Assay for the diagnosis of dermatophytosis

A nucleic acid probe, comprising:

• a modified nucleic acid comprising a 5′ C6-amino-linker,
• wherein the modified nucleic acid is capable of hybridizing specifically to a nucleic acid sequence from a single species of a human pathogenic dermatophyte that causes at least one of a skin, hair, and nail infection, the human pathogenic dermatophyte belonging to a Trichophyton genus, wherein the nucleic acid sequence from the human pathogenic dermatophyte comprises at least one of the sequence that is at least 98% identical to the full-length of SEQ ID NO:
• 1, a strand complementary to SEQ ID NO:
• 1, or SEQ ID NO:
• 1 in a vector or cell,
• wherein the modified nucleic acid is at least 95% identical to at least one of the full-length of SEQ ID NO:
• 1, the strand complementary to SEQ ID NO:
• 1, or SEQ ID NO:
• 1 in the vector or cell, over a length of at least 15 consecutive nucleotides, and
• wherein the length of the nucleic acid probe is no more than 200 nucleotides.

A method, comprising:

• a) obtaining a sample of at least one of nail material, hair material, or skin material, from a patient,
• b) amplifying, from the sample of the patient, a nucleic acid from a human pathogenic dermatophyte comprising at least one of the sequence that is at least 98% identical to the full-length of SEQ ID NO:
• 1, a strand complementary to SEQ ID NO:
• 1, or SEQ ID NO:
• 1 in a cell, the human pathogenic dermatophyte belonging to a Trichophyton genus, using a primer pair, thereby generating an amplicon, wherein the primer pair comprises:
• a forward primer having a length of 14 to 30 nucleotides, being at least 90% identical to Trichophyton sequence over the full-length of the forward primer, and having a final base of the forward primer no more than 200 bp away from a first base of SEQ ID NO:
• 1 in a 5′ to 3′ orientation, and
• a reverse primer having a length of 14 to 30 nucleotides, being at least 90% identical to Trichophyton sequence over the full-length of the reverse primer, and having a final base of the reverse primer no more than 200 bp away from a final base of SEQ ID NO:
• 1 in the 5′ to 3′ orientation,
• wherein each of the forward primer and the reverse primer are labeled with a label and form the primer pair, wherein the label is selected from the group consisting of a fluorescent label, a radioactive label, a colloidal gold label, and an enzymatically active label.

A primer pair, comprising:

• a forward primer having a length of 14 to 30 nucleotides, being at least 90% identical to Trichophyton sequence over the full-length of the forward primer, and having a final base of the forward primer no more than 200 bp away from a first base of SEQ ID NO:
• 1 in a 5′ to 3′ orientation, and
• a reverse primer having a length of 14 to 30 nucleotides, being at least 90% identical to Trichophyton sequence over the full-length of the reverse primer, and having a final base of the reverse primer no more than 200 bp away from a final base of SEQ ID NO:
• 1 in the 5′ to 3′ orientation,
• wherein each of the forward primer and the reverse primer are labeled with a label and form the primer pair, wherein the label is selected from the group consisting of a fluorescent label, a radioactive label, a colloidal gold label, and an enzymatically active label,
• wherein the forward and reverse primer, in combination, amplifies a nucleic acid comprising SEQ ID NO:
• 1 from a human pathogenic dermatophyte that causes at least one of a skin, hair, and nail infection, the human pathogenic dermatophyte belonging to a Trichophyton genus.

A kit, for the diagnosis of at least one of a skin, hair, and nail infection caused by a human pathogenic dermatophyte, comprising:

• a primer pair comprising:
• a forward primer having a length of 14 to 30 nucleotides, being at least 90% identical to Trichophyton sequence over the full-length of the forward primer, and having a final base of the forward primer no more than 200 bp away from a first base of SEQ ID NO:
• 1 in a 5′ to 3′ orientation,
• a reverse primer having a length of 14 to 30 nucleotides, being at least 90% identical to Trichophyton sequence over the full-length of the reverse primer, and having a final base of the reverse primer no more than 200 bp away from a final base of SEQ ID NO:
• 1 in the 5′ to 3′ orientation,
• wherein each of the forward primer and the reverse primer are labeled with a label and form the primer pair, wherein the label is selected from the group consisting of a fluorescent label, a radioactive label, a colloidal gold label, and an enzymatically active label,
• wherein the forward primer and reverse primer, in combination, amplify a nucleic acid comprising SEQ ID NO:
• 1 from the human pathogenic dermatophyte belonging to a Trichophyton genus, and
• a modified nucleic acid probe comprising a 5′ C6-amino-linker and capable of hybridizing specifically to the nucleic acid amplified by the forward primer and the reverse primer, wherein the modified nucleic acid is at least 98% identical to at least one of SEQ ID NO:
• 1, the strand complementary to SEQ ID NO:
• 1, or SEQ ID NO:
• 1 in the vector or cell, over a length of at least 15 consecutive nucleotides, wherein the length of the nucleic acid probe is no more than 200 nucleotides, wherein the modified nucleic acid probe is specific to a species of the human pathogenic dermatophyte belonging to the Trichophyton genus.

A modified nucleic acid amplified from a human pathogenic dermatophyte by a primer pair, comprising:

• a nucleic acid comprising at least 98% identity to the full-length of SEQ ID NO:
• 1 from the human pathogenic dermatophyte belonging to a Trichophyton genus that causes at least one of a skin, hair, and nail infection;
• wherein the nucleic acid is labeled with a label to form the modified nucleic acid, wherein the label is selected from the group consisting of a fluorescent label, a radioactive label, a colloidal gold label, and an enzymatically active label;
• and
• the primer pair comprising:
• a forward primer having a length of 14 to 30 nucleotides, being at least 90% identical to Trichophyton sequence over the full-length of the forward primer, and having a final base of the forward primer no more than 200 bp away from a first base of SEQ ID NO:
• 1 in a 5′ to 3′ orientation;
• and
• a reverse primer having a length of 14 to 30 nucleotides, being at least 90% identical to Trichophyton sequence over the full-length of the reverse primer, and having a final base of the reverse primer is no more than 200 bp away from a final base of SEQ ID NO:
• 1 in the 5′ to 3′ orientation.