Invasive aspergillosis : Update on conventional diagnosis Jacques Bille, MD Institut of Microbiology University Hospital Lausanne, Switzerland

Invasive aspergillosis :

Update on conventional diagnosis

Jacques Bille, MDInstitut of MicrobiologyUniversity HospitalLausanne, Switzerland

Invasive aspergillosis : update on conventional diagnosis

The problems

The disease is evolving

Clinical signs/ symptoms are non specific

Conventional diagnosis is insensitive or too late

Non conventional diagnosis is "promising"


The disease is evolving

A continuum of clinical presentations

New (?) hosts

Underdiagnosed ante-mortem


Hope, WW, Walsh TJ, Denning DW. The invasive and saprophytic syndromes due to Aspergillus species. Medical Mycology 2005, 43: S 207-238


Clinical signs/symptoms are non specific

feverdyspneanon productive coughhemoptysischest painpleural rib

In a proper hostrapidly progressive


Conventional diagnosis

• direct examination of tissue of indirect clinical specimens (sputum, BAL)

sputum/BAL tissue

unstained wet prep ± KOHroutine stains Gram HE fungal stains GMS, PAS GMS, PASfluorescent Calcofluor white

dyes Uvitex 2BBlankophor


In: Hope et al., Lancet Infectious Diseases 5: 609, 2005


Histopathologic diagnosis of Aspergillosis

Improvement by immunohistochemical detection

Monoclonal antibodies• WF-AF-1 (Dako) 1)

• EB-A1 2)

Sensitivity in culture-proven cases : 89-94%Genus or species specific results

1) Choi JK et al., Am J Clin Pathol 121: 18, 20042) Pierard GE et al., Am J Clin Pathol 96: 373, 1991 Verweij PE et al., Am J Clin Pathol 49: 798, 1996


In: Choi JK, Am J Clin Pathol 121: 18, 2004


Culture

Isolation: Sabouraud (+ antibiotics)Media blood agar, chocolate agar

Identification : malt-extract, corn-meal agarCzapek agar

Incubationtemperature 25-30°C improvement ?athmosphere aerobicduration 2-6 weeks


In: Andreoni et al., Medical Mycology Atlas


Culture

• Can we improve the sensitivity ? (in tissue)

• What is the best indirect specimen ?Sputum or BAL

• What are the PPV of a positive culture for IA, and the sensitivity of culture to diagnose IA ?


Performance of a diagnostic test

Caveat

– timing of sampling - evolution of the disease

– per test vs per episode analysis

– patient population

– Prior antifungal prophylaxis or therapy


Yield of culture for molds in histopathologic positive samples

n= culture positive

Autopsy samples 23 12 52%

Surgical or biopsy tissue 30 9 30%

Tarrand JJ et al., Am J Clin Pathol 2003; 119: 854


Culture

Can we do better ?

Mimic physiologic termperature and decreased oxygen environment : 35°C, 6% O2 -10% CO2

significant increase of Aspergillus spp from autopsy tissue and various clinical samples (+ 31%)

Tarrand JJ et al., J Clin Microbiol 2005; 43: 382


Culture : BAL is better than sputum (?)Overall sensitivity 50%

Adapted from Reichenberger et al., Bone Marrow Transplantation 1999; 24: 1195

Diagnostic yield of bronchoscopy specimen in histologically proven IPA

No. of cases Bronchoscopy diagnostic

Albeda 1984 15 14 5 36 %Treger 1985 16 12 8 67 %Kahn 1986 17 27 13 48 %Saito 1988 18 9 0 0 %Levy 1992 11 16 9 56 %Mc Whinney 1993 19 12 6 50 %Saugier-Weber 1993 2 10 3 30 %Von Eiff 1995 12 12 6 50 %Horvath 1996 8 29 11 38 %Caillot 1997 20 18 8 45 %Baron 1998 4 13 8 61 %Reichenberger 1999 9 23 7 30 %

Overall 195 84 43 %


The paradox of a positive sputum culture for Aspergillus

low sensitivity for IAlow specificity for IA

Can we improve ?


How to increase the PPV for IA of a positive sputum culture

• at (high) risk patient

• multiple positive samples 1)

• quantitative culture 1) 2)

• use of a score 2) 3)

1) Nalesnik et al., J Clin Microbiol 1980; 11: 3702) Greub and Bille, Clin Microbiol Infect 1998; 4: 7103) Bouza and Muñoz, J Clin Microbiol 2005; 43: 2075


Positive predictive value of a positive sputum culture for IA

Highly variable (15-77%)Depends on host factors

allo BMT 60%leukemia + neutropenia 70-80%SOT 20-60%HIV/AIDS 14-20%Corticosteroids 20%

Perfect et al. (MSG), Clin Infect Dis 2001; 33: 1824Yu et al., Am J Med 1986; 81: 249Horvath and Dummer, Am J Med 1996; 100: 171


Prospective assessment of the clinical signi- ficance of isolating A.fumigatus by culture

404 A.fumigatus positive cultures (260 patients)

90 (22.3%) from 31 (12%) patients with IA

6% if 1+ cult.18% 2+38% 3+

10% if 1-2 score40% 3-470% 5

Bouza et al., J Clin Microbiol 2005; 43: 2075

ScoreCriteriapointsInvasive procedure 1 2 + cultures 1Leukemia 2Corticosteroids 2Neutropenia 5


Radiology

2 interesting recent findings :

Systematic CT • is more sensitive than galactomannan for early detection of IA1)

• reduces the amount of antifungal therapy when combined to GM screening2)

Angio CT has a higher specificity than CT for IA3)

1) Weisser et al., Clinical Infectious Diseases 2005; 41: 11432) Maertens et al., Clinical Infectious Diseases 2005; 41: 12423) Sonnet et al., Am J Roentgenol 2005; 184: 746


Conclusions

Conventional diagnosis of IA is :

suboptimal indispensable

genus, species

AFST perfectible

Documents

Invasive aspergillosis : Update on conventional diagnosis Jacques Bille, MD Institut of Microbiology University Hospital Lausanne, Switzerland