Newly designed pentacyclic benzimidazole derivatives, featuring amino or amido side chains, were synthesized to assess their in vitro antiproliferative activity. Additionally, we conducted investigations into their direct interaction with nucleic acids, aiming to uncover potential mechanisms of biological action. These compounds were synthesized using conventional organic synthesis methodologies, alongside photochemical and microwave-assisted reactions. Upon synthesis, the newly derived compounds underwent in vitro testing for their antiproliferative effects on various human cancer cell lines. Notably, derivatives 6 and 9 exhibited significant antiproliferative activity within the submicromolar concentration range. Our biological assessment highlighted that the biological activity was strongly influenced by both the position of the N atom on the quinoline moiety and the position and nature of the side chain on the pentacyclic skeleton. Findings from fluorescence, circular dichroism spectroscopy, and thermal melting assays indicated a mixed binding mode—comprising intercalation and the binding of aggregated compounds along the polynucleotide backbone—of these pentacyclic benzimidazoles with DNA and RNA.

N6-methyladenosine (M6A) modification is the most abundant internal chemical modification on RNA molecules in eukaryotes and can affect mRNA metabolism and regulate RNA transcription, nuclear export, splicing, degradation, translation, ets, and has a significant impact on various aspects of physiology or pathobiology. Radiotherapy is the most common measure of tumor treatment, variety of cellular intrinsic mechanisms affect the responses of cells to ionizing radiation (IR) and effectiveness of cancer radiotherapy. In this review, we summarize and discuss the recent advances about the roles and mechanisms of RNA M6A methylation in cellular responses to radiation-induced DNA damage response, and outcomes of cancer radiotherapy. Insights into the RNA M6A methylation in radiation biology may facilitate the improvement of therapeutic strategies for cancer radiotherapy as well as radioprotection on normal tissues.

Fibrosis represents a process characterized by excessive deposition of extracellular matrix (ECM) proteins. It often represents the evolution of pathological conditions, causes organ failure, and can, in extreme cases, compromises the functionality of the organs to the point of causing death. In recent years, considerable efforts have been made to understand the molecular mechanisms underlying fibrotic evolution and to identify possible therapeutic strategies. Great interest has been aroused by the discovery of a molecular association between epithelial to mesenchymal plasticity (EMP), in particular epithelial to mesenchymal transition (EMT), and fibrogenesis, which has led to the identification of complex molecular mechanisms closely interconnected with each other, which could explain EMT-dependent fibrosis. However, the result remains unsatisfactory from a therapeutic point of view. In recent years, advances in epigenetics, which is based on chromatin remodeling through various histone modifications or through the intervention of non-coding RNAs (ncRNAs), have provided more information on the fibrotic process, and this could represent a promising path forward for the identification of innovative therapeutic strategies for organ fibrosis. In this review, we summarize current research on epigenetic mechanisms involved in organ fibrosis, with a focus on epigenetic regulation of EMP/EMT-dependent fibrosis.

In the last two decades, plant taxonomy has bloomed, following the development of a novel technique, namely DNA Barcoding. DNA barcodes are standardized sequences, ideally unique, coding or non-coding, either from the genome of the organism or from its organelles, that are used to identify/classify an organismal group; in short the method includes amplification of the DNA barcode, sequencing and comparison with a reference database containing the relevant sequences from different species. In plants, the use a universal DNA barcode, such as COI which is used in animals, has not been achieved so far. Several DNA barcodes from single loci as well as combinations of barcodes have been successfully used in various settings; additionally, as next generation sequencing becomes affordable, very long sequences- super barcodes, such as the chloroplast genome are being used successfully especially to distinguish between close species. In this review we summarize the work done on DNA barcoding in plants and discuss different approaches used so far.

Breast cancer is a serious disease and the second leading cause of cancer-related death among women in the U.S. New treatments for this aggressive disease are urgently needed. Repurposing FDA-approved drugs for cancer treatment is an alternative that saves time and lowers the costs needed for drug development. In this study, we investigated the effects of proguanil, an anti-malarial drug, in breast cancer cells. Proguanil exhibited a significant cytotoxic effect on breast cancer cell lines including patient derived cell lines. Proguanil caused apoptosis through increased production of ROS and consequent decrease of mitochondrial membrane potential, mitochondrial respiration, and ATP production rates. ROS generation by proguanil was up to 3-fold higher when compared to the control. Proguanil treatment increased the expression of Bax, p-H2AX, cleaved-caspase 9, cleaved PARP, and down-regulated bcl-2 and survivin in breast cancer cell lines. The enlargement of 4T1 breast tumors in female Balb/c mice was suppressed by 55% through daily oral administration of 20mg/kg of proguanil. Western blot analyses of proguanil- treated tumors showed increased levels of p-H2AX, Bax, c-PARP, and c-caspase3 compared to control. Proguanil proved to be an efficient in vitro and in-vivo inhibitor of breast cancer cells, hence should be considered further for clinical investigation against breast cancer.

Dengue is a pan-tropical arthropod-borne disease, endemic in over 100 countries. The dengue virus comprises four antigenic serotypes (DENV 1-4). The envelope (E) and non-structural 1 (NS1) proteins are highly immunogenic, and the E protein carries epitopes eliciting neutralizing antibodies. In previous studies, mice immunizations with two DNA vaccines encoding the E ectodomain (pE1D2) and NS1 proteins (pcTPANS1) from DENV2 induced high protection. Based on these findings, we constructed new plasmids containing the codon-optimized genes from these proteins for expression in mouse and human cells, named pEotmD2 and pNS1otmD2. Compared to the pE1D2, the pEotmD2 mediated lower expression of the recombinant protein in human and mouse cell lines, while the pNS1otmD2 plasmid was slightly more efficient than the pcTPANS1 in these cells. Immunogenicity and protection were evaluated in BALB/c mice immunized with the original and codon-optimized plasmids followed by challenged with DENV2. In accordance with the in vitro experiments, the antibody response and protection elicited by the pE1otmD2 plasmid was lower than those observed with the pE1D2. In contrast, the pNS1otmD2 vaccine enabled slightly higher levels of the humoral and protective responses than the pcTPANS1. Overall, our study revealed different effects of codon optimization on these two dengue DNA vaccines.

Biofilm matrix is akin to bacterial abodes, primarily composed of proteins, polysaccharides, extracellular DNA, and lipids, providing essential structural support to bacteria. While some biofilm matrix offers significant benefits, such as engineered biofilm reactors that assist in water pollutant removal, others pose risks to human health, potentially leading to conditions like lung infections. Realizing the potential of advantageous biofilm matrix and mitigating the harmful effects of detrimental ones necessitates a comprehensive and systematic exploration of this intricate domain. To facilitate this examination, our study leveraged a dataset of the 1000 most pivotal papers in the field and conducted an exhaustive bibliographic review. This paper serves as a valuable resource for understanding the current progress in biofilm matrix research and delving into vital avenues for future exploration in this dynamic field.

CXCL14 is one of the most evolutionarily conserved members of the chemokine family and is constitutionally expressed in multiple organs, suggesting that it is involved in the homeostasis maintenance of the system. CXCL14 is highly expressed in colon epithelial cells and shows obvious gene silencing in clinical colon cancer samples, suggesting that its silencing is related to the immune escape of cancer cells. In this paper, we analyzed the expression profiles of multiple human clinical colon cancer datasets and mouse colon cancer models to reveal the variation trend of CXCL14 expression during colitis, colon polyps, primary colon cancer, and liver metastases. The relationship between CXCL14 gene silencing and promoter hypermethylation was revealed through the colorectal carcinoma methylation database. The results suggest that CXCL14 is a tumor suppressor gene in colorectal carcinoma, which is activated first and then silenced during the process of tumor occurrence and deterioration. Promoter hypermethylation is the main cause of CXCL14 silencing. The methylation level of CXCL14 is correlated with the anatomic site of tumor occurrence and positively correlated with patient age, and associated with prognosis. Reversing the hypermethylation of CXCL14 may be an epigenetic therapy for colon cancer.

Background: Androgenetic alopecia (AGA) and alopecia areata (AA) are most common types of non-cicatricial alopecia. Both diseases have limited effective therapeutic options and affect patients' quality of life. Pharmacogenetic tests can help predict the most appropriate treatment option by evaluating the single nucleotide polymorphisms (SNPs) corresponding to genes related to alopecia. The objective of the study was to evaluate and compare selected SNPs and genes in AA and AGA patients from Romania and Brazil. Materials and methods: We investigated associations between AA and AGA and 45 tag SNPs of 13 genes in 287 Romanian and 882 Brazilian patients. The DNA samples were extracted from saliva using qPCR technique. Results: GR-alpha gene, GPR44-2 gene, SULT1A1 gene and CRABP2 gene were statistically significantly different in Brazil compared to Romania. Minoxidil may be recommended in half of the cases of AGA and AA. Patients with AGA and high expression of SRD5A1 or PTGFR-2 may benefit from Dutasteride, respectively Latanoprost treatment. Most of the studied genes showed no differences between the two population. Conclusion: The DNA analysis of the patients with alopecia may contribute to a successful treatment.

DNA damage response (DDR) pathways in keto-enol genotoxicity have not been characterized, and few studies have reported genotoxic effects in non-target organisms. The present study shows that concentrations of 11.2, 22.4, 37.3 mg/L of Movento® 240SC and 12.3, 24.6, 41.1 mg/L of Envidor® 240SC for 72 h oral exposure induced DSBs by significantly increasing the percentage of γH2AV expression in regions 2b and 3 from the germarium of wildtype females of Drosophila melanogaster Oregon R, compared to the control group (0.0 mg/L of insecticides), by confocal immunofluorescence microscopy. The comparison between both insecticides’ revels that only the Envidor® 240SC induces concentration-dependent DNA damage, as well as structural changes in the germarium. We determined that the DDR induced by Movento® 240SC depends on the activation of the ATMtefu, Chk1grp and Chk2lok kinases by significantly increasing the percentage of expression of γH2AX in regions 2b and 3 of the germarium, and that ATRmei-29D and p53dp53 kinases only respond at the highest concentration of 37.3 mg/L of Movento® 240SC. With the Envidor® 240SC insecticide, we determined that the DDR depends on the activation of the ATRmei-29D/Chk1grp and ATMtefu/Chk2lok kinases, and p53dp53 by significantly increasing the percentage of expression of γH2AX in the germarium.

Amphibian populations have been globally declining since at least 1990. In temperate forests of eastern North America, vernal pools offer important breeding habitats to many amphibians but are usually not considered for forest management because of their small size and temporary status. Effective monitoring and management of amphibians rely on accurate knowledge of their spatiotemporal distributions, which is expensive to collect due to the amount of fieldwork re-quired. In this study, we tested whether eDNA metabarcoding could identify the same amphibian communities as traditional inventory protocols. We collected eDNA samples in twelve vernal pools in the spring of 2019 and identified their communities by metabarcoding. At each pool, three traditional amphibian inventory methods were used in May and June 2019: call surveys using acoustic recorders, trapping, and active search surveys. In total, 13 amphibian species were detected, most of them being detected by both eDNA and traditional methods. We found that species ecology and behavior are key factors of its detectability by a specific method. As eDNA metabarcoding is comparatively inexpensive and presents a higher repeatability, we conclude that eDNA sampling should be considered for integration as a standard monitoring tool, after an initial assessment of amphibian diversity.

Genes encoding retinol binding protein 4 (RBP4), follicle stimulating hormone subunit beta (FSHB) and epidermal growth factor (EGF) have been proposed as candidate genes for reproduction traits in pigs. The study presented in this paper aimed to find associations between variants of these genes and reproduction traits in pigs reared in Poland. Investigation included Polish Large White x Polish Landrace (n=288) and Yorkshire (n=195) sows. Individual genotypes were determined by means of PCR-RFLP (single nucleotide polymorphism in RBP4, FSHB) or PCR (insertion/deletion in EGF) methods. Obtained result showed that RBP4 variants were related to total number born (TNB) and number born alive (NBA) in 1st, 2nd and 5th parities, FSHB to TNB and NBA in first two parities, however EGF with both traits in all parities (P≤0.05 or P≤0.01). In case of RBP4 gene, heterozygous genotype (AB) was favorable for analyzed traits, however in case of FSHB and EGF homozygous genotype, AA and BB respectively. Obtained results indicate that polymorphism in three analyzed genes is associated with important reproductive traits of pigs kept in Poland.

Background: Cell-free plasma DNA (cfDNA) is composed of circulating extracellular DNA originating from apoptosis or cell necrosis. It is generally present in healthy individuals, but its levels increase in different clinical conditions, such as kidney disease, sepsis, myocardial infarction, trauma and cancer. Caspase-3 plays a central role in apoptosis, a mechanism of programmed cell death involved in the pathogenesis of chronic kidney disease (CKD). Methods: We compared cfDNA and caspase-3 levels in 25 CKD patients and in 10 healthy subjects, evaluating the modification of their concentrations according to CKD stage. We also analyzed the correlation between cfDNA and caspase-3 levels in CKD patients and the correlation between cfDNA and caspase-3 levels and creatinine and urea. Results: CfDNA and caspase-3 levels were higher in CKD patients compared to healthy subjects, in particular in patients with advanced renal disease. A positive correlation between cfDNA and caspase-3 levels and between cfDNA and caspase-3 and creatinine and urea was also observed. Conclusion: Patients with chronic kidney disease present higher levels of cfDNA and caspase-3 levels compared with healthy subjects. The worsening of renal damage and the increase in uremic toxin concentration are associated to higher levels of cfDNA and caspase-3 levels, thus reflecting a central role of apoptosis in the pathogenesis of CKD.

Cell identity is determined by chromatin structure and the profiles of gene expression, which are dependent on chromatin accessibility and DNA methylation of regions critical for gene expression, such as enhancers and promoters. These epigenetic modifications are required for mammalian development and are essential for the establishment and the maintenance of the cellular identity. DNA methylation was once thought to be a permanent repressive epigenetic mark, but systematic analysis in various genomic contexts reveals a more dynamic regulation than previously thought. In fact, both active DNA methylation and demethylation occur during cell fate commitment and terminal differentiation. To link methylation signatures of specific genes to their expression profiles, we have determined the methyl-CpG configurations of the promoters of five genes switched on and off during murine postnatal brain differentiation by bisulfite-targeted sequencing. We report here the structure of significant, dynamic and stable methyl-CpG profiles associated with silencing or activation of expression of genes during neural stem cell and brain postnatal differentiation. Strikingly, these methylation cores mark different mouse brain areas and cell types derived from the same areas during differentiation.

Minute Virus of Mice (MVM) is an autonomous parvovirus of the Parvovirus family that replicates in mouse cells and transformed human cells. MVM genomes localize to cellular sites of DNA damage with the help of their essential non-structural phosphoprotein NS1 to establish viral replication centers. MVM replication induces a cellular DNA damage response that is mediated by signaling through the ATM kinase pathway, while inhibiting induction of the ATR kinase signaling pathway. However, the cellular signals regulating virus localization to cellular DNA damage response sites has remained unknown. Using chemical inhibitors to DNA damage response proteins, we have discovered that NS1 locallization to cellular DDR sites is independent of ATM or DNA-PK signaling, but is dependent on ATR signaling. Pulsing cells with an ATR inhibitor after S-phase entry leads to attenuated MVM replication. These observations suggest that the initial localization of MVM to cellular DDR sites depends on ATR signaling before it is inactivated by vigorous virus replication.

Free-living terrestrial mites (Acari) have persisted through numerous glacial cycles in Antarctica. Very little is known, however, of their genetic diversity and distribution, particularly within the Ross Sea region. To redress this gap, we sampled for mites throughout the Ross Sea region, East Antarctica, including Victoria Land and the Queen Maud Mountains (QMM), covering a latitudinal range of 72-85oS, as well as from Lauft Island near Mt Siple (73oS) in West Antarctica and Macquarie Island (54oS) in the sub-Antarctic. We assessed genetic diversity using mitochondrial cytochrome c oxidase subunit I gene sequences (COI-5P DNA barcode region), and also morphologically identified voucher specimens. We obtained 130 sequences representing four genera: Nanorchestes (n = 30 sequences), Stereotydeus (n = 46), Coccorhagidia (n = 18) and Eupodes (n = 36). Tree-based analyses (maximum likelihood) revealed 13 genetic clusters, representing as many as 23 putative species indicated by Barcode Index Numbers (BINs) from the Barcode of Life Datasystems (BOLD) database. We found evidence for geographically-isolated cryptic species, e.g. within Stereotydeus belli and S. punctatus, as well as unique genetic groups occurring in sympatry (e.g. Nanorchestes spp. in QMM). Collectively, these data confirm high genetic divergence as a consequence of geographic isolation over evolutionary timescales. From a conservation perspective, additional targeted sampling of understudied areas in the Ross Sea region should be prioritised, as further diversity is likely to be found for these short-range endemic mites.

Systematics plays the most crucial role in biodiversity conservation which is at stake due to anthropogenic activities and environmental degradations. The ever-increasing decline of classical taxonomic expertise drives the need to develop molecular marker-based tools for quick, efficient, and reliable detection of organisms, to assess their ecological impacts for deepening our understanding of systematic and evolutionary relationships between organisms which is central to biology. The pace of alpha taxonomy has quickened by its integration with an increasingly fashionable and novel concept called DNA barcoding which utilizes a short genetic marker or barcode to categorize species for enhanced biodiversity assessment. As a supplementary but not complete alternative of systematics research, DNA barcoding, however, not error-free, brings precision in identification by solving existing problems of classical taxonomy and phylogenetics, irrespective of the growth stage of organisms, particularly for known taxa rather than unknown ones. Mitochondrial gene Cytochrome C Oxidase 1 (COI) serves as a universal animal barcode but there is no such universal barcode for plants and developing a suitable one is more challenging. With the recent advancement of Next Generation Sequencing (NGS), DNA metabarcoding technology is advancing rapidly. Still, ambiguity and error prevail with the correct identification of species due to some problems. After extensive analysis of the existing DNA barcoding papers, this review discusses commonly used DNA barcodes in plants and animals, their roles, advantages, limitations to solve existing problems of conservation biology and add the author’s views and recommendations.

Plastid genomes are in general highly conserved given their slow evolutionary rate, thus large changes in their structure are unusual. However, when specific rearrangements are present, they are often phylogenetically informative. Asteraceae is a highly diverse family whose evolution is long driven by polyploidy (up to 48x) and hybridisation, both processes usually complicating systematic inferences. In this study, we have generated one of the most comprehensive plastome-based phylogenies of family Asteraceae, providing information about the structure, genetic diversity, and repeat composition of these sequences. By comparing the whole plastome sequences obtained, we confirmed the double inversion located in the long single copy region, for most of the species analysed (with the exception of basal tribes), a well-known feature for Asteraceae plastomes. We also show that genome size, gene order and gene content are highly conserved along the family. However, species representative of the basal subfamily Barnadesioideae -as well as in the sister family Calyceraceae - are lacking the pseudogene rps19 located in one inverted repeat. The phylogenomic analysis conducted here, based on 63 protein-coding genes, 30 transfer RNA genes and 21 ribosomal RNA genes from 36 species of Asteraceae, are overall consistent with the general consensus for the family’s phylogeny, while resolving the position of tribe Senecioneae and revealing some incongruences at tribe level between reconstructions based on nuclear and plastid DNA data.

Meniere Disease (MD) is a multifactorial disorder of the inner ear characterized by vertigo attacks associated with sensorineural hearing loss and tinnitus with a significant heritability. Although MD has been associated with several genes, no epigenetic studies have been performed in MD. Here we performed whole genome bisulfite sequencing in 14 MD patients and 6 healthy controls, with the aim of identifying a MD methylation signature and potential disease mechanisms. We observed a high number of differentially methylated CpGs (DMC) when comparing MD patients to controls (N= 9,545), several of them in hearing loss genes such as PCDH15, ADGRV1 and CDH23. Bioinformatic analyses of DMCs and cis-regulatory regions predicted phenotypes related to abnormal excitatory postsynaptic currents, abnormal NMDA-mediated receptor currents and abnormal glutamate-mediated receptor currents when comparing MD to controls. Moreover, we identified various DMCs in genes previously associated with cochleovestibular phenotypes in mice. We have also found 12 undermethylated regions (UMR) that were exclusive to MD, including 2 UMR in an inter CpG island in the PHB gene. We suggest that the DNA methylation signature allows to distinguish between MD patients and controls. The enrichment analysis confirms previous findings of a chronic inflammatory process underlying MD.

Background: Exposure in utero to particulate matter (PM2.5 and PM10) is associated with maladaptive health outcomes. Although exposure to prenatal PM2.5 and PM10 have cord blood DNA methylation signatures at birth, signature persistence into childhood and saliva cross-tissue applicability has not been tested. Methods: In the Fragile Families and Child Wellbeing Study, a United States 20-city birth cohort, average residential PM2.5 and PM10 during pregnancy was estimated using air quality monitors with inverse distance weighting. Saliva DNA methylation at ages 9 (n=749) and 15 (n=793) was measured using the Illumina HumanMethylation 450k BeadArray. Cumulative DNA methylation scores for particulate matter were estimated by weighting participant DNA methylation at each site by meta-analysis effect estimates from Gruzieva et al. 2019 and standardizing the sums. Using mixed effects regression analysis, we tested the associations between cumulative DNA methylation scores at ages 9 and 15 and PM exposure during pregnancy, adjusted for child sex, age, race/ethnicity, maternal income to needs ratio, nonmartial birth status, and saliva cell type proportions. Results: Our study sample was 50.5% male, 56.3% non-Hispanic Black, and 19.8% Hispanic, with median income to needs ratio of 1.4. In the third trimester, mean PM2.5 exposure levels were 27.9 ug/m3/day (standard deviation: 7.0, 23.7% of observations exceeded safety standards) and PM10 were 15.0 ug/m3/day (standard deviation: 3.1). An interquartile range increase in PM2.5 exposure (10.73 g/m3/day) was associated with -0.0287 standard deviation lower cumulative DNA methylation score for PM2.5 (95% CI: -0.0732, 0.0158, p=0.20) across all participants. An interquartile range increase in PM10 exposure (3.20 g/m3/day) was associated with -0.1472 standard deviation lower cumulative DNA methylation score for PM10 (95% CI: -0.3038, 0.0095, p=0.06) across all participants. The PM10 findings were driven by the age 15 subset where an interquartile range increase in PM10 exposure was associated with -0.024 standard deviation lower cumulative DNA methylation score for PM10 (95% CI: -0.043, -0.005, p=0.012). Findings were robust to adjustment for PM exposure at ages 1 and 3. Conclusion: In utero PM10 associated DNA methylation differences persist until age 15 and can be detected in saliva. Benchmarking the persistence and cell type generalizability is critical for epigenetic exposure biomarker assessment.

Epigenetics, specifically DNA methylation, allows for estimation of animal age from blood or remotely sampled skin. This multi tissue epigenetic aging clock uses 110 longitudinal samples from 34 Navy bottlenose dolphins (Tursiops truncatus), identifying 195 cytosine-phosphate-guanine sites associated with chronological aging via leave-one-individual-out-cross-validation (R2=0.95). With a median absolute error of 2.5 years this clock improves age estimation capacity in wild dolphins, expanding conservation efforts, enabling better understanding of population demographics.

Genetic diversity is inevitable in making any crop improvement program successful. DNA fingerprinting technology to assess the genetic relationship among the selected genotypes for identification and cataloging of different species and cultivars of roses is a promising tool for Rosa genomes. The inter-simple sequence repeats markers (ISSRs) were used to investigate the genetic diversity among twenty-one diverse Rosa genotypes belonging to two different species, Rosa hybrida and R. damascena, and three distinct groups of rose varieties, namely Hybrid Tea, Floribunda, and Damask roses. Twenty-four ISSR primers yielded a total of 280 scorable amplified fragments from 250-1800 bp in length, from which 244 were polymorphic, resulting in an average of 86.4% polymorphism. UPGMA cluster analysis based on Jaccard’s pairwise similarity coefficient values ranged from 0.264 to 0.818, clearly distinguished different species and genotypes, grouping them into three distinct clusters. The results confirmed a high degree of variation in the rose germplasm studied highlighting the potential of improvement in roses for the ornamental and perfume industry.

Determining the DNA sequencing of a small element in the mitochondrial DNA (DNA barcoding) makes it possible to easily identify individuals of different larval stages of marine crustaceans without the need for laboratory rearing. It can also be used to construct taxonomic trees, although it is not yet clear to what extent this barcode-based taxonomy reflects more traditional morphological or molecular taxonomy. Collections of zooplankton were made using conventional plankton nets in Newport Bay and the Pacific Ocean near Newport Beach, California (Lat. 33.628342, Long. -117.927933) between May 2013 and January 2020, and individual crustacean specimens were documented by videomicroscopy. Adult crustaceans were collected from solid substrates in the same areas. Specimens were preserved in ethanol and sent to the Canadian Centre for DNA Barcoding at the University of Guelph, Ontario, Canada for sequencing of the COI DNA barcode. From 1042 specimens, 544 COI sequences were obtained falling into 199 Barcode Identification Numbers (BINs), of which 76 correspond to recognized species. The results show the utility of DNA barcoding for matching life-cycle stages as well as for documenting the diversity of this group of organisms.

Epigenetics refers to the DNA chemistry changes that result in the modification of gene transcription and translation independently of the underlying DNA coding sequence. Epigenetic modifications are reported to involve various molecular mechanisms, including classical epigenetic changes affecting DNA methylation and histone modifications and small RNA-mediated processes, particularly that of microRNAs. Epigenetic changes are reversible and are closely interconnected. They are recognised to play a critical role as mediators of gene regulation, and any alteration in these mechanisms has been identified to mediate various pathophysiological conditions. Moreover, genetic predisposition and environmental factors, including dietary alterations, lifestyle or metabolic status, are identified to interact with the human epigenome, highlighting the importance of epigenetic factors as underlying processes in the etiology of various diseases such as MetS. This review will reflect on how both the classical and microRNA regulated epigenetic changes are associated with the pathophysiology of Metabolic syndrome. We would then focus on the various aspects of epigenetic-based strategies used to modify MetS outcomes, including epigenetic diet, epigenetic drugs, epigenome editing tools, and miRNA-based therapies.

Pythium insidiosum causes pythiosis, a fatal infectious disease of humans and animals worldwide. Prompt diagnosis and treatment are essential to improve the clinical outcome of pythiosis. Diagnosis of P. insidiosum relies on immunological, molecular, and proteomic assays. The main treatment of pythiosis aims to surgically remove all affected tissue to prevent recurrent infection. Due to the marked increase in case reports, pythiosis has become a public health concern. Thailand is an endemic area of human pythiosis. To obtain a complete picture of how the pathogen circulates in the environment, we surveyed the presence of P. insidiosum in urban (Bangkok) and rural areas of Thailand. We employed the hair-baiting technique to screen for P. insidiosum in 500 water samples. Twenty-seven culture-positive samples were identified as P. insidiosum by multiplex PCR, multi-DNA barcode (rDNA, cox1, cox2), and mass spectrometric analyses. These environmental strains of P. insidiosum fell into clade-I and -II genotypes and exhibited a close phylogenetic/proteomic relationship with Thai clinical strains. Biodiversity of the environmental strains also existed in a local habitat. In conclusion, P. insidiosum is widespread in Thailand. A better understanding of the ecological niche of P. insidiosum could lead to the effective prevention and control of this pathogen.

Nigella sativa (NS) is an effective medicinal plant possessing noteworthy antioxidant property. In NS, there are more than hundred phyto-chemicals reported, out of which thymoquinone is the utmost active phyto-constituent having sturdy antioxidative property. Thymoquinone is a cyclicdione, when reacts with sodium azide, converts into α-azido ketones i.e its analogs which are handy with extensive range of reactions. Sodium azide induces stress in plants thereby, modulating the antioxidant system. The present investigation was planned to elucidate the effect of sodium azide at different concentrations (5µM, 10µM, 20µM, 50µM, 100µM and 200µM) on its secondary metabolites (mainly thymoquinone) in NS callus culture extract (NSE). The results showed sodium azide effect on thymoquinone content and a concentration dependent boost in antioxidant property. It was also observed that thymoquinone content and percent yield (analyzed by RP-HPLC; Reverse Phase- High Performance Liquid Chromatography) were minimum (0.033±0.006% and 0.420±0.045%, respectively) at 200 µM sodium azide used. Whereas, antioxidant activity (analyzed by DPPH; 2, 2-diphenyl-1-picrylhydrazyl) was found to be maximum (3.873±0.402%) at same dose. Further, analysis was done for inhibition of oxidative DNA damage at different concentrations of sodium azide on NSE, maximum inhibition of DNA damage (0.243±0.017%) was found at 200 µM concentration of sodium azide. When correlated, strong positive correlation was observed between percent yield and percent thymoquinone, antioxidant property and inhibition of DNA damage. Whereas, strong negative correlation was observed between percent yield and antioxidant property, percent thymoquinone and antioxidant property, percent thymoquinone and inhibition of DNA damage. The findings evidently point out that the content of thymoquinone, antioxidant property and inhibition of DNA damage was affected by sodium azide.

The potential to affect gene expression via G-quadruplex stabilization has been extended to all domains of life, including viruses. Here, we investigate the polymorphism and structures of G-quadruplexes of the human papillomavirus type 52 with UV, CD and NMR spectroscopy and gel electrophoresis. We show that oligonucleotide with five G-tracts folds into several structures and that naturally occurring single nucleotide polymorphisms (SNPs) have profound effects on the structural polymorphism in the context of G-quadruplex forming propensity, conformational heterogeneity and folding stability. With help of SNP analysis, we were able to select one of the predominant forms, formed by G-rich sequence d(G3TAG3CAG4ACACAG3T). This oligonucleotide termed HPV52(1-4) adopts a three G-quartet snap back (3+1) type scaffold with four syn guanine residues, two edgewise loops spanning the same groove, a no-residue V loop and a propeller type loop. The first guanine residue is incorporated in the central G-quartet and all four-guanine residues from G4 stretch are included in the three quartet G-quadruplex core. Modification studies identified several structural elements that are important for stabilization of the described G-quadruplex fold. Our results expand set of G-rich targets in viral genomes and address the fundamental questions regarding folding of G-rich sequences.

Removal of the proliferation component of gene expression by PCNA adjustment has been addressed in numerous survival prediction studies for breast cancer and all cancers in the TCGA. These studies indicate that widespread co-regulation of proliferation upwardly biases survival prediction when gene selection is performed on a genome-wide basis. In addition, removal of the correlative effects of proliferation does not reduce the random bias associated with survival prediction using random gene selection. Since most cancers become addicted to DNA repair as a result of forced cellular replication, increased oxidation, and repair deficiencies from oncogenic loss or genetic polymorphisms, we pursued an investigation to remove the proliferation component of expression in DNA repair genes to determine survival prediction. This translational hypothesis-driven focus on DNA repair genes is directly amenable to finding new sets of DNA repair genes that could potentially be studied for inhibition therapy. Overall survival (OS) prediction was evaluated in 18 cancers by using normalized RNA-Seq data for 126 DNA repair genes with expression available in TCGA. Transformations for normality and adjustments for age at diagnosis, stage, and PCNA metagene expression were performed for all DNA repair genes. We also analyzed genomic event rates (GER) for somatic mutations, deletions, and amplification in driver genes and DNA repair genes. After performing empirical p-value testing with use of randomly selected gene sets, it was observed that OS could be predicted significantly by sets of DNA repair genes for 61% (11/18) of the cancers. Interestingly, PARP1 was not a significant predictor of survival for any of the 11 cancers. Results from cluster analysis of GERs indicates that the most opportunistic cancers for inhibition therapy may be AML, colorectal, and renal papillary, because of potentially less confounding due to lower GERs for mutations, deletions, and amplifications in DNA repair genes. However, the most opportunistic cancer for inhibition therapy is likely to be AML, since it showed the lowest GERs for mutations, deletions, and amplifications in DNA repair genes. In conclusion, our hypothesis-driven focus to target DNA repair gene expression adjusted for the PCNA metagene as a means of predicting OS in various cancers resulted in statistically significant sets of genes.

Radical oxygen species formed in human tissue cells by many endogenous and exogenous pathways, cause extensive oxidative damage, which has been linked to various human diseases. This review paper provides an overview of lipid peroxidation and focuses on the free-radicals initiated processes of LDL oxidative modification and DNA oxidative damage, which are widely associated to the initiation and development of atherosclerosis and carcinogenesis, respectively. The article subsequently provides an overview of the recent human trials or even in vitro investigations on the potential of natural antioxidant compounds (such as carotenoids, vitamins C and E) to monitor LDL and DNA oxidative changes.

The role of epigenetic alterations in the pathogenesis of age-related macular degeneration (AMD) has been pending so far. Our study investigated the effect of oxidative stress and inflammation on DNA methyltransferases (DNMTs) and Sirtuin 1 (SIRT1) functions, as well as on long interspersed nuclear element-1 (LINE-1) methylation, in human retinal pigment epithelial (ARPE-19) cells. Therefore, we evaluated whether treatment with resveratrol may restore changes in LINE-1 methylation by modulating DNMTs and SIRT1 functions. Cells were treated with 25 mU/ml glucose oxidase (GOx) or 10 µg/ml lipopolysaccharide (LPS) to mimic oxidative or inflammatory conditions, respectively. Oxidative stress decreased DNMT1, DNMT3a, DNMT3b and SIRT1 expression (p-values <0.05), as well as total DNMTs (-28.5%; p<0.0001) and SIRT1 (-29.0%;p<0.0001) activities. Similarly, inflammatory condition decreased DNMT1 and SIRT1 expression (p-values<0.05), as well as total DNMTs (-14.9%;p=0.007) and SIRT1 (-20.1%;p<0.002) activities. Interestingly, GOx- and LPS-treated cells exhibited lower LINE-1 methylation compared to controls (p-values<0.0001). We also demonstrated that treatment with 10 μM resveratrol for 24 hours counteracted the detrimental effect on LINE-1 methylation via increasing DNMTs and SIRT1 functions in cells upon oxidative and inflammatory conditions. However, further studies should explore the perspectives of resveratrol as a suitable strategy for the prevention and/or treatment of AMD.

A total of five protoclones were successfully cultured on PDA medium out of regenerated twenty two colonies of Termitomyces protoplast and further studied. Liquid MYG grown mycelial tissue is used for protoplast isolation by enzymatic digestionin a mixture containing Lysing enzyme 2% and Cellulase R10 2% in 0.6 M mannitol. The incubation conditions like temperature, shaking and time were standardized at 24ºC, 60 rpm and 10 hours, respectively for healthy protoplasts liberation. The purified protoplasts showed an average yield of 1.2 × 10⁷ cells/gm tissue with 31.60 ± 9.31% regeneration efficiency on specific medium and 77.12 ± 2.72% viability by FDA test. Four ISSR primers were used in this study resulting a total of 27 reproducible bands with mean value of 6.75. They showed similar banding pattern in all the lines with zero percent polymorphism ranged from 280 bp–2700 bp. The amplified rRNA-ITS gene showed ~600 bp size in gel and found a single restriction site for enzyme HaeIII in all the protoclones and parent with similar fragment size in all.

Dengue virus (DENV), the causative agent of dengue fever (DF), is one of the most important mosquito-borne viruses that can infect humans. Although much effort has been made on prevention and control of dengue, there are currently no anti-viral drugs or worldwide approved vaccines yet. In this study, we immunized six-week-old Balb/c mice with DNA vaccine candidates E and NS1-2a of DENV serotype 2 or the combination of them (E+NS1-2a) via an electroporation (EP)-assisted intramuscular gene delivery system and evaluated the immune response and protection. The highest specific antibody titres and cytokine levels secreted by splenocytes as well as the highest survival rate were observed in the E+NS1-2a group, followed by E group and NS1-2a group. Our data suggested that the combination of E and NS1-2a delivered by EP may be a superior preventive strategy against DENV.

Furazolidone (FZD) is a synthetic nitrofuran with the antiprotozoal and antibacterial activity. The proper mechanism of FZD induced toxicity is still unclear. This study aimed to investigate the protective effect of curcumin on FZD induced oxidative stress, DNA injury and apoptosis in human hepatocyte L02 cells. The results showed that curcumin treatment significantly ameliorated FZD induced cytotoxicity, characterized by decreasing the production of reactive oxygen species (ROS) and malondialdehyde, as well as increasing superoxide dismutase, catalase activities and glutathione contents. Moreover, curcumin pretreatment significantly inhibited FZD induced the loss of mitochondrial membrane potential, the activation caspase-9 and -3 and apoptosis. Comet assay showed that curcumin attenuated FZD induced DNA injury in a dose-dependent manner. Correspondingly, curcumin markedly reversed the up-regulation of p53, Bax, caspase-9 and -3 mRNA expressions and the down-regulation of Bcl-2 mRNA (all p<0.05 or 0.01). These results reveal that curcumin protects against FZD induced oxidative stress, DNA injury and cell apoptosis via inhibiting oxidative stress and mitochondrial pathway, which may be attributed to ROS scavenging and anti-oxidative ability of curcumin. Importantly, our study highlights that curcumin may be a potential way to prevent FZD-mediated oxidative DNA injury and apoptosis in human or animals.

Psoralens have been used in the treatment of several skin diseases such as psoriasis and atopic eczema through PUVA (psoralen + UVA), cutaneous T-cell lymphoma through ECP (extracorporeal photochemotherapy), and recently, the treatment of solid tumors through X-PACT (x-ray psoralen activated cancer therapy). Ingesting psoralens and exposing the molecules to UVA light allows for the flat-planar psoralens to intercalate or slide in-between base pairs and then to form photo-adducts with the DNA backbone which can stop or slow DNA replication. The intrinsic ability of psoralens to bind with DNA and their therapeutic effectiveness contribute to the increased interest in the study of these molecules. Several studies measuring psoralen-DNA binding capabilities have used a variety of spectroscopic techniques and different experimental conditions. In this study, spectrofluorometry was used to measure the DNA-binding capabilities of three different psoralen derivatives (6E, AMT and 8-MOP) under identical experimental conditions. The binding constants were 0.325x106 M-1 for 8-MOP, 0.516x106 M-1 for AMT, and 7.30x106 M-1 for 6E. This direct comparison of the three different psoralens demonstrated the impact of their molecular structure on the DNA-binding capacity. 6E’s significantly greater binding constant makes it the stronger candidate for further studies looking into potential applications for cancer treatment.

Background: Breast cancer incidence and mortality exhibit a rising trend globally among both premenopausal and postmenopausal women suggesting that there are serious errors in our preventive and therapeutic measures.Purpose: Providing a series of valuable, but misunderstood inventions highlighting the role of increasing estrogen signaling in prevention and therapy of breast cancer instead of its inhibition. Results: 1. Breast cells and breast cancer cells with germline BRCA1/2 mutations similarly show defect in liganded estrogen receptor (ER) signaling demonstrating its role in genomic instability and cancer initiation. 2. In breast cancers, the increased expression of certain receptor family is an effort for self directed improvement of genomic defects, while the weakness or loss of receptors indicates a defect requiring medical repair. 3. ER overexpression in breast cancer cells is capable of strengthening estrogen signaling and DNA repair, while in ER negative tumors, HER2 overexpression tries to upregulate unliganded ER activation and genome stabilization. 4. ER positive breast cancers responsive to endocrine therapy may show a compensatory ER overexpression resulting in a transient tumor response. Breast tumors, non responsive to endocrine therapy, show HER2 overexpression for compensation the complete blockade of liganded ER activation. 5. In breast tumors, somatic mutations serve upregulation of liganded or unliganded activation of ERs helping genome stabilization and apoptotic death. 6. The dynamic communication between breast cancer and its microenvironment is a wonderful collaboration among molecular players fighting for genomic repair and apoptosis in the tumor. 7. In breast cancers, there is no resistance to genotoxic or immune blocker therapies, but rather the non responsive tumor cells exhausted all compensatory possibilities against therapeutic damages. Conclusion: Understanding the behavior and ambition of breast cancer cells may achieve a turn in therapy via applying a supportive care instead of genotoxic measures.

Inflammatory bowel disease (IBD), including Crohn’s Disease (CD) and Ulcerative Colitis (UC) are chronic multifactorial disorders which affect the gastrointestinal tract with variable extent. Despite extensive research, their etiology and exact pathogenesis are still unknown. Cell-free DNAs (cfDNAs) are defined as any DNA fragments which are free from origin cell and able to circulate into bloodstream with or without microvescicles. CfDNAs are now being increasingly studied in different human diseases, like cancer or inflammatory diseases. However, to date it is unclear how IBD etiology is linked to cfDNAs in plasma. Extrachromosomal circular DNA (eccDNA) are non-plasmidic, nuclear, circular and closed DNA molecules found in all eukaryotes tested. CfDNAs appear to play an important role in autoimmune diseases, inflammatory processes, and cancer; recently, interest has also grown in IBD, and their role in the pathogenesis of IBD has been suggested. We now suggest that eccDNAs also plays a role in IBD. In this review we have comprehensively collected available knowledge in literature regarding cfDNA, eccDNA, and structures involving them such as neutrophil extracellular traps and exosomes, and their role in IBD. Finally, we focused on old and novel potential molecular therapies and drug delivery systems, such as nanoparticles, for IBD treatment.

We discuss the role of water bridging the DNA-enzyme interaction by resorting to recent 1 results showing that London dispersion forces between delocalized electrons of base pairs of DNA 2 are responsible for the formation of dipole modes that can be recognized by Taq polymerase. 3 We describe the dynamical origin of the high efficiency and precise targeting of Taq activity in 4 PCR. The spatiotemporal distribution of interaction couplings, frequencies, amplitudes, and phase 5 modulations comprise a pattern of fields instantiating the electromagnetic image of DNA in its 6 water environment, which is what the polymerase enzyme actually recognizes at long range. The 7 experimental realization of PCR amplification, achieved through replacement of the DNA template 8 by the treatment of pure water with electromagnetic signals recorded from viral and bacterial DNA 9 solutions, is found consistent with the gauge theory paradigm of quantum fields.

A ~3-kb deletion-type DNA copy-number variation (CNV, esv3587290) located at intron 7 of the VANGL1 gene (1p13.1, MIM*610132) has been proposed as a genetic factor for developing lupus nephritis (LN) in adult systemic lupus erythematosus (SLE) patients across European-descent populations, but its replication in other ethnicities has been inconsistent and its association with LN in childhood-onset SLE (cSLE) remains unknown. Here, we performed an exploratory association study in a sample of 66 unrelated cSLE Mexican patients (11 males, 55 females; ages 7.8 to 18.6 years). Two stratified groups were compared: cSLE patients with (N=39) or without (N=27) LN, as diagnosed by renal biopsy (N=17), proteinuria (N=33), urinary protein:creatinine ratio >0.2 (N=34), and erythrocyturia and/or granular casts in urinary sediment (N=16). For esv3587290 CNV genotyping, we performed an end-point PCR assay with breakpoint confirmation by Sanger sequencing. We also determined the allelic frequencies of the esv3587290 CNV in 181 deidentified ethnically-matched individuals (reference group). The obtained genotypes were tested for Hardy-Weinberg equilibrium (HWE) by c2 test. Associations between LN and esv3587290 CNV were tested by calculating the Odds Ratio (OR) and using Pearson's c² tests with a 95% confidence interval and P≤0.05. The esv3587290 CNV allele (OR 0.108, 95% CI 0.034-0.33, P=0.00003) and the heterozygous genotype (OR 0.04, 95% CI 0.119-0.9811, P=0.002) showed a significant protective effect against LN development. Finally, we characterized the precise breakpoint of the esv3587290 CNV as NG_016548.1(NM_138959.3):c.1314+1339_1315-897del (https://databases.lovd.nl/shared/variants/0000918418#00025811) in our population. This report supports the notion that a broad genetic heterogeneity underlies the susceptibility for developing LN.

Background: Glioblastoma remains a challenging brain tumor to treat due to its infiltrative nature. Accurately identifying tumor boundaries during surgery is crucial for successful tumor resection. This study introduces an innovative intraoperative visualization method utilizing a surgical fluorescence microscope to precisely locate tumor cells. The focus of the study is on IR-Glint, a novel preparation comprising Cy7.5-labeled aptamers Gli-233nt and Gli-55_3L specific to human glioblastoma. Methods: The aptamers were modified using molecular modeling and quantum chemical techniques. The effectiveness of the preparation was assessed using flow cytometry and microscopy on primary cultures of human glioblastoma. In vivo studies were conducted on mouse and rabbit models, employing orthotopic xenotransplantation of human brain glioblastoma with various imaging techniques, including PET/CT, in vivo fluorescence visualization, confocal laser scanning, and surgical microscopy. Results: The experiments validated the potential of IR-Glint for intraoperative visualization of glioblastoma using infrared imaging. Surface application of the aptamer-based formulation on the brain allowed clear visualization of the tumor, aiding surgeons in tumor resection. This approach also reduces the dosage required and mitigates potential toxic effects on the patient. Conclusions: This study demonstrates the promising potential of using in-frared dye-labeled aptamers for intraoperative visualization of glioblastoma. By improving surgical treatment outcomes in neurosurgery, this novel approach may pave the way for future advancements in the field.

Hepatitis B virus (HBV) remains a dominant cause of hepatocellular carcinoma (HCC). Recently it was shown that HBV and woodchuck hepatitis virus (WHV) integrate into hepatocyte genome minutes after invasion. Retrotransposons and transposable sequences were frequent sites of the initial insertions suggesting a mechanism for spontaneous HBV DNA disperse throughout hepatocyte genome. Several somatic genes were also identified as early insertional targets in infected hepatocytes and woodchuck livers. Head-to-tail joints (HTJs) dominated amongst fusions indicating their creation by non-homologous-end-joining (NHEJ). Their formation coincided with robust oxidative damage of hepatocyte DNA. This was associated with activation of the poly(ADP-ribose) polymerase 1 (PARP1)-mediated dsDNA repair as reflected by augmented transcription of PARP1 and XRCC1, the PARP1 binding partner, OGG1, a responder to oxidative DNA damage, and by increased activity of NAD+, a marker of PARP1 activation, and HO1, an indicator of cell oxidative stress. The engagement of the PARP1-mediated NHEJ repair pathway explains HTJ format of the initial merges. The findings showed that HBV and WHV are immediate inducers of oxidative DNA damage, hijack dsDNA repair to integrate into hepatocyte genome and by this may initiate pro-oncogenic process. Tracking initial integrations may uncover early markers of HCC and help to explain HBV-associated oncogenesis.

Abstract: DNA fragmentation factors 40 and 45 (DFF40 and DFF45) and B-cell lymphoma 2 (Bcl-2) expression were evaluated in the normal human endometrium and myometrium. DFF40, DFF45, and Bcl-2 expression was assessed via immunohistochemistry in the proliferative, secretory, and atrophic endometrium and myometrium collected postmenopausally and premenopausally during the proliferative and secretory phases of the menstrual cycle. The endometrium showed significantly higher DFF40 and DFF45 expression than that in the uterine myometrium; compared to the stroma, endometrial glands showed the highest expression in pre- and postmenopausal specimens. Glandular expression of DFF45 was dependent on the menstrual cycle, reaching its highest level in the secretory endometrium. The glandular expression of DFF40 and DFF45 was significantly lower in postmenopausal specimens than that in premenopausal tissue. No cycle-dependent changes were reported for stromal or myometrial DFF40 or DFF45 expression. Compared to the endometrial stroma and myometrium, Bcl-2 showed the highest expression in the glandular proliferative endometrium and the lowest expression in the stromal secretory endometrium and myometrium during the secretory phase of the cycle. DFF45 and Bcl-2 showed menstrual cycle-dependent expression, which was limited to the glandular layer of the endometrium.

Gastric cancer is a heterogeneous group of diseases with only a fraction of patients responds to immunotherapy. The relationships between tumor DNA damage response, the immune system and immunotherapy have recently attracted attention. Accumulating evidence indicate that DNA repair landscape is a significant factor in driving response to immune checkpoint blockade (ICB) therapy. In this study, to explore new prognostic and predictive biomarkers for gastric cancer patients who are sensitive and responsible to immunotherapy, we developed a novel 15-DNA repair gene signature (DRGS) and its related scoring system and evaluated the efficiency of DRGS in discriminating different molecular and immune characteristics and therapeutic outcomes of gastric adenocarcinoma. The results showed that DRGS high score patients showed significantly better therapeutic outcomes compared to DRGS low score patients (P < 0.001). Integrated analysis of multi-omics data demonstrated that the patients with high DRGS score were characteristic of high levels of anti-tumor lymphocytes infiltration, tumor mutation burden (TMB) and PD-L1 expression, and these patients exhibited a longer overall survival and may benefit more from ICB therapy, as compared to the low-score patients. Therefore, the DRGS and its scoring system may have implications in tailoring immunotherapy in gastric cancers.

Schistosoma infection in snails can be monitored by microscopy or indirectly by sentinel mice. As both these approaches sometimes miss infections, more sensitive tests are needed, particularly in low-level transmission settings. In this study, the loop-mediated isothermal amplification (LAMP) technique, designed to detect a specific 28S ribosomal S. japonicum gene with high sensitivity, was compared to microscopy using snail samples from 51 areas endemic for schistosomiasis in five Chinese provinces. The results were compared with those by polymerase chain reaction (PCR) adding DNA sequencing as a reference when needed. The testing of pooled snail samples showed that a dilution factor of 1/50, i.e., one infected snail plus 49 non-infected ones, would still result in a positive reaction after the recommended number of amplification cycles. Testing a total of 232 pooled samples, emanating from 4,006 snail specimens, with the LAMP assay showed a 6.5% rate of infection, while traditional microscopy found only 0.04% positive samples in the same materials. Parallel PCR analysis confirmed the diagnostic accuracy of the LAMP assay, with DNA sequencing even giving LAMP a slight lead. Microscopy and the LAMP test were carried out at local schistosomiasis-control stations demonstrating that the potential of the latter assay to serve as a point-of-care (POC) test with results available within 60–90 minutes, while the more complicated PCR test had to be carried out at the National Institute of Parasitic Diseases (NIPD) in Shanghai, China. In conclusion, LAMP was found to be clearly superior to microscopy and as good as, or better, than PCR. Application of LAMP testing would be useful for surveillance and risk prediction as it requires less time than other techniques and can be used under field conditions, which improves and accelerates schistosomiasis control.

Nucleocytoplasmic large DNA viruses (NCLDVs) have massive genomes and particle sizes compared to other known viruses. NCLDVs, including poxviruses, encode ATPases of the FtsK/HerA superfamily to facilitate genome encapsidation. However, their biochemical and structural characteristics are yet to be discerned. In this study, we demonstrate that the viral ATPases are significantly shorter than their bacterial homologs, representing the minimal ATPase core of the FtsK/HerA superfamily. We analyzed the sequence and structural features of the vaccinia virus A32 protein and determined their roles in the stability of the A32 protein and its ATPase activity. We sought to purify A32 by various techniques and noted that recombinant A32 expressed in E. coli is highly insoluble and unstable in solution, possibly because of the disordered C-terminus. N-terminal fusion with the thioredoxin solubility tag could alleviate this issue to some extent, but subsequent tag cleavage results in increased susceptibility to precipitation and degradation. We also predicted a conserved coiled-coil motif (CCM) towards the C-terminus of VV A32. ATPase activity of A32 is known to increase in the presence of DNA. Our comparative analysis of wildtype A32 versus CCM mutants suggests that this DNA dependence of A32’s ATPase activity is likely regulated by the CCM. Since CCM is also known to facilitate protein oligomerization, these findings provide new opportunities for further detailed characterization of A32.

Acute disseminated encephalomyelitis (ADEM) is an inflammatory demyelinating disorder of the white matter. Pathophysiology is thought to be immune mediated as in most cases the condition follows an infection or triggering incident. More recent literature has demonstrated that there may be a link between autoimmune conditions and ADEM. Here we present a case of ADEM in a middle-aged woman with systemic lupus erythematosus that recovered well after treatment with corticosteroids and Rituximab.

As the fast growth of information technology (IT), the safety of image transmission and the storing of images are increasingly concerned. Traditional image encryption algorithms have certain limitations in transmission and security, so there is an urgent need for a secure and reliable image encryption algorithm. A universal compression sensing (CS) image encryption (IE) algorithm based on DNA-triploid mutation (DTM) is presented in this paper. Firstly, by using CS algorithm an image is compressed while obtaining a range of chaotic sequences by iteration of chaotic map. Then DNA sequences are generated by encoding the image, and based on the DTM, new mutant DNA sequences are generated according to specific rules. Next, the chaotic sequences are operated at the DNA level to do confusion and diffusion operations on the image to ensure the security of the data. At last, DNA decoding is carried out to obtain the compressed-encrypted image. Both simulation experiments and performance tests fully show that a high level of security and reliability of the proposed algorithm in protecting image privacy is achieved.

Plant DNA extraction techniques are well-established, featuring a variety of protocols tailored to the needs of individual laboratories or research goals. Currently, there is a strong demand for plant DNA extraction methods that are rapid, cost-effective, and uncomplicated. We have devised an optimized protocol for plant DNA extraction that eliminates the need for liquid nitrogen. DNA was extracted from leaves of Inga feuilleei (pacae) in the Cañete Valley using a basic cetyltrimethylammonium bromide method. Subsequently, the extracted DNA was quantified and assessed for quality through agarose gel electrophoresis. This protocol holds the potential to significantly reduce costs in laboratories engaged in routine DNA extraction for genetic diversity and gene expression analyses. It addresses challenges encountered by other conventional methods, particularly in situations where they have proven ineffective due to issues related to explant, sample conditions, and the quantity and quality of nucleic acids. This is especially pertinent for numerous laboratories in developing countries where constraints related to the cost and availability of liquid nitrogen may be a limiting factor. Furthermore, this is the first study reported in this species for genomic studies.

Type 2 diabetes mellitus (T2DM) is a risk factor for male infertility, but the underlying molecular mechanisms remain unclear. Advanced glycation end products (AGEs) are pathogenic molecules for diabetic vascular complications. Here, we investigated the effects of DNA-aptamer raised against AGEs (AGE-Apt) on testicular and sperm abnormalities in a T2DM mouse model. KK-Ay (DM) and wild-type (non-DM) 4- and 7-week-old male mice were sacrificed to collect the testes and spermatozoa for immunofluorescence, RT-PCR, and histological analyses. DM and non-DM 7-week-old mice were subcutaneously infused with AGE-Apt or control aptamer for 6 weeks and then sacrificed. Plasma glucose, testicular AGEs, and RAGE gene expression in 4-week-old DM mice and plasma glucose, testicular AGEs, oxidative stress, and pro-inflammatory gene expressions in 7-week-old DM mice were higher than those in age-matched non-DM mice; the latter of which was associated with seminiferous tubular dilation. AGE-Apt did not affect glycemic parameters, but it inhibited the seminiferous tubular dilation, reduced the number of testicular macrophages and apoptotic cells, and restored the decrease in sperm concentration, motility, and viability of 13-week-old DM mice. Our findings suggest that AGEs-Apt may improve sperm abnormality by suppressing the AGE–RAGE-induced oxidative stress and inflammation in the testes of DM mice.

Much research has been conducted to determine how hair regeneration is regulated, as this could provide therapeutic, cosmetic, and even psychological interventions for hair loss. The current study focused on the hair growth effect and effective utilization of fatty oil obtained from Bryde's whales through high-throughput DNA microarray approach in conjunction with immunohistochemical observations. The research also examined the mechanisms and factors involved in hair growth. In an experiment using female C57BL/6J mice, the vehicle control group (VC: propylene glycol: ethanol: water), the positive control group (MXD: 3% minoxidil), and the experimental group (WO: 20% Whale Oil) were topically applied to the back of the mouse. The results showed that 3% MXD and 20% WO were more effective than VC in promoting hair growth, especially 20% WO. Furthermore, in hematoxylin and eosin-stained skin tissue, an increase in the number of hair follicles and subcutaneous tissue thickness was observed with 20% WO. Whole-genome transcriptomic analysis also showed that 20% WO may have lower stress- and inflammation-related responses than 3% MXD. Therefore, whale oil can be expected to be used as a safe hair growth agent.

L-Theanine, a unique non-protein amino acid, is an important bioactive component of green tea. Previous studies have shown that L-theanine has many potent health benefits, such as anti-anxiety effect, regulation of immune response, relaxing neural tension, reducing oxidative damage. However, little is known concerning whether L-theanine can improve the clearance of mitochondrial DNA (mtDNA) damage in organisms. Here, we reported that L-theanine treatment increased ATP production and improved mitochondrial morphology to extend the lifespan of UVC-exposed nematodes. Mechanistic investigations showed that L-theanine treatment enhanced the removal of mtDNA damage and extended lifespan by activating autophagy, mitophagy, mitochondrial dynamics, and mitochondrial unfolded protein response (UPRmt) in UVC-exposed nematodes. In addition, L-theanine treatment also up-regulated the expression of genes related to mitochondrial energy metabolism in UVC-exposed nematodes. Our study provided a theoretical basis for the possibility that tea drinking may prevent mitochondrial related diseases.

Abstract: Congenital cytomegalovirus (cCMV) infection poses significant risks to fetal development, particularly affecting the nervous system. This study reports a fetal autopsy case, examining cCMV infection and focusing on CMV DNA measurements in various fetal organs before formalin fixation, a novel approach for comprehensive CMV DNA evaluations in fetal organs affected by cCMV. A 20-week-old male fetus was diagnosed with cCMV following the detection of CMV DNA in ascites obtained via abdominocentesis in utero. After termination of pregnancy, multiple organs of the fetus, including the cerebrum, thyroid gland, heart, lungs, liver, spleen, kidneys, and adrenal glands, were extracted and examined for CMV DNA loads using real-time polymerase chain reaction. Histopathological examination involved hematoxylin-eosin and CMV-specific immunostaining. A correlation was found between CMV DNA loads and pathology, with higher CMV-infected cell numbers observed in organs positively identified with both staining methods, exhibiting CMV DNA levels of ≥1.0 × 104 copies/mL, compared to those detected solely by CMV-specific immunostaining, where CMV DNA levels ranged from 1.0 x 103 to 1.0 x 104 copies/mL. These results highlight a quantifiable relationship between organ infection extent and CMV DNA concentration, providing insights into cCMV pathogenesis and potentially informing future diagnostic and therapeutic strategies for cCMV infection.

The epigenetic mechanism of DNA methylation is essential for regulating gene expression and adaptation in plants, especially when exposed to various environmental stressors such as drought, heat, and nutrient availability. The impact of these stressors can lead to alterations in DNA methylation patterns and gene expression, which this study aims to investigate. Specifically, we are interested in understanding how DNA methylation plays a role in controlling the expression of stress-response genes, as well as genes involved in plant growth and development. To accomplish this, we are analyzing the changes in DNA methylation patterns and gene expression in response to different stressors like drought and heat. Our research findings suggest that DNA methylation is a vital regulator of plant adaptation to environmental stress, and we hope to gain further insight into these mechanisms by identifying genes that exhibit differential expression in response to stress and analyzing their DNA methylation status. This study offers valuable insights into the development of crops that can thrive under changing climate conditions.

The molecular mechanism how the infecting DNA of bacteriophage T4 is passing from the capsid through the bacterial cell wall and entering the cytoplasm is essentially unknown. After adsorption, the short tail fibers of the infecting phage extend from the baseplate and trigger the contraction of the tail sheath, leading to a puncturing of the outer membrane by the tail tip needle composed of the proteins gp5.4, gp5 and gp27. To explore the events that occur in the periplasm and at the inner membrane, we have constructed T4 phage that have a modified gp27 in their tail tip with a His-tag. Shortly after infection with these phages, the cells were chemically cross-linked, solubilised, and the cross-linked products were affinity purified on a nickel column. The co-purified proteins were identified by mass spectrometry, and we found that predominantly the inner membrane proteins DamX, SdhA and PpiD were cross-linked. The same partner proteins were identified when purified gp27 was added to Escherichia coli spheroplasts suggesting a direct protein-protein interaction.

The genus Colias Fabricius, 1807 includes numerous taxa and forms with uncertain status and taxonomic position. Among such taxa are Colias mongola Alphéraky, 1897 and Colias tamerlana Staudinger, 1897, interpreted in the literature either as conspecific forms, as subspecies of different but morphologically somewhat similar Colias species, or as distinct species-level taxa. Based on mitochondrial and nuclear DNA markers, we reconstruct a phylogeographic pattern of the taxa in question. We recover and include in our analysis DNA barcodes of the century-old type specimens, the lectotype of C. tamerlana deposited in the Natural History Museum (Museum für Naturkunde), Berlin, Germany (ZMHU), the paralectotype of C. tamerlana and the lectoptype of C. mongola deposited in the Zoological Institute, Russian Academy of Sciences, Saint-Petersburg, Russia (ZISP). Our analysis grouped all specimens within four (HP_I-HP-IV) deeply divergent but geographically poorly structured clades which did not support nonconspecifity of C. mongola–C. tamerlana. We also show that all studied females of the widely distributed haplogroup HP_II were infected with a single Wolbachia strain belonging to the supergroup B, while the males of this haplogroup, as well as all other investigated specimens, were not infected. Our data underline the relevance of large-scale sampling dataset analysis and the need of testing for Wolbachia infection to avoid erroneous phylogenetic reconstructions and species misidentification.

By definition, a cell is the smallest unit of biological life. This means that every living thing can be fundamentally narrowed down to a composition of cells working together for the furtherance of that being in existence. With the dawn dawn of the evolutionary theory that bespeaks of life commencing some 〜3.8 billion years ago with RNA strands that underwent evolution to become DNA, which led to what we have now as life in the world, or with the Panspermia hypothesis, assuming that life on Earth was seeded from space, what is being considered in this work pertains to the origin of life. In addition to the plethora of intuitive stances that concern the origin story of life, this work will only add to the these. The major hypothesis in this work is that the continuity of life, as pertaining to its procession from parent to offspring provides fundamental links to the origin of life and this is the major consideration that will be given. It is thus apparent that the cells that would be considered are germ cells and not somatic cells. A cursory look at the continuity of life, should give us clues as regards how life originated. We nonetheless, take into consideration the fact that some features due to its non-use have gone extinct, thus taking with them the needed knowledge to breaking even as regards our inquiry. Even at that, we propose that conjectures, hypothetically, can be made based on the physical evidence that is available to us today.

Bifenthrin (BF) is a new highly effective broad-spectrum insecticide that has gained widespread use. However, there is limited research on the potential toxic effects of bifenthrin pollution on amphibians. In this research, Chinese giant salamanders (CGS) were exposed to three different concentrations of BF (0, 0.04, and 4 g/L BF) for 1-week to investigate the toxic effects. Behavioral changes, liver pathology, oxidative stress factors, DNA damage, and transcriptome differences were observed and analyzed. The results showed that exposure to BF (0.04 and 4) significantly decreased the adenosine-triphosphate (ATP), superoxide dismutase (SOD), glutathione (GSH), catalase (CAT), and 8-hydroxydeoxyguanosine (8-OHdG) contents in the brain, liver, and kidney of CGS, whereas the malondialdehyde (MDA) and reactive oxygen species (ROS) contents were increased. The liver tissue showed considerable inflammatory reactions and structural malformations. RNAseq analysis of the liver showed that BF caused abnormal antioxidant indices of CGS, which affected molecular function genes such as catalytic activity, ATP-dependent activity, metabolic processes, signaling and immune system processes, behavior and detoxification significantly up-regulated, resulting the differential genes were significantly enriched in calcium signaling pathway, PPARα signaling pathway and NF-kB signaling pathway. These results indicated that BF induced the abnormal production of free radicals to exceed the potential of the self-defense system, causing different degrees of oxidative damage and lipid peroxidation in the body, resulting in oxidative stress, abnormal lipid metabolism, DNA damage, tissue inflammation, and abnormal behavior. This work provides a theoretical basis for the rational use of pyrethrin and environmental risks assessment, and scientific guidance for the conservation of amphibian populations.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease with a complex genetic architecture, showing monogenic, oligogenic, and polygenic inheritance. In this study, we describe the case of a 71 years-old man diagnosed with ALS with an atypical phenotype, due to the presence of a progressive ocular ptosis. Genetic analyses revealed two variants in SOD1 and TBK1 genes respectively and the LHON-associated m.14484T>C variant in the mitochondrial DNA (mtDNA). We discuss how all these variants may synergically impinge on mitochondrial function, contributing to the pathogenic mechanisms which might ultimately lead to the neurodegenerative process, possibly influencing the clinical ALS phenotype.

The life cycles and biodiversity of Pacific coast gastropods were analyzed by videomicroscopy and DNA barcoding of indi-viduals collected from tide pools and in plankton nets from a variety of shore stations. In many species (Families Calyptrae-idae, Cerithiopsidae, Strombidae, Vermetidae, Columbellidae, Nassariidae, Olivellidae, Hermaeidae, Onchidorididae, Gas-tropteridae, Haminoeidae), the free-swimming veligers were recovered from plankton collections; in Roperia poulsoni (family Muricidae) veligers were usually recovered from egg sacs where they had been retained although some escapees were found in plankton collections; in Pteropurpura festiva (family Muricidae) free-living veligers were also found; and in Atlanta californiensis (family Atlantidae) both veligers and adults were obtained from plankton collections making this a holoplank-tonic species. The results confirm that DNA barcoding using the COI barcode is a useful strategy to match life-cycle stages within species as well as to identify species and to document the level of biodiversity within the gastropods.

Alzheimer’s disease (AD) is a genetically complex senile neurodegeneration with unknown etiology. The first gene discovered to be mutated in early-onset AD, the amyloid precursor protein (APP), has been widely assumed as a causal factor in the disease cascade due to its generation of Aβ species. APP has an evolutionarily conserved biological role and activates a signaling program with notable similarities to integrin—a cell adhesion receptor with a wide array of functions. Intriguingly, several AD genome-wide association study (GWAS) candidate genes, including the SHARPIN locus recently reported by us and others, influence signaling of the integrin pathway. Integrins are focal adhesion regulators and serve in nervous system development, synaptic plasticity, and Tau phosphorylation. These observations suggest that the function of APP probably goes beyond Aβ generation in AD. Aging—the strongest risk factor for AD—is associated with various clock-like events in cells. For instance, neurons are continuously impacted by stochastic ‘hits’ to their genomes in aging, in the forms of DNA damage, insertion-deletions, copy-number variations (CNVs) and other types of somatic mutations. DNA damage and somatic mutations can result in neoplastic changes and cancer in mitotically active cells. However, their consequences in post-mitotic cells such as aging neurons are less defined. The current hypothesis holds that the stochastic loss of DNA sequence data at random loci in aging affects longer genes by chance more frequently. As a result, the biological processes coordinated by long genes may be more vulnerable to such random aging effects. Curiously, as shown by us and others, long genes are strongly enriched for synapse- and cell adhesion-related ontologies, more than any other biological process or cellular compartment. In addition, among various cell types, neurons possess the highest levels of long gene expression and are therefore more vulnerable to such harmful effects. The long gene vulnerability hypothesis provides a simple link between aging and the genetic landscape of AD and warrants new strategies for disease modification.

Genetic and sporadic forms of tauopathies, the most prevalent of which is Alzheimer’s Disease, are a scourge of the aging society, and in case of genetic forms, can also affect children and young adults. All tauopathies share ectopic expression, mislocalization, or aggregation of the microtubule associated protein TAU, encoded by the MAPT gene. As TAU is a neuronal protein widely expressed in the CNS, the overwhelming majority of tauopathies are neurological disorders. They are characterized by cognitive dysfunction often leading to dementia, and are frequently accompanied by movement abnormalities such as parkinsonism. Tauopathies can lead to severe neurological deficits and premature death. For some tauopathies there is a clear genetic cause and/ or an epigenetic contribution. However, for several others the disease etiology is unclear, with few tauopathies being environmentally triggered. Here we review current knowledge of tauopathies listing known genetic and important sporadic forms of this disease. Further, we discuss how DNA methylation as a major epigenetic mechanism emerges to be involved in the disease pathophysiology of Alzheimer’s, and related genetic and non-genetic tauopathies. Finally, we debate the application of epigenetic signatures in peripheral blood samples as diagnostic tool and usage of epigenetic therapy strategies for these diseases.

Single-stand selective monofunctional uracil DNA glycosylase 1 (SMUG1) works to remove uracil and certain oxidized bases from DNA during base excision repair (BER). This review provides a historical characterization of SMUG1 and 5-hydroxymethyl-2'-deoxyuridine (5-hmdU) one important substrate of this enzyme. Biochemical and structural analyses provide remarkable insight into the mechanism of this glycosylase revealing SMUG1 has a unique helical wedge which influences damage recognition during repair. Rodent studies suggest that, while SMUG1 shares substrate specificity with another uracil glycosylase UNG2, loss of SMUG1 can have unique cellular phenotypes. This review highlights the multiple roles SMUG1 may play in preserving genome stability, and how the loss of SMUG1 activity may promote cancer. Finally, we discuss recent studies indicating SMUG1 has moonlighting functions beyond BER, playing a critical role in RNA processing including the RNA component of telomerase.

Geckos demonstrate a remarkable variability in sex determination systems, but our limited knowledge prohibits accurate conclusions on the evolution of sex determination in this group. Eyelid geckos (Eublepharidae) are of particular interest, as they encompass species with both environmental and genotypic sex determination. We identified for the first time the X-specific gene content in the Yucatán banded gecko, Coleonyx elegans, possessing X1X1X2X2/X1X2Y multiple sex chromosomes by comparative genome coverage analysis between sexes. The X-specific gene content of Coleonyx elegans was revealed to be partially homologous to genomic regions linked to the chicken autosomes 1, 6 and 11. A qPCR-based test was applied to validate a subset of X-specific genes by comparing the difference in gene copy numbers between sexes, and to explore the homology of sex chromosomes across 11 eublepharid, two phyllodactylid and one sphaerodactylid species. Homologous sex chromosomes are shared between Coleonyx elegans and Coleonyx mitratus, two species diverged approximately 34 million years ago, but not with other tested species. As far as we know, the X-specific gene content of Coleonyx elegans / Coleonyx mitratus was never involved in the sex chromosomes of other gecko lineages, indicating that the sex chromosomes in this clade of eublepharid geckos evolved independently.

Drug abuse is a major issue worldwide. Methylphenidate (MPH) and alprazolam (ALZ) are commonly prescribed drugs for the treatment of ADHD and anxiety disorders, respectively. The limited studies suggest that abusers primarily use benzodiazepines to counteract adverse effects associated with methylphenidate usage. The main aim of this study was to investigate the interaction of drugs with DNA using spectroscopic methods. Female Wistar rats were administered with MPH (10, 20, 40 mg/kg) and ALZ (5, 10, 20 mg/kg) alone and in combination for a period of 28 days. The FT-IR and UV results reveal some spectral changes in a dose-dependent manner, which indicates interactions of drugs with DNA. Thus, the changes in spectral peaks provide some insight into the mechanism of the interaction of drugs with DNA.

Neutrophils play an important role as the first line of innate immune defense. One function of neutrophils, called neutrophil extracellular traps (NETs), has been discovered recently. NETs are extensive fibrous structures released extracellularly from activated neutrophils in response to infection. They are composed of cytosolic protein assembled on a scaffold of released chromatin. These structures suppress the dissemination of micro-organisms in blood by trapping them mechanically, and by exploiting coagulant function to segregate them within the circulation. In addition, NET components (DNA, histone, and granule proteins) also contribute to the triggering of an inflammatory process. NET function, however, can be regarded as a double-edged sword. On one hand, NET formation is an efficient strategy for neutralizing invading micro-organisms. On the other hand, NET can be harmful to the host, as its exposed by-products that are toxic to endothelial cells and parenchymal tissue. We present here the analogous biological and physiological features of the harmful positive amplification loop between inflammation and tissue damage induced by NETosis dysregulation and Coronavirus Disease-2019 (COVID-19) pathogenesis. Considering the rapid evolution of this disease symptoms and its lethality, we hypothesize that COVID-19 progresses under an amplifier loop, leading to an massive, uncontrolled inflammation process. We also describe the correlations of COVID-19 symptoms and biological features with those consecutive to uncontrolled NET formation causing various sterile or infectious diseases. General clinical conditions, and numerous pathological and biological features, are analogous with NETs deleterious effects. We postulate that Severe Acute Respiratory Syndrome-Coronavirus 2 (SARS-CoV2) induces a disproportionate virus-induced NET release, and that this plays a key role in COVID-19 pathogenesis. While neutrophils are the principal starting point for extracellular and circulating DNA release, targeting NETs rather than neutrophils themselves may stand for an effective strategy. This paper offers an in-depth review of NET formation, function and pathogenic dysregulation, as well as of current and future therapies to control NET unbalance. As such, it enables us also to suggest new therapeutic strategies to fight COVID-19. In combination with or independent of the latest tested approaches, we propose that, in the short term, deoxyribonuclease I (DNase-1) treatment should be evaluated; we also advocate a significant increase in research on the development of toll-like receptors (TLR) and C-type Lectin like receptors (CLEC) inhibitors, and on anti-IL26 therapies.

A decade after environmental scientists integrated high-throughput sequencing technologies in their toolbox, the genomics-based monitoring of anthropogenic impacts on biodiversity and ecosystems is yet to be implemented by regulatory frameworks. Despite the broadly acknowledged potential of environmental DNA and RNA to cost-efficiently and accurately monitor biodiversity, technical limitations and conceptual issues still stand in the way of its routine application by end-users. In addition, the multiplicity of potential implementation strategies may contribute to a perception of the methodology as being premature or “in development”, hence restraining regulators from binding these tools into legal frameworks. This review focuses on the strengths and limitations of genomics-based strategies that have emerged over the past ten years and have been classified for this purpose into three broad strategies: (A) Taxonomy-based approaches that focus on known bio-indicators or the diversity of taxonomically described taxa, (B) De novo approaches that do not require well-established taxonomy, and (C) Function-based approaches that rely on community-wide metrics, where taxa are interchangeable, or on functional profiles instead of compositional turnovers. We finally propose a roadmap for the implementation of environmental genomics into routine monitoring programs that leverage recent analytical advancements, upon which some critical limitations are alleviated.

Population genetic data underpin many studies of behavioral, ecological, and evolutionary processes in wild populations and contribute to effective conservation management. However, collecting genetic samples can be challenging when working with endangered, invasive, or cryptic species. Environmental DNA (eDNA) offers a way to sample genetic material non-invasively without requiring visual observation. While eDNA has been trialed extensively as a biodiversity and biosecurity monitoring tool with a strong taxonomic focus, it has yet to be fully explored as a means for obtaining population genetic information. Here, we review current research that employs eDNA approaches for the study of populations. We outline challenges facing eDNA-based population genetic methodologies, and suggest avenues of research for future developments. We advocate that with further optimizations, this emergent field holds great potential as part of the population genetics toolkit.

Micro-pollutants such as 17β-Estradiol (E2) have been detected in different water resources and their negative effects on the environment and organisms have been observed. Aptamers are established as a possible detection tool, but the underlying ligand binding is largely unexplored. In this study, a previously described 35-mer E2-specific aptamer was used to analyse the binding characteristics between E2 and the aptamer with a MD simulation in an aqueous medium. Because there is no 3D structure information available for this aptamer, it was modeled using coarse-grained modeling method. The E2 ligand was positioned inside a potential binding area of the predicted aptamer structure, the complex was used for an 25 ns MD simulation, and the interactions were examined for each time step. We identified E2-specific bases within the interior loop of the aptamer and also demonstrated the influence of frequently underestimated water-mediated hydrogen bonds. The study contributes to the understanding of the behavior of ligands binding with aptamer structure in an aqueous solution. The developed workflow allows generating and examining further appealing ligand-aptamer complexes.

Background: KDM5 enzymes are H3K4 specific histone demethylases involved in transcriptional regulation and DNA repair. These proteins are over-expressed in different kinds of cancer, including breast, prostate and bladder carcinoma, with positive effects on cancer proliferation and chemo-resistance. For these reasons, these enzymes are potential therapeutic cancer targets. Methods: In the present study, we analyzed the effects of three different inhibitors of KDM5 enzymes in MCF-7 breast cancer cells over-expressing JARID1B. In particular we tested H3K4 demethylation (western blot); target gene transcription (RNAseq and real time PCR); radio-sensitivity (citoxicity and clonogenic assays) and damage accumulation (kinetics of H2AX phosphorylation). Results: we show that two compounds with completely different chemical structure can selectively inhibit KDM5 enzymes and that both compounds are capable of increasing sensitivity of breast cancer cells to ionizing radiation and H2AX phosphorylation. Conclusions: These findings confirm the involvement of H3K4 specific demethylases in DNA damage signaling and repair and suggest new strategies for the therapeutic use of their inhibitors.

Zucchini (Cucurbita pepo subsp. pepo) is a seasonal vegetable with high nutritional and medical value. Many useful properties of this fruit are attributed to its bioactive compounds. Zucchini fruits (“Yellow” and “Light Green” varieties) and four distinctive components (lutein, β-carotene, zeaxanthin and dehydroascorbic acid)were selected. Firstly it was determinated the lutein, β-carotene, zeaxanthin and dehydroascorbic acid content in these fruits and then, in order to measure the safety and suitability of their use different assays were carried out: (i) Genotoxicity and antigenotoxicity tests to determine the safety and DNA-protection against hydrogen peroxide; (ii) cytotoxicity and (iii) DNA fragmentation assay to evaluate the proapoptotic effect. Results showed that: (i) all the substances were non-genotoxic, (ii) all the substances were antigenotoxic except the highest concentration of lutein, (iii) “Yellow” Zucchini epicarp and mesocarp exhibited the highest cytotoxic activity (IC₅₀ > 0.1 mg/mL and 0.2 mg/mL, respectively) and iv) “Light Green” Zucchini skin and pulp induced internucleosomal DNA fragmentation . To sump up, Zucchini fruit could play a positive role in human health and nutrition due to this fruit and its components were safe, able to inhibit significantly the H₂O₂-induced damage and exhibit antiproliferative and pro-apoptotic properties toward HL60 tumour cells. The information generated from this research should take into account to select potential accessions for breeding programs purposes.

The evidential value of an mtDNA match between biological remains and its potential donor is determined by the random match probability of the haplotype. This probability is based on the haplotype’s population frequency estimate. Consequently, implementing a population study representative of the population relevant to the forensic case is vital to correctly evaluating the evidence. The emerging number of poaching cases and the limited availability of such data emphasizes the need for improved fallow deer mtDNA population databank for forensic purposes, including targeting the entire mitochondrial control region. By sequencing the whole mitochondrial control region in 138 animals from five populations in Hungary we found four different haplotypes, including one which has not yet been described. Our results, supplemented with data already available from previous research do not support the possibility of determining the population of origin, although some patterns of geographical separation can be distinguished. Estimates of molecular diversity indicate approximately similarly low mtDNA diversity (Hd=0.565 and π = 0.002), compared with data from other countries. The calculated random match probability of 0.547 shows a high probability of coincidence and, therefore, a limited capacity for exclusion. Consequently, it may be necessary to search for polymorphic sites on other mitochondrial gene sections or to sequence the entire mitogenome of fallow deer.

The study presents a mathematical modeling for supporting the premise that our body is composed of blockchain systems. A cell population is considered a space with peer-to-peer communication networks for applying blockchain protocol. Transaction is defined as gene expression that constantly occurs for protein synthesis in each cell. The transaction process proceeds according to the blockchain protocol with sharing and recording the data on the blockchain ledger. The premise comes from the clues of the previous research such that the cell population is a complex structured network system having cell-to-cell connections. Although individual cells exhibit stochastic nonlinear dynamic behavior, cell population shows consensus behavior that reaches to ensemble through interaction among cells. It is inferred that gene expression is not regulated by the corresponding cell only nor determined by external intelligence such as the brain. This is achieved through a consensus through stochastic interactions between cells over the whole cell population. These findings imply that mathematical blockchain modeling is a suitable for gene expression process. The original contribution of the study is a methodology for applying mathematically the blockchain protocol to the real biological gene expression process. In other words, the DNA sequence is converted into a numeral bit sequence that makes it possible to implement the blockchain protocol. A new DNA sequence scheme is proposed with adding methylated cytosine and adenine as the 5th and 6th bases for including epigenetic information which has profound effect on gene expression and regulation. The methodology was applied to the real biological synthesis process of protein samples. The protein is composed of amino acids that are encoded by triplet codons of 216 kinds with 6 base RNA sequence. The gene expression information is traced backward from a synthesized protein sample, amino acids of codons, RNA transcript up to DNA sequence. One of the results is a numeric value in the form of a bit sequence with which mathematical blockchain modeling is applicable. The cryptographic authentication and the consensus process are mathematically proven to work properly by the blockchain protocol. It implies that the same protein is synthesized, but with different epigenetic data, then protein's latent material properties will certainly be different. Although the result has not been justified by the biologic experiment yet, it is sure that the biological hidden algorithm inside DNA sequence will be revealed by the binary bit-logic with physical on/off states which is mathematically proven. The research will greatly contribute to disease treatment and medicine development as well as epigenetics in the future.

Abstract: Background: Gender dimorphism and marital status on colorectal cancer mortality have 9 been previously documented, but the relationship between these factors and DNA mismatch repair 10 protein (MMRP) expression status is unknown. Methods: Colectomy specimens were reviewed ret- 11 rospectively for patients between 2018 to 2023 with demographics including race/ethnicity, gender, 12 marital status, faith, body mass index, pathologic staging, and MMRP expression status. Statistical 13 analyses were performed by using baseline characteristics tables and various programs in R-pack- 14 age. Results: A total 1018 colectomies were reviewed, and the tumor stages were significantly higher 15 in right colon (stage 3 and 4) than those in left colon and rectosigmoid colon (p<0.01). Marital status 16 was significantly associated with patients’ gender, age, tumor size and tumor stages (all p<0.01). 17 MMRP status were available in 775 cases, and 139 (17.9%) were MMRP deficient, and 636 (82%) 18 MMRP proficient. MMRP deficiency was significantly associated with older female patients, larger 19 tumor in size, higher tumor stages, higher histologic grade and more common in right colon (all 20 p<0.01). In addition, MMRP deficiency was statistically associated with higher percentage of di- 21 vorced and widowed patients (p<0.01). Multivariate linear regression analysis revealed persistent 22 association of MMRP deficiency with tumor size, tumor grade, tumor stage and nodal metastasis, 23 but the associations with gender and marital status no longer existed. Conclusion: The differences 24 in prevalence of CRC by gender and marital status and tumor MMRP status illustrate the im- 25 portance of these factors on tumor stages and nodal metastasis but these associations were more 26 complex with other confounding factors.

Bone injury is a common side effect of radiotherapy to tumors, which is a long-term response after damage to osteoblasts, especially reducing osteoblasts proliferation and differentiation. Currently, there are few studies on radiation-induced bone injury, the molecules involved in the ionizing radiation (IR) induced osteoblasts damage remain need to be excavated. In this study, the optimal IR damage conditions (8Gy, 2.22Gy/min) were firstly determined by measuring cell proliferation, cell cycle, cell apoptosis and further cell differentiation and mineralization abilities and the related genes (p-AKT, p-ERK1/2, cyclinB, BAX, BCL2, ALP, OPN, RUNX2, Collagen1) expression level changes in radiation-induced osteoblast injury model. Then, we screened 26 differentially expressed genes after the RNA-sequencing of the 8 Gy-irradiated MC3T3-E1 cells, and they were mainly involved in DNA damage and repair, cell apoptotic progress and cell cycle regulation, meanwhile, participated in several main pathway including PI3K-AKT signaling pathway, p53 signaling pathway and signaling pathway involved in cell cycle and cell senescence. We focused on verifying the differential expression genes and confirmed the MDM2, NOTCH1, CDKN1A and GCLC were upregulated after IR treatment, suggesting the key roles in the response of the IR. In addition, regarding on our sequencing results, the DNA damage and repair were also verified and the results suggested that IR induced DNA damage and repair in MC3T3-E1 cells. In summary, IR damaged MC3T3-E1 cells by inhibiting cell proliferation, impacting cell cycle process, inducing cell apoptosis and affecting the osteoblasts differentiation, which maybe due to the DNA damage and the differential expression of the key genes (MDM2, NOTCH1, CDKN1A and GCLC ).

Autism spectrum disorder (ASD) is a complex neurodevelopmental disorder, with a highly variable expression of phenotypes (restricted interest or activity and repetitive behavior in communication and social interactions), genes (mutation), markers (alteration of transcription) and pathways. Loss of function of the CC2D1A gene appears to primarily affect the brain, leading to a range of behavioral problems in humans. In our study published in 2020; we found that the expressions of miR-19a-3p, miR-361-5p, miR-150-5p, miR-3613-3p, miR-126-3p and miR-499a-5p were downregulated in the serum samples of autistic patients and their families. Here, acquired non-Mendelian hereditary character in a genetically defined mouse model of autism (Cc2d1a +/-) correlates with the transcriptional alteration of five miRNAs. We seek to test the hypothesis that miRNA levels are varying by changes in RNA/DNA structure during development, thereby creating transcription alteration. Behavioral tests were conducted on the offspring of Cc2d1a +/- mutant and control mice, such as novel object, social interaction, marble burying and tail suspension behavior. Two RNA fractions were isolated from mouse hippocampal tissues and sperm cells by standard TRIzol extraction: total RNA and fraction of RNA bound to DNA in the form of a DNA/RNA hybrid (R-loop). The expression levels of miR-19a-3p, miR-361-5p, miR-150-5p, miR-126-3p and miR-499a-5p were investigated by quantitative real-time RT-PCR. We report differences in the distribution of five miRNAs in the hippocampus between male and female mice in RNA fractions, particularly in colonies of Cc2d1a +/- mice. Furthermore, the number of miRNAs engaged in the DNA/RNA hybrid fraction is generally higher in the mutant pedigree than in the control group. On the other hand, in sperm both fractions are at lower levels than in controls. R-loops contribute to the physiology and pathology of organisms including human disease. Here we report a variation in five miRNA levels between gender and tissue. Our results suggest that the transcription levels of these five miRNAs are directly regulated by their RNA.

Understanding the molecular mechanism whereby an environmental chemical causes a disease is important for downstream purposes. In this study, an analytical workflow was designed to combine several publicly available datasets in order to identify CpG sites and genes that mediate the impact of exposure to environmental chemicals on cardiometabolic traits. Organophosphate and prenatal lead exposure were previously reported to change methylation level at the cg23627948 site. I found as cg23627948 site becomes methylated, the expression of GNA12 gene decreases and this leads to higher body fat percentage. Prenatal perfluorooctane sulfonate expo-sure was reported to increase the methylation level at cg21153102 site. The outcome of analyses revealed higher methylation at this site contributes to higher diastolic blood by changing the expression of CHP1 and GCHFR genes. Furthermore, HKR1 mediates the impact of B12 supple-mentation-cg05280698 hypermethylation on higher kidney function; while, CTDNEP1 mediates the impact of air pollution-cg03186999 hypomethylation on higher systolic blood pressure. This study reports CpG sites and genes that mediate the impact of environmental chemicals on cardiometabolic traits and describes a framework to investigate the underlying molecular paths using publicly available data.

The evolutionary cancer cell biology ECCB explains the deep homology existent between the life cycle of cancer and protists. Due to their origin and sensitivity to oxygen, all non-gametogenic germ lines and germ stem cells (GSCs), including human adult stem cells (ASCs) and cancer stem cells (CSCs), suffer severe DNADSB damage when exposed to stress oxygen content above 6.0% O2 (Urgermline hyperoxia) as occurring in tissue and bloodstream. As a result, GSCs/ASCs/ CSCs lose their stemness potential and convert from a functional asymmetric cyclic phenotype (ACD phenotype) to a dysfunctional symmetric phenotype capable of defective symmetric proliferation (DSCD) that is, however, unable to repair genomic degradation. Genome reconstruction occurs through evolutionary mechanisms evolved by the common ancestor of amoebozoans, metazoans, and fungi. They complete the cycle by removing the damage, reconstructing the functional genome, and reinstating the ASC phenotype. The stemness life cycle acts not only during carcinogenesis, tumorigenesis, and recurrence but also in the life cycle of protists.

Histone demethylases, enzymes responsible for removing methyl groups from histone proteins, have emerged as critical players in regulating gene expression and chromatin dynamics, thereby influencing various cellular processes. LSD2 and LSD1 have attracted considerable interest among these demethylases because of their associations with cancer. However, while LSD1 has received significant attention, LSD2 has not been recognized to the same extent. In this study, we conduct a comprehensive comparison between LSD2 and LSD1, with a focus on exploring LSD2's implications. While both share structural similarities, LSD2 possesses unique features. Functionally, LSD2 shows diverse roles, particularly in cancer, with tissue-dependent roles. Additionally, LSD2 extends beyond histone demethylation, impacting DNA methylation and DNA damage repair pathways. This study underscores the distinct roles of LSD2, providing insights into their contributions to cancer and other cellular processes.

Histone deacetylases inhibitors (HDACi) have shown promising therapeutic outcomes in haematological malignancies such as leu-kaemia, multiple myeloma and lymphoma, with dis-appointing results in solid tumours when used as monotherapy. As a result, combination therapies either with radiation or other deoxyribonucleic acid (DNA) damaging agents have been suggested as ideal strategy to improve their efficacy in solid tumours. Numerous in vitro and in vivo studies have demonstrated that HDACi can sensitise malignant cells to both electromagnetic and particle types of radiation by inhibiting DNA damage repair. Although the radiosensitising ability of HDACi has been reported as early as 1990s, the mechanisms of radiosensitisation are yet to be fully understood. This review brings forth the various protocols used to sequence the administration of radiation and HDACi treatments in the different studies. The possible contribution of these various protocols on the ambiguity that surrounds the mechanisms of radiosensitisation is also highlighted.

Diffuse midline gliomas are among the deadliest human cancers and have had little progress in treatment in the last 50 years. Cell cultures of these tumors have been developed recently, but it is unknown the degree to which such cultures retain the characteristics of the source tumors. DNA methylation profiling offers a powerful tool to look at genome-wide epigenetic changes that are biologically meaningful and can help assess the similarity of cultured tumor cells to their in vivo progenitors. Paraffinized diagnostic tissue from three diffuse intrinsic pontine gliomas with H3 K27M mutations was compared with subsequent passages of neurosphere cell cultures from those tumors. Each cell line was passaged 3-4 times and analyzed with DNA methylation arrays and standard algorithms that provided comparison of diagnostic classification and cluster analysis. All samples tested maintained high classifier scores and clustered within the reference group of H3 K27M-mutant diffuse midline gliomas. There was a gain of 1q in all cell lines, with two cell lines initially manifesting the gain of 1q only during culture. In vitro cell cultures of H3 K27M-mutant gliomas maintain high degrees of similarity in DNA methylation profiles to their source tumor, confirming their fidelity even with some chromosomal changes.

Our understanding of the divergence of bacteria, archaea, viruses, and mobile elements from the last universal common ancestor (LUCA) has not improved substantially since the crystallization hypothesis (explaining transition from cellular machinery to cellular life) was proposed by Woese. Here, we propose a hypothesis for the simultaneous emergence of bacteria, archaea, viruses, and mobile elements by sequential and concrete biochemical and cellular pathways. According to the hypothesis, the LUCA was a non-free-living pool of single operon type genomes composed of single-stranded (ss) RNA, double-stranded (ds) RNA, RNA/DNA hybrid, ssDNA, or dsDNA at an ancient submarine alkaline vent. Each dsDNA operon was transcribed by different systems in σ, TFIIB, or TBP genomes. Upon the fusion of multiple dsDNA operons by recombinase, the transcription system (in the σ, TFIIB, or TBP genome) was preferentially selected, leading to the first genetic linkage (described by Morgan). Vertical inheritance eventually led to Bacteria (σ genome) and Archaea (TBP genome). Eigen’s paradox (error catastrophe) can be overcome by the parallel gain of DNA replication and DNA repair mechanisms in both genomes. Enlarged DNA enabled efficient local biochemical reactions. Both genomes independently recruited lipids to facilitate reactions by forming coacervates (liquid droplets) at the chamber of the vent, leading to the lipid divide. Bilayer lipid membrane formation, proto-cell formation with a permeable membrane, proto-cell division, and the evolution of membrane-associated biochemistry are presented in detail. Simultaneous crystallization of systems in non-free-living bacteria and non-free-living archaea triggered the co-crystallization of primitive viruses and mobile elements. An arms race between non-free-living cells and primitive viruses finally led to free-living cells with a cell wall and mature viruses at the original vent. Cells and viruses spread to all vents on the planet, explaining the universality of the genetic code on Earth. The proposed scenario provides a plausible explanation for the origin of diverse taxa from the LUCA vertical and horizontal gene transfer.

Asian giant hornet, Vespa mandarinia, is an invasive species that could potentially destroy local honeybee industry in North America. It has been observed to nest in the coastal regions of British Columbia in Canada and Washington State in USA. What is the source population of the immigrant hornets? The identification of the source population can not only shed light on the route of immigration, but also on the similarity between the native habitat and the potential new habitat in the Pacific Northwest. We analyzed mitochondrial COX1 sequences of specimens sampled from multiple populations in China, South Korea, Japan and the Russian Far East. V. mandarinia exhibits phylogeographic patterns, forming monophyletic clades for 16 specimens from China, six specimens from South Korea and two specimens from Japan. The two mitochondrial COX1 sequences from Nanaimo, British Columbia are identical to the two sequences from Japan. The COX1 sequence from Blaine, Washington State, clustered with those from South Korea, and is identical to one sequence from South Korea (GenBank accession MN716828). Our geophylogeny, which allows visualization of genetic variation over time and space, provides evolutionary insights on the evolution and speciation of three closely related vespine species (V. tropica, V. soror and V. mandarinia). The geophylogeny also highlights a strong insufficiency in sample collection in China. The existing sequenced specimens from mainland China with geographic coordinates were represent only four provinces, missing populations along the coastal regions in East China. One therefore cannot exclude the possibility that the Asian giant hornet found in Canada and USA may also be identical to V. mandarinia in Eastern China. We strongly recommend more extensive DNA barcoding data with geographic information and longer DNA barcode (e.g., the entire mitochondrial genome) so that critical questions on invasive species and environmental conservation can be addressed more accurately than what is possible with existing data.

D-Aspartate and D-Serine serve as primary agonists at glutamate receptors, playing a crucial role in modulating synaptic plasticity and cell migration within the central nervous system. The pre-cise regulation of their levels is essential and intricately linked to the expression of their synthetic and catabolic enzymes, which are, in turn, primarily influenced by epigenetic modifications. In a comprehensive analysis, we examined the methylation profiles of the promoters and transcription start sites of critical genes associated with D-Aspartate and D-Serine metabolism in human post-mortem brain tissues sourced from normal individuals and those diagnosed with schizo-phrenia. Our approach involved a qualitative method capable of identifying specific families of methylated alleles known as epialleles, sharing a common pattern of methylated non-contiguous CpGs, referred to as cores. These methylated traits exhibit stability and consistency within com-plex populations of diverse DNA-methylated molecules. Our findings reveal brain area-specific methylation signatures to the DDO, DAO and DAOA genes serving as distinctive markers that differentiate normal brain areas from those affected by schizophrenia. These methylation patterns align with the reported high D-Aspartate and low D-Serine levels observed in schizophrenic brain areas. The study suggests a potential link between epigenetic modifications and the dysreg-ulation of these neurotransmitters in schizophrenia.

The COVID-19 pandemic caused by the RNA virus SARS-CoV-2, has had a significant impact not only on bodily metabolism but also at the genomic level. This study utilizes a bibliometric approach to map and identify the relationship between COVID-19 and DNA damage, genomic instability, and cell cycle deregulation. The research began with the development of in-depth research questions and conducted two rounds of literature search in the Scopus database with stringent filters to ensure relevance and accuracy. From an initial 1098 documents, 576 relevant documents were selected for further analysis. The collected data was then visualized using bibliometric methods, enabling the identification of important patterns, trends, and relationships among the research variables. Key findings indicate that DNA damage, which can induce apoptosis, is the worst impact of the infection on cells. Term frequency analysis and science mapping revealed a close relationship between "DNA damage" and "apoptosis". Furthermore, discussions on genomic instability highlight telomere regulation issues, while cell cycle deregulation during virus replication in mammalian cells indicates the importance of maintaining cell membrane integrity.This study confirms the importance of bibliometric analysis in detecting and understanding the impact of COVID-19 at the genomic level, proposing several potential research targets and enhancing our understanding of the biological response to viral infection.

Tyrosyl-DNA phosphodiesterase 1 (TDP1) is a human DNA repair protein. It is a member of the phospholipase D family based on the structure similarity. TDP1 is a key enzyme involved in repairing stalled topoisomerase 1 (TOP1)-DNA complexes. Previously we obtained with the CRISPR/Cas9 method HEK293A cells with the homozygous knockout of the Tdp1 gene and used knockout cells as a cellular model for studying the mechanisms of anticancer therapy. In this work we studied for the first time by transcriptomic analysis the effect of Tdp1 gene knockout on genes expression changes in human HEK293A cell line. We received original data that may indicate a role of TDP1 in other process besides repair of DNA-TOP1 complex. The differentially expressed genes (DEGs) analysis revealed that TDP1 could be involved in different processes such as cell adhesion and communication, spermatogenesis, mitochondrial function, neurodegeneration, cytokine response, and MAPK pathway signaling.

The Vitamin D Receptor (VDR) mediates the actions of vitamin D, which has important roles in bone homeostasis, growth/differentiation of cells, immune functions and reduction of inflammation. Emerging evidences suggest that epigenetic modifications of the VDR gene, particularly DNA methylation, may contribute to the onset and progression of many human disorders. This review aims to summarize the available information on the role of VDR methylation signatures in different pathological contexts, including autoimmune diseases, infectious diseases, cancer and others. The reversible nature of DNA methylation could enable the development of therapeutic strategies, offering new avenues for the management of these worldwide diseases.

Sperm DNA integrity and chromatin status serve as pivotal indicators of sperm quality, given their in-tricate link to sperm function, embryo development, and overall fertility. Defects in chromatin com-paction, which are often associated with compromised protamine content, can lead to damaged DNA strands. In this study, chromatin status and possible correlation with DNA damage was assessed in males of three mouse species: Mus musculus, M. spretus and M. spicilegus. We employed various staining methods, including aniline blue, methylene blue (Diff-Quik), toluidine blue and chromomy-cin A3, to assess chromatin compaction in cauda epididymal sperm. Samples were also analyzed by the sperm chromatin structure assay (SCSA) to estimate DNA fragmentation (%tDFI, %HDS). Analyses were carried out on freshly collected sperm and cells incubated for 3 h in a Hepes-buffered modified Tyrode's medium simulating conditions of the female reproductive tract. Notably, the analysis of chromatin status yielded minimal abnormal values across all three species, employing diverse meth-odologies. SCSA analyses revealed distinct variations in %tDFI between species. Following sperm incubation, the percentages of sperm stained with methylene blue exhibited differences among the species and were significantly correlated to the DNA fragmentation index. HDS demonstrated correla-tions with the percentages of sperm stained by aniline blue, methylene blue and chromomycin A3. Overall, chromatin compaction was high across all species, with limited differences among them. The relationship between chromatin status and DNA integrity appeared to be related to levels of sperm competition among species.

The development of biosensors for target detection plays a crucial role in advancing various fields of bioscience. This work presents the development of a genosensor that exploits the colorimetric phenol-sulfuric acid sugar reaction for the detection of DNA, and RNA as specific targets, and DNA intercalator molecules. The biosensor combines simplicity and reliability to create a novel bio-assay for accurate and rapid analysis. A 96-well microplate based on polystyrene polymer was used as the platform for an unmodified capture DNA immobilization via a silanization process and with (3-Aminopropyl) triethoxysilane (APTES). After that, a hybridization step was carried out to catch the target molecule, followed by adding phenol and sulfuric acid to quantify the amount of DNA or RNA sugar backbone. This reaction generates a yellow-orange color on the wells measured at 490 nm that was proportional to the target concentration. Under the optimum conditions, a calibration curve was obtained for each target. The developed biosensor demonstrated high sensitivity, good selectivity, and linear response over a wide concentration range for DNA and RNA targets. Additionally, the biosensor was successfully employed for the detection of DNA intercalator agents that inhibit the hybridization of DNA complementary to the immobilized capture DNA. The developed biosensor offers a potential tool for sensitive and selective detection in various applications, including virus diagnosis, genetic analysis, pathogenic bacteria monitoring, and drug discovery.

Infections with symbiotic single-celled organisms classified as Ciliophora, are commonly reported in various species of invertebrates. Freshwater mussels also play the role of a host for them, being responsible for their biodiversity. Knowledge about these relationships is insufficient. Freshwater mussels are among the most endangered groups of animals. Ciliate endosymbionts in mussels may be responsible for the diseases and increasingly frequent cases of mass mussel deaths and are particularly dangerous for commercially farmed mussels and for restocking of mussels as part of active conservation measures. On the other hand the Ciliophora parasites and commensals living in freshwater mussels is undescribed. Many of these species are likely to become extinct before they are known to science. Recently, two genera of Ciliophora, Conchophthirus sp. and Trichodina sp., have been described in the mantle cavity of U. crassus - an endangered and protected species in the EU. The basis of extensive research on the impact of endosymbiotic ciliates on U. crassus populations is the knowledge of their species composition, proportion of infected individuals, level of infection intensity and distribution in different types of rivers. Such studies were carried out in the mantle cavity of U. crassus from three rivers in three seasons. Cloning, sequencing and functional analysis of the genetic material of Ciliophora was carried out using NGS (Next-Generation Sequencing) analysis based on the hypervariable V4 and V9 regions of the 18SrRNA gene, which enables the identification of taxonomic groups, including genera and species. The most numerous OTUs are common and cosmopolitan species. But some commensals and potentially parasites were found too. Ciliophora associated with mantle cavity form an interesting, hierarchical biocomplex but their interactions with mussels need further studies.

To fill in the blanks for charophytes in the Caucasus to improve our knowledge of species distribution areas in Eurasia, a field survey and study of available specimens from Dagestan (North Caucasus, Russia) was conducted based on morphological observation and molecular genetic analysis. Seventeen new localities for six Chara species and one Tolypella species, with six new species and one new genus for the region studied, and one new species for the Caspian Sea region were found. Some species records changed the outline or filled in the gaps in the species distribution area. The presence of species with a distribution area mainly in the centre of Eurasia (C. globata, C. neglecta) with mainly Mediterranean – Middle East species (C. gymnophylla) is notable for this region, as well as for other studied regions of the Caucasus having a combination of species with different distribution areas. The frequent occurrence of C. gymnophylla points is similar to that in the Mediterranean region and the Middle East. Small brackish waterbodies at the coast of the Caspian Sea, freshwater mountain rivers, small waterbodies associated with them, and water reservoirs are the main habitats of charophytes in the regions. The habitat preference and distribution in the Caucasus allowed suggestion some species for protection. The absence of endemic species among charophytes from Dagestan and Caucasus contrasts with the flora of terrestrial magnoliophytes, which have a large number of endemic species.

Repetitive elements are a major component of DNA sequences due to their ability to propagate through the genome. Characterization of Metazoan repetitive profiles is picking up, however, current pipelines are failing to identify a significant proportion of divergent repeats in non-model organisms. The order Decapoda, for which repeat content analyses are largely lacking, is characterized by extremely variable genome sizes that suggest an important presence of repetitive elements. Here we developed a new standardized pipeline to annotate repetitive elements in non-model organisms, which we applied on 20 Decapoda and 6 other crustacean genomes. With our new tool we identify 10% more repetitive elements than standard pipelines. Repetitive elements were more abundant in Decapoda species than in other crustaceans, with an incredibly large number of highly repeated satDNA families. Moreover, we demonstrated a high correlation between assembly size and transposable elements and a different repeat dynamics between Dendrobranchiata and Reptantia. The patterns of repetitive elements largely reflect the phylogenetic relationships of Decapoda and the distinct evolutionary trajectories within Crustacea. In summary, our results highlight the impact of repetitive elements on genome evolution in Decapoda and the value of our novel annotation pipeline, which will provide a baseline for future comparative analyses.

Background: Changes in body weight are associated with the regulation of DNA methylation (DNAm). In this study, we investigated the associations between maternal gestational weight gain-related DNAm and foetal and neonatal body composition. Methods: Brazilian pregnant women from the Araraquara Cohort Study were followed up during pregnancy, delivery, and after hospital discharge. Women with normal pre-pregnancy BMI were allocated to two groups: adequate gestational weight gain (AGWG, n=45) and excessive gestational weight gain (EGWG, n=30). Foetal and neonatal body composition was evaluated by ultrasound and plethysmography, respectively. DNAm was assessed in maternal blood using Illumina Infinium MethylationEPIC BeadChip arrays. Linear regression models were used to explore the associations between DNAm and foetal and neonatal body composition. Results: Maternal weight, GWG, neonatal weight, and fat mass were higher in the EGWG group. Analysis of DNAm identified 46 differentially methyl-ated positions and 11 differentially methylated regions (DMRs) between the EGWG and AGWG groups. Nine human phenotypes were enriched for these 11 DMRs located in 13 genes (EMILIN1, HOXA5, CPT1B, CLDN9, ZFP57, BRCA1, POU5F1, ANKRD33, HLA-B, RANBP17, ZMYND11, DIP2C, TMEM232), highlighting the terms insulin resistance, and hyperglycaemia. Maternal DNAm was associated with foetal total thigh and arm tissues and subcutaneous thigh and arm fat, as well as with neonatal fat mass percentage and fat mass. Conclusion: The methylation pat-tern in the EGWG group indicated a risk for developing chronic diseases and involvement of ma-ternal DNAm in foetal lean and fat mass and in neonatal fat mass.

The polymerase chain reaction (PCR) is a widely used technique in the biosciences and has become increasingly popular in recent years. One of the key elements of this technique is the use of a DNA polymerase that is heat-stable and retains fidelity during the process. To this end, archaeal Fam-ily-B DNA polymerases are preferred due to their high thermostability and fidelity. In particular, the DNA polymerase from Thermus aquaticus (Taq DNApol) is widely utilized in PCR procedures. In this work, a novel in-silico structure-based methodology was employed to examine the most heat-tolerant DNA polymerase available. In spite of this, Thermococcus kodakarensis and Geobacillus stearothermophilus DNApol are more stable and heat-tolerant DNApols due to their high number of intra-protein interactions. Variations in the content of polar amino acids also played a significant role in the increase in heat stability. A further factor contributing to the stability of proteins is the stabilization of helix in secondary structure through the use of charged amino acids. DNApol from these organisms has been shown to be suitable for use in PCR, as well as in other biological processes able to withstand high temperatures. In this study, it has been demonstrated that im-provements in PCR performance can be easily obtained by blending elements from closely related archaeal polymerases, a strategy that may, in the future, be extended to other archaeal polymer-ases. This approach allowed for a comprehensive analysis of the enzyme's thermal stability and fidelity, leading to an improved understanding of the polymerase's properties and potential ap-plications

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of coronavirus disease 2019 (COVID-19), has caused more than 600 million cases and over 6 million deaths worldwide. Vaccination has been the main strategy used to contain the spread of the virus, and to avoid hospitalizations and deaths. Currently, there are two mRNA-based and one adenovirus vectored vaccines approved and available for use in the U.S. population. The versatility, low cost and rapid-to-manufacture attributes of DNA vaccines are important advantages over other platforms. However, DNA vaccination must meet higher efficiency levels for use in humans. Importantly, in vivo DNA delivery combined with electroporation (EP) has been successfully used in the veterinary field. Here we evaluated the safety, immunogenicity and protective efficacy of a novel linear SARS-CoV-2 DNA vaccine candidate for delivered by intramuscular injection followed by electroporation (Vet-ePorator™) in ferrets. The results demonstrated that the linear SARS-CoV-2 DNA vaccine candidate did not cause unexpected side effects, and was able to elicit neutralizing antibodies and T cell responses using a low dose of the linear DNA construct in prime-boost regimen, and significantly reduced shedding of infectious SARS-CoV-2 through oral and nasal secretions in a ferret model.

leaf beetle (BLB) (Ootheca mutabilis) has emerged as an important bean pest in Uganda, leading to devastating crop losses. There is limited information on the population genetic structure of BLB despite their importance. In this study, novel microsatellite markers and the partial mitochondrial cytochrome oxidase subunit I (mtCOI) gene sequences were used to analyze the spatial population genetic structure, genetic differentiation, gene flow and haplotype diversity of 87 O. mutabilis samples from five populations. We identified 19,356 simple sequence repeats (SSRs) (mono, di, tri, tetra, penta, and hexa-nucleotides) of which 81 di, tri and tetra-nucleotides were selected for primer synthesis. Five highly polymorphic SSR markers (4-21 alleles, heterozygosi-ty 0.59-0.84, polymorphic information content (PIC) 50.13-83.14%) were used for this study. Analyses of the five O. mutabilis populations with these five novel SSRs found 89% of genetic variation occurring within individuals, 9% among individuals and 2% among populations. Genetic differentiation was low but significant for SSR and insignificant for mtCOI partial sequence data while gene flow was high across the populations. There was no evidence of isolation by distance between geographical and genetic distances. Bayesian clustering identified signature of admixture that suggests genetic contributions from two ancestral genetic lineages, and the median-joining haplotype network showed low differentiation of many different haplotypes from the most common haplotype. Low genetic differentiation and high gene flow indicates unrestricted migrations between populations. This information will contribute to the design of BLB control strategies.

Exploring the contribution of proximal family risk factors on SLC6A4 DNA methylation in children with a history of maltreatment

The article is devoted to the new results of the author, which add his previously published ones, of studying hidden rules and symmetries in structures of long single-stranded DNA sequences in eukaryotic and prokaryotic genomes. The author uses the existence of different alphabets of n-plets in DNA: the alphabet of 4 nucleotides, the alphabet of 16 douplets, the alphabet of 64 triplets, etc. Each of such DNA alphabets of n-plets can serve for constructing a text as a chain of these n-plets. Using this possibility, the author represents any long DNA nucleotide sequence as a bunch of many so-called n-texts, each of which is written on the basis of one of these alphabets of n-plets. Each of such n-texts has its individual percents of different n-plets in its genomic DNA. But it turns out that in such multi-alphabetical or multilayer presentation of each of many genomic DNA, analyzed by the author, universal rules of probabilities and symmetry exist in interrelations of its different n-texts regarding their percents of n-plets. In this study, the tensor product of matrices and vectors is used as an effective analytical tool borrowed from the arsenal of quantum mechanics. Some additions to the topic of algebra-holographic principles in genetics are also presented. Taking into account the described genomic rules of probability, the author puts also forward a concept of the important role of stochastic resonances in genetic informatics.

Most Acinetobacter baumannii strains are naturally competent. Although some information is available about factors that enhance or reduce the frequency of transformation of this bacterium, the regulatory elements and mechanisms are barely understood. In this article, we describe studies on the role of H-NS in the regulation of expression of genes related to natural competency and the ability to uptake foreign DNA. The expression levels of the natural transformation-related genes pilA, pilT, pilQ, comEA, comEC, comF, and drpA were significantly increased in a Δhns derivative of Acinetobacter baumannii A118. Complementation of the mutant with a recombinant plasmid harboring hns restored expression levels of six of these genes (pilT remained expressed at high levels) to those of the wild-type strain. The transformation frequency of the A. baumannii A118 Δhns strain was significantly higher than that of the wild-type. Similar, albeit not identical, effects occurred when hns was deleted from the hypervirulent A. baumannii AB5075 strain. Reduction of gene expression in a few cases was not as pronounced as to reach wild-type levels, and expression of comEA was enhanced further. In conclusion, the expression of all seven transformation-related genes was enhanced after deleting hns in A. baumannii A118 and AB5075, and these modifications are accompanied by an increase in the cells’ transformability. The results demonstrate a role of H-NS in A. baumannii’s natural competence.

Tilapia lake virus (TiLV) is the main tilapia-infecting virus worldwide, causing serious economic losses. However, there is no vaccine for this viral disease. Here, TiLV ORF10 (TiLV-ORF10) encoding a protein with abundant epitopes was constructed into the eukaryotic expression vector pcDNA3.1, and used to evaluate the immune protective effects in Nile tilapia (Oreochromis niloticus). RT-PCR and western blot analyses confirmed vaccine plasmid expression in tilapia muscle tissues. Moreover, the transcription levels of immunoglobulin M, toll-like receptor 2, myeloid differentiation factor 88, interleukin 8, tumor necrosis factor alpha, gamma-IFN, and nuclear factor κB immune-related genes were statistically significantly upregulated in the spleen, liver, and kidney of vaccinated tilapias (P < 0.05). TiLV challenge experiments showed that relative percent survival (RPS) was significantly enhanced in fish by this DNA vaccine. Moreover, RPS was enhanced further when using a higher amount of the DNA vaccine (85.72% RPS at a DNA dose of 45 μg pcDNA3.1–ORF10). Vaccination with pcDNA3.1–ORF10 significantly reduced virus replication, as evidenced by the low amount of virus in the spleen, liver, and kidney of vaccinated tilapias after TiLV challenge. Thus, pcDNA3.1–ORF10 could induce protective immunity in tilapia and may be a potential vaccine candidate for controlling diseases caused by TiLV.

The Ramazzottius varieornatus tardigrade is an extremotolerant terrestrial invertebrate belonging to the phylum of Tardigrada. At a length of 0.1-1.0 mm, tardigrades are small animals with an exceptional tolerance to extreme conditions such as high pressure, chemicals and irradia-tion. These properties have been attributed to the recently-discovered Dsup protein. Dsup is a nucleosome-binding protein that prevents DNA damage against X-ray and oxidative stress without impairing cell life, also in Dsup-transfected animal and plant cells. However, the precise “protective” role of this protein is still under study. We performed experiments on human cells and shows that, as compared to control cells, Dsup+ cells are more resistant to UV-C exposure and H2O2. Real-time PCR identified different expression patterns of endogenous genes involved in apoptosis, cell survival and DNA damage repair in Dsup+ cells in response to H2O2 and UV-C. While H2O2 treatment in Dsup+ cells only marginally involved the activation of pathways responsible for DNA repair reinforcing the idea of a direct protective effect of the protein on DNA, in UV-C exposed cells, Dsup efficiently upregulates DNA damage repair genes. In conclusion, our data may help to delineate the different mechanisms by which the Dsup protein operates in response to different insults.

The urgent need for effective, safe and equitably accessible vaccines to tackle the ongoing spread of COVID-19 led researchers to generate vaccine candidates targeting varieties of immunogens of SARS-CoV-2. Because of its crucial role in mediating binding and entry to host cell and its proven safety profile, the subunit 1 (S1) of the spike protein represents an attractive immunogen for vaccine development. Here, we developed and assessed the immunogenicity of a DNA vaccine encoding the SARS-CoV-2 S1. Following in vitro confirmation and characterization, the humoral and cellular immune responses of our vaccine candidate (pVAX-S1) was evaluated in BALB/c mice using two different doses, 25 µg and 50 µg. Our data showed high levels of SARS-CoV-2 specific IgG and neutralizing antibodies in mice immunized with three doses of pVAX-S1. Analysis of the induced IgG subclasses showed a Th1-polarized immune response as demonstrated by the significant elevation of spike-specific IgG2a and IgG2b compared to IgG1. Furthermore, we found that immunization of mice with three doses of 50 µg of pVAX-S1 could elicit significant memory CD4+ and CD8+ T cell responses. Taken together, our data indicates that pVAX-S1 is immunogenic and safe in mice and is worthy of further preclinical and clinical evaluation.