DEVICES AND METHODS FOR DETECTION OF MICROORGANISMS

Abstract

A device for determining the presence of microorganisms in a sample and the antimicrobial sensitivity thereof, the device comprising multiple chambers including: a first control chamber; and a first test chamber; wherein said first control chamber comprises: media and/or nutrients that support or encourage microbial growth; and a tetrazolium indicator compound; and said first test chamber comprises: media and/or nutrients that support or encourage microbial growth; a tetrazolium indicator compound; and a first selective agent which inhibits the growth and/or replication of one or more specific microorganisms.

Claims

1. A device for determining the presence of microorganisms in a sample and the antimicrobial sensitivity thereof, the device comprising multiple chambers including: a first control chamber; and a first test chamber; wherein said first control chamber comprises: media and/or nutrients that support or encourage microbial growth; and a tetrazolium indicator compound; and said first test chamber comprises: media and/or nutrients that support or encourage microbial growth; a tetrazolium indicator compound; and a first selective agent which inhibits the growth and/or replication of one or more specific microorganisms.

2. A method of determining the presence of microorganisms in a sample and the antimicrobial sensitivity thereof, the method comprising: (a) contacting the sample with a first control composition comprising: media and/or nutrients that support or encourage microbial growth; and a tetrazolium indicator compound; and (b) contacting the sample with a first test composition comprising: media and/or nutrients that support or encourage microbial growth; a tetrazolium indicator compound; and a first selective agent which inhibits the growth and/or replication of one or more specific microorganisms; and (c) observing said first control composition and said first test composition.

3. A device according to claim 1 wherein the media and/or nutrients that support or encourage microbial growth used in each chamber or composition is selected from brain heart infusion and Wilkins Chalgren media.

4. A device according to claim 1 wherein the tetrazolium compound used in each chamber or composition is tetrazolium violet.

5. A device according to claim 1 wherein the first selective agent is selected from ciprofloxacin, nitrofurantoin, pivmecillnam, trimethoprim and cephalexin.

6. A device of claim 1 which further comprises one or more further test chambers wherein the or each further test chamber comprises media and/or nutrients that support or encourage microbial growth; a tetrazolium indicator compound; and a selective agent which inhibits the growth and/or replication of one or more specific microorganisms.

7. A method according to claim 2 which further comprises contacting the sample with one or more further test compositions wherein the or each further test composition comprises: media and/or nutrients that support or encourage microbial growth; a tetrazolium indicator compound; and a selective agent which inhibits the growth and/or replication of one or more specific microorganisms.

8. A device according to claim 6 wherein each chamber or each composition comprises the same media and/or nutrients that support and/or encourage microbial growth and the same indicator compound.

9. A device according to claim 6, wherein each test chamber or each test composition comprises a different selective agent or combination of selective agents which inhibits the growth and/or replication of one or more specific microorganisms.

10. A device according to claim 9 wherein each selective agent is selected from ciprofloxacin, nitrofurantoin, pivmecillinam, trimethoprim and cephalexin.

11. A method according to claim 2 wherein the sample is a urine sample.

12. A method according to claim 7 wherein step (c) involves observing the colour of the control composition(s) and test composition(s).

13. A method according to claim 2 wherein the compositions are incubated after steps (a) and (b) and before step (c).

14. A device according to claim 1 which further comprises a dilution reservoir.

15. A device for determining the presence of microorganisms in a sample and the antimicrobial sensitivity thereof, preferably a urine sample, the device comprising: a first control chamber and five test chambers wherein each chamber comprises media and/or nutrients that support or encourage microbial growth and a tetrazolium indicator compound and each test chamber comprises a selective agent; wherein the first test chamber comprises ciprofloxacin as the selective agent, the second test chamber comprises nitrofurantoin as the selective agent, the third test chamber comprises pivmecillinam as the selective agent, the fourth test chamber comprises trimethoprim as the selective agent and the fifth test chamber comprises cephalexin as the selective agent.

16. A method according to claim 2 which includes a step prior to step (a) and step (b) of diluting the sample.

17. (canceled)

18. An incubation apparatus comprising multiple recesses each adapted to receive a device of claim 1.

19. A kit comprising an incubation apparatus according to claim.

Description

[0203] The invention will now be further described with reference to the following non limiting examples and the accompanying drawings in which:

[0204] FIG. 1 is a perspective view of a device of the first aspect with the lid in place;

[0205] FIG. 2 is an expanded view of the device of FIG. 1 showing the lid removed;

[0206] FIG. 3 is a cross sectional view along the plane XX of the device of FIG. 1;

[0207] FIGS. 4A to 4F are the results of tested compositions which are later described.

[0208] FIG. 5 shows a device of the invention similar to that of FIGS. 1 to 3 which further comprises a dilution reservoir;

[0209] FIG. 6 shows a further device of the invention which comprises a dilution reservoir;

[0210] FIG. 7 shows a further device of the invention in which a dilution step is carried out in a syringe, prior to delivering the sample into the device;

[0211] FIG. 8 shows an incubation apparatus comprising recesses adapted to accommodate the device of FIGS. 1 to 3; and

[0212] FIG. 9 shows an incubation apparatus comprising recesses adapted to accommodate the device of FIG. 5.

[0213] FIGS. 1 to 3 show a device of the present invention including a transparent lower portion 1 for receiving a liquid sample and a lid 2. The lower portion is divided into 8 chambers 3 in an annular arrangement around a central core. An opaque insert 4 (suitably of white plastic) enables the colour of each chamber to be viewed individually without interference from the neighbouring chambers.

[0214] Lid 2 includes a central aperture 5. A cap 6 fits into the top of the core of the lower portion. In use when the lid is placed loosely onto the device a liquid poured through aperture 5 is directed by the cap into each of the chambers 3. Once the sample has been poured into the device the lid may be twisted closed by engagement of threads on the internal surface of the lid with threaded region 7 of the transparent lower portion of the device. Closing of the device by the twisting mechanism seals the aperture 5 with the cap 6. This prevents any transfer of liquid between the chambers.

[0215] In use each test/control chamber is provided with one or more capsules or other means comprising an indicator compound, nutrients and optionally one or more selective agents.

[0216] FIG. 5 shows a device similar to that of FIG. 1 but which further includes a reservoir. The reservoir 10 is fitted to the top of the device in a twistable and lockable manner. A diluent such as a buffer is provided within the reservoir. The reservoir is initially in a locked position (FIG. 5A) as demonstrated by a lock indicator 11. A sample, for example a sample of urine, is delivered into the device via syringe 12.

[0217] The reservoir is then twisted as indicated by arrow 13 (FIG. 5B) into a flowing position to allow the now diluted sample to be delivered into each of the chambers. Once the content of the reservoir has been emptied into the chambers the reservoir can be twisted back again (FIG. 5C) in the direction indicated by arrow 15 and the indicator 11 will again show this to be in a locked position. This device can now be incubated.

[0218] FIG. 6 shows an alternative device incorporating a reservoir 20 fixed to multiple chambers 21. In use each test/control chamber is provided with one or more capsules or other means comprising an indicator compound, nutrients and optionally one or more selective agents.

[0219] A valve 22 is provided at the top of the reservoir and a syringe 23 is used to deliver a sample into the reservoir (FIG. 6A). The reservoir will contain a diluent, for example a buffer solution. A tab 23 maybe used to lock the reservoir in a position such that the contents cannot flow into the chambers while the sample is added. Once the sample has been added to the reservoir the switch 24 is moved down into a flow position (FIG. 6B) to deliver the contents of the reservoir equally into the six chambers shown. When all of the liquid in the reservoir has been delivered into the chambers then the switch 24 is moved into a sealed position (FIG. 6C) in which the contents of the chamber are sealed within them.

[0220] FIG. 7 shows a syringe 30 comprising buffer 31 which not locked by a widget 32 to prevent accidental dispensing. A user can withdraw the sample 34 into the syringe (FIG. 7A).

[0221] The syringe can then be used to deliver a pre-mixed sample and into a device 33 which includes a portion 35 configured internally to direct flow equally into all of the chambers (FIG. 7B). Once all of the sample has been delivered the syringe can be removed (FIG. 7C).

[0222] FIG. 8 shows an incubation apparatus 41 comprising multiple recesses 40 each adapted to receive a device of FIG. 5. Each of the recesses includes multiple optical sensors 42 which measure the absorption at a particular wavelength in each individual chamber. An information screen 43 indicates which particular device unit and which chamber within the device has microbial growth.

[0223] FIG. 9 shows an incubation chamber 51 including multiple recesses 50 and an information screen 53. This device is similar to the one disclosed in FIG. 8 but shaped to accommodate the device of FIG. 7.

EXAMPLES

[0224] The device shown in FIGS. 1 and 2 includes 8 chambers. At least one of these is a control chamber which includes an indicator compound and nutrients. One or more of the remaining chambers may be test chambers. One chamber may contain no reagent to show the colour of the urine sample itself.

[0225] Each test chamber suitably includes an indicator compound (suitably the same indicator compound as the control chamber, nutrients and one or more selective agents.

[0226] The inventors have carried out some experiments in which the same urine sample has been tested (each time in triplicate) in test tubes with compositions suitable for inclusion in the control chamber and test chambers of the invention.

[0227] Each test tube contained 0.04 mg tetrazolium violet, 6.6 mg Wilkins Chalgren media. The test chambers also contained an amount of antibiotic compound as indicated in the tables below (as concentration in final volume of urine).

[0228] For each example, 0.2 mL of a urine sample was contaminated with 10.sup.5 cfu/mL of the specified bacteria was poured into each test tube. The resultant composition was incubated at 37 C. for 10 hours. FIGS. 3A to 3F show the test tubes after the incubation period.

TABLE-US-00001 Test A - P. aeruginosa Chamber Antibiotic Concentration (g/ml) Control None Test 1 Nitrofurantoin 64 Test 2 Pivmecillinam 4 Test 3 Trimethoprim 1 Test 4 Cephalexin 16 Test 5 Ciprofloxacin 0.25

[0229] The results shown in FIG. 4A indicate that the infection can be treated with ciprofloxacin, and is resistant to the other antibiotics.

TABLE-US-00002 Test B - E. coli 1 Chamber Antibiotic Concentration (g/ml) Control None Test 1 Nitrofurantoin 64 Test 2 Pivmecillinam 4 Test 3 Trimethoprim 4 Test 4 Cephalexin 16 Test 5 Ciprofloxacin 0.06

[0230] The results shown in FIG. 4B indicate that the infection can be treated with any of the tested antibiotics.

TABLE-US-00003 Test C - E. coli 2 Chamber Antibiotic Concentration (g/ml) Control None Test 1 Nitrofurantoin 64 Test 2 Pivmecillinam 32 Test 3 Trimethoprim 32 Test 4 Cephalexin Test 5 Ciprofloxacin 0.5

[0231] The results shown in FIG. 4C indicate that the infection can be treated with nitrofurantoin, and is resistant to the other antibiotics.

TABLE-US-00004 Test D - K. pneumoniae Chamber Antibiotic Concentration (g/ml) Control None Test 1 Nitrofurantoin 64 Test 2 Pivmecillinam 4 Test 3 Trimethoprim 8 Test 4 Cephalexin 32 Test 5 Ciprofloxacin 0.25

[0232] The results shown in FIG. 4D indicate that the infection can be treated with ciprofloxacin or trimethoprim, and is resistant to the other antibiotics.

TABLE-US-00005 Test E - E. coli 3 Chamber Antibiotic Concentration (g/ml) Control None Test 1 Nitrofurantoin 64 Test 2 Pivmecillinam 4 Test 3 Trimethoprim 1 Test 4 Cephalexin 32 Test 5 Ciprofloxacin 0.06

[0233] The results shown in FIG. 4E indicate that the infection can be treated with any of the tested antibiotics.

TABLE-US-00006 Test F - P. mirabilis Chamber Antibiotic Concentration (g/ml) Control None Test 1 Nitrofurantoin 64 Test 2 Pivmecillinam 16 Test 3 Trimethoprim 8 Test 4 Cephalexin Test 5 Ciprofloxacin 0.5

[0234] The results shown in FIG. 4F indicate that the infection cannot be treated with any of the tested antibiotics, and is resistant to all those tested.