Dendrobium candidum polysaccharide reduce atopic dermatitis symptoms and modulate gut microbiota in DNFB-induced AD-like mice.

Atopic dermatitis (AD) is a chronic inflammatory skin disease with a high prevalence worldwide. Increasing evidence suggests that the gut microbiota plays an important role in the pathogenesis of AD. In this study, we sought to verify the effect of Dendrobium candidum polysaccharides (DCP) on AD induced by 2,4-Dinitrofluorobenzene (DNFB) in Balb/c mice regarding its impact on the intestinal microbiome. We found that 2-week oral administration of DCP improved AD-like symptoms and histological damage of skin, reduced mast cell infiltration, down-regulated the level of serum total IgE and the expression of pro-inflammatory cytokines such as TNF-α, IFN-γ, IL-4 and IL-6, and increased the expression level of anti-inflammatory cytokine IL-10. The beneficial effect of DCP was attributed to the restoration of the intestinal microbiome composition and the unbalance of the intestinal homeostasis. Our results indicated that DCP might be used as a promising novel microbiota-modulating agent for the treatment of AD.

Analytical techniques for monitoring of toluene and xylene exposure biomarkers hippuric acid and methylhippuric acid in human urine samples.

Quantification of hippuric acid and methylhippuric acid in human urine matrices provides information on the toluene and xylene exposure conditions. High performance liquid chromatography coupled with UV detection is the preferable technique for hippuric acid and methylhippuric acid detection in human urine. This study was conducted to present analytical techniques developed for monitoring of hippuric acid and methylhippuric acid in human urine matrices during 2016-2021.

Review of Detection and Quantification of Rabies Virus Antibodies.

Rabies is an almost invariably fatal disease. According to the World Health Organization (WHO), rabies virus neutralizing antibody (RVNA) titers of ≥0.5 IU/mL are considered adequate for rabies protection. Therefore, detection and quantification of RABV antibodies are important. Many methods have been developed for detecting RABV antibodies. In the present study, we reviewed several methods of detecting RABV antibodies in human and animal samples and evaluated and compared their performance. Of 34 methods, 5 demonstrated unsatisfactory sensitivity or specificity. The others exhibited sensitivity and specificity of ≥75%. The correlation coefficient for five of eight methods was >0.8. The Bland-Altman mean bias of five of five methods was <±2.0. The kappa values of 25 of 28 methods were higher than 0.4, demonstrating at least moderate agreement. Analysis of the performance of these methods emphasized that any new technology should be considered carefully and objectively before being used as an appropriate and applicable alternative.

Magnetic molecularly imprinted polymers for extraction of S-phenylmercapturic acid from urine samples followed by high-performance liquid chromatography.

In this study, magnetic molecularly imprinted polymers (MMIPs) were prepared and used as sorbents for extraction of S-phenylmercapturic acid (S-PMA) from urine samples, followed by high-performance liquid chromatography ultraviolet-visible (HPLC-UV/Vis) analysis. The MMIPs were synthesized by the copolymerization reaction of (phenylthio) acetic acid (template molecule), methacrylic acid (functional monomers) and ethylene glycol dimethacrylate (cross-linkers). The morphology, structure property and surface groups of the prepared MMIPs were characterized by scan electron microscopy, transmission electron microscopy, infrared spectroscopy, X-ray diffraction pattern, thermogravimetric analyses, Brunauer-Emmett-Teller and vibrating sample magnetometer. The selectivity of the MMIPs was investigated in the presence of interferents. Various parameters affecting the S-PMA extraction efficiency were investigated, including MMIPs amount, pH, sample volume, desorption solvent, as well as extraction and desorption time. The obtained optimal parameters were as follows: MMIPs amount (20 mg), pH (3.0), sample volume (5 mL), desorption solvent (methanol/acetic acid [9/1, v/v]), extraction time (30 minutes) and desorption time (2 minutes). The method was validated according to the Food and Drug Administration Guidance for Industry on Bioanalytical Method Validation. The calibration curve for the analyte was linear in the concentration range of 0.030-1.0 mg/L (r = 0.9995). The LOD and LOQ of the method were 0.0080 and 0.0267 mg/L, respectively. The enrichment factor of the MMIPs was 5. The relative standard deviations of intra- and inter-day tests were in the range of 3.8-5.1% and 3.9-6.3%, respectively. The recoveries at three different concentrations of 0.10, 0.50 and 0.80 mg/L ranged between 95.2% and 98.6%. In addition, the MMIPs could be reused for at least eight times. The proposed method was successfully applied to the determination of S-PMA in urine samples. In addition, this developed method could be used as a tool in the early screening and clinical diagnosis of benzene intoxication.

Genome sequencing and comparative genome analysis of 6 hypervirulent Klebsiella pneumoniae strains isolated in China.

Hypervirulent Klebsiella pneumoniae (hvKP) has been increasingly reported over the past three decades and causes severe infections. To increase our understanding of hvKP at the genome level, genome sequencing and comparative genome analysis were performed on 6 hvKPs. The whole genome DNA from 6 hvKPs with different capsular serotypes isolated in China was extracted. The genome sequencing and assembly results showed the genome size of the six hvKPs and GC content. Comparative analyses of the genomes revealed the gene homology and genome rearrangement in the 6 hvKPs compared with Klebsiella pneumonia NTUH-K2044. The phylogenetic tree based on full-genome SNPs of the 7 hvKPs showed that NTUH-K2044 formed a single clade, showing distant evolutionary distances with the other six strains, and the non-K1 hvKP strains had a relatively closer phylogenetic relationship. BLAST comparison analysis found that some selected virulence genes had different degrees of deletion in the non-K1 hvKPs. SNP-based virulence gene mutation analysis showed that some virulence genes had different degrees of SNP mutations. The whole-genome sequencing and comparative genome analysis of six hvKP strains with NTUH-K2044 provide us with a basic understanding of the genome composition, genetic polymorphism, evolution and virulence genes of hvKP and a basis for further research on these genes and the pathogenesis of hvKP.

RcsAB and Fur Coregulate the Iron-Acquisition System via entC in Klebsiella pneumoniae NTUH-K2044 in Response to Iron Availability.

The iron acquisition system is an essential virulence factor for human infection and is under tight regulatory control in a variety of pathogens. Ferric-uptake regulator (Fur) is one of Fe2+-responsive transcription factor that maintains iron homeostasis, and the regulator of capsule synthesis (Rcs) is known to regulate exopolysaccharide biosynthesis. We speculate the Rcs may involve in iron-acquisition given the identified regulator box in the upstream of entC that participated in the biosynthesis of enterobactin. To study the coregulation by RcsAB and Fur of entC, we measured the ß-galactosidase activity and relative mRNA expression of entC in WT and mutant strains. The RcsAB- and Fur-protected regions were identified by an electrophoretic mobility shift assay (EMSA) and a DNase I footprinting assay. A regulatory cascade was identified with which Fur repressed rcsA expression and reduced RcsAB and entC expression. Our study demonstrated that entC was coregulated by two different transcriptional regulators, namely, RcsAB and Fur, in response to iron availability in Klebsiella pneumoniae.

Activated carbon/diatomite-based magnetic nanocomposites for magnetic solid-phase extraction of S-phenylmercapturic acid from human urine.

In this study, activated carbon/diatomite-based magnetic nancomposites (denoted as AC/DBMNs) were synthesized and applied as an adsorbent for magnetic solid-phase extraction of S-phenylmercapturic acid (S-PMA) from human urine prior to high-performance liquid chromatography. The surface morphologies and structures of AC/DBMNs were characterized by Fourier transform infrared spectroscopy, transmission and scanning electron microscopy, X-ray diffraction, Brunauer-Emmett-Teller surface area, vibrating sample magnetometer and ζ-potential measurements. The experimental parameters including sample volume, sample pH, adsorbent amount, extraction time, elution solvent and desorption time were investigated in detail. Under the optimum conditions, the method exhibited good linearity (r > 0.9993) within the concentration ranges of 0.03-1.0 mg/L. Moreover, the limits of detection and quantification were 0.01 and 0.03 mg/L, respectively. The enrichment factor was 5, and good recoveries (88.9-97.3%) with relative standard deviations in the range of 5.6-6.8% (n = 6) for inter-day and 6.3-8.1% (n = 6) for intra-day were achieved. The developed method was successfully applied to the analysis of S-PMA in urine samples. In addition, this accurate and sensitive method has great potential to be applied in the early screening and clinical diagnosis of the workers exposed to benzene.

Risk factors of bloodstream infections in recipients after liver transplantation: a meta-analysis.

PURPOSE: Bloodstream infection (BSI) is an important cause of adverse outcomes for recipients with liver transplantation (LT). This meta-analysis aimed to identify risk factors associated with post-LT BSI. METHODS: Relevant studies published up to June 2017 were searched from seven electronic databases. The studies were reviewed according to the inclusion and exclusion criteria. The Z test was used to determine the pooled odds ratio (OR) or standardized mean difference (SMD) of the risk factors. ORs and their corresponding 95% confidence intervals (CIs), or SMDs and their corresponding 95% CIs were used to identify the significant difference of risk factors. RESULTS: Seventeen studies enrolling 4410 recipients were included. Eleven risk factors were identified to be associated with BSI after LT: male recipient (OR = 1.28), ascites (OR = 1.68), model for end-stage liver disease (MELD) score (SMD = 0.20), Child-Pugh class C (OR = 1.69), operation time (SMD = 0.18), incompatible blood type (OR = 2.87), operative blood loss (SMD = 0.33), rejection (OR = 1.72), biliary complications (OR = 1.91), hemodialysis (OR = 3.37), and retransplantation (OR = 2.86). CONCLUSIONS: Although some risk factors were identified as significant factors for BSI after LT, which may provide a basis for clinical prevention, well-designed prospective studies should be done to overcome the limitations of this study.

Transcriptional regulation of galF by RcsAB affects capsular polysaccharide formation in Klebsiella pneumoniae NTUH-K2044.

RcsAB is an atypical two-component regulatory system that can regulate exopolysaccharide biosynthesis and is involved in the virulence of K. pneumoniae. The gene galF is well known as a gene involved in the biosynthesis of capsular polysaccharide (CPS). The specific DNA identification sequence for transcriptional regulation of RcsAB was found to be present in the promoter region of galF. This study aimed to detect the function of RcsAB in virulence and in biofilm and CPS formation. In addition, the transcriptional regulation of the galF gene in K. pneumoniae was studied. To determine the function of rcsAB gene, the wild-type K. pneumoniae strain NTUH-K2044 and the rcsAB knockout and complemented strains were used. The results showed decreased virulence, biofilm formation, and CPS levels in the rcsAB knockout strain. Complementation of the knockout by introducing an rcsAB fragment on an expression plasmid partially restored the virulence, biofilm, and CPS functions of the knockout strain. It indicated that the rcsAB genes might affect CPS formation and virulence of K. pneumonia. RT-qPCR, EMSA and DNase I footprinting assays were conducted to identify the transcriptional regulation of galF by RcsAB. RcsAB was seen to bind to the galF promoter-proximal region, and the binding site was further identified to be located from -177 bp to -152 bp upstream of the galF promoter. In conclusion, RcsAB could regulate the transcription of the galF gene positively by binding to the galF promoter DNA directly, and then affects the CPS formation of K. pneumonia.

Epidemiology of pathogens and antimicrobial resistanceof catheter-associated urinary tract infections in intensivecare units: A systematic review and meta-analysis.

BACKGROUND: In the intensive care unit (ICU), catheter-associated urinary tract infection (CAUTI) is the most common urinary tract infection. Nevertheless, there is no systematic review to investigate the epidemiology of pathogens and antimicrobial resistance of CAUTIs in ICUs. METHODS: Eight electronic databases were searched for eligible studies. A meta-analysis was performed to calculate the CAUTI incidence per 1,000 catheter days, the proportion of pathogen distribution, and the resistance rate with R3.3.2 software. RESULTS: Seventy-five studies were included. The total weighted CAUTI incidence per 1,000 catheter days was 7.78. Gram-negative bacteria (47.46%), fungi (27.81%), and gram-positive bacteria (19.06%) were isolated. Candida spp (27.4%), Escherichia spp (23.41%), and Enterococcus spp (15.0%) were the most frequent pathogens. Candida albicans, Candida tropicalis, and Candida glabrata were generally resistant to itraconazole, with resistance rates of 42.5%, 53.0%, and 59.7%, respectively. Escherichia spp displayed high rates of resistance to ampicillin (87.3%), ciprofloxacin (71.7%), and norfloxacin (71.2%). Enterococcus spp showed high rates of resistance to erythromycin (83.9%), penicillin (76.7%), and levofloxacin (73.8%). CONCLUSIONS: In ICUs, the CAUTI incidence per 1,000 catheter days is high. CAUTIs were mainly caused by gram-negative bacteria that were resistant to common antibiotics. There is a pressing demand for future research into CAUTI, including effective prevention, an understanding of antimicrobial resistance mechanisms, and development of new antibiotics for patient safety.

Genome-wide identification of genes regulated by RcsA, RcsB, and RcsAB phosphorelay regulators in Klebsiella pneumoniae NTUH-K2044.

Rcs phosphorelay system is a two-component signal transduction system, which can regulate the transcription of capsule polysaccharide and biofilm related genes in Enterobacteriaceae. In this study, microarray technology was used to investigate the overall genes regulated by RcsA, RcsB, and RcsAB and the regulation mechanism in Klebsiella pneumoniae, then COG analysis was performed to explore the functions of the differentially expressed genes. According to the microarray data result, a total of 45, 223 and 217 genes regulated by RcsA, RcsB, and RcsAB were screened. The result of COG analysis suggested that inorganic ion transport and metabolism related genes have a majority in RcsA regulating genes. Most of RcsB regulated genes were showed involved in energy production and conversion process. Besides Carbohydrate transport and metabolism genes were identified as the major components of the RcsAB regulated genes. 15 differentially expressed genes were confirmed by quantitative real-time PCR (RT-qPCR). The RT-qPCR results indicated that 13 genes consistent with microarray data. The results of this study provided important evidence for further research to investigate the influence of RcsA, RcsB, RcsAB regulators and further efforts to address the diseased caused by K.pneumoniae, such as pneumonia, bacteremia, and urinary tract infection.

Electrochemical aptasensor for thrombin using co-catalysis of hemin/G-quadruplex DNAzyme and octahedral Cu2O-Au nanocomposites for signal amplification.

In this work, novel octahedral Cu2O-Au nanocomposites were synthesized and first applied in an electrochemical aptasensor to detect thrombin (TB) with the aid of a DNAzyme for signal amplification. The octahedral Cu2O-Au nanocomposites have not only simultaneously served as signal amplifying molecules but have also been utilized as an ideal loading platform to immobilize a large number of electroactive substances and recognition probes. Gold nanoparticles (AuNPs) were grown directly on the surface of the octahedral Cu2O nanocrystals, and the Cu2O-Au nanocomposites obtained had the advantages of large surface areas and excellent biocompatibilities. The hemin/G-quadruplex, which was formed by intercalating hemin into the amino terminated thrombin binding aptamer (NH2-TBA), and the electroactive toluidine blue (Tb) were immobilized onto the Cu2O-Au nanocomposite surfaces through a stable Au-N bond. AuNPs, Cu2O and hemin/G-quadruplex co-catalyse the H2O2 in the working buffer to promote the electron transfer of Tb as a multiple signal amplification strategy in order to improve the performance of the electrochemical aptasensor. Under optimal conditions, the designed aptasensor exhibited sensitive detection of TB from 100 fM to 20nM with a lower detection limit of 23fM. This proposed aptasensor exhibited good sensitivity, high specificity and acceptable reproducibility and could be widely applied in bioassay analysis.

Risk Factors for Carbapenem-Resistant Klebsiella pneumoniae Infection: A Meta-Analysis.

AIMS: Carbapenem-resistant Klebsiella pneumoniae (CRKP) infection has been rapidly emerging as a life-threatening nosocomial disease in many countries. However, studies on the corresponding risk factors of CRKP infection showed inconsistent results. To resolve these inconsistencies, we conducted a meta-analysis of previous studies on the potential risk factors of CRKP infection. The results of this study could be used to develop CRKP infection prevention strategies. METHODS: Relevant works were systematically searched from five electronic databases up to September 2016. Z-test was used to determine the significance of the pooled odds ratios (ORs). ORs and 95% confidence intervals were utilized to compare the risk factors of CRKP infection. RESULTS: Sixteen studies that involved 3,627 participants were included in the meta-analysis. We identified the following risk factors that were associated with CRKP infection: (1) longer length of hospital stay (LOS) (OR = 12.92), (2) admission to intensive care unit (ICU) (OR = 2.48), (3) prior hospitalization (OR = 1.85), (4) longer days of ICU stay (OR = 4.58), (5) transplant recipient (OR = 2.01), (6) steroid use (OR = 1.43), (7) central venous catheter use (OR = 2.30), (8) mechanical ventilation (OR = 2.54), (9) presence of tracheostomy (OR = 3.63), (10) parenteral nutrition (OR = 2.38), (11) previous antibiotic use (OR = 3.31), and (12) exposure to carbapenems (OR = 4.01), (13) aminoglycosides (OR = 2.05), (14) glycopeptides (OR = 2.40), (15) quinolones (OR = 2.28), and (16) anti-pseudomonal penicillins (OR = 2.67). CONCLUSIONS: Sixteen risk factors including longer LOS, admission to ICU, previous antibiotic use, and exposure to carbapenems were associated with the development of CRKP infection. Identification of modifiable risk factors could play an important role in the prevention of CRKP infection.

Risk factors for infection and/or colonisation with extended-spectrum ß-lactamase-producing bacteria in the neonatal intensive care unit: a meta-analysis.

Extended-spectrum ß-lactamase (ESBL)-producing bacteria are an important cause of healthcare-associated infections in the neonatal intensive care unit (NICU). The aim of this meta-analysis was to identify risk factors associated with infection and/or colonisation with ESBL-producing bacteria in the NICU. Electronic databases were searched for relevant studies published from 1 January 2000 to 1 July 2016. The literature was screened and data were extracted according to the inclusion and exclusion criteria. The Z-test was used to calculate the pooled odds ratio (OR) of the risk factors. ORs and their 95% confidence intervals were used to determine the significance of the risk. A total of 14 studies, including 746 cases and 1257 controls, were identified. Thirteen risk factors were determined to be related to infection and/or colonisation with ESBL-producing bacteria in the NICU: birthweight [standardised mean difference (SMD) = 1.17]; gestational age (SMD = 1.36); Caesarean delivery (OR = 1.76); parenteral nutrition (OR = 7.51); length of stay in the NICU (SMD = 0.72); mechanical ventilation (OR = 4.8); central venous catheter use (OR = 2.85); continuous positive airway pressure (OR = 5.0); endotracheal intubation (OR = 2.82); malformations (OR = 2.89); previous antibiotic use (OR = 6.72); ampicillin/gentamicin (OR = 2.31); and cephalosporins (OR = 6.0). This study identified risk factors for infection and/or colonisation with ESBL-producing bacteria in the NICU, which may provide a theoretical basis for preventive measures and targeted interventions.

The KP1_4563 gene is regulated by the cAMP receptor protein and controls type 3 fimbrial function in Klebsiella pneumoniae NTUH-K2044.

Klebsiella pneumoniae (K. pneumoniae) is an opportunistic pathogen that can adhere to host cells or extracellular matrix via type 1 and type 3 fimbriae. KP1_4563 is a gene encoding a hypothetical protein in K. pneumoniae NTUH-K2044. KP1_4563 is located between the type 1 and type 3 fimbrial gene clusters and is likely associated with fimbrial function given its putative conserved domains of unknown function (DUF1471). Cyclic AMP receptor protein (CRP) regulates virulence-related gene expression and is a crucial transcriptional regulator in many bacteria. The predicted DNA recognition motif of CRP is present in the KP1_4563 promoter region. This study aimed to investigate the function of KP1_4563 in fimbriae and its transcriptional regulation mechanism by CRP. We generated Kp-Δ4563 mutant and complementation strains. We utilized phenotype and adhesion assays to evaluate the role of KP1_4563 in fimbriae. We conducted quantitative RT-PCR (qRT-PCR), LacZ fusion, electrophoretic mobility shift, and DNase I footprinting assays to study the transcriptional regulation of KP1_4563 gene by CRP. We found that KP1_4563 negatively regulates the function of type 3 fimbriae. Compared with NTUH-K2044, the absence of KP1_4563 enhanced the ability of Kp-Δ4563 to adhere to A549 cells. CRP negatively regulates KP1_4563 by directly binding to its promoter region. KP1_4563 plays an important role in type 3 fimbrial function. This novel insight will assist in the development of strategies for preventing K. pneumoniae infection.

Molecular characterisations of integrons in clinical isolates of Klebsiella pneumoniae in a Chinese tertiary hospital.

BACKGROUND: Integrons are mobile genetic elements that play an important role in the distribution of antibiotic-resistance genes among bacteria. This study aimed to investigate the distribution of integrons in clinical isolates of Klebsiella pneumoniae and explore the molecular mechanism of integron-mediated multiple-drug resistance in K. pneumoniae. METHODS: Class 1, 2, and 3 integrases were identified by polymerase chain reaction (PCR) among 178 K. pneumoniae clinical isolates. Antibiotic susceptibility was examined by disk-diffusion method. Conjugation experiments were conducted to evaluate the horizontal-transfer capability, and multilocus sequence typing (MLST) assays were conducted to explore the genetic relationships among the isolates. Highly virulent serotypes were identified by PCR from the 44 integron-positive isolates with variable regions. RESULTS: Class1 and 2 integrons were detected in 60.1% and 1.7% of isolates, respectively. One isolate carried both class 1 and 2 integrons. Class 3 integrons were not detected in all 178 isolates. Among the 44 integrons containing variable regions, 39 were located in conjugative plasmids. Dihydrofolate reductase (dfrA) and aminoglycoside adenyltransferase (aad) were found to be the most common in class 1 and 2 integrons. These gene cassettes encoded resistance to trimethoprim and aminoglycosides. Moreover, the association between integron carriage and antibiotic resistance was most significant for aminoglycosides, phenicols, and fluoroquinolones. Among the 44 integron-positive isolates with variable regions, 9 were classified as highly virulent serotypes (k1, k2, k20, and k54). In addition, MLST analysis detected 13 sequence types (STs), with the predominant ones being ST11 and ST15. The eBURST analysis revalued the existence of 11 singleton STs and one group, which is comprised of ST11 and ST437. CONCLUSIONS: The wide diversity of detected integrons suggested that the horizontal transfer by mobile genetic elements played a major role in the distribution of antimicrobial resistance genes, thereby indicating the urgent need to use effective means of avoiding the spread of drug-resistant bacteria.