Compositions, kits and related methods for the detection and/or monitoring of Salmonella

• US 8,703,421 B2
• Filed: 05/29/2009
• Issued: 04/22/2014
• Est. Priority Date: 05/30/2008
• Status: Active Grant

• Current Assignee: Gen-Probe Incorporated (Hologic Incorporated)
• Original Assignee: Gen-Probe Incorporated (Hologic Incorporated)
• Inventors: Reshatoff, Michael R., Ong, Edgar O., Hogan, James J.
• Primary Examiner(s): Horlick, Kenneth R.
• Assistant Examiner(s): Tung, Joyce
• Application Number:
  US12/995,168
• Publication Number:
  US 20110117564A1
• Time in Patent Office: 1,789 Days
• Field of Search: 435/6.12, 435/91.2, 536/24.33
• US Class Current: 435/6.12
• CPC Class Codes: C12Q 1/68 involving nucleic acids
• CPC Class Codes: C12Q 1/689 for bacteria
• CPC Class Codes: C12Q 2304/00 Chemical means of detecting microorganisms hydrolase substrates C12Q2334/00, peptidase substrates C12Q2337/00
• CPC Class Codes: Y02A 50/30 Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Abstract

Provided are compositions, kits, and methods for the identification of Salmonella. In certain aspects and embodiments, the compositions, kits, and methods may provide improvements in relation to specificity, sensitivity, and speed of detection.

Claims

1. A composition for use in a nucleic acid amplification assay to detect Salmonella in a sample, comprising a T7 provider oligonucleotide and a primer oligonucleotide that each target a sequence in a region of Salmonella nucleic acid,

wherein said T7 provider oligonucleotide and primer oligonucleotide used in said amplification assay target opposite strands of the Salmonella nucleic acid sequence to be amplified;

and

wherein said T7 provider oligonucleotide targets a sequence in a region of Salmonella nucleic acid corresponding to bases from about 268-320 of E. coli 23S rRNA and/or said primer oligonucleotide targets a sequence in a region of Salmonella nucleic acid primer oligonucleotide targets a sequence in a region of Salmonella nucleic acid corresponding to bases 338-395 of E. coli 23S rRNA.

2. The composition of claim 1, wherein said T7 provider oligonucleotide targets a sequence in a region of Salmonella nucleic acid corresponding to bases 268-302 or to bases 279-310 of E. coli 23S rRNA, and wherein said primer oligonucleotide targets a sequence in a region of Salmonella nucleic acid corresponding to bases 349-374 of E. coli 23S rRNA.

3. The composition of claim 1 or claim 2, wherein said T7 provider oligonucleotide and said primer oligonucleotide comprises 15-35 nucleotides that are at least 90% complementary to the targeted Salmonella nucleic acid sequence.

4. The composition of claim 1, wherein said T7 provider oligonucleotide is selected from the sequences of SEQ ID NOs:

1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, and 26.

5. The composition of claim 1, wherein said primer oligonucleotide is selected from the sequences of SEQ ID NOs:

35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, and 51.

6. The composition of claim 1, wherein said T7 provider oligonucleotide is the sequence of SEQ ID NO:

17 or complement thereof, and said primer oligonucleotide is the sequence of SEQ ID NO;

50.

7. The composition of claim 1, wherein said T7 provider oligonucleotide is the sequence of SEQ ID NO:

26 or complement thereof, and said primer oligonucleotide is the sequence of SEQ ID NO;

49.

8. The composition of claim 1 or claim 2, further comprising a blocker oligonucleotide.

9. A kit for use in a Salmonella nucleic acid amplification assay comprising the composition according to claim 1.

10. The kit of claim 9, further comprising a detection oligonucleotide.

11. The kit of claim 10, wherein said detection oligonucleotide is a torch oligonucleotide or molecular beacon.

12. A method for detecting Salmonella in a sample, said method comprising performing a nucleic acid amplification assay using the T7 provider oligonucleotide and the primer oligonucleotide composition of claim 1.

13. The method of claim 12, wherein detecting said amplified nucleic acid occurs in real-time.

14. The method of claim 12 or claim 13, wherein Salmonella nucleic acid is specifically detected in the presence of at least one of:

E. coli, Enterobacter, Citrobacter, Shigella, Pseudomonas and Listeria.

15. The method of claim 12, wherein Salmonella nucleic acid is specifically detected in the presence of at least one of:

E. coli, E. vulneris, E. hermannii, E. cloacae, E. aerogenes, E. hoshinae, P. mirabilis, C. brakii, P. fluorescens, S. flexneri, C. freundii, C. koseri/diversus, K. pneumoniae, S. marcescens, L. innocua, E. faecalis, C. jejuni, C. coli, and S. pneumoniae.