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Microscopic Observation and Culture of Aspergillus species: The Traditional Way Nancy McClenny, MPA, CLS/MT Associate Program Director, CLS Internship.

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Presentation on theme: "Microscopic Observation and Culture of Aspergillus species: The Traditional Way Nancy McClenny, MPA, CLS/MT Associate Program Director, CLS Internship."— Presentation transcript:

1 Microscopic Observation and Culture of Aspergillus species: The Traditional Way Nancy McClenny, MPA, CLS/MT Associate Program Director, CLS Internship Program, San Francisco State University San Francisco, CA

2 2 Whats the Role of the Clinical Laboratory? Who is a Clinical Laboratory Scientist (CLS)? Are microscopy and culture still the methods of choice for fungi? Who will do the work?

3 3 Diagnostic Specimen for Fungus Pathology Fungal stains Immunodiagnostics Clinical Laboratory Gram stain Wet mount (KOH) Culture Serology

4 4 MICROSCOPYMICROSCOPY

5 5 What fungus is shown in this Gram stain? Hyphae septate, 45° branching Septum cmazzanti, CMVL,Stanford

6 6 Microscopy in the Clinical Lab: The Critical Tools Fluorescent dye with KOH –Blankophor –Calcofluor –Uvitex 2B Calibrated micrometer Expert readers!

7 7 How do we know its Aspergillus? Accessory conidia A. nigerA. terreus Stipe Stanford Medical CenterWiley Schell, Duke Univ.

8 8 How do we know its NOT Aspergillus? Phialide Phialoconidium Fusarium species Annelloconidia Scedosporium apiospermum Wiley Schell, Duke Univ.Stanford Medical Center

9 9 How do we know its NOT Aspergillus? Dactylaria gallopava Methenamine SilverKOH Stanford Medical Center Dematiaceous hyphae Stanford Medical Center

10 10 Stipes of A. niger, not hyphae of Mucor. Conidia of A. niger, not Candida Wiley Schell, Duke Univ. Why do we need culture?

11 11 CULTURECULTURE

12 12 How can culture for Aspergilli be improved? Sporulation media for primary isolation Incubation at both 35-37°C and 30°C. Daily inspection of cultures Microaerophilic environment (Tarrand, Kontoyiannis, and Han. Abstract M- 909, 42 nd ICAAC, 2002.) A. versicolor on Sabouraud Agar nmcclenny A. versicolor on Potato Dextrose Agar nmcclenny

13 13 Rapid Identification of A. fumigatus ±1 hr 3-7 days hours at C

14 14 Poorly sporulating A. fumigatus Unusual or Atypical Isolates nmcclenny Neosartorya fischeri with masses of cleistothecia nmcclenny

15 15 Aspergillus terreus Jharris, Texas Dept. of Health nmcclenny Phenotypic Identification of Other Aspergillus Species nmcclenny

16 16 You just get used to how they look.

17 17 Who Will Do the Work? A 53-56% reduction in CLS training programs in the past 12 years 34% of professionals in microbiology laboratories 50 years old Available at:

18 18 Case Study CLS Internship Program S. F. State University April, 2004: $10.3 million is cut. CLS Internship Program must become self-supporting. August 2004: Director accepts a job offer outside of SFSU. September, 2004: CLS program is suspended.

19 19 Where Have All the Students Gone? Class 54 CLS Interns, SFSU Class??

20 20 Summary Microscopy and culture remain common, necessary diagnostic tools. Procedural enhancements to microscopy and culture are available. Recruitment and training of laboratory scientists must be concurrent with the integration of phenotypic and genotypic methods.


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