Method of detecting a Salmonella microorganism

Patent metadata

Field	Value
Publication number	US-10519481-B2
Application number	US-201715593598-A
Country	US
Kind code	B2
Filing date	May 12, 2017
Priority date	Dec 28, 2011
Publication date	Dec 31, 2019
Grant date	Dec 31, 2019

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 a microorganism, 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; and
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 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.;
detecting the presence of the first detectable product;
detecting the absence of the second detectable product; and
detecting the presence or absence third detectable product;
wherein observing the presence of the third detectable product indicates the presence of a Salmonella microorganism in the sample; and
wherein observing the absence of the third detectable product indicates the absence of a Salmonella microorganism in the sample.

2. The method of claim 1 , wherein observing the absence of the third detectable product indicates the presence of a non- Salmonella microorganism in the sample.

3. The method of claim 1 , wherein detecting the the third detectable product indicates the presence of a Salmonella microorganism that is selected from the group consisting of S. enterica, S. bongori , and combinations thereof.

4. The method of claim 3 , wherein detecting the presence of the third detectable product indicates the presence of a Salmonella microorganism of the species S. bongori.

5. The method of claim 1 , further comprising enumerating a number of colonies formed by microorganisms belonging to a group consisting of Salmonella microorganisms and β-galactosidase-producing microorganisms.

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

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

8. The method of claim 7 , further comprising analyzing the image using a processor.

9. 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.

10. 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.

11. The method of claim 1 , wherein the second differential indicator compound is selected from the group consisting of 5-bromo-4-chloro-3-indolyl-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.

12. 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.

13. 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.

14. The method of claim 13 , wherein the at least one second selective agent comprises a combination of nalidixic acid, streptomycin, and polymyxin B.

15. The method of claim 1 , 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.

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

17. The method of claim 1 , wherein the third differential indicator system comprises a pH indicator and a nutrient, a chromogenic enzyme substrate, or a combination thereof.

18. The method of claim 1 , wherein the third detectable product is optically distinguishable from the first detectable product and the second detectable product.

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

20. The method of claim 19 , wherein the enzyme substrate comprises a substance that 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 to detect caprylate esterase enzyme activity.

Assignees

• 3M Innovative Properties Co

Inventors

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

Classifications

• C12Q1/34

  involving hydrolase · 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

• C12Q2334/00

  O-linked chromogens for determinations of hydrolase enzymes, e.g. glycosidases, phosphatases, esterases · CPC title

• C12Q1/10 Primary

  Enterobacteria · CPC title