Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving viable micro-organism
Reexamination Certificate
2001-04-04
2004-08-03
Gitomer, Ralph (Department: 1651)
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving viable micro-organism
C435S030000
Reexamination Certificate
active
06770454
ABSTRACT:
FIELD OF THE INVENTION
This invention relates to a method and a kit for the detection and collection of microorganisms from the air and from surfaces utilizing a collection device that employs a dry collection methodology and a dry growth medium, and a container of a premeasured volume of liquid for hydrating the dry growth medium after microorganism collection on the dry growth medium has occurred.
BACKGROUND OF THE INVENTION
Many molds are considered to be harmful microorganisms. Mold spores are minute propagating units that facilitate the growth and spread of mold colonies. Inhalation of mold spores and other microorganisms into the lungs of humans has been found to be a significant factor in causing a number of different pneumoconioses and other health problems such as allergies, headaches, and fatigue. Most molds require oxygen and water to live and the aerobic molds of our environment are known to multiply and migrate by producing and releasing millions of mold spores into the air.
Methods for estimating the relative environmental mold spore bioburden count exist, but these methods involve the collection of environmental air mold spores on wet media such as Sabouraud's Dextrose Agar (an aqueous suspension of gelatin and nutrients that solidifies to a semi-solid, water-based gel below 45° C.). Exposing sterile Sabouraud Dextrose Agar in an open container such as a petri dish to air containing mold spores can result in mold spores settling on the agar on which they can then germinate and grow. After a number of days, the colonial growth resulting from the germination of a single spore may be so large that the colony may be visually discernable without the use of magnification equipment.
An example of the device described above is disclosed in U.S. Pat. No. 3,968,012 which provides a device for detecting bacteria and other microorganisms carried in the air inside of hospital respiratory machines, such as ventilators and anesthesia gas machines. The device comprises a culture medium dish, a cylindrical casing, and a sanitary cover where the dish and cover are substantially similar to a petri dish. In operation, the device is attached to the vent of a respiratory machine, and the cylindrical casing controls the flow of the air to the medium dish so that microbial particles are collected from the air.
Many other devices in the prior art combine machinery with wet culture media to sample air for testing. For example, U.S. Pat. No. 3,956,070 discloses a device comprising a casing to direct a flow of air over a cartridge containing a culture medium. The cartridge is comprised of a two-sided strip that is wound around two reels with a culture medium spread over each side of the strip. In operation, air is vented through the casing and over both sides of the culture medium strip. U.S. Pat. No. 3,980,524 discloses a device resembling the shape of a common flashlight which comprises a casing that houses batteries, a motor with a drive-shaft and flanges, and a culture medium cup. In this device, the motor is employed with the drive-shaft and flanges to direct a volume of air over the medium cup.
Dry growth media has been used to detect microorganisms in liquid samples. U.S. Pat. No. 4,565,783 discloses a device for growing microorganisms, comprising a self-supporting water-proof substrate, a layer of adhesive coated thereon, and a coating of dry, cold-water-soluble growth medium powder adhered uniformly to the surface of the adhesive. In accordance with the teachings of U.S. Pat. No. 4,565,783, when an aqueous test sample is placed on the substrate in contact with the dry growth medium powder, the growth medium is hydrated to a gel and germination of microorganisms present in the test sample may result.
Since environmental mold spores are and have been associated with adverse health effects in humans, it is desirable to monitor the environmental mold spore bioburden count in dwellings, office buildings, schools and other indoor areas inhabited by people. Various health organizations routinely monitor environmental air in specific locations for mold spores using passive and active agar impaction methods, however, these organizations generally do not test individual homes, businesses, churches, etc. There are health professionals who, for a fee, monitor the air for environmental mold spores in an individuals most frequently inhabited environments, but fees are high and anonymity may not be maintained during and/or after the testing.
Establishing the environmental mold spore count by moist agar impaction methods utilizing Sabouraud Dextrose Agar or other known nutrient formulations, for example trypticase soy agar as disclosed in U.S. Pat. No. 3,968,012, has many associated problems. The first problem is that sealed agar plates utilized in moist agar impaction methods have a relatively short shelf life, on the order of a few weeks at most. Further, moist agar can lose its moisture rapidly over time and, as a result, its ability to support microorganism germination and growth decreases rapidly to a point where germination may not occur at all.
In addition, the use of moist agar to collect environmental mold spores may create what can be described as a time lag problem. An environmental mold spore impacting an unsealed hydrated microbiological growth medium during the first moments of exposure, i.e., the first minute in a 60-minute exposure, may enjoy distinct survival advantages as compared to an environmental mold spore impacting a desiccated microbiological growth medium during the last moments of the exposure as a direct result of changes in moisture conditions of the growth medium over the exposure interval. The latter arriving spores may geminate only very slowly producing a micro-colony that may be too small to be visually enumerated, or may not even germinate at all. An undercount of the actual population of spores can result producing an inaccurate assessment of the true population of environmental mold spores in the location under analysis.
The known methods of the prior art typically employed in air quality sampling are the use of passive agars in petri dishes or agars which are inserted into active volumetric pump samplers. The use of agars in air quality sampling has multiple impediments for the individual consumer/user, including increased costs for preparation and storage and shipping, a very short shelf life and, if active volumetric pump samplers are used, an extremely high purchase cost or rental cost.
SUMMARY OF THE INVENTION
A useful method of environmental air and surface sampling which comprises the dry collection and subsequent hydration, growth and enumeration of microorganisms has been discovered. The use of such procedures has been found to overcome the problems of time-lag growth disparities and evaporation of the activating liquid during collection identified above. In addition, dry collection devices exhibit a relatively long shelf-life and provide the added advantage of being easier to handle, especially at the extremes of normal ambient temperatures. The dry collection method of the present invention is accomplished by means of an easy to use, low cost home test kit to quantify the environmental mold spore and microorganism count and represents an important step in the reduction of environmental microorganism levels. Such a kit is also useful to educators teaching concepts of microbiology and mathematics. The method and kit is also a tool for use in monitoring work environments including commercial/industrial facilities.
One aspect of the present invention is a method of detecting microorganisms comprising the steps of exposing a dry collection device containing a dry growth medium to environmental microorganisms, adding a premeasured volume of liquid to the dry growth medium, and allowing collected microorganisms to grow into colonies. In certain embodiments, the exposure step may be accomplished by placing the collection device on a surface for a predetermined interval of time so that microorganisms in the air can settle out onto the collection device
LaRocca Mary Anne Kunz
LaRocca Paul T.
Reilly Sean M.
Gitomer Ralph
Home Health Science Inc.
Synnestvedt & Lechner LLP
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