Profiling novel lateral gene transfer events in the human microbiome.

Lateral gene transfer (LGT) is an important mechanism for genome diversification in microbial populations, including the human microbiome. While prior work has surveyed LGT events in human-associated microbial isolate genomes, the scope and dynamics of novel LGT events arising in personal microbiomes are not well understood, as there are no widely adopted computational methods to detect, quantify, and characterize LGT from complex microbial communities. We addressed this by developing, benchmarking, and experimentally validating a computational method (WAAFLE) to profile novel LGT events from assembled metagenomes. Applying WAAFLE to >2K human metagenomes from diverse body sites, we identified >100K putative high-confidence but previously uncharacterized LGT events (~2 per assembled microbial genome-equivalent). These events were enriched for mobile elements (as expected), as well as restriction-modification and transport functions typically associated with the destruction of foreign DNA. LGT frequency was quantifiably influenced by biogeography, the phylogenetic similarity of the involved taxa, and the ecological abundance of the donor taxon. These forces manifest as LGT networks in which hub species abundant in a community type donate unequally with their close phylogenetic neighbors. Our findings suggest that LGT may be a more ubiquitous process in the human microbiome than previously described. The open-source WAAFLE implementation, documentation, and data from this work are available at http://huttenhower.sph.harvard.edu/waafle.

Functional profile of host microbiome indicates Clostridioides difficile infection.

Clostridioides difficile infection (CDI) is a gastro-intestinal (GI) infection that illustrates how perturbations in symbiotic host-microbiome interactions render the GI tract vulnerable to the opportunistic pathogens. CDI also serves as an example of how such perturbations could be reversed via gut microbiota modulation mechanisms, especially fecal microbiota transplantation (FMT). However, microbiome-mediated diagnosis of CDI remains understudied. Here, we evaluated the diagnostic capabilities of the fecal microbiome on the prediction of CDI. We used the metagenomic sequencing data from ten previous studies, encompassing those acquired from CDI patients treated by FMT, CDI-negative patients presenting other intestinal health conditions, and healthy volunteers taking antibiotics. We designed a hybrid species/function profiling approach that determines the abundances of microbial species in the community contributing to its functional profile. These functionally informed taxonomic profiles were then used for classification of the microbial samples. We used logistic regression (LR) models using these features, which showed high prediction accuracy (with an average AUC≥0.91), substantiating that the species/function composition of the gut microbiome has a robust diagnostic prediction of CDI. We further assessed the confounding impact of antibiotic therapy on CDI prediction and found that it is distinguishable from the CDI impact. Finally, we devised a log-odds score computed from the output of the LR models to quantify the likelihood of CDI in a gut microbiome sample and applied it to evaluating the effectiveness of FMT based on post-FMT microbiome samples. The results showed that the gut microbiome of patients exhibited a gradual but steady improvement after receiving successful FMT, indicating the restoration of the normal microbiome functions.

The role of connectivity on COVID-19 preventive approaches.

Preventive and modeling approaches to address the COVID-19 pandemic have been primarily based on the age or occupation, and often disregard the importance of heterogeneity in population contact structure and individual connectivity. To address this gap, we developed models based on Erdos-Rényi and a power law degree distribution that first incorporate the role of heterogeneity and connectivity and then can be expanded to make assumptions about demographic characteristics. Results demonstrate that variations in the number of connections of individuals within a population modify the impact of public health interventions such as lockdown or vaccination approaches. We conclude that the most effective strategy will vary depending on the underlying contact structure of individuals within a population and on timing of the interventions.

Algorithmic approaches to clonal reconstruction in heterogeneous cell populations.

BACKGROUND: The reconstruction of clonal haplotypes and their evolutionary history in evolving populations is a common problem in both microbial evolutionary biology and cancer biology. The clonal theory of evolution provides a theoretical framework for modeling the evolution of clones. RESULTS: In this paper, we review the theoretical framework and assumptions over which the clonal reconstruction problem is formulated. We formally define the problem and then discuss the complexity and solution space of the problem. Various methods have been proposed to find the phylogeny that best explains the observed data. We categorize these methods based on the type of input data that they use (space-resolved or time-resolved), and also based on their computational formulation as either combinatorial or probabilistic. It is crucial to understand the different types of input data because each provides essential but distinct information for drastically reducing the solution space of the clonal reconstruction problem. Complementary information provided by single cell sequencing or from whole genome sequencing of randomly isolated clones can also improve the accuracy of clonal reconstruction. We briefly review the existing algorithms and their relationships. Finally we summarize the tools that are developed for either directly solving the clonal reconstruction problem or a related computational problem. CONCLUSIONS: In this review, we discuss the various formulations of the problem of inferring the clonal evolutionary history from allele frequeny data, review existing algorithms and catergorize them according to their problem formulation and solution approaches. We note that most of the available clonal inference algorithms were developed for elucidating tumor evolution whereas clonal reconstruction for unicellular genomes are less addressed. We conclude the review by discussing more open problems such as the lack of benchmark datasets and comparison of performance between available tools.

Insertion sequence elements-mediated structural variations in bacterial genomes.

Mobile genetic elements (MGEs) impact the evolution and stability of their host genomes. Insertion sequence (IS) elements are the most common MGEs in bacterial genomes and play a crucial role in mediating large-scale variations in bacterial genomes. It is understood that IS elements and MGEs in general coexist in a dynamical equilibrium with their respective hosts. Current studies indicate that the spontaneous movement of IS elements does not follow a constant rate in different bacterial genomes. However, due to the paucity and sparsity of the data, these observations are yet to be conclusive. In this paper, we conducted a comparative analysis of the IS-mediated genome structural variations in ten mutation accumulation (MA) experiments across eight strains of five bacterial species containing IS elements, including four strains of the E. coli. We used GRASPER algorithm, a denovo structural variation (SV) identification algorithm designed to detect SVs involving repetitive sequences in the genome. We observed highly diverse rates of IS insertions and IS-mediated recombinations across different bacterial species as well as across different strains of the same bacterial species. We also observed different rates of the elements from the same IS family in different bacterial genomes, suggesting that the distinction in rates might not be due to the different composition of IS elements across bacterial genomes.

STRScan: targeted profiling of short tandem repeats in whole-genome sequencing data.

BACKGROUND: Short tandem repeats (STRs) are found in many prokaryotic and eukaryotic genomes, and are commonly used as genetic markers, in particular for identity and parental testing in DNA forensics. The unstable expansion of some STRs was associated with various genetic disorders (e.g., the Huntington disease), and thus was used in genetic testing for screening individuals at high risk. Traditional STR analyses were based on the PCR amplification of STR loci followed by gel electrophoresis. With the availability of massive whole genome sequencing data, it becomes practical to mine STR profiles in silico from genome sequences. Software tools such as lobSTR and STR-FM have been developed to address these demands, which are, however, built upon whole genome reads mapping tools, and thus may not be sensitive enough. RESULTS: In this paper, we present a standalone software tool STRScan that uses a greedy algorithm for targeted STR profiling in next-generation sequencing (NGS) data. STRScan was tested on the whole genome sequencing data from Venter genome sequencing and 1000 Genomes Project. The results showed that STRScan can profile 20% more STRs in the target set that are missed by lobSTR. CONCLUSION: STRScan is particularly useful for the NGS-based targeted STR profiling, e.g., in genetic and human identity testing. STRScan is available as open-source software at http://darwin.informatics.indiana.edu/str/ .

Inter-Strain Differences in LINE-1 DNA Methylation in the Mouse Hematopoietic System in Response to Exposure to Ionizing Radiation.

Long Interspersed Nuclear Element 1 (LINE-1) retrotransposons are the major repetitive elements in mammalian genomes. LINE-1s are well-accepted as driving forces of evolution and critical regulators of the expression of genetic information. Alterations in LINE-1 DNA methylation may lead to its aberrant activity and are reported in virtually all human cancers and in experimental carcinogenesis. In this study, we investigated the endogenous DNA methylation status of the 5' untranslated region (UTR) of LINE-1 elements in the bone marrow hematopoietic stem cells (HSCs), hematopoietic progenitor cells (HPCs), and mononuclear cells (MNCs) in radioresistant C57BL/6J and radiosensitive CBA/J mice and in response to ionizing radiation (IR). We demonstrated that basal levels of DNA methylation within the 5'-UTRs of LINE-1 elements did not differ significantly between the two mouse strains and were negatively correlated with the evolutionary age of LINE-1 elements. Meanwhile, the expression of LINE-1 elements was higher in CBA/J mice. At two months after irradiation to 0.1 or 1 Gy of 137Cs (dose rate 1.21 Gy/min), significant decreases in LINE-1 DNA methylation in HSCs were observed in prone to radiation-induced carcinogenesis CBA/J, but not C57BL/6J mice. At the same time, no residual DNA damage, increased ROS, or changes in the cell cycle were detected in HSCs of CBA/J mice. These results suggest that epigenetic alterations may potentially serve as driving forces of radiation-induced carcinogenesis; however, future studies are needed to demonstrate the direct link between the LINE-1 DNA hypomethylation and radiation carcinogenesis.

Densely ionizing radiation affects DNA methylation of selective LINE-1 elements.

Long Interspersed Nucleotide Element 1 (LINE-1) retrotransposons are heavily methylated and are the most abundant transposable elements in mammalian genomes. Here, we investigated the differential DNA methylation within the LINE-1 under normal conditions and in response to environmentally relevant doses of sparsely and densely ionizing radiation. We demonstrate that DNA methylation of LINE-1 elements in the lungs of C57BL6 mice is dependent on their evolutionary age, where the elder age of the element is associated with the lower extent of DNA methylation. Exposure to 5-aza-2'-deoxycytidine and methionine-deficient diet affected DNA methylation of selective LINE-1 elements in an age- and promoter type-dependent manner. Exposure to densely IR, but not sparsely IR, resulted in DNA hypermethylation of older LINE-1 elements, while the DNA methylation of evolutionary younger elements remained mostly unchanged. We also demonstrate that exposure to densely IR increased mRNA and protein levels of LINE-1 via the loss of the histone H3K9 dimethylation and an increase in the H3K4 trimethylation at the LINE-1 5'-untranslated region, independently of DNA methylation. Our findings suggest that DNA methylation is important for regulation of LINE-1 expression under normal conditions, but histone modifications may dictate the transcriptional activity of LINE-1 in response to exposure to densely IR.

Radiation-induced changes in DNA methylation of repetitive elements in the mouse heart.

DNA methylation is a key epigenetic mechanism, needed for proper control over the expression of genetic information and silencing of repetitive elements. Exposure to ionizing radiation, aside from its strong genotoxic potential, may also affect the methylation of DNA, within the repetitive elements, in particular. In this study, we exposed C57BL/6J male mice to low absorbed mean doses of two types of space radiation-proton (0.1 Gy, 150 MeV, dose rate 0.53 ± 0.08 Gy/min), and heavy iron ions ((56)Fe) (0.5 Gy, 600 MeV/n, dose rate 0.38 ± 0.06 Gy/min). Radiation-induced changes in cardiac DNA methylation associated with repetitive elements were detected. Specifically, modest hypomethylation of retrotransposon LINE-1 was observed at day 7 after irradiation with either protons or (56)Fe. This was followed by LINE-1, and other retrotransposons, ERV2 and SINE B1, as well as major satellite DNA hypermethylation at day 90 after irradiation with (56)Fe. These changes in DNA methylation were accompanied by alterations in the expression of DNA methylation machinery and affected the one-carbon metabolism pathway. Furthermore, loss of transposable elements expression was detected in the cardiac tissue at the 90-day time-point, paralleled by substantial accumulation of mRNA transcripts, associated with major satellites. Given that the one-carbon metabolism pathway can be modulated by dietary modifications, these findings suggest a potential strategy for the mitigation and, possibly, prevention of the negative effects exerted by ionizing radiation on the cardiovascular system. Additionally, we show that the methylation status and expression of repetitive elements may serve as early biomarkers of exposure to space radiation.

Combined exposure to protons and (56)Fe leads to overexpression of Il13 and reactivation of repetitive elements in the mouse lung.

Interest in deep space exploration underlines the needs to investigate the effects of exposure to combined sources of space radiation. The lung is a target organ for radiation, and exposure to protons and heavy ions as radiation sources may lead to the development of degenerative disease and cancer. In this study, we evaluated the pro-fibrotic and epigenetic effects of exposure to protons (150 MeV/nucleon, 0.1 Gy) and heavy iron ions ((56)Fe, 600 MeV/nucleon, 0.5 Gy) alone or in combination (protons on Day 1 and (56)Fe on Day 2) in C57BL/6 male mice 4 weeks after irradiation. Exposure to (56)Fe, proton or in combination, did not result in histopathological changes in the murine lung. At the same time, combined exposure to protons and (56)Fe resulted in pronounced molecular alterations in comparison with either source of radiation alone. Specifically, we observed a substantial increase in the expression of cytokine Il13, loss of expression of DNA methyltransferase Dnmt1, and reactivation of LINE-1, SINE B1 retrotransposons, and major and minor satellites. Given the deleterious potential of the observed effects that may lead to development of chronic lung injury, pulmonary fibrosis, and cancer, future studies devoted to the investigation of the long-term effects of combined exposures to proton and heavy ions are clearly needed.

In Vitro Toxicity and Epigenotoxicity of Different Types of Ambient Particulate Matter.

Exposure to ambient particulate matter (PM) has been associated with adverse health effects, including pulmonary and cardiovascular disease. Studies indicate that ambient PM originated from different sources may cause distinct biological effects. In this study, we sought to investigate the potential of various types of PM to cause epigenetic alterations in the in vitro system. RAW264.7 murine macrophages were exposed for 24 and 72 h to 5- and 50-µg/ml doses of the water soluble extract of 6 types of PM: soil dust, road dust, agricultural dust, traffic exhausts, biomass burning, and pollen, collected in January-April of 2014 in the area of Little Rock, Arkansas. Cytotoxicity, oxidative potential, epigenetic endpoints, and chromosomal aberrations were addressed. Exposure to 6 types of PM resulted in induction of cytotoxicity and oxidative stress in a type-, time-, and dose-dependent manner. Epigenetic alterations were characterized by type-, time-, and dose-dependent decreases of DNA methylation/demethylation machinery, increased DNA methyltransferases enzymatic activity and protein levels, and transcriptional activation and subsequent silencing of transposable elements LINE-1, SINE B1/B2. The most pronounced changes were observed after exposure to soil dust that were also characterized by hypomethylation and reactivation of satellite DNA and structural chromosomal aberrations in the exposed cells. The results of our study indicate that the water-soluble fractions of the various types of PM have differential potential to target the cellular epigenome.

Long-term epigenetic effects of exposure to low doses of 56Fe in the mouse lung.

Despite significant progress, the long-term health effects of exposure to high charge (Z) and energy (E) nuclei (HZEs) and the underlying mechanisms remain poorly understood. Mouse studies show that space missions can result in pulmonary pathological states. The goal of this study was to evaluate the pro-fibrotic and pro-carcinogenic effects of exposure to low doses of heavy iron ions ((56)Fe) in the mouse lung. Exposure to (56)Fe (600 MeV; 0.1, 0.2 and 0.4 Gy) resulted in minor pro-fibrotic changes, detected at the beginning of the fibrotic phase (22 weeks post exposure), which were exhibited as increased expression of chemokine Ccl3, and interleukin Il4. Epigenetic alterations were exhibited as global DNA hypermethylation, observed after exposure to 0.4 Gy. Cadm1, Cdh13, Cdkn1c, Mthfr and Sfrp1 were significantly hypermethylated after exposure to 0.1 Gy, while exposure to higher doses resulted in hypermethylation of Cdkn1c only. However, expression of these genes was not affected by any dose. Congruently with the observed patterns of global DNA methylation, DNA repetitive elements were hypermethylated after exposure to 0.4 Gy, with minor changes observed after exposure to lower doses. Importantly, hypermethylation of repetitive elements coincided with their transcriptional repression. The findings of this study will aid in understanding molecular determinants of pathological states associated with exposure to (56)Fe, as well as serve as robust biomarkers for the delayed effects of irradiation. Further studies are clearly needed to investigate the persistence and outcomes of molecular alterations long term after exposure.