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An Investigation of Microbials In Hospital Air Environments

The microbiological quality of indoor air, long a concern among academics and researchers, has become an issue to the general public as well. Concerns have been driven by a combination of factors, including increased time spent indoors, superinsulated building construction, and increased media attention to sick building syndrome and building-related illness. Legislative and regulatory bodies have begun paying attention as well; during 1994, H.R. 2919, The Indoor Air Quality Act, passed the House of Representatives, while the Occupational Health and Safety Administration (OSHA) released proposed rules on indoor air quality for public comment.

Despite increasing public pressure, statutory or regulatory guidelines, by most estimates, are unlikely in the immediate future. There is a general acceptance of the cause-and-effect relationship between aeroallergen exposure and allergic and infectious disease; however, threshold levels or permissible exposure limits (PELs) have not been developed. It appears that building managers, industrial hygienists and indoor environment consultants will continue to be challenged to provide guidance in an unregulated arena. Compounding this difficulty is a lack of readily available data regarding the types and concentrations of microorganisms commonly recovered from indoor air environments.

The microbiological quality of indoor air in hospitals is as much of an issue as in any other type of building, with increased emphasis because of the potential severity of the consequences of nosocomial infection. It seems somewhat counter-intuitive, but many patients are actually at increased risk of infection while in the hospital. For example, transplant patients are routinely given immunosuppressant drugs to reduce the likelihood of organ rejection, and radiation therapy weakens the immune system. The samples included in this paper were collected in hospital environments by our staff or were submitted to our laboratory for analysis by clients from around the country.

This article will review 16 microbiological aerosol sampling events from different hospitals. Five types of sample sites were selected for inclusion: operating rooms; intensive and critical care units; patient rooms; nurses' stations; and corridors. Site selection was based upon representation of typical regions within a hospital, sampling methodology and the amount of data available for review. These data are intended to demonstrate the types and concentrations of microorganisms which may be recovered within the hospital environment. Data reported herein are not meant to serve as guidelines for prescriptive purposes.

All of the data included for review in this article are from the analysis of samples collected using a two stage Grasby-Andersen 10-850 microbial impact sampler. Fungi were recovered using Sabouraud Dextrose Agar (SDA) and incubated for 21 days at 25E C. Fungal identification was performed by microscopic evaluation, gross visual inspection and, when necessary, gas chromatographic techniques. Bacteria were recovered using Plate Count Agar (PCA) or Trypticase Soy Agar (TSA). Samples were incubated for five to seven days at 28E C. Bacterial identification was performed using the MIDI/Sherlock Microbial Identification System (MIS), microscopic evaluation and colony morphology. The MIS uses gas chromatography to generate a profile of the unknown microorganism's cellular fatty acids (CFA) and statistical pattern recognition software to compare the profile to the CFA databases for identification.

RESULTS

Operating Rooms: Air samples were collected from 18 operating rooms (ORs) during 16 different sampling events. Results obtained from analysis of these samples indicate that fungi were recovered in 83.3 percent of the samples, while bacteria were recovered in all of the samples. Less prevalent were yeasts and actinomycetes, recovered in 33.3 percent and 16.6 percent of the samples, respectively.

The concentrations of fungi recovered ranged from 0 colony forming units per cubic meter of air (CFU/m³) to 104 CFU/m³. Penicillium spp. was recovered most frequently, followed by Aspergillus spp. and Cladosporium spp. in 52.4 percent, 23.9 percent and 14.3 percent of the samples, respectively. Most of the fungi recovered from samples included in this article were identified to genus level only. However, it is significant to note that 4.7 percent of the Aspergillus spp. recovered from ORs were identified as A.fumigatus, which is known to be the primary cause of aspergillosis.

Bacteria were recovered in all of the samples collected in ORs. Bacterial concentrations ranged from 15-225 CFU/m³ with a mean concentration of 74 CFU/m³. About 75 percent of the bacteria recovered were gram positive. Staphylococcus spp. and Micrococcus spp. occurred most frequently, 59 percent and 47 percent, respectively. The most common species of these bacteria were S.epidermidis, S.hominis, S.capitis, S.haemolyticus, M.luteus, M.roseus and M.kristinae. Bacillus spp., Arthrobacter spp. and Pseudomonas spp. were each recovered in approximately 18 percent of the samples. Streptococcus sp., Methylobacterium sp. and Corynebacterium sp. were each recovered in approximately 12 percent of the samples. Flavobacterium sp., Clavibacter sp., Rathayibacter sp. and Rhodococcus spp. were also recovered, but at a very low frequencies.

Surprisingly, yeasts were recovered in 33.3 percent of the samples collected in hospital operating rooms. However, the concentrations were quite low. The most frequently isolated yeasts were Cryptococcus albidus and Candida spp., while Rhodotorula rubra, Aureobasidium spp. and Zygosacchomyces sp. were much less prevalent. The survival of yeasts is usually dependent on the moisture content of the surrounding air and the temperature. Dry warm conditions tend to eliminate viable yeast.

Actinomycetes were rarely recovered from the operating room air samples (6 percent of the samples). Low recovery may be because of the use of agar poorly suited for actinomycete recovery. Of the actinomycetes recovered, 75 percent were Streptomyces while 25 percent were similar to Streptoverticillium.

Intensive Care and Critical Care Units: Air samples were collected from 17 intensive care unit (ICU) and Critical Care Unit (CCU) sites and analyzed according to the techniques described above. Fungi were recovered at a mean concentration of 23 CFU/m³ and a range of 0-90 CFU/m³. Seven of the 17 samples were negative for fungi after 21 days of incubation of 25E C. Cladosporium spp., Aspergillus spp. and Penicillium spp. were the most frequently isolated fungi from the samples which exhibited fungal growth. One sample contained very low concentrations of Acremonium sp., while another contained moderate levels of Aspergillus fumigatus. The significance of A. fumigatus as the primary cause of aspergillosis has been discussed briefly above. Lacellina sp. and Gliocephalotrichum sp. were observed in two of the samples. In our experience, these organisms are rare in indoor environments and their presence may be related to the intake of outdoor air.

Yeasts were also isolated from the ICU/CCU samples, but at very low concentrations. The types isolated were similar to those isolated from the operating room samples, primarily Candida spp. and Rhodotorula spp.

Nurses' Stations: One type of area within the hospital that experiences constant activity is the nurses' station. Although these areas may not seem as critical as operating rooms or intensive care units, it must be recognized that "microbial aerosol communication" does occur between rooms and areas within a hospital. Microbes are passively aerosolized by mechanical disturbance of spores or microbe-containing dust.

A total of 11 samples collected from different nurses' stations. Fungi were recovered from 73 percent of the samples. All but one of the samples were collected as part of routine monitoring programs. These routine samples had a mean concentration of 23 CFU/m³ and range of 0-70 CFU/m³. One sample, collected from a problem or complaint area had 565 CFU/m³. Unfortunately, we performed only the laboratory analysis and do know whether the elevated levels of fungi were determined to be the cause of the complaints.

The most frequently isolated fungi were Penicillium spp. and Cladosporium spp. which were each recovered in 55 percent of the samples. Aspergillus spp. was recovered from 33 percent of the samples collected at nurses' stations and all of the aspergilli were identified to species level. A.fumigatus was isolated from only one sample. Chrysosporium sp. and Gliocephalotrichum sp. were also isolated, but at very low concentrations and from only one sample.

Low concentrations of yeasts were recovered from 25 percent of the samples collected at nurses' stations. Rhodotorula rubra and Aureobasidium spp. were identified.

Bacteria were recovered from all of the samples at the nurses' stations. Excluding one outlier, the mean bacterial concentration was 52 CFU/m³ and the range 7-88 CFU/m³. One sample, collected in an area from which several complaints had been registered, contained 511 CFU/m³. To our knowledge, it was not determined whether bacteria was the cause of the problem.

Bacillus spp. was isolated from 75 percent of the samples. The most common species of Bacillus were B.mycoides, B. megaterium and B.licheniformis. We were unable to identify some of the species of Bacillus recovered because of their unique CFA profiles. Staphylococcus spp. and Micrococcus spp. were each recovered from 63 percent of the samples, with the most common species identified as S.haemolyticus, S.simulans, S.cohnii, S.hominis, S.aureus, M.luteus and M.varians. Arthrobacter spp. were recovered from 25 percent of the samples, however, not all of the isolates were successfully identified to species level.

Patient Rooms: A limited number of air samples collected from patient rooms are included for review. Fungi were recovered at a concentration ranging from 14-70 CFU/m³ and at mean of 43 CFU/m³ of air. Bacteria were recovered at concentrations ranging from 15-226 CFU/m³ and at a mean of 104 CFU/m³.

Only selected samples were subjected to fungal and bacterial identification procedures and quantitative data was not obtained from all samples. Common fungi recovered include Cladosporium spp., Penicillium spp. and Paecilomyces spp. The most common types of bacteria were Staphylococcus spp. and Bacillus spp. Less common were Pseudomonas spp., Rhodococcus sp., Corynebacterium sp., Nocardia sp., Micromonospora sp., and Aureobacterium sp.

Hospital Corridors: Air samples collected from corridors and hallways yielded concentrations of fungi ranging from 0-273 CFU/m³ with a mean of 84 CFU/m³. Penicillium spp. was detected in all of the samples, while Aspergillus spp. and Cladosporium spp. were each recovered in 50 percent of the samples. Aspergillus niger, A.flavus and A.glaucus were identified, but not all isolates were subject to identification. Also, Sporobolomyces sp. and Gliocephalotrichum sp. were each detected in one sample at very low concentrations.

Bacteria were recovered at concentrations ranging from 95-462 CFU/m³ with a mean concentration of 207 CFU/m³. This level of bacteria is somewhat higher than the other sites, but is not unusual considering the type of sample site and the level of human activity associated with corridors. Bacteria recovered from samples collected in hospital corridors were not identified.

DISCUSSION

Substantial research is needed to develop a database which reflects typical microbial levels in a variety of indoor air environments. Such projects should ideally include repeated sampling at established sites (including outdoor samples), should be conducted throughout the year, and should incorporate a variety of media types to enhance recovery. Microbial data should be correlated with measurement of physical parameters, HVAC system information, incidence of disease and other factors. Development of such a database would allow consultants to begin to make prudent recommendations regarding microbiological quality of indoor air.

We do not routinely recommend sampling indoor air environments for bioaerosols without a thorough site investigation. However, despite the limitations of this data as discussed above, it is clear that facilities managers of hospitals with operating rooms, intensive care units and/or critical care units which are lacking HEPA-quality air filtration would be justified in performing routine sampling in these and other areas of the hospital which house immuno-compromised patients.

We suggest immediate investigation of the HVAC system be triggered if samples collected during static periods in these areas yield more than 100 CFU/m³ of either fungi or bacteria.

At the time of publication, Scott Tighe was the senior microbiologist and Paul Warden was the manager of client services at Analytical Services, Inc., a commercial microbiology laboratory in Williston, VT.

Published in the May 1995 issue of Indoor Air Review. Reprinted by permission of Indoor Environment Review, a division of IAQ Publications.