Compositions and methods for detecting or quantifying Human Parainfluenza Virus 4
Compositions, methods, kits, and uses are provided for detecting or quantifying an Human Parainfluenza virus 1 (HPIV-1), HPIV-2, HPIV-3, and/or HPIV-4 nucleic acid, e.g., using nucleic acid amplification and hybridization assays. In some embodiments, the compositions, methods, kits, and uses target the HN gene of HPIV-1, HPIV-2, and/or HPIV-3 and/or the NP gene of HPIV-4.
1. A composition comprising at least a first HPIV-4 amplification oligomer and a second HPIV-4 amplification oligomer, wherein:
the first HPIV-4 amplification oligomer and second HPIV-4 amplification oligomer are configured to amplify an HPIV-4 amplicon, wherein the first HPIV-4 amplification oligomer comprises a target-hybridizing region consisting of the sequence of SEQ ID NO: 78 and the second HPIV-4 amplification oligomer comprises a target-hybridizing region consisting of the sequence of SEQ ID NO: 56, and
wherein the composition further comprises a third HPIV-4 oligomer configured to hybridize to the HPIV-4 amplicon, and the third oligomer comprises a chemiluminescent label or a fluorescent label.
2. The composition of claim 1 , wherein the third HPIV-4 oligomer comprises the sequence of SEQ ID NO: 71 or a sequence with one or two mismatches to SEQ ID NO: 71.
3. The composition of claim 2 , wherein the third HPIV-4 oligomer comprises the sequence of SEQ ID NO: 71.
4. The composition of claim 1 , wherein at least one of the first, second, and third HPIV-4 oligomers comprises at least one 5-methylcytosine.
5. A method of detecting HPIV-4 in a sample, comprising:
contacting the sample with at least a first HPIV-4 amplification oligomer and a second HPIV-4 amplification oligomer, wherein the first HPIV-4 amplification oligomer and second HPIV-4 amplification oligomer are configured to amplify an HPIV-4 amplicon, wherein the first HPIV-4 amplification oligomer comprises a target-hybridizing region consisting of the sequence of SEQ ID NO: 78 and the second HPIV-4 amplification oligomer comprises a target-hybridizing region consisting of the sequence of SEQ ID NO: 56,
performing a nucleic acid amplification reaction which produces the HPIV-4 amplicon in the presence of an HPIV-4 nucleic acid, and
detecting the presence or absence of the HPIV-4 amplicon.
6. The method of claim 5 , wherein the nucleic acid amplification reaction comprises PCR with a polymerase with 5′-to-3′ exonuclease activity, and detecting the HPIV-4 amplicon using a probe oligomer comprising a fluorophore and a quencher, wherein exonucleolysis of the probe by the polymerase reduces quenching of fluorescence by the quencher.
7. A reaction mixture for detecting HPIV-4 in a sample, comprising:
(i) at least a first HPIV-4 amplification oligomer and a second HPIV-4 amplification oligomer, wherein the first HPIV-4 amplification oligomer and second HPIV-4 amplification oligomer are configured to amplify an HPIV-4 amplicon, wherein the first HPIV-4 amplification oligomer comprises a target-hybridizing region consisting of the sequence of SEQ ID NO: 78 and the second HPIV-4 amplification oligomer comprises a target-hybridizing region consisting of the sequence of SEQ ID NO: 56, and
(ii) a third HPIV-4 oligomer configured to hybridize to the HPIV-4 amplicon, wherein the third oligomer comprises a detectable chemiluminescent label or a fluorescent label,
wherein the reaction mixture is aqueous or lyophilized.
8. The method of claim 6 , wherein the probe oligomer comprises the sequence of SEQ ID NO: 71 or a sequence with one or two mismatches to SEQ ID NO: 71.
9. The method of claim 8 , wherein the probe oligomer comprises the sequence of SEQ ID NO: 71.
10. The reaction mixture of claim 7 , wherein the reaction mixture further comprises at least one enzyme selected from the group consisting of a DNA polymerase, a reverse transcriptase, and an RNA polymerase.
11. The reaction mixture of claim 7 , wherein the third HPIV-4 oligomer comprises the sequence of SEQ ID NO: 71 or a sequence with one or two mismatches to SEQ ID NO: 71.
12. The reaction mixture of claim 11 , wherein the third HPIV-4 oligomer comprises the sequence of SEQ ID NO: 71.
13. The reaction mixture of claim 7 , wherein at least one of the first, second, and third HPIV-4 oligomers comprises at least one 5-methylcytosine.