fungi

• fungi are eukaryotic, heterotrophic organisms that obtain nutrients by absorbing them from their environment
• the cell wall is made up of chitin (similar to the exoskeleton of insects)
• Fungi are non-photosynthetic aerobic eukaryotic organisms that absorb nutrients from their environment
• Fungi are definitely not plants
• Fungi could be multicellular, unicellular, or dimorphic
• There are over 100,000 species of fungi, with 500 associated with human disease and around 100 capable of causing infection in normal individuals
• Fungal cells have a rigid cell wall composed of chitin, mannans, glucans, and sometimes cellulose
• Fungi are heterotrophic, lack chlorophyll, and obtain nutrition by secreting enzymes for external digestion
• Fungi are simpler in structure than plants or animals, with the basic structural unit being a chain of tubular, filament-like cells (hypha) or an independent single cell (yeast)
• Fungi reproduce by means of microscopic propagules called conidia or spores, with asexual and sexual stages
• Spores are very resistant to heat, cold, acids, bases, and other chemicals, and are potent allergens
• Fungi are classified into Kingdom Fungi with 7 phyla, where only Glomeromycota, Ascomycota, and Basidiomycota are pathogenic to humans
• Glomeromycota are usually aseptate with branching hyphae, producing asexual spores inside a sporangium
• Ascomycota are mostly septate filamentous, with asexual and sexual reproduction methods
• Basidiomycota are mostly septate filamentous, with some yeasts, and reproduce asexually and sexually
• Other anamorphic fungi include Hyphomycetes with genera like Aspergillus, Cladophialophora, Fusarium, Penicillium, and Trichophyton
• Fungi can be classified based on ultrastructural cellular form into yeasts (unicellular), molds (multicellular), and dimorphics (both unicellular and multicellular)
• Fungal infections can be classified based on the area of the body affected into superficial, cutaneous, subcutaneous, systemic, and opportunistic mycosis
• Presence of fungi in clinical samples does not necessarily confirm true infection, and severity depends on the immunologic status of the host
• Most pathogenic fungi do not produce toxins, show physiologic modifications during parasitic infection, are thermotolerant, and can resist host defenses
• Clinical syndromes of fungal infections include mycotoxicosis, hypersensitivity diseases, and colonization with eventual disease
• Mycotoxicosis is secondary to ingestion of fungal toxins, causing tissue inflammation, necrosis, gangrene, liver damage, and carcinogenesis
• Hypersensitivity diseases are usually due to fungal spores in the air, triggering asthmatic attacks, rhinitis, pneumonitis, and alveolitis
• Laboratory procedures for diagnosing fungal infections involve a combination of clinical observation and investigation, with different approaches for diagnosis including microscopic detection, culture isolation, and serologic response detection
• Quality specimens, good storage, and relevant information are crucial for mycotic laboratory investigations
• New diagnostic procedures based on the detection of fungal DNA in clinical material are currently being developed
• Microscopic examination of skin specimens is part of the direct microscopic examination of clinical specimens for fungal diagnosis
• Direct Microscopic Examination of Clinical Specimens:

  • Microscopic examination can reveal a fungal organism in minutes
  • Guides treatment decisions
  • Guides whether an organism recovered later in culture is a contaminant or a pathogen
  • Helps in selecting the most appropriate culture conditions to recover organisms visualized on direct smear
  • Drawbacks include:
  • Quality of the specimen
  • Age of the specimen
  • Extent of the disease process
  • Nature of the tissue being examined
  • Experience of the microscopist
• Direct Microscopic Examination of Clinical Specimens:

  • Keratinised dermatologic specimens, sputum, and minced tissue samples can be examined after treatment with 10–20% Potassium hydroxide (KOH)
  • Wet preparation without stain or with stain like lactophenol cotton blue, methylene blue, or calcofluor white
  • India ink can reveal encapsulated Cryptococcus neoformans cells in CSF
  • Gram stain, Giemsa stain, and Wright’s stain can be used to detect Histoplasma capsulatum
  • Papanicolaou stain
• Microscopic Examination of Clinical Specimens:

  • Histopathologic examination is reliable for establishing the diagnosis of subcutaneous and systemic fungal infections
  • Stains like Hematoxylin and eosin, Methenamine-silver, Periodic acid-Schiff (PAS), and Mucicarmine can be used for staining
• Microscopic Examination of Clinical Specimens:

  • Drawback: seldom permits the precise fungal genus involved to be identified
  • Immunohistochemical methods with Monoclonal antibodies have more specificity, useful for Aspergillus species or members of the order Mucorales
• Culture:

  • Isolation in culture permits identification of most pathogenic fungi
  • Most pathogenic fungi are not fastidious and can grow on any media
  • Culture media include Sabouraud’s dextrose agar, Potato dextrose agar, Potato flakes, Brain heart infusion for Histoplasma capsulatum
  • Additives like chloramphenicol, gentamicin, and cycloheximide are used to suppress bacteria
• Culture:

  • Chromogenic agars can be used for fungal identification
  • Optimum growth temperature for most pathogenic fungi is around 30°C
  • Some fungi are obligate pathogens like H. capsulatum or Trichophyton rubrum, while some are opportunistic like A. fumigatus or C. albicans
• Culture:

  • No isolate should be dismissed as a contaminant without careful consideration
  • Blood culture can help recover Candida species more readily than dimorphic fungi and moulds
  • Different methods like lysis-centrifugation and semi-automated agitation blood culture systems are used
• Fungal Identification:

  • Phenotypic identification includes macroscopic and microscopic morphology
  • Spores can be induced for identification
  • Biochemical tests like assimilation and fermentation of sugars are used for identification
  • Commercial identification systems and rapid tests like germ tube test and urease test are employed
• Fungal Identification:

  • Skin testing is discouraged as it may interfere with serological studies
  • Serologic testing assists in diagnosing fungal infections and determining efficacy of therapy
  • Different tests like immunodiffusion, complement fixation, latex agglutination, and ELISA are used
• Serology:

  • Antibodies testing is useful in diagnosing endemic fungal infections like histoplasmosis and coccidioidomycosis
  • Antigen detection methods are used for cryptococcosis, histoplasmosis, and aspergillosis
• Molecular diagnosis:

  • DNA sequences are used for determining genetic relatedness among fungi
  • Polymerase chain reaction (PCR) and other methods are used for rapid detection and identification of fungi
• Molecular diagnosis:

  • Molecular sub-typing methods like MLST and whole-genome sequencing are employed
  • Antifungal Drug Susceptibility Testing:
  • Minimum inhibitory concentration (MIC) is used to determine in-vitro susceptibility pattern
  • Methods like micro and macro-dilution are used
  • Drugs like Amphotericin B and voriconazole are tested for Candida and cryptococcus
• yeast cells reproduce through budding