Bioaerosol Sampling
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arunkdevassyBioaerosol Sampling
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arunkdevassyBIOAEROSOL SAMPLING
TOPICS COVERED:
1. Need for sampling
2. Devices
3. Active sampling devices
4. Assay methods
5. Disadvantages of Colony counting method
Bioaerosol monitoring: It is carried out because of three important reasons
1. to meet legal requirements or to comply with guidelines
2. to collect epidemiological data, possibly with a view to setting occupational
exposure limits
3. For general scientific interest and research.
SAMPLING DEVICES
Samplers must collect a representative sample of the required fraction of the
bioaerosol with the minimum of stress, so that the biological activity of the aerosol
is not too much impaired. There are no standard protocols available for monitoring
bioaerosols. The performance of a sampler designed to collect airborne dusts can
be described in terms of a number of parameters
1. Aspiration efficiency: efficiency with which particles directly enter the
sampler
2. overall efficiency: efficiency by which the particles reach the collection
surface
3. bioefficiency – ability of the sampler to collect microorganisms “alive”. This
will depend on type of microorganism involved.
4. impaction onto solid or semi-solid surfaces, impingement into liquid, and
There are two principle means of monitoring the microbiological population of the
air,
1. Passive monitoring: Passive monitoring is usually done using ‘settle plates’
– standard Petri dishes containing appropriate culture media that are opened
and exposed for a given time and then incubated to allow visible colonies to
develop and be counted
2. Active sampling: Active monitoring requires the use of a microbiological air
sampler to physically draw a known volume of air over, or through, a
particle collection device and there are three main types
Active sampling equipments
Impactor: Impactor samplers use a solid or adhesive medium, such as agar, for
particle collection.
1. Casella Slit Sampler: It samples air at high speed through a narrow slit(s)
onto a rotating nutrient agar plate, and relies on the growth of the
microorganisms for their detection. Since the particles are collected onto a
single agar plate the CSS does not give any indication of size distribution
2. Andersen Microbial Sampler: The Andersen Microbial Sampler (AMS)
(Andersen, t9:58) is a cascade impactor which collects airborne particles
onto a series of nutrient agar plates. It can be used to gain information on the
size distribution within the sampled aerosol. From that we can enumerate the
collection at upper respiratory track, the bronchioles, the aleoveli.The
collection efficiency of the AMS is more affected by wind speed than by
particle size. The collection efficiency fells rapidly at high wind speeds for
higher aerodynamic particle.
3. Biotest RCS Sampler. The Biotest Reuter Centrifugal Aerosol Sampler
(RCS) is a portable hand-held instrument, much used in the biotechnology
industry, which collects particles onto an agar strip and relies on the growth
of microorganisms for their detection. The sampler does not give any
indication of the size of particles collected. An agar strip is inserted into the
drum around the impeller blades. Air is drawn into the sampler at a rate of
40 1 min- 1 by the impeller. Airborne particles are subjected to a centrifugal
acceleration and impacted onto the agar strip at high velocity
4. Spore traps: Particles are impacted on an adhesive-coated transparent plastic
tape supported on a rotating drum. The air sampling rate is 10 1 min- 1
Impingers: Impingers use a liquid medium for particle collection. Typically,
sampled air is drawn by a suction pump through a narrow inlet tube into a small
flask containing the collection medium. This accelerates the air towards the surface
of the collection medium and the flow rate is determined by the diameter of the
inlet tube. When the air hits the surface of the liquid, it changes direction abruptly
and any suspended particles are impinged into the collection liquid. Once the
sampling is complete the collection liquid can be cultured to enumerate viable
microorganisms. E.g. Porton All Glass Impinger (AGI), Multistage Liquid
Impinger. Another important impinger is cyclone impinger
Filter systems: A known volume of air is sent through membrane or gelatin filters.
One of the main disadvantages of using filters to collect microorganisms is that
they afford little protection to cells so the large volumes of air passing over the
filters may cause desiccation. The collected microorganisms can be washed off the
filters for detection or detected directly on the filter using staining techniques and
epifluorescence microscopy.
ASSAY METHODS TESTED & VERIFIED FOR BIOAEROSOLS
1. Colony counting: Collect directly onto agar plate or spread a sample of
collection fluid. The number of colony forming units in a litre of sampled air
is calculated. It is slow &can underestimate total no: of cells. It has been
estimated that culturable counts account for only between 0.0001 and 10% of
the total population within environmental samples. The same bioaerosol
particle collected into a liquid, e.g. in an impinger or cyclone, may break up
to release the cells into the collection fluid so that, when plated onto agar, a
larger number of colonies are formed and a higher CFUI 1 of sampled air is
obtained.
2. ATP-bioluminescence: Enumeration based on luminescent output when ATP
reacts with luciferin.It is fast. It is not specific. Viable count is measured.
3. Fluoresce microscopy: Cells are stained with fluorescent dye & counted with
microscope. Total count is possible
DISADVANTAGES OF COLONY-COUNTING
Sampling can affect the viability of microorganisms due to physical stress, osmotic
shock. The organism species, growth conditions for the culture, method of aerosol
generation, sampler, and the airborne environment. Desiccation, radiation, oxygen,
ozone and its reaction products and various pollutants, described below, can affect
the viability of microorganisms’ .Colony counting therefore underestimates total
count. It has been estimated that culturable counts account for only between 0.0001
and 10% of the total population within environmental samples. The only
information from colony counting is the proportion of the sampled population
which has the ability to form cultures. In a nutrient agar medium one species can
compete and suppress the growth of other species. One possible solution is by
using selective media. Collection liquids also influence the viability and are
frequently based upon simple salt solutions (for example, phosphates) with
additives such as proteins, antifoams, etc., or are complex media, plus antifoams.
Transportation: To maintain collected microorganisms in a stable and viable state,
refrigeration or packing samples in ice may be the best approach, but there may
also be a need to formulate transportation media capable of neutralising toxic by-
products that could decrease viability.
the assayed sample may be less than the "true" value in the aerosol.
The following criteria should be used to determine a sampling strategy:
1. Type of sampling required; i.e. personal or static (area) (the position and
number of static samplers used also need to be considered)
2. Specificity required which microorganism to be monitored
3. Level of sensitivity
4. Speed with which a result is obtained
5. Importance of Total Cell Count vs. viable
6. Particle size range of interest this will depend on choice of sampler
For example for a assessing an agriculture worker exposure you need a sampling
method which measures total count, inhalable. The Total Count is important as
non viable cell can also precipitate allergic reaction. Bioaerosol and general dust
levels are likely to be high and the sampler used must be able to collect without
blocking. For a sewage treatment worker exposure assessment, gram negative
bacteria (e.g. coliforms), gram positive bacteria (e.g. streptococci and most
clostridia), pathogens (e.g. Salmonella and Vibrio) and viruses (e.g. poliomyelitis
and hepatitis) are of most interest.
References
1. J. DOUWES Bioaerosol Health Effects and Exposure Assessment: Progress and
Prospects Ann. occup. Hyg., Vol. 47, No. 3, pp. 187–200, 2003 © 2003 British
Occupational Hygiene Society
2. w. D. Griffiths THE ASSESSMENT OF BIOAEROSOLS: A CRITICAL REVIEW
aerosolscience vol .25 No:8 pp 1425-1458 1994
3. Martı´ Nadal Health risks of the occupational exposure to microbiological and chemical
pollutants in a municipal waste organic fraction treatment plant Int. J. Hyg. Environ.
Health 212 (2009) 661–669
