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Theileria parasites are known to induce the transformation of host bovine leukocytes, involved in rapid proliferation, evasion from apoptotic mechanisms, and increased dissemination. In this study, we reveal the involvement of m 6 A RNA modification in T. annulata infection-induced transformation of bovine leukocytes. We conducted m 6 A sequencing and bioinformatics analysis to map the mRNA methylation patterns of T. annulata-infected host leukocytes. We observe specific mRNA modifications for T. annulata-infected leukocytes and a strong correlation between the proliferation rate of the infected Leukocytes with m 6 A modifications We observe that the increased amounts of m 6 A seem to impact some cell cycle dynamics, potentially via modifications of E2F4 mRNA. Moreover, we further identify HIF-1α as a possible driver of these m 6 A RNA modifications that have clear relevance to cellular proliferation dynamics. Overall, our results provide insights into the role of m6A mRNA methylation in the molecular crosstalk between Theileria and their host leukocytes, emphasizing the critical role of mRNA methylation in host-parasite interaction., (Copyright © 2025 The Authors. Published by Elsevier B.V. All rights reserved.)

In microelectronics, one of the main 3D integration strategies consists of vertically stacking and electrically connecting various functional chips using through-silicon vias (TSVs). For the fabrication of the TSVs, one of the challenges is to conformally deposit a low dielectric constant insulator thin film at the surface of the silicon. To date, there is no universal technique that can address all types of TSV integration schemes, especially in the case requiring a low deposition temperature. In this work, an organosilicate polymer deposited by initiated chemical vapor deposition (iCVD) was developed and integrated as an insulating layer for TSVs. Process studies have shown that poly(1,3,5-trivinyl-1,3,5-trimethyl cyclotrisiloxane) (P(V 3 D 3 )) can present good conformality on high aspect ratio features by increasing the substrate temperature up to 100 °C. The trade-off is a moderate deposition rate. The thermal stability of the polymer has been investigated, and we show that a thermal annealing at 400 °C (with or without ultraviolet exposure) allows the stabilization of the dielectric films by removing residual oligomers. Then, P(V 3 D 3 ) was integrated in high aspect ratio TSV (10 × 100 μm) on 300 mm silicon wafers using a standard integration flow for TSV metallization. Functional devices were successfully fabricated (including daisy chains of 754 TSVs) and electrically characterized. Our work shows that the metallization barrier should be carefully selected to eliminate the appearance of voids at the top corner of the TSV after the Cu annealing step. Moreover, an appropriate integration process should be used to avoid the appearance of cohesive cracks in the liner. This work constitutes a first proof of concept of the use of an iCVD polymer in a quasi-industrial microelectronic environment. It also highlights the benefit of iCVD as a promising technique to deposit conformal dielectric thin films in a microelectronic pilot line environment.

The organelle paralogy hypothesis (OPH) aims to explain the evolution of non-endosymbiotically derived organelles. It predicts that lineage-specific pathways or organelles should result when identity-encoding membrane-trafficking components duplicate and co-evolve. Here, we investigate the presence of such lineage-specific membrane-trafficking machinery paralogs in Apicomplexa, a globally important parasitic lineage. We are able to identify 18 paralogs of known membrane-trafficking machinery, in several cases co-incident with the presence of new endomembrane organelles in apicomplexans or their parent lineage, the Alveolata. Moreover, focused analysis of the apicomplexan Arf-like small GTPases (i.e., ArlX3) revealed a specific post-Golgi trafficking pathway. This pathway appears involved in delivery of proteins to micronemes and rhoptries, with knockdown demonstrating reduced invasion capacity. Overall, our data have identified an unforeseen post-Golgi trafficking pathway in apicomplexans and are consistent with the OPH mechanism acting to produce endomembrane pathways or organelles at various evolutionary stages across the alveolate lineage., Competing Interests: Declaration of interests The authors have no competing interests., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Malaria-associated pathogenesis such as parasite invasion, egress, host cell remodelling and antigenic variation requires concerted action by many proteins, but the molecular regulation is poorly understood. Here we have characterized an essential Plasmodium-specific Apicomplexan AP2 transcription factor in Plasmodium falciparum (PfAP2-P; pathogenesis) during the blood-stage development with two peaks of expression. An inducible knockout of gene function showed that PfAP2-P is essential for trophozoite development, and critical for var gene regulation, merozoite development and parasite egress. Chromatin immunoprecipitation sequencing data collected at timepoints matching the two peaks of pfap2-p expression demonstrate PfAP2-P binding to promoters of genes controlling trophozoite development, host cell remodelling, antigenic variation and pathogenicity. Single-cell RNA sequencing and fluorescence-activated cell sorting revealed de-repression of most var genes in Δpfap2-p parasites. Δpfap2-p parasites also overexpress early gametocyte marker genes, indicating a regulatory role in sexual stage conversion. We conclude that PfAP2-P is an essential upstream transcriptional regulator at two distinct stages of the intra-erythrocytic development cycle., (© 2023. The Author(s).)

Background: The excessive inflammatory responses provoked by SARS-CoV-2 infection are critical factors affecting the severity and mortality of COVID-19. Previous work found that two adjacent co-occurring mutations R203K and G204R (KR) on the nucleocapsid (N) protein correlate with increased disease severity in COVID-19 patients. However, links with the host immune response remain unclear., Methods: Here, we grouped nasopharyngeal swab samples of COVID-19 patients into two cohorts based on the presence and absence of SARS-CoV-2 nucleocapsid KR mutations. We performed nasopharyngeal transcriptome analysis of age, gender, and ethnicity-matched COVID-19 patients infected with either SARS-CoV-2 with KR mutations in the N protein (KR patients n = 39) or with the wild-type N protein (RG patients n = 39) and compared to healthy controls (n = 34). The impact of KR mutation on immune response was further characterized experimentally by transcriptomic and proteomic profiling of virus-like-particle (VLP) incubated cells., Results: We observed markedly elevated expression of proinflammatory cytokines, chemokines, and interferon-stimulated (ISGs) genes in the KR patients compared to RG patients. Using nasopharyngeal transcriptome data, we found significantly higher levels of neutrophils and neutrophil-to-lymphocyte (NLR) ratio in KR patients than in the RG patients. Furthermore, transcriptomic and proteomic profiling of VLP incubated cells confirmed a similar hyper-inflammatory response mediated by the KR variant., Conclusions: Our data demonstrate an unforeseen connection between nucleocapsid KR mutations and augmented inflammatory immune response in severe COVID-19 patients. These findings provide insights into how mutations in SARS-CoV-2 modulate host immune output and pathogenesis and may contribute to more efficient therapeutics and vaccine development., (© 2023. The Author(s).)

Malaria pathogenicity results from the parasite's ability to invade, multiply within and then egress from the host red blood cell (RBC). Infected RBCs are remodeled, expressing antigenic variant proteins (such as PfEMP1, coded by the var gene family) for immune evasion and survival. These processes require the concerted actions of many proteins, but the molecular regulation is poorly understood. We have characterized an essential Plasmodium specific Apicomplexan AP2 (ApiAP2) transcription factor in Plasmodium falciparum (PfAP2-MRP; Master Regulator of Pathogenesis) during the intraerythrocytic developmental cycle (IDC). An inducible gene knockout approach showed that PfAP2-MRP is essential for development during the trophozoite stage, and critical for var gene regulation, merozoite development and parasite egress. ChIP-seq experiments performed at 16 hour post invasion (h.p.i.) and 40 h.p.i. matching the two peaks of PfAP2-MRP expression, demonstrate binding of PfAP2-MRP to the promoters of genes controlling trophozoite development and host cell remodeling at 16 h.p.i. and antigenic variation and pathogenicity at 40 h.p.i. Using single-cell RNA-seq and fluorescence-activated cell sorting, we show de-repression of most var genes in Δpfap2-mrp parasites that express multiple PfEMP1 proteins on the surface of infected RBCs. In addition, the Δpfap2-mrp parasites overexpress several early gametocyte marker genes at both 16 and 40 h.p.i., indicating a regulatory role in the sexual stage conversion. Using the Chromosomes Conformation Capture experiment (Hi-C), we demonstrate that deletion of PfAP2-MRP results in significant reduction of both intra-chromosomal and inter-chromosomal interactions in heterochromatin clusters. We conclude that PfAP2-MRP is a vital upstream transcriptional regulator controlling essential processes in two distinct developmental stages during the IDC that include parasite growth, chromatin structure and var gene expression.

MicroRNAs (miRNAs) are small noncoding RNAs that can play critical roles in regulating various cellular processes, including during many parasitic infections. Here, we report a regulatory role for miR-34c-3p in cAMP-independent regulation of host cell protein kinase A (PKA) activity in Theileria annulata -infected bovine leukocytes. We identified prkar2b (cAMP-dependent protein kinase A type II-beta regulatory subunit) as a novel miR-34c-3p target gene and demonstrate how infection-induced upregulation of miR-34c-3p repressed PRKAR2B expression to increase PKA activity. As a result, the disseminating tumorlike phenotype of T. annulata -transformed macrophages is enhanced. Finally, we extend our observations to Plasmodium falciparum-parasitized red blood cells, where infection-induced augmentation in miR-34c-3p levels led to a drop in the amount of prkar2b mRNA and increased PKA activity. Collectively, our findings represent a novel cAMP-independent way of regulating host cell PKA activity in infections by Theileria and Plasmodium parasites. IMPORTANCE Small microRNA levels are altered in many diseases, including those caused by parasites. Here, we describe how infection by two important animal and human parasites, Theileria annulata and Plasmodium falciparum, induce changes in infected host cell miR-34c-3p levels to regulate host cell PKA kinase activity by targeting mammalian prkar2b . Infection-induced changes in miR-34c-3p levels provide a novel epigenetic mechanism for regulating host cell PKA activity independent of fluxes in cAMP to both aggravate tumor dissemination and improve parasite fitness.

The outcome of transplant recipients is variable depending on the study population, vaccination status and COVID-19 variants. Our aim was to study the impact of Omicron subvariants on the mortality of transplant recipients. We reviewed the results of SARS-CoV-2 whole genome sequence of random isolates collected from 29 December 2021 until 17 May 2022 in King Faisal Specialist Hospital and Research center, Jeddah (KFSHRC-J), Saudi Arabia performed as hospital genomic surveillance program for COVID-19 variants. We included 25 transplant patients infected with confirmed Omicron variants.17 (68%) and 8 (32%) patients had Omicron BA.1 and BA.2, respectively. 12 (68%) patients had renal transplants. Only 36% of patients received three doses of COVID-19 vaccines. 23 (92%) patients required hospitalization. 20 (80%) patients survived and 6 (25%) required intensive care unit (ICU) admission. Among ICU patients, 66.7% were more than 50 years, 50% had two to three comorbidities and 5 out of 6 (83%) died. The mortality of transplant patients infected with Omicron variants in our cohort was higher than other centers as a limited number of patients received booster vaccines. Optimizing booster vaccination is the most efficient method to improve the mortality of COVID-19 in transplant recipients recognizing the inefficacy of monoclonal antibodies in the presence of SARS-CoV-2 emerging variants. We did not show a difference in mortality in transplant patients infected with Omicron BA.1 and BA.2 knowing the limitation of our sample size., (© 2023. The Author(s).)

A fungal metabolite, FR235222, specifically inhibits a histone deacetylase of the apicomplexan parasite Toxoplasma gondii and TgHDAC3 has emerged as a key factor regulating developmental stage transition in this species. Here, we exploited FR235222 to ask if changes in histone acetylation regulate developmental stage transition of Theileria annulata, another apicomplexan species. We found that FR235222 treatment of T. annulata-infected transformed leukocytes induced a proliferation arrest. The blockade in proliferation was due to drug-induced conversion of intracellular schizonts to merozoites that lack the ability to maintain host leukocyte cell division. Induction of merogony by FR235222 leads to an increase in expression of merozoite-marker (rhoptry) proteins. RNA-seq of FR235222-treated T. annulata-infected B cells identified deregulated expression of 468 parasite genes including a number encoding parasite ApiAP2 transcription factors. Thus, similar to T. gondii, FR235222 inhibits T. annulata HDAC (TaHDAC1) activity and places parasite histone acetylation as a major regulatory event of the transition from schizonts to merozoites., (© 2022. The Author(s).)

Monitoring SARS-CoV-2 spread and evolution through genome sequencing is essential in handling the COVID-19 pandemic. Here, we sequenced 892 SARS-CoV-2 genomes collected from patients in Saudi Arabia from March to August 2020. We show that two consecutive mutations (R203K/G204R) in the nucleocapsid (N) protein are associated with higher viral loads in COVID-19 patients. Our comparative biochemical analysis reveals that the mutant N protein displays enhanced viral RNA binding and differential interaction with key host proteins. We found increased interaction of GSK3A kinase simultaneously with hyper-phosphorylation of the adjacent serine site (S206) in the mutant N protein. Furthermore, the host cell transcriptome analysis suggests that the mutant N protein produces dysregulated interferon response genes. Here, we provide crucial information in linking the R203K/G204R mutations in the N protein to modulations of host-virus interactions and underline the potential of the nucleocapsid protein as a drug target during infection., (© 2022. The Author(s).)

Background: Plasmodium simium, a malaria parasite of non-human primates (NHP), was recently shown to cause zoonotic infections in humans in Brazil. We sequenced the P. simium genome to investigate its evolutionary history and to identify any genetic adaptions that may underlie the ability of this parasite to switch between host species., Results: Phylogenetic analyses based on whole genome sequences of P. simium from humans and NHPs reveals that P. simium is monophyletic within the broader diversity of South American Plasmodium vivax, suggesting P. simium first infected NHPs as a result of a host switch of P. vivax from humans. The P. simium isolates show the closest relationship to Mexican P. vivax isolates. Analysis of erythrocyte invasion genes reveals differences between P. vivax and P. simium, including large deletions in the Duffy-binding protein 1 (DBP1) and reticulocyte-binding protein 2a genes of P. simium. Analysis of P. simium isolated from NHPs and humans revealed a deletion of 38 amino acids in DBP1 present in all human-derived isolates, whereas NHP isolates were multi-allelic., Conclusions: Analysis of the P. simium genome confirmed a close phylogenetic relationship between P. simium and P. vivax, and suggests a very recent American origin for P. simium. The presence of the DBP1 deletion in all human-derived isolates tested suggests that this deletion, in combination with other genetic changes in P. simium, may facilitate the invasion of human red blood cells and may explain, at least in part, the basis of the recent zoonotic infections., (© 2021. The Author(s).)

DNA sequencing technologies provide unprecedented opportunities to analyze within-host evolution of microorganism populations. Often, within-host populations are analyzed via pooled sequencing of the population, which contains multiple individuals or "haplotypes." However, current next-generation sequencing instruments, in conjunction with single-molecule barcoded linked-reads, cannot distinguish long haplotypes directly. Computational reconstruction of haplotypes from pooled sequencing has been attempted in virology, bacterial genomics, metagenomics, and human genetics, using algorithms based on either cross-host genetic sharing or within-host genomic reads. Here, we describe PoolHapX, a flexible computational approach that integrates information from both genetic sharing and genomic sequencing. We demonstrated that PoolHapX outperforms state-of-the-art tools tailored to specific organismal systems, and is robust to within-host evolution. Importantly, together with barcoded linked-reads, PoolHapX can infer whole-chromosome-scale haplotypes from 50 pools each containing 12 different haplotypes. By analyzing real data, we uncovered dynamic variations in the evolutionary processes of within-patient HIV populations previously unobserved in single position-based analysis., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.)

The extensive sequence data generated from SARS-CoV-2 during the 2020 pandemic has facilitated the study of viral genome evolution over a brief period of time. This has highlighted instances of directional mutation pressures exerted on the SARS-CoV-2 genome from host antiviral defense systems. In this brief review we describe three such human defense mechanisms, the apolipoprotein B mRNA editing catalytic polypeptide-like proteins (APOBEC), adenosine deaminase acting on RNA proteins (ADAR), and reactive oxygen species (ROS), and discuss their potential implications on SARS-CoV-2 evolution., Competing Interests: Declaration of competing interest None declared., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

Apicomplexan parasites cause major human disease and food insecurity. They owe their considerable success to highly specialized cell compartments and structures. These adaptations drive their recognition, nondestructive penetration, and elaborate reengineering of the host's cells to promote their growth, dissemination, and the countering of host defenses. The evolution of unique apicomplexan cellular compartments is concomitant with vast proteomic novelty. Consequently, half of apicomplexan proteins are unique and uncharacterized. Here, we determine the steady-state subcellular location of thousands of proteins simultaneously within the globally prevalent apicomplexan parasite Toxoplasma gondii. This provides unprecedented comprehensive molecular definition of these unicellular eukaryotes and their specialized compartments, and these data reveal the spatial organizations of protein expression and function, adaptation to hosts, and the underlying evolutionary trajectories of these pathogens., Competing Interests: Declaration of Interests The authors declare no competing interests., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

Cell cycle transitions are generally triggered by variation in the activity of cyclin-dependent kinases (CDKs) bound to cyclins. Malaria-causing parasites have a life cycle with unique cell-division cycles, and a repertoire of divergent CDKs and cyclins of poorly understood function and interdependency. We show that Plasmodium berghei CDK-related kinase 5 (CRK5), is a critical regulator of atypical mitosis in the gametogony and is required for mosquito transmission. It phosphorylates canonical CDK motifs of components in the pre-replicative complex and is essential for DNA replication. During a replicative cycle, CRK5 stably interacts with a single Plasmodium -specific cyclin (SOC2), although we obtained no evidence of SOC2 cycling by transcription, translation or degradation. Our results provide evidence that during Plasmodium male gametogony, this divergent cyclin/CDK pair fills the functional space of other eukaryotic cell-cycle kinases controlling DNA replication., Competing Interests: AB, MZ, ER, CP, LB, TM, AS, NK, PA, RP, DB, SV, AH, AP, DF, AH, RT, MB No competing interests declared, (© 2020, Balestra et al.)

Kinesin-8 proteins are microtubule motors that are often involved in regulation of mitotic spindle length and chromosome alignment. They move towards the plus ends of spindle microtubules and regulate the dynamics of these ends due, at least in some species, to their microtubule depolymerization activity. Plasmodium spp. exhibit an atypical endomitotic cell division in which chromosome condensation and spindle dynamics in the different proliferative stages are not well understood. Genome-wide shared orthology analysis of Plasmodium spp. revealed the presence of two kinesin-8 motor proteins, kinesin-8X and kinesin-8B. Here we studied the biochemical properties of kinesin-8X and its role in parasite proliferation. In vitro, kinesin-8X has motility and depolymerization activities like other kinesin-8 motors. To understand the role of Plasmodium kinesin-8X in cell division, we used fluorescence-tagging and live cell imaging to define its location, and gene targeting to analyse its function, during all proliferative stages of the rodent malaria parasite P. berghei life cycle. The results revealed a spatio-temporal involvement of kinesin-8X in spindle dynamics and an association with both mitotic and meiotic spindles and the putative microtubule organising centre (MTOC). Deletion of the kinesin-8X gene revealed a defect in oocyst development, confirmed by ultrastructural studies, suggesting that this protein is required for oocyst development and sporogony. Transcriptome analysis of Δkinesin-8X gametocytes revealed modulated expression of genes involved mainly in microtubule-based processes, chromosome organisation and the regulation of gene expression, supporting a role for kinesin-8X in cell division. Kinesin-8X is thus required for parasite proliferation within the mosquito and for transmission to the vertebrate host., Competing Interests: The authors have declared that no competing interests exist.

Background: Viruses and other infectious agents cause more than 15% of human cancer cases. High-throughput sequencing-based studies of virus-cancer associations have mainly focused on cancer transcriptome data., Methods: In this study, we applied a diverse selection of presequencing enrichment methods targeting all major viral groups, to characterize the viruses present in 197 samples from 18 sample types of cancerous origin. Using high-throughput sequencing, we generated 710 datasets constituting 57 billion sequencing reads., Results: Detailed in silico investigation of the viral content, including exclusion of viral artefacts, from de novo assembled contigs and individual sequencing reads yielded a map of the viruses detected. Our data reveal a virome dominated by papillomaviruses, anelloviruses, herpesviruses, and parvoviruses. More than half of the included samples contained 1 or more viruses; however, no link between specific viruses and cancer types were found., Conclusions: Our study sheds light on viral presence in cancers and provides highly relevant virome data for future reference., (© The Author(s) 2019. Published by Oxford University Press for the Infectious Diseases Society of America.)

In recent years, the availability of reduced representation library (RRL) methods has catalysed an expansion of genome-scale studies to characterize both model and non-model organisms. Most of these methods rely on the use of restriction enzymes to obtain DNA sequences at a genome-wide level. These approaches have been widely used to sequence thousands of markers across individuals for many organisms at a reasonable cost, revolutionizing the field of population genomics. However, there are still some limitations associated with these methods, in particular the high molecular weight DNA required as starting material, the reduced number of common loci among investigated samples, and the short length of the sequenced site-associated DNA. Here, we present MobiSeq, a RRL protocol exploiting simple laboratory techniques, that generates genomic data based on PCR targeted enrichment of transposable elements and the sequencing of the associated flanking region. We validate its performance across 103 DNA extracts derived from three mammalian species: grey wolf (Canis lupus), red deer complex (Cervus sp.) and brown rat (Rattus norvegicus). MobiSeq enables the sequencing of hundreds of thousands loci across the genome and performs SNP discovery with relatively low rates of clonality. Given the ease and flexibility of MobiSeq protocol, the method has the potential to be implemented for marker discovery and population genomics across a wide range of organisms-enabling the exploration of diverse evolutionary and conservation questions., (© 2018 John Wiley & Sons Ltd.)

The mitochondrial F-type ATP synthase, a multisubunit nanomotor, is critical for maintaining cellular ATP levels. In T. gondii and other apicomplexan parasites, many subunit components necessary for proper assembly and functioning of this enzyme appear to be missing. Here, we report the identification of 20 novel subunits of T. gondii F-type ATP synthase from mass spectrometry analysis of partially purified monomeric (approximately 600 kDa) and dimeric (>1 MDa) forms of the enzyme. Despite extreme sequence diversification, key FO subunits a, b, and d can be identified from conserved structural features. Orthologs for these proteins are restricted to apicomplexan, chromerid, and dinoflagellate species. Interestingly, their absence in ciliates indicates a major diversion, with respect to subunit composition of this enzyme, within the alveolate clade. Discovery of these highly diversified novel components of the apicomplexan F-type ATP synthase complex could facilitate the development of novel antiparasitic agents. Structural and functional characterization of this unusual enzyme complex will advance our fundamental understanding of energy metabolism in apicomplexan species., Competing Interests: The authors have declared that no competing interests exist.

After publication of the original article [1] it was identified that order of the author list had been presented incorrectly. The author Robert Gniadecki's surname was also incorrect in the original article.

Background: Cutaneous basal cell carcinoma (BCC) is the commonest cancer worldwide. BCC is locally invasive and the surrounding stromal microenvironment is pivotal for tumourigenesis. Cancer associated fibroblasts (CAFs) in the microenvironment are essential for tumour growth in a variety of neoplasms but their role in BCC is poorly understood., Methods: Material included facial BCC and control skin from the peritumoural area and from the buttocks. With next-generation sequencing (NGS) we compared mRNA expression between BCC and peritumoural skin. qRT-PCR, immunohistochemical and immunofluorescent staining were performed to validate the NGS results and to investigate CAF-related cyto-and chemokines., Results: NGS revealed upregulation of 65 genes in BCC coding for extracellular matrix components pointing at CAF-related matrix remodeling. qRT-PCR showed increased mRNA expression of CAF markers FAP-α, PDGFR-β and prolyl-4-hydroxylase in BCC. Peritumoural skin (but not buttock skin) also exhibited high expression of PDGFR-β and prolyl-4-hydroxylase but not FAP-α. We found a similar pattern for the CAF-associated chemokines CCL17, CCL18, CCL22, CCL25, CXCL12 and IL6 with high expression in BCC and peritumoural skin but absence in buttock skin. Immunofluorescence revealed correlation between FAP-α and PDGFR-β and CXCL12 and CCL17., Conclusion: Matrix remodeling is the most prominent molecular feature of BCC. CAFs are present within BCC stroma and associated with increased expression of chemokines involved in tumour progression and immunosuppression (CXCL12, CCL17). Fibroblasts from chronically sun-exposed skin near tumours show gene expression patterns resembling that of CAFs, indicating that stromal fibroblasts in cancer-free surgical BCC margins exhibit a tumour promoting phenotype.

A novel human protoparvovirus related to human bufavirus and preliminarily named cutavirus has been discovered. We detected cutavirus in a sample of cutaneous malignant melanoma by using viral enrichment and high-throughput sequencing. The role of cutaviruses in cutaneous cancers remains to be investigated.

In a screen for unexplained mutation events we identified a previously unrecognized mechanism generating clustered DNA polymorphisms such as microindels and cumulative SNPs. The mechanism, short-patch double illegitimate recombination (SPDIR), facilitates short single-stranded DNA molecules to invade and replace genomic DNA through two joint illegitimate recombination events. SPDIR is controlled by key components of the cellular genome maintenance machinery in the gram-negative bacterium Acinetobacter baylyi. The source DNA is primarily intragenomic but can also be acquired through horizontal gene transfer. The DNA replacements are nonreciprocal and locus independent. Bioinformatic approaches reveal occurrence of SPDIR events in the gram-positive human pathogen Streptococcus pneumoniae and in the human genome., Competing Interests: The authors declare no conflict of interest.