Candida orba sp. nov., a new cactus-specific yeast species from Queensland, Australia.

A new species of yeast from decaying cladodes of Opuntia cactus, Candida orba, is described. This species is a member of a four-species clade of cactophilic yeasts. The new species has only been found in one region of Queensland, Australia, where it was presumably introduced during attempts to eradicate prickly pear cactus. DNA-DNA relatedness, phylogenetic analysis, physiological differences, killer-sensitivity profiles and mating reactions establish the distinctness of the taxon as a new species. C. orba is most closely related to Phaffomyces thermotolerans, a species found associated with columnar cacti in the North American Sonoran Desert. The type strain of C. orba, isolated from rotting cladodes of Opuntia stricta in the State of Queensland, Australia, is strain UCD-FST 84-833.1T (= CBS 8782T = NRRL Y-27336T = ATCC MYA-341). Only the h- mating type of the species has been recovered. The lack of the opposite mating type could be the result of a bottleneck during its introduction to Australia. The original geographic/host distribution of this species in the Americas is unknown.

Phylogenetic structure of the Sporopachydermia cereana species complex.

A large number of isolates previously referred to as members of the 'Sporopachydermia cereana species complex' were examined by various DNA characterization methods, leading to the conclusion that the complex is in fact made up of 10 species, one of which contains three varieties. The sequences of the internal transcribed spacer (ITS) region and the D1/D2 divergent domains of the large subunit rDNA were determined for representatives of each taxon and specific primers based on differences in the ITS were designed for rapid identification of five of the taxa. Whereas the data provide additional elements for the calibration of the ITS as a criterion for species delineation, the emerging pattern is that the ITS region does not function as well as the D1/D2 domains as an evolutionary clock. Some taxa appear to be specific for the geographical regions where they were isolated, and the distribution of many taxa is mutually exclusive.

Pichia hawaiiensis sp. nov., occurring in decaying bark of Charpentiera trees in the Hawaiian archipelago.

A description is given for Pichia hawaiiensis sp. nov., a nitrate-utilizing member of the genus Pichia E. C. Hansen emend. Kurtzman. Seven strains of the new species were isolated during the years 1972, 1973 and 1978 from rotting bark of the Hawaiian tree genera Charpentiera, Pisonia and Cheirodendron. P. hawaiiensis is heterothallic but appears to occur in nature mainly in the diploid state. Asci are deliquescent and produce up to four hat-shaped spores per ascus. Phylogenetic analysis of the 600 nucleotide D1/D2 domain of the 26S rDNA showed that P. hawaiiensis is most closely related to Pichia populi and Williopsis californica (syn. Hansenula californica). The type strain of P. hawaiiensis, isolated on the island of Hawaii from the rotting bark of Charpentiera sp. containing insect larvae, is strain UCD-FST 72-181T (= ATCC MYA-137T = CBS 8760T = NRRL Y-27270T).

Ribosomal DNA, species structure, and biogeography of the cactophilic yeast Clavispora opuntiae.

The ribosomal DNA of the cactophilic yeast species Clavispora opuntiae was studied in order to clarify the global distribution of the yeast. Over 500 strains, including isolates from several new localities worldwide, were characterized by rDNA restriction mapping. An unusual restriction pattern previously encountered only in one strain, from Conception Island in the Bahamas, was found in several Brazilian isolates. Sequences of the D1/D2 and D7/D8 divergent domains of the large subunit (LSU) and of the intergenic spacers (IGS) confirmed that these strains represent a genetically distinct variety of Clavispora opuntiae. This divergence had previously been hypothesized on the basis of reduced genetic recombination in inter-varietal crosses and the presence of a polymorphic ApaI restriction site located in the LSU. The exact position of the ApaI site in the D8 divergent domain and the nature of the variation that it reveals were determined. The complete sequences of 12 intergenic spacers clarified the significance of the species-wide variation uncovered by restriction mapping. Most of the polymorphic sites occur in the IGS1 and IGS2 regions, on either side of the 5S gene, and the variation is largely due to differences in the numbers and the sequences of internal repeats. Two other polymorphic sites are located in the external transcribed spacer (ETS) region. The reliability of various sites as indicators of overall spacer sequence divergence differed from one case to another. Variety-specific probes were devised and used to screen 120 strains for the presence of recombinant rDNA spacers. Three strains gave ambiguous results, but these did not constitute evidence that inter-varietal recombination has taken place in nature. The hypothesis that the global movement of Clavispora opuntiae has been influenced by the worldwide biological control of prickly pear with Cactoblastis cactorum, a moth of Argentinian origin, has received additional support from the demonstration that Argentinian strains have rDNAs similar to those found where the moth has been introduced. A dramatic founder effect was identified in a yeast population collected in cacti (Maui, Hawaii) in a site where the moth had been recently introduced.

Outbreak of Eremothecium coryli Fruit Rot of Tomato in California.

A ripe fruit rot of tomato (Lycopersicon esculentum) associated with stink bug (Euschistus conspersus) feeding caused extensive losses in the Sacramento Valley of California in 1998. Portions of several fields were abandoned at harvest due to fruit rot levels >30%. Symptoms included a soft, water-soaked rot often surrounding a firm, dark green island. Entire fruit sometimes became watery and soft. The ascomycetous yeast, Eremothecium coryli, characterized by spindle-shaped ascospores with long, thin, whip-like terminal appendages (1), was consistently isolated from diseased tissue plated on V8 or potato dextrose agar amended with tetracycline. Ripe tomato fruit were inoculated with a 0.1-ml suspension of 1.5 × 105 ascospores per ml injected to a depth of ≈4 mm to simulate stink bug probing. Fruit were placed in plastic containers and, in initial tests, were held at room temperature. In subsequent tests, inoculated fruit were maintained at constant temperatures of 20, 25, 30, or 35°C. Controls included inoculum either placed on the fruit surface or injected with sterilized water. Each treatment consisted of one to four fruit in two to four replications. All experiments were repeated. Lesions were visible within 5 days after inoculation and were >25 mm in diameter after 7 days. Progression of rot occurred at all temperatures but was fastest at 30 and 35°C. Controls remained symptomless. E. coryli was reisolated from infected fruit. This is the first documentation of field-level losses caused by E. coryli associated with stink bug feeding. Isolates are maintained in the Herman J. Phaff Collection of Yeast and Yeast-like Microorganisms at the University of California, Davis. Reference: (1) G. S. de Hoog et al. 1998. Eremothecium Borzi emend. Kurtzman. Pages 201-208 in: The Yeasts-A Taxonomic Study. C. P. Kurtzman and J. W. Fell, eds. Elsevier Scientific Publishers, Amsterdam.

Pichia lachancei sp. nov., associated with several Hawaiian plant species.

A description is given of Pichia lachancei sp. nov., a new species of yeast that occurs in association with several Hawaiian plant species of the genera Tetraplasandra, Cheirodendron and Clermontia. The new species is heterothallic and occurs in nature in the haploid as well as the diploid state. Upon conjugation of complementary mating types, zygotes are formed that reproduce by budding as diploid cells. When placed on sporulation medium, four hat-shaped spores are produced which are rapidly released from the ascus. Phylogenetic analysis showed that P. lachancei is most closely related to Pichia rhodanensis and Pichia jadinii. The diploid type strain of P. lachancei, isolated from rotting bark of Tetraplasandra hawaiiensis on the island of Hawaii, is strain UCD-FST 79-9T (= ATCC 201914T = CBS 8557T = NRRL Y-27008T).

Dipodascus starmeri sp. nov., a new species of yeast occurring in cactus necroses.

In a previous publication describing the geographic distribution of yeasts associated with cactus necroses (W. T. Starmer, M.-A. Lachance, H. J. Phaff, and W. B. Heed, Evol. Biol. 24:253-296, 1990), 127 isolates were identified as strains of Candida ingens van der Walt et van Kerken on the basis of morphology and certain phenotypic characteristics. Here we show by using DNA hybridization and additional phenotypic properties that these strains were misidentified and that they represent a minimum of three separate species that can be differentiated from C. ingens and from each other by utilization of 2-propanol or acetone, sensitivity to digitonin, utilization of L-lysine as a sole nitrogen source, vitamin dependence, NaCl tolerance, lipolytic activity, and habitat. One of the new species is haploid and heterothallic, and its teleomorph represents the genus Dipodascus. We describe Dipodascus starmeri sp. nov. The phylogenetic relationship of D. starmeri with other members of the genus Dipodascus and its anamorph, the genus Geotrichum, was estimated from ribosomal DNA nucleotide sequence divergence. The type strain, a heterothallic haploid isolate, is UCD-FST 72-316 (= CBS 780.96 = ATCC 200546 = NRRL Y-17816). The complementary mating type is UCD-FST 81-513.3 (= CBS 781.96 = ATCC 200547 = NRRL Y-17817).

Ultrastructure of the cell wall of Schizosaccharomyces pombe following treatment with various glucanases.

The ultrastructure of isolated cell walls of Schizosaccharomyces pombe was studied by electron microscopy after treatment with the following purified enzymes: endo-beta-(1-->3)-glucanase, endo-beta-(-->6)-glucanase, and endo-alpha-(1-->3)-glucanase produced by Bacillus circulans; exo-beta-(1-->3)-glucanase and endo-beta-(1-->3)-glucanase produced by Schizosaccharomyces japonicus var. versatilis. The exo-beta-(1-->3)-glucanase had no detectable effect on the walls, but amorphous wall material was removed by action of the endo-beta-(1-->3)- and endo-beta-(1-->6)-glucanases of B. circulans to reveal a wall component consisting of densely interwoven microfibrils. The fibrils were hydrolyzed by treatment with the Schiz. japonicus endo-beta-(1-->3)-glucanase followed by B. circulans endo-alpha-(1-->3)-glucanase--suggesting that they were composed of -beta-(1-->3)-linked glucan and alpha-(1-->3)-linked glucan. The presence of a fibrillar component in untreated walls was evident after negative staining.

Candida caseinolytica sp. nov., a new species of yeast occurring in necrotic tissue of Opuntia and Stenocereus species in the southwestern United States and Baja California, Mexico.

We describe Candida caseinolytica, a new yeast species which occurs in rotting tissues of opuntias and other cacti in the North American Sonoran Desert and a few other localities. This small-celled, slowly growing yeast does not ferment any sugar and assimilates a limited number of carbon compounds, including 2- and 5-ketogluconic acids. It exhibits strong extracellular proteolytic activity on casein at pH 6.5, but gelatin is not hydrolyzed or is only weakly hydrolyzed by a few strains. The type strain of C. caseinolytica is strain UCD-FST 83-438.3 (= ATCC 90546 = CBS 7781).

Metschnikowia hawaiiensis sp. nov., a heterothallic haploid yeast from Hawaiian morning glory and associated drosophilids.

A new haploid, heterothallic yeast species was isolated repeatedly from morning glory (Ipomoea acuminata) flowers and from two associated drosophilid species, Scaptomyza calliginosa and Drosophila floricola, in a Hawaiian kipuka. Haploid strains of this organism multiply asexually by budding and, under nutrient deprivation, by the formation of long germ tubes that develop into branching true mycelia. Mating compatibility is controlled by two alleles of a single locus. Plasmogamy between compatible strains is followed by the development of very large elongate asci bearing vestiges of the zygotes and the formation in each ascus of two unusually large aciculate ascospores similar to those formed by members of the genus Metschnikowia. Membership in the genus Metschnikowia is supported by the physiological profile of the yeast, which is typical of the genus but not identical to the profile of any previously described species. The name Metschnikowia hawaiiensis is proposed to emphasize the geographic origin of the new species, not its habitat, which has not been determined precisely. The holotype strain of M. hawaiiensis is strain UWO(PS) 87-2167.2 (= ATCC 76059 = CBS 7432), and the isotype strain is strain UWO(PS) 87-2203.2 (= ATCC 76058 = CBS 7433).

The ecological role of killer yeasts in natural communities of yeasts.

The killer phenomenon of yeasts was investigated in naturally occurring yeast communities. Yeast species from communities associated with the decaying stems and fruits of cactus and the slime fluxes of trees were studied for production of killer toxins and sensitivity to killer toxins produced by other yeasts. Yeasts found in decaying fruits showed the highest incidence of killing activity (30/112), while yeasts isolated from cactus necroses and tree fluxes showed lower activity (70/699 and 11/140, respectively). Cross-reaction studies indicated that few killer-sensitive interactions occur within the same habitat at a particular time and locality, but that killer-sensitive reactions occur more frequently among yeasts from different localities and habitats. The conditions that should be optimal for killer activity were found in fruits and young rots of Opuntia cladodes where the pH is low. The fruit habitat appears to favor the establishment of killer species. Killer toxin may affect the natural distribution of the killer yeast Pichia kluyveri and the sensitive yeast Cryptococcus cereanus. Their distributions indicate that the toxin produced by P. kluyveri limits the occurrence of Cr. cereanus in fruit and Opuntia pads. In general most communities have only one killer species. Sensitive strains are more widespread than killer strains and few species appear to be immune to all toxins. Genetic study of the killer yeast P. kluyveri indicates that the mode of inheritance of killer toxin production is nuclear and not cytoplasmic as is found in Saccharomyces cerevisiae and Kluyveromyces lactis.

A comparison of yeast communities found in necrotic tissue of cladodes and fruits ofOpuntia stricta on Islands in the Caribbean Sea and where introduced into Australia.

Yeast communities growing in the decaying tissues (cladodes and fruits) ofOpuntia stricta (prickly pear cactus) and associated yeast vectors (Drosophila species) were compared in two geographic regions (Caribbean and eastern Australia). The Australian yeast community provides an interesting comparison to the Caribbean community, because the host plantO. stricta was introduced to Australia over 100 years ago. Many of the yeasts found in the Australian system also were introduced during a period of biological control (1926-1935) when they accompanied rotting prickly pear cladodes and insects shipped to Australia from the Americas. The yeast community composition (proportion of each species) is compared at several levels of organization: (1) within and between regions, (2) across seasons and years, and (3) within and between tissue types. The yeast species composition of the cladode communities are similar from locality to locality, season to season, and year to year, with the region-to-region similarity slightly less. The composition of the fruit-yeast communities are distinct from region to region and only show some overlap with the cladodes within regions when collected simultaneously in the same locality. It is suggested that the cladode-microorganism-Drosophila system is relatively closed (little extrinsic influence) whereas the fruit-microorganism-Drosophila system is open (large extrinsic influence).

Adaptations of Drosophila and yeasts: their interactions with the volatile 2-propanol in the cactus-microorganism-Drosophila model system.

The interactions of yeasts growing in decaying cactus tissue with and without 2-propanol were studied with respect to the costs and benefits provided to three cactophilic Drosophila species (D. mojavensis, D. arizonensis and D. buzzatii). Two common cactus yeasts, Candida sonorensis and Cryptococcus cereanus, which can tolerate and metabolize 2-propanol, provide benefits to the three Drosophila species in the presence of the alcohol, as compared with another common cactus yeast, Pichia cactophila, which has less tolerance and cannot metabolize 2-propanol. Because 2-propanol is commonly found in decaying cactus tissue and C. sonorensis and Cr. cereanus are also frequently recovered from the rotting tissue being utilized by the Drosophila species, the interactions described here are viewed as a possible adaptation in which the yeast provides benefits to one of its vectors by metabolism of 2-propanol in the habitat.

DNA relatedness among aquatic yeasts of the genus Metschnikowia and proposal of the species Metschnikowia australis comb. nov.

DNA hybridization studies were conducted to determine the taxonomic status of the aquatic group of Metschnikowia species and their varieties. Among the DNAs of the four varieties of Metschnikowia bicuspidata, that of Metschnikowia bicuspidata var. australis showed 37 to 51% relative binding with the DNAs of Metschnikowia bicuspidata var. bicuspidata and of the varieties chathamia and californica. On this basis, low intervarietal fertility, and unique habitat in antarctic seawater, we have proposed to raise M. bicuspidata var. australis to the rank of species, Metschnikowia australis, comb. nov. DNA complementarity of Metschnikowia bicuspidata var. californica and Metschnikowia bicuspidata var. chathamia DNA was greater than 80% with that of M. bicuspidata var. bicuspidata. Metschnikowia australis can be differentiated from other Metschnikowia species and varieties by its inability to form chlamydospores, the formation of two needle-shaped ascospores per ascus, lack of glucose fermentation, and lack of assimilation of both methyl-alpha-D-glucoside and glucono-delta-lactone. Other DNA-DNA reassociation experiments showed that Metschnikowia zobellii is a distinct species when compared with both aquatic and terrestrial species of the genus.

DNA base composition and DNA relatedness among species of Trichosporon Behrend.

The nuclear DNA of 28 species (30 strains investigated) of yeasts classified currently or previously in the genus Trichosporon. was analysed for its molar percentage of guanine + cytosine (mol% G + C). This criterion, together with biochemical characteristics, suggested the separation of the organisms studied into two groups. The first group, which appears related to the Ascomycetes, includes thirteen species with a G + C content lower than 50 mol% (34.7-48.8), and lacks urease (except T. margaritiferum). The second group appears related to the Basidiomycetes and includes fifteen species with a G + C content higher than 50 mol% (57-64) and has the ability to hydrolyse urea. A DNA homology experiment with T. beigelii and twelve other species of the second group showed very low values of complementarity with T. beigelii-labeled DNA. All these species must be considered as taxa other than T. beigelii.

The ecology of the yeast flora in necroticOpuntia cacti and of associatedDrosophila in Australia.

A survey was made of the yeast communities isolated from necrotic tissue of 4 species of prickly-pear cacti (Opuntia stricta, O. tomentosa, O. monacantha, andO. streptacantha) which have colonized in Australia. Yeast communities were sampled from a number of localities and at different times. Cactus specific yeasts accounted for 80% of the total isolates, and the 3 most common species contributed 63% of the total. Comparisons of the species compositions of the yeast communities indicated that the differences among communities were greater betweenOpuntia species than between different localities within a single cactus species, and also that differences between years were greater than average differences between localities within years. Multivariate statistical tests of association between yeast community and physical features of rots indicated that temperature, pH, and age of rot all exerted some influence on the structure of the yeast community. Similar analyses involvingDrosophila species inhabiting these cactus rots suggested the existence of complex associations betweenDrosophila community, yeast community, and physical and chemical attributes of the cactus necroses.