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Journal of Clinical Microbiology, December 2001, p. 4520-4522, Vol. 39, No. 12
Department of Diagnostic Sciences and
Pathology1 and Department of
OCBS,3 Dental School, University of
Maryland, and Department of Pathology, The Johns Hopkins
University,4 Baltimore, Maryland, and
Department of Oral Diagnostic and Oral Pathology, Faculdade de
Odontologia da U. F. do Rio de Janeiro, and F.O.U. Gama Filho,
Rio de Janeiro, Brazil2
Received 4 June 2001/Returned for modification 7 August
2001/Accepted 2 September 2001
Oral and subgingival samples from periodontal lesions were
collected from 54 human immunodeficiency virus (HIV)-positive and 20 HIV-negative patients and cultured for yeast species. Of the 54 samples
cultured from HIV-positive patients, 44 (82%) were positive for yeast
species, of which 29 (66%) were subgingival. A total of 19 (48%)
patients were positive for Candida dubliniensis, of which
15 (79%) were colonized in subgingival sites. Seven isolates of
Candida glabrata, two isolates of Candida
parapsilosis, and one isolate of Saccharomyces
cerevisiae were recovered. This study reports for the first time
the recovery of C. dubliniensis from subgingival intraoral
sites and confirms the presence of Candida species in sites
of periodontal disease associated with HIV.
Fungal species have gained
considerable importance as opportunistic pathogens of immunocompromised
individuals, particularly those infected with human immunodeficiency
virus (HIV) (12, 28). Although Candida albicans
is the most common etiologic agent of oral candidosis, Candida
dubliniensis has emerged as another pathogen, noted for its in
vitro potential for azole resistance and its enhanced in vitro
adherence to human buccal epithelial cells (9, 14, 16, 17, 20,
23, 30).
The human oral cavity offers a unique environment with a multitude of
ecological niches for microbial colonization. Periodontal diseases have
gained attention in HIV-positive individuals as a result of the rapidly
progressive and often destructive nature of the diseases
(10). Periodontal diseases are characterized by an
inflammatory, degenerative, and necrotic response in the gingival and
underlying connective tissues, elicited by microbial colonization in
periodontal pockets (3, 13). The initiation and
progression of periodontal diseases are the result of the colonization
and multiplication of microorganisms in subgingival sites, stimulation
of the host immune system, and the release of host immune factors
causing tissue damage (10). HIV-positive individuals and
AIDS patients in particular often suffer from somewhat unique forms of
periodontal diseases, the result of altered immune response in the
periodontal tissues with changes in cellular and humoral immune
responses (1, 2, 24, 27).
The cultivable microflora from HIV-positive patients with periodontal
diseases has been found to be comprised of periodontopathic species
similar to those found in HIV-negative subjects with periodontitis, mainly gram-negative bacteria (5, 24). Although the
presence of yeasts was a striking feature of the subgingival plaque
samples from some AIDS patients, follow-up studies confirming these
preliminary data have been lacking (11, 27, 28, 32). The
present investigation sought to begin to determine any association
between the recovery of yeasts and the presence of advanced periodontal
lesions. The study was designed to recover and identify yeast isolates
from the oral tissues and subgingival sites of HIV-positive patients with advanced periodontal disease and to compare the results to those
obtained from a group of 20 HIV-negative subjects with comparable clinical periodontal diseases.
Fifty-four adult patients with positive histories of HIV infection and
20 healthy nonimmunocompromised control subjects were recruited, and
informed consent was obtained. Each patient had clinical evidence of at
least one deep periodontal lesion (pocket depth, >5 mm). Periodontal
evaluations of all patients included clinical examination and
radiographs. The subjects ranged in age from 17 to 58 years (24%
female and 76% male). At the time of collection, 14 patients had
evidence of clinical mucosal oral lesions that were suggestive of
candidosis. The HIV-negative control subject group was comparable with
regard to age and sex, without clinical signs of fungal infection. With
a sterile curette, subgingival samples were collected from periodontal
sites (pocket depth, >5 mm), identified by measurement with a Marquis
probe and by periodontal attachment loss as ascertained by clinical
and/or radiological examination. In addition to subgingival samples,
tongue or buccal mucosal (TBM) samples were obtained with a sterile
swab. Subgingival samples were immediately streaked onto Sabouraud
dextrose agar (Difco Laboratories, Detroit, Mich.) for isolation;
similarly, oral swabs were rolled on Sabouraud dextrose agar and
streaked for isolation. Samples were collected from all patients by the same procedures. Agar plates were incubated at 35 to 37°C, in a
non-CO2 atmosphere, for 48 to 72 h and checked daily for
growth. Fungal growth was subjectively rated as ranging from light to heavy.
All isolates growing on primary culture were identified by conventional
mycology methods. Briefly, isolates that were germ tube positive were
tested for their ability to grow at 45°C (C. albicans
grows at 45°C, whereas C. dubliniensis fails to grow at
this temperature). Germ tube-positive isolates that failed to grow at
45°C were tested with the coaggregation assay, as described previously (15). CHROMagar Candida (CHROMagar, Paris,
France) was used to identify yeast species that yielded negative
results in germ tube culture and C. dubliniensis based on
colony color. All isolates were tested for substrate assimilation
profiles by the API 20C Aux system (bioMérieux Vitek, Inc.,
Hazelwood, Mo.) to identify germ tube-negative isolates to the species
level and to confirm the identification of C. dubliniensis. All germ tube culture-positive isolates that failed
to grow at 45°C were positive with the coaggregation assay and
produced a dark green colony color in testing with CHROMagar
Candida; these results were consistent with C. dubliniensis identification. The API 20C system confirmed the
identification of C. dubliniensis and identified to the
species level the germ tube-negative isolates.
The results of fungal cultures from the HIV-positive patients are
summarized in Table 1. Among the 54 patient samples, a total of 44 (82%) were positive for yeasts; 29 (66%) of these were subgingival
samples. C. albicans, C. dubliniensis,
Candida glabrata, Candida parapsilosis,
and Saccharomyces cerevisiae were the yeast species
recovered. Two or three species were coisolated from eight patients,
and subgingival and TBM samples of nine patients indicated colonization
by different species. A total of 21 (48%) patients were found to be
harboring C. dubliniensis; 15 (79%) samples from these
patients were recovered from periodontal lesions, with seven
exclusively in the subgingiva (Table 1).
The 20 HIV-negative patients yielded six positive TBM samples and one
positive subgingival sample, with light growth of C. albicans. C. dubliniensis was not recovered from the HIV-negative patients.
0095-1137/01/$04.00+0 DOI: 10.1128/JCM.39.12.4520-4522.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
Recovery of Candida dubliniensis and Other Yeasts from
Human Immunodeficiency Virus-Associated Periodontal Lesions
ABSTRACT
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TABLE 1.
Yeast species recovered from HIV-positive periodontal
patients
The oral cavity is a prime site for a variety of tissue lesions associated with HIV infection, including mucocutaneous and esophageal candidosis, sometimes as the first clinical expression of HIV infection (4, 18, 22, 25, 31, 32). Many studies have reported subgingival C. albicans in adult periodontitis, yet it is not known whether subgingival fungal colonization or infection participates in the pathogenesis of destructive periodontal disease (11, 29).
Early in the AIDS epidemic, some studies found that the microflora of oral subgingival sites includes C. albicans in as many as 50 to 60% of diseased sites of AIDS patients (10, 25, 26, 32). It was suggested that when C. albicans gains access to underlying periodontal tissues, damage results from the metabolites produced by the yeast (10). Yeast species in the subgingival tissue of AIDS patients may also be the source of disseminated candidosis. The most recent study performed by Hannula et al. (11) on isolation of subgingival yeasts reported that the mean proportion of total microbiota was higher for C. albicans-positive samples than for C. albicans-negative samples from selected subjects, indicating the need for further analysis to determine the relationship between subgingival occurrence of C. albicans and periodontal diseases.
C. dubliniensis has been shown to have, in addition to greater adherence to human buccal epithelial cells, mucin, and the oral bacterium Fusobacterium nucleatum, significantly higher proteinase activity, which was shown to degrade mucins, the major constituents of mucus (7, 15, 16, 20, 21). With interactions among microorganisms shown to be an important step in infectious disease processes in the oral cavity, the ability of C. dubliniensis to adhere to F. nucleatum may aid in colonization of deep sulci of periodontal pockets (6, 8, 19).
In this investigation, 44 of the 54 patients sampled were positive for fungus, with 21 (48%) positive for C. dubliniensis. Of the 44 positive patients, 29 (66%) had positive subgingival cultures; 15 (39%) of these samples were positive for C. dubliniensis. In addition to C. dubliniensis and C. albicans, C. glabrata, C. parapsilosis, and S. cerevisiae were the other species recovered from the subgingival samples (Table 1). In contrast, results of fungal cultures of the control, HIV-negative group demonstrated one patient with a subgingival culture of light growth of C. albicans. No other species were recovered.
This study reports, for the first time, the recovery of C. dubliniensis from subgingival periodontal lesions and confirms the presence of Candida in the subgingival sites of HIV-positive patients with periodontal diseases. The high frequency of recovery of Candida from subgingival samples of immunocompromised individuals, in comparison to that of a control group, supports the need for further investigations to determine the role that Candida species, and in particular C. dubliniensis, may play in the progression of periodontal lesions.
This research was conducted in partial fulfillment of a Fogarty fellowship (awarded to Sonia Ferreira) supported by the Institute of Human Virology and the AIDS International Training and Research Program.
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FOOTNOTES |
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* Corresponding author. Mailing address: Department of Oral Medicine, Dental School, UMAB, 666 W. Baltimore St., Baltimore, MD 21201. Phone: (410) 706-7628. Fax: (410) 706-0519. E-mail: mrizk{at}umaryland.edu.
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Journal of Clinical Microbiology, December 2001, p. 4520-4522, Vol. 39, No. 12
0095-1137/01/$04.00+0 DOI: 10.1128/JCM.39.12.4520-4522.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
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