Emerging Infectious Diseases
[Volume 5 No.1 / January - February 1999]

Suggested Citation

Dispatches

Candida dubliniensis Candidemia in Patients with Chemotherapy-Induced
Neutropenia and Bone Marrow Transplantation

Jacques F. G. M. Meis,* Markus Ruhnke, Ben E. De Pauw,* Frank C. Odds,
Wolfgang Siegert, and Paul E. Verweij*
*University Hospital Sint Radboud, Nijmegen, The Netherlands; Humbold
University, Berlin, Germany; and Janssen Research Foundation, Beerse,
Belgium

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      The recently described species Candida dubliniensis has been
      recovered primarily from superficial oral candidiasis in
      HIV-infected patients. No clinically documented invasive
      infections were reported until now in this patient group or in
      other immunocompromised patients. We report three cases of
      candidemia due to this newly emerging Candida species in
      HIV-negative patients with chemotherapy-induced
      immunosuppression and bone marrow transplantation.

Although Candida albicans remains the most common opportunistic yeast
pathogen in patients with AIDS and other immunocompromised persons, species
less susceptible to fluconazole are becoming more common (1). Recently, a
newly described species, Candida dubliniensis, was isolated from
oropharyngeal lesions in patients with AIDS living in Dublin, Ireland (2).
C. dubliniensis, phenotypically very similar to C. albicans in producing
both germ tubes and chlamydospores, has since been recovered from the oral
washings of approximately 25% of 94 HIV-positive Irish patients with or
without AIDS and 3% of 150 HIV-negative Irish persons (3,4), which suggests
that this species belongs to the indigenous microflora of the oral cavity,
albeit in a minority of healthy persons. Subsequent reports indicate that
the species has a worldwide distribution (4). The role of C. dubliniensis
as a pathogen has been limited to oral candidiasis. We now report three
cases of candidemia due to C. dubliniensis in patients not infected with
HIV. The yeasts were initially identified as C. albicans because each
produced germ tubes and chlamydospores; this identification became suspect
when equivocal carbohydrate assimilation patterns were obtained.

Case 1

Graft-versus-host disease of the skin, liver, and digestive tract developed
in a 39-year-old woman with chronic myelogenous leukemia after an
allogeneic hematopoietic stem cell transplant in September 1995, during
which she was treated with cyclosporine and high-dose prednisolone. Germ
tubeproducing Candida spp., later identified as C. dubliniensis, were
isolated from stool samples obtained for routine testing. The white-cell
count was 2.7 x 10(sup 9)/L (72% granulocytes); 4 days later fever and ascites
developed, and C. dubliniensis was cultured from three separate blood
cultures (two sets obtained by venipuncture and one by the central venous
line) taken on the same day (MIC fluconazole, 0.25 g/ml). Ascitic fluid
obtained by a sterile puncture also grew C. dubliniensis. Ascites was
probably related to hypoalbuminemia. An echogram showed no radiologic
evidence of liver candidiasis, although alkaline phosphatase was elevated
(222 U/L; normal < 120 U/L). Treatment was started intravenously with
fluconazole, 800 mg/day; 3 days later, C. dubliniensis were still recovered
in one of five blood cultures taken over 2 days, but from then on, blood
cultures were yeast-negative. Because the patient was in stable condition,
the central line was not removed. Cytomegalovirus (CMV) disease also
developed, which could explain the elevated alkaline phosphatase, with
severe thrombopenia (< 20 x 10(sup 9)/L). The patient was given ganciclovir and
hyperimmune gammaglobulin (Cytotect); Staphylococcus epidermidis bacteremia
also developed, and the patient died 3 weeks after the onset of candidemia,
with severe graft-versus-host disease stage IV, complicated by candidemia
and CMV disease. Permission for autopsy was not granted.

Case 2

In July 1995, a 5-year-old boy was treated with cytotoxic chemotherapy for
relapsed nasopharyngeal rhabdomyosarcoma. Two episodes of bacteremia caused
by Streptococcus mitis and S. epidermidis followed, and the boy was treated
with ceftazidime and later ciprofloxacin (combined with vancomycin) for 3
weeks. Cultures of stools and oral specimens yielded germ tubeproducing
Candida spp. later identified as C. dubliniensis. Four days before the
onset of candidemia the patient became febrile; Staphylococcus aureus and
C. dubliniensis were cultured from sputum. At this time, the child was not
aplastic (leukocyte count 2 x 10(sup 9)/L). Flucloxacillin and ceftazidime were
started. When fungal blood cultures were taken, the patient was very ill,
had profuse diarrhea and high fever, and was leukopenic with a total
leukocyte count of 0.3 x 10(sup 9)/L (granulocytes <0.1 x 10(sup 9)/L; thrombocytes 12
x 10(sup 9)/L); 1 day later, three blood cultures, taken over 24 hours through a
central line, yielded C. dubliniensis (MIC fluconazole, 0.5 g/ml).
Treatment with 12 mg/kg fluconazole was started immediately. C.
dubliniensis were still being recovered from two blood cultures 2 days
after treatment began, but after that, cultures remained sterile, and the
patient gradually improved. The central line was removed 20 days after the
last positive blood culture but was not submitted for culture. The patient
was treated with fluconazole for 1 month (3 weeks intravenously, and 1 week
orally) and was discharged 2 months after the onset of candidemia. No
yeasts were recovered from fecal cultures and oral washes, but 1 month
after discharge, oral washes again sporadically grew C. dubliniensis. The
patient received further radiotherapy without any evidence of candidemia
but died 1 year later of relapsed rhabdomyosarcoma.

Case 3

An 8-year-old girl with sickle cell disease combined with -thalassemia and
recurrent hemolytic crisis received an allogeneic hematopoietic stem cell
transplant in January 1995. One year before, she had a splenectomy because
of hypersplenism. The conditioning regimen for the transplant consisted of
busulfan, cyclophosphamid, and antithymocyte globulin administered with a
Hickman catheter. Cyclosporine and methotrexate were given as prophylaxis
against graft-versus-host disease. A suspension of cotrimoxazole and
amphotericin B was given as antiinfective prophylaxis. Seventeen days after
transplant, sepsis syndrome and renal failure developed while the patient
was still profoundly granulocytopenic (< 0.1 x 10(sup 9)/L). Germ tubeproducing
Candida spp. later identified as C. dubliniensis were isolated from two
sets of blood cultures drawn 6 hours apart from a peripheral vein and from
two sets through the central venous catheter (MIC fluconazole, 0.25 g/ml).
The patient had no signs of oral candidiasis, and yeasts were not recovered
from cultures (oral washes and stools). At this time the patient had
already been treated for 72 hours with imipenem and vancomycin. Because of
persistent fever unresponsive to broad-spectrum antibacterial agents,
intravenous amphotericin B (30 mg) was empirically added. Once the results
of the positive blood cultures became known, 5-flucytosin (100 mg/kg) was
added to the regimen. After initiation of amphotericin B, later blood
cultures remained negative for yeasts. The Hickman catheter was removed 14
days later when the patient had recovered from neutropenia. Catheter tip
cultures remained negative. However, low grade fever persisted.
Nonetheless, because the patient's condition was stable, treatment was
changed to oral fluconazole (50 mg t.i.d.) for another 2 weeks and the
patient was discharged. The cause of persistent fever was not identified,
but approximately 6 months later, the patient recovered.

Microbiologic Results

All yeast isolates were initially identified by germ tube and chlamydospore
formation as C. albicans, but carbohydrate assimilation patterns by
commercial test kits (Auxacolor, Sanofi Pasteur, Paris and API 20C,
Analytab Products, Plainview, New York) gave equivocal results.
Furthermore, the isolates did not elaborate -glucosidase, grew very weakly
at 42C, and failed to grow at 45C (6); they produced dark green colonies
on CHROMagar Candida plates (Becton Dickinson, Etten-Leur, The Netherlands)
typical of C. dubliniensis (4,5) and abundant chlamydospores on rice-cream
agar after 24 hours (2). In contrast with C. albicans, the yeasts isolated
from our patients' specimens hybridized poorly with the C.
albicansspecific Ca3 fingerprinting probe (5) and gave characteristic
arbitrary primer phosphatase-polymerase chain reaction patterns for C.
dubliniensis with primer RP02 (5'-GCGATCCCCA-3'). Each C. dubliniensis
isolate yielded two major bands at 0.4 kb and 1.0 kb, with up to five weak
bands ranging from 0.9 kb to 1.3 kb. In contrast, with C. albicans, the two
major bands were never observed. Instead, each C. albicans isolate yielded
approximately 15 bands of various intensity, ranging from 0.65 kb to 2.4
kb. Furthermore, banding patterns obtained with RP02 were clearly different
from C. glabrata, C. krusei, C. tropicalis, and C. parapsilosis. In vitro
susceptibility testing for fluconazole (powder provided by Pfizer BV,
Capelle a/d IJssel, The Netherlands) was performed by the broth
microdilution method with RPMI-1640 with L-glutamine, buffered with MOPS
incubated at 35C, and read after 48 hours according to NCCLS M-27A (7). C.
parapsilosis ATCC 22019 and C. krusei ATCC 6258 were included as quality
control strains. The isolates from patients 1 and 2 were deposited as CBS
8500 and CBS 8501 at the yeast division, Centraalbureau voor
Schimmelcultures (CBS), Delft, The Netherlands.

C. dubliniensis was first described 3 years ago (2) and is genetically and
phylogenetically distinct from C. albicans (8). Hitherto, its pathogenic
role has been mainly restricted to oropharyngeal infections in HIV-infected
persons and AIDS patients (3,5). In a recent study of C. dubliniensis, one
isolate recovered from a blood culture and one from postmortem lung tissue
was examined (6); however, no clinical data were described to allow
determination of the pathogenic role. The cases we have described show that
C. dubliniensis can cause candidemia in immunocompromised patients.
However, these may not be the first cases of invasive disease due to this
yeast. Identification and differentiation from other germ tubeproducing
yeasts on the basis of phenotypic characteristics has been problematic (8);
therefore, the incidence and prevalence of this organism and its role in
invasive disease have been difficult to determine. For instance, a strain
of C. stellatoidea originally isolated in 1957 from the sputum of a patient
with bronchopneumonia and deposited in the British Culture Collection of
Pathogenic Fungi has been shown to be C. dubliniensis (2,3), and an isolate
of C. albicans (from sputum of a Dutch patient) deposited in the culture
collection of CBS in 1952 has been shown to be C. dubliniensis (Meis,
unpub. obs.). In both cases, it has not been established whether the C.
dubliniensis isolates were responsible for invasive infections.

Fluconazole appears to be less active against C. dubliniensis than against
C. albicans (4) since C. dubliniensis is usually associated with recurrent
episodes of candidiasis and protracted exposure to azole antifungal drugs
in patients with AIDS. Fluconazole showed excellent in vitro activity
against each of the C. dubliniensis isolated from the blood cultures of our
patients; each patient responded well clinically. Nevertheless, it is too
early to estimate the true susceptibility of this species to fluconazole.
This requires the correct identification of the species, which now seems
necessary, given its ability to cause invasive disease in patients treated
for malignant diseases.

Acknowledgments

      We thank T. Rijs, L. Van Nuffel, and G. Dams for excellent technical
assistance and J.P. Donnelly for discussion.

      Dr. Meis is head of the Division of Bacteriology and Mycology,
University of Nijmegen, The Netherlands, and consultant for medical
microbiology and infectious diseases, University Hospital and Clinics. He
is a member of the Nijmegen Mycological Research Group, and his research
focuses on infections in immunocompromised patients, with particular
interest in the management and diagnosis of invasive fungal infections.

      Address for correspondence: Jacques F.G.M. Meis, Department of
Medical Microbiology (440), University Hospital Nijmegen, P.O. Box 9101,
6500 HB Nijmegen, The Netherlands; fax: 31-24-354-0216; e-mail:
j.meis@mmb.azn.nl.

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Emerging Infectious Diseases
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Centers for Disease Control and Prevention
Atlanta, GA

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