1. Challenges in the diagnosis of Invasive Mould Diseases
What are we looking at?
Challenges in the diagnosis of Invasive Mould Diseases
Alessandro C. Pasqualotto Porto Alegre, Brazil

2. Potential conflicts of interest
Research Grants
Myconostica, Pfizer, Merck, Sigma-Tau, CAPES, CNPq, Fungal Research Trust
Travel Grants
Pfizer, United Medical, Schering (now Merck), Bagó, Merck
Speaker honoraria
Pfizer, United Medical, Merck, Schering (now Merck), Biometrix
Este slide e’ rapido

3. IFD are highly lethal diseases
First assumption:
IFD are highly lethal diseases
Este slide fala por si, em especial se considerarmos infeccoes por fungos filamentosos

4. Incidence 13.3% in lung transplant recipients
Apresento entao a sobrevida entre tx de pulmao que tiveram ou nao aspergilose.
Incidence 13.3% in lung transplant recipients
Xavier MO, Pasqualotto AC, et al. ECCMID 2009

5. Rapidly evolving diseases
Dizer entao que a evolucao e’ muito rapida, em paciente imunocomprimido, particularmente neutropenicos. Usar o termo agressivo ja desde o inicio, para acostumar a plateia com o que vira a seguir (a importancia de sermos agressivos).
4 days later

6. Disseminated infection
Sem tratamento e com persistencia de neutropenia, as infeccoes podem disseminar para o SNC, momento em que a mortalidade e’ entre %.

7. We need to intervene asap
Second assumption:
We need to intervene asap
Este slide fala por si

8. Early versus late intervention
Mortality rate (%)
Entao precisamos de um dx precoce, o que pode mudar o desfecho. No entanto, e’ preciso que saibamos o que estamos tratando
Von Eiff, et al. Respiration 1995; 62: 241-7

9. Early versus late intervention
Mortality rate (%)
Von Eiff, et al. Respiration 1995; 62: 241-7

10. But how can we achieve such an early diagnosis?
O que queremos e’ a verdade! Dizer com certeza que o paciente tem IFD. Mas isto requer biopsia e cultura, o que nao e’ alcancavel em geral na pratica clinica

11. A small black scar + serum GM Patient died 2 days earlier
1 day after this picture was taken
Vejam por ex este caso, de um paciente com leucemia aguda. Doenca com rapida evolucao para obito. GM positiva, mas que adianta, uma vez que o paciente ja esta morto neste estagio?

12. Zygo + A. flavus A small black scar + serum GM Patient died
2 days earlier
+ serum GM
Patient died
1 day after this picture was taken
Zygo + A. flavus

13. MD Anderson Cancer Centre Necropsy study over a 15-years period
IFD detected in 31% over 1,017 necropsies
Antemortem diagnosis in only 25%
Em hospital de referencia, e’ incrivel que a maioria dos dx ocorram pos-mortem apenas
Chamilos G, et al. Haematologica 2006; 91: 986-9

14. MD Anderson Cancer Centre Necropsy rate has reduced over time%
Mas infelizmente estamos fazendo menos e menos necropsia
Chamilos G, et al. Haematologica 2006; 91: 986-9

15. By the way, what is the necropsy rate in your institution?
>40%
10-39%
1-10%
<1%
Are you kidding?
Comentar aqui que histopatologia e’ a unica maneira de comprovar a presenca da infeccao

16. MD Anderson Cancer Centre Trends in the prevalence of IFD%
Chamilos G, et al. Haematologica 2006; 91: 986-9

17. CT scan and galactomannan
No need to worry!
CT scan and galactomannan
are there to help us out!

18. ‘Halo sign’ surrounding a nodule
Day 0:
halo
Day 4:
size, halo
Day 7:
air crescent
Tanto o halo como o sinal do ar crescente sao manifest altammente sugestivas. No entanto, o halo e’ fugaz e precoce. Se nao tivermos politica agressiva de TC, nao faremos o dx
Caillot, et al. J Clin Oncol 1997; 15:

19. The sign is not specific for IA
Vasculitis
Metastasis
Pseudomonas infections
Zygomycosis and other angio-invasive infections
Esta tambem presente em muitas lesoes vasculares, incluindo zigomicose!
Greene RE, et al. Clin Infect Dis 2007; 44: 373-9

20. Absence of typical findings at chest CT scan
COPD
Steroids
Other non-neutropenic patients / ICU
Lung transplant recipients
? Monoclonal antibodies

21. Organising cryptogenic pneumonia
‘Reversed halo sign’
Organising cryptogenic pneumonia
Importante lembrar tambem do sinal do halo reverso, que pode sugerir zigo
Wahba H, et al. Clin Infect Dis 2008; 46:

22. ‘Reversed halo sign’ Review of 189 cases of invasive mould disease
Overall frequency 4%
Wahba H, et al. Clin Infect Dis 2008; 46:

23. ‘Reversed halo sign’ Review of 189 cases of invasive mould disease
Overall frequency 4%
Zygomycosis 19%
Aspergillosis <1%
Fusariosis 0% (p<0.01)
Wahba H, et al. Clin Infect Dis 2008; 46:

24. Other predictors of zygomycosis
>10 nodules
Pleural effusion
Concomitant sinusitis
Treatment with voriconazole
Lembrar do paciente que tivemos – bem como da nossa estatistica do HCPA
Chamilos G, et al. Clin Infect Dis 2005; 41: 60-6

25. Meta-analysis of GM testing
Low PPV High NPV
Pfeiffer CD, et al. Clin Infect Dis 2006; 42:

26. Caution with low +ve indexes!
Reproducibility
Caution with low +ve indexes!
Ja acumulamos alguma experiencia com o teste e observamos variacao importante … nao se pode usar na presenca de ictericia …
Upton A, et al. J Clin Microbiol 2005; 43:

27. GM release by non-Aspergillus fungi
Penicillium marneffei
Geotricum capitatum
Acremonium species
Alternaria alternata
Rhodotorula rubra
Trichophyton species
Paecilomyces variotii
Botrytis tulipae
Cladosporium species
Exophiala dermatitidis
Agora discutir GM
Aquino VR, Goldani LZ, Pasqualotto AC. Mycopathologia 2007; 163:

28. Cross-reaction with GM testing%
Xavier MO, Pasqualotto AC, Severo LC. Clin Vaccin Immunol 2009; 16: 132-3

29. Clinical case 19 year-old man, refractory leukaemia
Febrile neutropenia
Amox-clav for E. Coli bacteremia
Maertens J, et al. Clin Infect Dis 2004; 39:

30. Clinical case 19 year-old man, refractory leukaemia
Febrile neutropenia
Amox-clav for E. Coli bacteremia
Daily GM determination
D1 after antibiotic: GM index of >1.5
5 +ve tests afterwards
Maertens J, et al. Clin Infect Dis 2004; 39:

31. Clinical case 19 year-old man, refractory leukaemia
Febrile neutropenia
Amox-clav for E. Coli bacteremia
Daily GM determination
D1 after antibiotic: GM index of >1.5
5 +ve tests afterwards
Fluoroquinolone: gradual reduction in GM index
Maertens J, et al. Clin Infect Dis 2004; 39:

32. Clinical case 1 wk later Pipe-tazo for appendicitis
GM >2.5; bilateral nodular infiltrate
Maertens J, et al. Clin Infect Dis 2004; 39:

33. Clinical case 1 wk later Probable IA (EORTC / MSG)
Pipe-tazo for appendicitis
GM >2.5; bilateral nodular infiltrate
Probable IA (EORTC / MSG)
Antifungal therapy + meropenem
Gradual ↓ in GM index
Maertens J, et al. Clin Infect Dis 2004; 39:

34. Clinical case 1 wk later Probable IA (EORTC / MSG)
Pipe-tazo for appendicitis
GM >2.5; bilateral nodular infiltrate
Probable IA (EORTC / MSG)
Antifungal therapy + meropenem
Gradual ↓ in GM index
Necropsy: leukaemia infiltrate
Absence of IA
Maertens J, et al. Clin Infect Dis 2004; 39:

35. Meta-analysis of GM testing
Proven or probable IA
Sensitivity
Specificity
Haematological malignancies
0.58 (52-64)
0.95 (94-96)
Solid organ transplantation
0.41 (21-64)
0.85 (80-89)
Para estes casos e’ preciso testar LBA; e ai nao temos ponto de corte
Pfeiffer CD, et al. Clin Infect Dis 2006; 42:

36. (particularly for sensitivity)
Specificity 94%
Sensitivity 79%
Marked heterogeneity
(particularly for sensitivity)
BAL PCR testing
Tuon FF. Rev Iberoam Micol 2007; 24: 89-94

37. PCR Critical points Variable sensitivity / specificity
Lack of standardised targets / reagents
Extraction method
Platform (conventional PCR vs Real time)
Poor understanding of DNA kinetics
Not yet part of the EORTC/MSG criteria
De Pauw B, et al. Clin Infect Dis 2008; 46:

38. Beta-Glucan Detected in IFDs caused by Candida and Aspergillus Fungal
cell wall
Phospholipid bilayer
of the fungal cell
membrane
-(1,3)-glucan
-(1,3)-glucan synthase
Ergosterol
Yoshida M, et al. J Med Veter Mycol 1997; 35: 371-4

39. Beta-Glucan Detected in IFDs caused by Candida and Aspergillus
Trichosporon
Fusarium
Acremonium
Saccharomyces
Pneumocystis
Yoshida M, et al. J Med Veter Mycol 1997; 35: 371-4

40. speaking about a test that nobody uses in Brazil …
There he is again,
speaking about a test that nobody uses in Brazil …

41. The reality in Brazil A survey performed in collaboration with ANVISA
140 hospitals
>42,000 beds
65% teaching hospitals
90% belonging to the Sentinel Network

42. Complexity in hospital care%

43. Is IFD a problem in your centre?
38.7%
n=140

44. Do you know your local epidemiology?
40.1%
n=140

45. Specialised media for fungi
19.7%
n=140

46. Aspergillus identification
at the species level
58.5%
n=140

47. Fungal staining - biopsies
51.1%
n=140

48. Biopsy specimens are sent in formalin only?
26.0%
n=140

49. Access to high resolution CT
34.3%
n=140

50. Galactomannan
83.6%
n=140

51. The appropriateness or inappropriateness of feelings is relative to the ground and to the circumstances of those feelings Aristotle The Doctrine of the Mean, BC
Aristoteles ja dizia que nao ha sentimento que nao possa ser expresso … Por favor, nao se sintam culpados por usar de agressividade …

52. Fight fire with fire Metallica Ride the Lightning, 1984 AD
Mais recentemente …

53. Invasive diagnostic interventions
CT-guided percutaneous lung biopsy: yield of %
Dizer que a Cornelia tem feito um otimo trabalho neste campo, tentando ao maximo fazer o dx ainda em vida para pacientes com infeccoes invasivas por fungos filamentosos
Nosari A, et al. Haematologica 2003; 88:
Crawford SW, et al. Transplantation 1989; 48:
Hoffer FA, et al. Pediatr Radiol 2001; 31:
Lass-Florl C, et al. Clin Infect Dis 2007; 45: e1001-4

54. Invasive diagnostic interventions
CT-guided percutaneous lung biopsy: yield of %
Platelets >60,000/ml are required
Pneumothorax 18%
Haemoptysis 3%
Nosari A, et al. Haematologica 2003; 88:
Crawford SW, et al. Transplantation 1989; 48:
Hoffer FA, et al. Pediatr Radiol 2001; 31:
Lass-Florl C, et al. Clin Infect Dis 2007; 45: e1001-4

55. Invasive diagnostic interventions
Transbronchial biopsies:
False-negative results are frequently seen
Lass-Florl C, Freund MC. In: Aspergillosis: from diagnosis to prevention. Pasqulaotto AC, ed. Springer, 2009

56. Invasive diagnostic interventions
Open lung biopsies:
Provide larger samples of tissue with improved accuracy and specificity
Lass-Florl C, Freund MC. In: Aspergillosis: from diagnosis to prevention. Pasqulaotto AC, ed. Springer, 2009

57. Invasive diagnostic interventions
Open lung biopsies:
Provide larger samples of tissue with improved accuracy and specificity
Contradictory results
Complication rate of 10-15%
Severe thrombocytopenic patients who urgently require histological identification should undergo open lung biopsy
Lass-Florl C, Freund MC. In: Aspergillosis: from diagnosis to prevention. Pasqulaotto AC, ed. Springer, 2009

58. Invasive diagnostic interventions
Needle biopsy /
Surgical resection
Peripheral lesions
Bilateral /
multifocal disease
BAL
Focal lesions near the hilum and the great vessels in the lung, particularly in neutropenic patients, urgent thoracotomy and resection should be considered because of the risk of life-threatening haemoptysis
Focal lesions near the
hilum / great vessels
Urgent thoracotomy and resection
Lass-Florl C, Freund MC. In: Aspergillosis: from diagnosis to prevention. Pasqulaotto AC, ed. Springer, 2009

59. Conclusion It is mostly but not all about IA
Invasive mould diseases have to be differentiated from each other

60. Conclusion It is mostly but not all about IA
Invasive mould diseases have to be differentiated from each other
Diagnosis is the most challenging step in infectious diseases

61. Conclusion It is mostly but not all about IA
Invasive mould diseases have to be differentiated from each other
Diagnosis is the most challenging step in infectious diseases
We need a better understanding on the performance of the available tests

62. Acknowledgments Mycology team Infection Control Dept
Luiz Carlos Severo
Valerio R Aquino
Cecilia B Severo
Luciana Guazelli
Melissa Xavier
Infection Control Dept
Teresa Sukiennik