I. LABORATORY SAFETY: PRECAUTIONS AND PRACTICES 



Apparently Legionnaires" disease is acquired by tlie airborne route, presumably after droplet 

 nuclei containing the Legionnaires' disease bacterium (LDB) are iniialed. Laboratories must also 

 consider a second route - the puncture of the skin with equipment or broken glass contaminated 

 with the LDB. The following general precautions and laboratory practices are minimal safety 

 standards for handling infectious material. In addition, most of the contributors to this manual 

 included in their procedures or discussions safety measures that are pertinent to the techniques 

 detailed in their chapters. 



A. Aerosols 



Many laboratory manipulations generate potentially infectious aerosols; every effort 

 should be made to minimize aerosol formation and to contain any such aerosols formed. 

 Appropriate precautions can significantly reduce aerosols, which are generated whenever a 

 liquid surface is disrupted, e.g., the liquid film between the cap and the wall of a container 

 stretching and breaking when a screw cap or cover is removed: bubbles of lluid bursting at 

 the tip of a pipette or on the surface of a fiuid after vigorous shaking or mixing; fluids 

 spattering from heated wire loops; wire loops vibrating after inadvertently touching the sides 

 of tubes or containers; tubes breaking or leaking in a centrifuge; streaking agar media in 

 tubes or plates, especially when medium surface is rough (resulting from bubbles in the 

 medium when it was poured). The following practices help minimize and/or contain aerosols. 



1. Always perform operations such as transferring cultures and preparing microscope 

 smears for examination in a biological safety cabinet. 



2. Perform any procedure which produces aerosols but cannot readily be done in a 

 safety cabinet (such as centrifuging in a large centrifuge, shaking broth cultures during 

 incubation, or using a high-speed blender) with safety equipment that will prevent the 

 dissemination of infectious aerosols if the primary container breaks or leaks. 



3. Streak agar plates which have a smooth instead of rougli agar surface and thus 

 reduce aerosol formation by 99''r. 



4. Insert a cool rather than hot inoculating loop into media. This will reduce aerosols 

 90%-95%. 



5. .'Mlow the last drop in the pipette to drain out instead of blowing it out. This will 

 reduce aerosol formation 67%. 



6. Mix cultures in a tube instead of a pipette and thus reduce droplet nuclei by 

 1 00':^. 



7. Work over a disinfectant-soaked gauze sheet (rather than on the hard surface of a 

 bench top) to absorb falling drops of culture material and reduce aerosols by 90%. 



8. Insert a needle wrapped with an alcohol pledget througli the top of a stoppered 

 vaccine bottle to release the pressure. This reduces aerosol production by 99%. 



9. Leave screw caps on centrifuge tubes to allow contents to settle before opening 

 caps, and reduce aerosols considerably. 



10. Centrifuge in safety cups to eliminate essentially all aerosol formation in the event 

 of tube breakage. 



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