Search Results (816)

The diet of dairy calves can be contaminated with mycotoxins, posing a potential risk to animal health. This case study report aimed to make the first assessment of the presence of multiple mycotoxins in concentrates fed to dairy calves in Brazil. A total of 19 concentrate samples intended for dairy calves were analyzed using liquid chromatography coupled with mass spectrometry. Aflatoxins, deoxynivalenol, and T-2 toxin were not detected in any samples, whereas fumonisins B₁ (FB₁) and B₂ (FB₂) were present in 100% of the samples, with mean concentrations of 2750.1 μg/kg and 834.9 μg/kg, respectively. Zearalenone (ZEN) was detected in 36.8% of samples, with a mean concentration of 929.9 μg/kg. Significant correlations were observed between FB₁ and FB₂ (ρ = 0.978; p < 0.001) and between FB₂ and ZEN (ρ = 0.735; p = 0.05). While the physical form of the concentrate did not influence (p > 0.05) mycotoxin concentrations, a trend was observed for FB₁ (ρ = −0.417; p = 0.07) and FB2 (ρ = −0.395; p = 0.09). These findings highlight the frequent occurrence of Fusarium mycotoxins, likely due to pre-harvest contamination, emphasizing the potential risk of additive or synergistic effects in dairy calves. Full article

Surveillance of swine influenza A virus (swIAV) traditionally focuses on respiratory matrices, yet emerging evidence suggests that fecal shedding and secondary environmental contamination may also contribute to viral dissemination. In this study, we collected and analyzed nasal, rectal, environmental, milk, and colostrum samples from naturally infected pigs in a commercial farm in Minas Gerais, Brazil. IAV RNA was detected in 25% of samples, including 42% from asymptomatic animals, with nasal swabs showing higher detection rates (30%) than rectal swabs (20%), though rectal Ct values were consistently higher, indicative of lower viral loads. We successfully isolated viable viruses from feces and effluent samples. Whole-genome sequencing revealed co-circulation of enzootic pH1N1 clade #2 (HA) and pN1 clade #4 (NA), alongside human-origin H3N2 sequences clustering within clade 3C.2a1b.2a.2a.1, and N2 segments related to pre-3C human lineages from 2001 to 2002. Phylogenetic and p-distance analyses support both recent reverse zoonosis and historical transmission events. Detection of complete HA/NA sequences from rectal swabs and treated effluent further emphasizes the surveillance value of non-respiratory matrices. The integration of respiratory and fecal/environmental sampling appears important to achieve more comprehensive IAV monitoring in swine herds and may have significant implications for One Health strategies in Brazil and beyond. Full article

Background: Polymerase chain reaction (PCR) is highly sensitive and specific for the rapid diagnosis of invasive fungal disease (IFD) but is not yet widely implemented due to concerns regarding limited standardisation between assays, the lack of commercial options and the absence of clear guidance on interpreting results. Objectives and Methods: This review provides an update on technical and clinical aspects of PCR for the diagnosis of the most pertinent fungal pathogens, including Aspergillus, Candida, Pneumocystis jirovecii, Mucorales spp., and endemic mycoses. Summary: Recent meta-analyses have demonstrated that quantitative PCR (qPCR) offers high sensitivity for diagnosing IFD, surpassing conventional microscopy, culture and most serological tests. The reported specificity of qPCR is likely underestimated due to comparison with imperfect reference standards with variable sensitivity. Although the very low limit of detection of qPCR can generate false positive results due to procedural contamination or patient colonisation (particularly in pulmonary specimens), the rates are comparable to those observed for biomarker testing. When interpreting qPCR results, it is essential to consider the pre-test probability, determined by the patient population, host factors, clinical presentation and risk factors. For patients with low to moderate pre-test probability, the use of sensitive molecular tests, often in conjunction with serological testing or biomarkers, can effectively exclude IFD when all tests return negative results, reducing the need for empirical antifungal therapy. Conversely, for patients with high pre-test probability and clinical features of IFD, qPCR testing on invasive specimens from the site of infection (such as tissue or bronchoalveolar lavage fluid) can confidently rule in the disease. The development of next-generation sequencing methods to detect fungal infection has the potential to enhance the diagnosis of IFD, but standardisation and optimisation are essential, with improved accessibility underpinning clinical utility. Full article

Background: Urine microbial analysis is a frequently requested test that is often associated with contamination during specimen collection or storage, which leads to false-positive diagnoses and delayed reporting. In the era of digitalization, machine learning (ML) can serve as a valuable tool to support clinical decision-making. Methods: This study investigates the application of a simple artificial neural network (ANN) to pre-identify negative and contaminated (false-positive) specimens. An ML model was developed using 8181 urine samples, including cytology, dipstick tests, and culture results. The dataset was randomly split 2:1 for training and testing a multilayer perceptron (MLP). Input variables with a normalized importance below 0.2 were excluded. Results: The final model used only microbial and either urine color or urobilinogen pigment analysis as inputs; other physical, chemical, and cellular parameters were omitted. The frequency of positive and negative specimens for bacteria was 6.9% and 89.6%, respectively. Contaminated specimens represented 3.5% of cases and were predominantly misclassified as negative by the MLP. Thus, the negative predictive value (NPV) was 96.5% and the positive predictive value (PPV) was 87.2%, leading to 0.82% of the cultures being unnecessary microbial cultures (UMC). Conclusions: These results suggest that the MLP is reliable for screening out negative specimens but less effective at identifying positive ones. In conclusion, ANN models can effectively support the screening of negative urine samples, detect clinically significant bacteriuria, and potentially reduce unnecessary cultures. Incorporating morphological information data could further improve the accuracy of our model and minimize false negatives. Full article

Background: Cell-free DNA (cfDNA) has become a cornerstone of liquid biopsy applications, offering promise for early disease detection and monitoring. However, its widespread clinical adoption is limited by variability in pre-analytical processing, especially during isolation. Current extraction methods face challenges in yield, purity, and reproducibility. Methods: We developed and optimized SafeCAP 2.0, a novel magnetic bead-based cfDNA extraction kit, focusing on efficient recovery, minimal genomic DNA contamination, and PCR compatibility. Optimization involved systematic evaluation of magnetic bead chemistry, buffer composition, and reagent volumes. Performance was benchmarked against a commercial reference kit (Apostle MiniMax) using spiked oligonucleotides and plasma from patients with stage IV NSCLC. Results: The optimized protocol demonstrated superior recovery with a limit of detection (LoD) as low as 0.3 pg/µL and a limit of quantification (LoQ) of 1 pg/μL with no detectable PCR inhibition. In comparative studies, SafeCAP 2.0 showed equivalent or improved performance over the commercial kit. Clinical validation using 47 patient plasma samples confirmed robust cfDNA recovery and fragment integrity. Conclusions: SafeCAP 2.0 offers a cost-effective, high-performance solution for cfDNA extraction in both research and clinical workflows. Its design and validation address key pre-analytical barriers, supporting integration into routine diagnostics and precision medicine platforms. Full article

Nowadays, there is special interest in promoting the consumption of ancestral crops and minimally processed foods with high nutritional value. However, besides nutritional issues, safety assessments must be addressed. This study aimed to evaluate mycotoxin contamination in five minimally processed traditional Ecuadorian foods: ochratoxin A (OTA), fumonisin B₁ (FB₁), and aflatoxins (AFs) in brown rice, lupin, and quinoa; OTA, FB₁, and deoxynivalenol (DON) in whole-wheat flour; and OTA and AFs in peanuts. Samples (45 samples of peanuts and whole-wheat flour, 47 of brown rice, 46 of quinoa, and 36 of lupin) were collected from local markets and supermarkets in the three most populated cities in Ecuador. Mycotoxins were determined by RP-HPLC with fluorescence and detection. Results were compared with the maximum permitted levels (MPLs) of European Regulation 2023/915/EC. Overall contamination reached up to 59.8% of the analyzed samples (38.4% with one mycotoxin and 21.5% with co-occurrence). OTA was the most prevalent mycotoxin (in 82.6% of quinoa, 76.7% of whole-wheat flour, 53.3% of peanuts, 48.6% of lupin, and 25.5% of brown rice), and a modest number of quinoa (17%) and lupin (5.7%) samples surpassed the MPLs. DON was found in 82.2% of whole-wheat flour (28.9% > MPL). FB₁ was detected in above 25% of brown rice and whole-wheat flour and in 9% of the quinoa samples. FB₁ levels were above the MPLs only for whole-wheat flour (17.8%). AFB₁ and AFG₁ showed similar prevalence (about 6.5 and 8.5%, respectively) in quinoa and rice and about 27% in peanuts. Overall, these findings underscore the importance of enhancing fungal control in the pre- and post-harvest stages of these foods, which are recognized for their high nutritional value and ancestral worth; consequently, the results present key issues related to healthy diet promotion and food sovereignty. This study provides compelling insights into mycotoxin occurrence in minimally processed Ecuadorian foods and highlights the need for further exposure assessments by combining population consumption data. Full article

This study investigated individuals’ emotional responses to bed bugs and how these were influenced by sex, proximity, and educational intervention. Using a pre-post experimental design, participants (n = 157) completed emotional assessments before and after viewing an educational video about bed bugs. Contrary to our initial hypothesis that only fear and disgust would be observed, participants also exhibited high levels of anxiety and anger. Following the educational intervention, disgust, fear, and anger toward bed bugs increased significantly. Participants experienced greater disgust and fear when imagining encounters with bed bugs in closer proximity, with home infestations giving stronger responses than workplace scenarios. The educational video reduced disgust toward bed bugs in the home but increased fear of them in public spaces, potentially promoting vigilance that could limit bed bug spread. Females reported higher levels of disgust and fear than males across all proximity conditions, supporting evolutionary theories regarding sex-specific disgust sensitivity. The educational video successfully increased participants’ knowledge about bed bugs while simultaneously shifting emotional responses from contamination-based disgust to threat-specific fear. These findings suggest that educational interventions can effectively modify emotional responses to bed bugs, potentially leading to more rational management behaviors by transforming vague anxiety into actionable awareness of specific threats. Full article

The CRISPR/Cas system has attracted increasing attention in accurate nucleic acid detection. Herein, we reported a CRISPR/Cas12a-chemiluminescence cascaded bioassay (CCCB) for the amplification-free and sensitive detection of human papillomavirus type 16 (HPV-16) and parvovirus B19 (PB-19). A magnetic bead (MB)-linking single-stranded DNA (LssDNA)-alkaline phosphatase (ALP) complex was constructed as the core component of the bioassay. During the detection process, the single-stranded target DNA was captured and enriched by LssDNA and then activated the trans-cleavage activity of Cas12a. Due to the Cas12a-mediated cleavage of LssDNA, ALP was released from the MB, subsequently catalyzing the substrate to generate a chemiluminescence (CL) signal. Given the cascade combination of CRISPR/Cas12a with the CL technique, the limits of detection for HPV-16 and PB-19 DNA were determined as 0.14 pM and 0.37 pM, respectively, and the whole detection could be completed within 60 min. The practicality and reliability of the platform were validated through target-spiked clinical specimens, and the recovery rate was 93.4–103.5%. This dual-amplification strategy—operating without target pre-amplification—featured high specificity, low contamination risk, facile preparation, and robust stability. It provides a novel approach for sensitive nucleic acid detection, with the potential for rapid extension to the diagnosis of various infectious diseases. Full article

Introduction: Despite the approval of novel agents that have significantly improved long-term survival rates for multiple myeloma (MM) patients undergoing autologous stem cell transplant (ASCT), most patients eventually relapse. The failure to achieve or maintain bone marrow (BM) minimal residual disease (MRD) negativity is a recognised adverse prognostic factor for progression-free survival (PFS) and overall survival (OS). Contamination of stem cell apheresis by clonal plasma cells may also affect prognosis, though data remain limited. Methods: We conducted a prospective, single-centre observational study including 100 newly diagnosed MM patients eligible for ASCT and treated with bortezomib-based triplet induction. MRD was assessed both on BM and apheresis samples using multiparameter flow cytometry (MFC-MRD) with a sensitivity of 10⁻⁵. Results: Clonal plasma cells were detected in 22 apheresis samples (aMRD+), all of which were associated with BM MRD positivity. Patients with aMRD+ had inferior pre-ASCT responses (≥VGPR: 10% vs. 63%, p = 0.005) and worse post-ASCT BM MRD negativity rates (4% vs. 49%, p = 0.048). After a median follow-up of 52.4 months, aMRD+ was associated with shorter progression-free survival (median 38.5 vs. not reached, p = 0.007) and overall survival (median 60 months vs. not reached, p = 0.003). Conclusions: Contamination of the apheresis product is associated with persistent BM disease and poorer outcomes. Combined MRD assessment in both bone marrow and apheresis may improve risk stratification in MM patients undergoing ASCT. Full article

The global plastic crisis has generated significant interest in repurposing waste plastics as asphalt modifiers, presenting both environmental and engineering advantages. This study offers a comprehensive review of the applications of waste plastics in asphalt, focusing on their types, modification mechanisms, incorporation techniques, and environmental impacts, alongside proposed mitigation strategies. Commonly utilized plastics include polyethylene (PE), polypropylene (PP), polystyrene (PS), polyvinyl chloride (PVC), and polyethylene terephthalate (PET), each affecting asphalt performance differently—enhancing high-temperature stability and fatigue resistance while exhibiting varying levels of compatibility and environmental risks. The incorporation techniques, namely wet and dry processes, differ in terms of efficiency, cost, and environmental footprint: the wet process enhances durability but requires more energy, whereas the dry process is more cost-effective but may lead to uneven dispersion. Environmental concerns associated with these practices include toxic emissions (such as polycyclic aromatic hydrocarbons and volatile organic compounds) during production, microplastic generation through abrasion and weathering, and ecological contamination of soil and water. Mitigation strategies encompass optimizing plastic selection, improving pre-treatment and compatibilization methods, controlling high-temperature processing, and monitoring the spread of microplastics. This review highlights the need for balanced adoption of waste plastic-modified asphalt, emphasizing sustainable practices to maximize benefits while minimizing risks. Full article

Arsenic contamination in groundwater is a significant public health concern, with As(III) posing a greater and more challenging risk than As(V) due to its higher toxicity, mobility, and weaker adsorption affinity. Fe-Mn-based adsorbents offer a promising solution, simultaneously oxidizing As(III) to As(V), enhancing its adsorption. This study evaluates an Fe-Mn nanocomposite across typical batch (20 mg of adsorbent), fixed-bed column (28 g), and pilot-scale (2.5 kg) studies, bridging the gap between laboratory and real-world applications. Batch experiments yielded maximum adsorption capacities of 6.25 mg/g and 4.71 mg/g in a synthetic matrix and real groundwater, respectively, demonstrating the impact of the water matrix on adsorption. Operational constraints and competing anions led to a lower capacity in the pilot (0.551 mg/g). Good agreement was observed between the breakthrough curves in the pilot (breakthrough at 475 bed volumes) and the fixed-bed column studies (365–587 bed volumes) under similar empty bed contact times (EBCTs). The Thomas, Adams–Bohart, and Yoon–Nelson models demonstrated that lower flow rates and extended EBCTs significantly enhance arsenic removal efficiency, prolonging the operational lifespan. Our findings demonstrate the necessity of continuous-flow experiments using real contaminated water sources and the importance of optimizing flow conditions, EBCTs, and pre-treatment in order to successfully scale up Fe-Mn-based adsorbents for sustainable arsenic removal. Full article

The conservation of small and genetically vulnerable brown bear populations, such as the Cantabrian subpopulation in Spain, depends on developing species-specific assisted reproductive technologies and genetic resource banks. However, the lack of standardized sperm collection and cryopreservation protocols hinders their application. This study provides the first comparative analysis of sperm quality and proteomic profiles from three different origins: epididymal, pre-ejaculated, and ejaculated. Sperm quality parameters —motility and kinetic, viability, apoptosis, and oxidative stress— and protein expression were assessed. Although yields were similar, marked differences were observed in sperm quality and protein profiles. Sixty-three proteins involved in metabolism, stress response, and oxidative balance were differentially expressed depending on sperm origin. Epididymal sperm showed the highest viability and motility, lowest apoptosis, and a proteomic profile indicative of active spermatogenesis and enhanced oxidative stress defense. In contrast, ejaculated sperm had increased oxidative stress and reduced expression of metabolic proteins, while pre-ejaculated sperm exhibited lower motility, likely due to urine contamination and mitochondrial protein alterations, despite comparable viability and apoptosis. These findings offer novel insights into brown bear sperm biology and highlight the importance of sperm origin in developing optimized assisted reproduction strategies, ultimately supporting ex situ conservation efforts for this species. Full article

Donor human milk (DHM) can harbor microbial contaminants that cause serious infections in premature infants. Bacillus cereus is a pathogen frequently found in DHM, capable of forming spores that can resist Holder pasteurization (62.5 °C, 30 min). Since no microbial growth is acceptable in post-pasteurized DHM, microbiological testing of pre-pasteurized DHM provides information about its contamination level to determine if it should be accepted for pasteurization. Culture is the gold standard in microbiological control but it requires 24–48 h to provide results. In this study we developed and validated a non-commercial real-time PCR assay for the detection of Bacillus cereus (BC test) in DHM specimens on a fully automated high-throughput platform, the cobas^® 6800 system. The BC test showed excellent sensitivity and specificity, repeatability and linearity over an 8-log range and a low limit of detection in milk specimens, as well as good agreement with selective culture methods. BC test was then used to systematically control all milk donations (3439) over a 24-month period. Bacillus cereus was detected in 14.2% of DHM, with monthly rates ranging from 6 to 29% and a significantly higher incidence in warmer months. Incorporating this assay into our laboratory workflow improved efficiency and reduced turnaround time. Full article

Circulating cell-free DNA (cfDNA) is an important biomarker for various cancer types, enabling a non-invasive testing approach. However, pre-analytical variables, including sample collection, tube type, processing conditions, and extraction methods, can significantly impact the yield, integrity, and overall quality of cfDNA. This study presents a comprehensive analytical validation of a magnetic bead-based, high-throughput cfDNA extraction system, with a focus on assessing its efficiency, reproducibility, and compatibility with downstream molecular applications. The validation was performed using a range of sample types: synthetic cfDNA spiked into DNA-free plasma, multi-analyte ctDNA plasma controls, Seraseq ctDNA reference material in a plasma-like matrix, extraction specificity controls, residual clinical specimen from patients, and samples from healthy individuals stored at room temperature or 4 °C for up to 48 h to assess stability. Extracted cfDNA was analyzed for concentration, percentage, and fragment size, using the Agilent TapeStation. Variant detection was evaluated using a next-generation sequencing (NGS) assay on the Seraseq ctDNA reference material. The results demonstrated high cfDNA recovery rates, consistent fragment size distribution (predominantly mononucleosomal and dinucleosomal), minimal genomic DNA (gDNA) contamination, and strong concordance between detected and expected variants in reference materials. The workflow also showed robust performance under different study parameters, variable sample conditions, including sample stability and integrity. Together, these findings confirm the efficiency and reliability of the evaluated cfDNA extraction system and underscore the importance of standardized pre-analytical workflows for the successful implementation of liquid biopsy for early cancer detection, therapeutic monitoring, and improved patient outcomes. Full article

Aflatoxin (AF) contamination of crops during the pre-harvest period is a significant global concern for food and feed safety (FFS). In Europe, climate change presents a growing threat to agricultural products by increasing the risk of AF contamination. This umbrella review evaluates the scope and quality of pre-harvest data on climate-related AF contamination in Europe, addressing key questions: What insights do researchers provide on the relationship between climate change and pre-harvest AF contamination, and what data are lacking? Which crops are the focus of current research, and where in Europe are these studies concentrated? How is the data presented, and is it standardized? We conducted an umbrella literature review, extracting relevant studies from PubMed and Scopus up to 14 October 2024. Our findings indicate that rising temperatures, droughts, and shifting rainfall patterns increasingly favor the growth of aflatoxigenic fungi and pre-harvest AF contamination in European crops, posing risks to FFS and agricultural stability. However, inconsistencies in data collection and reporting limit cross-regional comparisons and hinder the development of effective mitigation strategies. Standardizing methodologies and improving data accessibility will enhance predictive modeling, strengthen risk assessments, and support targeted adaptation efforts, providing actionable insights for policymakers and agricultural stakeholders. Full article