Method of detecting a Salmonella microorganism

Patent metadata

Field	Value
Publication number	US-9677111-B2
Application number	US-201214368525-A
Country	US
Kind code	B2
Filing date	Dec 18, 2012
Priority date	Dec 28, 2011
Publication date	Jun 13, 2017
Grant date	Jun 13, 2017

Abstract

A method of detecting a Salmonella microorganism is provided. The method includes the use of a selective growth medium, a first indicator system that is converted to a first detectable product by a Salmonella microorganism, and a second indicator system that is converted to a second detectable product by β-galactosidase enzyme activity. The method further comprises inoculating the growth medium and incubating the inoculated growth medium at a temperature higher than 40 degrees C.

First claim

The invention claimed is:

1. A method of detecting Salmonella microorganisms, the method comprising:
providing,
a sample to be tested;
a culture device;
a nutrient medium that facilitates growth of a Gram-negative enteric microorganism;
a first selective agent that inhibits the growth of Gram-positive microorganisms;
a first differential indicator system comprising a first differential indicator compound that can be converted to a first detectable product by a member of a group of Salmonella microorganisms that includes a microorganism of the species Salmonella bongori;
a second differential indicator system comprising a second differential indicator compound that can be converted to a second detectable product by a β-galactosidase enzyme activity;

contacting in the culture device the nutrient medium, the first selective agent, the first differential indicator system, and the second differential indicator system with a sample to form an inoculated culture device;
incubating the inoculated culture device for a first period of time at a temperature greater than 40 degrees C.;
observing the culture device to detect a presence or an absence of the first detectable product; and observing the culture device to detect a presence or an absence of the second detectable product;
wherein observing the presence of the first detectable product indicates a presence in the sample of a Salmonella microorganism;
wherein observing the presence of the first detectable product juxtaposed with the second detectable product indicates a presence in the sample of a microorganism other than a β-galactosidase-producing member of the species Salmonella bongori.

2. The method of claim 1 , wherein the culture device is provided as a thin film culture device with the nutrient medium, the first selective agent, the first differential indicator system, and the second differential indicator system disposed therein in a dehydrated form.

3. The method of claim 1 , wherein observing the absence of the first detectable product indicates an absence of a Salmonella microorganism in the sample.

4. The method of claim 1 , wherein observing the nutrient medium to detect the presence of the first detectable product comprises observing the culture device to detect a first detectable color.

5. The method of claim 1 , wherein the first differential indicator compound comprises an enzyme substrate.

6. The method of claim 5 , wherein the enzyme substrate comprises an enzyme substrate to detect caprylate esterase enzyme activity or to detect α-galactosidase enzyme activity.

7. The method of claim 6 , wherein the enzyme substrate is selected from the group consisting of 5-bromo-6-chloro-3-indolyl caprylate, 4-nitrophenyl caprylate, 2-naphthyl caprylate, 5-bromo-4-chloro-3-indoxyl-α-D-galactopyranoside, resorufinyl-α-D-galactopyranoside, and 4-nitrophenyl-α-D-galactopyranoside.

8. The method of claim 5 , wherein the first detectable product is water-insoluble.

9. The method of claim 1 , wherein the first differential indicator system comprises a pH indicator and at least one carbohydrate selected from the group consisting of melibiose, 2-deoxy-D-ribose, mannitol, L-arabinose, dulcitol, maltose, L-rhamnose, trehalose, D-xylose, and sorbitol.

10. The method of claim 1 , wherein the first detectable product is water-soluble, wherein observing the first detectable product comprises observing a colored zone adjacent to a microbial colony.

11. The method of claim 1 , wherein observing the second detectable product comprises observing the culture device to detect a second detectable color.

12. The method of claim 1 , wherein the second differential indicator compound is selected from the group consisting of 5-bromo-4-chloro-3-indolyl-1-β-D-galactopyranoside, 5-bromo-3-indolyl-β-D-galactopyranoside, 5-bromo-6-chloro-3-indolyl-β-D-galactopyranoside, 2-nitrophenyl-β-D-galactopyranoside, and 4-nitrophenyl-β-D-galactopyranoside.

13. The method of claim 1 , wherein the first detectable product is water-soluble and the second detectable produce is water-insoluble or, alternatively, wherein the first detectable product is water-insoluble and the second detectable produce is water-soluble.

14. The method of claim 1 , wherein the first selective agent is selected from the group consisting of an antibiotic, bile salts, bile salts No. 3, deoxycholic acid, cholic acid, crystal violet, novobiocin, nalidixic acid, polymyxin B, streptomycin, methicillin, cefsoludin, or a combination of any two or more of the foregoing selective agents.

15. The method of claim 1 , wherein the culture device further comprises at least one second selective agent, wherein the at least one second selective agent inhibits the growth of at least one Gram-negative enteric microorganism that is not a member of the genus Salmonella.

16. The method of claim 15 , wherein the second selective agent is selected from the group consisting of a β-lactam antibiotic, an aminoglycoside antibiotic, a quinolone antibiotic, a sulfa antibiotic, a polymyxin antibiotic, and a combination of any two or more of the foregoing antibiotics.

17. The method of claim 16 , wherein the at least one second selective agent comprises a combination of naladixic acid, streptomycin, and polymyxin B.

18. The method of claim 1 , wherein incubating the culture device comprises incubating the culture device at a temperature between 41 to 44 degrees C.

19. The method of claim 1 , further comprising:
providing an article with a third differential indicator system comprising a third differential indicator compound that can be converted by a Salmonella microorganism to a third detectable product;
contacting the article with the inoculated culture device;
incubating the inoculated culture device for a second period of time; and
observing the culture device to detect the third detectable product;
wherein observing the third detectable product juxtaposed with the first detectable product indicates the presence of a Salmonella microorganism in the sample.

20. The method of claim 19 , further comprising:
providing a nondifferential indicator compound that can be converted by an enteric microorganism to a fourth detectable product; and
observing the culture device to detect a presence or an absence of the fourth detectable product;
wherein contacting in the culture device the nutrient medium, the first selective agent, the first differential indicator system, and the second differential indicator system with a sample to form an inoculated culture device further comprises contacting in the culture device the nutrient medium, the first selective agent, the first differential indicator system, the second differential indicator system, and the nondifferential indicator compound with the sample to form the inoculated culture device.

21. The method of claim 1 , wherein observing the culture device comprises observing the culture device visually.

22. The method of claim 1 , wherein observing the culture device comprises creating an image of the culture device using an imaging device.

23. The method of claim 22 , further comprising analyzing the image using a processor.

24. The method of claim 1 , further comprising enumerating a number of colonies formed by microorganisms belonging to the group.

25. The method of claim 1 , wherein the culture device comprises a gelling agent.

26. The method of claim 1 ,

Assignees

• 3M Innovative Properties Co

Inventors

• Binsfeld Chirstine A
• Mach Patrick A
• Celt Mara S
• Stanenas Adam J

Classifications

• C12Q1/10 Primary

  Enterobacteria · CPC title

• C12Q1/34 Primary

  involving hydrolase · CPC title

• C12Q2304/00

  Chemical means of detecting microorganisms · CPC title

• C12Q1/04 Primary

  Determining presence or kind of microorganism; Use of selective media for testing antibiotics or bacteriocides; Compositions containing a chemical indicator therefor {(C12Q1/6897 takes precedence)} · CPC title

• C12Y302/01023

  Beta-galactosidase (3.2.1.23), i.e. exo-(1-->4)-beta-D-galactanase · CPC title