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The long-term persistence of a population which has suffered a bottleneck partly depends on how historical demographic dynamics impacted its genetic diversity and the accumulation of deleterious mutations. Here we provide genomic evidence for the detrimental genetic effect of a recent population bottleneck in the endangered black-faced spoonbill (Platalea minor) even after its rapid population recovery. Our population genomic data suggest that the bird’s effective population size, N, had been relatively stable (7,500-9,000) since the end of the last glacial maximum; however, a recent brief yet severe bottleneck (N= 20) around the 1940s wiped out more than 99% of its historical N in roughly three generations. By comparing it with its sister species, the royal spoonbill (P. regia) whose conservation status is of lesser concern, we found that despite a more than 15-fold population recovery since 1988, genetic drift has led to higher levels of inbreeding (7.4 times more runs of homozygosity longer than 100 Kb) in the black-faced spoonbill than in the royal spoonbill genome. Although the two spoonbills have similar levels of genome-wide nucleotide diversity and heterozygosity, because of relaxed purifying selection, individual black-faced spoonbills carry 3% more nonsynonymous substitutions than royal spoonbills each of which is 7% more deleterious. Our results imply that the persistence of a threatened species cannot be inferred from a recovery in its population. They also highlight the necessity of continually using genomic indices to monitor its genetic health and employing all possible measures to assure its long-term persistence in the ever-changing environment.

Demographic bottlenecks generally reduce genetic diversity through more intense genetic drift, but their net effect may vary along the genome due to the random nature of genetic drift and to local effects of recombination, mutation, and selection. Here, we analyzed the changes in genetic diversity following a bottleneck by comparing whole-genome diversity patterns in populations with and without severe recent documented declines of Iberian (Lynx pardinus, n = 31) and Eurasian lynx (Lynx lynx, n = 29). As expected, overall genomic diversity correlated negatively with bottleneck intensity and/or duration. Correlations of genetic diversity with divergence, chromosome size, gene or functional site content, GC content, or recombination were observed in nonbottlenecked populations, but were weaker in bottlenecked populations. Also, functional features under intense purifying selection and the X chromosome showed an increase in the observed density of variants, even resulting in higher θW diversity than in nonbottlenecked populations. Increased diversity seems to be related to both a higher mutational input in those regions creating a large collection of low-frequency variants, a few of which increase in frequency during the bottleneck to the point they become detectable with our limited sample, and the reduced efficacy of purifying selection, which affects not only protein structure and function but also the regulation of gene expression. The results of this study alert to the possible reduction of fitness and adaptive potential associated with the genomic erosion in regulatory elements. Further, the detection of a gain of diversity in ultra-conserved elements can be used as a sensitive and easy-to-apply signature of genetic erosion in wild populations.

Biotechnology has the potential to deliver solutions to many global problems in medicine, materials science, nutrition, agriculture, natural resource preservation, and energy. Engineered cells and enzymes can perform chemical transformations that are rare or unknown in nature, and even catalyze reactions not accessible to traditional synthetic chemistry while also operating at gentler, more environmentally friendly conditions. Decreases in the price of DNA sequencing and synthesis has led to generation of vast databases that can be screened for any imaginable function. These sequence databases are even more powerful now due to the development of software to enable rapid generation of 3D protein structures, like AlphaFold2. However, tools to predict the function of these proteins or their performance in engineered cells are not yet robust, leading to long development times and limited successful applications to date. New tools and methods must be developed for the true potential of biotechnology to be unlocked.For my thesis, I explore metabolic pathway construction and screening, protein design, and protein sequence-structure-function relationships across broad contexts with the objective of tool and knowledge development for future efforts in biotechnology. My first chapter discusses the introduction of the triazine degradation pathway, of interest for remediation of contaminated sites, into E. coli and methods for characterizing pathway flux and identifying bottlenecks to guide engineering efforts and limit accumulation of metabolic intermediates. My second chapter focuses on methods for the design of supercharged proteins, which have many interesting potential applications, and parameters that increase the likelihood of successful design of these proteins. My third chapter regards the generation and characterization of a library of single point mutations in the enzyme B-glucosidase B and use of kinetic data to predict the effects of changes in sequence on enzyme function. These seemingly disparate topics all serve to improve tools for protein screening, production, functional prediction, and application, addressing several gaps toward improved development timelines and success rates for biocatalysts.

Understanding of the present-day genetic diversity, population structure, and evolutionary history of tree species can inform resource management and conservation activities, including response to pressures presented by a changing climate. Cercis canadensis (Eastern Redbud) is an economically valuable understory tree species native to the United States (U.S.) that is also important for forest ecosystem and wildlife health. Here, we document and explain the population genetics and evolutionary history of this deciduous tree species across its distributed range. In this study, we used twelve microsatellite markers to investigate 691 wild-type trees sampled at 74 collection sites from 23 Eastern U.S. states. High genetic diversity and limited gene flow were revealed in wild, natural stands of C. canadensis with populations that are explained by two major genetic clusters. These findings indicate that an ancient population bottleneck occurred coinciding with the last glacial maximum (LGM) in North America. The structure in current populations likely originated from an ancient population in the eastern U.S. that survived LGM and then later diverged into two contemporary clusters. Data suggests that populations have expanded since the last glaciation event from one into several post-glacial refugia that now occupy this species’ current geographic range. Our enhanced understanding benchmarks the genetic variation preserved within this species and can direct future efforts in conservation, and resource utilization of adaptively resilient populations that present the greatest genetic and structural diversity.

During antibiotic treatment, the evolution of bacterial pathogens is fundamentally affected by bottlenecks and varying selection levels imposed by the drugs. Bottlenecks—that is, reductions in bacterial population size—lead to an increased influence of random effects (genetic drift) during bacterial evolution, and varying antibiotic concentrations during treatment may favour distinct resistance variants. Both aspects influence the process of bacterial evolution during antibiotic therapy and thereby treatment outcome. Surprisingly, the joint influence of these interconnected factors on the evolution of antibiotic resistance remains largely unexplored. Here we combine evolution experiments with genomic and genetic analyses to demonstrate that bottleneck size and antibiotic-induced selection reproducibly impact the evolutionary path to resistance in pathogenic Pseudomonas aeruginosa, one of the most problematic opportunistic human pathogens. Resistance is favoured—expectedly—under high antibiotic selection and weak bottlenecks, but—unexpectedly—also under low antibiotic selection and severe bottlenecks. The latter is likely to result from a reduced probability of losing favourable variants through drift under weak selection. Moreover, the absence of high resistance under low selection and weak bottlenecks is caused by the spread of low-resistance variants with high competitive fitness under these conditions. We conclude that bottlenecks, in combination with drug-induced selection, are currently neglected key determinants of pathogen evolution and outcome of antibiotic treatment., Strong population bottlenecks in combination with weak antibiotic selection consistently favours the evolution of resistance across independently performed Pseudomonas aeruginosa evolution experiments.

With the rapid development of genetic analysis techniques and crop population size, phenotyping has become the bottleneck restricting crop breeding. Breaking through this bottleneck will require phenomics, defined as the accurate, high-throughput acquisition and analysis of multi-dimensional phenotypes during crop growth at organism-wide levels, ranging from cells to organs, individual plants, plots, and fields. Here we offer an overview of crop phenomics research from technological and platform viewpoints at various scales, including microscopic, ground-based, and aerial phenotyping and phenotypic data analysis. We describe recent applications of high-throughput phenotyping platforms for abiotic/biotic stress and yield assessment. Finally, we discuss current challenges and offer perspectives on future phenomics research.

As neuroscience literature suggests, extreme capsule is considered a whiter matter tract. Nevertheless, it is not clear whether extreme capsule itself is an association fiber pathway or only a bottleneck for other association fibers to pass. Via our review, investigating anatomical position, connectivity and cognitive role of the bundles in extreme capsule, and by analyzing data from the dissection, it can be argued that extreme capsule is probably a bottleneck for the passage of uncinated fasciculus (UF) and inferior fronto-occipital fasciculus (IFOF), and these fasciculi are responsible for the respective roles in language processing.

Population bottlenecks can reduce genetic diversity and may lead to inbreeding depression. However, some studies have provided evidence that long lifespans buffer negative genetic effects of bottlenecks. Others have cautioned that longevity might merely mask the effects of genetic drift, which will still affect long-term population viability. We used microsatellite data from actual populations of tuatara (Sphenodon punctatus) and eastern massasaugas (Sistrurus catenatus) as a starting point for simulated population declines to evaluate the performance of bottleneck tests under a range of scenarios. We quantified losses in genetic diversity for each scenario and assessed the power of commonly used tests (i.e., M-ratio, heterozygosity excess, and mode-shift) to detect known bottlenecks in these moderate- to long-lived species. Declines in genetic diversity were greater in bottlenecks simulated for eastern massasaugas, the shorter-lived species, and mode-shift and heterozygosity excess tests were more sensitive to population declines in this species. Conversely, M-ratio tests were more sensitive to bottlenecks simulated in tuatara. Despite dramatic simulated population declines, heterozygosity excess and mode-shift tests often failed to detect bottlenecks in both species, even when large losses in genetic diversity had occurred (both allelic diversity and heterozygosity). While not eliminating type II error, M-ratio tests generally performed best and were most reliable when a critical value (Mc) of 0.68 was used. However, in tuatara simulations, M-ratio tests had high rates of type I error when Mc was calculated assuming θ = 10. Our results suggest that reliance on these tests could lead to misguided species management decisions.

Metabolic engineering is a key technology for cell factories construction by rewiring cellular resources to achieve efficient production of target chemicals. However, the existence of bottlenecks in synthetic pathway can seriously affect production efficiency, which is also one of the core issues for metabolic engineers to solve. Therefore, developing an approach for diagnosing potential metabolic bottlenecks in a faster and simpler manner is of great significance to accelerate cell factories construction. The cell-free reaction system based on cell lysates can transfer metabolic reactions from in vivo to in vitro, providing a flexible access to directly change protein and metabolite variables, thus provides a potential solution for rapid identification of bottlenecks. Here, bottleneck diagnosis of the N-acetylneuraminic acid (NeuAc) biosynthesis pathway in industrially important chassis microorganism Bacillus subtilis was performed using cell-free synthesis system. Specifically, a highly efficient B. subtilis cell-free system for NeuAc de novo synthesis was firstly constructed, which had a 305-fold NeuAc synthesis rate than that in vivo and enabled fast pathway dynamics analysis. Next, through the addition of all potential key intermediates in combination with substrate glucose respectively, it was found that insufficient phosphoenolpyruvate supply was one of the NeuAc pathway bottlenecks. Rational in vivo metabolic engineering of NeuAc-producing B. subtilis was further performed to eliminate the bottleneck. By down-regulating the expression level of pyruvate kinase throughout the growth phase or only in the stationary phase using inhibitory N-terminal coding sequences (NCSs) and growth-dependent regulatory NCSs respectively, the maximal NeuAc titer increased 2.0-fold. Our study provides a rapid method for bottleneck diagnosis, which may help to accelerate the cycle of design, build, test and learn cycle for metabolic engineering.

The research was aimed at determination of genetic variability of Prestice Black-Pied (PC) pig breed (Czech national breed and genetic resource) and to evaluate possible presence of recent bottleneck in this closed small pig population. One hundred and eighty of breeding boars were analysed by eleven tetramer Short Tandem Repeats (STR) panel specifically developed for the genotyping of breeding livestock. Despite the fact that appearance of rare alleles, which may be relatively increased after recent bottleneck, was discovered, the heterozygosity excess was not significant. The PC breed has not undergone recent bottleneck and remained at mutation-drift equilibrium.

The peregrine falcon (Falco peregrinus) has been affected by persecution, pollution, trade and habitat degradation, but it is considered a flagship conservation success story because of successful reintroductions. However, in the UK there were never formal reintroduction programmes for peregrine falcons, and it appears that UK populations – and specifically the Sussex peregrines of the English south coast – recently recovered from a population crash unassisted. To study this, we obtained samples from contemporary populations in southern England, Ireland, continental Europe, domestic-bred peregrine falcons, and from England pre-population crash. Using microsatellite and mtDNA control region data, the genetic diversity and structure, signatures of genetic bottlenecks, and potential origin of the Sussex peregrines was investigated. We found low levels of genetic diversity across all peregrine falcon populations, low but significant genetic differentiation among all populations, and a few private alleles, indicating some level of genetic structure in European peregrines. Although we could not pinpoint the origin of the Sussex peregrines, the data suggests that it is not likely to have originated from escaped domestic birds or from adjacent European populations. The results obtained here parallel other studies on peregrines elsewhere showing low genetic diversity but genetic structure. We conclude that not enough time elapsed for genetic erosion to occur due to the population bottleneck, and that at least for the Sussex peregrines there is no need for genetic conservation by wild-take and subsequent captive breeding programmes as long as current protection measures remain in place.

International audience; The Mascarene Petrel Pseudobulweria aterrima is a critically endangered seabird endemic to Reunion Island, with an extremely small population suffering several threats. Fifteen polymorphic microsatellite loci were isolated from this species to analyse genetic diversity, estimate contemporary effective population size, search for evidence of a population bottleneck and see whether results support the hypothesis that life history traits could preserve allelic diversity in small populations. Results from 22 individuals found grounded as a consequence of light pollution highlight a surprisingly high genetic diversity, an absence of inbreeding, a contemporary effective population size estimated at approximately 1211 individuals and a probable bottleneck around 10 000 generations ago. Additional studies on genetic diversity and structure from a larger number of samples are thus required to evaluate the evolutionary potential of this critically endangered species.

Evolution is a key process by which populations can adapt to novel conditions, but it is not well understood how predictable this process is. Predictability is expected to depend on the ratio of deterministic and stochastic processes that contribute to evolutionary change and this ratio is modulated by the effective population size. Smaller effective populations harbor less genetic diversity and stochastic processes are generally expected to play a larger role, leading to less repeatable evolutionary trajectories. Empirical insight into the relationship between effective population size and repeatability is limited and biased towards asexual unicellular organisms. Here, we used populations of obligately outcrossing Caenorhabditis elegans to test whether fitness increase and selection response were more heterogeneous after a moderate or strong population bottleneck compared to a scenario without bottleneck. Nematodes were exposed to a novel bacterial prey and lower temperature. Population sizes after one week of growth (as a proxy of fitness) were measured before and after 15 generations of evolution. We found that replicates across all (no/moderate/strong bottleneck) treatments evolved higher fitness and no significant difference in average or maximum fitness was found among treatments. Partitioning fitness variance among effects from selection and effects from chance showed that a strong (but not a moderate) bottleneck reduced the relative contribution of selection effects to fitness variation. However, the reduced contribution from selection did not translate to a significant reduction in the repeatability of fitness evolution. Thus, although a strong bottleneck reduced the contribution of deterministic evolutionary change, we found only marginal effects on quantitative measurements of repeatability in evolution. We conclude that the extent to which evolution is predictable may not universally depend on effective population size. IMPACT SUMMARYEvolution is a key process by which populations can adapt to novel conditions. The predictability of evolution is elusive, but the extent to which evolution is predictable is central to our understanding of evolutionary processes and to emergent applications in medicine, agriculture, and conservation. Predictability is expected to be high when evolutionary change is mainly driven by selection, because this leads to repeatable evolutionary trajectories. However, chance effects, e.g. from genetic drift, reduce repeatability. In populations with smaller effective sizes (and thus reduced genetic diversity), chance effects are generally thought to be more prevalent than in large effective populations, suggesting that effective population size influences evolutionary repeatability. Recent theoretical insights cast doubt on the universality of this relationship between evolutionary repeatability and effective population size. Thus far, empirical work testing these theoretical insights is limited to asexual species, leaving a gap in our knowledge. Therefore, we performed an evolutionary experiment using sexually reproducing populations of the nematode Caenorhabditis elegans. Populations either went through a strong or moderate bottleneck or through no bottleneck at all to create populations with different degrees of genetic variation. These populations with different histories (but similar census size) were then exposed to novel conditions. Our results showed that neither average fitness in the novel environment or the repeatability of fitness evolution were significantly lower following a bottleneck. This was despite a reduced contribution from selection and increased contribution from chance to fitness variation in populations that experienced a strong bottleneck. We therefore show that in sexually reproducing species, increased contributions from chance in populations with lower effective sizes do not necessarily affect evolutionary repeatability. This is an important novel insight that is relevant both for our fundamental understanding as well as for evolutionary forecasting applications in agriculture and conservation, which are primarily targeted at sexual species.

Genetic diversity is important for long‐term viability of a population. Low genetic diversity reduces persistence and survival of populations and increases susceptibility to diseases. Comparisons of the neutral markers with functional loci such as immune genes [Toll‐like receptors; TLR] can provide useful insights into evolutionary potential of a species and how the diversity of pathogens and selection pressures on their hosts are directly linked to their environment. In this study, we compare genetic diversity in neutral (eleven microsatellite loci) and adaptive (seven TLR loci) loci to determine genetic variation in a nonmigratory western Himalayan passerine, the black‐throated tit (Aegithalos concinnus), distributed across an elevation gradient with varying degree of pathogen‐mediated selection pressure. We further compare the diversity in TLR loci with a high‐elevation sister species, the white‐throated tit (Aegithalos niveogularis). Our results indicate a lack of population genetic structure in the black‐throated tit and signatures of a past bottleneck. In contrast, we found high diversity in TLR loci and locus‐specific (TLR7) signatures of pathogen‐mediated selection, which was comparable to diversity in the white‐throated tit. Levels of diversity at TLR5 locus corresponded very closely with neutral microsatellite variation. We found evidence of positive selection in TLR1LA, TLR5, and TLR7 loci highlighting the importance in pathogen recognition. Our finding demonstrates that reduction in neutral variation does not necessarily lead to reduction in functional genetic diversity and probably helps in revival of population in a widespread species., Comparisons of allelic richness (size corrected) using eight neutral loci and seven TLR loci (*p

Inheritance of the mitochondrial genome does not follow the rules of conventional Mendelian genetics. The mitochondrial DNA (mtDNA) is present in many copies per cell and is inherited through the maternal germline. In addition, mutations in the mtDNA will give rise to heteroplasmy, the coexistence of different mtDNA variants within a single cell, whose levels can vary considerably between cells, organs or organisms. The inheritance and subsequent accumulation of deleterious variants are the cause of severe progressive mitochondrial disorders and play a role in many other conditions, including aging, cancer and neurodegenerative disorders. Here, we discuss the processes that give rise to cell-to-cell variability in mtDNA composition, focussing on somatic mtDNA segregation and on less conventional sources of heteroplasmy: non-maternal inheritance and mtDNA recombination. Understanding how mtDNA variants and mutations emerge and evolve within an organism is of crucial importance to prevent and cure mitochondrial disease and can potentially impact more common aging-associated conditions.

Historic climate changes had always driven geographical populations of coastal plants to contract and recover dynamically, even die out completely. Species suffering from such bottlenecks usually lose intraspecific genetic diversity, but how do these events influence population subdivision patterns of coastal plants? We investigated this question in the typical coastal plant: mangrove species Aegiceras corniculatum. Inhabiting the intertidal zone of the tropical and subtropical coast of the Indo-West Pacific oceans, its populations are deemed to be greatly shaped by historic sea-level fluctuations. Using dual methods of Sanger and Illumina Solexa sequencing, we found that the 18 sampled populations were structured into two groups, namely, the “Indo-Malayan” group, comprising three subgroups (the northern South China Sea, Gulf of Bengal, and Bali), and the “Pan-Australasia” group, comprising the subgroups of the southern South China Sea and Australasia. Based on simulations using the approximate Bayesian computation method, we inferred that the southern South China Sea subgroup, which penetrates the interior of the “Indo-Malayan” group, originated from the Australasia subgroup, accompanied by a severe bottleneck event, with a spot of gene flow from both the Australasia and “Indo-Malayan” groups. Geographical barriers such as the Sundaland underlie the genetic break between Indian and Pacific Oceans, but the discontinuity between southern and northern South China Sea was originated from genetic drift in the bottleneck event. Hence, we revealed a case evidencing that the bottleneck event promoted population subdivision. This conclusion may be applicable in other taxa beyond coastal plants.

Key message Taxus baccata remnants established recently tend to contribute less to the species’ overall genetic variation than historical populations because they are subjected to a greater impact of the founder effect and genetic isolation. As tree trunk perimeter is a rough indicator of genetic variation in a population, this measure should be considered in conservation programs. Context Genetic variation within Taxus baccata (L.) populations is not associated with the current census size but correlates well with the effective size, suggesting that genetic drift intensity reflects variation in demographic histories. Aims We hypothesize that recently established populations are subjected to greater bottleneck than old remnants. Using the mean trunk perimeter as a surrogate of tree age, we test whether the demographic history and genetic variation are associated with the mean tree age. Methods Using 18 microsatellite markers, we analyze the genetic diversity and demographic history of 11 yew populations in Poland to assess the relationship between the mean trunk perimeter and the inferred genetic parameters. Results Populations reveal significant differences in levels of genetic variation and in the intensity and time of genetic bottleneck. After excluding an apparent outlier, the genetic variation is significantly greater while the bottleneck intensity lower in populations with a greater perimeter. Conclusion Due to continuous species decline and increasing fragmentation, the non-uniform contribution of yew remnants to the overall genetic variation tends to decrease together with the mean tree age. Germplasm collections for the species should take into account tree perimeter as a rough indicator of the genetic variation of a population.

Significance Mitochondria frequently carry different DNA—a state called heteroplasmy. Heteroplasmic mutations can cause mitochondrial diseases and are involved in cancer and aging, but they are also common in healthy people. Here, we study heteroplasmy in 96 multigenerational healthy families. We show that mothers effectively transmit very few mitochondrial DNA to their offspring. Because of this bottleneck, which intensifies with increasing maternal age at childbirth, mutation frequencies can change dramatically between a mother and her child. Thus, a child might inherit a disease-causing mutation at high frequency from an asymptomatic carrier mother and might develop a disease. We also demonstrate that natural selection acts against disease-causing mutations during germline development. Our study has important implications for genetic counseling of mitochondrial diseases., Heteroplasmy—the presence of multiple mitochondrial DNA (mtDNA) haplotypes in an individual—can lead to numerous mitochondrial diseases. The presentation of such diseases depends on the frequency of the heteroplasmic variant in tissues, which, in turn, depends on the dynamics of mtDNA transmissions during germline and somatic development. Thus, understanding and predicting these dynamics between generations and within individuals is medically relevant. Here, we study patterns of heteroplasmy in 2 tissues from each of 345 humans in 96 multigenerational families, each with, at least, 2 siblings (a total of 249 mother–child transmissions). This experimental design has allowed us to estimate the timing of mtDNA mutations, drift, and selection with unprecedented precision. Our results are remarkably concordant between 2 complementary population-genetic approaches. We find evidence for a severe germline bottleneck (7–10 mtDNA segregating units) that occurs independently in different oocyte lineages from the same mother, while somatic bottlenecks are less severe. We demonstrate that divergence between mother and offspring increases with the mother’s age at childbirth, likely due to continued drift of heteroplasmy frequencies in oocytes under meiotic arrest. We show that this period is also accompanied by mutation accumulation leading to more de novo mutations in children born to older mothers. We show that heteroplasmic variants at intermediate frequencies can segregate for many generations in the human population, despite the strong germline bottleneck. We show that selection acts during germline development to keep the frequency of putatively deleterious variants from rising. Our findings have important applications for clinical genetics and genetic counseling.

In many germinating orchids that rely on fungal partners for their nutrition, fungal diversity decreases during development. We document this pattern in the in situ development of Dendrobium moniliforme, where the diversity of cultivable Tulasnella species drops from the early germinating stage (6 Tulasnella species) to that of emergence of the second leaf (2 species), with species discontinuance and no new species appearing. We investigated the functional aspects of this decay by germinating seeds in vitro with the different strains available and observed, over 60 and 120 days, a perfect match between the stages at which Tulasnella spp. occur in situ and the ability to support in vitro development to this stage. The taxonomic bottleneck during germination may result, at least in D. moniliforme, from inability of the fungus to support seedling growth beyond a specific stage. Moreover, the isolated Tulasnella strains that best supported D. moniliforme development did not cluster together phylogenetically. Thus, the interaction between partners, rather than intrinsic fungal traits, may be involved in bottleneck of fungal symbionts during orchid germination.

By the beginning of the twentieth century, many brown bear populations in Europe were on the brink of extinction due to relentless hunting pressure and habitat loss. The situation was critical also in Estonia, where in the 1920s the population went through a severe demographic bottleneck. Thanks to the protective measures implemented in the 1930s, the population started to recover. However, the process has been slow, especially in the western and southern areas of the country. To study the effects of the demographic bottleneck, we analysed 216 brown bear samples from throughout their main range in Estonia. In combination with widely used methods of population genetics, a recently developed spatially explicit analysis (distribution of residual dissimilarity, DResD) was also applied. Three genetic clusters were revealed, of which two were most likely founded by the survivors of the bottleneck. The DResD analysis revealed several contact zones near the Estonian-Russian border, suggesting that the third cluster was influenced by gene flow from the neighbouring population in Russia. The DResD analysis revealed also a male biased movement corridor along the forested south-north axis in the central part of Estonia. In comparison to other European populations, the genetic diversity of the Estonian population is relatively low and is comparable with other populations that have gone through a severe bottleneck. This work has important implications for brown bear conservation and highlights once again the dangers of excessive hunting.

We study the optimal single-step coarse toll design problem for the bottleneck model where the toll level and tolling duration have maximum acceptable upper bounds and the unconstrained optimal solution exceeds the upper bounds. We consider proportional user heterogeneity where users’ values of time and schedule delay vary in fixed proportions. Three classic coarse tolling models are studied, the ADL, Laih and braking models. In the ADL model, untolled users form a mass arrival at the bottleneck following the last tolled user. In the Laih model, there is a separated waiting facility for untolled users to wait until the toll ends. In the braking model, untolled users can choose to defer their arrival at the bottleneck to avoid paying the toll. We find that, in the ADL and the Laih models, the optimal solution chooses the maximum acceptable toll level and tolling duration. The ADL model further requires the tolling period to be started as late as possible to eliminate the queue at the toll ending moment. In the braking model, if the upper bound of the tolling duration is too small, no toll should be charged. Otherwise the optimal solution chooses the maximum acceptable tolling duration and may choose a toll price less than the maximum acceptable level.

Arctic foxes, Vulpes lagopus living on Mednyi Island suffered a drastic decline in population size in the late 1970s due to an outbreak of mange epizootic. This dramatic fall in numbers rendered the subspecies endangered, and the concomitant loss of variability resulted in a population bottleneck. Here, we investigate whether differences in cranial morphology between Mednyi Island Arctic foxes and Bering Island Arctic foxes could be attributed to the severe population bottleneck suffered by the Mednyi population in the 1970s. We used morphometric traits as proxies for genetic data to provide estimates of FST. Results show higher FST estimates for the Mednyi population than for the Bering population, which we interpret as a bottleneck signature. FST results also indicate a pattern of divergence between the two populations consistent with random genetic drift. Bottleneck detection is critical for the interpretation of the demographic history of the endangered Mednyi Island Arctic fox, with consequences for conservation management.

Prochlorococcus are the most abundant photosynthetic organisms in the modern ocean. A massive DNA loss event occurred in their early evolutionary history, leading to highly reduced genomes in nearly all lineages, as well as enhanced efficiency in both nutrient uptake and light absorption. The environmental landscape that shaped this ancient genome reduction, however, remained unknown. Through careful molecular clock analyses, we established that this Prochlorococcus genome reduction occurred during the Neoproterozoic Snowball Earth climate catastrophe. The lethally low temperature and exceedingly dim light during the Snowball Earth event would have inhibited Prochlorococcus growth and proliferation, and caused severe population bottlenecks. These bottlenecks are recorded as an excess of deleterious mutations accumulated across genomic regions and inherited by descendant lineages. Prochlorococcus adaptation to extreme environmental conditions during Snowball Earth intervals can be inferred by tracing the evolutionary paths of genes that encode key metabolic potential. Key metabolic innovation includes modified lipopolysaccharide structure, strengthened peptidoglycan biosynthesis, the replacement of a sophisticated circadian clock with an hourglass-like mechanism that resets daily for dim light adaption and the adoption of ammonia diffusion as an efficient membrane transporter-independent mode of nitrogen acquisition. In this way, the Neoproterozoic Snowball Earth event may have altered the physiological characters of Prochlorococcus , shaping their ecologically vital role as the most abundant primary producers in the modern oceans.

Mitochondrial DNA (mtDNA) is present in multiple copies within an organism. Since these copies are not identical, a single individual carries a heterogeneous population of mtDNAs, a condition known as heteroplasmy. Several factors play a role in the dynamics of the within-organism mtDNA population: among them, genetic bottlenecks, selection, and strictly maternal inheritance are known to shape the levels of heteroplasmy across mtDNAs. In Metazoa, the only evolutionarily stable exception to the strictly maternal inheritance of mitochondria is the doubly uniparental inheritance (DUI), reported in 100+ bivalve species. In DUI species, there are two highly divergent mtDNA lineages, one inherited through oocyte mitochondria (F-type) and the other through sperm mitochondria (M-type). Having both parents contributing to the mtDNA pool of the progeny makes DUI a unique system to study the dynamics of mtDNA populations. Since, in bivalves, the spermatozoon has few mitochondria (4–5), M-type mtDNA faces a tight bottleneck during embryo segregation, one of the narrowest mitochondrial bottlenecks investigated so far. Here, we analyzed the F- and M-type mtDNA variability within individuals of the DUI species Ruditapes philippinarum and investigated for the first time the effects of such a narrow bottleneck affecting mtDNA populations. As a potential consequence of this narrow bottleneck, the M-type mtDNA shows a large variability in different tissues, a condition so pronounced that it leads to genotypes from different tissues of the same individual not to cluster together. We believe that such results may help understanding the effect of low population size on mtDNA bottleneck.

The evolutionary mechanisms by which SARS-CoV-2 viruses adapt to mammalian hosts and, potentially, undergo antigenic evolution depend on the ways genetic variation is generated and selected within and between individual hosts. Using domestic cats as a model, we show that SARS-CoV-2 consensus sequences remain largely unchanged over time within hosts, while dynamic sub-consensus diversity reveals processes of genetic drift and weak purifying selection. We further identify a notable variant at amino acid position 655 in Spike (H655Y), which was previously shown to confer escape from human monoclonal antibodies. This variant arises rapidly and persists at intermediate frequencies in index cats. It also becomes fixed following transmission in two of three pairs. These dynamics suggest this site may be under positive selection in this system and illustrate how a variant can quickly arise and become fixed in parallel across multiple transmission pairs. Transmission of SARS-CoV-2 in cats involved a narrow bottleneck, with new infections founded by fewer than ten viruses. In RNA virus evolution, stochastic processes like narrow transmission bottlenecks and genetic drift typically act to constrain the overall pace of adaptive evolution. Our data suggest that here, positive selection in index cats followed by a narrow transmission bottleneck may have instead accelerated the fixation of S H655Y, a potentially beneficial SARS-CoV-2 variant. Overall, our study suggests species- and context-specific adaptations are likely to continue to emerge. This underscores the importance of continued genomic surveillance for new SARS-CoV-2 variants as well as heightened scrutiny for signatures of SARS-CoV-2 positive selection in humans and mammalian model systems., Author summary Through ongoing human adaptation, spill-back events from other animal intermediates, or with the distribution of vaccines and therapeutics, the landscape of SARS-CoV-2 genetic variation is certain to change. The evolutionary mechanisms by which SARS-CoV-2 will continue to adapt to mammalian hosts depend on genetic variation generated within and between hosts. Here, using domestic cats as a model, we show that within-host SARS-CoV-2 genetic variation is predominantly influenced by genetic drift and purifying selection. Transmission of SARS-CoV-2 between hosts is defined by a narrow transmission bottleneck, involving 2–5 viruses. We further identify a notable variant at amino acid position 655 in Spike (H655Y), which arises rapidly and is transmitted in cats. Spike H655Y has been previously shown to confer escape from human monoclonal antibodies and is currently found in over 1,000 human sequences. Overall, our study suggests species- and context-specific adaptations are likely to continue to emerge, underscoring the importance of continued genomic surveillance in humans and non-human mammalian hosts.

SUMMARYThe demographic history is a foundation of human evolutionary studies. However, the ancient demographic history during the Mid-Pleistocene is poorly investigated while it is essential for understanding the early origin of humankind. Here we present the fast infinitesimal time coalescent (FitCoal) process, which allows the analytical calculation of the composite likelihood of a site frequency spectrum and provides the precise inference of demographic history. We apply it to analyze 3,154 present-day human genomic sequences. We find that African populations have passed through a population super bottleneck, a small effective size of approximately 1,280 breeding individuals between 930 and 813 thousand years ago. Further analyses confirm the existence of the super bottleneck on non-African populations although it cannot be directly inferred. This observation, together with simulation results, indicates that confounding factors, such as population structure and selection, are unlikely to affect the inference of the super bottleneck. The time interval of the super bottleneck coincides with a gap in the human fossil record in Africa and possibly marks the origin ofHomo heidelbergensis. Our results provide new insights into human evolution during the Mid-Pleistocene.

Background Current patterns of population genetic variation may have been shaped by long-term evolutionary history and contemporary demographic processes. Understanding the underlying mechanisms that yield those patterns is crucial for informed conservation of endangered species. The critically endangered white-headed langur, Trachypithecus leucocephalus, is endemic to a narrow range in southwest China. This species shows very low genetic diversity in its 2 main relict populations, Fusui and Chongzuo. Whether this has been caused by a short evolutionary history or recent population declines is unknown. Therefore, we investigated the contributions of historical and recent population demographic changes to population genetic diversity by using 15 nuclear microsatellite markers and mitochondrial DNA (mtDNA) control region sequences. Results Using genetic data from 214 individuals we found a total of 9 mtDNA haplotypes in the Fusui population but only 1 haplotype in the Chongzuo population, and we found an overall low genetic diversity (haplotype and nucleotide diversities: h = 0.486 ± 0.036; π = 0.0028 ± 0.0003). The demographic history inferred from mtDNA and microsatellite markers revealed no evidence for historical population size fluctuations or recent population bottlenecks. Simulations of possible population divergence histories inferred by DIYABC analysis supported a recent divergence of the Chongzuo population from the Fusui population and no population bottlenecks. Conclusions Despite severe population declines caused by anthropogenic activities in the last century, the low genetic diversity of the extant white-headed langur populations is most likely primarily due to the species’ shallow evolutionary history and to a recent, local population founder event. Electronic supplementary material The online version of this article (10.1186/s12862-019-1451-y) contains supplementary material, which is available to authorized users.

An early analysis of SARS-CoV-2 deep-sequencing data that combined epidemiological and genetic data to characterize the transmission dynamics of the virus in and beyond Austria concluded that the size of the virus’s transmission bottleneck was large – on the order of 1000 virions. We performed new computational analyses using these deep-sequenced samples from Austria. Our analyses included characterization of transmission bottleneck sizes across a range of variant calling thresholds and examination of patterns of shared low-frequency variants between transmission pairs in cases where de novo genetic variation was present in the recipient. From these analyses, among others, we found that SARS-CoV-2 transmission bottlenecks are instead likely to be very tight, on the order of 1-3 virions. These findings have important consequences for understanding how SARS-CoV-2 evolves between hosts and the processes shaping genetic variation observed at the population level.

Genetic bottlenecks can limit the success of populations colonizing new ranges. However, successful colonizations can occur despite bottlenecks, a phenomenon known as the genetic paradox of invasion. Eusocial Hymenoptera such as bumblebees (Bombusspp.) should be particularly vulnerable to genetic bottlenecks, since homozygosity at the sex-determining locus leads to costly diploid male production (DMP). The Tree Bumblebee (Bombus hypnorum) has rapidly colonized the UK since 2001 and has been highlighted as exemplifying the genetic paradox of invasion. Using microsatellite genotyping, combined with the first genetic estimates of DMP in UKB. hypnorum, we tested two alternative genetic hypotheses (‘bottleneck’ and ‘gene flow’ hypotheses) forB. hypnorum's colonization of the UK. We found that the UK population has not undergone a recent severe genetic bottleneck and exhibits levels of genetic diversity falling between those of widespread and range-restrictedBombusspecies. Diploid males occurred in 15.4% of reared colonies, leading to an estimate of 21.5 alleles at the sex-determining locus. Overall, the findings show that this population is not bottlenecked, instead suggesting that it is experiencing continued gene flow from the continental European source population with only moderate loss of genetic diversity, and does not exemplify the genetic paradox of invasion.

This paper chooses car travel and bus travel as the research objects, establishes a dual-mode equilibrium model based on the bottleneck model, and compares the travel characteristics of the no-toll and fine-toll schemes. We find that the fine-toll scheme can eliminate the queuing time at the bottleneck, but it also increases the congestion risk cost of bus travel. In order to eliminate the queuing time at the bottleneck and reduce the congestion risk cost of bus travel without increasing the car travel cost and bus travel cost, we propose an optimization scheme of fine toll and bus departure quantity and analyze its travel characteristics theoretically. Through the numerical example, we calculate and analyze the equilibrium results of no-toll scheme, fine-toll scheme, and optimization scheme of fine toll and bus departure quantity. The results indicate that the optimization scheme of fine toll and bus departure quantity can help travelers to choose a reasonable travel mode and travel time to travel in the rush hour.

A pivotal debate on biodiversity conservation is whether the scarce budgets must be invested in critically endangered taxa or in those with higher chances to survive due to larger population sizes. Addressing the fate of extremely bottlenecked taxa is an ideal way to test this idea, but empirical cases are surprisingly limited. The reintroduction of the extinct-in-the-wild Alagoas curassow (Pauxi mitu) by Brazilian scientists in September 2019 added to the two other known cases of survival to bottlenecks of only two or three individuals. We exploit the reasons why this species has survived, and we report how investments to rescue the Alagoas curassow resulted in the protection of many other taxa, suggesting that in the face of the dramatic number of extinctions expected for the Anthropocene, integration must prevail over a choice.

Viral populations undergo a repeated cycle of within-host growth followed by transmission. Viral evolution is affected by each stage of this cycle. The number of viral particles transmitted from one host to another, known as the transmission bottleneck, is an important factor in determining how the evolutionary dynamics of the population play out, restricting the extent to which the evolved diversity of the population can be passed from one host to another. Previous study of viral sequence data has suggested that the transmission bottleneck size for influenza A transmission between human hosts is small. Reevaluating these data using a novel and improved method, we largely confirm this result, albeit that we infer a slightly higher bottleneck size in some cases, of between 1 and 13 virions. While a tight bottleneck operates in human influenza transmission, it is not extreme in nature; some diversity can be meaningfully retained between hosts., The transmission bottleneck is defined as the number of viral particles that transmit from one host to establish an infection in another. Genome sequence data have been used to evaluate the size of the transmission bottleneck between humans infected with the influenza virus; however, the methods used to make these estimates have some limitations. Specifically, viral allele frequencies, which form the basis of many calculations, may not fully capture a process which involves the transmission of entire viral genomes. Here, we set out a novel approach for inferring viral transmission bottlenecks; our method combines an algorithm for haplotype reconstruction with maximum likelihood methods for bottleneck inference. This approach allows for rapid calculation and performs well when applied to data from simulated transmission events; errors in the haplotype reconstruction step did not adversely affect inferences of the population bottleneck. Applied to data from a previous household transmission study of influenza A infection, we confirm the result that the majority of transmission events involve a small number of viruses, albeit with slightly looser bottlenecks being inferred, with between 1 and 13 particles transmitted in the majority of cases. While influenza A transmission involves a tight population bottleneck, the bottleneck is not so tight as to universally prevent the transmission of within-host viral diversity. IMPORTANCE Viral populations undergo a repeated cycle of within-host growth followed by transmission. Viral evolution is affected by each stage of this cycle. The number of viral particles transmitted from one host to another, known as the transmission bottleneck, is an important factor in determining how the evolutionary dynamics of the population play out, restricting the extent to which the evolved diversity of the population can be passed from one host to another. Previous study of viral sequence data has suggested that the transmission bottleneck size for influenza A transmission between human hosts is small. Reevaluating these data using a novel and improved method, we largely confirm this result, albeit that we infer a slightly higher bottleneck size in some cases, of between 1 and 13 virions. While a tight bottleneck operates in human influenza transmission, it is not extreme in nature; some diversity can be meaningfully retained between hosts.

The present study was performed to reveal genetic diversity and bottleneck of Turkish and Algerian autochthonous sheep breeds using fifteen microsatellites marker recommended by FAO (2011). Animal material for the study was consisted of 180 head sheep raised in different location in Turkey and Algeria. A total of 349 alleles were detected from fifteen markers studied. The mean number of alleles (23.26), observed heterozygosity (0.76) and polymorphic information content (0.89) findings indicated that noticeable genetic variability in sheep population studied. Fourteen out of the sixteen microsatellite markers studied had a positive FIS value. The mean value of FIS was 0.061.The infinite allele model (IAM), two-phase mutation model (TPM) and stepwise mutation model (SMM) in the Bottleneck software were used to check genetic bottleneck. The Lshaped curve obtained from analyze indicates absence of bottleneck in studied sheep population raised in Turkey and Algeria. Consequently, it can be said that these results will help to develop conservation and breeding strategies for the sheep population.

Giant clams are an important ecological component of coral reefs in the Red Sea, as they enhance the reef’s productivity and provide habitat that can increase diversity. Three species of giant clams, namely Tridacna maxima, T. squamosa, and T. squamosina have been described within the Red Sea. However, due to its scarcity, information about the distribution and ecology of T. squamosina in the Saudi Arabian Red Sea is still lacking. This study used DNA barcoding to confirm the identity of the rare T. squamosina in the Farasan Banks. Six mtCOI fragments (500 bp) of T. squamosina were successfully amplified using the SQUA-primers for the first time. We used our data along with 18 reference sequences (16S) from the online database to assess the genetic diversity and population structure of T. squamosina. Low genetic diversity among the T. squamosina populations inferred from the 16S sequences implies a recent bottleneck for this species, which is supported by their historically higher diversity based on the coalescent-based estimator. Given the small population abundance and limited genetic variation of T. squamosina, it may warrant immediate local protections such as biobanking and fertility preservation programs as well as effective integrated coastal zone management plans.

If they undergo new mutations at each replication cycle, why are RNA viral genomes so fragile, with most mutations being either strongly deleterious or lethal? Here we provide theoretical and numerical evidence for the hypothesis that genetic fragility is partly an evolutionary response to the multiple population bottlenecks experienced by viral populations at various stages of their life cycles. Modelling within-host viral populations as multi-type branching processes, we show that mutational fragility lowers the rate at which Muller’s ratchet clicks and increases the survival probability through multiple bottlenecks. In the context of a susceptible-exposed-infectious-recovered epidemiological model, we find that the attack rate of fragile viral strains can exceed that of more robust strains, particularly at low infectivities and high mutation rates. Our findings highlight the importance of demographic events such as transmission bottlenecks in shaping the genetic architecture of viral pathogens., Funding for this work was provided by the Alexander von Humboldt Foundation in the framework of the Sofja Kovalevskaja Award endowed by the German Federal Ministry of Education and Research to M.S. Work in València was supported by Spain Agencia Estatal de Investigación - FEDER grant PID2019-103998GB-I00 and Generalitat Valenciana grant PROMETEO2019/012 to S.F.E.

Many positive-sense RNA viruses, especially those infecting plants, are known to experience stringent, stochastic population bottlenecks inside the cells they invade, but exactly how and why these populations become bottlenecked are unclear. A model proposed ten years ago advocates that such bottlenecks are evolutionarily favored because they cause the isolation of individual viral variants in separate cells. Such isolation in turn allows the viral variants to manifest the phenotypic differences they encode. Recently published observations lend mechanistic support to this model and prompt us to refine the model with novel molecular details. The refined model, designated Bottleneck, Isolate, Amplify, Select (BIAS), postulates that these viruses impose population bottlenecks on themselves by encoding bottleneck-enforcing proteins (BNEPs) that function in a concentration-dependent manner. In cells simultaneously invaded by numerous virions of the same virus, BNEPs reach the bottleneck-ready concentration sufficiently early to arrest nearly all internalized viral genomes. As a result, very few (as few as one) viral genomes stochastically escape to initiate reproduction. Repetition of this process in successively infected cells isolates viral genomes with different mutations in separate cells. This isolation prevents mutant viruses encoding defective viral proteins from hitchhiking on sister genome-encoded products, leading to the swift purging of such mutants. Importantly, genome isolation also ensures viral genomes harboring beneficial mutations accrue the cognate benefit exclusively to themselves, leading to the fixation of such beneficial mutations. Further interrogation of the BIAS hypothesis promises to deepen our understanding of virus evolution and inspire new solutions to virus disease mitigation.

As neuroscience literature suggests, the extreme capsule is considered a white matter tract. Nevertheless, it is not clear whether the extreme capsule itself is an association fiber pathway or only a bottleneck for other association fibers to pass. Via our review, investigating anatomical position, connectivity, and cognitive role of the extreme capsule, and by analyzing data from the dissection, it can be argued that extreme capsule is probably a bottleneck for the passage of uncinated fasciculus (UF) and inferior fronto occipital fasciculus (IFOF), and its different role of language processing is due to various tracts that pass it through.

Mixed infections provide opportunities for viruses to increase genetic diversity by facilitating genomic reassortment or recombination, and they may lead to the emergence of new virus species. Mixed infections of two economically important orthotospoviruses, Tomato spotted wilt orthotospovirus (TSWV) and Impatiens necrotic spot orthotospovirus (INSV), were found in recent years, but no natural reassortants between INSV and TSWV were ever reported. The goal of this study was to establish how vector preferences and the ability to transmit INSV and TSWV influence transmission and establishment of mixed infections. Our results demonstrate that thrips prefer to oviposit on TSWV and INSV mixed-infected plants over singly infected or healthy plants, providing young nymphs with the opportunity to acquire both viruses. Conversely, we observed that thrips served as a bottleneck during transmission and favored transmission of one of the two viruses over the second one, or over transmission of both viruses simultaneously. This constraint was relaxed in plants, when transmission of TSWV and INSV occurred sequentially, demonstrating that plants serve as orthotospovirus permissive hosts, while thrips serve as a bottleneck. Viral fitness, as measured by virus replication, transmission, and competition with other viral strains, is not well studied in mixed infection. Our study looks at the success of transmission during mixed infection of orthotopoviruses, enhancing the understanding of orthotospovirus epidemiology and evolution.

RNA viruses exist as populations of closely related genomes, characterized by a high diversity of low-frequency variants. As viral genomes from one population disperse to establish new sites of replication, the fate of these low-frequency variants depends to a large extent on the size of the founding population. Focusing on foot-and-mouth disease virus (FMDV) we conjecture that variants are more likely to be transmitted through wide bottlenecks, but more likely to approach fixation in new populations following narrow bottlenecks; therefore, the longer-term rate of accumulation of ‘nearly neutral’ variants at high frequencies is likely to be inversely related to the bottleneck size. We examine this conjecturein vivoby estimating bottleneck sizes relating ‘parent’ and ‘daughter’ populations observed at different scales ranging from within host to between host (within the same herd, and in different herds) using a previously established method. Within hosts, we find bottleneck sizes to range from 5 to 20 viral genomes between populations transmitted from the pharynx to the serum, and from 4 to 54 between serum and lesion populations. Between hosts, we find bottleneck sizes to range from 2 to 39, suggesting inter-host bottlenecks are of a similar size to intra-host bottlenecks. We establish a statistically significant negative relationship between the probability of genomic consensus level change and bottleneck size, and present a simple sampling model that captures this empirical relationship. We also present a novelin vitroexperiment to investigate the impact of bottleneck size on the frequency of mutations within FMDV populations, demonstrate that variant frequency in a population increases more rapidly during small population passages, and provide evidence for positive selection during the passage of large populations.

Heteroplasmy is the presence of variable mitochondrial DNA (mtDNA) within the same individual. The dynamics of heteroplasmy allele frequency among tissues of the human body is not well understood. Here, we measured allele frequency at heteroplasmic sites in two to eight hairs from each of 11 humans using next-generation sequencing. We observed a high variance in heteroplasmic allele frequency among separate hairs from the same individual—much higher than that for blood and cheek tissues. Our population genetic modelling estimated the somatic bottleneck during embryonic follicle development of separate hairs to be only 11.06 (95% confidence interval 0.6–34.0) mtDNA segregating units. This bottleneck is much more drastic than somatic bottlenecks for blood and cheek tissues (136 and 458 units, respectively), as well as more drastic than, or comparable to, the germline bottleneck (equal to 25–32 or 7–10 units, depending on the study). We demonstrated that hair undergoes additional genetic drift before and after the divergence of mtDNA lineages of individual hair follicles. Additionally, we showed a positive correlation between donor's age and variance in heteroplasmy allele frequency in hair. These findings have important implications for forensics and for our understanding of mtDNA dynamics in the human body. This article is part of the theme issue ‘Linking the mitochondrial genotype to phenotype: a complex endeavour’.