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Risk Assessment in the Clinical Microbiology Laboratory
Nellie DumasBacteriology Laboratory
Wadsworth CenterNew York State Department of Health
Risk Assessment WebinarJune 15, 2011
Clinical Laboratory PRIORITIES
Safety
Accuracy
Timeliness
Lab-Acquired Infections in NYS 1999-2010
• E. coli O157
• Neisseria meningitidis
• Salmonella Typhi
• Campylobacter jejuni
• Brucella sp.
Biohazard Risk Assessment
CLEP Safety Sustaining Standard of Practice 1: Biohazard Risk Assessment and Biosafety Program
Minimize risk for lab-acquired infections by addressing:
• Aerosol-generating specimen/culture procedures (e.g., vortexing, centrifuging, pipetting, mixing)
• Contamination / Cross-contamination potential
• Sharps
Biohazard Risk Assessment
• Identification of hazardous characteristics of agents worked with in lab
• Identification of lab procedure hazards: agent concentration, suspension volume, equipment, aerosol-generating procedures, use of sharps
• Determine appropriate biosafety level and select additional precautions
• Evaluate proficiency of staff regarding safe practices and safe operation of equipment
• Review risk assessment with a biosafety professional
Clinical specimen types known to be potential sources for agents of laboratory-acquired infection
Specimen Type Agents that could be presentBlood HBV, HCV, HIV, SARS, WNV, Brucella, N. meningitidis, Francisella,
Enteric pathogens, B. pertussis, Leptospira, other select agents such as B. anthracis, Y. pestis, Burkholderia mallei and B. pseudomallei
Serum WNV, Botulism toxin, HBV, HCV, HIV
CSF HBV, WNV, HIV, Brucella, N. meningitidis, Francisella, M. tuberculosis, B. anthracis, Y. pestis
Saliva N. meningitidis, HBV, HIV
Urine HBV, HIV, SARS, Brucella, Francisella, Salmonella, STEC, M. tuberculosis, Y. pestis, Leptospira, B. anthracis
Feces Salmonella, Shigella, Campylobacter, Y. pestis, Vibrio species, H. pylori, STEC, HBV, SARS, Polio virus, B. anthracis, Botulism toxin
Semen Brucella, HBV, HIV
Nasopharyngeal N. meningitidis, B. pertussis, C. diphtheriae, H. pylori, Polio virus, SARS, HBV
Respiratory sites SARS, Brucella, B. anthracis, M. tuberculosis, B. pertussis, Legionella, Francisella, Burkholderia mallei, B. pseudomallei, Y. pestis
Tissues SARS, Polio virus, WNV, Brucella, M. tuberculosis, Campylobacter, Leptospira, B. mallei, B. pseudomallei
Gastric lavage M. tuberculosis, H. pylori
Wound/Skin lesion exudates Francisella, B. anthracis, Burkholderia mallei, B. pseudomallei
Biorisk characteristics of viruses reported to cause laboratory-acquired infections
Biological Agent
Infective Dose
Potential mode of transmission
RecommendedContainment
Hepatitis B 108-109 particles/ml Percutaneous or mucocutaneous
BSL-2 for handling body fluid and tissue specimens
Hepatitis C 102-103 particles/mlPercutaneous, rarely
mucocutaneousBSL-2 for handling body fluid and
tissue specimens
HIV 100-104 particles/mlPercutaneous or mucocutaneous
BSL-2 for handling body fluid and tissue specimens
Influenza Viruses Varies by strain Inhalation of aerosols; mucocutaneous
H1N1: Splash protection if performing rapid immunoassay; biosafety cabinet
if performing IFA, DFA, culture or molecular assays
HPAI: requires BSL-3 conditions
SARS-Corona virus Unknown Inhalation of aerosols Untreated specimens processed in biological safety cabinet
West Nile Virus UnknownInhalation of aerosols;
percutaneous or mucocutaneous
BSL-3 for manipulation of cultures
Biorisk characteristics of bacteria reported to cause laboratory-acquired infections
Biological Agent
Infective Dose Potential mode of transmission
RecommendedContainment
Brucella species 10 to 100 orgs Inhalation of aerosols BSL-3 for manipulation of cultures
Campylobacterspecies
500 orgs or less Ingestion due to cross-contamination
BSL-2
Coxiella burnetii 10 orgs Inhalation of aerosols BSL-3 for manipulation of cultures
Escherichia coli O157:H7
10 to 100orgs Ingestion due to cross-contamination
BSL-2
Francisellatularensis
Aerosols: 5 to 10 orgs
Ingestion: 108 orgs
Inhalation of aerosols BSL-3 for manipulation of cultures
Biological Agent
Infective Dose
Potential mode of transmission
RecommendedContainment
Mycobacterium tuberculosis
1 to 10 orgs Inhalation of aerosols BSL-3 for manipulation of cultures
Neisseriameningitidis
Not known Inhalation of aerosols Sterile site isolates should be manipulated in biological safety
cabinet
Salmonella species
105 to 109
orgsIngestion due to cross-
contaminationBSL-2 for non-typhi; BSL-3
recommended for manipulation of Salmonella Typhi if aerosols are likely
Shigella species 10 to 100 orgs Ingestion due to cross-contamination
BSL-2
Staphylococcus aureus
Virulence varies greatly
between strains
Inhalation of aerosols; percutaneous or mucocutaneous
BSL-2
Biorisk characteristics of bacteria reported to cause laboratory-acquired infections
Protect from Aerosol and Cross-contamination Transmission
Always work with suspect specimens in BSC (ref: BMBL5 Agent Summary Statements):
• Neisseria meningitidis!!!!!!!!!• Salmonella Typhi• Shiga toxin-producing E. coli• Shigella• B. anthracis, Brucella, B. mallei,
B. pseudomallei, F. tularensis, Y. pestis
High Risk Procedures for High Risk Agents
Outside of Biosafety Cabinet
• Opening culture plates• Picking colonies off culture plates• Vortexing tubes • Setting up automated identification
systems• Performing catalase• Performing oxidase• Performing motility test
Biohazard Risk Assessment
Method or Procedure:• Risk Factor(s)• Probable route(s) of transmission• Required practices, primary barriers, and
safety equipment• Triggers for implementation of enhanced
precautions for high risk agents• Required enhancements in practices,
primary barriers, or safety equipment
Aerosol-generating Procedures
• Vortexing
• Centrifuging
• Pipetting
• Mixing
• Grinding
• Blending
• Automated system preparations
• Fixing slides with a flame
• Taking off disposable gloves improperly
Risk assessment of laboratory activitiesLaboratory activity Causes
aerosolsPotential
for splashes
Potential for
ingestion due to
splashes
Potential for skin
inoculation
Processing primary specimens
Removing caps or swabs from culture containers Yes Yes Yes
Grinding tissues Yes
Blending food specimens Yes
Centrifuging Yes
Pouring or decanting fluids Yes Yes Yes
Using needles or syringes:
Aspirating fluid from a sealed bottle Yes Yes
Withdrawing needles from stopper Yes Yes
Inoculating culture plates with fluid from a syringe Yes Yes Yes
Disposal of contaminated sharps Yes
Laboratory activity Causes aerosols
Potential for
splashes
Potential for
ingestion due to
splashes
Potential for skin
inoculation
Bacterial suspensions
Using a swab to make a bacterial suspension Yes Yes Yes
Vortexing , sonicating Yes Yes Yes
Aspirating bacterial suspension with pipette Yes Yes Yes
Using autoinoculater for automated identification systems
Yes Yes Yes
Inoculating identification cards using vacuum system Yes
Inoculating suspension onto plate with a swab Yes Yes Yes
Risk assessment of laboratory activities
Laboratory activity Causes aerosols
Potential for
splashes
Potential for
ingestion due to
splashes
Potential for skin
inoculation
Manipulating bacterial culture plates
Opening plate to observe growth Possibly
“Sniffing” plate as part of bacterial identification Yes
Using inoculation loops:
Flaming loop with bacterial growth on it Yes
Cooling loop in culture plate Yes
Manipulating bacterial growth with hot loop or needle Yes Yes
Disposal of specimens and culture plates
Roughly discarding bacterial suspensions Yes Yes Yes
Roughly discarding culture plates or specimens into receptacle
Yes Yes Yes
Risk assessment of laboratory activities
Procedure Hazards & Controls
Risk Factor(s) Hazards Required practices and safety equipment
Triggers for implementation of
enhanced precautions
Required enhancements in practices and safety
equipment
Removing caps or swabs from culture containers
Aerosols/droplets, splashes
Handle and process specimen with facial barrier (BSC, face or bench shield), lab coat, disposable gloves
Suspect botulism specimens
Suspect select agents
Mouse Bioassay – use animal testing PPE & handling SOP
Suspect select agents – deliver to Biodefense Lab
Grinding tissues Aerosols/droplets Same as above Same as above Same as above
Blending food samples
Aerosols/droplets Same as above Same as above Same as above
Centrifuging Aerosols/droplets Same as above; use capped cups and open in BSC
Same as above Same as above
Pouring or decanting fluids
Aerosols/droplets, splashes
Same as above Same as above Same as above
Method or Procedure: Processing primary specimens
Risk Factor(s) Hazards Required practices and safety equipment
Triggers for implementation of
enhanced precautions
Required enhancements in
practices and safety equipment
Aspirating fluid from a sealed bottle
Aerosols/droplets, percutaneous
Handle and process specimen with facial barrier (BSC, face or bench shield), lab coat, disposable glove; use safe sharps practices; do not recap; use sharps disposal container
Suspect botulism specimens
Suspect select agents
Mouse Bioassay – use animal testing PPE & handling SOP
Suspect select agents –deliver to Biodefense Lab
Withdrawing needles from stopper
Aerosols/droplets, percutaneous
Same as above Same as above Same as above
Inoculating culture plates with fluid from a syringe
Aerosols/droplets, splashes, percutaneous
Same as above Same as above Same as above
Disposal of contaminated sharps
Percutaneous Follow sharps handling and disposal SOP; do not recap; use sharps disposal container
Same as above Same as above
Procedure Hazards & Controls
Method or Procedure: Using needles or syringes
Procedure Hazards & Controls
Risk Factor(s) Hazards Required practices and safety equipment
Triggers for implementation of
enhanced precautions
Required enhancements in
practices and safety equipment
Using a swab to make a bacterial suspension
Aerosols/droplets, splashes
Perform initial testing of unknown Gram negs in BSC until SA R/O; wear appropriate PPE
Suspect select agents
Suspect N. meningitidis
Suspect select agents –deliver to Biodefense Lab
Suspect N. mening/use BSC
Vortexing, sonicating Aerosols/droplets, splashes
Perform in BSC; use capped/enclosed materials; allow settling time
Same as above Same as above
Aspirating bacterial suspension with pipette
Aerosols/droplets, splashes
Perform initial testing of unknown Gram negs in BSC until SA R/O; wear approp PPE
Same as above Same as above
Using autoinoculaterfor automated identification system
Aerosols/droplets, splashes
NA NA NA
Inoculating identification cards using vacuum system
Aerosols/droplets NA NA NA
Inoculating suspension onto plate with a swab
Aerosols/droplets, splashes
Same as above Same as above Same as above
Method or Procedure: Bacterial suspensions
Procedure Hazards & Controls
Risk Factor(s) Hazards Required practices and safety equipment
Triggers for implementation of
enhanced precautions
Required enhancements in practices and safety
equipment
Roughly discarding bacterial suspensions
Aerosols/droplets, splashes
Place carefully in bleach/biohazard bag bucket on bench
Any suspensions made in BSC
Carefully discard in bleach/biohazard bag in BSC; follow SOP for bag disposal
Roughly discarding culture plates or specimens
Aerosols/droplets, splashes
Place carefully in biohazard bag/autoclave stock pot
Any culture plates within BSC Carefully discard in bleach/biohazard bag in BSC; follow SOP for bag disposal
Method or Procedure: Disposal of specimens and culture plates
Primary specimens for bacterial culture
Inoculate culture plates wearing lab coat and gloves and face shield, or inside biological safety cabinet
Growth of gram-negative
diplococcisuggestive of
Neisseria meningitidis
Work in BSC until Neisseria meningitidis
is ruled out
Source: Respiratorysite or sterile site such
as blood or CSFSlow-growing colonies
on blood agarGram stain:
gram-negative coccobacilli
Any specimen source
Work in BSC
Oxidase –Urea –
No hemolysisNo pigment
Rule out Francisellatularensis
Oxidase +Urea +
No hemolysisNo pigment
Rule out Brucella sp.
Other bacterial growth
Work on bench top
usingappropriate
sterile technique
Primary specimen received with request to test for or
confirm any of the following:
Bacillus anthracisBrucella species
Burkholderia malleiBurkholderia pseudomallei
Coxiella speciesFrancisella species
Q-feverRickettsia prowazekiiRickettsia rickettsii
Yersinia pestis
Begin work in BSL-3 facilityuntil select agents are
ruled out
Flowchart for processing primary specimens
Biorisk Assessment
Establish a PPE Program, including
• Use of Biosafety Cabinet
• Use of Facial Barriers
• Appropriate Use of Disposable Gloves
Biosafety Containment Risk Assessment
Recommended Use of Biosafety Cabinet includes::
• Primary specimen handling/processing
• Aerosol-generating procedures
• Open capped centrifuge cups in BSC
• All vortexing: Use closed items only
(Allow settling time before opening or manipulating vortexed material)
Disposable Gloves
• Caution should be observed in removing gloves: Snapping or stretching gloves may result in aerosol formation!
• Immediate glove removal upon leaving each work station is intended to prevent inadvertent contamination of communal objects (e.g., phones, pens, keyboards, etc.)
Clinical Microbiology Lab Reality
Budgetary Issues:• Space constraints - new technologies to
reduce exposure potential, but no space• PPE – off brands; inferior grade; use of
appropriate PPE reduces risk, but does not eliminate (staff training!)
• Laboratory equipment –upgrading/replacing existing equipment; maintenance contracts
Clinical Microbiology Lab Reality
Education/Training:
• Trigger points!!
• Indicate when work should be performed in BSC
• How to properly perform tasks in BSC
• Annual risk assessment – clearly identifies potential aerosol and exposure operations
Clinical Microbiology Lab Reality
Suspect/Unknown Specimens:
• Perform within your risk assessment reality!
• Perform testing within your capability!
• Slow-growing organisms - FLAGS
• PT specimens - FLAGS
• Select agents – rule/out and refer - FLAGS
Clinical Laboratory PRIORITIES
Safety
Accuracy
Timeliness
Contact Info
• Nellie Dumas: nbd01@health.state.ny.us
• Christina Egan: eganc@wadsworth.org
• David Hill: djh08@health.state.ny.us
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