Culture medium for microorganisms including para-aminobenzoic acid as a selective agent

Abstract

The field of the invention is the analysis of target microorganisms in a complex sample. The present invention more particularly relates to a culture medium for the detection of at least one target microorganism including: at least one natural or synthetic fermentation or enzymatic activity substrate, and at least one selective agent, which inhibits non-target microorganisms, constituted by para-amino benzoic acid, one of its derivatives or one of their salts, at a concentration of between 0.05 and 1 g/L.

Claims

1. A culture medium for the detection of at least one target microorganism comprising: at least one natural or synthetic fermentation or enzymatic substrate of the at least one target microorganism; and selective agents, wherein the selective agents consist of: bile salts, at least one agent selected from the group consisting of para-amino benzoic acid, a derivative of para-amino benzoic acid, a salt thereof, and combinations thereof, and optionally at least one of Novobiocin, Vancomycin, Amphotericin, and Cefsulodin; wherein a concentration of the para-amino benzoic acid, a derivative of para-amino benzoic acid, and/or a salt thereof ranges from 0.05 to 1 g/L; the derivative of para-amino benzoic acid comprises a para-amino benzoic skeleton; the at least one target microorganism is a Salmonella genus bacterium; the at least one natural or synthetic fermentation or enzymatic substrate comprises at least one chromogenic or fluorogenic substrate; and a hydrolysation of the at least one natural or synthetic fermentation or enzymatic substrate distinguishes the at least one target microorganism from non-target microorganisms in the culture medium by causing a change in color or fluorescence, respectively, in the target microorganism.

2. The culture medium according to claim 1, wherein the culture medium contains at least one component that promotes a culturing of the Salmonella genus bacterium.

3. The culture medium according to claim 1, wherein the non-target microorganisms are selected from the group consisting of enterobacteria, except a Salmonella genus bacterium, and the culture medium contains at least one component that inhibits or slows a culturing of the non-target microorganisms.

4. The culture medium according to claim 1, wherein the derivative of para-amino benzoic acid is selected from the group consisting of 4-(methylamino) benzoic acid, 4-amino-2-chlorobenzoic acid, 4-amino-3-nitro benzoic acid, and 4-aminosalicylic acid.

5. The culture medium according to claim 1, wherein the bile salts comprise at least sodium deoxycholate.

6. The culture medium according to claim 1, wherein the selective agents include the at least one of Novobiocin, Vancomycin, Amphotericin, and Cefsulodin.

7. The culture medium according to claim 1, wherein the at least one natural or synthetic fermentation or enzymatic substrate is a metabolic substrate.

8. The culture medium according to claim 7, wherein a degradation product of the metabolic substrate varies a pH of the culture medium.

9. The culture medium according to claim 1, wherein the concentration of the para-amino benzoic acid, a derivative of para amino benzoic acid, and/or a salt thereof ranges from 0.3 to 1 g/L.

10. The culture medium according to claim 1, wherein the concentration of the para-amino benzoic acid, a derivative of para-amino benzoic acid, and/or a salt thereof ranges from 0.5 to 1 g/L.

11. A culture medium for the detection of at least one target microorganism comprising: at least three chromogenic substrates; and selective agents, wherein the selective agents consist of: bile salts, at least one agent selected from the group consisting of para-amino benzoic acid, a derivative of para-amino benzoic acid, a salt thereof, and combinations thereof, and optionally at least one of Novobiocin, Vancomycin, Amphotericin, and Cefsulodin; wherein a concentration of the at least one agent ranges from 0.05 to 1 g/L; the derivative of para-amino benzoic acid comprises a para-amino benzoic skeleton; the at least one target microorganism is a Salmonella genus bacterium; and a hydrolysation of one or more of the at least three chromogenic substrates distinguishes the at least one target microorganism from non-target microorganisms in the culture medium by causing a change in color in the target microorganism.

12. A process for detecting a target microorganism, in a mixed population of target and non-target microorganisms, comprising: placing a sample, in which it is desired to detect the target microorganism in the sample, in contact with the culture medium according to claim 1; incubating the culture medium at a temperature that promotes reproduction of the target microorganism; and observing the microorganisms on the culture medium, wherein the target microorganism is a Salmonella genus bacterium.

13. A method comprising: detecting in vitro at least one target microorganism with the culture medium according to claim 1, wherein the at least one target microorganism is a Salmonella genus bacterium.

14. A method comprising: detecting at least one target microorganism with the culture medium of claim 1.

Description

EXAMPLES

Example 1Culture Medium for the Detection of Salmonella Genus Bacteria Including 3 Chromogenic Substrates, a Combination of Antibiotics (Novobiocin, Cefsulodin, Vancomycin, Amphotericin) and Different Concentrations of Para-Aminobenzoic Acid (PABA)

(1) 1. Preparation of the Tested Media

(2) The tested media are as follows: Control medium: chromID Salmonella medium (ref. 43621) marketed by the applicant, containing Novobiocin, Vancomycin, Amphotericin and Cefsulodin and the three chromogenic substrates Medium 1: Control medium, with addition of 0.1 g/L PABA, Medium 2: Control medium, with addition of 0.3 g/L PABA, Medium 3: Control medium, without antibiotics and with addition of 0.5 g/L PABA,
These media are media in 90-millimeter Petri dishes.

(3) 2. Seeding and Reading of the Media

(4) Strains of different Salmonella species or serovars (S. enteritidis, S. thyphimurium, S. cabana, S. gallinarum, S. paratyphi for example) and other microbe genera such as the gram-negative bacteria (E. coli, Enterobacter, Citrobacter, Proteus, Hafnia), all from the Applicant's collection, were suspended in physiological water, then seeded on the media, in accordance with the 4-quadrant technique. The strains were incubated at 37 C. for 24 hours. The colonies formed were examined visually after 24 hours of incubation.

(5) 3. Results:

(6) The results obtained are presented in table 1 below.

(7) TABLE-US-00001 TABLE 1 Gram strains - other than Salmonella with Salmonella growth/Gram strains - detected/Salmonella other than Salmonella Medium tested tested Control 9/9 8/10 1 9/9 6/10 2 9/9 7/10 3 9/9 1/5

(8) These results show a good degree of sensitivity of detection of the Salmonella strains by all of the culture media. However, only media 1 to 3 decrease the growth of other Gram-negative bacteria.

Example 2Culture Medium for the Detection of Salmonella Genus Bacteria Including 3 Chromogenic Substrates, a Combination of Selective Elements (Novobiocin, Cefsulodin, Vancomycin, Amphotericin, Bile Salts, Sodium Deoxycholate) and Para-Aminobenzoic Acid (PABA)

(9) 1. Preparation of the Medium According to the Invention

(10) The media tested in the experiments were as follows: Control Medium: chromID Salmonella medium (ref. 43621) containing Novobiocin, Vancomycin, Amphotericin, Cefsulodin, 1.5 g/L bile salts and 3 chromogenic substrates Medium 1: Control medium, with addition of 0.1 g/L PABA, bile salts increased to 3 g/L Medium 2: Control medium, with addition of 0.1 g/L PABA, bile salts increased to 3 g/L and 5 g/L sodium deoxycholate

(11) 2. Seeding and Reading of the Media

(12) Strains belonging to different species or serovars of Salmonella (S. enteritidis, S. thyphimurium, S. cubana, S. gallinarum, S. paratyphi, S. arizonae for example) and other microbe genera, such as the gram-negative bacteria (Serratia, Enterobacter, Citrobacter, Proteus, Hafnia) all from the Applicant's collection, were suspended in physiological water, then seeded on the media, in accordance with the 4-quadrant technique. The dishes were incubated at 37 C. for 24 hours. The colonies formed were examined visually after 24 hours of incubation.

(13) 3. Results:

(14) The results obtained are presented in table 2 below.

(15) TABLE-US-00002 TABLE 2 Salmonella Bacteria genera other than detected/Salmonella Salmonella with Medium tested growth/other bacteria genera tested C 9/10 5/6 1 9/10 3/6 2 10/10 1/6

(16) These results show an excellent degree of sensitivity of all of the media for seeking Salmonella strains, with detection of all of the 10 Salmonella strains for formula 2.

(17) However, only media 1 and 2 make it possible to decrease the growth of bacteria genera other than Salmonella. The best selectivity is obtained by associating PABA with bile salts such as sodium deoxycholate.