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Previous Article | Next Article 
Journal of Clinical Microbiology, April 1998, p. 1035-1038, Vol. 36, No. 4
0095-1137/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.
Intergenic Transcribed Spacer PCR Ribotyping for Differentiation
of Saccharomyces Species and Interspecific
Hybrids
Michael J.
McCullough,1,2,3
Karl V.
Clemons,1,2,3
John H.
McCusker,4 and
David A.
Stevens1,2,3,*
Department of Medicine, Division of
Infectious Diseases and Geographic Medicine, Stanford University School
of Medicine, Stanford, California 943051;
California Institute for Medical
Research2 and
Department of Medicine,
Division of Infectious Diseases,3 Santa Clara
Valley Medical Center, San Jose, California 95128; and
Department of Microbiology, Duke University Medical Center,
Durham, North Carolina 277104
Received 23 July 1997/Returned for modification 18 November
1997/Accepted 14 January 1998
 |
ABSTRACT |
The taxonomy of the genus Saccharomyces has undergone
significant changes recently with the use of genotypic rather
than phenotypic methods for the identification of strains to the
species level. The sequence of rRNA genes has been utilized for the
identification of a variety of fungi to the species level. This
methodology, applied to species of Saccharomyces, allows
unknown Saccharomyces isolates to be assigned to the type
strains. It was the aim of the present study to assess whether typing
of the intergenic spacer region by using restriction fragment
length polymorphisms of PCR products (intergenic transcribed spacer PCR
[ITS-PCR] ribotyping) could distinguish among type strains of the 10 accepted species of Saccharomyces and further to assess if
this method could distinguish strains that were interspecific hybrids.
Cellular DNA, isolated after the lysis of protoplasts, was amplified by
PCR using ITS1 and ITS4 primers, purified by liquid chromatography, and
digested with restriction endonucleases. Ribotyping patterns using the restriction enzymes MaeI and HaeIII could
distinguish all species of Saccharomyces from each other,
as well as from Candida glabrata, Candida
albicans, and Blastomyces dermatitidis. The only
exception to this was the inability to distinguish between
Saccharomyces bayanus and S. pastorianus
(S. carlsbergensis). Furthermore, interspecific hybrids
resulting from the mating of sibling species of
Saccharomyces were shown to share the ITS-PCR ribotyping
patterns of both parental species. It should now be possible, by this
simple PCR-based technique, to accurately identify these strains to the
species level, thereby allowing an increase in our understanding of the
characteristics required by these interspecific hybrids for
their particular ecological niches.
| |
INTRODUCTION |
The taxonomy of the genus
Saccharomyces has undergone significant changes
recently with more importance being placed on genotypic rather
than phenotypic methods for the identification of strains to the
species level. This has resulted in a change in the number of
recognized species of Saccharomyces moving from 41 (15) to 7 (3) or 10 (20).
The DNA that encodes the ribosomal RNA (rDNA) has been utilized by many
investigators for the determination of species of a wide variety of
yeasts and fungi (1, 7, 11, 14, 21, 27). These methods rely
on the conserved nature of rDNA such that isolates from the same
species maintain the same sequence, whereas the more phylogenetically
diverse the species is, the greater the difference in the sequences of
rDNA is. The present methodology has utilized restriction fragment
length polymorphisms (RFLPs) of PCR products obtained by using primers
that specifically amplify the region spanning from the 3' end of the
18S rDNA to the 5' end of the 25S rDNA. This region includes the 5.8S
rDNA and the two intergenic transcribed spacer (ITS) regions ITS1 and ITS2 (26).
This methodology has been applied to Saccharomyces species
for the authentication of strains in the Saccharomyces
cerevisiae complex (10). These authors found that they
could separate strains of S. cerevisiae from those of the
S. bayanus-S. pastorianus (S. carlsbergensis) cluster; however strains in this cluster could not
be separated from each other by this method (10). No other Saccharomyces species were investigated in this previous
work.
More recently, Messner and Prillinger (17) have used a
similar yet more thorough method for the differentiation of 10 genotypically distinct Saccharomyces species. In their
study, the complete chromosomal ribosomal repeat was amplified in two
parts, the 18S rDNA, including both ITS regions and the 5.8S rDNA
(total length, 2,600 bp), and separately the 25S rDNA (length, 3,300 bp) (17). Restriction fragments generated from these two PCR
products by nine restriction endonucleases yielded characteristic
patterns by which unknown isolates of Saccharomyces could be
assigned to the type strains (17).
Isolates of Saccharomyces are becoming of increasing medical
importance due to their recently appreciated pathogenic potential (4, 5, 16). It was the aim of the present study to assess if
the simpler PCR-based method (10) could distinguish among type strains of the 10 accepted species of Saccharomyces
(20). A further aim was to assess whether this method could
distinguish isolates which were interspecific hybrids resulting from
the mating of two closely related yet distinct species of
Saccharomyces. Isolates that do not sporulate or do not form
viable spores cannot be identified to the species level by classical
genetic techniques (19), as these strains cannot be
separated from nonmating asporogenous strains of either parental
species.
| |
MATERIALS AND METHODS |
The yeast strains examined in this study are listed in Table
1. All strains were obtained from the
American Type Culture Collection, except for the interspecific hybrid
strains and the Candida glabrata and Candida
albicans isolates, which were wild-type (clinical) isolates
collected by the California Institute for Medical Research laboratory
and identified to the species level by routine phenotypic methods.
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TABLE 1.
Lanes, species, and reference numbers of the strains
utilized for the assessment of ITS-PCR ribotyping for the
identification of Saccharomyces isolates to the
species level
|
|
Cellular DNA was isolated by previously described methods (22,
23).
Primers for the amplification of the 5.8S rDNA were ITS1 (5'-TCC
GTA GGT GAA CCT GCG G-3') and ITS4 (5'-TCC TCC GCT TAT TGA TAT GC-3'). DNA was amplified in the buffer supplied by the
Taq polymerase manufacturer (Gibco BRL, Grand Island, N.Y.)
in a 100-µl volume containing 1 µM primers, 1.5 mM
MgCl2, 2.5 U of Taq polymerase, 200 µM dATP,
200 µM dCTP, 200 µM dGTP, and 200 µM dTTP. The reactions were
performed in an automated thermal cycler (GeneMate; Lab-Line Instruments, Melrose Park, Ill.). DNA samples were denatured by incubation for 3 min at 94°C before 30 cycles of 94°C for 1 min, 60°C for 1 min, and 72°C for 2.5 min. After PCR, amplified DNA was
purified by liquid chromatography (Wizard PCR Preps; Promega, Madison,
Wis.).
For RFLP analysis, the purified PCR products were digested with one of
the following restriction enzymes: MaeI, HaeIII,
CfoI, DdeI, BglII, BamHI,
HindIII, EcoRI, SmaI, or
PstI (Boehringer Mannheim, Indianapolis, Ind.). Digestions
were done by overnight incubation with 10 U of enzyme at 37°C.
Aliquots (10 µl) of the PCR products, with and without endonuclease
digestion, were then analyzed by electrophoresis through a 3% (wt/vol)
agarose gel (2% Nusieve GTG, 1% SeaKem Gold [FMC BioProducts,
Philadelphia, Pa.]) in TAE buffer (40 mM Tris-acetate, 0.2 mM EDTA)
for 2 h at 10 V/cm and visualized by UV transillumination at 302 nm after ethidium bromide staining.
| |
RESULTS |
The ITS region amplified by PCR showed a product of
approximately 850 bp for S. cerevisiae, S. pastorianus (S. carlsbergensis), S. paradoxus, and S. bayanus and for the
interspecific hybrids S. paradoxus/S. cerevisiae
and S. bayanus/S. cerevisiae (Fig. 1, lanes 1 to 10). The ITS-PCR products
for the remainder of the Saccharomyces species varied
between approximately 660 and 760 bp (Fig. 1, lanes 11 to 16). The size
of the ITS-PCR products for C. glabrata, C. albicans, and Blastomyces dermatitidis were approximately 850, 520, and 640 bp, respectively (Fig. 1, lanes 17 to
19).
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FIG. 1.
Photograph of ethidium bromide-stained,
UV-transilluminated PCR products after electrophoresis within a 3%
agarose gel. The order of the isolates is given in Table 1.
|
|
Of the 10 restriction endonucleases used, only two were able to
discriminate among all species tested. Utilization of the RFLPs
generated by these enzymes, MaeI and HaeIII, made
it possible to distinguish all species of Saccharomyces from
each other as well as from C. glabrata, C. albicans, and B. dermatitidis, with the exception of
the nonseparation of S. pastorianus (S. carlsbergensis) from S. bayanus (Fig.
2 and 3,
lanes 7 and 8 and lanes 4 and 5, respectively).
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FIG. 2.
Photograph of ethidium bromide-stained,
UV-transilluminated MaeI-digested PCR
products after electrophoresis within a 3% agarose gel. The DNA from
the PCR (Fig. 1) was purified by the Wizard PCR Preps purification
system prior to overnight digestion with 10 U of the restriction
endonuclease MaeI. The lane order is the same as that in
Fig. 1.
|
|
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FIG. 3.
Photograph of ethidium bromide-stained,
UV-transilluminated, HaeIII-digested PCR products after
electrophoresis within a 3% agarose gel. The DNA from the PCR (Fig. 1)
was purified by the Wizard PCR Preps purification system prior to
overnight digestion with 10 U of the restriction endonuclease
HaeIII. The lane order is the same as that in Fig. 1.
|
|
The ITS-PCR ribotyping pattern obtained with the restriction
enzyme MaeI was found to separate all species of
Saccharomyces with the exception of three pairs,
S. cerevisiae-S. bayanus, S. cerevisiae-S. pastorianus (S. carlsbergensis), and S. pastorianus (S. carlsbergensis)-S. bayanus (Fig.
2).
The ITS-PCR ribotyping pattern obtained with HaeIII could
separate all species of Saccharomyces with the exception of
three pairs of species, S. cerevisiae-S.
paradoxus, S. pastorianus (S. carlsbergensis)-S. bayanus, and S. castellii-S. dairensis (Fig. 3).
Therefore, the combined information from both of these restriction
enzymes, MaeI and HaeIII, made it possible to
separate all species of Saccharomyces except
S. pastorianus (S. carlsbergensis) and S. bayanus (Fig. 2
and 3) from each other. The inability to separate these two species was
found with all 10 restriction endonucleases assessed.
The ITS-PCR ribotyping of interspecific hybrids was then determined.
The RFLPs generated by MaeI showed that the interspecific hybrid S. paradoxus/S. cerevisiae shared banding
patterns with both of the parental species (Fig. 2) and was therefore
distinguishable from both these parental species as well as the other
species of Saccharomyces. The RFLPs generated by
HaeIII showed that the interspecific hybrid S. bayanus/S. cerevisiae shared banding patterns with both of
these species (Fig. 3) and was therefore distinguishable from both its
parental species as well as from other species.
| |
DISCUSSION |
The results of the present study show that ITS-PCR ribotyping is a
simple method that can distinguish among all Saccharomyces species other than S. bayanus-S. pastorianus (see
below). These findings extend the usefulness of this technique from
previous investigations (10) and confirm the findings of
Messner and Prillinger (17) while simplifying the
methodology by markedly decreasing the size of the PCR amplicon
required to successfully differentiate these species of
Saccharomyces.
By this method it was not possible to distinguish between the type
strains of S. pastorianus (S. carlsbergensis) and S. bayanus. This finding is
consistent with earlier studies showing that these two species are very
closely related (2, 10, 24). Our results do not support the
previously proposed hypothesis that S. pastorianus (S. carlsbergensis) is a separate species or an
interspecific hybrid of S. cerevisiae with another
sibling species (8, 25). Previous research has shown that
S. pastorianus (S. carlsbergensis) carries two copies of certain genes, one that is identical to that of
S. cerevisiae and a second gene, which is distinct. The genes that have been most extensively studied are MET2,
which encodes homoserine acetyltransferase (9), the gene
encoding acetyl-coenzyme A-binding protein (6), and
ATF1, which encodes alcohol acetyltransferase
(13). The results from this previous research are consistent
with the proposal that S. pastorianus (S. carlsbergensis) originated as a hybrid between
S. cerevisiae and a sibling species, such as
S. monacensis or S. bayanus
(6, 9, 13). However, the results of the present study show a clear difference between the ITS-PCR ribotyping pattern of the interspecific hybrid S. cerevisiae/S. bayanus and
the ITS-PCR ribotyping patterns of S. pastorianus,
S. bayanus, and S. cerevisiae. Other
studies support such differentiation (18). It may well be
that deletion of one set of the rDNA has occurred in the type strains
of S. pastorianus (S. carlsbergensis)
used in the present study, resulting in an ITS-PCR ribotyping pattern
identical to that of S. bayanus. This process has been
previously postulated to occur in species of Saccharomyces
(12). Part of the difficulty in interpreting these data
arises from the terminology used by various authors. For example, the
species S. monacensis has been abolished and replaced
by S. pastorianus (S. carlsbergensis)
(3). Future studies, utilizing sequence data from a number
of regions of the rDNA, are necessary to definitively establish the
taxonomic relationship between the species of Saccharomyces.
The finding of the present study that interspecific hybrids resulting
from the mating of two closely related yet distinct species of
Saccharomyces share the ITS-PCR ribotyping patterns of both
parental species allows the previously not possible differentiation of
isolates (19). It may well be that some strains of
Saccharomyces occurring in nature, used commercially in
industry, promoted for human consumption for health reasons, or acting
as human pathogens are the result of interspecific mating. These hybrid
strains may be phenotypically better adapted for certain ecological
niches and have hitherto been designated as S. cerevisiae due to the difficulty in the conventional methodologies
required for correct identification to the species level. It should now
be possible, by this simple PCR-based species identification technique,
to accurately delineate these strains, thereby allowing an increase in
our understanding of the characteristics required for each of these
particular ecological niches.
| |
ACKNOWLEDGMENTS |
This research was funded in part by a Fellowship from the
Commonwealth AIDS Research Grants Committee of the National Health and
Medical Research Council of the Australian Federal Government.
| |
FOOTNOTES |
*
Corresponding author. Mailing address: Division of
Infectious Diseases, Santa Clara Valley Medical Center, 751 South
Bascom Ave., San Jose, CA 95128. Phone: (408) 885-4313. Fax: (408)
885-4306. E-mail:
oflahren{at}wpgate.hhs.co.santa-clara.ca.us.
| |
REFERENCES |
| 1.
|
Baleiras Couto, M. M.,
J. T. Vogels,
H. Hofstra,
J. H. Huis in't Veld, and J. M. Vossen.
1995.
Random amplified polymorphic DNA and restriction enzyme analysis of PCR amplified rDNA in taxonomy: two identification techniques for food-borne yeasts.
J. Appl. Bacteriol.
79:525-535[Medline].
|
| 2.
|
Banno, I., and Y. Kaneko.
1989.
A genetic analysis of taxonomic relation between Saccharomyces cerevisiae and Saccharomyces bayanus.
Yeast
5:S373-S377.
|
| 3.
|
Barnett, J. A.
1992.
The taxonomy of the genus Saccharomyces Meyen ex Rees: a short review for the non-taxonomists.
Yeast
8:1-23.
|
| 4.
|
Clemons, K. V.,
J. H. McCusker,
R. W. Davis, and D. A. Stevens.
1994.
Comparative pathogenesis of clinical and nonclinical isolates of Saccharomyces cerevisiae.
J. Infect. Dis.
169:859-867[Medline].
|
| 5.
|
Clemons, K. V.,
J. H. McCusker,
R. W. Davis, and D. A. Stevens.
1997.
Saccharomyces cerevisiae and the host-fungus interplay, p. 193-198.
In
H. Vanden Bossche, D. A. Stevens, and F. C. Odds (ed.), Host-fungal interplay. National Foundation for Infectious Diseases, Bethesda, Md.
|
| 6.
|
Fujii, T.,
N. Nagasawa,
A. Iwamatsu,
T. Bogaki,
Y. Tamai, and M. Hamachi.
1994.
Molecular cloning, sequence analysis, and expression of the yeast alcohol acetyltransferase gene.
Appl. Environ. Microbiol.
60:2786-2792[Abstract/Free Full Text].
|
| 7.
|
Fujita, S.,
B. A. Lasker,
T. J. Lott,
E. Reiss, and C. J. Morrison.
1995.
Microtitration plate enzyme immunoassay to detect PCR-amplified DNA from Candida species in blood.
J. Clin. Microbiol.
33:962-967[Abstract].
|
| 8.
|
Hammond, J. R. M.
1993.
Brewer's yeasts, p. 8-11.
In
A. H. Rose, and J. S. Harrison (ed.), The yeasts. Academic Press, New York, N.Y.
|
| 9.
|
Hansen, J., and M. C. Kielland-Brandt.
1994.
Saccharomyces carlsbergensis contains two MET2 alleles similar to homologues from S. cerevisiae and S. monacensis.
Gene
140:33-40[Medline].
|
| 10.
|
Huffman, J. L.,
F. I. Molina, and S. C. Jong.
1992.
Authentication of ATCC strains in the Saccharomyces cerevisiae complex by PCR fingerprinting.
Exp. Mycol.
16:316-319.
|
| 11.
|
Johnston, C. G., and S. D. Aust.
1994.
Detection of Phanerochaete chrysosporium in soil by PCR and restriction enzyme analysis.
Appl. Environ. Microbiol.
60:2350-2354[Abstract/Free Full Text].
|
| 12.
|
Kielland-Brandt, M. C.,
T. Nilsson-Tillgren,
C. Gjermansen,
S. Holmberg, and M. B. Pedersen.
1995.
Genetics of brewing yeasts, p. 223-254.
In
A. E. Wheals, A. H. Rose, and J. S. Harrison (ed.), The yeasts. Academic Press, New York, N.Y.
|
| 13.
|
Knudsen, J.,
N. J. Faergeman,
H. Skott,
R. Hummel,
C. Borsting, and T. M. Rose.
1994.
Yeast acyl-CoA-binding protein:acyl-CoA-binding affinity and effect on intracellular acyl-CoA pool size.
J. Biochem.
302:479-485.
|
| 14.
|
Kumeda, Y., and T. Asao.
1996.
Single-strand conformation polymorphism analysis of PCR-amplified ribosomal DNA internal transcribed spacers to differentiate species of Aspergillus Section Flavi.
Appl. Environ. Microbiol.
62:2947-2952[Abstract].
|
| 15.
|
Lodder, J., and N. J. W. Kreger-van Rij.
1952.
The yeasts, a taxonomic study.
North Holland Publishing Company, Amsterdam, The Netherlands.
|
| 16.
|
McCusker, J. H.,
K. V. Clemons,
D. A. Stevens, and R. W. Davis.
1994.
Saccharomyces cerevisiae virulence phenotype as determined with CD-1 mice is associated with the ability to grow at 42°C and form pseudohyphae.
Infect. Immun.
62:5447-5455[Abstract/Free Full Text].
|
| 17.
|
Messner, R., and H. Prillinger.
1995.
Saccharomyces species assignment by long range ribotyping.
Antonie Leeuwenhoek
67:363-370.
|
| 18.
|
Molnar, O.,
R. Messner,
H. Prillinger,
U. Stahl, and E. Slavikova.
1995.
Genotypic identification of Saccharomyces species using random amplified polymorphic DNA analysis.
Syst. Appl. Microbiol.
18:136-145.
|
| 19.
|
Naumov, G. I.
1987.
Genetic basis for classification and identification of the ascomycetous yeast.
Studies Mycol.
30:469-475.
|
| 20.
|
Naumov, G. I.,
E. S. Naumov, and C. Gailadrin.
1993.
Genetic and karyotypic identification of wine Saccharomyces bayanus yeast isolated in France and Italy.
Syst. Appl. Microbiol.
16:274-279.
|
| 21.
|
O'Donnell, K., and L. E. Gray.
1995.
Phylogenetic relationships of the soybean sudden death syndrome pathogen Fusarium solani f. sp. phaseoli inferred from rDNA sequence data and PCR primers for its identification.
Mol. Plant-Microbe Interact.
8:709-716[Medline].
|
| 22.
|
Philippsen, P.,
A. Stotz, and C. Scherf.
1991.
DNA of Saccharomyces cerevisiae.
Methods Enzymol.
194:169-182[Medline].
|
| 23.
|
Scherer, S., and D. A. Stevens.
1987.
Application of DNA typing methods to epidemiology and taxonomy of Candida species.
J. Clin. Microbiol.
25:675-679[Abstract/Free Full Text].
|
| 24.
|
Vaughan-Martini, A., and A. Martini.
1987.
Three newly delimited species of Saccharomyces sensu stricto.
Antonie Leeuwenhoek
53:77-84.
|
| 25.
|
Vaughan-Martini, A.,
A. Martini, and G. Cardinali.
1993.
Electrophoretic karyotyping as a taxonomic tool in the genus Saccharomyces.
Antonie Leeuwenhoek
63:145-156[Medline].
|
| 26.
|
White, T. J.,
T. Bruns,
S. Lee, and J. Taylor.
1996.
Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics, p. 315-322.
In
M. A. Innis, D. H. Gelfand, J. J. Sninsky, and T. J. White (ed.), PCR protocols: a guide to methods and applications. Academic Press, San Diego, Calif.
|
| 27.
|
Williams, D. W.,
M. J. Wilson,
M. A. Lewis, and A. J. Potts.
1995.
Identification of Candida species by PCR and restriction fragment length polymorphism analysis of intergenic spacer regions of ribosomal DNA.
J. Clin. Microbiol.
33:2476-2479[Abstract].
|
Journal of Clinical Microbiology, April 1998, p. 1035-1038, Vol. 36, No. 4
0095-1137/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.
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Top
Abstract
Introduction
