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1.
Gómez-Raja J Andaluz E Magee B Calderone R Larriba G 《Fungal genetics and biology : FG & B》2008,45(4):527-541
Candida albicans is a diploid organism that exhibits high levels of heterozygosity. Although the precise manner by which this heterozygosity provides advantage for the commensal/pathogenic life styles of C. albicans is not known, heterozygous markers are themselves useful for studying genomic rearrangements, which occur frequently in C. albicans. Treatment of CAI-4 with UV light yielded histidine auxotrophs which could be complemented by HIS4, suggesting that strain CAI-4 is heterozygous for HIS4. These auxotrophs appeared to have undergone mitotic recombination and/or chromosome loss. As expected from a heterozygote, disruption of the functional allele of HIS4 resulted in a his4::hisG-URA3-hisG strain that is auxotrophic for histidine. Sequencing of random clones of the HIS4 ORF from CAI-4 and its precursor SC5314 revealed the presence of 11 SNPs, seven synonymous and four non-synonymous. Site-directed mutagenesis indicates that only one of those SNPs, T929G (Gly310Val), is responsible for the non-functionality of the encoded enzyme. HIS4 analysis of five commonly used laboratory strains is reported. This study provides a new, easily measured nutritional marker that can be used in future genetic studies in C. albicans. 相似文献
2.
Andaluz E Bellido A Gómez-Raja J Selmecki A Bouchonville K Calderone R Berman J Larriba G 《Molecular microbiology》2011,79(6):1462-1482
RAD52 is required for almost all recombination events in Saccharomyces cerevisiae. We took advantage of the heterozygosity of HIS4 in the Candida albicans SC5314 lineage to study the role of Rad52 in the genomic stability of this important fungal pathogen. The rate of loss of heterozygosity (LOH) at HIS4 in rad52-ΔΔ strains was ~10(-3) , at least 100-fold higher than in Rad52(+) strains. LOH of whole chromosome 4 or truncation of the homologue that carries the functional HIS4 allele was detected in all 80 rad52-ΔΔ His auxotrophs (GLH -GL lab His(-)) obtained from six independent experiments. Isolates that had undergone whole chromosome LOH, presumably due to loss of chromosome, carried two copies of the remaining homologue. Isolates with truncations carried centric fragments of broken chromosomes healed by de novo telomere addition. GLH strains exhibited variable degrees of LOH across the genome, including two strains that became homozygous for all the heterozygous markers tested. In addition, GLH strains exhibited increased chromosomal instability (CIN), which was abolished by reintroduction of RAD52. CIN of GLH isolates is reminiscent of genomic alterations leading to cancer in human cells, and support the mutator hypothesis in which a mutator mutation or CIN phenotype facilitate more mutations/aneuploidies. 相似文献
3.
Bougnoux ME Pujol C Diogo D Bouchier C Soll DR d'Enfert C 《Fungal genetics and biology : FG & B》2008,45(3):221-231
Candida albicans is a diploid yeast that can undergo mating and a parasexual cycle, but is apparently unable to undergo meiosis. Characterization of the population structure of C. albicans has shown that reproduction is largely clonal and that mating, if it occurs, is rare or limited to genetically related isolates. Because molecular typing has delineated distinct clades in C. albicans, we have tested whether recombination was common within clades, but rare between clades. Two hundred and three C. albicans isolates have been subjected to multilocus sequence typing (MLST) and the haplotypes at heterozygous MLST genotypes characterized. The C. albicans isolates were distributed among nine clades, of which five corresponded to those previously identified by Ca3 fingerprinting. In each of these clades with more than 10 isolates, polymorphic nucleotide positions located on between 3 and 4 of the six loci were in Hardy-Weinberg disequilibrium. Moreover, each of these polymorphic sites contained excess heterozygotes. This was confirmed by an expanded analysis performed on a recently published MLST dataset for 1044 isolates. On average, 66% of polymorphic positions in the individual clades were in significant excess of heterozygotes over the five clades. These data indicate that mating within clades as well as self-fertilization are both limited and that C. albicans clades do not represent a collection of cryptic species. The study of haplotypes at heterozygous loci performed on our dataset indicates that loss of heterozygosity events due to mitotic recombination is moderately common in natural populations of C. albicans. The maintenance of substantial heterozygosity despite relatively frequent loss of heterozygosity could result from a selective advantage conferred by heterozygosity. 相似文献
4.
Marie-Elisabeth Bougnoux Claude Pujol Dorothée Diogo Christiane Bouchier David R Soll Christophe d'Enfert 《Fungal genetics and biology : FG & B》2008,45(3):221-231
Candida albicans is a diploid yeast that can undergo mating and a parasexual cycle, but is apparently unable to undergo meiosis. Characterization of the population structure of C. albicans has shown that reproduction is largely clonal and that mating, if it occurs, is rare or limited to genetically related isolates. Because molecular typing has delineated distinct clades in C. albicans, we have tested whether recombination was common within clades, but rare between clades. Two hundred and three C. albicans isolates have been subjected to multilocus sequence typing (MLST) and the haplotypes at heterozygous MLST genotypes characterized. The C. albicans isolates were distributed among nine clades, of which five corresponded to those previously identified by Ca3 fingerprinting. In each of these clades with more than 10 isolates, polymorphic nucleotide positions located on between 3 and 4 of the six loci were in Hardy-Weinberg disequilibrium. Moreover, each of these polymorphic sites contained excess heterozygotes. This was confirmed by an expanded analysis performed on a recently published MLST dataset for 1044 isolates. On average, 66% of polymorphic positions in the individual clades were in significant excess of heterozygotes over the five clades. These data indicate that mating within clades as well as self-fertilization are both limited and that C. albicans clades do not represent a collection of cryptic species. The study of haplotypes at heterozygous loci performed on our dataset indicates that loss of heterozygosity events due to mitotic recombination is moderately common in natural populations of C. albicans. The maintenance of substantial heterozygosity despite relatively frequent loss of heterozygosity could result from a selective advantage conferred by heterozygosity. 相似文献
5.
A Sarachek 《Zeitschrift für allgemeine Mikrobiologie》1983,23(6):385-391
Nalidixic acid induces segregation of auxotrophs from prototrophic hybrids of Candida albicans artifically produced by fusing complementing auxotrophic protoplasts. The auxotrophies recovered are limited to those introduced through the fusion process, and patterns of segregations for linked auxotrophic markers demonstrate the segregants are products mitotic crossing-over. Nalidixic acid does not induce auxotrophies of any sort in clinical isolates of C. albicans. These findings are contrary to a current hypothesis that natural strains of C. albicans are diploid and heterozygous for a variety of auxotrophic mutations. 相似文献
6.
Molecular typing of Candida albicans is important for studying the population structure and epidemiology of this opportunistic yeast, such as population dynamics, nosocomial infections, multiple infections and microevolution. The genetic diversity of C. albicans has been rarely studied in China. In the present study, multilocus sequence typing (MLST) was used to characterize the genetic diversity and population structure of 62 C. albicans isolates collected from 40 patients from Huashan Hospital in Shanghai, China. A total of 50 diploid sequence types (DSTs) were identified in the 62 C. albicans isolates, with 41 newly identified DSTs. Based on cluster analysis, the 62 isolates were classified into nine existing clades and two new clades (namely clades New 1 and New 2). The majority of the isolates were clustered into three clades, clade 6 (37.5%), clade 1 (15.0%) and clade 17 (15.0%). Isolates of clade New 2 were specifically identified in East Asia. We identified three cases of potential nosocomial transmission based on association analysis between patients’ clinical data and the genotypes of corresponding isolates. Finally, by analyzing the genotypes of serial isolates we further demonstrated that the microevolution of C. albicans was due to loss of heterozygosity. Our study represents the first molecular typing of C. albicans in eastern China, and we confirmed that MLST is a useful tool for studying the epidemiology and evolution of C. albicans. 相似文献
7.
Interspecific complementation analysis by protoplast fusion of Candida tropicalis and Candida albicans adenine auxotrophs. 总被引:1,自引:0,他引:1
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A protocol employing inositol starvation was used to isolate proline and adenine auxotrophs of Candida tropicalis. Interspecific hybrids between red adenine auxotrophs of C. tropicalis and Candida albicans were formed by protoplast fusion. These C. tropicalis red adenine auxotrophs were shown to fall into two complementation groups by crossing them with a known C. albicans ade1 tester strain. It is suggested that these two groups correspond to the ade1 and ade2 mutants of Saccharomyces cerevisiae and C. albicans and that these defined mutants may be useful in attempts to develop transformation systems for C. tropicalis. 相似文献
8.
A single pair of primers, deduced from the intron nucleotide sequence of the Candida albicans CaYST1 gene, was used in PCR analysis performed with both genomic DNA and whole cells of clinical isolates of Candida species and other microorganisms. All the clinical C. albicans isolates generated the expected 310 bp amplicon; other Candida species as well as laboratory strains belonging to other fungal genera failed to amplify any DNA fragment, except for Candida pseudotropicalis (amplicon of 1200 bp), Kluyveromices marxianus (amplicon of 1250 bp) and Cryptococcus neoformans (several amplicons longer than 1200 bp). Unusual C. albicans isolates from Africa also yielded the expected 310 bp amplicon. These results indicate that genes containing intron sequences may be useful to design species-specific primers for identification of fungal strains by PCR. The sensitivity of the method was evaluated for C. albicans genomic DNA by using both various DNA concentrations (224 ng to 2.7 pg) and different cell amounts (10(7); to 5 cells). The results obtained may be useful in earlier detection of candidiasis. 相似文献
9.
《Experimental mycology》1983,7(4):328-335
Ultraviolet light readily induced both auxotrophic and morphological mutants in isolates ofF. moniliforme from corn and sorghum. Sexual analysis of these mutants yielded a genetic map with several instances of linkage. Forced heterokaryons formed readily between auxotrophs of the same strain but not between auxotrophs of different strains. At least four nuclear gene loci, but not the mating-type locus, controlled heterokaryon formation in this fungus. In the auxotroph pairings, heterokaryosis was confined to the anastomosed cells; there was no evidence for nuclear migration through established hyphae. Genetic complementation occurred in anastomosed cells, and the products of this complementation were then translocated to the homokaryotic cells of the colony. No heterozygous diploids were recovered from the heterokaryons, although there was evidence for rare somatic gene recombination. Heterokaryosis in isolates ofF. moniliforme in nature is probably rare. 相似文献
10.
Shin JH Og YG Cho D Kee SJ Shin MG Suh SP Ryang DW 《Journal of microbiology (Seoul, Korea)》2005,43(6):546-554
We assessed the genetic relations and epidemiological links among bloodstream isolates of Candida albicans, which were obtained from a university hospital over a period of five years. The 54 bloodstream isolates from the 38 patients yielded 14 different karyotypes, 29 different patterns after digestion with SfiI (REAG-S), and 31 different patterns after digestion with BssHII (REAG-B) when analyzed using three different pulsed-field gel electrophoresis (PFGE) typing methods. In 11 patients with serial bloodstream isolates, all strains from each patient had the same PFGE pattern. The dendrograms for all of the strains revealed that the distribution of similarity values ranged from 0.70 to 1.0 in the REAG-S patterns, and from 0.35 to 1.0 in the REAG-B patterns. Overall, the combination of the three different PFGE methods identified 31 distinct types, reflecting the results obtained using the REAG-B alone different. different Five PFGE types were shared among 22 isolates from 12 patients. These types of strains were more frequently associated with central venous catheter-related fungemia than the other 26 type strains (92% versus 31%; P < 0.005). Of five PFGE types, four isolates were determined to be epidemiologically related: each of these types was primarily from two or three patients who had been hospitalized concurrently within the same intensive care unit. Our results suggest that the REAG-B constitutes perhaps the most useful PFGE method for investigating C. albicans candidemia and also shows that a relatively high proportion of C. albicans candidemia may be associated with exogenous acquisition of clonal strains. 相似文献
11.
This study evaluated the phenotypic tests used to differentiate Candida albicans from Candida dubliniensis. A total of 55 isolates from vaginal secretions, oral cavity and hemoculture were studied. They were originally identified as C. albicans, based on their morphological and physiological characteristics. These isolates were tested for colony color development on CHROMagar Candida medium, growth at 45 degrees C on Sabouraud Dextrose agar, lipolytic activity on Tween 80 Agar medium and colony morphology and chlamydoconidia formation on Staib agar medium. Of the 55 isolates studied, seven yielded one or more phenotypic characteristics suggestive of Candida dubliniensis. These isolates were tested by PCR with specific primers for Candida dubliniensis and API ID 32. The seven isolates were confirmed as Candida albicans. All of these finding indicate that DNA based tests should be used for definitive identification of Candida dubliniensis. 相似文献
12.
Mariano Pde L Milan EP da Matta DA Colombo AL 《Memórias do Instituto Oswaldo Cruz》2003,98(4):533-538
We investigated the presence of Candida dubliniensis among isolates previously identified as Candida albicans and maintained in a yeast stock collection from 1994 to 2000. All isolates were serotyped and further evaluated for antifungal susceptibility profile. After doing a screening test for C. dubliniensis isolates based on the capability of colonies to grow at 42 C, its final identification was obtained by randomly amplified polymorphic DNA (RAPD) analysis using three different primers. A total of 46 out of 548 screened isolates did not exhibit growth at 42 C and were further genotyped by RAPD. Eleven isolates were identified as C. dubliniensis with RAPD analysis. Regarding serotypes, 81.5% of C. albicans and all C. dubliniensis isolates belonged to serotype A. Of note, 9 out of 11 C. dubliniensis isolates were obtained from patients with acquired immunodeficiency syndrome (Aids) and all of them were susceptible to azoles and amphotericin B. We found 17 (3%) C. albicans isolates that were dose-dependent susceptibility or resistant to azoles. In conclusion, we found a low rate of C. dubliniensis isolates among stock cultures of yeasts previously identified as C. albicans. Most of these isolates were recovered from oral samples of Aids patients and exhibited high susceptibility to amphotericin B and azoles. C. albicans serotype A susceptible to all antifungal drugs is the major phenotype found in our stock culture. 相似文献
13.
Auxotrophic mutants of Candida albicans FC18 were induced by a combination of treatments with nitrous acid and UV irradiation. Arginine (Arg-), histidine (His-) and methionine/cysteine (MetA-) auxotrophs were recovered by this means. The Arg- auxotrophs lacked active argininosuccinate lyase (EC 4.3.2.1), the enzyme catalysing the final step in arginine biosynthesis. Thus the locus may be designated arg-4. The mutant strains bearing this mutation did not form germ tubes unless the germination medium contained arginine. 相似文献
14.
Comparative pathogenicity of auxotrophic mutants of Candida albicans 总被引:11,自引:0,他引:11
An induced mutant of Candida albicans with greatly decreased virulence for mice is described. The mutant was one of five auxotrophic mutants obtained by ultraviolet irradiation of a clinical isolate (strain MY 1044). The five mutants included two methionine auxotrophs, one methionine-cysteine auxotroph, one temperature-sensitive serine auxotroph, and one auxotroph with unknown growth requirements. Each of the mutants produced normal mycelium and had a normal profile of susceptibility to four antifungal drugs. The virulence of each mutant was compared with the parent strain by LD50 determination in mice. Four of the five auxotrophs exhibited LD50's that were not significantly different from the parent strain (mean LD50 = 7.5 x 10(5) cells). However, the temperature-sensitive serine auxotroph was significantly less virulent than the parent strain (LD50 greater than 10(7) cells), even though it grew well in vivo and in mouse serum at 37 degrees C in vitro. Use of this mutant in conjunction with its "isogenic" parent should help to elucidate true virulence factors in C. albicans. 相似文献
15.
16.
Micrococcus glutamicus ATCC 13032, a glutamic acid-producing organism, was treated with 0.2M ethylmethane sulfonate, the auxotrophs isolated showing varied patterns of extracellular amino acids. Eighty auxotrophic strains were obtained, out of which 31 excreted 1.0-4.0 mg threonine per ml and all the auxotrophs required biotin for growth and production of the amino acid. Eleven auxotrophs produced 1.5 to 3.0 mg alanine per ml and these auxotrophs required amino acids for their growth. Other auxotrophs lost their excretion capacity in subsequent fermentation trials. Further mutation of the biotin-requiring auxotroph Micrococcus glutamicus EM with gamma rays resulted in the isolation of 89 auxotrophic strains, out of which 28 excreted threonine (up to 5.0 mg per ml) higher than the parent auxotroph. Exposure to X-rays yielded 97 auxotrophs, out of these 35 producing 1.0-3.0 mg methionine per ml and requiring biotin for growth and production of the amino acid. Other auxotrophs produced alanine (0.5 to 2.0 mg per ml) and threonine (2.0 to 3.3 mg per ml). Irradiation with gamma rays favoured the development of threonine producing auxotrophs while X-rays favoured methionine-producing auxotrophs. 相似文献
17.
Bacteriophage P1 as a vehicle for Mu mutagenesis of Salmonella typhimurium. 总被引:10,自引:8,他引:2
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We developed a procedure using bacteriophage P1 as a vector for transferring Mu phage deoxyribonucleic acid into Salmonella typhimurium. Mu phage transferred in this manner yielded lysogenic auxotrophs, and we demonstrated that specific deletions and lac gene fusions can be selected. 相似文献
18.
Value of morphotyping for the characterization of Candida albicans clinical isolates 总被引:1,自引:0,他引:1
Giammanco GM Lopes MM Coimbra RS Pignato S Grimont PA Grimont F Freitas G Giammanco G 《Memórias do Instituto Oswaldo Cruz》2005,100(5):483-490
Until recently, morphotyping, a method evaluating fringe and surface characteristics of streak colonies grown on malt agar, has been recommended as a simple and unexpensive typing method for Candida albicans isolates. The discriminatory power and reproducibility of Hunter's modified scheme of Phongpaichit's morphotyping has been evaluated on 28 C. albicans isolates recovered from the oral cavity of asymptomatic human immunodeficiency virus-positive subjects, and compared to two molecular typing methods: randomly amplified polymorphic DNA (RAPD) fingerprinting, and contour clamped homogeneous electric field (CHEF) electrophoretic karyotyping. Morphological features of streak colonies allowed to distinguish 11 different morphotypes while RAPD fingerprinting yielded 25 different patterns and CHEF electrophoresis recognized 9 karyotypes. The discriminatory power calculated with the formula of Hunter and Gaston was 0.780 for morphotyping, 0.984 for RAPD fingerprinting, and 0.630 for karyotyping. Reproducibility was tested using 43 serial isolates from 15 subjects (2 to 6 isolates per subject) and by repeating the test after one year storage of the isolates. While genetic methods generally recognized a single type for all serial isolates from each of the subjects studied, morphotyping detected strain variations in five subjects in the absence of genetic confirmation. Poor reproducibility was demonstrated repeating morphotyping after one year storage of the isolates since differences in at least one character were detected in 92.9% of the strains. 相似文献
19.
Chronic mucocutaneous candidiasis (CMC) is a primary immunodeficiency disorder characterised by susceptibility to chronic Candida and fungal dermatophyte infections of the skin, nails and mucous membranes. Molecular epidemiology studies of CMC infection are limited in number and scope and it is not clear whether single or multiple strains inducing CMC persist stably or are exchanged and replaced. We subjected 42 C. albicans individual single colony isolates from 6 unrelated CMC patients to multilocus sequence typing (MLST). Multiple colonies were typed from swabs taken from multiple body sites across multiple time points over a 17-month period. Among isolates from each individual patient, our data show clonal and persistent diploid sequence types (DSTs) that were stable over time, identical between multiple infection sites and exhibit azole resistant phenotypes. No shared origin or common source of infection was identified among isolates from these patients. Additionally, we performed C. albicans MLST SNP genotype frequency analysis to identify signatures of past loss of heterozygosity (LOH) events among persistent and azole resistant isolates retrieved from patients with autoimmune disorders including CMC. 相似文献
20.
Auxotrophy of Pseudomonas aeruginosa in cystic fibrosis 总被引:4,自引:0,他引:4
Seventy-four of 403 (18.4%) sputum isolates of Pseudomonas aeruginosa from 49 of 136 (36.0%) adults with cystic fibrosis (CF) were auxotrophic mutants. Two of 11 (18.2%) isolates of P. aeruginosa taken from patients with non-CF bronchiectasis were also auxotrophic. All 99 strains taken from non-bronchiectatic sources were prototrophic. Forty-six of 55 (83.6%) CF auxotrophs required one or more of 36 growth factors tested; the requirements for the remaining 9 isolates were not identified. Methionine was the sole factor required by 17 of 22 (77.3%) isolated which depended on a single factor. We conclude that auxotrophy is a feature of P. aeruginosa infection in cystic fibrosis. 相似文献