.to provide information and assistance.

Biosecurity

Biosecurity vs. Biosafety

BiosecurityBiosecurity refers to ensuring the refers to ensuring the security security of biological materialsof biological materials to prevent theft, to prevent theft, illicit use or release. illicit use or release.

BiosafetyBiosafety focuses on focuses on reducing exposure toreducing exposure to and and release of biological materialsrelease of biological materials..

Integrating biosecurity and biosafety Integrating biosecurity and biosafety programs is important for work with select programs is important for work with select agents.agents.

Inspection

Inspection Elements

Laboratory IdentificationLaboratory Identification Containment Facility/EquipmentContainment Facility/Equipment Work PracticesWork Practices Hazard CommunicationHazard Communication Biohazardous Waste HandlingBiohazardous Waste Handling Laboratory Personnel Laboratory Personnel

Documents

Principles and Practices of Biosafety