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Sampling For Airborne Fungal Spores

Reasons for Conducting Air Sampling For Fungal Spores

Air sampling pump
Air sampling

Sampling for airborne fungal spores may be conducted for a number of reasons including assessing the air quality, detection of pathogenic organisms, epidemiological surveys and predictions, detection of mycotoxin producing fungi, or detection and measurement of exposure to allergenic fungi and actinomycetes. Before sampling for airborne fungal spores it’s important to have a clear objective and the data that would meet that objective. Some of these objectives require counting and identification of the airborne fungal particulates. A well calibrated pump is therefore required. The method of sampling, the equipment and the media are determined by the data required. For example if one is interested in species identification, then they would collect culturable air samples using an Andersen or RCS sampler.

Methods for Sampling for Airborne Fungal Spores

Culturable Air Samples

If the air was sampled by impaction onto a growth media, then the samples are first incubated at suitable temperature followed by counting and identification of resulting colonies (if any). A number of factors may affect the reliability of the colony counts data. These include the suitability of the agar media used for sampling, colony density, and presence of fast growing isolates. Some agar media are highly selective and therefore would not suitable when the objective was to determine the diversity of fungal species in the air. If the colony density is very high, then counting becomes difficult. In the presence of fast growing isolates, slow growing isolates are masked and hence difficult to count.

Non-culturable Air Samples

The second method of air sampling is where the air is impacted on inert medium, usually a filter membrane or adhesive-coated slide. The collection medium is then analyzed by transmitted light microscopy (popularly known as direct microscopic examination), typically at 600–1000 × magnification. A number of different collection devices may be used for spore trap sampling of which the most common are slit or circular impactors such as the Air-O-Cell®, Allergenco D, Micro 5, Cyclex D and other similar cassettes. Mixed cellulose ester membrane (MCEM) filters are also occasionally used.

Choose Your Spore Trap Analytical Labs Wisely

Identification of airborne fungal spores to species can only be accomplished by an experienced mycologist. Similarly, spore trap analysis requires considerable skill and experience on the part of the analyst to identify spores accurately, and to differentiate them from other airborne particulate. In most cases identification of airborne fungal spores by direct microscopic examination is limited to genus. In some cases identification of spores to specific genus is difficult. A good spore trap analyst requires several years of experience. A recent multi-laboratory comparative study of spore trap analyses found that only 75% of commercial accredited labs could consistently identify correctly Cladosporium spores and only 50% could identify Aspergillus/Penicillium-like spores consistently. The level of experience required for species identification or spore trap analysis cannot be attained in a short time, and it cannot easily be acquired by individuals lacking advanced training in mycology or plant pathology.

Mold & Bacteria Consulting Laboratories is accredited by CALA to the Internationally recognized ISO/IEC 17025:2005. 

How To Take Bacteria Samples

Sampling Air For Bacteria
Bacteria air samples are taken by impacting air onto some suitable growth media. Two of the commonly used samplers are Andersen (N6 Single Stage) and Reuter Centrifugal sampler (RCS). The Andersen sampler uses petri-dishes while the RCS uses agar strips. For environmental bacteria, a general purpose media such as Tryptic (Trypticase) Soy Agar (TSA) can be used.

Sampling Surfaces For Bacteria
Sterile cotton swabs can be used to sample for bacteria contaminating surfaces. The swabs are then sent immediately to a laboratory for culturing and identification of the bacteria to genus or species.

Taking Air Samples For Mold Testing: Settle Plate Method

A number of methods can be used to test air for mold or other microbial contamination. One of the oldest methods of testing air for microbial contamination is the settle plates method. Though the method is semi-quantitative, it is still considered a useful method. In industries such as food, pharmaceutical and cosmetics the method is used to assess the likely number of microorganisms depositing onto the product or surface in a given time. The method involves opening and exposing petri dishes containing agar medium suitable for growth of microorganisms of interest. If one is interested in testing for mold, agar plates containing malt extract agar (MEA) supplemented with some antibiotics to suppress bacterial growth would be used. The agar plates are left open at table-top level at selected points in the room for half-hour to 4 hours. This allows mold spores and fragments to settle onto agar media by gravity. Mold test kits (involving growth media) are settle plates.

Settle Plates Results

The number of microorganisms deposited onto the agar surface of the plate over the period of exposure is determined by incubation of the agar plates at 25ºC for 5- 7 days and counting colonies that develop. The results can be expressed as number of colony forming units (CFUs) per unit time. The counted colonies can then be further characterised to genera or species. Higher numbers of CFUs and/or presence of potential pathogenic or toxigenic molds such Aspergillus fumigatus and Stachybotyrs chartarum are indicators of a problem.

Disadvantages of Settle Plates

Settle plate method is an extremely useful method for assessing air contamination by microorganisms. It is easy to conduct and very cost effective. However, only viable microorganisms would be detected by this method and hence it may give a false impression that the air is “clean” if most of the airborne microorgainisms are dead. False negatives may also be obtained from buildings with:

  • very restricted mold growths.
  • very still air in undisturbed rooms.
  • species of poorly culturable molds (e.g., Stachybotrys chartarum).
  • molds consisting of species with poor airborne dissemination (e.g., Aureobasidium on windowsills, Cladosporium on painted cold air vents, Fusarium and many other wet-spored fungi).

Indoor Air Quality: Testing for Mold Contamination in Buildings

Testing for mold contamination in buildings may involve collection and analysis of the following types of samples:

Air Samples

Air samples may be collected for total spore/hyphal fragment count (non-culture analysis) or for culture analysis. The analysis includes enumeration and identification of significant species. Both non-culture and culture analyses have some limitations. For example with non-culture analysis, identification of mold spores to species is difficult and for culture analysis only the viable spores/hyphal fragments are detected. In some situations, a combination of the 2 types of samples is recommended.

Tape, Bulk, Swab, Dust Samples for Direct Microscopic and Culture Analysis

Tape, swab, and dust samples are taken from surfaces suspected of mold contamination. For bulk, a piece building material such as a piece of carpet, drywall, insulation material or wood is collected. The analyses of these samples by direct microscopy allows identification to genus but experienced mycologists may identify some molds to species.

If identification to species is required, the samples are cultured in some suitable media. However, as was mentioned for air samples, only viable mold would grow in media. It is therefore recommended that both direct microscopy and culturing are used when identification of species is required.

For details on types of mold testing, click Services.

Indoor Mould: What are Mould Spores?

Chaetomium and Penicillium/Aspergillus spores from indoor airMould spores are tiny structures produced by moulds for the purpose of propagation. The term spore is general. There specific terms used for spores depending on how they are produced or the group of fungi producing them. Ascospores for example are spores produced by a group of fungi called Ascomycetes while Basidiospores are spores produced by Basidiomycetes which include the mushrooms.

How would spores affect human health?
Spores are tiny (range between 2-100 micrometers) and therefore are easily inhaled into the lungs. Susceptible individuals react to the protein component of the cell wall of the spores. Some moulds such as Aspergillus fumigatus may also grow in the lungs causing what is referred to as Aspergillosis.

How would you tell if you are inhaling potentially harmful levels of spore concentration?
The way to tell if occupants of a building were inhaling potentially harmful levels of spore concentration is to take air samples. Air may be taken to be analysed by culture methods or by direct microscopic examination. Culture methods may significantly underestimate the total airborne mould concentration in the air because only the viable spores or fragments of the mould can be detected in culture. The spores/fragments may also not be detected if the media used for culturing were not suitable for the types of moulds present in the air. Samples taken for direct microscopic examination (also referred to as non-viable air samples) allow the analyst to count all the spores and fragments regardless of whether the spores were viable or dead. Since susceptible individuals can also react to dead spores, non-viable samples would be the best to give an idea of whether occupants were inhaling potentially hazardous levels of mould spores.

Ulocladium and Stachybotrys spores from indoor airThe pictures show spores trapped from highly contaminated indoor air. The first picture shows spores of Chaetomium and Aspergillus/Penicillium. The second photo shows Ulocladium spores and Stachybotrys spores. Chaetomium, Stachybotrys and Ulocladium species are indicators of serious water damage. Spores of these moulds and those of Aspergillus and Penicillium pose inhalation risk.

Do you have a mould question? Send it to us at Contact Us.

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