Compositions and Methods for Detecting Group A Streptococcus

1 . A composition or kit comprising at least first and second amplification oligomers, wherein:

(a) the first amplification oligomer and second amplification oligomer are configured to amplify a Group A Streptococcus (GAS) amplicon;

(b) the first amplification oligomer:

(i) comprises a first sequence which is SEQ ID NO: 1, 4, 7, 10, 13, 16, 19, 22, 27, 30, 31, 34, or 37;

(ii) comprises a sequence with 0, 1, or 2 mismatches to a first sequence which is SEQ ID NO: 1, 4, 7, 10, 13, 16, 19, 22, 27, 30, 31, 34, or 37;

(iii) competes for hybridization to a Group A Streptococcus (GAS) nucleic acid under stringent conditions with an amplification oligomer whose sequence consists of a first sequence which is SEQ ID NO: 1, 4, 7, 10, 13, 16, 19, 22, 27, 30, 31, 34, or 37; or

(iv) comprises at least 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 contiguous nucleotides of a first sequence which is SEQ ID NO: 1, 4, 7, 10, 13, 16, 19, 22, 27, 30, 31, 34, or 37; and

(c) the second amplification oligomer:

(i) comprises a second sequence which is SEQ ID NO: 3, 6, 9, 12, 15, 17, 20, 23, 28, 32, 35, or 38;

(ii) comprises a sequence with 0, 1, or 2 mismatches to a second sequence which is SEQ ID NO: 3, 6, 9, 12, 15, 17, 20, 23, 28, 32, 35, or 38;

(iii) competes for hybridization to a Group A Streptococcus (GAS) nucleic acid under stringent conditions with an amplification oligomer whose sequence consists of a second sequence which is SEQ ID NO: 3, 6, 9, 12, 15, 17, 20, 23, 28, 32, 35, or 38; or

(iv) comprises at least 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 contiguous nucleotides of a second sequence which is SEQ ID NO: 3, 6, 9, 12, 15, 17, 20, 23, 28, 32, 35, or 38.

2 . A method of detecting GAS in a sample, the method comprising:

contacting the sample with at least first and second amplification oligomers, thereby forming a composition,

performing a nucleic acid amplification reaction in the composition which produces a GAS amplicon in the presence of a GAS nucleic acid, and

detecting the presence or absence of the at least one amplicon,

wherein (a) the first amplification oligomer:

(i) comprises a first sequence which is SEQ ID NO: 1, 4, 7, 10, 13, 16, 19, 22, 27, 30, 31, 34, or 37;

(ii) comprises a sequence with 0, 1, or 2 mismatches to a first sequence which is SEQ ID NO: 1, 4, 7, 10, 13, 16, 19, 22, 27, 30, 31, 34, or 37;

(iii) competes for hybridization to a Group A Streptococcus (GAS) nucleic acid under stringent conditions with an amplification oligomer whose sequence consists of a first sequence which is SEQ ID NO: 1, 4, 7, 10, 13, 16, 19, 22, 27, 30, 31, 34, or 37; or

(iv) comprises at least 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 contiguous nucleotides of a first sequence which is SEQ ID NO: 1, 4, 7, 10, 13, 16, 19, 22, 27, 30, 31, 34, or 37; and

(b) the second amplification oligomer:

(i) comprises a second sequence which is SEQ ID NO: 3, 6, 9, 12, 15, 17, 20, 23, 28, 32, 35, or 38;

(ii) comprises a sequence with 0, 1, or 2 mismatches to a second sequence which is SEQ ID NO: 3, 6, 9, 12, 15, 17, 20, 23, 28, 32, 35, or 38;

(iii) competes for hybridization to a Group A Streptococcus (GAS) nucleic acid under stringent conditions with an amplification oligomer whose sequence consists of a second sequence which is SEQ ID NO: 3, 6, 9, 12, 15, 17, 20, 23, 28, 32, 35, or 38; or

(iv) comprises at least 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 contiguous nucleotides of a second sequence which is SEQ ID NO: 3, 6, 9, 12, 15, 17, 20, 23, 28, 32, 35, or 38.

3 . The composition or kit of claim 1 , wherein the first amplification oligomer competes for hybridization to a Group A Streptococcus (GAS) nucleic acid under stringent conditions with an amplification oligomer whose sequence consists of a first sequence which is SEQ ID NO: 1, 4, 7, 10, 13, 16, 19, 22, 27, 30, 31, 34, or 37.

4 . The composition or kit of claim 1 , wherein the first sequence is the sequence of SEQ ID NO: 1.

5 .- 16 . (canceled)

17 . The composition or kit of claim 1 , wherein the second sequence is the sequence of SEQ ID NO: 3.

18 .- 28 . (canceled)

29 . The composition or kit of claim 1 , wherein the composition or kit further comprises a third oligomer, and the third oligomer:

(i) comprises a third sequence which is SEQ ID NO: 2, 5, 8, 11, 14, 18, 25, 26, 21, 24, 29, 33, 36, or 39;

(ii) comprises a sequence with 0, 1, or 2 mismatches to a third sequence which is SEQ ID NO: 2, 5, 8, 11, 14, 18, 25, 26, 21, 24, 29, 33, 36, or 39;

(iii) competes for hybridization to a Group A Streptococcus (GAS) nucleic acid under stringent conditions with an oligomer whose sequence consists of a third sequence which is SEQ ID NO: 2, 5, 8, 11, 14, 18, 25, 26, 21, 24, 29, 33, 36, or 39; or

(iv) comprises at least 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 contiguous nucleotides of a third sequence which is SEQ ID NO: 2, 5, 8, 11, 14, 18, 25, 26, 21, 24, 29, 33, 36, or 39.

30 .- 33 . (canceled)

34 . The composition or kit of claim 29 , wherein the third oligomer comprises a detectable label.

35 . The composition or kit of claim 34 , wherein the detectable label is fluorescent.

36 . The composition or kit of claim 35 , wherein the third oligomer comprising a fluorescent label further comprises a quencher.

37 . The composition or kit of claim 35 , wherein at least one oligomer comprising a fluorescent label is a non-extendable oligomer.

38 . The composition or kit of claim 1 , wherein at least one oligomer comprises one or more 5-methyl-dC residues.

39 . The composition or kit of claim 38 , wherein the one or more 5-methyl-dC residues comprise residues corresponding to one or more, or all, of positions 8, 18, and 24 of SEQ ID NO: 4, and/or wherein the one or more 5-methyl-dC residues comprise residues corresponding to one or more, or all, of positions 2, 4, 15, 19, and 21 of SEQ ID NO: 5, and/or wherein the one or more 5-methyl-dC residues comprise residues corresponding to one or more, or all, of positions 7, 8, 11, and 13 of SEQ ID NO: 6, and/or wherein the one or more 5-methyl-dC residues comprise residues corresponding to one or more, or all, of positions 7, 8, 10, and 21 of SEQ ID NO: 16, and/or wherein the one or more 5-methyl-dC residues comprise residues corresponding to one or more, or all, of positions 6, 7, 9, 11, 12, 15, and 19 of SEQ ID NO: 18, and/or wherein the one or more 5-methyl-dC residues comprise residues corresponding to one or more, or all, of positions 17 and 23 of SEQ ID NO: 17, and/or wherein the one or more 5-methyl-dC residues comprise residues corresponding to one or more, or all, of positions 5, 7, 11, 13, and 19 of SEQ ID NO: 23, and/or wherein the one or more 5-methyl-dC residues comprise residues corresponding to one or more, or all, of positions 17, 23, 25, 26, and 28 of SEQ ID NO: 25, and/or wherein the one or more 5-methyl-dC residues comprise residues corresponding to one or more, or all, of positions 8, 14, 15, 16, 17, and 28 of SEQ ID NO: 26.

40 . The method of claim 2 , wherein:

the presence or absence of the amplicon is detected according to the occurrence or non-occurrence of hybridization of the third oligomer to the amplicon.

41 . The method of claim 2 , wherein the nucleic acid amplification reaction comprises PCR.

42 . The method of claim 2 , wherein the nucleic acid amplification reaction comprises PCR with a polymerase with 5′-to-3′ exonuclease activity, and detecting an amplicon using a probe oligomer comprising a fluorophore and a quencher, wherein exonucleolysis of the probe by the polymerase reduces quenching of fluoresence by the quencher.

43 . The composition or kit of claim 1 , wherein the GAS amplicon is about 100-200 nucleotides in length.

44 . (canceled)

45 . (canceled)

46 . The composition of claim 1 , wherein the composition is aqueous, frozen, or lyophilized.

47 . A method for detecting or quantifying a GAS nucleic acid in a sample, comprising utilizing the composition or kit of claim 1 to detect or quantify the GAS nucleic acid.

48 . A method of making an amplification oligomer recited in claim 1 , comprising the step of chemically synthesizing the amplification oligomer using sequential solid phase oligonucleotide synthesis.

49 . (canceled)