Saccharomyces sensu stricto as a model system for evolution and ecology

Baker's yeast, Saccharomyces cerevisiae, is not only an extensively used model system in genetics and molecular biology, it is an upcoming model for research in ecology and evolution. The available body of knowledge and molecular techniques make yeast ideal for work in areas such as evolutionary and ecological genomics, population genetics, microbial biogeography, community ecology and speciation. As long as ecological information remains scarce for this species, the vast amount of data that is being generated using S. cerevisiae as a model system will remain difficult to interpret in an evolutionary context. Here we review the current knowledge of the evolution and ecology of S. cerevisiae and closely related species in the Saccharomyces sensu stricto group, and suggest future research directions.     

Mitochondria--tool for taxonomic identification of yeasts from Saccharomyces sensu stricto complex

Mitochondrial genomes of Saccharomyces and close relatives previously used for transplacement of mitochondria to S. cerevisiae were examined. The origins of replication in mitochondrial DNA, the presence of nuclear and mitochondrial polymorphic loci and the ability to produce mitochondrial respiration-deficient mutants were used to reclassify some collection yeasts and to assign others into four separate subgroups. The first included isolates identical to Saccharomyces cerevisiae (S. italicus, S. oviformis, S. chevalieri and S. capensis) which possess 5 or more replication origins. The second group consists of S paradoxus (var douglasii) mitochondrial genome with the equal number of ori sequences but incompatible mitochondria. The third group represents Saccharomyces sensu stricto petite-positive species (S. carlsbergensis, S. heterogenicus, S. uvarum, S. willianus) with 1-2 origins of replication significantly different from S. cerevisiae. In addition, the locus between tRNA(fMet) and tRNA(Pro) is about one-half of the 1400 bp members of S. cerevisiae complex. The last group includes isolates that do not belong to Saccharomyces sensu stricto group as they are petite-negative and devoid of any S. cerevisiae-like replication origins.     

Karyomorphology of the genusLycopodium sensu stricto and relationships among species

Karyomorphological comparisons were made of six species of JapaneseLycopodium sensu stricto. There were no marked differences at interphase and prophase among the six species.Lycopodium annotinum had 2n=68 and the formula of its metaphase karyotype was 18m(median centromeric chromosomes)+12sm(submedian)+12st(subterminal)+26t(terminal).Lycopodium casuarinoides had 2n=68=16m+10sm+18st+24t,L. clavatum 2n=68=22m+12sm+18st+16t, andL. obscurum 2n=68=10m+22sm+20st+16t. Each of these species, which belong to different sections, displayed several karyomorphological differences. Among themL. casuarinoides differs largely from the others in its mean chromosome length, ratio of the longest chromosome to the shortest, and frequency of m+sm chromosomes. BothLycopodium complanatum andL. nikoense, belonging to sectionComplanata, had a common karyotype 2n=46=10m+12sm+18st+6t. This section displayed a low differentiation in its karyotype. In the wholeLycopodium s.s., the ratios of m+sm in a complement varied from 38 to 50%, being higher among pteridophytes.     

Genetic identification of naturalSaccharomyces sensu stricto yeasts from Finland,Holland and Slovakia

Genetic and karyotypic studies of naturalSaccharomyces sensu stricto yeasts from Finland, Holland and Slovakia revealed three wild sibling-species:Saccharomyces cerevisiae, Saccaromyces bayanus andSaccharomyces paradoxus.     

Identification of Saccharomyces sensu stricto and Torulaspora yeasts by PCR ribotyping

18S rDNA + ITS1 and 25S rDNA PCR products covering more than 95% of the nuclear ribosomal DNA repeat unit of 28 Saccharomyces sensu stricto and Torulaspora yeasts and their anamorph forms were digested with Hae III, Msp I, Hinf I and Cfo I. Using combinations of two restriction enzymes, specific ribotyping patterns of six species were found. PCR ribotyping offers a convenient tool for quick identification of yeast isolates, but specificity of ribotyping patterns should be checked with a larger number of strains to avoid misidentification because of lack of variation within different taxa or because of strain-specific ribotyping patterns of species type strains.     

Pangenome inventory of Burkholderia sensu lato,Burkholderia sensu stricto,and the Burkholderia cepacia complex reveals the uniqueness of Burkholderia catarinensis

Here the pangenome analysis of Burkholderia sensu lato (s.l.) was performed for the first time, together with an updated analysis of the pangenome of Burkholderia sensu stricto, and Burkholderia cepacia complex (Bcc) focusing on the Bcc B. catarinensis specific features of its re-sequenced genome. The pangenome of Burkholderia s.l., Burkholderia s.s., and of the Bcc was open, composed of more than 96% of accessory genes, and more than 62% of unknown genes. Functional annotations showed that secondary metabolism genes belonged to the variable portion of genomes, which might explain their production of several compounds with varied bioactivities. Taken together, this work showed the great variability and uniqueness of these genomes and revealed an underexplored unknown potential in poorly characterized genes. Regarding B. catarinensis 89^T, its genome harbors genes related to hydrolases production and plant growth promotion. This draft genome will be valuable for further investigation of its biotechnological potentials.     

Molecular and metabolic characterization of cold-tolerant alpine soil Pseudomonas sensu stricto

Alpine soils undergo dramatic temporal changes in their microclimatic properties, suggesting that the bacteria there encounter uncommon shifting selection gradients. Pseudomonads constitute important members of the alpine soil community. In order to characterize the alpine Pseudomonas community and to assess the impact of shifting selection on this community, we examined the ability of cold-tolerant Pseudomonas isolates to grow on a variety of carbon sources, and we determined their phylogenetic relationships based on 16S ribosomal DNA sequencing. We found a high prevalence of Pseudomonas in our soil samples, and isolates from these soils exhibited extensive metabolic diversity. In addition, our data revealed that many of our isolates form a unique cold-adapted clade, representatives of which are also found in the Swedish tundra and Antarctica. Our data also show a lack of concordance between the metabolic properties and 16S phylogeny, indicating that the metabolic diversity of these organisms cannot be predicted by phylogeny.     

Bacteriolytic activity of selected vertebrate sera for Borrelia burgdorferi sensu stricto and Borrelia bissettii

An in vitro assay to evaluate the bacteriolytic activity of the complement pathway was applied to 2 strains of Borrelia bissettii, CO501 and DN127, and compared with that of B. burgdorferi sensu stricto B31. Sera from mule deer (Odocoileus hemionus) and the Western Fence lizard (Sceloporus occidentalis) were completely borreliacidal for B. burgdorferi and for both strains of B. bissettii. Serum from Bobwhite quail (Colinus virginianus) was nonlytic for B. burgdorferi and partially lytic for B. bissettii strains, CO-501 and DN127. Serum from a New Zealand White rabbit (Oryctolagus cuniculus) was partially lytic for all 3 strains of Borrelia, whereas serum from white-footed mice (Peromyscus leucopus) were nonlytic for all 3 Borrelia strains. The spectrum of complement sensitivity of B. bissettii appears to be similar to that of European B. afzelii in that tested rodent serum is not lytic to these 2 genospecies. Interestingly, both B. bissettii and B. afzelii have been found to be closely associated with rodents. Complement sensitivity demonstrated in these experiments may suggest and possibly predict specific reservoir-host associations.     

RAPD fingerprinting for the distinction of Prevotella intermedia sensu stricto from Prevotella nigrescens

A collection of 70 oral strains including reference strains and clinical isolates identified as Prevotella intermedia sensu lato was constituted to cover a large clinical and geographical diversity. Electrophoresis of the enzyme malate dehydrogenase allowed the identification of the 70 study strains as Prevotella intermedia sensu stricto (n= 36), Prevotella nigrescens (n= 31) and three unclassified strains. By using four primers, DNA fingerprints were generated from 20 strains as random amplified polymorphic DNA (RAPD). Matching co-migrating amplicon positions by pairwise comparison allowed the clustering of the fingerprints as two groups coincident with the P. intermedia/P. nigrescens assignment by enzyme electrophoresis of malate dehydrogenase. Our data suggest that isolates identified asP. intermedia sensu lato by conventional criteria can be speciated asP. intermedia sensu stricto or P. nigrescens by RAPD fingerprinting.     

Bread, beer and wine: yeast domestication in the Saccharomyces sensu stricto complex

Yeasts of the Saccharomyces sensu stricto species complex are able to convert sugar into ethanol and CO(2) via fermentation. They have been used for thousands years by mankind for fermenting food and beverages. In the Neolithic times, fermentations were probably initiated by naturally occurring yeasts, and it is unknown when humans started to consciously add selected yeast to make beer, wine or bread. Interestingly, such human activities gave rise to the creation of new species in the Saccharomyces sensu stricto complex by interspecies hybridization or polyploidization. Within the S. cerevisiae species, they have led to the differentiation of genetically distinct groups according to the food process origin. Although the evolutionary history of wine yeast populations has been well described, the histories of other domesticated yeasts need further investigation.     

Leaf anatomy of the genus Psoralea sensu stricto (Psoraleeae, Papilionoideae, Leguminosae)

Leaf anatomy is described from 17 species of Psoralea sensu stricto which includes four species of the genus Hallia Thunberg. Species of the two genera share many characters including the presence of rod-shaped crystals, similar stipule structure and a possible continuum in secretory cavity anatomy ranging from a small and non-trabeculate to a large and trabci ulate form. Hallia species are distinguished b the presence of large tannin cells in the bundle sheaths and a narrow lcngllv.width ratio of palisade cells. In the light of other floral evidence this is considered insufficient to separate the two genera, a view confirmed by cluster analysis. Thus, Salter's proposal of 1939 for their amalgamation is supported. Leaf anatomy of herbaceous species is compared with that of seedlings of Psoralea sp. to assess the possibility that the herbaceous species may have arisen through neoteny from species that are trees or shrubs. Comparative venation studies between stipules and scale leaves suggest that the scale leaf form arose from pinnate-leaved ancestors by leaf reduction.     

Phytochemistry and bioactivity of Acacia sensu stricto (Fabaceae: Mimosoideae)

Phytochemistry Reviews - Acacia sensu lato (Fabaceae: Mimosoideae) was recently retypified and divided into five genera worldwide: Acacia, Acaciella, Mariosousa, Senegalia and Vachellia. Acacia...     

Multiplex PCR for the identification of Anisakis simplex sensu stricto, Anisakis pegreffii and the other anisakid nematodes

A multiplex PCR method was established for the rapid identification of Anisakis simplex sensu stricto, A. pegreffii, A. physeteris, Pseudoterranova decipiens, Contracaecum osculatum and Hysterothylacium aduncum. The sequence alignment of the internal transcribed spacer 1 region (ITS-1) between A. simplex s. str. and A. pegreffii showed a high degree of similarity, but only two C-T transitions were observed. To differentiate A. simplex s. str. from A. pegreffii, an intentional mismatch primer with an artificial mismatched base at the second base from the primer 3' end was constructed. This intentional mismatch primer, which produced a PCR band only from A. pegreffii DNA, was able to differentiate the two morphologically indistinguishable sibling species of A. simplex. Specific forward primers for other anisakid species were also designed based on the nucleotide sequences of the ITS region. The multiplex PCR using these primers yielded distinct PCR products for each of the anisakid nematodes. The multiplex PCR established in this study would be a useful tool for identifying anisakid nematodes rapidly and accurately.     

Natural polyploidization of some cultured yeast Saccharomyces sensu stricto: auto- and allotetraploidy

Using genetic and flow cytometric analyses, we showed that wine strain S6U is an allotetraploid of S. cerevisiae x S. bayanus. Hybrid constitution of the strain and its meiotic segregants was confirmed by Southern hybridization analysis of their chromosomal DNAs using four S. cerevisiae cloned genes: LYS2 (chr. II), TRK1 (chr. X), ARG4 (chr. VIII), ACT1 (chr. VI) and PCR/RFLP analysis of the MET2 gene (chr. XIV). Monosporic progeny of strain S6U was highly viable in first generation but completely nonviable in the second one. According to the genetic analysis, sherry strain S. cerevisiae SBY 2592 was found to be an autotetraploid heterozygous for homo-heterothallism.     

Genetic and phenotypic diversity of Saccharomyces sensu stricto strains isolated from Amarone wine

Individual yeast strains belonging to the Saccharomyces sensu stricto complex were isolated from Amarone wine produced in four cellars of the Valpolicella area (Italy) and characterized by conventional physiological tests and by RAPD-PCR and mtDNA restriction assays. Thirteen out of 20 strains were classified as Saccharomyces cerevisiae (ex S. cerevisiae p.r. cerevisiae and p.r. bayanus) and the remaining as Saccharomyces bayanus (ex S. cerevisiae p.r. uvarum). RAPD-PCR method proved to be a fast and reliable tool for identification of Saccharomyces sensu stricto strains and also gave intraspecific differentiation. Restriction analysis of mtDNA permitted to distinguish S. cerevisiae and S. bayanus species and to discern polymorphism among S. cerevisiae isolates. The assessment of the phenotypic diversity within the isolates by gas-chromatographic analysis of secondary fermentation products was explored. Small quantities of isobutanol were produced by most of the strains and higher amounts by some S. cerevisiae strains with phenotypes Gal^- and Mel^-; all S. bayanus strains produced low amounts of amilyc alcohols. From this study it appears that each winery owns particular strains, with different genetic and biochemical characteristics, selected by specific environmental pressures during the Amarone winemaking process carried out at low temperature in presence of high sugar content.     

Physiological and genetic stability of hybrids of industrial wine yeasts Saccharomyces sensu stricto complex

Aims: The aim of this study was to examine the physiological and genetic stability of hybrids of industrial wine yeasts Saccharomyces sensu stricto complex subjected to acidic stress during fermentation. Methods and Results: Laboratory‐constructed yeast hybrids, one intraspecific Saccharomyces cerevisiae × S. cerevisiae and three interspecific S. cerevisiae ×Saccharomyces bayanus, were subcultured in aerobic or anaerobic conditions in media with or without l ‐malic acid. Changes in the biochemical profiles, karyotypes and mitochondrial DNA profiles of the segregates were assessed after 50–190 generations. All yeast segregates showed a tendency to increase the range of the tested compounds utilized as sole carbon sources. Interspecific hybrids were alloaneuploid and their genomes tended to undergo extensive rearrangement especially during fermentation. The karyotypes of segregates lost up to four and appearance up to five bands were recorded. The changes in their mtDNA patterns were even broader reaching 12 missing and six additional bands. These hybrids acquired the ability to sporulate and significantly changed their biochemical profiles. The alloaneuploid intraspecific S. cerevisiae hybrid was characterized by high genetic stability despite the phenotypic changes. l ‐malic acid was not found to affect the extent of genomic changes of the hybrids, which suggests that their demalication ability is combined with resistance to acidic stress. Conclusions: The results reveal the plasticity and extent of changes of chromosomal and mitochondrial DNA of interspecific hybrids of industrial wine yeast especially under anaerobiosis. They imply that karyotyping and restriction analysis of mitochondrial DNA make it possible to quickly assess the genetic stability of genetically modified industrial wine yeasts but may not be applied as the only method for their identification and discrimination. Significance and Impact of the Study: Laboratory‐constructed interspecific hybrids of industrial strains may provide a model for studying the adaptive evolution of wine yeasts under fermentative stress.     

Vector competence of Ixodes angustus (Acari: Ixodidae) for Borrelia burgdorferi sensu stricto

The vector competence of Ixodes angustus for Borrelia burgdorferi sensu stricto (s.s.) was investigated in the laboratory. The larval progeny of female ticks from Washington State were placed on Swiss-Webster mice that had been inoculated intravenously with 10⁸ spirochetes each of a Californian isolate of B. burgdorferi. Spirochetes were detected in 6 (12%) of 50 nymphs derived from larvae that had fed on these animals. Ten nymphs from the same cohort of experimentally infected ticks were placed on each of 4 naive deer mice (Peromyscus maniculatus). One of the mice seroconverted to B. burgdorferi and spirochetes were isolated from its ear tissues 4 weeks after exposure to ticks. Further vector competence trials were conducted with I. angustus ticks from California. Larvae were fed on deer mice that had been inoculated intradermally with B. burgdorferi along with larvae of I. spinipalpis as a comparison group. There was no significant difference in the prevalence of infection in nymphs of I. angustus (8.2%) versus those of I. spinipalpis (12.1%). We conclude that I. angustus is a competent experimental vector of B. burgdorferi s.s. and its efficiency for acquiring and transstadially passing such spirochetes is similar to that of I. spinipalpis.     

Interaction and transmission of two Borrelia burgdorferi sensu stricto strains in a tick-rodent maintenance system

In the northeastern United States, the Lyme disease agent, Borrelia burgdorferi sensu stricto, is maintained by enzoonotic transmission, cycling between white-footed mice (Peromyscus leucopus) and black-legged ticks (Ixodes scapularis). B. burgdorferi sensu stricto is genetically variable and has been divided into three major genotypes based on 16S-23S ribosomal DNA spacer (RST) analysis. To better understand how genetic differences in B. burgdorferi sensu stricto may influence transmission dynamics in nature, we investigated the interaction between an RST1 and an RST3 strain in a laboratory system with P. leucopus mice and I. scapularis ticks. Two groups of mice were infected with either BL206 (RST1) or B348 (RST3). Two weeks later, experimental mice were challenged with the opposite strain, while control mice were challenged with the same strain as that used for the primary infection. The transmission of BL206 and B348 from infected mice was then determined by xenodiagnosis with uninfected larval ticks at weekly intervals for 42 days. Mice in both experimental groups were permissive for infection with the second strain and were able to transmit both strains to the xenodiagnostic ticks. However, the overall transmission efficiencies of BL206 and B348 were significantly different. BL206 was more efficiently transmitted than B348 to xenodiagnostic ticks. Significantly fewer double infections than expected were detected in xenodiagnostic ticks. The results suggest that some B. burgdorferi sensu stricto strains, such as BL206, may be preferentially maintained in transmission cycles between ticks and white-footed mice. Other strains, such as B348, may be more effectively maintained in different tick-vertebrate transmission cycles.