Diagnosis and management of Aspergillus diseases: executive summary of the 2017 ESCMID-ECMM-ERS guideline

  • A.J. Ullmann
    Affiliations
    Department of Infectious Diseases, Haematology and Oncology, University Hospital Würzburg, Würzburg, Germany

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • J.M. Aguado
    Affiliations
    Infectious Diseases Unit, University Hospital Madrid, Madrid, Spain

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • S. Arikan-Akdagli
    Affiliations
    Department of Medical Microbiology, Hacettepe University Medical School, Ankara, Turkey

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • D.W. Denning
    Affiliations
    The National Aspergillosis Centre, Wythenshawe Hospital, Mycology Reference Centre Manchester, Manchester University NHS Foundation Trust, ECMM Excellence Centre of Medical Mycology, Manchester, UK

    The University of Manchester, Manchester, UK

    Manchester Academic Health Science Centre, Manchester, UK

    European Confederation of Medical Mycology (ECMM)
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  • A.H. Groll
    Affiliations
    Department of Paediatric Haematology/Oncology, Centre for Bone Marrow Transplantation, University Children's Hospital Münster, Münster, Germany

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • K. Lagrou
    Affiliations
    Department of Microbiology and Immunology, ECMM Excellence Centre of Medical Mycology, University Hospital Leuven, Leuven, Belgium

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • C. Lass-Flörl
    Affiliations
    Institute of Hygiene, Microbiology and Social Medicine, ECMM Excellence Centre of Medical Mycology, Medical University Innsbruck, Innsbruck, Austria

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • R.E. Lewis
    Affiliations
    Infectious Diseases Clinic, Sant’Orsola-Malpighi Hospital, University of Bologna, Bologna, Italy

    ESCMID Fungal Infection Study Group (EFISG)
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  • P. Munoz
    Affiliations
    Department of Medical Microbiology and Infectious Diseases, Hospital General Universitario Gregorio Marañón, Madrid, Spain

    CIBER Enfermedades Respiratorias - CIBERES (CB06/06/0058), Madrid, Spain

    Medicine Department, School of Medicine, Universidad Complutense de Madrid, Madrid, Spain

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • P.E. Verweij
    Affiliations
    Department of Medical Microbiology, Radboud University Medical Centre, Centre of Expertise in Mycology Radboudumc/CWZ, ECMM Excellence Centre of Medical Mycology, Nijmegen, Netherlands

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • A. Warris
    Affiliations
    MRC Centre for Medical Mycology, Institute of Medical Sciences, University of Aberdeen, Aberdeen, UK

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • F. Ader
    Affiliations
    Department of Infectious Diseases, Hospices Civils de Lyon, Lyon, France

    Inserm 1111, French International Centre for Infectious Diseases Research (CIRI), Université Claude Bernard Lyon 1, Lyon, France

    European Respiratory Society (ERS)
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  • M. Akova
    Affiliations
    Department of Medicine, Section of Infectious Diseases, Hacettepe University Medical School, Ankara, Turkey

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • M.C. Arendrup
    Affiliations
    Department Microbiological Surveillance and Research, Statens Serum Institute, Copenhagen, Denmark

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • R.A. Barnes
    Affiliations
    Department of Medical Microbiology and Infectious Diseases, Institute of Infection and Immunity, School of Medicine, Cardiff University, Cardiff, UK

    European Confederation of Medical Mycology (ECMM)
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  • C. Beigelman-Aubry
    Affiliations
    Department of Diagnostic and Interventional Radiology, Centre Hospitalier Universitaire Vaudois (CHUV), Lausanne, Switzerland

    European Respiratory Society (ERS)
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  • S. Blot
    Affiliations
    Department of Internal Medicine, Ghent University, Ghent, Belgium

    Burns, Trauma and Critical Care Research Centre, University of Queensland, Brisbane, Australia

    European Respiratory Society (ERS)
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  • E. Bouza
    Affiliations
    Department of Medical Microbiology and Infectious Diseases, Hospital General Universitario Gregorio Marañón, Madrid, Spain

    CIBER Enfermedades Respiratorias - CIBERES (CB06/06/0058), Madrid, Spain

    Medicine Department, School of Medicine, Universidad Complutense de Madrid, Madrid, Spain

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • R.J.M. Brüggemann
    Affiliations
    Radboud Centre for Infectious Diseases, Radboud University Medical Centre, Centre of Expertise in Mycology Radboudumc/CWZ, ECMM Excellence Centre of Medical Mycology, Nijmegen, Netherlands

    ESCMID Fungal Infection Study Group (EFISG)
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  • D. Buchheidt
    Affiliations
    Medical Clinic III, University Hospital Mannheim, Mannheim, Germany

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • J. Cadranel
    Affiliations
    Department of Pneumology, University Hospital of Tenon and Sorbonne, University of Paris, Paris, France

    European Respiratory Society (ERS)
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  • E. Castagnola
    Affiliations
    Infectious Diseases Unit, Istituto Giannina Gaslini Children's Hospital, Genoa, Italy

    ESCMID Fungal Infection Study Group (EFISG)
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  • A. Chakrabarti
    Affiliations
    Department of Medical Microbiology, Postgraduate Institute of Medical Education & Research, Chandigarh, India

    European Confederation of Medical Mycology (ECMM)
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  • M. Cuenca-Estrella
    Affiliations
    Instituto de Salud Carlos III, Madrid, Spain

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • G. Dimopoulos
    Affiliations
    Department of Critical Care Medicine, Attikon University Hospital, National and Kapodistrian University of Athens, Medical School, Athens, Greece

    European Respiratory Society (ERS)
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  • J. Fortun
    Affiliations
    Infectious Diseases Service, Ramón y Cajal Hospital, Madrid, Spain

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • J.-P. Gangneux
    Affiliations
    Univ Rennes, CHU Rennes, Inserm, Irset (Institut de Recherche en santé, environnement et travail) – UMR_S 1085, Rennes, France

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • J. Garbino
    Affiliations
    Division of Infectious Diseases, University Hospital of Geneva, Geneva, Switzerland

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • W.J. Heinz
    Affiliations
    Department of Infectious Diseases, Haematology and Oncology, University Hospital Würzburg, Würzburg, Germany

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • R. Herbrecht
    Affiliations
    Department of Haematology and Oncology, University Hospital of Strasbourg, Strasbourg, France

    ESCMID Fungal Infection Study Group (EFISG)
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  • C.P. Heussel
    Affiliations
    Diagnostic and Interventional Radiology, Thoracic Clinic, University Hospital Heidelberg, Heidelberg, Germany

    European Confederation of Medical Mycology (ECMM)
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  • C.C. Kibbler
    Affiliations
    Centre for Medical Microbiology, University College London, London, UK

    European Confederation of Medical Mycology (ECMM)
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  • N. Klimko
    Affiliations
    Department of Clinical Mycology, Allergy and Immunology, North Western State Medical University, St Petersburg, Russia

    European Confederation of Medical Mycology (ECMM)
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  • B.J. Kullberg
    Affiliations
    Radboud Centre for Infectious Diseases, Radboud University Medical Centre, Centre of Expertise in Mycology Radboudumc/CWZ, ECMM Excellence Centre of Medical Mycology, Nijmegen, Netherlands

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • C. Lange
    Affiliations
    International Health and Infectious Diseases, University of Lübeck, Lübeck, Germany

    Clinical Infectious Diseases, Research Centre Borstel, Leibniz Center for Medicine & Biosciences, Borstel, Germany

    German Centre for Infection Research (DZIF), Tuberculosis Unit, Hamburg-Lübeck-Borstel-Riems Site, Lübeck, Germany

    European Respiratory Society (ERS)
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  • T. Lehrnbecher
    Affiliations
    Division of Paediatric Haematology and Oncology, Hospital for Children and Adolescents, Johann Wolfgang Goethe-University, Frankfurt, Germany

    European Confederation of Medical Mycology (ECMM)
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  • J. Löffler
    Affiliations
    Department of Infectious Diseases, Haematology and Oncology, University Hospital Würzburg, Würzburg, Germany

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • O. Lortholary
    Affiliations
    Department of Infectious and Tropical Diseases, Children's Hospital, University of Paris, Paris, France

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • J. Maertens
    Affiliations
    Department of Haematology, ECMM Excellence Centre of Medical Mycology, University Hospital Leuven, Leuven, Belgium

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • O. Marchetti
    Affiliations
    Infectious Diseases Service, Department of Medicine, Lausanne University Hospital, Lausanne, Switzerland

    Department of Medicine, Ensemble Hospitalier de la Côte, Morges, Switzerland

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • J.F. Meis
    Affiliations
    Department of Medical Microbiology and Infectious Diseases, Canisius-Wilhelmina Hospital, Centre of Expertise in Mycology Radboudumc/CWZ, ECMM Excellence Centre of Medical Mycology, Nijmegen, Netherlands

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • L. Pagano
    Affiliations
    Department of Haematology, Universita Cattolica del Sacro Cuore, Roma, Italy

    European Confederation of Medical Mycology (ECMM)
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  • P. Ribaud
    Affiliations
    Quality Unit, Pôle Prébloc, Saint-Louis and Lariboisière Hospital Group, Assistance Publique-Hôpitaux de Paris, Paris, France
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  • M. Richardson
    Affiliations
    The National Aspergillosis Centre, Wythenshawe Hospital, Mycology Reference Centre Manchester, Manchester University NHS Foundation Trust, ECMM Excellence Centre of Medical Mycology, Manchester, UK

    The University of Manchester, Manchester, UK

    Manchester Academic Health Science Centre, Manchester, UK

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • E. Roilides
    Affiliations
    Infectious Diseases Unit, 3rd Department of Paediatrics, Faculty of Medicine, Aristotle University School of Health Sciences, Thessaloniki, Greece

    Hippokration General Hospital, Thessaloniki, Greece

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • M. Ruhnke
    Affiliations
    Department of Haematology and Oncology, Paracelsus Hospital, Osnabrück, Germany

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • M. Sanguinetti
    Affiliations
    Institute of Microbiology, Fondazione Policlinico Universitario A. Gemelli – Università Cattolica del Sacro Cuore, Rome, Italy

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • D.C. Sheppard
    Affiliations
    Division of Infectious Diseases, Department of Medicine, Microbiology and Immunology, McGill University, Montreal, Canada

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • J. Sinkó
    Affiliations
    Department of Haematology and Stem Cell Transplantation, Szent István and Szent László Hospital, Budapest, Hungary

    ESCMID Fungal Infection Study Group (EFISG)
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  • A. Skiada
    Affiliations
    First Department of Medicine, Laiko Hospital, National and Kapodistrian University of Athens, Athens, Greece

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • M.J.G.T. Vehreschild
    Affiliations
    Department I of Internal Medicine, ECMM Excellence Centre of Medical Mycology, University Hospital of Cologne, Cologne, Germany

    Centre for Integrated Oncology, Cologne-Bonn, University of Cologne, Cologne, Germany

    German Centre for Infection Research (DZIF) partner site Bonn-Cologne, Cologne, Germany

    European Confederation of Medical Mycology (ECMM)
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  • C. Viscoli
    Affiliations
    Ospedale Policlinico San Martino and University of Genova (DISSAL), Genova, Italy

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)
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  • O.A. Cornely
    Correspondence
    Corresponding author. O.A. Cornely, FECMM, Department I for Internal Medicine, ECMM Excellence Centre of Medical Mycology, University Hospital, Kerpener Strasse 62, 50937 Cologne, Germany.
    Affiliations
    First Department of Medicine, Laiko Hospital, National and Kapodistrian University of Athens, Athens, Greece

    German Centre for Infection Research (DZIF) partner site Bonn-Cologne, Cologne, Germany

    CECAD Cluster of Excellence, University of Cologne, Cologne, Germany

    Clinical Trials Center Cologne, University Hospital of Cologne, Cologne, Germany

    ESCMID Fungal Infection Study Group (EFISG)

    European Confederation of Medical Mycology (ECMM)

    ESCMID European Study Group for Infections in Compromised Hosts (ESGICH)
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Open ArchivePublished:March 12, 2018DOI:https://doi.org/10.1016/j.cmi.2018.01.002

      Abstract

      The European Society for Clinical Microbiology and Infectious Diseases, the European Confederation of Medical Mycology and the European Respiratory Society Joint Clinical Guidelines focus on diagnosis and management of aspergillosis. Of the numerous recommendations, a few are summarized here. Chest computed tomography as well as bronchoscopy with bronchoalveolar lavage (BAL) in patients with suspicion of pulmonary invasive aspergillosis (IA) are strongly recommended. For diagnosis, direct microscopy, preferably using optical brighteners, histopathology and culture are strongly recommended. Serum and BAL galactomannan measures are recommended as markers for the diagnosis of IA. PCR should be considered in conjunction with other diagnostic tests. Pathogen identification to species complex level is strongly recommended for all clinically relevant Aspergillus isolates; antifungal susceptibility testing should be performed in patients with invasive disease in regions with resistance found in contemporary surveillance programmes. Isavuconazole and voriconazole are the preferred agents for first-line treatment of pulmonary IA, whereas liposomal amphotericin B is moderately supported. Combinations of antifungals as primary treatment options are not recommended. Therapeutic drug monitoring is strongly recommended for patients receiving posaconazole suspension or any form of voriconazole for IA treatment, and in refractory disease, where a personalized approach considering reversal of predisposing factors, switching drug class and surgical intervention is also strongly recommended. Primary prophylaxis with posaconazole is strongly recommended in patients with acute myelogenous leukaemia or myelodysplastic syndrome receiving induction chemotherapy. Secondary prophylaxis is strongly recommended in high-risk patients. We strongly recommend treatment duration based on clinical improvement, degree of immunosuppression and response on imaging.

      Keywords

      Introduction

      This is the third fungal diagnosis and management clinical guideline published in cooperation with various European scientific societies [
      • Arendrup M.C.
      • Boekhout T.
      • Akova M.
      • Meis J.F.
      • Cornely O.A.
      ESCMID and ECMM joint clinical guidelines for the diagnosis and management of rare invasive yeast infections.
      ,
      • Chowdhary A.
      • Meis J.F.
      • Guarro J.
      • de Hoog G.S.
      • Kathuria S.
      • Arendrup M.C.
      • et al.
      ESCMID and ECMM joint clinical guidelines for the diagnosis and management of systemic phaeohyphomycosis: diseases caused by black fungi.
      ,
      • Cornely O.A.
      • Arikan-Akdagli S.
      • Dannaoui E.
      • Groll A.H.
      • Lagrou K.
      • Chakrabarti A.
      • et al.
      ESCMID and ECMM joint clinical guidelines for the diagnosis and management of mucormycosis 2013.
      ,
      • Cornely O.A.
      • Bassetti M.
      • Calandra T.
      • Garbino J.
      • Kullberg B.J.
      • Lortholary O.
      • et al.
      ESCMID* guideline for the diagnosis and management of Candida diseases 2012: non-neutropenic adult patients.
      ,
      • Cuenca-Estrella M.
      • Verweij P.E.
      • Arendrup M.C.
      • Arikan-Akdagli S.
      • Bille J.
      • Donnelly J.P.
      • et al.
      ESCMID* guideline for the diagnosis and management of Candida diseases 2012: diagnostic procedures.
      ,
      • Hope W.W.
      • Castagnola E.
      • Groll A.H.
      • Roilides E.
      • Akova M.
      • Arendrup M.C.
      • et al.
      ESCMID* guideline for the diagnosis and management of Candida diseases 2012: prevention and management of invasive infections in neonates and children caused by Candida spp.
      ,
      • Lortholary O.
      • Petrikkos G.
      • Akova M.
      • Arendrup M.C.
      • Arikan-Akdagli S.
      • Bassetti M.
      • et al.
      ESCMID* guideline for the diagnosis and management of Candida diseases 2012: patients with HIV infection or AIDS.
      ,
      • Tortorano A.M.
      • Richardson M.
      • Roilides E.
      • van Diepeningen A.
      • Caira M.
      • Munoz P.
      • et al.
      ESCMID and ECMM joint guidelines on diagnosis and management of hyalohyphomycosis: Fusarium spp., Scedosporium spp. and others.
      ,
      • Ullmann A.J.
      • Akova M.
      • Herbrecht R.
      • Viscoli C.
      • Arendrup M.C.
      • Arikan-Akdagli S.
      • et al.
      ESCMID* guideline for the diagnosis and management of Candida diseases 2012: adults with haematological malignancies and after haematopoietic stem cell transplantation (HCT).
      ]. This part of the guideline regarding invasive and chronic aspergillosis is a condensation of all the recommendations made by the guideline subcommittees and is presented in tables for easier and faster reading. More details on how the recommendations were arrived at are planned in supplementary publications. This Aspergillus guideline will follow the style of other guidelines by including diagnostic and therapeutic guidance. Other scientific groups have published guidelines on this topic previously and all follow the common goal to provide clinicians with best guidance in their everyday working environment. Our goal was to provide a comprehensive European guideline focusing on the life-threatening diseases caused by Aspergillus spp.

      Methods

      Author panel recruitment and organization was similar to what was done previously [
      • Ullmann A.J.
      • Cornely O.A.
      • Donnelly J.P.
      • Akova M.
      • Arendrup M.C.
      • Arikan-Akdagli S.
      • et al.
      ESCMID* guideline for the diagnosis and management of Candida diseases 2012: developing European guidelines in clinical microbiology and infectious diseases.
      ]. In brief, experts in the field were nominated by the three societies: European Society for Clinical Microbiology and Infectious Diseases (ESCMID), the European Confederation of Medical Mycology (ECMM) and the European Respiratory Society (ERS). The total of 53 authors were grouped into their special fields of expertise. Subgroup coordinators were responsible for the first draft of recommendations. There were two face-to-face meetings followed by numerous electronic exchanges. Some of the first recommendations were presented at ECCMID 2014. This summary was reviewed and approved by all authors and sent to the ESCMID guideline director for public review. Then the final version was submitted to Clinical Microbiology and Infection for additional peer review and subsequent publication. Only the rationale of the chronic pulmonary aspergillosis (CPA) guideline was published ahead of time [
      • Denning D.W.
      • Cadranel J.
      • Beigelman-Aubry C.
      • Ader F.
      • Chakrabarti A.
      • Blot S.
      • et al.
      Chronic pulmonary aspergillosis: rationale and clinical guidelines for diagnosis and management.
      ].
      Questions were predefined and modified where appropriate and the strength of recommendation and quality of evidence was slightly modified (Table 1) [
      • Cornely O.A.
      • Cuenca-Estrella M.
      • Meis J.F.
      • Ullmann A.J.
      European Society of Clinical Microbiology and Infectious Diseases (ESCMID) Fungal Infection Study Group (EFISG) and European Confederation of Medical Mycology (ECMM) 2013 joint guidelines on diagnosis and management of rare and emerging fungal diseases.
      ]. Diagnostic tests are regarded as interventions.
      Table 1Strength of recommendation and quality of evidence
      Strength of recommendationDefinition
      Grade ASocieties strongly support a recommendation for use
      Grade BSocieties moderately support a recommendation for use
      Grade CSocieties marginally support a recommendation for use
      Grade DSocieties support a recommendation against use
      Quality of evidenceDefinition
      Level IEvidence from at least one properly designed randomized, controlled trial (oriented on the primary end point of the trial)
      Level IIEvidence from at least one well-designed clinical trial (including secondary end points), without randomization; from cohort or case–controlled analytic studies (preferably from more than one centre); from multiple time series; or from dramatic results of uncontrolled experiments
      Level IIIEvidence from opinions of respected authorities, based on clinical experience, descriptive case studies, or reports of expert committees
      Added indexSource of Level II evidence
      rMeta-analysis or systematic review of randomized controlled trials
      tTransferred evidence, i.e. results from different patients‘ cohorts, or similar immune-status situation
      hComparator group: historical control
      uUncontrolled trials
      aPublished abstract presented at an international symposium or meeting
      ∗ Poor quality of planning, inconsistency of results, indirectness of evidence etc. would lower the Strength of recommendation.

      Summary of recommendations

       Diagnostic procedures

      Early diagnosis of invasive aspergillosis (IA) is a challenge and should be based on the integration of clinical, radiological and microbiological data.

       Thoracic imaging

      In patients at risk for IA with fever of unknown origin or clinical symptoms of lower respiratory tract infection who remain febrile despite broad-spectrum antibacterial treatment, thin-section chest computed tomography (multidetector (MDCT), multislice (MSCT), spiral CT, high resolution CT) at optimized dose (according to the ALARA (‘As Low As Reasonably Achievable’) principle) is the imaging modality of choice (AII) [
      • Marchiori E.
      • Irion K.L.
      Commentary on: "Analysis of initial and follow-up CT findings in patients with invasive pulmonary aspergillosis after solid organ transplantation".
      ,
      • Bergeron A.
      • Porcher R.
      • Sulahian A.
      • de Bazelaire C.
      • Chagnon K.
      • Raffoux E.
      • et al.
      The strategy for the diagnosis of invasive pulmonary aspergillosis should depend on both the underlying condition and the leukocyte count of patients with hematologic malignancies.
      ,
      • Lim C.
      • Seo J.B.
      • Park S.Y.
      • Hwang H.J.
      • Lee H.J.
      • Lee S.O.
      • et al.
      Analysis of initial and follow-up CT findings in patients with invasive pulmonary aspergillosis after solid organ transplantation.
      ,
      • Wingard J.R.
      New approaches to invasive fungal infections in acute leukemia and hematopoietic stem cell transplant patients.
      ,
      • Greene R.E.
      • Schlamm H.T.
      • Oestmann J.W.
      • Stark P.
      • Durand C.
      • Lortholary O.
      • et al.
      Imaging findings in acute invasive pulmonary aspergillosis: clinical significance of the halo sign.
      ,
      • Greene R.
      The radiological spectrum of pulmonary aspergillosis.
      ,
      • Heussel C.P.
      • Kauczor H.U.
      • Heussel G.
      • Fischer B.
      • Mildenberger P.
      • Thelen M.
      Early detection of pneumonia in febrile neutropenic patients: use of thin-section CT.
      ,
      • Heussel C.P.
      • Kauczor H.U.
      • Heussel G.E.
      • Fischer B.
      • Begrich M.
      • Mildenberger P.
      • et al.
      Pneumonia in febrile neutropenic patients and in bone marrow and blood stem-cell transplant recipients: use of high-resolution computed tomography.
      ,
      • Caillot D.
      • Casasnovas O.
      • Bernard A.
      • Couaillier J.F.
      • Durand C.
      • Cuisenier B.
      • et al.
      Improved management of invasive pulmonary aspergillosis in neutropenic patients using early thoracic computed tomographic scan and surgery.
      ,
      • Caillot D.
      • Mannone L.
      • Cuisenier B.
      • Couaillier J.F.
      Role of early diagnosis and aggressive surgery in the management of invasive pulmonary aspergillosis in neutropenic patients.
      ,
      • Chamilos G.
      • Marom E.M.
      • Lewis R.E.
      • Lionakis M.S.
      • Kontoyiannis D.P.
      Predictors of pulmonary zygomycosis versus invasive pulmonary aspergillosis in patients with cancer.
      ]. Pulmonary CT angiography may be of interest in the early diagnosis of IA by depicting directly vessel occlusion at the level of a suspicious fungal lesion with a potential high negative predictive value regarding imaging evaluation [
      • Stanzani M.
      • Battista G.
      • Sassi C.
      • Lewis R.E.
      • Tolomelli G.
      • Clissa C.
      • et al.
      Computed tomographic pulmonary angiography for diagnosis of invasive mold diseases in patients with hematological malignancies.
      ,
      • Stanzani M.
      • Sassi C.
      • Lewis R.E.
      • Tolomelli G.
      • Bazzocchi A.
      • Cavo M.
      • et al.
      High resolution computed tomography angiography improves the radiographic diagnosis of invasive mold disease in patients with hematological malignancies.
      ,
      • Sonnet S.
      • Buitrago-Tellez C.H.
      • Tamm M.
      • Christen S.
      • Steinbrich W.
      Direct detection of angioinvasive pulmonary aspergillosis in immunosuppressed patients: preliminary results with high-resolution 16-MDCT angiography.
      ], and is required in case of haemoptysis (AII). In selected patients where CT is not wanted or not feasible, magnetic resonance imaging (MRI) of the lungs may represent an alternative imaging to thin-section MSCT [
      • Sodhi K.S.
      • Khandelwal N.
      • Saxena A.K.
      • Singh M.
      • Agarwal R.
      • Bhatia A.
      • et al.
      Rapid lung MRI in children with pulmonary infections: time to change our diagnostic algorithms.
      ,
      • Maschmeyer G.
      • Carratala J.
      • Buchheidt D.
      • Hamprecht A.
      • Heussel C.P.
      • Kahl C.
      • et al.
      Diagnosis and antimicrobial therapy of lung infiltrates in febrile neutropenic patients (allogeneic SCT excluded): updated guidelines of the infectious diseases working party (AGIHO) of the German Society of Hematology and Medical Oncology (DGHO).
      ,
      • Rieger C.
      • Herzog P.
      • Eibel R.
      • Fiegl M.
      • Ostermann H.
      Pulmonary MRI—a new approach for the evaluation of febrile neutropenic patients with malignancies.
      ,
      • Araz O.
      • Karaman A.
      • Ucar E.Y.
      • Bilen Y.
      • Durur Subasi I.
      DCE-MRI findings of invasive aspergillosis in patient with acute myeloid leukemia.
      ,
      • Blum U.
      • Windfuhr M.
      • Buitrago-Tellez C.
      • Sigmund G.
      • Herbst E.W.
      • Langer M.
      Invasive pulmonary aspergillosis. MRI, CT, and plain radiographic findings and their contribution for early diagnosis.
      ,
      • Yan C.
      • Tan X.
      • Wei Q.
      • Feng R.
      • Li C.
      • Wu Y.
      • et al.
      Lung MRI of invasive fungal infection at 3 tesla: evaluation of five different pulse sequences and comparison with multidetector computed tomography (MDCT).
      ], positron emission tomography-CT being of modest interest in the diagnostics of IA [
      • Hot A.
      • Maunoury C.
      • Poiree S.
      • Lanternier F.
      • Viard J.P.
      • Loulergue P.
      • et al.
      Diagnostic contribution of positron emission tomography with [18F]fluorodeoxyglucose for invasive fungal infections.
      ,
      • Camus V.
      • Edet-Sanson A.
      • Bubenheim M.
      • Hitzel A.
      • Becker S.
      • David M.
      • et al.
      (1)(8)F-FDG-PET/CT imaging in patients with febrile neutropenia and haematological malignancies.
      ].
      No CT scanning technique is 100% sensitive or specific for pulmonary IA [
      • Desoubeaux G.
      • Bailly E.
      • Chandenier J.
      Diagnosis of invasive pulmonary aspergillosis: updates and recommendations.
      ,
      • Bruno C.
      • Minniti S.
      • Vassanelli A.
      • Pozzi-Mucelli R.
      Comparison of CT features of aspergillus and bacterial pneumonia in severely neutropenic patients.
      ,
      • Nucci M.
      • Nouer S.A.
      • Grazziutti M.
      • Kumar N.S.
      • Barlogie B.
      • Anaissie E.
      Probable invasive aspergillosis without prespecified radiologic findings: proposal for inclusion of a new category of aspergillosis and implications for studying novel therapies.
      ]. Classical CT findings of angioinvasive aspergillosis include macronodule(s) >1 cm, which may be surrounded by a halo of ground-glass attenuation (halo sign, early phase, inconstant) [
      • Bruno C.
      • Minniti S.
      • Vassanelli A.
      • Pozzi-Mucelli R.
      Comparison of CT features of aspergillus and bacterial pneumonia in severely neutropenic patients.
      ,
      • Caillot D.
      • Couaillier J.F.
      • Bernard A.
      • Casasnovas O.
      • Denning D.W.
      • Mannone L.
      • et al.
      Increasing volume and changing characteristics of invasive pulmonary aspergillosis on sequential thoracic computed tomography scans in patients with neutropenia.
      ,
      • Kami M.
      • Kishi Y.
      • Hamaki T.
      • Kawabata M.
      • Kashima T.
      • Masumoto T.
      • et al.
      The value of the chest computed tomography halo sign in the diagnosis of invasive pulmonary aspergillosis. An autopsy-based retrospective study of 48 patients.
      ,
      • Kuhlman J.E.
      • Fishman E.K.
      • Burch P.A.
      • Karp J.E.
      • Zerhouni E.A.
      • Siegelman S.S.
      Invasive pulmonary aspergillosis in acute leukemia. The contribution of CT to early diagnosis and aggressive management.
      ], pleural based wedge-shaped areas of consolidation [
      • Franquet T.
      • Gimenez A.
      • Hidalgo A.
      Imaging of opportunistic fungal infections in immunocompromised patient.
      ], alveolar consolidations [
      • Bruno C.
      • Minniti S.
      • Vassanelli A.
      • Pozzi-Mucelli R.
      Comparison of CT features of aspergillus and bacterial pneumonia in severely neutropenic patients.
      ,
      • Horger M.
      • Hebart H.
      • Einsele H.
      • Lengerke C.
      • Claussen C.D.
      • Vonthein R.
      • et al.
      Initial CT manifestations of invasive pulmonary aspergillosis in 45 non-HIV immunocompromised patients: association with patient outcome?.
      ,
      • Althoff Souza C.
      • Muller N.L.
      • Marchiori E.
      • Escuissato D.L.
      • Franquet T.
      Pulmonary invasive aspergillosis and candidiasis in immunocompromised patients: a comparative study of the high-resolution CT findings.
      ], masses (especially in solid organ transplant (SOT) recipients) [
      • Lim C.
      • Seo J.B.
      • Park S.Y.
      • Hwang H.J.
      • Lee H.J.
      • Lee S.O.
      • et al.
      Analysis of initial and follow-up CT findings in patients with invasive pulmonary aspergillosis after solid organ transplantation.
      ,
      • Caillot D.
      • Couaillier J.F.
      • Bernard A.
      • Casasnovas O.
      • Denning D.W.
      • Mannone L.
      • et al.
      Increasing volume and changing characteristics of invasive pulmonary aspergillosis on sequential thoracic computed tomography scans in patients with neutropenia.
      ], internal low attenuation [
      • Horger M.
      • Einsele H.
      • Schumacher U.
      • Wehrmann M.
      • Hebart H.
      • Lengerke C.
      • et al.
      Invasive pulmonary aspergillosis: frequency and meaning of the "hypodense sign" on unenhanced CT.
      ], reverse halo sign [
      • Marchiori E.
      • Godoy M.C.
      • Zanetti G.
      • Hochhegger B.
      • Rodrigues R.S.
      The reversed halo sign. Another CT finding useful for distinguish invasive pulmonary aspergillosis and pulmonary lymphoma.
      ], cavity or air-crescent sign (delayed finding), ground glass opacities and pleural effusion [
      • Greene R.E.
      • Schlamm H.T.
      • Oestmann J.W.
      • Stark P.
      • Durand C.
      • Lortholary O.
      • et al.
      Imaging findings in acute invasive pulmonary aspergillosis: clinical significance of the halo sign.
      ,
      • Desoubeaux G.
      • Bailly E.
      • Chandenier J.
      Diagnosis of invasive pulmonary aspergillosis: updates and recommendations.
      ,
      • Kojima R.
      • Tateishi U.
      • Kami M.
      • Murashige N.
      • Nannya Y.
      • Kusumi E.
      • et al.
      Chest computed tomography of late invasive aspergillosis after allogeneic hematopoietic stem cell transplantation.
      ]. Bronchoinvasive forms may appear as tracheal or bronchial wall thickening, centrilobular nodules with tree in bud appearance [
      • Bergeron A.
      • Porcher R.
      • Sulahian A.
      • de Bazelaire C.
      • Chagnon K.
      • Raffoux E.
      • et al.
      The strategy for the diagnosis of invasive pulmonary aspergillosis should depend on both the underlying condition and the leukocyte count of patients with hematologic malignancies.
      ] in a patchy distribution, predominant peribronchial areas of consolidation [
      • Logan P.M.
      • Primack S.L.
      • Miller R.R.
      • Muller N.L.
      Invasive aspergillosis of the airways: radiographic, CT, and pathologic findings.
      ] or bronchopneumonia [
      • Kojima R.
      • Tateishi U.
      • Kami M.
      • Murashige N.
      • Nannya Y.
      • Kusumi E.
      • et al.
      Chest computed tomography of late invasive aspergillosis after allogeneic hematopoietic stem cell transplantation.
      ] (Table 2).
      Table 2Recommendations for imaging and bronchoalveolar lavage
      PopulationIntentionIntervention
      a Diagnostic tests are interventions.
      SoRQoECommentRef.
      Neutropenia, fever or clinical symptoms of pneumonia, empiric antibiotics failing to achieve defervescence, e.g. FUOTo detect pulmonary infiltratesChest CT and thin section multi-detector CT (MDCT)AIIDose optimization recommended
      • Caillot D.
      • Casasnovas O.
      • Bernard A.
      • Couaillier J.F.
      • Durand C.
      • Cuisenier B.
      • et al.
      Improved management of invasive pulmonary aspergillosis in neutropenic patients using early thoracic computed tomographic scan and surgery.
      ,
      • Blum U.
      • Windfuhr M.
      • Buitrago-Tellez C.
      • Sigmund G.
      • Herbst E.W.
      • Langer M.
      Invasive pulmonary aspergillosis. MRI, CT, and plain radiographic findings and their contribution for early diagnosis.
      ,
      • Desoubeaux G.
      • Bailly E.
      • Chandenier J.
      Diagnosis of invasive pulmonary aspergillosis: updates and recommendations.
      ,
      • Pasmans H.L.
      • Loosveld O.J.
      • Schouten H.C.
      • Thunnissen F.
      • van Engelshoven J.M.
      Invasive aspergillosis in immunocompromised patients: findings on plain film and (hr)CT.
      To identify vessel occlusionChest angio-CT/pulmonary CT angiographyBII
      • Stanzani M.
      • Battista G.
      • Sassi C.
      • Lewis R.E.
      • Tolomelli G.
      • Clissa C.
      • et al.
      Computed tomographic pulmonary angiography for diagnosis of invasive mold diseases in patients with hematological malignancies.
      ,
      • Stanzani M.
      • Sassi C.
      • Lewis R.E.
      • Tolomelli G.
      • Bazzocchi A.
      • Cavo M.
      • et al.
      High resolution computed tomography angiography improves the radiographic diagnosis of invasive mold disease in patients with hematological malignancies.
      ,
      • Sonnet S.
      • Buitrago-Tellez C.H.
      • Tamm M.
      • Christen S.
      • Steinbrich W.
      Direct detection of angioinvasive pulmonary aspergillosis in immunosuppressed patients: preliminary results with high-resolution 16-MDCT angiography.
      HaemoptysisTo identify vessel erosionChest angio-CT/pulmonary CT angiographyAII
      • Heussel C.P.
      • Kauczor H.U.
      • Heussel G.
      • Mildenberger P.
      • Dueber C.
      Aneurysms complicating inflammatory diseases in immunocompromised hosts: value of contrast-enhanced CT.
      ,
      • Didier M.
      • Guedin P.
      • Staub F.
      • Catherinot E.
      • Rivaud E.
      • Scherrer A.
      • et al.
      Pulmonary arterial mycotic pseudoaneurysms in a patient with invasive pulmonary aspergillosis. Successful occlusion by coils.
      Any, with infiltrateTo identify possible underlying fungal or other infectious diseaseBALAII
      • Caillot D.
      • Casasnovas O.
      • Bernard A.
      • Couaillier J.F.
      • Durand C.
      • Cuisenier B.
      • et al.
      Improved management of invasive pulmonary aspergillosis in neutropenic patients using early thoracic computed tomographic scan and surgery.
      ,
      • Azoulay E.
      • Mokart D.
      • Lambert J.
      • Lemiale V.
      • Rabbat A.
      • Kouatchet A.
      • et al.
      Diagnostic strategy for hematology and oncology patients with acute respiratory failure: randomized controlled trial.
      ,
      • Hummel M.
      • Rudert S.
      • Hof H.
      • Hehlmann R.
      • Buchheidt D.
      Diagnostic yield of bronchoscopy with bronchoalveolar lavage in febrile patients with hematologic malignancies and pulmonary infiltrates.
      ,
      • Boersma W.G.
      • Erjavec Z.
      • van der Werf T.S.
      • de Vries-Hosper H.G.
      • Gouw A.S.
      • Manson W.L.
      Bronchoscopic diagnosis of pulmonary infiltrates in granulocytopenic patients with hematologic malignancies: BAL versus PSB and PBAL.
      ,
      • Cornillet A.
      • Camus C.
      • Nimubona S.
      • Gandemer V.
      • Tattevin P.
      • Belleguic C.
      • et al.
      Comparison of epidemiological, clinical, and biological features of invasive aspergillosis in neutropenic and nonneutropenic patients: a 6-year survey.
      ,
      • Jain P.
      • Sandur S.
      • Meli Y.
      • Arroliga A.C.
      • Stoller J.K.
      • Mehta A.C.
      Role of flexible bronchoscopy in immunocompromised patients with lung infiltrates.
      ,
      • Peikert T.
      • Rana S.
      • Edell E.S.
      Safety, diagnostic yield, and therapeutic implications of flexible bronchoscopy in patients with febrile neutropenia and pulmonary infiltrates.
      Any, with infiltrateTo obtain appropriate specimens for microscopy, culture and PCRCT-guided BALAIII
      • Ramila E.
      • Sureda A.
      • Martino R.
      • Santamaria A.
      • Franquet T.
      • Puzo C.
      • et al.
      Bronchoscopy guided by high-resolution computed tomography for the diagnosis of pulmonary infections in patients with hematologic malignancies and normal plain chest X-ray.
      ,
      • Becker M.J.
      • Lugtenburg E.J.
      • Cornelissen J.J.
      • Van Der Schee C.
      • Hoogsteden H.C.
      • De Marie S.
      Galactomannan detection in computerized tomography-based broncho-alveolar lavage fluid and serum in haematological patients at risk for invasive pulmonary aspergillosis.
      Abbreviations: BAL, bronchoalveolar lavage; CT, computed tomography; FUO, fever of unknown origin; PCR, polymerase chain reaction; QoE, Quality of evidence; SoR, Strength of recommendation.
      a Diagnostic tests are interventions.

       Bronchoalveolar lavage and biopsies

      Other diagnostic procedures include early bronchoalveolar lavage (BAL) (AII) [
      • Liss B.
      • Vehreschild J.J.
      • Bangard C.
      • Maintz D.
      • Frank K.
      • Gronke S.
      • et al.
      Our 2015 approach to invasive pulmonary aspergillosis.
      ,
      • Azoulay E.
      • Mokart D.
      • Lambert J.
      • Lemiale V.
      • Rabbat A.
      • Kouatchet A.
      • et al.
      Diagnostic strategy for hematology and oncology patients with acute respiratory failure: randomized controlled trial.
      ,
      • Hummel M.
      • Rudert S.
      • Hof H.
      • Hehlmann R.
      • Buchheidt D.
      Diagnostic yield of bronchoscopy with bronchoalveolar lavage in febrile patients with hematologic malignancies and pulmonary infiltrates.
      ,
      • Boersma W.G.
      • Erjavec Z.
      • van der Werf T.S.
      • de Vries-Hosper H.G.
      • Gouw A.S.
      • Manson W.L.
      Bronchoscopic diagnosis of pulmonary infiltrates in granulocytopenic patients with hematologic malignancies: BAL versus PSB and PBAL.
      ,
      • Cornillet A.
      • Camus C.
      • Nimubona S.
      • Gandemer V.
      • Tattevin P.
      • Belleguic C.
      • et al.
      Comparison of epidemiological, clinical, and biological features of invasive aspergillosis in neutropenic and nonneutropenic patients: a 6-year survey.
      ,
      • Jain P.
      • Sandur S.
      • Meli Y.
      • Arroliga A.C.
      • Stoller J.K.
      • Mehta A.C.
      Role of flexible bronchoscopy in immunocompromised patients with lung infiltrates.
      ,
      • Peikert T.
      • Rana S.
      • Edell E.S.
      Safety, diagnostic yield, and therapeutic implications of flexible bronchoscopy in patients with febrile neutropenia and pulmonary infiltrates.
      ], guided by CT findings [
      • Ramila E.
      • Sureda A.
      • Martino R.
      • Santamaria A.
      • Franquet T.
      • Puzo C.
      • et al.
      Bronchoscopy guided by high-resolution computed tomography for the diagnosis of pulmonary infections in patients with hematologic malignancies and normal plain chest X-ray.
      ,
      • Becker M.J.
      • Lugtenburg E.J.
      • Cornelissen J.J.
      • Van Der Schee C.
      • Hoogsteden H.C.
      • De Marie S.
      Galactomannan detection in computerized tomography-based broncho-alveolar lavage fluid and serum in haematological patients at risk for invasive pulmonary aspergillosis.
      ], and less frequently CT-guided transthoracic biopsies, video-assisted thoracoscopic surgery, open lung biopsies, transbronchial biopsies or convex endobronchial ultrasound transbronchial needle aspiration, the latter technique appearing to be a promising procedure in this setting [
      • Maschmeyer G.
      • Carratala J.
      • Buchheidt D.
      • Hamprecht A.
      • Heussel C.P.
      • Kahl C.
      • et al.
      Diagnosis and antimicrobial therapy of lung infiltrates in febrile neutropenic patients (allogeneic SCT excluded): updated guidelines of the infectious diseases working party (AGIHO) of the German Society of Hematology and Medical Oncology (DGHO).
      ,
      • de Bazelaire C.
      • Coffin A.
      • Cohen-Zarade S.
      • de Margerie-Mellon C.
      • Scemama A.
      • Sabatier F.
      • et al.
      CT-guided biopsies in lung infections in patients with haematological malignancies.
      ,
      • Reinwald M.
      • Spiess B.
      • Heinz W.J.
      • Heussel C.P.
      • Bertz H.
      • Cornely O.A.
      • et al.
      Aspergillus PCR-based investigation of fresh tissue and effusion samples in patients with suspected invasive aspergillosis enhances diagnostic capabilities.
      ,
      • Gupta S.
      • Sultenfuss M.
      • Romaguera J.E.
      • Ensor J.
      • Krishnamurthy S.
      • Wallace M.J.
      • et al.
      CT-guided percutaneous lung biopsies in patients with haematologic malignancies and undiagnosed pulmonary lesions.
      ,
      • Shi J.M.
      • Cai Z.
      • Huang H.
      • Ye X.J.
      • He J.S.
      • Xie W.Z.
      • et al.
      Role of CT-guided percutaneous lung biopsy in diagnosis of pulmonary fungal infection in patients with hematologic diseases.
      ,
      • Lass-Florl C.
      • Resch G.
      • Nachbaur D.
      • Mayr A.
      • Gastl G.
      • Auberger J.
      • et al.
      The value of computed tomography-guided percutaneous lung biopsy for diagnosis of invasive fungal infection in immunocompromised patients.
      ,
      • Rickerts V.
      • Mousset S.
      • Lambrecht E.
      • Tintelnot K.
      • Schwerdtfeger R.
      • Presterl E.
      • et al.
      Comparison of histopathological analysis, culture, and polymerase chain reaction assays to detect invasive mold infections from biopsy specimens.
      ,
      • Carrafiello G.
      • Lagana D.
      • Nosari A.M.
      • Guffanti C.
      • Morra E.
      • Recaldini C.
      • et al.
      Utility of computed tomography (ct) and of fine needle aspiration biopsy (fnab) in early diagnosis of fungal pulmonary infections. Study of infections from filamentous fungi in haematologically immunodeficient patients.
      ,
      • Nosari A.
      • Anghilieri M.
      • Carrafiello G.
      • Guffanti C.
      • Marbello L.
      • Montillo M.
      • et al.
      Utility of percutaneous lung biopsy for diagnosing filamentous fungal infections in hematologic malignancies.
      ,
      • Manhire A.
      • Charig M.
      • Clelland C.
      • Gleeson F.
      • Miller R.
      • Moss H.
      • et al.
      Guidelines for radiologically guided lung biopsy.
      ,
      • Georgiadou S.P.
      • Sampsonas F.L.
      • Rice D.
      • Granger J.M.
      • Swisher S.
      • Kontoyiannis D.P.
      Open-lung biopsy in patients with undiagnosed lung lesions referred at a tertiary cancer center is safe and reveals noncancerous, noninfectious entities as the most common diagnoses.
      ,
      • Armenian S.H.
      • Hoffman J.A.
      • Butturini A.M.
      • Kapoor N.
      • Mascarenhas L.
      Invasive diagnostic procedures for pulmonary infiltrates in pediatric hematopoietic stem cell transplant recipients.
      ,
      • Zihlif M.
      • Khanchandani G.
      • Ahmed H.P.
      • Soubani A.O.
      Surgical lung biopsy in patients with hematological malignancy or hematopoietic stem cell transplantation and unexplained pulmonary infiltrates: improved outcome with specific diagnosis.
      ,
      • Wingard J.R.
      • Hiemenz J.W.
      • Jantz M.A.
      How I manage pulmonary nodular lesions and nodular infiltrates in patients with hematologic malignancies or undergoing hematopoietic cell transplantation.
      ,
      • Choi Y.R.
      • An J.Y.
      • Kim M.K.
      • Han H.S.
      • Lee K.H.
      • Kim S.W.
      • et al.
      The diagnostic efficacy and safety of endobronchial ultrasound-guided transbronchial needle aspiration as an initial diagnostic tool.
      ,
      • Casal R.F.
      • Adachi R.
      • Jimenez C.A.
      • Sarkiss M.
      • Morice R.C.
      • Eapen G.A.
      Diagnosis of invasive aspergillus tracheobronchitis facilitated by endobronchial ultrasound-guided transbronchial needle aspiration: a case report.
      ,
      • Aragaki-Nakahodo A.
      • Benzaquen S.
      • Kirschner M.
      Coinfection by Nocardia beijingensis and Nocardia arthritidis in an immunocompromised patient diagnosed by endobronchial ultrasound guided transbronchial needle aspiration (EBUS-TBNA).
      ]. Contraindications to these techniques need to be considered.

       Imaging of other sites

      Moreover, according to clinical symptoms, paranasal CT, CT or MRI of the central nervous system (CNS) as well as abdominal CT may also be required. In particular, findings of sinusitis with bone erosion may be observed, intracranial and/or intraorbital extension of the disease being best evaluated by MRI [
      • Thery A.
      • Espitalier F.
      • Cassagnau E.
      • Durand N.
      • Malard O.
      Clinical features and outcome of sphenoid sinus aspergillosis: a retrospective series of 15 cases.
      ,
      • Miyamoto Y.
      • Sakamoto Y.
      • Ohuchi M.
      • Tokunaga R.
      • Shigaki H.
      • Kurashige J.
      • et al.
      Orbital apex syndrome caused by invasive aspergillosis as an adverse effect of systemic chemotherapy for metastatic colorectal cancer: a case report.
      ,
      • Yuan L.
      • Prayson R.A.
      Optic nerve aspergillosis.
      ]. In the brain, due to direct spread from paranasal sinuses or haematogenous dissemination, meningeal enhancement or empyema, cerebral abscess, mycotic aneurysms as well as haemorrhagic lesions and rarely stroke may be seen [
      • Marzolf G.
      • Sabou M.
      • Lannes B.
      • Cotton F.
      • Meyronet D.
      • Galanaud D.
      • et al.
      Magnetic resonance imaging of cerebral aspergillosis: imaging and pathological correlations.
      ,
      • Ashdown B.C.
      • Tien R.D.
      • Felsberg G.J.
      Aspergillosis of the brain and paranasal sinuses in immunocompromised patients: CT and MR imaging findings.
      ,
      • DeLone D.R.
      • Goldstein R.A.
      • Petermann G.
      • Salamat M.S.
      • Miles J.M.
      • Knechtle S.J.
      • et al.
      Disseminated aspergillosis involving the brain: distribution and imaging characteristics.
      ,
      • Guermazi A.
      • Gluckman E.
      • Tabti B.
      • Miaux Y.
      Invasive central nervous system aspergillosis in bone marrow transplantation recipients: an overview.
      ].

       Microscopy and culture

      Both microscopy and culture should be attempted on appropriate specimens from patients at risk for IA (AII) with a priority for culture in most cases where insufficient material is available. Demonstrating tissue invasion by hyphae through microscopic examination of biopsy or autopsy material provides a diagnosis of proven invasive fungal infection. However, the sensitivity of microscopy for IA is 50% at best [
      • Rüchel R.
      • Schaffrinski M.
      Versatile fluorescent staining of fungi in clinical specimens by using the optical brightener blankophor.
      ]. Specimens may be examined as a wet mount preparation with or without the addition of 10% potassium hydroxide. Fluorescent dyes such as Calcofluor White™ or Blancophor™ have the advantages of increased sensitivity, rapid turnaround time and broad applicability but are not specific for Aspergillus (AII). Gomori's methenamine silver stain (GMS) and periodic acid-Schiff (PAS) can be applied to histological sections and smears and should be conducted in all cases in which IA is considered a possibility (Table 3). Respiratory secretions from patients with suspected aspergillosis must be processed rapidly for culture to prevent overgrowth by bacteria and yeasts. To achieve optimal recovery of Aspergillus from BAL fluid, centrifugation of the sample is advised with investigation of the sediment (AIII). It is recommended that cultures of high volume untreated sputum and BAL should be performed as opposed to culturing small volumes of digested, liquefied samples [
      • Fraczek M.G.
      • Kirwan M.B.
      • Moore C.B.
      • Morris J.
      • Denning D.W.
      • Richardson M.D.
      Volume dependency for culture of fungi from respiratory secretions and increased sensitivity of Aspergillus quantitative PCR.
      ] (Table 4). Specific media to support fungal growth are recommended. Species identification to the complex level should be carried out for clinically relevant isolates from patients who need antifungal treatment, and for epidemiological purposes (AIII) (Table 5).
      Table 3Microscopic examinations
      PopulationIntentionInterventionSoRQoECommentRef.
      AnyTo identify fungal elements in histological sections and stainsHistological examination

      Gomori's methenamine silver stain Periodic acid–Schiff
      AIIIHistopathology is an essential investigation

      Inability to definitively distinguish other filamentous fungi

      GMS: removes cellular background; more sensitive to hyphal elements

      PAS: advantage of counter stain to check cellular detail
      • Lass-Florl C.
      • Resch G.
      • Nachbaur D.
      • Mayr A.
      • Gastl G.
      • Auberger J.
      • et al.
      The value of computed tomography-guided percutaneous lung biopsy for diagnosis of invasive fungal infection in immunocompromised patients.
      ,
      • Rüchel R.
      • Schaffrinski M.
      Versatile fluorescent staining of fungi in clinical specimens by using the optical brightener blankophor.
      ,
      • Denning D.W.
      • Kibbler C.C.
      • Barnes R.A.
      British Society for Medical M
      British society for medical mycology proposed standards of care for patients with invasive fungal infections.
      ,
      • Vyzantiadis T.A.
      • Johnson E.M.
      • Kibbler C.C.
      From the patient to the clinical mycology laboratory: how can we optimise microscopy and culture methods for mould identification?.
      AnyTo identify fungal elements in histological sections and stainsFluorescent dyes: Calcofluor white™, Uvitex 2B, Blancophor™AIINot specific to Aspergillus but high sensitivity and the micromorphology may provide information on the fungal class (e.g. Aspergillus: typically dichotomous and septate, Mucorales: pauci-septate and 90° angle branching, yeast: budding)

      Rapid turnaround time

      Broad applicability

      May be applied to frozen sections, paraffin-embedded tissue
      • Choi J.K.
      • Mauger J.
      • McGowan K.L.
      Immunohistochemical detection of Aspergillus species in pediatric tissue samples.
      ,
      • Kaufman L.
      • Standard P.G.
      • Jalbert M.
      • Kraft D.E.
      Immunohistologic identification of Aspergillus spp. and other hyaline fungi by using polyclonal fluorescent antibodies.
      ,
      • Verweij P.E.
      • Smedts F.
      • Poot T.
      • Bult P.
      • Hoogkamp-Korstanje J.A.
      • Meis J.F.
      Immunoperoxidase staining for identification of Aspergillus species in routinely processed tissue sections.
      ,
      • Hayden R.T.
      • Isotalo P.A.
      • Parrett T.
      • Wolk D.M.
      • Qian X.
      • Roberts G.D.
      • et al.
      In situ hybridization for the differentiation of Aspergillus, Fusarium, and Pseudallescheria species in tissue section.
      ,
      • Sundaram C.
      • Umabala P.
      • Laxmi V.
      • Purohit A.K.
      • Prasad V.S.
      • Panigrahi M.
      • et al.
      Pathology of fungal infections of the central nervous system: 17 years' experience from southern India.
      AnyTo identify fungal elements in histological sections and stainsImmunohistochemistry

      Monoclonal antibody WF-AF-1 or EB-A1

      In situ hybridization
      BIIHave the potential to provide genus- and species-specific data

      Commercially available monoclonal antibodies

      WF-AF-1 is specific for Aspergillus fumigatus, Aspergillus flavus, and Aspergillus niger

      Time consuming and not broadly available
      • Choi J.K.
      • Mauger J.
      • McGowan K.L.
      Immunohistochemical detection of Aspergillus species in pediatric tissue samples.
      ,
      • Kaufman L.
      • Standard P.G.
      • Jalbert M.
      • Kraft D.E.
      Immunohistologic identification of Aspergillus spp. and other hyaline fungi by using polyclonal fluorescent antibodies.
      ,
      • Verweij P.E.
      • Smedts F.
      • Poot T.
      • Bult P.
      • Hoogkamp-Korstanje J.A.
      • Meis J.F.
      Immunoperoxidase staining for identification of Aspergillus species in routinely processed tissue sections.
      ,
      • Hayden R.T.
      • Isotalo P.A.
      • Parrett T.
      • Wolk D.M.
      • Qian X.
      • Roberts G.D.
      • et al.
      In situ hybridization for the differentiation of Aspergillus, Fusarium, and Pseudallescheria species in tissue section.
      ,
      • Sundaram C.
      • Umabala P.
      • Laxmi V.
      • Purohit A.K.
      • Prasad V.S.
      • Panigrahi M.
      • et al.
      Pathology of fungal infections of the central nervous system: 17 years' experience from southern India.
      AnyTo identify fungal elements in fresh clinical specimens (e.g. BAL)Application of fluorescent dyes Calcofluor white™ or Uvitex 2B or Blancophor™AIIEssential investigation

      Not specific for Aspergillus species

      High sensitivity

      Rapid turn-around time

      Broad applicability

      No species identification but the micromorphology may provide information on the fungal class (e.g. Aspergillus: typically dichotomous and septate, Mucorales: pauci-septate and 90° angle branching, yeast: budding)
      • Lass-Florl C.
      • Resch G.
      • Nachbaur D.
      • Mayr A.
      • Gastl G.
      • Auberger J.
      • et al.
      The value of computed tomography-guided percutaneous lung biopsy for diagnosis of invasive fungal infection in immunocompromised patients.
      ,
      • Rüchel R.
      • Schaffrinski M.
      Versatile fluorescent staining of fungi in clinical specimens by using the optical brightener blankophor.
      ,
      • Chander J.
      • Chakrabarti A.
      • Sharma A.
      • Saini J.S.
      • Panigarhi D.
      Evaluation of calcofluor staining in the diagnosis of fungal corneal ulcer.
      Abbreviations: BAL, bronchoalveolar lavage; CNS, central nervous system; GMS, Gomori's methenamine silver stain; HE, haematoxylin-eosin; PAS, Periodic acid–Schiff; QoE, Quality of evidence; SoR, Strength of recommendation.
      Table 4Sample selection and pre-analytical respiratory sample treatment
      PopulationIntentionInterventionSoRQoECommentRef.
      AnyTo achieve a homogeneous sample of viscous samples such as sputumLiquefaction using a mucolytic agent, e.g. Pancreatin®, Sputolysin®, or using sonication and 1,4-dithiothreitolAIIIEssential investigation

      High-volume sputum culture (entire sample) shown to significantly increase recovery
      • Fraczek M.G.
      • Kirwan M.B.
      • Moore C.B.
      • Morris J.
      • Denning D.W.
      • Richardson M.D.
      Volume dependency for culture of fungi from respiratory secretions and increased sensitivity of Aspergillus quantitative PCR.
      ,
      • Baxter C.G.
      • Jones A.M.
      • Webb K.
      • Denning D.W.
      Homogenisation of cystic fibrosis sputum by sonication—an essential step for Aspergillus PCR.
      AnyTo achieve optimal recovery of Aspergillus from BAL by centrifugation and investigation of the sedimentCentrifugation of BALs or bronchial aspiratesAIIIEssential investigation

      Isolation of Aspergillus dependent on volume cultured
      • Fraczek M.G.
      • Kirwan M.B.
      • Moore C.B.
      • Morris J.
      • Denning D.W.
      • Richardson M.D.
      Volume dependency for culture of fungi from respiratory secretions and increased sensitivity of Aspergillus quantitative PCR.
      Abbreviations: BAL, bronchoalveolar lavage; PCR, polymerase chain reaction; QoE, Quality of evidence; SoR, Strength of recommendation.
      Table 5Culture and Aspergillus species identification
      PopulationIntentionInterventionSoRQoECommentRef.
      AnyPrimary isolation from deep sites samples (e.g. biopsies, blood, CSF)Culture on SDA, BHI agar, PDA at 30°C and 37°C for 72 hAIIIBlood inhibits conidiation; BHI can help to recover some isolates; isolation of several colonies or isolation of the same fungus from a repeat specimen enhance significance
      • Fraczek M.G.
      • Kirwan M.B.
      • Moore C.B.
      • Morris J.
      • Denning D.W.
      • Richardson M.D.
      Volume dependency for culture of fungi from respiratory secretions and increased sensitivity of Aspergillus quantitative PCR.
      ,
      • Cuenca-Estrella M.
      • Bassetti M.
      • Lass-Flîrl C.
      • R†cil Z.
      • Richardson M.
      • Rogers T.R.
      Detection and investigation of invasive mould disease.
      ,
      • Richardson M.
      • Ellis M.
      Clinical and laboratory diagnosis.
      Primary isolation from non-sterile samples, e.g. sputum, respiratory aspiratesCulture on SDA, BHI agar, PDA with gentamicin plus chloramphenicol at 30°C and 37°C for 72 hAIIIHigh-volume sputum culture (entire sample) shown to significantly increase recovery; quantitative cultures are not discriminative for infection or colonization
      Identification of species complexMacroscopic and microscopic examination from primary culturesAIIColony colour, conidium size, shape and septation. Colour of conidia and conidiophore and conidiogenesis (tease or tape mounts are preferred); expertise needed for interpretation

      Thermotolerance test (growth at 50°C for species confirmation of A. fumigatus)
      Identification of species complex (and species identification of A. fumigatus specifically)Culture on identification media at 25–30°C, 37°C and 50°C (2% MEA and Czapek-Dox Agar) and microscopic examinationAII
      Identification at species levelMALDI-TOF MS identificationBIIIn-house databases are often used to improve identification rates
      • Alanio A.
      • Beretti J.L.
      • Dauphin B.
      • Mellado E.
      • Quesne G.
      • Lacroix C.
      • et al.
      Matrix-assisted laser desorption ionization time-of-flight mass spectrometry for fast and accurate identification of clinically relevant Aspergillus species.
      ,
      • Bille E.
      • Dauphin B.
      • Leto J.
      • Bougnoux M.E.
      • Beretti J.L.
      • Lotz A.
      • et al.
      Maldi-tof ms andromas strategy for the routine identification of bacteria, mycobacteria, yeasts, Aspergillus spp. and positive blood cultures.
      ,
      • De Carolis E.
      • Vella A.
      • Florio A.R.
      • Posteraro P.
      • Perlin D.S.
      • Sanguinetti M.
      • et al.
      Use of matrix-assisted laser desorption ionization-time of flight mass spectrometry for caspofungin susceptibility testing of Candida and Aspergillus species.
      ,
      • Lau A.F.
      • Drake S.K.
      • Calhoun L.B.
      • Henderson C.M.
      • Zelazny A.M.
      Development of a clinically comprehensive database and a simple procedure for identification of molds from solid media by matrix-assisted laser desorption ionization-time of flight mass spectrometry.
      Identification at species levelSequencing of ITS, β-tubulin and calmodulinAIIINot necessary in organisms with typical growth, but in cases of atypical growth
      • Balajee S.A.
      • Borman A.M.
      • Brandt M.E.
      • Cano J.
      • Cuenca-Estrella M.
      • Dannaoui E.
      • et al.
      Sequence-based identification of Aspergillus, Fusarium, and Mucorales species in the clinical mycology laboratory: where are we and where should we go from here?.
      ,
      • Samson R.A.
      • Hong S.
      • Peterson S.W.
      • Frisvad J.C.
      • Varga J.
      Polyphasic taxonomy of Aspergillus section fumigati and its teleomorph neosartorya.
      To study outbreaksMicrosatellite and CSP analysisCIITo study outbreaks (which in general may comprise more than one genotype)
      • Caramalho R.
      • Gusmão L.
      • Lackner M.
      • Amorim A.
      • Araujo R.
      Snapafu: a novel single nucleotide polymorphism multiplex assay for Aspergillus fumigatus direct detection, identification and genotyping in clinical specimens.
      ,
      • Hurst S.F.
      • Kidd S.E.
      • Morrissey C.O.
      • Snelders E.
      • Melchers W.J.
      • Castelli M.V.
      • et al.
      Interlaboratory reproducibility of a single-locus sequence-based method for strain typing of Aspergillus fumigatus.
      ,
      • Guinea J.
      • García de Viedma D.
      • Peláez T.
      • Escribano P.
      • Munoz P.
      • Meis J.F.
      • et al.
      Molecular epidemiology of Aspergillus fumigatus: an in-depth genotypic analysis of isolates involved in an outbreak of invasive aspergillosis.
      BIITo study colonization patterns
      • Rougeron A.
      • Giraud S.
      • Razafimandimby B.
      • Meis J.F.
      • Bouchara J.P.
      • Klaassen C.H.
      Different colonization patterns of Aspergillus terreus in patients with cystic fibrosis.
      Abbreviations: BHI, brain–heart infusion; CSF, cerebrospinal fluid; CSP, cell surface protein; ITS, internal transcribed spacer; MALDI-TOF MS, matrix-assisted laser desorption/ionization time-of-flight mass spectometry identification; MEA, malt extract agar; PDA, potato dextrose agar; QoE, Quality of evidence; SDA, Sabouraud dextrose agar; SoR, Strength of recommendation.

       Non-culture based assays

      Galactomannan (GM) detection in fluids (especially BAL) is more sensitive than culture for diagnosis of IA. GM is reported as optical density index (ODI). In serum samples an ODI cut-off of 0.5 results in high sensitivity in haematological patients in the absence of mould-active prophylaxis (AI) (Table 6). Serial screening for serum GM in prolonged neutropenia and in allogeneic stem cell transplantation recipients during the early engraftment phase has a high sensitivity and negative predictive value for IA (AII) [
      • Maertens J.A.
      • Klont R.
      • Masson C.
      • Theunissen K.
      • Meersseman W.
      • Lagrou K.
      • et al.
      Optimization of the cutoff value for the Aspergillus double-sandwich enzyme immunoassay.
      ]. Serial screening is not recommended in patients on mould-active prophylaxis [
      • Duarte R.F.
      • Sanchez-Ortega I.
      • Cuesta I.
      • Arnan M.
      • Patino B.
      • Fernandez de Sevilla A.
      • et al.
      Serum galactomannan-based early detection of invasive aspergillosis in hematology patients receiving effective antimold prophylaxis.
      ].
      Table 6Galactomannan testing in blood samples
      PopulationIntentionInterventionSoRQoECommentRef.
      Patients with prolonged neutropenia or allogeneic stem cell transplantation recipients not on mould-active prophylaxisProspective screening for IAGM in blood
      a Serum or plasma.
      AIHighest test accuracy requiring two consecutive samples with an ODI ≥0.5 or retesting the same sample

      Prospective monitoring should be combined with HRCT and clinical evaluation
      • Maertens J.A.
      • Klont R.
      • Masson C.
      • Theunissen K.
      • Meersseman W.
      • Lagrou K.
      • et al.
      Optimization of the cutoff value for the Aspergillus double-sandwich enzyme immunoassay.
      ,
      • Springer J.
      • Morton C.O.
      • Perry M.
      • Heinz W.J.
      • Paholcsek M.
      • Alzheimer M.
      • et al.
      Multicenter comparison of serum and whole-blood specimens for detection of aspergillus DNA in high-risk hematological patients.
      ,
      • Maertens J.
      • Van Eldere J.
      • Verhaegen J.
      • Verbeken E.
      • Verschakelen J.
      • Boogaerts M.
      Use of circulating galactomannan screening for early diagnosis of invasive aspergillosis in allogeneic stem cell transplant recipients.
      ,
      • Leeflang M.M.
      • Debets-Ossenkopp Y.J.
      • Visser C.E.
      • Scholten R.J.
      • Hooft L.
      • Bijlmer H.A.
      • et al.
      Galactomannan detection for invasive aspergillosis in immunocompromised patients.
      ,
      • Furfaro E.
      • Mikulska M.
      • Miletich F.
      • Viscoli C.
      Galactomannan: testing the same sample twice?.
      ,
      • Morrissey C.O.
      • Chen S.C.
      • Sorrell T.C.
      • Milliken S.
      • Bardy P.G.
      • Bradstock K.F.
      • et al.
      Galactomannan and PCR versus culture and histology for directing use of antifungal treatment for invasive aspergillosis in high-risk haematology patients: a randomised controlled trial.
      ,
      • Maertens J.
      • Verhaegen J.
      • Lagrou K.
      • Van Eldere J.
      • Boogaerts M.
      Screening for circulating galactomannan as a noninvasive diagnostic tool for invasive aspergillosis in prolonged neutropenic patients and stem cell transplantation recipients: a prospective validation.
      Draw samples every 3–4 daysCIII
      Patients with prolonged neutropenic or allogeneic stem cell transplantation recipients on mould active prophylaxisProspective screening for IAGM in blood
      a Serum or plasma.
      DIILow prevalence of IA in this setting with consequently low PPV of blood GM test

      Prophylaxis may have a negative impact on sensitivity of the test or the low yield may be due to decreased incidence of IA
      • Hoenigl M.
      • Seeber K.
      • Koidl C.
      • Buzina W.
      • Wîlfler A.
      • Duettmann W.
      • et al.
      Sensitivity of galactomannan enzyme immunoassay for diagnosing breakthrough invasive aspergillosis under antifungal prophylaxis and empirical therapy.
      ,
      • Marr K.A.
      • Laverdiere M.
      • Gugel A.
      • Leisenring W.
      Antifungal therapy decreases sensitivity of the Aspergillus galactomannan enzyme immunoassay.
      Patients with a haematological malignancyTo diagnose IAGM in blood
      a Serum or plasma.
      Significantly lower sensitivity in non-neutropenic patients
      • Pfeiffer C.D.
      • Fine J.P.
      • Safdar N.
      Diagnosis of invasive aspergillosis using a galactomannan assay: a meta-analysis.
      ,
      • Leeflang M.M.
      • Debets-Ossenkopp Y.J.
      • Visser C.E.
      • Scholten R.J.
      • Hooft L.
      • Bijlmer H.A.
      • et al.
      Galactomannan detection for invasive aspergillosis in immunocompromised patients.
      ,
      • Cordonnier C.
      • Botterel F.
      • Ben Amor R.
      • Pautas C.
      • Maury S.
      • Kuentz M.
      • et al.
      Correlation between galactomannan antigen levels in serum and neutrophil counts in haematological patients with invasive aspergillosis.
      ,
      • Maertens J.
      • Theunissen K.
      • Verhoef G.
      • Verschakelen J.
      • Lagrou K.
      • Verbeken E.
      • et al.
      Galactomannan and computed tomography-based preemptive antifungal therapy in neutropenic patients at high risk for invasive fungal infection: a prospective feasibility study.
      • Neutropenic patients
      AII
      • Non-neutropenic patients
      BII
      ICU patientsTo diagnose IAGM in blood
      a Serum or plasma.
      CIIBetter performance in neutropenic than in non-neutropenic patients
      • Meersseman W.
      • Lagrou K.
      • Maertens J.
      • Wilmer A.
      • Hermans G.
      • Vanderschueren S.
      • et al.
      Galactomannan in bronchoalveolar lavage fluid: a tool for diagnosing aspergillosis in intensive care unit patients.
      ,
      • Guinea J.
      • Jensen J.
      • Peláez T.
      • Gijón P.
      • Alonso R.
      • Rivera M.
      • et al.
      Value of a single galactomannan determination (platelia) for the diagnosis of invasive aspergillosis in non-hematological patients with clinical isolation of Aspergillus spp.
      Solid organ recipientsTo diagnose IAGM in blood
      a Serum or plasma.
      CIILow sensitivity, good specificity

      Most data for lung SOT
      • Pfeiffer C.D.
      • Fine J.P.
      • Safdar N.
      Diagnosis of invasive aspergillosis using a galactomannan assay: a meta-analysis.
      ,
      • Husain S.
      • Kwak E.J.
      • Obman A.
      • Wagener M.M.
      • Kusne S.
      • Stout J.E.
      • et al.
      Prospective assessment of platelia Aspergillus galactomannan antigen for the diagnosis of invasive aspergillosis in lung transplant recipients.
      ,
      • Tabarsi P.
      • Soraghi A.
      • Marjani M.
      • Zandian P.
      • Baghaei P.
      • Najafizadeh K.
      • et al.
      Comparison of serum and bronchoalveolar lavage galactomannan in diagnosing invasive aspergillosis in solid-organ transplant recipients.
      Any other patientTo diagnose IAGM in blood
      a Serum or plasma.
      CIIPiperacillin/tazobactam may no longer be responsible for false-positive results according to recent studies

      Cross-reactivity in case of histoplasmosis, fusariosis, talaromycosis (formerly: penicilliosis)

      False-positive results reported due to ingestion of ice-pops, transfusions, antibiotics, Plasmalyt® infusion
      • Maertens J.
      • Theunissen K.
      • Verhoef G.
      • Verschakelen J.
      • Lagrou K.
      • Verbeken E.
      • et al.
      Galactomannan and computed tomography-based preemptive antifungal therapy in neutropenic patients at high risk for invasive fungal infection: a prospective feasibility study.
      ,
      • Guigue N.
      • Menotti J.
      • Ribaud P.
      False positive galactomannan test after ice-pop ingestion.
      ,
      • Petraitiene R.
      • Petraitis V.
      • Witt 3rd, J.R.
      • Durkin M.M.
      • Bacher J.D.
      • Wheat L.J.
      • et al.
      Galactomannan antigenemia after infusion of gluconate-containing plasma-lyte.
      ,
      • Martin-Rabadan P.
      • Gijon P.
      • Alonso Fernandez R.
      • Ballesteros M.
      • Anguita J.
      • Bouza E.
      False-positive aspergillus antigenemia due to blood product conditioning fluids.
      ,
      • Mikulska M.
      • Furfaro E.
      • Del Bono V.
      • Raiola A.M.
      • Ratto S.
      • Bacigalupo A.
      • et al.
      Piperacillin/tazobactam (tazocintm) seems to be no longer responsible for false-positive results of the galactomannan assay.
      ,
      • Vergidis P.
      • Walker R.C.
      • Kaul D.R.
      • Kauffman C.A.
      • Freifeld A.G.
      • Slagle D.C.
      • et al.
      False-positive Aspergillus galactomannan assay in solid organ transplant recipients with histoplasmosis.
      ,
      • Huang Y.T.
      • Hung C.C.
      • Liao C.H.
      • Sun H.Y.
      • Chang S.C.
      • Chen Y.C.
      Detection of circulating galactomannan in serum samples for diagnosis of Penicillium marneffei infection and cryptococcosis among patients infected with human immunodeficiency virus.
      ,
      • Nucci M.
      • Carlesse F.
      • Cappellano P.
      • Varon A.G.
      • Seber A.
      • Garnica M.
      • et al.
      Earlier diagnosis of invasive fusariosis with Aspergillus serum galactomannan testing.
      ,
      • King S.T.
      • Stover K.R.
      Considering confounders of the galactomannan index: the role of piperacillin-tazobactam.
      Cancer patientsTo monitor treatmentGM in blood
      a Serum or plasma.
      AII
      • Nouér S.A.
      • Nucci M.
      • Kumar N.S.
      • Grazziutti M.
      • Barlogie B.
      • Anaissie E.
      Earlier response assessment in invasive aspergillosis based on the kinetics of serum Aspergillus galactomannan: proposal for a new definition.
      ,
      • Bergeron A.
      • Porcher R.
      • Menotti J.
      • Poirot J.L.
      • Chagnon K.
      • Vekhoff A.
      • et al.
      Prospective evaluation of clinical and biological markers to predict the outcome of invasive pulmonary aspergillosis in hematological patients.
      ,
      • Chai L.Y.
      • Kullberg B.J.
      • Johnson E.M.
      • Teerenstra S.
      • Khin L.W.
      • Vonk A.G.
      • et al.
      Early serum galactomannan trend as a predictor of outcome of invasive aspergillosis.
      Abbreviations: GM, galactomannan; IA, invasive aspergillosis; ICU, intensive care unit; ODI, optical density index; PPV, positive predictive value; QoE, Quality of evidence; SoR, Strength of recommendation; SOT, solid organ transplantation.
      a Serum or plasma.
      Sensitivity of serum GM testing is significantly lower in non-neutropenic versus neutropenic patients [
      • Teering S.
      • Verreth A.
      • Peeters A.
      • Van Regenmortel N.
      • De Laet I.
      • Schoonheydt K.
      • et al.
      Prognostic value of serum galactomannan in mixed ICU patients: a retrospective observational study.
      ]. Decrease of the ODI during the first 2 weeks of antifungal therapy is a reliable predictor of a satisfactory response in cancer patients [
      • Nouér S.A.
      • Nucci M.
      • Kumar N.S.
      • Grazziutti M.
      • Barlogie B.
      • Anaissie E.
      Earlier response assessment in invasive aspergillosis based on the kinetics of serum Aspergillus galactomannan: proposal for a new definition.
      ]. GM detection in BAL specimens has an excellent performance with evidence that ODI of 0.5–1.0 has decreased predictive values compared with results of >1.0 [
      • D'Haese J.
      • Theunissen K.
      • Vermeulen E.
      • Schoemans H.
      • De Vlieger G.
      • Lammerijn L.
      • et al.
      Detection of galactomannan in bronchoalveolar lavage fluid samples of patients at risk for invasive pulmonary aspergillosis: analytical and clinical validity.
      ] (AII) (Table 7). The test also has diagnostic value in patients undergoing lung transplantation or who are in intensive care [
      • Pasqualotto A.C.
      • Xavier M.O.
      • Sanchez L.B.
      • de Oliveira Costa C.D.
      • Schio S.M.
      • Camargo S.M.
      • et al.
      Diagnosis of invasive aspergillosis in lung transplant recipients by detection of galactomannan in the bronchoalveolar lavage fluid.
      ,
      • Luong M.L.
      • Clancy C.J.
      • Vadnerkar A.
      • Kwak E.J.
      • Silveira F.P.
      • Wissel M.C.
      • et al.
      Comparison of an aspergillus real-time polymerase chain reaction assay with galactomannan testing of bronchoalvelolar lavage fluid for the diagnosis of invasive pulmonary aspergillosis in lung transplant recipients.
      ,
      • Meersseman W.
      • Lagrou K.
      • Maertens J.
      • Wilmer A.
      • Hermans G.
      • Vanderschueren S.
      • et al.
      Galactomannan in bronchoalveolar lavage fluid: a tool for diagnosing aspergillosis in intensive care unit patients.
      ]; a sensitivity of 100% and a specificity of 90.4% was defined at cut-off of 1.5 [
      • Pasqualotto A.C.
      • Xavier M.O.
      • Sanchez L.B.
      • de Oliveira Costa C.D.
      • Schio S.M.
      • Camargo S.M.
      • et al.
      Diagnosis of invasive aspergillosis in lung transplant recipients by detection of galactomannan in the bronchoalveolar lavage fluid.
      ].
      Table 7Galactomannan testing in samples other than blood
      PopulationIntentionInterventionSoRQoECommentRef.
      AnyTo diagnose pulmonary IATo apply GM test on BAL fluidAIIGM in BAL is a good tool to diagnose, optimal cut-off to positivity 0.5 to 1.0
      • D'Haese J.
      • Theunissen K.
      • Vermeulen E.
      • Schoemans H.
      • De Vlieger G.
      • Lammerijn L.
      • et al.
      Detection of galactomannan in bronchoalveolar lavage fluid samples of patients at risk for invasive pulmonary aspergillosis: analytical and clinical validity.
      ,
      • Luong M.L.
      • Clancy C.J.
      • Vadnerkar A.
      • Kwak E.J.
      • Silveira F.P.
      • Wissel M.C.
      • et al.
      Comparison of an aspergillus real-time polymerase chain reaction assay with galactomannan testing of bronchoalvelolar lavage fluid for the diagnosis of invasive pulmonary aspergillosis in lung transplant recipients.
      ,
      • Reinwald M.
      • Spiess B.
      • Heinz W.J.
      • Vehreschild J.J.
      • Lass-Flîrl C.
      • Kiehl M.
      • et al.
      Diagnosing pulmonary aspergillosis in patients with hematological malignancies: a multicenter prospective evaluation of an Aspergillus PCR assay and a galactomannan elisa in bronchoalveolar lavage sample.
      ,
      • Heng S.C.
      • Morrissey O.
      • Chen S.C.
      • Thursky K.
      • Manser R.L.
      • Nation R.L.
      • et al.
      Utility of bronchoalveolar lavage fluid galactomannan alone or in combination with PCR for the diagnosis of invasive aspergillosis in adult hematology patients: a systematic review and meta-analysis.
      ,
      • Zou M.
      • Tang L.
      • Zhao S.
      • Zhao Z.
      • Chen L.
      • Chen P.
      • et al.
      Systematic review and meta-analysis of detecting galactomannan in bronchoalveolar lavage fluid for diagnosing invasive aspergillosis.
      ,
      • Fisher C.E.
      • Stevens A.M.
      • Leisenring W.
      • Pergam S.A.
      • Boeckh M.
      • Hohl T.M.
      The serum galactomannan index predicts mortality in hematopoietic stem cell transplant recipients with invasive aspergillosis.
      AnyTo diagnose cerebral IATo apply GM test on cerebrospinal fluidBIINo validated cut-off
      • Verweij P.E.
      • Brinkman K.
      • Kremer H.P.
      • Kullberg B.J.
      • Meis J.F.
      Aspergillus meningitis: diagnosis by non-culture-based microbiological methods and management.
      ,
      • Viscoli C.
      • Machetti M.
      • Gazzola P.
      • De Maria A.
      • Paola D.
      • Van Lint M.T.
      • et al.
      Aspergillus galactomannan antigen in the cerebrospinal fluid of bone marrow transplant recipients with probable cerebral aspergillosis.
      AnyTo detect GM in tissueTo apply GM test on lung biopsiesBIIUsing a cut-off 0.5 resulted in a sensitivity of 90 % and a specificity of 95%; specimens need to be sliced, precondition for doing so is that sufficient material is available; dilution in isotonic saline
      • Lass-Florl C.
      • Resch G.
      • Nachbaur D.
      • Mayr A.
      • Gastl G.
      • Auberger J.
      • et al.
      The value of computed tomography-guided percutaneous lung biopsy for diagnosis of invasive fungal infection in immunocompromised patients.
      ,
      • Klont R.R.
      • Mennink-Kersten M.A.
      • Verweij P.E.
      Utility of Aspergillus antigen detection in specimens other than serum specimens.
      Abbreviations: BAL, bronchoalveolar lavage; GM, galactomannan; IA, invasive aspergillosis; QoE, Quality of evidence; SoR, Strength of recommendation.
      A constituent of the cell wall of many species and genera of fungi, (1-3)-β-d-glucan (BDG), is released into body fluids in association with fungal infection. A limited role is given for the exclusive testing of the BDG in diagnosing IA (BII) (Table 8); however, the combination with GM or PCR improves specific detection [
      • Karageorgopoulos D.E.
      • Vouloumanou E.K.
      • Ntziora F.
      • Michalopoulos A.
      • Rafailidis P.I.
      • Falagas M.E.
      β-d-glucan assay for the diagnosis of invasive fungal infections: a meta-analysis.
      ].
      Table 8β-d-glucan assays
      PopulationIntentionInterventionSoRQoECommentRef.
      Mixed population: adult ICU, haematological disorders, SOTTo diagnose IFDDiagnostic assayCIIFive different assays

      Overall sensitivity of 77% and specificity of 85%

      Specificity limits its value in this setting
      • Karageorgopoulos D.E.
      • Vouloumanou E.K.
      • Ntziora F.
      • Michalopoulos A.
      • Rafailidis P.I.
      • Falagas M.E.
      β-d-glucan assay for the diagnosis of invasive fungal infections: a meta-analysis.
      ,
      • Lu Y.
      • Chen Y.Q.
      • Guo Y.L.
      • Qin S.M.
      • Wu C.
      • Wang K.
      Diagnosis of invasive fungal disease using serum (1-3)- β-d-glucan: a bivariate meta-analysis.
      Screening assaysCIITwo or more consecutive samples: sensitivity: 65%; specificity: 93%

      Studies included once to thrice weekly. Varies with assay and cut-off: Wako assay sensitivity: 40%–97%, specificity: 51%–99%
      • Karageorgopoulos D.E.
      • Vouloumanou E.K.
      • Ntziora F.
      • Michalopoulos A.
      • Rafailidis P.I.
      • Falagas M.E.
      β-d-glucan assay for the diagnosis of invasive fungal infections: a meta-analysis.
      ,
      • Lu Y.
      • Chen Y.Q.
      • Guo Y.L.
      • Qin S.M.
      • Wu C.
      • Wang K.
      Diagnosis of invasive fungal disease using serum (1-3)- β-d-glucan: a bivariate meta-analysis.
      Adult haematological malignancy and HSCTTo diagnose IFDDiagnostic assayCIIOverall sensitivity: 50%–70%, specificity: 91%–99%
      • Liss B.
      • Cornely O.A.
      • Hoffmann D.
      • Dimitriou V.
      • Wisplinghoff H.
      1,3-β-d-glucan contamination of common antimicrobials-authors' response.
      ,
      • Liss B.
      • Cornely O.A.
      • Hoffmann D.
      • Dimitriou V.
      • Wisplinghoff H.
      1,3-β-d-glucan contamination of common antimicrobials.
      ,
      • Liss B.
      • Cornely O.A.
      • Hoffmann D.
      • Dimitriou V.
      • Wisplinghoff H.
      1,3-β-d-glucan concentrations in blood products predict false positive post-transfusion results.
      ,
      • Lamoth F.
      • Cruciani M.
      • Mengoli C.
      • Castagnola E.
      • Lortholary O.
      • Richardson M.
      • et al.
      β-glucan antigenemia assay for the diagnosis of invasive fungal infections in patients with hematological malignancies: a systematic review and meta-analysis of cohort studies from the third European Conference on Infections in Leukemia (ECIL-3).
      ,
      • Senn L.
      • Robinson J.O.
      • Schmidt S.
      • Knaup M.
      • Asahi N.
      • Satomura S.
      • et al.
      1,3-β-d-glucan antigenemia for early diagnosis of invasive fungal infections in neutropenic patients with acute leukemia.
      ,
      • Ellis M.
      • Al-Ramadi B.
      • Finkelman M.
      • Hedstrom U.
      • Kristensen J.
      • Ali-Zadeh H.
      • et al.
      Assessment of the clinical utility of serial β-d-glucan concentrations in patients with persistent neutropenic fever.
      ,
      • Kawazu M.
      • Kanda Y.
      • Nannya Y.
      • Aoki K.
      • Kurokawa M.
      • Chiba S.
      • et al.
      Prospective comparison of the diagnostic potential of real-time PCR, double-sandwich enzyme-linked immunosorbent assay for galactomannan, and a (1→3)- β-d-glucan test in weekly screening for invasive aspergillosis in patients with hematological disorders.
      ,
      • Odabasi Z.
      • Mattiuzzi G.
      • Estey E.
      • Kantarjian H.
      • Saeki F.
      • Ridge R.J.
      • et al.
      β-d-glucan as a diagnostic adjunct for invasive fungal infections: validation, cutoff development, and performance in patients with acute myelogenous leukemia and myelodysplastic syndrome.
      ,
      • Ostrosky-Zeichner L.
      • Alexander B.D.
      • Kett D.H.
      • Vazquez J.
      • Pappas P.G.
      • Saeki F.
      • et al.
      Multicenter clinical evaluation of the (1→3) β-d-glucan assay as an aid to diagnosis of fungal infections in humans.
      ICU—mixed adult immunocompromised patients (haematology, SOT, cancer, immunosuppressive therapy, liver failure, HIV)To diagnose IADiagnostic assayCIIOverall sensitivity: 78%–85%, specificity: 36%–75%, NPV: 85%–92%

      Specificity increased at higher cut-off values
      • De Vlieger G.
      • Lagrou K.
      • Maertens J.
      • Verbeken E.
      • Meersseman W.
      • Van Wijngaerden E.
      β-d-glucan detection as a diagnostic test for invasive aspergillosis in immunocompromised critically ill patients with symptoms of respiratory infection: an autopsy-based study.
      ,
      • Del Bono V.
      • Delfino E.
      • Furfaro E.
      • Mikulska M.
      • Nicco E.
      • Bruzzi P.
      • et al.
      Clinical performance of the (1,3)- β-d-glucan assay in early diagnosis of nosocomial Candida bloodstream infections.
      ICU—mixed adult population: SOT, liver failure, immunosuppressedScreening assaysCIIISensitivity: 91%, specificity: 58%, PPV: 25%, NPV: 98%.

      Positive mean of 5.6 days before positive mould culture

      High false-positive rate in early ICU admission
      • Acosta J.
      • Catalan M.
      • del Palacio-Pérez-Medel A.
      • Montejo J.C.
      • De-La-Cruz-Bértolo J.
      • Moraques M.D.
      • et al.
      Prospective study in critically ill non-neutropenic patients: diagnostic potential of (1,3)- β-d-glucan assay and circulating galactomannan for the diagnosis of invasive fungal disease.
      Adult haematological malignancy and HSCTTo diagnose IADiagnostic assayCIIOverall sensitivity: 57%–76%, specificity: 95%–97%
      • Lu Y.
      • Chen Y.Q.
      • Guo Y.L.
      • Qin S.M.
      • Wu C.
      • Wang K.
      Diagnosis of invasive fungal disease using serum (1-3)- β-d-glucan: a bivariate meta-analysis.
      ,
      • Lamoth F.
      • Cruciani M.
      • Mengoli C.
      • Castagnola E.
      • Lortholary O.
      • Richardson M.
      • et al.
      β-glucan antigenemia assay for the diagnosis of invasive fungal infections in patients with hematological malignancies: a systematic review and meta-analysis of cohort studies from the third European Conference on Infections in Leukemia (ECIL-3).
      ,
      • De Vlieger G.
      • Lagrou K.
      • Maertens J.
      • Verbeken E.
      • Meersseman W.
      • Van Wijngaerden E.
      β-d-glucan detection as a diagnostic test for invasive aspergillosis in immunocompromised critically ill patients with symptoms of respiratory infection: an autopsy-based study.
      Screening assaysCIIOverall sensitivity: 46%, specificity: 97%

      Confirmation with GM increases specificity

      Data suggest BDG is unsuitable for ruling out diagnosis of IA
      Abbreviations: BDG, β-d-glucan test; GM, galactomannan; HSCT, haematopoietic stem cell transplantation; IA, invasive aspergillosis; ICU, intensive care unit; IFD, invasive fungal disease; NPV, negative predictive value; PPV, positive predictive value; QoE, Quality of evidence; SoR, Strength of recommendation; SOT, solid organ transplantation.
      The Aspergillus lateral flow device assay can be performed on serum and on BAL samples, but at the time of writing this assay is not commercially available [
      • Prattes J.
      • Lackner M.
      • Eigl S.
      • Reischies F.
      • Raggam R.B.
      • Koidl C.
      • et al.
      Diagnostic accuracy of the aspergillus-specific bronchoalveolar lavage lateral-flow assay in haematological malignancy patients.
      ] (Table 9).
      Table 9Lateral flow device antigen test for invasive aspergillosis
      PopulationIntentionInterventionSoRQoECommentRef.
      Haematological malignancy and solid organ transplantTo diagnose IALFD applied on BAL samplesBIIRetrospective study. Sensitivity and specificity of BAL LFD tests for probable IPA were 100% and 81% (PPV 71%, NPV 100%), five patients with possible IPA had positive LFD, no proven IA
      • Hoenigl M.
      • Koidl C.
      • Duettmann W.
      • Seeber K.
      • Wagner J.
      • Buzina W.
      • et al.
      Bronchoalveolar lavage lateral-flow device test for invasive pulmonary aspergillosis diagnosis in haematological malignancy and solid organ transplant patients.
      Haematopoietic stem cell transplantationTo diagnose IALFD applied on serum samplesBIIProspective screening in 101 patients undergoing allogeneic HSCT
      • Held J.
      • Schmidt T.
      • Thornton C.R.
      • Kotter E.
      • Bertz H.
      Comparison of a novel Aspergillus lateral-flow device and the platelia© galactomannan assay for the diagnosis of invasive aspergillosis following haematopoietic stem cell transplantation.
      Immunocompromised patientsTo diagnose IALFD applied on BAL samplesBIIRetrospective study. Sensitivities for LFD, GM, BDG and PCR were between 70% and 88%. Combined GM (cut-off >1.0 OD) with LFD increased the sensitivity to 94%, while combined GM (cut-off >1.0 OD) with PCR resulted in 100% sensitivity (specificity for probable/proven IPA 95%–98%).
      • Hoenigl M.
      • Prattes J.
      • Spiess B.
      • Wagner J.
      • Prueller F.
      • Raggam R.B.
      • et al.
      Performance of galactomannan, β-d-glucan, Aspergillus lateral-flow device, conventional culture, and PCR tests with bronchoalveolar lavage fluid for diagnosis of invasive pulmonary aspergillosis.
      Abbreviations: BAL, bronchoalveolar lavage; BDG, β-D-glucan test; GM, galactomannan; HSCT, haematopoietic stem cell transplantation; IA, invasive aspergillosis; IFD, invasive fungal diseases; LFD, lateral device flow; NPV, negative predictive value; PCR, polymerase chain reaction; PPV, positive predictive value; QoE, Quality of evidence; SoR, Strength of recommendation.
      Aspergillus PCR has been applied mostly to blood and BAL fluid. For both sample types, a combination with other biomarkers increases the likelihood of IA [
      • Boch T.
      • Reinwald M.
      • Postina P.
      • Cornely O.A.
      • Vehreschild J.J.
      • Heussel C.P.
      • et al.
      Identification of invasive fungal diseases in immunocompromised patients by combining an aspergillus specific PCR with a multifungal DNA-microarray from primary clinical samples.
      ,
      • Boch T.
      • Spiess B.
      • Cornely O.A.
      • Vehreschild J.J.
      • Rath P.M.
      • Steinmann J.
      • et al.
      Diagnosis of invasive fungal infections in haematological patients by combined use of galactomannan, 1,3-β-d-glucan, aspergillus PCR, multifungal DNA-microarray, and aspergillus azole resistance PCRs in blood and bronchoalveolar lavage samples: results of a prospective multicentre study.
      ]. The performance of serum PCR is not significantly different from that of whole blood [
      • Springer J.
      • Morton C.O.
      • Perry M.
      • Heinz W.J.
      • Paholcsek M.
      • Alzheimer M.
      • et al.
      Multicenter comparison of serum and whole-blood specimens for detection of aspergillus DNA in high-risk hematological patients.
      ,
      • Springer J.
      • White P.L.
      • Hamilton S.
      • Michel D.
      • Barnes R.A.
      • Einsele H.
      • et al.
      Comparison of performance characteristics of aspergillus PCR in testing a range of blood-based samples in accordance with international methodological recommendations.
      ,
      • White P.L.
      • Wiederhold N.P.
      • Loeffler J.
      • Najvar L.K.
      • Melchers W.
      • Herrera M.
      • et al.
      Comparison of nonculture blood-based tests for diagnosing invasive aspergillosis in an animal model.
      ,
      • White P.L.
      • Barnes R.A.
      • Springer J.
      • Klingspor L.
      • Cuenca-Estrella M.
      • Morton C.O.
      • et al.
      Clinical performance of aspergillus PCR for testing serum and plasma: a study by the European aspergillus PCR initiative.
      ]. Prospective screening of high-risk haematological patients by a combination of GM and PCR improves the diagnostic accuracy and is associated with an earlier diagnosis [
      • Rogers T.R.
      • Morton C.O.
      • Springer J.
      • Conneally E.
      • Heinz W.
      • Kenny C.
      • et al.
      Combined real-time PCR and galactomannan surveillance improves diagnosis of invasive aspergillosis in high risk patients with haematological malignancies.
      ,