Search Results (227)

Background: Colorectal cancer (CRC), a leading cause of cancer-related mortality worldwide, necessitates improved non-invasive diagnostic and monitoring tools. Circulating tumor cells (CTCs), as intact cellular biomarkers in liquid biopsies, offer valuable morphological and genetic information and hold significant clinical potential for early detection, prognosis, therapy monitoring, and drug development. Bio-ferrography is a non-invasive immunomagnetic separation technique that isolates magnetically labeled entities from fluid samples onto a glass substrate via a focused external magnetic field. Methods: This study employs, for the first time, bio-ferrography for isolation, counting, and microscopic characterization of circulating tumor cells (CTCs) expressing the human epidermal growth factor receptor (EGFR) from blood biopsies taken from patients in the hospital. Magnetic beads conjugated with anti-EGFR antibodies were used to selectively capture CTCs from peripheral blood samples of patients with metastatic CRC and other epithelial malignancies. The method enabled both enumeration and microscopic characterization of isolated cells. Results: Preliminary clinical results demonstrate that bio-ferrography achieves a sensitivity of 90% in stage-IV patients and exhibits higher true positive detection rates compared to conventional tumor biomarkers, including carbohydrate antigen 19-9 (CA 19-9) and carcinoembryonic antigen (CEA). Conclusions: These findings highlight the potential of bio-ferrography as a robust platform for CTC isolation and analysis. Full article

In this study, an exploratory transcriptomic investigation was conducted to evaluate the feasibility of using feather transcriptomics to detect sex differences and gene responses to physiological changes in chickens. Feathers represent a promising non-invasive biological source of RNA, as the feather pulp of growing feathers contains living cells capable of active transcription. Growing feathers were collected from 150-day-old male and female chickens (Bionda Piemontese, a slow-growing breed) raised under a free-range system and fed two finisher diets differing in lipid content: low-lipid (LL, ether extract 3.6%) and high-lipid (HL, ether extract 9.3%) diets. RNA was extracted from feather pulp, and 12 pools were subjected to whole RNA-Seq analysis. The study was designed as 2 × 2 factorial experiments investigating the effects of diet and sex on gene expression. A total of 17,360 transcripts were detected and used for downstream analyses. Differential gene expression and functional enrichment analyses were performed. The main effects of diet and sex were estimated with an additive design using the DEseq2 package, while for the sex-specific diet analyses, subgroup comparisons were conducted on the RaNA-Seq platform. The analysis of the main effect of diet reveals that three genes associated with ether lipid metabolism (PLA2G10, PLA2G4F, and ENPP6) were upregulated in chickens fed the HL diet. In roosters, HL feeding significantly altered the expression of APOA1 and SLC27A4, suggesting an effect on lipid transport and metabolic regulation within the PPAR signaling pathway. In contrast, hens showed differential expression primarily in pathways related to apelin signaling, extracellular matrix remodeling, and cardiovascular function, rather than classical lipid metabolism pathways; additionally, gene set enrichment analysis indicated a limited enrichment of linoleic acid metabolism, suggesting secondary involvement of lipid metabolic processes. These findings are consistent with those in the literature reporting sex-related differences between males and females. The results further suggest that transcriptomic responses to dietary lipid supplementation can be investigated through the expression of selected candidate genes in feather pulp. Among the genes identified, PLA2G10, PLA2G4F, ENPP6, APOA1, and SLC27A4 emerged as potential molecular markers associated with dietary treatment, and the importance of sex-dependent transcriptional responses was highlighted. In conclusion, this study demonstrates the potential of feather pulp as a viable source of RNA for transcriptomic analyses in live chickens, providing a minimally invasive alternative to conventional tissue sampling. These preliminary results also support the hypothesis that feathers represent a practical and ethically favorable tissue for future nutrigenomic and genetic improvement studies, ultimately supporting more sustainable poultry production. Full article

Scent marking has been discussed as an important component of communication in brown bears (Ursus arctos Linnaeus, 1758). However, the environmental factors influencing the occurrence of rub trees and their value for non-invasive genetic sampling remain poorly understood. This study examined the patterns of rub tree occurrence in the eastern High Tatra Mountains (Slovakia) at two spatial scales. At the tree scale, paired-design generalized linear mixed models showed that rub trees were more frequently recorded on large-diameter coniferous trees, indicating an association with visually prominent and chemically suitable substrates. At the landscape scale, logistic regression models revealed that the probability of rub tree occurrence increased with elevation and distance from human settlements, identifying high-elevation forests as areas of higher predicted rub tree occurrence. The best-supported model was used to produce a predictive map of rub tree occurrence across the study area. We also evaluated whether rub trees are reliable sources of biological material for non-invasive sampling. Hair collected during repeated field visits provided DNA suitable for genotyping and individual identification. Overall, the results show that rub trees exhibit non-random spatial patterns and represent effective focal points for systematic genetic sampling, linking patterns of rub tree occurrence to the spatial targeting of non-invasive genetic sampling in mountain landscapes. Full article

Background/Objectives: Malignant transformation of soft tissue lesions is uncommon but represents a significant diagnostic challenge with substantial clinical consequences. This spectrum encompasses four interrelated processes but biologically distinct processes: (1) true malignant transformation of benign lesions; (2) dedifferentiation of low-grade or intermediate malignancies; (3) secondary malignancy arising in chronic inflammatory or non-neoplastic conditions; and (4) apparent progression related to tumor heterogeneity and sampling error. Although these four entities involve biologically distinct mechanisms, they are grouped under “malignant progression” for conceptual clarity. While this umbrella approach has limitations due to biological heterogeneity, this unified radiologic framework aims to supplement, rather than oversimplify, their distinct biological behaviors. Representative examples include neurofibroma and epidermal inclusion cyst among benign lesions; atypical lipomatous tumor/well-differentiated liposarcoma, dermatofibrosarcoma protuberans, and solitary fibrous tumor among lesions showing dedifferentiation or malignant progression; and chronic inflammatory or scar-related conditions and previously irradiated tissue associated with secondary malignancy. Some lesions that appear to progress during follow-up may represent initial underdiagnosis rather than true biologic progression. Methods: This narrative review summarizes current imaging features, underlying pathologic mechanisms, and clinical risk factors associated with these processes in soft tissue lesions. Particular emphasis is placed on radiologic–pathologic correlation and conditions prone to histopathologic misinterpretation. Results: Imaging red flags—including interval or rapid growth, deep fascial invasion, heterogeneous enhancement, perilesional edema, and necrosis—should raise concern for malignant progression across these categories. However, overlapping imaging features and sampling errors may result in pathologic misdiagnosis and delayed treatment. Particularly, atypical lipomatous tumors are frequently misdiagnosed as simple lipomas, while fibrosarcomas may be erroneously interpreted as aggressive fibromatosis. Advanced imaging and multidisciplinary review may help reduce diagnostic errors. Patients with predisposing factors such as genetic syndromes, chronic inflammation, prior burns, or previous radiation exposure warrant close surveillance. Conclusions: Accurate diagnosis of soft tissue lesions with true malignant transformation, dedifferentiation, or secondary malignancy—as well as recognition of diagnostic pitfalls—is essential for appropriate management. Integrated radiologic–pathologic assessment may help improve diagnostic accuracy and clinical decision-making in soft tissue oncology. Full article

Small, isolated, and fragmented populations often exhibit low levels of genetic diversity as a result of genetic drift, limited gene flow, and inbreeding. Huemul (Hippocamelus bisulcus) is a medium-sized South American deer categorized as endangered by the International Union for Conservation of Nature (IUCN). Huemul in its northernmost range was previously distributed in Central Chile between 36° S and 37° S, but its current distribution and conservation status in the region are poorly documented. We used non-invasive genetic approaches to assess the genetic diversity, population connectivity, and demographic history of the huemul’s northernmost population using nuclear microsatellite markers from fecal samples of wild individuals. Observed nuclear DNA genetic variation (Ho = 0.2958 ± 0.0318) was moderate, and allelic richness was low (Ar = 3.43–4.01), consistent with the theoretical expectation that isolated populations may retain heterozygosity while losing allelic richness more rapidly. However, the estimated nuclear DNA effective population size was low (Ne = 47; 95% CI: 19.2–∞). Demographic simulations project continued loss of genetic diversity under all scenarios modeled. Our results provide a foundation for further study of this population and provide the genetic data necessary to design detailed management plans to ensure the persistence of healthy populations of this rare and elusive deer. Full article

The Fusobacterium nucleatum complex, which comprises oral lineage 1 (L1) strains, is strongly associated with colorectal cancer (CRC). NusG (N-utilisation substance G) is a transcription elongation factor that is universally conserved. This study aimed to develop and validate a novel nusG-directed polymerase chain reaction (PCR) assay to specifically and sensitively detect ectopic Fusobacterium L1 colonisation in clinical CRC patient samples. Four L1-specific primer pairs targeting the nusG gene were designed using MEGA11 software (Molecular Evolutionary Genetics Analysis, version 11.0.13) and successfully employed in 40 stool samples from CRC patients and healthy controls (HC). Additionally, five species-specific primer pairs were designed for the L1 species F. animalis clades 1 and 2, F. nucleatum, F. polymorphum, and F. vincentii, and were successfully applied to stool and saliva samples. Their specificity was verified via Sanger sequencing. Two L1-specific primer pairs (NusG5-F/NusG6-R and NusG2a-F/NusG5-R) demonstrated robust performance in our cohort, showing statistical significance (padj < 0.05) and a large effect size (|r| ≥ 0.5) in the difference in Ct values and absolute cell counts between CRC patients and the HC group. These primer pairs also exhibited promising preliminary diagnostic potential, with respective area under the curve (AUC) values of 0.909 and 0.883. However, Fusobacterium L1 abundance in saliva samples did not differ significantly between groups, indicating that definitive conclusions cannot be drawn due to the limited power of the salivary sub-cohort. The data indicates that nusG-based PCR primers could be used as reliable, non-invasive biomarkers as a complementary tool for early CRC diagnostics. While potentially applicable in the context of other Fusobacterium-implicated diseases, further validation in larger, ethnically diverse cohorts remains essential. Full article

Celiac disease (CD) is an autoimmune enteropathy of the small intestine affecting genetically susceptible individuals, characterized by an aberrant immune response to gliadin and sustained IgA-driven inflammation. IgA exists in two main subclasses, IgA1 and IgA2, which differ in distribution and function, but their profile in CD remains poorly characterized. Circulating free light chains (FLCs) are markers of B-cell activation and immune dysregulation, yet their role in CD has not been fully explored. The aim of this study was to characterize IgA subclasses and FLC profiles in newly diagnosed celiac patients. We analyzed sera from 108 CD patients and 29 healthy controls, assessing conventional serological markers (anti-tissue transglutaminase and anti-endomysial antibodies), together with total IgA, IgA1, IgA2, and FLC levels using a turbidimetric method. CD patients exhibited higher total IgA levels and an increased IgA1/IgA2 ratio, alongside a decreased k/λ ratio; these differences remained significant after adjustment for age and sex. When combined in a multivariable logistic model, these biomarkers yielded an AUC of 0.827, suggesting that the parameters identified in the univariate analyses provide complementary, non-redundant information that jointly highlights a reorganization of the humoral immune response. Due to the limited sample size, our results need confirmation in larger cohorts. However, our findings suggest a reorganization of the IgA compartment in CD, with selective expansion of IgA1 and preferential λ light chain usage, highlighting coordinated alterations in the humoral immune response. The integration of such markers, potentially in combination with -omics approaches, may contribute to a more refined and less invasive characterization of celiac disease. Full article

Molecular genetic approaches have become indispensable in veterinary medicine for early tumour detection, confirming histomorphological diagnoses, and non-invasive diagnosis using urine samples. In canine urothelial carcinoma (UC), molecular testing includes BRAF^V595E mutation analysis and the identification of specific copy number alterations (CNAs). CNAs affect multiple genes simultaneously, leading to their overexpression or underexpression. Recurrent gene gains on canine chromosomes (CFA)13 and CFA36 and losses on CFA19 are highly prevalent in canine UCs. This study evaluates CNA test performance and the influence of tumour purity on CNA test results using 76 histologically confirmed UC tissue samples. The tumour region of each histopathological section was measured, and its proportion relative to the total tissue area was calculated. In total, 58/76 cases (76.3%) were CNA-positive, defined by a ratio greater than 1.23 for both CFA13/19 and CFA36/19. A total of 14/18 negative cases had a ratio > 1.23 for CFA13/19 or CFA36/19. The sensitivity of this CNA assay depends on the applied thresholds, with a tumour-to-total tissue ratio of ≥20% yielding positive CNA results in 84% of cases, rising to 89% at ≥40%. These findings indicate that <20% tumour content and a moderate to high proportion of copy number-neutral cells may be associated with reduced CNA detectability. Full article

Background: Urinary cellular biomarkers detected by flow cytometry have emerged as promising non-invasive tools for the diagnosis, prognosis, and monitoring of renal and urological diseases. However, a comprehensive synthesis of their clinical applicability is lacking. Objectives: This review aimed to systematically evaluate and summarize the evidence of urinary cellular biomarkers identified through flow cytometry in human populations with renal or urological diseases. Methods: A systematic search of PubMed, Scopus and Web of Science was conducted from inception to January 2025. Observational studies analyzing urinary samples by flow cytometry were included, whereas animal studies, genetic-only analyses and studies with incomplete data were excluded. Data extraction and risk of bias assessment were performed independently by two reviewers using a standardized form and the Newcastle-Ottawa Scale. Due to heterogeneity in study populations, designs, and cytometry methods, results were synthesized narratively. Results: Of 3938 records screened, 23 studies met the inclusion criteria. All studies applied flow cytometry to characterize urinary cellular biomarkers across renal diseases. Several studies reported promising diagnostic and monitoring applications, but substantial heterogeneity in study design, cytometry protocols, and marker panels limited comparability. Longitudinal analyses and robust prognostic validation were infrequently reported. Conclusions: Urinary cellular biomarkers assessed by flow cytometry represent a promising non-invasive approach for evaluating renal and urological diseases. However, clinical implementation remains constrained by heterogeneity and insufficient validation. Future research should focus on standardized methodologies and large prospective studies to establish their diagnostic and prognostic utility. Full article

Background/Objectives: Child malnutrition is a global health challenge linked to poor growth, impaired development, weakened immunity, and adverse outcomes. Early risk detection is essential, but current screening tools differ in accuracy and feasibility. This study aimed to develop and validate new bedside pediatric malnutrition screening models based on machine learning and evolutionary computation methods that can capture complex patterns in non-invasive clinical indicators while remaining practical for routine ward use. Methods: We conducted a cross-sectional study including 180 hospitalized children (1 month–18 years) recruited consecutively from six pediatric wards. The required sample size (minimum 138 participants) was calculated a priori using national prevalence estimates of pediatric undernutrition (4–9.5%) to ensure adequate precision at a 95% confidence level. Data collection included a questionnaire, anthropometry, subjective malnutrition risk assessment, and the Subjective Global Nutritional Assessment (SGNA) tool. Screening models were developed using decision trees, random forests, XGBoost, lasso regression, artificial neural networks, ANFIS, and genetic programming. Their performance was evaluated against the SGNA tool and physician-based subjective malnutrition risk assessment using sensitivity, specificity, AUC, and Cohen’s κ. Results: Machine learning and intelligent evolutionary models (GP, ANN, and ANFIS) showed the best performance in this sample, with substantial to high agreement (κ = 0.81–1.00) and high diagnostic accuracy (AUC = 0.92–1.00) with the subjective malnutrition risk assessment. The GP model demonstrated the highest apparent accuracy in this dataset, but also higher complexity, whereas simpler models such as decision trees showed lower accuracy but greater interpretability and feasibility for routine clinical use. However, validation was performed on a relatively small independent sample, and no external validation was conducted, which may limit the generalizability of the findings. Conclusions: While complex models may serve as digital assessment instruments, simpler models are rapid and more suitable for bedside screening. All developed models are non-invasive and cost-effective and show potential for supportive approaches for early detection of malnutrition risk at hospital admission. However, given the limited validation sample and the absence of external validation, these findings should be interpreted with caution, and further large-scale, multicenter studies are required to confirm generalizability and clinical applicability. Full article

This scientometric review examines the evolution and scope of invasive (blood and tissue) and non-invasive (faeces, hair, and saliva) sampling in terrestrial mammal population genetics, with particular emphasis on the comparability of observed heterozygosity (Ho), expected heterozygosity (He), and the inbreeding coefficient (Fis) between studies published from 1985 to 2026. Searches in Web of Science and Scopus, filtered under PRISMA/PRISMA-S criteria, yielded a compendium of articles analysed with Bibliometrix and VOSviewer 1.6.20 to quantify temporal production, keyword evolution, collaborative networks, and publication outlets. Searches in Web of Science and Scopus, filtered under PRISMA/PRISMA-S criteria, yielded a broad corpus of 145 articles for general scientometric analyses, of which 85 met the eligibility criteria for the focused analysis of Ho, He, and Fis. The field shows steady growth (annual rate ≈ 6.1%), substantial authorship and international collaboration, and increasing thematic diversity. Adoption of non-invasive sampling has accelerated, broadening spatial and taxonomic coverage, but also increasing exposure to DNA degradation and genotyping error when laboratory quality control is insufficient. Across the literature, reporting of quality control practices (e.g., extraction blanks, negative PCR controls, multi-tube replication, and error-rate estimation) has improved over time but remains inconsistent. Comparisons indicate that differences in Ho, He, and Fis between invasive and non-invasive sampling are generally modest once marker system and species are taken into account. These findings indicate that quality control and transparency in reporting, rather than invasiveness per se, are the main factors determining comparability among studies. The scientometric patterns also reveal a methodological transition from microsatellites to SNP-based and reduced representation approaches, with implications for synthesis across marker types. Overall, this review identifies geographic and taxonomic biases in research effort and highlights the need for standardised reporting of DNA quality indicators, inclusion thresholds, and validation protocols to strengthen genetic monitoring in mammalian conservation. Full article

Misiones, Argentina, holds one of the largest remnants of the Atlantic Forest, with almost 1.4 million hectares of native forest, representing a critical landscape for sustainable biodiversity conservation. However, connectivity across this ecoregion is increasingly threatened by habitat conversion, landscape fragmentation, and poaching pressures that extend beyond protected area boundaries, undermining long-term sustainability of wildlife populations. Using conservation detection dogs, we located, collected, and genetically confirmed 198 scats belonging to four game species: 20 lowland tapir (Tapirus terrestris), 72 white-lipped peccary (Tayassu pecari), 55 collared peccary (Pecari tajacu), and 51 Azara’s agouti (Dasyprocta azarae). Analyses examining species-specific habitat associations emphasized the importance of extending inference beyond point locations to encompass species’ home ranges, with native forest consistently identified as a key component of habitat use. The high prevalence of scats in mosaics of human-modified habitats outside protected areas, especially along their borders, underscores the importance of managing these areas as part of a broader sustainable landscape matrix. While native forest fragments outside of protected areas may serve as important refugia supporting species persistence, their contribution to sustainable management depends on reducing poaching pressure across these landscapes. There is an urgent need to expand antipoaching efforts beyond protected areas and across the Atlantic Forest in the Green Corridor of Misiones while preventing ongoing deforestation and the expansion of monoculture plantations. Achieving sustainable wildlife management in this region will require integrated strategies that promote sustainable land use, conservation planning, and rural development. Full article

Nitrate (NO₃⁻) serves as a pivotal molecule with dual functions in nutrient supply and signaling during plant growth and development. Precise monitoring of its spatiotemporal dynamics in planta is therefore essential for dissecting the regulatory mechanisms underlying plant nitrogen metabolism. However, conventional nitrate detection methods suffer from inherent limitations, including destructive sampling, insufficient spatiotemporal resolution, and an inability to achieve real-time whole-plant monitoring. Here, we report a genetically encoded nitrate biosensor, designated NitNRCL1, constructed using a split firefly luciferase complementation system. Functional validation in both prokaryotic and eukaryotic systems demonstrates that NitNRCL1 responds to changes in nitrate availability and generates stable chemiluminescent signals in bacteria and diverse plant species. Importantly, NitNRCL1 enables non-invasive, real-time, and whole-plant monitoring of nitrate levels in living plants. Using NitNRCL1, we successfully imaged the spatiotemporal dynamics of nitrate signaling in Arabidopsis thaliana. Collectively, our findings establish NitNRCL1 as a robust and novel tool for investigating nitrate transport, signaling, and metabolic pathways in plants. This biosensor advances our mechanistic understanding of plant nitrate biology and provides a technical foundation for breeding nitrogen-use-efficient crops and developing precision fertilization strategies. Full article

Background: Pangolins are critically endangered mammals, and a comprehensive understanding of their genetic diversity is crucial for effective conservation. The mitochondrial genome serves as a vital molecular marker for phylogenetic and population genetic studies. Obtaining genetic material from these elusive animals non-invasively remains a challenge. This study aimed to sequence and characterize the complete mitochondrial genome of Manis javanica and explore the phylogenetic relationships among pangolin species. Methods: The complete mitochondrial genome was sequenced from a saliva-derived sample. Standard procedures for DNA extraction, amplification, and sequencing were employed. The genome was assembled and annotated using bioinformatic tools. Phylogenetic analysis was conducted based on the cytochrome c oxidase subunit I (COXI) gene sequences from nine pangolin species, with the resulting tree constructed using the maximum-likelihood method. Results: The complete mitochondrial genome of M. javanica (GenBank accession: PP110760) is a circular molecule of 16,573 bp, containing 13 protein-coding genes, 22 tRNA genes, 2 rRNA genes, and a control region. The overall base composition showed a lower GC content (43.83%) than AT content (56.17%). Phylogenetic analysis based on COXI sequences delineated the nine species into three distinct genera: Manis, Phataginus, and Smutsia. Within the genus Manis, Manis pentadactyla was identified as the closest relative to M. javanica. The newly described species Manis mysteria was found to be closer to Manis culionensis and Manis crassicaudata than to other congeners. Furthermore, the analysis indicated that African pangolins diverged earlier than Asian pangolins. Conclusions: This study successfully demonstrates the feasibility of extracting and sequencing the complete mitochondrial genome from saliva samples, providing a valuable non-invasive method for future genetic studies on pangolins. The genomic data and phylogenetic results offer significant molecular insights that will benefit the genetic management and conservation of critically endangered pangolin resources. Full article

Background/Objectives: Non-indigenous species are increasingly reshaping Mediterranean marine ecosystems, particularly under ongoing climate warming. The rayed pearl oyster Pinctada radiata, a thermophilic species originating from the Indo-Pacific region, is one of the earliest and most successful invaders in the Mediterranean Sea and has recently established populations in the Adriatic Sea. Methods: This study integrates preliminary shell morphometric data with molecular genetic analyses based on mitochondrial cytochrome c oxidase subunit I (COI) and nuclear internal transcribed spacer 2 (ITS2) markers to confirm species identity and examine patterns of genetic variation in comparison with other Mediterranean Sea regions and the Persian Gulf. Results: Phylogenetic analyses based on COI confirmed P. radiata as a distinct and well-supported monophyletic lineage, whereas the nuclear ITS2 marker showed limited resolution and interspecific overlap. Mediterranean and Adriatic populations showed low COI haplotype and nucleotide diversity and weak genetic structuring, consistent with recent colonization and secondary expansion, whereas Persian Gulf populations were more genetically diverse. Conclusions: Future studies should employ larger sample sizes and broader geographic sampling across both the Mediterranean Sea and the full native range of P. radiata, combined with high-resolution genome-wide nuclear markers, to better resolve connectivity and invasion dynamics. Full article