aero biology

8/10/2019 Aero Biology

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Aerobiology

Microbiology of the Atmosphere


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Aerobiology

The interdisciplinary science that deals with

the movement and dispersal of bioaerosols

The movement of bioaerosols is generallypassive and is greatly influenced by the

environment

The survival of viable bioaerosols is also

dependent on the environmental conditions


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Bioaerosols

Biological agents carried in the air as large

molecules, volatile compounds, single

particles, or clusters of particles that are living

or were released from a living organism

Particles sizes - 0.5m to 100 m

Capable of eliciting diseases that may be

infectious, allergic, or toxigenic with theconditions being acute or chronic


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Bioaerosols in Our Environment

Outdoor Sources

Fungal Spores

Pollen Bacteria

Indoor Sources

Viruses

Bacteria Fungal Spores

Dust mites

Cockroaches Animal Dander -

especially cats


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Outdoor Aerobiology

Atmosphere

Transport

Fungal Spores Pollen


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Spores, Pollen and Gravity

In still airspores (and pollen) fall to the

ground at a rate (based on Stokes law)

that is proportional to the square of itsradius

i.e. the bigger the spore the faster it will fall

Aerodynamics also influenced by

non-spherical shape or irregular shape

ornamentation and aggregation

these increase drag and delay deposition


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Stokes Law

V =2 s r

9

gr2

V = terminal velocity in cm/s

s= density of sphere

r = density of air

g = acceleration due to gravity

= viscosity of air )1.8 x 10-4g/cm/sec

r = radius of sphere


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0.5 - 14.0


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Settling rates of particles


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Airborne Transport


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Effects of

environmental

factorsImpact

Deposition

Dispersal

Take-off

Source

The Aerobiological Pathway


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Airborne Transport

Perfectly still air seldom occurs

Prevailing air currents delay deposition by

gravity

Air flow is complex allowing bioaerosols to be

transported over short ranges to global

distances

Transport occurs in the turbulent layer of the

atmosphere


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Turbulent Layer of Atmosphere

Air movement shifting and unpredictable

Depends on wind speed, direction,temperature, and local eddies caused byroughness of the terrain

Fungal spores are a normal component of

the turbulent layer possibly up to 200,000spores/m3of air; however, pollen levelsnormally two orders of magnitude lower


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Long distance transport

Well documented in palynology literature as

well as in aerobiology literature

Typically these are reports of one time

incursions of exotic pollen types or preseason

pollen

Exoticpollen in sediments in Canada

Preseason birch pollen in Scandinavian countriesfrom southern areas in Europe


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Well studied examples of long

distance transport

Wheat rust sporesStarting in the 1920s the

movement of Pucciniagraminisspores has

been demonstrated

Tobacco blue moldSince late 1980s, themovement of Peronospora tabaccina spores

has been followed

Mountain cedar pollenMy lab has been

following the LDT since 1980

African dust and associated microorganisms

have been studied over past decade


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Juniperus asheipollen

Juniperus ashei (mountain cedar) has a limiteddistribution Arbuckle Mountains of Oklahoma

Edwards Plateau of Texas

Scattered areas of Ozarks

Highly allergenic (cedar fever in Texas) Tulsa allergist reported 15% patient sensitivity

Mid-winter pollination (Dec. and Jan.) Tulsa pollen season Feb through early Nov

Evidence of pollen in Tulsa atmosphere Cedar pollen detected since 1980


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Juniperus asheigrows amongst the dissected

slopes of Cretaceous limestone of the Edwards

Plateau. The species co-occurs withJ. virginiana

to the east and J. pinchotii otherJuniperus spp. to

the west.

Distribution of Juniperus ashei


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Incursion of J. asheipollen into Tulsa

Each winter since 1980 J. asheipollen has

been registered by our Tulsa air samplers

Pollen recorded on 20% to 60% of the days

in Dec and Jan

Concentrations typically low however very

high concentrations have been registered on

several occasions (based on NAB level ofvery high >1500 pollen grains/m3)


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Mountain Cedar Forecasting

Started in Dec 1998

Daily forecasts in Dec and Jan

Forecasts posted on internet athttp://pollen.utulsa.edu

During past 6 season 812 forecasts issued Three sites in TexasAustin, Junction, San

Angelo One site in OklahomaArbuckle Mountains

One site in ArkansasOzark Mountains (5 yrs)
http://pollen.utulsa.edu/http://pollen.utulsa.edu/


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Forecast Components

Release forecastbased on meteorologicalconditions and phenology of the plant Temperatures above 45o F

R.H. below 50%

Sunshine

No rain in previous 24 hours

Pollen cone maturity

Downwind forecast based on model projections

using: HY-SPLIT model trajectories

Meteorological conditions along the path


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University of Tulsa

Mountain Cedar Pollen Forecast

Date Issued: 15 January 2000

Mountain Cedar location(s): Arbuckle Mountains, OK

Regional weather: Friday/Saturday, January 14/15 - TX/OK: High pressure will control

the weather for the weekend. Moist flow off the Gulf of Mexico is causing some clouds

and showers in deep South TX and some variable cloudiness in south-central TX.

Cloudiness will increase today for other parts of southern TX and there may be somedrizzle tonight. Otherwise, fair weather will prevail, and the chance of rain will diminish

for southern TX as the center of the High moves away to the east. Late-night/early

morning clouds in some areas Saturday night are expected. Warming trend through the

weekend. Highs today 50's and 60's, lows tonight 30's to 50's, highs Saturday 60's and

70's.

Trajectory weather: Mostly sunny today, high in the 60's. Partly cloudy tonight, low

near 40. Dry with a high around 60 on Sunday.

Trajectory confidence: High

OUTLOOK: *** Serious Threat*** Conditions are favorable for pollen release.

Strong release expected. Temperatures and humidities are favorable much of the day.

Airborne pollen will be moving into northeast OK (including the Tulsa area) later

this afternoon.... and across southern MO and southern IL tonight. Residents of

western KY may also be affected. TK

T j t St t( ) ( h b th * ) S lf Okl h


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Trajectory Start(s)(shown by the *on map): Sulfur, Oklahoma


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1998-2004 Tulsa Cedar Profile

During Dec and Jan of the 6 yearscedarpollen present on 233 days

Most days lowlevels

Over 50 days moderate to very highlevels 12 days with highlevels

One day had very highlevel (2,109 pollen/m3)

Trajectories crossed Tulsa >150 times with

the majority of trajectories from the ArbuckleMts.


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London, Ontario

Air sampling data from Jim Anderson from

London, Ontario

Juniperuspollen recorded 22 times in Dec

and Jan from 1998-2004

Maximum was 58 pollen grains/m3


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Continental transport

27 Jan 99, Jim Anderson inLondon, Ontario reportedatmospheric Juniperuspollen - 58 pollen grains/m3

Trajectories show that the

source of this pollen wasTexas population ofJuniperusashei

Our Jan 26 forecastindicated that the pollenhas the potential to travelvery long distances.


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Fungal Bioaerosols


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THE FUNGI

Kingdom Protista

Div Myxomycota

Div Oomycota

Kingdom Eumycota

Div Chytridiomycota

Div Zygomycota

Div Ascomycota

Div Basidiomycota

Asexual Fungi


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FUNGAL SPORES

Majority of spore types adapted for airborne

dispersal

Spores unicellular to multicellular from 1 to

100 m

Main sources in the environment

Leaf surfaces (phylloplane fungi)

Soil


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Spore Release Mechanisms

Passive

Wind - generally the most abundant airborne

spores in the atmosphere - include membersof the Dry Air Spora which peak in afternoon

Rain - rain splash as well as tap and puff

Active

Generally require moisture

Common mechanism for ascospores and

basidiospores


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Wind Dispersed Spores

Frequently related to wind speed and

turbulence

Typically borne on erect conidiophores or

sporangiophores that elevate spores above

the substrate

Dry Air Spora Cladosporium, Alternaria,

Epicoccum, Drechslera, Pithomyces,Curvularia, smut spores


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Dry Air Spora


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Diurnal Rhythm of Cladospor ium

Hourly CladosporiumLevels

Tulsa May 24, 1999

0

5,000

10,000

15,000

20,000

25,000

30,000

12:00

AM

2:00

AM

4:00

AM

6:00

AM

8:00

AM

10:00

AM

12:00

PM

2:00 PM 4:00 PM 6:00 PM 8:00 PM 10:00

PM

S

pores/m3


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Cladospor iumspores peak hourly concentration of

>120,000 spores/m3 during a spore plume

Cladosporium ,

30 September 1998

Tulsa

0

20,000

40,000

60,000

80,000

100,000

120,000

140,000

0:00 2:00 4:00 6:00 8:00 10:00 12:00 14:00 16:00 18:00 20:00 22:00

Spores/m3


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Cladospor ium

Common fungal genusoccurring both indoorsand outdoors

Most abundant outdoorspore type with aworldwide distribution

Normally exists as asaprobe or weak plant

pathogen Spores are known to be

allergenic


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Cladospor iumspp.


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Passive Discharge during Rain

Vibration and shaking as raindrops hit leaf

May explain increases in Cladosporium

during rainfall

Release of basidiospores by puffballs shows

similar puffing when raindrop strikes mature

fruiting body


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Rain Splash

Generally spores surrounded by mucilage

Mucilage protects from desiccation but also

prevent dispersal by wind

First raindrops dissolve mucilage

Resulting spore suspension dispersed by

subsequent raindrops

Spores commonly thin, colorless, elongate


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Active Discharge - Ascospores

Hygroscopic material within ascus absorbs

moisture

Ascus swells and develops high osmotic

pressure

Pressure causes ascus to burst, explosively

shooting spores into the atmosphere

Need for rainfall will often override diurnalrhythm of release


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Basidiospore discharge

from basidium

Moisture condense around

crystal of manitol

Bullers drop enlarges

Fuses with a film of water

around spore

Shifts center of gravity

Result of discharge

Spores shot only a fractionof a millimeter

Spores freed from gill or

pore and able to fall free

and reach turbulent layer


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Basidiospore Rhythm

Need for moisture

confines spore release

to periods of highhumidity

Typically peak levels in

pre-dawn hours and

low levels in afternoon

May of 1998

0

500

1000

1500

2000

2500

2:00AM

4:00AM

6:00AM

8:00AM

10:00AM

12:

00PM

2:

00PM

4:

00PM

6:

00PM

8:

00PM

10:

00PM

12:00AM

spores/m

3


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Airborne Fungal Spore

Concentrations in Tulsa 2002

0

10,000

20,000

30,000

40,000

50,000

60,000

J F M A M J J A S O N D

Spores/m3

0

10,000

20,000

30,000

40,000

50,000

60,000


Spores/m3

2002

23 T id tifi d Cl d i A


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23 Taxa identified: Cladospor ium, Ascospores,Basidiospores, and Alternar iaComprised 90% of Total

Basidiospores

0

500

1000

1500

2000

2500

3000

3500

4000

4500

5000

1/1 2/1 3/1 4/1 5/1 6/1 7/1 8/1 9/1 10/1 1 1/1 12/1

spores/m3

Ascospores

0

2000

4000

6000

8000

10000

12000

1/1 2/1 3/1 4/1 5/1 6/1 7/1 8/1 9/1 10/1 11/1 12/1

spores/m

3

Cladospor ium

0

5000

10000

15000

20000

25000

30000

35000

40000

45000

1 /1 2 /1 3 /1 4 /1 5 /1 6 /1 7 /1 8 /1 9 /1 10/1 11 /1 12/1

spores/m3

A l t ernari a

0

500

1000

1500

2000

2500

1/1 2/1 3/1 4/1 5/1 6/1 7/1 8/1 9/1 10/1 11/1 12/1

spores/m3


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Indoor Aerobiology

Outdoor spores enter readily

Many fungi can also amplify indoors

anytime moisture is available fungi can grow

on many indoor substrates

Penicillium, Aspergillus,andCladosporium

are most common indoors

Many can form mycotoxinsAspergillussppand Stachybotrys

E i t l f t th t i fl


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Environmental factors that influence

indoor fungal contamination

Outdoor concentration and type

Type and rate of ventilation

Activity levels Indoor moisture levels

Modern building materials


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Typical Yearly Spore Levels 1998

0

10,000

20,000

30,000

40,000

50,000

60,000



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Spore plumesshow the influence of

environmental conditions

Total Spore Concentration and Wind Speed

Hectorville, OK - Sept 21, 1998

0

20000

40000

60000

80000

100000

120000

140000

160000

180000

200000

12:00

a.m.

2:00

a.m.

4:00

a.m.

6:00

a.m.

8:00

a.m.

10:00

a.m.

12:00

p.m.

2:00

p.m.

4:00

p.m.

6:00

p.m.

8:00

p.m.

10:00

p.m.

Time

Tota

lSporeConcentration

s

(spores/m^3)

0

1

2

3

4

5

6

7

WindSpeed(m/s)


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Water Leak


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Humidity

Indoor relative humidity Below 30% R.H. no mold growth

Above 70% optimal for mold

Usually mold growth can occur above 50% Humid air condenses on cool surfaces

Cold windows in winter - molding and sills become wetand suitable for fungal growth

Cold floors in winter

Cooling coils in AC units in summer Humid air allows hygroscopic materials to absorb

water

Hi h h idit i h h


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High humidity in home where

subslab ducts failed


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Moisture Problems

Become worse in past 20 - 30 yrs

increased use of washing machines, dishwashers

vaporizers and humidifiers actively spray droplets

into the air (often contaminated)

tighter buildings for energy conservation trap

moisture

Anytime moisture available fungi will grow


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Cladosporium one of the most common

indoor fungi growing on a diffuser


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Penic i l l ium

One of the mostcommon soil fungi innatural environment

Over 250 species

Well known allergen

Some species producemycotoxins

Some species produceantibiotics

Produce VOCs


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Penic i l l ium in culture


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Aspergi l lus

Also common soil fungi

Cause rot of stored grain

Over 150 species

Well known allergens

Several species form

mycotoxins

Some species can grow at

high temperatures

Several species causeinfections in lung, sinuses,

and hypersensitivity

pneumonitis


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Aspergi l lus niger


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Asperg i llus fum igatus


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Penic i ll ium and Asperg i llus

Small spores passively

aerosolized when spore

clusters disturbed

Spores extremely buoyant,remain airborne for

extended time

PenicilliumandAspergillus

spores look alikedistinguished in culture


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Stachybotrys chartarum

Soil fungus in nature

Commonly found indoors on

wet materials containing

cellulose, such as

wallboard, jute, wicker,straw baskets, and paper

materials

Spores in slimy mass

Thought to be allergenicalthough little is known

May produce potent

mycotoxins


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Indoor air sample


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Air SamplingTechniques


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Principal Collection Methods

Gravity

Impaction

Impingement

Filtration


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Gravity

Simplest but least accurate method ofcollecting airborne biological samples

Coated microscope slide or open petri dish

containing agar exposed to atmosphere Non quantitative for atmospheric

concentrations

Affected by particle size and shape and airmovement

Biased deposition of large particles


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IMPACTIONSAMPLERS

Separate particles from the air stream by

using inertia of particles

Forces deposition onto a solid or agar

surface

Most commonly used method

Instruments available for culturable sampling

and total spore sampling

Impaction Samplers


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Impaction SamplersTotal Spore Samplers

Outdoor Samplers

Hirst Spore TrapBurkard and others

Rotorod Sampler

Samplair MK-3 Sampler

Indoor Samplers

Burkard Continuous Recording Air Sampler

Samplair MK-3 Sampler

Burkard Personal Volumetric Sampler

Air-O-Cell Sampler

Cyclex-D Sampler


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Burkard Spore Trap


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Advantages of Burkard Spore Trap

High efficiency down to less than 5 m

Allows for accuracy for small fungal spores such

as basidiospores and small ascospores

Time discrimination Permits analysis for diurnal rhythms

Permanent slides for future reference


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Burkard 7-day sampler head

Standard is the 7-day sampling head

Sampler drum mounted on 7-day clock

Drum moves by orifice at 2 mm per hr

Melenex tape mounted on drum and greased(Lubriseal, High Vacuum Grease, other)

Air is brought in at 10 l/min and impacts on greased

Melenex tape

Drum changed each week


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Seven Day Sampling Head


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Processing the 7-day drum

Melenex tape removed from drum

Tape cut into seven 24 hour segments each 48 mmlong

Segments mounted on microscope slides in 10%gelvatol (polyvinyl alcohol) and dried

Glycerin-jelly mounting medium added and a 50 mmcover slip

Mounting medium contains pollen stain - either basicfuchsin or phenosafarin


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Melenex tape on cutting board


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24 hour sampling head

Outdoor Air Sample from Burkard Spore Trap


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Outdoor Air Sample from Burkard Spore Trap


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Rotorod Samplers


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Rotorod Samplers

Models most often used have retracting rodsfor intermittent operation (10% sampling timetypical)

Head rotates at 2400 rpm Leading edge of rod coated with grease

Pollen and spores impacted on greasedsurface

Efficient for pollen and spores >10 m


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Rotorod Sampler

Rods spin at 2400 rpm. Particles (spores and pollen) are impacted on

leading face of the rod which is greased. Efficiency decreases dramatically

below 10 m. Intermittent head spins 10% of time.


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Allergenco Samplair


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Continuous Recording Burkard

24 hour sampling onto a coated microscope slide


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Burkard Personal Sampler


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Air-O-Cell Slide


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Cyclex-d

Andersen (N-6) Single Stage


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( ) g g

Sampler


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Andersen 6-Stage and 2-Stage Impactors


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Biocassette Sampler

Burkard Portable Air Sampler for Agar


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Burkard Portable Air Sampler for Agar

Plates


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Inpingement Samplers

Separate particles from the air stream by

using inertia of particles

Forces deposition into liquid collection

medium (usually a dilute buffer)

Aggregates of cells can be broken apart

Allows for several possible analytic

applications: culture, microscopy,biochemistry, immunoassays, PCR


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Impingers

AGI-30 BioSamplerBurkard

Multi-stage


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Filtration

Separates particles from airstream by

passage through a porous substrate, usually

a membrane filter

Collection depends on filter pore size andflow rate

Loss of viability may occur due to desiccation

Adaptable for a variety of assays


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Sampler Performance

Collection efficiency is the ability to capture

the particles onto the collection medium

Physical characteristics of the inlet and the

airflow rate used to calculate d50 Particle diameter at which 50% of particles are

collected

Because of sharp cut-off it is generally acceptedthat all particles above this size are collected

d50 values for some samplers (CUT SIZE)


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d50values for some samplers (CUT SIZE)

Sampler d50

AGI-30 Impinger 0.30 m

Air-O-Cell Cassette 2.30 m

Andersen Single Stage 0.65 m

BioSampler Impinger 0.30 m

Burkard 7-Day Spore Trap 3.70 m

Burkard Personal Sampler 2.52 m

Buttner MP, Willeke K, Grinshpun SA. Sampling and Analysis of Airborne

Microorganisms. In: Manual of Environmental Microbiology 2nded. ASM Press,

Washington, 2002.


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Analytic Methods

Microscopy

Culture

Biochemistry Molecular Biology


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Microscopy

Identification of total spores (both culturable

and non-culturable) along with pollen and

other particulates

Identification to species level usually notpossible

Identification of morphologically similar

spores not possible


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Culture

Only viable bacteria and fungi

Limited to those taxa able to grow in culture

on medium used

Success based on medium and incubation

time and temperature

Permits speciation when required

Results expressed as CFU/m3


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Biochemistry

Detection of specific compounds

May not be specific for a genus or species

Assay examples:

Ergosterol

B 1,3-glucan

Endotoxins

MycotoxinsStachybotrys toxins and ochratoxin

Various types of assays such as HPLC

I h i


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Immunochemistry

Specific allergen molecules

Not useful for routine air sampling

Typically used for filter samples and impinger

samples but have been used for spore trapsamples

Requires prior development of an antibody

Widely used for dust mite, cockroach, cat,etc.

Few fungal assays available

M l l Bi l


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Molecular Biology

Detection of specific genetic elements

Highly specific and sensitive

Currently restricted to a few organisms

aero biology

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