Sequence Divergence Research Articles

Testing the evolutionary theory of inversion polymorphisms in the yellow monkeyflower (Mimulus guttatus)

Chromosomal inversions have been implicated in a remarkable range of natural phenomena, but it remains unclear how much they contribute to standing genetic variation. Here, we evaluate 64 inversions that segregate within a single natural population of the yellow monkeyflower (Mimulus guttatus). Nucleotide diversity patterns confirm low internal variation for the derived orientation (predicted by recent origin), elevated diversity between orientations (predicted by natural selection), and localized fluctuations (predicted by gene flux). Sequence divergence between orientations varies idiosyncratically by position, not following the suspension bridge pattern predicted if the breakpoints are the targets of selection. Genetic variation in gene expression is not inflated close to inversion breakpoints but is clearly partitioned between orientations. Like sequence variation, the pattern of expression variation suggests that the capture of coadapted alleles is more important than the breakpoints for the fitness effects of inversions. This work confirms several evolutionary predictions for inversion polymorphisms, but clarity emerges only by synthesizing estimates across many loci.

Aposematic color polymorphism is a poor indicator of species boundaries in North American Paranthrene (Lepidoptera: Sesiidae) as evidenced by a multi-gene phylogeny.

Color polymorphism among animal species can influence speciation. Factors such as natural and sexual selection, genetic drift and gene flow contribute to the maintenance of color polymorphism within the species or spur speciation. The evolutionary and ecological mechanisms for color polymorphism are taxon specific and only a few species have been studied. A phylogeny provides an evolutionary framework to understanding the association between color polymorphism and species. Paranthrene species are day flying aposematic moths that mimic wasps in both appearance and behavior. The genus has several polymorphic species and some color forms were originally described as species. Paranthrene presents the opportunity to test for an association between color polymorphism and species boundaries. We reconstructed a phylogeny using DNA sequence from COI, EF-1alpha, and Wingless genes from 67 specimens representing all North American Paranthrene species, nine color variants, and two outgroups. Parsimony and Bayesian analyses generally agreed in topology and support values. Paranthrene simulans (Grote, 1881) was polyphyletic, and monophyly of P. pellucida Greenfield and Karandinos, 1979 was not recovered. Paranthrene robiniae Hy. Edwards, 1880 was polyphyletic and genitalic and genetic differences among the three clades supported the recognition of two new pseudocryptic species, Paranthrene oasis Smith, Taft and, Cognato, new species and Paranthrene gilaensis Smith, Taft and, Cognato, new species. Paranthrene color variants did not overwhelming associate with species boundaries. Of the nine color forms we examined, only two were monophyletic, had DNA sequence divergence comparable to other species, and associated with species diagnostic morphology. It is likely that genetic drift and allopatric isolation explains the fixation of color variants with species. The mechanisms maintaining color polymorphisms within Paranthrene species will remain unknown until experimentation concerning the degree of aposematic protection in reference to wasp models is conducted.

Functions of PMS2 and MLH1 important for regulation of divergent repeat-mediated deletions

Repeat-mediated deletions (RMDs) are a type of deletion rearrangement that utilizes two repetitive elements to bridge a DNA double-strand break (DSB) that leads to loss of the intervening sequence and one of the repeats. Sequence divergence between repeats causes RMD suppression and indeed this divergence must be resolved in the RMD products. The mismatch repair factor, MLH1, was shown to be critical for both RMD suppression and a polarity of sequence divergence resolution in RMDs. Here, we sought to study the interrelationship between these two aspects of RMD regulation (i.e., RMD suppression and polar divergence resolution), by examining several mutants of MLH1 and its binding partner PMS2. To begin with, we show that PMS2 is also critical for both RMD suppression and polar resolution of sequence divergence in RMD products. Then, with six mutants of the MLH1-PMS2 heterodimer, we found several different patterns: three mutants showed defects in both functions, one mutant showed loss of RMD suppression but not polar divergence resolution, whereas another mutant showed the opposite, and finally one mutant showed loss of RMD suppression but had a complex effect on polar divergence resolution. These findings indicate that RMD suppression vs. polar resolution of sequence divergence are distinct functions of MLH1-PMS2.

Structural characterization of human monoclonal antibodies targeting uncommon antigenic sites on spike glycoprotein of SARS-CoV.

The function of the spike protein N terminal domain (NTD) in coronavirus (CoV) infections is poorly understood. However, some rare antibodies that target the SARS-CoV-2 NTD potently neutralize the virus. This finding suggests the NTD may contribute in part to protective immunity. Pan-sarbecovirus antibodies are desirable for broad protection, but the NTD region of SARS-CoV and SARS-CoV-2 exhibit a high level of sequence divergence, and therefore, cross-reactive NTD-specific antibodies are unexpected, and there is no structure of a SARS-CoV NTD-specific antibody in complex with NTD. Here we report a monoclonal antibody COV1-65 encoded by the IGHV1-69 gene that recognizes the NTD of SARS-CoV S protein. A prophylaxis study showed the MAb COV1-65 prevented disease when administered before SARS-CoV challenge of BALB/c mice, an effect that requires intact Fc effector functions for optimal protection in vivo. The footprint on the S protein of COV1-65 is near to functional components of the S2 fusion machinery, and the selection of COV1-65 escape mutant viruses identified critical residues Y886H and Q974H, which likely affect the epitope through allosteric effects. Structural features of the mAb COV1-65-SARS-CoV antigen interaction suggest critical antigenic determinants that should be considered in the rational design of sarbecovirus vaccine candidates.

Examining the molecular clock hypothesis for the contemporary evolution of the rabies virus.

The molecular clock hypothesis assumes that mutations accumulate on an organism's genome at a constant rate over time, but this assumption does not always hold true. While modelling approaches exist to accommodate deviations from a strict molecular clock, assumptions about rate variation may not fully represent the underlying evolutionary processes. There is considerable variability in rabies virus (RABV) incubation periods, ranging from days to over a year, during which viral replication may be reduced. This prompts the question of whether modelling RABV on a per infection generation basis might be more appropriate. We investigate how variable incubation periods affect root-to-tip divergence under per-unit time and per-generation models of mutation. Additionally, we assess how well these models represent root-to-tip divergence in time-stamped RABV sequences. We find that at low substitution rates (<1 substitution per genome per generation) divergence patterns between these models are difficult to distinguish, while above this threshold differences become apparent across a range of sampling rates. Using a Tanzanian RABV dataset, we calculate the mean substitution rate to be 0.17 substitutions per genome per generation. At RABV's substitution rate, the per-generation substitution model is unlikely to represent rabies evolution substantially differently than the molecular clock model when examining contemporary outbreaks; over enough generations for any divergence to accumulate, extreme incubation periods average out. However, measuring substitution rates per-generation holds potential in applications such as inferring transmission trees and predicting lineage emergence.

The Growing Phenomenon of ‘Frozen’ Virus Genome Sequences and Their Likely Origin in Research Facility Escapes

‘Frozen’ virus genome sequences are sampled from outbreaks and have unusually low sequence divergence when compared to genome sequences from historical strains. A growing number of ‘frozen’ virus genome sequences are being reported as virus genome sequencing becomes more common. Examples of ‘frozen’ sequences include the 1977 H1N1 ‘Russian’ flu; Venezuelan Equine Encephalitis Virus from Venezuela and Colombia in 1995; E71 sequences from a Hand, Foot and Mouth outbreak in 2007–2009 in China; and a polio strain isolated in 2014 from Anhui, China. The origin of these ‘frozen’ sequences has been attributed to escapes from research facilities and often appears to be associated with vaccine work. Consequently, a new paradigm for pathogen emergence appears in operation, that involves laboratory research or vaccine production which utilizes ‘live’ virus isolates of historical strains. The accidental release and re-emergence of such strains are straightforward to detect from their genome sequences and should spur the routine sequencing and publication of all known pathogenic viral strains undergoing experimentation, or being used for vaccine manufacture, in order to facilitate tracing. However, it is noted that novel pathogenic viruses accidentally released into the population from research facilities are harder to detect if their sequence has first not been made public, which should prompt the routine sequencing and reporting of all novel pathogenic viruses before experimentation.

Chromosome-level genome assemblies and genetic maps reveal heterochiasmy and macrosynteny in endangered Atlantic Acropora

BackgroundOver their evolutionary history, corals have adapted to sea level rise and increasing ocean temperatures, however, it is unclear how quickly they may respond to rapid change. Genome structure and genetic diversity contained within may highlight their adaptive potential.ResultsWe present chromosome-scale genome assemblies and linkage maps of the critically endangered Atlantic acroporids, Acropora palmata and A. cervicornis. Both assemblies and linkage maps were resolved into 14 chromosomes with their gene content and colinearity. Repeats and chromosome arrangements were largely preserved between the species. The family Acroporidae and the genus Acropora exhibited many phylogenetically significant gene family expansions. Macrosynteny decreased with phylogenetic distance. Nevertheless, scleractinians shared six of the 21 cnidarian ancestral linkage groups as well as numerous fission and fusion events compared to other distantly related cnidarians. Genetic linkage maps were constructed from one A. palmata family and 16 A. cervicornis families using a genotyping array. The consensus maps span 1,013.42 cM and 927.36 cM for A. palmata and A. cervicornis, respectively. Both species exhibited high genome-wide recombination rates (3.04 to 3.53 cM/Mb) and pronounced sex-based differences, known as heterochiasmy, with 2 to 2.5X higher recombination rates estimated in the female maps.ConclusionsTogether, the chromosome-scale assemblies and genetic maps we present here are the first detailed look at the genomic landscapes of the critically endangered Atlantic acroporids. These data sets revealed that adaptive capacity of Atlantic acroporids is not limited by their recombination rates. The sister species maintain macrosynteny with few genes with high sequence divergence that may act as reproductive barriers between them. In the Atlantic Acropora, hybridization between the two sister species yields an F1 hybrid with limited fertility despite the high levels of macrosynteny and gene colinearity of their genomes. Together, these resources now enable genome-wide association studies and discovery of quantitative trait loci, two tools that can aid in the conservation of these species.

Energy Budgeting of Different Cropping Sequences in the Indian Upper Gangetic Plains

Background: This study is meant to examine the energy requirement and energy input- output of different cropping sequences .The crucial objective to conduct experiment to understand energy efficiency using inputs and outputs which were disbursed in cropping sequences. Hence, it is need of the hour to identify the most remunerative and cost effective cropping sequence with high energy efficient for UGP of India. Methods: This study was carried out at the research farm of ICAR-Indian Institute of Farming Systems Research, Modipuram, during 2017-2021.The divergent cropping sequences viz. sugarcane-ratoon-wheat (CS1); rice-wheat-dhaincha (CS2); pigeonpea +maize-chickpea-okra (CS3); maize-berseem-black gram (CS4); sorghum-mustard-green gram (CS5) and Napier+cowpea/berseem (CS6) were compared in reference to curtail higher energy inputs through selected alternate cropping sequences. The obtained energy values were calculated by multiplying the amount of inputs and outputs by using energy conversion factors. Result: Maximum inputs energy consumed by sugarcane crop alone (33.14´103 MJ ha-1). Results shown that irrigation, seed, fertilizers and diesel required higher energy for the completion of cultural operations. However, higher inputs energy was used in irrigation followed by seed and fertilizers, respectively. In regard to per cent energy intake through inputs, the highest energy spent was for irrigation (35.30 MJ ha-1) and fertilizer (23.80 MJ ha-1).The wheat equivalent yield was higher in sugarcane-ratoon-wheat (125.58 t ha-1). Maximum output energy was with above system (596.70×103 MJ ha-1). Highest net energy returns was counted with sugarcane-ratoon-wheat (549.37´103 MJ ha-1), energy ratio (12.60) and energy profitability (11.60). Indeed, energy efficiency was highest in same system (1657.50) followed by maize-berseem-black gram (1421.96).

Structural diversity and distribution of NMCP-class nuclear lamina proteins in streptophytic algae.

Nuclear Matrix Constituent Proteins (NMCPs) in plants function like animal lamins, providing the structural foundation of the nuclear lamina and regulating nuclear organization and morphology. Although they are well-characterized in angiosperms, the presence and structure of NMCPs in more distantly related species, such as streptophytic algae, are relatively unknown. The rapid evolution of NMCPs throughout the plant lineage has caused a divergence in protein sequence that makes similarity-based searches less effective. Structural features are more likely to be conserved compared to primary amino acid sequence; therefore, we developed a filtration protocol to search for diverged NMCPs based on four physical characteristics: intrinsically disordered content, isoelectric point, number of amino acids, and the presence of a central coiled-coil domain. By setting parameters to recognize the properties of bona fide NMCP proteins in angiosperms, we filtered eight complete proteomes from streptophytic algae species and identified strong NMCP candidates in six taxa in the Classes Zygnematophyceae, Charophyceae, and Klebsormidophyceae. Through analysis of these proteins, we observed structural variance in domain size between NMCPs in algae and land plants, as well as a single block of amino acid conservation. Our analysis indicates that NMCPs are absent in the Mesostigmatophyceae. The presence versus absence of NMCP proteins does not correlate with the distribution of different forms of mitosis (e.g., closed/semi-closed/open) but does correspond to the transition from unicellularity to multicellularity in the streptophytic algae, suggesting that an NMCP-based nucleoskeleton plays important roles in supporting cell-to-cell interactions.

Phylogenetic placement of Mt Bamboutos endemic skink, Trachylepis mekuana (Chirio and Ineich, 2001)

African skinks of the genus Trachylepis is one of the most diverse genera of lizards in Africa. Although, many species have not been validated phylogenetically in recent years. In this study we evaluate the phylogenetic status of the Cameroon Volcanic Line endemic montane skink, Trachylepis mekuana. We recover this species as part of the larger Trachylepis varia group, with sister relationship to the morphological similar skinks from Eastern Africa Trachylepis megalura and central Africa Trachylepis raymondlaurenti. Low sequence divergence (&lt;1.6% 16S) have been observed among the three species. Based on some morphological and colouration differences, as well as mostly allopatric distribution we regard them as good species.

The risk of inbreeding versus outbreeding depression in managing an endangered and locally adapted population of a sedentary bird

AbstractA debate in conservation genomics centers on whether to conserve small, fragmented populations independently or blend them through translocations from larger populations. Translocations of red grouse (Lagopus scotica) from Great Britain to supplement the Irish population have been suggested. We incorporate a variety of genetic datasets to address this. We used genome wide data from 23 contemporary and historic red grouse from Great Britain and Ireland. We also investigate microsatellite data, sequence candidate pigmentation genes, and assess phenotypic color variation. Genomic data indicate higher inbreeding in Irish grouse relative to an English population and significant divergence for genomic (FST = 0.095) and microsatellite (FST = 0.03) markers. Contemporary Ne was seven times smaller in the Irish population compared to the English. We identified divergent regions linked to pigmentation, immune response, and food intake. We show phenotypic differences in plumage color and sequence divergence among coding regions in the melanin pathway including MC1R (FST from genomic data of 0.3). The two populations thus appear locally adapted and this divergence between the source and target population when used for conservation translocations can swamp locally adapted alleles and/or introduce maladapted genotypes, leading to outbreeding depression. While it is important to avoid inbreeding by sustaining larger populations, our research emphasizes the need for practitioners to consider population divergence and local adaptation. We advocate against translocations between Ireland and Britain as a conservation strategy in this particular case and underscore the importance of prioritizing local populations where possible.

Substrate-Multiplexed Assessment of Aromatic Prenyltransferase Activity.

An increasingly effective strategy to identify synthetically useful enzymes is to sample the diversity already present in Nature. Here, we construct and assay a panel of phylogenetically diverse aromatic prenyltransferases (PTs). These enzymes catalyze a variety of C-C bond forming reactions in natural product biosynthesis and are emerging as tools for synthetic chemistry and biology. Homolog screening was further empowered through substrate-multiplexed screening, which provides direct information on enzyme specificity. We perform a head-to-head assessment of the model members of the PT family and further identify homologs with divergent sequences that rival these superb enzymes. This effort revealed the first bacterial O-Tyr PT and, together, provide valuable benchmarking for future synthetic applications of PTs.

Genomic analysis provides insights into the origin and divergence of fruit flavor and flesh color of pummelo.

Pummelo (Citrus maxima) is one of the most important citrus crops and have genetically contributed sweet orange, lemon and most citrus cultivars. It has been cultivated for c. 4000 years in China and is also distributed in many Southeast Asian countries. Nevertheless, the origin and dispersal of pummelo remain elusive. We conducted whole-genome sequencing for 290 pummelo accessions from China and Southeast Asia (SEA). Our findings indicated that pummelo was originated in Yunnan province. The divergence of the China-SEA accessions occurred c. 2000 years ago and the divergence was likely facilitated through the Maritime Silk Road. We detected the divergence of genomic regions associated with fruit flavor and color, indicating different selection by human activities in different regions. A gene encoding lycopene cyclase 2 (LCYB2) exhibited a high degree of divergence in expression and sequence between red-flesh and white-flesh pummelos. A SNP in the coding region of LCYB2 resulted in a reduction in lycopene β-cyclizing enzyme activity, leading to the accumulation of lycopene and the development of the red-flesh trait. This study reveals the origin and evolutionary history of pummelo and provides insights into the genomic basis for the divergence of fruit flavor and color.

Continuous evolution of user-defined genes at 1 million times the genomic mutation rate.

When nature evolves a gene over eons at scale, it produces a diversity of homologous sequences with patterns of conservation and change that contain rich structural, functional, and historical information about the gene. However, natural gene diversity accumulates slowly and likely excludes large regions of functional sequence space, limiting the information that is encoded and extractable. We introduce upgraded orthogonal DNA replication (OrthoRep) systems that radically accelerate the evolution of chosen genes under selection in yeast. When applied to a maladapted biosynthetic enzyme, we obtained collections of extensively diverged sequences with patterns that revealed structural and environmental constraints shaping the enzyme's activity. Our upgraded OrthoRep systems should support the discovery of factors influencing gene evolution, uncover previously unknown regions of fitness landscapes, and find broad applications in biomolecular engineering.

Divergent Tandem Acyl Carrier Proteins Necessitate In-Series Polyketide Processing in the Leinamycin Family.

The leinamycin family of polyketides are promising antitumor antibiotics, yet several aspects of their biosynthesis remain elusive. All leinamycin family members bear a sulfur-containing moiety which is essential for the anticancer activity exhibited by leinamycin. The key building blocks required for the incorporation of these functionalities are introduced in the final module of the polyketide synthase (PKS), which elegantly combines β-branching and thiocysteine incorporation to generate a diverse library of sulfur-based molecular scaffolds. Two acyl carrier proteins (ACPs) form a key didomain component of this module, but their amino acid sequence divergence has brought into question the common notion of functional equivalence. Here, we provide unprecedented functional evidence that these tandem ACPs play distinct roles in the final module of polyketide assembly. Using the weishanmycin biosynthetic pathway as a template, the in vitro reconstitution of key polyketide chain extension and β-branching steps in this module has revealed strict functional selectivity for a single ACP. Furthermore, we propose a cryptic transacylation step must occur prior to polyketide off-loading and cyclization. Altogether, these mechanistic investigations suggest that an atypical in-series mechanism underpins sulfur incorporation in the leinamycin family, and provides significant progress towards delineating their late-stage assembly.

Divergent Tandem Acyl Carrier Proteins Necessitate In‐Series Polyketide Processing in the Leinamycin Family

AbstractThe leinamycin family of polyketides are promising antitumor antibiotics, yet several aspects of their biosynthesis remain elusive. All leinamycin family members bear a sulfur‐containing moiety which is essential for the anticancer activity exhibited by leinamycin. The key building blocks required for the incorporation of these functionalities are introduced in the final module of the polyketide synthase (PKS), which elegantly combines β‐branching and thiocysteine incorporation to generate a diverse library of sulfur‐based molecular scaffolds. Two acyl carrier proteins (ACPs) form a key didomain component of this module, but their amino acid sequence divergence has brought into question the common notion of functional equivalence. Here, we provide unprecedented functional evidence that these tandem ACPs play distinct roles in the final module of polyketide assembly. Using the weishanmycin biosynthetic pathway as a template, the in vitro reconstitution of key polyketide chain extension and β‐branching steps in this module has revealed strict functional selectivity for a single ACP. Furthermore, we propose a cryptic transacylation step must occur prior to polyketide off‐loading and cyclization. Altogether, these mechanistic investigations suggest that an atypical in‐series mechanism underpins sulfur incorporation in the leinamycin family, and provides significant progress towards delineating their late‐stage assembly.

DNA sequencing of topotypes helps delineate species distributions in the Ischnocnema verrucosa complex (Anura, Brachycephalidae)

As many new evolutionary lineages are being discovered and formally named, sequencing topotypes when holotypes are not available becomes essential for taxonomy. This study uses a DNA-taxonomy approach to sequence new populations of the Ischnocnema verrucosa species complex (Brazilian Wart Frogs) from different locations, including, for the first time, individuals from the type localities. Phylogenetic analysis of the mitochondrial 16S gene recovered a monophyletic Ischnocnema verrucosa species series composed of three main clades. The most recent common ancestor was estimated to be 33.76 million years ago, and diversification within the three main clades occurred primarily during the Miocene. We delimited eight species-level lineages with high levels of sequence divergence (7% to 16%). Our study highlights the importance of DNA taxonomy and the necessity of protecting and sequencing topotypes in taxonomic studies. Our study also contributes to the conservation and understanding of the genus Ischnocnema and the biodiversity of the Brazilian Atlantic Forest region.

Sheeppox virus genome sequences from the European outbreaks in Spain, Bulgaria, and Greece in 2022–2024

In 2022-2024, three outbreaks of sheeppox (SPP) were reported in the European Union. These occurred in Spain, Bulgaria, and Greece and had serious economic consequences due to animal losses and trade restrictions. Five sheeppox virus (SPPV) whole-genome sequences (WGSs) were determined from samples collected during these outbreaks and analyzed in the context of all other published WGSs. Sheeppox virus strains can be divided in two, or possibly three, main groups. The isolates from the recent outbreaks belong to clade A2, which includes strains historically circulating in the Middle East and Northern Africa. Sequence divergence was low among the isolates that caused the recent European outbreaks. These results highlight the need for more regular and dense surveillance in under-sampled areas and the use of WGS to increase the chance of pinpointing the origin of an introduction, identifying potential introduction routes, and providing insights into SPPV evolution.

Convergence and divergence of diploid and tetraploid cotton genomes.

Polyploidy is an important driving force in speciation and evolution; however, the genomic basis for parallel selection of a particular trait between polyploids and ancestral diploids remains unexplored. Here we construct graph-based pan-genomes for diploid (A2) and allotetraploid (AD1) cotton species, enabled by an assembly of 50 genomes of genetically diverse accessions. We delineate a mosaic genome map of tetraploid cultivars that illustrates genomic contributions from semi-wild forms into modern cultivars. Pan-genome comparisons identify syntenic and hyper-divergent regions of continued variation between diploid and tetraploid cottons, and suggest an ongoing process of sequence evolution potentially linked to the contrasting genome size change in two subgenomes. We highlight 43% of genetic regulatory relationships for gene expression in diploid encompassing sequence divergence after polyploidy, and specifically characterize six underexplored convergent genetic loci contributing to parallel selection of fiber quality. This study offers a framework for pan-genomic dissection of genetic regulatory components underlying parallel selection of desirable traits in organisms.

Vishniacozyma floricola sp. nov., a flower-related tremellomycetous yeast species from Europe.

During the course of two independent studies conducted in Hungary and Spain, four conspecific yeast strains were isolated from flowers of different plant species. DNA sequences of two barcoding regions, the D1/D2 domain of the LSU rRNA gene and the internal transcribed spacer (ITS) region (ITS1-5.8S rRNA gene-ITS2), revealed that the four strains represent an undescribed Vishniacozyma (family Bulleribasidiaceae, Basidiomycota) species. In terms of pairwise sequence similarities and according to our phylogenetic analyses of the concatenated DNA sequences of the ITS region and the D1/D2 domain of the LSU rRNA gene, the undescribed species is most closely related to Vishniacozyma melezitolytica, a yeast species of phylloplane origin. The novel species differs from the type strain of V. melezitolytica by 8 substitutions and 3 insertion/deletion (indels) and 11 substitutions and 5 indels along the D1/D2 domain of the LSU rRNA gene and the ITS region, respectively. In addition to the DNA sequence divergences, the two species differ in some physiological characters as well. We propose the species Vishniacozyma floricola sp. nov. to accommodate the above-noted strains (holotype, NCAIM Y.02320; isotype, CBS 18939; MycoBank number, 856028).