Serotype-Dependent Inhibition of Streptococcus pneumoniae Growth by Short-Chain Fatty Acids.

Streptococcus pneumoniae (pneumococcus) is an opportunistic pathogen that can cause severe infectious diseases such as pneumonia, meningitis, and otitis media. Despite the availability of antibiotics and pneumococcal vaccines against some invasive serotypes, pneumococcal infection remains a tremendous clinical challenge due to the increasing frequency of infection by antimicrobial resistant, nonencapsulated, and/or non-vaccine serotype strains. Short-chain fatty acids (SCFAs), which are produced at various mucosal sites in the body, have potent antimicrobial activity, including inhibition of pathogen growth and/or bacterial biofilm formation. In this study, we investigated the antimicrobial activity of SCFAs (acetate, propionate, and butyrate) against various serotypes pneumococci. Propionate generally inhibited the growth of S. pneumoniae serotypes included in the pneumococcal conjugate vaccine (PCV) 13, except for serotypes 3 and 7F, though butyrate and acetate showed no or low inhibition, depending on the serotypes. Of note, butyrate showed strong inhibition against serotype 3, the most prevalent invasive strain since the introduction of the PCV. No SCFAs showed inhibitory effects against serotype 7F. Remarkably, the nonencapsulated pneumococcal strain had more sensitivity to SCFAs than encapsulated parental strains. Taken together, these results suggest that propionate showing the most potent inhibition of pneumococcal growth may be used as an alternative treatment for pneumococcal infection, and that butyrate could be used against serotype 3, which is becoming a serious threat.

Lipoproteins are key immunostimulatory components of Bacillus species for dendritic cell maturation and activation.

Dendritic cells (DCs) play an important role in immunity by sensing and responding to invasive microbes. Bacillus species are rod-shaped sporulating bacteria that include the pathogenic Bacillus cereus and commensal Bacillus subtilis. Although the interaction between DC and these two Bacillus species has been studied, their key structural component that prompts DC activation is poorly understood. Here, we investigated the two Bacillus species in DC activation by whole cells and their representative microbe-associated molecular patterns (MAMPs). MAMPs including lipoteichoic acid (LTA), lipoprotein (LPP), and peptidoglycan (PGN) were purified from the two Bacillus species. Among the MAMPs, LPP from both species most potently induced the maturation and activation of DCs while PGN, but not LTA, moderately stimulated DCs. LPPs from both Bacillus species enhanced the expression of DC maturation markers including CCR7, CD40, CD80, CD83, CD86, CD205, MHC-I, and MHC-II. Among the MAMPs from B. cereus, PGN most considerably lowered the endocytic capacity of DCs implying DC maturation whereas PGN from B. subtilis lowered it to a similar degree to its LPP. Furthermore, DCs sensitized with LPPs from both Bacillus species and PGN from B. subtilis moderately induced TNF-α and IL-6 production. Notably, a combination of MAMPs did not show any synergistic effect on DC activation. Taken together, our results demonstrate that LPP is the key structural component in B. cereus and B. subtilis that leads to DC activation.

Adversarial Data Augmentation and Transfer Net for Scrap Metal Identification Using Laser-Induced Breakdown Spectroscopy Measurement of Standard Reference Materials.

In this study, we propose a transfer learning-based classification model for identifying scrap metal using an augmented training dataset consisting of laser-induced breakdown spectroscopy (LIBS) measurement of standard reference material (SRMs) samples, considering varying experimental setups and environmental conditions. LIBS provides unique spectra for identifying unknown samples without complicated sample preparation. Thus, LIBS systems combined with machine learning methods have been actively studied for industrial applications such as scrap metal recycling. However, in machine learning models, a training set of the used samples may not cover the diversity of the scrap metal encountered in field measurements. Moreover, differences in experimental configuration, where laboratory standards and real samples are analyzed in situ, may lead to a wider gap in the distribution of training and test sets, dramatically reducing the performance of the LIBS-based fast classification system for real samples. To address these challenges, we propose a two-step Aug2Tran model. First, we augment the SRM dataset by synthesizing spectra of unobserved types through attenuation of dominant peaks corresponding to sample composition and generating spectra depending on the target sample using a generative adversarial network. Second, we used the augmented SRM dataset to build a robust real-time classification model with a convolutional neural network, which is further customized for the target scrap metal with limited measurements through transfer learning. For evaluation, SRMs of five representative metal types, including aluminum, copper, iron, stainless steel, and brass, are measured with a typical setup to form the SRM dataset. For testing, scrap metal from actual industrial fields is experimented with three different configurations, resulting in eight different test datasets. The experimental results show that the proposed scheme produces an average classification accuracy of 98.25% for the three experimental conditions, as high as the results of the conventional scheme with three separately trained and executed models. Additionally, the proposed model improves the classification accuracy of arbitrarily shaped static or moving samples with various surface contaminations and compositions, and even for differing ranges of charted intensities and wavelengths. Therefore, the proposed Aug2Tran model can be used as a systematic model for scrap metal classification with generalizability and ease of implementation.

Effect of Ultrasonic Shot Peening and Laser Shock Peening on the Microstructure and Microhardness of IN738LC Alloys.

IN738LC is a conventional-cast Ni-based superalloy intended for power generation and aerospace applications. Typically, ultrasonic shot peening (USP) and laser shock peening (LSP) are utilized to enhance cracking, creep, and fatigue resistance. In this study, the optimal process parameters for USP and LSP were established by observing the microstructure and measuring the microhardness of the near-surface region of IN738LC alloys. The LSP impact region (modification depth) was approximately 2500 µm, which was much higher than the USP impact depth of 600 µm. The observation of the microstructural modification and resulting strengthening mechanism revealed that the build-up of dislocations upon peening with plastic deformation was crucial for alloy strengthening in both methods. In contrast, significant strengthening via γ' shearing was observed only in the USP-treated alloys.

A Phase II/III, Multicenter, Observer-blinded, Randomized, Non-inferiority and Safety, study of typhoid conjugate vaccine (EuTCV) compared to Typbar-TCV® in healthy 6 Months-45 years aged participants.

The typhoid conjugate vaccine (TCV) ensures a long-lasting protective immune response, requires fewer doses and is fit for children under 2 years of age. From Phase I study, EuTCV displayed considerable immunogenicity and reliable safety, thus endorsing further examination in Phase II/III trials. Therefore, a clinical Phase II/III study (NCT04830371) was conducted to evaluate its efficacy in healthy Filipino participants aged 6 months to 45 years through administration of the test vaccine (Arm A, B, and C) or comparator vaccine Typbar-TCV® (Arm D). Sera samples were collected pre-vaccination (Visit 1) and post-vaccination (Visit 4, Day 28) to assess the immunogenicity of EuTCV and Typbar-TCV®. During the study, participants were regularly monitored through scheduled visits to the clinic to report any adverse events associated with the vaccine. For vaccine safety, the proportion of solicited and unsolicited Treatment-Emergent Adverse Events was all comparable between EuTCV and Typbar-TCV® groups. A single dose of EuTCV produced seroconversion in 99.4% of treated participants, with seroconversion rates non-inferior to that of Typbar-TCV®. Batch-to-batch consistency was concluded based on the 90% Confidence Interval of the geometric mean ratio (EuTCV Arm A, B, and C) at Week 4, lying within the equivalence margin of 0.5 to 2.0 for all batches. Results from this Phase II/III clinical trial of EuTCV in healthy volunteers show comparable safety and considerable immunogenicity, compared to Typbar-TCV®, meeting the objectives of this pivotal study. ClinicalTrials.gov registration number: NCT04830371.

Propionate Attenuates Growth of Oral Streptococci through Enhancing Methionine Biosynthesis.

Oral streptococci are considered as an opportunistic pathogen associated with initiation and progression of various oral diseases. However, since the currently-available treatments often accompany adverse effects, alternative strategy is demanded to control streptococci. In the current study, we investigated whether short-chain fatty acids (SCFAs), including sodium acetate (NaA), sodium propionate (NaP), and sodium butyrate (NaB), can inhibit the growth of oral streptococci. Among the tested SCFAs, NaP most potently inhibited the growth of laboratory and clinically isolated strains of Streptococcus gordonii under anaerobic culture conditions. However, the growth inhibitory effect of NaP on six different species of other oral streptococci was different depending on their culture conditions. Metabolic changes such as alteration of methionine biosynthesis can affect bacterial growth. Indeed, NaP enhanced intracellular methionine levels of oral streptococci as well as the mRNA expression level of methionine biosynthesis-related genes. Collectively, these results suggest that NaP has an inhibitory effect on the growth of oral streptococci, which might be due to alteration of methionine biosynthesis. Thus, NaP can be used an effective bacteriostatic agent for the prevention of oral infectious diseases caused by oral streptococci.

Practical Approaches to Apply Ultra-Thick Graphite Anode to High-Energy Lithium-Ion Battery: Carbonization and 3-Dimensionalization.

Lithium-ion batteries with ultra-thick electrodes have high energy density and low manufacturing costs because of the reduction of the inactive materials in the same battery volume. However, the partial usage of the full capacity and the low rate capability are caused by poor ionic and electronic conduction. In this work, the effects of two approaches, such as electrode binder carbonization by heat treatment and 3-dimensionalization by the laser structuring of ultra-thick graphite anodes to lithium-ion batteries for high energy density, are investigated. During the heat treatment, the polyvinylidene fluoride (PVDF) binder is carbonized to form fluorinated graphitic carbons, thereby increasing the number of lithium-ion storage sites and the improvement of the electrode capacity by 14% (420 mAh g-1 and 20 mAh cm-2). Further, the carbonization improves the rate capability by 31% at 0.1 C by simultaneously reducing the ionic and electronic resistances. Furthermore, after the laser structuring of the carbonized electrode, the areal discharge capacity increases to 50% at the increasing current rates, resulting from drastically improved ionic conduction. In addition to the electrochemical characteristics, these two approaches contribute considerably to the fast wetting of the electrolyte into the ultra-thick electrode. The carbonization and laser structuring of the ultra-thick graphite anodes are practical approaches for high-energy batteries to overcome the thickness limitation.

Lactobacillus plantarum Lipoteichoic Acids Possess Strain-Specific Regulatory Effects on the Biofilm Formation of Dental Pathogenic Bacteria.

Bacterial biofilm residing in the oral cavity is closely related to the initiation and persistence of various dental diseases. Previously, we reported the anti-biofilm activity of Lactobacillus plantarum lipoteichoic acid (Lp.LTA) on a representative dental cariogenic pathogen, Streptococcus mutans. Since LTA structure varies in a bacterial strain-specific manner, LTAs from various L. plantarum strains may have differential anti-biofilm activity due to their distinct molecular structures. In the present study, we isolated Lp.LTAs from four different strains of L. plantarum (LRCC 5193, 5194, 5195, and 5310) and compared their anti-biofilm effects on the dental pathogens, including S. mutans, Enterococcus faecalis, and Streptococcus gordonii. All Lp.LTAs similarly inhibited E. faecalis biofilm formation in a dose-dependent manner. However, their effects on S. gordonii and S. mutans biofilm formation were different: LRCC 5310 Lp.LTA most effectively suppressed the biofilm formation of all strains of dental pathogens, while Lp.LTAs from LRCC 5193 and 5194 hardly inhibited or even enhanced the biofilm formation. Furthermore, LRCC 5310 Lp.LTA dramatically reduced the biofilm formation of the dental pathogens on the human dentin slice infection model. Collectively, these results suggest that Lp.LTAs have strain-specific regulatory effects on biofilm formation of dental pathogens and LRCC 5310 Lp.LTA can be used as an effective anti-biofilm agent for the prevention of dental infectious diseases.

Short-chain fatty acids inhibit the biofilm formation of Streptococcus gordonii through negative regulation of competence-stimulating peptide signaling pathway.

Streptococcus gordonii, a Gram-positive commensal bacterium, is an opportunistic pathogen closely related to initiation and progression of various oral diseases, such as periodontitis and dental caries. Its biofilm formation is linked with the development of such diseases by enhanced resistance against antimicrobial treatment or host immunity. In the present study, we investigated the effect of short-chain fatty acids (SCFAs) on the biofilm formation of S. gordonii. SCFAs, including sodium acetate (NaA), sodium propionate (NaP), and sodium butyrate (NaB), showed an effective inhibitory activity on the biofilm formation of S. gordonii without reduction in bacterial growth. SCFAs suppressed S. gordonii biofilm formation at early time points whereas SCFAs did not affect its preformed biofilm. A quorum-sensing system mediated by competence-stimulating peptide (CSP) is known to regulate biofilm formation of streptococci. Interestingly, SCFAs substantially decreased mRNA expression of comD and comE, which are CSP-sensor and its response regulator responsible for CSP pathway, respectively. Although S. gordonii biofilm formation was enhanced by exogenous synthetic CSP treatment, such effect was not observed in the presence of SCFAs. Collectively, these results suggest that SCFAs have an anti-biofilm activity on S. gordonii through inhibiting comD and comE expression which results in negative regulation of CSP quorum-sensing system. SCFAs could be an effective anti-biofilm agent against S. gordonii for the prevention of oral diseases.

Regulation of Bone Cell Differentiation and Activation by Microbe-Associated Molecular Patterns.

Gut microbiota has emerged as an important regulator of bone homeostasis. In particular, the modulation of innate immunity and bone homeostasis is mediated through the interaction between microbe-associated molecular patterns (MAMPs) and the host pattern recognition receptors including Toll-like receptors and nucleotide-binding oligomerization domains. Pathogenic bacteria such as Porphyromonas gingivalis and Staphylococcus aureus tend to induce bone destruction and cause various inflammatory bone diseases including periodontal diseases, osteomyelitis, and septic arthritis. On the other hand, probiotic bacteria such as Lactobacillus and Bifidobacterium species can prevent bone loss. In addition, bacterial metabolites and various secretory molecules such as short chain fatty acids and cyclic nucleotides can also affect bone homeostasis. This review focuses on the regulation of osteoclast and osteoblast by MAMPs including cell wall components and secretory microbial molecules under in vitro and in vivo conditions. MAMPs could be used as potential molecular targets for treating bone-related diseases such as osteoporosis and periodontal diseases.

Immune Responses to Irradiated Pneumococcal Whole Cell Vaccine.

Streptococcus pneumoniae (pneumococcus) can cause respiratory and systemic diseases. Recently, γ-irradiation-inactivated, non-encapsulated, intranasal S. pneumoniae (r-SP) vaccine has been introduced as a novel serotype-independent and cost-effective vaccine. However, the immunogenic mechanism of r-SP is poorly understood. Here, we comparatively investigated the protective immunity and immunogenicity of r-SP to the heat-(h-SP) or formalin-inactivated vaccine (f-SP) without adjuvants. Mice were intranasally immunized with each vaccine three times and then challenged with a lethal dose of S. pneumoniae TIGR4 strain and then subsequently evaluated for their immune responses. Immunization with r-SP elicited modestly higher protection against S. pneumoniae than h-SP or f-SP. Immunization with r-SP enhanced pneumococcal-specific IgA in the nasal wash and IgG in bronchoalveolar lavage fluid. Immunization with r-SP enhanced S. pneumoniae-specific IgG, IgG1, and IgG2b in the serum. r-SP more potently induced the maturation of dendritic cells in the cervical lymph nodes than h-SP or f-SP. Interestingly, populations of follicular helper T cells and IL-4-producing cells were potently increased in cervical lymph nodes of r-SP-immunized mice. Collectively, r-SP could be an effective intranasal, inactivated whole-cell vaccine in that it elicits S. pneumoniae-specific antibody production and follicular helper T cell activation leading to protective immune responses against S. pneumoniae infection.

Streptococcus gordonii: Pathogenesis and Host Response to Its Cell Wall Components.

Streptococcus gordonii, a Gram-positive bacterium, is a commensal bacterium that is commonly found in the skin, oral cavity, and intestine. It is also known as an opportunistic pathogen that can cause local or systemic diseases, such as apical periodontitis and infective endocarditis. S. gordonii, an early colonizer, easily attaches to host tissues, including tooth surfaces and heart valves, forming biofilms. S. gordonii penetrates into root canals and blood streams, subsequently interacting with various host immune and non-immune cells. The cell wall components of S. gordonii, which include lipoteichoic acids, lipoproteins, serine-rich repeat adhesins, peptidoglycans, and cell wall proteins, are recognizable by individual host receptors. They are involved in virulence and immunoregulatory processes causing host inflammatory responses. Therefore, S.gordonii cell wall components act as virulence factors that often progressively develop diseases through overwhelming host responses. This review provides an overview of S. gordonii, and how its cell wall components could contribute to the pathogenesis and development of therapeutic strategies.

Micropatterned conductive hydrogels as multifunctional muscle-mimicking biomaterials: Graphene-incorporated hydrogels directly patterned with femtosecond laser ablation.

Multifunctional biomaterials that can provide physical, electrical, and structural cues to cells and tissues are highly desirable to mimic the important characteristics of native tissues and efficiently modulate cellular behaviors. Especially, electrically conductive biomaterials can efficiently deliver electrical signals to living systems; however, the production of conductive biomaterials presenting multiple cell interactive cues is still a great challenge. In this study, we fabricafed an electrically conductive, mechanically soft, and topographically active hydrogel by micropatterning a graphene oxide (GO)-incorporated polyacrylamide hydrogel (GO/PAAm) with femtosecond laser ablation (FLA) and subsequent chemical reduction. FLA parameters were optimized to efficiently produce distinct line patterns on GO/PAAm hydrogels to induce myoblast alignment and maturation. The line patterns distances (PD) were varied to have different topographies (20-80 µm PD). In vitro studies with C2C12 myoblasts revealed that the micopatterned hydrogels are superior to the unpatterned substrates in inducing myogenesis and myotube alignment. Reduced GO/PAAm with 50 µm PD, i.e., PD50/r(GO/PAAm), showed the best results among the various features for differentiation and myotube alignment. Electrical stimulation of myoblasts on the micropatterned conductive hydrogels further promoted the differentiation of myoblasts. In vivo implantation studies indicated good tissue compatibility of PD50/r(GO/PAAm) samples. Altogether, we successfully demonstrated that the micropatterned r(GO/PAAm) may offer multiple properties capable of positively affecting myoblast responses. This hydrogel may serve as an effective multifunctional biomaterial, which possesses the topography for cell alignment/maturation, mechanical properties of the native skeletal muscle tissue, and desirable electrical conductivity for delivering electrical signals to cells, for various biomedical applications such as muscle tissue scaffolds. STATEMENT OF SIGNIFICANCE: Micropatterned conductive hydrogels were created by polymerization of a graphene oxide-incorporated polyacrylamide hydrogel, micropatterning with femtosecond laser ablation, and chemical reduction, which can mimic important characteristics of native skeletal muscle tissues. The micropatterned conductive hydro-gels promoted myogenesis/alignment, enabled electrical stimulation of myoblasts, and displayed good tissue compatibility, which can therefore serve as a multifunctional biomaterial that is topographically active, mechanically soft, and electrically conductive for delivering multiple cell stimulating signals for potential skeletal muscle tissue engineering applications.

Mapping of cutaneous melanoma by femtosecond laser-induced breakdown spectroscopy.

Surgical excision (Mohs micrographic surgery) is the standard procedure to treat a melanoma, in which an in situ histologic examination of sectioned skin is carried out repeatedly until no cancer cells are detected. The possibility to identify melanoma from the surrounding skin by femtosecond laser-induced breakdown spectroscopy (fs-LIBS) is investigated. For experiments, melanoma induced on a hairless mouse by injection of B16/F10 murine melanoma cell was sampled in the form of frozen tissue sections as in Mohs surgery and analyzed by fs-LIBS (λ = 1030 nm, τ = 550 fs). For analysis, the magnesium signal normalized by carbon intensity was utilized to construct an intensity map around the cancer, including both melanoma and surrounding dermis. The intensity map showed a close match to the optically observed morphological and histological features near the cancer region. The results showed that when incorporated into the existing micrographic surgery procedure, fs-LIBS could be a useful tool for histopathologic interpretation of skin cancer possibly with significant reduction of histologic examination time.

Analysis of major elements in pigmented melanocytic chicken skin using laser-induced breakdown spectroscopy.

The concentration difference of major elements in melanocytic skin with respect to pigmentation level is analysed by laser-induced breakdown spectroscopy (LIBS) to investigate the applicability of LIBS as an in situ feedback tool for selective and complete laser removal of melanocytic skin tissue like nevus. The skin of black silkie chicken which had a characteristic darkly pigmented perifollicular skin surrounded by lightly pigmented extrafollicular skin was used as the sample. The results showed higher LIBS signal intensities of Ca2+ and Mg2+ but lower intensities of Na+ , Cl- and K+ in the perifollicular skin than in the extrafollicular skin, which demonstrated the feasibility to use LIBS as a reliable method to distinguish skin tissues with difference in pigmentation level. Plasma emission of biochemical elements generated with a laser irradiation on melanocytic skin lesion.

Subdermal Flexible Solar Cell Arrays for Powering Medical Electronic Implants.

A subdermally implantable flexible photovoltatic (IPV) device is proposed for supplying sustainable electric power to in vivo medical implants. Electric properties of the implanted IPV device are characterized in live animal models. Feasibility of this strategy is demonstrated by operating a flexible pacemaker with the subdermal IPV device which generates DC electric power of ≈647 µW under the skin.

Elemental analysis of tissue pellets for the differentiation of epidermal lesion and normal skin by laser-induced breakdown spectroscopy.

By laser induced breakdown spectroscopy (LIBS) analysis of epidermal lesion and dermis tissue pellets of hairless mouse, it is shown that Ca intensity in the epidermal lesion is higher than that in dermis, whereas Na and K intensities have an opposite tendency. It is demonstrated that epidermal lesion and normal dermis can be differentiated with high selectivity either by univariate or multivariate analysis of LIBS spectra with an intensity ratio difference by factor of 8 or classification accuracy over 0.995, respectively.

Differentiation of cutaneous melanoma from surrounding skin using laser-induced breakdown spectroscopy.

Laser-induced breakdown spectroscopy (LIBS) has the potential to be used as a surgical tool for simultaneous tissue ablation and elemental analysis of the ablated tissue. LIBS may be used to distinguish melanoma lesions from the surrounding dermis based on the quantitative difference of elements within melanoma lesions. Here, we measured the elements in homogenized pellets and real tissues from excised skin samples of melanoma-implanted mice. In addition, statistical analysis of LIBS spectra using principal component analysis and linear discriminant analysis was performed. Our results showed that this method had high detection sensitivity, highlighting the potential of this tool in clinical applications.

Comprehensive Comparison of Various Techniques for the Analysis of Elemental Distributions in Thin Films: Additional Techniques.

In a recent publication by Abou-Ras et al., various techniques for the analysis of elemental distribution in thin films were compared, using the example of a 2-µm thick Cu(In,Ga)Se2 thin film applied as an absorber material in a solar cell. The authors of this work found that similar relative Ga distributions perpendicular to the substrate across the Cu(In,Ga)Se2 thin film were determined by 18 different techniques, applied on samples from the same identical deposition run. Their spatial and depth resolutions, their measuring speeds, their availabilities, as well as their detection limits were discussed. The present work adds two further techniques to this comparison: laser-induced breakdown spectroscopy and grazing-incidence X-ray fluorescence analysis.

Influence of water content on the ablation of skin with a 532 nm nanosecond Nd:YAG laser.

This work reports that the ablation volume and rate of porcine skin changed significantly with the change of skin water content. Under the same laser irradiation conditions (532 nm Nd:YAG laser, pulse width = 11.5 ns, pulse energy = 1.54 J, beam radius = 0.54 mm), the ablation volume dropped by a factor of 4 as the skin water content decreased from 40 wt. % (native) to 19 wt. % with a change in the ablation rate below and above around 25 wt. %. Based on the ablation characteristics observed by in situ shadowgraph images and the calculated tissue temperatures, it is considered that an explosive rupture by rapid volumetric vaporization of water is responsible for the ablation of the high water content of skin, whereas thermal disintegration of directly irradiated surface layer is responsible for the low water content of skin.