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OBJECTIVES

MYCOLOGY CASES FROM 1. Describe clinical and laboratory findings in cases NEJM caused by fungi. 2. Review typical presentations of the fungi in this Lynda Britton, Ph.D. MLS(ASCP)CM presentation. [email protected] LSUHSC Shreveport

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Case 1 DISSEMINATED FUSARIUM PANCYTOPENIA

• 8-year-old boy presented with subcutaneous CBC Result Reference Range nodules Hemoglobin 7.5 g/dL 11.5-14.5 g/dL • 1-week history of fever WBC 200/mm3 5000-11,000/mm3 • 2 months after starting treatment for relapsing B-cell leukemia Platelets 12,000/mm3 150,000-400,000/mm3 • Fevers persisted despite the initiation of broad- spectrum intravenous antibiotic and treatment

3 4 N Engl J Med 2020; 382:e64

DISSEMINATED FUSARIUM BIOPSY SPECIMEN SHOWED SEPTATE HYPHAE

1 week later subcutaneous nodules developed

on the trunk, arms, and legs 5 6

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FUSARIUM DIMERUM FUSARIUM DIMERUM

• Colonies are salmon-colored, moist and -like, producing very little aerial mycelium • Only slow-growing, clinically significant Fusarium spp. • Macroconidiophores are short, simple (unbranched) or branched • Macroconidia are small in comparison to other species and have 0-3 septa. • Microconidia are absent.

• Chlamydoconidia may be present but are usually rare. 7 8

FUSARIUM

FUSARIUM • Fusarium DIMERUM • Plant pathogens that cause COMPLEX root and stem rot, vascular wilt, or fruit rot • Cause mycotic keratitis and , especially in burn victims and bone marrow transplant patients; Also leukopenic particularly for those on broad- spectrum antibiotics

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Case 2 Fusarium CANDIDA ESOPHAGITIS

• A 72-year-old man presented with a 2- week history of odynophagia • 16 months before diffuse panbronchiolitis treated with macrolide antibiotics • Linear, white, mucosal, plaquelike lesions on the esophagus • Endoscopic findings confirmed by a culture of esophageal brushing positive for 11 12 N Engl J Med 2017; 376:1574 • HIV negative

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CANDIDA AT RISK FOR HEAD AND ESOPHAGITIS NECK

• Wear dentures • Oral antifungal • Have diabetes therapy • Have cancer • 2 weeks later • Have HIV/AIDS • Take antibiotics or corticosteroids, including inhaled corticosteroids for conditions like asthma • Have dry mouth • Smoke

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GROWTH CHARACTERISTICS OF CANDIDA ALBICANS CANDIDA ALBICANS

• Grow on almost all common laboratory media including blood agar, EMB, CNA, PEA, etc. Sabouraud agar with antibacterial antimicrobials (Mycosel) is recommended • Some species of Candida (tropicalis, krusei, and parapsilosis) are susceptible to cycloheximide but C. albicans is not

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CANDIDA ENDOCARDITIS AFTER CANDIDA ALBICANS TAVR

• 76-year-old man with congestive heart failure • Worsening shortness of breath • Undergone transcatheter aortic-valve replacement (TAVR) 9 months earlier • Temperature of 39.0°C and grade 5 systolic murmur

17 Case 3 N Engl J Med 2019; 380:e1 18

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LABORATORY FINDINGS

• WBC 16,000/mm3 • CRP 120 mg/L • 6 consecutive blood cultures positive for Candida parapsilosis ECHOCARDIOGRAM AND VEGETATIONS • Prosthetic-valve endocarditis

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SUSCEPTIBILITY RISK FACTORS FOR

• Critical illness with a prolonged intensive care unit stay • Presence of central venous catheters • Use of broad-spectrum antibiotics or total parenteral nutrition • Having hematologic or solid organ malignancy, stem cell transplantation, neutropenia, or recent abdominal surgery (especially in the presence of an anastomotic leak) • Being a pre-term infant with a very low birth weight • Having renal failure or hemodialysis Candida albicans, C. glabrata, C. parapsilosis, C. tropicalis, and C. • Injection drug use krusei 21 22

CANDIDA CANDIDA PARAPSILOSIS PARAPSILOSIS

• Glucose yeast extract peptone broth • Ovoid cells, cylindrical cells 20 µm • Cornmeal agar • Pseudohyphae in branched chains

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Case 4 GOMORI METHENAMINE SILVER FUNGAL STAIN ENDOPHTHALMITIS

• 34-year-old man with a history of intravenous drug use and infection with HCV • 1-week history of pain and decreased vision in his right eye • Blood cultures and echocardiography negative • Mass adherent to optic nerve • Necrotizing granuloma

N Engl J Med 2020; 382:e48 25 26

FUNGAL ENDOPHTHALMITIS FUNGAL ENDOPHTHALMITIS

• Completed 6 weeks of oral voriconazole • 2 to 15% of all endophthalmitis cases are endogenous • Visual acuity improved to 20/30 • 1% of patients with candidemia develop Candida • Stable at 6-month follow-up visit with no endophthalmitis evidence of recurrence. • Candida species most common cause of endogenous infections • Rare complication of eye injury or eye surgery • Occurs as a post-surgical complication in approximately 0.1% of all cataract surgeries

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CRYPTOCOCCUS Case 5 NEOFORMANS MENINGOENCEPHALI TIS

• A 36-year-old man, sexually active MSM • 2-week history of fever, headache, drowsiness, and photophobia • Opening pressure on lumbar puncture was 29, <20 cm GRAM STAIN AND INDIA INK • HIV viral load was 300,000 copies/ml

• CD4+ count was 7N cells/µL Engl J Med 2018; (reference 379:281 range, 500 to 29 30 1450)

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LABORATORY FINDINGS • Liposomal amphotericin B and flucytosine • Resolution of symptoms and CRYPTOCOC • CSF cell count 340 cells/µL (reference range, 0 to negative results on CSF after 2 CUS 10) weeks of treatment NEOFORMAN • 90% mononuclear cells predominantly lymphocytes • Consolidation therapy with S MENINGO- ENCEPHALITI • Glucose 46 mg per dL (reference range, 40 to 70 • Antiretroviral therapy S mg per deciliter • Protein 0.80 g/L (reference range, 0.15 to 0.45) • Cryptococcal antigen titer -1:128, 31 32 • CSF culture grew

CRYPTOCOCCUS NEOFORMANS CRYPTOCOCCUS NEOFORMANS EPIDEMIOLOGY EPIDEMIOLOGY • Incidence is fairly low: many people exposed without • Found commonly in the droppings of fowl, particularly pigeons symptoms who do not contract the disease, but excrete it in their feces • An opportunist that primarily affects those with a • Lives for prolonged periods of time in pigeon droppings if not compromised immune state exposed to sunlight • Humans inhale dust contaminated with pigeon or fowl fecal • Causes respiratory and systemic disease such as material containing the viable yeast forms meningitis

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MOST SUSCEPTIBLE TO CRYPTOCOCCUS VIRULENCE FACTOR • Defective cell mediated immunity • AIDS and other immune diseases • Large polysaccharide • Hodgkin's disease capsule that prevents • Lymphomas phagocytosis and the • Leukemia production of antibody • Sarcoidosis • Systemic erythematosis

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CRYPTOCOCCUS DIAGNOSTIC FEATURES OF NEOFORMANS COLONIES C. NEOFORMANS

• Mucoid and creamy white • Encapsulated budding yeast • Positive Cryptococcus latex • Cycloheximide is inhibitory to it • Phenoloxidase produced— • Brown/black colonies on bird or niger • Formation of melanin-like seed agar pigments on bird seed agar • Urease positive • Nitrate negative • Susceptible to cycloheximide 37 38

CRYPTOCOCCAL LATEX DISSEMINATED ANTIGEN TEST Case 6 CRYPTOCOCCOSIS

• Very sensitive and specific test better than India ink prep • 60-year-old man presented with a 4-week history of • Capsular material is solubilized in the body fluids and can progressive lesions. be titered with a specific antiserum • Latex particles are coated with specific rabbit • For the past 6 years, he had been treated for chronic immunoglobulin to Cryptococcus and mixed with dilutions lymphocytic leukemia (CLL) of patient's serum or CSF. • Positive agglutination is diagnostic • CD4+ T-cell count was 20 per cubic millimeter (1%) • Titer for prognosis • Multiple located mainly on his face but also on • CSF or serum for detection of early, asymptomatic his shoulders, arms, and legs cryptococcal infection in HIV-infected patients • Higher sensitivity than microscopy or culture. 39 N Engl J Med 2014; 370:1741 40

DISSEMINATED CRYPTOCOCCOSIS

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CANDIDA AURIS IN AN CRYPTOCOC- Case 7 INTENSIVE CARE SETTING COSIS • Reusable skin-surface axillary temperature probes • Rare since • persisted despite cleaning with quaternary ammonium compounds HAART: • 70 patients in UK neurosciences intensive care unit decreased • Risk factors: 52 median age 90% • 67% male with lower serum albumin levels (P=0.06) • Case-fatality • Higher body temperature (P=0.08) ratio ~12% • Higher serum sodium levels (P=0.07) • Systemic fluconazole treatment

Leading cause of death in HIV in Africa> TB43 N Engl J Med 2019; 380:890-891 44

CANDIDA AURIS C. AURIS

•Ist multidrug-resistant species of yeast • Emerging, multidrug-resistant pathogen associated • Hospital-acquired with outbreaks worldwide, often in ICUs • Serious bloodstream infections, wound, ear • 2009 first reported, traced to 2004 • May spread between patients • October 26, 2020: Case count 1,394 • 1 in 4 die: 30--60% of patients infected died • Survive for extended periods on common hospital room surfaces • Difficult to ID

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IDENTIFICATION OF CANDIDA AURIS

• No pseudohyphae • Sometimes aggregates of cells • Grows well at 42ᵒC • MALDI-TOF with high scores • Sequencing the 28s rDNA • Send to CDC AR lab network

• Antifungal testing white, pink, red or purple 48

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LABORATORY MISIDENTIFICATION OF C. AURIS C. AURIS SUSCEPTIBILITY dentificationMethod Organism C. auris can be misidentified as Vitek 2 YST Candida haemulonii Candida duobushaemulonii • All isolates were resistant to fluconazole (MIC >32 mg/L) API 20C Rhodotorula glutinis (characteristic red color not present) • 30% were nonsusceptible to voriconazole (MIC >2 mg/L). Candida sake • 40% were resistant to amphotericin B (MIC >2 mg/L) BD Phoenix yeast identification system Candida haemulonii Candida catenulata • 60% were resistant to 5-flucytosine (MIC >32 mg/L) MicroScan Candida famata * • 70% were multidrug resistant, including 30% (n = 3) that Candida guilliermondii * Candida lusitaniae were resistant to 3 classes of drugs * Candida parapsilosis • All isolates were susceptible to echinocandins * RapID Yeast Plus Candida parapsilosis

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AT RISK OF C. AURIS

• Recent surgery, diabetes, broad-spectrum antibiotic and antifungal use • Recently spent time in nursing homes and have lines and tubes (such as breathing tubes, feeding tubes and central venous catheters) • Found in patients of all ages, from preterm infants to the elderly

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MULTIDRUG-RESISTANT CANDIDA AURIS INFECTIONS IN CRITICALLY ILL CORONAVIRUS DISEASE PATIENTS, INDIA, APRIL–JULY 2020 • Candidemia affected 15 critically ill ICU coronavirus disease patients • 10 C. auris; 8 elderly and male • All had indwelling invasive central venous and urinary catheters • C. auris colonizes skin, persists in environments, causes nosocomial outbreaks

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Case 8 DISSEMINATED

• 34-year-old man with HIV and CD4 count was 39 • Viral load was 197,000 copies • 1-week history of headache, fever, and confusion • Large, ulcerative lesion was noted on his tongue MULTIPLE FUNGAL ORGANISMS CONSISTENT WITH • Patchy infiltrates in both lungs SPHERULES • IgG antibodies detected in the blood and CSF

N Engl J Med 1947; 237:610-616 55 56

COCCIDIOIDES WAS ALSO NOTED DISSEMINATED IN FUNGAL CULTURES OF BAL COCCIDIOIDOMYCOSIS FLUID

• Liposomal amphotericin B and fluconazole • HIV genotype testing revealed resistance to 1 drug • Antiretroviral adjusted • 3 month later viral load undetectable and coccidioidomycosis resolved

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COCCIODIODES IMMITIS

• Grows fast on Sabouraud agar, with the mycelium covering the slant or plate within 3-7 days • Young culture is cobweb-like, with fluffy areas alternating with areas adherent to the agar COCCIDIOIDOMYCOSIS

2018 15,611 cases in U.S. ~200 deaths

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COCCIDIOIDOMYCOSIS

• Acute respiratory disease • Inhalation of Coccidioides fungal spores prevalent in soil of Southwestern U.S., San Joaquin Valley and desert areas of Arizona • Incidence peaks in fall and winter months • 40% of infections manifest as fever, cough, fatigue, chest pain, shortness of breath, and • Self-limited or progressive

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PATHOGENSIS OF COCCIDIOIDES AT RISK OF COCCIDIOIMYCOSIS • Arthroconidia reach the alveoli of the lungs where they develop • People who have weakened immune into spherules within 48 to 72 hours systems: • Endospores are disseminated through the lymphatics and blood • Have HIV/AIDS • Pulmonary disease is pneumonic and necrosis and cavitation • Have had an organ transplant may occur after hypersensitivity develops • Are taking corticosteroids • If patient has cellular immunity, process will be controlled and • >60 years old resolved • Pregnant women • If not, may become disseminated • People who have diabetes • People who are Black or Filipino

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COCCIDIOIDOMYCOSIS IN ARIZONA

• 2017–2018 fall and winter incidence of coccidioidomycosis largest increase in cases in the last 5 years • The 4827 reported cases reported from October 2017 to March 2018 represented a 58.3% increase over the previous year. • Weather such as precipitation, which enhances fungal replication • Followed by extreme heat which can facilitate spread • Increased population and construction in Maricopa County • Area with highest incidence, might have also contributed to the VALLEY FEVER increase. 65 66

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VALLEY FEVER (COCCIDIOIDOMYCOSIS) DISSEMINATED AWARENESS — CALIFORNIA, 2016– Case 9 2017 • 97% of U.S. cases are reported from Arizona and • 30-year-old man with HIV California • 2-week history of fever, bloody diarrhea, and generalized • Increased 213% from 2014 to 2018 weakness • Only 25% of persons living in a high-incidence • Febrile, pale, and cachectic region and 3.0% of persons living in a moderate- • Discontinued antiretroviral therapy 3 months before incidence region were • Spent 2 months in French Guiana

N Engl J Med 2019; 380:e13 MMWR Morb Mortal Wkly Rep 2020;69:1512–1516 67 68

LABORATORY FINDINGS

• CD4+ cell count of 6 per/mm • C-reactive protein level of 239 mg/L • Pancytopenia GIEMSA STAINING OF • H. capsulatum in peripheral blood PERIPHERAL BLOOD and bone marrow aspirate PCR

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HISTO- PLASMOSIS

• Tuberculate • Causes histoplasmosis, cave disease, and Darling’s disease macroconidia • Endemic in the Ohio and Mississippi River Valleys; resides • Convert to yeast at in nitrogen-rich soil and organic matter 37o C • Acquired by inhalation • Differentiated from the saprophyte, • Primary infection is pulmonary Sepedonium that fails • Disseminate to the reticuloendothelial system to grow on media with cycloheximide 71 72

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HISTOPLASMA CAPSULATUM PATHOGENESIS

• Microconidia are inhaled from an exogenous source and penetrate the alveoli • Convert to small budding yeast cells • Yeast phagocytized by alveolar macrophages • Cleared through the upper respiratory tract or may disseminate by the blood stream

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HISTOPLASMOSIS AT RISK

• People in endemic areas: occupational or participate in activities exposing them to soil with bird or bat droppings • Disseminated: • Immunosuppressed persons (HIV/AIDS, organ transplant, or use of immunosuppressive medications , infants, or adults age HISTOPLASMOSIS 55 years and older.

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DISSEMINATED • 39-year-old man with acute BLASTOMYCOSI hemoptysis and chills S ONE MONTH Case 10 • CT--infiltrate in the right upper lobe LATER • Treated with levofloxacin • Son pulled on his left ring DISSEMINATED • Gone hiking in northern Minnesota finger caused pain and BLASTOMYCO during the preceding summer swelling SIS • Osteomyelitis • Persistent infiltrate in the right upper lobe • Treated with oral itraconazole for 1 year

N Engl J Med 2017; 376:e9 77 78

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DISSEMINATE D BLASTOMYCO SIS • KOH revealed yeast with broad- based budding • Cultures DISSEMINATED grew Blastomyce s dermatitidis • Broad-based large budding yeast

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DISSEMINATED BLASTO-

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Case 11 DISSEMINATED CUTANEOUS BLASTOMYCOSIS DISSEMINATED • 44-year-old man with multiple nodules over his body, including CUTANEOUS face, chest, back, arms, and legs BLASTOMYCOSIS • 6 months earlier, he had had fever, cough, and vomiting for 1 week • Worked in construction and reported no recent travel, exposure to pets, or contact with sick persons • Normal WBC count, clear lungs and no lymphadenopathy • Negative HIV and 83 84 N Engl J Med 2018; 379:74

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Case 12 TALAROMYCES INFECTIO MARNEFFEI INFECTIO N N

• 29-year-old man with 2-month history • A biopsy specimen obtained of fever, cough, dyspnea, and weight from a showed loss, skin lesions numerous round fungal • Extensive umbilicated papules • Tissue culture grew Talaromyces structures in histiocytes marneffei (formerly Penicillium • Methenamine silver stain marneffei) • HIV positive with CD4+ count of 25

N Engl J Med 2017; 377:2580 85 86

TALAROMYCES MARNEFFEI INFECTIO N TALAROMYCES MARNEFFEI • Treatment with intravenous amphotericin B was initiated and 2 weeks later, antiretroviral therapy • endemic in southern China, was started. Southeast Asia, and northeastern India • 4 months later, the cutaneous • Causes infection in people with HIV infection who live papules had diminished in these regions substantially, with residual • Incidence decreased with the increased use of superficial atrophic scars HAART • Increased incidence among other persons with 87 impaired cell-mediated immunity such as organ 88 transplantation

Case 13 TALARO- MYCES • A 65-year-old man otherwise healthy MARNEFFEI • 5-month history of a progressively enlarging annular rash over his right hand and forearm • Worked as a farmer and developed after trauma to his hand from agricultural work • Large, nontender, annular areas of over the dorsum of the right hand and forearm with ulceration and 89 crusting 90

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SPOROTRICHOSIS SPOROTRICHOSIS

• The patient was treated with itraconazole and and local thermotherapy for 2.5 months • 4 months after the completion of treatment, the erythema and crusting had resolved

91 92

SPOROTRICHOSIS

• Common in plants and animals • Subacute-to-chronic infection • Localized lymphangitic form • Disseminated cutaneous form • Extracutaneous form 93 94 • Increasing frequency

Case 14 IN A SOUDANENSE NEWBORN • Scalp scrapings and a swab • 3-week-old male newborn cultured on Sabouraud’s dextrose agar grew Trichophyton brought to pediatrician with 3 large scalp lesions • PCR revealed Trichophyton soudanense • Mother had similar skin • Closely related to T. violaceum lesions on her upper trunk • Anthropophilic tinea capitis in N Engl J Med 2018; 378:2022 northern Africa

95 • Slow-growing 96

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Case 15 HIV MANIFESTING AS MAYO STUDY PROXIMAL WHITE ONYCHOMYCOSIS • T. violaceum and T. soudanense most common among • 48-year-old-man with whitish patients of African descent discoloration and hyperkeratosis of • Underrecognized because rare in U.S. the first toenail • 1997 to 2014: 81 children • No fever, weight loss, malaise, • 67 patients (82.7%) African, 4 African‐American, 3 white, 2 lymphadenopathy, or rash Asian and 5, unknown • KOH revealed numerous septate • Median time to clinical cure was 2.3 months hyphae

97 N Engl J Med 2017; 377:e26 98

HIV MANIFESTING AS PROXIMAL HIV MANIFESTING AS WHITE PROXIMAL WHITE ONYCHOMYCOSIS ONYCHOMYCOSIS • isolated in a • Proximal white onychomycosis with rapid extension from potato dextrose agar culture of the proximal to the distal is more unusual and can be suggestive of HIV the nail scrapings and had the characteristic red-wine color • Viral load of 17,510 copies • CD4+ count of 40 • LPCB revealed pyriform microconidia in “birds on a wire” • Treatment with antiretroviral therapy and antifungal appearance therapy was initiated • Good clinical response and resolution of onychomycosis

99 100

Case 16 TINEA FACIEI

• Healthy 5-year-old girl with dry, scaly, erythematous, and annular lesions on • Scrapings revealed numerous hyaline septate hyphae her face • PCR positive for Trichophyton mentagrophytes • Raised outer border and a pale center • Initially treated with topical terbinafine and • Similar lesions on her left earlobe and right arm olamine • Her mother and sister had similar- • After 14 days of treatment with oral terbinafine lesions appearing skin lesions on the arms completely resolved and neck 101 102 N Engl J Med 2014; 370:e31

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TRYCHOPHYTON DIFFERENTIATING MENTAGROPHYTES TRICHOPHYTON SPP. • T. rubrum is red on the reverse when grown on cornmeal agar and T. mentagrophytes is yellow • T. mentagrophytes is urease positive within one week. T. rubrum takes 2 weeks or is negative for urease • T. mentagrophytes perforates in vitro and is positive on BCP milk solids agar • T. rubrum is able to assimilate sorbitol

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TRYCHOPHYTON MENTAGROPHYTES Case 17 TINEA VERSICOLOR

• T. mentagrophytes = Positive • 24-year-old woman presented with a 12-year history of a • T. rubrum = Negative depigmenting rash • Rash was most notable in the summer months, with remission during cooler seasons. • She reported prominent scaling of her skin, particularly after showering. • Over the preceding 2 to 3 years, the rash had spread to include her entire torso with extension down her arms. She was concerned that facial involvement might occur. • Previous therapies with multiple topical antifungal agents had not regenerated skin pigmentation 106 105 N Engl J Med 2016; 374:e11

TINEA TINEA VERSICOLOR VERSICOLOR

• Large, coalescing, hypopigmented patches and macules on her torso and upper arms • Yeast in a “spaghetti and meatball” pattern in the superficial • Tinea versicolor 107 108

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Case 18

TINEA VERSICOLOR

• Scaling that results from stretching or scraping of the • 27-year-old man with a 3-month history of progressive obstruction skin suggestive of tinea versicolor and bleeding from the right naris • Patient treated with a course of oral fluconazole and • Red, granular, pedunculated, topical nonpulsatile mass that obstructed the right nasal cavity and bled on • Process of skin repigmentation may take months once touch the fungal cause is eliminated

N Engl J Med 2019; 380:1359 109 N Engl J Med 2019; 380:1359 110

RHINOSPORIDIOSIS

• Multiple thick-walled sporangia in various stages of maturation, surrounded by dense inflammation • Chronic granulomatous disease caused by the aquatic protistan parasite

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RHINOSPORIDIOSIS

RHINO- • Rare aquatic protistan parasite of fish SPORIDIOSIS • Oropharynx, conjunctiva, rectum, and external genitalia affected • Endemic in India and Sri Lanka, Africa and South America • No effective antimicrobial therapy • Follow-up 8 months later--recurrence in the nasopharynx that were removed

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QUESTIONS?

CODE:

115

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