Methods of detecting sars-cov-2, influenza, and rsv

A composition comprising sets of primers that detect the presence of an influenza A gene, an influenza B gene, a RSV gene, and a SARS-CoV-2 gene,

• a. wherein the set of primers that detects the presence or absence of influenza A comprises at least one of:
• i. a set of primers that detects influenza A PB2 selected from:
• 1. a forward and reverse primer for detecting a sequence of the influenza PB2 gene;
• 2. a forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 1, and a reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% complementary to at least 15 contiguous nucleotides of SEQ ID NO:
• 1;
• and
• 3. a forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 17, and a reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 18;
• ii. a set of primers that detects influenza A PA selected from:
• 1. a forward and reverse primer for detecting a sequence of the influenza PA gene;
• 2. a forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 2, and a reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% complementary to at least 15 contiguous nucleotides of SEQ ID NO:
• 2;
• and
• 3. a forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 20, and a reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 21;
• iii. a set of primers that detects influenza A MP selected from:
• 1. a forward and reverse primer for detecting a sequence of the influenza A MP gene;
• 2. a forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 3, and a reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% complementary to at least 15 contiguous nucleotides of SEQ ID NO:
• 3;
• and
• 3. a forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 23, and a reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 24;
• iv. a set of primers that detects avian influenza MP selected from:
• 1. a forward and reverse primer for detecting a sequence of the avian influenza MP gene;
• 2. a forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 4, and a reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% complementary to at least 15 contiguous nucleotides of SEQ ID NO:
• 4;
• and
• 3. a forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 26, and a reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 27;
• b. wherein the set of primers that detects the presence or absence of influenza B comprises at least one of:
• i. a set of primers that detects influenza B MP selected from:
• 1. a forward and reverse primer for detecting a sequence of the influenza B MP gene;
• 2. a forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 6, and a reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% complementary to at least 15 contiguous nucleotides of SEQ ID NO:
• 6;
• and
• 3. a forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 32, and a reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 33;
• ii. a set of primers that detects influenza B NS selected from:
• 1. a forward and reverse primer for detecting a sequence of the influenza B NS gene;
• 2. a forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 7, and a reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% complementary to at least 15 contiguous nucleotides of SEQ ID NO:
• 7;
• and
• 3. a forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 35, and a reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 36;
• c. wherein the set of primers that detects the presence or absence of RSV comprises at least one of:
• i. a set of primers that detects RSV A selected from:
• 1. at least one forward and at least one reverse primer for detecting a sequence of the RSV A gene;
• and
• 2. at least one forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 38 and/or SEQ ID NO:
• 67, and at least one reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% complementary identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 39, SEQ ID NO:
• 68, and/or SEQ ID NO:
• 69;
• and
• ii. a set of primers that detects RSV B selected from:
• 1. a forward and reverse primer for detecting a sequence of the RSV B gene;
• and
• 2. a forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 41, and a reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 42;
• and
• d. wherein the set of primers that detects the presence or absence of SARS-CoV-2 comprises at least one of:
• i. a set of primers that detects SARS-CoV-2 E selected from:
• 1. a forward and reverse primer for detecting a sequence of the SARS-CoV-2 E gene;
• 2. a forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 44, and a reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% complementary to at least 15 contiguous nucleotides of SEQ ID NO:
• 44;
• 3. a forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 70, and a reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 71;
• and
• 4. a forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 48, and a reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 49;
• ii. a set of primers that detects SARS-CoV-2 N2 selected from:
• 1. a forward and reverse primer for detecting a sequence of the SARS-CoV-2 N2 gene;
• 2. a forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 45, and a reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% complementary to at least 15 contiguous nucleotides of SEQ ID NO:
• 45;
• and
• 3. a forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 73 or SEQ ID NO:
• 51, and a reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 52
• iii. a set of primers that detects SARS-CoV-2 RdRP selected from:
• 1. a forward and at least one reverse primer for detecting a sequence of the SARS-CoV-2 RdRP gene;
• 2. a forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 76, and at least one reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 61 and/or SEQ ID NO:
• 78.

A method of detecting the presence or absence of SARS-CoV-2 in a biological sample from a subject and/or determining whether a subject has COVID-19, comprising:

• a. contacting a biological sample from the subject with sets of primers that detect a SARS-CoV-2 E, SARS-CoV-2 N2 gene, and/or SARS-CoV-2 RdRP gene;
• b. conducting one or more polymerase chain reaction (PCR);
• and
• c. detecting an amplicon that is produced by the PCR;
• wherein the set of primers that detects the presence or absence of SARS-CoV-2 E, SARS-CoV-2 N2 gene, and/or SARS-CoV-2 RdRP genes comprises at least one of:
• a) a set of primers that detects SARS-CoV-2 E selected from:
• i. a forward and reverse primer for detecting a sequence of the SARS-CoV-2 E gene;
• ii. a forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 44, and a reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% complementary to at least 15 contiguous nucleotides of SEQ ID NO:
• 44;
• and
• iii. a forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 70, and a reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 71;
• and
• iv. a forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 48, and a reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 49;
• b) a set of primers that detects SARS-CoV-2 N2 selected from:
• i) a forward and reverse primer for detecting a sequence of the SARS-CoV-2 N2 gene;
• ii) a forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 45, and a reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% complementary to at least 15 contiguous nucleotides of SEQ ID NO:
• 45;
• and
• iii) a forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 73 or SEQ ID NO:
• 51, and a reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 52;
• and
• c) a set of primers that detects SARS-CoV-2 RdRP selected from:
• i) a forward and at least one reverse primer for detecting a sequence of the SARS-CoV-2 RdRP gene;
• ii) a forward primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 76, and at least one reverse primer comprising a region of at least 15 contiguous nucleotides having a sequence that is at least 85% identical to at least 15 contiguous nucleotides of SEQ ID NO:
• 61 and/or SEQ ID NO:
• 78.