Nucleic acid probes for the detection and identification of fungi

Claim 1

A method of determining whether one or more fungal species selected from the group of fungal species consisting of Acremonium species, Aspergillus clavatus, Aspergillus flavus, Aspergillus fumigatus, Aspergillus glaucus, Aspergillus nidulans, Aspergillus niger, Aspergillus ochraceus, Aspergillus terreus, Aspergillus unguis, Aspergillus ustus, Beauveria species, Bipolaris species, Blastochizomyces species, Blastomyces dermatitidis, Chrysoporium species, Cladosporium species, Curvularia species, Candida albicans, Candida glabrata, Candida guilliermondii, Coccidioides immitis, Candida kefyr, Candida brusei, Cryptoccus laurentii, Candida lusitaniae, Cryptococcus neoformans var gattii, Cryptococcus neoformans, Candida parapsilosis, Cryptococcus terreus, Candida tropicalis, Fusarium species, Filobasidium capsuligenum, Filobasidiella neoformans var bacillispora, Filobasidiella neoformans var neoformans, Filobasidium uniguttulam, Geotrichum species, Histoplasma capsulatum, Malbranchea species, Mucor species, Paecilomyces species, Penicillium species, Pseudallescheria boydii, Rhizopus species (NO: 1), Rhizopus species (NO: 2), Rhizopus species (NO: 3), Sporothrix species, Scopulariopsis brevicaulis, Scopulariopsis brumptii, Saccharomyces cerevisiae and Trichosporon beigelii is present in a sample of fungi, said method comprising the following steps: a) extracting nucleic acid material from fungi contained in said sample; b) adding two known oligonucleotide primers, one of said primers being (SEQ ID NO: 1) or (SEQ ID NO: 2), said primers bracketing a hypervariable region on the 28S rDNA or rRNA present in the fungal species of said group; c) amplifying the sequence between said primers; and d) using one or more labeled probes directed to a portion of the hypervariable region bracketed by said primers, each said labeled probe being specific for one of said fungal species from said group, wherein each said one or more labeled probes is fully complementary to a species-unique nucleotide sequence in said hypervariable region, to determine whether said fungal species identified by each said labeled probe is present in said sample, wherein, furthermore, said one or more labeled probes each has a nucleotide residue sequence consisting of from 10 to 50 consecutive nucleotide residues from a sequence selected from the group consisting of (SEQ ID NO: 24) through (SEQ ID NO: 74) and the complements thereof.

Claim 2

A method of determining whether one or more fungal species selected from the group of fungal species consisting of Acremonium species, Aspergillus clavatus, Aspergillus flavus, Aspergillus fumigatus, Aspergillus glaucus, Aspergillus nidulans, Aspergillus niger, Aspergillus ochraceus, Aspergillus terreus, Aspergillus unguis, Aspergillus ustus, Beauveria species, Bipolaris species, Blastochizomyces species, Blastomyces dermatitidis, Chrysoporium species, Cladosporium species, Curvularia species, Candida albicans, Candida glabrata, Candida guilliermondii, Coccidioides immitis, Candida kefyr, Candida brusei, Cryptoccus laurentii, Candida lusitaniae, Cryptococcus neoformans var gattii, Cryptococcus neoformans, Candida parapsilosis, Cryptococcus terreus, Candida tropicalis, Fusarium species, Filobasidium capsuligenum, Filobasidiella neoformans var bacillispora, Filobasidiella neoformans var neoformans, Filobasidium uniguttulam, Geotrichum species, Histoplasma capsulatum, Malbranchea species, Mucor species, Paecilomyces species, Penicillium species, Pseudallescheria boydii, Rhizopus species (NO: 1), Rhizopus species (NO: 2), Rhizopus species (NO: 3), Sporothrix species, Scopulariopsis brevicaulis, Scopulariopsis brumptii, Saccharomyces cerevisiae and Trichosporon beigelii is present in a sample of fungi, said method comprising the following steps: a) extracting nucleic acid material from fungi contained in said sample; b) adding two known oligonucleotide primers, one of said primers being (SEQ ID NO: 1) or (SEQ ID NO: 2), said primers bracketing a hypervariable region on the 28S rDNA or rRNA present in the fungal species of said group; c) amplifying the sequence between said primers; and d) using one or more labeled probes directed to a portion of the hypervariable region bracketed by said primers, each said labeled probe being specific for one of said fungal species from said group, wherein each said one or more labeled probes is fully complementary to a species-unique nucleotide sequence in said hypervariable region, to determine whether said fungal species identified by each said labeled probe is present in said sample, wherein, furthermore, said one or more labeled probes each has a nucleotide residue sequence consisting of from 10 to 50 consecutive nucleotide residues from a sequence selected from the group consisting of (SEQ ID NO: 24) through (SEQ ID NO: 74) and the complements thereof.

Claim 19

A method of determining whether one or more fungal species selected from the group of fungal species consisting of Acremonium species, Aspergillus clavatus, Aspergillis flavus, Aspergillus glaucus, Aspergillus nidulans, Aspergillus niger, Aspergillus ochraceus, Aspergillus terreus, Aspergillus unguis, Aspergillus ustus, Beauveria species, Bipolaris species, Blastochizomyces species, Blastomyces dermatitidis, Chrysoporium species, Cladosporium species, Curvularia species, Candida albicans, Candida glabrata, Candida guilliermondii, Coccidioides immitis, Candida kefyr, Candida brusei, Cryptoccus laurentii, Candida lusitaniae, Cryptococcus neoformans var gattii, Cryptococcus neoformans, Candida parapsilosis, Cryptococcus terreus, Candida tropicalis, Fusarium species, Filobasidium capsuligenum, Filobasidiella neoformans var bacillispora, Filobasidiella neoformans var neoformans, Filobasidium uniguttulam, Geotrichum species, Histoplasma capsulatum, Malbranchea species, Mucor species, Paecilomyces species, Penicillium species, Pseudallescheria boydii, Rhizopus species (NO: 1), Rhizopus species (NO: 2), Rhizopus species (NO: 3), Sporothrix species, Scopulariopsis brevicaulis, Scopulariopsis brumptii, Saccharomyces cerevisiae and Trichosporon beigelii is present in a sample of fungi, said method comprising the following steps: a) extracting the nucleic acid material from the fungi contained in said sample; b) adding a universal fungal probe selected from the group consisting of (SEQ ID NO: 1), (SEQ ID NO: 2) and the complements thereof; c) using one or more second probes, each said second probe being specific for one of said fungal species from said group, wherein each said one or more second probes is fully complementary to a species-unique nucleotide sequence in said hypervariable region and wherein, further, said one or more second probes each has a nucleotide residue sequence consisting of from 10 to 50 consecutive nucleotide residues from a sequence selected from the group consisting of (SEQ ID NO: 24) through (SEQ ID NO: 74) and the complements thereof; and d) determining whether said fungal species identified by each said second probe is present in said sample, wherein at least one of said probes is connected to a signal moiety and at least one of said probes is connected to a moiety that allows separation of said probes.

Claim 20

A method of determining whether one or more fungal species selected from the group of fungal species consisting of Acremonium species, Aspergillus clavatus, Aspergillis flavus, Aspergillus glaucus, Aspergillus nidulans, Aspergillus niger, Aspergillus ochraceus, Aspergillus terreus, Aspergillus unguis, Aspergillus ustus, Beauveria species, Bipolaris species, Blastochizomyces species, Blastomyces dermatitidis, Chrysoporium species, Cladosporium species, Curvularia species, Candida albicans, Candida glabrata, Candida guilliermondii, Coccidioides immitis, Candida kefyr, Candida brusei, Cryptoccus laurentii, Candida lusitaniae, Cryptococcus neoformans var gattii, Cryptococcus neoformans, Candida parapsilosis, Cryptococcus terreus, Candida tropicalis, Fusarium species, Filobasidium capsuligenum, Filobasidiella neoformans var bacillispora, Filobasidiella neoformans var neoformans, Filobasidium uniguttulam, Geotrichum species, Histoplasma capsulatum, Malbranchea species, Mucor species, Paecilomyces species, Penicillium species, Pseudallescheria boydii, Rhizopus species (NO: 1), Rhizopus species (NO: 2), Rhizopus species (NO: 3), Sporothrix species, Scopulariopsis brevicaulis, Scopulariopsis brumptii, Saccharomyces cerevisiae and Trichosporon beigelii is present in a sample of fungi, said method comprising the following steps: a) extracting the nucleic acid material from the fungi contained in said sample; b) adding a universal fungal probe selected from the group consisting of (SEQ ID NO: 1), (SEQ ID NO: 2) and the complements thereof; c) using one or more second probes, each said second probe being specific for one of said fungal species from said group, wherein each said one or more second probes is fully complementary to a species-unique nucleotide sequence in said hypervariable region and wherein, further, said one or more second probes each has a nucleotide residue sequence consisting of from 10 to 50 consecutive nucleotide residues from a sequence selected from the group consisting of (SEQ ID NO: 24) through (SEQ ID NO: 74) and the complements thereof; and d) determining whether said fungal species identified by each said second probe is present in said sample, wherein at least one of said probes is connected to a signal moiety and at least one of said probes is connected to a moiety that allows separation of said probes.