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MICROBIOLOGY

SAFETY ON THE CLINICAL SAFETY ON THE LABORATORY CLINICAL MICROBIOLOGY MODULE/UNIT MICROBIOLOGY LABORATORY JOSETTE RUBE S. VALENZUELA MODULE/UNIT JOSETTE RUBE S. VALENZUELA

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OUTLINE Laboratory Safety A. Possible Routes for Infection Exposure Control Plan Universal and Standard VIII. Precautions A. Safety Guidelines for the Laboratory Personal Protective Equipment Engineering Controls IX. A. Biosafety Levels B. Biosafety Cabinets Postexposure Plan Disposal of Hazardous Waste A. Autoclave

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LEARNI NG OUTCOMES Define histology and other terms related to the study of tissues. Differentiate the common techniques used in the study of histology. Enumerate the steps in the preparation of tissue sections. Correlate the study of histology in the practice of the medical technology profession I.

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 B. Sterilization C. Disinfection D. Antiseptics E. Biocides F. Liquid Decontaminants Fire Safety A. Type A B. Type B C. Type C D. Type D E. Class K Chemical Safety A. Material Safety Data Sheet B. Hazardous Chemicals

LABORATORY SAFETY

A. POSSIBLE ROUTES FOR INFECTION Airborne o Aerosols from centrifugation of unstopped tubes or from heating cultures or specimens too rapidly, removing stoppers from tubes, leakage from a container that holds contaminated specimens Ingestion o Infection as a result of failure to wash hands or eating, drinking, smoking, applying cosmetics, or pipetting w the mouth Direct inoculation o Infection from needlesticks, broken glass, animal bites, small scratches on the dingers Mucous membrane contact o Infection if the organism can directly enter through the mucous membranes (conjunctiva of the eye) Arthropod vectors o Infectious sources include ticks, fleas, and mosquitoes

II. EXPOSURE CONTROL PLAN Exposure control plan  The risk of exposure to infectious agents for all job classifications and explains exposure-reduction methods must include: o Safety education o Universal precautions and safety standard precautions o Engineering controls o PPE o Disposal of hazardous waste o Postexposure procedures III.

UNIVERSAL AND STANDARD PRECAUTIONS

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Centers for Disease Control (CDC) o Universal precautions as recommendations that describe handling of clinical specimens by health care personnel; 1987 Clinical Laboratory and Standards Institute (CLSI) o Universal precautions as set of preventive measures designed to reduce the risk of transferring HIV, hepa B virus, and other bloodborne pathogens in the health care setting o Apply to human blood and other body fluids w visible blood but does not apply to feces, nasal secretions, saliva (unless they contain visible blood) Standard precautions o Set of preventive measures applied to all patients that are designed to reduce the risk of infection in the health care setting Potentially infectious o Blood, tissue, body fluids, secretions, excretions (except sweat)

A. SAFETY GUIDELINES FOR THE LABORATORY No food or drink No smoking No cosmetics Protective eyewear must be worn Wear face shields Clothing: gowns, aprons lab coats, closed shoes Hair should be tied up Wash hands frequently and thoroughly Eyewash stations: 100 feet or 10 secs away from any area in the lab 10. Emergency stations: shower 11. Respirators: available where there is air contamination 12. Mouth pipetting: ingestion of microorganisms; use pipette bulbs/automated devices 13. Glassware and sharps: in puncture-resistant containers 14. Centrifuges: minimize the production of aerosols 1. 2. 3. 4. 5. 6. 7. 8. 9.

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IV. PERSONAL PROTECTIVE EQUIPMENT Protective laboratory clothing disposable gloves, eye protection, face masks Barrier protection: to prevent skin and mucous membrane contamination Gloves: worn when there is potential for skin contact w potentially infectious materials; worn in phlebotomy and capillary puncture Facial protection and body covering: worn when splashes w blood and other body fluids High-efficiency particulate air respirators: should be fit tested for each person; those who encounter mycobacteria through contact w either patient or specimen Lab employees who come in contact w body fluids should be offered hepa B vaccinations for free V. ENGINEERING CONTROLS To protect employees from the hazards that may occur during the performance of lab procedures All labs must adhere to a minimum of Biosafety Level 2 guidelines

A. BIOSAFETY LEVELS FOR INFECTIOUS AGENTS Table 1. Biosafety Levels BS Function 1

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No known pathogenic potential for immunocompetent individuals Bacillus subtilis Most undergraduate lab courses operate at this level Adherence to std lab techniques Level 1 + lab coats, protective gloves, limited access, decontamination of all infectious waste, biohazard signs Apparatus includes partial containment equipment Lab acquired infections: HBV, HIV, Staphylococcus, enteric pathogens such as Salmonella and Shigella Level 2 + special lab clothing, controlled access Mycobacterium tuberculosis, Brucella, Coccidiodes immitis, Rickettsia, arbovirus Air movement must be carefully contained Level 3 + entrance through a separate room, street clothing  lab clothing Maximum containment Filovirus, arenavirus

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Biohazard label  Used to identify those areas of the lab where infectious specimens or cultures are stored/present

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      Figure 1. Biohazard label Sterilization  Done by heat, UV light, passage of air through HEPA filter i.

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BIOSAFETY LEVEL 2 CRITERIA FOR MICROBIOLOGY PRACTICES Limit access to lab when working w infectious agents Decontaminate work surfaces at least once a day and after any spill Use mechanical pipetting devices Do not eat, drink smoke, or use cosmetics. Do not store food in biological refrigerator. Wash hands thoroughly Minimize the creation of aerosols Wear lab coats, gowns, smocks and remove before leaving the lab Avoid skin contamination by covering cuts w occlusive bandages and using gloves Do not use needles that are bent, cut, or recapped Handle M. tuberculosis at BSL 3 in Class II BSC while wearing a solid-front gown and personal respirator (or facefolding mask); use sealed safety cups for centrifugation Handle mild-lime fungi in BSC; plated media for myco should be sealed w a cellulose band or cellophane or labelling tape (prevent dispersion of spores) B.

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Vertical laminar-flow biological cabinets Open-fronted; have HEPA filter, recirculated airflow w/in the workspace Exhaust air is filtered, too, by the HEPA filter Type II-A o Self-contained w 70% of the air recirculated o Not required to be vented; acceptable for low- to moderate-risk agents o Most hospital microbio labs use this type Type II-B o Must be vented w 30% of the air exhausted from the cabinet and 70% recirculated back into the room iii. CLASS III BSC Provide the highest level of safety All air entering and leaving the cabinet is sterilized w a HEPA filter Supply air: drawn through the HEPA filter Exhaust air: filtered through 2 HEPA filters System is entirely enclosed Infectious materials: handled w rubber gloves VI. POST EXPOSURE PLAN All lab accidents or safety incidents: reported to the lab safety officer or supervisor Immediate and appropriate medical treatment Administration of immunizations, hepa B immune immunoglobulin, hepa B vaccinations Collection of serum samples at 6 weeks, 3 mos, and 6 mos for antibodies to HIV or abraded skin exposure to HBV or HIV Documentation of the accident w a report Corrective actions VII. DISPOSAL OF HAZARDOUS WASTE Microbiological waste must be decontaminated before disposal Needles and sharps should be placed in puncture-resistant containers Contaminated materials should be placed into two leak-proof plastic bags Contaminated pipettes, swabs, and glass should be placed into puncture-resistant burn boxes A. AUTOCLAVE Decontaminates that uses saturated steam at 121oC and 15 psi of pressure for 1 hr to sterilize Infectious medical waste: 132oC for 1/2 -1 hr Monitored periodically Bacillus stearothermophilus: spore indicator; weekly monitoring C. STERILIZATION Physical or chemical process that kills all microorganisms, including spores Moist heat or steam under pressure: autoclave Dry heat: 171oC for 1 hr, 160oC for 2 hrs, 121oC for 6 hrs Liquid: glutaraldehyde, 6-30% hydrogen peroxide, 6-8% formaldehyde, chlorine dioxide

BIOSAFETY CABINETS

i. CLASS I BSC Provide partial containment for procedures involving moderate- and high-risk microorganisms (BSL 2 and 3) Open-fronted, negative-pressure, ventilated cabinets Unsterilized air enters and circulates w/in the cabinet Exhaust air from cabinet is filter by HEPA filter ii. CLASS II BSC Provide partial containment for procedures involving moderate- and high-risk microorganisms (BSL 2 and 3) Sterilize bot air entering and circulating w/in the cabinet and the exhaust air

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D. DISINFECTION Destroys most microbes but does not kill spores Moist heat: 75-100oC Liquid: glutaraldehyde, 3-6% hydrogen peroxide, 1-8% formaldehyde, chlorine compounds, 70% isopropyl alcohol Liquid household bleach Quaternary ammonium compounds: benzalkonium chloride E. ANTISEPTICS For skin or tissue; not to be substituted for a disinfectant Alcohols: 70% ethyl or isopropyl alcohol; not against spores Iodophors: iodine combined w an organic carrier molecule; povidone-iodine and poloxamer-iodine 2

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Hexachlorophene: bactericidal and broad spectrum but may be toxic

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F. BIOCIDES Chemical agents that inactivate microorganisms; either -static and inhibit growth of the microbe or -cidal and kill the target organism G. LIQUID DECONTAMINANTS 70% EtOH or 10% bleach (NaClO) For 10 mins to increase ability to decontaminate VIII. FIRE SAFETY Ignition sources: open flames; heating elements; spark gaps, from light switches or static electricity; electrical instrumentation A. TYPE A Water Fire Extinguisher Used on combustible materials for fires in paper, wood, rubber, sloth, plastics Extinguish fire w cooling effect

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REVIEW QUESTIONS

Question 1 Question 2 Question 3 Answer 1 Answer 2 Answer 3 XI.

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REFERENCES

APA Format

H. TYPE B Carbon dioxide fire extinguisher Used on extremely flammable liquids or electrical fires including fires of oil, kerosene, gasoline, paints, fats, grease, solvents Extinguish fire by eliminating oxygen I. TYPE C Dry chemical extinguishers Used in electrical fires in wiring and other electrical sources or equipment Extinguish fire bc chemicals do not conduct electricity J. TYPE D Combustible metals Used for fires involving combustible metals: Na, K, Mg, Na-K alloys NaCl salt wc forms a metal layer over the fire wc occludes the oxygen Graphite, sodium carbonate based, copper based

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K. CLASS K Dry and wet chemical extinguishers Used for kitchen fires involving combustible materials such as oil or fat Extinguish fire by using various wet or dry chemical agents

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IX. CHEMICAL SAFETY Every compound or reagent must have precautionary labels

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A. MATERIAL SAFETY DATA SHEET Characteristics of hazardous compound chemicals

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Corrosive: causes visible destruction or irreversible damage to human skin on contact Toxic: serious biological effects after inhalation, ingestion, or skin contact w relatively small amts Carcinogenic: ability of chemical to induce a malignant tumor Ignitable: any chemical that can burn and includes both combustible and flammable liquids Explosive: reactive and unstable substances that readily undergo violent chemical change

Table 2. Sections of MSDS Section Information I Manufacturer’s name and address, emergency tel. no., tel no. for info about preparation II Hazardous ingredient’s identity info, components, specific chemical identity common name, PEL III Physical/chemical characteristics IV Fire and explosion hazard data V Reactivity data VI Health hazard data: routes of entry VII Precautions of safe handling and use VIII Control measures L.

HAZARDOUS CHEMICALS

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