Habitat fragmentation is one of the main threats to semi-natural grasslands all over Europe, causing reductions in both numbers of populations and numbers of individuals withinpopulations. Small isolated populations are particularly vulnerable to fluctuations that reducespopulation size and could lead to loss of genetic variation due to inbreeding depression, orfixation of harmful alleles due to genetic drift. The aim of this study was to investigate if thecritically endangered eyebright Euphrasia stricta var. suecica experiences pollen limitation,inbreeding depression and heterosis or outbreeding depression. Low pollen limitation waspredicted as well as low inbreeding depression due to high selfing rate. Outbreedingdepression or heterosis was expected based on previous studies indicating strong geneticdifferentiation among populations. This study was conducted in two traditionally managedmeadows in the central part of Gotland, Sweden, using supplemental hand-pollinations andcontrolled crosses. Individuals were collected when the flowering period was over and seedswere counted and weighed. There were no signs of either pollen limitation, inbreedingdepression, heterosis or outbreeding depression in any of the two wooded hay meadows. Thedegree of autonomous selfing was high, indicating that E. stricta var. suecica is not dependenton pollinators for seed production. A significant difference between the meadows was foundin proportion of viable seeds. As there were no signs of local adaptation, individuals could betranslocated between the two studied populations or seed sowings could be used to increasepopulation size. However, before such conservation efforts are undertaken, it is important toconduct further studies that investigate several populations including more life-stages.

<p> </p> <p>The degree of self-fertilization in a population determines levels of genetic variation and high selfing rates could thus limit future adaptive potential. Theory predicts that intermediate selfing rates should not persist, yet many plants exhibit mixed mating. Cleistogamy is a floral heteromorphism where individuals produce both showy potentially outcrossing chasmogamous flowers and closed obligately selfing cleistogamous flowers. Reproduction via cleistogamous flowers is thought to be beneficial because of their greater energetic economy compared to chasmogamous flowers but can be costly if selfing leads to inbreeding depression or accelerates the fixation of deleterious mutations within populations. Cleistogamy has evolved independently multiple times and can be used to study the maintenance of adaptive mixed mating. To investigate this, I estimated the costs and benefits of selfing in three populations of <em>Ruellia humilis </em>Nutt (Acanthaceae) in greenhouse common garden experiments. To quantify the costs, I performed hand pollinations and quantified fitness components of progeny resulting from selfing and outcrossing within- and between-populations. To quantify the relative energetic advantage of cleistogamous flowers, I measured dry flower mass, fertility, seed number per fruit, and pollen-ovule ratios of both types of flowers. I found negative cumulative inbreeding depression in all populations, indicative of selfed progeny outperforming outcrossed progeny. While the fitness consequences of between population outcrossing ranged from heterosis to outbreeding depression. When looking at the energetic benefits of selfing, I found that the cost of reproduction via cleistogamous flowers was between 3 and 14-fold less than the cost for outcrossing flowers. Finally, I combined data on inbreeding depression and the energetic costs of reproduction and found that chasmogamous flowers of <em>R. humilis </em>must provide between a 3 to a 45-fold fitness advantage to be maintained, the magnitude of which was dependent upon maternal population. Overall, I conclude that none of the existing hypotheses are sufficient enough to provide the selective advantage needed to explain the persistence of chasmogamous flowers in <em>R. humilis</em>. Without any supported explanations for the maintenance of mixed mating, the exploration of genetic constraints on the loss of chasmogamous flowers could solve this long-standing mystery. </p>