DEEP FUNGAL INFECTIONS-Candida, Aspergillus

Morphology, pathology, microbiology

1. PRESENTER:Dr.SHWETHA.B MODERATOR:Dr.PRAKASH.C.J DEEP FUNGAL INFECTIONS

2. FUNGAI Fungi are eukaryotes which grow in two basic forms: YEASTS MOULDS/MOLDS

3. Dimorphic fungi Filamentous fungi unicellular fungus molds(Mycelia) yeasts depending on environmental conditions . saprophytic stage. parasitic stage Asexually (mitosis), by Yeasts reproduce by a of hyphal growth and tip process budding extension, or with the help of asexual spores . Sexual reproduction (meiosis) , produces sexual spores.

4. Reproduction in Fungi Asexual spores have number of morphological types:conidia,sporangiospores,arthrospores,and blastospores . The morphology of the asexual spores of fungi is an important identification characteristic. Sexual spores:zygospores,ascospores,and basidiospores .

5. Fungal infections also called mycoses, are of four major types: • Superficial and cutaneous mycoses are common and limited to superficial or keratinized layers of skin, hair, and nails. 1.Dermatophytoses 2. Tinea versicolour 3.Tinea Nigra 4.Piedra

6. Subcutaneous mycoses involve the skin, subcutaneous tissues,lymphatics and rarely disseminate systemically. Mycetoma Sporotrichosis Chromoblastomycosis Lobomycosis Endemic mycoses are caused by dimorphic fungi that can produce serious systemic illness in healthy individuals. Histoplasmosis Blastomycosis Coccidioidomycosis Paracoccidioidomycosis

7. Opportunistic mycoses can cause life-threatening systemic diseases in individuals who are immunosuppressed . Candidiasis Aspergillosis Cryptococcosis Zygomycosis Pneumocystosis Penicilliosis Marneffei

8. RISK FACTORS Most fungi are opportunist. Chemotherapeutic agents Irradiation Immunosuppressive agents Hyperalimmentation Use of broad spectrum antibiotics Malignancies Organ transplant Metabolic disease AIDS

9. SPECIMEN COLLECTION AND TRANSPORT Swabs are decidedly inferior swab fibers may be mistaken for hyphal elements. The best specimens for mycologic diagnosis are scrapings, curettings , aspirates, and lesion biopsies. Hair, nails, scalp, and skin scrapings should be sent to the laboratory in a clean dry container. Tissue samples should be sent in a sterile container with a small amount of sterile, preservative free saline or a transport medium to prevent drying of the sample for culture

10. DIAGNOSIS 4 appoaches to the diagnosis of mycotic diseases: Clinical Mycologic Mycological culture and histopathology should complement each other whenever possible,because few of fungai are not yet isolated in culture. Pathologic Immunologic

11. Pathologic: Microscopy: Wet preparation: briefly heat material under coverslip with 10% KOH. Stained preparation: H and E and methylene blue, lactophenol blue, periodic acid-Schiff (PAS), indian ink,calcofluor white stain etc. Mycologic : Culturing: Universal and selective medium- Sabouraud dextrose agar Main identifing structures are morphological, in particular the asexual and, if present, sexual reproductive structures.

12. Stains Diagnostic applications Special stains for fungi Gomori’s methenamine –silver Periodic acid- schiff Detecting most of fungi. Mucin stains Mayer’s mucicarmine Alcian blue Mucoid capsule of cryptococcus neoformans . Gram stains Actinomycetes Botryomycosis candida Giemsa stain Leishmania Pneumocystis carinii

13. IMMUNOHISTOLOGY Formaline fixed paraffin embedded tissue can be used. They help to confirm presumptive histopathologic diagnosis especially when atypical forms of fungus are present.

14. DEEP FUNGAL INFECTIONS 1. Subcutaneous 2. Endemic and 3. Opportunistic fungal infection

15. SUBCUTANEOUS MYCOSES MYCETOMA/MADURA FOOT/MADUROMYCOSIS: A mycetoma is a slowly progressive, localized and often disfiguring infection of the skin, soft tissues and bone produced by inoculation of various soil-dwelling fungi and filamentous bacteria.

16. Causal organisms: Fungi ( eumycetoma )- Madurella mycetomatis , M. grisea , Scedosporium apiospermum , Leptosphaeria senegalensis . Actinomycetes ( actinomycetoma )- Actinomadura madurae,Streptomyces somaliensis and Nocardia brasiliensis . Mycetoma usually occurs in the tropics among immunocompetent farmers and outdoor laborers whose skin is exposed to trauma. The foot is a common site of infection.

17. CLINICALLY : Single or multiple sinus tracts. discharge pus containing grains onto skin surface. PATHOLOGY: The colonies of organisms, called “grains,” resemble the “sulfur granules” of actinomycosis . Dermis and subcutis shows abscess containing grains in the centre and surrounded by neutrophils and an outer layer of granulomatous inflammation. Histological differentiation between the grains/granules formed by actinomycetes and fungi is crucial in determining treatment and prognosis.

18. EUMYCETOMA ACTINOMYCETOMA Grains Broad , septate,fungal hyphae Gram positive, branched,bacterial filaments. Black and white white The granules are surrounded by intense eosinophilic material ( Splendore-Hoeppli phenomenon) .

19. Mycologic : Culture on standard fungal media- Sabouraud dextrose agar, takes 4 weeks or longer to grow. Once isolated, the fungal species is identified using gross colony morphology, pigmentation, and micromorphologic characteristics of reproductive structures following sporulation . In cases in which sporulation does not occur: Physiologic tests - carbohydrate and nitrate utilization may aid in identification.

20. Terminal conidia of the  Madurella mycetomatis  fungus. ( Lactophenol cotton blue stain, 400×.) Treatment:Surgical excision and systemic antifungal chemotherapy.

21. CHROMOBLASTOMYCOSIS /CHROMOMYCOSIS Chronic, localized infection caused by pigmented ( dematiaceous ) fungi:botryomyces caespitosus,cladosporium carrionii,fonsecaea compacta . Clinically : locally spreading verrucous plaque or solid nodule at the site of traumatic implantation.

22. PATHOLOGY: Clusters of small, round, thick-walled, brown muriform , nonhyphal cells: sclerotic bodies ( medular bodies or “copper pennies”). They are grouped in the dermis which elicit a granulomatous and suppurative inflammatory reactions.

23. An additional characteristic is that mycetoma result in tissue necrosis, whereas chromoblastomycosis infections lead to excessive proliferation of host tissue.

24. SPOROTRICHOSIS Sporotrichosis is a chronic infection of the skin, subcutaneous tissues and regional lymph nodes caused by Sporothrix schenckii -dimorphic fungus. The mode of entry - usually traumatic implantation and inhalation, the disease produced is usually a localized systemic infection (vs. pulmonary). The patients at risk for sporotrichosis are “alcoholic rose gardener.”

25. Clinically: Ulceronodular lesions. The infection frequently spreads along subcutaneous lymphatic channels, resulting in a chain of similar nodular skin lesions- lymphocutaneous sporotrichosis

26. Pathology: The periphery of the nodules is granulomatous and the center is suppurative . Surrounding skin shows exuberant pseudoepitheliomatous hyperplasia. The yeast cells of Sporothrix are rarely seen in tissue, The sporothrix appears as spherical,oval or elongated (cigar shaped) yeast like cells Some yeasts are surrounded by an eosinophilic , spiculated zone(“ Splendore-Hoeppli substance”) and are termed “asteroid bodies”

27. The fungal cells often bear elongated buds with narrow based attachment .

28. Mycologic : Primary isolation media with cycloheximide incubated at 25° to 30° C. This organism produces two types of conidia: 1.Thinwalled hyaline conidia arranged as a rosette around the apex of a conidiophore , that arise at right angles from the hyphae . 2. Thick-walled, dark, sessile conidia attached directly to the hyphae . They may be arranged sympodially around an expanded vesicle at the tip of the conidiophore , producing an arrangement that described as a floret.

29. Sympodial conidia of  Sporothrix schenckii  are borne in clusters  at the tips of lateral conidiophores. ( Lactophenol cotton blue stain, 400×.)

30. LOBOMYCOSIS Rare, chronic infection of skin and subcutaneous tissue due to Loboa loboi Clinically : Papule or a small nodule, slowly proliferates to form extensive keloidal or verrucous lesions in dermis. Pathology: thick walled cells with uniform size and shape ,reproduce by budding in chains resembles string of pearls. Surrounding dermis shows dispersed epithelioid and gaint cell granulomatous inflammatory reaction.

31. SYSTEMIC MYCOSES HISTOPLASMOSIS Caused by Histoplasma capsulatum a dimorphic fungus. Bat nests, caves and soil beneath trees are foci of exposure. Portal of entry is by inhalation of conidia of dimorphic fugus .

32. PATHOPHYSIOLOGY: Histoplasmosis resembles tuberculosis in many ways. Primary infection begins with phagocytosis of microconidia by alveolar macrophages. As it grow, additional macrophages are recruited, producing an area of pulmonary consolidation. A few macrophages carry organisms first to hilar and mediastinal lymph nodes and then throughout the body. The organisms proliferate in cells until the onset of hypersensitivity and cell-mediated immune responses. Activated macrophages destroy the phagocytosed yeasts, forming necrotizing granulomas at sites of infection.

33. Clinically: The magnitude of the exposure and the immune status of the host influence the clinical manifestations of disease-asymptomatic infectious process to disseminated life-threatening disease Acute Pulmonary Infection: Flulike syndrome with high fever, chills, fatigue, cough, and pleuritic or retrosternal chest pain. Self limiting.

34. Granulomatous and Fibrosing Mediastinitis : Granulomatous disease is characterized by enlarged lymph nodes, which may obstruct the airways, pulmonary vessels, or esophagus. Fistulas can form within the lymph nodes and adjacent mediastinal structures. Fibrosing disease is a reaction that occurs in the mediastinum among individuals predisposed to an excessive response to Histoplasma antigens.

35. The cellular components of the granuloma largely disappear and the caseous material calcifies, forming a “ fibrocaseous nodule”

36. Chronic Pulmonary Histoplasmosis : This can occurs primarily in patients with chronic obstructive pulmonary disease (COPD). In this disease, calcification and cavitations may occur, which may mimic chronic pulmonary tuberculosis and pulmonary neoplasia .

37. Disseminated histoplasmosis : Dissemates - reticuloendothelial system can occur as part of acute pulmonary histoplasmosis , resulting in healed granulomas that often calcify, especially in the spleen. Clinically disseminated infection occurs in two classes of patients. The first group consists of individuals at the extremes of age, immune system that is incompletely developed . The second group comprises patients with recognized immunosuppressive diseases Progression of disease may be rapid or insidious

38. Compromised patients may develop an infection of the reticuloendothelial system, resulting in lymphadenopathy , hepatosplenomegaly , or thrombocytopenia . Involves adrenal cortex -hormonal insufficiency. Central nervous system- chronic meningitis, intracerebral granulomas , or both. Endovascular infection- endocarditis with large, bulky vegetations. Any part of the GI tract may be affected, and ulcerating lesions may suggest a neoplasm macroscopically.

39. Pathology: Facultative intracellular pathogen, H. capsulatum is found predominantly in macrophages. They are yeast cell, spherical to oval reproduce by budding basophillic cytoplasm is retracted to form rigid but thin poorly stained cell wall creating clear space/halo.

40. PAS and GMS stains are more sensitive. DD: The yeasts of H. capsulatum in caseating granulomas are distinctive to provide a presumptive diagnosis, but unusual granulomatous presentations of P. ( carinii ) jiroveci must be differentiated. Poorly encapsulated cryptococci can resemble yeast form of H.capsulatum,but cryptococci are carminophilics . Culture is generally required for disease confirmation.

41. Mycologic : Enriched agar, such as brain-heart infusion agar supplemented with sheep blood, which is incubated at 25° to 30° C. The colonies are fluffy and vary from white to buff-brown. Diagnostic are asexual forms include microconidia and macroconidia . The more characteristic macroconidia have roughened projections from the periphery of the conidia, a configuration referred to as tuberculate .

42. Tuberculate   macroconidia   with  microconidia   of  Histoplasma capsulatum  in culture. These structures are characteristic of the environmental mold form of this dimorphic fungus. ( Lactophenol cotton blue stain, 400×.)

43. Antigen detection, on the other hand, has been shown to be useful for the diagnosis and management of histoplasmosis . The sensitivity of urine antigen detection for diagnosis is greatest in patients with disseminated disease (up to 92%) or acute pulmonary histoplasmosis (75% to 80%). Treatment: Amphotericin B Ketoconazole -disseminated infections.

44. BLASTOMYCOSIS Blastomycosis is a chronic granulomatous and suppurative pulmonary disease, which is often followed by dissemination to other body sites, principally the skin and bone. The causative organism, Blastomyces dermatitidis , is a dimorphic fungus that grows as a mold in warm moist soil, rich in decaying vegetable matter. Blastomycosis is acquired by inhalation of spores from the soil.

45. Clinical: Chronic pulmonary blastomycosis - low-grade fever, weight loss, and localized pulmonary infiltrates. Dissemination of yeast from the lung most commonly results in cutaneous or skeletal infection. Cutaneous lesions often are hypertrophic or ulcerative and may be locally destructive.

46. Pathology: The characteristic histologic response to B. dermatitidis -mixture of acute inflammation with microabscess formation and granulomatous inflammation. In cutaneous lesions, pseudoepitheliomatous hyperplasia of the epidermis overlying the inflammation is characteristic. Yeast cells are present in microabscesses or within multinucleated giant cells can be demonstrated with H&E, but readily visible when stained with PAS or GMS.

47. yeasts of B. dermatitidis , which are spherical and 8–14 μm across, with broad-based bud ,thick double contoured wall and multiple basophillic nuclei in a central body .

48. Broad-based budding is diagnostic helps to differentiate it from histoplasma capsulatum and cryptococcus neoformans . Mycology: The lollipop appearance of the conidium on a conidiophore is characteristic of the environmental mold. ( Lactophenol cotton blue stain, 400×.) Treatment:Amphotericin B,2-hydroxystilbamidine

49. COCCIDIOIDOMYCOSIS Coccidioidomycosis is a chronic, necrotizing mycotic infection that clinically and pathologically resembles tuberculosis. The disease caused by Coccidioides immitis , dimorphic fungus. Portal of entry:inhalation of airborne arthroconidia Clinical:Most cases are mild and asymptomatic and are limited to the lungs and regional lymph nodes. Occasionally, C. immitis infections spread outside the lungs to produce lifethreatening disease.

50. Pathology: The tissue response to Coccidioides spp. is granulomatous , with and without caseation . Primary pulmonary coccidioidomycosis : Acute suppurative and granulomatous pneumonitis . Residual pulmonary coccidioidal nodule / coccidioidoma : is a peripheral sharply circumscribed and centrally necrotic granuloma Few of these nodules are centrally cavitated

51. Developing spherules are typically found in macrophages and multinucleated giant cells. Endospores (2 to 5 µm) within the spherules (up to 250 µm) and developing spherules have a large range of sizes.

52. Spherules of Coccidioides spp. in tissue showing release of  endospores . which, in turn, mature to become spherules. ( Gomori’s methenamine silver stain, 1000×.) Spherules of C.immitis should be distinguished from sporangia of rhinospoidium seeberi by their inherent brown color,size,wall thickness and morphology of endospores .

53. Arthroconidia  of  Coccidioides  species. Alternating barrel-shaped arthroconidia are separated by thin-walled, empty disjunctor cells within portions of the hyphae . ( Lactophenol cotton blue stain, 400×.) Treatment: Systemic antifungal chemotherapy.

54. OPPORTUNISTIC MYCOSES Candidiasis Many Candida species are endogenous human flora, well adapted to the human body. However, they can cause disease when host defenses are compromised. Usually C. albicans , are the most frequent cause of human fungal infections.

55. Pathogenesis: Candida can shift between different phenotypes. Phenotypic switching involves regulation of phase-specific genes and provides a way for Candida to adapt to changes in the host environment. Enzymes that contribute to invasiveness: Aspartyl proteinases : which may promote tissue invasion by degrading extracellular matrix proteins Catalases:which may enable the organism to resist oxidative killing by phagocytic cells.

56. Candida produces adhesins that mediate adherence to host cells and contribute to virulence. These adhesins include Integrin -like protein, which binds to fibrinogen, fibronectin , and laminin ; Protein that binds to epithelial cells; and Several agglutinins that bind to endothelial cells or fibronectin .

57. The ability of C. albicans to grow as biofilms also contributes to its capacity to cause disease. On implanted medical devices they escape from immune responses and antifungal drug therapy. Neutrophils , macrophages and TH17 cells are important for protection against Candida infection. Phagocytosis and oxidative killing. Filamentous forms, but not yeast, can escape from phagosomes and enter the cytoplasm and proliferate.

58. β-1,3-glucan expressed by the yeast engages dectin on dendritic cells induces IL-6 and IL-23 production, which promotes TH17 responses. The TH17 responses elicited by Candida are responsible for recruiting neutrophils and monocytes

59. Clinical: Oral Candidiasis ( Thrush): Most commonly  candidiasis  takes the form of a superficial  infection on mucosal surfaces of the oral cavity.  Florid  proliferation of the fungi creates gray-white, dirty-looking  pseudomembranes  composed of matted organisms and inflammatory debris.  Deep to the surface, there is mucosal hyperemia  and inflammation.  

60. Gastrointestinal Candidiasis : Most frequently as esophagitis and less commonly as gastritis Commonly seen in AIDS patients and in those with hematolymphoid malignancies.  They present with dysphagia and retrosternal pain. Endoscopy demonstrates white plaques and pseudomembranes  resembling oral thrush on the esophageal mucosa. The diagnosis can be made by brush cytology.  

61. Candida vaginitis :   It is common in women who are diabetic,pregnant,or on oral contraceptive pills. It is associated with intense itching and a thick, curdlike discharge. Cutaneous candidiasis :   Present in many different forms,including infection of the nail proper( onychomycosis ),nail folds ( paronychia ),hair follicles( folliculitis ), moist,intertriginous skin,such as armpits or webs of the fingers and toes ( intertrigo ),and penile skin( balanitis ). 

62. Invasive candidiasis :   Is caused by bloodborne dissemination of organisms to various tissues or organs.Common patterns include:   (1)Renal  abscesses,  (2)Myocardial abscesses and endocarditis ,  (3) Brain microabscesses and meningitis,    (4) Endophthalmitis ,  and  (5)Hepatic abscesses.

63. Pathology: The tissue response to Candida is regularly purulent, resembling lesions of bacterial infection with abscess formation. On occasion, granulomatous . In tissue, most Candida organisms appear as mats of budding yeasts measuring 3 to 5 µm in diameter mixed with pseudohyphae . C.glabrata , produces the smaller-sized yeast cells , does not produce pseudohyphae and must be differentiated from Histoplasma in tissue. Vascular invasion, is uncommon in yeast infection. Yeast more easily recognized with GMS or PAS.

64. Filamentous  extensions  from  the  periphery  of  colonies  of  Candida albicans  are known colloquially as “feet.” Thickwalled asexual spores ( Chlamydoconidia )that occur at the ends of pseudohyphae are characteristic for this species and for Candida dubliniensis . (Cornmeal agar plate, 400×.)

65. ASPERGILLOSIS The term aspergillosis describes infections caused by moulds belonging to the genus Aspergillus -Hyaline fungus. Aspergillus fumigatus is the most common pathogenic species of the fungus. Saprotrophic , found in decaying organic matter. Acquired by inhalation of spores. The risk factors are neutropenia and use of corticosteroids.

66. Pathogenesis: Air borne conidia, Reach alveoli, alveolar macrophages recognize Aspergillus through TLR2 and the lectin dectin1, which recognizes β-1,3- glucan . TLRs can recognize products of the fungal hyphae and trigger the release of pro-inflammatory mediators- TNF-α, IL-β, and chemokines . Neutrophils produce reactive oxygen intermediates that kill hyphae . Virulence factors- adhesins , antioxidants, enzymes, and toxins.

67. Clinical: Otomycosis : caused by Aspergillus niger or A. fumigatus species complex. Pain, decreased hearing, and a discharge are accompanied by a fluffy green or black growth in the ear canal. Ocular aspergillosis : Keratomycosis usually follows trauma to the eye, if treated with topical steroids. Pain and blurring of vision follow the traumatic episode, and if not treated, the infection may extend into the anterior chamber.

68. Sinus Infection: Invasive fungal sinusitis can be classified as an acute fulminant invasive disease or as a chronic indolent invasive disease . Cutaneous Infection : Aspergillus spp. may infect the skin after dissemination from u pulmonary, or the primary infection after direct cutaneous exposure to a contaminated source . Dissemination from another primary source in the suppressed patient is difficult to treat and generally results in death. This condition is usually recognized by multiple lesions appearing over different areas of the body.

69. Primary Pulmonary Infection: Primary pulmonary aspergillosis is the most common invasive mold infection in immunocompromised patients. Less invasive and noninvasive diseases that affect the lungs such as 1. Aspergilloma (fungus ball), 2.Chronic necrotizing pulmonary aspergillosis , 3.Semi-invasive aspergillosis , 4.Chronic invasive pulmonary aspergillosis , 5.Symptomatic pulmonary aspergillosis , and 6.Aspergillus pseudotuberculosis

70. The predilection of aspergilli to invade blood vessels may involve hematogenous spread to other organs. Disseminated Infection: Dissemination from the lung or any other primary site of infection may affect any other organ system within the body. Single or multiple organs, including the brain, skin, kidneys, pleura, heart, esophagus, liver, or any other site, may be involved

71. Pathology: The tissue reaction may be granulomatous but is more commonly dominated by polymorphonuclear neutrophils unless severely neutropenic . Vascular invasion, thrombosis, and infarction often are prominent features. The hyphae are thin (2 to 5 µm) and septate and branch at acute angles (dichotomous) .

72. Mycologic : Fluffy colonies with granular areas produced by fruiting heads  of  Aspergillus flavus . The development of yellow or green coloration within the white colonies is characteristic. ( Sabouraud dextrose agar.)

73. A fruiting head of  Aspergillus  is demonstrated within a pulmonary  mycetoma  (fungus ball).Demonstration of the fruiting head documents the presence of this genus

74. Gold standard for diagnosis of invasive aspergillosis is a positive culture from sterile tissue with histologic evidence of mycelial invasion. Serologic tests are available for the detection of galactomannan to rapidly diagnose invasive aspergillosis and for the detection of (1-3)-beta-d- glucan for nonspecific diagnosis of an invasive fungal infection .

75. MUCORMYCOSIS/ ZYGOMYCOSIS/PHYCOMYCOSIS Opportunistic infection. caused by bread mold fungi, including Mucor , Rhizopus , Lichtheimia , and Cunninghamella . The two major clinical presentations for mucormycosis are sinus infection with or without cerebral or orbital involvement and localized pulmonary infection that may lead to disseminated disease.

76. Clinical: Paranasal sinuses :An invasive infection known as rhinosinus , rhinocerebral , sinus-orbital, or craniofacial mucormycosis . The hyphae may rapidly break through the thin walls of the sinus and extend up into the orbit, forward into the skin of the face, and back up into the cranial cavity. Predilection for vascular invasion. Pulmonary disease: followed by hematogenous dissemination. Infection begins as an undifferentiated pneumonia, which may be complicated by hemoptysis and cavitation .

77. Pathology: The tissue response is dominated by necrosis.. The propensity for these molds to vessels, producing acute thrombosis, leads to extensive coagulative necrosis. The hyphae appear as ribbons(thin walled) of broad hyphae that are often twisted and collapsed with a variable width from 5 to 20 µm. Septa are sparse and usually are not observed in tissue. Branching tends to occur at right angles in a haphazard pattern.

78. Gomori’s methenamine silver stain, 400×

79. CRYPTOCOCCOSIS Infection with the encapsulated yeast Cryptococcus neoformans . Found in soil and pigeon droppings Most infections occur in immunocompromised patients, especially those with AIDS. Clinical: Pulmonary cryptococcosis Cerebromeningeal cryptococcosis

80. Pulmonary cryptococcosis Immunologically competent patients may exhibit no symptoms Immunocompromised patients on the other hand may have extensive infection Extrapulmonary disease may appear weeks after a pulmonary infection has been documented. Cerebromeningeal cryptococcosis Cryptococcal meningitis is the most frequent and most serious focus of disseminated cryptococcal infection Headache and changes in mental status and personality often dominate the clinical picture.

81. Pathology: The encapsulated yeast in variable size with a decidedly round nature and narrow-based budding are clues to the identification of this organism in tissue. The presence of a thick polysaccharide capsule gives these organisms the characteristic appearance of having a clear space around the yeast cells Differentiation in tissue sections can be accomplished by staining of cryptococcal mucopolysaccharide with mucin stains demonstration of melanin pigment with the Fontana-Masson stain-capsule deficient cryptococcus .

82. A. In a section of the lung stained with hematoxylin and eosin, Cryptococcus neoformans appears as holes or bubbles. B. The same section stained with mucicarmine illustrates the capsule of the organism.

83. Poorly encapsulated strains of C. neoformans elicit a granulomatous inflammatory response, and yeasts are found predominantly in the cytoplasm of macrophages. Mycology: Colonies  of  Cryptococcus neoformans   usually  appear  mucoid . Some strains are poorly encapsulated and lack the mucoid appearance. ( Sabouraud dextrose agar.)

84. The polysaccharide antigen of the C. neoformans species complex can be detected in cerebrospinal fluid and serum, using the latex agglutination test

85. PNEUMOCYSTIS CARINII PNEUMONIA Infection with Pneumocystis carinii usually presents as pneumonitis . Infection is through droplets inhalation. It occurs in immunosuppressed or debilitated patients and is the commonest cause of pneumonia in AIDS. Clinical: Acute or insidious, symptomatic adults frequently present with dyspnea , a nonproductive cough, an inability to breathe deeply, chest tightness, and night sweats. The most common extrapulmonary sites of infection are the thyroid gland, liver, bone marrow, lymph nodes, and spleen

86. Pathology: The definitive diagnosis -demonstration of cysts or trophozoites within clinical specimens These cysts, which are 2 to 6 µm in diameter, stain brown to black and have the characteristic cup-shaped or crescent-shaped morphology. Gomori’s methenamine silver stain, 400×

87. Cysts do not bud; this feature can be used to distinguish this organism from other fungi found in tissue. Use of fluorescein -conjugated monoclonal antibodies to detect P. jiroveci in clinical specimens has proved to be a specific and sensitive method for identification . Molecular assays-the mitochondrial large subunit rRNA target using a PCR platform was shown to be the most sensitive assay,

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