The Saccharomyces sensu stricto group comprises species that are of industrial importance as well as species that are confined to natural environments. Recent discoveries of various wild Saccharomyces species have increased research interest in the study of yeast ecology and biogeography. In this PhD thesis, the newly discovered wild species S. jurei was characterised both genetically and phenotypically. Also, the biodiversity of Saccharomyces species that inhabit tree bark and the surrounding soil at different altitude zones in the Italian Carnic Alps was investigated using classical culturing methods and a newly developed targeted metagenomics approach. Genetic crossing between the two S. jurei strains (NCYC 3947T and NCYC 3962) and other species in the Saccharomyces genus revealed that the S. jurei strains are reproductively isolated and, therefore, constitute a distinct biological species. Both strains were able to grow well in a wide range of temperatures, demonstrating a generalist growth thermoprofile. Under environmental stresses, the fitness of NCYC 3962 was greater than that of NCYC 3947T, particularly under the condition of 0.6% acetic acid. The impact of mitochondrial DNA (mtDNA) on phenotypic variation was tested using hybrids between S. jurei and S. cerevisiae or S. uvarum. Upon the crossing of two yeast species, the mtDNA is initially biparetally inherited, but, after ~20 generations, the hybrids retain only one mitotype. Here, I constructed hybrids under different environmental conditions to study the effects on mitochondrial inheritance. In glycerol media at 35Â°C, S. cerevisiae mtDNA was preferentially retained, and at 10Â°C, S. jurei mtDNA was retained in both crosses. The results revealed that the carbon source and temperature have significant effect on mtDNA inheritance. The fitness of the hybrids at different temperatures correlated with mtDNA inheritance from either the cryotolerant or thermotolerant parent. The application of different sugars in the enrichment media used to isolate the Saccharomyces species from oak bark and soil revealed that sucrose is the most efficient sugar for the isolation of S. paradoxus. The use of melezitose allowed the isolation of S. kudriavzevii strains oak bark incubated at 10Â°C. Phylogenetic and variant analyses of the representative strains of this species revealed a distinct Italian group that was closely related to the European population. However, the Italian group was more reproductively isolated from the European strains than the Asian strains. This result may be due to undetected chromosomal rearrangements. Since the enrichment culturing method can introduce biases for the isolation of S. paradoxus, a targeted high-throughput sequencing of DNA extracted from the soil surrounding trees at different altitudes was performed. A selectivity step was conducted prior to the sequencing that was based on the amplification of the internal transcribed spacer (ITS) rRNA, which has a size that is specific to the Saccharomyces genus. Sequencing of the ITS1 region from the selected ITS resulted in the detection of most Saccharomyces species. Interestingly, sequences belonging to both S. eubayanus and S. mikatae were retrieved, providing proof of their presence in Europe. The detected Saccharomyces sequenced differed by up to 3 bp in comparison to sequences of known populations which may indicate new distinct populations. The results of this PhD thesis provided a preliminary phenotypic characterisation of novel S. jurei strains in response to environmental conditions, highlighting the strainsâ potential for use in biotechnological and industrial applications. Also, the results of this study support the use of both culturing methods and targeted metagenomics to capture the diversity of Saccharomyces species in wild environments.
|Date of Award||1 Aug 2019|
- The University of Manchester
|Supervisor||Daniela Delneri (Supervisor) & Chris Knight (Supervisor)|