Methods and Compositions for Identifying Yeast

Abstract

The invention relates to a method of identifying a specific yeast species in patient tissue or body fluid. The method comprises the steps of extracting and recovering DNA of the yeast species from the patient tissue or body fluid, amplifying the DNA, hybridizing a probe to the DNA to specifically identify the yeast species, and specifically identifying the yeast species. The invention also relates to a method of identifying a yeast mycotoxin in patient tissue or body fluid. The method comprises the steps of extracting and recovering the yeast mycotoxin from the patient tissue or body fluid, contacting the yeast mycotoxin with an antibody directed against the yeast mycotoxin, and identifying the yeast myocotoxin. Both of these methods can be used to determine if a patient is at risk for or has developed a disease state related to a yeast infection, and to develop an effective treatment regimen for the patient.

1 . A method of identifying a specific yeast species in patient tissue or body fluid, the method comprising the steps of:
 extracting and recovering DNA of the yeast species from the patient tissue or body fluid;   amplifying the DNA;   hybridizing a probe to the DNA to specifically identify the yeast species; and   specifically identifying the yeast species.   
     
     
         2 . The method of  claim 1  wherein the amplifying step is performed with primers that hybridize to the DNA. 
     
     
         3 . The method of  claim 1  wherein the body fluid are selected from the group consisting of urine, nasal secretions, nasal washes, bronchial lavages, bronchial washes, spinal fluid, sputum, gastric secretions, seminal fluid, other reproductive tract secretions, lymph fluid, whole blood, serum, and plasma. 
     
     
         4 . The method of  claim 1  wherein the DNA is amplified using PCR. 
     
     
         5 . The method of  claim 4  wherein the PCR is real-time PCR. 
     
     
         6 . The method of  claim 1  wherein the probe is fluorescently labeled. 
     
     
         7 . The method of  claim 5  wherein the probe is fluorescently labeled. 
     
     
         8 . The method of  claim 1  wherein the yeast species are selected from the group consisting of  Candida albicans, Candida glabrata, Candida kruseii , and  Candida tropicalis.    
     
     
         9 . The method of  claim 2  wherein the probe, a forward primer, and a reverse primer are used during the amplification step and the probe comprises the sequence of SEQ ID NO: 1, the forward primer comprises the sequence of SEQ ID NO: 2, and the reverse primer comprises the sequence of SEQ ID NO: 3. 
     
     
         10 . The method of  claim 2  wherein the probe, a forward primer, and a reverse primer are used during the amplification step and the probe comprises the sequence of SEQ ID NO: 4, the forward primer comprises the sequence of SEQ ID NO: 5, and the reverse primer comprises the sequence of SEQ ID NO: 6. 
     
     
         11 . The method of  claim 2  wherein the probe, a forward primer, and a reverse primer are used during the amplification step and the probe comprises the sequence of SEQ ID NO: 7, the forward primer comprises the sequence of SEQ ID NO: 8, and the reverse primer comprises the sequence of SEQ ID NO: 9. 
     
     
         12 . The method of  claim 2  wherein the probe, a forward primer, and a reverse primer are used during the amplification step and the probe comprises the sequence of SEQ ID NO: 10, the forward primer comprises the sequence of SEQ ID NO: 11, and the reverse primer comprises the sequence of SEQ ID NO: 12. 
     
     
         13 . The method of  claim 2  wherein the amplified sequence is internal transcribed spacer regions of nuclear ribosomal DNA. 
     
     
         14 . The method of  claim 1  wherein the probe is bound to a bead dyed with a fluorochrome. 
     
     
         15 . A method of identifying a yeast mycotoxin in patient tissue or body fluid, the method comprising the steps of:
 extracting and recovering the mycotoxin from the patient tissue or body fluid;   contacting the mycotoxin with an antibody directed against the mycotoxin; and   identifying the myocotoxin.   
     
     
         16 . The method of  claim 15  further comprising the step of quantifying the mycotoxin. 
     
     
         17 . The method of  claim 15  wherein the body fluid is selected from the group consisting of urine, nasal secretions, nasal washes, bronchial lavages, bronchial washes, spinal fluid, sputum, gastric secretions, seminal fluid, other reproductive tract secretions, lymph fluid, whole blood, serum, and plasma. 
     
     
         18 . The method of  claim 15  wherein the mycotoxin is selected from the group consisting of a gliotoxin and a patulin. 
     
     
         19 . The method of  claim 15  wherein the tissue is derived from a patient tissue biopsy and is in a 10% formalin solution or is in a paraffin block. 
     
     
         20 . The method of  claim 15  wherein the antibody is bound to a bead dyed with a fluorochrome. 
     
     
         21 . A kit comprising a purified nucleic acid with a sequence selected from the group consisting of SEQ ID NO: 1 to SEQ ID NO: 12 or with a complement of a sequence selected from the group consisting of SEQ ID NO: 1 to SEQ ID NO: 12. 
     
     
         22 . A kit comprising components for the extraction and recovery of a yeast mycotoxin from body fluid or tissue of a patient. 
     
     
         23 . The kit of  claim 22  further comprising components for identification of the mycotoxin. 
     
     
         24 . The kit of  claim 23  wherein the components for identification of the mycotoxin include beads dyed with a fluorochrome and coupled to antibodies to the mycotoxin or to the mycotoxin or to a mycotoxin antigen. 
     
     
         25 . A purified nucleic acid comprising a sequence of SEQ ID NO: 1 to SEQ ID NO: 12 or a sequence that hybridizes under highly stringent conditions to a sequence consisting of SEQ ID NO: 1 to SEQ ID NO: 12.