Microbiology Case Study: Immunocompromised 65 Year Old Man

Case History:

A 65 year old man presents to the emergency room with acute onset back pain. Of note, the man was diagnosed with Burkitt’s lymphoma two months prior and had recently received a course of chemotherapy. During the workup for his back pain, a chest CT is obtained and reveals a 2 cm pulmonary nodule in the left upper lobe with a surrounding “groundglass halo” highly suspicious for invasive fungal involvement. A fine needle aspiration (FNA) of the nodule is performed and tissue is sent for histopathologic examination as well as bacterial and fungal culture.

Chest CT showing a 2 cm nodule in the left upper lobe
Chest CT showing a 2 cm nodule in the left upper lobe
Silver stain of the FNA specimen showing ribbon-like fungal elements with rare septations
Silver stain of the FNA specimen showing ribbon-like fungal elements with rare septations
Scotch-tape preparation reveals rarely septate hyphae with internodal rhizoids and pyriform sporangia supported by funnel-shaped apophyses.
Scotch-tape preparation reveals rarely septate hyphae with internodal rhizoids and pyriform sporangia supported by funnel-shaped apophyses.
Scotch-tape preparation reveals rarely septate hyphae with internodal rhizoids and pyriform sporangia supported by funnel-shaped apophyses.
Scotch-tape preparation reveals rarely septate hyphae with internodal rhizoids and pyriform sporangia supported by funnel-shaped apophyses.

 

Laboratory Identification:

One rapidly growing white colony was identified that became grey over time. The colony was a “lid-lifter” that began pushing at the lid after only a few days. Microscopically, the organisms had broad hyphae with single and branching sporangiophores. At the ends of the sporangiophores there were pyriform, or pear-shaped, sporangia sitting atop funnel-shaped apophyses. Rhizoids were found to be internodal, or arising from the hyphae between the sporangiophores. Based on this morphology, the fungal organisms were identified as Lichtheimia corymbifera complex (formally Absidia corymbifera).

Discussion:

Lichtheimia corymbifera is an organism within the phylum Zygomycota and is one of two pathogenic species in the genus Lichtheimia. This organism is known as an uncommon cause of Zygomycosis and is only implicated in approximately 5% of cases. As in most cases of Zygomycosis, exemplified in our patient, Lichtheimia corymbifera most often affects immunocompromised patients. It is ubiquitous in the environment and is associated with decaying plant matter and soil. Disease is caused by inhalation of spores.

Important points for laboratory identification:

Lichtheimia

  • Growth at 35-37°C (capable of growth up to 50°C)
  • Inhibited by media containing cycloheximide
  • Internodal rhizoids
  • Pyriform sporangia
  • Apophysis

Compared to other common Zygomycetes:

Mucor

  • No rhizoids
  • Round sporangia
  • No apophysis

Rhizopus

  • Nodal rhizoids (directly opposite of the sporangiophores)
  • Round sporangia
  • No apophysis

 

-Britni Bryant, MD is a 2nd year anatomic and clinical pathology resident at the University of Vermont Medical Center.

 

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-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.

Microbiology Case Study: 64 Year Old Male with Swollen Finger

A 64 year old male presented with one week of swelling in his right 4th finger. He was initially treated for a suspected bacterial infection, but did not respond to treatment and the finger was aspirated. The specimen was positive for fungal organisms.

Fungal plates grew the following:

Note the flat powdery/velvety colony growth.  It is common for the colonies to have a purple or lavender color with a white border.
Note the flat powdery/velvety colony growth. It is common for the colonies to have a purple or lavender color with a white border.

Scotch tape prep revealed the following morphology:

Note that the elongated Phialides, and taper to a long slender tube, resembling bowling pins.  Sometimes this morphology has also been referred to as "skeleton hands."
Note that the elongated phialides, and taper to a long slender tube, resembling bowling pins. Sometimes this morphology has also been referred to as “skeleton hands.”

Discussion:
Purpureocillium lilacinum (formerly Paecilomyces lilacinus) is a fungus that is found ubiquitously within our environment, but has rarely been associated with disease in humans. A review paper in 2004 found 119 reported cases that implicated P. lilacinum from 1964 to 2004. It has been most commonly associated with ocular infections, often linked to intraocular lens implantations. There is scarce data concerning its susceptibility. In the event of a cutaneous infection such as the one presented in this case, it is recommended that posaconazole be used as first line therapy. In the event of treatment failure, or intolerance, there is little data about which antifungals to treat with, though voriconazole has been shown to have successful in-vitro.

The patient in this scenario had a history of undifferentiated spondylarthropathy, and was on methotrexate for a monoarthropathy in same finger in which the fungal growth occurred. He also received a cortisone injection into the joint adjacent to, but not directly into the site of the infection approximately a month prior to presentation. It could be possible that the cortisone injection had allowed the fungus to be inoculated into the finger, but we may never be certain. The patient also was gardening prior to his infection, and that could have also possibly contributed to his fungal infection.

P. lilacinum is found readily in the environment, and should be considered on the differential of cutaneous infections. Though immunocompromised patients have historically been more susceptible, it has been reported in immunocompetent individuals and should be considered, especially in the event of failure of response to antibiotic treatment. More research needs to be done to better understand treatment regimens for this organism, though this is difficult as it has been difficult to test in animal models.

Reference:

Clinical manifestations, treatment and outcome of Paecilomyces lilacinus infections. F.J. Pastor and J. Guarro. Volume 12, Issue 10, pages 948–960, October 2006 DOI: 10.1111/j.1469-0691.2006.01481.x

-Rich Smith is a Pathology Student Fellow at University of Vermont Medical Center.

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-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.

Microbiology Case Study: Immunocompromised Boy with Skin Nodules

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.

Septate hyphae on GMS stained section of the skin punch biopsy.
Septate hyphae on GMS stained section of the skin punch biopsy.
Septate hyphae on GMS stained section of the skin punch biopsy.
Septate hyphae on GMS stained section of the skin punch biopsy.
Microscopic identification of Fusarium by lactophenol cotton blue stain.
Microscopic identification of Fusarium by lactophenol cotton blue stain.
Colony of Fusarium growing on inhibitory mold agar (IMA).
Colony of Fusarium growing on inhibitory mold agar (IMA).

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.

Microbiology Case Study: 76 Year Old Female with Upper Back Pain

Case History:
A 76 year old female presents with a two year history of worsening upper back pain. Imaging revealed compression fractures of the first three thoracic vertebrae (T1-T3). Fine needle aspiration and a core biopsy of the T3 vertebral body were examined in surgical pathology. There was acute and chronic granulomatous inflammation with fungal organisms observed on histologic examination. Surgery for decompression and fusion of C5-T6 vertebrae was performed and tissue was sent for fungal culture.

Potato flake agar shows a tan-brown fungus.
Potato flake agar shows a tan-brown fungus.
Mycosel agar shows beige-white fungal growth.
Mycosel agar shows beige-white fungal growth.
Scotch tape prep shows septate hyphae with unbranched conidiophores and single, terminal, "lollipop" conidia.
Scotch tape prep shows septate hyphae with unbranched conidiophores and single, terminal, “lollipop” conidia.
Silver stain of involved bone with fungal organisms exhibiting broad-based budding.
Silver stain of involved bone with fungal organisms exhibiting broad-based budding.

Laboratory Identification:
The workup revealed a thermally dimorphic fungus with a mold form growing in the laboratory at 25°C and a yeast form present in the surgical pathology specimen. The mold form is moderately slow growing and has septate hyphae with small, round, terminal conidia often described as “lollipops.” The yeast form is large (8-15 microns) with broad based buds and double contoured cell walls. The immune system reacts to the presence of the fungus by forming granulomas and leads to acute and chronic inflammation within the involved tissue. The organisms can occasionally be seen within giant cells in histologic sections. The silver stain, as seen above, highlights the organisms.

Discussion:
The fungus described above exhibits the features of Blastomyces dermatitidis. This organism resides in soil and decaying plant matter and is endemic to eastern North America including the Mississippi and Ohio River Valleys as well as areas surrounding the Great Lakes and St. Lawrence River. The most common primary sites of involvement for Blastomyces are cutaneous and pulmonary. Following a primary infection, the disease can progress to disseminated blastomycosis which involves other sites such as bone.

The primary site of infection in this case is unknown. There was no history of cutaneous ulcers and chest imaging was unremarkable. The patient did have a remote history of bloody sputum production which she had attributed to “dental difficulties” that she was experiencing and has since resolved. This may have been evidence of a primary pulmonary infection preceding the vertebral involvement; however it is difficult to say with certainty.

The classic double contoured cell walls are not evident on the silver stain of the surgical pathology specimen in this case. This may be due to the fact that the bone required decalcification before histologic sections could be taken. The decalcification process may have caused an artifactual loss of the double contour. Despite the fact that this classic finding was not seen, the macroscopic and microscopic morphology is most consistent with Blastomyces.

The patient is being treated with long-term itraconazole and is currently doing well.

-Britni Bryant, MD is a 2nd year anatomic and clinical pathology resident at the University of Vermont Medical Center.

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-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.

Microbiology Case Study

Patient History:

81 year old man with a history of systemic vasculitis (present for the past 10 years ANCA negative, ANA negative, Rheumatoid factor <20) on immunosuppression (plaquenil with prednisone 40mg for flares about every 6 months), type 2 diabetes, and hypertension presented to an outside hospital with weakness and dyspnea. He was found to have a widespread purpura, ulcerative lesions, acute kidney injury (creatinine 4.7), and 3 days of hematochezia. He was started on 7 days of levoquin and zosyn for a presumed pneumonia and with no improvement was transferred to our institution. On admission, a CT scan of the chest demonstrated bilateral multifocal pneumonia and multiple cavitary nodules within the lungs. A thoracentesis was performed and was transudative (wbc 1883, N 63%, protein 2.6).

Laboratory findings:

  • WBC 7000/cmm
  • Hemoglobin 9 g/dL
  • Platelet count 104 K/cmm
  • Bacterial culture blood, no growth
  • Cryptococcal antigen negative
  • Pleural fluid bacterial culture and smear negative
  • Pleural fluid AFB culture and smear – no acid fast bacilli, modified acid fast bacilli seen from bottle
  • Pleural fluid fungal culture and smear – no fungi seen, rare modified acid fast bacilli growing
  • Histoplasma urinary antigen positive
  • Histoplasma antibodies negative
  • Blastomyces urinary antigen negative
Gram stain of growth from the AFB bottle showing beaded, branch Gram positive bacilli.
Gram stain of growth from the AFB bottle showing beaded, branch Gram positive bacilli.
Modified acid fast stain of growth from the AFB bottle showing modified acid fast bacilli.
Modified acid fast stain of growth from the AFB bottle showing modified acid fast bacilli.
Isolated growth on BCYE media.
Isolated growth on BCYE media.

Discussion:

Based on Gram stain and modified acid fast stain, modified acid fast bacilli suggestive of Nocardia species was reported. Nocardia are strict aerobic, gram positive, filamentous rods that stain partially acid fast. This is due to the mycolic acids in the cell wall which are shorter than those of mycobacteria. Nocardia species produce many virulence factors including Cord factor (prevents intracellular killing), catalase and superoxide dismutase (which inactivate reactive oxygen species that would otherwise prove toxic to the bacteria).

Nocardia grow well on buffered charcoal yeast extract agar and at 30oC. They produce aerial hyphae and can have a chalky colony appearance. Species level identification is best done with molecular methods. This isolate was identified as Nocardia farcinica at a reference laboratory.

Nocardia species are ubiquitous in the soil. They can cause infections in immunocompromised hosts usually after inhalation or direct inoculation. Infections include bronchopulmonary disease and cutaneous infections. With bronchopulmonary disease, cavitation and spread to the pleura is common, which fits with our patient. Dissemination is also seen with common sites being brain and subcutaneous tissue.

Our patient had a positive Histoplasma urinary antigen, but negative Histoplasma antibodies. The working diagnosis was disseminated Histoplasmosis and he was being treated with amphotericin B. He expired and no postmortem exam was performed. Fungal cultures from the pleural fluid were not growing fungus at the time of this post. Fungal cultures were not obtained from sputum and a BAL was not performed.

-Dan Olsen, MD is a 4th year anatomic and clinical pathology resident at the University of Vermont Medical Center.

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-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.

There’s a Fungus Among Us

A 53 year old man with history of stroke, alcoholism, heart failure, hypertension, and atrophic right kidney presented to the ED with acute urinary retention and complained of dysuria and frequency. He was afebrile, denied nausea/vomiting or headaches. His labs at admission are listed below:

  • WBC: 21 k
  • Na: 122
  • Cr: 3 (baseline 1.2)

Urinalysis showed innumerable white blood cells, leukocyte esterase 3+ and negative nitrite.

A catheter was placed and drained 1 L of yellow cloudy urine. The patient refused admission and he was prescribed ciprofloxacin 500 mg BID empirically and was sent home with a foley catheter in place with plans to follow up with Urology. He returned to the ED the following day because his foley catheter was not draining urine and he noted leaking around his catheter. CT scan was obtained and showed ill-defined areas of increased and decreased attenuation within the urinary bladder lumen and left hydroureteronephrosis.

fungusball1

Urine cultures obtained during his initial presentation grew >100,000 yeast and he was treated with fluconazole. The patient was taken to the operating room 11 days after first presentation to diagnose and treat the mass in the bladder. A tan-brown mass was removed and send to surgical pathology. Representative section (H&E stain) of the specimen is shown below:

fungusball2

Which of the following statements regarding Candiduria is true?

  1. Most patients with candiduria are asymptomatic and the yeasts merely represent colonization
  2. The presence of pseudohyphae in the urine or the number of colonies growing in culture help to distinguish colonization from infection
  3. The most commonly involved organ in disseminated candidiasis is the heart
  4. There is a higher propensity for fungal ball formation in adults than children

The correct answer is 1. Most patients with candiduria are asymptomatic and the yeast merely represent colonization. Infected patients may have symptoms (dysuria, frequency, suprapubic discomfort) while others might not. Pyuria is so common in patients with a chronic indwelling bladder catheter that it cannot be used to indicate infection.

Neither the presence of pseudohyphae in the urine nor the number of colonies growing in culture (unlike bacterial cultures) help to distinguish colonization from infection. Ascending infections are rare but usually subacute or chronic, unilateral and can cause perinephric abscesses.

Fungus balls in adults are uncommon with less than 10 adult cases reported in the literature. Risk factors include uncontrolled diabetes, prolonged use of antibiotics or steroids and immune compromise. Classic laboratory findings include marked leukocytosis, pyuria, hematuria and a concomitant bacterial urinary tract infection. Most cases are caused by Candida species although Aspergillus has been implicated in a few cases.

The kidneys are the most commonly involved organ in disseminated candidiasis and there is a higher propensity of fungus ball formation in neonates.

-Agnes Balla, MD is a 1st year anatomic and clinical pathology resident at the University of Vermont Medical Center.

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-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.