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

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

Documents

invasive aspergillosis

conventional diagnosis

positive culture

switzerland slide

progressive slide

promising slide

positive sputum culture

sensitivity of culture