Commercial Kits Research Articles

Quantum dot-based biomimetic fluorescence immunoassays for enrofloxacin detection in animal-derived foods.

Long-term consumption of foods with excessive enrofloxacin (ENRO) residues may cause the accumulation of ENRO in the human body, thus damaging human health. In this study, quantum dot-based biomimetic fluorescence immunoassays were used for enrofloxacin detection in food of animal origin. Under the most suitable conditions, the detection limit (IC15) of the method in standard solution was 0.13 ± 0.02 ng L-1 and the sensitivity (IC50) was 0.13 ± 0.18 μg L-1. The recoveries for ENRO from four fortified food samples, including chicken, eggs, shrimp, and milk, ranged from 85.74% to 114.19% and the coefficients of variation were 0.01-18.09%. The established method shows good agreement with the results obtained using commercial ELISA kits, with a correlation coefficient of 0.997. The proposed method shows the advantages of high sensitivity, specificity and wide detection range. It can be used as an alternative method for the rapid and sensitive detection of ENRO in food of animal origin.

Antibody isotyping and cytokine profiling in natural cases of Burkholderia mallei infection (glanders) in equines.

Immunological aspects of B. mallei infection were rarely studied in natural cases of equines. The present study was conducted to determine IgG, IgM, IgA and IgG (T) titre against recombinant Hcp1, TssA and TssB proteins and PilA-Hcp1-TssN-BipD and BpaB-BpaC- BMAA0553 chimeras and cytokine responses in glanders affected equine serum. The study was conducted on serum samples collected from 151 glanders positive equines which include horses (n=134), mules (n=16), and donkeys (n=01). The IgM, IgG, IgA and IgG (T) titre were determined against recombinant antigens by indirect ELISA and interleukin(IL)-1β, IL-17, IL-6, tumor necrosis factor alpha(TNF-α), monocyte chemoattractant protein 1 (MCP-1) and interferon gamma (IFN-γ) responses were measured by commercial ELISA kits. The study showed that glanders affected equines elicited strong antibody response against Hcp1, moderate responses against TssA and TssB, and weak responses against two chimeras. Among the cytokines, IL-1β, MCP-1, IL-17 and IL-6 concentration were significantly higher in glanders affected equine serum. We found that IgG antibody titre was higher than IgM, IgG (T) and IgA isotypes and Hcp1 was most predominant antibody inducers in comparison to TssA, TssB, PilA-Hcp1-TssN-BipD and BpaB-BpaC-T2SS proteins. The elevated level of IL-1β, MCP-1, IL-17, IL-6, IFN-γ and TNF-α observed in this study support the important role of this cytokines for augmenting cellular defence by recruitment of macrophages, neutrophil and dendritic cells against B. mallei infection. Further studies should be conducted to determine memory cell responses in natural cases of equine glanders using recombinant B. mallei proteins for identifying well-characterized immuno-protective vaccine candidates.

Detection of Antimicrobial-Induced Survival/Dead Bacteria via mEos4b Photoconversion: A Preliminary Study.

The escalating prevalence of hospital-acquired infections poses a critical challenge for healthcare systems worldwide. Effective management requires rapid identification of pathogens and their antibiotic resistance profiles. In this study, we utilized the photoconvertible mEos4b protein, which transitions from green to red fluorescence upon blue light exposure, to distinguish live from dead bacteria. The mEos4b gene was cloned into a prokaryotic vector and expressed in Escherichia coli BL21. The Minimum Inhibitory Concentration (MIC) of the transgenic bacteria was determined for five antibiotics, followed by a post-antibiotic effect assessment over a two-hour exposure period. The optimal photoconversion time for mEos4b was established as 90 seconds, and confocal microscopy was used to visualize live (green) and dead (red) cells post-exposure. The mEos4b-TR system proved highly specific, accurately distinguishing live and dead bacteria without producing false positives, even in control groups, which is a common issue in commercial live-dead kits. By relying on cellular metabolic activity rather than dyes, this system minimizes nonspecific interactions and contamination, making it more reliable than traditional methods prone to false readings. These results highlight the potential of the mEos4b-TR system as a superior alternative for rapid, precise bacterial viability assessments, particularly in determining antibiotic susceptibility. This preliminary study demonstrates the system's differentiation of viable and non-viable cells, suggesting its potential application in future studies involving novel antibacterial agents to refine antibiotic sensitivity testing.

Culture-Independent Quantitative PCR Detected Mobilized Colistin Resistance Genes (mcr-1, mcr-2, mcr-3, mcr-4, and mcr-5) in Chicken Gut Contents in Bangladesh

Inappropriate antimicrobial use in food animal farming propels antimicrobial resistance (AMR) that affects all health domains. Colistin is a ‘Reserve’ antibiotic for human treatment to be conserved for multidrug-resistant pathogens; however, it is being used as an animal growth promoter in many developing countries. The evolution of mobilized colistin resistance (mcr) gene-mediated colistin resistance has been reported to be associated with rampant colistin use. This study investigated the current variants of the mcr gene in chicken gut contents in Bangladesh. A cross-sectional study was designed to assess the mcr-1 to mcr-5 genes in 80 fresh poultry droppings from commercial poultry farms and 40 poultry droppings from household farms. DNA was extracted from each poultry dropping using commercial kits (Qiagen GmbH, Hilden, Germany). Real-time quantitative polymerase chain reaction (RT-qPCR) was employed using the qTOWER3 thermal cycler (Analytik Jena GmbH, Jena, Germany) to analyze the mcr gene variants in the extracted DNA. This study observed that 47.5% (57/120) of the samples exhibited the presence of at least one mcr gene out of the five variants investigated. The individual detection rates of the mcr-1, mcr-2, mcr-3, mcr-4, and mcr-5 genes were 42.5% (51/120), 2.5% (3/120), 1.7% (2/120), 5% (6/120), and 9.2% (11/120), respectively. The co-carriage of two or more genes was found in over 10% (10/57) of the samples. The triple occurrence of mcr genes was identified in three samples with the combination of mcr-1+mcr-2+mcr-4, mcr-1+mcr-3+mcr-5, and mcr-1+mcr-4+mcr-5. Overall, a significantly higher number of mcr genes were identified in the commercial farm chicken droppings compared to the household chicken droppings (p = 0.007). The existence of mcr genes in poultry feces in Bangladesh emphasizes the importance of proper poultry waste disposal and good hygiene practices in poultry livestock and its value chain. The potential impact of environmental ARGs should be considered in national and global policy documents. An integrated and combined approach to the One Health concept should be applied in all domains to understand and control the environment’s role in the evolution and transmission of AMR.

Comparative evaluation of commercial DNA isolation approaches for nanopore-only bacterial genome assembly and plasmid recovery.

The advent of Oxford Nanopore Technologies has undergone significant improvements in terms of sequencing costs, accuracy, and sequencing read lengths, making it a cost-effective, and readily accessible approach for analyzing microbial genomes. A major challenge for bacterial whole genome sequencing by Nanopore technology is the requirement for a higher quality and quantity of high molecular weight DNA compared to short-read sequencing platforms. In this study, using eight pathogenic bacteria, we evaluated the quality, quantity, and fragmented size distribution of extracted DNA obtained from three different commercial DNA extraction kits, and one automated robotic platform. Our results demonstrated significant variation in DNA yield and purity among the extraction kits. The ZymoBIOMICS DNA Miniprep Kit (ZM) provided a higher purity of DNA compared to other kit-based extractions. All kit-based DNA extractions were successfully performed on all twenty-four samples using a single MinION flow cell, with the Nanobind CBB Big DNA kit (NB) yielding the longest raw reads. The Fire Monkey HMW-DNA Extraction Kit (FM) and the automated Roche MagNaPure 96 platform (RO) outperformed in genome assembly, particularly in gram-negative bacteria. Based on our finding, we recommend a minimum read coverage and raw read N50, obtained from the appropriate DNA extraction kit for each bacterial species, to optimize genome assembly and plasmid recovery. This approach will assist end-users in selecting the most effective kit-based extraction method for bacterial whole-genome assembly using only long-read nanopore sequences.

Surface monitoring of L. monocytogenes by real-time fluorescence and colorimetric LAMP.

Listeria monocytogenes is a major foodborne pathogen affecting developing, and developed countries. The analysis of food contact surfaces in food industries is key for better controlling this pathogen. The current study focused on the development, optimization, and evaluation of a rapid and simple method for the detection of L. monocytogenes on stainless steel surfaces, suitable for decentralized setups, taking advantage of Loop-mediated isothermal amplification (LAMP). This was accomplished using a general pre-enrichment broth (TSB), with a simple DNA extraction based on a chelating resin, and final isothermal amplification. Two different detection strategies were tested, real-time fluorescence and naked-eye colorimetric, which were evaluated after 5, 7, and 24h of pre-enrichment. Regardless the detection chemistry selected, after 5-7h of pre-enrichment, 103-104CFU/cm2 were needed to obtain a positive result, while after 24h, it was possible to detect concentrations below 10CFU/cm2. Within each given time, all the performance parameters calculated, relative sensitivity, specificity, and accuracy, reached values higher than 80-90%; likewise, a Cohen's k of concordance with a culture-based approach higher than 0.8. Overall, the most sensitive assay can be performed in roughly 25h. This time-to-result outperforms commercial kits with the added value of specifically detecting L. monocytogenes instead of Listeria spp. KEY POINTS: • Real-time fluorescence and naked-eye colorimetric, were compared for the novel assay. • An LOD50 of 3.4CFU/cm2 and 4.2CFU/cm2 was calculated for the two assays. • Three pre-enrichment times were compared providing 24h better results.

Optimizing Genomic DNA Extraction from Avian Feathers: A Modified Phenol-Chloroform Approach for Enhanced Efficiency and Cost-Effectiveness.

The procurement of blood and tissue samples for DNA extraction in avian species intended for molecular studies is associated with the induction of discomfort and pain in the subjects, compounded by practical challenges in application and ethical considerations. Consequently, feathers have emerged as a more prevalent source for molecular investigations, particularly in the fields of poultry and ornithology. However, the effective extraction of DNA from feathers necessitates the breakdown of the hard keratinized tissue within the feather structure. This study aimed to devise a highly efficient, cost-effective, and easily adaptable Modified Phenol-Chloroform (MPC) approach for genomic DNA extraction from feathers, addressing shortcomings identified in previous studies on feather-based DNA isolation. The MPC method was employed to extract genomic DNA from feather samples obtained from six distinct avian species (chicken, guinea fowl, canary, pigeon, emu, and goose). Comparative evaluation of DNA isolation efficiency was conducted by employing two different commercial DNA kits alongside the MPC method. The results showed significantly higher DNA concentrations (ng/ml) from chicken feathers using the MPC method compared to those obtained with commercial kits (p < 0.05), along with high DNA purity (1.83 ± 0.11). Subsequent PCR experiments, employing nuclear and mitochondrial DNA-specific primers, illustrated the effective amplification of short and long fragments from MPC-isolated DNA samples. In contrast to commercial kits, the findings underscore the successful application of the MPC method in isolating high-quality genomic DNA from feathers characterized by elevated keratin content.

A novel ionic liquid-based approach for DNA and RNA extraction simplifies sample preparation for bacterial diagnostics.

DNA- and RNA-based diagnostics play a pivotal role in accurately detecting and characterizing health-relevant bacteria, offering insights into bacterial presence, viability and treatment efficacy. Herein, we present the development of a novel extraction protocol for both DNA and RNA, designed to enable simple and rapid molecular diagnostics. The extraction method is based on the hydrophilic ionic liquid (IL) 1-ethyl-3-methylimidazolium acetate and silica-coated magnetic beads. First, we developed an IL-based cell lysis protocol for bacteria that operates at room temperature. Subsequently, we established a magnetic bead purification procedure to efficiently and reproducibly extract DNA and RNA from the IL-lysates. The IL not only lyses the cells, but also facilitates the adsorption of nucleic acids (NAs) onto the surface of the magnetic beads, eliminating the need for a chaotropic binding buffer and allowing for purification of NAs without significant effort and materials required. Lastly, we combined the cell lysis step and the purification step and evaluated the novel IL-based extraction method on periopathogenic bacterial cultures, comparing it to commercial DNA and RNA extraction kits via (RT)-qPCR. In comparison to the reference methods, the IL-based extraction protocol yielded similar or superior results. Furthermore, costs are lower, required materials and equipment are minimal and the process is fast (30min), simple and automatable. These characteristics favour the developed method for use in routine and high-throughput testing as well as in point-of-care, on-site and low-resource settings, thereby advancing the field of molecular diagnostics.

Fluorescent primers amplification refractory mutation system qPCR (FP ARMS-qPCR) for MTHFR C677T SNP genotyping.

The currently used methods for the detection of methylene tetrahydrofolic acid reductase (MTHFR) C677T single nucleotide polymorphism (SNP) are either time-consuming or expensive. In this study, we devised an accurate, rapid and easy-to-use SNP detection system based on fluorescent primers amplification refractory mutation system qPCR, known as FP ARMS-qPCR. Fluorescent primers (FPs) modified by fluorescent dye or quencher near the 3' terminal thymine were designed. The reaction conditions were optimized and the performance was evaluated. Using commercial kits as controls, 242 samples were tested in parallel to verify the feasibility of the FP ARMS-qPCR assay. We demonstrated the good sensitivity and specificity of the FP ARMS-qPCR with optimized conditions. The assay was able to accurately distinguish between different SNP sites of MTHFR C677T in less than 2h using as low as 50 pg of template genomic DNA. Completely consistent genotyping results reveal that FP ARMS-qPCR is concordant with commercial kits. We established a specific, sensitive, and rapid FP ARMS-qPCR method for the detection of MTHFR C677T genotype. This could also serve as a potential diagnostic tool for a variety of diseases.

Total RNA Extraction from Rice Vegetative Tissues Using Magnetic Beads.

Ribonucleic acid (RNA) extraction is the first critical step in gene expression analysis. In this chapter, we describe a high-throughput RNA extraction method using guanidine thiocyanate and isopropyl alcohol (HighGI). The use of carboxyl-coated paramagnetic beads, instead of silica membrane columns, enables semi-automation using a liquid handling system and high-throughput RNA extraction for large-scale transcriptome studies. Homemade mixes of paramagnetic beads and buffers make HighGI inexpensive. In addition, HighGI-extracted RNA retains low molecular weight RNA molecules less than 200bp, which is typically lost in commercial column-based kits.

Seroprevalence of Mycobacterium avium subsp. paratuberculosis Infections in Small Ruminants in Europe

Paratuberculosis, also known as Johne’s disease, is a granulomatous enteritis in both domestic and wild ruminants caused by the bacterium Mycobacterium avium subsp. paratuberculosis. Understanding the prevalence of this disease in small ruminants is essential for disease control and prevention strategies. A systematic review of the literature was conducted using the PubMed, ScienceDirect and Scopus databases to identify all articles reporting Mycobacterium avium subsp. paratuberculosis (MAP) seroprevalence in sheep and goats in Europe, published from January 2006 to December 2023. The initial search for existing publications reporting systematic reviews and primary studies was carried out by searching the available databases. For the final selection of studies, an initial screen for basic eligibility and a detailed appraisal of quality were performed. After the study selection, the relevant data was extracted. The detailed appraisal generated 21 publications that reported 55 studies, 22 (40.0%) from sheep (12 at the animal-level and 10 at the flock-level) and 28 (50.9%) from goats (17 at the animal-level and 11 at the flock-level), and 5 (9.1%) from mixed small ruminant species at the animal level. In total, 34 (61.8%) were animal-level studies and 21 (38.2%) were flock-level studies. Population and inclusion criteria were highly variable among studies. Sample sizes ranged from 291 to 15,585 animals. Most studies reported testing adult animals (over 24 months of age). Commercial ELISA kits were used in most studies. The highest prevalence was obtained in sheep (100%) in Türkiye, and the lowest was found also in sheep (0.7%) in Austria. Overall, the results suggest that MAP antibodies have been frequently detected among small ruminants in some countries and there is a need for standardisation of case definitions to improve the accuracy of prevalence estimates. Further research is needed to understand the risk factors associated with MAP infection in small ruminants and to develop effective control and prevention strategies.

Shedding Light on the Molecular Diversities of miRNA in Cancer- an Exquisite Mini Review.

Short non-coding ribonucleic acids are also known as "Micro ribonucleic acids (miRNAs)". The miRNAs make a contribution to the regulation of genes and mitigation of cancer cell growth in humans. miRNAs play a significant role in several BPs, namely apoptosis, cell cycle progression, and development. It is well-recognized that miRNAs are crucial for the tumors' growth and also serve as Tumor Suppressors (TSs) or oncogenes. As miRNAs also act as an effective tumor suppressor, studying the molecular diversities of the miRNAs makes way to minimize cancer progression and the corresponding death rates in the future. Therefore, miRNAs along with their Biological Processes (BPs) and molecular diversities are thoroughly researched in this paper. Consequently, miRNAs particularly target their 3' UnTranslated Region (3'-UTR) for controlling the target mRNAs' stability and protein translation. So, this study also expresses the impact of microRNA variants in various cancer cells, namely Breast cancer, Gastric or stomach cancer, ovarian cancer, and lymphocytic leukemia. Furthermore, the database named PhenomiR and commercial kits that are used in the miRNA data analysis are discussed in this article to provide extensive knowledge about the molecular diversity analysis of miRNA and their influences on cancerous cells.

Watch Out for the Inherent Vulnerabilities in Developing Multi-tenant Cloud-FPGA: Communication Protocols

As FPGAs are being deployed in the cloud infrastructure for acceleration, the technology of multi-tenant FPGA has emerged as a topic of interest. This development has drawn considerable attention to its security issues. While previous research primarily focused on the security of applications, there has been limited exploration of the vulnerabilities inherent in FPGA IPs. In our work, we examine the vulnerabilities of two widely used data transmission protocols in modern FPGAs: the Advanced eXtensible Interface (AXI) and Peripheral Component Interconnect Express (PCIe). Our experiments, conducted with commercial FPGA development kits, launched fault injection attacks through the shared power distribution network (PDN). Through non-invasive electromagnetic (EM) trace measurement, we characterize the voltage fluctuation across various attack patterns. Subsequently, we simulate real-world data transfers using two crafted datasets with different statistical characteristics. The experimental results demonstrate the unique security vulnerabilities of the current AXI and PCIe protocols in the context of a multi-tenant cloud-FPGA. In response to such vulnerability, we further propose two defense strategies: InChAXI that utilizes integrity checking for AXI-based data, and FCPCIe that employs frequency scaling for PCIe-based data. The performance evaluation demonstrates that our proposed defenses can significantly reduce the fault injections on the AXI-based data transmission by 705 times with small overheads – 0.5% in hardware footprint and 7.9% in latency, respectively. On the other hand, FCPCIe effectively prevents the fault injection attack during the PCIe-based data transmission by reducing the user clock frequency, while incurring a 10.13% overhead on data throughput.

Comparison of the Diagnostic Performance of Antigen B Purified from Sheep Hydatid Cyst Fluid (HCF) with Commercial ELISA Kit.

Cystic echinococcosis (CE) is a zoonotic parasitic disease caused by the metacestode of Echinococcus granulosus. CE is a health problem in Middle Eastern countries, such as Iran. The purpose of this study was to purify subunit 8 KDa antigen B from crude sheep hydatid cyst fluid (HCF) and compare its sensitivity and specificity with a commercial human ELISA kit (PT-Hydatid-96). 28 sera samples were collected from hydatid cyst patients who had surgery for a hydatid cyst and had their disease confirmed by pathology after the surgery. Furthermore, 35 samples of healthy individuals with no history of hydatid cysts were collected, as were nine serum samples from parasite-infected non-CE patients. HCF was obtained from sheep fertile cysts at a Sari slaughterhouse and used as an antigen. In an indirect ELISA test, the B antigen was employed, and the results were compared to those from a commercial ELISA kit. The results of this study were analyzed using the Kappa test. The commercial ELISA kit showed 17 cases (23.6%) positive, 44 cases (61.1%) negative, and 11 cases (15.3%) borderline. B antigen showed that 18 (25%), 43 (59.7 %), and 11 (15.3%) were positive, negative, and borderline, respectively. One sample (1.4% of 72 total samples) of 35 serum samples from healthy individuals was positive using B antigen-based ELISA. In addition, all nine serum samples from parasite-infected non-CE patients were negative for both tests. The sensitivity and specificity of the commercial ELISA kit have been evaluated at 60.7% and 100%, respectively. For B antigenbased ELISA, these values are 64.3 and 97.7%, respectively. Antigen B produced from hydatid cyst fluid is a promising option for serological identification of hydatid cysts in both infected and healthy individuals. In an indirect ELISA test, hydatid fluid antigen could be used as a precise source of detection.

Leukocyte Platelet-Rich Plasma-Derived Exosomes Restrained Macrophages Viability and Induced Apoptosis, NO Generation, and M1 Polarization.

Chronic refractory wounds refer to wounds that cannot be repaired timely. Platelet-rich plasma (PRP) has significant potential in chronic wound healing therapy. The exosomes isolated from PRP were proved to exhibit more effectiveness than PRP. However, the therapeutic potential of exosomes from PRP on chronic refractory wounds remained elusive. Hence, this study aimed to clarify the action of exosomes from PRP on chronic refractory wounds by evaluating the response of macrophages to exosomes. Pure platelet-rich plasma (P-PRP) and leukocyte platelet-rich plasma (L-PRP) were prepared from the fasting venous blood of healthy volunteers. Exosomes were extracted from P-PRP and L-PRP using ultracentrifugation and identified by transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA), and western blot. Macrophages were obtained by inducing THP-1 cells with phorbol-12-myristate-13 acetate (PMA). The internalization of exosomes into macrophages was observed utilizing confocal laser scanning microscopy after being labeled with PKH67. Cell viability was determined by CCK-8 assay. Cell apoptosis was measured utilizing a flow cytometer. The polarization status of M1 and M2 macrophages were evaluated by detecting their markers. Nitric oxide (NO) detection was conducted using the commercial kit. Exosomes from P-PRP and L-PRP were absorbed by macrophages. Exosomes from L-PRP restrained viability and induced apoptosis of macrophages. Besides, exosomes from P-PRP promoted M2 polarization, and exosomes from L-PRP promoted M1 polarization. Furthermore, exosomes from L-PRP promoted NO generation of macrophages. Exosomes from L-PRP restrained viability, induced apoptosis and NO generation of macrophages, and promoted M1 polarization, while exosomes from P-PRP increased M2 polarization. The exosomes from L-PRP presented a more effective effect on macrophages than that from P-PRP, making it a promising strategy for chronic refractory wound management.

Beta-elemene alleviates cigarette smoke-triggered inflammation, apoptosis, and oxidative stress in human bronchial epithelial cells, and refrains the PI3K/AKT/mTOR signaling pathway.

Chronic obstructive pulmonary disease (COPD) is a grievous disease that adversely affects human health and life. β-elemene is a type of sesquiterpenoid extracted from Curcuma wenyujin (Zingiberaceae) and displays effects on suppressing tumor growth. However, the regulatory impact of β-elemene in COPD development is not reported. This study explored the functioning of β-elemene in the progression of COPD. The cell survival rate was confirmed through Cell Counting Kit-8 (CCK-8) assay. The cell apoptosis was evaluated through flow cytometry. The protein expressions were examined through western blot. The levels of malondialdehyde (MDA), superoxide dismutase (SOD) and reactive oxygen species (ROS) were examined through the corresponding commercial kits. The levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and IL-1β were inspected through Enzyme-Linked Immunosorbent Assay (ELISA). The study demonstrated that β-elemene exaggerated cell viability and reduced cell apoptosis in BEAS-2B human bronchial epithelial cell line stimulated by cigarette smoke extract (CSE). Oxidative stress was heightened after 5% CSE induction, but this impact was counteracted by β-elemene treatment. In addition, enhancive inflammation induced by cigarette smoke was attenuated by β-elemene treatment. Finally, our results indicated that the triggered the phosphatidylinositol 3-kinase-protein kinase B-mechanistic target of rapamycin (PI3K/AKT/mTOR) pathway mediated by cigarette smoke was refrained by β-elemene treatment. It was concluded that β-elemene reduced cigarette smoke-triggered inflammation, apoptosis, and oxidative stress in human bronchial epithelial cell line, and refrained PI3K/AKT/mTOR signaling pathway. This study proposed that β-elemene could act as a hopeful drug for the treatment of COPD.

An antibody nanopore-enabled microsensor for detection of osteoprotegerin

This paper introduces an antibody-based nanopore thin film sensor for detecting osteoprotegerin (OPG) in buffer solutions and serum samples, offering significant improvements over current commercial enzyme-linked immunosorbent assay kits in terms of cost, specificity, and selectivity. Capable of detecting OPG concentrations as low as 0.25 pg ml−1—far below the limits of traditional lab equipment—this sensor requires only about 1 μl of serum for effective measurement. The utilization of reference sensors helps reduce non-specific binding, enhancing the sensor’s accuracy. Its affordability and operational simplicity make it ideal for point-of-care monitoring of OPG levels in real-time patient care.

Enhanced detection of Opisthorchis viverrini infection: A comparative evaluation of modified one-step FECT and conventional diagnostic methods in low-intensity setting

The formalin-ethyl acetate concentration technique (FECT) is one of the most sensitive diagnostic method not only for all helminths, but also for Opisthorchis viverrini infections in stool examinations. However, it remains a diagnostic problem for light infections. We modified the one-step FECT to determine the low-intensity of O. viverrini infection and compared with various conventional detection methods. The study utilized 160 egg-positive and 160 randomly negative stool samples for O. viverrini eggs by conventional FECT (cFECT) to compare the methods, including the simple smear, the Kato-Katz method, the two commercial stool examination kits, and the one-step FECT. Our results showed that the one-step FECT method had the highest sensitivity (95.6 %), followed by cFECT (87.9 %), the Kato-Katz (55.5 %), Aquisfek SF-FIX (48.3 %), simple smear (42.3 %), and Mini Parasep® SF (35.1 %). The ability of one-step FECT exhibited better ability to detect low parasite intensities compared to the cFECT (18 eggs per gram (e.p.g.) versus 34 e.p.g.) and the other conventional diagnostic methods. In addition, the investigation of O. viverrini infection in endemic regions in northeastern Thailand based on 3900 fecal samples revealed that the one-step FECT with an intensity of 66.8 e.p.g. (range 18–226) was significantly higher in sensitivity than cFECT, which had an intensity of 58.0 e.p.g. (range 34–214). Interestingly, fecal samples with less than 50 e.p.g. could not be detected by cFECT in 67 % of cases, and 69 out of 3900 samples were negative. In conclusion, one-step FECT improves the detection of low-intensity O. viverrini infection, which is suitable for parasites screening, especially for low-intensity infections in the community.

Prevalence and Subtypes Distribution (ST10, ST14, ST25, ST26) of Blastocystis spp. in Anatolian Water Buffalo (Bubalus bubalis) in Van, Türkiye.

Blastocystis spp. is one of the most common protozoa worldwide, living in the gastrointestinal tract of humans and many other animals. On the basis of the genetic heterogeneity of small subunit ribosomal RNA, at least 28 subtypes (ST1-ST17, ST21 and ST23-ST32) are reported to exist in mammals and birds. This study was carried out to determine the prevalence and subtypes of Blastocystis spp. in Anatolian buffaloes (Bubalus bubalis) in Van province in the Eastern Anatolia Region of Turkey. DNA was extracted using commercial GeneMATRIX Stool DNA Purification Kit and then stored at -20°C until PCR amplification. After PCR amplification of the SSU rRNA gene region positive Blastocystis spp., amplicons from buffalo faeces were sequenced and then deposited in GenBank (OR576949.1, OR576950.1, OR576970.1, OR576971.1, OR577019.1, PP837943.1, PP837940.1, PP837939.1, PP837604.1, PP837937.1, PP837934.1, PP837601.1, PP837936.1 and PP837603.1). PCR analysis of 120 faecal samples showed a total prevalence of 11.67% (14/120). The prevalence was higher in females and young animals (p>0.05). Sequence analysis revealed Blastocystis spp., ST10, ST14, ST25 and ST26 subtypes. To our knowledge, Blastocystis subtypes ST25 and ST26 in buffaloes were reported for the first time in this study. It is thought that more large-scale studies should be carried out to determine the zoonotic subtype potential of this protozoan in the region.

Seroprevalence and associated risk factors for Fasciola hepatica in sheep in Nile Delta of Egypt

Fascioliasis is a globally distributed zoonotic parasitic disease that affects ruminants, including sheep. This study conducted from January to December 2023, aimed to determine the prevalence of Fasciola hepatica in sheep across three governorates in Egypt's Nile Delta and to assess associated risk factors. A total of 455 serum samples were analyzed using a commercial ELISA kit, revealing antibodies against F. hepatica in 22.2 % of the tested sheep. There was no significant association between locality or sex and the seroprevalence of F. hepatica in sheep; however, the highest prevalence was observed in Kafr Sheikh and in female sheep.Concerning risk factors, poor conditioned sheep aged between 1 and 2 years showed 2.1 and 3.8 times higher of infection probability than others. In addition, the risk of F. hepatica infection in sheep increased significantly in winter season (OR = 6.6, 95 %CI: 2.6–16.8), in absence of prophylactic treatment (OR = 2.2, 95 %CI: 1.3–3.3) and in presence of snail (OR = 3, 95 %CI: 3–5.4). The existence of antibodies against F. hepatica in examined sheep raising in Nile Delta indicating that the disease is reported in the studied areas and needs to be managed on farms through control and preventative measures.