Diagnostics and Antifungals Flashcards
How do you prove mycoses
- microscopy shows fungal elements associated with tissue damage or the fungus itself
- culture shows fungus in normally sterile site
How do you find probable mycoses
- has host factor of immunodeficiency
- clinical evidence such as CT
- mycological evidence such as culture or fungal antigen detection
What do you look for with microscopy?
- stain for chitin
- stain candidiaphores
- reverse stain capsule
- Gram stain - yeast usually G+
- histological staining to see example spherule
What are the disadvantages of microscopy?
- not definitive species ID
- requires tissue or serum or invasive procedures
- not very sesitive
- need large amount of fungi
Whats the advantages of cultures?
- CHROMagar to ID candida species
- show thermal dimorphism
Whats the disadvantages to cultures?
- clinical relevance uncertain (false positives, esp. candida)
- requires sample with viable organism
- long time to grow
How can you use immunological memory to detect mycoses?
- if had previous infection then you have reactive T cells
- T cells produce IFNgamma and IL8 which cause inflammation and a big red spot
How can you use fungal antibodies to test for mycoses?
- separate antigens on gel
- add patient serum
- if light up = infection
ELISA (Enzyme-Linked ImmunoSorbent Assay)
- bind antigen to bottom of tube
- add patient serum
- wash
- add florescent antibody to constant region of patient antibody
- wash
- if light up = infection
Whats the disadvantage of ELISA
antibodies are also evidence of past infections
ELISA/EIA (Enzyme-linked Immuno Assay)
- binds antibody to bottom of tube
- add patient serum
- wash
- add florescent antibody that binds antigen
- wash
- if light up = antigen in patient serum = active infection
What are the advantages to EIA
- detects active infection
- indicative of levels of fungemia
What are the disadvantages to EIA
- low sensitivity (low antigen levels until dissemination)
- preblems with specificity (esp. Histo/Blasto/Cocc because cross reactivity)
How do you detect beta glucan (horseshoe crab version)
- horseshoe crab sacrifice
- their Factor G is activated by beta glucan
- this activates clotting enzymes OR you can use a color changing molecule
How do you detect beta glucan (catheter version)
- beta glucan is shed from biofilm
- can detect beta glucans peripherally from catheter biofilm
How do you detect mycoses with DNA?
- look at ITS1 and ITS2 polymorphisms
- can analyze ITS sizes and also look at point mutations
What are the advatages to using fungal DNA to detect?
- highly specific
- only have fungal DNA if active infection
What are the advatages to using fungal DNA to detect?
- highly specific
- only have fungal DNA if active infection
What are the disadvantages to fungal DNA to detect?
- requires molecular laboratory
- clinical relevance uncertain (normal flora candida for example)
allylamines
inhibit sterol biosynthesis via squalene to lanosterol
azoles (imidazoles and triazoles)
inhibit sterol biosynthesis via lanosterol and 14-demethyllanosterol
polyenes
inhibit ergosterols themselves
How do polyenes kill?
act like AMPs to create pores
How do polyenes kill?
act like AMPs to create pores
Whats the issue with polyenes?
cross reactivity with mammalian cholesterol
Are azoles and allylamines fungicidal or fungistatic and why?
fungistatic becuase inhibit ergosterol production so can’t grow
nikkomycin
inhibits chintin synthase
echinocandins
inhibit beta glucan synthase
Why is it important to know species before treating?
Sensitivity and resistance to antifungals differ amongst species
How does antifungal resistance occur?
- eeffluc pump activation (common)
- loss of target susceptibility via mutations (less common)
