Opportunistic fungal agents: Major fungal agents cause respiratory infections Pneumocystis jirovecii pneumonia Zygomycoses Aspergillosis Penicillosis. Fungi causing systemic mycoses: Blastomyces dermatitidis Histoplasma capsulatum Paracoccidioides brasiliensis Coccidioides immitis. Yeast: Cryptococcus neoformans Recently, the taxonomy of Pneumocystis has been changed (2002). Once thought to be a protozoan, now under fungus based on nucleic acid sequence studies. Taxonomists renamed the human species of Pneumocystis as Pneumocystis jirovecii. Two known species: P. carinii & P. jirovecii Pneumocystis pneumonia is one of the common opportunistic infections in AIDS Pneumocystis exists in cyst and trophozoite forms. The Cysts - found in the environment; in human tissues, both cysts and trophozoites (containing 4–8 sporozoites) are found. Once inhaled, the cysts are carried to – the lungs - transform into trophozoite Trophozoites induce - inflammatory response – recruitment of plasma cells -frothy exudate - also called plasma cell pneumonia Infection is transmitted by respiratory droplets In immunocompetent individuals: Asymptomatic In immunocompromised patients: Fatal pneumonia Specimens: Induced sputum, BAL or lung biopsy Microscopy Trophozoites can be demonstrated by Giemsa, toluidine blue, Grocott’s methenamine silver stain The cyst wall stains black with methenamine silver stain The organism cannot be cultured Serology Complement fixation test & Latex agglutination test Histopathological examination of lung tissue - reveals cysts. Gomori’s methenamine silver (GMS) staining method-demonstrate the cysts of P. jirovecii. Cysts – black-colored crushed ping-pong balls against the green background Histopathological examination of lung tissue - reveals cysts. Gomori’s methenamine silver (GMS) staining method-demonstrate the cysts of P. jirovecii. Cysts – black-colored crushed ping-pong balls against the green background Histopathological examination of lung tissue - reveals cysts. Gomori’s methenamine silver (GMS) staining method-demonstrate the cysts of P. jirovecii. Cysts – black-colored crushed ping-pong balls against the green background Radiology: Chest X-ray - classical finding of bilateral diffuse infiltrates. CT of the lung - ground-glass opacities at the early stage. Atypical manifestations - nodular densities, cavitary lesions PCR - developed for detection of P. jirovecii specific genes Detection of 1, 3 β-D-glucan in serum Cotrimoxazole (trimethoprim/sulfamethoxazole) - drug of choice for Pneumocystis pneumonia. Given for 14 days in non-HIV patients and 21 days in patients with HIV. Also the recommended drug for primary and secondary prophylaxis in patients with HIV Life-threatening infections caused by aseptate fungi belonging to the phylum Zygomycota 1. Order Mucorales (causes mucormycosis) Rhizopus (R. arrhizus and R. microsporus) Mucor racemosus, Rhizomucor pucillus Lichtheimia corymbifera , Apophysomyces elegans 2. Order ento

1. FUNGAL INFECTIONS
OF RESPIRATORY
TRACT
Dr. Mayuri Bhise
AIIMS RAJKOT

2. Opportunistic fungal agents: Major fungal
agents cause respiratory infections
 Pneumocystis jirovecii pneumonia
 Zygomycoses
 Aspergillosis
 Penicillosis.

3. CONT…
Fungi causing systemic mycoses:
 Blastomyces dermatitidis
 Histoplasma capsulatum
 Paracoccidioides brasiliensis
 Coccidioides immitis.
 Yeast: Cryptococcus neoformans

4. PNEUMOCYSTIS PNEUMONIA
 Recently, the taxonomy of Pneumocystis has been changed (2002).
 Once thought to be a protozoan, now under fungus based on nucleic acid
sequence studies.
 Taxonomists renamed the human species of Pneumocystis as Pneumocystis
jirovecii.
 Two known species: P. carinii & P. jirovecii
 Pneumocystis pneumonia is one of the common opportunistic infections in
AIDS

5. Pathogenesis
 Pneumocystis exists in cyst and trophozoite forms. The
 Cysts - found in the environment; in human tissues, both cysts and
trophozoites (containing 4–8 sporozoites) are found.
 Once inhaled, the cysts are carried to – the lungs - transform into trophozoite
 Trophozoites induce - inflammatory response – recruitment of plasma cells -
frothy exudate - also called plasma cell pneumonia

6. Mode of Transmission
 Infection is transmitted by respiratory droplets
 In immunocompetent individuals: Asymptomatic
 In immunocompromised patients: Fatal pneumonia

7. Laboratory Diagnosis
 Specimens: Induced sputum, BAL or lung biopsy
 Microscopy
 Trophozoites can be demonstrated by Giemsa, toluidine blue,
Grocott’s methenamine silver stain
 The cyst wall stains black with methenamine silver stain
 The organism cannot be cultured
 Serology
 Complement fixation test & Latex agglutination test

8. Laboratory Diagnosis
 Histopathological examination of lung tissue - reveals cysts.
 Gomori’s methenamine silver (GMS) staining method-demonstrate the cysts of
P. jirovecii.
 Cysts – black-colored crushed ping-pong balls against the green background

9. CONT…
Pneumocystis jirovecii: (a) X-ray shows interstitial lung infiltrates and (b)
Grocott’s methenamine silver stain— fungal elements in the alveolar
spaces

10. CONT…
 Radiology: Chest X-ray - classical finding of
bilateral diffuse infiltrates.
 CT of the lung - ground-glass opacities
at the early stage.
 Atypical manifestations - nodular densities,
cavitary lesions
CT scan of lungs suggestive of Pneumocystis
pneumonia (soft tissue density nodule with
reverse halo sign in the right upper lobe).

11. CONT…
 PCR - developed for detection of P. jirovecii specific genes
 Detection of 1, 3 β-D-glucan in serum

12. Treatment
 Cotrimoxazole (trimethoprim/sulfamethoxazole) - drug of choice for
Pneumocystis pneumonia.
 Given for 14 days in non-HIV patients and 21 days in patients with HIV.
 Also the recommended drug for primary and secondary prophylaxis in patients
with HIV

13. ZYGOMYCOSIS

14. Introduction
 Life-threatening infections caused by aseptate fungi belonging
to the phylum Zygomycota
1. Order Mucorales (causes mucormycosis)
 Rhizopus (R. arrhizus and R. microsporus)
 Mucor racemosus, Rhizomucor pucillus
 Lichtheimia corymbifera , Apophysomyces elegans

15. CONT….
2. Order entomophthorales (causes entomophthoromycosis)
 Basidiobolus ranarum
 Conidiobolus coronatus

16. Mucormycosis - Pathogenesis
 Spores found ubiquitously in the environment
 Transmission - inhalation, inoculation, or rarely ingestion of spores
 Spores - mycelial form which is angioinvasive – resulting in the spread of
infection

17. Predisposing factors:
 Conditions with increased iron load
 Diabetic ketoacidosis
 End stage renal disease
 Iron therapy or deferoxamine
 Defects in phagocytic functions

18. Mucormycosis – Clinical Manifestations
1. Rhino cerebral mucormycosis: Most common form, Orbital cellulitis,
proptosis, and vision loss
2. Pulmonary mucormycosis - in patients with leukaemia
3. Cutaneous mucormycosis
4. Gastrointestinal mucormycosis – necrotizing enterocolitis
5. Disseminated mucormycosis: Brain
6. Miscellaneous forms

19. Laboratory Diagnosis
 Histopathological staining or methenamine silver stain of
tissue biopsies shows broad aseptate hyaline hyphae with
wide-angle branching
 Culture on SDA at 25°C: White cottony woolly colonies with
tube filling growth (hence called lid lifters).
 Rhizopus - colonies become brown black later, due to
sporulation giving rise to salt and pepper appearance
Zygomycoses—histopathology of tissue
section shows aseptate broad hyphae
(Methenamine silver stain).
A B C D
A. Rhizopus colony on SDA shows
white cottony woolly colonies with black
spores (salt and pepper appearance);
B. Mucor on SDA—white cottony woolly
colonies

20. Cont….

21. Laboratory Diagnosis
 Microscopic appearance: LPCB mount - broad aseptate hyaline hyphae, from
which sporangiophore – sporangium – sporangiospores.
 Rhizoid: Root-like growth arising from hyphae
 Rhizopus - nodal rhizoids
 Lichtheimia - Internodal rhizoids
 Mucor - Rhizoids absent.

22. CONT…
C D
C. LPCB mount of colonies of Rhizopus shows sporangium with
rhizoid present;
D. LPCB mount of colonies of Mucor shows sporangium
(absence of rhizoid).
 Mucor show branched
sporangiophores arising
randomly along aerial
mycelium; rhizoids are absent
 Rhizopus have rhizoids and
sporangiophores, which arise
in groups directly above the
rhizoids

23. CONT…
A. Rhizopus; B. Lichtheimia; C.
Mucor

24. Treatment of Zygomycoses
 Amphotericin B deoxycholate - drug of choice for all forms of mucormycosis.
 Posaconazole or Isavuconazole - given alternatively.
 For mild localized skin lesions in immunocompetent patients, which can be
removed surgically.

25. Entomophthoromycosis
 Subcutaneous lesions produced by members of the order Entomophthorales,
i.e. Conidiobolus and Basidiobolus.
 Latter is also associated with visceral involvement.

26. ASPERGILLOSIS

27. Introduction
 Aspergillosis refers to the invasive and allergic diseases caused by a hyaline
mold named Aspergillus.
 There are nearly 35 pathogenic and allergenic species of Aspergillus,
important ones being— A. fumigatus, A. flavus, and A. niger

28. Pathogenesis
 Widely distributed in nature - decaying plants
 Transmission – inhalation of airborne conidia.

29. Risk factors for invasive aspergillosis are:
 Glucocorticoid use (the most important risk factor)
 Profound neutropenia
 Neutrophil dysfunction
 Underlying pneumonia, chronic obstructive pulmonary disease, tuberculosis
or sarcoidosis
 Anti-tumor necrosis factor therapy.

30. Clinical Manifestations
Incubation period - 2 to 90 days.
 Pulmonary aspergillosis: A most common form
 Allergic bronchopulmonary aspergillosis (ABPA)
 Severe bronchial asthma
 Aspergilloma (fungal ball)
 Acute angioinvasive pulmonary aspergillosis
 Chronic cavitary pulmonary aspergillosis

31. CONT…
 Invasive sinusitis
 Chronic granulomatous sinusitis
 Maxillary fungal ball
 Allergic fungal sinusitis
 Cardiac aspergillosis: Endocarditis (native or prosthetic) and
pericarditis
 Cerebral aspergillosis: Brain abscess, haemorrhagic infarction, and
meningitis

32. CONT…
 Ocular aspergillosis: Keratitis and endophthalmitis
 Ear infection: Otitis externa
 Cutaneous aspergillosis
 Nail bed infection: Onychomycosis
 Mycotoxicosis

33. Laboratory Diagnosis - Direct
Examination
 Specimens - sputum and tissue biopsies
 KOH (10%) mount or histopathological staining of specimens - narrow septate
hyaline hyphae with acute angle branching
 Culture: SDA and incubated at 25°C
 Species identification - based on the macroscopic and microscopic (LPCB
mount) appearance of the colonies

34. Identification features of Aspergillus species
Aspergillus
species
Macroscopic appearance of
colony
Microscopic appearance of colony (LPCB mount)
A. fumigatus Colonies—smoky green, velvety to
powdery,
reverse is white
 Vesicle is conical-shaped
 Phialides are arranged in single row
 Conidia arise from upper third of vesicle
 Conidia are hyaline
A. flavus Colonies—yellow green, velvety,
reverse is white
 Vesicle is globular-shaped
 Phialides in one or two rows
 Conidia arise from upper two-third to entire vesicle
 Conidia are hyaline
A. niger Colonies—black, cottony type,
reverse is white
 Vesicle is globular-shaped
 Phialides in two rows
 Conidia arise from entire vesicle
 Conidia are black in color

35. Conidiation of various Aspergillus
species
A. A. fumigatus; B. A. flavus; C. A. niger

36. Aspergillus (colonies on SDA)
A. Aspergillus fumigatus; B. Aspergillus flavus; C. Aspergillus niger

37. Aspergillus microscopic picture (LPCB
mount)
A. Aspergillus fumigatus; B. Aspergillus flavus; C. Aspergillus niger

38. Laboratory Diagnosis - Antigen Detection
 β-d-Glucan antigen assay: Marker of invasive fungal infections
 Raised in most invasive fungal infection – invasive
aspergillosis
 Galactomannan antigen: Detected by ELISA in patient’s sera or
urine.

39. Laboratory Diagnosis - Antibody
Detection
 Useful for chronic invasive aspergillosis and aspergilloma, where the culture is
usually negative
 In allergic syndromes such as ABPA and severe asthma, specific serum IgE
levels are elevated.

40. Treatment of Aspergillosis
 For invasive aspergillosis—voriconazole is the drug of choice
 For ABPA—itraconazole is the drug of choice
 For single aspergilloma—surgery is indicated
 For chronic pulmonary aspergillosis—itraconazole or voriconazole
 For prophylaxis - Posaconazole is indicated.

41. PENICILLIOSIS

42. Clinical Significance
 Penicillium has more than 250 species, found as saprophytes in the
environment
 Penicillium marneffei - dimorphic fungus produces wart-like skin lesions
 Mycotoxicosis – toxins released by certain species of Penicillium, such as P.
cyclopium, P. verrucosum, and P.puberulum

43. Laboratory Diagnosis
 Invasive penicilliosis: endophthalmitis and endocarditis
 Superficial disease: otomycosis, keratitis and Onychomycosis
 Allergic disease: asthma and allergic pneumonitis

44. Laboratory Diagnosis
 P. marneffei - dimorphic fungus
 Other Penicillium - only as Molds, grow easily on SDA at 25°C
 Colonies - rapid growing, flat with velvety to powdery texture and greenish in
color

45. CONT….
 Microscopic appearance - LPCB
mount of the colonies
 Hyaline thin septate - conidiophore
and its branches - elongated metulae
- flask-shaped phialides originate – a
chain of conidia - brush border
appearance

46. PULMONARY
CRYPTOCOCCOSIS

47. PULMONARY CRYPTOCOCCOSIS
 Cryptococcus neoformans - capsulated yeast causes meningitis in HIV-infected
individuals.
 Acquired by inhalational route
 In immunocompetent individuals - lungs exhibit defense mechanisms.
 People with low immunity, pulmonary infection occurs first, followed by
dissemination through blood to distant sites such as CNS.

48. Cryptococcus neoformans var. neoformans:
(a) India ink preparation of spinal fluid showing yeast cells surrounded by a large
capsule and (b) mucoid colony

49. Treatment
 Amphotericin B
 5-fluorocytosine
 Imidazoles (miconazole, ketoconazole)
 Triazoles (itraconazole, fluconazole, voriconazole)
 Echinocandins (caspofungin, micafungin)

50. Q 1
A 45-year-old known HIV seropositive patient was admitted in the ICU as a case of
pneumonia. His CD4 count was 80 cells/mm. GMS stain smear from his broncho-alveolar
lavage (BAL) fluid showed cysts measuring 4–6 microns. What is the organism likely
associated with this disease?
a) T. gondii
b) P. jirovecii
c) H. nana
d) C. sinensis

51. Q 2
A 55-year-old known asthmatic farmer was admitted to evaluate for chronic productive
cough. On admission, he had a high-grade fever, and his chest X-ray showed lobular
infiltrates. On Gram staining, his sputum showed numerous eosinophils and fungal
filaments with septate hyphae showing 45° angle branching. Name the organism associated
with this infection:
 a) Aspergillus fumigatus
 b) Candida albicans
 c) Mucor
 d) Cryptococcus

52. Q3
A high case fatality due to vascular invasion in an immunocompromised host is the
characteristic feature of:
 a) Candidiasis
 b) Mucormycosis
 c) Blastomycosis
 d) Sporotrichosis

53. Q 4
The drug of choice for Pneumocystis pneumonia is:
a. Amphotericin B
b. Flucytosine
c. Cotrimoxazole
d. Voriconazole

54. Q 5
 Azole active against mucormycosis is:
a. Voriconazole
b. Fluconazole
c. Itraconazole
d. Posaconazole