Over on her blog, Maryn McKenna discusses the latest emerging microscopic threat: drug-resistant fungal infections. It mentions the organism Candida auris and states: “The Centers for Disease Control and Prevention (CDC) is so concerned that it recently sent an alert to U.S. hospitals, even though only one possible case of the resistant fungus has been identified in the United States so far.”
Case History
A 44 year old woman presented to an outside hospital with a chief complaint of abdominal tenderness and increased abdominal girth over the past few days. Her past medical history was significant for type II diabetes mellitus and associated complications including retinopathy and chronic kidney disease. As a result of her renal failure, she was currently undergoing peritoneal dialysis. Due to concern for infectious peritonitis, a paracentesis was performed and the resulting fluid obtained was sent to the Microbiology laboratory for Gram stain and bacterial culture. Because of difficulty in identifying the isolate, it was sent to our mycology section for further work up.
Laboratory Identification
Figure 1. Many sporangia of various sizes containing endospores arranged in a “soccerball” like pattern or morula configuration (Lactophenol cotton blue prep, 400x).
The isolate was received on potato dextrose agar (PDA) and appeared as discrete cream colored colonies, resembling a yeast. Upon transfer to cornmeal agar and incubation at 30°C, the isolate grew rapidly over the course of two days. The organism did not grow on media containing cycloheximide. A lactophenol cotton blue prep was performed (Figure 1) and showed many sporangia, ranging in size from 4-15 µm. In the larger forms, individual endospores were able to visualized and they were arranged in a “soccerball” like pattern. No budding or hyphae were present. Given these characteristics, the organism was identified as Prototheca wickerhammii.
Discussion
Prototheca wickerhammii is classified as an achlorophyllous algae and is known to cause human infections involving the skin and subcutaneous tissues, bursa of the elbow joint (olecranon bursitis) and rarely, systemic infections. P. wickerhammii is ubiquitous in nature and infection is usually the result from traumatic inoculation. Both immunocompetent and immunocompromised hosts can be affected, although more severe or systemic disease occurs in those who have defects in cell mediated immunity.
Prototheca spp. grow rapidly on PDA after incubation at 30°C for 2-3 days. Initially, it may be confused with a yeast based on plate morphology as they are cream colored and have a yeast-like consistency. When a lactophenol cotton blue prep is viewed under the microscope, sporangia of various sizes are identified (3-30 µm) that contain endospores arranged in a “soccerball” or symmetric daisy like pattern. Size of the sporangia, assimilation tests (such as the API 20C) and automated yeast identification systems (such as the Vitek yeast ID by bioMérieux) are helpful in identifying Prototheca to a species level. Both P. wickerhammii (4-11 µm in diameter) and P. zopfii (9-28 µm in diameter) can cause disease in humans, but P. wickerhammii is more common. While P. wickerhammii has symmetric internal divisions as seen in the above image, P. zopfii has random internal divisions.
The treatment of P. wickerhammii usually includes a combination of surgical management and anti-fungals depending on the site of involvement. For superficial skin infections, localized excision and the use of topical or systemic anti-fungals (azoles and amphotericin B) has been shown to have success. The treatment for olecranon bursitis focuses on bursectomy. In the case of systemic infections, amphotericin B has been the most successful treatment modality. Susceptibility testing is not routinely recommended to guide treatment of initial infections, as studies are few and results don’t always correlate with outcome.
Lisa Stempak, MD, is an Assistant Professor of Pathology at the University of Mississippi Medical Center in Jackson, MS. She is certified by the American Board of Pathology in Anatomic and Clinical Pathology as well as Medical Microbiology. Currently, she oversees testing performed in both the Chemistry and Microbiology Laboratories. Her interests include infectious disease histology, process and quality improvement and resident education.
Case History:
A previously healthy 30 year old man presents with a painful right eye. Three weeks before he had been cutting concrete with an electric saw when a piece of hard concrete hit him in the eye. The eye became painful the following day. He was treated with empiric antibiotics but the eye pain failed to improve. He was eventually seen by ophthalmology, where ophthalmologic exam demonstrated findings suspicious for fungal keratitis. Corneal scraping was performed and sent for bacterial and fungal culture.
Laboratory Identification:
Bacterial culture showed no growth. Fungal culture demonstrated rapid growth of multiple white, cottony molds on potato flake agar. Over time these white colonies turned dark grey/brown. The reverse surface of the agar was white at first but also turned dark grey/brown over time.
Microscopically (scotch tape preparation), there were thin hyphae with single conidia arising directly off the tips of tapered conidiophores of variable length. Occasional conidia were also observed arising directly off of the sides of hyphae. The conidia were small and oval in shape with a truncated base, and somewhat darker as compared to the surrounding hyphae. Sexual forms were not observed.
Discussion:
The clinical history and laboratory findings are characteristic of Pseudallescheria boydii/Scedosporium apiospermum.
Pseudallescheria boydii/Scedosporium apiospermum is an environmental mold which can be isolated from rural soils, polluted waters, manure and compost. Infection occurs secondary to local trauma. In immunocompetent individuals, infection is limited to the site of trauma, with some of the more common presentations being fungal keratitis, endophthalmitis, eumycotic mycetoma, sinusitis and pneumonia in the setting of near drowning. In immunocompromised individuals, infection can disseminate and involve any organ.
Pseudallescheria boydii/Scedosporium apiospermum is the most common cause of eumycotic mycetoma. Mycetoma is a chronic granulomatous infection of the subcutaneous tissue, usually involving the distal lower extremities, which can be caused by either a fungus (eumycotic mycetoma) or an actinomyces species bacteria (actinic mycetoma). On clinical exam there are multiple draining sinus tracts. The causative microorganisms aggregate into macroscopically visible groups (“granules”) which can be white, yellow or brown in color. Mycetoma may progress over time to involve underlying soft tissue, muscle, fascia and bone. Other less common causes of eumycotic mycetoma include Madurella spp., Acremonium spp., Fusarium spp., and Curvularia spp.
This organism has two names because historically different names were assigned to the sexual state (Pseudallescheria boydii) and to the asexual state (Scedosporium apiospermum). Morphologically, Pseudallescheria and Scedosporium are identical, the only difference being the presence of the sexual form, cleistothecia, in Pseudallescheria. Cleistothecia, when present, can be recognized as very large, dark-brown asci containing numerous ascospores. Technically, in the absence of cleistothecia the correct diagnosis would be Scedosporium boydii, however in clinical practice both names are usually listed regardless of whether cleistothecia are seen.
Pseudallescheria boydii/Scedosporium apiospermum is resistant to amphotericin B, but should be susceptible to azole therapy. The patient is being treated with topical voriconazole and oral fluconazole and currently (two weeks of antifungal therapy) has near complete resolution of symptomatology.
-Javier De Luca-Johnson, MD is a 3rd year anatomic and clinical pathology resident at the University of Vermont Medical Center.
-Christi Wojewoda, MD, is the Director of Clinical Microbiology at the University of Vermont Medical Center and an Assistant Professor at the University of Vermont.
A 7 year old Congolese male presented with pruritic, erythematous, non-flaky rash on top of his scalp for the past 3 weeks. The rash in non-painful, but continues to spread. His mother has been applying hydrocortisone cream nightly, with no improvements.
Laboratory Identification
A hair sample was obtained for fungal culture. Colonies were yellow and waxy with feet-like projections. Microscopic morphology on lactophenol analine blue scotch tape prep revealed broad hyphae with tortuous branches. The hyphae lacked obvious micro and macro conidia, raising the suspicion for Trichophyton violaceum.
Discussion
Trichophyton violaceum is an anthropophilic fungus seen predominantly in North Africa, East Asia and parts of the Middle East. It forms slow growing with glabrous colonies. Microscopically, broad tortuous hyphae are seen. Microconidia and Macroconidia are notably absent. T. violaceum causes Tinea Capitis, which can be acquired through scalp contact with the dermatophyte, either with direct contact with an infected individual or an object. It can also affect skin, nails and beards. It manifests clinically as pruritic scaly patches with alopecia, often producing black dots. Affected hairs demonstrate an endothrix infection.
-Mustafa Mohammed, MD is a 2nd year anatomic and clinical pathology resident at the University of Vermont Medical Center.
-Christi Wojewoda, MD, is the Director of Clinical Microbiology at the University of Vermont Medical Center and an Assistant Professor at the University of Vermont.
An elementary school aged boy with a history of pre-B cell acute lymphocytic leukemia with a failed bone marrow transplant was transferred to a regional children’s hospital for leukodepletion and participation in an experimental clinical trial. At that time, his CBC was significant for 10% polymorphonuclear cells and 50% blasts. He was subsequently transferred to the ICU in respiratory failure and developed papulonecrotic lesions on his face, trunk, and bilateral legs. Prior to this, he was pancytopenic with no blasts present with cell counts of 100 WBC, hemoglobin 8.3 and 37,000 platelets. His Fungitell assay, which detects (1-3)-β-d-glucan, was positive.
Routine blood culture, fungal culture from the endotracheal tube, and fungal culture from the skin lesion biopsy specimens all had fungal elements on KOH stain. Young growth of a whitish, fluffy mold was present on all cultures within two days. Histopathology on the punch biopsy of a skin lesion on the thigh showed septate hyphae within the dermis, epidermis, and invading the vasculature that was particularly apparent with GMS stain (Figure 1a and 1b). Within a few days, the fungal cultures showed septate hyphae with microconidia using lactophenol cotton blue tape preparation, and shortly thereafter the mold developed into macroconidia with multiple septations taking on canoe-like forms (Figure 2). The white, cotton-like colonies developed a pink tinge (Figure 3). These characteristics allowed for the identification of the growth as Fusarium sp.
Fusarium is an opportunistic hyaline mold with infection most commonly seen in immunocompromised hosts. It can cause keratitis through contamination of contact lenses, penetration due to trauma, or use of immunosuppressive steroid ophthalmic solution. It is increasingly becoming the cause of disseminated infection in neutropenic hosts with a broader spectrum of disease, which includes: skin lesions, fungemia, rhinocerebral involvement and pneumonia. In these cases, without an immune system to fight the infection, mortality is high. Inhalation of airborne conidia, ingestion from water sources or access through mucosal membranes are all potential points of entry.
The colony growth on plated fungal media is rapid, usually maturing within four days. On microscopic examination, Fusarium hyphae are septate, approximately 3-6 microns wide with acute angle branching. Microconidia are small, oval-shaped, and no larger than 4 x 8 microns in size. These can look like Acremonium sp. Macroconidia are canoe- or sickle-shaped with the largest dimension being about 80 microns in length, exhibiting 3-5 septatations.
–Jodi Music, MD, is an AP/CP resident at UT Southwestern Medical Center.
-Erin McElvania TeKippe, Ph.D., D(ABMM), is the Director of Clinical Microbiology at Children’s Medical Center in Dallas Texas and an Assistant Professor of Pathology and Pediatrics at University of Texas Southwestern Medical Center.
A 51 year old woman with a significant smoking history presented with 8-9 weeks of fever and cough. Shortly after the beginning of her illness, she developed pleuritic left-sided chest pain and hemoptysis. She was treated with amoxicillin and then prednisone without improvement. She had progressively worse pain and hemoptysis as well as fevers and night sweats, with weight loss. A chest x-ray and CT scan showed a left upper lobe mass- like infiltrate suspicious for a carcinoma. She underwent transbronchial fine needle aspiration biopsy of the lesion which showed the following morphology.
The specimen was also sent for fungal culture.
Laboratory diagnosis:
Blastomyces dermatidis was identified by microscopy and colony morphology. Septate, delicate hyphae with single, circular-to-pyriform condia on short conidiophores (lollipops) were seen on a scotch tape prep. The colonies appeared waxy and wrinkled, with a cream-tan color. Large, thick-walled yeast with buds attached by a broad base, 8-15 um with double-contoured walls, were demonstrated in tissue. Additionally, this patient had a positive urine antigen test for Blastomyces.
Discussion:
Blastomyces’ natural habitat is unknown but the organism is thought to reside in soil or wood, particularly in the Ohio, Mississippi, and Missouri River valley regions. It takes 2-30 days to grow in the lab. The infectious form is the conidia which are transmitted by inhalation. Common sites of infection include skin, lungs, and bone. The typical presentation in an immunocompetent individual is a pulmonary infection with associated acute or chronic suppurative and granulomatous lesions. Blastomyces infection may also cause osteomyelitis, prostatitis, urethritis, CNS infection, and disseminated infection. Immunocompromised patients may present with disseminated infection with involvement of skin, bone, and multiple organs. Infection may be confirmed by exoantigen testing or by nucleic acid probe testing.
-Lauren Pearson, D.O. is a 2nd year anatomic and clinical pathology resident at the University of Vermont Medical Center.
-Christi Wojewoda, MD, is the Director of Clinical Microbiology at the University of Vermont Medical Center and an Assistant Professor at the University of Vermont.
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