Gel Electrophoresis Research Articles

Recurrent Outbreak of Carbapenem-Resistant IMP-1-Producing Pseudomonas aeruginosa in Kidney Transplant Recipients: The Impact of Prolonged Patient Colonization.

Infections by carbapenem-resistant Pseudomonas aeruginosa (CRPA) have been associated with high morbidity and mortality among solid organ recipients. To delineate the epidemiological and molecular characteristics of a recurrent outbreak of imipenem (IMP)-producing P. aeruginosa (CRPA) among kidney transplant (KT) recipient METHODS: We described a recurring CRPA outbreak in a KT ward, divided into two periods: before unit closure (Feb 2019-2020) and after reopening (Aug 2020-Dec 2023). Routine surveillance cultures (SCs) were performed using axillary-perineum-rectal swabs with immunochromatographic tests. A case-control study identified risk factors for CRPA acquisition. Pulsed-field gel electrophoresis and whole genome sequencing characterized the strains. After reopening, new cases arose from patients previously colonized, peaking 18 months later. A total of 67 KT recipients with CRPA-IMP-producing strains were identified. All except one sequenced strain belonged to the ST446 clone, differing by a maximum of 110 single nucleotide polymorphisms. Forty-five (67.2%) cases were identified through SC, with 45.7% showing intermittent SC positivity. Patients remained colonized for up to 623 days. Twenty-four (35.8%) patients had infections, with the most common site being the urinary tract. Identified risk factors included older age, deceased donor, re-transplantation, reoperation, carbapenem or quinolone use, lymphopenia, hospital stay >10 days, and the first 60 days post-KT. KT recipients can harbor CRPA for extended periods, and detecting CRPA-colonized patients is challenging. These characteristics highlight the patient as the major source and a critical point in outbreak control.

Phosphate and two-stage cooking: Impact on lipid oxidation and tenderness in sous-vide beef semitendinosus.

The study investigated phosphate's effects on tenderness and lipid oxidation in beef cooked sous-vide. Semitendinosus beef cuts were treated with 0.15% and 0.3% sodium tripolyphosphate (STPP), and plain water (0% STPP). These cuts were then sous-vide cooked at two different temperatures, 60 °C and 70 °C, for two different durations, 3 h and 6 h. Analytical techniques including Warner-Bratzler shear force (WBSF), pH, thiobarbituric acid reactive substances (TBARS), sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), scanning electron microscopy (SEM), and Fourier-transform infrared spectroscopy (FTIR) were employed. STPP concentration did not yield a statistically significant impact on the pH and FTIR spectrum of sous-vide beef (p > 0.05). However, STPP markedly enhanced tenderness by loosening the myofibril structure (observed in SEM) and weakening myosin and actin bands (observed in SDS-PAGE). This improvement led to lower WBSF values with 0.3% STPP, particularly noticeable at 60 °C for 6 h. Additionally, 0.3% STPP delayed lipid oxidation, evidenced by lower oxidation values after 4 weeks of refrigerated storage at 4 °C. These findings highlight STPP's potential to improve tenderness and lipid oxidation stability in sous-vide beef.

Study on Internal Standards Applicable to the Detection of Recombinant Erythropoietin.

With the discovery of the variant c.577del in the EPO gene, the procedure for detecting the presence of recombinant erythropoietin (rEPO) has become complicated and time-consuming. To address this situation, an rEPO confirmation method that uses reverse-normal immunopurification coupled with western blotting (WB) and sodium N-lauroylsarcosinate polyacrylamide gel electrophoresis (SAR-PAGE), which can detect the EPO variant (VAR-EPO) and rEPO with anti-VAR-EPO and anti-EPO antibodies, has been developed. Therefore, it is necessary to develop an internal standard (IS) that can be recognized by an anti-VAR-EPO antibody to monitor reverse immunopurification, ensuring the reliability and accuracy of the analysis. In this study, we constructed an IS based on VAR-EPO modified with polyethylene glycol (PEG) and then assessed its reliability and applicability in doping analysis. The data provided here show that the obtained PEGylated VAR-EPO can be used as an IS to monitor the detection of not only rEPO and VAR-EPO but also EPO receptor agonists in urine samples.

The low enoyl-acyl carrier protein reductase activity of FabI2 is responsible for the high unsaturated fatty acid composition in Sinorhizobium meliloti

BackgroundSinorhizobium meliloti is noted for its exceptional capacity to produce unsaturated fatty acids (UFAs). Earlier studies have indicated that S. meliloti primarily employs the FabA-FabB pathway for UFA synthesis, however, the mechanisms remain elusive. This study was conducted to elucidate these mechanisms responsible for the significant UFA production in S. meliloti.MethodsThe genes encoding enoyl-acyl carrier protein (ACP) reductase (ENR) were disrupted using the suicide plasmid pK18mobsacB, followed by the creation of single-crossover and double-crossover mutants. The ENR proteins were expressed in Escherichia coli BL21(DE3) strains and subsequently purified. Their enzymatic activities were assessed through gel electrophoresis and NADH oxidation assays. Additionally, the fatty acid composition was determined using gas chromatography-mass spectrometry (GC-MS) and thin-layer chromatography.ResultsOur findings demonstrate that the heterologous expression of fabI2 in a temperature-sensitive E. coli fabI mutant results in a significant enhancement of UFA production. Genetic analyses confirmed that fabI2 is an indispensable gene in the S. meliloti genome, as it cannot be disrupted. Interestingly, we observed that fabI2 could only be functionally replaced by the Enterococcus faecalis fabI gene and not by the homologous fabI1 from S. meliloti, E. coli fabI, or Pseudomonas aeruginosa fabV. Furthermore, we validated that the deletion of fabI1 in S. meliloti triggered an increase in UFA production compared to the wild-type strain Rm1021.ConclusionsIn this study, we identified the ENR, encoded by the S. meliloti SMc00326 gene (fabI2), as playing a pivotal role in the biosynthesis of UFAs. Additionally, the FabI1 enzyme, encoded by SMc00005, was found to modulate the fatty acid composition within S. meliloti. Together, these discoveries establish a foundation for the development of a model that explains the significant contribution of FabI2 to the robust synthesis of UFAs in S. meliloti.

Diagnostic potential of rhinovirus C using reverse-transcription loop-mediated isothermal amplification (RT-LAMP).

Rhinoviruses (RVs), particularly RV-C, frequently cause acute respiratory infections and asthma exacerbations. However, there is a lack of routine detection methods. Thus, this study aims to develop a rapid molecular and differential diagnostic detection method for RV-C using the reverse transcription (RT) loop-mediated isothermal amplification (LAMP) approach. The RT-LAMP assay targeting the 5'UTR region of RV-C genome was optimized by varying the reaction temperature, magnesium sulphate, betaine concentrations, and reaction time. Compared with conventional RT-PCR with a sensitivity of 106 copies of RNA, RT-LAMP demonstrated a significant increase in efficiency and sensitivity with a quantifiable viral load of at least 101 copies of RNA by gel electrophoresis and colour change, and 104 copies of RNA for end-point detection with a turbidimeter for 40min. The assay is also specific without amplifying RV-A16 and RV-B72 genomic RNA. In the proof-of-concept assay using 30 clinical respiratory samples with known etiological agents, it detected all RV-C isolates, of which its accuracy was confirmed by sequencing. The newly developed RT-LAMP assay demonstrated good analytical sensitivity and specificity toward RV-C. The assay provides an alternative for improved RV-C diagnosis.

Dominance of dengue virus serotype-2 in Pakistan (2023-2024): Molecular characterization of the envelope gene and exploration of antiviral targets.

Dengue virus infection, caused by a single positive-stranded RNA virus from the Flaviviridae family, represents a significant public health challenge in tropical and subtropical regions. This virus has four serotypes (DENV-1, 2, 3, and 4), primarily transmitted by Aedes mosquitoes. Despite extensive research, effective antiviral treatments and vaccines remain elusive due to the viral diversity and the complex mechanisms such as antibody-dependent enhancement (ADE). In the current study, NS1-positive serum samples from dengue cases in Pakistan (2023-2024), were analyzed to determine the predominant serotype and characterize the envelope (E) gene for further exploration of antiviral targets. Out of 100 samples, 63 (63%) tested positive for DENV-2, indicating its predominance during this period, while two samples showed mixed infections with DENV-2 and DENV-3. The envelope gene was successfully amplified using nested PCR, validated through gel electrophoresis and sanger sequencing. Phylogenetic analysis revealed high similarity of the DENV-2 isolates to strains from China and India. Computational modeling of the envelope protein structure identified potential antiviral binding sites and further molecular docking studies suggested that specific antiviral compounds like Arbidol and Quercetin can inhibit early steps in viral infection. Additionally, BepiPred-3.0 predicted several B-cell epitopes, which could be useful for vaccine development. These findings enhance our understanding of dengue epidemiology in Pakistan and contribute to the development of targeted antiviral therapies, potentially informing future vaccination strategies and outbreak management.

ROS responsive nanozyme loaded with STING silencing for the treatment of sepsis-induced acute lung injury

Acute lung injury (ALI) is a common complication of sepsis and a leading cause of mortality in septic patients. Studies indicate that STING may play a crucial role in the pathogenesis of sepsis-induced ALI by interacting with the PARP-1/NLRP3 pathway. Therefore, targeting STING inhibition has potential as a novel therapeutic strategy for ALI. However, effective inhibition remains challenging due to the widespread expression of STING across various tissues. In this study, we developed a nanozyme-based drug delivery system, DSPE-TK-mPEG-MnO2@siSTING (abbreviated as DTmM@siSTING), using DSPE-TK-mPEG-MnO2 as the carrier, and characterized it via scanning electron microscopy, dynamic light scattering, nanoparticle size analysis, and gel electrophoresis. To evaluate the therapeutic effects of DTmM@siSTING, an in vitro ALI cell model and an in vivo ALI mouse model were established, assessing the nanozyme's impact on ROS levels, inflammatory responses, and the PARP-1/NLRP3 pathway in sepsis-induced ALI. Results demonstrated that DTmM@siSTING exhibited good physiological stability. In vitro, DTmM@siSTING significantly reduced ROS levels, myeloperoxidase activity, and expression of inflammatory cytokines, while also inhibiting PARP-1/NLRP3 pathway activation. In vivo experiments further revealed that DTmM@siSTING effectively delivered siSTING to the lungs, mitigating sepsis-induced ALI and associated inflammatory responses. Additionally, DTmM@siSTING displayed excellent biocompatibility. In summary, our findings suggest that DTmM@siSTING significantly enhances the therapeutic efficacy of siSTING, alleviating ALI by inhibiting ROS production, inflammatory responses, and activation of the PARP-1/NLRP3 pathway. This novel approach presents a promising therapeutic avenue for sepsis-induced ALI.

Synthesis of nanocellulose from Acalypha hispida leaves through enzymatic hydrolysis

The enzyme-hydrolysis of Acalypha hispida plant leaves for the synthesis of nanocellulose remains underexplored, yet it holds significant potential due to its biocompatibility, eco-friendliness, and the exceptional physical and biochemical properties. This innovative method employed the enzyme xylanase, produced by an indigenous strain of Bacillus pumilus, isolated from a compost pile sample and identified through 16S rRNA sequencing. The xylanolytic activity of the isolate was confirmed by the clear hydrolysis zones around bacterial colonies on xylan agar. The enzyme was purified using ammonium sulfate precipitation followed by dialysis, with a molecular weight of 45 kDa determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The purified xylanase was then utilized for enzyme-hydrolyzed nanocellulose (EHNC) synthesis. EHNC stands out for its green processing, biocompatibility, biodegradability, and low production cost. The resulting EHNC exhibited particle diameters ranging from 15 to 40 nm, with a zeta potential of −22.3 mV. The minimal enzyme concentrations and short incubation periods suggest significant potential for enzymatic applications in the production of nanofibrillated cellulose, offering an efficient and sustainable approach to manufacturing advanced nanocellulose-based materials.

Different polyphenols on cold plasma-induced covalent ovalbumin-polyphenols conjugates and affecting the antigenicity of OVA and its functionalities

The study investigated cold plasma (CP) treatment to modulate the antigenicity and functionality of ovalbumin (OVA) through CP-induced covalent conjugation with five structurally distinct dietary polyphenols, gallic acid (GA), syringic acid (SA), (-)-epigallocatechin gallate (EGCG), (-)-epicatechin gallate (EGC), and (-)-epicatechin (EC). Results indicated that a maximum polyphenol content of 176.63 ± 1.83 μmol/g in OVA-EGCG conjugates was obtained following CP treatment (60 s). The generation of covalent OVA-polyphenol conjugates was confirmed through sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis. The binding sites of the five polyphenols in OVA were identified using LC-MS/MS. Different regions and quantities of binding sites were observed for the various polyphenols. Seventeen binding sites were detected in OVA-EC conjugates, whereas only one site (His371) of EGC was found in OVA-EGC conjugates. Additionally, a maximum reduction in IgE binding capacity (63.13 ± 0.87%) of OVA was achieved after GA grafting, supported by western blotting (WB) and enzyme-linked immunosorbent assay (ELISA) analyses. The masking and steric-hindrance effect on linear epitopes by polyphenol grafting play key roles in reducing OVA’s antigenicity. Moreover, the emulsifying properties and antioxidant activity of OVA were substantially improved following polyphenol grafting. Correlation analysis revealed that the hydroxyl group (-OH) content in polyphenol molecules was the critical factor determining covalent binding efficiency between OVA and polyphenols, consequently resulting in antigenicity elimination and functionality improvement of OVA. This study demonstrates that CP-induced covalent grafting of polyphenols is an effective strategy for eliminating antigenicity and modulating the functionalities of proteins.

Effect of pre-acidification induction on the physicochemical features, myofibrillar protein microstructure, and headspace volatiles of ready-to-cook goose meat

This study examined the impact of pre-acidification induction on the quality attributes and flavor retention of ready-to-cook (RTC) goose meat products. The results demonstrated that pre-acidification could influence the eating qualities of RTC goose meat by effectively regulating the physicochemical properties of goose myofibrillar proteins (MP) including solubility and water-holding capacity. Elevated carbonyl content indicated an enhanced gel-forming capacity in RTC goose meat during storage, coupled with reduced total sulfhydryl content from enhanced protonation pretreatment and augmented lipid oxidation. Structural characterization of MP via sodium dodecyl sulfate–polyacrylamide gel electrophoresis, circular dichroism spectroscopy, and intrinsic fluorescence revealed the formation of a dense protein matrix under highly acidic conditions. Furthermore, the headspace concentration of aldehydes increased by 3.23 times upon enhancing the pre-acidification intensity, resulting in the production of esters and acidic flavor compounds with favorable aromas. Correlation analysis demonstrated the dependence of headspace concentrations of volatile constituents on the acidification-enhanced surface hydrophobicity of MP, attributed to the modified binding sites of proteins after pre-acidification. Current results have indicated both the positive and negative influence of pre-acidulation induction on the eating quality of goose meat products, suggesting the necessity of introducing extra processes to modulate the quality of prefabricated products.

Artificial trimerization of β-glucosidase for enhanced thermostability and activity via computational redesign

The improvement of enzyme thermostability is of great significance in the biotechnology industry. Herein, we modified the C-terminus of β-glucosidase via the computer-aided rational design and successfully obtained six variants that exhibit improved optimal temperatures and thermal inactivation half-lives at 65 °C. Among these variants, the thermal inactivation half-life and hydrolytic activity of TrCel1b-H13 at 65 °C were increased by 416 and 3223 folds, respectively, compared to those of the wild type. The optimal catalytic temperature and melting temperature (Tm) of TrCel1b-H13 were elevated by 55 °C and 20 °C respectively, compared to those of the wild type. The Kcat/Km value of TrCel1b-H13 was improved by 13.3-fold, compared to that of the wild type. Furthermore, we found that the modified β-glucosidase, TrCel1b-H13, self-assembles into a trimeric structure, which was confirmed by X-ray crystallography and blue native polyacrylamide gel electrophoresis. The amino acid residues, including E28, H60, E99, K106, and K471, located at the trimeric interface of the adjacent subunits, were found to play an important role in maintaining the crystal structure of the protein trimer. Thus, protein multimerization via the computer-aided rational design method provides a cost-effective tool for simultaneously improving the thermostability and activity of enzymes.

Molecular characterization of ST15 carbapenem-resistant Klebsiella pneumoniae isolated in a single patient

The carbapenem-resistant K. pneumoniae (CRKP) poses a serious threat to antibiotic applicability and public health. During treatment, K. pneumoniae (KP) frequently exhibits shifts in drug-resistant phenotypes, complicating clinical treatment as it transitions from sensitivity to resistance. In this study, we analyzed the clinical and molecular characteristics of drug resistance changes in KP strains isolated from a single patient, and the potential mechanisms underlying these resistance changes. Antimicrobial susceptibility test and string test were conducted to evaluate the resistant and virulent characterization of the strains. Pulsed field gel electrophoresis (PFGE) was used to investigate the homology relationship between the strains. The whole genome sequencing and phylogenetic analysis of 9 representative isolates was also performed. The transfer ability of the drug-resistant plasmid was studied by plasmid conjugation experiment. The transconjugants were verified by PCR amplification of specific genes, antimicrobial susceptibility test and PFGE. Our results revealed that 9 KP strains, isolated from the same patient, exhibited 'resistance-sensitivity-resistance-sensitivity' alternately to carbapenems. The differences in DNA fingerprint bands among the nine KP isolates were ≤ 3, which can be classified as the same PFGE type. Phylogenetic analysis showed that these 9 strains constituted a distinct branch within the phylogenetic tree. All nine KP strains belonged to the ST15-KL19 clone. Six of the strains were classified as CRKP, all of which carried eleven drug resistance genes: oqxB, oqxA, fosA6, aac(3)-lld, blaSHV-28, blaKPC-2, blaOXA-1, mph(A), tet(A), catB3 and aac(6')-lb-cr, mediating drug resistance to quinolones, fosfomycin, aminoglycosides, β-lactam, carbapenems, macrolides and chloramphenicol, belonging to multi-drug resistant bacteria. The carbapenem-resistant plasmid p2-KP3762-1 was found to transfer within species, from CRKP to hypervirulent K. pneumoniae (hvKP) RJF293HA, carbapenem-sensitive K. pneumoniae (CSKP) KP3657 and E. coli C600 at a frequency of (1.19±1.58) ×10-6, (1.09±1.38) ×10-7 and (10.9±9.53) ×10-6 respectively, resulting in the dissemination of carbapenem resistance genes. In this study, KP strains isolated from a single patient exhibited an alternating phenotype of resistant-sensitive-resistant-sensitive to carbapenems. The 9 KP isolates share a high degree of genetic similarity. The plasmid p2-KP3762-1, harboring the carbapenem resistance gene blaKPC-2, may undergo inter-strain and inter-clone transfer via conjugation in the patient during treatment. Furthermore, our findings suggest that the pathogens in this patient are likely to have a common ancestral origin.

Menaquinone-7 and its therapeutic potential in type 2 diabetes mellitus based on a Zucker diabetic fatty rat model

BackgroundType 2 diabetes mellitus (T2DM) is marked by insulin resistance, low grade chronic inflammation, and endothelial dysfunction. Vitamin K2, especially menaquinone-7 (MK-7), might delay T2DM progression and alleviate its consequences. Hence, this study evaluated the effects of MK-7 on serum and urine markers of diabetes in an animal model of T2DM. MethodsHetero- (fa/+, control) and homozygous (fa/fa, diabetic) male Zucker diabetic fatty (ZDF) rats were supplemented or not with MK-7 for 12 weeks. After euthanasia, vitamin K1, menaquinone-4 and MK-7 serum concentrations were analyzed via reversed phase high pressure liquid chromatography. Glucose (serum), fructosamine (serum) and creatinine (serum and urine) levels were assessed photometrically, serum cystatin C and urinary total protein were turbidimetrically determined. Serum transforming growth factor beta1 (TGF-β1) and procollagen type III N-terminal peptide (PIIINP) were quantified with enzyme-linked immunosorbent assay. Urinary marker proteins were analyzed via sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Nephropathy was assessed histologically. ResultsSupplementation led to significantly elevated MK-7 serum levels and a significant reduction of PIIINP serum levels in both hetero- and homozygous ZDF rats. Additionally, not statistically significant reductions of TGF-β1 serum levels, proteinuria as well as the nephropathy score were observed. In vivo body mass, serum fructosamine, glucose, cystatin C and creatinine levels were unaffected. ConclusionMK-7 reduced serum markers of fibrosis, histological features of nephropathy and urinary protein excretion, but failed to affect serum markers of T2DM. The therapeutic potential of MK-7 in T2DM and its mode of action should be further investigated in more detail.

Evaluation of a commercial Immunochromatographic Strip Assay (ICT) for rapid detection of bovine, porcine and human Rotavirus A

The most frequent diarrhoeal pathogen “Rotavirus” is commonly detected by Ribo Nucleic Acid- Polyacrylamide Gel Electrophoresis (RNA-PAGE), Reverse Transcription Polymerase Chain Reaction (RT-PCR) and Immunoassay (Enzyme Linked Immunosorbent Assay) in humans. However, use of these tests is limited in animals therefore, penside test like Immunochromatographic test/Lateral Flow Assay (ICT/LFA) is necessary. The present study was carried out to compare the diagnostic efficacy of rapid commercial ICT strip assay, RNA-PAGE and RT-PCR for specific detection of Rotavirus A (RVA) from stool samples of calves, piglets and children. A total of 313 faecal samples were collected between November 2022 to April 2023 from children below 5years of age (n=100), calves (n=100) and piglets (n=113) ≤ 3 months of age and were screened by RNA-PAGE, RT-PCR and commercial ICT kit for rotavirus A.The overall positivity of RVA from human, calves and piglets using RNA-PAGE, RT-PCR and LFA was found to be 35% (35/100), 8% (8/100) and 14.16% (16/113), respectively. Higher positivity of rotavirus was recorded in male children (44.26%, 27/61) than in female children (20.51%, 8/39). The kappa agreement between LFA and RT-PCR was 0.79 (substantial agreement); between LFA and RNA-PAGE was 0.61 (substantially low agreement) and between RNA-PAGE and RT-PCR was found to be 0.75 (substantial agreement). Relative diagnostic sensitivity and specificity of LFA in comparison with RT-PCR was 74.54% and 98.44%, respectively and in comparison with RNA-PAGE was 72.97% and 93.47%, respectively. RNA-PAGE in comparison with RT-PCR is having diagnostic sensitivity and specificity of 65.45% and 99.6%, respectively. In conclusion, the ICT strip assay is a rapid, convenient and effective method with satisfactory efficacy for detection of RVA from children, calves and piglets. ICT fulfilled the WHO ASSURED (Affordable, Sensitive, Specific, User-friendly, Rapid and robust, Equipment-free and Delivered) criteria for point-of-care testing. It can be useful in determining rotavirus outbreaks in resource-limited settings.

Variation in Lipoprotein(a) Response to Potent Lipid Lowering: The Role of Apolipoprotein (a) Isoform size

BackgroundLipoprotein(a) [Lp(a)] is a driver of residual cardiovascular risk. Proprotein Convertase Subtilisin/Kexin Type 9 inhibitors (PCSK9i) decrease Lp(a) with significant heterogeneity in response. We investigated contributors to the heterogeneous response. MethodsCHOlesterol Reduction and Residual Risk in Diabetes (CHORD) was a prospective study examining lipid lowering in participants with an LDL-C >100 mg/dl with and without diabetes (DM) on lipid lowering therapy (LLT) for 30-days with evolocumab 140 mg every 14 days combined with either atorvastatin 80 mg or ezetimibe 10 mg daily. Lp(a) level was measured by immunoturbidometry, and the apolipoprotein a [apo(a)] isoform size was measured by denaturing agarose gel electrophoresis and western blotting. We examined the change in Lp(a) levels from baseline to 30 days. ResultsAmong 150 participants (mean age 50 years, 58% female, 50% non-White, 17% Hispanic, 50% diabetes), median (IQR) Lp(a) was 27.5 (8-75) mg/dL at baseline and 23 (3-68) mg/dL at 30 days, leading to a 10% (0-36) median reduction (P<0.001). Among 73 (49%) participants with Lp(a) ≥30 mg/dL at baseline, there was a 15% (3-25) median reduction in Lp(a) (P<0.001). While baseline Lp(a) level was not correlated with change in Lp(a) (r=0.04, P=0.59), apo(a) size directly correlated with Lp(a) reduction (P<0.001). After adjustment for age, sex, race/ethnicity, DM, and type of LLT, apo(a) size remained positively associated with a reduction in Lp(a) (Beta 0.95, 95%CI, 0.93-0.97, p<0.001). ConclusionOur data demonstrate variation in Lp(a) reduction with potent LLT. Change in Lp(a) was strongly associated with apo(a) isoform size.

Real-time monitoring of cattle (Bos taurus) tissue using a novel point-of-care (POC) polymerase spiral reaction (PSR) colorimetric assay

Adulteration of cattle meat with other species raises critical public health concerns, as existing detection methods lack sensitivity, specificity and practicality. We address this gap by introducing a novel Point-of-Care (POC) polymerase spiral reaction (PSR) assay. This rapid, on-site tool enables highly specific and sensitive identification of cattle tissue, even in resource-limited settings. This innovation paves the way for improved food safety and consumer protection. The cattle-specific PSR assay employs COX3 gene-targeting primers and a chromogenic indicator triad (HNB, Malachite Green and Phenol Red) for rapid visual detection. Cross-amplification absence with other meats (buffalo, sheep, goat and pig) and concordance with SYBR Green I and agarose gel electrophoresis confirm specificity. PSR at 62 °C for 60minutes exhibits a 100 fg detection limit and identifies 0.1% meat admixing. Efficacy is demonstrated across a diverse range of cattle meat samples, blind samples and processed cattle muscle samples. Compatibility with both conventional (snap chill) and commercial DNA extraction kits enables on-site analysis within 90minutes, facilitating rapid field sample analysis.

Characterization of Orexin Gene in Tilapia (Oreochromis niloticus):Regulator Feeding Appetite, and Correlation with Reproductive Factors

The mechanism of energy allocation related to increased appetite and feed intake in fish involves several neuropeptides encoded by specific genes. Orexin (OX) is a peptide secreted in the pars tuberalis of the hypophysis that acts as a stimulator in appetite increase (orexigenic factor). However, the expression of different orexigenic factors varies among vertebrate species, reflecting their unique types and lineages. Therefore, this study aimed to identify genes encoding appetite in Tilapia. RNA isolation, complementary Deoxyribose Nucleic Acid (cDNA) cloning, and DNA amplification were performed from brain samples of gonadally mature Tilapia. The PCR products were subsequently sent to Macrogen.Inc for sequencing. The amplification results of Orexin with β-Actin (positive control) observed using agarose gel electrophoresis showed that the size of the nucleotide base product of each gene was 196 bp and 197 bp. Confirmation of sequencing results carried out using the Basic Local Alignment Search Tool (BLAST) method - National Center for Biotechnology Information (NCBI) for Orexin and β-Actin were 97% and 100%. Based on these results, it can be concluded that each target gene isolated from the tilapia brain showed homology/similarity with the sequence available in the NCBI database

Determination of antimicrobial activities and DNA protection properties of Scytosiphon lomentaria extracts

Scytosiphon lomentaria is a brown algae that has many promising biological activities in medicine as well as being consumed as food in many countries. This study aims to investigate the antimicrobial, antimutagenic and DNA damage protective activity of S. lomentaria by obtaining hexane, methanol and water extracts. To evaluate the antimicrobial effect of the extracts, minimum inhibitory concentration (MIC) tests were performed on both yeasts and bacteria. The antimutagenic activities of the extracts were investigated by the Ames test, and their protective effects against oxidative DNA damage were investigated by agarose gel electrophoresis using pBR322 plasmid DNA. Antimicrobial results showed that hexane methanol and aqueous extract were more effective on Candia albicans (5, 2.5 and 2.5 mg/mL, respectively) compared to all strains used. All concentrations of the methanol extract exhibited strong antimutagenicity (more than 40% inhibition) on both TA98 and TA100 mutant strains. It was determined that the extracts protected DNA against oxidative DNA damage. The results show that S. lomentaria extracts have antimicrobial activity and DNA protection properties, making them have the potential to be evaluated as new natural resources that can be used in antimicrobial and anticancer formulations

How much do we know about nanobacteria?

Background/PurposeNanobacteria, known to date as self-replicating, nano-scale size organisms, smaller than bacteria. However, whether these are living organisms or agglomeration of biomolecules was one of the most controversial issues for many years. One of the reasons for debate is their lack of any genetic material. Although many studies investigate whether nanobacteria are the cause of diseases that affect human health, the discussions did not reach a conclusion. While most studies try to unravel the structures of nanobacteria, some studies investigated their effects on diseases. It has been proposed that pathological calcifications in kidney and dental pulp stones, as well as calcifications in heart valves and arteries, might be caused by nanobacteria. In this study, we investigated the effects and molecular structures of nanobacteria on the formation of atheroma plaques that cause atherosclerosis. MethodsNanobacteria grown from atheroma plaques in cell culture medium were analysed by scanning electron microscope (SEM) and energy dispersive X-ray spectroscopy (EDS). Macromolecules of nanobacteria were separated by SDS-polyacrylamide gel electrophoresis and their sensitivity to protease and nucleases was tested. After determining their resistance to nucleases and susceptibility to proteinase K, protein profiles were determined by quadrupole time-of-flight mass spectrometry (QTOF-MS). ResultsIn SEM, spherical structures with sizes ranging from 80 to 900 nm were visualized. EDS analyses showed that very low concentration of phosphorus (P: 0.3 %) indicated absence of nucleic acids. The resistance was observed to proteases. Also, protein profiles indicated human proteins, with a high percentage of albumin. ConclusionIt was determined that nanobacteria derived from atheroma plaque were structures composed of human proteins. The findings confirmed the theory that nanobacteria are non-living structures formed by a mechanism called biocrystallization. Although the accumulation of nanobacteria may be a contributing element to the creation of atheroma plaque, it is possible that the etiological components that initiate the formation of the plaque are not nanobacteria.

Mastication of Carrots with Different Shapes Affects the Composition of the Salivary Proteome—A Pilot Study

Chewing is a sensorimotor activity that aims to prepare food for swallowing, in which saliva intervenes, moistening food particles, assisting in bolus formation, enzymatic digestion, and swallowing. This study investigated the effect of chewing carrots of different shapes on salivary secretion and protein profiles using 2-DE gel electrophoresis. Fifteen participants chewed sliced or grated carrots, or parafilm (non-food control), in a crossover design, with saliva samples collected before and after mastication. The results showed significant differences in salivation and saliva composition when chewing carrots vs. the control, with sliced carrots inducing greater changes: 37 protein spots, including amylase and immunoglobulin spots, 9 protein spots, and 1 protein spot were significantly altered after chewing sliced carrots, grated carrots, and parafilm, respectively. Mastication combined with the sensory properties of food had a greater effect on saliva secretion and salivary protein levels than mastication alone. Among carrot shapes, the results suggest that harder food textures, which require more chewing effort, lead to more pronounced changes in salivary protein profiles. These findings contribute to the understanding of how food shape and texture influence salivation and salivary proteome dynamics, with potential implications for oral digestion, food-related sensory experiences, and the personalisation of diets for individuals with a compromised chewing capacity.