The extraordinary spectrum of diseases caused by Aspergillus David W. Denning Wythenshawe Hospital...

The extraordinary spectrum of diseases caused by Aspergillus

David W. DenningWythenshawe Hospital

University of Manchester

The genus The genus AspergillusAspergillus - importance to - importance to humanityhumanity

www.aspergillus.org.uk

cause invasive and allergic diseasein humans and other animals:

A. fumigatus

cause plant and food spoilage and produce mycotoxins:

A. flavus and A. parasiticus

on the negative side:on the negative side:

The genus The genus AspergillusAspergillus - importance to - importance to humanityhumanity


on the positive side:on the positive side:

composting

well-established model organism in cell biology and genetics:A. nidulans

food production:enzymes and organic acids: A. niger East Asian foods: A. oryzae and A. sojae

pharmaceuticals:echinocandins: A. nidulans and A. sydowilovastatin: A. terreusfumagillin: A. fumigatus

Spores inhaled Germination

Mass of hyphae (plateau phase)

Hyphal elongation and branching

Aspergillus Life-cycle


A. nidulans – may be amphotericin B resistant

The genus The genus AspergillusAspergillus – – ~180 species, ~180 species,

38 have caused disease (able to grow at 37C)38 have caused disease (able to grow at 37C)

Common in the environmentCommon in the environment

A. nigerA. terreus – resistant to AmBA. flavus -sometimes amphotericin B resistant


A. fumigatus low frequency of azole resistance

Aspergillus fumigatus

conidial head

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)

Immunosuppression and infection

• Inhalation of aspergillus spores is a common daily occurrence. A healthy immune system would normally remove the spores and no symptoms or infection would occur.

• In individuals whose immune system may be suppressed either because of illness eg AIDS, cancer patients or drugs, spores may germinate and resulting tissue or systemic aspergillus invasion can result.

• Individuals with allergies such as asthma, can also be vulnerable to aspergillus disease.

Interaction of Interaction of AspergillusAspergillus with the host with the host

A unique microbial-host interactionA unique microbial-host interaction

Immune dysfunction

Frequency

of a

sperg

illosis

Immune hyperactivity

Frequency

of

asp

erg

illosi

s

Acute IA

Subacute IA

Tracheobronchitis AspergillomaChronic cavitaryChronic fibrosing

ABPAAllergic sinusitis

. www.aspergillus.org.uk

Normal immune function

Changing incidence of fatal invasive mycoses in non-HIV

patients in USA

Rate

per

100,0

00 p

opula

tion

0.0

0.2

0.4

0.6

0.8

1981 1986 1991 1996

CandidiasisAspergillosis

McNeil et al, Clin Infect Dis 2001;33:641

Invasive pulmonary aspergillosis


Normal lungIPA

IPA occurs in ~7% of acute

leukaemia patients, 10-15% allogeneic BMT

patients

Unequivocal ‘Halo sign’ surrounding a noduleUnequivocal ‘Halo sign’ surrounding a nodule

Herbrecht, Denning et al, NEJM 2002;347:408-15.

Halo sign

Recent examples of the frequency of invasive aspergillosis

Underlying condition Incidence Reference/yearAcute myeloid leukaemia

8% Cornet, 2002

Acute lymphatic leukaemia

6.3% Cornet, 2002

Allogeneic HSCT 11-15% Grow, 2002; Marr, 2002

Lung transplantation 6.2-12.8% Minari, 2002; Singh,2003

Heart-lung transplantation

11% Duchini, 2002

Small bowel tranplantation

11% Duchini, 2002

AIDS 2.9% Libanore, 2002

Gillies & Campbell, www.aspergillus.org.uk

Bleeding as an aspect of disseminated invasive aspergillosis

Fumagillin is anti-angiogenic

A haemolysin described from Aspergillus fumigatus

Other factors that contribute to thrombosis or a coagulopathy?

How does Aspergillus fumigatus cause thrombosis (clotting of vessels) and also bleeding?

Filler et al, Blood 2004;103:2134; Paris et al, Infect Immun 1997;65:1510.

Interaction of conidia and

endothelial cell projections

Internalisation of conidia (and hyphae) by

endothelial cells with injury

apparent at 4 hours


Cerebral aspergillosis (abscess) in chronic lymphocytic leukaemia

Dissemination via the blood stream

to the brain occurs in ~5% of cases of

invasive aspergillosis, and

in ~40% of allogeneic bone marrow (HSCT)

recipients

Early diagnosis of invasive aspergillosis is important

Treatment started <10d >11dMortality 40% 90%

Von Eiff et al, Respiration 1995;62:241-7.

Sputum Cultures for Fungus

Bacteriological media inferior to fungal media – 32% higher yield on fungal

media

A four day A. fumigatus culture on malt extract agar (above). Light microscopy

pictures are taken at 1000x, stained with lacto-phenol cotton blue.

Aspergillus Antigen Test

• Diagnosis or surveillance? • Only blood, or BAL, CSF etc• Best OD cut-off - 0.7• False positives in kids / antibiotics• False negative with antifungal

prophylaxis• Not as useful for non-hematology• Not useful if pre-existing antibody

Herbrecht et al, J Clin Microbiol 2002;20:1898-906; and others

Outcome from invasive aspergillosis – amphotericin B therapy

Survival Functions by Site of Infection

Days

3603303002702402101801501209060300

Cu

mu

lative

Su

rviv

al R

ate

1.0

.9

.8

.7

.6

.5

.4

.3

.2

.1

0.0

CNS or Disseminated

Pulmonary (n=83)

Aspergilloma (n=10)

Multi-site (n=11)

Sinusitis (n=17)

(n=35)

Lin et al, Clin Infect Dis 2001;32:358

Sub-acute invasive aspergillosis in AIDSSub-acute invasive aspergillosis in AIDS


Sub-acute invasive aspergillosisSub-acute invasive aspergillosis

• Less immunocompromised patients• Slower progression of disease (> 1 month)• Cavitary or nodular pulmonary disease typical • Vascular invasion less common• Dissemination less common• Antigen testing less useful• Antibody testing may be helpful in diagnosis


Chronic necrotizing aspergillosis(CNPA)

Chronic necrotizing pulmonary aspergillosis (CNPA) is a subacute process usually found in patients with some degree of immunosuppression.

Usually it is associated with underlying lung disease, alcoholism, or chronic corticosteroid therapy. Because it is uncommon, CNPA often remains unrecognized for weeks or months and causes a progressive cavitary pulmonary infiltrate.

Right upper lobe. Patient has diabetes and pulmonary

mycobacterium avium- shows small cavitary lesion PT MS 1995.

Chronic necrotising pulmonary aspergillosis

Denning, Clin Microbiol Infect 2001;7(Suppl 2):25-31.

Right upper lobe showing circular shadow partly filled by a mass. PT

MS 1996

Same lobe shows expansion of the shadow, still partially

filled with a mass. Pt MS 1998

Right lobe shows huge cavity containing some

debris, with +ve aspergillus precipitins.Pt

MS 1999

Aspergillus and airways

Langley, ATS 2004

Types of aspergillosis of the airways• Colonisation (no disease – could be at risk)• Obstructing Aspergillus tracheobronchitis /Mucus

impaction (non-invasive)• Aspergillus bronchitis/tracheobronchitis (superficially invasive only)• Ulcerative Aspergillus tracheobroncitis (locally

invasive) (lung transplants – at anastomosis)• Pseudomembranous Aspergillus tracheobronchitis

(Extensive disease, locally invasive, associated with IPA and may disseminate)

Aspergillus tracheobronchitis

Autopsy drawing of a ‘normal’ 3 year old who died over 10 days

Wheaton, Path Trans 1890; 41:34-37

Aspergillus tracheobronchitis

Review of 58 patients in literature for normal and immuno compromised patients - risk factors

%None (ie normal) 25Heart / Lung transplant 18Solid tumour 15BMT 13Leukaemia 13HIV/AIDS 8Other 8

Kemper et al, Clin Infect Dis 1993; 17: 344

AspergillomaAspergilloma

Patient RTDecember 2002

Fungus ball

http://www.aspergillus.man.ac.uk/secure/image_library/agillom/brazil6.htm

Chronic pulmonary aspergillosis – pre-Chronic pulmonary aspergillosis – pre-existing diseaseexisting disease

All 18 patients had prior pulmonary disease

9 TB, 5 with atypical mycobacteria

13 smokers or ex-smokers

All 18 non-immunocompromised

3 excess alcohol

Denning DW et al, Clin Infect Dis 2003; 37:S265

Chronic pulmonary aspergillosis - Chronic pulmonary aspergillosis - presentationpresentation

Weight loss 16 / 18 (89%)Cough 15 / 18 (83%)Shortness of breath 9 / 18 (50%)Haemoptysis 9 / 18 (50%)Fatigue / malaise 5 / 18 (28%)Chest pain 3 / 18 (17%)Sputum production ++ 3 / 18 (17%)Fever 2 / 18 (11%)


Chronic pulmonary aspergillosis - serology

All 18 patients had positive Aspergillus precipitins (1+ - 4+)

All 18 patients had elevated inflammatory markers, CRP, PV and / or ESR

14 of 18 (78%) had elevated total IgE (>20), 13 >200 and 7 >400

9 of 14 (67%) had Aspergillus specific IgE (RAST)


Chronic cavitary pulmonary aspergillosis Chronic cavitary pulmonary aspergillosis (CCPA)(CCPA)

Patient RWSeptember 1992 Relapse in normal lung


Patient RWDecember 1991 Pre surgical resection

Chronic cavitary pulmonary Chronic cavitary pulmonary aspergillosisaspergillosis

Patient RWJuly 1993


Chronic Cavitary Pulmonary Chronic Cavitary Pulmonary AspergillosisAspergillosis

Patient JAJan 2001


Patient JAFeb 2002


Patient JAApril 2003


Patient JAJuly 2003

Chronic cavitary pulmonary Chronic cavitary pulmonary aspergillosisaspergillosis

Patient JP June 1999


Chronic Cavitary Pulmonary Aspergillosis, Chronic Cavitary Pulmonary Aspergillosis, with aspergillomawith aspergilloma

Patient JP July 2001


Chronic Fibrosing Pulmonary Chronic Fibrosing Pulmonary AspergillosisAspergillosis

Patient JPApril 2002


Mannose Binding Lectin (MBL)- a key Mannose Binding Lectin (MBL)- a key part of the innate immune systempart of the innate immune system

Disulphide bondInteraction with

collectin receptor

Interaction withMASP

Exon 1

Exon 2

Exon 3

Exon 4

CRD

Ca binding site2+

Crosdale et al J Infect Dis 2001;184:653

Mannose Binding ProteinMannose Binding Protein

5 mutations described 2 in promoter region (less important)3 in open reading frame (M52, M54, M57)

Codon 54 mutation present in 16% of Caucasian homozygous in 2%

Defects associated with bacterial infections in children and hepatitis B carriage

MutationsMutations

Eisen & Minchinton Clin Infect Dis 2003;37:1496

CCPA and human gene defectsCCPA and human gene defects

• 8 of 11 (72%) had low MBL genotypes p=<0.05 (compared to normal controls)

• 8 of 17 (47%) had low MBL genotypes p=0.0002

• 32% and 21.5% frequency of 2 SPA2 mutations, compared with normals (18% and 11%) (p=0.021 and p=0.044)

• not related to coeliac disease (<1 in 30)

Crosdale et al J Infect Dis 2001;184:653; Vaid et al, unpublished.

ALLERGIC BRONCHOPULMONARY ASPERGILLOSIS – Key diagnostic criteria

• Asthma• Blood eosinophilia (>1,000 / cu

mm)• History of pulmonary infiltrates• Central bronchiectasis

Rickett et al. Arch Intern Med 1983; 143: 1553; Patterson, Chest 2000;118:7

ABPA possibleABPA possibleABPA probable

ABPA almost certain

• Precipitins against A. fumigatus positive• Aspergillus IgE antibody >2x asthma control• Aspergillus IgG antibody >2x asthma control• Total serum IgE concentration, >1000 iu/mL

If 3 tests +ve, then ABPA very likely, If all 4 +ve the diagnosis established

ABPA


Before bronchoscopy

After bronchoscopy

ABPA mucous plugging


ABPA - CT showing central bronchiectasis


ABPA and surfactant

5 surfactant proteins in man, SPA1, SPA2, SPB, SPC and SPD – all ‘collectin’ family

Mason et al, Am J Physiol 1998;275:L1-13.

ABPA – surfactant defects

2 exonic polymorphisms, and 2 intronic polymorphisms in SP-A2 associated with ABPA

A1660G = OR of 4.78; or if combined with G1649C = OR 10.4

Also associated with higher peripheral eosinophilia

Saxena et al, J Allergy Clin Immunol 2003;111:1001-7.

Eosinophilic fungal rhinosinusitis or allergic fungal sinusitis

Patient with chronic symptoms of nasal obstruction, loss of smell and nasal polyps

Ponikau et al, Mayo Clinic Proc 1999;74:877 & WWW.aspergillus.org.uk

Eosinophilic fungal rhinosinusitis(link with airborne fungi - ?which most important

= Myelin basic protein, highly toxic to local epithelium

Ponikau et al, Mayo Clinic Proc 1999;74:877

A link between Aspergillus and asthma?

Fungal-associated asthma – evidence

Fungal-associated asthma

ABPA

Treatment of ABPAand pilot data

Severe asthma linked with fungal

sensitisation

Frequency of fungal sensitisation

High spore counts and asthmatic attacks

Spore counts and asthma attacks and admission to hospital

All circumstantial evidence

• Thunderstorm asthma – linked to Alternaria

• Asthma deaths (Chicago) linked to high ambient spores counts and season (summer autumn) when spore counts highest

• Asthma hospital admission linked to high ambient spore counts (Derby, New Orleans, Ottawa

• Asthma hospital attendance linked to high spore counts , but not pollen counts (Canada)

• Asthma symptoms increased on days of high spore counts (California, Pennsylvania)

O'Hollaren, N Engl J Med 1991; 324: 359; Newson, Occup Environ Med 2000; 57: 786-92.

Fungus at homeEnvironmental data

• Mouldy housing associated with worse asthma, with a correlation between asthma severity and degree of dampness in the home and separately with visible mould growth

• In Germany bronchial reactivity in children was associated with damp housing

• Mouldy and damp school associated with asthma symptoms and emergency room visits

• Highest concentration of Aspergillus fumigatus is at home

Williamson, Thorax 1997;52:229. Taskinen, Acta Paediatr 1999; 88:1373.

Severe asthma and moulds

Mild asthma – 564 (50%)

Moderate asthma – 333 (29%)

Severe asthma – 235 (21%) – linked with fungus skin test positivity

Zureik et al, Br Med J 2002;325:411

Asthma severity, house dust mites, cats and moulds

Langley, ATS 2004

Allergen No asthman= 111

Mild asthma FEV1 >75%

<90%n= 67

Moderate asthma FEV1

>60% <75%n= 42

Severe asthma FEV1

>60% n= 42

House dust mite

61% 71% 45% 77%

Cats* 49% 51% 38% 35%

Moulds# 17% 19% 36% 31%

* P = 0.05# p = 0.01

The extraordinary spectrum of diseases caused by Aspergillus David W. Denning Wythenshawe Hospital...

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