Jordi Morata

Introduction: Brown trout (Salmo trutta L.) is a salmonid fish considered a genetically and geographically highly structured species. The species is catalogued as vulnerable in the Spanish Red List since 1992. Based on morphological differentiation, some authors have proposed splitting Salmo trutta into many taxa and in recent years, several binomials involving morphologically differentiated Western Mediterranean brown trout morphs have regained relevance as distinct species and are now assessed in the IUCN Red List. However, Spanish legislation still considers a single species, S. trutta, inhabiting Atlantic and Mediterranean rivers. Analyses of mitochondrial DNA variation across the species’ native range have revealed numerous matrilineal lineages and helped disentangle the phylogenetic relationships and biogeographic history of brown trout. Four largely distant evolutionary lineages defined by sequencing the mitochondrial control region (Adriatic, AD, Mediterranean, ME, Atlantic, AT and Duero, DU) are native among the Iberian brown trout populations. Stocking activities have resulted in introgressive hybridization of these native populations with non-native northern Atlantic stocks throughout the Iberian rivers and compromise the evolutionary singularities of brown trout in the Iberian Peninsula. Methodology: In this study, we compare complete mitogenomes from Iberian brown trout specimens with those from other regions, using both newly sequenced samples and whole mitogenomes from the GenBank database and estimate their divergence times relative to those observed among species of other Salmoninae genera, such as Oncorhynchus and Salvelinus, to assess patterns of species-specific divergence among Iberian brown trout populations. Results: The evolutionary relationships of the specimens using their mitogenomes fitted with previous analyses using partial mitochondrial sequences and confirmed the strong differentiation among the Iberian lineages. All the branches involving distant Iberian mitogenomes are sister branches with taxa historically described under different binomials (such as Salmo macrostigma, S. multipunctatus, S. pallaryi). Conclusions: The results provide evidence supporting the existence of potentially distinct and undescribed native Salmo species in the Iberian rivers.

Two new diagnostic classifications of acute myeloid leukemia (AML) were published in 2022 to update current knowledge on disease biology. In previous 2017-edition categories of AML with myelodysplasia-related changes, AML was not otherwise specified, but AML with mutated RUNX1 experienced profound changes. We performed whole exome sequencing on a cohort of 69 patients with cytogenetic intermediate-risk AML that belonged to these diagnostic categories to correlate their mutational pattern and copy-number alterations with their new diagnostic distribution. Our results show that 45% of patients changed their diagnostic category, being AML myelodysplasia-related the most enlarged, mainly due to a high frequency of myelodysplasia-related mutations (58% of patients). These showed a good correlation with multilineage dysplasia and/or myelodysplastic syndrome history, but at the same time, 21% of de novo patients without dysplasia also presented them. RUNX1 was the most frequently mutated gene, with a high co-occurrence rate with other myelodysplasia-related mutations. We found a high prevalence of copy-neutral loss of heterozygosity, frequently inducing a homozygous state in particular mutated genes. Mild differences in current classifications explain the diagnostic disparity in 10% of patients, claiming a forthcoming unified classification.

Germ line predisposition in acute myeloid leukemia (AML) has gained attention in recent years because of a nonnegligible frequency and an impact on management of patients and their relatives. Risk alleles for AML development may be present in patients without a clinical suspicion of hereditary hematologic malignancy syndrome. In this study we investigated the presence of germ line variants (GVs) in 288 genes related to cancer predisposition in 47 patients with available paired, tumor-normal material, namely bone marrow stroma cells (n = 29), postremission bone marrow (n = 17), and saliva (n = 1). These patients correspond to 2 broad AML categories with heterogeneous genetic background (AML myelodysplasia related and AML defined by differentiation) and none of them had phenotypic abnormalities, previous history of cytopenia, or strong cancer aggregation. We found 11 pathogenic or likely pathogenic variants, 6 affecting genes related to autosomal dominant cancer predisposition syndromes (ATM, DDX41, and CHEK2) and 5 related to autosomal recessive bone marrow failure syndromes (FANCA, FANCM, SBDS, DNAJC21, and CSF3R). We did not find differences in clinical characteristics nor outcome between carriers of GVs vs noncarriers. Further studies in unselected AML cohorts are needed to determine GV incidence and penetrance and, in particular, to clarify the role of ATM nonsense mutations in AML predisposition.

Antimicrobial peptides (AMPs) are widely distributed molecules secreted mostly by cells of the innate immune system to prevent bacterial proliferation at the site of infection. As with classic antibiotics, continued treatment with AMPs can create resistance in bacteria. However, whether AMPs can generate tolerance as an intermediate stage towards resistance is not known. Here, we show that the treatment of Escherichia coli with different AMPs induces tolerance by lag, particularly for those peptides that have internal targets. This tolerance can be detected as different morphological and physiological changes, which depend on the type of peptide molecule the bacterium has been exposed to. In addition, we show that AMP tolerance can also affect antibiotic treatment. The genomic sequencing of AMP-tolerant strains shows that different mutations alter membrane composition, DNA replication, and translation. Some of these mutations have also been observed in antibiotic-resistant strains, suggesting that AMP tolerance could be a relevant step in the development of antibiotic resistance. Monitoring AMP tolerance is relevant vis-á-vis the eventual therapeutic use of AMPs and because cross-tolerance might favor the emergence of resistance against conventional antibiotic treatments.

Antimicrobial peptides (AMPs) are widely distributed molecules secreted mostly by cells of the innate immune system to prevent bacterial proliferation at the site of infection. As with classic antibiotics, continued treatment with AMPs can create resistance in bacteria. However, whether AMPs can generate tolerance as an intermediate stage towards resistance is not known. Here we show that treatment of Escherichia coli with different AMPs induces tolerance by lag, particularly for those peptides that have internal targets. This tolerance can be detected as different morphological and physiological changes, which depend on the type of peptide molecule the bacterium has been exposed to. In addition, we show that AMP tolerance can also affect antibiotic treatment. Genomic sequencing of AMP-tolerant strains shows that different mutations alter membrane composition, DNA replication, and translation. Some of these mutations have also been observed in antibiotic-resistant strains, suggesting that AMP tolerance could be a relevant step in the development of antibiotic resistance. Monitoring AMP tolerance is relevant with a view to eventual therapeutic use of AMPs and because cross-tolerance might favor the emergence of resistance against conventional antibiotic treatments.

Summary: The precisionFDA Truth Challenge V2 aimed to assess the state-of-the-art of variant calling in difficult-to-map regions and the Major Histocompatibility Complex (MHC). Starting with FASTQ files, 20 challenge participants applied their variant calling pipelines and submitted 64 variant callsets for one or more sequencing technologies (~35X Illumina, ~35X PacBio HiFi, and ~50X Oxford Nanopore Technologies). Submissions were evaluated following best practices for benchmarking small variants with the new GIAB benchmark sets and genome stratifications. Challenge submissions included a number of innovative methods for all three technologies, with graph-based and machine-learning methods scoring best for short-read and long-read datasets, respectively. New methods out-performed the 2016 Truth Challenge winners, and new machine-learning approaches combining multiple sequencing technologies performed particularly well. Recent developments in sequencing and variant calling have enabled benchmarking variants in challenging genomic regions, paving the way for the identification of previously unknown clinically relevant variants.

Eukaryotic genome replication depends on thousands of DNA replication origins (ORIs). A major challenge is to learn ORI biology in multicellular organisms in the context of growing organs to understand their developmental plasticity. We have identified a set of ORIs of Arabidopsis thaliana and their chromatin landscape at two stages of post-embryonic development. ORIs associate with multiple chromatin signatures including transcription start sites (TSS) but also proximal and distal regulatory regions and heterochromatin, where ORIs colocalize with retrotransposons. In addition, quantitative analysis of ORI activity led us to conclude that strong ORIs have high GC content and clusters of GGN trinucleotides. Development primarily influences ORI firing strength rather than ORI location. ORIs that preferentially fire at early developmental stages colocalize with GC-rich heterochromatin, but at later stages with transcribed genes, perhaps as a consequence of changes in chromatin features associated with developmental processes. Our study provides the set of ORIs active in an organism at the post-embryo stage that should allow us to study ORI biology in response to development, environment, and mutations with a quantitative approach. In a wider scope, the computational strategies developed here can be transferred to other eukaryotic systems.

Transposable elements are one of the main drivers of plant genome evolution. Transposon insertions can modify the gene coding capacity or the regulation of their expression, the latter being a more subtle effect, and therefore particularly useful for evolution. Transposons have been show to contain transcription factor binding sites that can be mobilized upon transposition with the potential to integrate new genes into transcriptional networks. Miniature inverted-repeat transposable elements (MITEs) are a type of noncoding DNA transposons that could be particularly suited as a vector to mobilize transcription factor binding sites and modify transcriptional networks during evolution. MITEs are small in comparison to other transposons and can be excised, which should make them less mutagenic when inserting into promoters. On the other hand, in spite of their cut-and-paste mechanisms of transposition, they can reach very high copy numbers in genomes. We have previously shown that MITEs have amplified and redistributed the binding motif of the E2F transcription factor in different Brassicas. Here, we show that MITEs have amplified and mobilized the binding motifs of the bZIP60 and PIF3 transcription factors in peach and Prunus mume, and the TCP15/23 binding motif in tomato. Our results suggest that MITEs could have rewired new genes into transcriptional regulatory networks that are responsible for important adaptive responses and breeding traits in plants, such as stress responses, flowering time, or fruit ripening. The results presented here therefore suggest a general impact of MITEs in the evolution of transcriptional regulatory networks in plants.