Effect of resource availability on evolution of virulence and competition in an environmentally transmitted pathogen.

Understanding ecological and epidemiological factors driving pathogen evolution in contemporary time scales is a major challenge in modern health management. Pathogens that replicate outside the hosts are subject to selection imposed by ambient environmental conditions. Increased nutrient levels could increase pathogen virulence by pre-adapting for efficient use of resources upon contact to a nutrient rich host or by favouring transmission of fast-growing virulent strains. We measured changes in virulence and competition in Flavobacterium columnare, a bacterial pathogen of freshwater fish, under high and low nutrient levels. To test competition between strains in genotype mixtures, we developed a quantitative real-time PCR assay. We found that a virulent strain maintained its virulence and outcompeted less virulent strains independent of the nutrient level and resource renewal rate while a less virulent strain further lost virulence in chemostats under low nutrient level and over long-term serial culture under high nutrient level. Our results suggest that increased outside-host nutrient levels might maintain virulence in less virulent strains and increase their contribution to epidemics in aquaculture. The results highlight a need to further explore the role of resource in the outside-host environment in maintaining strain diversity and driving evolution of virulence among environmentally growing pathogens.

A multilocus sequence analysis scheme for characterization of Flavobacterium columnare isolates.

BACKGROUND: Columnaris disease caused by Flavobacterium columnare is a serious problem in aquaculture, annually causing large economic losses around the world. Despite considerable research, the molecular epidemiology of F. columnare remains poorly understood. METHODS: We investigated the population structure and spatiotemporal changes in the genetic diversity of F. columnare population in Finland by using a multilocus sequence typing (MLST) and analysis (MLSA) based on DNA sequence variation within six housekeeping genes. A total of 83 strains of F. columnare were collected from eight different areas located across the country between 2003 and 2012. RESULTS: Partial sequencing of six housekeeping genes (trpB, tuf, atpA, rpoD, gyrB and dnaK) revealed eight sequence types and a moderate level of genetic diversity (H=0.460). Phylogenetic analysis of the concatenated protein-encoding gene sequence data (ca. 3,509 nucleotides) formed two lineages, which could be further divided into five clusters. All analysed F. columnare strains appeared to have a genetic origin distinct from that of another important fish pathogen form the genus Flavobacterium, F. psychrophilum. Although the value of the index of association between alleles, 0.292 (P<0.001), supports some degree of clonality for this species in Finland, recombination has introduced molecular diversity to the population almost three times more than mutation. CONCLUSION: The results suggest that Finnish F. columnare strains have an epidemic population structure followed by clonal expansion of successful genotypes. Our study with reproducible methodology and comparable results establishes a robust framework for the discrimination and phylogenetic analysis of F. columnare isolates, which will help to improve our understanding about geographic distribution and epidemiology of columnaris disease.

The effect of inbreeding rate on fitness, inbreeding depression and heterosis over a range of inbreeding coefficients.

Understanding the effects of inbreeding and genetic drift within populations and hybridization between genetically differentiated populations is important for many basic and applied questions in ecology and evolutionary biology. The magnitudes and even the directions of these effects can be influenced by various factors, especially by the current and historical population size (i.e. inbreeding rate). Using Drosophila littoralis as a model species, we studied the effect of inbreeding rate over a range of inbreeding levels on (i) mean fitness of a population (relative to that of an outbred control population), (ii) within-population inbreeding depression (reduction in fitness of offspring from inbred versus random mating within a population) and (iii) heterosis (increase in fitness of offspring from interpopulation versus within-population random mating). Inbreeding rate was manipulated by using three population sizes (2, 10 and 40), and fitness was measured as offspring survival and fecundity. Fast inbreeding (smaller effective population size) resulted in greater reduction in population mean fitness than slow inbreeding, when populations were compared over similar inbreeding coefficients. Correspondingly, populations with faster inbreeding expressed more heterosis upon interpopulation hybridization. Inbreeding depression within the populations did not have a clear relationship with either the rate or the level of inbreeding.

The benefits of interpopulation hybridization diminish with increasing divergence of small populations.

Interpopulation hybridization can increase the viability of small populations suffering from inbreeding and genetic drift, but it can also result in outbreeding depression. The outcome of hybridization can depend on various factors, including the level of genetic divergence between the populations, and the number of source populations. Furthermore, the effects of hybridization can change between generations following the hybridization. We studied the effects of population divergence (low vs. high level of divergence) and the number of source populations (two vs. four source populations) on the viability of hybrid populations using experimental Drosophila littoralis populations. Population viability was measured for seven generations after hybridization as proportion of populations facing extinction and as per capita offspring production. Hybrid populations established at the low level of population divergence were more viable than the inbred source populations and had higher offspring production than the large control population. The positive effects of hybridization lasted for the seven generations. In contrast, at the high level of divergence, the viability of the hybrid populations was not significantly different from the inbred source populations, and offspring production in the hybrid populations was lower than in the large control population. The number of source populations did not have a significant effect at either low or high level of population divergence. The study shows that the benefits of interpopulation hybridization may decrease with increasing divergence of the populations, even when the populations share identical environmental conditions. We discuss the possible genetic mechanisms explaining the results and address the implications for conservation of populations.

Inbreeding rate modifies the dynamics of genetic load in small populations.

The negative fitness consequences of close inbreeding are widely recognized, but predicting the long-term effects of inbreeding and genetic drift due to limited population size is not straightforward. As the frequency and homozygosity of recessive deleterious alleles increase, selection can remove (purge) them from a population, reducing the genetic load. At the same time, small population size relaxes selection against mildly harmful mutations, which may lead to accumulation of genetic load. The efficiency of purging and the accumulation of mutations both depend on the rate of inbreeding (i.e., population size) and on the nature of mutations. We studied how increasing levels of inbreeding affect offspring production and extinction in experimental Drosophila littoralis populations replicated in two sizes, N = 10 and N = 40. Offspring production and extinction were measured over 25 generations concurrently with a large control population. In the N = 10 populations, offspring production decreased strongly at low levels of inbreeding, then recovered only to show a consistent subsequent decline, suggesting early expression and purging of recessive highly deleterious alleles and subsequent accumulation of mildly harmful mutations. In the N = 40 populations, offspring production declined only after inbreeding reached higher levels, suggesting that inbreeding and genetic drift pose a smaller threat to population fitness when inbreeding is slow. Our results suggest that highly deleterious alleles can be purged in small populations already at low levels of inbreeding, but that purging does not protect the small populations from eventual genetic deterioration and extinction.

Laboratory relationships between adult lifetime reproductive success and fitness surrogates in a Drosophila littoralis population.

The difficulties in measuring total fitness of individuals necessitate the use of fitness surrogates in ecological and evolutionary studies. These surrogates can be different components of fitness (e.g. survival or fecundity), or proxies more uncertainly related to fitness (e.g. body size or growth rate). Ideally, fitness would be measured over the lifetime of individuals; however, more convenient short-time measures are often used. Adult lifetime reproductive success (adult LRS) is closely related to the total fitness of individuals, but it is difficult to measure and rarely included in fitness estimation in experimental studies. We explored phenotypic correlations between female adult LRS and various commonly used fitness components and proxies in a recently founded laboratory population of Drosophila littoralis. Noting that survival is usually higher in laboratory conditions than in nature, we also calculated adjusted adult LRS measures that give more weight to early reproduction. The lifetime measures of fecundity, longevity, and offspring viability were all relatively highly correlated with adult LRS. However, correlations with short-time measures of fecundity and offspring production varied greatly depending on the time of measurement, and the optimal time for measurement was different for unadjusted compared to adjusted adult LRS measures. Correlations between size measures and adult LRS varied from weak to modest, leg size and female weight having the highest correlations. Our results stress the importance of well-founded choice of fitness surrogates in empirical research.