Laser Desorption Ionization-time Of Flight Research Articles

Inanimate Surfaces and Air Contamination with Multidrug Resistant Species of Staphylococcus in the Neonatal Intensive Care Unit Environment

Background: Contamination of the hospital environment with multi-resistant (MDR) Staphylococcus increases the risk of infection. The aim of this study is to identify the MDR species of Staphylococcus on inanimate surfaces, in air, and in clinical samples, and analyze the risk factors that correlate with the occurrence of infections in a Neonatal Intensive Care Unit. Methods: Samples of inanimate surfaces and air were taken using a premoistened swab (0.9% sodium chloride) and spontaneous air sedimentation, respectively. The clinical isolates were recovered from infected neonates. The isolates (environmental and clinical) were identified by matrix-assisted laser desorption ionization-time of flight and the resistance profile was calculated using the disk diffusion agar technique. Results: In total, 181 isolates were obtained, 93 from (surfaces), 18 from the air, and 70 clinical samples. S. epidermidis was the most frequent species (66.8%), and the failure rate in air cleaning was 100%. More than 60% of the isolates were MDR, and the majority of clinical isolates (60.4%) had a resistance profile identical to that of the environmental isolates. Conclusion: Staphylococcus spp. were found in most of the analyzed samples, with a high frequency of MDR isolates, demonstrating the importance of the hospital environment as a reservoir, and the need for infection control measures, and rational use of antimicrobials.

Genetic Characterization of Antibiotic Resistant Enterobacteriaceae Isolates From Bovine Animals and the Environment in Nigeria.

There is a link between antibiotic resistance in humans, livestock and the environment. This study was carried out to characterize antibiotic resistant bovine and environmental Enterobacteriaceae isolates from Edo state, Nigeria. A total of 109 consecutive isolates of Enterobacteriaceae were isolated from March–May 2015 from 150 fecal samples of healthy bovine animals from three farms at slaughter in Edo state Nigeria. Similarly, 43 Enterobacteriaceae isolates were also obtained from a total of 100 environmental samples from different sources. Isolates were recovered and identified from samples using standard microbiological techniques. Recovered isolates were pre-identified by the Microbact Gram-Negative identification system and confirmed with Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry and ribosomal multilocus sequence typing (rMLST). Antibiotic susceptibility testing was carried out by Kirby-Bauer method for 14 antibiotics. Whole genome sequencing (WGS) was carried out for isolate characterization and identification of resistance determinants. Out of 109 animal and 43 environmental Enterobacteriaceae isolates, 18 (17%) and 8 (19%) isolates based on selection criteria showed antibiotic resistance and were further investigated by whole genome sequencing (WGS). Resistance genes were detected in all (100%) of the resistant bovine and environmental Enterobacteriaceae isolates. The resistance determinants included β-lactamase genes, aminoglycoside modifying enzymes, qnr genes, sulfonamide, tetracycline and trimethoprim resistance genes, respectively. Out of the 18 and 8 resistant animal and environmental isolates 3 (17%) and 2 (25%) were multidrug resistant (MDR) and had resistance determinants which included efflux genes, regulatory systems modulating antibiotic efflux and antibiotic target alteration genes. Our study shows the dissemination of antibiotic resistance especially MDR strains among Nigerian bovine and environmental Enterobacteriaceae isolates. The presence of these resistant strains in animals and the environment constitute a serious health concern indicated by the difficult treatment options of the infections caused by these organisms. To the best of our knowledge we report the first detailed genomic characterization of antibiotic resistance in bovine and environmental Enterobacteriaceae isolates for Nigeria.

The Association of ERCC1 and ERCC5 Polymorphisms with Lung Cancer Risk in Han Chinese.

Background: Polymorphisms in DNA damage repair genes are important determinants for cancer susceptibility, clinical phenotype diversity, and therapy. However, their relationship with lung cancer remains unclear. This study aimed to investigate the role of DNA damage repair gene polymorphisms in the risk of lung cancer.Methods: The matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectroscopy-based genotyping system was used to genotype 601 individuals (200 lung cancer patients and 401 age- and sex-matched healthy controls) for polymorphisms in excision repair cross-complementing group 1 (ERCC1) and ERCC5 genes.Results: The ERCC5 rs4771436 GG genotype, recessive model (GG vs. GT+TT), and the ERCC5 rs1047768 recessive model (CC vs. CT+TT) were associated with significantly increased risks of lung cancer (P=0.029, P=0.014, and P=0.044, respectively), especially in men and individuals aged 60 years or younger.Conclusion: ERCC5 rs4771436 and rs1047768 genotypes were associated with an increased risk of lung cancer, suggesting that polymorphisms in DNA repair genes are significantly related to the risk of lung cancer, and play an important role in the occurrence of lung cancer.

Biosafety and Proteome Profiles of Different Heat Inactivation Methods for Mycobacterium tuberculosis.

ABSTRACTStudies involving the pathogenic organism Mycobacterium tuberculosis routinely require advanced biosafety laboratory facilities, which might not be readily available in rural areas where tuberculosis burdens are high. Attempts to adapt heat inactivation techniques have led to inconsistent conclusions, and the risk of protein denaturation due to extensive heating is impractical for subsequent mass spectrometry (MS)-based protein analyses. In this study, 240 specimens with one or two loops of M. tuberculosis strain H37Rv biomass and specific inactivated solutions were proportionally assigned to six heat inactivation methods in a thermal block at 80°C and 95°C for 20, 30, and 90 min. Twenty untreated specimens served as a positive control, and bacterial growth was followed up for 12 weeks. Our results showed that 90 min of heat inactivation was necessary for samples with two loops of biomass. Further protein extraction and a matrix-assisted laser desorption ionization–time of flight (MALDI-TOF) MS assay demonstrated adequate scores for bacterial identification (≥1.7), with the highest score achieved in the 80°C/90 min and 95°C/30 min treatment groups. A proteomics study also confidently identified 648 proteins with ∼93% to 96% consistent protein abundances following heating at 95°C for 20, 30, and 90 min. Heat inactivation at 95°C for 90 min yielded the most quantifiable proteins, and a functional analysis revealed proteins located in the ribosomal subunit. In summary, we proposed a heat inactivation method for the M. tuberculosis strain H37Rv and studied the preservation of protein components for subsequent bacterial identification and protein-related assays.IMPORTANCE Inactivation of Mycobacterium tuberculosis is an important step to guarantee biosafety for subsequent M. tuberculosis identification and related research, notably in areas of endemicity with minimal resources. However, certain biomolecules might be denatured or hydrolyzed because of the harsh inactivation process, and a standardized protocol is yet to be determined. We evaluated distinct heating conditions to report the inactivation efficiency and performed downstream mass spectrometry-based M. tuberculosis identification and proteomics study. The results are important and useful for both basic and clinical M. tuberculosis studies.

1234. Can Susceptibility to One Carbapenem be Conferred to Another? Frequency of Discordance in Gram-negative Clinical Isolates

BackgroundCarbapenem-Resistant Enterobacterales (CRE) and Carbapenem-Resistant Pseudomonas aeruginosa (CRPA) can exhibit resistance to one carbapenem while remaining susceptible to another. While case reports describing discrepant carbapenem susceptibilities are available, the authors are unaware of any literature reporting aggregate carbapenem susceptibility discrepancies at a hospital level. MethodsSusceptibility data from April 1, 2017 - December 31, 2017 was extracted through an antibiogram report for a 706-bed hospital. Ertapenem, imipenem-cilastatin, and meropenem susceptibilities were captured and compared for common Enterobacterales and Pseudomonas aeruginosa. Organism identification was performed using Matrix Assisted Laser Desorption Ionization-Time of Flight (MALDI-TOF) mass spectrometry. Antibiotic susceptibility testing was performed using BD PhoenixTM. Carbapenem susceptibilities were interpreted using the most updated Clinical and Laboratory Standards Institute (CLSI) breakpoints at the time of assessment (2021). Carbapenem discordance was defined as an organism being susceptible to one carbapenem and non- susceptible (intermediate or resistant) to another. Approval was obtained from the institution’s Institutional Review Board.ResultsMeropenem proved to be the most active antimicrobial for all organisms (Figure 1). Carbapenem susceptibility discordance ranged from 0%-23.8% (Table 1). There was a significant difference in the incidence of discordance between Enterobacterales and Pseudomonas aeruginosa isolates (2.6% vs. 6.1%, p < 0.001). Of the 20 Pseudomonas aeruginosa isolates with discordant carbapenem susceptibilities, 70% were meropenem susceptible/imipenem non-susceptible and 30% were imipenem susceptible/meropenem non-susceptible. The most common site for discordance was urine for both Enterobacterales and Pseudomonas aeruginosa. However, while there was a significant rate of discordance between sites for Pseudomonas isolates, this was not the case for Enterobacterales (Table 2).Figure 1: Carbapenem Susceptibility by Isolate Table 1: Frequency of Carbapenem Discordance Table 2: Frequency of Carbapenem Discordance by Site ConclusionDue to the wide range of susceptibility discordance, clinical implications can be drastic if an institution is relying on susceptibility of one carbapenem to confer susceptibility to another carbapenem.Disclosures All Authors: No reported disclosures

Simultaneous Production of Multiple Antimicrobial Compounds by Bacillus velezensis ML122-2 Isolated From Assam Tea Leaf [Camellia sinensis var. assamica (J.W.Mast.) Kitam.

Bacillus velezensis ML122-2 is an antimicrobial-producing strain isolated from the leaf of Assam tea or Miang [Camellia sinensis var. assamica (J.W.Mast.) Kitam.]. The cell-free supernatant (CFS) of strain ML122-2 exhibits a broad-spectrum antimicrobial activity against various Gram-positive and Gram-negative bacteria as well as the mold Penicillium expansum. The genome of B. velezensis ML122-2 was sequenced and in silico analysis identified three potential bacteriocin-associated gene clusters, that is, those involved in the production of mersacidin, amylocyclicin, and LCI. Furthermore, six gene clusters exhibiting homology (75–100% DNA sequence identity) to those associated with the secondary metabolites bacilysin, bacillibactin, surfactin, macrolactin H, bacillaene, and plipastatin were identified. Individual antimicrobial activities produced by B. velezensis ML122-2 were purified and characterized by Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry analysis, revealing three antimicrobial peptides with molecular masses corresponding to surfactin, plipastatin, and amylocyclicin. Transcriptional analysis of specific genes associated with mersacidin (mrsA), amylocyclicin (acnA), plipastatin (ppsA), and surfactin (srfAA) production by B. velezensis ML122-2 showed that the first was not transcribed under the conditions tested, while the latter three were consistent with the presence of the associated peptides as determined by mass spectrometry analysis. These findings demonstrate that B. velezensis ML122-2 has the genetic capacity to produce a wide range of antimicrobial activities that may support a specific community structure and highlight the biotechnological properties of Assam tea.

Residual Monoclonal Free Light Chain Positivity By Mass Spectrometry Identifies Patients at Increased Risk of Early Relapse Following First-Line Anti-Myeloma Treatment

Residual Monoclonal Free Light Chain Positivity By Mass Spectrometry Identifies Patients at Increased Risk of Early Relapse Following First-Line Anti-Myeloma Treatment

Quantitative analysis of galactose using LDI-TOF MS based on a TiO2 nanowire chip

A novel method for quantifying galactose was developed to serve as a newborn screening test for galactosemia using laser desorption/ionization time-of-flight (LDI-TOF) mass spectrometry (MS) with a TiO2 nanowire chip. Herein, phosphate citrate buffer, serum, and dried blood spot (DBS) were employed for the quantitative analysis of galactose. To quantitatively analyze galactose, its reduction potential was used to oxidize o-phenylene diamine (OPD) into 2,3-diaminophenazine (DA), which were both detected using LDI-TOF MS with a TiO2 nanowire chip according to the concentration of galactose. The reproducibility and the interference of glucose were determined to demonstrate the applicability of this method. Moreover, mixtures of galactose, phenylalanine, and 17 α-OHP were analyzed to determine the interference induced by other biomarkers of metabolic disorders. The OPD oxidation of galactose was found to be selectively achieved under high-glucose conditions, similar to human blood, thereby showing good reproducibility. The intensities of the mass peaks of OPD and DA based on LDI-TOF MS with a TiO2 nanowire chip were linearly correlated in the galactose concentration range of 57.2–220.0 μg/mL (r2 = 0.999 and 0.950, respectively) for serum samples and 52.5–220.0 μg/mL (r2 = 0.993 and 0.985, respectively) for DBS after methanol precipitation/extraction. The enzyme immunoassay and LDI-TOF MS analysis results were statistically analyzed, and a mixture of phenylalanine, 17 α-OHP, and galactose was simultaneously investigated quantitatively at the cutoff level.

Determination of Uropathogenic Escherichia coli in Urine by an Immunobiosensor Based Upon Antigen-Antibody Biorecognition with Fluorescence Detection and Bead-Injection Analysis

Uroinfections, which are mainly caused by pathogenic bacteria, especially uro-pathogenic E. coli (UPEC), are among the major health problems in the modern world. The current gold standard for the identification of uroinfection causing pathogens in urine is microbiological cultivation, but uro-pathogens are also determined with quantitative polymerase chain reaction (qPCR) and matrix assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry. However, it takes several hours or days to obtain the results. A promising option for rapid identification of pathogens is the application of biosensors. The aim of the current study was to develop a biosensor for the fast detection of UPEC in urine samples allowing achieving a limit of detection below 103 CFU/mL, essential for the diagnostics of recurrent uroinfections and related diseases. First, the pathogens were attached onto a single use column and specifically detected using E. coli antibodies conjugated with a fluorescence marker. The bead-injection analysis platform for fluidics provided E. coli limits of detection and quantification in 150 µL urine samples <3 and <5 cells/mL, respectively. The time of analysis was 17 min. The biosensor results were in good correlation with the results obtained with other methods, indicating that the complex urine matrix of UPEC patients did not affect the biosensor measurements.

Alternating copolymerization of CO2 and cyclohexene oxide initiated by rare-earth metal complexes stabilized by o-phenylenediamine-bridged tris(phenolate) ligand

A series of o-phenylenediamine bridged tris(phenolate) ligand-stabilized rare-earth metal complexes were synthesized and characterized. Lanthanum (complex 1), neodymium (complex 2tBu), and yttrium (complex 3) complexes stabilized by ligand of bulky tert-butyl substituents are mononuclear, whereas neodymium complexes 2Me and 2Cl bearing smaller methyl and chloro substituents are dinuclear, respectively. They were applied in the alternating copolymerization of CO2 and cyclohexene oxide. The addition of benzyl alcohol is beneficial to improving yields (up to 86%) and selectivity of poly(carbonate) (up to 99%). Stoichiometric reaction of benzyl alcohol with rare-earth metal complexes 1 and 2tBu results in formation of complexes 4 and 5 as dinuclear complexes with benzyl alcohol coordination, which was confirmed via single crystal X-ray diffraction analysis, and nuclear magnetic resonance (NMR) spectroscopy (for 4). Complex 5 shows equally good activity and selectivity of that of complex 2tBu and benzyl alcohol, consolidating that complex 5 is indeed generated during polymerization. The neodymium tris(phenolate) 5 is one of the few examples of rare-earth metal complex as a single-component initiator for copolymerization of epoxide and CO2. Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry analysis proves benzyloxy group as the chain end. 1H NMR monitoring suggests that the insertion of CO2 may be the first step in the growth of the polymer chain.

Dysgonomonas capnocytophagoides Bacteremia in a Patient With Stage IV Colon Adenocarcinoma

Dysgonomonas capnocytophagoides bacteremia is a rare clinical entity described in only five case reports. Difficulties in the identification and intrinsic multidrug resistance (MDR) of the organism make diagnosis and treatment challenging. We present a case of D. capnocytophagoides bacteremia which highlights the diagnostic and treatment challenges posed by this organism. The case also contributes to the nascent understanding of the clinical profile of patients with D. capnocytophagoides infection and the antimicrobial susceptibility of the organism.A 56-year-old male with advanced colon adenocarcinoma on palliative fluorouracil, leucovorin, and irinotecan (FOLFIRI) presented with abdominal pain. He had been discharged recently following an ICU admission for neutropenic fever with diarrhea and polymicrobial bacteremia resulting in sepsis. Diarrhea resolved during hospitalization. Mediport was retained, surveillance blood cultures remained negative, and he completed 14 days of levofloxacin. Upon readmission for abdominal pain, vital signs were normal and neutropenia had resolved. A Gram-negative rod grew from blood cultures drawn peripherally and from the port with no differential time-to-positivity. Multiple testing platforms were used in an attempt to identify the organism, to include the VERIGENE® Gram-negative blood culture test, matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry, VITEK ® 2 GN ID, and the Thermo Scientific™ RapID™ NH System (Thermo Scientific, Waltham, MA). Test results from all platforms were either inconclusive or contradictory in their identification of the organism, making the determination of appropriate treatment difficult. Given inconsistent results on multiple testing platforms, the isolate was sent for whole genome sequencing (WGS). Additional workup performed during the hospitalization included a diagnostic paracentesis without evidence of spontaneous bacterial peritonitis, transesophageal echocardiogram without evidence of infective endocarditis, and dental evaluation without evidence of the infectious source. Abdominal CT showed nonspecific terminal ileitis. He was treated for presumed HACEK bacteremia and was transitioned from piperacillin-tazobactam to ceftriaxone to complete a two-week course at hospital discharge. He also received a seven-day course of doxycycline for concomitant, mild lower extremity cellulitis which resolved during hospitalization. Ultimately, antimicrobial susceptibility testing which resulted following discharge was not consistent with the HACEK organism. Testing demonstrated resistance to multiple antimicrobials including ceftriaxone, as well as susceptibility to trimethoprim-sulfamethoxazole (TMP/SMX). WGS ultimately identified the organism as D. capnocytophagoides. Despite ceftriaxone resistance, he reported feeling well at follow-up with negative surveillance blood cultures.This patient shares several features with the few patients previously identified with D. capnocytophagoides bacteremia, including malignancy, recent neutropenia, and presumed gastrointestinal source. As in the small number of prior reported cases, the organism was difficult to identify leading to delay in diagnosis and treatment. The case demonstrates the importance of critical thinking in the face of contradictory test results. Additionally, based on susceptibility profiles described in prior literature, we suspect doxycycline treated his bacteremia.

Li+ and Na+ attachment to some dipeptides via LDI-TOF mass spectrometry: Fragmentation patterns

Laser desorption ionization-time of flight (LDI-TOF) mass spectrometry is used for studying the attachment of Na+ and Li+ ions to four dipeptides including phenylalanyl-alanine (Phe-Ala), tyrosyl-alanine (Tyr-Ala), L-Phenylalanyl-L-Phenylalanine (Phe-Phe), and alanyl-glutamine (Ala-Gln) dipeptides. The LiCl, NaOH, and NaF salts are used as the source of Li+ and Na+ ions in the LDI of the dipeptides. Our aim is the investigation of the difference between the fragmentation patterns of the selected dipeptides in the presence of Na+ and Li+ ions due to the laser radiation and providing information for the fragmentation of larger peptides in the same conditions. The characteristic peak, related to [dipeptide-H + 2Na]+ species, is observed in the mass spectrum of Phe-Ala and Tyr-Ala dipeptides in the presence of NaF, while the breaking of the peptide bond (OC-NH) occurs for the Phe-Phe in the presence of the aforementioned salts. The characteristic peak of Ala-Gln dipeptide ([(Ala-Gln)-H + 2Na]+) is observed in the absence of any salt. The mass spectra of the dipeptides, recorded in the presence of LiCl, are crowded compared to those recorded in the presence of NaF and NaOH showing the effect of the type of alkali salt on the dipeptide fragmentation. The theoretical calculations are employed to investigate the ability of the interaction sites of dipeptides for the attachment of one and two Na+ and determine the most stable structure of the [dipeptide-H + 2Na]+ species for each dipeptide.

Sulfamethoxazole-trimethoprim-induced liver injury and genetic polymorphisms of NAT2 and CYP2C9 in Taiwan.

Sulfamethoxazole-trimethoprim (SMX-TMP) is one of the most frequently used antibiotics. SMX is metabolized by N-acetyltransferase (NAT) and cytochrome P450 2C9 (CYP2C9) to nontoxic or toxic intermediates. Little is known about the association between genetic variations of these enzymes and SMX-TMP-induced liver injury (SILI). The aim of this study was to explore the genetic polymorphisms of NAT2 and CYP2C9 and the susceptibility to SILI in a Han Chinese population. A total of 158 patients with SILI and 145 controls were recruited in this study. PCR-based genotyping with matrix-assisted laser desorption ionization-time of flight was used to assay the major NAT2 and CYP2C9 genotypes including NAT2 rs1495741, rs1041983, rs1801280, CYP2C9 rs1799853, rs1057910 and rs4918758. The SILI group had a higher frequency of the NAT2 rs1495741 variant AA genotype and rs1041983 variant TT genotype than the controls (42.4 vs. 25.5%; P = 0.008, and 40.5 vs. 25.5%; P = 0.022, respectively). The SILI group had more slow acetylators than the controls (43.7 vs. 25.5%; P = 0.001). There were no significant differences in the genetic variations of CYP2C9 between the SILI and control groups. After adjusting for confounding factors, the NAT2 slow acetylators still had an increased risk of SILI (adjusted OR: 2.49; 95% confidence interval: 1.46-4.24; P = 0.001), especially in those with hepatocellular and mixed type SILI. NAT2 slow acetylators are associated with a higher risk of SILI in the Han Chinese population. However, CYP2C9 genetic polymorphisms are not associated with the susceptibility to SILI.

돼지 난자난구세포 복합체의 난구세포 분포에 따른 단백질체 차이 분석

Porcine cumulus cell layers are thought to influence various fundamental functions during oocyte maturation. The expansion of cumulus-oocyte complexes (COCs) is known to highly correlate with oocyte maturation, fertilization, and embryonic development. In this study, we performed two dimensional electrophoresis (2-DE) and matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) analysis using porcine COCs proteins. Porcine COCs were collected from ovary follicles and classified as the good-quality group and the poor-quality group based on cumulus cell layers around oocyte. The COCs protein samples were separated in a pH 3.0–10.0 IPG gels for the first dimension, and then 8%–16% SDS-PAGE gel for the second dimension. Gel image analysis was performed in protein spots between good- and poor-quality groups, and then differential protein spots were identified using MALDI-TOF analysis. There were more than 300 protein spots in the 2-DE gel image obtained from 1 mg protein samples of COCs. Five differential protein spots were identified by gel image analysis with more than 1.5-fold differences. All 5 protein spots were up-regulated in good quality group and revealed as transferrin, tight junction protein ZO-2, tubulin beta chain, meiosis-specific nuclear structural protein 1 and dynamin 1-like isoform 4 in good. Our results indicated the identification of 5 differentially expressed proteins in the porcine good-quality COCs and these protein profiles will make available important guides for porcine good oocyte selection for the production of in vitro embryos.

Proteomic Imaging of Vestibular Schwannomas and Normal Nerves. Histopathologic Correlations.

Proteomic analysis of vestibular schwannoma (VS), non-vestibular schwannoma (NVS), and normal nerve (NN) using mass spectrometry and imaging of matrix assisted laser desorption ionization-time of flight (MALDI-TOF). Retrospective, qualitative, and descriptive study on VS, NVS, and NN. Samples were provided by our Tumor Bank. They were analyzed histologically then sprayed by acid matrix. The laser beam of MALDI performed desorption-ionization of the sample. A mass spectrogram (MS) was drawn depending on time of flight of ionized peptides, and MALDI-imaging was obtained which is a summation color spectrum depending on sample's peptide content. The slice was reexamined histologically and results compared with MALDI-imaging. Fifty schwannomas were sampled, of which 27 exploitable: 22 VS (17 Antoni type A and five type B) and five NVS (all Antoni type B). Eleven NN were analyzed. Among the 22 VS, near-total correlation between MALDI-imaging and pathology was found in two cases (9.1%), partial correlation in four (18.2%), and no correlation in 16 (72.7%); correlations were more frequent in VS of the Antoni type B. MS showed a peptide spike at 2,000 m/z in 7 (31.8%) and 5,000 m/z in 21 (95.5%). Among the five NVS, near-total correlation was found in three cases (60%), partial correlation in one (20%), and no correlation in one (20%). MS showed a peptide spike at 2,000 m/z in two (40%) and 5,000 m/z in all (100%). Among the 11 NN, near-total correlation was found in nine cases (81.8%), partial correlation in one (9.1%), and no correlation in one (9.1%). MS showed no peptide spike at 2,000 or 5,000 m/z. Behind homogeneous areas on histology, there was great heterogeneity on MALDI-imaging and MS, regarding VS and NVS, but not NN. There was a lack of correlation between MALDI-imaging and pathology in VS (except Antoni type B) as compared with NVS and NN. The lack of correlation in VS of the type A as compared with type B VS and NVS could be attributed to the overexpression of degeneration-associated proteins/peptides in VS of the type B as well as NVS that are better correlated with histologic findings. The two peptide spikes detected in schwannoma and not in NN opens up the prospect of tumor biomarkers identifiable by sequencing. The proteomic polymorphism found in VS and NVS was absent on histology which is a new morphologic characteristic of schwannoma. Further studies should be performed in the future to confirm the benefit and usefulness of the MALDI in the analysis of VS and NVS.

Prevalence and Characterization of Extended-Spectrum Beta-Lactamase-Producing Enterobacteriaceae in Major Hospitals in Gabon.

In Gabon, few data exist on extended-spectrum beta-lactamases-producing Enterobacteriaceae (ESBL-PE). This study investigated ESBL-PE prevalence and the associated resistance genes in clinical samples (n = 5,956) and anal swabs (n = 78) analyzed in eight hospitals and a medical analysis laboratory in Gabon from January 2016 to March 2018. Matrix-Assisted Laser Desorption Ionization-Time Of Flight (MALDI-TOF) mass spectrometry analysis identified 790 Enterobacteriaceae isolates (n = 712 clinical samples and n = 78 fecal samples). ESBL-PE prevalence (Müller-Hinton agar disk diffusion method and double-disk synergy test) was 11.8% (84/712) in clinical samples (15.5% from inpatients and 7.1% from outpatients; p < 0.05) and 16.7% (13/78) in carriage isolates. Most ESBL-PE were isolated from urine samples (46/84). In clinical and carriage ESBL-PE isolates, Escherichia coli was predominant (42.8% and 61.5%; phylogroups A, B1, B2, and D), followed by Klebsiella pneumoniae (41.7% and 23.1%). Multiplex PCR and bi-directional sequencing showed that CTX-M group 1 (blaCTX-M-15) was predominant in clinical and carriage ESBL-PE (94% and 92.3%) among which 85.7% and 92.3% also harbored one to three β-lactamase-encoding genes (blaTEM-1, blaOXA-1, or blaSHV-1). Resistance genes were detected in all hospitals in Gabon. ESBL-PE prevalence in Gabon has not reached alarming levels yet, but corrective and monitoring measures are needed to curb their emergence.

Multicenter evaluation of the VITEK MS matrix-assisted laser desorption/ionization-time of flight mass spectrometry system for identification of bacteria, including Brucella, and yeasts.

The use of matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry has proven to be rapid and accurate for the majority of clinical isolates. Some gaps remain concerning rare, emerging, or highly pathogenic species, showing the need to continuously expand the databases. In this multicenter study, we evaluated the accuracy of the VITEK MS v3.2 database in identifying 1172 unique isolates compared to identification by DNA sequence analysis. A total of 93.6% of the isolates were identified to species or group/complex level. A remaining 5.2% of the isolates were identified to the genus level. Forty tests gave a result of no identification (0.9%) and 12 tests (0.3%) gave a discordant identification compared to the reference identification. VITEK MS is also the first MALDI-TOF MS system that is able to delineate the four members of the Acinetobacter baumannii complex at species level without any specific protocol or special analysis method. These findings demonstrate that the VITEK MS v3.2 database is highly accurate for the identification of bacteria and fungi encountered in the clinical laboratory as well as emerging species like Candida auris and the highly pathogenic Brucella species.

Study of the Molecular-Weight Distribution of Binder Pitches for Carbon Blocks

The present study aimed to identify the required characteristics of binder pitches in the filler-binder mixing process to effectively manufacture graphite blocks. To this end, a binder pitch was separated into pitch fractions of varying molecular-weight segments. The role and effectiveness of each pitch fraction were then analyzed with respect to their molecular-weight distribution. As a result, the optimal molecular-weight distribution was determined. More specifically, a coal-tar pitch was separated into solvent-soluble and solvent-insoluble fractions. The molecular-weight distribution was determined according to this classification, and the characteristics of each pitch fraction were examined. The pitch separation process was conducted using three solvents: hexane, toluene, and quinoline. The resulting pitch was separated into the following pitch fractions: hexane-soluble (HS), hexane-insoluble-toluene-soluble (HI-TS), toluene-insoluble-quinoline-soluble (TI-QS), and quinoline-insoluble (QI). Fourier transform infrared (FT-IR) spectrum, matrix-assisted laser desorption ionization-time of flight (MALDI-TOF), and softening point of each pitch fraction were measured. Also, pitch samples were refabricated while varying the mixing ratio of these pitch fractions, and carbon blocks were then prepared using them. The compressive strength and porosity of these blocks were measured and compared. The P154_B pitch with a high content of TI-QS was used to fabricate a green block. Due to the high viscosity of the binder used, the fluidity was not sufficiently high, and thus, the green block made of this pitch had relatively low strength. The other blocks had similar levels of strength. After the carbonization process, the carbon block with a high content of HS (P352_B-C) and the carbon block with the HS content removed (P073_B-C) showed lower compressive strength than their respective green-block counterparts (P352_B and P073_B). However, their strength was higher compared to those of the other carbon blocks. In the case of carbon block P073_B-C, the HS content was completely removed, and thus, the content of TI-QS (β-resin) was relatively high. Accordingly, this carbon block ended up with large amounts of components that had high coking values (CVs), and this contributed to limiting the formation of pores. Therefore, the compressive strength of this carbon block was high. In the case of the carbon block with a high content of HS (P352_B-C), a suitable level of viscosity was achieved because the HS components ensured high fluidity. As a result, blocks with higher density and compressive strength could be fabricated. The major findings of the present study confirm that producing carbon blocks with high mechanical properties requires binder pitches with a balanced combination of suitable viscosity to ensure sufficiently high fluidity and a proper level of CV to effectively suppress the formation of pores in the mixing and molding process.

Re-188-tricarbonyl tamoxifen as a theranostic radiopharmaceutical for estrogen receptor expressing breast cancers: radiolabeling, characterization and in-vitro cytotoxic assessment.

Development of a novel theranostic radiopharmaceutical for estrogen receptor, expressing unresectable primary and metastatic breast cancers. Tamoxifen was radiolabeled with Rhenium-188 (Re-188) through tricarbonyl core. Radiolabeled complex was characterized by 1proton nuclear magnetic resonance spectroscopy and Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF). Various quality control tests such as sterility, apyrogenicity, and radiochemical purity (RCP) were performed to assess the suitability of the radiopharmaceutical for intravenous administration. In-vitro cell culture studies were performed for cytotoxic assessment. In addition to this, exposure due to different doses of Re-188-tricarbonyl tamoxifen was also calculated. Re-188-tricarbonyl and Re-188-tricarbonyl tamoxifen showed more than 99% RCP. Sample was found to be sterile and pyrogens levels were within the permissible limit. Re-188-tricarbonyl tamoxifen was successfully characterized by MALDI-TOF and 1H-NMR spectroscopy. Re-188 (1.480 MBq) and tamoxifen (0.027 or 0.054 µM) individually showed 36 and 70% cell death, respectively. However, radiolabeled complex (Re-188-tricarbonyl tamoxifen) with the same amount of radioactivity (1.480 MBq) increased the cell death to more than 90% with one-fifth to one-tenth molar concentration of tamoxifen (0.0054 μM). Re-188-tricarbonyl tamoxifen can be synthesized in-house in radiopharmacy lab. Radionuclide therapy with Re-188-tricarbonyl tamoxifen can be given using 10 times less amount of tamoxifen as compared to cold tamoxifen.

Investigation of Phylogeny and Drug Resistance Mechanisms of Elizabethkingia anophelis Isolated from Blood and Lower Respiratory Tract

Elizabethkingia species are environmental bacteria associated with opportunistic infections in vulnerable populations. Traditionally, Elizabethkingia meningoseptica was considered the predominant pathogenic species. However, commercial identification systems have routinely misidentified Elizabethkingia anophelis as E. meningoseptica, leading to a mischaracterization of clinical strains and an underestimation of the role of E. anophelis in human disease. Elizabethkingia spp. harbor multidrug resistance (MDR) genes that pose challenges for treatment. Differentiation between Elizabethkingia spp. is particularly important due to differences in antimicrobial resistance (AMR) and epidemiological investigation. In this study, we describe a case of MDR E. anophelis isolated from the blood and lower respiratory tract of a patient who was successfully treated with minocycline. These isolates were initially misidentified by matrix assisted laser desorption ionization-time of flight as E. meningoseptica, whereas whole genome sequencing (WGS) confirmed the isolates as E. anophelis with the closest related strain being E. anophelis NUHP1, which was implicated in a 2012 outbreak in Singapore. Several AMR genes (blaBlaB, blaBlaGOB, blaCME, Sul2, erm(F), and catB) were identified by WGS, confirming the mechanisms for MDR. This case emphasizes the utility of WGS for correct speciation, elucidation of resistance genes, and relatedness to other outbreak strains. As E. anophelis is associated with a high mortality and has been found in hospital system sinks, WGS is critically important for determining strain relatedness and tracking outbreaks in the hospital setting.