Oral carriage of candida in healthy and HIV-seropositive persons

Oral carriage of candida in healthy and HIV-seropositive

persons

Catharina Hester Johanna Hauman, BMedsci, M.Medsci, BCHD,” Ivor Oscar Christopher Thompson, BCHD, (Hons) BCHD, MCHD,” Francois Theunissen, NHD (Med Tech),b and Pieter Wolfaardt, BCHD, MCHD,C Tygerberg, South Africa FACULTY

OF DENTISTRY,

UNIVERSITY

OF STELLENBOSCH

The prevalence of oral colonization with Candida species was studied in 28 HIV-seropositive and 28 healthy persons. Candida was cultured from 75% and 68% of HIV-positive and control persons, respectively, with a significantly higher density carriage in the HIV-seropositive group. Positive smears were seen in 39% of all patients. Candida albicans was the most frequently isolated species with Biotype 1 accounting for 56 % of the isolates. Resistance to antifungal agents was seen in Candida strains isolated from both groups. (ORAL SURC ORAL MED ORAL PATHOL 1993;76:570-2)

Candida species are opportunistic yeastlike fungi that commonly colonize human mucosal surfaces. Systemic and oral changes within the host may promote a proliferation of the patient’s own commensal oral mycoflora, with an ensuing risk of host tissue invasion.’ Oral candidiasis becomes clinically apparent in the prodromal stages of AIDS2 with more than 75% of infected patients presenting with candidiasis during the course of the disease.3 Oral candidiasis is a potential reservoir of organisms for spreading the disease locally in the compromised host, exemplified by esophageal candidiasis in AIDS.’ Appropriate knowledge of oral candidal colonization and infection is thus of concern in the management of patients with AIDS and related disorders. The aim of the present study was to compare the oral carrier rate of Candida species between confirmed HIV-positive and healthy persons in a South African population. MATERIAL Subjects

AND METHODS

Fifty-six patients were screened for the presence of oral Candida species and anti-HIV antibodies. Twenty-eight patients tested positive for the presence of anti-HIV antibodies by enzyme-linked immunosorbent assay plus Western blotting. They were routinely monitored as part of the outpatient population at Tygerberg and Somerset Hospitals. This group consisted of 25 men and three women, mean age 33.36 years f 11.67, age range 21 to 60 years. Twenty-two patients were HIV-seropositive with%enior Lecturer, Department of Oral Pathology. bPrincipal Technologist, Department of Oral Pathology. CDepartment of Periodontology and Oral Medicine. Copyright B 1993 by Mosby-Year Book, Inc. 0030-4220/93/$1 .OO+ ,lO 7/13/50085

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out any symptoms (CDC stage I) while six had persistent generalized lymphadenopathy (CDC stage IX). All 28 patients had CD4 counts higher than 400. Only one patient was receiving AZT therapy at the time of sampling. He had also been treated with ketoconazole for oral candidiasis. One other person from this group had received antifungal therapy previously. The remaining 28 patients were “healthy” persons attending the Oral and Dental Teaching Hospital at Tygerberg for dental treatment. For ethical and financial reasons they were not tested for the presence of HIV antibodies. A thorough medical history was taken of each person, and all patients with a history of diabetes mellitus, xerostomia, or any form of immunosuppression or any persons on immunosuppressive medication were excluded from this study. This group consisted of 15 men and 10 women, mean age 30.18 years i 9.48, age range 18 to 61 years. There was therefore no significant difference between the ages of the HIV and the control group (KruskalWallis test; p = 0.31). With the exception of the work done by Arendorf and Walker,4 results from previous studies showed no difference in the Candida carrier rate between smokers and nonsmokers.5-9 Smokers were therefore not excluded from this study. Two patients wore dentures, one from each group, and none of the patients had clinical signs of oral candidiasis at the time of sampling. Collection

of samples

Each person rinsed his or her mouth with 5 ml of sterile phosphate buffered saline solution for 1 minute.” The rinse was collected in sterile containers, and a 100 ~1 aliquot was plated onto Sabouraud dextrose agar. After 48-hours incubation at 37” C, the number of colony-forming units (CFU) per milliliter of rinse was determined. All the morphologically dif-

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Table 1. Culture of ICandida species and epithelial

Table III. C. albicans biotypes as determined by the

smears from the oral cavity of HIV-seropositive and healthy control persons

API 20 C system6 Biotypes

T$igir~

1 3 4 5 6 13 BS*

Candida Positive culture Mean CFUs Positive smears

21 (75%) 815.4 (SD = 1696.7) 11 (39%)

19 (68%) 239.3 (SD = 654.4) 11 (39%)

0.5 0.02

Candida albicans Candida stellatoidea Candida krzwei Candida tropicalis Torulopsis glabrata Torulopsis candida

HIV-positive (n = 21) 20 (95%) 1 (5%) 0 0 0 0

13 (65%) 0 1(5%) 0 4 (20%) 1 (5%) 1 (5%)

Corztrol (rz = 22) 12 (55%) P (5%) 1(5%) II (5%) 2 (9%) 0 5t (22%)

*BS = Special biotypes not described by Williamson et aL6 tAll five strains showed a similar carbohydrate assimilation pattern.

Table II. Candida species identification Species

HIV-positive (n = 20)

Control (n = 291% 22 (76%) 1 (3%) 1 (3%) 1 (3%) 2 (7%) 2 (7%)

*More than one Candido species/strain were isolated from five of the control persons.

ferent colonies were identified to species level with the aid of the germ-tube” and disk diffusion test.12 Biotype designation of Candida albicans isolates was done according to the method of Williamson et a1.13 and antifungal sensitivities were determined with the Candifast antifungal sensitivity kit (International Mycoplasma, ‘Toulon, France). Epithelial sIrnear, were taken from the posterior third of the tongue, stained with periodic acid-Schiff14 and examined microscopically for the presence of Candida spores and hyphae. RESULTS

Candida could be cultured from 21 of 28 (75%) HIV-positive patients and from 19 of 28 (68%) healthy persons (Table I). Positive smears were present in 11 persons from both groups (Table I). The mean CFU was significantly higher in the HIV-positive group (~1= 0.02) compared with the control group (Table I). Candida albicans represented 95% of the species isolated from the HIV-positive group and 76% of those from the healthy group (Table II) with biotype 1 the most prevalent phenotypic strain in both groups (Tagle III). Antifungal agent susceptibility patterns as det.ermined by the Candifast susceptibility kit are shown in Table IV. DISCUSSION

No significant difference in carrier rate was detected between the HIV seropositiveand the“healthy”

Table IV. Candida susceptibility patterns as determined by the Candifast susceptibility kit

L Amphotericin B Nystatin Fluorocytosine Econazole Ketokonazole Miconazole Fluconazole

19 (95%) 20 (100%) 19 (95%) 9 (45%) 20 (100%) 16 (80%) 16 (80%)

28 (97%) 29 (100%) 24 (83%) 13 (45%) 26 (90%) 20 (69%) 20 (69%)

control groups (p = 0.5). This latter finding is contrary to the results of Torssander et al.,’ who found the carrier rate of Candida in HIV-seropositive homosexual men significantly higher than the rates in HIV-seronegative homosexual and heterosexual men. The density (CFU) carriage for Candida in our study was, however, significantly (p = 0.02) higher in the HIV-seropositive group. Previous results showed a greater incidence of candidal carriers among healthy women than healthy men.4, 7,9, 15-17This could explain the relatively high carrier rate in our control group that had more women than the test group (10 versus 3). The higher density Candida carriage in the HIVpositive group suggests that HIV seropositivity may lead to changes in the oral environment and in local immunologic reactivity thus allowing greater numbers of Candida to colonize the oral cavity. This may explain the high incidence of oral candidiasis in HIVseropositive patients while still in the early stages of the disease. Mycelial findings in smears of subjects from both groups without any clinical symptoms of candidiasis confirm the results of Torssander et al.’ who suggested that mycelial carriers may possibly develop oral candidiasis more frequently in cases of impaired host defense. Another explanation is that

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the mycelial forms may indicate a transition from commensalismto parasitism in these specific subjects, as Torssander et al.’ reported positive smearsin orally symptomlessimmunocompromised patients followed by the development of clinically manifested infections. As in previous studies,18,l9 biotype 1 isolates of Candida albicans were the most frequently recovered phenotype, accounting for 65% and 55% of the HIVseropositive and control group, respectively. A percentage of the strains typed in the present study (14.3%) revealed distinctive carbohydrate assimilation patterns that were not described by Williamson et a1.13Korting et al. l8 also found strains with carbohydrate assimilation patterns that did not fit into the schemeof Williamson et a1.13To evaluate the importance of this feature and to compare results, Korting et a1.18suggestedthat additional biotyping should be done with systems having higher discriminatory power. Alternatively, it may be worthwhile to expand the scheme of Williamson et a1.,t3 a scheme that proved to be easy to use. Contrary to expectations, Candida isolates from both the HIV-seropositive and control groups showed resistanceto many of the antifungal drugs. Enormous discrepancies between azole anti-Candida MICS in vitro and in vivo have been reported previously.20-23 The Candifast antifungal sensitivity kit is relatively new on the market and not yet well-tested. It, however, appears as if this system is not a reliable in vitro test system for antifungal sensitivity. Results from this study suggest no link between biotypes as determined by the API 20 C system, antifungal agent sensitivity as determined by the Candifast antifungal sensitivity kit, and HIV-seropositivity. Asignificantly higherdensitycarriagefor Candida was seen in this group. We thank Mrs. I. Stander of the Medical Research Council for statistical analysis of the data, Dr. Spracklen of Somerset Hospital for his contributions, and Mrs. R. Blaauw for typing the manuscript. REFERENCES 1. Torssander J, Morfeldt-Manson L, Biberfeld G, Karlsson A, Putkonen PO, Wasserman J. Oral Candida albicans in HIV infection. Stand J Infect Dis 1987;19:291-5. 2. Klein RS, Harris CA, Small CB, Moll B, Lesser M, Freedland GH. Oral candidiasis in high-risk patients as the initial manifestation of the acquired immunodeficiency syndrome. N Eng J Med 1984;311:354-8. 3. Farmon J, Tavitian A, Rosenthal LE, et al. Focal esophageal

ORAL SURGERY ORAL MEDICINE ORAL PATHOLOGY November 1993 candidiasis in acquired immune deficiency syndrome. Gastrointest Radio1 1986;11:213-7. 4. Arendorf TM, Walker DM. Prevalence and intraoral distribution of Candida albicans in man. Arch Oral Biol 1980;25:110. 5. Gergely L, Uri J. Day by day variation in the mycotic flora of the mouth. Arch Oral Biol 1966;11:15-9. 6. Beasley JD. Smoking and oral moniliasis. J Oral Med 1969; 24:83-6. 7. Bastiaan RJ, Reade PC. The prevalence of Candida albicans in the mouths of tobacco smokers with and without oral mucous membrane keratoses. ORAL SURG ORAL MED ORAL PATHOL 1982;53:148-57. 8. Oliver DE, Shillitoe EJ. Effects of smoking on the prevalence and intraoral distribution of Candida albicans. J Oral Path 1984;13:265-70. 9. Masipa JN, Hauman CHJ, Raubenheimer EJ. Oral carriage of Candida species in patients visiting the Medunsa Dental Clinic. Tydsk Tandheelk Ver S. Afr. 1992;47:407-9. 10. Samaranayake LP, Lamb AB, McFarlane TW. Oral carriage of Candida species and coliforms in patients with burning mouth syndrome. J Oral Path Med 1986;18:233-5. 11. Rippon JW. Medical Mycology. Philadelphia: WB Saunders, 1988:566. 12. Sobczak H. A simple disk diffusion test for differentiation of veast soecies. J Med Microbial 1985;20:307-16--. 13. Williamson MI, Samaranayake LP, MacFarlane TW. Biotypes of Candida albicans using the API 20 C system. FEMS Microbial Lett 1986;37:27-9. 14. Culling CFA. Handbook of histopathologicai and histochemical techniques. London: Butterworths, 1974:268. 15. Todd RL. Studies on yeast-like organisms isolated from the mouths and throats of normal persons. Am J Hyg 1937;25:21220. 16. Lilienthal B. Studies of flora of the mouth. HI: Yeastlike organisms-some observations on their incidence in the mouth. Aust J Exper Biol & M SC 1950;28:279-86. 17. Barlow AJE, Chattaway FW. Observations on the carriage of Candida albicans in man. Br J Dermatol 1969:X1 :I 03-6. 18. Korting HC, Allert M, Georgic A, Frosch M. In vitro susceptibilities and biotypes of Candida albicans isolates from the oral cavities of patients infected with human immunodeficiency virus. J Clin Microbial 1988;26:2626-3 1. 19. Franker CK, Lucartorto FM, Johnson BS, Jacobson JJ. Characterization of the mycoflora from oral mucosal surfaces of some HIV-infected patients. ORAL SURG ORAL MED ORAL PATHOL 1990;69:683-7. 20. Heeres J, Backz LJJ, Van Cutzen J. Antimycotic azoles 7: Synthesis and antifungal properties of a series of recent triazol-3-ones. J Med Chem 1984;27:894-900. 2 1. Heel RC. Ketoconazole: a review of its therapeutic efficacy in superficial and systemic fungal infections. Drugs 1982;9:42447. 22. Ryley JF, Wilson RG, Gravestock MB, Poyser JP. Experimental approaches to antifungal chemotherapy. Adv Pharmaco1 Chemother 1981;18:49-176. 23. Odds FC. Laboratory tests for the activity of imidazole and triazole antifungal agents in vitro. Dermatologica 1985;4:26070. Reprint requests: C.H.J. Hauman, B. Medsci, M. Medsci, BCHD Department of Oral Pathology Private Bag Xl Tygerberg, South Africa 7505