[Drug resistance and genomic characteristics of a strain of O139 Vibrio cholerae isolated from human bloodstream infection].

Objective: To analyze the drug resistance and genomic characteristics of a strain of serogroup O139 Vibrio cholerae producing cholera toxin isolated from the bloodstream of a person with bacteremia. Methods: The broth dilution method and automatic drug sensitivity analyzer were used to determine the antibiotic sensitivity of the strain. The complete genome sequence of the strain was obtained by using second-generation gene sequencing and nanopore sequencing. BLAST software was used for comparison and analysis with CARD, Resfinder, ISfinder, VFDB, and other databases. The drug-resistant genes, insertion sequences and virulence genes carried by the strain were identified. MEGA 5.1 software was used to construct a genetic phylogenetic tree based on the core genomic single nucleotide polymorphisms. Results: V. cholerae SH400, as the toxigenic strain, carried multiple virulence-related genes and four virulence islands. The strain was resistant to streptomycin, tetracycline and cotrimoxazole, carrying corresponding drug-resistant genes. The strain also carried IncA/C plasmid with the size of 172914 bp and contained 10 drug-resistant genes. Combined with the genomic evolutionary relationship, this study found that the drug-resistant genes and drug-resistant plasmids carried among strains showed certain aggregation. The traditional ST type of strain SH400 was ST69, and the cgMLST type was a new type highly similar to cgST-252. Conclusion: This strain of serogroup O139 V. cholerae carries the ctxAB gene, multiple drug-resistant genes and IncA/C plasmid, and there are multiple drug-resistant islands.

[Construction of spvD gene deletion mutant and compensation strains in Salmonella enteritidis and its effects on Caco-2 cells].

Objective: To analyze the effects of spvD gene on invasion and intracellular proliferation of Caco-2 cells and in order to provide insight into the function of that gene and the underlying mechanism of Salmonella caused infection. Methods: Functional verification of spvD gene deletion mutant and compensation strain. The deletion mutant strain was constructed through a suicide plasmid-mediated homologous recombination. The compensation plasmid constructed by cloning the coding sequence of spvD by PCR into plasmid pBAD33 was mobilized into the deletion mutant by conjugation and the pBAD33 was introduced into wild strains and deleted mutant strains as control. The relative expression of spvD mRNA was detected by quantitative reverse transcription PCR. In order to analyze the virulence of spvD against Caco-2 cells, Caco-2 cells was cocultured with wild type Salmonella enteritidis carrying spvD gene, the deletion mutant strain and compensation strain respectively. The expression level of spvD mRNA and the the number of Salmonella enteritidis after Caco-2 cells intervention were compared between the three groups by LSD-t test, and the invasion rate was compared by χ2 test. Results: The expression level of spvD mRNA in wild type Salmonella enteritidis was set as unit "1", the deletion mutant strain was "0.00", and the compensation strain was "2.60" (LSD-twild, deleted=1.11, P=0.31; LSD-twild, compensation=-1.77, P=0.13; LSD-t deleted, compensation=-2.88, P=0.03), which confirmed the successful construction of the deletion mutant strain and the compensation strain. The invasion experiment results of the above three Salmonella enteritidis strains on Caco-2 cells showed that the invasion rate of wild strain was 0.23%, the invasion rate of deleted mutant strain was 0.16%, and the invasion rate of compensation strain was 0.16%, with no statistical significance (χ2=1.13, P=0.570). By comparing the number of Salmonella enteritidis at different time points after Caco-2 cells intervention, it was discovered that the number of Salmonella enteritidis in wild strains (6.50×106 CFU/ml) and compensation strains (7.25×106 CFU/ml) was significantly increased than that in deletion mutant strain (1.90×106 CFU/ml) after 16 h coculture (LSD-twild, deleted=7.95, P=0.00; LSD-twild, compensation=-1.27, P=0.25; LSD-t deleted, compensation=-9.22, P=0.00). Conclusion: It is not considered that spvD gene can affect the invasion of Salmonella enteritidis on Caco-2 cells, but the gene can promote the reproduction of Salmonella enteritidis in Caco-2 cells.

Juvenile hormone regulates the reproductive diapause through Methoprene-tolerant gene in Galeruca daurica.

Juvenile hormone (JH) signalling plays an important role in regulation of reproductive diapause in insects. However, its underlying molecular mechanism has been unclear. Methoprene-tolerant (Met), as a universal JH receptor, is involved in JH action. To gain some insight into its function in the reproductive diapause of Galeruca daurica, a serious pest on the Inner Mongolia grasslands undergoing obligatory summer diapause at the adult stage, we cloned the complete open-reading frame (ORF) sequences of Met and other 7 JH signalling-related genes, including JH acid methyltransferase (JHAMT), JH esterase (JHE), JH epoxide hydrolase (JHEH), Krüppel homologue 1 (Kr-h1), vitellogenin (Vg), forkhead box O (FOXO) and fatty acid synthase 2 (FAS2), from this species. GdMet encoded a putative protein, which contained three domains typical of the bHLH-PAS family. Expression patterns of these eight genes were developmentally regulated during adult development. Topical application of JH analogue (JHA) methoprene into the 3-day-old and 5-day-old adults induced the expression of GdMet. Silencing GdMet by RNAi inhibited the expression of JHBP, JHE, Kr-h1 and Vg, whereas promoted the FAS2 expression, which enhanced lipid accumulation and fat body development, and finally induced the adults into diapause ahead. Combining with our previous results, we conclude that JH may regulate reproductive diapause through a conserved Met-dependent pathway in G. daurica.

[Comparison and application of postprandial and fasting blood lipid levels in 839 physical examinees in Beijing].

Objective: To investigate the feasibility of application of non-fasting dyslipidemia cutoff values in community population. Methods: Self-control study was used. 839 physical examinees (292 males and 547 females) were recruited in clinical laboratory of Guang'an men Hospital from January to October 2018. The median (interquartile range) of age was 60 (54, 66) years. Blood samples were collected before and at 4 h after a standard breakfast. Comparison of fasting and postprandial lipoprotein levels was performed using Paired-Samples T Test or Two-Related-Samples Wilcoxon. The changes of 4-hour postprandial blood lipid levels and the percentages of postprandial dyslipidemia according to different stratification of fasting dyslipidemia were performed using one-way ANOVA and χ2 test, respectively. Results: Compared with fasting, 4-hour postprandial total cholesterol (TC), low density lipoprotein cholesterol (LDL-C), high density lipoprotein cholesterol (HDL-C), non-high density lipoprotein cholesterol (non-HDL-C), apolipoprotein A1 (ApoA1) and apolipoprotein B (ApoB) decreased slightly, postprandial triglyceride (TG) increased by 0.72 mmol/L, and postprandial remnant-like lipoprotein cholesterol (RLP-C) increased by 0.27 mmol/L (t or Z values = 10.26,22.94,24.22,4.71,16.61,26.92,-23.58,-19.35, P<0.05, respectively). According to the non-fasting dyslipidemia cut-off values recommended by the European consensus, there were 10%, 16.6%, 10.1%, 12.3%, 30% and 34.9% of the population in the appropriate levels of fasting TC, LDL-C, HDL-C, non-HDL-C, TG and RLP-C distributed in elevated levels of postprandial, respectively. The changes of 4-hour postprandial TC, LDL-C, non-HDL-C and HDL-C increased with the elevation of fasting level (F=9.50,6.18,8.07,3.86,P<0.01), and the maximum changes of TC≤3.5%, LDL-C≤6.8%, non-HDL-C≤2.9%, HDL-C≤6.3%; the change of 4-hour postprandial TG increased slightly first and then decreased significantly (51.3% vs. 57.9% vs. 39.2%, F=19.05, P<0.01); the change of 4-hour postprandial RLP-C decreased (50.8% vs. 33.2%, F=10.40, P<0.01). The cut-off values of 4-hour postprandial dyslipidemia were TC ≥5.1 mmol/L, LDL-C ≥3.2 mmol/L, HDL-C ≤0.9 mmol/L, non-HDL-C ≥4.0 mmol/L and RLP-C ≥1.0 mmol/L. The cut-off values of borderline elevated and elevated TG levels were ≥2.2 mmol/L and ≥3.4 mmol/L, respectively. Conclusions: The cut-off values of postprandial dyslipidemia including TC, LDL-C, HDL-C, non-HDL-C and RLP-C were preliminarily established in community population, which could be applied to the routine lipid profile evaluation in the physical examination population. And it might be needed that postprandial TG was managed hierarchically according to different cut-off values.

[Genomic recombination of the vibrio cholerae serogroup O1 El Tor pandemic strains].

Objective: To analyze the genomic recombination of the vibrio cholerae serogroup O1 El Tor pandemic strains. Methods: A total of 292 complete or draft genome sequences of Vibrio cholerae O1 serogroup El Tor strains isolated from 1937 to 2015 were selected from National Biotechnology Information Center database. The genome alignment of strains was computed by snippy software by using N16961 as reference sequence. Then ClonalFrameML software was used to do the recombinant analysis. The wilcox.test function in agricolae package was used to compare the number recombinant segments and the total length of recombinant regions between small and large chromosomes. The kruskal function was used to compare the number recombinant segments and the total length of recombinant regions among different isolation continents. The KOBAS tool was used to do the gene ontology enrichment analysis of recombinant hotspot genes. Results: Of all 292 strains of Vibrio cholerae, 163 strains (55.8%) were recombined. The median of normalized recombinant segment number of small chromosome was 4.7×10(-6) (9.3×10(-7), 2.0×10(-5)), which was significantly larger than that of large chromosome [2.4×10(-6) (3.4×10(-7), 5.7×10(-6))] (P<0.001). The median (P(25),P(75)) of recombinant segment number of strains isolated from Africa, Asia, Europe, North America and South America were 23(1.0,33.0), 1.0(0.0,34.0), 6.0(2.0,13.0), 0.0(0.0,1.0) and 29.5(6.8,56.8), respectively, and the difference was statistically significant (P<0.001). The median (P(25),P(75)) of total length of recombinant regions of strains isolated from Africa, Asia, Europe, North America and South America were 233.0(4.0, 461.0), 11.0(0.0, 695.5), 56.0(4.0,111.0), 0.0(0.0,9.0) and 347.5(132.8,1 323.5) bp, respectively, and the difference was statistically significant (P<0.001). Gene ontology Enrichment analysis showed that the functions of 62 recombinant hotspot genes were mainly enrichment in chemotaxis, taxis, response to external stimulus, receptor activity and molecular transducer activity. Conclustion: In this study, we found that there were significant differences in the number of recombinant fragments and the length of recombinant regions between large and small chromosomes of Vibrio cholerae El Tor. We also found significant differences in the number of recombinant fragments and the total length of recombinant regions among different continents.

[Prediction of syncope with nonlinear dynamic analysis during head-up tilt in vasovagal syncope patients].

OBJECTIVE: To quantify the relationship between cerebral blood flow velocity and peripheral blood pressure during hypotension period, aiming to predict the brain hypotension before symptomatic occurrence. METHODS: Twenty vasovagal syncope (VVS) patients who had a previous clinical history were selected in groups and 20 pair-matched control subjects underwent 70° tilt-up test. The subjects remained supine for 30 minutes before recordings when Doppler probes, electrodes and Finapres device were prepared. After continuous baseline recordings for 10 min, the subjects underwent head up tilt (HUT) test (70°), and were standing upright for 30 minutes or until syncope was imminent. For ethical reasons, the subjects were turned back to supine position immediately after SBP dropped to ≥20 mmHg, when their consciousness persisted. The point of syncope was synchronized for all the subjects by the point SBP reached the minima. Their beat-to-beat blood pressures (BP) were recorded continuously and bilateral middle cerebral artery (MCA) flow velocities were obtained with two 2 MHz Doppler probes from a transcranial Doppler ultrasonography (TCD) system. A nonlinear dynamic method--multimodal pressure flow (MMPF) analysis was introduced to access cerebral autoregulation during different time intervals. We introduced a new indicator--syncope index (SI), which was extracted from blood flow velocity (BFV) signal to evaluate the variation of cerebral vascular tension, and could reflect the deepness of dicrotic notch in BFV signal. RESULTS: Compared with the syncope index of the baseline value at the beginning of the tilt test, SI in VVS group showed significantly lower when the VVS occurred (0.16±0.10 vs.0.27±0.10,P<0.01),while there was no significant difference in syncope index between the control group at the end of the tilt test and the baseline value at the beginning of the tilt test. For those VVS patients, pulse index and resistance index had no significant change. Syncope index decreased significantly 3 minutes before the point of syncope (0.23±0.07 vs.0.29±0.07,P<0.01). CONCLUSION: Dynamic regulation is exhausted when vasovagal syncope occurred. Tension decrease of small vessels could have some relationship with loss of the cerebral autoregulation capability. The proposed syncope index could be a useful parameter in predicting syncope of VVS patients since it decreased significantly up to 3 minutes earlier from the point of syncope.

[Patients with anti-GABA (B) receptor encephalitis: 8 cases report].

Objective: To discuss the clinical features, diagnosis, therapy and prognosis of anti-GABA (B) receptor encephalitis. Methods: Clinical data of patients with anti-gaba B receptor encephalitis admitted to the department of neurology of the second hospital of hebei medical university from April 2016 to February 2018 were collected and retrospectively analyzed. Results: Eight patients with anti-GABA (B) receptor encephalitis were included. All the eight cases presented with epileptic seizures. The initial symptoms were clinical seizures in six cases, memory deficit in one case, mental and behavior disorder in one case. Cerebrospinal fluid (CSF) test indicated high intracranial pressure in four patients, increased protein in two patients and lymphocytosis in four patients. Long-term electroencephalogram (EEG) showed abnormal in all patients. Brain MRI showed limbic system lesions in four patients. The anti-GABA (B) receptor antibody was positive in both serum and CSF in all patients. Four patients had small cell lung cancer. All the eight patients received immunotherapy and symptoms were improved. After 6-24 months' follow-up, 1 patient recovered completely, and in the remaining 7 patients, mild symptoms were left behind. Conclusions: The anti-GABA (B) receptor encephalitis is characterized by refractory epilepsy. Epilepsy is the most common initial symptom. About 50% patients are accompanied by tumors. Anti-GABA (B) receptor antibodies testing and tumor screening should be actively carried out.

Validation of a Metastatic Assay using biopsies to improve risk stratification in patients with prostate cancer treated with radical radiation therapy.

Background: Radiotherapy is an effective treatment of intermediate/high-risk locally advanced prostate cancer, however, >30% of patients relapse within 5 years. Clinicopathological parameters currently fail to identify patients prone to systemic relapse and those whom treatment intensification may be beneficial. The purpose of this study was to independently validate the performance of a 70-gene Metastatic Assay in a cohort of diagnostic biopsies from patients treated with radical radiotherapy and androgen deprivation therapy. Patients and methods: A bridging cohort of prostate cancer diagnostic biopsy specimens was profiled to enable optimization of the Metastatic Assay threshold before further independent clinical validation in a cohort of diagnostic biopsies from patients treated with radical radiotherapy and androgen deprivation therapy. Multivariable Cox proportional hazard regression analysis was used to assess assay performance in predicting biochemical failure-free survival (BFFS) and metastasis-free survival (MFS). Results: Gene expression analysis was carried out in 248 patients from the independent validation cohort and the Metastatic Assay applied. Ten-year MFS was 72% for Metastatic Assay positive patients and 94% for Metastatic Assay negative patients [HR = 3.21 (1.35-7.67); P = 0.003]. On multivariable analysis the Metastatic Assay remained predictive for development of distant metastases [HR = 2.71 (1.11-6.63); P = 0.030]. The assay retained independent prognostic performance for MFS when assessed with the Cancer of the Prostate Assessment Score (CAPRA) [HR = 3.23 (1.22-8.59); P = 0.019] whilst CAPRA itself was not significant [HR = 1.88, (0.52-6.77); P = 0.332]. A high concordance [100% (61.5-100)] for the assay result was noted between two separate foci taken from 11 tumours, whilst Gleason score had low concordance. Conclusions: The Metastatic Assay demonstrated significant prognostic performance in patients treated with radical radiotherapy both alone and independent of standard clinical and pathological variables. The Metastatic Assay could have clinical utility when deciding upon treatment intensification in high-risk patients. Genomic and clinical data are available as a public resource.

Reinfection hazard of hand-foot-mouth disease in Wuhan, China, using Cox-proportional hazard model.

Hand-foot-mouth disease (HFMD) is an acute infectious disease caused by serotypes of the enterovirus (EV) family. HFMD reinfection occurs commonly in lack of cross-protection between different EV serotypes. In this study, we investigated the hazards of HFMD reinfection using Cox-proportional hazard model. Retrospective data of 95 209 HFMD cases in Wuhan during 2008-2015 was used. Kaplan-Meier survival methods and Cox-proportional hazard model were used to estimate the hazard probabilities. Of the all HFMD cases, about 2% experienced reinfection (1842/95 209). Kaplan-Meier curves revealed the reinfection risk sharply increased before 40 months from first infection. Higher hazards of reinfection were detected among those who were males, aged 3 years and below, scattered children, belonging to urban areas and first infected with coxsackievirus (CV)-A16 compared with their respective counterparts. Cox-proportional hazard model suggested that gender, age, group, living area and serotypes of first infection had significant effect on reinfection even after adjusting for potential confounding effects of other selected factors considered in the study. These results indicate that boys aged 3 years and below, especially those living in urban areas and first infected with CV-A16 are more prone to reinfection. Interventions should be imposed on these high-risk populations.

Molecular cloning of heat shock protein 10 (Hsp10) and 60 (Hsp60) cDNAs from Galeruca daurica (Coleoptera: Chrysomelidae) and their expression analysis.

Galeruca daurica (Joannis) is a new outbreak pest in the Inner Mongolia grasslands in northern China. Heat shock protein 10 and 60 (Hsp10 and Hsp60) genes of G. daurica, designated as GdHsp10 and GdHsp60, were cloned by rapid amplification of cDNA ends techniques. Sequence analysis showed that GdHsp10 and GdHsp60 encoded polypeptides of 104 and 573 amino acids, respectively. Sequence alignment and phylogenetic analysis clearly revealed that the amino acids of GdHsp10 and GdHsp60 had high homology and were clustered with other Hsp10 and Hsp60 genes in insects which are highly relative with G. daurica based on morphologic taxonomy. The mRNA expression analysis by real-time PCR revealed that GdHsp10 and GdHsp60 were expressed at all development stages and in all tissues examined, but expressed highest in eggs and in adults' abdomen; both heat and cold stresses could induce mRNA expression of GdHsp10 and GdHsp60 in the 2nd instar larvae; the two Hsp genes were expressed from high to low with the extension of treatment time in G. daurica eggs exposed to freezing point. Overall, our study provides useful information to understand temperature stress responses of Hsp60 and Hsp10 in G. daurica, and provides a basis to further study functions of Hsp60/Hsp10 relative to thermotolerance and cold hardiness mechanism.

Neutron flux measurements on a mock-up of a storage cask for high-level nuclear waste using 2.5 MeV neutrons.

To store and dispose spent nuclear fuel, shielding casks are employed to reduce the emitted radiation. To evaluate the exposure of employees handling such casks, Monte Carlo radiation transport codes can be employed. Nevertheless, to assess the reliability of these codes and nuclear data, experimental checks are required. In this study, a neutron generator (NG) producing neutrons of 2.5 MeV was employed to simulate neutrons produced in spent nuclear fuel. Different configurations of shielding layers of steel and polyethylene were positioned between the target of the NG and a NE-213 detector. The results of the measurements of neutron and γ radiation and the corresponding simulations with the code MCNP6 are presented. Details of the experimental set-up as well as neutron and photon flux spectra are provided as reference points for such NG investigations with shielding structures.

Reference gene selection and evaluation for expression analysis using qRT-PCR in Galeruca daurica (Joannis).

Quantitative real-time PCR (qRT-PCR) has been used extensively to analyze gene expression and decipher gene function. To obtain the optimal and stable normalization factors for qRT-PCR, selection and validation of reference genes should be conducted in diverse conditions. In insects, more and more studies confirmed the necessity and importance of reference gene selection. In this study, eight traditionally used reference genes in Galeruca daurica (Joannis) were assessed, using qRT-PCR, for suitability as normalization genes under different experimental conditions using four statistical programs: geNorm, Normfinder, BestKeeper and the comparative ΔC t method. The genes were ranked from the most stable to the least stable using RefFinder. The optimal suite of recommended reference genes was as follows: succinate dehydrogenase (SDHA) and tubulin-alpha (TUB-α) for temperature-treated larvae; ribosomal protein L32, SDHA and glutathione S-transferase were best for all developmental stages; ACT and TUB-α for male and female adults; SDHA and TUB-α were relatively stable and expressed in different tissues, both diapause and non-diapause adults. Reference gene evaluation was validated using expression of two target genes: the P450 CYP6 gene and the heat shock protein gene Hsp70. These results confirm the importance of custom reference gene selection when studies are conducted under diverse experimental conditions. A standardized qRT-PCR analysis procedure for gene functional studies is provided that could be useful in studies on other insect species.

Identification of odorant-binding protein genes in Galeruca daurica (Coleoptera: Chrysomelidae) and analysis of their expression profiles.

Odorant-binding proteins (OBPs) play a fundamental role in insect olfaction. In recent years, Galeruca daurica (Joannis) (Coleoptera: Chrysomelidae) has become one of the most important insect pests in the Inner Mongolian grasslands of China. This pest only feeds on the species of Allium plants, implying the central role of olfaction in its search for specific host plants. However, the olfaction-related proteins have not been investigated in this beetle. In this study, we identified 29 putative OBP genes, namely GdauOBP1-29, from the transcriptome database of G. daurica assembled in our laboratory by using RNA-Seq. All 29 genes had the full-length open reading frames except GdauOBP29, encoding proteins in length from 119 to 202 amino acids with their predicted molecular weights from 12 to 22 kDa with isoelectric points from 3.88 to 8.84. Predicted signal peptides consisting of 15-22 amino acid residues were found in all except GdauOBP6, GdauOBP13 and GdauOBP29. The amino acid sequence identity between the 29 OBPs ranged 8.33-71.83%. GdauOBP1-12 belongs to the Classic OBPs, while the others belong with the Minus-C OBPs. Phylogenetic analysis indicated that GdauOBPs are the closest to CbowOBPs from Colaphellus bowringi. RT-PCR and qRT-PCR analyses showed that all GdauOBPs were expressed in adult antennae, 11 of which with significant differences in their expression levels between males and females. Most GdauOBPs were also expressed in adult heads (without antennae), thoraxes, abdomens, legs and wings. Moreover, the expression levels of the GdauOBPs varied during the different development stages of G. daurica with most GdauOBPs expressed highly in the adult antennae but scarcely in eggs and pupae. These results provide insights for further research on the molecular mechanisms of chemical communications in G. daurica.

Phase I and biomarker study of OPB-51602, a novel signal transducer and activator of transcription (STAT) 3 inhibitor, in patients with refractory solid malignancies.

BACKGROUND: The aim of this study was to determine the maximum-tolerated dose (MTD), safety, pharmacokinetics, and pharmacodynamics of OPB-51602, an oral, direct signal transduction activator of transcription 3 (STAT3) inhibitor, in patients with refractory solid tumors. PATIENTS AND METHODS: Three cohorts were studied: cohort A, a sequential dose escalation of OPB-51602 administered intermittently (days 1-14 every 21 days); cohort B, an expansion cohort evaluating the dose lower than the MTD; cohort C, evaluating continuous daily dosing. RESULTS: Fifty-one patients were studied at 2, 4, and 5 mg per day dosing. The MTD was 5 mg; first-cycle dose-limiting toxicities (DLTs) were grade 3 hyponatremia in one patient, and grade 3 dehydration in another. Intermittent dosing of both 2 and 4 mg doses were tolerable, and the recommended phase II dose was 4 mg. Cohort B investigated 4 mg intermittently, whereas cohort C investigated 4 mg continuously. Common toxicities included fatigue, nausea/vomiting, diarrhea, anorexia, and early-onset peripheral neuropathy. Drug-induced pneumonitis occurred in two patients in cohort C. Continuous dosing was associated with a higher incidence of peripheral neuropathy and a lower mean relative dose intensity, compared with intermittent dosing. Steady-state pharmacokinetics was characterized by high oral clearance, mean elimination half-life ranging from 44 to 61 h, and a large terminal-phase volume of distribution. An active metabolite, OPB-51822, accumulated to a greater extent than OPB-51602. Flow cytometry of peripheral blood mononuclear cells demonstrated pSTAT3 (Tyr(705)) inhibition following exposure. Two patients achieved partial responses at 5 mg intermittently and 4 mg continuously; both had epidermal growth factor receptor (EGFR) mutation-positive non-small-cell lung cancer (NSCLC) with prior EGFR tyrosine kinase inhibitor exposure. CONCLUSION: OPB-51602 demonstrates promising antitumor activity, particularly in NSCLC. Its long half-life and poorer tolerability of continuous dosing, compared with intermittent dosing, suggest that less frequent dosing should be explored. CLINICALTRIALSGOV IDENTIFIER: NCT01184807.

KRAS and BRAF mutations are rare and related to DNA mismatch repair deficiency in gastric cancer from the East and the West: results from a large international multicentre study.

BACKGROUND: Inhibitors of the epidermal growth factor (EGFR) signaling pathway have a major role in the treatment of KRAS wild-type colorectal cancer patients. The EGFR pathway has been shown to be activated in gastric cancer (GC). However, published data on KRAS and BRAF mutation status is limited in GC and has not been compared between GC from different geographic regions. METHODS: The prevalence of KRAS and BRAF mutations was established in 712 GC: 278 GC from the United Kingdom, 230 GC from Japan and 204 GC from Singapore. The relationship between KRAS/BRAF mutation status, DNA mismatch repair (MMR) status, clinicopathological variables and overall survival was analysed. RESULTS: Overall, 30 (4.2%) GC carried a KRAS mutation. In total, 5.8% of the UK GC, 4% of Japan GC and 1.5% of Singapore GC were KRAS mutant. KRAS mutant GC had fewer lymph node metastases in the UK cohort (P=0.005) and were more frequent in elderly patients in the Japan cohort (P=0.034). KRAS mutations were more frequent in MMR-deficient GC in the UK and the Japanese cohort (P<0.05). A BRAF mutation was only detected in a single Japanese GC. CONCLUSIONS: This large multicentre study demonstrated that KRAS mutations and DNA MMR deficiency have a role in a small subgroup of GC irrespective of country of origin, suggesting that this subgroup of GC may have developed along a common pathway. Further studies need to establish whether concomitant mutations or amplifications of other EGFR signalling pathway genes may contribute to the activation of this pathway in GC.

[Lithocholic acid decreases mRNA stability of nuclear receptor PPARα by upregulating miR-21 expression in hepatoma HepG2 cells].

OBJECTIVE: To investigate the regulatory effects of lithocholic acid (LCA) on nuclear receptor peroxisome proliferatoractivated receptor-alpha (PPARα) mRNA stability at the post-transcriptional level. METHODS: PPARα 3'UTR luciferase reporter gene vectors were constructed and transfected into HepG2 cells to observe the changes in cellular luciferase activity in response to LCA treatments. Bioinformatic prediction and miRNA PCR array technique were used to identify the differentially expressed miRNAs induced by LCA and their potential binding sites on the 3'UTR. The binding sites (Mut1, Mut2 and Mut1+Mut2) were mutated to compare the changes in cellular luciferase activity following LCA treatment. Western blotting and RTqPCR were used to detect the activated signaling pathway and the expression levels of its downstream transcription factors in LCA-treated cells. The changes in PPARα protein expression level were detected in the cells following overexpression of the transcription factors. RESULTS: Treatment with 100 µmol/L LCA significantly reduced luciferase activity of PPARα 3'UTR1 and 3'UTR2 in HepG2 cells by more than 50% (P<0.01) and induced significant upregulation of miR-21 and miR-22, especially the former (by 2.35 folds, P<0.05). Two predicted miR-21-binding sites in the 3'UTR1 were mutated to construct Mut1, Mut2 and Mut1+Mut2 reporter gene vectors. LCA treatment down-regulated 3'UTR1 luciferase activity by 51%, while Mut1, Mut2, and Mut1+Mut2 were down-regulated by 37%, 39%, and 13%, respectively. LCA caused ERK1/2 phosphorylation and activation of the ERK1/2 signaling pathway, and treatment with 100 µmol/L LCA upregulated the expression of transcription factor early growth response 1 (EGR1) by 5.83 folds (P<0.01). Transient overexpression of EGR1 significantly decreased cellular PPARα protein levels (P<0.05). CONCLUSION: LCA reduces PPARα mRNA stability and thus decreases PPARα mRNA and protein expressions in hepatocytes by activating the ERK1/2 signaling pathway and upregulating EGR1 and miR-21, which targets 3'UTR regulatory region of PPARα mRNA.

[Etiological analysis on a foodborne disease outbreak caused by two serotypes of Salmonella].

Objective: To understand the etiological characteristics of 2 serotypes of Salmonella strains isolated from a foodborne disease outbreak. Methods: A total of 11 anal swabs of the cases, 13 suspected contaminated food and 10 environmental samples were collected from a foodborne disease outbreak occurred on September 8, 2022 in a school. The anal swabs were enriched with selenite brilliant green enrichment broth (SBG) and brain heart infusion broth (BHI) respectively. PCR detection and culture of common intestinal pathogens were carried out. The suspected food samples were tested according to national standards for food safety. Multiple suspected Salmonella colonies were obtained and selected for serotype determination and whole genome sequencing. Serotypes were determined based on the whole-genome sequence, and clustering analysis was performed based on core genome single nucleotide polymorphism (SNP). Results: The positive rates of Salmonella in anal swabs and suspected food samples were 9/11 and 5/13 respectively. Both Salmonella Uganda and Salmonella Idikan were isolated from 4 anal swabs and 4 suspected food samples. For the remaining samples, only Salmonella Uganda or Salmonella Idikan was isolated in each sample. The positive rate of Salmonella in 11 anal swabs of the cases after BHI enrichment for 12 h and 24 h were all 9/11 in real-time PCR, same to the culture results. Salmonella Uganda and Salmonella Idikan formed two independent and genetically distant lineages in the clustering tree based on core genome SNP, and 0-14 and 0-23 SNP were observed in Salmonella Uganda and Salmonella Idikan respectively. Conclusions: This foodborne disease outbreak was probably caused by Salmonella Uganda and Salmonella Idikan, which both exhibited strong genetic diversity. The PCR based pathogen screening strategy plus pathogen enrichment for cases' annal swabs can be used in the routine outbreak investigation.

Utility of atrial temporary pacing as an acute treatment for bradyarrhythmias and tachyarrhythmias in the intensive care setting with preservation of atrioventricular synchrony.

Temporary pacing is often used as an acute treatment of bradyarrhythmias and suppression of tachyarrhythmias. In patients with cardiogenic shock, loss of atrioventricular synchrony may worsen cardiovascular haemodynamics. We present a series where temporary right atrial pacing was used as an acute treatment for both bradyarrhythmias and tachyarrhythmias.