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Journal of Clinical Microbiology, January 1998, p. 216-218, Vol. 36, No. 1
0095-1137/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.
Confirmation of the Distinct Genotype Groups
within the Form Species Candida parapsilosis
Basabi
Roy and
Sally A.
Meyer*
Department of Biology, Georgia State
University, Atlanta, Georgia 30303
Received 19 June 1997/Returned for modification 18 August
1997/Accepted 13 October 1997
 |
ABSTRACT |
The DNA relatedness and restriction fragment length
polymorphism patterns of whole-cell DNA and the general phenotypic
properties of 14 isolates of the form species Candida
parapsilosis representing three diverse genetic groups were
compared. The data confirm the unrelatedness of the three groups at the
species level.
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INTRODUCTION |
In recent years, Candida
parapsilosis, a form species of the Fungi Imperfecti defined by
morphological and physiological properties, has been increasingly
implicated as a cause of fungemia, endocarditis in neutropenic or
severely debilitated patients, and candidal vaginitis (22).
The species has been commonly associated with the skin and fingernails
of humans (18), and it has been isolated from soil and fresh
and marine waters (1, 3, 13).
Genetic heterogeneity has been reported within this species in several
studies. Scherer and Stevens (16) divided their clinical isolates of C. parapsilosis into three groups on the basis
of restriction fragment length polymorphisms (RFLPs). The data from this study suggested that C. parapsilosis is genotypically
more heterogeneous than Candida albicans. Lehmann et al.
(6), on the basis of randomly amplified polymorphic DNA
profiles, also observed three distinct groups among isolates of
C. parapsilosis. Group I and II isolates of the latter study
shared the API 20C (Analytab, Plainview, N.Y.) code 6756171, whereas
the two isolates in group III had the biotype code 6776171. Further
studies of representative strains from these three groups indicated
that each group included isolates with distinctive karyotypes
(9). Lin et al. (8) examined 45 clinical
isolates that were physiologically similar to C. parapsilosis as determined with the API 20C kit and reported the
presence of three distinct isoenzyme-defined groups that were also
distinguishable on the basis of sequence analysis of the internally
transcribed spacer sequences flanking the 5.8S RNA gene.
In this investigation, we examined the nuclear DNA base composition,
DNA-DNA reassociation, and DNA RFLPs of 13 isolates of C. parapsilosis from diverse sources (including representatives of
groups I to III mentioned above) and the type strain.
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MATERIALS AND METHODS |
Yeast strains.
The strains examined in this study are listed
in Table 1. They include the type strain of C. parapsilosis
and representative strains of the three groups reported by Lin et al.
(8). The standard morphological and physiological
characterizations of the yeasts were performed according to the methods
described by van der Walt and Yarrow (21).
DNA base composition and reassociation studies.
DNA was
isolated and purified by the method of Marmur (10) as
modified by Meyer and Phaff (12). DNA base composition
(mol% G+C) was determined by the thermal denaturation method as
described by Marmur and Doty (11). The optical renaturation
method of Seidler and Mandel (17) as modified by Kurtzman et
al. (5) was employed for the DNA reassociation studies.
RFLPs.
DNA was extracted and purified by the methods of Su
and Meyer (19) and Scherer and Stevens (16), and
restriction enzyme analysis was performed with HinfI,
HindIII, PvuII, and HaeIII restriction endonucleases (Boehringer Mannheim GmbH, Mannheim, Germany). The DNA fragments were separated in a 1% agarose gel in
Tris-acetate-EDTA buffer (40 mM Tris-OH, 20 mM acetic acid, 2 mM
Na2EDTA [pH 8.1]) (2).
| |
RESULTS |
Percent DNA relatedness divided the isolates into three
distinct groups, each of which agreed with the molecular
characteristics of the representative cultures of the groups
established by Lin et al. (8). There was greater than 95%
DNA relatedness among isolates within a group and less than 25%
relatedness between isolates from different groups (Table
2). The G+C contents of the isolates were
40.2 to 41.5 mol% (group I), 38.5 to 40.5 mol% (group II), and 39.4 to 40.7 mol% (group III) (Table 1).
Group I isolates all had similar RFLP patterns after digestion with
HinfI (Fig. 1). Polymorphisms
were evident in this group after double digestion with
HindIII and HaeIII (Fig.
2) and HindIII and
PvuII (Fig. 3). The RFLP
patterns of representative isolates of each group after digestion with
HinfI (data not shown) and the double digestions
demonstrated RFLP distinctions among the three groups (Fig.
4 and 5).
No polymorphisms were observed with the restriction digest patterns of
isolates of group II and group III.
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FIG. 1.
C. parapsilosis group I RFLP patterns from
HinfI digests. Lanes: M,  DNA digested with
HindIII; A, ATCC 22019; B, NRRL Y-10022; C, NRRL
Y-10208; D, NRRL Y-9282; E, NRRL Y-10518; F, CBS 2215; G, CBS 8181.
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FIG. 2.
C. parapsilosis group I RFLP patterns from
HindIII-HaeIII digests. Lanes: M, DNA
digested with HindIII; A, ATCC 22019; B, NRRL Y-10022;
C, NRRL Y-10208; D, NRRL Y-10518; E, CBS 2215; F, NRRL Y-9282; G, CBS
8050; H, CBS 8181.
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FIG. 3.
C. parapsilosis group I RFLP patterns from
HindIII-PvuII digests. Lanes: M, DNA
digested with HindIII; A, ATCC 22019; B, NRRL Y-10022;
C, NRRL Y-10208; D, NRRL Y-10518; E, CBS 2215; F, NRRL Y-9282; G, CBS
8050; H, CBS 8181.
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FIG. 4.
C. parapsilosis RFLP profiles from
HindIII-HaeIII digests. Lanes: M, DNA
digested with HindIII; A and B, group I ATCC 22019 and
CBS 8050, respectively; C and D, group II MCO 452 and NRRL Y-9536,
respectively; E and F, group III MCO 429 and MCO 448, respectively.
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FIG. 5.
C. parapsilosis RFLP profiles from
HindIII-PvuII digests. Lanes: M, DNA
digested with HindIII; A and B, group I ATCC 22019 and
CBS 8050, respectively; C and D, group II MCO 452 and NRRL Y-9536,
respectively; E and F, group III MCO 429 and MCO 448, respectively.
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DISCUSSION |
Our DNA relatedness and RFLP analyses support the earlier
divisions of isolates classified as C. parapsilosis into
three separate groups. The low degree of DNA relatedness among the
groups indicates that they represent distinct species. Although two of
these groups (II and III) have been recognized only within the past
decade, one of the group II cultures was originally isolated in 1966 from the Antarctic Peninsula (possibly from human or animal pollution). Our recent clinical isolates (1996) represented group I and group III.
We do not have sufficient information for the phenotypic differentiation of these genetically distinct yeasts, nor can we
currently discern if differences in their epidemiology exist.
The finding of genetically diverse groups among the form species of the
imperfect yeasts is not unusual. Lehmann et al. (7) found
two distinct genetic groups within Candida haemulonii. The phenotype of Candida famata is found among strains of
Candida guilliermondii and several species of
Debaryomyces (14). A variety of distinct
genotypes are present within the general phenotype of
Trichosporon cutaneum (15). Most recently,
C. albicans was found to include a diverse genetic group
which was designated a distinct species, Candida
dubliniensis (20). The application of molecular
procedures to yeast systematics has resulted in major reclassifications
of many species and genera (4, 13). It is obvious that
understanding the genetic diversity of the yeasts associated with human
infections requires further study.
| |
ACKNOWLEDGMENT |
We offer special thanks to D. G. Ahearn, Georgia State
University, Atlanta, Ga., for his guidance and recommendations in this research and the preparation of the manuscript.
| |
FOOTNOTES |
*
Corresponding author. Mailing address: Dept. of
Biology, Georgia State University, Atlanta, GA 30303. Phone: (404)
651-2260. Fax: (404) 651-2509. E-mail:
biosam{at}panther.gsu.edu.
| |
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Journal of Clinical Microbiology, January 1998, p. 216-218, Vol. 36, No. 1
0095-1137/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.
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Abstract
Introduction
