Division of Infectious Diseases, Department of Medicine, Santa Clara Valley ... Geographic Medicine, Department of Medicine, Stanford University School of ..... High- prevalence of Candida spp. on hands of health care workers (HCWs) in.
JOURNAL OF CLINICAL MICROBIOLOGY, Aug. 1995, p. 2224–2226 0095-1137/95/$04.0010 Copyright 1995, American Society for Microbiology
Vol. 33, No. 8
Genotypic Analysis of Pseudoepidemic Due to Contamination of Hanks’ Balanced Salt Solution with Candida parapsilosis STANLEY C. DERESINSKI,1,2,3,4,5* KARL V. CLEMONS,1,2,5 CAROL A. KEMPER,1,2,3,4,5 KARI ROESCH,4 BARBARA WALTON,4 AND DAVID A. STEVENS1,2,5 Division of Infectious Diseases, Department of Medicine, Santa Clara Valley Medical Center,1 and California Institute for Medical Research,5 San Jose, California 95128; Division of Infectious Diseases and Geographic Medicine, Department of Medicine, Stanford University School of Medicine, Stanford, California 943052; and AIDS Community Research Consortium3 and Sequoia District Hospital,4 Redwood City, California 94063 Received 30 January 1995/Returned for modification 3 March 1995/Accepted 13 May 1995
A cluster of isolates of Candida parapsilosis recovered from clinical specimens was demonstrated, by both classical and molecular epidemiological techniques, to have resulted from contamination in the laboratory. The source of the pseudoepidemic was a repeatedly utilized contaminated container of Hanks’ balanced salt solution used in specimen processing. The patterns of restriction fragment length polymorphisms of DNA extracted from the clinical and environmental isolates were identical to each other but composed a newly identified unique C. parapsilosis DNA type. effusion (Table 1). No other organisms were recovered from any of the specimens. In each case, no organism was seen on Gram stain and small numbers (1 to 10 colonies) of yeast cells grew in culture. Culture of specimens from two additional sites, fluid from a pacemaker generator pocket and pleural fluid from patient C, yielded no organisms. Epidemiological investigation revealed that the medical procedures, location, and physician and nursing staff differed in each case. No break in surgical or laboratory technique was
Fungal diseases, particularly nosocomially acquired candidiasis, are recognized with increasing frequency as an important cause of hospital-acquired infection (6). While most such infections are caused by Candida albicans, other species are also commonly involved. Among these is Candida parapsilosis, an organism which has caused a wide variety of hospital-acquired infections, including a number of multipatient outbreaks (22). We recently encountered a cluster of isolates of C. parapsilosis recovered from clinical specimens that proved to be the result of contamination during laboratory processing rather than of patient infection. (These data were presented in part at the 34th Interscience Conference on Antimicrobial Agents and Chemotherapy, Orlando, Fla., 7 October 1994.) Materials and methods. The isolates (6 clinical isolates from 5 patient specimens and 1 environmental isolate) were identified by using the Baxter MicroScan Yeast Identification Panel (12). Restriction enzyme analysis. Isolates were characterized by restriction enzyme analysis of restriction fragment length polymorphisms (RFLP) in comparison with each other and with archival isolates of C. parapsilosis as previously described (1, 16). Results. C. parapsilosis was isolated on routine primary isolation media from six clinical specimens obtained over a period of 20 days from five patients (Table 1). One patient (patient C), who was febrile and in whom infection had been suspected, was initially treated with amphotericin B for 5 days, at which time it was apparent to the clinician that the isolation of the organism was not indicative of infection but was, in fact, part of a pseudoepidemic, and administration of amphotericin B was discontinued. No other patient received antifungal therapy. None of the five patients had clinical evidence of C. parapsilosis infection. Five of the specimens were of solid material; a sixth was of 0.1 ml of bloody material in a syringe used to aspirate a hip
FIG. 1. Gel electrophoresis of EcoRI restriction digests of DNA from isolates of C. parapsilosis. Lanes: M, HindIII digest of l DNA; 1, 2, 10, and 11, C. parapsilosis DNA types described previously (16): group VII (subgroup 1), group VI (subgroup 2), group V (subgroup 1), and group V (subgroup 2), respectively; 3, from patient B; 4, from patient E; 5, from patient D; 6, from patient A; 7 and 8, from patient C; and 9, HBSS.
* Corresponding author. Mailing address: Division of Infectious Diseases, Santa Clara Valley Medical Center, 751 S. Bascom Ave., San Jose, CA 95128. Phone: (408) 885-4301. Fax: (408) 885-4306. 2224
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TABLE 1. Clinical data Patient
Diagnosis
Procedure
Place of procedure
C. parapsilosis-positive specimen
A B C D E
Aortic valve disease Poor healing, postoperative wound Suspected pacemaker infection Suspected hip sepsis Lymphadenopathya
Aortic valve replacement Wound closure Removal of pacemaker, thrombectomy Needle aspiration Lymph node excision
Operating room Operating room Operating room Patient room Ambulatory surgery
Aortic valve Shoulder tissue Clot, tissue from subclavian vein Bloody fluid Lymph node
a
Due to toxoplasmosis.
reported. Aseptic technique was used, including sterile gloves, scissors, forceps, and disposable pipettes. Solid specimens were ground in a sterile tissue grinder. Hanks’ balanced salt solution (HBSS) was added to the tissue for grinding. HBSS had also been used to flush material from the syringe in the single case in which a liquid specimen was cultured. HBSS was ordinarily purchased by the laboratory in 50-ml bottles. However, because of unavailability of this product size, a 500-ml bottle was purchased instead and had been in use for an estimated 1 month at the time of the incident. Culture of the contents of this bottle yielded heavy growth of C. parapsilosis. This isolate and the six obtained from the patient specimens were all MicroScan ID Biotype 544-020-00. There were no other isolations of C. parapsilosis by this laboratory in the 6 months before or after the outbreak. The RFLP patterns resulting from EcoRI digestion of the cellular DNA of the seven C. parapsilosis isolates revealed them to be identical to each other, thus indicating a common origin (Fig. 1). The identity of these isolates was further confirmed by restriction with a second enzyme, HindIII (data not shown). Comparison of the EcoRI-generated RFLP patterns of the seven outbreak isolates with those from archival isolates of C. parapsilosis allowed their placement within the previously described DNA typing scheme of Scherer and Stevens (16). The comparisons demonstrated that the seven outbreak isolates were identical but unique in their RFLP pattern when compared with previously described strains (16). Of the DNA types initially reported (16), the outbreak isolates appear to be a member of group VII. In this reference system (16), groups are defined by the brightly stained bands and subgroups are differentiated by comparisons using the entire RFLP pattern. Three subgroups of group VII have now been identified (2, 16), of which the outbreak isolates are a newly described DNA type (group VII, subgroup 3). Discussion. This cluster of C. parapsilosis recovered from clinical specimens was demonstrated by both classical and molecular epidemiological techniques to represent a pseudoepidemic. These results are compatible with the hypothesis that the isolates tested arose from a point source and that that source was the HBSS used to process the cultures. A number of pseudoepidemics involving a variety of contaminating microorganisms have been reported, including some due to yeast-like organisms, including uncommonly isolated ones such as Trichosporon beigelii and Rhodotorula rubra (5, 20). In one instance, visualization of nonviable yeast-like structures on Gram stain of surgical specimens proved to be due to the presence of nonviable organisms in sterilized surgical glove powder that had been left in the surgical site (3). A number of true hospital outbreaks of fungemia due to C. parapsilosis have been reported, including ones due to contamination of intravenous device equipment such as pumps and pressure transducers (17, 18). An outbreak of endophthalmitis
due to C. parapsilosis in patients who had undergone cataract extraction and intraocular lens implantation resulted from contamination of a balanced salt solution used intraoperatively from irrigation of the surgical site. The isolates from the patients and the balanced salt solution were found to be identical by antifungal susceptibility pattern and enzyme profile (7, 10). The source of contamination of the HBSS in our pseudoepidemic is unknown. C. parapsilosis, in contrast to C. albicans, can often be isolated from natural environmental sources, especially soil, water, and plants (4, 13, 21). In one instance, contaminated carbon dioxide incubators were the probable source of culture contamination with C. parapsilosis (15). Laboratory and other health-care workers are potential sources of contamination of microbiological specimens. C. parapsilosis is commonly found on integumentary structures of humans (9). In one study, yeasts were isolated from the subungual space of the hands of 69.0% of 26 adult volunteers; 51.3% of the yeasts were identified as C. parapsilosis (8). Another study found that 29 to 39% of health-care workers in two intensive care units had hand colonization with Candida spp., 46% of which were C. parapsilosis (11). Hand carriage was implicated, along with environmental contamination, in the acquisition of C. parapsilosis, which were demonstrated by restriction enzyme analysis to be identical, by patients in a bone marrow transplant unit and an intensive care unit (14). The DNA RFLP technique utilized in our study is sufficiently sensitive to detect multiple polymorphisms of C. parapsilosis (16). With this method, and discounting multiple isolates of the same type from an epidemiological cluster, a total of eight different types of C. parapsilosis have now been distinguished among 13 epidemiologically unrelated clusters or individual isolates (2, 16, 19). The present study strongly indicated the common identity of the isolates, since their C. parapsilosis genotype had not been encountered in previous studies and presumably represents a less common type of C. parapsilosis. We demonstrated, by both classical and molecular epidemiological techniques, that a cluster of isolates of C. parapsilosis recovered from clinical specimens resulted from contamination in the laboratory. Rapid recognition of the event described here as a pseudoepidemic prevented the unnecessary administration of antifungal therapy to all but one patient. This episode demonstrates the potential for contamination of ordinarily sterile container contents which are repeatedly utilized. Such solutions, whether used for culture processing or otherwise, should be stored in small volumes to limit opportunities for contamination. We acknowledge the bibliographic assistance of Karen W. Moody, Sequoia Hospital, Redwood City, Calif., as well as the contribution of Pilsang Park, California Institute for Medical Research, San Jose. Salary support (C.A.K.) was in part provided by the State of California’s Universitywide AIDS Research Program (UARP DD93-SD132) through the California Collaborative Treatment Group.
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