Maintenance of Intraspecific Diversity in Response to Species Competition and Nutrient Fluctuations

Intraspecific diversity is a substantial part of biodiversity, yet little is known about its maintenance. Understanding mechanisms of intraspecific diversity shifts provides realistic detail about how phytoplankton communities evolve to new environmental conditions, a process especially important in times of climate change. Here, we aimed to identify factors that maintain genotype diversity and link the observed diversity change to measured phytoplankton morpho-functional traits Vmax and cell size of the species and genotypes. In an experimental setup, the two phytoplankton species Emiliania huxleyi and Chaetoceros affinis, each consisting of nine genotypes, were cultivated separately and together under different fluctuation and nutrient regimes. Their genotype composition was assessed after 49 and 91 days, and Shannon’s diversity index was calculated on the genotype level. We found that a higher intraspecific diversity can be maintained in the presence of a competitor, provided it has a substantial proportion to total biovolume. Both fluctuation and nutrient regime showed species-specific effects and especially structured genotype sorting of C. affinis. While we could relate species sorting with the measured traits, genotype diversity shifts could only be partly explained. The observed context dependency of genotype maintenance suggests that the evolutionary potential could be better understood, if studied in more natural settings including fluctuations and competition.

Treatment of wound infections in a mouse model using Zn2+-releasing phage bound to gold nanorods

Infections caused by drug-resistant bacteria, particularly gram-negative organisms, are increasingly difficult to treat using antibiotics. A potential alternative is phage therapy, in which phages infect and lyse the bacterial host. However, phage therapy poses serious drawbacks and safety concerns, such as the risk of genetic transduction of antibiotic resistance genes, inconsistent pharmacokinetics, and unknown evolutionary potential. In contrast, metallic nanoparticles possess precise, tunable properties, including efficient conversion of electronic excitation into heat. In this work, we demonstrate that engineered phage-nanomaterial conjugates that target the gram-negative pathogen P. aeruginosa, are highly effective as a treatment of infected wounds in mice. Photothermal heating, performed as a single treatment (15 min) or as two treatments on consecutive days, rapidly reduced the bacterial load and released Zn2+ to promote wound healing. The phage-nanomaterial treatment was significantly more effective than systemic fluoroquinolone antibiotics in reducing both bacterial load and wound size, and was notably effective against a P. aeruginosa strain resistant to polymyxins, a last-line antibiotic therapy. Unlike these antibiotics, the phage-nanomaterial showed no detectable toxicity or systemic effects in mice, consistent with the short duration and localized nature of phage-nanomaterial treatment. Our results demonstrate that phage therapy controlled by inorganic nanomaterials can be a safe and effective antimicrobial strategy in vivo.

Chikungunya virus: Molecular epidemiology of nonstructural proteins in Pakistan

Chikungunya virus (CHIKV) is considered a public health problem due to its rapid spread and high morbidity. In 2016–2017 an outbreak of CHIKV was occurred in Pakistan but the data regarding the genomic diversity of CHIKV was not reported. Hence, the current study aimed to determine the genetic diversity of CHIKVs in Pakistan. A cross sectional study was carried out using sera of infected CHIKV patients (n = 1549) during the outbreak in Pakistan (2016–2018). Nucleotide sequencing of non-structural genes of CHIKV from eight isolates were performed followed by phylogenetic analysis using Bayesian method. Phylogenetic analysis suggested that the Pakistani CHIKV strains belonged to Indian Ocean Lineage (IOL) of genotype ECSA and C1.3a clade. Furthermore, the Pakistani isolates showed several key mutations (nsP2-H130Y, nsP2-E145D, nsP4-S55N and nsP4- R85G) corresponding to mutations reported in 2016 Indian strains of CHIKV. The molecular analysis revealed high evolutionary potential of CHIKV strains as well as better understanding of enhanced virulence and pathogenesis of this outbreak. The study highlights the need to continue surveillance in order to understand viral diversity over time and to devise preventive measures to limit diseases transmission in the region.

Conservation genetics as a management tool: The five best-supported paradigms to assist the management of threatened species

About 50 y ago, Crow and Kimura [An Introduction to Population Genetics Theory (1970)] and Ohta and Kimura [Genet. Res. 22, 201–204 (1973)] laid the foundations of conservation genetics by predicting the relationship between population size and genetic marker diversity. This work sparked an enormous research effort investigating the importance of population dynamics, in particular small population size, for population mean performance, population viability, and evolutionary potential. In light of a recent perspective [J. C. Teixeira, C. D. Huber, Proc. Natl. Acad. Sci. U.S.A. 118, 10 (2021)] that challenges some fundamental assumptions in conservation genetics, it is timely to summarize what the field has achieved, what robust patterns have emerged, and worthwhile future research directions. We consider theory and methodological breakthroughs that have helped management, and we outline some fundamental and applied challenges for conservation genetics.

Seed sourcing for climate-resilient grasslands: the role of seed source diversity during early restoration establishment

Restoration often advocates for the use of local seed in restoration, however increasingly new strategies have been proposed to incorporate diverse sources to maintain evolutionary potential within seed mixes. Increasing seed sources per species within a seed mix should increase genetic variation, however, few empirical studies have evaluated how seed source diversity impacts plant community composition following restoration. Thus, the goal of this research was to compare the use of single or multi-source seed mix treatments to plant community diversity following restoration. Using 14 species commonly applied in grassland restoration, we examined plant community diversity following restoration comparing seed mixes with either one or five sources per species across two restoration sites in Minnesota and South Dakota, United States. Following seeding, species establishment and abundance were recorded to calculate plant diversity for each seed mix treatment. There were no major effects of seed mix treatment on community emergence and diversity observed, with the majority of plant establishment reflecting non-seeded species. However, site-specific differences were observed. Heterogeneous land-use history associated with the Minnesota site likely contributed to differences across the restoration treatments. In contrast, community diversity at the South Dakota site was homogeneous across seed mix treatments with changes in plant community influenced solely by early season species establishment. This suggests land-use history irrespective of seed mix treatment influences establishment and persistence, particularly in the first year following restoration. Future monitoring across seasons will be needed to evaluate if community diversity changes in response to seed mix treatment.

HIV-1 Vif gained breadth in APOBEC3G specificity after cross-species transmission of its precursors

APOBEC3G (A3G) is a host-encoded cytidine deaminase that potently restricts retroviruses, such as HIV-1, and depends on its ability to package into virions. As a consequence of this, HIV-1 protein Vif has evolved to antagonize human A3G by targeting it for ubiquitination and subsequent degradation. There is an ancient arms-race between Vif and A3G highlighted by amino acids 128 and 130 in A3G that have evolved under positive selection due to Vif-mediated selective pressure in Old World primates. Nonetheless, not all possible amino acid combinations at these sites have been sampled by nature and it is not clear the evolutionary potential of species to resist Vif antagonism. To explore the evolutionary space of positively selected sites in the Vif-binding region of A3G, we designed a combinatorial mutagenesis screen to introduce all 20 amino acids at sites 128 and 130. Our screen uncovered mutants of A3G with several interesting phenotypes, including loss of antiviral activity and resistance of Vif antagonism. However, HIV-1 Vif exhibited remarkable flexibility in antagonizing A3G 128 and 130 mutants, which significantly reduces viable Vif resistance strategies for hominid primates. Importantly, we find that broadened Vif specificity was conferred through Loop 5 adaptations that were required for cross-species adaptation from Old World monkey A3G to hominid A3G. Our evidence suggests that Vif adaptation to novel A3G interfaces during cross-species transmission may train Vif towards broadened specificity that can further facilitate cross-species transmissions and raise the barrier to host resistance. Importance APOBEC3G (A3G) is an antiviral protein that potently restricts retroviruses like HIV. In turn, the HIV-1 protein Vif has evolved to antagonize A3G through degradation. Two rapidly evolving sites in A3G confer resistance to unadapted Vif and act as a barrier to cross-species transmission of retroviruses. We recently identified a single amino acid mutation in an SIV Vif that contributed to the cross-species origins of SIV infecting chimpanzee, and ultimately the HIV-1 pandemic. This mutation broadened specificity of this Vif to both antagonize the A3G of its host while simultaneously overcoming the A3G barrier in the great apes. In this work, we explore the evolutionary space of human A3G at these rapidly evolving sites to understand if the broadened Vif specificity gained during cross-species transmission confers an advantage to HIV-1 Vif in its host-virus arms race with A3G.

Turtle Nest-Site Choice, Anthropogenic Challenges, and Evolutionary Potential for Adaptation

Oviparous animals, such as turtles, lay eggs whose success or demise depends on environmental conditions that influence offspring phenotype (morphology, physiology, and in many reptiles, also sex determination), growth, and survival, while in the nest and post-hatching. Consequently, because turtles display little parental care, maternal provisioning of the eggs and female nesting behavior are under strong selection. But the consequences of when and where nests are laid are affected by anthropogenic habitat disturbances that alter suitable nesting areas, expose eggs to contaminants in the wild, and modify the thermal and hydric environment experienced by developing embryos, thus impacting hatchling survival and the sexual fate of taxa with temperature-dependent sex determination (TSD) and genotypic sex determination (GSD). Indeed, global and local environmental change influences air, water, and soil temperature and moisture, which impact basking behavior, egg development, and conditions within the nest, potentially rendering current nesting strategies maladaptive as offspring mortality increases and TSD sex ratios become drastically skewed. Endocrine disruptors can sex reverse TSD and GSD embryos alike. Adapting to these challenges depends on genetic variation, and little to no heritability has been detected for nest-site behavior. However, modest heritability in threshold temperature (above and below which females or males develop in TSD taxa, respectively) exists in the wild, as well as interpopulation differences in the reaction norm of sex ratio to temperature, and potentially also in the expression of gene regulators of sexual development. If this variation reflects additive genetic components, some adaptation might be expected, provided that the pace of environmental change does not exceed the rate of evolution. Research remains urgently needed to fill current gaps in our understanding of the ecology and evolution of nest-site choice and its adaptive potential, integrating across multiple levels of organization.

Micro- and macroevolutionary change of colony-level traits in bryozoans

The evolution of trait variation among populations of animals is difficult to study due to the many overlapping genetic and environmental influences that control phenotypic expression. In a group of animals, bryozoans, it is possible to isolate genetic contributions to phenotypic variation, due to the modular nature of bryozoan colonies. Each bryozoan colony represents a snapshot of the phenotypes that correspond to a single genotype, which can be summarized as a phenotypic distribution. We test whether these phenotypic distributions are heritable across generations of colonies in two sister species of the bryozoan Stylopoma, grown and bred in a common garden breeding experiment. We find that components of phenotypic distributions, specifically median trait values of colony members, are heritable between generations of colonies. Furthermore, this heredity has macroevolutionary importance because it correlates with the morphological distance between these two species. Because parts of phenotypic distributions are heritable, and this heritability corresponds to evolutionary divergence between species, we infer that these distributions have the potential to evolve. The evolutionary potential of these phenotypic distributions may underpin the emergence of colony-level traits, like division of labor in colonies.

Intra-genomic variation in symbiotic dinoflagellates: Recent divergence or natural hybridization?

<p>The perpetuity of coral reefs will ultimately depend on the ability of corals to adapt to changing conditions. Inter-specific hybridization can provide the raw genetic material necessary for adaptation, and stimulate macro-evolutionary leaps during periods of environmental upheaval. Though well-documented in corals, hybridization has yet to be identified in their dinoflagellate symbionts (genus Symbiodinium), despite growing evidence of sexual reproduction in this genus. The integral roles that these symbiotic algae play in coral productivity, reef accretion and ‘coral bleaching’ emphasize the need to better understand their short-term evolutionary potential. In this thesis, I develop new molecular and statistical methodology, and combine lab- and field-based analysis to explore the potential for hybridization between divergent Symbiodinium taxa. To screen for putative Symbiodinium hybrids, intra-genomic variation was examined within individual symbionts isolated from the reef-building coral Pocillopora damicornis at Lord Howe Island (Australia). A nested quantitative PCR (qPCR) assay was developed to quantify polymorphic internal transcribed spacer 2 (ITS2) sequences within the genome of each symbiont cell. Three genetically distinct Symbiodinium populations were detected co-existing within the symbiont consortium of P. damicornis. Mixed populations of ‘pure’ Symbiodinium types C100 and C109 coexisted with a population of cells hosting co-dominant C100 and C109 ITS2 repeats. Genetically heterogeneous Symbiodinium cells were more common than homogeneous symbionts in four of the six colonies analysed, with a maximum proportional abundance of 89%. Morphological, functional and ecological attributes of heterogeneous Symbiodinium cells were characterized to assess their candidacy as putative hybrids. The proportional abundance of genetically heterogeneous symbionts was spatially and temporally conserved within colonies, indicating a lack of competition between Symbiodinium populations. However, this abundance ratio varied considerably between colonies separated by metres to tens of metres, and to a greater extent between sites isolated by hundreds to thousands of metres. The local thermal maximum emerged as a significant predictor of the proportional abundance of genetically heterogeneous Symbiodinium cells, suggesting that the distribution of these ‘putative hybrids’ is influenced by a reduced affinity for thermal stress. Genetically heterogeneous Symbiodinium cells were around 50% larger (by volume) than homogeneous cells, occupied tissue of the coral host at reduced densities, and showed relatively poor light-harvesting efficiency. Colonies hosting a higher proportion of these symbionts suffered a reduction in overall photosynthetic performance (maximum gross photosynthesis normalised to respiration; P:R) at the ambient temperature of 25 °C. This disparity was maintained when the temperature was elevated to simulate the maximum experienced within the LHI lagoon (29 °C). Under these stressful conditions, colonies dominated by putative Symbiodinium hybrids were only marginally capable of net oxygen production. The influence of putative Symbiodinium hybrids on the growth and survival of P. damicornis was tested by reciprocally transplanting coral colonies between reef sites featuring distinct temperature regimes. Neither calcification nor mortality was influenced by the proportional abundance of genetically heterogeneous cells in the symbiont consortium. This uncoupling of symbiont performance and host fitness may be explained by stochastic events such as predation and disease, which substantially increase variation in growth and mortality in field experiments. Alternatively, it may represent some unknown benefit associated with hosting hybrid symbionts, belying their relatively poor photosynthetic performance, and explaining the widespread abundance of these heterogeneous Symbiodinium cells on the Lord Howe Island reef. Our inability to maintain many clade C Symbiodinium types in culture prevents direct observations of hybridization between C100 and C109. Unequivocal evidence of this phenomenon will therefore likely remain elusive until high-resolution, single-copy nuclear markers can be developed, since the incomplete displacement of ancestral polymorphisms can leave a similar genomic signature to that of hybridization. However, this study serves to provide an initial proof-of-principle for hybridization between divergent Symbiodinium taxa. In doing so, it highlights the need to better understand the evolutionary processes underpinning coral- and symbiont-adaptation in a changing climate.</p>

Intra-genomic variation in symbiotic dinoflagellates: Recent divergence or natural hybridization?

<p>The perpetuity of coral reefs will ultimately depend on the ability of corals to adapt to changing conditions. Inter-specific hybridization can provide the raw genetic material necessary for adaptation, and stimulate macro-evolutionary leaps during periods of environmental upheaval. Though well-documented in corals, hybridization has yet to be identified in their dinoflagellate symbionts (genus Symbiodinium), despite growing evidence of sexual reproduction in this genus. The integral roles that these symbiotic algae play in coral productivity, reef accretion and ‘coral bleaching’ emphasize the need to better understand their short-term evolutionary potential. In this thesis, I develop new molecular and statistical methodology, and combine lab- and field-based analysis to explore the potential for hybridization between divergent Symbiodinium taxa. To screen for putative Symbiodinium hybrids, intra-genomic variation was examined within individual symbionts isolated from the reef-building coral Pocillopora damicornis at Lord Howe Island (Australia). A nested quantitative PCR (qPCR) assay was developed to quantify polymorphic internal transcribed spacer 2 (ITS2) sequences within the genome of each symbiont cell. Three genetically distinct Symbiodinium populations were detected co-existing within the symbiont consortium of P. damicornis. Mixed populations of ‘pure’ Symbiodinium types C100 and C109 coexisted with a population of cells hosting co-dominant C100 and C109 ITS2 repeats. Genetically heterogeneous Symbiodinium cells were more common than homogeneous symbionts in four of the six colonies analysed, with a maximum proportional abundance of 89%. Morphological, functional and ecological attributes of heterogeneous Symbiodinium cells were characterized to assess their candidacy as putative hybrids. The proportional abundance of genetically heterogeneous symbionts was spatially and temporally conserved within colonies, indicating a lack of competition between Symbiodinium populations. However, this abundance ratio varied considerably between colonies separated by metres to tens of metres, and to a greater extent between sites isolated by hundreds to thousands of metres. The local thermal maximum emerged as a significant predictor of the proportional abundance of genetically heterogeneous Symbiodinium cells, suggesting that the distribution of these ‘putative hybrids’ is influenced by a reduced affinity for thermal stress. Genetically heterogeneous Symbiodinium cells were around 50% larger (by volume) than homogeneous cells, occupied tissue of the coral host at reduced densities, and showed relatively poor light-harvesting efficiency. Colonies hosting a higher proportion of these symbionts suffered a reduction in overall photosynthetic performance (maximum gross photosynthesis normalised to respiration; P:R) at the ambient temperature of 25 °C. This disparity was maintained when the temperature was elevated to simulate the maximum experienced within the LHI lagoon (29 °C). Under these stressful conditions, colonies dominated by putative Symbiodinium hybrids were only marginally capable of net oxygen production. The influence of putative Symbiodinium hybrids on the growth and survival of P. damicornis was tested by reciprocally transplanting coral colonies between reef sites featuring distinct temperature regimes. Neither calcification nor mortality was influenced by the proportional abundance of genetically heterogeneous cells in the symbiont consortium. This uncoupling of symbiont performance and host fitness may be explained by stochastic events such as predation and disease, which substantially increase variation in growth and mortality in field experiments. Alternatively, it may represent some unknown benefit associated with hosting hybrid symbionts, belying their relatively poor photosynthetic performance, and explaining the widespread abundance of these heterogeneous Symbiodinium cells on the Lord Howe Island reef. Our inability to maintain many clade C Symbiodinium types in culture prevents direct observations of hybridization between C100 and C109. Unequivocal evidence of this phenomenon will therefore likely remain elusive until high-resolution, single-copy nuclear markers can be developed, since the incomplete displacement of ancestral polymorphisms can leave a similar genomic signature to that of hybridization. However, this study serves to provide an initial proof-of-principle for hybridization between divergent Symbiodinium taxa. In doing so, it highlights the need to better understand the evolutionary processes underpinning coral- and symbiont-adaptation in a changing climate.</p>