Deep phylogenetic-based clustering analysis uncovers new and shared mutations in SARS-CoV-2 variants as a result of directional and convergent evolution

Nearly two decades after the last epidemic caused by a severe acute respiratory syndrome coronavirus (SARS-CoV), newly emerged SARS-CoV-2 quickly spread in 2020 and precipitated an ongoing global public health crisis. Both the continuous accumulation of point mutations, owed to the naturally imposed genomic plasticity of SARS-CoV-2 evolutionary processes, as well as viral spread over time, allow this RNA virus to gain new genetic identities, spawn novel variants and enhance its potential for immune evasion. Here, through an in-depth phylogenetic clustering analysis of upwards of 200,000 whole-genome sequences, we reveal the presence of not previously reported and hitherto unidentified mutations and recombination breakpoints in Variants of Concern (VOC) and Variants of Interest (VOI) from Brazil, India (Beta, Eta and Kappa) and the USA (Beta, Eta and Lambda). Additionally, we identify sites with shared mutations under directional evolution in the SARS-CoV-2 Spike-encoding protein of VOC and VOI, tracing a heretofore-undescribed correlation with viral spread in South America, India and the USA. Our evidence-based analysis provides well-supported evidence of similar pathways of evolution for such mutations in all SARS-CoV-2 variants and sub-lineages. This raises two pivotal points: the co-circulation of variants and sub-lineages in close evolutionary environments, which sheds light onto their trajectories into convergent and directional evolution (i), and a linear perspective into the prospective vaccine efficacy against different SARS-CoV-2 strains (ii).

New Avenues for Old Travellers: Phenotypic Evolutionary Trends Meet Morphodynamics, and Both Enter the Global Change Biology Era

Evolutionary trends (ETs) are traditionally defined as substantial changes in the state of traits through time produced by a persistent condition of directional evolution. ETs might also include directional responses to ecological, climatic or biological gradients and represent the primary evolutionary pattern at high taxonomic levels and over long-time scales. The absence of a well-supported operative definition of ETs blurred the definition of conceptual differences between ETs and other key concepts in evolution such as convergence, parallel evolution, and divergence. Also, it prevented the formulation of modern guidelines for studying ETs and evolutionary dynamics related to them. In phenotypic evolution, the theory of morphodynamics states that the interplay between evolutionary factors such as phylogeny, evo-devo constraints, environment, and biological function determines morphological evolution. After introducing a new operative definition, here we provide a morphodynamics-based framework for studying phenotypic ETs, discussing how understanding the impact of these factors on ETs improves the explanation of links between biological patterns and processes underpinning directional evolution. We envisage that adopting a quantitative, pattern-based, and multifactorial approach will pave the way to new potential applications for this field of evolutionary biology. In this framework, by exploiting the catalysing effect of climate change on evolution, research on ETs induced by global change might represent an ideal arena for validating hypotheses about the predictability of evolution.

Construction and directional evolution of anti-enrofloxacin ScFv antibody for the immunoassay of fluoroquinolones

A recombinant anti-enrofloxacin single-chain antibody (scFv) was produced for the detection of enrofloxacin. An immunized mouse phage display scFv library with a capacity of 2.35×109 CFU/mL was constructed and used for anti-enrofloxacin scFv screening. After four rounds of bio-panning, 10 positives were isolated and identified successfully. The highest positive scFv was expressed in E. coli BL21. Then, its recognition mechanisms were studied using the molecular docking method. The result showed the amino acid residues Leu121 were the key residues for the binding of ScFv to ENR. Based on the results of virtual mutation, the ScFv antibody was evolved by directional mutagenesis of contact amino acid residue Leu121 to Asn. After the expression and purification, an indirect competitive enzyme-linked immunosorbent assay (IC-ELISA) based on the parental and mutant ScFv were established for enrofloxacin respectively. The IC50 value of the assay established with the ScFv mutant was 1.63 ng/mL, while the parental ScFv was 21.08 ng/mL, this result showed highly increased affinity with up to 12.9-folds improved sensitivity. The mean recovery for ENR ranged from 71.80% to 117.35% with 10.46% relative standard deviation between the intra-assay and the inter-assay. The results indicate that we have obtained a highly sensitive anti-ENR scFv by the phage library construction and directional evolution, and the scFv-based IC-ELISA is suitable for the detection of ENR residue in animal derived edible tissues and milk.

Construction and directional evolution of anti-enrofloxacin ScFv antibody for the immunoassay of fluoroquinolones

A recombinant anti-enrofloxacin single-chain antibody (scFv) was produced for the detection of enrofloxacin. An immunized mouse phage display scFv library with a capacity of 2.35×109 CFU/mL was constructed and used for anti-enrofloxacin scFv screening. After four rounds of bio-panning, 10 positives were isolated and identified successfully. The highest positive scFv was expressed in E. coli BL21. Then, its recognition mechanisms were studied using the molecular docking method. The result showed the amino acid residues Leu121 were the key residues for the binding of ScFv to ENR. Based on the results of virtual mutation, the ScFv antibody was evolved by directional mutagenesis of contact amino acid residue Leu121 to Asn. After the expression and purification, an indirect competitive enzyme-linked immunosorbent assay (IC-ELISA) based on the parental and mutant ScFv were established for enrofloxacin respectively. The IC50 value of the assay established with the ScFv mutant was 1.63 ng/mL, while the parental ScFv was 21.08 ng/mL, this result showed highly increased affinity with up to 12.9-folds improved sensitivity. The mean recovery for ENR ranged from 71.80% to 117.35% with 10.46% relative standard deviation between the intra-assay and the inter-assay. The results indicate that we have obtained a highly sensitive anti-ENR scFv by the phage library construction and directional evolution, and the scFv-based IC-ELISA is suitable for the detection of ENR residue in animal derived edible tissues and milk.

Genomic recombination events may reveal the evolution of coronavirus and the origin of SARS-CoV-2

To trace the evolution of coronaviruses and reveal the possible origin of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which causes the coronavirus disease 2019 (COVID-19), we collected and thoroughly analyzed 29,452 publicly available coronavirus genomes, including 26,312 genomes of SARS-CoV-2 strains. We observed coronavirus recombination events among different hosts including 3 independent recombination events with statistical significance between some isolates from humans, bats and pangolins. Consistent with previous records, we also detected putative recombination between strains similar or related to Bat-CoV-RaTG13 and Pangolin-CoV-2019. The putative recombination region is located inside the receptor-binding domain (RBD) of the spike glycoprotein (S protein), which may represent the origin of SARS-CoV-2. Population genetic analyses provide estimates suggesting that the putative introduced DNA within the RBD is undergoing directional evolution. This may result in the adaptation of the virus to hosts. Unsurprisingly, we found that the putative recombination region in S protein was highly diverse among strains from bats. Bats harbor numerous coronavirus subclades that frequently participate in recombination events with human coronavirus. Therefore, bats may provide a pool of genetic diversity for the origin of SARS-CoV-2.

The genomic recombination events may reveal the evolution of coronavirus and the origination of 2019-nCoV

To trace the evolution of coronavirus and reveal the possible origination of the novel pneumonia coronavirus (2019-nCoV), we collected and thoroughly analyzed 2966 publicly available coronavirus genomes, including 182 2019-nCoVs strains. We observed 3 independent recombination events with statistical significance between some isolates from bats and pangolins. In consistence with previous records, we also detected the putative recombination between Bat-CoV-RaTG13 and Pangolin-CoV-2019 covering the receptor bind domain (RBD) of the spike glycoprotein (S protein), which may lead to the origination of 2019-nCoV. Population genetic analyses give estimations indicating that the recombinant region around RBD is possibly undergoing directional evolution. This may result to the adaption of the virus to be infectious in hosts. Not surprisingly, we find that the S protein of coronavirus keeps high diversity among bat isolates, which may provide a genetic pool for the origination of 2019-nCoV.

GENE-52. ANALYSIS OF RECURRENT MUTATIONS WITHIN CANCERS REVEALS PATTERNS OF DIRECTIONAL EVOLUTION IN BRAIN TUMORS

INTRODUCTION Intratumor genetic heterogeneity is commonly caused by a stochastic evolution, but can also show remarkable selectivity resulting in directional evolution. We hypothesized that directional evolution is driven by its genetic onset-stage. This process could enforce already existing traits resulting in an enhancement of linear pathways rather than the complementary/parallel evolution. METHODS We generated a prediction model for directional evolution based on its genetic onset-stage. We performed a comprehensive analysis of 16 different tumor types of ~10,000 patients of whole-exome sequencing and copy number variation, obtained from The Cancer Genome Atlas. RESULTS We found in 5% of the patients that directional evolution occurs on a fixed genetic onset stage. There is a strong relationship between the frequencies of directional evolution and the frequencies of commonly co-occurring mutations in the onset stage. These co-occurring mutations are frequently observed in the same chromosomal region and occasionally in the same pathway. Brain tumors show a frequent directional evolution towards EGFR mutations and to a lesser extent PIK3CA mutations. CONCLUSION Given these strong correlations and since directional evolution is apparently a highly selective process, our prediction model could predict more effective therapies by choosing therapies targeting both the mutated gene that is affected by directional evolution as well as the commonly occurring co-mutated genes.