Parasexuality and Ploidy Change in Candida tropicalis

Candida species exhibit a variety of ploidy states and modes of sexual reproduction. Most species possess the requisite genes for sexual reproduction, recombination, and meiosis, yet only a few have been reported to undergo a complete sexual cycle including mating and sporulation. Candida albicans, the most studied Candida species and a prevalent human fungal pathogen, completes its sexual cycle via a parasexual process of concerted chromosome loss rather than a conventional meiosis. In this study, we examine ploidy changes in Candida tropicalis, a closely related species to C. albicans that was recently revealed to undergo sexual mating. C. tropicalis diploid cells mate to form tetraploid cells, and we show that these can be induced to undergo chromosome loss to regenerate diploid forms by growth on sorbose medium. The diploid products are themselves mating competent, thereby establishing a parasexual cycle in this species for the first time. Extended incubation (>120 generations) of C. tropicalis tetraploid cells under rich culture conditions also resulted in instability of the tetraploid form and a gradual reduction in ploidy back to the diploid state. The fitness levels of C. tropicalis diploid and tetraploid cells were compared, and diploid cells exhibited increased fitness relative to tetraploid cells in vitro, despite diploid and tetraploid cells having similar doubling times. Collectively, these experiments demonstrate distinct pathways by which a parasexual cycle can occur in C. tropicalis and indicate that nonmeiotic mechanisms drive ploidy changes in this prevalent human pathogen.     

Virulence phenotypes result from interactions between pathogen ploidy and genetic background

Studying fungal virulence is often challenging and frequently depends on many contexts, including host immune status and pathogen genetic background. However, the role of ploidy has often been overlooked when studying virulence in eukaryotic pathogens. Since fungal pathogens, including the human opportunistic pathogen Candida albicans, can display extensive ploidy variation, assessing how ploidy impacts virulence has important clinical relevance. As an opportunistic pathogen, C. albicans causes nonlethal, superficial infections in healthy individuals, but life‐threatening bloodstream infections in individuals with compromised immune function. Here, we determined how both ploidy and genetic background of C. albicans impacts virulence phenotypes in healthy and immunocompromised nematode hosts by characterizing virulence phenotypes in four near‐isogenic diploid and tetraploid pairs of strains, which included both laboratory and clinical genetic backgrounds. We found that C. albicans infections decreased host survival and negatively impacted host reproduction, and we leveraged these two measures to survey both lethal and nonlethal virulence phenotypes across the multiple C. albicans strains. In this study, we found that regardless of pathogen ploidy or genetic background, immunocompromised hosts were susceptible to fungal infection compared to healthy hosts. Furthermore, for each host context, we found a significant interaction between C. albicans genetic background and ploidy on virulence phenotypes, but no global differences between diploid and tetraploid pathogens were observed.     

Ploidy effects on the relationship between floral phenotype,reproductive investment,and fitness in an autogamous species complex

Premise

The relationships between reproductive investment, phenotype, and fitness have been broadly studied in cross-pollinated plants in contrast to selfing species, which are considered less interesting in this area because they are supposed to be a dead end in any evolutionary pathway. Still, selfing plants are unique systems to study these questions since the position of reproductive structures and traits related to flower size play an important role in female and male pollination success.

Methods

Erysimum incanum s.l. is a selfing species complex that has three levels of ploidy (diploids, tetraploids, and hexaploids) and traits that are typically associated with the selfing syndrome. Here, we used 1609 plants belonging to these three ploidies to characterize the floral phenotype and spatial configuration of reproductive structures, reproductive investment (pollen and ovule production), and plant fitness. Then, we used structural equation modelling to analyze the relationship between all these variables across ploidy levels.

Results

An increase in ploidy level leads to bigger flowers with anthers exserted farther and more pollen and ovules. In addition, hexaploid plants had higher absolute values for herkogamy, which is positively correlated with fitness. Ovule production significantly mediated the natural selection acting on different phenotypic traits and pollen production, a pattern that is maintained across ploidies.

Conclusions

The changes in floral phenotypes, reproductive investment, and fitness with ploidy level suggest that genome duplication can be a driver for transitions in reproductive strategy by modifying the investment in pollen and ovules and linking them with plant phenotype and fitness.     

SPONTANEOUS MUTATION ACCUMULATION IN MULTIPLE STRAINS OF THE GREEN ALGA,CHLAMYDOMONAS REINHARDTII

Estimates of mutational parameters, such as the average fitness effect of a new mutation and the rate at which new genetic variation for fitness is created by mutation, are important for the understanding of many biological processes. However, the causes of interspecific variation in mutational parameters and the extent to which they vary within species remain largely unknown. We maintained multiple strains of the unicellular eukaryote Chlamydomonas reinhardtii, for approximately 1000 generations under relaxed selection by transferring a single cell every ～10 generations. Mean fitness of the lines tended to decline with generations of mutation accumulation whereas mutational variance increased. We did not find any evidence for differences among strains in any of the mutational parameters estimated. The overall change in mean fitness per cell division and rate of input of mutational variance per cell division were more similar to values observed in multicellular organisms than to those in other single‐celled microbes. However, after taking into account differences in genome size among species, estimates from multicellular organisms and microbes, including our new estimates from C. reinhardtii, become substantially more similar. Thus, we suggest that variation in genome size is an important determinant of interspecific variation in mutational parameters.     

Do ploidy level and nuclear genome size and latitude of origin modify the expression of <Emphasis Type="Italic">Phragmites australis</Emphasis> traits and interactions with herbivores?

We studied the relationship between genome size and ploidy level variation and plant traits for the reed grass Phragmites australis. Using a common garden approach on a global collection of populations in Aarhus, Denmark, we investigated the influence of monoploid genome size and ploidy level on the expression of P. australis growth, nutrition and herbivore-defense traits and whether monoploid genome size and ploidy level play different roles in plant trait expression. We found that both monoploid genome size and latitude of origin contributed to variation in traits that we studied for P. australis, with latitude of origin being generally a better predictor of trait values and that ploidy level and its interaction with monoploid genome size and latitude of origin also contributed to trait variation. We also found that for four traits, tetraploids and octoploids had different relationships with the monoploid genome size. While for tetraploids stem height and leaf water content showed a positive relationship with monoploid genome size, octoploids had a negative relationship with monoploid genome size for stem height and no relationship for leaf water content. As genome size within octoploids increased, the number of aphids colonizing leaves decreased, whereas for tetraploids there was a quadratic, though non-significant, relationship. Generally we found that tetraploids were taller, chemically better defended, had a greater number of stems, higher leaf water content, and supported more aphids than octoploids. Our results suggest trade-offs among plant traits mediated by genome size and ploidy with respect to fitness and defense. We also found that the latitude of plant origin is a significant determinant of trait expression suggesting local adaptation. Global climate change may favor some genome size and ploidy variants that can tolerate stressful environments due to greater phenotypic plasticity and to fitness traits that vary with cytotype which may lead to changes in population genome sizes and/or ploidy structure, particularly at species’ range limits.     

Reversed selection responses in small populations of the housefly (Musca domestica L.)

We compared the efficacy of artificial and natural selection processes in purging the genetic load of perpetually small populations. We subjected replicate lines of the housefly (Musca domestica L.), recently derived from the wild, to artificial selection for increased mating propensity (i.e., the proportion of male–female pairs initiating copulation within 30 min) in efforts to cull out the inbreeding depression effects of long-term small population size (as determined by a selection protocol for increased assortative mating). We also maintained parallel non-selection lines for assessing the spontaneous purge of genetic load due to inbreeding alone. We thus evaluated the fitness of artificially and ‘naturally’ purging populations held at census sizes of 40 individuals over the course of 18 generations. We found that the artificially selected lines had significant increases in mating propensity (up to 46% higher from the beginning of the protocol) followed by reversed selection responses back to the initial levels, resulting in non-significant heritabilities. Nevertheless, the ‘naturally’ selected lines had significantly lower fitness overall (a 28% reduction from the beginning of the protocol), although lower effective population sizes could have contributed to this effect. We conclude that artificial selection bolstered fitness, but only in the short-term, because the inadvertent fixation of extant genetic load later resulted in pleiotropic fitness declines. Still, the short-term advantage of the selection protocol likely contributed to the success of the speciation experiment since our recently-derived housefly populations are particularly vulnerable to inbreeding depression effects on mating behavior.     

Ploidy level and genome size of locally adapted populations of <Emphasis Type="Italic">Silene ciliata</Emphasis> across an altitudinal gradient

Silene ciliata Poiret is a small perennial that presents several ploidy levels and inhabits the mountain ranges of the European Mediterranean basin. Recent studies have shown evidence of local adaptation in populations located across an altitudinal gradient in Sierra de Guadarrama (Central Spain) at the species’ southernmost distribution limit. In this study, we assessed whether the existence of local adaptation in these populations was related to differences in karyological features (ploidy level or chromosome number modification) or in nuclear DNA amount. Optical microscope (phase contrast and epifluorescence after DAPI staining) and flow cytometry were used to estimate the ploidy level and genome size of several family lines in three populations across the altitude gradient. With a sampling three times higher than usual in genome size assessments, all individuals showed a constant diploid set (2n = 24), so that polyploidy or other chromosome number modifications were discarded. The small genome size found (mean ± SD; 2C = 1.76 ± 0.06 pg) was within the range of those found in other Silene species. Significant differences in genome size were found when the three populations of S. ciliata were compared. The largest genome size found at the intermediate population may be associated to lower environmental stress at the mid elevation, in line with the recent studies in this area.     

Aurora kinase Ipl1 facilitates bilobed distribution of clustered kinetochores to ensure error‐free chromosome segregation in Candida albicans

Candida albicans, an ascomycete, has an ability to switch to diverse morphological forms. While C. albicans is predominatly diploid, it can tolerate aneuploidy as a survival strategy under stress. Aurora kinase B homolog Ipl1 is a critical ploidy regulator that controls microtubule dynamics and chromosome segregation in Saccharomyces cerevisiae. In this study, we show that Ipl1 in C. albicans has a longer activation loop than that of the well‐studied ascomycete S. cerevisiae. Ipl1 localizes to the kinetochores during the G1/S phase and associates with the spindle during mitosis. Ipl1 regulates cell morphogenesis and is required for cell viability. Ipl1 monitors microtubule dynamics which is mediated by separation of spindle pole bodies. While Ipl1 is dispensable for maintaining structural integrity and clustering of kinetochores in C. albicans, it is required for the maintenance of bilobed distribution of clustered kinetochores along the mitotic spindle. Depletion of Ipl1 results in erroneous kinetochore‐microtubule attachments leading to aneuploidy due to which the organism can survive better in the presence of fluconazole. Taking together, we suggest that Ipl1 spatiotemporally ensures bilobed kinetochore distribution to facilitate bipolar spindle assembly crucial for ploidy maintenance in C. albicans.     

Mutation accumulation in populations of varying size: large effect mutations cause most mutational decline in the rotifer Brachionus calyciflorus under UV‐C radiation

Theory predicts that fitness decline via mutation accumulation will depend on population size, but there are only a few direct tests of this key idea. To gain a qualitative understanding of the fitness effect of new mutations, we performed a mutation accumulation experiment with the facultative sexual rotifer Brachionus calyciflorus at six different population sizes under UV‐C radiation. Lifetime reproduction assays conducted after ten and sixteen UV‐C radiations showed that while small populations lost fitness, fitness losses diminished rapidly with increasing population size. Populations kept as low as 10 individuals were able to maintain fitness close to the nonmutagenized populations throughout the experiment indicating that selection was able to remove the majority of large effect mutations in small populations. Although our results also seem to imply that small populations are effectively immune to mutational decay, we caution against this interpretation. Given sufficient time, populations of moderate to large size can experience declines in fitness from accumulating weakly deleterious mutations as demonstrated by fitness estimates from simulations and, tentatively, from a long‐term experiment with populations of moderate size. There is mounting evidence to suggest that mutational distributions contain a heavier tail of large effects. Our results suggest that this is also true when the mutational spectrum is altered by UV radiation.     

Flowering‐mediated root‐fungus symbiosis loss is related to jasmonate‐dependent root soluble sugar deprivation

The role of flowering in root‐fungal symbiosis is not well understood. Because flowering and fungal symbionts are supported by carbohydrates, we hypothesized that flowering modulates root‐beneficial fungal associations through alterations in carbohydrate metabolism and transport. We monitored fungal colonization and soluble sugars in the roots of Arabidopsis thaliana following inoculation with a mutualistic fungus Phomopsis liquidambari across different plant developmental stages. Jasmonate signalling of wild‐type plants, sugar transport, and root invertase of wild‐type and jasmonate‐insensitive plants were exploited to assess whether and how jasmonate‐dependent sugar dynamics are involved in flowering‐mediated fungal colonization alterations. We found that flowering restricts root‐fungal colonization and activates root jasmonate signalling upon fungal inoculation. Jasmonates reduce the constitutive and fungus‐induced accumulation of root glucose and fructose at the flowering stage. Further experiments with sugar transport and metabolism mutant lines revealed that root glucose and fructose positively influence fungal colonization. Diurnal, jasmonate‐dependent inhibitions of sugar transport and soluble invertase activity were identified as likely mechanisms for flowering‐mediated root sugar depletion upon fungal inoculation. Collectively, our results reveal that flowering drives root‐fungus cooperation loss, which is related to jasmonate‐dependent root soluble sugar depletion. Limiting the spread of root‐fungal colonization may direct more resources to flower development.     

JOINT ALLELIC EFFECTS ON FITNESS AND METRIC TRAITS

Theoretical explanations of empirically observed standing genetic variation, mutation, and selection suggest that many alleles must jointly affect fitness and metric traits. However, there are few direct demonstrations of the nature and extent of these pleiotropic associations. We implemented a mutation accumulation (MA) divergence experimental design in Drosophila serrata to segregate genetic variants for fitness and metric traits. By exploiting naturally occurring MA line extinctions as a measure of line‐level total fitness, manipulating sexual selection, and measuring productivity we were able to demonstrate genetic covariance between fitness and standard metric traits, wing size, and shape. Larger size was associated with lower total fitness and male sexual fitness, but higher productivity. Multivariate wing shape traits, capturing major axes of wing shape variation among MA lines, evolved only in the absence of sexual selection, and to the greatest extent in lines that went extinct, indicating that mutations contributing wing shape variation also typically had deleterious effects on both total fitness and male sexual fitness. This pleiotropic covariance of metric traits with fitness will drive their evolution, and generate the appearance of selection on the metric traits even in the absence of a direct contribution to fitness.     

Fitness decline in spontaneous mutation accumulation lines of Caenorhabditis elegans with varying effective population sizes

The rate and fitness effects of new mutations have been investigated by mutation accumulation (MA) experiments in which organisms are maintained at a constant minimal population size to facilitate the accumulation of mutations with minimal efficacy of selection. We evolved 35 MA lines of Caenorhabditis elegans in parallel for 409 generations at three population sizes (N = 1, 10, and 100), representing the first spontaneous long-term MA experiment at varying population sizes with corresponding differences in the efficacy of selection. Productivity and survivorship in the N = 1 lines declined by 44% and 12%, respectively. The average effects of deleterious mutations in N = 1 lines are estimated to be 16.4% for productivity and 11.8% for survivorship. Larger populations (N = 10 and 100) did not suffer a significant decline in fitness traits despite a lengthy and sustained regime of consecutive bottlenecks exceeding 400 generations. Together, these results suggest that fitness decline in very small populations is dominated by mutations with large deleterious effects. It is possible that the MA lines at larger population sizes contain a load of cryptic deleterious mutations of small to moderate effects that would be revealed in more challenging environments.     

Quantification and decomposition of environment‐selection relationships

In nature, selection varies across time in most environments, but we lack an understanding of how specific ecological changes drive this variation. Ecological factors can alter phenotypic selection coefficients through changes in trait distributions or individual mean fitness, even when the trait‐absolute fitness relationship remains constant. We apply and extend a regression‐based approach in a population of Soay sheep (Ovis aries) and suggest metrics of environment‐selection relationships that can be compared across studies. We then introduce a novel method that constructs an environmentally structured fitness function. This allows calculation of full (as in existing approaches) and partial (acting separately through the absolute fitness function slope, mean fitness, and phenotype distribution) sensitivities of selection to an ecological variable. Both approaches show positive overall effects of density on viability selection of lamb mass. However, the second approach demonstrates that this relationship is largely driven by effects of density on mean fitness, rather than on the trait‐fitness relationship slope. If such mechanisms of environmental dependence of selection are common, this could have important implications regarding the frequency of fluctuating selection, and how previous selection inferences relate to longer term evolutionary dynamics.     

Does sibling competition have a sex‐specific effect on offspring growth and development in the burying beetle Nicrophorus vespilloides?

During offspring growth, reduction in resource availability through competitive interactions will restrict how large individuals can become. Given that selection to mature at a large size will be greater for the sex in which large size provides the highest fitness return, sex‐specific differences in response to a decrease in resource availability may be expected. Using Nicrophorus vespilloides Herbst (Coleoptera: Silphidae) we examined the sex‐specific response of offspring to resource availability through sibling competition. We found that males and females were affected similarly by an increase in the level of sibling competition as brood size increased. Interestingly, although male N. vespilloides were consistently heavier than females, over a range of brood sizes, they were only significantly heavier than females at intermediate brood sizes. At present, the causes behind this finding remain unclear.     

Polyploidy does not control all: Lineage‐specific average chromosome length constrains genome size evolution in ferns

Recent studies investigating the evolution of genome size diversity in ferns have shown that they have a distinctive genome profile compared with other land plants. Ferns are typically characterized by possessing medium‐sized genomes, although a few lineages have evolved very large genomes. Ferns are different from other vascular plant lineages as they are the only group to show evidence for a correlation between genome size and chromosome number. In this study, we aim to explore whether the evolution of fern genome sizes is not only shaped by chromosome number changes arising from polyploidy but also by constraints on the average amount of DNA per chromosome. We selected the genus Asplenium L. as a model genus to study the question because of the unique combination of a highly conserved base chromosome number and a high frequency of polyploidy. New genome size data for Asplenium taxa were combined with existing data and analyzed within a phylogenetic framework. Genome size varied substantially between diploid species, resulting in overlapping genome sizes among diploid and tetraploid spleenworts. The observed additive pattern indicates the absence of genome downsizing following polyploidy. The genome size of diploids varied non‐randomly and we found evidence for clade‐specific trends towards larger or smaller genomes. The 578‐fold range of fern genome sizes have arisen not only from repeated cycles of polyploidy but also through clade‐specific constraints governing accumulation and/or elimination of DNA.     

Design of a Simple Model of <Emphasis Type="Italic">Candida albicans</Emphasis> Biofilms Formed under Conditions of Flow: Development,Architecture, and Drug Resistance

Candida albicans biofilms on most medical devices are exposed to a flow of body fluids that provide water and nutrients to the fungal cells. While C. albicans biofilms grown in vitro under static conditions have been exhaustively studied, the same is not true for biofilms developed under continuous flow of replenishing nutrients. Here, we describe a simple flow biofilm (FB) model that can be built easily with materials commonly available in most microbiological laboratories. We demonstrate that C. albicans biofilms formed using this flow system show increased architectural complexity compared to biofilms grown under static conditions. C. albicans biofilms under continuous medium flow grow rapidly, and by 8 h show characteristics similar to 24 h statically grown biofilms. Biomass measurements and microscopic observations further revealed that after 24 h of incubation, FB was more than twofold thicker than biofilms grown under static conditions. Microscopic analyses revealed that the surface of these biofilms was extremely compact and wrinkled, unlike the open hyphal layer typically seen in 24 h static biofilms. Results of antifungal drug susceptibility tests showed that C. albicans cells in FB exhibited increased resistance to most clinically used antifungal agents.