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bioRxiv2023

The Pathfinder plasmid toolkit for genetically engineering newly isolated bacteria enables the study of Drosophila-colonizing Orbaceae

Toolkits of plasmids and genetic parts streamline the process of assembling DNA constructs and engineering microbes. Many of these kits were designed with specific industrial or laboratory microbes in mind. For researchers interested in non-model microbial systems, it is often unclear which tools and techniques will function in newly isolated strains. To address this challenge, we designed the Pathfinder toolkit for quickly determining the compatibility of a bacterium with different plasmid components. Pathfinder plasmids combine three different broad-host-range origins of replication with multiple antibiotic resistance cassettes and reporters, so that sets of parts can be rapidly screened through multiplex conjugation. We first tested these plasmids in Escherichia coli, a strain of Sodalis praecaptivus that colonizes insects, and a Rosenbergiella isolate from leafhoppers. Then, we used the Pathfinder plasmids to engineer previously unstudied bacteria from the family Orbaceae that were isolated from several fly species. Engineered Orbaceae strains were able to colonize Drosophila melanogaster and could be visualized in fly guts. Orbaceae are common and abundant in the guts of wild-caught flies but have not been included in laboratory studies of how the Drosophila microbiome affects fly health. Thus, this work provides foundational genetic tools for studying new host-associated microbes, including bacteria that are a key constituent of the gut microbiome of a model insect species. IMPORTANCETo fully understand how microbes have evolved to interact with their environments, one must be able to modify their genomes. However, it can be difficult and laborious to discover which genetic tools and approaches work for a new isolate. Bacteria from the recently described Orbaceae family are common in the microbiomes of insects. We developed the Pathfinder plasmid toolkit for testing the compatibility of different genetic parts with newly cultured bacteria. We demonstrate its utility by engineering Orbaceae strains isolated from flies to express fluorescent proteins and characterizing how they colonize the Drosophila melanogaster gut. Orbaceae are widespread in Drosophila in the wild but have not been included in laboratory studies examining how the gut microbiome affects fly nutrition, health, and longevity. Our work establishes a path for genetic studies aimed at understanding and altering interactions between these and other newly isolated bacteria and their hosts.

microbiology↗

bioRxiv2023

Testing the efficacy of different molecular tools for parasite conservation genetics: a case study using horsehair worms (Phylum Nematomorpha)

In recent years, parasite conservation has become a globally significant issue. Because of this, there is a need for standardised methods for inferring population status and possible cryptic diversity. However, given the lack of molecular data for some groups, it is challenging to establish procedures for genetic diversity estimation. Therefore, universal tools, such as double digest restriction-site associated DNA sequencing (ddRADseq), could be useful when conducting conservation genetic studies on rarely studied parasites. Here, we generated a ddRADseq dataset that includes all three described Taiwanese horsehair worms (Phylum Nematomorpha), possibly one of the most understudied animal groups. Additionally, we produced data for a fragment of the cytochrome c oxidase subunit I (COXI) for said species. We used the COXI dataset in combination with previously published sequences of the same locus for inferring the effective population size (Ne) trends and possible population structure. We found that a larger and geographically broader sample size combined with more sequenced loci resulted in a better estimation of changes in Ne. We were able to detect demographic changes associated with Pleistocene events in all the species. Furthermore, the ddRADseq dataset for Chordodes formosanus did not reveal a genetic structure based on geography, implying a great dispersal ability, possibly due to its hosts. We showed that different molecular tools can be used to reveal genetic structure and demographic history at different historical times and geographical scales, which can help with conservation genetic studies in rarely studied parasites.

zoology↗

bioRxiv2023

Whole-genome sequencing of ethnolinguistic diverse northwestern Chinese Hexi Corridor people from the 10K_CPGDP project suggested the differentiated East-West genetic admixture along the Silk Road and their biological adaptations

The ancient Silk Road served as the main connection between East and West Eurasia for several centuries. At any rate, the genetic exchange between populations along the ancient Silk Road was likely to leave traces on the contemporary gene pool of local people in Northwest China, which was the passage of the Northern Silk Road. However, genetic sources from northwestern China are under-represented in the current population-scale genomic database. To characterize the genetic architecture and adaptative history of the Northern Silk Road ethnic populations, we performed whole-genome sequencing on 126 individuals from six ethnolinguistic groups (Tibeto-Burman (TB)-speaking Tibetan, Mongolic (MG)-speaking Dongxiang/Tu/eastern Yugur, and Turkic (TK)-speaking Salar/western Yugur) living in Gansu and Qinghai in the 10K Chinese people Genomic Diversity Project (10K_CPGDP). We observed ethnicity-related differentiated population structures among these geographically close Northwest Chinese populations, that is, Salar and Tu people showed a close affinity with southwestern TB groups, and other studied populations shared more alleles with MG and Tungusic groups. Overall, the patterns of genetic clustering were not consistent with linguistic classifications. We estimated that Dongxiang, Tibetan, and Yugur people inherited more than 10% West Eurasian ancestry, much higher than that of Salar and Tu people (<7%). Hence, the difference in the proportion of West Eurasian ancestry has primarily contributed to the genetic divergence of geographically close Northwest Chinese populations. The signatures of natural selection were identified in genes associated with cardiovascular system diseases or lipid metabolism related to triglyceride levels (e.g., PRIM2, PDE4DIP, NOTCH2, DDAH1, GALNT2, and MLIP) and developmental and neurogenetic diseases (e.g., NBPFs 8/9/20/25P, etc.). Moreover, the EPAS1 gene, a transcription factor regulating hypoxia response, showed relatively high PBS values in our studied groups. The sex-biased admixture history, in which the West Eurasian ancestry was introduced primarily by males, was identified in Dongxiang, Tibetan, and Yugur populations. We determined that the eastern-western admixture occurred [~]783-1131 years ago, coinciding with the intensive economic and cultural exchanges during the historic Trans-Eurasian cultural exchange era.

genomics↗

bioRxiv2023

Revealing the chassis-effect on a broad-host-range genetic switch and its concordance with interspecies bacterial physiologies

Broad-host-range synthetic biology is an emerging frontier that aims to expand our current engineerable domain of microbial hosts for biodesign applications. As more novel species are brought to "model status", synthetic biologists are discovering that identically engineered genetic circuits can exhibit different performances depending on the organism it operates within, an observation referred to as the "chassis-effect". It remains a major challenge to uncover which genome encoded and physiological biological determinants will underpin chassis effects that govern the performance of engineered genetic devices. In this study, we compared model and novel bacterial hosts to ask whether phylogenomic relatedness or similarity in host physiology is a better predictor of toggle switch performance. This was accomplished using comparative framework based on multivariate statistical approaches to systematically demonstrate the chassis-effect and characterize the performance dynamics of a genetic toggle switch operating within six Gammaproteobacteria. Our results solidify the notion that genetic devices are significantly impacted by host-context. Furthermore, we formally determined that hosts exhibiting more similar metrics of growth and molecular physiology also exhibit more similar toggle switch performance, indicating that specific bacterial physiology underpins measurable chassis effects. The result of this study contributes to the field of broad-host-range synthetic biology by lending increased predictive power to the implementation of genetic devices in less-established microbial hosts.

synthetic biology↗

bioRxiv2023

Epigenetic then genetic variations underpin rapid adaptation of oyster populations (Crassostrea gigas) to Pacific Oyster Mortality Syndrome (POMS)

Disease emergence is accelerating in response to human activity-induced global changes. Understanding the mechanisms by which host populations can rapidly adapt to this threat will be crucial for developing future management practices. Pacific Oyster Mortality Syndrome (POMS) imposes a substantial and recurrent selective pressure on oyster populations (Crassostrea gigas). Rapid adaptation to this disease may arise through both genetic and epigenetic mechanisms. In this study, we used a combination of whole exome capture of bisulfite-converted DNA, next-generation sequencing, and (epi)genome-wide association mapping, to show that natural oyster populations differentially exposed to POMS displayed signatures of selection both in their genome (single nucleotide polymorphisms) and epigenome (CG-context DNA methylation). Consistent with higher resistance to POMS, the genes targeted by genetic and epigenetic variations were mainly related to host immunity. By combining correlation analyses, DNA methylation quantitative trait loci, and variance partitioning, we revealed that a third of the observed phenotypic variation was explained by interactions between the genetic sequence and epigenetic information, [~]14% by the genetic sequence, and up to 25% by the epigenome alone. Thus, as well as genetic adaptation, epigenetic mechanisms governing immune responses contribute significantly to the rapid adaptation of hosts to emerging infectious diseases.

evolutionary biology↗

bioRxiv2023

Ultra-high throughput mapping of genetic design space

Massively parallel genetic screens have been used to map sequence-to-function relationships for a variety of genetic elements. However, because these approaches only interrogate short sequences, it remains challenging to perform high throughput (HT) assays on constructs containing combinations of multiple sequence elements arranged across multi-kb length scales. Overcoming this barrier could accelerate synthetic biology; by screening diverse gene circuit designs and learning "composition-to-function" mappings that reveal genetic part composability rules and enable rapid identification of behavior-optimized variants. Here, we introduce CLASSIC, a genetic screening platform that combines long- and short-read next-generation sequencing (NGS) modalities to quantitatively assess pools of constructs of arbitrary length containing diverse part compositions. We show that CLAS-SIC can measure expression profiles of >105 gene circuit designs (from 5-20 kb) in a single experiment in human cells. The resulting datasets can be used to train ML models that accurately predict circuit behavior across expansive circuit design landscapes, revealing part composability rules that govern circuit performance. Our work shows that by expanding the throughput of each design-build-test-learn (DBTL) cycle, CLASSIC enhances the pace and scale of synthetic biology and establishes an experimental basis for data-driven design of complex genetic systems.

synthetic biology↗

bioRxiv2023

Determinants of genetic diversity in sticklebacks

Understanding what determines species and population differences in levels of genetic diversity has important implications for our understanding of evolution, as well as for the conservation and management of wild populations. Previous comparative studies have emphasized the roles of linked selection, life-history trait variation and genomic properties, rather than pure demography, as important determinants of genetic diversity. However, these findings are based on coarse estimates across a range of highly diverged taxa, and it is unclear how well they represent the processes within individual species. We assessed genome-wide genetic diversity ({pi}) in 45 nine-spined stickleback (Pungitius pungitius) populations and found that{pi} varied 15-fold among populations ({pi}min{approx}0.00015,{pi} max{approx}0.0023) whereas estimates of recent effective population sizes varied 122-fold. Analysis of inbreeding coefficients (FROH) estimated from runs of homozygosity revealed strong negative association between{pi} and FROH. Genetic diversity was also negatively correlated with mean body size and longevity, but these associations were not statistically significant after controlling for demographic effects (FROH). The results give strong support for the view that populations demographic features, rather than life history differences, are the chief determinants of genetic diversity in the wild.

evolutionary biology↗

bioRxiv2023

Evolutionary rescue through gene flow despite genetic incompatibilities shaped diversity of the pseudo-cereal grain amaranth

Crop domestication and the subsequent expansion of crops have long been thought of as a linear process from a wild ancestor to a domesticate. However, evidence of gene flow from locally adapted wild relatives that provided adaptive alleles into crops has been identified in multiple species. Yet, little is known about the evolutionary consequences of gene flow during domestication and the interaction of gene flow and genetic load in crop populations. We study the pseudo-cereal grain amaranth that has been domesticated three times in different geographic regions of the Americas. We quantify the amount and distribution of gene flow and genetic load along the genome of the three grain amaranth species and their two wild relatives. Our results show ample gene flow between crop species and between crops and their wild relatives. Gene flow from wild relatives decreased genetic load in the three crop species. This suggests that wild relatives could provide evolutionary rescue by replacing deleterious alleles in crops. We assess experimental hybrids between the three crop species and found genetic incompatibilities between one Central American grain amaranth and the other two crop species. These incompatibilities might have created recent reproductive barriers and maintained species integrity today. Together, our results show that gene flow played an important role in the domestication and expansion of grain amaranth, despite genetic species barriers. The domestication of plants was likely not linear and created a genomic mosaic by multiple contributors with varying fitness effects for todays crops.

evolutionary biology↗

bioRxiv2023

Genetic and Environmental interactions contribute to immune variation in rewilded mice

The relative and synergistic contributions of genetics and environment to inter-individual immune response variation remain unclear, despite its implications for understanding both evolutionary biology and medicine. Here, we quantify interactive effects of genotype and environment on immune traits by investigating three inbred mouse strains rewilded in an outdoor enclosure and infected with the parasite, Trichuris muris. Whereas cytokine response heterogeneity was primarily driven by genotype, cellular composition heterogeneity was shaped by interactions between genotype and environment. Notably, genetic differences under laboratory conditions can be decreased following rewilding, and variation in T cell markers are more driven by genetics, whereas B cell markers are driven more by environment. Importantly, variation in worm burden is associated with measures of immune variation, as well as genetics and environment. These results indicate that nonheritable influences interact with genetic factors to shape immune variation, with synergistic impacts on the deployment and evolution of defense mechanisms.

immunology↗

bioRxiv2023

Host genetic variation guides hepacivirus clearance, chronicity, and liver fibrosis in mice

Background & AimsHuman genetic variation is thought to guide the outcome of hepatitis C virus (HCV) infection but model systems within which to dissect these host genetic mechanisms are limited. Norway rat hepacivirus (NrHV), closely related to HCV, causes chronic liver infection in rats but causes acute self-limiting hepatitis in typical strains of laboratory mice, which resolves in two weeks. The Collaborative Cross (CC) is a robust mouse genetics resource comprised of a panel of recombinant inbred strains, which model the complexity of the human genome and provide a system within which to understand diseases driven by complex allelic variation. Approach & ResultsWe infected a panel of CC strains with NrHV and identified several that failed to clear virus after 4 weeks. Strains displayed an array of virologic phenotypes ranging from delayed clearance (CC046) to chronicity (CC071, CC080) with viremia for at least 10 months. Body weight loss, hepatocyte infection frequency, viral evolution, T-cell recruitment to the liver, liver inflammation and the capacity to develop liver fibrosis varied among infected CC strains. ConclusionsThese models recapitulate many aspects of HCV infection in humans and demonstrate that host genetic variation affects a multitude of virus and host phenotypes. These models can be used to better understand the molecular mechanisms that drive hepacivirus clearance and chronicity, the virus and host interactions that promote chronic disease manifestations like liver fibrosis, therapeutic and vaccine performance, and how these factors are affected by host genetic variation.

microbiology↗

bioRxiv2023

Recombulator-X: a fast and user-friendly tool for estimating X chromosome recombination rates in forensic genetics

Background and ObjectiveGenetic markers (especially short tandem repeats or STRs) located on the X chromosome are a valuable resource to solve complex kinship cases in forensic genetics in addition or alternatively to autosomal STRs. Groups of tightly linked markers are combined into haplotypes, thus increasing the discriminating power of tests. However, this approach requires precise knowledge of the recombination rates between adjacent markers. Recombination rates vary across the human genome and cannot be automatically derived from linkage physical maps. The International Society of Forensic Genetics recommends that recombination rate estimation on the X chromosome is performed from pedigree genetic data while taking into account the confounding effect of mutations. However, the only existing implementations that satisfy these requirements have several drawbacks: they were never publicly released, they are very slow and/or need cluster-level hardware and strong computational expertise to use. In order to address these key concerns, we developed Recombulator-X, a new open-source Python tool. MethodsThe most challenging issue, namely the running time, was addressed with dynamic programming techniques to greatly reduce the computational complexity of the algorithm, coupled with JIT compilation to further increase performance. We also extended the statistical framework from STR to any polymorphic marker. ResultsCompared to the previous methods, Recombulator-X reduces the estimation times from weeks or months to less than one hour for typical datasets. Moreover, the estimation process, including preprocessing, has been streamlined and packaged into a simple command-line tool that can be run on a normal PC. Where previous approaches were limited to small panels of STR markers (up to 15), our tool can handle greater numbers (up to 100) of mixed STR and non-STR markers. ConclusionsIn the genetic forensic community, state-of-the-art estimation methods for X chromosome recombination rates have seen limited usage due to the technical hurdles posed by previous implementations. Recombulator-X makes the process much simpler, faster and accessible to researchers without a computational background, hopefully spurring increased adoption of best practices. Moreover, it extends the estimation framework to larger panels of genetic markers (not only STRs), allowing analyses of sequencing-based data.

bioinformatics↗

bioRxiv2023

Genetic basis of expression and splicing underlying spike architecture in wheat (Triticum aestivum L.)

IntroductionWheat is one of the most important staple crops worldwide, and an important source of human protein and mineral element intake. Continuously increasing stable production of wheat is critical for global food security under the challenge of population growth and limited resource input. ObjectiveSpike architecture determines the potential grain yield of wheat. However, the mechanisms of transcriptional regulation of spike architecture in wheat remain largely unknown, limiting further genetic improvement of wheat yield. In this study we explored the genetic basis of spike architecture in wheat. MethodsPopulation RNA-seq methods were used to identify the eQTLs and sQTLs associated with spike architecture and applied this to dissection of the genetic basis of gene expression and splicing controlling these complex yield-related traits. ResultsIn total, 4,143 expression quantitative trait loci (eQTLs) and 12,933 splice QTLs (sQTLs) were identified in wheat based on 178 RNA-seq samples, revealing 774 cis-eQTLs and 321 cis-sQTLs for 86 eGenes and 73 sGenes, respectively. Integration of eQTLs and sQTLs with genome-wide association study (GWAS) identified dozens of additional novel candidate genes that may contribute to spike-related traits. Gene network analysis showed that eQTLs and sQTLs were widely involved in the co-expression modules that regulate wheat spike architecture. Notably, the eQTL locus AX-108754757 regulated the expression of 5 eGenes that negatively controled grain number per spike. AX-111592099 regulated both the splicing and expression of TraesCS7B02G442100, encoding an E3 ubiquitin ligase, and playing a central role in regulating spike length. ConclusionThis study provides new insights into the genetic basis of spike architecture. This improved understanding of spike-related traits in wheat will contribute to more rapid genetic improvement.

plant biology↗

bioRxiv2023

Influence of oceanography and geographic distance on genetic structure: how varying the sampled domain influences conclusions in Laminaria digitata

Understanding the environmental processes shaping connectivity can greatly improve management and conservation actions which are essential in the trailing edge of species distributions. In this study, we used a dataset built from 32 populations situated in the southern limit of the kelp species Laminaria digitata. By extracting data from 11 microsatellite markers, our aim was to (1) refine the analyses of population structure, (2) compare connectivity patterns and genetic diversity between island and mainland populations and (3) evaluate the influence of sampling year, hydrodynamic processes, habitat discontinuity, spatial distance and sea surface temperature on the genetic structure using a distance-based redundancy analysis (db-RDA). Analyses of population structure enabled to identify well connected populations associated to high genetic diversity, and others which appeared genetically isolated from neighboring populations and showing signs of genetic erosion verifying contrasting ecological (and demographic) status in Brittany and the English Channel. By performing db-RDA analyses on various sampling sizes, geographic distance appeared as the dominant factor influencing connectivity between populations separated by great distances, while hydrodynamic processes were the main factor at smaller scale. Finally, Lagrangian simulations enabled to study the directionality of gene flow which has implications on source-sink dynamics. Overall, our results have important significance in regard to the management of kelp populations facing pressures both from global warming and their exploitation for commercial use.

evolutionary biology↗

bioRxiv2023

Natural genetic variation underlying the negative effect of elevated CO2 on ionome composition in Arabidopsis thaliana

The elevation of atmospheric CO2 leads to a decline in the plant mineral content, which might pose a significant threat to food security in the coming decades. To date, very few genes have been identified as having a role in the negative effect of elevated CO2 on plant mineral composition. Yet, several studies have shown a certain degree of diversity in the ionomes response to elevated CO2, associated with genotypic variation. This suggests the existence of genetic factors controlling the effect of CO2 on ionome composition. However, no large-scale studies have been carried out to date to explore the genetic diversity of the ionome responses to elevated CO2. Here, we used six hundred Arabidopsis thaliana accessions, representing geographical distributions ranging from worldwide to regional and local environments, to analyze the natural genetic variation underlying the negative effect of elevated CO2 on the ionome composition in plants. We show that the growth under elevated CO2 leads to a global and important decrease of the ionome content whatever the geographic distribution of the population. We also observed a high range of genetic diversity in the response of the ionome composition to elevated CO2, and we identified sub-populations, showing effects on their ionome ranging from the most pronounced to resilience or even to a benefit in response to elevated CO2. Using genome-wide association mapping on the response of each mineral element to elevated CO2 or on integrative traits, we identified a large set of QTLs and genes associated with the ionome response to elevated CO2. Finally, we demonstrate that the function of one of these genes is associated to the negative effect of elevated CO2 on the plant mineral composition. This resource will contribute to understand the genetic mechanisms underlying the negative effect of elevated CO2 on plant mineral nutrition, and could help towards the development of crops adapted to a high-CO2 world.

plant biology↗

bioRxiv2023

Using a P. falciparum genetic cross to dissect the relative contributions of pfcrt and plasmepsin II/III to piperaquine response-related traits

Piperaquine (PPQ) is widely used in combination with dihydroartemisinin (DHA) as a first-line treatment against malaria parasites. Multiple genetic drivers of PPQ resistance have been reported, including mutations in the Plasmodium falciparum chloroquine resistance transporter (pfcrt) and increased copies of plasmepsin II/III (pm2/3). We generated a cross between a Cambodia-derived multi-drug resistant KEL1/PLA1 lineage isolate (KH004) and a drug susceptible parasite isolated in Malawi (Mal31). Mal31 harbors a wild-type (3D7-like) pfcrt allele and a single copy of pm2/3, while KH004 has a chloroquine-resistant (Dd2-like) pfcrt allele with an additional G367C substitution and four copies of pm2/3. We recovered 104 unique recombinant progeny and examined a targeted set of progeny representing all possible combinations of variants at pfcrt and pm2/3 for detailed analysis of competitive fitness and a range of PPQ susceptibility phenotypes, including PPQ survival assay (PSA), area under the dose-response curve (AUC), and a limited point IC50 (LP-IC50). We find that inheritance of the KH004 pfcrt allele is required for PPQ resistance, whereas copy number variation in pm2/3 further enhances resistance but does not confer resistance in the absence of PPQ-R-associated mutations in pfcrt. Deeper investigation of genotype-phenotype relationships demonstrates that progeny clones from experimental crosses can be used to understand the relative contributions of pfcrt, pm2/3, and parasite genetic background, to a range of PPQ-related traits and confirm the critical role of the PfCRT G367C substitution in PPQ resistance. ImportanceResistance to PPQ used in combination with DHA has emerged in Cambodia and threatens to spread to other malaria-endemic regions. Understanding the causal mutations of drug resistance and their impact on parasite fitness is critical for surveillance and intervention, and can also reveal new avenues to limiting the evolution and spread of drug resistance. An experimental genetic cross is a powerful tool for pinpointing the genetic determinants of key drug resistance and fitness phenotypes and have the distinct advantage of assaying the effects of naturally evolved genetic variation. Our study was significantly strengthened because the full a range of copies of KH004 pm2/3 was inherited among the progeny clones, allowing us to directly test the role of pm2/3 copy number on resistance-related phenotypes in the context of a unique pfcrt allele. Our multi-gene model suggests an important role for both loci in the evolution of this ACT resistant parasite lineage.

genomics↗

bioRxiv2023

Evaluating the ileal and cecal microbiota composition of a 1940 heritage genetic line and a 2016 commercial line of white leghorns fed representative diets from 1940 and 2016

This study was conducted to identify and evaluate the differences between the microbiome composition of the ileum and ceca of 1940 and 2016 genetic strains of white leghorns fed representative contemporary diets from those times. Ileal and cecal samples were aseptically collected from both genetic lines at 69 weeks of age. The genomic DNA of the ileal and cecal contents were extracted and the V4 region of the 16S rDNA was sequenced on an Illumina Miseq. Microbiota data were filtered and aligned using the QIIME2 2020.2 pipeline. Alpha and beta diversity metrics were generated and the Analysis of Composition of Microbiomes (ANCOM) was utilized to determine significantly different taxa. Data were considered significant at P [≤] 0.05 for main effects and Q [≤] 0.05 for pairwise differences. Alpha diversity of the ileum and ceca were significantly different (P = 0.001; Q = 0.001; however, no differences between genetic lineage were observed (P > 0.05; Q > 0.05). The beta diversity between the ileum and ceca, as well as between the genetic lines (1940 vs. 2016) were significantly different from one another (P = 0.001; Q = 0.001). Using ANCOM, Proteobacteria and Actinobacteriota were significantly different than other phyla (P 0.05) with a higher relative abundance of Proteobacteria being observed among treatment groups 2 and 3, while Actinobacteriota had higher relative abundance in treatment groups 1 and 4. Among the significantly different genera in the ileum, Pseudomonas, Rhizobiaceae, Leuconostoc, and Aeriscardovia were different (P 0.05) with treatment groups 1 and 4 having a higher relative abundance of Aeriscardovia, while treatment groups 2 and 3 had higher relative abundance in both Pseudomonas and Leuconostoc. In the ceca, Proteobacteria, Firmicutes, Actinobacteriota, and Euryarchaeota were significantly different phyla (P 0.05) with Firmicutes having the highest relative abundance across all treatment groups. Among the significantly different genera (Pseudomonas, Leuconostoc, Alloprevotella, and Aeriscardovia), Alloprevotella had the highest relative abundance across all treatment groups 1 and 2, while Leuconostoc and Pseudomonas had the highest relative abundance in treatment group 4. Results from this study suggest that genetic makeup in conjunction with the nutritional composition of laying hens influences the cecal and ileal microbiota of corresponding hens.

microbiology↗

bioRxiv2023

Machine learning and phylogenetic models identify predictors of genetic variation in Neotropical amphibians

AimIntraspecific genetic variation is key for adaptation and survival in changing environments and is known to be influenced by many factors, including population size, migration, and life history traits. We investigated genetic variation within Neotropical amphibian species to provide insights into how natural history traits, phylogeny, climatic, and geographic characteristics influence intraspecific diversity. LocationNeotropics. TaxonAmphibians. MethodsWe assembled datasets using open-access databases for natural history traits, genetic sequences, phylogenetic trees, climatic, and geographic data. For each species, we calculated overall nucleotide diversity ({pi}) and tested for isolation by distance (IBD) and isolation by environment (IBE). We then identified predictors of {pi}, IBD, and IBE using Random Forest (RF) regression or RF classification. To incorporate phylogenetic relationships, we fitted phylogenetic generalized linear mixed models (PGLMMs) to predict {pi}, IBD, and IBE. ResultsWe compiled 4,052 mitochondrial DNA sequences from 256 amphibian species (230 frogs and 26 salamanders), georeferencing 2,477 sequences from 176 species that were not linked to occurrence data. RF regressions and PGLMMs were congruent in identifying range size and precipitation ({sigma}) as the most important predictors of {pi}. RF classification and PGLMMs identified minimum elevation as an important predictor of IBD, and maximum latitude and precipitation ({sigma}) as the best predictors of IBE. Main conclusionsThis study unified machine learning and phylogenetic methods and identified predictors of genetic variation in Neotropical amphibians. This approach was valuable to determine which predictors were congruent between methods. We found that species with small ranges or living in zones with less variable precipitation tended to have low genetic diversity. We also showed that Western Mesoamerica, Andes, and Atlantic Forest biogeographic units harbor high diversity across many species that should be prioritized for protection. These results could play a key role in the development of conservation strategies for Neotropical amphibians.

evolutionary biology↗

bioRxiv2023

1LocusSim a mobile-friendly simulator for teaching population genetics

SummaryBiology students often struggle with the fundamental concepts of evolutionary genetics, including genetic drift, mutation, and selection. To address this problem, 1LocusSim was developed to simulate the interaction of different factors, such as population size, mutation, selection, and dominance, to study their effect on allelic frequency during evolution. With 1LocusSim, students can compare theoretical results with simulation outputs and solve and analyze different problems of population genetics. The 1LocusSim web has a responsive design which means that it has been specifically designed to be used on smartphones. To demonstrate its use, I review the classical overdominance model of population genetics and highlight a characteristic that is often not explicitly stated. Specifically, it is emphasized that the equilibrium of the model does not depend on the homozygous selection coefficients but rather on the ratio of the selection coefficients. This is already clear from the classical formula but maybe not so much for students. Also it implies that the equilibrium can be expressed solely in terms of the dominance coefficient h. To verify these theoretical prediction, I utilize the simulator and calculate the equilibrium for the well-known case of sickle cell anaemia. Simulating basic population genetic models on smartphones can be a powerful learning aid that fosters critical skills and opens up new opportunities for Biology students. By utilizing this tool, students can learn at their own pace and convenience, anywhere and anytime. Software and data availability1LocusSim if freely available at https://1LocusSim-biosdev.pythonanywhere.com/. Website implemented under the Bottle micro web-framework for Python, with all major browsers supported. Contactacraaj@uvigo.es Supplementary informationThe manual and examples are available from the help button of the program or directly at https://acraaj.webs.uvigo.es/1LocusSim/1LocusSim_EN.html. Both the program and the manual pages are in English and Spanish.

scientific communication and education↗