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. 2021 Dec 1;173(3):103915. doi: 10.1016/j.resmic.2021.103915

Table 1.

Overview of advantages and disadvantages of the most commonly used methodologies for mycological evidences.

Methodologies Advantages Disadvantages References
Proven diagnosis Fungal culture
  • -

    Detection of the fungal pathogen;

  • -

    Detection of antifungal resistance;

  • -

    Identification at species level.
  • -

    Long turn-around-time; in case of yeasts, up to five days, and moulds up to four weeks;

  • -

    Qualification to select the proper medium;

  • -

    Long-delayed targeted treatment;

  • -

    Prone to contaminations;

  • -

    Low sensitivity for candidemia and aspergillosis.

 [35,50,173]
Microscopy
  • -

    Visualization of fungal structures;

  • -

    Analysis of shape, tracking of motion, and classification of microorganisms;

  • -

    Visualization of fungal biofilm formation.
  • -

    Does not allow fungal genus or species identification;

  • -

    Similar microscopic appearance of several fungus.

 [34,35,174]
Histopathology
  • -

    Detection of tissues invasion by fungi;

  • -

    Detection of the host response or tissue necrosis.
  • -

    Similar histopathologic appearance of several fungus;

  • -

    The use of stains does not always provide an accurate identification at species level;

  • -

    Limited sensitivity.

 [34,35]
Chromogenic mediaa
  • -

    Detection in polymicrobial samples;

  • -

    Several commercially available chromogenic media;

  • -

    Detection and identification of Candida at the species level;

  • -

    Fast and cost-efficient.
  • -

    Difficult distinction between Candida non-albicans species;

  • -

    Difficult differentiation of emergent pathogenic Candida species, such as C. auris;

  • -

    Phenotypic similarities between related species may hamper the distinction at specie level.

 [59,62,63]
Fluorescence in situ hybridizationa(FISH)
  • -

    Accurate identification of Candida spp. infections;

  • -

    Time saving, comparing with conventional methods;

  • -

    High specificity and sensitivity.
  • -

    High detection limit;

  • -

    Reduced number of peptide nucleic acid (PNA) probes commercially available.

 [34,175]
Matrix-Assisted Laser Desorption Ionization-Time of Flight - Mass spectrometrya(MALDI-TOF)
  • -

    Identification of the pathogen at the genus, species, and strain levels;

  • -

    Accurate and rapid identification of Candida spp. and Aspergillus spp.;

  • -

    High concordance with conventional methods;

  • -

    Easy performance;

  • -

    Reduced cost per analysis;

  • -

    Applicability for a wide range of microorganisms;

  • -

    Differentiation of closely related species;

  • -

    Great potential for antifungal susceptibility testing.
  • -

    Prior extraction step is required;

  • -

    Incapable of performing quantification;

  • -

    High initial instrument cost;

  • -

    Limit species coverage in the fungal reference databases of available MALDI-TOF MS systems;

  • -

    Databases require continuous updates to cover rarest and emergent fungal species.

 [34,35,77,[79], [80], [81],176]
Biochemical Phenotypic Identification Systemsa
  • -

    Reduced costs;

  • -

    Allows quantitative and qualitative information;

  • -

    Accurate identification of an unknown sample;

  • -

    Several commercially available platforms.
  • -

    Laborious methods and time-consuming;

  • -

    Results are only provided after a few days;

  • -

    Low sensitivity to identify and distinguish emergent pathogenic species, such as C. auris.

[64,[66], [67], [68], [69],72,73,177]
Probable diagnosis (Serological methods) 1,3 β-D-glucan
  • -

    Detection of relevant fungal pathogens;

  • -

    Non-invasive;

  • -

    Fast results;

  • -

    High NPV is excellent for invasive fungal infections screening;
  • -

    Nonspecific panfungal test;

  • -

    Lower sensitivity and high number of false-positive results;

  • -

    Certain fungus produce less β-D-glucan (Cryptococcus spp.) or do not produce any (Blastomyces spp. and mucoraceous moulds);

  • -

    Lack of specificity for endemic mycosis diagnosis.

 [35,106,107,166,178]
Mannan antigen and antimannan antibody
  • -

    Best performance when the two biomarkers are combined;

  • -

    Non-invasive;

  • -

    Economic;

  • -

    Deliver fast results.
  • -

    Limited specificity due to normal commensalism or colonization by Candida species;

  • -

    Limited sensitivity of antibody assays for immunocompromised patients;

  • -

    Lower sensitivity for C. krusei and C. parapsilosis.

[35,166,178,179]
Galactomannan
  • -

    Good biomarker for the detection of invasive aspergillosis;

  • -

    Useful for assessing the response to antifungal therapy.
  • -

    Low sensitivity for early diagnosis.

  • -

    Cross reactivity with Fusarium spp. and Histoplasma spp.;

  • -

    Lower sensitivity for Aspergillus fumigatus.

[35,[124], [125], [126],178,179]
Antibody-based (Immunofluorescence, ELISA, Lateral flow assay, Latex agglutination assay)
  • -

    Higher accuracy than the standard serologic markers mentioned above;

  • -

    Low cost;

  • -

    Easy and fast performance (can be applied to POC).
  • -

    Reduced sensitivity for immunocompromised patients;

  • -

    Limited specificity;

  • -

    Careful analysis regarding patients with no clinical signs;

  • -

    Antigen-antibody methods still not available for some fungal pathogens (mucormycosis, fusariosis, and scedosporiosis).

 [35,127,130,131,133,174]
Probable diagnosis Invasive Fungal Infections Imaging
  • -

    Several imaging options available;

  • -

    Overall information at few time;

  • -

    Monitor the progression of the infection, as well as patient's response to antifungal therapy.
  • -

    Need for a multimodal imaging method;

  • -

    The use of each imaging method is limited and specific.

 [93,95,96]
PCR-based methods
  • -

    Short turn-around time;

  • -

    High sensitivity and specificity;

  • -

    qPCR allows quantification of amplified DNA in real-time;

  • -

    Allows species identification and intraspecies differentiation;

  • -

    Panfungal PCRs can exclude or generally diagnose, invasive fungal infections;

  • -

    New PCR-platforms are capable of including steps from samples preparation, DNA extraction and results, in an entire closed system, free of contamination.
  • -

    Traditional PCR does not allow quantification of the amplified DNA;

  • -

    Lack of standardization of the fungal DNA isolation techniques;

  • -

    Contaminations;

  • -

    Careful selection of primers and optimization of the reaction conditions;

  • -

    Panfungal PCRs does not allow a targeted therapeutic plan;

  • -

    PCR-based methods can be difficultly applied to some samples, especially non-sterile samples, such as BAL.

 [34,35,82,136,143,149]
a

Methodologies to achieve specific identification after the isolation of pathogenic fungi.