Rare disease

Case report

Isolated cerebral mucormycosis caused by
Rhizomucor pusillus
Saira Farid,1 Omar AbuSaleh,1 Rachael Liesman,2 Muhammad Rizwan Sohail1
1

Department of Infectious
Diseases, Mayo Clinic,
Rochester, Minnesota, USA
2
Department of Clinical
Microbiology, Mayo Clinic,
Rochester, Minnesota, USA
Correspondence to
Dr Muhammad Rizwan Sohail,
​sohail.​muhammad@​mayo.​edu
Accepted 19 September 2017

Summary
A 61-year-old man with relapsing chronic lymphocytic
leukaemia, status post allogeneic stem cell transplant
and multiple chemotherapy regimens presented to the
emergency room after suffering a grand mal seizure.
His evaluation revealed a 1.5–2 cm ring-enhancing left
temporal lobe brain lesion on the CT scan. This brain
lesion was resected and the histopathology revealed
an invasive fungal organism resembling mucormycosis.
Amplification and sequencing of the 28S ribosomal RNA
gene identified the organism as Rhizomucor pusillus.
The patient was treated with liposomal amphotericin
B 5 mg/kg every 24 hours for 4 weeks, and then was
transitioned to oral posaconazole. Serial brain imaging
at 1 and 3 months, while on therapy, showed significant
improvement.

Background
We report a rare case of isolated cerebral mucormycosis due to unusual Mucorales sp, Rhizomucor
pusillus, in an immunocompromised host. Diagnosis
was established by histopathological examination of
the resected mass showing fungal element consistent with mucormycosis. The causative organism
was identified with amplification and sequencing
gene targets that lead to the final identification of
R. pusillus. The patient had excellent outcome with
combined surgical and medical therapy.

Case presentation

To cite: Farid S, AbuSaleh O,
Liesman R, et al. BMJ Case
Rep Published Online First:
[please include Day Month
Year]. doi:10.1136/bcr-2017221473

A 61-year-old man presented to the emergency
department with acute grand mal seizure activity. He
had a complicated medical history most remarkable
for relapsing chronic lymphocytic leukaemia (CLL),
diagnosed 20 years prior to his current presentation, for which he underwent multiple treatments,
including an allogeneic peripheral blood stem cell
transplant 3 years prior to his presentation. His
post-transplant was complicated by chronic cutaneous graft versus host disease (GVHD) for which
he was maintained on tacrolimus and prednisone
along with photopheresis. After he experienced a
CLL relapse, he was started on ibrutinib. He was
kept on trimethoprim–sulfamethoxazole for pneumocystis prophylaxis and fluconazole for fungal
prophylaxis, and received monthly intravenous
immunoglobulin (Ig) for hypogammaglobulinaemia. The patient had no prior history of seizures
and was afebrile at presentation. He had no history
of intravenous drug use. He lived in the Midwest
region of USA and wintered in Arizona state.

On neurological examination, patient displayed
expressive aphasia but had no other focal or global
neurological deficits. The skin examination had
findings consistent with cutaneous GVHD. Respiratory, cardiovascular, gastrointestinal and genitourinary examinations were unremarkable.

Investigations
Initial laboratory evaluations included a complete
blood count revealing anaemia (haemoglobin of
8.5 g/dL and haematocrit of 26.8%), lymphopaenia (absolute lymphocyte count of 0.28×109/L)
and thrombocytopaenia (platelet count of 149/L).
Serum creatinine at presentation level was 1 mg/
dL. Additional tests included normal chemistries,
prostate-specific antigen, lipids and thyroid-stimulating hormone level. Blood PCR for cytomegalovirus was negative. 1–3 Beta-D-glucan testing
was performed and was negative (36 pg/mL;
ref: <60 pg/mL=negative).
A CT scan of the head revealed a 1.5–2 cm ring
enhancing lesion in the left temporal lobe (figure 1).
Subsequent MRI showed a 1.8 cm ring-enhancing
lesion in the left temporal lobe with vasogenic
oedema, which was thought to be an abscess or
malignancy. CT of the chest, sinuses and abdomen–
pelvis revealed no evidence of extracranial disease.
Bacterial and fungal blood cultures remained negative after 5 and 30 days of incubation, respectively,
and extensive serological workup for infectious
aetiologies was negative, including serum Cryptococcus antigen, Histoplasma complement fixation
and immunodiffusion antibodies, Histoplasma
urine antigen, Blastomyces antibody enzyme immunoassay (EIA), Coccidioides antibody EIA and
serum galactomannan.
A left temporal craniotomy with mass resection was performed and the initial intraoperative
impression was of a metastatic tumour. Consequently, no tissue was sent to microbiology for
culture. However, histopathological examination of
the tissue showed necrosis with acute and chronic
inflammation. Grocott-Gomori methenamine silver
(GMS) stain highlighted numerous broad pauciseptate fungal hyphae, suggestive of a Mucorales
sp (figure 2). To confirm the identity of the causative pathogen, amplification and sequencing of
the 28S ribosomal RNA gene and the internal transcribed spacer 1 and 2 (ITS1 and 2) gene regions
were performed on the formalin-fixed paraffin-embedded (FFPE) tissue and sequencing results identified the organism as R. pusillus.

Farid S, et al. BMJ Case Rep 2017. doi:10.1136/bcr-2017-221473

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Rare disease

Figure 1 (A) Head CT with contrast, coronal section, showing a left
temporal lobe ring-enhancing lesion surrounded by vasogenic oedema
(arrow head). (B) Brain MRI, flair sequence, showing left temporal lobe
mass surrounded by vasogenic oedema.

Treatment

The patient received induction therapy with liposomal amphotericin B 5 mg/kg every 24 hours for 4 weeks, and then was transitioned to oral posaconazole delayed-release tablets. Serial brain
imaging at 1 and 3 months, while on therapy, showed significant
improvement with decreased oedema and further contraction of
the surgical cavity.

Outcome and follow-up

Clinically, patient made almost complete recovery and was maintained on oral posaconazole for secondary prophylaxis given the
need for ongoing immunosuppression for GVHD. At 1-year
follow-up, the patient was doing well and there were no signs of
disease recurrence.

Discussion

The term mucormycosis is often used to imply an angioinvasive infection caused by moulds that belong to the order Mucorales, a ubiquitous environmental mould found in decaying
organic substrates. Members of the Mucorales order important
in human disease include Rhizopus sp, Mucor sp, Rhizomucor
sp, Lichtheimia sp and Cunninghamella sp, and are characterised by pauciseptate, irregularly branching, ribbon-like hyphae.

Figure 2 Brain tissue with GMS stain. Numerous broad, ribbon-like,
pauciseptate or pauciseptate hyphae are stained, consistent with
Mucorales sp. GMS, Grocott-Gomori methenamine silver.
2

In a review of 929 reported cases of confirmed mucormycosis,
47% were attributed to the genus Rhizopus, 18% by pathogens
belonging to the genus Mucor, 7% by the genus Cunninghamella
and 4% by pathogens in the genus Rhizomucor.1
Mucormycosis is most often associated with immunocompromised hosts as an opportunistic infection. Risk factors for
the development of invasive mucormycosis include diabetic
ketoacidosis, neutropaenia, corticosteroid use, haematological
malignancy and solid organ or bone marrow transplantation.
Invasive mucormycosis in immunocompetent individuals has
been reported following trauma associated with natural disasters. Mucormycosis can be fatal in up to 96% of disseminated
cases and delays in microbial diagnosis may contribute to high
mortality. Site of infection, underlying predisposing factors
and the species involved are known predictors for mortality
as well.1
The genus Rhizomucor, first described in 1978, is a thermophilic saprophytic fungus in the order Mucorales. R. pusillus, R.
miehei and R. variabilis are the pathogenic species associated
with human disease. Due to the small size (3–5 µm), the spores
of Rhizomucor are easily disseminated via the airborne route and
aerosolised sporangiospores are readily inhaled. R. pusillus has a
wide geographic distribution and, although a frequent cause of
invasive Mucorales infection in animals, has been rarely associated with human disease.2
R. pusillus is reported to be less virulent and have lower
minimal inhibitory concentrations (MIC) to antifungal agents
compared with more common Mucorales sp, such as Cunninghamella, which is associated with increased mortality.2 However,
infection with R. pusillus causes significant morbidity and
mortality, and patients have a 30% risk for disseminated infection at presentation and 46% risk of mortality.2 Individuals with
haematological malignancies are at highest risk of acquiring this
infection.2
Diagnosis of invasive mucormycosis may be clinically challenging. The Mucorales are associated with disease presentation
involving multiple organ systems, including pulmonary, rhinocerebral, cutaneous, central nervous system (CNS) and the gastrointestinal tract.
Rhinocerebral disease is characterised by rapid invasion into
surrounding facial structures and is classically associated with
poorly controlled diabetes and ketoacidosis. Pulmonary mucormycosis is more commonly described in patients with neutropaenia. Cutaneous mucormycosis is most often associated with
traumatic disruption of the skin barrier. Skin is also a common
site of disseminated infection.
Cerebral mucormycosis is most often diagnosed in the context
of concomitant sinus disease or disseminated disease. However,
isolated cerebral mucormycosis may account for up to 8% of
all cases of mucormycosis.1 Intravenous drug use is the most
important risk factor for isolated cerebral disease.1 3 Radiographic features cannot distinguish between mucormycosis and
other pyogenic cerebral infections. However, a rapidly enlarging
brain abscess in the basal ganglia in the context of intravenous
drug use should raise the suspicion for isolated cerebral mucormycosis.4 5
The diagnosis of mucormycosis often relies on histopathological examination of surgically removed tissue and visualisation of broad, ribbon-like, pauciseptate hyphae with wide-angle
branching suggests a diagnosis of Mucorales. Angioinvasion is
common in Mucorales infections, and hyphae may be visualised within blood vessels. Acute inflammation with neutrophilic
predominance is often present, but may be diminished in patients
with neutropaenia.
Farid S, et al. BMJ Case Rep 2017. doi:10.1136/bcr-2017-221473

Rare disease
Distinguishing between Aspergillus and Mucorales may be
complicated when hyphae are not abundant in tissue. Artefactual folds or breakdown of hyphae may also be confused with
septations, leading to an incorrect identification. Additionally,
histopathological identification of mucormycosis does not
provide genus and species information or susceptibility data and
has limited ability to detect mixed or dual fungal infections.6 7
Culture of Mucorales from infected tissue is important for
species identification, yet the recovery of Mucorales in culture
is often difficult. In a large case series of histopathologically
confirmed mucormycosis, cultures were positive in only 50%
(465/929) of cases.1
Processing in the microbiology laboratory may damage the
fragile pauciseptate hyphae, decreasing culture yield from tissue
sources, and mincing of tissue, rather than grinding, is recommended. Growth is rapid (1–7 days) on routine fungal selective
media (eg, Inhibitory mould agar) and isolates often have erect
aerial mycelium reaching the lid of the petri dish, leading to the
colloquial designation of this group as ‘lid lifters’. Despite the
angioinvasive nature of Mucorales, blood cultures are rarely
positive.8
Molecular methods are increasingly used for detection and
identification of fungi in fresh and FFPE tissue and, most recently,
in plasma specimens. Broad-range fungal PCR, using primers
targeting the ITS gene regions or the 28S or 18S ribosomal
RNA gene regions, followed by sequencing has demonstrated
good agreement with culture results, and sensitivity is increased
when applied to fresh over FFPE tissue. Molecular detection
may allow for the identification of the infecting organism to the
genus and species level, a distinct advantage over histopathological diagnosis, which may facilitate the choice of appropriate
antimicrobial therapy based on known susceptibility patterns.8 9
Advances in the diagnosis of Mucorales are needed to accurately and rapidly diagnosis patients with invasive disease. PCR
assays targeting the most common species of Mucorales have
been designed for use in tissue samples or serum/plasma. In one
study, frozen plasma specimens from a cohort of 44 patients with
proven or probable mucormycosis were retrospectively tested
using a set of three Mucorales PCR assays targeting Rhizomucor
sp, Lichtheimia sp and Mucor/Rhizopus sp. Interestingly, 81%
(36/44) of patients had at least one positive PCR, often preceding
the date of radiological findings or microbiological confirmation.10 Clinical use of molecular assays for the detection of
Mucorales in plasma specimens may allow for earlier diagnosis
and initiation of effective therapy in patients. While promising,
evaluation of the performance of these assays in larger prospective studies is needed.
Early diagnosis and initiation of Mucorales active antifungal
therapy in known to improve mortality, and delay in therapy of
>1 week has been associated with increased mortality.11 Antifungal
agents active against the Mucorales include amphotericin B, isavuconazole and posaconazole, and the two active azoles are commonly
used as maintenance therapy after initial treatment with amphotericin B. Surgical resection, if feasible, and modification of immunosuppression are also known to improve mortality.12

Farid S, et al. BMJ Case Rep 2017. doi:10.1136/bcr-2017-221473

Learning points
►► Diagnosis of mucormycosis may be challenging, and

often requires a high index of clinical suspicion and
histopathological visualisation of broad, ribbon-like,
pauciseptate hyphae with wide-angle branching in biopsy
specimens.
►► Tissue processing at the microbiology laboratory can affect
the yield of cultures. Early communication with laboratory
staff regarding clinical suspicion of mucormycosis may permit
modified tissue processing steps to optimise culture yield.
►► Molecular methods may help to established the diagnosis
and identifying causative species, especially when fungal
cultures are not sent or are negative.
►► Early diagnosis and initiation of antifungal therapy along
with surgery, if feasible, are associated with improved patient
survival.
Contributors SF and MRS: case report planning. SF, OAS andRL: crafting of the
manuscript. SF, OAS and MRS: critical revision of themanuscript for important
intellectual content. MRS: case report supervision. Allauthors: acquisition of data;
analysis and interpretation of data.
Competing interests None declared.
Patient consent Obtained.
Provenance and peer review Not commissioned; externally peer reviewed.
© BMJ Publishing Group Ltd (unless otherwise stated in the text of the article)
2017. All rights reserved. No commercial use is permitted unless otherwise expressly
granted.

References

1 Roden MM, Zaoutis TE, Buchanan WL, et al. Epidemiology and outcome of
zygomycosis: a review of 929 reported cases. Clin Infect Dis 2005;41:634–53.
2 Gomes MZ, Lewis RE, Kontoyiannis DP. Mucormycosis caused by unusual
mucormycetes, non-Rhizopus, -Mucor, and -Lichtheimia species. Clin Microbiol Rev
2011;24:411–45.
3 Verma A, Brozman B, Petito CK. Isolated cerebral mucormycosis: report of a case and
review of the literature. J Neurol Sci 2006;240:65–9.
4 Hopkins RJ, Rothman M, Fiore A, et al. Cerebral mucormycosis associated with
intravenous drug use: three case reports and review. Clin Infect Dis 1994;19:1133–7.
5 Sundaram C, Mahadevan A, Laxmi V, et al. Cerebral zygomycosis. Mycoses
2005;48:396–407.
6 Sangoi AR, Rogers WM, Longacre TA, et al. Challenges and pitfalls of morphologic
identification of fungal infections in histologic and cytologic specimens: a ten-year
retrospective review at a single institution. Am J Clin Pathol 2009;131:364–75.
7 Guarner J, Brandt ME. Histopathologic diagnosis of fungal infections in the 21st
century. Clin Microbiol Rev 2011;24:247–80.
8 Lackner M, Caramalho R, Lass-Flörl C. Laboratory diagnosis of mucormycosis: current
status and future perspectives. Future Microbiol 2014;9:683–95.
9 Bialek R, Zelck UE. [PCR-based diagnosis of mucormycosis in tissue samples].
Pathologe 2013;34:511–8.
10 Millon L, Herbrecht R, Grenouillet F, et al. Early diagnosis and monitoring of
mucormycosis by detection of circulating DNA in serum: retrospective analysis of 44
cases collected through the French Surveillance Network of Invasive Fungal Infections
(RESSIF). Clinical Microbiology and Infection 2016;22:810.e1–810.e8.
11 Chamilos G, Lewis RE, Kontoyiannis DP. Delaying amphotericin B-based frontline
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2016;30:143–63.

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Farid S, et al. BMJ Case Rep 2017. doi:10.1136/bcr-2017-221473