Interaction of Aspergillus with the host A unique microbial-host interaction Immune dysfunction Frequency of aspergillosis Immune hyperactivity Frequency of aspergillosis Acute IA Subacute IA Aspergilloma Chronic pulmonary ABPA Severe asthma with fungal sensitisation Allergic sinusitis. After Casadevall & Pirofski, Infect Immun 1999;67:3703

Size of fungal disease problem globally 1.Invasive aspergillosis - ? 70,000 cases/year in EU, >5M at risk; new problems COPD, ICU etc - ~50% mortality 2.Candidaemia in UK – 2,000 cases, rising, many more at risk, ~40% mortality 3.Cryptococcal meningitis - ~1M worldwide annually 4.Chronic pulmonary aspergillosis after TB – 1.1M cases prevalence 5.Chronic pulmonary aspergillosis total - ~3M 6.Asthma 197M in adults, of which ~10-20% severe, UK and USA have very high prevalence rates 7.Allergic bronchopulmonary aspergillosis in asthma - ~3M worldwide (2.1% of adults with asthma) 8.Severe asthma with fungal sensitisation - ~13M worldwide (33% of 20% (severe only))

CLASSIFICATION OF ASPERGILLOSIS Persistence without disease - colonisation of the airways or nose/sinuses Airways/nasal exposure to airborne Aspergillus Invasive aspergillosis Acute (<1 month course) Subacute/chronic necrotising (1-3 months) Chronic aspergillosis (>3 months) Chronic cavitary pulmonary Aspergilloma of lung Chronic fibrosing pulmonary Chronic invasive sinusitis Maxillary (sinus) aspergilloma Allergic Allergic bronchopulmonary (ABPA) Extrinsic allergic (broncho)alveolitis (EAA) Asthma with fungal sensitisation Allergic Aspergillus sinusitis (eosinophilic fungal rhinosinusitis)

Risk factors for invasive aspergillosis Major Neutropenia (+ monocytopenia) Corticosteroid treatment Minor CD4penia Inherited immunodeficiency (ie CGD) Lung or sinus damage/disease Severe liver disease Exposure to high inocula

Examples of at-risk patients and pace of progression Degree of immunocompromise Risk of acquisition (and pace of progression) ‘Normal immunity’, high inoculum HIV infection Chronic leukaemia Short course glucocorticoids Acute respiratory infection, ie influenza Temporary neutropenia Long term glucocorticoids etc Solid organ transplant + rejection + CMV AIDS Leukemia and profound neutropenia Allogeneic stem cell transplant + GVHD Relapsed/uncontrolled leukemia 5% 10% 15% 20% 25% Medical ICU, COPD + sepsis

Where in the hospital does invasive aspergillosis occur? Cornillet et al, Clin Infect Dis 2006;43:577

Risk factors for invasive aspergillosis in AIDS Stage of AIDSCDC Group II 4 (1%) CDC Group IV 289 (72%) Neutropenia <1000 x 10 6 /L92/202 (46%) Corticosteroid therapy79/202 (39%) Prior pulmonary infection124/169 (73%) Khoo & Denning, Clin Infect Dis 1994; 19 (S1) 541

Lymphoma and corticosteroids 4 days later

CT scan showing nodules with halo – lung cancer and neutropenia

Aspergillus, IPA and COPD Guinea et al, Clin Microbiol Infect 2010;16:870 ~ 22% of Aspergillus in COPD = invasive aspergillosis

Aspergillus, IPA and COPD Guinea et al, Clin Microbiol Infect 2010;16:870

Aspergillus, IPA and COPD Guinea et al, Clin Microbiol Infect 2010;16:870 Clues to the diagnosis of IA GOLD stage 3 or 4. Excess wheezing (consider tracheobronchitis) Worsening infiltrates in an ‘exacerbation’ (66%) Bilateral infiltrates (55%) Culture of Aspergillus High corticosteroid exposure recently Do NOT expect fever (38%), chest pain or haemoptysis

Invasive aspergillosis in COPD Bulpa, Clin Infect Dis 2007;30:782

Invasive aspergillosis in ICU 127 of 1850 (6.9%) consecutive medical ICU admissions with IA or colonisation (micro/histol). 89/127 (70%) did not have haematological malignancy 67/89 proven/probable IA, 33 of 67 (50%) COPD Meersemann et al, Am J Resp Med Crit Care 2004;170:621.

Radiology completely unhelpful in suspecting the diagnosis Meersseman, Clin Infect Dis 2007;45:205

Risk factors for invasive aspergillosis in ICU Meersseman, Clin Infect Dis 2007;45:205

Invasive aspergillosis in ICU Vandewoude et al, Critical Care 2006;10:R31. Aspergillus detected, no infection N = 89 Invasive aspergillosis + treatment N = 73 Invasive aspergillosis no treatment N = 12

Invasive Aspergillosis in Children in the USA in 2000 Incidence of 437 cases per 100,000 immunocompromised children MORTALITY RATES Disease Patients without Invasive Aspergillosis (151,537 children) Patients with Invasive Aspergillosis (666 children) Relative Risk of Death Acute Leukemia (ALL) 1%21%14.9 Acute Leukemia (AML) 3%20%5.0 Brain Tumor 2%69%21.6 Bone Marrow Transplant 8%44%3.8 Zaoutis TE, et al. Pediatrics 2006;117:e711

Tait, Thorax 1993;48: 1285 Pseudomembranous Aspergillus tracheobronchitis Wheezing 4 days before death, immunocompromised

Pseudomembranous Aspergillus tracheobronchitis

Bulpa Eur Resp J 2007;30:782 Pseudomembranous Aspergillus tracheobronchitis with IPA in COPD

Myelodysplasia with clinical evidence of acute invasive fungal rhinosinusitis after chemotherapy – biopsy showed hyphal invasion of bone Pre-treatment 6 months later after initial caspofungin then voriconazole

Cultures for Aspergillus from sputum and BAL Horvath & Dummer, Am J Med 1996;100:171 Bacteriological media inferior to fungal media – 32% higher yield on fungal media Yield in IA from BAL and sputum ~30% Cultures take 1-10 days to grow + time to identification

Modalities for early diagnosis of invasive aspergillosis CT scanning Microscopy Antigen (blood or respiratory fluid) [PCR (blood or respiratory fluid)]

Pediatric Galactomannan Prospective study from –450 adult allogeneic HSCT patients (3883 samples) –347 children with hematologic malignancies (2376 samples) GM > 1.5 in at least two sequential samples AdultPediatric –Sensitivity88.6%100% –Specificity97.5%89.9% False-positive antigenemia –Adult patients 2.5% (10/406) –Pediatric patients10.1% (34/338) Sulahian Cancer 2001;91:311.

Pediatric GM in Oncology Patients St. Jude & NCI –56 pediatric oncology patients –39 pts without IA; 17 pts with proven/probable IA –Most done retrospectively (frozen samples) At least one GM positive in 11/17 patients with IA (sensitivity 65.7%) False-positive rate 1% (all ≤ 0.8) (per sample) At least one false-positive in 12.8% patients Piperacillin-tazobactam was not used in any patients in this study No association with accuracy and patient age Hayden R Pediatr Infect Dis J 2008;27:815

Unequivocal ‘Halo sign’ surrounding a nodule Herbrecht, Denning et al, NEJM 2002;347: Small vessel angioinvasion Halo

IPA

Pulmonary nodules a useful feature if invasive pulmonary aspergillosis CT features in 48 CTs of which 17 IPA IPAOther Halo13/17 0/31 Nodules 14/1711/31 Masses 6/17 2/31 Kami, Mycoses 2002;45:

Microscopy Ruchel R, Fluorescent brighteners such as Calcufluor white, Blankophor increase sensitivity and speed

Cultures for Aspergillus from sputum and BAL Horvath & Dummer, Am J Med 1996;100: Bacteriological media inferior to fungal media – 32% higher yield on fungal media Yield in IA from BAL and sputum ~30%

13/17 (76%) in acute leukaemia with CT abnormality 20/20 (100%) in haem-onc pts with IPA 37/49 (76%) in HSCT & haem-onc with IPA 6 of 11 (55%) immunocompromised (8 of 11 +ve by PCR) 5/20 (25%) in suspected IFIs Becker, Br J Haem 2003;121:448; Sanguinetti, JCM 2003;41:3922; Musher, JCM 2004;42:5517. Aspergillus Antigen in BAL 17/17 (100%) in neutropenic patients before antifungal Rx, 0% after 3d antifungal therapy

Diagnosis of IPA in leukaemia using blood PCR Barnes et al, J Clin Pathol 2009;62: haematology patients Itraconazole prophylaxis for AML and HSCT Fluconazole prophylaxis for others (ALL, lymphoma etc) EORTC/MSG criteria applied 2x weekly sampling

Diagnosis of IPA in leukaemia using blood PCR and Aspergillus antigen (EIA) Barnes et al, J Clin Pathol 2009;62:64

Chronic pulmonary aspergillosis Single fungal ball or aspergilloma in a pre- existing cavity Infection of the lung by Aspergillus

Simple (single) aspergilloma Patient RK Haempotysis, nil else Positive Aspergillus antibodies in blood Lobectomy Wythenshawe Hospital

Simple (single) aspergilloma Patient NM Positive Aspergillus antibodies in blood Lobectomy Wythenshawe Hospital August 2006 May 2009 Community acquired New cough pneumonia requiring ICU care

Chronic pulmonary aspergillosis Single fungal ball or aspergilloma in a pre- existing cavity Infection of the lung by Aspergillus Invasive aspergillosis /community acquired infection Chronic cavitary pulmonary aspergillosis +/- fungal ball Chronic fibrosing pulmonary aspergillosis +/- fungal ball

‘Multicavity’ disease is the hallmark of chronic cavitary pulmonary aspergillosis (CCPA) Wythenshawe Hospital + Aspergillus IgG antibodies (precipitins) + symptoms

Chronic cavitary pulmonary aspergillosis (CCPA) – sputum production Wythenshawe Hospital Aspergillus cultures positive in CCPA in 10-40% of cases only

Chronic cavitary pulmonary aspergillosis (CCPA) – coughing up blood (haemoptysis) Wythenshawe Hospital

Underlying diseases in patients with CPA (%) Smith, Eur Resp J 2010 In press Classical tuberculosis Atypical tuberculosis ABPA COPD/emphysema Pneumothorax Lung cancer survivor Pneumonia Sarcoidosis (stage II/III) Thoracic surgery Rheumatoid arthritis Asthma / SAFS Ankylosing spondylitis None

Frequency of chronic pulmonary aspergillosis after TB Anonymous. Tubercle 1970;51:227; Sonnenberg et al, Lancet 2001;358: % of patients with TB are left with a cavity ~10% of all cases of pulmonary TB get CPA

Global CPA cases per region related to TB 5 year prevalence estimates Denning, Pleuvry & Cole, Bull WHO 2011 in press

Chronic pulmonary aspergillosis Chronic cavitary pulmonary aspergillosis complicating ABPA Chronic cavitary pulmonary aspergillosis with bilateral aspergillomas complicating sarcoidosis

ABPA and development of CPA

Bronchoscopy in an ABPA patient on no treatment UHSM, unpublished

Bronchoscopy in an ABPA patient on no treatment UHSM, unpublished BAL Abundant mixed inflammatory cells with ciliated columnar cells and a few fungal hyphae, in keeping with Aspergillus. A few Charcot leyden crystals. No maligant cells.

Routine versus high volume culture versus real time PCR for Aspergillus Fraczek, ECCMID Abstract submitted Samplen Aspergillus positive samples (%) Aspergillus cultureMycAssay Aspergillus real time PCR RoutineHigh volume Pre-bronch sputum404 (100) Post-bronch sputum401 (25)4 (100) First trap aspiration302 (67)3 (100) First BAL (10-20mL)5004 (80) Second BAL (10-50mL5004 (80)

Molecular detection of Aspergillus spp. in sputum Denning et al. Clin Infect Dis 2011; Laboratory resultABPACPANormals Culture positive for A. fumigatus 0/19 7/42 (16.7%) 0/11 qPCR positive for Aspergillus spp 15/19 (78.9%) 30/42 (71.4%) 4/11 (36.4%)

Colonisation in ‘normal’ lungs Lass-Florl et al, Br J Haematol 1999;104: of 30 (73%) grew a fungus in both lung samples taken 10/30 (33%) grew >1 species

Antifungal therapy IDSA guidelines. Walsh et al. Clin Infect Dis 2008;46:327

Time to initial response with posaconazole therapy 6 months 12 months Mean 95% confidence interval Felton et al. Clin Infect Dis In press.