The extraordinary spectrum of diseases caused by Aspergillus

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

2. Several species of Aspergillus can cause disease in humans and animals particularly when the host is immunosuppressed, eg in cancer patients, transplant patients and AIDS patients. Aspergillus spores are airborne and occur naturally in the environment, but are often more abundant around building sites. Near compost or where damp conditions prevail. Invasive disease usually only occurs in immunosuppressed patients with inhalation being the primary route of infection. Allergic aspergillosis occurs in patients with asthma, atopy or cystic fibrosis. In many underdeveloped countries contamination of crops such as maize, groundnuts etc has lead to loss of crops and severe illness in both humans and farm stock where contaminated food is ingested (due to mycotoxins which harm the body). Several species of Aspergillus can cause disease in humans and animals particularly when the host is immunosuppressed, eg in cancer patients, transplant patients and AIDS patients. Aspergillus spores are airborne and occur naturally in the environment, but are often more abundant around building sites. Near compost or where damp conditions prevail. Invasive disease usually only occurs in immunosuppressed patients with inhalation being the primary route of infection. Allergic aspergillosis occurs in patients with asthma, atopy or cystic fibrosis. In many underdeveloped countries contamination of crops such as maize, groundnuts etc has lead to loss of crops and severe illness in both humans and farm stock where contaminated food is ingested (due to mycotoxins which harm the body).

3. 1) Fungi play an essential role in both the Nitrogen and Carbon cycle by breaking down dead organic material. 2) The detailed genetic study of A.nidulans has lead to some important discoveries, such as the parasexual cycle in fungi, and the genetic defect causing alkaptonuria 3) Most of us use fungi every day without knowing it. We eat mushrooms and Quorn™ (a vegetarian fungal protein made from Fusarium graminarium), but we also prepare many other foods using fungi. The yeast Saccharomyces cerevisiae is used to ferment sugar to alcohol and carbon dioxide – the process used to make beer and wine and also to make bread rise. All citric acid is produced by Apergillus niger. The fungi Aspergillus oryzae and Aspergillus sojae are used in the production of the oriental foods soy sauce and miso. We also use fungi to produce flavourings, vitamins and enzymes and to mature many cheeses.4) Apart from penicillin, the most important antibiotics from fungi are the cephalosporins (beta-lactams with similar mode of action to penicillin, but with less allergenicity) and griseofulvin (from Penicillium griseofulvum and related species) which is used to treat athlete's foot and related fungal infections of the skin. The echinocandins anidulafungin and aminocandin (HMR 327) are semi-synthetic antifungals originally derived from fermentation of A. nidulans var restrictus and A. sydowi respectively. Lovastatin a cholesterol lowering drug is produced by Aspergillus terreus. And the group of antifungal drugs called echinocandins have been produced using A nidulans and A sydowi. Fumagillin is a secondary metabolite that inhibits angiogenesis and antibiotic properties, especially against protozoa such as Entamoeba histolytica. LD50 in mice 800 mg/kg body-weight subcutaneously although as much as 2 mg/kg body-weight could be tolerated orally. Fumigillin or analogue TNP-470 have been used in the treatment of microsporidiosis, usually caused by the microscopic fungus Enterocytozoon bieneusi (Fumagillin treatment of intestinal microsporidiosis. New Engl J Med 2002;346:1963-9.). 1) Fungi play an essential role in both the Nitrogen and Carbon cycle by breaking down dead organic material. 2) The detailed genetic study of A.nidulans has lead to some important discoveries, such as the parasexual cycle in fungi, and the genetic defect causing alkaptonuria 3) Most of us use fungi every day without knowing it. We eat mushrooms and Quorn™ (a vegetarian fungal protein made from Fusarium graminarium), but we also prepare many other foods using fungi. The yeast Saccharomyces cerevisiae is used to ferment sugar to alcohol and carbon dioxide – the process used to make beer and wine and also to make bread rise. All citric acid is produced by Apergillus niger. The fungi Aspergillus oryzae and Aspergillus sojae are used in the production of the oriental foods soy sauce and miso. We also use fungi to produce flavourings, vitamins and enzymes and to mature many cheeses.4) Apart from penicillin, the most important antibiotics from fungi are the cephalosporins (beta-lactams with similar mode of action to penicillin, but with less allergenicity) and griseofulvin (from Penicillium griseofulvum and related species) which is used to treat athlete's foot and related fungal infections of the skin. The echinocandins anidulafungin and aminocandin (HMR 327) are semi-synthetic antifungals originally derived from fermentation of A. nidulans var restrictus and A. sydowi respectively. Lovastatin a cholesterol lowering drug is produced by Aspergillus terreus. And the group of antifungal drugs called echinocandins have been produced using A nidulans and A sydowi. Fumagillin is a secondary metabolite that inhibits angiogenesis and antibiotic properties, especially against protozoa such as Entamoeba histolytica. LD50 in mice 800 mg/kg body-weight subcutaneously although as much as 2 mg/kg body-weight could be tolerated orally. Fumigillin or analogue TNP-470 have been used in the treatment of microsporidiosis, usually caused by the microscopic fungus Enterocytozoon bieneusi (Fumagillin treatment of intestinal microsporidiosis. New Engl J Med 2002;346:1963-9.).

4. Aspergillus Life-cycle Short movie clips showing germination of spores and growth of hyphae can be viewed on this website at the following link: http://www.aspergillus.man.ac.uk/secure/educationsection/movies/af65hyphae.htmlShort movie clips showing germination of spores and growth of hyphae can be viewed on this website at the following link: http://www.aspergillus.man.ac.uk/secure/educationsection/movies/af65hyphae.html

5. The relative frequency of species associated with acute invasive aspergillosis is 85-90% A. fumigatus, 5-10% A. flavus, 3-7% A. niger and 1% A. nidulans. There are approximately 25 -30 other species of Aspergillus which have caused disease in humans.. ( see http://www.aspergillus.man.ac.uk/ view the species section within the image bank). The few species which cause disease in man and animals are also known to have some resistance to antifungal drugs.The relative frequency of species associated with acute invasive aspergillosis is 85-90% A. fumigatus, 5-10% A. flavus, 3-7% A. niger and 1% A. nidulans. There are approximately 25 -30 other species of Aspergillus which have caused disease in humans.. ( see http://www.aspergillus.man.ac.uk/ view the species section within the image bank). The few species which cause disease in man and animals are also known to have some resistance to antifungal drugs.

6. CLASSIFICATION OF ASPERGILLOSIS Persistence without disease - colonisation of the airways or nose/sinuses Exposure to individual Aspergillus spores or conidia is almost constant. If eradicated immediately, as is usual in normal people, no disease results. If colonisation occurs, it may be short or long term. The pattern of disease is mostly determined by the host group (see slide 8), with probably a component of the inoculum size contributing to invasive disease. Acute and subacute disease is usually associated with immunocompromised patients eg AIDS, chemotherapy, transplant etc. and these will be discussed first.Exposure to individual Aspergillus spores or conidia is almost constant. If eradicated immediately, as is usual in normal people, no disease results. If colonisation occurs, it may be short or long term. The pattern of disease is mostly determined by the host group (see slide 8), with probably a component of the inoculum size contributing to invasive disease. Acute and subacute disease is usually associated with immunocompromised patients eg AIDS, chemotherapy, transplant etc. and these will be discussed first.

7. 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.

8. Interaction of Aspergillus with the hostA unique microbial-host interaction Objective: The risk of developing invasive aspergillosis increases with increasing levels of immune deficiency. In contrast the risk of allergic aspergillosis is enhanced in individuals whose immune system is hyper-reactive (atopic). Objective: The risk of developing invasive aspergillosis increases with increasing levels of immune deficiency. In contrast the risk of allergic aspergillosis is enhanced in individuals whose immune system is hyper-reactive (atopic).

9. Changing incidence of fatal invasive mycoses in non-HIV patients in USA This slide contrasts the reduction in mortality related to candidaemia and invasive candidiasis with invasive aspergillosis in the US. Other figures from Europe indicate 4% of patients dying in teaching hospitals have invasive aspergillosis, including medical intensive care in patients not normally considered susceptible.This slide contrasts the reduction in mortality related to candidaemia and invasive candidiasis with invasive aspergillosis in the US. Other figures from Europe indicate 4% of patients dying in teaching hospitals have invasive aspergillosis, including medical intensive care in patients not normally considered susceptible.

10. Chest X ray showing large lesion due to invasive pulmonary aspergillosis. Chest radiograph with ‘classical’ appearance of a pulmonary infarction – a wedge-shaped lesion peripherally set against the pleura. This patient was receiving chemotherapy including corticosteroids, who had had a splenectomy previously presented with fever and right-sided pleuritic chest pain. Blood cultures grew Aspergillus fumigatus and he responded to amphotericin B and flucytosine. (Case published in Denning DW, Williams A H. Invasive pulmonary aspergillosis diagnosed by blood culture and successfully treated. Br J Dis Chest 1987, 81: 300. See also case 38 in the case history section www.aspergillus.man.ac.uk)Chest X ray showing large lesion due to invasive pulmonary aspergillosis. Chest radiograph with ‘classical’ appearance of a pulmonary infarction – a wedge-shaped lesion peripherally set against the pleura. This patient was receiving chemotherapy including corticosteroids, who had had a splenectomy previously presented with fever and right-sided pleuritic chest pain. Blood cultures grew Aspergillus fumigatus and he responded to amphotericin B and flucytosine. (Case published in Denning DW, Williams A H. Invasive pulmonary aspergillosis diagnosed by blood culture and successfully treated. Br J Dis Chest 1987, 81: 300. See also case 38 in the case history section www.aspergillus.man.ac.uk)

11. CT scan showing characteristic halo sign - in a cavity with fungus ball there is a crescent shaped semi-translucent space along the upper portion, of a density giving the appearance of a halo. Angioinvasion is the pathological hallmark of acute IPA in the neutropenic setting. Two patterns of pathology are discernable: invasion of major proximal pulmonary arteries with resultant thrombosis and distal tissue infarction and a well circumscribed spherical nodule with a vessel in the centre of the lesion invaded by hyphae. Such nodules have a pale centre of coagulative necrosis with extensive permeation of tissue by hyphal elements but few inflammatory cells or haemorrhage. Surrounding this necrotic centre is haemorrhagic parenchyma. In the former, the radiological appearance is one of a wedge shaped lesion with the base abutting the visceral pleura. [Fraser]. The latter lesion is seen on CT as a nodule with or without an associated halo sign [Fraser, Meziane]. If the lesion cavitates, the area of central necrosis (sequestrum) contracts with replacement by an air-cap, and an air-crescent sign may be visible. Fraser RS. Pulmonary aspergillosis: pathologic and pathogenetic features. Pathol Annu 1993; 28 Pt 1: 231-277. Meziane MA, Hruban RH, Zerhouni EA, et al. High resolution CT of the lung parenchyma with pathological correlation. Radiographics 1988; 8: 27-54.CT scan showing characteristic halo sign - in a cavity with fungus ball there is a crescent shaped semi-translucent space along the upper portion, of a density giving the appearance of a halo. Angioinvasion is the pathological hallmark of acute IPA in the neutropenic setting. Two patterns of pathology are discernable: invasion of major proximal pulmonary arteries with resultant thrombosis and distal tissue infarction and a well circumscribed spherical nodule with a vessel in the centre of the lesion invaded by hyphae. Such nodules have a pale centre of coagulative necrosis with extensive permeation of tissue by hyphal elements but few inflammatory cells or haemorrhage. Surrounding this necrotic centre is haemorrhagic parenchyma. In the former, the radiological appearance is one of a wedge shaped lesion with the base abutting the visceral pleura. [Fraser]. The latter lesion is seen on CT as a nodule with or without an associated halo sign [Fraser, Meziane]. If the lesion cavitates, the area of central necrosis (sequestrum) contracts with replacement by an air-cap, and an air-crescent sign may be visible. Fraser RS. Pulmonary aspergillosis: pathologic and pathogenetic features. Pathol Annu 1993; 28 Pt 1: 231-277. Meziane MA, Hruban RH, Zerhouni EA, et al. High resolution CT of the lung parenchyma with pathological correlation. Radiographics 1988; 8: 27-54.

12. Recent examples of the frequency of invasive aspergillosis Recent examples of the incidence of IA in different at risk groups. Actual numbers are unit dependant and diagnostic criteria dependent.Recent examples of the incidence of IA in different at risk groups. Actual numbers are unit dependant and diagnostic criteria dependent.

13. Bleeding as an aspect of disseminated invasive aspergillosis Fumagillin -  a product of Aspergillus fumigatus is an inhibitor of endothelial cell proliferation and angiogenesis. Yet angiogenesis is a hallmark for invasive aspergillosis. (see previous slide). Certain proteases are probably important in contributing to a coagulation disorder. Fumagillin -  a product of Aspergillus fumigatus is an inhibitor of endothelial cell proliferation and angiogenesis. Yet angiogenesis is a hallmark for invasive aspergillosis. (see previous slide). Certain proteases are probably important in contributing to a coagulation disorder.

14. How does Aspergillus fumigatus cause thrombosis (clotting of vessels) and also bleeding?  These images show the interaction of swollen Aspergillus fumigatus conidia interacting with endothelial cells (the innermost layer of cells in blood vessels). The images direct interaction of conidia with the cells, phagocytosis by the endothelial cell, and then cellular damage, all in a 4 hour time-frame. The precise mechanisms are not clear.These images show the interaction of swollen Aspergillus fumigatus conidia interacting with endothelial cells (the innermost layer of cells in blood vessels). The images direct interaction of conidia with the cells, phagocytosis by the endothelial cell, and then cellular damage, all in a 4 hour time-frame. The precise mechanisms are not clear.

15. Cerebral aspergillosis (abscess) in chronic lymphocytic leukaemia Patient diagnosed with Stage C Chronic Lymphocytic Leukaemia, treated in the MRC CLL4 study, with prednisolone for 4 weeks followed by oral chlorambucil for 7 days. Patient developed severe pneumonia due to pseudomonas and staphylococcus. Following treatment with broad spectrum antibiotics and 1 week of Abelcet, patient was readmitted with headache, disorientation and fever. CT brain scans showed 3 ring enhancing lesions, aspirated material showed neutrophils but grew aspergillus. Patient now improved on Abelcet (4 mg/kg) and oral itrconazole suspension 200mg b.d. Patient diagnosed with Stage C Chronic Lymphocytic Leukaemia, treated in the MRC CLL4 study, with prednisolone for 4 weeks followed by oral chlorambucil for 7 days. Patient developed severe pneumonia due to pseudomonas and staphylococcus. Following treatment with broad spectrum antibiotics and 1 week of Abelcet, patient was readmitted with headache, disorientation and fever. CT brain scans showed 3 ring enhancing lesions, aspirated material showed neutrophils but grew aspergillus. Patient now improved on Abelcet (4 mg/kg) and oral itrconazole suspension 200mg b.d.

16. Early diagnosis of invasive aspergillosis is important Treatment started <10d >11d Mortality 40% 90%

17. Sputum Cultures for Fungus Bacteriological media inferior to fungal media – 32% higher yield on fungal media At least 3 sputum specimens should be submitted for fungal culture whenever fungal infection is suspected.On a survey of routine bacterial culture versus fungal culture, many positive cultures for Aspergillus were found on the routine cultures, but 32% of instances of Aspergillus were only seen in the fungal culture medium. Horvath & Dummer, Am J Med 1996;100:171-8.At least 3 sputum specimens should be submitted for fungal culture whenever fungal infection is suspected.On a survey of routine bacterial culture versus fungal culture, many positive cultures for Aspergillus were found on the routine cultures, but 32% of instances of Aspergillus were only seen in the fungal culture medium. Horvath & Dummer, Am J Med 1996;100:171-8.

18. There are substantial data on aspergillus antigen (galactomannan) using the commercial ELISA format published. Essentially, it can be detected in blood in most patients with acute invasive aspergillosis. False positives arise because of intestinal absorption of galactomannan which is found in many foods (see http://www.aspergillus.man.ac.uk/indexhome.htm?secure/diagnosis/index.php~main (oldest article), and contamination of some antibiotics batches with antigen. Most Aspergillus species are detected, as all the common pathogenic species produce galactomannan. False negatives do arise, probably partly because of complexing by antibody. If the absolute incidence of IA is high (ie >6%) the balance of argument is to use a screening approach, if less frequent than this, a diagnostic approach. However, anti-aspergillus prophylaxis (such as itraconazole solution) probably reduces the test’s utility. Once positive, rising titres on treatment predict failure, and usually death. CSF antigen is useful for diagnosing cerebral aspergillosis, and BAL antigen is probably useful for pulmonary aspergillosis, but the assay cut-off and performance is not yet clear.There are substantial data on aspergillus antigen (galactomannan) using the commercial ELISA format published. Essentially, it can be detected in blood in most patients with acute invasive aspergillosis. False positives arise because of intestinal absorption of galactomannan which is found in many foods (see http://www.aspergillus.man.ac.uk/indexhome.htm?secure/diagnosis/index.php~main (oldest article), and contamination of some antibiotics batches with antigen. Most Aspergillus species are detected, as all the common pathogenic species produce galactomannan. False negatives do arise, probably partly because of complexing by antibody. If the absolute incidence of IA is high (ie >6%) the balance of argument is to use a screening approach, if less frequent than this, a diagnostic approach. However, anti-aspergillus prophylaxis (such as itraconazole solution) probably reduces the test’s utility. Once positive, rising titres on treatment predict failure, and usually death. CSF antigen is useful for diagnosing cerebral aspergillosis, and BAL antigen is probably useful for pulmonary aspergillosis, but the assay cut-off and performance is not yet clear.

19. Outcome from invasive aspergillosis – amphotericin B therapy Note 60% death rate in 30 days from acute invasive pulmonary aspergillosis treated with amphotericin B.Note 60% death rate in 30 days from acute invasive pulmonary aspergillosis treated with amphotericin B.

20. This patient, thought initially to have pulmonary tuberculosis in AIDS, has bilateral upper-lobe cavities, more marked on the left. He presented with fever, non-productive cough and dyspnoea. Bronchoscopy yielded Aspergillus fumigatus. He refused therapy and died with progressive disease. He is reported as patient 8 in (Denning DW, Follansbee S, Scolaro M, Norris S, Edelstein D, Stevens DA. Pulmonary aspergillosis in AIDS. N Engl J Med 1991; 324: 654-662.) This patient, thought initially to have pulmonary tuberculosis in AIDS, has bilateral upper-lobe cavities, more marked on the left. He presented with fever, non-productive cough and dyspnoea. Bronchoscopy yielded Aspergillus fumigatus. He refused therapy and died with progressive disease. He is reported as patient 8 in (Denning DW, Follansbee S, Scolaro M, Norris S, Edelstein D, Stevens DA. Pulmonary aspergillosis in AIDS. N Engl J Med 1991; 324: 654-662.)

21. Sub-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

22. 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.

23. This man was an insulin dependent diabetic, otherwise well. The first panel (chest X-ray) shows the original infection with mycobacterial infection (M. avium intracellulare (atypical)). The second shows a much larger cavity, with a fungal mass in it. An isolated film would allow a diagnosis of aspergilloma, but later X-rays show progression. The third chest X-ray shows an expanding cavity, with a larger fungal ball in it. The final X-ray shows a very large thin-walled cavity containing fluid. Aspiration of the fluid grew Staphylococcus aureus.This man was an insulin dependent diabetic, otherwise well. The first panel (chest X-ray) shows the original infection with mycobacterial infection (M. avium intracellulare (atypical)). The second shows a much larger cavity, with a fungal mass in it. An isolated film would allow a diagnosis of aspergilloma, but later X-rays show progression. The third chest X-ray shows an expanding cavity, with a larger fungal ball in it. The final X-ray shows a very large thin-walled cavity containing fluid. Aspiration of the fluid grew Staphylococcus aureus.

24. CLASSIFICATION OF ASPERGILLOSIS Persistence without disease - colonisation of the airways or nose/sinuses Exposure to individual Aspergillus spores or conidia is almost constant. If eradicated immediately, as is usual in normal people, no disease results. If colonisation occurs, it may be short or long term. The pattern of disease is mostly determined by the host group (see next slide), with probably a component of the inoculum size contributing to invasive disease. However chronic disease if usually seen in patients with apparently normal immune systems.Exposure to individual Aspergillus spores or conidia is almost constant. If eradicated immediately, as is usual in normal people, no disease results. If colonisation occurs, it may be short or long term. The pattern of disease is mostly determined by the host group (see next slide), with probably a component of the inoculum size contributing to invasive disease. However chronic disease if usually seen in patients with apparently normal immune systems.

25. The categories of invasive aspergillosis of the the airways and whether the disease is likely to be invasive or not, is described.The categories of invasive aspergillosis of the the airways and whether the disease is likely to be invasive or not, is described.

26. This drawing shows the appearance of the tracheobronchial tree at autopsy. It shows white plaques of fungal growth on the trachea and main bronchi. This is the first case of Aspergillus tracheobronchitis reported, and emphasises that some cases are in ‘normal’ patients, ie no immunosuppression.This drawing shows the appearance of the tracheobronchial tree at autopsy. It shows white plaques of fungal growth on the trachea and main bronchi. This is the first case of Aspergillus tracheobronchitis reported, and emphasises that some cases are in ‘normal’ patients, ie no immunosuppression.

27. Aspergillus tracheobronchitis Review of 58 patients in literature for normal and immuno compromised patients - risk factors % None (ie normal) 25 Heart / Lung transplant 18 Solid tumour 15 BMT 13 Leukaemia 13 HIV/AIDS 8 Other 8

28. Pulmonary aspergilloma is a mass caused by a fungal infection that usually grows in pre-existing lung cavities. An aspergilloma can also form in the maxillary sinus, often related to dental work on the upper teeth. In the lung, it must be distinguished from chronic cavitary pulmonary aspergillosis (complex aspergilloma), characterised by multicavity disease. Aspergillomas are single, and always have substantial pleural thickening over them, and these patients have positive serum aspergillus precipitins.Pulmonary aspergilloma is a mass caused by a fungal infection that usually grows in pre-existing lung cavities. An aspergilloma can also form in the maxillary sinus, often related to dental work on the upper teeth. In the lung, it must be distinguished from chronic cavitary pulmonary aspergillosis (complex aspergilloma), characterised by multicavity disease. Aspergillomas are single, and always have substantial pleural thickening over them, and these patients have positive serum aspergillus precipitins.

29. Chronic pulmonary aspergillosis – pre-existing 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 Studied symptoms in 18 patients .Chronic cavitary pulmonary aspergillosis (complex aspergilloma),is characterised by multicavity disease.Studied symptoms in 18 patients .Chronic cavitary pulmonary aspergillosis (complex aspergilloma),is characterised by multicavity disease.

30. Chronic pulmonary aspergillosis - presentation 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%)

31. 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)

32. Close up view of right upper-lobe of the lung in a 45 year old man who smoked cigarettes showing an ill-defined shadow behind the clavicle and additional abnormalities inferior to this in the right upper-lobe. The lesions were considered to be possibly malignant and surgically resected. The specimen revealed a 2cm cavity with necrotic contents associated with local bronchiectasis and thickening of the parietal pleura. The lung specimen showed severe emphysema with fibrosis. Microscopically, the cavity was in an area of cystic bronchiectasis with erosion of the mucosa. The cavity contents were purulent and contained a fungus ball without apparent invasion or tissue eosinophilia. A fibrosing (necrotising) granuloma was seen superior to the cavity. Stains for acid fast organisms and cultures for TB were negative and fungal cultures were not done.. B. June 1992 Recurrence of disease. Chest radiograph demonstrating cavitary invasive aspergillosis. Despite resection of part of the right upper-lobe, invasive aspergillosis recurred six months later. Sputum cultures grew A. fumigatus and Aspergillus antibodies were detected in serum. He responded to itraconazole but subsequently progressed.  Close up view of right upper-lobe of the lung in a 45 year old man who smoked cigarettes showing an ill-defined shadow behind the clavicle and additional abnormalities inferior to this in the right upper-lobe. The lesions were considered to be possibly malignant and surgically resected. The specimen revealed a 2cm cavity with necrotic contents associated with local bronchiectasis and thickening of the parietal pleura. The lung specimen showed severe emphysema with fibrosis. Microscopically, the cavity was in an area of cystic bronchiectasis with erosion of the mucosa. The cavity contents were purulent and contained a fungus ball without apparent invasion or tissue eosinophilia. A fibrosing (necrotising) granuloma was seen superior to the cavity. Stains for acid fast organisms and cultures for TB were negative and fungal cultures were not done.. B. June 1992 Recurrence of disease. Chest radiograph demonstrating cavitary invasive aspergillosis. Despite resection of part of the right upper-lobe, invasive aspergillosis recurred six months later. Sputum cultures grew A. fumigatus and Aspergillus antibodies were detected in serum. He responded to itraconazole but subsequently progressed.  

33. This CT scan of the thorax illustrates the formation of multiple cavities, without any fungal ball formation (aspergilloma) developing in ‘normal’ (emphysematous) lung a non immunocompromised patient. The appearances are reminiscent of tuberculosis and coccidioidomycosis. Pleural involvement is seen as well.This CT scan of the thorax illustrates the formation of multiple cavities, without any fungal ball formation (aspergilloma) developing in ‘normal’ (emphysematous) lung a non immunocompromised patient. The appearances are reminiscent of tuberculosis and coccidioidomycosis. Pleural involvement is seen as well.

34. Young 33 year-old female smoker (no other risk factors) spontaneously developed these bilateral pulmonary shadows that then cavitated, and the cavities expanded in size of the next 3 years. Young 33 year-old female smoker (no other risk factors) spontaneously developed these bilateral pulmonary shadows that then cavitated, and the cavities expanded in size of the next 3 years.

38. Case 004 (also published) in case histories section.Case 004 (also published) in case histories section.

39. Case 004 (also published) in case histories section.Case 004 (also published) in case histories section.

40. Case 004 (also published) in case histories section.Case 004 (also published) in case histories section.

41. Mannose-binding lectin (MBL) is a serum protein of hepatic origin belonging to a family of Ca2+-dependent collagenous lectins, most of which are components of the innate immune system). MBL is able to bind through multiple sites to various carbohydrate structures and, on binding to its ligands, is able to activate complement in an antibody- and C1q-independent manner using MBL-associated serine protease 1 (MASP-1) and MASP-2). In humans, low levels of MBL are caused by one of three structural mutations found within exon 1 of the MBL gene. One - M54 - effects 16% of caucasians.Mannose-binding lectin (MBL) is a serum protein of hepatic origin belonging to a family of Ca2+-dependent collagenous lectins, most of which are components of the innate immune system). MBL is able to bind through multiple sites to various carbohydrate structures and, on binding to its ligands, is able to activate complement in an antibody- and C1q-independent manner using MBL-associated serine protease 1 (MASP-1) and MASP-2). In humans, low levels of MBL are caused by one of three structural mutations found within exon 1 of the MBL gene. One - M54 - effects 16% of caucasians.

42. Mannose 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

43. CCPA 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) Mannose binding protein genetic defects are common in CCPA patients (47-72%). Less common are defects in superfactant A2 protein, one of the 5 main surfactants found in the lung. Cytokine defects are also likely.Mannose binding protein genetic defects are common in CCPA patients (47-72%). Less common are defects in superfactant A2 protein, one of the 5 main surfactants found in the lung. Cytokine defects are also likely.

44. CLASSIFICATION OF ASPERGILLOSIS Persistence without disease - colonisation of the airways or nose/sinuses Exposure to individual Aspergillus spores or conidia is almost constant. If eradicated immediately, as is usual in normal people, no disease results. If colonisation occurs, it may be short or long term. The pattern of disease is mostly determined by the host group (see next slide), with probably a component of the inoculum size contributing to invasive disease.Exposure to individual Aspergillus spores or conidia is almost constant. If eradicated immediately, as is usual in normal people, no disease results. If colonisation occurs, it may be short or long term. The pattern of disease is mostly determined by the host group (see next slide), with probably a component of the inoculum size contributing to invasive disease.

45. ALLERGIC BRONCHOPULMONARY ASPERGILLOSIS – Key diagnostic criteria Asthma Blood eosinophilia (>1,000 / cu mm) History of pulmonary infiltrates Central bronchiectasis Key criteria for the diagnosis of ABPAKey criteria for the diagnosis of ABPA

46. Remarkably oedematous bronchial mucosa, as seen in ABPA. Before and after image shows remarkable obstruction by a mucus plug, subsequently removed by bronchoscopy.Remarkably oedematous bronchial mucosa, as seen in ABPA. Before and after image shows remarkable obstruction by a mucus plug, subsequently removed by bronchoscopy.

47. There was a long mucous plug in the anterior segment of the RUL. Half of this was aspirated and sent for microscopy and culture. The second half "fell into" the bronchus intermedius (which feeds the right middle lobe) and was only partially aspirated. There was a long mucous plug in the anterior segment of the RUL. Half of this was aspirated and sent for microscopy and culture. The second half "fell into" the bronchus intermedius (which feeds the right middle lobe) and was only partially aspirated.

48. ABPA - CT showing central bronchiectasis Central bronchiectasis is very characteristic of ABPA. It is also seen in cystic fibrosis. It is one of the diagnostic criteria for the disease.Central bronchiectasis is very characteristic of ABPA. It is also seen in cystic fibrosis. It is one of the diagnostic criteria for the disease.

49. ABPA and surfactant 5 surfactant proteins in man, SPA1, SPA2, SPB, SPC and SPD – all ‘collectin’ family Structures of 2 important surfactants.Structures of 2 important surfactants.

50. 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 Examples of polymorphisms conferring susceptibility to ABPA in India.Examples of polymorphisms conferring susceptibility to ABPA in India.

52. These panels are the same/adjacent sections stained with H&E (left) and an immunofluorescent antibody to MBP. It shows the sinus cavity full of mucus which contains substantial amount of MBP. These panels are the same/adjacent sections stained with H&E (left) and an immunofluorescent antibody to MBP. It shows the sinus cavity full of mucus which contains substantial amount of MBP.

58. This study shows a high frequency of house dust allergy irrespective of the severity of asthma, a decreasing frequency of cat allergy with worsening asthma (perhaps these people stop keeping cats at home), and a higher frequency of mould (Cladosporium, Aspergillus, Alternaria and Penicillium) allergy as asthma gets measurably worse. Asthma is assessed based on FEV1 (forced expiratory volume in 1 second) a measure how open the airways are.This study shows a high frequency of house dust allergy irrespective of the severity of asthma, a decreasing frequency of cat allergy with worsening asthma (perhaps these people stop keeping cats at home), and a higher frequency of mould (Cladosporium, Aspergillus, Alternaria and Penicillium) allergy as asthma gets measurably worse. Asthma is assessed based on FEV1 (forced expiratory volume in 1 second) a measure how open the airways are.