COMPOSITIONS AND METHODS FOR DETECTING CORONAVIRUS NUCLEIC ACID
Disclosed are nucleic acid oligomers, including amplification oligomers and detection probes, for amplification and/or detection of human coronavirus OC43,HKU1, NL63, and/or 229E nucleic acid. Also disclosed are methods of nucleic acid amplification and/or detection using the disclosed oligomers, as well as corresponding reaction mixtures and kits.
1 . (canceled)
2 . (canceled)
3 . A composition for determining the presence or absence of human coronavirus OC43 (OC43) in a sample, said composition comprising a first amplification oligomer combination comprising first and second human coronavirus OC43-specific amplification oligomers capable of amplifying a target region of an OC43 target nucleic acid, wherein the first OC43-specific amplification oligomer comprises a first OC43-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:5, SEQ ID NO:38, SEQ ID NO:39, or SEQ ID NO:24 and including from 0-7 nucleotide analogs, and wherein the second OC43-specific amplification oligomer comprises a second OC43-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:6, SEQ ID NO:43, SEQ ID NO:41, SEQ ID NO:32, SEQ ID NO:44, or SEQ ID NO:51 and including from 0-7 nucleotide analogs.
4 . The composition of claim 3 , further comprising an OC43-specific detection probe comprising a target-hybridizing sequence selected from the group consisting of SEQ ID NO:7, SEQ ID NO:47, SEQ ID NO:33, or SEQ ID NO:34, or an RNA equivalent or DNA/RNA chimeric thereof and including from 0-7 nucleotide analogs.
5 . The composition of claim 4 further comprising a second amplification oligomer combination for determining the presence or absence of human coronavirus HKU1 (HKU1) in a sample, said second amplification oligomer combination comprising first and second human coronavirus HKU1-specific amplification oligomers capable of amplifying a target region of an HKU1 target nucleic acid, wherein the first HKU-specific amplification oligomer comprises a first HKU1-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:8, SEQ ID NO:20, SEQ ID NO:26, SEQ ID NO:40, or SEQ ID NO:25 and including from 0-7 nucleotide analogs, and wherein the second HKU1-specific amplification oligomer comprises a second HKU1-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:9, SEQ ID NO:31, or SEQ ID NO:48 and including from 0-7 nucleotide analogs.
6 . The composition of claim 5 , further comprising an HKU1-specific detection probe comprising a target-hybridizing sequence selected from the group consisting of SEQ ID NO:10, SEQ ID NO:27, or SEQ ID NO:19, or an RNA equivalent or DNA/RNA chimeric thereof and including from 0-7 nucleotide analogs.
7 . The composition of claim 5 further comprising a third amplification oligomer combination for determining the presence or absence of human coronavirus NL63 (NL63) in a sample, said third amplification oligomer combination comprising first and second NL63-specific amplification oligomers capable of amplifying a target region of an NL63 target nucleic acid, wherein the first NL63-specific amplification oligomer comprises a first NL63-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:11, SEQ ID NO:37, SEQ ID NO:50, or SEQ ID NO:30 and including from 0-7 nucleotide analogs, and wherein the second NL63-specific amplification oligomer comprises a second NL63-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:12, SEQ ID NO:49, SEQ ID NO:45, or SEQ ID NO:23 and including from 0-7 nucleotide analogs.
8 . The composition of claim 7 , further comprising an NL63-specific detection probe comprising a target-hybridizing sequence selected from the group consisting of SEQ ID NO:13, SEQ ID NO:18, SEQ ID NO:22, or SEQ ID NO:17, or an RNA equivalent or DNA/RNA chimeric thereof and including from 0-7 nucleotide analogs.
9 . The composition of claim 8 further comprising a fourth amplification oligomer combination for determining the presence or absence of human coronavirus 229E (229E) in a sample, said fourth amplification oligomer combination comprising first and second 229E-specific amplification oligomers capable of amplifying a target region of a 229E target nucleic acid, wherein the first 229E-specific amplification oligomer comprises a first 229E-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:14 or SEQ ID NO:42 and including from 0-7 nucleotide analogs, and wherein the second 229E-specific amplification oligomer comprises a second 229E-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:15 or SEQ ID NO:35 and including from 0-7 nucleotide analogs.
10 . The composition of claim 9 , further comprising a 229E-specific detection probe comprising a target-hybridizing sequence selected from the group consisting of SEQ ID NO:16 or SEQ ID NO:36, or an RNA equivalent or DNA/RNA chimeric thereof and including from 0-7 nucleotide analogs.
11 . (canceled)
12 . (canceled)
13 . A kit for determining the presence or absence of human coronavirus OC43 (OC43) in a sample, said kit comprising a first amplification oligomer combination comprising first and second human coronavirus OC43-specific amplification oligomers capable of amplifying a target region of an OC43 target nucleic acid, wherein the first OC43-specific amplification oligomer comprises a first OC43-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:5, SEQ ID NO:38, SEQ ID NO:39, or SEQ ID NO:24 and including from 0-7 nucleotide analogs, and wherein the second OC43-specific amplification oligomer comprises a second OC43-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:6, SEQ ID NO:43, SEQ ID NO:41, SEQ ID NO:32, SEQ ID NO:44, or SEQ ID NO:51 and including from 0-7 nucleotide analogs.
14 . The kit of claim 13 , further comprising an OC43-specific detection probe comprising a target-hybridizing sequence selected from the group consisting of SEQ ID NO:7, SEQ ID NO:47, SEQ ID NO:33, or SEQ ID NO:34, or an RNA equivalent or DNA/RNA chimeric thereof and including from 0-7 nucleotide analogs.
15 . The kit of claim 14 further comprising a second amplification oligomer combination for determining the presence or absence of human coronavirus HKU1 (HKU1) in a sample, said second amplification oligomer combination comprising first and second human coronavirus HKU1-specific amplification oligomers capable of amplifying a target region of an HKU1 target nucleic acid, wherein the first HKU-specific amplification oligomer comprises a first HKU1-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:8, SEQ ID NO:20, SEQ ID NO:26, SEQ ID NO:40, or SEQ ID NO:25 and including from 0-7 nucleotide analogs, and wherein the second HKU1-specific amplification oligomer comprises a second HKU1-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:9, SEQ ID NO:31, or SEQ ID NO:48 and including from 0-7 nucleotide analogs.
16 . The kit of claim 15 , further comprising an HKU1-specific detection probe comprising a target-hybridizing sequence selected from the group consisting of SEQ ID NO:10, SEQ ID NO:27, or SEQ ID NO:19, or an RNA equivalent or DNA/RNA chimeric thereof and including from 0-7 nucleotide analogs.
17 . The kit of claim 16 further comprising a third amplification oligomer combination for determining the presence or absence of human coronavirus NL63 (NL63) in a sample, said third amplification oligomer combination comprising first and second NL63-specific amplification oligomers capable of amplifying a target region of an NL63 target nucleic acid, wherein the first NL63-specific amplification oligomer comprises a first NL63-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:11, SEQ ID NO:37, SEQ ID NO:50, or SEQ ID NO:30 and including from 0-7 nucleotide analogs, and wherein the second NL63-specific amplification oligomer comprises a second NL63-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:12, SEQ ID NO:49, SEQ ID NO:45, or SEQ ID NO:23 and including from 0-7 nucleotide analogs.
18 . The kit of claim 17 , further comprising an NL63-specific detection probe comprising a target-hybridizing sequence selected from the group consisting of SEQ ID NO:13, SEQ ID NO:18, SEQ ID NO:22, or SEQ ID NO:17, or an RNA equivalent or DNA/RNA chimeric thereof and including from 0-7 nucleotide analogs.
19 . The kit of claim 18 further comprising a fourth amplification oligomer combination for determining the presence or absence of each of human coronavirus 229E (229E) in a sample, said fourth first amplification oligomer combination comprising first and second 229E-specific amplification oligomers capable of amplifying a target region of a 229E target nucleic acid, wherein the first 229E-specific amplification oligomer comprises a first 229E-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:14 or SEQ ID NO:42 and including from 0-7 nucleotide analogs, and wherein the second 229E-specific amplification oligomer comprises a second 229E-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:15 or SEQ ID NO:35 and including from 0-7 nucleotide analogs.
20 . The kit of claim 19 , further comprising a 229E-specific detection probe comprising a target-hybridizing sequence selected from the group consisting of SEQ ID NO:16 or SEQ ID NO:36, or an RNA equivalent or DNA/RNA chimeric thereof and including from 0-7 nucleotide analogs
21 . A method for determining the presence or absence of each of human coronaviruses OC43, HKU1, NL63, and 229E in a sample, said method comprising:
(a) contacting a sample containing or suspected of containing human coronavirus OC43 (OC43), human coronavirus HKU1 (HKU1), human coronavirus NL63 (NL63), and/or human coronavirus 229E (229E) with first, second, third, and fourth amplification oligomer combinations, wherein
(i) the first amplification oligomer combination comprises first and second OC43-specific amplification oligomers capable of amplifying a target region of an OC43 target nucleic acid, wherein the first OC43-specific amplification oligomer comprises a first OC43-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:5, SEQ ID NO:38, SEQ ID NO:39, or SEQ ID NO:24 and including from 0-7 nucleotide analogs, and wherein the second OC43-specific amplification oligomer comprises a second OC43-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:6, SEQ ID NO:43, SEQ ID NO:41, SEQ ID NO:32, SEQ ID NO:44, or SEQ ID NO:51 and including from 0-7 nucleotide analogs;
(ii) the second amplification oligomer combination comprises first and second HKU1-specific amplification oligomers capable of amplifying a target region of an HKU1 target nucleic acid, wherein the first HKU-specific amplification oligomer comprises a first HKU1-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:8, SEQ ID NO:20, SEQ ID NO:26, SEQ ID NO:40, or SEQ ID NO:25 and including from 0-7 nucleotide analogs, and wherein the second HKU1-specific amplification oligomer comprises a second HKU1-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:9, SEQ ID NO:31, or SEQ ID NO:48 and including from 0-7 nucleotide analogs;
(iii) the third amplification oligomer combination comprises first and second NL63-specific amplification oligomers capable of amplifying a target region of an NL63 target nucleic acid, wherein the first NL63-specific amplification oligomer comprises a first NL63-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:11, SEQ ID NO:37, SEQ ID NO:50, or SEQ ID NO:30 and including from 0-7 nucleotide analogs, and wherein the second NL63-specific amplification oligomer comprises a second NL63-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:12, SEQ ID NO:49, SEQ ID NO:45, or SEQ ID NO:23 and including from 0-7 nucleotide analogs; and
(iv) the fourth amplification oligomer combination comprises first and second 229E-specific amplification oligomers capable of amplifying a target region of a 229E target nucleic acid, wherein the first 229E-specific amplification oligomer comprises a first 229E-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:14 or SEQ ID NO:42 and including from 0-7 nucleotide analogs, and wherein the second 229E-specific amplification oligomer comprises a second 229E-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:15 or SEQ ID NO:35 and including from 0-7 nucleotide analogs;
(b) performing an in vitro nucleic acid amplification reaction, wherein any OC43, HKU1, NL63, and/or 229E target nucleic acids, if present in the sample, are used as a template for generating amplicons corresponding the OC43, HKU1, NL63, and/or 229E target regions present in the sample; and
(c) detecting the presence or absence of the amplicons, thereby determining the presence or absence of OC43, HKU1, NL63, and 229E in the sample.
22 . The method of claim 21 , further comprising one or more of
(a) an OC43-specific detection probe comprising a target-hybridizing sequence selected from the group consisting of SEQ ID NO:7, SEQ ID NO:47, SEQ ID NO:33, or SEQ ID NO:34, or an RNA equivalent or DNA/RNA chimeric thereof and including from 0-7 nucleotide analogs;
(b) an HKU1-specific detection probe comprising a target-hybridizing sequence selected from the group consisting of SEQ ID NO:10, SEQ ID NO:27, or SEQ ID NO:19, or an RNA equivalent or DNA/RNA chimeric thereof and including from 0-7 nucleotide analogs;
(c) an NL63-specific detection probe comprising a target-hybridizing sequence selected from the group consisting of SEQ ID NO:13, SEQ ID NO:18, SEQ ID NO:22, or SEQ ID NO:17, or an RNA equivalent or DNA/RNA chimeric thereof and including from 0-7 nucleotide analogs; and
(d) a 229E-specific detection probe comprising a target-hybridizing sequence selected from the group consisting of SEQ ID NO:16 or SEQ ID NO:36, or an RNA equivalent or DNA/RNA chimeric thereof and including from 0-7 nucleotide analogs.
23 - 35 . (canceled)
36 . The composition of claim 10 , wherein
a) the first amplification oligomer combination comprises:
(i) the first OC43-specific amplification oligomer comprising the first OC43-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:5 and the second OC43-specific amplification oligomer comprising the second OC43-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:6 or SEQ ID NO:43;
(ii) the first OC43-specific amplification oligomer comprising the first OC43-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:38 or SEQ ID NO:30 and the second OC43-specific amplification oligomer comprising a second OC43-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:41 or SEQ ID NO:32; or
(iii) the first OC43-specific amplification oligomer comprising the first OC43-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:24 and the second OC43-specific amplification oligomer comprising the second OC43-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:44 or SEQ ID NO:51;
(b) the second amplification oligomer combination comprises:
(i) the first HKU1-specific amplification oligomer comprising the first HKU1-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:8, SEQ ID NO:20, or SEQ ID NO:26 and the second HKU1-specific amplification oligomer comprising the second HKU1-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:9;
(ii) the first HKU1-specific amplification oligomer comprising the first HKU1-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:40 and the second HKU1-specific amplification oligomer comprising the second HKU1-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:31; or
(iii) the first HKU1-specific amplification oligomer comprising the first HKU1-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:25 and the second HKU1-specific amplification oligomer comprising the second HKU1-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:48;
(c) the third amplification oligomer combination comprises:
(i) the first NL63-specific amplification oligomer comprising the first NL63-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:11, and the second NL63-specific amplification oligomer comprising the second NL63-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:12;
(ii) the first NL63-specific amplification oligomer comprising the first NL63-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:37, and the second NL63-specific amplification oligomer comprising the second NL63-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:49; or
(iii) the first NL63-specific amplification oligomer comprising the first NL63-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:50 or SEQ ID NO:30 and the second NL63-specific amplification oligomer comprising the second NL63-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:45 or SEQ ID NO:23;
(d) a fourth amplification oligomer combination comprises:
(i) the first 229E-specific amplification oligomer comprising the first 229E-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:14 and the second 229E-specific amplification oligomer comprising the second 229E-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:15; or
(ii) the first 229E-specific amplification oligomer comprising the first 229E-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:42 and the second 229E-specific amplification oligomer comprising the second 229E-specific target-hybridizing sequence substantially corresponding to the nucleotide sequence of SEQ ID NO:35.