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  • 1. INFECTIOUS DISEASE OF PATHOLOGY Fungus Dr. Naila Awal (Postgraduate student)
  • 2. Definition • Fungi are the eukaryotic, saprophytic, parasitic micro organism, unicellular or multicellular having chitin in their cell wall & ergosterol & zymosterol in their cell membrane.
  • 3. Classification Morphological classification1) Mold- fungi which are structurally long filamentous & multicellular & form a network of hyphae/mycelium. • Ex- Dermatophytes Aspergillus Mucor --Aseptate Septate
  • 4. 2)Yeast- unicellular round/oval like fungi reproduction by budding. 3) Yeast like fungusFungus that reproduction by budding, but bud fails to separate from parent cell & ultimately form a structure that looks like a chain of elongated cell. They form pseudohyphae. • Ex- Cryptococcus neoformans. • Ex- Candida
  • 5. • 4) Dimorphic fungus • Fungi that can remain either mold/ yeast form depending upon the temperature of the environment. • Ex- Histoplasma capsulatum • 37C-->change to yeast form • 25C --> mold form
  • 6. Depending upon the site of infection 1) Superficial mycosesThose fungal infection ,caused by the fungus that lies on the surface of the stratum corneum. 2) Cutaneous mycosescaused by the fungus that liberate keratinase enzyme & therefore can invade deep into keratin layer of skin & also nail &hair. • ExMalassezia furfur --> • Pityriasis versicolor • • ExDermatophytes--> dermatophytoses (Ring worm)
  • 7. 3) Sub cutaneous mycoses- caused by the fungus that lies on the surface of the stratum corneum. 4)Deep mycoses- caused by the fungus that infect internal organ • Ex• Rhinosporidium seeberi– Rhinosporidioses. • Ex• Candida–Candidiasis. • Histoplasma capsulatumHistoplasmosis.
  • 8. Superficial mycoses (Pityriasis versicolor) • Disease- Malassezia furfur • Clinical manifestationLesions are macular, non-inflammatory, well demarcated & hypopigmented.
  • 9. Lab diagnosis • Sample- Skin scrapping • Lab procedure• The scrapping material is placed on glass slide few drops of KOH is added cover slip wait for 20 mins. • Observation-Cluster of rounded yeast cell & short ,stout, curve, septate hyphae.
  • 10. Cutaneous Mycoses (Dermatophytes) • Genus1)Trichophyton- Nail & Hair 2)Epidermatophyton- Nail 3)Microsporum- Hair C/FSkin-->Ring like lesion. Ring- Scaly, hypopigmented & inflammatory. Nail yellow, brittle & thick. Hair brittle.
  • 11. Lab diagnosis • SampleSkin- Skin scrapping Hair- Hair clipping. If infection present in scalp, then skin adjoining scalp is taken. Nail Nail shaving at the margin between healthy & unhealthy area & from the deeper portion of nail.
  • 12. Lab procedure 1)The material is placed on glass slide few drops of KOH is added cover slip wait for 30 mins (skin)/10mins (hair)/several hours. • Observation- long thin, branched & septate hyphae. 2) Culture- Sabouraud's dextrose agar media25-28 C for 3-4 weeks. • Colony morphology- colony is picked up place on glass slide Lacto phenol cotton blue dye is added cover slip examine under microscope.
  • 13. • Spiral Hyphae Tinea mentagrophytes • Anthar hyphaeTinea schoenlenii • Macroconidia Microsporum • Microconidia • Trichophyton rubrum
  • 14. Sub cutaneous mycoses Mycetoma • Definition- It is a granulomatous disease of • • • • subcutaneous tissue caused by various fungi & bacteria where there is gradual destruction of tissue leading to loss of function of the affected area. Disease- Maduromycosis /Madura foot Actinomycetoma when madura foot is caused by bacteria. Eumycetoma when madura foot is caused by fungus. Site- 1)Foot most common site 2)Lower extremities 3)Back
  • 15. • Clinical manifestation1)Suppuration 2)Abscess formation 3)Granuloma 4)Tumor like swelling containing multiple draining sinus.
  • 16. Pathogenesis • Local trauma of skin Micro organism enters to the body Inflammatory response Granulomatous inflammation.
  • 17. Lab diagnosis • Sample- muco pus • Lab procedure• Muco pus is taken in 3 separate glass slides. • 1st slide- 1 drop of normal saline is added cover slip viewed by hand glass • Observation• If granules are black- Madurella grisea red-Madurella mycetomatis yellow white-Pseudallescheria boydii
  • 18. 2nd glass slide • Add few drops of 20% KOH in muco pus wait for 10 mins • Observation• If fine fillamentous branch with no chlamydospore Bacteria • If thick septate branched hyphae with chlamydospore Fungus
  • 19. 3rd glass slide• Granules are crushed on a slide Z-N stain decolorized with 1% H2SO4 Red branching filament Nocardia
  • 20. • HistopathhologySuppurative granuloma• Center abundant polymorphonuclear cells & granules • Surrounded by --lymphocytes, plasma cell, histiocytes & fibrosis.
  • 21. Rhinosporidiosis • Definition- It is a chronic granulomatous disease characterized by production of large polyp / wart like lesion. • Causative organism- Rhinosporidium seeberi • Site- Common site –Nose-78% Nasophaynx-68% Tonsil-3% Eye-1%.
  • 22. Pathogenesis Local trauma Fungus is inoculated into the mucosal epithelium of nasal cavity Replication of fungus Hyperplasic growth of host tissue & immune response Granulomatous disease
  • 23. Morphology • The lesions are polypoid, reddish & granular. • May be multiple & pedunculated. • Highly vascular & bleed on touch. M/E• Papillomatous hyperplesia of nasal mucosa. • Multiple mature & immature cyst (sporangia) are packed with spores. • Infiltration of chronic inflammatory cells into normal tissue.
  • 24. Deep Mycoses • ClassificationDeep mycoses Primary deep mycoses Opportunistic mycoses (Fungal infection that infect healthy Individual) Ex Coccidioidomycosis Paracoccidioidomycosis Histoplasmosis Blastomycosis (Fungal infection that infect immunosuppressive individual) ExCandidiasis Cryptococcosis Aspergillosis Mucormycosis
  • 25. Histoplasmosis • Causative organism- Histoplasma capsulatum. • Pathogenesis• Source of infection- Soil • Mode of transmission- Inhalation. Through inhalation macro & microconidia enters into lung engulf by alveolar macrophage through macrophage they spread to the various tissue. (Liver, spleen, lymph node)
  • 26. • C/F1)Asymptomatic 2)Pulmonary histoplasmosis- fever, dry cough. More severe case- granulomatous lesion in lung. 3)Localized lesion in extrapulmonary sitemediastinum, adrenal,liver, meninges. 4) Disseminated histoplasmosis- Immunosuppressed person.
  • 27. Lab diagnosis • Specimen1)Pulmonary histoplasmosis- Sputum. 2)Buffy coat of blood. 3)Bone marrow aspirate. 4)Biopsy specimen from different internal organ. • Lab procedure• 1) Direct microscopy- Specimen is taken on the glass slide Giemsa staining Microscopic examination. • Findings- Yeast like fungus within the cytoplasm of macrophage.
  • 28. 2)CultureSabouraud’s dextrose agar media- incubate at 37 C for 4 weeks. Observation of colony morphologyhyphae with macro & micro conidia. 3) SerologyPatient’s serum-Anti histoplasma capsulatum . 4)SkinHistoplasmin skin test.
  • 29. 5) HistopathologyPulmonary histoplasmosisEpithelioid granuloma with central caseous necrosis, which coalesce to form areas of consolidation. Disseminated histoplasmosis-->Mononuclear phagocytes filled with fungal yeast. -->Epithelioid granuloma are not formed.
  • 30. Cryptococcosis • Definition- It is a true yeast surrounded by thick polysaccharide capsule. • Causative organism- Cryptococcus neoformans. • Nice to knowCapsulated bacteriaStreptococcus pneumoniae Klebsiella Haemophillus influenzae Neisseria meningitidis
  • 31. Mode of transmission- Inhalation of yeast . • Pathogenesis• It is asymptomatic / produce influenza like symptoms which resolves automatically in healthy person. But in immunocompromised person Through inhalation enters into lung via blood meninges cryptococcal meningitis.
  • 32. Lab Diagnosis • Specimen1)CSF 2)Sputum 3)Tissue sample Lab procedure1)Wet film microscopy- circular/oval yeast cell. 2)India ink preparation- CSF is centrifuged deposit is taken on glass slide1 drop of india ink is added cover slip examine under microscope. Observation- against dark background darkly illuminated single circular/oval yeast with budding.
  • 33. 3) Culture- Blood agar media- 37 C for 24 hours Saborauds dextrose agar media-37 C for 2 days. • 2 days after, colonies are picked up on glass slide examine under the microscope • ObservationSpherical yeast cell with budding. 4)Serological test- Anti cryptococcal Ab +
  • 34. Histopathological findings • CNS• In immunosuppressed personSoap bubble lesion- gelatinous masses of fungi in meninges or may expand the perivascular VirchowRobin space within the grey matter. • In non immunosuppressed personChronic granulomatous lesion composed of macrophage, lymphocyte & foreign body giant cell. Suppuration may also occur. • Lung- Solitary pulmonary granuloma.
  • 35. Aspergillosis • Causative agent- Aspergillus fumigatus. • Mode of transmission- By the inhalation of Aspergillus spore. • Spectrum of disease1) No infection- b/c alveolar macrophage engulf & destroy the conidia. 2)Person who have allergic to Aspergillus antigen, manifest as Allergic bronchopulmonary Aspergillosis (ABPA). 3)Person who have cavity to lung (due to TB, sarcoidosis), conidia after reaching the cavity they germinate & produce abundant hyphae in lung cavity. This clinical condition is known as Aspergilloma.
  • 36. • They manifest asHaemoptysis Dyspnoea Asthma
  • 37. 4)Person who are in steroid therapy (leukemia, bone marrow transplantation) inhale conidia produce severe manifestation of disease Invasive Aspergillosis which is clinically manifest as pneumonia. • After lung infection hyphae invade blood vessels haematogenous spread to different internal organ Abscess (brain, liver, kidney).
  • 38. Lab diagnosis • Sample1)Sputum 2)Blood 3)Lung biopsy. • Lab procedure• 1) Gram staining- Septate hyphae with dichotomous branching. • 2)CultureBlood agar Sabouraud's dextrose agar media
  • 39. • Observation of colony- Aspergillus colonies are usually fast growing, white, yellow, yellow-brown, brown to black or shades of green. • 3) Serological test- Immunodiffusion tests for the detection of antibodies to Aspergillus species . • 4) Histopathology• AspergillomaProliferative mass of hyphae form fungal ball which lies freely within the cavities.  Surrounding inflammatory reaction may be sparse/chronic inflammation/ fibrosis.
  • 40. Invassive aspergillosis • 1) lungAspergillus form fruiting bodies (usually in lung cavities) & Septate filaments, branching at right angles (40 degrees)
  • 41. Mucormycosis Definition- It is an opportunistic infection caused by ‘bread mould fungi’ including- Mucor, Rhizopus, Abscidia & Cunninghamella. Predisposing factors• • • • • Neutropenia DM Corticosteroid use Iron overload Breakdown of cutaneous barrier as a result of burning, surgical wound/ trauma.
  • 42. Major route of infection- 1) Inhalation 2) Ingestion 3) Traumatic inoculation. Clinical presentation• 5 clinical form of Mucormycosis- Rhino cerebral, pulmonary, gastrointestinal, primary cutaneous & Disseminated. • Rhino cerebral MucormycosisInitial symptoms- acute sinusitis, congestion, purulent nasal discharge, fever, unilateral headache, peri orbital edema, proptosis, facial numbness, cranial nerve palsy.
  • 43. • Pulmonary MucormycosisFever, haemoptysis • Gastrointestinal Mucormycosis• Bowel perforation, peritonitis, GIT hemorrhage. • Severe immunocompromised person manifest as primary cutaneous lesion. • They do not harm in immunocompetent individual but infect immunosuppressed person.
  • 44. Pathogenesis • It is transmitted by air borne asexual spore. • After inhalation of spore Colonize into nasal sinus Orbit Brain Rhino cerebral Mucormycosis Lung Engulf by alveolar macrophage & oxidative killing
  • 45. Lab diagnosis • Specimen- Biopsy from suspicious areas. • Lab procedure1) Staining• H&E stain- often hard to see – GMS, PAS stains better Observation-Non septate, irregularly wide fungal hyphae with frequent right angle branching.
  • 46. 2) Histopathology- Common site• nasal sinus, lung &GIT Rhino cerebral Mucormycosis• Local tissue necrosis • Invade arterial wall • Penetrate peri orbital tissue. • Pulmonary Mucormycosis• Areas of hemorrhagic pneumonia with • Vascular thrombi & • Distal infraction.
  • 47. Candidiasis • Species1)Candida albicans. 2)Candida tropicalis. 3)Candida parapsilosis. • Features1) Dimorphic fungus• • 25 C-True hyphae. 37C-Pseudohyphae with yeast. 2) May produce chlamydospore- Sexual spore. 3) Causes opportunistic infection.
  • 48. • Predisposing factor of candidiasis1) Immunosuppression due to prolong use of – Anti cancer drug, steroid. 2)AIDS 3)Prolong use of antibiotic 4)DM 5)Pregnancy 6)Bone marrow transplant recipient 7)Leukaemia, lymphoma 8)Very young & very old age.
  • 49. Clinical spectrum of candidiasis• 1) Mucosal candidiasis• • • Oral thrush Candida esophagitis Vulvo vaginal candidiasis • • • • • • • 2) Cutaneous CandidiasisNail proper- Onychomycosis Nail fold- Paronychia Armpit/ web of the fingers & toes- Intertrigo Hair follicle- Folliculitis Penile skin- Balanitis Perineum of infant- Diaper rash
  • 50. 3) Chronic muco cutaneous candidiasis• It involves superficial skin & mucous membrane. 4)Invasive candidiasis-
  • 51. Oral thrush • Definition- It is the superficial fungal infection on the mucous membrane of oral cavity. • Seen inNewborn Debilitated people Children receiving oral steroid  HIV positive patient for asthma & following a course of broad spectrum antibiotics that destroy normal bacterial flora. • Morphology- They form grey-white dirty looking pseudo membrane, composed of matted organism & inflammatory debris. • Deep to the surface there is mucosal hyperemia & inflammation.
  • 52. Candida esophagitis • Commonly seen inAIDS patient Hematolymphoid malignancy. • Clinical presentationDysphagia Retrosternal pain. • Endoscopic findingsEsophageal mucosa-- plaque & pseudo membrane.
  • 53. Vaginal candidiasis • Common in• Women withDM Pregnancy Taking OCP • Clinical manifestationIntense itching Thick curd like discharge
  • 54. Invasive candidiasis • Definition- It is a candidiasis which is caused by blood borne dissemination of organisms into various tissue / organs. • Persons risk for developing candidiasis1) ICU patient 2) Surgical patient 3)Patient with central venous catheter 4) Immunocompromised person. 5) Very LBW infant
  • 55. • Common pattern/ Effect1) Kidney- Renal abscess 2) Heart- Myocardial abscess & endocarditis. 3)CNS- Brain micro abscess & meningitis 4) Liver- Hepatic abscess 5) Eye- Endopthalmitis Findings Inflammatory response  Occasionally granuloma.
  • 56. Lab diagnosis of candidiasis • Sample-->Swab -->Scrapping from superficial lesion -->Tissue biopsy -->Exudates -->Blood, CSF, Urine -->Material from I/V catheter
  • 57. Lab procedure 1) Wet film preparation- From Swab, exudates, CSF (centrifuge), urine (centrifuge) • FindingsSpherical/ oval yeast cell with budding.
  • 58. 2)StainingA) Gram staining- Gram + round/oval yeast cell with budding. B) Immuno fluorescence staining- calcoflor white stain. 3) CultureA) Sabouraud's dextrose agar media B) Blood agar media • 37 C for 48 hours in aerobic condition.
  • 59. Colony morphology• 2-4 mm in diameter, circular, white/creamy, soft with mucor odor. 4)Confirmation – Gram staining from the culture plate. • Gram + yeast with budding.
  • 60. 5)Germ tube test (Confirmatory test for Candida albicans) • 0.5ml human serum is taken in a test tube • Test fungal colony is added • Incubate at 37C for 4 hours • Then 1 drop of serum is collected & place on glass slide • Cover slip • Microscopic examination
  • 61. • Observation- Yeast like cell with finger like projection. • Comment+ for Candida albicans 7) Sugar fermentation test• It differs for different species.