Search Results (323)

Carbapenem resistance is considered one of the greatest current threats to public health, particularly in the management of infections in clinical settings. Carbapenem resistance in bacteria is mainly due to mechanisms such as the production of carbapenemases (such as the imipenemase IMP, or other enzymes like VIM, NDM, and KPC), that can be detected by several laboratory tests, including immunochromatography and automated real-time PCR (qPCR). Methods: As part of local studies to monitor carbapenem-resistant bacteria in Costa Rica, two cases were initially identified with inconsistent IMP detection results. A possible gene drop-out in the automated qPCR test was suggested based on the negative result, contrasting with the positive result by immunochromatography and whole-genome sequencing. We hypothesized that molecular testing could be optimized through the development of tailored assays to improve the detection of IMP genes. Thus, using IMP gene sequences from the local isolates and regional sequences in databases, primers were redesigned to extend the detection of IMP alleles of regional relevance. Results: The tailored qPCR was applied to a local collection of 119 carbapenem-resistant isolates. The genomes of all 14 positive cases were sequenced, verifying the results of the custom qPCR, despite the negative results of the automated testing. Conclusions: Guided by whole-genome sequencing, it was possible to extend the molecular detection of IMP alleles circulating in Latin America using a tailored qPCR to overcome IMP gene drop-out and false-negative results in an automated qPCR. Full article

Objective: To investigate the neuroprotective effects of a Rho-associated kinase (ROCK) inhibitor on retinal ganglion cells (RGCs) in vitro and in vivo. Methods: For in vivo studies, a unilateral optic nerve crush mouse model was established. Then, 100 mM Y-27632 (a ROCK inhibitor) or saline was applied to the experimental eyes once a day for 14 days. The effects of the ROCK inhibitor were evaluated by counting the surviving RGCs in the enucleated flat retina tissues and measuring the inner retinal thickness using optical coherence tomography (OCT), the amplitude of the electroretinogram (ERG), and the change in intraocular pressure (IOP). For the in vitro study, RGCs were isolated from five-day-old mice using a modified immunopanning method with magnetic beads. The isolated RGCs were incubated for 72 h with various concentrations of Y-27632, after which TUNEL assays were performed to determine the number of surviving RGCs. Results: Y-27632 has neuroprotective effects, as it significantly increased the number of surviving RGCs by approximately 6.3%. OCT and ERG data also revealed that Y-27632 induced neuroprotective effects in vivo; furthermore, Y-27632 reduced IOP by approximately 18.3%. The in vitro study revealed the dose-dependent neuroprotective effects of Y-27632, with the highest dose of Y-27632 (1000 nM) increasing the RGC survival rate after 72 h of incubation compared with that of the control. Conclusions: The ROCK inhibitor Y-27632 may exert some neuroprotective effects on RGCs when it is used as an eye drop through an IOP-independent mechanism. Full article

Antibiotic resistance is one of the most significant public health issues today. As a consequence, there is an urgent need for novel classes of antibiotics. This necessitates the development of highly efficient screening methods for the rapid identification of antibiotic-producing bacteria. Here, we describe a new method for high-throughput screening of antimicrobial compounds (AMC) producing halophilic bacteria. Our methodology used a newly designed 3D-printed Petri plate replicator used for drop deposition and colony replication. We employed this device in combination with a modified agar overlay assay to screen more than 7400 bacterial colonies. A total of 54 potential AMC producers were discovered at a success rate of 0.7%. Although 40% of them lost their antibacterial activity during the secondary screening, 22 strains retained inhibitory activity and were able to suppress the growth of one or more safe relatives of the ESKAPE group pathogens. The ethyl acetate extract from the most potent strain, Virgibacillus salarius POTR191, demonstrated moderate antibacterial activity against Enterococcus faecalis, Acinetobacter baumanii, and Staphylococcus epidermidis with minimal inhibitory concentrations of 128 μg/mL, 128 μg/mL, and 512 μg/mL, respectively. We propose that our replica plate assay could be used for target-based antimicrobial screening of various extremophilic bacteria. Full article

Sustainable deepwater drilling for oil and gas offers significant potential. In this work, we synthesized a nanoscale collapse-prevention agent by grafting didecyldimethylammonium chloride onto spherical nano-silica and characterized it using Fourier-transform infrared spectroscopy, thermogravimetric analysis, zeta-potential, and particle-size measurements, as well as SEM and TEM. Adding 1 wt% of this agent to a bentonite slurry only marginally alters its rheology and maintains acceptable low-temperature flow properties. Microporous-membrane tests show filtrate passing through 200 nm pores drops to 55 mL, demonstrating excellent plugging. Core-immersion studies reveal that shale cores retain integrity with minimal spalling after prolonged exposure. Rolling recovery assays increase shale-cutting recovery to 68%. Wettability tests indicate the water contact angle rises from 17.1° to 90.1°, and capillary rise height falls by roughly 50%, reversing suction to repulsion. Together, these findings support a synergistic plugging–adsorption–hydrophobization mechanism that significantly enhances wellbore stability without compromising low-temperature rheology. This work may guide the design of high-performance collapse-prevention additives for safe, efficient deepwater drilling. Full article

Background: We investigated the role of a high-sensitive cardiac troponin T (hsTnT) increase below the upper limit of normal (ULN) in patients with breast cancer (BC). hsTnT assays accurately quantify very low plasma troponin concentrations and enable the early detection of cardiomyocyte injury before a drop in the left ventricular ejection fraction (LVEF). The increase in hsTnT below the ULN in response to chemotherapy has not previously been studied. Method: This was an open-label pilot study. Female patients with newly diagnosed BC scheduled to receive systemic cancer treatment were recruited. Blood sampling and echocardiography were performed at baseline, at 3 and 6 months of cancer treatment. hsTnT concentrations were measured using the Elecsys TnT hs assay (Roche Diagnostics). The limit of blank and 99th percentile cutoff values for the hsTnT assay were 3 and 14 ng/L. We calculated the rise in hsTnT (ΔhsTnT) by the difference (%) between its baseline level and during follow-up (FU) in each patient. Results: Among eligible subjects, we excluded 4 patients before the start of treatment and 17 patients during the follow-up with values for the hsTnT >14 ng/L. Finally, 60 women with a median age of 48.6 ± 1.3 years were included in the study. The median baseline hsTnT concentration was 5.5 ± 1.4 ng/L. During 6 months of cancer treatment, hsTnT increased in all patients by up to 10–305% from baseline, with an average of 94.2%. LV EF was normal at baseline and decreased significantly compared to the value before cancer treatment (61.9 ± 3.3% vs. 56.3 ± 7.0%; p < 0.045). We correlated the hsTnT rise with a drop in LV EF ≥ 10% from its baseline level. Logistic regression analysis showed that Δ hsTnT has a good predictive value for LV dysfunction, 0.78 (p = 0.05), 95% CI (0.67–0.90). The increase in hsTnT > 81% was determined as the optimal threshold value for detecting early biochemical cardiotoxicity. Conclusion: It was investigated that hsTnT rise within the cutoff < 14 ng/L can be used as a marker of early biochemical cardiotoxicity and is valuable for predicting LV drop in 6 months of FU. We conclude that BC patients with increased hsTnT plasma concentration > 81% from the baseline value should be considered as high-risk patients for cardiotoxicity and need more precise cardiac monitoring and early preventive medical intervention strategies. Full article

Renal cell carcinoma (RCC) presents significant therapeutic challenges due to its resistance to conventional treatments. Natural compounds with photodynamic properties, such as chlorophyll derivatives, offer potential for novel interventions. This study investigates the apoptotic effects of a chlorophyll-enriched green propolis compound activated by daylight-mediated photodynamic therapy (PDT) on RCC cells. A novel compound formulated from standardized ethanol extracts of Taiwanese green propolis, wheatgrass, and mulberry leaves was characterized using high-performance liquid chromatography (HPLC). Human RCC 786-O cells were treated with varying concentrations of the compound, with or without daylight PDT (570 nm). Cell viability was assessed via MTT assay, and median effective concentrations (EC₅₀) were calculated. HPLC analysis identified Artepillin C as the major constituent. The compound induced dose-dependent cytotoxicity, which was significantly enhanced by daylight PDT. EC₅₀ values dropped from 3.027 µL (compound alone) to 1.728 µL (with PDT), indicating synergistic efficacy. Cell viability significantly decreased in PDT-treated cells compared to non-treated controls (p < 0.05) indicating apoptosis. Daylight-activated PDT significantly amplifies the anticancer efficacy of the compound against RCC cells. Preliminary data suggest the potential of chlorophyll-enriched green propolis photodynamic activation (GPDT) as a natural adjunctive strategy for RCC, warranting further in vivo investigation. Full article

Innate immune cells appear to have an important implication in the resolution and/or the aggravation of the COVID-19 pathogenesis after infection with SARS-CoV-2. To better appreciate the role of these cells during COVID-19, changes in blood eosinophil, the neutrophil and monocyte count, and levels of surface protein markers have been reported. However, analyses at several timepoints of multiple surface markers on granulocytes and monocytes over a period of one month after a SARS-CoV-2 infection are missing. Therefore, in this study, we performed blood eosinophil, neutrophil, and monocyte phenotyping using a list of surface proteins and flow cytometry during a period of 30 days after the hospitalization of patients with severe SARS-CoV-2 infections. Blood cell counts were reported at seven different timepoints over the 30-day period as well as measures of multiple mediators in serum using a targeted multiplex assay approach. Our results indicate a 95% drop in the blood eosinophil count by D1, with eosinophils displaying a phenotype defined as CD69/CD63/CD125^high and CCR3/CD44^low during the early phases of hospitalization. Conversely, by D7 the neutrophil count increased significantly and displayed an immature, activated, and immunosuppressive phenotype (i.e., 3% of CD10/CD16^low and CD10^lowCD177^high, 6.7% of CD11b^highCD62L^low, and 1.6% of CD16^highCD62L^low), corroborated by enhanced serum proteins that are markers of neutrophil activation. Finally, our results suggest a rapid recruitment of non-classical monocytes leaving CD163/CD64^high and CD32^low monocytes in circulation during the very early phase. In conclusion, our study reveals potential very early roles for eosinophils and monocytes in the pathogenesis of COVID-19 with a likely reprogramming of eosinophils in the bone marrow. The exact roles of the pro-inflammatory neutrophils and the functions of the eosinophils and the monocytes, as well as these innate immune cell types, interplays need to be further investigated. Full article

Edible flowers are gaining recognition as rich sources of nutrients and phytochemicals. In Mexico, the flower of Agave inaequidens has been traditionally consumed since pre-Hispanic times. This study investigated its nutritional profile and the in vitro gastrointestinal bioaccessibility of its phenolic fraction. During in vitro digestion (oral, gastric, and intestinal), the total phenolic content of A. inaequidens significantly decreased from 138 to 21 mg GAE/100 g DW (15.22% bioaccessibility), while total flavonoid content dropped from 8 to 4.6 mg CE/100 g DW (57.5% bioaccessibility). Consequently, antioxidant activity, assessed by ABTS, DPPH, and hemolysis inhibition assays, also declined post-digestion. Interestingly, the digestive process modulated the flower’s inhibitory activity against digestive enzymes before and after in vitro digestion: α-amylase inhibition slightly decreased (IC₅₀ 1.8 to 2.1 mg/mL), but α-glucosidase (IC₅₀ 2.7 to 1.6 mg/mL) and lipase (IC₅₀ > 3 to 1.4 mg/mL) inhibition increased. The A. inaequidens flower is a good source of fiber and low in fat. These findings underscore its potential as a functional food ingredient, offering bioaccessible phenolic compounds with antioxidant and enzyme inhibitory properties. Full article

The rising demand for vegan products calls for new plant-based antimicrobial preservation methods. This study evaluates an antifungal ingredient obtained by fermenting oat drink with lactic acid bacteria to extend vegan mortadella’s shelf life. In vitro tests showed antimicrobial effects against Aspergillus flavus, Penicillium commune, and Listeria monocytogenes (inhibition zones: 2–5 mm). The enrichment of the oat drink culture medium with additional nutrients enhanced fermentation performance and increased antifungal activity. The fermented culture medium with the highest antimicrobial activity was used to develop a bioactive ingredient for the preservation of vegan mortadella conservation. Adding 3% of this ingredient to vegan mortadella improved microbial stability, reducing mesophilic bacteria by 2.5 Log₁₀ CFU/g and increasing lactic acid bacteria. Lower pH and water activity changes were observed but remained within quality standards. Contamination assays showed a consistent reduction of A. flavus over 7 days, while P. commune and L. monocytogenes dropped below detection within 2 days. In contrast, control samples maintained contamination levels near 3.0 Log₁₀ CFU/g. These findings support the potential of fermented oat-based ingredients as effective, natural preservatives for vegan foods. Full article

Microglia, the resident immune cells of the central nervous system (CNS), play essential roles in maintaining brain homeostasis. While transient activation is protective, chronic microglial reactivity contributes to neuroinflammatory damage and neurodegeneration. The mitochondrial mechanisms underlying this shift remain poorly understood. Here, we investigated whether lipopolysaccharide (LPS) induces coordinated mitochondrial and metabolic alterations in BV-2 microglial cells. LPS stimulation (100 ng/mL, 24 h) induced a reactive phenotype, with increased Iba1 (+82%), F4/80 (+132%), and Cd68 (+44%), alongside elevated hydrogen peroxide (~6-fold) and nitrite (~45-fold). Cytotoxicity increased by 40% (LDH assay), and cell viability dropped to ~80% of the control (MTT). Extracellular lactate increased, indicating glycolytic reprogramming. However, LPS-primed cells showed greater ATP depletion under antimycin A challenge, reflecting impaired metabolic flexibility. Hoechst staining revealed a ~4-fold increase in pyknotic nuclei, indicating apoptosis. Mitochondrial dysfunction was confirmed by a 30–40% reduction in membrane potential (TMRE, JC-1), a ~30% loss of Tomm20, and changes in dynamics: phospho-Drp1 increased (+23%), while Mfn1/2 decreased (33%). Despite a ~70% rise in Lamp2 signal, Tomm20–Lamp2 colocalization decreased, suggesting impaired mitophagy. High-resolution respirometry revealed decreased basal (−22%), ATP-linked (24%), and spare respiratory capacity (41%), with increased non-mitochondrial oxygen consumption. These findings demonstrate that LPS induces mitochondrial dysfunction, loss of metabolic adaptability, and increased apoptotic susceptibility in microglia. Mitochondrial quality control and energy flexibility emerge as relevant targets to better understand and potentially modulate microglial responses in neuroinflammatory and neurodegenerative conditions. Full article

Background: Endophthalmitis, associated with intraocular procedures, is an infection of the eye that can rapidly result in substantial irreversible loss of vision and may even lead to removal of the eye. Prevention strategies vary globally and often include antibiotic use—often consisting of a broad-spectrum mono-therapeutic agent. The purpose of this study is to test the efficacy and value of combinations of cefuroxime (cefu), cefazolin (cefa), azithromycin (azith), and/or tobramycin (tob) as alternatives to the use of moxifloxacin. We looked for synergism since these antimicrobials used different mechanisms of inhibition. Methods: Initially, we examined individual antimicrobials to determine the minimal bactericidal concentrations (MBC) of each individual treatment against Klebsiella pneumonia, Pseudomonas aeruginosa, Staphylococcus aureus, two clinical isolates of MRSA, and Staphylococcus epidermidis, by both the Zone of Inhibition (ZOI) and the Colony Forming Unit (CFU) assays. We then used these data in a combinatorial study. Results: We found combinations that were synergistic against all the bacteria tested, resulting in total eradication [8 logs] of all bacteria. We found that the ZOI assay provided less consistent results than the CFU assays. Conclusions: We have found combinations of these antimicrobials that were synergistic in the total eradication (8 logs) of all bacteria tested. These three combinations were: cefuroxime/azithromycin; azithromycin/tobramycin; and cefuroxime/tobramycin. Moxifloxacin (Vigamox) did not completely eradicate Staphylococcus epidermidis. These combinations can then be used as eye drops to serve as a prophylactic for endophthalmitis after eye injections and eye surgery. Full article

Norovirus, a foodborne pathogen, causes a significant economic and health burden globally. Although detection methods exist, they are expensive and non-field deployable. A flow-based dielectrophoretic biosensor was designed for the detection of foodborne pathogenic viruses and was tested using bacteriophage MS2 as a norovirus surrogate. The flow-based MS2 sensor comprises a concentrator and a detector. The concentrator is an interdigitated electrode array designed to impart dielectrophoretic effects to manipulate viral particles toward the detector in a fluidic channel. The detector is made of a silver electrode conjugated with anti-MS2 IgG to allow for antibody–antigen biorecognition events and is supplied with the electrical current for the purpose of measurement. Serially diluted MS2 suspensions were continuously injected into the fluidic channel at 0.1 mL/min. A cyclic voltammogram indicated that current measurements from single-walled carbon nanotube (SWCNT)-coated electrodes increased compared to uncoated electrodes. Additionally, a drop in the current measurements after antibody immobilization and MS2 capture was observed with the developed electrodes. Antibody immobilization at the biorecognition site provided greater current changes with the antibody-MS2 complexes vs. the assays without antibodies. The electric field applied to the fluidic channel at 10 V_pp and 1 MHz contributed to an increase in current changes in response to MS2 bound on the detector and was dependent on the MS2 concentrations in the sample. The developed biosensor was able to detect MS2 with a sensitivity of 10² PFU/mL within 15 min. Overall, this work demonstrates a proof of concept for a rapid and field-deployable strategy to detect foodborne pathogens. Full article

The stability of Cotinus coggygria extract-loaded liposomes (non-treated and UV-irradiated) was determined after 60 days through an investigation of the impact of storage on liposomal physical and antioxidant properties. The liposome size varied in a narrow range for 60 days; PDI was 0.273–0.313 (non-treated) and 0.829–0.911 (UV-irradiated). The zeta potential ranged from −28.2 to −29.6 mV (non-treated) and from −21.5 to −22.0 mV (UV-irradiated). The obtained liposomes with the extract neutralized 81.9% of free DPPH radicals before UV irradiation and 80.9% after irradiation. In the ABTS assay, UV irradiation also significantly reduced the antioxidant capacity, from 12.02 to 10.55 µmol Trolox equivalent (TE)/mL. The ABTS and DPPH radical scavenging activity of the UV-irradiated liposomes significantly decreased after the 60-day storage (8.93 µmol TE/mL and 75.4%, respectively), whereas in the non-treated sample, the mentioned drop in antioxidant capacity was not noticed. Liposomal formulations of C. coggygria extract can exhibit significant potential for further development as a functional food or dermo-cosmetic ingredient. Full article

Thelazia callipaeda (T. callipaeda) is a zoonotic ocular parasite that poses a public health risk. Current treatment depends on mechanical parasite extraction and specific prophylactic anthelmintics, but direct ocular deworming agents are limited and expensive, and mechanical removal is often incomplete, causing animal stress and ocular injury. Levamisole eye drops have been applied; however, their efficacy and safety remain undefined. We established a standardized T. callipaeda viability assessment system (TVAS) and an animal infection model to evaluate candidate drugs. In vivo, 5 mg/mL levamisole was administered at escalating frequencies. Ocular symptoms, complete blood count (CBC), and serum biochemistry were measured, and cytotoxicity was assessed using Cell Counting Kit-8 (CCK-8) and lactate dehydrogenase (LDH) assays on rabbit conjunctival epithelial cells (RCECs).Four doses of 5 mg/mL levamisole applied at 30 min intervals cleared T. callipaeda from infected eyes within 2 h without significant changes in CBC, serum biochemistry, or ocular symptom scores. CCK-8 and LDH assays indicated minimal cytotoxicity in RCECs within 4 h. However, prolonged exposure (6–12 h) led to a significant decrease in RCEC viability, suggesting potential cytotoxicity with extended use and highlighting the need for further safety evaluation. A regimen of four topical administrations of 5 mg/mL levamisole at 30 min intervals cleared T. callipaeda from infected eyes with minimal cytotoxicity, supporting its safety and efficacy as a topical treatment. Full article

Background: Glioblastoma (GBM) remains almost uniformly fatal, owing in part to therapy-resistant cancer stem-like cells (CSCs) and to temozolomide (TMZ) resistance driven by O⁶-methylguanine-DNA methyltransferase (MGMT). Differentiation therapy with all-trans retinoic acid (ATRA) has the potential to attenuate stemness and sensitize GBM to TMZ. We therefore asked whether ATRA reduces expression of key CSC markers and MGMT in established GBM lines. Methods: Two established human GBM cell lines, U87-MG and A172, were cultured under neurosphere-promoting conditions to enrich for potential stem-like subpopulations. Cells were treated with either 1 µM ATRA or vehicle control (DMSO) for 5 days. Total RNA was extracted, and cDNA was synthesized. Quantitative Real-Time PCR (qPCR) assessed relative mRNA expression levels of key stemness transcription factors (SOX2, NES) and the DNA repair gene MGMT and corresponding protein levels were measured by an Enzyme-Linked Immunosorbent Assay (ELISA). Gene expression was normalized to the geometric mean of two validated housekeeping genes (GAPDH, ACTB). Relative quantification was calculated using the ΔΔCt method, and statistical significance was determined using Student’s t-tests. Results: ATRA markedly suppressed stemness and MGMT in both lines. In U87-MG, SOX2 mRNA fell 3.7-fold (p = 0.0008) and protein 2.99-fold (148.3 ± 6.0 → 49.7 ± 2.7 pg µg⁻¹; p = 0.0002); Nestin dropped 4.1-fold (p = 0.0005) and 3.51-fold (450.0 ± 17.3 → 128.3 ± 4.4 pg µg⁻¹; p = 0.00008). MGMT decreased 2.6-fold at transcript level (p = 0.0065) and 2.11-fold at protein level (81.7 ± 4.4 → 38.7 ± 1.8 pg µg⁻¹; p = 0.0005). In A172, SOX2 was reduced 2.9-fold (p = 0.0041) and 2.31-fold (p = 0.0007); Nestin 3.3-fold (p = 0.0028) and 2.79-fold (p = 0.00009). MGMT declined 2.2-fold (p = 0.0132) and 1.82-fold (p = 0.0015), respectively. Conclusions: Five-day exposure to ATRA diminishes SOX2, Nestin, and MGMT at both mRNA and protein levels in stem-enriched GBM cultures, supporting the premise that ATRA-induced differentiation can concurrently blunt CSC traits and TMZ-resistance mechanisms. These data provide a molecular rationale for testing ATRA in combination regimens aimed at improving GBM therapy. Full article