Building Services Management – Air Sampling - “Smells moldy in here”: The basics of air sampling.
Jason K. Dobranic, Ph.D.
Maintenance professionals are often the first responders to mold complaints in their facilities. building occupants can be exposed to fungal (mold) spores through direct contact with contaminated uilding materials and contents. More commonly, exposure occurs by inhalation of spores found in reathable air spaces. Air monitoring or sampling may be a valuable tool to determine exposure levels to mold spores by the building’s occupants. In most circumstances, indoor air quality specialists will be alled in to perform the air sampling.
Air sampling methods can be divided into two different categories: methods generating “total spore counts,” typically reported as counts per cubic meter (m3) and methods that determine “culturable fungi,” typically reported as colony forming units (CFU) per cubic meter. Total spores counts are an amalgamate of both living and nonliving spores. These methods do not allow for the discrimination between living or dead spores. Culturable fungi results, tabulate only living fungal particles, which can include hyphal fragments as well as spores.
Sampling for total spore counts can be carried out using specially-greased slides and a Burkard or Allergenco sampler. Initial costs for these devices are high but subsequent purchases of greased slides are relatively inexpensive. The principal behind each of these devices is the same; air is vacuumed through and condensed onto the greased slide. Particles in the air are forced onto the grease, become embedded, and stick to its surface.
A more common approach would be to use your own vacuum pump with specially-designed, single-use spore-trap cassettes. Some of the popular cassettes on the market include the Cyclex-d, Air-O-Cell, Microcell5, and the Laro100. Particles in the air are trapped in the cassette either on a proprietary adhesive or, as in the case of the Laro100, a 0.8-micron (mm) filter. The cassettes are opened, prepped and examined under a microscope in a laboratory by properly trained analysts who determine the total spore count. Depending upon your specific complaint or scenario, it may not be important to know the viability of a spore, since dead spores are also potentially allergenic or toxigenic to susceptible individuals.
Culturable, or viable fungi, are those fungi that are able to grow on laboratory media, under laboratory-set conditions. Laboratory media are composed of agar, a gelatin-like substance derived from algae, and a nutrient source, such as malt extract. In order for the fungus to grow, it must develop from a viable spore or fragment of hyphae. Hyphae are tubular or filamentous structures that make up the body of the fungus. Since it is impossible to determine if the colony grew specifically from a spore or a hyphal fragment, it is reported as a colony-forming unit. However, not all fungi are able to grow on all laboratory media, so it is important to communicate with an experienced laboratory who can recommend the most suitable media for your project. Typically, malt extract agar (MEA) is recommended as a general isolating medium, whereas, cornmeal agar or cellulose agar are often recommended when Stachybotrys chartarum is of primary concern. Dozens of other specialized media are also available.
You can sample for culturable fungi using an Andersen impactor, SAS sampler, RCS Biotest sampler, or an all-glass impinger. The Andersen impactor and SAS sampler are similar devices in that both use agar plates. Air is vacuumed through a sieve-like cover thereby forcing particles to adhere to the agar’s surface. The Andersen impactor requires an external vacuum pump whereas, the SAS sampler is a small self-contained, portable unit. The RCS Biotest sampler works differently by using centrifugal force and a specially-made agar strip. Air is pulled through and spun around causing particles suspended in it to collide onto a strip of agar media. The all-glass impinger is, as the name implies, a device composed of glass. It works by vacuuming air on a tangent through a liquid medium causing it to vortex and particles become trapped in the liquid. This liquid can then be poured or mixed with an agar medium to check for fungal growth. These fungal cultures must be incubated (grown) for a period of time in the laboratory; typically 6-10 days before being analyzed. If the aerosolized particles include viable fungal spores or hyphal fragments, and the media is suitable for a particular mold, then they will grow into a colony. If you utilize one type of media, you should realize that you are likely to underestimate the total amount of culturable fungi in the air.
Many investigators choose to sample on several media in order to maximize the identification of all possible culturable fungi.
All of the above mentioned methods has its pitfalls, thus effecting the quality of the data reported. One common case is overloading of samples. In the case of spore traps, overloading occurs when air with a heavy saturation of particles is sampled for a long period. Inside the cassette, these particles are concentrated until they become compacted, one on top of the other. It is difficult for the laboratory analyst to count spores through this heavy background level. For culture-based methods, overloading occurs when a large quantity of air is sampled resulting in many fungal particles impacting on the media. This leads to a high concentration of colony forming units growing on the plate in close proximity to each other and makes it difficult to accurately count single colonies.
Whatever method you use, stick to the manufacturers’ recommended sampling rates while adjusting the total volume. The total volume is adjusted by varying the sampling time. Depending on the conditions under which you are sampling; time adjustments may be required when the air is visibly dusty or when sampling between walls (wall checks). The investigator’s sampling protocol is crucial in the ultimate reliability of the results.
These protocols should include calibration of sampling equipment, aseptic technique, properly packaging samples for shipment to the laboratory, and accurately filling out a chain-of-custody. “Aseptic technique” is a microbiology term referring to general techniques that help minimize the chance of contaminating the sample with microbes from sources other than from the test source (either air or surface). Proper protocols should reduce the probability of cross-contaminating samples. Destructive surface sampling techniques should not be performed before air sampling, because spores will be liberated into the air, thereby artificially inflating air spore counts. Contaminants may also be introduced from the environment, equipment, supplies, previous samples, and personnel. Wiping down sampling equipment with alcohol pads is effective in reducing cross-contamination through equipment. A great way to monitor your sampling technique is to use field blanks or negative controls. For example, during the normal course of air sampling for a project, set-up your equipment with sampling medium or cassette, but stop right before the step where you would turn the vacuum on.
Remove the media or cassette and ship to the laboratory. Results from a field blank should be negative for growth and spores or you might have a cross-contamination issue. Air sampling is not a complicated endeavor, but there are enough small details that must be carried out properly, or your samples will be compromised. Unknowingly using compromised results to solve your IAQ problems will only lead to compounded problems and wasted money. I hope this primer gives everyone some insight on the ins and outs of air sampling.