1.  Candidemia: species involved, virulence factors and antimycotic susceptibility Concetta De Luca, Maria Guglielminetti, Antonella Ferrario, Maria Calabrò, Erminia Casari NEW MICROBIOLOGICA, 35, 459-468, 2012 生科三乙 黃建凱 指導老師 : 藍清隆

2.  Introduction

3. Candida species  In the immunocompetent host, Candida albicans is a benign member of the human microbiota, and colonizes the human gastrointestinal, respiratory and reproductive tracts. Candida albicans

4. Candidemia  Candidemia is one of Bloodstream Infection(BSI), which are caused by Candida.  Candidemia are common causes of the nosocomial infections with the highest mortality.  Proteinase and phospholipase production can lead to dysfunction or even rupture of cell membranes, which facilitate adhesion of the microorganism to the host.

5.  Recent reports indicate a trend towards an increasing prevalence of infections caused by species of Candida other than C. albicans.  Moreover, less common species may emerge as important opportunistic pathogens in the future.

6. Aim  To investigate the characteristics of the Candida species involved in BSI episodes in our Institute to define the optimal therapeutic approach.  We therefore studied the species involved, and determined the phopholipase and protease activity of Candida species isolated from blood infection and the susceptibility pattern towards the main antifungal agents currently available.

7.  Material & Methods

8. CHROMagar TM Candida  Composition in g / L: Agar 15.0, peptone 10.2; Mix chromogenic 22.0, Chloramphenicol 0.5, pH: 6.1 + / - 0.2  C. albicans - green  C. tropicalis – dark blue  C. krusei – light red  Other species – white to violet

9. API 20C AUX  API ® Gram negative Identification  API 20E – 18-24 hour identification of Enterobacteriacae and other non-fastidious gram negative bacteria  API Rapid 20E – 4-hour identification of Enterobacteriaceae  API 20NE – 24 to 48-hour identification of Gram negative non-Enterobacteriaceae  API NH – 2-hour identification of Neisseria Haemophilus andBranhamella catarrhalis   API ® Gram positive Identification  API Staph – Overnight identification of clinical staphylococci andmicrococci  RAPIDEC ® Staph – 2-hour identification of the commonly occurring staphylococci  API 20 Strep – 4 or 24-hour identification of streptococci and enterococci  API Coryne – 24-hour identification of Corynebacteria and coryne-like organisms   API ® Anaerobe Identification  API 20A ® – 24-hour identification of anaerobes  Rapid ID 32A – 4-hour identification of anaerobes   API ® Yeast Identification  API 20C AUX – 48 to 72-hour identification of yeasts   Others  API ® 50 CH – Performance of carbohydrate metabolism tests  API ZYM ® – Semiquantitation of enzymatic activities

10. Sensititre YeastOne panel  All isolates were tested for antifungal susceptibility.  It contains Anidulafungin (AND), Micafungin (MF), Caspofungin (CAS), 5-flucytosine (FLU), Voriconazole (VOR), Itraconazole (IT), Fluconazole (FLU) and Amphotericin B (AB).

11. 抗菌劑  Fluconazole 抑菌劑  Anidulafungin  Caspofungin  Amphotericin B 殺菌劑

12. Gibco® RPMI Media 1640  The isolates were tested in accordance with the manufacturer’s instructions, using an inoculum at the concentration of 1x10 6 to 5x10 6 cells per ml in RPMI 1640 broth.

13. Patients’ blood culture 37 o C, 5 days Identity MIC CHROMagar & API 20C AUX Culture in RPMI Media 1640 In twofold serial 12 dilutions Sensititre YeastOne panel Incubated at 37 o C, 24 h

14. MIC  Minimal Inhibitory Concentration(MIC) is determined as the lowest concentration of antifungal agent preventing development of colonies.

15. Test for Proteinase & Phospholipase activity YEPD(yeast extract, peptone and glucose) 10  l BSAagar, pH=5.0 30 o C, 5 days sterile saline 5  l Agar contain egg yolk, pH=4.3 37 o C, 5 days

16.  Results

17. Distribution of Candida species FIGURE 1

18. Distribution of Candida species according to hospital Unit FIGURE 2 48% 36% 4% 12% 50% 55% 27% 70% 20%

19. Protease and phospholipase activity of Candida strains FIGURE 3

20. No significant relationship between production of these enzymes and antifungal susceptibility

21. Fluconazole MIC distribution In C. albicans In C. glabrata

22. Anidulafungin MIC distribution In C. albicans In C. glabrata

23. Caspofungin MIC distribution In C. albicans In C. glabrata

24.  Discussion

25. Geographical shift  An equivalent distribution of isolates between C. albicans species (48%) and C. non- albicans species (52%) in our Institute  C. albicans are north drift, C. non- albicans species are south drift in other literature  Most common speicies of the northern C. non- albicans is C. parapsilosis  Most common C. non- albicans strains is C. glabrata in our Institute

26. Empirical antibiotic therapy  比方某個病人發燒、胸片看到浸潤，在肺炎的「疑似診斷」 之下，抗生素的選擇就要看臨床醫師的「經驗」囉！  目前是有所謂的「治療指引」；有的把病人是否「典型」 來分，「典型」的肺炎通常症狀明顯而突發，白血球超標； 「非典型」肺炎則症狀較緩、較輕，白血球不是那麼高。  也有「治療指引」把病人分成「社區型」、「安養機構型」 和「院內感染型」，通常後者代表的可能是多重抗藥菌 種。  在確診之前（事實上很難確診，約有一半的肺炎是查不出 確切致病菌的），抗生素的選擇就是「憑經驗」的，寫作 empirical antibiotics ！