DNA Analysis Research Articles

Antimicrobial resistance and epidemiological patterns of Streptococcus pyogenes in Türkiye.

Drug-resistant Group A beta-hemolytic streptococci remain significant infectious agents globally. This study investigated the major S. pyogenes strains responsible for infections in Türkiye and their susceptibility to beta-lactam and macrolide antibiotics. We determined the minimum inhibitory concentration using the penicillin gradient test and performed emmtyping and DNA fingerprinting via pulsed-field gel electrophoresis (PFGE) to analyze the clonal spread of 92 S. pyogenes strains isolated from two hospitals in Türkiye between 2020 and 2022. Our findings revealed the predominant S. pyogenes strains causing infections in the population and provided insights into the epidemiological relatedness of these drug-resistant strains. This study also evaluated the correlation between emm typing and PFGE in tracking S. pyogenes epidemiology. In this study, the current resistance patterns of S. pyogenes strains in Türkiye identified erythromycin resistance in a few strains, but no resistance to penicillin was detected. This study revealed that emm types 1, 12 and 89 as S. pyogenes strain genotypes were responsible for epidemic infections in Türkiye. PFGE genotyping and emm typing were found to provide better phylogenetic classification in the investigation of S. pyogenes epidemiology.

Comprehensive landscape of FGFR variations and prognosis revelance in colorectal cancer from circulating tumor DNA and tissue gene analyses in 2083 patients.

278 Background: FGFR genomic alterations have been identified, and several FGFR inhibitors are approved for various malignancies, including bladder and cholangiocarcinoma. However, limited studies have shown FGFR alterations and their prognostic impact in colorectal cancer (CRC). Methods: This retrospective analysis involved tissue and blood samples from a single-institute cohort (The Sixth Affiliated Hospital of Sun Yat-sen University) of stage I-IV CRC patients from 2014 to 2024. FGFR genomic alterations were analyzed using tissue NGS and plasma ctDNA. Disease-free survival (DFS) for stages I-III and progression-free survival (PFS) for stage IV were compared between FGFR-altered and negative CRC patients, matched in a 1:2 ratio. Validation of genomic alterations and their prognostic impact was conducted using cBioPortal and TCGA data. Results: FGFR alterations were found in 7.89% of 608 tissue samples and 10.95% of 2083 ctDNA samples. The most frequent alteration was FGFR1 amplification in all cohorts, along with FGFR2 p.R165W and FGFR3 p.A634T mutations in ctDNA. FGFR alterations correlated with younger age, left-sided colon tumors, adenocarcinomas, poor differentiation, advanced TNM staging, and multi-organ metastasis. Prognostic analysis showed a higher proportion of FGFR alterations in stage IV patients (66.23% vs. 59.14%), lower rates of no evidence of disease (NED) (6.62% vs. 17.68%), and significantly shorter PFS (median 17 vs. 20 months, HR 2.184, p=0.007). In stages I-III, FGFR-altered patients also had a lower NED rate (79.22% vs. 82.19%) and significantly shorter DFS (median 13 vs. 29 months, HR 2.833, p=0.002). Among frequent alterations, FGFR1 amplification was significantly associated with poor prognosis (PFS 17 vs. 20 months, HR 2.221, p=0.014; DFS 13 vs. 29 months, HR 2.964, p=0.002), while others showed no significant correlation. In stage IV patients receiving bevacizumab, FGFR1 alteration was linked to significantly shorter PFS (p=0.0021). KEGG analysis indicated FGFR1 was associated with angiogenesis pathways. FGFR alteration profiles and prognostic outcomes were consistent across tissue samples, ctDNA analyses, and databases. Conclusions: Our study outlines the genomic landscape and prognostic significance of FGFR in CRC, highlighting FGFR1 amplification as the dominant alteration. FGFR alterations were associated with poorer outcomes, as reflected in lower NED rates and shorter DFS and PFS. FGFR1 amplification was notably linked to reduced DFS and PFS, and it correlated with shorter PFS in patients receiving bevacizumab, indicating a potential link to resistance against bevacizumab. ctDNA n=2083 Tissue genen=608 cBioPortal Databasen=6998 FGFR Total 228/10.95% 48/7.89% 528/8.0% FGFR1 76/3.65% 29/4.77% 264/4.0% FGFR2 78/3.74% 10/1.64% 125/1.9% FGFR3 104/4.99% 10/1.64% 139/2.1%

Phylogenetic and structural characterization of Kentomonas inusitatus n. sp.: Unique insect trypanosomatid of the Strigomonadinae subfamily naturally lacking bacterial endosymbiont.

All insect trypanosomatids of the subfamily Strigomonadinae harbor a proteobacterial symbiont in their cytoplasm and unique ultrastructural cell organization. Here, we report an unexpected finding within the Strigomonadinae subfamily: the identification of a new species lacking bacterial symbiont, represented by two isolates obtained from Calliphoridae flies in Brazil and Uganda. This species is hereby designated as Kentomonas inusitatus n. sp. Molecular investigations targeting symbiont DNA, cell proliferation, and ultrastructural analyses agreed with the absence of bacterial symbionts in cultured flagellates. PCR-screening specifically targeting symbiont DNA corroborated the absence of symbionts in K. inusitatus present in the intestine of the respective host flies. K. inusitatus exhibited forms varying in size and shape. While displaying overall ultrastructural features of the Strigomonadinae, the novel species showed mitochondrial branches juxtaposed to the plasma membrane in locations both without and notable, with subpellicular microtubules. The discovery of the first Strigomonadinae species naturally lacking a symbiont and closely related to K. sorsogonicus, suggests a unique evolutionary history for the genus Kentomonas. Our findings provide novel insights into the complex relationships between trypanosomatids and their symbionts.

Plasma shallow whole-genome sequencing profiling in patients with RAS WT metastatic colorectal cancer (mCRC) undergoing first-line anti-EGFR therapy.

214 Background: Acquired resistance (AR) to first-line (1L) chemotherapy (CT) + anti-EGFR inhibitors is a major challenge in the management of RAS WT mCRC patients (pts). While AR in later lines has been explained with point mutations in the EGFR pathway, its real mechanisms in 1L remain poorly understood. Tumor molecular biology can be dynamically studied through circulating tumor DNA (ctDNA) analysis, a real-time, non-invasive approach to detect AR. In this context, we conducted the multicenter prospective PLATFORM-B study, which enrolled 100 RAS WT mCRC pts receiving 1L CT + cetuximab across 15 Spanish Medical Oncology Departments, alongside 30 RAS MUT pts (control) treated with 1L CT + bevacizumab. Peripheral blood samples have been collected at baseline (BL), week 8, and disease progression (PD). Next-generation sequencing analysis (Ion S5 system) of ctDNA at PD revealed AR mutations in only 10% of pts, suggesting that alterations in the EGFR pathway may only partially explain AR in 1L. Therefore, we conducted shallow whole-genome sequencing (shWGS) to further characterize genomic profiles possibly related with novel mechanisms of AR. Methods: Baseline and PD samples with sufficient remaining plasma were considered for this analysis. shWGS was performed with a coverage ranging 0.5X to 2.0X and analyzed using ichorCNA pipeline to evaluate various genomic characteristics such as copy number alterations (CNA), tumor fraction (TF), subclonal fraction (SF), and ploidy. Molecular results were correlated to clinical-pathological features and outcomes. Results: Overall, 35 mCRC pts (27 RAS WT and 8 RAS MUT ) were included in the study. At BL ( N = 26), median TF (mTF) was 0.19, median ploidy 2, median SC fraction 0.5, median SC genome fraction 0.21, and median subclonal CNA fraction 0.41. At PD ( N = 33), mTF was 0.08, median ploidy 2, median SC fraction 0.5, median SC genome fraction 0.17, and median subclonal CNA fraction 0.38. Of all the features analyzed, only the TF showed potential prognostic value. At BL, pts with TF levels above the median had poorer overall survival (OS) compared to those with lower levels, approaching statistical significance ( P = .055). At PD, higher TF levels were statistically significantly associated with worse OS ( P = .02). No meaningful differences between the RAS WT and RAS MUT groups were observed. Specific variations in CNA, TF, SF, or ploidy under treatment are being correlated to clinical data to seek for predictive value. Conclusions: This preliminary analysis of the PLATFORM-B using shWGS showed the prognostic value of TF. Further analyses are undergoing to better understand the molecular landscape of acquired resistance to 1L anti-EGFR-based therapy in RAS WT mCRC pts.

Utility of circulating tumor DNA in patients with unresectable pancreatic cancer using a patient-specific panel: ARTEMIS-PC study.

684 Background: Circulating tumor DNA (ctDNA) analysis has proven to be a highly accurate method for assessing treatment efficacy and detecting molecular residual disease (MRD) in cancer patients, outperforming tumor markers and imaging modalities. However, the ctDNA detection rate is reported to be low, and its utility remains unclear in pancreatic cancer (PC). We conducted the prospective “ARTEMIS-PC” study to evaluate the utility of a personalized, tumor-informed MRD assay in patients with unresectable PC. Methods: Eligible patients had histopathologically confirmed unresectable PC, were previously untreated, and had disease measurable per Response Evaluation Criteria in Solid Tumors v1.1. A patient-specific tumor-informed assay (Invitae Personalized Cancer Monitoring) was used for the detection of ctDNA. Blood samples were collected at structured pre- and post-systemic therapy timepoints. Results: A total of 99 patients were eligible for the ARTEMIS-PC study, and personalized panels were successfully created for 92 patients. The median age of patients was 70 years (range 45-81 years), with 43/92 being female. Thirty patients were stage III, and 62 stage IV. Median size of the primary tumor was 38 mm (range 13-100 mm). MRD positivity at enrollment was 88.0%, with 73.3% in stage III patients, and 95.2% in stage Ⅳ (p=0.007). During follow-up, ctDNA clearance was achieved in 33 patients (40.7%). ctDNA clearance was associated with a significantly higher objective response rate (61.5% vs. 17.6%, p=0.001) and disease control rate (100% vs. 64.7%, p=0.002) compared to those who did not achieve clearance. Patients who achieved ctDNA clearance had significantly longer progression-free survival (PFS) than those who did not (9.0 vs. 3.5 months, hazard ratio 0.2, p<0.001). The predictive performance of select biomarkers (variant allele frequency [VAF], CEA, CA19-9) was evaluated using area under the curve (AUC) for treatment response and disease control. For disease control, VAF showed superior predictive performance with AUCs of 0.84 at enrollment and 0.97 at both Week 4 and Week 8, compared to CEA (AUCs: 0.65, 0.73, 0.60) and CA19-9 (AUCs: 0.51, 0.57, 0.56). The predictive performance for treatment response was lower than AUC 0.75 for any of the biomarkers. Among patients who were MRD negative before treatment or became negative during follow-up (n=44), 45.5% (n=20) subsequently turned MRD positive. Of these, disease progression was confirmed by CT in 14 patients. The median time from MRD turning positive to CT-confirmed progressive disease was 88.5 days (range: -28 to 385 days). Conclusions: ctDNA clearance is a useful predictor of improved objective response, disease control, and PFS, with VAF emerging as the more accurate biomarker for disease control, underscoring its potential utility in treatment monitoring in patients with unresectable PC. Clinical trial information: UMIN000043561 .

Assessment of Genetic Diversity and Construction of Core Germplasm in Populations of Acorus tatarinowii Based on SNP Markers

Acorus tatarinowii is a natural medicinal plant integral to traditional aromatic therapies. It is commonly employed in the treatment of depression, epilepsy, and Alzheimer's disease due to its significant medicinal and aromatic properties. However, the genetic diversity of wild A. tatarinowii resources has declined due to over-exploitation and habitat destruction. This study aims to assess the genetic diversity of the natural populations of A. tatarinowii, establish a core germplasm bank, explore its genetic richness and uniqueness, prevent genetic erosion, and identify beneficial genes. In this study, for the first time, 429A. tatarinowii samples from 40 populations were analyzed for genetic diversity and population structure using Hyper-Seq technology. A total of 4,772,850 high-quality Single Nucleotide Polymorphisms (SNPs) and 1,563,000 Insertions and Deletions (InDels) variant loci were identified, with C/T as the predominant variant type and a Ts/Tv ratio of 1.079. Annotation of these loci indicated that the majority of variants occurring in intergenic regions, accounting for 50.59% of the total. Moreover, the heterozygosity, nucleotide diversity, and FST of A. tatarinowii suggested low genetic diversity within this species within the populations. The analysis of molecular variance (AMOVA) revealed that the population variation of A. tatarinowii is mainly caused by the variation between populations (72.06%), while the variation within populations only contributes a small part (27.94%) Through NJ tree, PCA, and ADMIXTURE analyses, the 429A. tatarinowii samples were classified into five subgroups, with some genetic exchange observed. A total of 7,163 high-quality polymorphic SNPs were identified, and a core germplasm consisting of 85 samples was established, achieving genotype retention rates similar to those of the original germplasm. This indicates that a smaller number of germplasm resources can effectively represent the majority of the genetic diversity. Additionally, PCA analysis further confirmed the representativeness and validity of the constructed core germplasm resources. Furthermore, the DNA fingerprints of the 429 accessions were established using the most effective combinations of 26 SNP markers, which served as specific markers to effectively distinguish all samples. In conclusion, these findings offer valuable insights into the genetic structure of A. tatarinowii, facilitating the identification of high-quality genes and providing a scientific foundation for the development of breeding programs and conservation strategies for A. tatarinowii.

An open-label, phase IB/II study of abemaciclib with paclitaxel for tumors with CDK4/6 pathway genomic alterations.

Disruption of cyclin D-dependent kinases (CDKs), particularly CDK4/6, drives cancer cell proliferation via abnormal protein phosphorylation. This open-label, single-arm, phase Ib/II trial evaluated the efficacy of the CDK4/6 inhibitor, abemaciclib, combined with paclitaxel against CDK4/6-activated tumors. Patients with locally advanced or metastatic solid tumors with CDK4/6 pathway aberrations were included. Based on phase Ib, the recommended phase II doses were determined as abemaciclib 100 mg twice daily and paclitaxel 70 mg/m2 on days 1, 8, and 15, over 4-week-long cycles. The primary endpoint for phase II was the overall response rate (ORR). The secondary endpoints included the clinical benefit rate (CBR), progression-free survival (PFS), overall survival (OS), and safety. Tissue-based next-generation sequencing and exploratory circulating tumor DNA analyses were carried out. Between February 2021 and April 2022, 30 patients received abemaciclib/paclitaxel (median follow-up: 15.7 months), and 27 were included in the efficacy analysis. CDK4/6 amplification (50%) and CCND1/3 amplification (20%) were common activating mutations. The ORR was 7.4%, with two partial responses, and the CBR was 66.7% (18/27 patients). The median OS and PFS were 9.9 months [95% confidence interval (CI) 5.7-14.0 months] and 3.5 months (95% CI 2.6-4.3 months), respectively. Grade 3 adverse events (50%, 21 events) were mainly hematologic. Genetic analysis revealed a 'poor genetic status' subgroup characterized by mutations in key signaling pathways (RAS, Wnt, PI3K, and NOTCH) and/or CCNE amplification, correlating with poorer PFS. Abemaciclib and paclitaxel showed moderate clinical benefits for CDK4/6-activated tumors. We identified a poor genetic group characterized by bypass signaling pathway activation and/or CCNE amplification, which negatively affected treatment response and survival. Future studies with homogeneous patient groups are required to validate these findings.

Natural history study of patients with pancreatic acinar cell carcinoma: A 3-year prospective longitudinal analysis.

764 Background: Pancreatic acinar cell carcinoma (PACC) is a rare exocrine tumor of the pancreas that comprises 0.2-2% of all pancreatic malignancies. There are no prospective randomized studies in this disease and most data about PACC comes from retrospective database analyses and case reports. Methods: The Cancer Moonshot-funded My Pediatric and Adult Rare Tumor (MyPART) network’s Rare Solid Tumor Natural History study (NCT03739827) is a prospective, single-institution observational study that enrolls patients of all ages with rare solid tumors (<15 cases per 100,000 people per year), including PACC. Patients can participate remotely (field cohort) or visit the NIH for annual evaluations (in-person cohort). All participants complete medical and family histories, patient reported outcomes (PROs) measures, and provide saliva for DNA analysis. Relevant data such as clinical and family histories, surgical and molecular pathology, and imaging results are extracted from patient medical records and entered in the central study database. Archival tumor samples (when available) are analyzed using a 500+ gene panel (TruSight500, Illumina) and discussed in a molecular tumor board. Patients participating in the in-person cohort undergo additional evaluations including clinical examination, imaging studies, genetic counseling, and blood sample collection for clinical and research purposes (standard clinical labs, germline DNA/RNA, immune phenotypes, cytokines) as indicated. Results: From 09/2021 until 09/2024, 17 PACC patients (n=13 for the field cohort, n=4 for the in-person cohort) were accrued. The mean age of diagnosis is 64.7 (48-78) years, and the male to female ratio is 14:3. At the time of diagnosis, 6 patients had Stage I or II, 1 had Stage III, and 10 had Stage IV disease. The median follow-up time since accrual is 7.6 months, and 4 patients have died. No patients have been lost to follow-up. Of the 6 patients diagnosed with early-stage disease, 5 had surgery and all received adjuvant chemotherapy, while 1 patient is receiving neoadjuvant chemotherapy. Among the 9 patients diagnosed with advanced disease, 8 received platinum-based regimens as a first-line therapy. Tumor molecular profiling was available for 16 patients. The most common tumor mutations observed were in CDKN2A (29.4%), BRCA2 (29.4%), and SMAD4 (23.5%). Six patients received targeted therapies matched to their tumor molecular profile. Conclusions: The demographic profile of our cohort aligns with previous studies, with a mean age of diagnosis in the early-to-mid 60s and a high male to female ratio. Our cohort highlights a shift in first-line treatment to platinum-based therapies over the past decade. Many PACC tumors have targetable mutations found in molecular profiling, and the efficacy of matched therapy warrants further exploration. Clinical trial information: NCT03739827 .

Circulating tumor DNA analysis for prediction of prognosis and molecular insights in patients with resectable gastric cancer: results from a prospective study.

This study aimed to evaluate the prognostic value of plasma circulating tumor DNA (ctDNA) level in patients with resectable gastric cancer (GC). A total of 59 patients were prospectively enrolled, with their ctDNA detected and paired tumor tissue collectedat various peri-operative time points. Patients with higher 1-month post-operative ctDNA levels demonstrated shorter overall survival status (hazard ratio [HR]=5.30, p=0.0022) and a higher risk of recurrence (HR=3.85, p=0.011).The model combining ctDNA with conventional serum tumor markers for GC, including carcinoembryonic antigen, carbohydrate antigen 19-9, and CA72-4, shows high predictive effectiveness for GC prognosis with an area under the curve of 0.940 (p=0.002), which is higher than net ctDNA and other models without ctDNA. Patients with lower ctDNA levels were more likely to have positive stromal programmed cell death ligand 1 expression (p=0.046). Additionally, DCAF4L2 mutation was identified as the crucial gene mutation in ctDNA suggesting poor prognosis of patients with GC. Overall, this study highlights that post-operative ctDNA can serve as an effective biomarker for prognostic prediction and recurrence surveillance in resectable GC.

Fetal hematological phenotypes of various hemoglobinopathies and demonstration of embryonic hemoglobins on capillary electrophoresis: a large cohort data from prenatal screening program.

This study reported a large cohort of fetal blood analysis of various hemoglobinopathies. A total of 371 fetal blood specimens were recruited. Complete blood count and hemoglobin (Hb) analysis using capillary electrophoresis were performed. Genotypes were defined by DNA analysis. Among 371 fetuses, 36 were non-thalassemic and 29 thalassemia genotypes were identified in the remaining 335 fetuses. Fetuses with β-thalassemia and Hb E traits, homozygous Hb E, and Hb E-β0-thalassemia had similar hematological parameters as those of non-thalassemic. However, the levels of Hb A in β-thalassemia and Hb E traits were approximately half of that observed in the non-thalassemic fetuses. As for Hb E, fetuses with a single copy of the βE-globin gene in the Hb E trait and Hb E-β0-thalassemia had lower Hb E levels as compared to that of the homozygous Hb E. For α-thalassemia, fetuses with one or two α-globin gene defects had small changes in hematological parameters, but variable Hb Bart's levels were observed. Fetuses with Hb H and Hb H-CS diseases had moderate anemia, whereas those with homozygous Hb CS and Hb Bart's hydrops fetalis had severe anemia. Identification of the fetuses with Hb Bart's hydrops fetalis with various genetic interactions allows the exact re-location of electrophoretic mobilities of various embryonic Hbs. This study confirmed the genetic heterogeneity of hemoglobinopathies among the fetuses and fetal blood analysis are useful for presumptive diagnosis of hemoglobinopathies. The results should facilitate a prevention and control program of hemoglobinopathies in the region.

Morphological and DNA analysis of pollen grains on butterfly individuals reveal their flower visitation history.

Many butterfly species are conspicuous flower visitors. However, understanding their flower visitation patterns in natural habitats remains challenging due to the difficulty of tracking individual butterflies. Therefore, we aimed at establishing a protocol to solve the problem using the Common five-ring butterfly, Ypthima argus (Nymphalidae: Satyrinae). Focusing on the pollen grains attached the butterfly's body surface, we examined validities of two pollen analyses based on pollen morphology and DNA markers (ITS1 and ITS2), in addition to the classical route census method. We captured thirty-nine butterflies from mid-April to early July and collected pollen grains from each individual. Morphological and DNA analyses of collected pollens identified eighteen and thirty-four taxa of insect pollinated plants respectively, including woody plants such as Castanopsis. The DNA analysis detected as many as thirteen plant taxa from a single butterfly, indicating its high sensitivity for detecting flower visitation. We detected more plant taxa in May when many individuals were flying. This is assumingly related to the post emergence days of the butterflies with more foraging experience. We also found that fluctuations of pollen grain numbers of Leucanthemum vulgare and Erigeron philadelphicus on individual butterflies depend on their flowering periods overlapping partly. Consequently, we conclude that pollen morphology and DNA barcoding analysis, and field observations are mutually complementary techniques, providing an integrated pollen analysis method to study the pollination ecology of butterflies.

Artificial Intelligence and Identification of the Deceased: a Narrative Review With Implications in Forensic Science.

Identification of the dead is of utmost importance in mass disasters, war crimes, and forensic examinations. The biological profile, established by a forensic anthropologist is one the necessary steps involved in the identification of the dead. Several parameters can be estimated such as sex, age, stature, biogeographical affinity, and DNA profile of the unknown person. It is crucial to estimate these parameters of identification which may narrow down the investigation process. On the other hand, Artificial Intelligence (AI) in the modern world is showing magical uses in different fields. This communication aims to highlight the uses of AI tools for predicting parameters such as sex, age, stature, biogeographical affinity, and DNA profile of unknown persons with more accuracy and in less time. A literature search was conducted using databases PubMed, Scopus, Web of Science, and ScienceDirect for analyzing the use of artificial intelligence, machine learning, and deep learning algorithms for establishing the biological profile in disaster victim identification (DVI) and forensic casework. Moreover, this research foresees a paradigm shift in investigative techniques as technology advances, highlighting the convergence of AI and anthropological ideas for an improved understanding of the biological profiles of unknown deceased individuals.

Beyond genomics: using RNA-seq from dried blood spots to unlock the clinical relevance of splicing variation in a diagnostic setting.

We aimed to assess the impact of splicing variants reported in our laboratory to gain insight into their clinical relevance. A total of 108 consecutive individuals, for whom 113 splicing variants had been reported, were selected for RNA-sequencing (RNA-seq), considering the gene expression in blood. A protocol was developed to perform RNA extraction and sequencing using the same sample (dried blood spots, DBS) provided for the DNA analysis, including library preparation and bioinformatic pipeline analysis. Splicing in genes of interest was inspected using IGV, with at least three unaffected individuals as controls. From the 113 variants, we confirmed an abnormal splicing in 64 variants (57%). In 15 variants (13%), we did not observe a splicing alteration. In the remaining 34 variants, no decision could be made on the splicing effect due to insufficient sample quality (21%) or a low number of reads (9%). The most common event leading to aberrant splicing was exon skipping, identified in 31 variants (48%). Other events included cryptic donor/acceptor site usage (n = 25; 39%), intron retention (n = 4; 6%), and other complex events (n = 4; 6%). In three patients, pathologically reduced enzymatic activity (measured using the same DBS) served as additional confirmation of the abnormal splicing caused by variants in HEXA, GAA, and GLA. We implemented an RNA-seq pipeline using the same sample provided for genomic testing. This multiomic approach, as implemented in our routine diagnostic processes, clarifies the clinical relevance of most of the analyzed variants and delivers more comprehensive genetic testing.

Enhancing trace DNA profile recovery in forensic casework using the amplicon RX post-PCR clean-up kit

This study evaluated the effectiveness of the amplicon RX post-PCR clean-up kit in enhancing trace DNA profile recovery from forensic casework samples amplified using the GlobalFiler PCR amplification kit. The impact of post-PCR clean-up on allele recovery and signal intensity was assessed in both trace casework samples and control samples across a range of DNA concentrations. The results showed that the amplicon RX method significantly improved allele recovery compared to the 29-cycle protocol (p = 8.30 × 10−12) and achieved slightly better results than the 30-cycle protocol (p = 0.019). Additionally, the Amplicon RX method demonstrated a significant increase in signal intensity (p = 2.70 × 10−4), reflecting improved sensitivity in detecting trace DNA profiles compared to the 30-cycle protocol. In the evaluation of control samples, the amplicon RX method consistently outperformed both the 29- and 30-cycle protocols, especially at lower DNA concentrations (D3: 0.001 ng/µL). While the performance of all methods declined at the lowest concentration (D4: 0.0001 ng/µL), the Amplicon RX method still demonstrated superior allele recovery (p = 0.014 compared to 29 cycles; p = 0.011 compared to 30 cycles). Therefore, the Amplicon RX method should be widely adopted in forensic laboratories to enhance the analysis of extremely low-template and compromised samples. These findings highlight the potential of the amplicon RX post-PCR clean-up kit to improve trace DNA analysis in forensic casework. Further research is recommended to validate these results and explore its broader application in forensic DNA analysis, particularly in complex DNA mixtures and extremely low-template samples.

Increased interferon I signaling, DNA damage response and evidence of T-cell exhaustion in a patient with combined interferonopathy (Aicardi-Goutières Syndrome, AGS) and cohesinopathy (Cornelia de Lange Syndrome, CdLS)

BackgroundType I interferonopathies including Aicardi-Goutiéres Syndrome (AGS) represent a heterogeneous group of clinical phenotypes. Herein, we present a Case with combined AGS and Cornelia de Lange Syndrome (CdLS)—a cohesinopathy—with comprehensive analysis of the immune and genomic abnormalities.Case and methodsA 20-year old man presented with chilblain lesions and resorption of distal phalanges of fingers and toes, somatic and psychomotor retardation, microcephaly, synophrys, hearing losing and other aberrancies consistent with the phenotype of CdLS. We used whole exome sequencing to genetically map the associated mutations and performed transcriptome profiling and enrichment analysis in CD14+ monocytes of the patient and immune phenotyping by mass cytometry (CyToF), comparing to healthy individuals and lupus patients as disease controls. DNA damage response was assayed by confocal microscopy in the peripheral blood of this patient.ResultsNext generation exome sequencing confirmed a homozygous SAMHD1 gene mutation and a hemizygous non-synonymous mutation on SMC1A gene, responsible for the AGS and CdLS, respectively. Transcriptome profiling of CD14+ monocytes of the patient showed enrichment of type I IFN signaling and enhanced DNA damage response pathway. Broad immune phenotype of the peripheral blood of the patient revealed absence of activated T cell populations, increased frequency of NK cells and plasmablasts and enhanced granulocytic lineage. Further analysis suggested activation of the ATM branch of DNA damage response and increased apoptosis in the periphery of the patient.ConclusionsA rare case of a patient bearing two genetic lesions (responsible for AGS/CdLS syndromes) exhibits distinctive features of genomic damage and interferon responses. Immune phenotype revealed granulocytic skewing and absence of activated T cells compatible with chronic antigenic stimulation and/or homing of these cells at sites of inflammation.

Genomic analysis of three medieval parchments from German monasteries

In the last two decades there has been growing interest in the analysis of ancient DNA obtained from the parchment used in historic documents. The genetic insight that this data provides makes collections of historic documents an invaluable source for studying the development and spread of historical livestock populations. Additionally, the biological data may provide new information for the historical analysis that could be used to determine the provenance as well as the authenticity of these documents. In this study, we extracted DNA from three medieval parchments that were written in German monasteries in the twelfth century. The source animal of the parchments could be identified as cattle and we compared their genome sequences with those of modern populations that are part of the 1000 Bull Genomes Project. The three animals were found to carry mtDNA haplogroup T3 and show a closer genetic relationship to other historic animals than to modern breeds. We further identified 39 haplotypes and 132 SNPs variants, which are rare (<0.1) or even non-existent in modern breeds. Finally, the genetic distances between the parchment samples show a putative association with the dates when the documents were written, indicating the usefulness of genetic analysis for provenance research.

Autosomal and Y-STR genetic database from a population of the Spanish Civil War (1936-1939) and the postwar period.

Under the initiative of the "Direcció General de Memòria democràtica-Departament de Justícia" (Generalitat of Catalonia, Spain), a multi-disciplinar project was funded to identify the remains of people disappeared in Catalonia during and after the Spanish Civil War (1936-1939). Samples were officially sent by Autonomous Government of Catalonia to the Laboratory of Forensic and Population Genetics at Complutense University, Madrid, Spain, to be genotyped. Our study presents a database of 343 victims genotyped for STRs comprised in GlobalFiler™ PCR Amplification Kit (Thermofisher Scientific) and a subset of 292 typed with Y-STRs from Yfiler™ Plus PCR Amplification Kit (Thermofisher Scientific). Complete profiles amounted to 116 (33.81%) and 89 (30.48%) for autosomal and Y-STRs respectively. Allelic/haplotype frequencies, forensic parameters and HW equilibrium were calculated with STRAF software. Drop-out frequencies were also calculated for each locus. All the markers were in HW equilibrium (p > 0.05). Allelic drop-out frequencies were larger in the case of CSF1PO (1.5340E-01) and TPOX (1.2640E-01), probably due to extremely low DNA preservation combined with a less efficient PCR for these loci, among other causes. The comparison of our autosomal database with a set of modern European populations from STRidER database (ENFSI) reveals that the present study sample clusters with other West European samples. This is also the case for Y-STR when our study is compared with European and North African samples. Our results seem to suggest that modern Spanish forensic databases can be used to identify deceased persons during the Spanish Civil War and later, from their living descendants.

Development and validation of rapid eDNA detection method for yellow mud turtle, Kinosternon flavescens: a field study in South Texas, USA

The conservation of freshwater turtle species depends on precise and effective monitoring techniques. Environmental DNA (eDNA) analysis is a potential method for identifying cryptic and elusive turtle species in aquatic ecosystems. eDNA analysis can help to identify key regions for conservation efforts and monitor changes in population levels over time. This study aims to evaluate the effectiveness of a rapid eDNA detection method for the yellow mud turtle (Kinosternon flavescens, an indicator species that is endangered in some states in the USA), which inhabits local oxbow lakes (e.g., resacas) in Cameron County, South Texas. A species-specific nested PCR assay was designed to enhance the detection of yellow mud turtle species. Water samples were collected from five locations within Cameron County for the detection of yellow mud turtle eDNA. Our results revealed the presence of yellow mud turtles in two out of the five surveyed locations. Our study shows great potential for eDNA monitoring for yellow mud turtle species. This study also provides insights on using eDNA monitoring to protect yellow mud turtle species and recommendations for future research and conservation initiatives.

Population genomics and genetic diversity of the invasive chrysanthemum lace bug (Corythucha marmorata) across its invasive range in Japan

The escalating global movement of alien species, facilitated by increased trade and travel, poses a pressing need to comprehend their invasive potential and the consequent ecological and economic ramifications. Despite a growing body of evidence on rapid evolutionary shifts in invasive species, comprehensive insights into the genetic variability underlying these adaptations are constrained by limited genomic resources. Understanding the role of genetic variation in the success or failure of biological invaders is thus crucial. This study focuses on the chrysanthemum lace bug, Corythucha marmorata, as a model system to investigate the interplay of genetic variation and invasion dynamics. Our analysis reveals moderate genetic structure among countries, with significant genetic differentiation observed within populations. Mitochondrial COI DNA and haplotype network analysis revealed shared haplotypes between Japan and North America, indicating recent events of introduction, while exclusive Japanese haplotypes and significant FST and GST values suggest local divergence. Phylogenetic and STRUCTURE analyses show genetic clusters unique to Japan, with populations like SAG and CER displaying higher divergence. Bottlenecks followed by divergence, as indicated by the DIYABC-RF analysis, point to a complex evolutionary history involving multiple introductions and subsequent local divergence in Japan.

"DNA Doesn't Lie:" Genetic Essentialism and Determinism in Law & Order: Special Victims Unit.

Law and Order: Special Victims Unit (SVU) (1999-present) is a popular primetime drama that spotlights the use of genetic information to solve crimes. Despite the show's heavy reliance on the forensic use of DNA evidence, the role of genetics in defining family and identity arises in complex ways. Many episodes wrestle with social, ethical, and legal questions that reflect assumptions about genetic essentialism and genetic determinism, but counterarguments about the importance of non-biological relationships, social factors, and legal entitlements are given equal or greater weight. For this study, we identified and viewed 38 episodes from SVU's first twenty seasons centered on genetic themes in non-forensic contexts. Two recurring themes emerged: (1) that the role DNA plays is only one factor in a complex web of biological and social considerations that shape our understanding of kinship; and (2) that genetic predispositions to behavioral traits such as mental illness or violence should not be seen as obscuring the responsibility of personal choice. By treating genetics as a complex source of information requiring social context to be understood, SVU allows audiences to play an active role in interpreting its meaning.