MICROORGANISMS

K C C A P U L O N G | Z H E R R E R A | K D I N F A N T E

I C P A C I A | J P S E V I L L A

M C B 1 5 0 M I C R O B I A L E C O L O G Y

AIRIN

N

OBJECTIVES:

• Identify different microorganisms present in the air;

• Understand and demonstrate the sedimentation procedures, the gravity plate method and impingement technique;

• Compare the number and types of airborne microorganisms present in the intramural and extramural environments;

• Learn to compute for the determination of the microbial load present in the air using the formulas given.

GRAVITY PLATE METHOD• also called as settle plate

method

• plates were exposed in the assigned areas for 15 minutes

• microorganisms were collected by sedimentation or by gravity to deposit air particles (microorganisms) on the plates

ADVANTAGES

inexpensive and easily performed

very useful for directly monitoring airborne contamination of specific surfaces

Useful for qualitative analysis of airborne microorganisms

very limited since this method is only capable of monitoring viable biological particles that sediment into air and settle at time of exposure

it will not detect smaller particles or droplets suspended in the air and it cannot sample specific volumes of air

vulnerable to interference and contamination from non-airborne sources and growth medium might deteriorate if exposed for too long

microorganisms may easily become overgrown in heavily contaminated conditions

DISADVANTAGES

The laboratory is the area expected to have the most number of microorganisms since microorganisms in

air are most likely to spread in areas where pathogens are handled for research purpose (Pepper et al.,

2014).

IMPINGEMENT METHOD

When air is dispersed in the liquid, particles in the air  are trapped.

The usual volume of collection medium is 20 mL and the typical sampling duration is approximately 20 minutes which prevents evaporation during the sampling of warm climates or freezing of the liquid medium when sampling at lower temperatures.

Quantification of airborne microorganisms is accomplished by plating the collection fluid.

A suitable collecting medium for liquid impingement samplers must preserve the viability of the microorganism while inhibiting its multiplication.

ADVANTAGES

gives quantitative results where sample volume can be calculated using the flow rate and sampling timemore accurate representation of present microorganisms in air will be obtained

DISADVANTAGES

the technique may damage some microbial cells and affect viability

no particle size discrimination which prevents accurate characterization of the sizes of the airborne particles that are collected, and at overlong sampling times it allow multiplication of cells in the liquid medium

DISCUSSIONS

More bacterial air spores grew on PCA compared to that of the NA plate mainly because PCA has glucose, making it richer in terms of nutrient content and carbon source. (In samples obtained from lab and rooftop).

The glass beads acted as dispersants. They were used to prevent bubble formation which could have affected the microbial cell diffusion. It breaks aggregates of microorganisms thus uniformly releasing them into the solution and further trap the microorganisms in the solution because of its pores.

RESULTS

SITE IMPINGEMENT TECHNIQUE GRAVITY PLATE METHOD

bacteria (PCA) fungi (PDA) NA bacteria(PCA)

fungi(PDA)

100 10-1 100 10-1

Lab 21,68 52,81 0,1 1,1 58,55 68,29 0,4

Street 86,121 122,60 1,3 2,0 50,49 32,28 5,13

Rooftop 86,144 65,spreader

2,0 1,0 17,15 26,34 1,3

Table 3.1. Bacterial and fungal counts recorded from air sampled in different environments.

Table 3.2. Computed microbial load (cfu / ft3 air) for the various environments sampled.

* sampling time= 20 mins; ft3 of air sampled = 7.1429; Vi= 50mL

SITE cfu / mL of impingement liquid

cfu / ft3 air*

bacteria fungi bacteria fungi

Lab 1.68 x 10^3 <100 ESPC 1.18 X 10^4 <700

Street 1.77 x 10^3 <100 ESPC 1.24 X 10^4 <700

Rooftop 1.40 x 10^3 <100 ESPC 9.80 x 10^3 <700

RESULTS

Table 3.3. Cultural and morphological characteristics of predominant bacteria and fungi present in air sampled from different environments.

SITE DOMINANT BACTERIA DOMINANT FUNGI

cultural(PCA)

microscopic(OIO)

cultural(PDA)

microscopic(HPO)

Lab Circular, white, flat, entire margination

G(-), cocci cottony, circular, white

filamentous

Street White colony (spreader), opaque

G(-), long rods, in chains

Green mold, cottony, compact

conidia borne on a vesicle, septate, possibly Aspergillus

Rooftop circular, entire margin, flat, yellow-pigmented colony

G(+), circular shape

a) cottony, filamentous form and margin, white

club-shaped conidiophore, septate, possibly Aspergillus

RESULTS

COMPUTATIONS

cfu / ft3 air

 

Given:

sampling time = 20min

capacity of limiting orifice = 10 L / min

volume of impingement liquid after sampling = 50mL

Number of liters per sample = [sampling time (in minutes)] x [capacity of limiting orifice]

= 20min x 10 L / min

= 200 L

Cubic feet of air sample = liters of sample / 28

= 200 L / 28

= 7. 1429 ft3

FOR BACTERIA

CFU per cubic feet of air =

N= CFU/ml of impingement

Vi = 50ml

Va = 7. 1429 ft3

= 1.18

= 1.24

= 9

Laboratory

Street

Rooftop

FOR FUNGI

𝐶𝐹𝑈𝑐𝑢𝑏𝑖𝑐 𝑓𝑒𝑒𝑡 𝑜𝑓 𝑎𝑖𝑟

=¿100(50𝑚𝑙 )7.1429 ft 3

¿<700

𝐶𝐹𝑈=¿100𝐸𝑆𝑃𝐶

REFERENCES

Compendium of Methods for the Microbiological Examination of Foods (Frances Pouch Downes, Keith Ito) American Public Health Association, Apr 1, 2001

Food Quality Magazine: Air Sampling 101. Judie Buddemeyer. June/July 2005 issue. http://www.foodquality.com/details/article/878155/Air_Sampling_101.html?tzcheck=1

Pepper, I.L., Gerba, C.P. and Gentry, T.J. 2014. Environmental Microbiology. 3rd edition. Londom: Elsevier Inc.