Search Results (81)

In the Chan Buddhism koan (gong’an 公案) tradition, the act of “slaying the snake” functions as a signature gesture imbued with complex, historically layered cultural meanings. Rather than merely examining its motivations, this paper emphasizes tracing the semantic transformations that this motif has undergone across different historical contexts. It argues that “snake-slaying” operated variously as an imperial narrative strategy reinforcing ruling class ideology; as a form of popular resistance by commoners against flood-related disasters; as a dietary practice among aristocrats and literati seeking danyao (elixirs) 丹藥 for reclusion and transcendence; and ultimately, within the Chan tradition, as a method of spiritual cultivation whereby masters sever desires rooted in attachment to both selfhood and the Dharma. More specifically, first, as an imperial narrative logic, snake-slaying embodied exemplary power: both Liu Bang 劉邦 and Guizong 歸宗 enacted this discursive strategy, with Guizong’s legitimacy in slaying the snake deriving from the precedent set by Liu Bang. Second, as a folk strategy of demystification, snake-slaying acquired a moral aura—since the snake was perceived as malevolent force, their slaying appeared righteous and heroic. Finally, as a mode of self-cultivation among the aristocracy, snake-slaying laid the groundwork for its later internalization. In Daoism, slaying the snake was a means of cultivating the body; in Chan Buddhism, the act is elevated to a higher plane—becoming a way of cultivating the mind. This transformation unfolded naturally, as if predestined. In all cases, the internalization of the snake-slaying motif was not an overnight development: the cultural genes that preceded its appearance in the Chan tradition provided the fertile ground for its karmic maturation and discursive proliferation. Full article

E. coli attaches to, and forms biofilms on various surfaces, including latex and polystyrene, contributing to nosocomial spread. E. coli responds to both exogenous and endogenous insulin, which induces behavioral changes. Human insulin, a quorum signal surrogate for microbial insulin, may affect the ability of E. coli to interact with latex and polystyrene in the presence of various sugars. E. coli ATCC 25923 was grown in peptone (1%) yeast nitrogen base broth to either the logarithmic or stationary growth phase. Adherence to latex was determined using 6 × 6 mm latex squares placed in a suspension of washed cells (10³ CFU/mL; 30 min; 37 °C) in buffer containing insulin at 2, 20, and 200 µU/mL (Humulin^® R; Lilly) with and without mannose, galactose, fructose, sorbose, arabinose, xylose, lactose, maltose, melibiose, glucose-6-phosphate, glucose-1-phosphate, and glucosamine at concentrations reported to affect behavioral response. Attachment levels to latex were determined by the press plate method. Biofilm levels were measured in a similar fashion but with overnight cultures in flat bottom uncoated polystyrene plates. Controls were media, insulin, sugar, or buffer alone. Glucose served as the positive control. Overall, the stationary phase cells’ adherence to latex was greater, regardless of the test condition, than was measured for the logarithmic phase cells. The effect of insulin on adherence to latex was insulin and sugar concentration dependent. The addition of insulin (200 µU/mL) resulted in a significantly (p < 0.05) increased adherence to latex and biofilm formation on polystyrene compared with sugar alone for 12 of the 13 sugars tested with stationary phase bacteria and 10 of the 13 sugars tested with logarithmic phase bacteria. Adherence in response to sorbose was the only sugar tested that was unaffected by insulin. These findings show that insulin enhances E. coli’s association with materials in common usage in medical environments in a nutrition-dependent manner. Full article

Historical monuments enrich lives by connecting people to their community and heritage. They are key to understanding population movements and local development. This study analyzes how the presence of castles classified as National Monuments or of Public Interest can attract tourists, extend their stay, and motivate local authorities to invest in cultural activities, in particular in low-density areas. Heritage and tourism connect through economic, cultural, social, and territorial goals. They boost local economies, preserve cultural traditions, empower communities, and protect landscapes. Thoughtful planning ensures that regional character is maintained and a balance is struck between growth, preservation, equity, and safeguarding for the future. To evaluate the economic and touristic impact of classified castles in Portuguese municipalities, we started by analyzing the ‘Existence of Castle’ variable against three key factors: tourism flows (number of guests and average stay) and municipal expenditure on cultural activities Additionally, we considered other important explanatory variables from alternative studies, including the following: the number of parishes, maximum and minimum altitude, cultural heritage and sports activity expenses, resident population, density, monthly earnings, commercial firms, Tourism Carrying Capacity, tourism revenues, and overnight stays. Our empirical results reveal that medieval castles generate diverse effects on cultural activities managed by municipalities. Quantile regressions demonstrate that municipalities with fewer resources but with a castle invest more in cultural activities, regardless of how expenditure is measured. Additionally, the presence of a castle attracts more tourists and extends their stay, indicating significant potential for economic and tourist development which is not yet fully realized. Full article

Spermatogenesis is a process of self-renewal of spermatogonial stem cells and their proliferation and differentiation to generate mature sperm. This process involves interactions between testicular somatic (mainly Sertoli cells) and spermatogonial cells at their different stages of development. The functionality of Sertoli cells is regulated by hormones and testicular autocrine/paracrine factors. In this study, we investigated the effects of follicle-stimulating hormone (FSH) and testosterone addition on Sertoli cell cultures that undergo hypotonic shock, with a primary focus on Sertoli cell activity. Cells were enzymatically isolated from testicular seminiferous tubules of 7-day-old mice. These cells were cultured in vitro for 3 days. Thereafter, some cultures were treated with hypotonic shock to remove germ cells. After overnight, fresh media without (control; CT) or with FSH, testosterone (Tes), or FSH+T were added to the hypotonic shock-treated or untreated (CT) cultures for 24 h. The morphology of the cultures and the presence of Sertoli cells and germ cells were examined. The expression of growth factors (CSF-1, LIF, SCF, GDNF) or other specific Sertoli cell factors [transferrin, inhibin b, androgen receptor (AR), androgen binding protein (ABP), FSH receptor (FSHR)] was examined by qPCR. Our immunofluorescence staining showed depletion/major reduction in VASA-positive germ cells in Sertoli cell cultures following hypotonic shock (HYP) treatment compared to untreated cultures (WO). Furthermore, the expression of the examined growth factors and other factors was significantly increased in HYP cultures compared to WO (in the CT). However, the addition of hormones significantly decreased the expression levels of the growth factors in HYP cultures compared to WO cultures under the same treatment. In addition, the expression of all other examined Sertoli cell factors significantly changed following HYP treatment compared to WO and following treatment with FSH and or T. However, the expression levels of some factors remained normal following the treatment of Sertoli cell cultures with one or both hormones (transferrin, Fsh-r, Abp, Ar). Thus, our results demonstrate the crucial role of germ cells in the functionality of Sertoli cells and the possible role of FSH and T in maintaining, at least partially, the normal activity of Sertoli cells following germ cell depletion in vitro by hypotonic shock treatment. Full article

Background: Prompt administration of optimal antibiotic therapy is essential in the management of bacteraemia to reduce morbidity and mortality and to facilitate antibiotic stewardship. To identify the most effective therapy, rapid and accurate antimicrobial susceptibility testing (AST) is essential. ASTar is an automated AST system that delivers minimum inhibitory concentrations (MICs) for 23 antimicrobials and is designed for testing Gram-negative bacteria directly from positive blood cultures, with results available after 6 h. Methods: The ASTar system was evaluated with 64 positive blood cultures from patients with bacteraemia and 56 simulated blood cultures inoculated with a range of antibiotic-resistant isolates. The ASTar results for 12 antibiotics commonly used in our hospitals were compared with the results derived from three different methods of disc susceptibility testing and MICs determined by broth microdilution (BMD). Results: For 121 isolates of Gram-negative bacteria, ASTar showed an average essential agreement of 87.2% and an average categorical agreement of 94%, when compared with BMD. Very major errors (false susceptibility) and major errors (false resistance) were associated with 0.9% and 3.4% of results, respectively. The results were at least as accurate as those obtained from EUCAST disc susceptibility methods (both rapid and overnight methods). Conclusions: The ASTar system is an effective method for delivering accurate and rapid (6 h) AST results for Gram-negative bacteria by the direct testing of positive blood cultures. Full article

The development of biological pesticides is rapidly becoming an integral aspect of pest management in sustainable agriculture. This study was conducted to evaluate the effectiveness of Bacillus halotolerans strain B33 against three common seedborne fungal pathogens—Fusarium graminearum, Alternaria alternata, and Aspergillus flavus. B33 strain identity was determined using the 16S rRNA and tuf gene sequences. Commercial wheat, barley, oat, and rye seeds were artificially infected by fungal isolates and then treated with B33 overnight culture in Nutrient Broth. The obtained results indicate high efficacy against F. graminearum (83.55–94.38%) and A. alternata (85.05–96.70%), whereby the highest efficacy was noted on wheat seed and the lowest was detected on rye seed. On the other hand, B33 achieved 100% efficacy against A. flavus on barley, rye, and oat seeds, while being 96.24% effective against this pathogen on wheat. Principal component analysis indicated the highest treatment influence on A. flavus. The effect of all tested treatments on seed germination was statistically significant compared to the controls, whereby the number of germinated seeds declined as the seed infection rate increased. B. halotolerans strain B33 effectively managed seedborne fungal pathogens, thereby enhancing seed germination. Full article

This study evaluated the impact of a single variation in the etching time of H₂SO₄/H₂O₂-treated titanium (Ti) surfaces on the adsorption of growth and differentiation factor-5 (GDF-5) and their effects on the acquisition of the osteogenic phenotype in vitro. Rat primary calvarial osteogenic cells were grown for up to 14 days on the following Ti surfaces: (1) 30 min: nanotopography obtained with a 1:1 mixture of H₂SO₄/H₂O₂ for 30 min (control); (2) 30 min + GDF-5: a 30 min-etched Ti sample adsorbed with recombinant human (rh) GDF-5; (3) 4 h: nanotopography obtained with a 1:1 mixture of H₂SO₄/H₂O₂ for 4 h (control); (4) 4 h + GDF-5: a 4 h-etched Ti sample adsorbed with rhGDF-5. The GDF-5 adsorption procedure was carried out on the day before cell plating using 200 ng/mL rhGDF-5 overnight at 4 °C. The 30 min- and 4 h-etched Ti samples exhibited a high hydrophilic network of nanopits with a tendency towards larger nanopits for the 4 h group, which corresponded to an enhanced GDF-5 adsorption. For both etching times, coating with GDF-5 resulted in less hydrophilic surfaces that supported (1) a reduction in the proportion of spread cells and an enhanced extracellular osteopontin labeling at early time points of culture, and (2) increased alkaline phosphatase activity preceding an enhanced mineralized matrix formation compared with controls, with a tendency towards higher osteogenic activity for the 4 h + GDF-5 group. In conclusion, the osteogenic potential induced by the GDF-5 coating can be tailored by subtle changes in the nanotopographic characteristics of Ti surfaces. Full article

Various scholars have noted Mesopotamian impact on various aspects of Israelite religious beliefs and practices and extrapolated from these on the broader nature of the relations between the regions. Indeed, no society is an island, and influences are inevitable, especially when a small, peripheral society is in contact with a powerful center. Still, insufficient attention has been paid to the social and cultural contexts of the interaction, and studies have often extrapolated from examples that are exceptional. A systematic examination of the cultural and social reactions in Judah to the intensifying interaction with Assyria reveals that avoidance, subversion, and resistance were far more prevalent than emulation. The large-scale death and deportations that accompanied the destruction of sites and regions by the Assyrian armies in the last third of the 8th century (mostly outside Judah) resulted in an understanding that nothing was secure anymore, that complete kinship groups could be annihilated overnight, and that long-held traditions could simply vanish. This gave rise to a mentality of “life in the shadow of the bomb”, which explains many subsequent developments in Judah, providing the context for various religious changes. Full article

Using a murine osteomyelitis model, we recently demonstrated that Staphylococcus aureus sarA and sarA/agr mutants generated in the USA300 strain LAC are attenuated to a greater extent than an isogenic agr mutant and that this can be attributed to a significant extent to the increased production of extracellular proteases in both mutants. Based on this, we used a mass-based proteomics approach to compare the proteomes of LAC, its isogenic agr, sarA, and sarA/agr mutants, and isogenic derivatives of all four of these strains unable to produce the extracellular proteases aureolysin, SspA, SspB, ScpA, or SplA-F. This allowed us to identify proteins that were present in reduced amounts in sarA, and sarA/agr mutants owing to the increased production of extracellular proteases. A total of 1039 proteins were detected in conditioned media (CM) from overnight cultures of LAC, and protease-mediated degradation was shown to contribute to the reduced abundance of 224 of these (21.6%) in CM from the sarA and sarA/agr mutants. Among these were specific proteins previously implicated in the pathogenesis and therapeutic recalcitrance of S. aureus osteomyelitis. This demonstrates that the ability of sarA to limit protease production plays a key role in post-translational remodeling of the S. aureus proteome to a degree that can be correlated with reduced virulence in our osteomyelitis model, and that it does so irrespective of the functional status of agr. This also suggests that at least some of these 224 proteins may be viable targets for prophylactic or therapeutic intervention. Full article

Background/Objectives: Bacteraemia can be fatal without antibiotic intervention. Antibiotic Susceptibility Testing (AST) provides the necessary information for targeted antibiotic therapy; however, the traditional method using disc diffusion can take over two days from a positive blood culture. Inappropriate empiric therapy is associated with increased mortality and increased antibiotic resistance, highlighting the need for more rapid turnaround times for AST. By making changes to an established method, turnaround times can be reduced. Methods: Eighty-two patient positive blood culture samples were collected from January to April 2022, representing the range of common bacteria causing sepsis. This followed the normal methodology in the laboratory of inoculating agar from positive blood cultures in preparation for European Committee on Antimicrobial Susceptibility Testing (EUCAST) disc diffusion AST method. EUCAST methodology outlines that disc diffusion should be performed on isolates from an overnight culture of 16–24 h. This study looked at comparing disc diffusion results from cultures with 6 h of incubation to those with incubation times of 24 h, after organism identification by MALDI-ToF. Results from 6-h and 24-h cultures were compared by disc zone sizes and by interpreted susceptibility reading following EUCAST guidelines of sensitive, resistant, susceptible with increased exposure, or an area of technical uncertainty. Results: A total of 99.65% interpreted susceptibility readings matched across all organisms to all relevant antibiotics, with an average zone size difference of 1.08 mm between results from 6 h versus 24 h cultures. Conclusions: This method offers a non-automated way of using the traditional disc diffusion method, reducing turnaround times while still producing reliable and accurate results. This would mean validated ASTs can be set up in the same day as a blood culture flags positive rather than waiting for a longer culture. As this method is widely used within the laboratory already, it would mean that additional training is not required, as the process is the same, and only incubation time varies. This would positively impact patient outlook due to the shorter use of empiric therapy, and benefit antimicrobial stewardship (AMS). Full article

Bloodstream infections (BSIs) are life-threatening infections for which a timely initiation of appropriate antimicrobial therapy is critical. Antibiotic susceptibility testing (AST) directly performed on positive blood culture broths can help initiate targeted antibiotic therapy sooner than the standard AST performed on colonies isolated on solid media after overnight incubation. Faster antimicrobial susceptibility testing (AST) results can improve clinical outcomes, and reduce broad-spectrum antimicrobial consumption and healthcare-associated costs in sepsis. In this study, we evaluated the accuracy of a direct AST inoculation method on the BD Phoenix M50 system using serum separator tubes to harvest bacteria from positive pediatric blood culture bottles. Direct AST was performed on 132 monomicrobial pediatric blood culture bottles that were positive for Enterobacterales (65; 49.2%), Staphylococcus aureus (46; 34.8%), and non-fermenting Gram-negative bacilli (21; 16%). Overall, the categorical and essential agreements between the direct method and standard method were 99.6% and 99.8%, respectively. Very major, major, and minor error rates were 0.1%, 0.09%, and 0.20% respectively. Direct AST performed on pediatric blood culture bottles using BD Phoenix M50 can quickly provide accurate susceptibility information to guide antimicrobial therapy in patients with BSI. Full article

Fibres in the micro- and nanometre scale are suited to a broad range of applications, including drug delivery and tissue engineering. Electrospinning is the manufacturing method of choice, but it has some limitations. Novel pressure-driven fibre-forming techniques, like pressurised gyration (PG), overcome these limitations; however, the compatibility of PG with biological materials has not yet been evaluated in detail. For the first time, this limitation of PG was investigated by optimising PG for microbial cell processing and incorporating bacterial cultures into fibrous polymeric scaffolds for sustained release. Multiple polymer–solvent systems were trialled, including polyvinylpyrrolidone (PVP)/phosphate-buffered saline (PBS) 25% w/v, polyethylene oxide (PEO)/PBS 20% w/v, and PVP/ethanol 20% w/v. Rheological studies revealed the surface tension of the PVP/PBS, PEO/PBS, and PVP/ethanol polymer–solvent systems to be 73.2, 73.9, and 22.6 mN/m, respectively. Scanning electron microscopy showed the median fibre diameters to be between 9.8 μm and 26.1 μm, with PVP producing larger fibres. Overnight Bacillus subtilis cultures were then incorporated into the chosen polymeric solutions and processed into fibres using PG. The produced cell-loaded fibres were incubated in LB broth to assess the cell viability of the encapsulated cells. Colony counts post-incubation showed the PVP/PBS 25% fibres resulted in 60% bacterial growth, and PEO/PBS 20% fibres led to 47% bacterial growth, whereas PVP/ethanol 20% fibres did not lead to any bacterial growth. Based on the results gathered during this study, it can be concluded that PG offers a promising way of encapsulating cells and other sensitive biological products while having many notable advantages compared to electrospinning. This research demonstrates proof of concept research-based evidence and showcases the potential of pressurised gyration as a key disruptive innovation in probiotic delivery system design and manufacturing. Full article

Staphylococcus aureus is frequently highlighted as a priority for novel drug research due to its pathogenicity and ability to develop antibiotic resistance. Coagulase-negative staphylococci (CoNS) are resident flora of the skin and nares. Previous studies have confirmed their ability to kill and prevent colonization by S. aureus through the production of bioactive substances. This study screened a bank of 37 CoNS for their ability to inhibit the growth of methicillin-resistant S. aureus (MRSA). Deferred antagonism assays, growth curves, and antibiofilm testing performed with the cell-free supernatant derived from overnight CoNS cultures indicated antimicrobial and antibiofilm effects against MRSA indicators. Whole genome sequencing and BAGEL4 analysis of 11 CoNS isolates shortlisted for the inhibitory effects they displayed against MRSA led to the identification of two strains possessing complete putative bacteriocin operons. The operons were predicted to encode a nukacin variant and a novel epilancin variant. From this point, strains Staphylococcus hominis C14 and Staphylococcus epidermidis C33 became the focus of the investigation. Through HPLC, a peptide identical to previously characterized nukacin KQU-131 and a novel epilancin variant were isolated from cultures of C14 and C33, respectively. Mass spectrometry confirmed the presence of each peptide in the active fractions. Spot-on-lawn assays demonstrated both bacteriocins could inhibit the growth of an MRSA indicator. The identification of natural products with clinically relevant activity is important in today’s climate of escalating antimicrobial resistance and a depleting antibiotic pipeline. These findings also highlight the prospective role CoNS may play as a source of bioactive substances with activity against critical pathogens. Full article

The problem that this study addresses is to understand how microwave radiation is able to degrade genomic DNA of E. coli. In addition, a comparative study was made to evaluate the suitability of a high-throughput automated electrophoresis platform for quantifying the DNA degradation under microwave radiation. Overall, this study investigated the genomic DNA degradation of E. coli under microwave radiation using automated gel electrophoresis. To examine the viable organisms and degradation of genomic DNA under microwave exposure, we used three methods: (1) post-microwave exposure, where E. coli was enumerated using modified mTEC agar method using membrane filtration technique; (2) extracted genomic DNA of microwaved sample was quantified using the Qubit method; and (3) automated gel electrophoresis, the TapeStation 4200, was used to examine the bands of extracted DNA of microwaved samples. In addition, to examine the impacts of microwaves, E. coli colonies were isolated from a fecal sample (dairy cow manure), these colonies were grown overnight to prepare fresh E. coli culture, and this culture was exposed to microwave radiation for three durations: (1) 2 min; (2) 5 min; and (3) 8 min. In general, Qubit values (ng/µL) were proportional to the results of automated gel electrophoresis, TapeStation 4200, DNA integrity numbers (DINs). Samples from exposure studies (2 min, 5 min, and 8 min) showed no viable E. coli. Initial E. coli levels (at 0 min microwave exposure) were 5 × 10⁸ CFU/mL, and the E. coli level was reduced to a non-detectable level within 2 min of microwave exposure. The relationships between Qubit and TapeStation measurements was linear, except for when the DNA level was lower than 2 ng/µL. In 8 min of microwave exposure, E. coli DNA integrity was reduced by 61.7%, and DNA concentration was reduced by 81.6%. The overall conclusion of this study is that microwave radiation had a significant impact on the genomic DNA of E. coli, and prolonged exposure of E. coli to microwaves can thus lead to a loss of genomic DNA integrity and DNA concentrations. Full article

KH-type splicing regulatory protein (KSRP) is a single-stranded nucleic acid-binding protein with multiple functions. It is known to bind AU-rich motifs within the 3′-untranslated region of mRNA species, which in many cases encode dynamically regulated proteins like cytokines. In the present study, we investigated the role of KSRP for the immunophenotype of macrophages using bone marrow-derived macrophages (BMDM) from wild-type (WT) and KSRP^−/− mice. RNA sequencing revealed that KSRP^−/− BMDM displayed significantly higher mRNA expression levels of genes involved in inflammatory and immune responses, particularly type I interferon responses, following LPS stimulation. In line, time kinetics studies revealed increased levels of interferon-γ (IFN-γ), interleukin (IL)-1β and IL-6 mRNA in KSRP^−/− macrophages after 6 h subsequent to LPS stimulation as compared to WT cultures. At the protein level, KSRP^−/− BMDM displayed higher levels of these cytokines after overnight stimulation. Matching results were observed for primary peritoneal macrophages of KSRP^−/− mice. These showed higher IL-6, tumor necrosis factor-α (TNF-α), C-X-C motif chemokine 1 (CXCL1) and CC-chemokine ligand 5 (CCL5) protein levels in response to LPS stimulation than the WT controls. As macrophages play a key role in sepsis, the in vivo relevance of KSRP deficiency for cytokine/chemokine production was analyzed in an acute inflammation model. In agreement with our in vitro findings, KSRP-deficient animals showed higher cytokine production upon LPS administration in comparison to WT mice. Taken together, these findings demonstrate that KSRP constitutes an important negative regulator of cytokine expression in macrophages. Full article