In situ metabolomic analysis of osteonecrosis of the femoral head (ONFH) using MALDI MSI.

Osteonecrosis of the femoral head (ONFH) is a common orthopedic disease characterized by disability and deformity. To better understand ONFH at molecular level and to explore the possibility of early diagnosis, instead of diagnosis based on macroscopic spatial characteristics, a matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI MSI) method was developed for ONFH disease for the first time. The most challenging step for ONFH MSI is to deal with human bone tissues which are much harder than the other biological samples studied by the reported MSI studies. In this work, the MSI sectioning method of hard bone tissues was established using tender acids and a series of test criteria. Small-molecule metabolites, such as lipids and amino acids, were detected in bone sections, realizing the in situ detection of spatial distribution of biometabolites. By comparing the distribution of metabolites from different regions of normal femoral head, ONFH bone tissue (ONBT), and adjacent ONFH bone tissue (ANBT), the whole process of femoral head from normal stage to necrosis was monitored and visualized at molecular level. Moreover, this developed MSI method was used for metabolomics study of ONFH. 72 differential metabolites were identified, suggesting that disturbances in energy metabolism and lipid metabolism affected the normal life activities of osteoblasts and osteoclasts. This study provides new perspectives for future pathological studies of ONFH.

Mapping small metabolite changes after traumatic brain injury using AP-MALDI MSI.

Traumatic brain injury (TBI) is an alteration of brain function caused by a sudden transmission of an external force to the head. The biomechanical impact induces acute and chronic metabolic changes that highly contribute to injury evolution and outcome. TBI heterogeneity calls for approaches allowing the mapping of regional molecular and metabolic changes underpinning disease progression, with mass spectrometry imaging (MSI) as an efficient tool to study the spatial distribution of small metabolites. In this study, we applied an innovative targeted atmospheric pressure-MALDI mass spectrometry imaging (AP-MALDI MSI) approach, starting from an extensive list of metabolites, representative of different metabolic pathways, individually validated on the tissue under analysis with original standards using 2,5-dihydroxybenzoic acid (DHB), to characterize the impact of TBI on regional changes to small metabolites in the brain. Brains from sham and TBI mice obtained 21 days post-injury were analyzed to examine the spatial metabolic profile of small metabolites belonging to different metabolic pathways. By a whole brain analysis, we identified four metabolites (alanine, lysine, histidine, and inosine) with higher abundance in TBI than sham mice. Within the TBI group, lysine, histidine, and inosine were higher in the hemisphere ipsilateral to the biomechanical impact vs. the contralateral one. Images showed a major involvement of the ipsilateral thalamus characterized by the increase of arginine, lysine, histidine, and inosine and a significant reduction of glutamic acid, and N-acetylaspartic acid compared to the contralateral thalamus. These findings indicate high-resolution imaging mass spectrometry as a powerful tool to identify region-specific changes after a TBI to understand the metabolic changes underlying brain injury evolution.

Modelling the rapid detection of Carbapenemase-resistant Klebsiella pneumoniae based on machine learning and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

In this study, 80 carbapenem-resistant Klebsiella pneumoniae (CR-KP) and 160 carbapenem-susceptible Klebsiella pneumoniae (CS-KP) strains detected in the clinic were selected and their matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) peaks were collected. K-means clustering was performed on the MS peak data to obtain the best "feature peaks", and four different machine learning models were built to compare the area under the ROC curve, specificity, sensitivity, test set score, and ten-fold cross-validation score of the models. By adjusting the model parameters, the test efficacy of the model is increased on the basis of reducing model overfitting. The area under the ROC curve of the Random Forest, Support Vector Machine, Logistic Regression, and Xgboost models used in this study are 0.99, 0.97, 0.96, and 0.97, respectively; the model scores on the test set are 0.94, 0.91, 0.90, and 0.93, respectively; and the results of the ten-fold cross-validation are 0.84, 0.81, 0.81, and 0.85, respectively. Based on the machine learning algorithms and MALDI-TOF MS assay data can realize rapid detection of CR-KP, shorten the in-laboratory reporting time, and provide fast and reliable identification results of CR-KP and CS-KP.

The first case of urosepsis caused by Corynebacterium aurimucosum in an immunocompetent patient.

Non-diphtheroid Corynebacterium sepsis is rare and has affected only immunocompromised or particularly predisposed patients so far. We present the first case of urosepsis caused by Corynebacterium aurimucosum in a 67-year-old woman, without any known immunodeficiencies and in absence of any immunosuppressive therapy, admitted to the hospital for fever and acute dyspnea. This work suggests a new approach in evaluating the isolation of Corynebacteria, especially if isolated from blood. In particular, it highlights the potential infectious role of C. aurimucosum (often considered a contaminant and only rarely identified as an etiological agent of infections) and its clinical consequences, detailing also interesting aspects about its microbiological diagnosis and relative therapy and clarifying contrasting data of literature.

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Flavonifractor plautii Bacteremia With Generalized Peritonitis: A Case Report and Literature Review.

Flavonifractor plautii is an obligate anaerobic rod bacterium that is part of the human gut microbiota. We describe a case of bacteremia caused by F. plautii in a mildly immunocompromised patient with acute generalized peritonitis. The patient is an 83-year-old male, with a history of stage III hepatocellular carcinoma 11 months prior, stage I gastric cancer, and cerebral infarction three months prior. He visited the emergency room of our hospital with a chief complaint of right-sided abdominal pain. A partial resection of the colon was performed due to stenosis of the transverse colon. Due to increasing abdominal pain, the patient underwent surgery for acute generalized peritonitis on the 11th postoperative day. F. plautii was detected in blood cultures collected prior to surgery, and the patient was treated with piperacillin/tazobactam 2.25 g four times a day for 11 days. The patient resumed eating and was discharged with no recurrence. This species may also stain gram-negative, and caution should be exercised in reporting results due to the potential impact on initial antimicrobial therapy. Gram staining showed variation in the length of the bacterium, which is considered a characteristic of this species. Appropriate antimicrobial therapy for F. plautii has yet to be established, and further accumulation of cases is needed to understand the resistance mechanism and confirm the effectiveness of different antimicrobials.

A Case Report of a Rare Pulmonary Opportunistic Infection in an Infant Caused by Tsukamurella tyrosinosolvens.

This case report describes the case of a female infant hospitalized for severe pneumonia. During the treatment process, various antibiotics are used to treat and prevent further infection. The child had a weak physical condition, combined with neuroblastoma, paraneoplastic syndrome, and low immune function, leading to Tsukamurella tyrosinosolvens infection. The treatment was eventually abandoned owing to poor prognosis. This study aims to through the medium, dyeing, electron microscope observation, 16s rRNA and high-throughput sequencing investigated the morphological characteristics, staining properties, electron microscope morphology, antibiotic resistance, and genomic characteristics of Tsukamurella tyrosinosolvens. The aim of the study is to provide data reference for clinical laboratory staff in bacteria identification research, and to provide relevant help for clinicians in diagnosis and treatment.

Impact of Ultrafiltration on the Physicochemical Properties of Bovine Lactoferrin: Insights into Molecular Mass, Surface Morphology, and Elemental Composition.

The large-scale isolation of bovine lactoferrin (bLF) typically involves using large amounts of concentrated eluents, which might introduce impurities to the final product. Sometimes, protein pre-concentration is required for the greater accuracy of experimental results. In this research, the supplied bLF sample was subjected to additional ultrafiltration (UF) to eliminate possible small impurities, such as salts and peptides of bLF. Beforehand, the basic characterization of native bLF, including surface-charge properties and the structural sensitivity to the various pH conditions, was performed. The study aimed to evaluate the difference in molecular mass, primary structure, surface morphology, and elemental composition of the protein before and after UF. The research was provided by application of spectroscopic, spectrometric, electrophoretic, and microscopic techniques. The evident changes in the surface morphology of bLF were observed after UF, while the molecular masses of both proteins were comparable. According to MALDI-TOF/MS results, UF had a positive impact on the bLF sample representation, improving the identification parameters, such as sequence coverage and intensity coverage.

RhizoMAP: a comprehensive, nondestructive, and sensitive platform for metabolic imaging of the rhizosphere.

BACKGROUND: Elucidating the intricate structural organization and spatial gradients of biomolecular composition within the rhizosphere is critical to understanding important biogeochemical processes, which include the mechanisms of root-microbe interactions for maintaining sustainable plant ecosystem services. While various analytical methods have been developed to assess the spatial heterogeneity within the rhizosphere, a comprehensive view of the fine distribution of metabolites within the root-soil interface has remained a significant challenge. This is primarily due to the difficulty of maintaining the original spatial organization during sample preparation without compromising its molecular content. RESULTS: In this study, we present a novel approach, RhizoMAP, in which the rhizosphere molecules are imprinted on selected polymer membranes and then spatially profiled using matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI). We enhanced the performance of RhizoMAP by combining the use of two thin (< 20 µm) membranes (polyester and polycarbonate) with distinct MALDI sample preparations. This optimization allowed us to gain insight into the distribution of over 500 different molecules within the rhizosphere of poplar (Populus trichocarpa) grown in rhizoboxes filled with mycorrhizae soil. These two membranes, coupled with three different sample preparation conditions, enabled us to capture the distribution of a wide variety of molecules that included phytohormones, amino acids, sugars, sugar glycosides, polycarboxylic acids components of the Krebs cycle, fatty acids, short aldehydes and ketones, terpenes, volatile organic compounds, fertilizers from the soil, and others. Their spatial distribution varies greatly, with some following root traces, others showing diffusion from roots, some associated with soil particles, and many having distinct hot spots along the plant root or surrounding soil. Moreover, we showed how RhizoMAP can be used to localize the origin of the molecules and molecular transformation during root growth. Finally, we demonstrated the power of RhizoMAP to capture molecular distributions of key metabolites throughout a 20 cm deep rhizosphere. CONCLUSIONS: RhizoMAP is a method that provides nondestructive, untargeted, broad, and sensitive metabolite imaging of root-associated molecules, exudates, and soil organic matter throughout the rhizosphere, as demonstrated in a lab-controlled native soil environment.

Expansion of MALDI-TOF MS database as a strategy for identification of Haemophilus species other than H. influenzae.

OBJECTIVES: This study aimed to evaluate an expanded matrix-assisted laser desorption-ionization-time of flight mass spectrometry (MALDI-TOF MS) database for the identification of Haemophilus species other than H. influenzae (Hi). METHODS: A total of 144 Haemophilus species, cultured from respiratory samples from people (living) with cystic fibrosis, were identified with MALDI-TOF MS and 16S rRNA sequencing. Of these, 99 Haemophilus strains showed >99% similarity with the best matching strain in the National Center for Biotechnology Information (NCBI) database and were assigned to a single Haemophilus subspecies using both MALDI-TOF MS and 16S rRNA sequencing. The MS profiles of a subset of strains (n = 58/99) were added to the Bruker MALDI-TOF MS database. Subsequently, 270 different strains that were analyzed previously in a routine setting were re-analyzed. RESULTS: 16S rRNA sequencing reliably identified 99/144 Haemophilus strains (>99% similarity). H. haemolyticus 16S rRNA identification was suboptimal since only 3/21 H. haemolyticus strains attained a similarity of >99% with H. haemolyticus 16S rRNA sequence in the NCBI database. Expansion of the MALDI-TOF MS database improved the number of reliable identifications only moderately for H. haemolyticus, H. influenzae and H. paraphrohaemolyticus (<10%). By contrast, improved identification was more outspoken for H. parahaemolyticus, H. parainfluenzae, H. sputorum and H. pittmaniae (>85%). CONCLUSION: 16S rRNA sequencing is a valuable method for the identification of Haemophilus sp. other than Hi. Expansion of the MALDI-TOF MS database, based on 16S rRNA sequencing results, increased the proportion of reliable identifications and in this study resulted in an increase of 10% of Haemophilus sp. other than Hi strain identifications.

Candida albicans in the oral cavities of pets: biofilm formation, putative virulence, antifungal resistance profiles and classification of the isolates.

The study aimed to investigate Candida albicans presence, antifungal resistance, biofilm formation, putative virulence genes, and molecular characterization in oral samples of dogs and cats. A total of 239 oral samples were collected from cats and dogs of various breeds and ages at Erciyes University, Faculty of Veterinary Medicine Clinics, between May 2017 and April 2018. Among 216 isolates obtained, 15 (6.95%) were identified as C. albicans, while 8 (3.7%) were non-albicans Candida species. Antifungal susceptibility testing revealed sensitivities to caspofungin, fluconazole, and flucytosine in varying proportions. Molecular analysis indicated the presence of fluconazole and caspofungin resistance genes in all C. albicans isolates. Additionally, virulence genes ALS1, HWP1, and HSP90 showed variable presence. Biofilm formation varied among isolates, with 46.7% strong, 33.3% moderate, and 20% weak producers. PCA analysis categorized isolates into two main clusters, with some dog isolates grouped separately. The findings underscore the significance of oral care and protective measures in pets due to C. albicans prevalence, biofilm formation, virulence factors, and antifungal resistance in their oral cavity, thereby aiding clinical diagnosis and treatment in veterinary medicine.

Effect of dynamic exclusion and the use of FAIMS, DIA and MALDI-mass spectrometry imaging with ion mobility on amyloid protein identification.

Amyloidosis is a disease characterized by local and systemic extracellular deposition of amyloid protein fibrils where its excessive accumulation in tissues and resistance to degradation can lead to organ failure. Diagnosis is challenging because of approximately 36 different amyloid protein subtypes. Imaging methods like immunohistochemistry and the use of Congo red staining of amyloid proteins for laser capture microdissection combined with liquid chromatography tandem mass spectrometry (LMD/LC-MS/MS) are two diagnostic methods currently used depending on the expertise of the pathology laboratory. Here, we demonstrate a streamlined in situ amyloid peptide spatial mapping by Matrix Assisted Laser Desorption Ionization-Mass Spectrometry Imaging (MALDI-MSI) combined with Trapped Ion Mobility Spectrometry for potential transthyretin (ATTR) amyloidosis subtyping. While we utilized the standard LMD/LC-MS/MS workflow for amyloid subtyping of 31 specimens from different organs, we also evaluated the potential introduction in the MS workflow variations in data acquisition parameters like dynamic exclusion, or testing Data Dependent Acquisition combined with High-Field Asymmetric Waveform Ion Mobility Spectrometry (DDA FAIMS) versus Data Independent Acquisition (DIA) for enhanced amyloid protein identification at shorter acquisition times. We also demonstrate the use of Mascot's Error Tolerant Search and PEAKS de novo sequencing for the sequence variant analysis of amyloidosis specimens.

Formic acid sandwich method is well-suited for filamentous fungi identification and improves turn around time using Zybio EXS2600 mass spectrometry.

OBJECTIVES: Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is extensively employed for the identification of filamentous fungi on MALDI Biotyper (Bruker Daltonics) and Vitek MS (biomerieux), but the performance of fungi identification on new EXS2600 (Zybio) is still unknow. Our study aims to evaluate the new EXS2600 system's (Zybio) ability to rapidly identify filamentous fungi and determine its effect on turnaround time (TAT) in our laboratory. METHODS: We tested 117 filamentous fungi using two pretreatment methods: the formic acid sandwich (FA-sandwich) and a commercial mold extraction kit (MEK, Zybio). All isolates were confirmed via sequence analysis. Laboratory data were extracted from our laboratory information system over two 9-month periods: pre-EXS (April to December 2022) and post-EXS (April to December 2023), respectively. RESULTS: The total correct identification (at the species, genus, or complex/group level) rate of fungi was high, FA-sandwich (95.73%, 112/117), followed by MEK (94.02%, 110/117). Excluding 6 isolates not in the database, species-level identification accuracy was 92.79% (103/111) for FA-sandwich and 91.89% (102/111) for MEK; genus-level accuracy was 97.29% (108/111) and 96.39% (107/111), respectively. Both methods attained a 100% correct identification rate for Aspergillus, Lichtheimia, Rhizopus Mucor and Talaromyces species, and were able to differentiate between Fusarium verticillioides and Fusarium proliferatum within the Fusarium fujikuroi species complex. Notably, high confidence was observed in the species-level identification of uncommon fungi such as Trichothecium roseum and Geotrichum candidum. The TAT for all positive cultures decreased from pre EXS2600 to post (108.379 VS 102.438, P < 0.05), and the TAT for tissue decreased most (451.538 VS 222.304, P < 0.001). CONCLUSIONS: The FA-sandwich method is more efficient and accurate for identifying filamentous fungi with EXS2600 than the MEK. Our study firstly evaluated the performance of fungi identification on EXS2600 and showed it is suitable for clinical microbiology laboratories use.

Assessing earthworm exposure to a multi-pharmaceutical mixture in soil: unveiling insights through LC-MS and MALDI-MS analyses, and impact of biochar on pharmaceutical bioavailability.

In the European circular economy, agricultural practices introduce pharmaceutical (PhAC) residues into the terrestrial environment, posing a potential risk to earthworms. This study aimed to assess earthworm bioaccumulation factors (BAFs), the ecotoxicological effects of PhACs, the impact of biochar on PhAC bioavailability to earthworms, and their persistence in soil and investigate earthworm uptake mechanisms along with the spatial distribution of PhACs. Therefore, earthworms were exposed to contaminated soil for 21 days. The results revealed that BAFs ranged from 0.0216 to 0.329, with no significant ecotoxicological effects on earthworm weight or mortality (p > 0.05). Biochar significantly influenced the uptake of 14 PhACs on the first day (p < 0.05), with diminishing effects over time, and affected significantly the soil-degradation kinetics of 16 PhACs. Moreover, MALDI-MS analysis revealed that PhAC uptake occurs through both the dermal and oral pathways, as pharmaceuticals were distributed throughout the entire earthworm tissue without specific localization. In conclusion, this study suggests ineffective PhAC accumulation in earthworms, highlights the influence of biochar on PhAC degradation rates in soil, and suggests that uptake can occur through both earthworm skin and oral ingestion.

Genotyping and Phenotyping of Indigenous Saccharomyces cerevisiae from a New Zealand Organic Winery and Commercial Sources Using Inter-Delta and MALDI-TOF MS Typing.

We used inter-delta typing (IDT) and MALDI-TOF profiling to characterize the genetic and phenotypic diversity of 45 commercially available winemaking Saccharomyces cerevisiae strains and 60 isolates from an organic winemaker from Waipara, New Zealand, as a stratified approach for predicting the commercial potential of indigenous isolates. A total of 35 IDTs were identified from the commercial strains, with another 17 novel types defined among the Waipara isolates. IDT 3 was a common type among strains associated with champagne production, and the only type in commercial strains also observed in indigenous isolates. MALDI-TOF MS also demonstrated its potential in S. cerevisiae typing, particularly when the high-mass region (m/z 2000-20,000) was used, with most indigenous strains from each of two fermentation systems distinguished. Furthermore, the comparison between commercial strains and indigenous isolates assigned to IDT 3 revealed a correlation between the low-mass data (m/z 500-4000) analysis and the recommended use of commercial winemaking strains. Both IDT and MALDI-TOF analyses offer useful insights into the genotypic and phenotypic diversity of S. cerevisiae, with MALDI-TOF offering potential advantages for the prediction of applications for novel, locally isolated strains that may be valuable for product development and diversification.

Application of MALDI-TOF MS to Identify and Detect Antimicrobial-Resistant Streptococcus uberis Associated with Bovine Mastitis.

Streptococcus uberis is a common bovine mastitis pathogen in dairy cattle. The rapid identification and characterization of antimicrobial resistance (AMR) in S. uberis plays an important role in its diagnosis, treatment, and prevention. In this study, matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) was used to identify S. uberis and screen for potential AMR biomarkers. Streptococcus uberis strains (n = 220) associated with bovine mastitis in northern Thailand were identified using the conventional microbiological methods and compared with the results obtained from MALDI-TOF MS. Streptococcus uberis isolates were also examined for antimicrobial susceptibility using a microdilution method. Principal component analysis (PCA) and the Mann-Whitney U test were used to analyze the MALDI-TOF mass spectrum of S. uberis and determine the difference between antimicrobial-resistant and -susceptible strains. Using MALDI-TOF MS, 73.18% (161/220) of the sampled isolates were identified as S. uberis, which conformed to the identifications obtained using conventional microbiological methods and PCR. Using PCR, antimicrobial-resistant strains could not be distinguished from antimicrobial-susceptible strains for all three antimicrobial agents, i.e., tetracycline, ceftiofur, and erythromycin. The detection of spectral peaks at 7531.20 m/z and 6804.74 m/z was statistically different between tetracycline- and erythromycin-resistant and susceptible strains, respectively. This study demonstrates a proteomic approach for the diagnosis of bovine mastitis and potentially for the surveillance of AMR among bovine mastitis pathogens.

Comparison of Culture-Dependent and Culture-Independent Methods for Routine Identification of Airborne Microorganisms in Speleotherapeutic Caves.

The effective identification of bacterial and fungal isolates is essential for microbiological monitoring in environments like speleotherapeutic caves. This study compares MALDI-TOF MS and the OmniLog ID System, two high-throughput culture-based identification methods. MALDI-TOF MS identified 80.0% of bacterial isolates to the species level, while the OmniLog ID System identified 92.9%. However, species-level matches between the methods were only 48.8%, revealing considerable discrepancies. For discrepant results, MALDI-TOF MS matched molecular identification at the genus level in 90.5% of cases, while the OmniLog ID System matched only in 28.6%, demonstrating MALDI-TOF MS's superiority. The OmniLog ID System had difficulties identifying genera from the order Micrococcales. Fungal identification success with MALDI-TOF MS was 30.6% at the species level, potentially improvable with a customised spectral library, compared to the OmniLog ID System's 16.7%. Metagenomic approaches detected around 100 times more microbial taxa than culture-based methods, highlighting human-associated microorganisms, especially Staphylococcus spp. In addition to Staphylococcus spp. and Micrococcus spp. as indicators of cave anthropisation, metagenomics revealed another indicator, Cutibacterium acnes. This study advocates a multi-method approach combining MALDI-TOF MS, the OmniLog ID System, culture-based, and metagenomic analyses for comprehensive microbial identification. Metagenomic sampling on nitrocellulose filters provided superior read quality and microbial representation over liquid sampling, making it preferable for cave air sample collection.

Antimicrobial Resistance of Enterococcus Isolates from Poultry Farms in the Republic of Serbia (Autonomous Province of Vojvodina).

Enterococcus species are significant intestinal commensals of animals, including poultry. However, they have emerged as important opportunistic infective agents in both veterinary and human medicine as well as major nosocomial pathogens, owing to their increasing antimicrobial resistance. This research aimed to investigate the prevalence and antimicrobial resistance patterns of Enterococcus spp. isolated from poultry farms in the north of Serbia. A total of 40 samples of overshoes or feces were collected from 40 poultry farms and analyzed for the presence of Enterococcus spp. using PCR or MALDI-TOF mass spectrometry for their identification. The number of isolates was 40 and included 11 isolates from laying hens, 2 isolates from turkeys, 3 from broiler breeders, and 24 from broilers. The Kirby-Bauer disk diffusion method was used to test for antibiotic susceptibility in accordance with the Clinical and Laboratory Standards Institute and EUCAST guidelines. The results showed that Enterococcus faecalis was isolated from 37.5% farms, and E. faecium from 42.5%. E. hirae was identified in 15% of poultry establishments, and E. durans and E. thialandicus on 2.5%. Notably, resistance to erythromycin, streptomycin, fluoroquinolones, and tetracyclines among the frequently used antibiotics was found. Furthermore, 35% of the isolates had multidrug resistance (MDR). In order to prevent the spread of antibiotic resistance in chicken farming and protect the health of the public and animals alike, our findings highlight the critical need for improved surveillance and control measures. To effectively establish a containment strategy for Enterococcus spp. isolated from poultry farms, more research into the processes behind their antibiotic resistance is required.

Mass Spectrometry Imaging Combined with Sparse Autoencoder Method Reveals Altered Phosphorylcholine Distribution in Imipramine Treated Wild-Type Mice Brains.

Mass spectrometry imaging (MSI) is essential for visualizing drug distribution, metabolites, and significant biomolecules in pharmacokinetic studies. This study mainly focuses on imipramine, a tricyclic antidepressant that affects endogenous metabolite concentrations. The aim was to use atmospheric pressure matrix-assisted laser desorption/ionization (AP-MALDI)-MSI combined with different dimensionality reduction methods to examine the distribution and impact of imipramine on endogenous metabolites in the brains of treated wild-type mice. Brain sections from both control and imipramine-treated mice underwent AP-MALDI-MSI. Dimensionality reduction methods, including principal component analysis, multivariate curve resolution, and sparse autoencoder (SAE), were employed to extract valuable information from the MSI data. Only the SAE method identified phosphorylcholine (ChoP) as a potential marker distinguishing between the control and treated mice brains. Additionally, a significant decrease in ChoP accumulation was observed in the cerebellum, hypothalamus, thalamus, midbrain, caudate putamen, and striatum ventral regions of the treated mice brains. The application of dimensionality reduction methods, particularly the SAE method, to the AP-MALDI-MSI data is a novel approach for peak selection in AP-MALDI-MSI data analysis. This study revealed a significant decrease in ChoP in imipramine-treated mice brains.

Characteristics of Umami Taste of Soy Sauce Using Electronic Tongue, Amino Acid Analyzer, and MALDI-TOF MS.

The objective of this study was to investigate the umami characteristics of soy sauce using electronic tongue evaluation and amino acid composition and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) analysis. The soy sauce peptides were isolated from soy sauce using XAD-16 macroporous resin combined with ethanol solution. The results showed that the soy sauce peptide fraction eluted by 60% ethanol (SS-60%) exhibited a prominent umami taste, and the umami scores were highly positively correlated with the amino acid nitrogen contents of soy sauces. The umami scores of SS-60% were significantly positively correlated with the contents of free amino acids. Especially, Phe showed the highest positive correlation with the umami scores. In addition, five characteristic ion peaks with m/z at 499, 561, 643, 649, and 855 were identified in the peptide mass fingerprinting. Therefore, this study provides new insights into the umami characteristics for the taste evaluation and reality identification of soy sauce.

Occurrence of Enterococci in the Process of Artisanal Cheesemaking and Their Antimicrobial Resistance.

Enterococci are a group of microorganisms that have a controversial position from some scientific points of view. The species of the greatest clinical importance are E. faecalis and E. faecium, which are common agents of nosocomial infections. However, enterococci also have important applications in the dairy industry, as they are used as non-starter lactic acid bacteria (NSLAB) in a variety of cheeses, especially artisanal cheeses. The aim of this study was to determine the presence of representatives from the Enterococcus genus using PCR and MALDI-TOF MS methods on samples of raw milk, processing environment swabs, and cheese from four different artisanal dairy plants in Slovakia. Among the 136 isolates of enterococci, 9 species of genus Enterococci (E. faecalis, E. faecium, E. durans, E. devriesi, E. hirae, E. italicus, E. casseliflavus, E. malodoratus, and E. gallinarum) were identified and were tested for their antimicrobial resistance (AMR) to 8 antibiotics (amoxicillin, penicillin, ampicillin, erythromycin, levofloxacin, vancomycin, rifampicin, and tetracycline); most of them were resistant to rifampicin (35.3%), ampicillin (22.8%), and tetracycline (19.9%). A PCR analysis of vanA (4.41%) and tetM (14.71%) revealed that antimicrobial resistance genes were present in not only phenotypic resistant isolates of enterococci but also susceptible isolates. The investigation of antimicrobial resistance in enterococci during the cheesemaking process can be a source of valuable information for public health in the concept of "One Health".