We have demonstrated that a normal laboratory strain of Candida albicans spontaneously produces mutants which acquire the ability to assimilate certain carbon sources that are not utilized by the parental strain. The examination of mutants acquiring the ability to utilize either sorbose or D-arabinose revealed a few additional phenotypic changes, including the gain and loss of the capacity to assimilate other carbon sources. The change of assimilation patterns resembled the polymorphic variation of assimilation patterns found among different wild-type strains of C. albicans. Most importantly, these sorbose-and D-arabinose-positive mutants were associated with chromosomal rearrangements, with each class of positive mutants having alterations of specific chromosomes. These findings demonstrated for the first time that chromosomal alterations in C. albicans are involved in genetic variation of fundamental functions of this asexual microorganism.We previously reported that standard laboratory strains of Candida albicans spontaneously gave rise to various types of single and multiple chromosomal rearrangements at a frequency of about 1.4% (40), and we suggested that this high frequency of chromosomal aberrations provides a means for genetic variation in this asexual microorganism (38,40). Although the chromosomal aberrations were associated with alterations of colonial and cellular morphologies, rates of growth, pseudohyphae, chlamydospore production, germ tube formation, growth at extreme temperatures, and color differences on BiGGY and phloxine B media (38-40), we were unable to discern any specific relationships between the various types of chromosomal rearrangements and phenotypic variability.Recently, Wickes et al. (48) reported that certain Candida stellatoidea strains spontaneously gave rise to sucrose-positive (Suc+) colonies that exhibited chromosomal rearrangements.In addition, we observed that all of a large number of spontaneously derived colony morphology mutants having chromosomal alterations were also associated with different patterns of carbon and nitrogen assimilation (37).Because of the obvious selective importance of carbon and nitrogen sources, these findings prompted us to investigate possible relationships between the assimilation patterns, chromosomal rearrangements, and other phenotypic traits. Furthermore, changes in assimilation provide an opportunity to investigate discrete and easily assayable phenotypes.In this paper, we describe the isolation and characterization of positive mutants, i.e., mutants that have gained the ability to assimilate substrates. In particular, independently derived spontaneous mutants that have acquired the ability to utilize either sorbose or D-arabinose were isolated from the normal strain 3153A. Of considerable importance, and in contrast to the previous report (39), these positive mutants contained altered electrophoretic karyotypes, with each class of positive mutants having alterations of specific chromosomes. This is the
Four strains of Cryptococcus neoformans var. gattii originating from Eucalyptus camaldulensis, three from Australia and one from San Francisco, were tested for their serotype, virulence for mice, and a number of genetic and molecular characteristics. All were found to be serotype B and showed significantly higher virulence for mice than did the type strains of C. neoformans var. gattii and Filobasidiella neoformans var. bacilispora, which were obtained from human cryptococcosis cases. Electrophoretic karyotypes of the strains from Australia were identical, although they were collected from sites at least 15 to 500 km apart. The electrophoretic karyotype of the strain from San Francisco was the same as that of the Australian isolates except for the mobility of one chromosome. On the contrary, no two isolates of serotype B (of a total of 11) from clinical sources were the same, regardless of their geographic origin. Furthermore, none of the clinical isolates showed a chromosomal banding pattern identical to that of Eucalyptus-originated strains. The Eucalyptusoriginated strains failed to form dikaryons when crossed with the tester strains of the two varieties of F. neoformans. Hybridization analysis with a nucleic acid probe (AccuProbe C. neoformans Culture Confirmation Test; Gen-Probe Inc., San Diego, Calif.), however, showed signals of equal intensity for clinical strains and the Eucalyptus-originated strains. Various fungi phylogenetically related to C. neoformans, including a phenol oxidase-positive strain of Cryptococcus laurentii obtained from E. camaldulensis, were negative in the nucleic acid hybridization test. These observations confirm that, in spite of karyotypic differences and the lack of dikaryon formation with the tester strains ofF. neoformans, Eucalyptus-originated C. neoformans var. gattii is the same organism as those isolated from cases of human infection. Furthermore, the C. neoformans culture confirmation test using a commercial nucleic acid probe is specific for C. neoformans. * Corresponding author. clinical strains of serotype B from various geographic areas were compared with Eucalyptus-originated strains for their karyotypes. MATERIALS AND METHODS Strains of C. neoformans var. gattii from E. camakdulensis. Three of the Eucalyptus-originated strains, B-4506, B-4507, and B-4508, were isolated by one of the investigators (D. Ellis) in Australia. B-4506 and B-4507 were isolated from the Barossa Valley, South Australia, at sites approximately 15 km apart. B-4508 was isolated from E. camaldulensis debris collected at Balranald, southwestern New South Wales. The site for B-4508 is approximately 500 km from the sites for B-4506 and B-4507. Strain B-4534 was isolated by T. Pfeifer from E. camaldulensis growing near Fort Point, San Francisco. These strains were cultured on yeast extract-peptoneglucose (YEPD) agar, as well as niger seed (birdseed) agar (26), to observe their colony characteristics. Serotyping. Serotyping of C. neoformans was performed by previously described methods (15). Mating te...
Recently developed helper virus-free methods of herpes simplex virus (HSV) amplicon vector packaging provide stocks that are virtually devoid of the cytotoxic component normally associated with traditional helper virus-based packaging methods. These approaches involve cotransfection of amplicon plasmid DNA with either a five-cosmid set or a bacterial artificial chromosome (BAC) that contains the HSV genome without its cognate pac signals. Helper virus-free amplicon packaging produces low-titer stocks (Ͻ10 5 expressing particles/ml) that exhibit a high frequency of pseudotransduction. In an effort to enhance amplicon titers, we introduced in trans a genomic copy of the virion host shutoff (vhs) protein-encoding gene UL41 into both cosmid-
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