Study On Rapid Methods For Detection Of Deep-seated Pathogenic Fungi By PCR And Their Clinical Application

Because of the improvement of the medical sciences and the development of new treatments for hematologic malignancies and organ transplants, the number of immunocompromised patients was steadily increased in recent 20 years. Immunosuppression is occasionally accompanied with an insidious onset of fungal infections, with some cases being nosocomial. Invasive mycoses have become a major cause of infectious morbidity and mortality in patients receiving immunosuppressive chemotherapy for cancer or organ transplantation or in immunocompromised patients, such as individuals with AIDS, cancer, drug abusers, and so on. More than 150 species of fungi are known to be responsible for infections in human. All of tissues and organs can be involved. Since opportunistic mycoses are often grave, the early, rapid, and accurate identification of the pathogenic fungus is critical for timely and appropriate management. Fungi are traditionally identified by morphology and metabolic characteristics and may require days to weeks to be isolated on cultures. Indentification may sometime be difficult, and these would impede the selection of antimicrobial agents in cases in which species identification of the organism is necessary. Therefore, it is desirable to have a rapid and accurate method of identification. In this article, we developed a universal fungal primer for panfungal PCR and the multiplex primers for Candida albicans, Aspergillus fumigatus and Cryptococcus neoformans for the triplex PCR. The results of detection to 60 clinical specimens and successful construction of invasive pulmonary aspergillosis rabbit models showed that the PCR methods was faster, simpler than the conventional morphological identification.Chapter I Developing universal fungal primer PCR for detection of familiar deep-seated pathogenic fungi. First, A 260bp PCR product was successfully amplified from all 78 strains of 55 fungal species in 9 fungal genus studied with universal fungal primer PCR to testify the specificity of the PCR. All of fungi yielded a 260-bp amplified fragment in panfungal PCR, while 5 strains of bacteria and human whole blood were negtive. Second, serial diluted fungal DNA of C. albicans was apmlified with panfungal PCR to detect the sensitivity of the PCR. The sensitivity of PCR for fungi was 10~2...