Standard Operating Procedure for
Environmental Monitoring of
Microbial and Dust Contamination i n Kennesaw State University’s
Microbiology Lab, Prep, and Research Areas
By
Wysteria Hart
Golriz Khadem
Tim Shirley

The Standard Operating Procedure should be used to monitor microbial and dust contamination in the microbiology lab areas at Kennesaw State
University; including the teaching micro lab, its prep area and research lab. This monitoring is preformed to assess the microbial activities and possible contamination in these areas. Contamination to these areas could be attributed to personnel, student use, and general traffic through these areas. Contamination in these areas could possibly impede the progress of learning by students and researchers. These contaminates include Coryneforms from dust, fungal growth, and skin contaminants. There are a number of methodologies used for sampling possible microorganisms. These include swabbing of specific areas and passive microbial air sampling that uses plates of nutrient agar where microbes in the air can settle.
One important consideration must be the selection of sampling sites. The frequency of sampling depends upon the degree of risk associated with the contamination level at each sampling location. Areas where product is exposed to air or surfaces are important locations and included in the sampling plan. Biweekly is chosen to be done during normal work activity in each area of Prep,
Teaching, and Research. This bi-weekly testing has been developed to be flexible and user friendly. A flow chart is included in appendix A.
Areas to be tested include air supply vents, air exhaust vents, light switch, refrigerator door handle, floor, exhaust hoods, bench tops and general area, and any touch keys located on computers or instrumentation.

Environmental Monitoring Standard Operating Procedures for Preparation,
Teaching, and Research Areas Located At Kennesaw State University
Microbiology Laboratory:
 Environmental sampling shall be performed bi-weekly with follow up of confirmatory testing as indicated.
 The areas shall be closed to janitorial personnel at least 24 hours prior to testing.
 The Laboratory Director of Environmental Monitoring should be notified of test results

Record the date, location an d technician’s name where random samples were swabbed on room layout charts and/or log charts.

Guidelines specific to the Teaching and Research lab are listed last.
BENCH TOP
1. Monitor for aerobic gram positive & negative Coryneform organisms

Choose random section of lab bench top in each room
 Log location being swabbed on chart and drawing

Moisten a sterile cotton swab with dionized water
 Swab one (1) centimeter square area to be tested
 Streak sample onto Tryptic Soy Agar plates (follow technique and common Good Lab Practices outlined in appendix B)
 Perform streaks in triplicate

Allow to incubate at 37
˚C for 24-48 hours


Remove plates and record results into log
2. Monitor for fungal organisms (yeast and mold)
 Choose random section of bench top in each room
 Log location being swabbed on chart and drawing
 Moisten a sterile cotton swab with di-ionized water

Swab one (1) centimeter square area to be tested
 Streak sample onto Sabouraud Dextrose Agar plates (follow technique and common Good Lab Practices outlined in appendix B)
 Perform streaks in triplicate
 Allow to incubate at 37˚C for 24 to 48 hours

Remove plates and record results into log
3. Monitor for skin contaminants

Choose random section of bench top in each room
 Log location being swabbed on chart and drawing
 Moisten a sterile cotton swab with di-ionized water

Swab one (1) centimeter square area to be tested
 Streak sample onto Sheep Blood Agar plates (follow technique and common Good Lab Practices outlined in appendix B)
 Perform streaks in triplicate
 Allow to incubate at 37 ˚C for 24 to 48 hours
 Remove plates and record results into log
*Results are determined by performing SOP’s for microbial classification

**If more than 10 colonies grow on consecutive tests or if colonies appear to be gram negative, conduct follow up confirmatory testing.
***If follow up confirms level of contamination, take remedial actions
INSTRUMENTATION (COMPUTER KEYBOARD, pH METER, GLASSWARE,
LIGHT SWITCHES, REFRIGERATOR DOOR HANDLE, AND FLOOR)
* It is recommended that at least one to two of the areas listed above are sampled during each testing phase.
1. Monitor for aerobic gram positive & negative Coryneform organisms
 Choose random section of any one or all of the areas listed above

Log location or item being swabbed on chart and drawing
 Moisten a sterile cotton swab with di-ionized water

Swab one (1) centimeter square area to be tested
 Streak sample onto Tryptic Soy Agar plates (follow technique and common Good Lab Practices outlined in appendix B)

Perform streaks in triplicate
 Allow to incubate at 37 ˚C for 24-48 hours

Remove plates and record results into log
2. Monitor for fungal organisms (yeast and mold)
 Choose random section of any one or all of the areas listed above
 Log location or item being swabbed on chart and drawing
 Moisten a sterile cotton swab with di-ionized water

Swab one (1) centimeter square area to be tested


Streak sample onto Sabouraud Dextrose Agar plates (follow technique and common Good Lab Practices outlined in appendix B)
 Perform streaks in triplicate
 Allow to incubate at 37˚C for 24 to 48 hours
 Remove plates and record results into log
3. Monitor for skin contaminants
 Choose random section of any one or all of the areas listed above

Log location being swabbed on chart and drawing
 Moisten a sterile cotton swab with di-ionized water
 Swab one (1) centimeter square area to be tested

Streak sample onto Sheep Blood Agar plates (follow technique and common Good Lab Practices outlined in appendix B)

Perform streaks in triplicate
 Allow to incubate at 37˚C for 24 to 48 hours
 Remove plates and record results into log
*Results are determined by performing SOP’s for microbial classification
**If more than 10 colonies grow on consecutive tests or if colonies appear to be gram negative, conduct follow up confirmatory testing.
***If follow up confirms level of contamination, take remedial actions
HVAC SUPPLY VENT
1. Monitor for aerobic gram positive & negative Coryneform organisms

Choose random section of air supply vent


Log location being swabbed on chart and drawing
 Moisten a sterile cotton swab with di-ionized water
 Swab one (1) centimeter square area to be tested
 Streak sample onto Tryptic Soy Agar plates (follow technique and common Good Lab Practices outlined in appendix B)

Perform streaks in triplicate
 Allow to incubate at 37˚C for 24 to 48 hours

Remove plates and record results into log
2. Monitor for fungal organisms
 Choose random section of air supply vent

Log location being swabbed on chart and drawing
 Moisten a sterile cotton swab with di-ionized water

Swab one (1) centimeter square area to be tested
 Streak sample onto Sabouraud Dextrose Agar plates (follow technique and common Good Lab Practices outlined in appendix B)

Perform streaks in triplicate
 Allow to incubate at 37˚C for 24 to 48 hours

Remove plates and record results into log
*Results are determined by performing SOP’s for microbial classification
**If more than 10 colonies grow on consecutive tests or if colonies appear to be gram negative, conduct follow up confirmatory testing.
***If follow up confirms level of contamination, take remedial actions

HVAC EXHAUST VENT AND HOOD
1. Monitor for aerobic gram positive & negative Coryneform organisms
 Choose random section of air exhaust vent
 Log location being swabbed on chart and drawing
 Moisten a sterile cotton swab with di-ionized water

Swab one (1) centimeter square area to be tested
 Streak sample onto Tryptic Soy Agar plates (follow technique and common Good Lab Practices outlined in appendix B)
 Perform streaks in triplicate
 Allow to incubate at 37˚C for 24 to 48 hours

Remove plates and record results into log
*Results are determined by performing SOP’s for microbial classification
**If more than 10 colonies grow on consecutive tests or if colonies appear to be gram negative, conduct follow up confirmatory testing.
***If follow up confirms level of contamination, take remedial actions
PASSIVE OPEN AIR MONITORING
1. Monitor for aerobic gram positive & negative Coryneform organisms
 In prep, teaching and research area choose random area
 Log location being swabbed on chart and drawing
 Open a Tryptic Soy Agar plates and leave to open air overnight while room is closed to cleaning personnel

Cover and a llow to incubate at 37˚C for 24 to 48 hours


Remove plate and record results into log
2. Monitor for fungal organisms (yeast and mold)
 In prep, teaching and research area choose random area
 Log location being swabbed on chart and drawing
 Open a Sabouraud Dextrose Agar plates and leave to open air overnight while room is closed to cleaning personnel
 Cover and a llow to incubate at 37˚C for 24 to 48 hours

Remove plate and record results into log
3. Monitor for skin contaminants
 In prep, teaching, and research area choose random area

Log location being swabbed on chart and drawing
 Open a Sheep Blood Agar plates and leave to open air overnight while room is closed to cleaning personnel
 Cover and a llow to incubate at 37˚C for 24 to 48 hours
 Remove plate and record results into log
*Results are determined by performing SOP’s for microbial classification
**If more than 10 colonies grow on consecutive tests or if colonies appear to be gram negative, conduct follow up confirmatory testing.
***If follow up confirms level of contamination, take remedial actions

Appendix A
FLOW CHART
To be performed in each room
Coryneform
Bench Top Fungus
if + rx take remedial action
if + rx take remedial action
if + rx take remedial action Skin
Coryneform
Instrumentation Fungus if + rx take remedial action
if + rx take remedial action
Skin if + rx take remedial action
Coryneform if + rx take remedial action
HVAC Supply
Fungus if +rx take remedial action
HVAC Exhaust Coryneform if + rx take remedial action
Coryneform
Open air Fungus
if + rx take remedial action
if + rx take remedial action
Skin if + rx take remedial action

Works Cited:
Biological Environmental Control Laboratories, 1999. “Supplement 8 of USP 23
Serves As Guide For Establishing A Microbial Monitoring Program.” http://www.beclabs.com/baciss4.htm
(April 5, 2004).
Sterigenics, 2004. “Environmental monitoring.” http://www.iba-sni.com/lenv-monitoring.asp
(April 5, 2004).
Dr. Jerald Hendrix. Professor of Biology. Kennesaw State University.
Rm SC 332. jhendrix@kennesaw.edu
(April 8, 2004)
Furr, Keith A. Laboratory Safety, 5 th Edition, (Bocca Raton: CRC Press LLC,
2000) 603-645.
Birge, Edward A. Modern Microbiology, (Dubuque: Wm. C. Brown Publishers,
1992) 347-428.
Kaufman, James A. A Kinder, Gentler Laboratory Standard, (Natick, MA,
Laboratory Safety Institute) 19-32