FIELD OF THE INVENTION
The present invention relates to detecting microbes. More particularly, the present
invention relates to a system and a method for detecting airborne microbes in an
enclosed chamber.
BACKGROUND OF THE INVENTION
Micro-organisms are omnipresent, in the air, water, soil, in and on living
organisms. Micro-organisms are known to survive extreme conditions like hot
springs and frigid climate. They can survive in acidic and alkaline environment.
Under unfavourable conditions of temperature and water, some of them form hard
and tough coverings called as cysts. As and when the favourable conditions return,
they break open their cysts and continue their normal life cycles. The term
"microbes" or "micro-organisms" used herein mean the same thing and can be
used interchangeably. Also, the term "microbe" used hereinafter may refer to
bacteria, fungi or viruses.
Microbes are responsible for numerous maladies in mankind, for example,
Escherichia. coli, which causes watery diarrhoea, abdominal cramps and
vomiting; Staphylococcus aureus, which causes sudden onset of severe nausea
and vomiting, abdominal cramps, diarrhoea and fever; Salmonella typhimurium,
which causes diarrhoea, fever, abdominal cramps, and vomiting; and Salmonella
typhi which causes typhoid, diarrhoea, fever, abdominal cramps and vomiting.
These disease causing bacterial strains are widely present in bathrooms, toilets, air
and on floors and walls of living areas. Consequently, there is a need to accurately
determine the number of microbes in enclosed spaces as a means to monitor the
cleanliness in said spaces, microbial air pollution and like purposes. There exists a
wealth of information in patent and non-patent literature pertaining to the methods
for determining the microbial load or density in the environment or enclosed
spaces.
( h~nesep atent C'h 1005 1971I ( discloses an apparatus which sucks and filters
airborne microorganisms. The filtered microorganisms were collected and
identified using microbiological techniques.
Further, European patent EP 0450850 B1 discloses a method for determining air
borne bacteria which comprises successive steps of passing a volume of air
through a membrane filter to collect air borne bacteria on the surface of said
membrane filter, incubating any bacteria collected on said filter; and measuring
the number of the colonies which may develop on the surface of said membrane
filter.
Still further, PCT publication WO 2008105893 A2 discloses a system and method
for detecting airborne or waterborne particles and classifying the detected
particles. The invention comprises a optical system for measuring an individual
particle size; a second optical system to detect a UV laser-induced intrinsic
fluorescence signal from an individual particle, a data recording format for
assigning both particle size and fluorescence intensity to an individual particle and
computer readable program code for differentiatingmicrobes from nonmicrobes
(e.g. inert dust particles).
Yet another disclosure, US patent US 5766958 A describes a method and device
for detecting airborne, infectious inicroorganisms in indoor air and collecting
them for rapid identification. Air containing microorganisms is drawn into an
enclosed chamber where it is percolated through a liquid such that many of the
microorganisms become encapsulated in the liquid. The air is released into the
room, while the microorganism-containing liquid is directed to a reservoir from
which samples may be extracted for analysis.
Likewise, United States publication US 20040171 137 A1 discloses a method and
apparatus for the detection of microbes in liquids, in air and on non-living surfaces
in which samples are exposed to electromagnetic radiation of specific energies
capable of exciting various metabolites within the microbial cells to be sampled.
The m~crobtal content of the 5amplc IS measured h! analyrlng tluorescencc
signais from the sample.
The inventors of the present invention sought to develop a comparable system and
method for detecting and determining microbes present in an enclosed space and
ambient air. Remarkably, they discovered that recovery of micro-organisms from
ambient air, according to methods discussed in the prior art, did not co-relate well
with their actual numbers present therein. Hence, the prior art devices or systems
lacked the ability to accurately detect and determine microbes in an enclosed
space. In order to overcome this drawback, they have developed a novel system
and a method that recovers microbes from ambient air in greater numbers than
those recovered by devices and methods revealed in prior art.
SUMMARY OF THE INVENTION
The present invention is broadly related to a system and a method for detecting
airborne microbes in an enclosed chamber.
In accordance with an embodiment of the invention, there is provided a system for
detecting airborne microbes in an enclosed chamber, comprising:
(a) an enclosed chamber 10;
(b) an air suction port 20 on the enclosed chamber 10;
(c) a filter 45 removably attached with the air suction port 20, wherein the
filter isolates the interior of the chamber from the external
environment;
(d) a pumping means 15 operatively associated with the interior of the
enclosed chamber for sucking air out of the enclosed chamber through
the filter 45;
(e) a means 30 for sanitization;
(f) at least one sealable opening in the enclosed chamber 10;
(g) a means 55 for dispersing microbes;
(h) a means 60 for dispersing a composition;
(i) a means 50 for dispersing humectant; and
Cj) a means 35 for circulating air inside the enclosed chamber 10;
wherein the means 55 for dispersing microbes, the means 60 for dispersing the
composition, the means 50 for dispersing humectant and the means 35 for
circulating air inside the enclosed chamber can be removably introduced into the
enclosed chamber 10 through the at least one sealable opening.
In accordance with an embodiment of the invention a conduit means 40
operatively connects the pumping means 15 to the interior of the enclosed
chamber.
Preferably, the filter 45 useful for the present invention is a filter paper or a
membrane filter.
Further, the composition may be an air-freshener, an insect repellent, an
insecticide, a deodorant, an essential oil, a fragrance, an aromatic substance, or a
sublimable composition.
Still further, the humectant is selected from the group consisting of water,
glycerine and combinations thereof. Preferably, the humectant comprises water
and glycerine in ratios ranging fiom 1 : 100 to 100: 1 and more preferably in a ratio
of 1:3.
Preferably, the microbes that may be detected and determined by the system of the
present invention are selected from the group consisting of bacteria, hngi and
viruses.
In accordance with another embodiment of the invention there is provided a
method for detecting airborne microbes in an enclosed chamber, comprising the
steps of:
(a) sanitizing an enclosed chamber;
(b) measuring a blank reading;
(c) dispersing a pre-calculated quantity of microbes into the enclosed
chamber;
(d) dispersing a pre-calculated quantity of humectant into the enclosed
chamber;
(e) dispersing a pre-calculated quantity of a composition into the enclosed
chamber;
(f) mixing the microbes, the humectant and the composition for a precalculated
period of time;
(g) sucking the air in the enclosed chamber through a filter;
(h) placing the filter in a culture medium and incubating;
(i) counting surviving microbe colonies after incubation; and
(j) calculating log reduction value and percentage efficacy of the
composition.
BRIEF DESCRIPTION OF DRAWINGS
Figure 1. is a top schematic view of a system for determining and detecting
airborne microbes according to an exemplary embodiment of the invention.
The drawing is illustrative and not intended to be limiting, but is an example of
the preferred embodiment, simplified for explanatory purposes, and not drawn to
scale.
DETAILED DESCRIPTION OF THE INVENTION
Discussed below are some representative embodiments of the current invention.
The invention in its broader aspects is not limited to the specific details and
representative methods. The illustrative examples are described in this section in
connection with the embodiments and methods provided. The invention according
to its various aspects is particularly pointed out and distinctly claimed in the
attached claims read in view of this specification, and appropriate equivalents.
It is to be noted that, as used in the specification and the appended claims, the
singular forms "a," "an," and "the" include plural referents unless the context
clearly dictates otherwise. Thus, for example, reference to a composition
containing "a compound" includes a mixture of two or more compounds. It should
also be noted that the term "or" is generally employed in its sense including
"and/or" unless the content clearly dictates otherwise.
In the following description, the system of the present invention will be described
in reference to a preferred embodiment. This embodiment is merely illustrative,
and is not meant to be limiting.
Figure 1 is a drawing showing an example of a system of the present invention.
The system consists of an enclosed chamber 10 and an air suction port 20 on the
enclosed chamber.
A filter 45 is removably attached with the air suction port 20, wherein the filter 45
isolates the interior of the chamber from the external environment, i.e. prevents
microbes from passing through it while allowing free flow of air. The filter may
be a membrane filter or a filter paper. The membrane filter is a film made up of
nitrocellulose, cellulose acetate, polyether sulfone, polycarbonate or the like and
having a multiplicity of dense minute holes with nearly uniform diameter. Filter
papers on the other hand are formed by entanglement of a multiplicity of filaments
and hence have minute holes having irregular shapes. In contrast, membrane
filters have uniform-shape minute holes and hence characterized by their very
sharp separatability of bacteria. For the purpose of collecting airborne bacteria,
membrane filters with a hole diameter of 0.2 to 0.8 pm are generally used. Some
membrane filters are provided with a grating mark with a pitch of for example 2 to
3 mm, so that the number of colonies, after the bacteria collected have been
incubated, can readily be counted.
The system is also provided with a pumping means 15 that sucks air out of the
chamber 10 through the filter 45. In a preferred embodiment, the pumping means
15 is a vacuum pump, which is operatively connected to the enclosed chamber 10
by a conduit means 40 for evacuating the enclosed chamber 10, i.e. sucking air out
of the enclosed chamber.
Further, the enclosed chamber is provided with a means for sanitization. The term
"sanitization" used herein refers to the process of eliminating all microbes from a
defined space, such as the enclosed chamber 10. Preferably, the means 30 for
sanitization is a source of ultra-violet radiation, also called an ultra-violet lamp,
preferably attached with the ceiling of the enclosed chamber.
Preferably, the enclosed chamber 10 comprises a floor, a ceiling, four walls 25,
26, 27, and 28 and a sealable opening 29 on a wall 26 having open and closed
conditions; the open condition allowing transfer of materials in and out of the
enclosed chamber 10 and the closed condition of the sealable opening 29 limiting
access to the enclosed chamber. However, in alternate embodiments, the chamber
may be of any suitable shape and size wherein the floor, ceiling or walls may not
be clearly defined. In a preferred embodiment, the sealable opening 29 is a door
which can be in an open or closed position.
In the preferred embodiment, a means 55 for dispersing a pre-calculated amount
of microbes and a means 60 for dispersing a pre-calculated amount of the
composition are mounted on the wall 28 opposite to the wall 26 bearing the door.
However, the means for dispersing a pre-calculated amount of the microbes and a
pre-calculated amount of the composition may be mounted on any suitable surface
within the enclosed chamber. The term "composition" used herein refers to any
substance or a mixture whose anti-microbial activity may be evaluated by the
system and method of the present invention. The means 55 for dispersing
microbes, the means 60 for dispersing the composition and the means 50 for
dispersing humectants may be an ultrasonic nebulizer, a micropump or any
pressure activated device.
In order to evaluate the efficacy of a composition against air borne microbes, it is
desirable to thoroughly mix a suspension of the microbes and the composition for
a pre-determined period of time, in air. However, in the absence of a suitable
carrier, the microbes settle down faster on the floor or surface of the enclosed
space or chamber than in its presence. The term "carrier" used herein refers to
humectants such as water, glycerine and so forth. Although, water and glycerine
have been used in the present invention, any other humectant, which enables the
microbes to be suspended in air for a longer period can be used. Consequently, a
system for detecting and determining air-born microbes and measuring the
efficacy of an anti-microbial composition, without humectants (carrier), would
detect less than the actual number of microbes. Hence, to prevent the microbes
from settling on the floor or surface of the enclosed chamber, the air is humidified
by humectants, which are released by a means 50 for dispersing pre-calculated
amount of humectant. Examples of humectants include, but not limited to,
glycerine, water and combinations thereof. The humectant comprises water and
glycerine preferably in ratios ranging from 1 : 100 to 100: 1 and more preferably in
a ratio of 1:3. The inventors of the present invention have found that the bacterial
recovery on dispersing the humectant into the chamber in the form of a fine mist
was greater than in the absence of humectants. The term "bacterial recovery" used
herein refers to the number of colonies observed on incubating the filter on a
nutrient medium, after sucking the microbe spiked air through it. The means 50
for releasing pre-calculated amount of humectant into the enclosed chamber in the
form of a fine mist is removably introduced into the enclosed chamber through the
door.
Additionally, the enclosed chamber is provided with a means 35 for circulating air
and efficient mixing of the dispersed microbes, humectant and composition. In a
preferred embodiment of the invention the means 35 for circulating air is either a
fan or a blower.
Now described is a method for determining the efficacy of the anti-bacterial
composition by the use of the system of the present invention.
Firstly, the enclosed chamber 10, with the door closed, is sanitized by switching
on the source of ultraviolet radiation, i.e. an ultraviolet lamp, for pre-determined
amount of time, i.e. 10 to 60 min. After that, the source of ultraviolet radiation is
switched off and a means 35 for circulating air mounted in the enclosed chamber
is switched on. The fan distributes the microbial culture uniformly across the
entire chamber and enables efficient mixing of the composition, microbes and
humectant. A blank reading is taken to check for the presence of any bacteria after
sanitization. The blank reading is taken by switching on the pumping means 15
and sucking the air inside the enclosed chamber 10 through a filter 45 covering the
air suction port 20. Thereafter, the vacuum pump is switched off, the filter is
demounted and incubated on a nutrient agar plate, to record the blank reading.
Next, a known quantity of microbes, i.e. the test organism is dispersed into the
enclosed chamber by a means 55 for dispersing pre-calculated amount of
microbes. In a preferred embodiment the means 55 for dispersing the microbes is
a nebulizer. Preferably, 1 o4 cWmL are nebulized.
Simultaneously, humectant is dispersed into the chamber in the form of a fine mist
through a means 50 for dispersing humectant. Examples of humectants include,
but not limited to, glycerine, water and combinations thereof. The humectant used
in the present invention comprises water and glycerine preferably in ratios ranging
fi-om 1 : 100 to 100: 1 and more preferably in a ratio of 1 :3.
Thereafter, a known quantity of the composition is sprayed into the enclosed
chamber by means 60 for dispersing pre-calculated amount of the composition
into the chamber. The composition, humectant, and the microbes are allowed to
mix uniformly throughout the enclosed chamber for a pre-calculated period of
time. After that, the pumping means 15 is switched on and the air inside the
enclosed chamber 10 is sucked through a filter 45 covering the air suction port 20
to collect the microbes. The filter may be a membrane filter or a filter paper. The
temperature of the enclosed chamber is maintained at 25 OC and relative humidity
is maintained at 50% for the entire duration of the process.
s Next, the vacuum pump is switched off and the filter paper is demounted and
incubated on a nutrient agar plate at 30 O C for 72 h. After incubation, the number
of colony forming units of the microbes is determined and the percentage efficacy
of the composition and log reduction value is calculated.
As described heretofore, the system and method of the present invention can
detect and determine airborne microbes, and evaluate the efficacy of a
composition against airborne microbes with high reproducibility. The composition
may be an air-freshener, an insect repellent, an insecticide, a deodorant, an
essential oil, a fragrance, an aromatic substance, or a sublimable composition.
Other features of the invention will become apparent in the course of the
following descriptions of exemplary embodiments which are given for illustration
of the invention and are not intended to be limiting thereof.
Example 1.
Bacterial recovery count in the absence of a humectant
The enclosed chamber (Length xWidth height: 9 ft x 4 ft x6 ft) was sanitized by
switching on the source of ultraviolet radiation (254 nm) for a period of 1 h. A
blank reading was taken to check for the presence of any bacteria after
sanitization. The blank reading was taken by switching on the pumping means 15
and sucking the air inside the enclosed chamber 10 through a filter 45 covering the
air suction port 20 at the rate of 28.3 Llmin for a duration of 2 min. Thereafter, the
vacuum pump was switched off and the filter was demounted and incubated on a
nutrient agar plate (prepared as per IS5402:2012), to record the blank reading.
This was done to ensure proper sanitization of the enclosed chamber. The source
of ultraviolet radiation was switched off. Culture spiking solution of 300 cWmL
was prepared by McFarland dilution method. The culture was dispersed by means
of a nebulizer at the rate of 1 mL1min. The fan was switched on for the entire
duration of the process, for efficient distribution of the microbial culture
throughout the enclosed chamber. Thereafter, the vacuum pump was switched on
and the air inside the enclosed chamber was sucked outside through a filter paper
(Merk: 0.45 pm, 47 mm) to capture the microbes. The air was drawn out through
the filter paper at the rate of 28.3 Llmin for a duration of 2 min. The temperature
inside the enclosed chamber was maintained at 25 OC and the relative humidity
was 50%. Finally, the vacuum pump was switched off, the filter paper demounted
and incubated in a nutrient agar plate at 30 OC for 72 h, and thereafter the number
of colony forming units on the filter paper was determined. The above method
was performed on the following bacterial cultures: Escherichia. coli (MTCC-443),
Staplzylococcus aureus (MTCC-96), Salmonella tvphimuvizlin (MTCC-733) and
Salmonella typhi (NCTC-786) and the results are illustrated in table 1. However, it
should be noted that the above method may be used to estimate any other
speciesof microbes.
Table 1. Microbial count obtained on nutrient agar plate, in the absence of
humectant
Example 2.
Bacterial recovery count in the presence of humectant
The enclosed chamber (Length xWidth height: 9 ft x 4 ft x6 ft) was sanitized by
switching on the ultraviolet lamp (254 nm) for a period of 1 h. A blank reading
was taken to check for the presence of any bacteria after sanitization. The blank
reading was taken by switching on the pumping means 15 and sucking the air
inside the enclosed chamber 10 through a filter 45 covering the air suction port 20
at the rate of 28.3 Llmin for a duration of 2 min. Thereafter, the vacuum pump is
switched off and the filter is demounted and incubated on a nutrient agar plate
(prepared as per IS5402:2012), to record the blank reading. This was done to
ensure proper sanitization of the enclosed chamber. Thereafter the ultraviolet lamp
was switched off. Microbial culture containing 300 cWmL was prepared by
Salmonella
typhimurium
3 00
230
Staphylococcus
aureus
300
240
Bacterial
Culture
Spiking
range in
cfdml
Count
obtained on
plate in
cfu/ml
Salmonella
Q!hi
3 00
245
Esclzerichia
coli
300
260
McFarland dilution method. The culture was dispersed by means of a nebulizer at
the rate of 1 mL/min and a humectant comprising water and glycerine in a ratio of
1 :3, was simultaneously released into the chamber in the form of a fine mist by a
fogging machine. The fan was switched on for the entire duration of the process
for efficient mixing of the microbes, and humectant throughout the enclosed
chamber. After mixing for two minutes, the vacuum pump was switched on and
the air inside the enclosed chamber was sucked outside through a filter paper
(Merk: 0.45 pm, 47 rnm) to capture the microbes. The air was drawn out through
the filter paper at the rate of 28.3 Llmin for a duration of 2 min. The temperature
inside the enclosed chamber was maintained at 25 OC and the relative humidity
was 50%. Finally, the vacuum pump was switched off, the filter paper demounted
and incubated in a nutrient agar plate at 30 "C for 72 h, and the number of colony
forming units on the filter paper was determined. The above method was
performed on the following bacterial cultures: Escherichia. coli (MTCC-443),
Staphylococcus aureus (MTCC-96), Salmonella typhimurium (MTCC-733) and
Salmonella typhi (NCTC-786); and the results are illustrated in table 2. However,
it should be noted that the above method may be used to estimate any other
species of microbes.
Table 2. Microbial count obtained on nutrient agar plate, in the presence of
humectant
Salmonella
@phi
300
285
Salmonella
typhimurium
3 00
280
Bacterial
Culture
Spiking
range,
cfu/mL
Count
obtained on
plate,
cfu/ml
Escherichia.
coli
300
285
Staphylococcus
aureus
300
290
A comparison of the microbial count obtained on the agar plate in the presence
and absence of humectant reveals that the recovery of microbes is greater in a
method comprising the step of dispersing humectants into the enclosed chamber.
Example 3
The efficacy of commercial sample "Composition 1" against airborne
microbes
The enclosed chamber (Length xWidth height: 9 ft x 4 ft x6 ft) was sanitized by
switching on the ultra-violet lamp (254 nrn) for a period of 1 h. A blank reading
was taken to check for the presence of any bacteria after sanitization. The blank
reading was taken by switching on the pumping means 15 and sucking the air
inside the enclosed chamber 10 through a filter 45 covering the air suction port 20
at the rate of 28.3 Llmin for a duration of 2 min. Thereafter, the vacuum pump
was switched off and the filter was demounted and incubated on a nutrient agar
plate (prepared as per IS5402:2012), to record the blank reading. Microbial
culture containing lo4 cWmL was prepared by McFarland dilution method. The
culture was dispersed by means of a nebulizer at the rate of 1 mllmin and a
humectant comprising water and glycerine in a ratio of 1:3 was simultaneously
released into the chamber by a fogging machine. After that, known quantity of the
composition was introduced into the chamber with the help of product dispenser
provided along with the sample. The fan was switched on for the entire duration
of the process for efficient mixing of the microbial culture, humectant and
composition throughout the enclosed chamber. After mixing for two min, the
vacuum pump was switched on and the air inside the enclosed chamber was
sucked outside through a filter paper (Merk: 0.45 pm, 47 mm) to capture the
microbes. The air was drawn out through the filter paper at the rate of 28.3 Llmin
for 2 min. The temperature inside the enclosed chamber was maintained at 25 "C
and the relative humidity was 50%. Finally, the vacuum pump was switched off,
the filter paper demounted and incubated in a nutrient agar plate at 30 "C for 72 h,
and the number of colony forming units on the filter paper was determined. The
above method was performed on the following bacterial cultures: Escherichia.
coli (MTCC-443), Staphylococcus aureus (MTCC-96), Salmonella typhimurium
(MTCC-733) and Salmonella typhi (NCTC-786). However, it should be noted that
the above method may be used to estimate any other species of microbes. The log
reduction values and efficacies (percentage reductions) of "Composition 1" on the
above bacterial cultures, on releasing said composition into the enclosed space by
means of a dispenser provided with the composition were calculated and
presented in table 3.
Table 3. Log reduction and Efficacy (Percentage reduction) after spraying
known quantity of "Composition 1" into the enclosed chamber
From the data illustrated in table 3, it is evident that treatment of a sterile
chamber, artificially inoculated with known quantity of Escherichia. coli culture,
and known quantity of Composition 1, with the help of product dispenser
provided along with the sample, afforded a 99.02% reduction in Escherichia. coli
q
?3
.5- Y X
%
&
M S
2.01
2.08
2.10
2.04
Bacterial
culture
Escherichia
coli
Staphylococcus
aureus
Salmonella
@phi
Salmonella
typhimurium
. ..
E
2 . 2
3;0
3qi 2& Ma2
II a e 5 a o
5 gh
W
99.02
99.16
99.20
99.07
Ca,
-m a EP = E
% , a a a
h 4 z 3
"E .C -
a1 --a
1 .OO
1 .OO
1 .OO
1 .OO
.9 3 m o
8 ..3
cl;;
1 ,
8
O h 2 e3 M
$ V1
M S
4.00
4.00
4.00
4.00
g 5
Y
9.80
8.40
8.00
9.30
1.99
1.92
1.90
1.96
count, in 2 min. Likewise. the percentage reductions observed in case of
Staphylococcus aureus culture was 99.16%, Salmonella typhi culture was 99.20%,
and Salmonella typhimurium culture was 99.07%.
Example 4
The efficacy of commercial sample "Composition 2" against airborne
microbes.
The efficacy of Composition 2 against air borne microbes was determined
according to the method provided in Example 3. The log reduction values and
efficacies (percentage reductions) of Composition 2 on the bacterial cultures, i.e.
Escherichia. coli (MTCC-443), Staphylococcus aureus (MTCC-96), Salmonella
typhimurium (MTCC-733) and Salmonella typhi (NCTC-786), on releasing a
known quantity of said composition into the enclosed space by means of a
dispenser provided with the composition were calculated and presented in table 4.
However, it should be noted that the above method may be used to estimate any
other species of microbes.
Table 4. Log reduction and Efficacy (Percentage reduction) after releasing
known quantity of "Composition 2" into the enclosed chamber
. rn
E: a C 7 2 =
q 3 'pI?: 8
M
II 5
5 5
3 E
ew g
99.21
99.15
F
5
.3- C
U s
'I?
p:
M S
2.10
2.07
Bacterial
culture
Escherichia.
coli
Staphylococcus
aureus
.-2-
V)
8
C a 3 3
1 ..I h a u
C
1 .OO
1 .OO
E: L
ug
7.90
8.50
S
4.00
4.00
s 5
1.90
1.93
chamber, artificially inoculated with known quantity of Escherichia. coli culture,
and a known quantity of Composition 2, with the help of product dispenser
provided along with the sample, afforded a 99.2 1% reduction in Escherichia. coli
count, in 2 min. Likewise, the reduction observed in case of Staphylococcus
aureus culture was 99.124, Salmonella typhi culture was 99.04%, and Salmonella
typhimurium culture was 99.20%.
Salmonella
typhimurium
While particular embodiments of the present invention have been illustrated and
described, it would be obvious to those skilled in the art that various other changes
and modifications can be made without departing from the spirit and scope of the
invention. It is thereof intended to cover in the appended claims such changes and
modifications that are within the scope of the invention.
From the results illustrated in table 4, it is evident that treatment of a sterile
1 .OO 4.00 8.00 1.90 2.10 99.20

We claim:
1. A system for detecting airborne microbes in an enclosed chamber,
comprising:
(a) an enclosed chamber 10;
(b) an air suction port 20 on the enclosed chamber 10;
(c) a filter 45 removably attached with the air suction port 20, wherein the
filter isolates the interior of the chamber from the external
environment;
(d) a pumping means 15 operatively associated with the interior of the
enclosed chamber for sucking air out of the enclosed chamber through
the filter 45;
(e) a means 30 for sterilization;
(f) at least one sealable opening in the enclosed chamber 10;
(g) a means 55 for dispersing microbes;
(h) a means 60 for dispersing the composition;
(i) a means 50 for dispersing humectant; and
Cj) a means 35 for circulating air inside the enclosed chamber 10;
wherein the means 55 for dispersing microbes, the means 60 for dispersing
the composition, the means 50 for dispersing humectant and the means 35
for circulating air inside the enclosed chamber can be removably
introduced into the enclosed chamber 10 through the at least one sealeable
opening.
2. The system for detecting airborne microbes in an enclosed chamber, as
claimed in claim 1, wherein a conduit means 40 operatively connects the
pumping means 15 to the interior of the enclosed chamber.
3. The system for detecting airborne microbes in an enclosed chamber, as
claimed in claim 1, wherein the filter 45 is a filter paper or a membrane
filter.
4. The system for detecting airborne microbes in an enclosed chamber, as
claimed in claim 1, wherein the composition is an air-freshener, an insect
repellent, an insecticide, a deodorant, an essential oil, a fragrance, an
aromatic substance, or a sublimable composition.
5. The system for detecting airborne microbes in an enclosed chamber, as
claimed in claim 1, wherein the humectant is selected from the group
consisting of water, glycerine and combinations thereof.
6. The system for detecting airborne microbes in an enclosed chamber, as
claimed in claim 5, wherein the humectant comprises water and glycerine
preferably in ratios ranging from 1 : 100 to 100: 1 and more preferably in a
ratio of 1:3.
7. The system for detecting airborne microbes in an enclosed chamber, as
claimed in claim 1, wherein the microbes are selected from the group
consisting of bacteria, hngi and viruses.
8. A method for detecting airborne microbes in an enclosed chamber,
comprising the steps of:
(a) sanitizing an enclosed chamber;
(b) measuring a blank reading;
(c) dispersing a pre-calculated quantity of microbes into the enclosed
space;
(d) dispersing a pre-calculated quantity of humectant into the enclosed
chamber;
(e) dispersing a pre-calculated quantity of the composition into the
enclosed space;
(f) mixing the microbes, the humectant and the composition for a precalculated
amount of time;
(g) sucking the air in the enclosed chamber through a filter;
(h) placing the filter in a culture medium and incubating;
(i) counting surviving microbe colonies after incubation; and
Cj) calculating log reduction value and percentage efficacy of the
composition.
9. The method for detecting airborne microbes in an enclosed chamber, as
claimed in claim 8, wherein the filter is a filter paper or a membrane filter.
10. The method for detecting airborne microbes in an enclosed chamber, as
claimed in claim 8, wherein the microbes are selected from the group
consisting of bacteria, fungi and viruses.
1 I . A method to determinc the anti-microbial efficacy of a con~position,
comprising the steps of:
(a) sanitizing an enclosed chamber;
(b) measuring a blank reading;
(c) dispersing a pre-calculated quantity of microbes into the enclosed
space;
(d) dispersing a pre-calculated quantity of humectant into the enclosed
chamber;
(e) dispersing a pre-calculated quantity of the composition into the
enclosed space;
(f) mixing the microbes, the humectant and the composition for a precalculated
amount of time;
(g) sucking the air in the enclosed chamber through a filter;
(h) placing the filter in a culture medium and incubating;
(i) counting surviving microbe colonies after incubation; and
(j) calculating log reduction value and percentage efficacy of the
composition.