MXPA98002030A - A method for determining the sensitivity of antimicrobial agent of a non-parafinophilic microorganism and an associated apparatus - Google Patents

Abstract

A method for determining the sensitivity of at least one non-parafinophilic microorganism from a sample obtained from a patient for a different antimicrobial agent and predetermined amounts thereof. The method includes providing at least one container containing an aqueous solution and inoculating the solution with the sample. The method further includes placing within the container (i) a slide coated with a carbon source and (ii) a predetermined amount of an antimicrobial agent to be tested. By observing the growth of the non-paraffinophilic microorganism or the lack thereof on the slide, it can be determined whether the predetermined quantity of the antimicrobial agent is effective to inhibit the growth of the non-paraffinophilic microorganism on the slide. An associated apparatus is also described

Description

A METHOD TO DETERMINE THE SENSITIVITY OF ANTIMICROBIAL AGENT OF A NON-PARAFHMOFILIC MICROORGANISM AND AN ASSOCIATED APPARATUS BACKGROUND OF THE INVENTION This invention relates to a method for determining the sensitivity of an antimicrobial agent of a non-paraffinophilic microorganism and an associated apparatus. The treatment of infections often involves the educated assumptions of medical personnel about the nature of the microorganism involved and the correct antimicrobial agent and the amount thereof necessary to effectively treat the microorganism present in the infected tissue. Frequently there is a need to treat a mixed flora of several microorganisms. The medical staff is really interested in determining quickly which antimicrobial agents and in what doses, are necessary to ensure the effective inhibition of the growth of all the microorganisms present in the patient. There is currently no efficient, effective and economical way for a doctor to quickly determine which antimicrobial agent and what dose is needed in order to treat the patient. A doctor simply does not have available for the type of information regarding the sensitivity of the antimicrobial agent that it would make possible a more accurate selection of an antimicrobial agent and, once an appropriate antimicrobial agent is selected, facilitates a more precise dose of treatment. If this information is available, a doctor can treat the infection more effectively. In addition, because the antimicrobial agents are expensive, the information could be used so that only the amount of antimicrobial agent needed could be used to treat the infection. Finally, and more importantly, since antimicrobial agents can have undesirable side effects, the information can be used to find a more effective antimicrobial agent and the dose thereof, which will limit the undesirable side effects. As used herein, the term "nonparaffinophilic microorganism" means any microorganism supported by a carbon source other than paraffin. Examples of such nonparaffinophilic microorganisms include, but are not limited to the following: Mycobacterium tuberculosis; Mycobacterium paratuberculosis; Mycobacterium leprae; Staphylococcus; Streptococcus; E. Coli; Listeria; Brucellae; Humemophilus; Treponema; Pneumococcus; Clostridium: Cryptococcus; Coccidioides; and Histoplasma. Also, as used herein, the term "patient" refers to a member of the animal kingdom, including humans, whose body sample is being processed by the method and apparatus of the invention.

U.S. Patent Nos. 5,153,119 and 5,316,918 describe methods and apparatus for identifying and testing the antibiotic sensitivity of Mycobacterium avium-intracellulare ("MAI"), a nonparaffinophilic microorganism. The inventor named in those patents is Robert A. Ollar, one of the co-inventors of the invention described herein. This method involves providing a container containing an aqueous solution and injecting a sample into the solution. After this, a slide coated with paraffin is placed inside the container. The slide is then observed for the presence or absence of growth of MAI. Despite the existence of Dr. Ollar's patents, there is still a need for a method of testing the sensitivity of antimicrobial agent of one or more nonparaffinophilic microorganisms in a manner that minimizes the effectiveness of the antimicrobial agent used to inhibit the growth of one or more nonparaffinophilic microorganisms that may be present in a patient.

BRIEF DESCRIPTION OF THE INVENTION The invention has met or exceeded the aforementioned needs as well as others. The method to determine the sensitivity of at least one non-parafinophilic microorganism of a sample obtained from a patient to different antimicrobial agents and predetermined amounts thereof comprises providing at least one container containing an aqueous solution and inoculating the solution with the sample. The method further includes placing within the container (i) a slide coated with a carbon source and (ii) a predetermined amount of an antimicrobial agent to be tested. Observing the growth of the non-paraffinophilic microorganism or the lack thereof on the slide, it can be determined whether the predetermined amount of the antimicrobial agent is effective in inhibiting the growth of the non-paraffinophilic microorganism on the slide. An associated apparatus is also described. The apparatus comprises a container adapted to contain an aqueous solution, a quantity of antimicrobial agent to be tested and the sample. The apparatus further includes a slide covered with a carbon source, the slide being adapted to be placed in the container. Again, observation of the growth of the non-parafinophilic microorganism from the sample on the slide can be used to determine the concentration of the antimicrobial agent necessary to resist the growth of the non-parafinophilic microorganism on the slide.

BRIEF DESCRIPTION OF THE DRAWING A complete understanding of the invention can be achieved from the following detailed description of the invention when read in conjunction with the accompanying accompanying drawing which shows one embodiment of the antimicrobial agent sensitivity apparatus.

DETAILED DESCRIPTION The method and apparatus of the invention provide an efficient, effective and economical way to determine the sensitivity of at least one non-parafinophilic microorganism to different antimicrobial agents and the predetermined amounts thereof. Referring now to the single Figure, the sensitivity method of the antimicrobial agent will be explained with reference to one embodiment of the antimicrobial agent sensitivity apparatus 50. The apparatus 50 consists of six containers in the form of test tubes 60, 61, 62 , 63, 64, 65 each containing an amount of an aqueous solution, such as Czapek's broth 70, 71, 72, 73, 74, 75. It will be appreciated that the aqueous solution does not contain a carbon source, as desired for provide a single source of carbon in the slide (described below) to effectively culture the non-paraffinophilic microorganism to be tested on the slide and not in the aqueous solution. The aqueous solutions in test tubes 61-65 contain uniform ranges of increasing concentrations of an antimicrobial agent to be tested. The test tube 60 is used as a control tube that does not contain any antimicrobial agent. The sample from the patient is then inoculated into each of the test tubes 60-65. The sample can be a blood sample; any tissue biopsy; stomach fluid; urine; cerebrospinal fluid; Nasopharyngeal mucosa or saliva. These samples can be obtained from patients in the doctor's office or in the emergency room of a hospital, for example, by known techniques. The slides 80, 81, 82, 83, 84 and 85 coated with a carbon source are then placed inside the respective test tubes 60, 61, 62, 63, 64 and 65. The slides are incubated for a period of one hour. minimum of twenty-four (24) hours. Observing the growth of the nonparaffinophilic microorganism 90, 91, 92, 93, 94 on the 80-85 slides, the minimum inhibitory concentration ("M IC") of the antimicrobial agent necessary to prevent the growth of the nonparaffinophilic microorganism can be determined, the concentration of MIC in test tube 75 is therefore found. It will be appreciated that a sample may sometimes have more than one non-paraffinophilic microorganism present in the same. For example, if a patient has an abscess in the brain associated with bacterial endocarditis, the patient will be more likely to have an individual pathogen (ie, staphylococcus). However, there may be a mixed flora that has grown on the slide (ie, staphylococcus and meningococci). However, it is imperative to treat all bacterial flora, since any bacteria present is the cause of pathogenicity in the patient. This invention allows a physician to specify an antimicrobial agent and a particular dose thereof which will inhibit all the growth of flora on the slide and which is therefore effective to treat all non-parafinophilic microorganisms that are pathogenic in the patient. It will be appreciated that although the apparatus 50 is shown with multiple containers and multiple slides 80-85, that the invention is not limited to multiple containers and multiple slides, but that it also covers a single container and a single slide. The carbon source and slides 80-85 may include a gelatinous matrix containing a carbon source. A carbon source may be one or more of those selected from the group consisting of glucose, fructose, glycerol, mannitol, asparagine and casein among others. Another embodiment may include providing a slide and coating the slide with an adhesive and securing a plurality of gel beds to the adhesive. The carbon source can then be ionically or affinity bound to the gel beds.

Slides 80-85 with the gelatinous matrix containing a carbon source can be prepared by the following method. A container, such as a beaker for analysis, is first filled with 100 ml of distilled water. Two (2) grams of agar (the gelatinous matrix) and three (3) grams of the carbon source (such as glucose) are placed in the beaker for analysis. This mixture is boiled and the steam sterilized and the molten gelatinous matrix is poured into a petri dish, which is seated on a hot plate. In this way the gelatinous matrix / carbon source remains fused. After this, a sterile slide such as the slide 80 is introduced into the molten gelatinous matrix / carbon source and coated therewith. The now coated slide is removed from the petri dish and allowed to stand for a minute or two to solidify the coating therein. The slide with the coating of a gelatinous matrix containing a carbon source is easily placed in one of the test tubes 60-65 containing the aqueous solution and the sample. An alternative method of preparation of the slide involves first coating the slide with an adhesive, such as collodion and then applying a plurality of gel beds (commercially available from Pharmacia of Parsippany, New Jersey) to the adhesive. The gel beds are approximately one meter in diameter. The slide containing the coating of the gel beds is now immersed in a pH regulating agent containing the carbon source (such as glucose) to bind the carbon source to the gel beds either ionically or by affinity. Non-paraffinophilic microorganisms that can be identified using the method of the invention include any microorganism supported by a carbon source other than paraffin. Non-paraffinophilic microorganisms include, but are not limited to Mycobacterium tuberculosis; Mycobacterium paratuberculosis; Mycobacterium leprae; Staphylococcus; Streptococcus; E. Coli; Listeria; Brucellae; Humemophilus; Treponema; Pneumococcus; Clostridium; Cryptococcus; Coccidioides; and Histoplasma. It will be appreciated that a method for determining the sensitivity of at least one non-parafinophilic microorganism in a sample has been described and an associated apparatus has been described. The method is effective and efficient and does not involve the use of expensive and complicated equipment. An associated apparatus is also described. While the specific embodiments of the invention have been described, those skilled in the art will appreciate that various modifications and alterations to those details could be developed in light of the general teachings of the description. Accordingly, the particular provisions described are intended to be illustrative only and do not limit the scope of the invention to be given by the broad scope of the appended claims and any and all equivalents thereof.

Claims (19)

1. CLAIMS 1 . A method for determining the sensitivity of at least one non-paraffinophilic microorganism from a sample obtained from a patient for different antimicrobial agents and the predetermined amounts thereof, the method comprising: providing at least one container containing an aqueous solution; inoculate the solution with the sample; placing within the container (i) a slide coated with a carbon source and (ii) a predetermined amount of an antimicrobial agent; and observing the growth of the non-parafinophilic microorganism or the lack thereof on the slide to determine whether the predetermined amount of the antimicrobial agent is effective to inhibit the growth of the nonparaffinophilic microorganism on the slide.
2. 2. The method of claim 1, including employing as said slide one coated with a gelatinous matrix containing the carbon source.
3. 3. The method of claim 1, which includes providing the slide by first adhering a plurality of gel beds to the slide and then bonding the carbon source to the gel beds.
4. 4. The method of claim 3, wherein the carbon source is ionically bound to the gel beds.
5. The method of claim 3, wherein the carbon source is affinity bound to the gel beds.
6. The method of claim 3, including adhering the gel beds to the slide by means of an adhesive.
7. The method of claim 6, which includes employing as said adhesive a gelatinous matrix with the carbon source.
8. The method of claim 1, which includes employing as said carbon source one or more of the group comprising glucose, fructose, glycerol, mannitol, asparagine and casein.
9. The method of claim 1, including employing as said sample one selected from the group comprising blood, stomach fluid, urine, cerebrospinal fluid, nasopharyngeal mucosa and saliva.
10. The method of claim 1, which includes providing a plurality of containers each containing an aqueous solution; inoculate each container with a quantity of the sample; placing (i) a separate slide covered with a carbon source and (ii) a quantity of antimicrobial agent to be tested in each container, each container containing a predetermined amount different from the antimicrobial agent; and observing the growth of the non-paraffinophilic microorganism or the lack thereof on the slides to determine the minimum inhibitory concentration of the antimicrobial agent to inhibit the growth of the non-paraffinophilic microorganism.
11. 11. An apparatus for determining the sensitivity of at least one non-paraffinophilic microorganism from a sample obtained from a patient to different antimicrobial agents and predetermined amounts thereof comprising: a container adapted to contain (i) an aqueous solution; (ii) an amount of an antimicrobial agent to be tested; and (iii) said sample; and a slide coated with a carbon source, the carbon source being adapted to be placed in the container, whereby observation of the growth of the non-parafinophilic microorganism from the sample on the slide can be used to determine the concentration of the agent antimicrobial necessary to resist the growth of the non-parafinophilic microorganism on the slide.
12. The apparatus of claim 11, wherein the slide is coated with a gelatinous matrix containing the carbon source.
13. The apparatus of claim 11, wherein the slide is coated with a plurality of gel beds that have bonded thereto to the carbon source
14. 14. The apparatus of claim 13, wherein the carbon source is ionically bound to the gel beds.
15. 15. The apparatus of claim 13, where the carbon source is bound by affinity to the gel beds.
16. 16. The apparatus of claim 13, wherein the gel beds are adhered to the slide by an adhesive
17. 17. The apparatus of claim 16, wherein the adhesive is collodion. The apparatus of claim 11, wherein the carbon source is one or more selected from the group comprising glucose, fructose, glycerol, mannitol, asparagine, urea, casein and hydrolyzate. The apparatus of claim 11, which includes a plurality of containers each adapted to contain (i) an aqueous solution; (ii) a quantity of antimicrobial agent to be tested; (iii) said sample; and a plurality of slides each coated with a carbon source, each of which is adapted to be placed in one of the containers.