Novel insights into the quorum sensing system of Yersina enterocolitica 8081

Y. enterocolitica possesses an N-acyl-homoserine lactone (AHL)-dependent quorum sensing (QS) system which consists of the luxRI homologues, yenRI. The objective of this project was to increase our understanding of the QS system(s) in Y. enterocolitica by characterising the AHL-dependent system on a genus wide basis. Species-wide analysis of an Y. enterocolitica multi-strain genome database revealed an additional luxR homologue, we termed yeoR, which is present in the fully sequenced, highly virulent Y. enterocolitica 8081 strain and other USA strains but absent in UK strains. This indicates genetic differences in the QS systems of 'New World' (North America)and 'Old World' (Europe and Japan) strains of Y. enterocolitica. yeoR is an 'orphan' luxR homologue which is not associated with an adjacent luxI homologue. Consequent work described in this study focused on the Y. enterocolitica 8081 strain. LC-mass spectrometry of spent culture supernatants from Y. enterocolitica 8081 grown at 30°C identified 16 different AHLs, mainly N-(3-oxohexanoyl)-L-homoserine lactone (62.4%) and N-hexanoyl-L-homoserine lactone (27.4%). 11 of the 16 AHLs had not previously been documented for this bacterium, including N-(3-oxoheptanoyl)-Lhomo serine lactone (with an odd number of carbons in the acyl chain) which constituted 5.1 % and has rarely been documented previously. The lambda red recombinase method of mutagenesis was used for the rapid generation of QS mutants in Y. enterocolitica 8081. Seven QS mutants were generated, three were single mutants: deltayenI deltayenR and deltayeoR, three were double mutants: deltayenIyenR, deltayenIyeoR and deltayenRyeoR, and one was a triple mutant: deltayenIyenRycoR. Analysis of the QS mutants revealed that the level of AHLs synthesised at 30°C by the R mutants: deltayenR, deltayeoR and deltayenRyeoR, were similar to the wildtype while for AHL synthase mutants: deltayenI, deltayenIyenR, deltayenIycoR and deltayenIyenRyeoR, levels of AHL were greatly reduced compared with the wildtype but were not entirely abolished. Transcriptomic analysis of Y.enterocolitica 8081 wildtype and its QS mutants using microarray technology revealed many possible QS-related regulatory networks including motility, type III secretion system and High Pathogenicity Island. Microarray data, together with RQ-PCR, phenotypic studies and promoter fusion studies showed that QS is correlated with virulence regulation (expression of virF and tyeA), virulence factors (expression of yadA and invA) and maintenance of the pYVe plasmid (expression of repA and spyA), which have not been previously documented. In conclusion, this study revealed that Y. enterocolitica 8081 has a sophisticated QS system which consists of the AHL synthase, YenI and two LuxR-type regulators, YenR and Y coR working in tandem and sharing overlapping regulon. This system maybe linked to the hypervirulence in the 8081 strain but further work is needed to confirm this

Novel insights into the quorum sensing system of Yersina enterocolitica 8081

Y. enterocolitica possesses an N-acyl-homoserine lactone (AHL)-dependent quorum sensing (QS) system which consists of the luxRI homologues, yenRI. The objective of this project was to increase our understanding of the QS system(s) in Y. enterocolitica by characterising the AHL-dependent system on a genus wide basis. Species-wide analysis of an Y. enterocolitica multi-strain genome database revealed an additional luxR homologue, we termed yeoR, which is present in the fully sequenced, highly virulent Y. enterocolitica 8081 strain and other USA strains but absent in UK strains. This indicates genetic differences in the QS systems of 'New World' (North America)and 'Old World' (Europe and Japan) strains of Y. enterocolitica. yeoR is an 'orphan' luxR homologue which is not associated with an adjacent luxI homologue. Consequent work described in this study focused on the Y. enterocolitica 8081 strain. LC-mass spectrometry of spent culture supernatants from Y. enterocolitica 8081 grown at 30°C identified 16 different AHLs, mainly N-(3-oxohexanoyl)-L-homoserine lactone (62.4%) and N-hexanoyl-L-homoserine lactone (27.4%). 11 of the 16 AHLs had not previously been documented for this bacterium, including N-(3-oxoheptanoyl)-Lhomo serine lactone (with an odd number of carbons in the acyl chain) which constituted 5.1 % and has rarely been documented previously. The lambda red recombinase method of mutagenesis was used for the rapid generation of QS mutants in Y. enterocolitica 8081. Seven QS mutants were generated, three were single mutants: deltayenI deltayenR and deltayeoR, three were double mutants: deltayenIyenR, deltayenIyeoR and deltayenRyeoR, and one was a triple mutant: deltayenIyenRycoR. Analysis of the QS mutants revealed that the level of AHLs synthesised at 30°C by the R mutants: deltayenR, deltayeoR and deltayenRyeoR, were similar to the wildtype while for AHL synthase mutants: deltayenI, deltayenIyenR, deltayenIycoR and deltayenIyenRyeoR, levels of AHL were greatly reduced compared with the wildtype but were not entirely abolished. Transcriptomic analysis of Y.enterocolitica 8081 wildtype and its QS mutants using microarray technology revealed many possible QS-related regulatory networks including motility, type III secretion system and High Pathogenicity Island. Microarray data, together with RQ-PCR, phenotypic studies and promoter fusion studies showed that QS is correlated with virulence regulation (expression of virF and tyeA), virulence factors (expression of yadA and invA) and maintenance of the pYVe plasmid (expression of repA and spyA), which have not been previously documented. In conclusion, this study revealed that Y. enterocolitica 8081 has a sophisticated QS system which consists of the AHL synthase, YenI and two LuxR-type regulators, YenR and Y coR working in tandem and sharing overlapping regulon. This system maybe linked to the hypervirulence in the 8081 strain but further work is needed to confirm this

The quorum sensing system of Yersinia enterocolitica 8081 regulates swimming motility, host cell attachment, and virulence plasmid maintenance

Although Yersinia enterocolitica genomes are highly heterogeneous, they contain a conserved N-acylhomoserine lactone-dependent (AHL) quorum sensing (QS) system consisting of the luxR and luxI orthologs yenR and yenI respectively. Certain hypervirulent strains also contain a putative orphan luxR gene, ycoR, that is not linked to an AHL synthase. To explore the contribution of yenR/yenI/ycoR to QS-dependent phenotypes in Yersinia enterocolitica strain 8081, single and multiple mutants were constructed. AHL profiling identified N-(3-oxohexanoyl) homoserine lactone, N-hexanoylhomoserine lactone, and N-(3-oxoseptanoyl) homoserine lactone as the most abundant. The AHL profiles of the yenR, ycoR and yenR/ycoR mutants were similar to the parent suggesting that the two LuxR homologues do not regulate AHL production while the yenI mutants were AHL-negative. A role for QS in swimming motility and cell attachment was demonstrated. Down-regulation of the virulence plasmid partition gene, spyA, in yenI and yenI/yenR/ycoR mutants is consistent with the greater loss of the Y. enterocolitica pYVe virulence plasmid in the yenI mutant during serial passage at 37 C but not at 22 C. A role for QS-regulated spyA in virulence plasmid maintenance is suggested

Molecular Cloning of a New Immunomodulatory Protein from Anoectochilus formosanus which Induces B Cell IgM Secretion through a T-Independent Mechanism

An immunomodulatory protein (IPAF) was purified and cloned from Anoectochilus formosanus, an Orchidaceae herbal plant in Asia. The major targeting immune cells of IPAF and its modulating effects toward B lymphocytes were investigated. Rapid amplification of cDNA ends (RACE) was conducted to clone the IPAF gene, and the obtained sequence was BLAST compared on the NCBI database. MACS-purified mouse T and B lymphocytes were stimulated with IPAF and the cell proliferation, activation, and Igs production were examined. IPAF comprised a 25 amino acids signal peptide and a 138 amino acids protein which was homologous to the lectins from Orchidaceae plant. IPAF selectively induced the cell proliferation in mouse splenic B lymphocytes but not T lymphocytes. The IPAF-induced B cells exhibited increased CD69 and MHC class II expression, and a dose- and time-dependent enhancement in IgM production. These results suggested potential benefits of IPAF to strengthen the humoral immunity

Intravenous alteplase for stroke with unknown time of onset guided by advanced imaging: systematic review and meta-analysis of individual patient data

Background: Patients who have had a stroke with unknown time of onset have been previously excluded from thrombolysis. We aimed to establish whether intravenous alteplase is safe and effective in such patients when salvageable tissue has been identified with imaging biomarkers. Methods: We did a systematic review and meta-analysis of individual patient data for trials published before Sept 21, 2020. Randomised trials of intravenous alteplase versus standard of care or placebo in adults with stroke with unknown time of onset with perfusion-diffusion MRI, perfusion CT, or MRI with diffusion weighted imaging-fluid attenuated inversion recovery (DWI-FLAIR) mismatch were eligible. The primary outcome was favourable functional outcome (score of 0–1 on the modified Rankin Scale [mRS]) at 90 days indicating no disability using an unconditional mixed-effect logistic-regression model fitted to estimate the treatment effect. Secondary outcomes were mRS shift towards a better functional outcome and independent outcome (mRS 0–2) at 90 days. Safety outcomes included death, severe disability or death (mRS score 4–6), and symptomatic intracranial haemorrhage. This study is registered with PROSPERO, CRD42020166903. Findings: Of 249 identified abstracts, four trials met our eligibility criteria for inclusion: WAKE-UP, EXTEND, THAWS, and ECASS-4. The four trials provided individual patient data for 843 individuals, of whom 429 (51%) were assigned to alteplase and 414 (49%) to placebo or standard care. A favourable outcome occurred in 199 (47%) of 420 patients with alteplase and in 160 (39%) of 409 patients among controls (adjusted odds ratio [OR] 1·49 [95% CI 1·10–2·03]; p=0·011), with low heterogeneity across studies (I2=27%). Alteplase was associated with a significant shift towards better functional outcome (adjusted common OR 1·38 [95% CI 1·05–1·80]; p=0·019), and a higher odds of independent outcome (adjusted OR 1·50 [1·06–2·12]; p=0·022). In the alteplase group, 90 (21%) patients were severely disabled or died (mRS score 4–6), compared with 102 (25%) patients in the control group (adjusted OR 0·76 [0·52–1·11]; p=0·15). 27 (6%) patients died in the alteplase group and 14 (3%) patients died among controls (adjusted OR 2·06 [1·03–4·09]; p=0·040). The prevalence of symptomatic intracranial haemorrhage was higher in the alteplase group than among controls (11 [3%] vs two [<1%], adjusted OR 5·58 [1·22–25·50]; p=0·024). Interpretation: In patients who have had a stroke with unknown time of onset with a DWI-FLAIR or perfusion mismatch, intravenous alteplase resulted in better functional outcome at 90 days than placebo or standard care. A net benefit was observed for all functional outcomes despite an increased risk of symptomatic intracranial haemorrhage. Although there were more deaths with alteplase than placebo, there were fewer cases of severe disability or death. Funding: None

Clinical Characteristics and In-Hospital Outcomes in Dialysis Patients with Septic Arthritis

Background and Objectives: Septic arthritis is a medical emergency associated with high morbidity and mortality. The incidence rate of septic arthritis among dialysis patients is higher than the general population, and dialysis patients with bacteremia frequently experience adverse outcomes. The aim of this study was to identify the clinical features and risk factors for longer hospital length of stay (LOS), positive blood culture, and in-hospital mortality in dialysis patients with septic arthritis. Materials and Methods: The medical records of 52 septic arthritis dialysis patients admitted to our hospital from 1 January 2009 to 31 December 2020 were analyzed. The primary outcomes were bacteremia and in-hospital mortality. Variables were compared, and risk factors were evaluated using linear and logistic regression models. Results: Twelve (23.1%) patients had positive blood cultures. A tunneled cuffed catheter for dialysis access was used in eight (15.4%) patients, and its usage rate was significantly higher in patients with positive blood culture than in those with negative blood culture (41.7 vs. 7.5%, p = 0.011). Fever was present in 15 (28.8%) patients, and was significantly more frequent in patients with positive blood culture (58.3 vs. 20%, p = 0.025). The most frequently involved site was the hip (n = 21, 40.4%). The most common causative pathogen was Gram-positive cocci, with MRSA (n = 7, 58.3%) being dominant. The mean LOS was 29.9 ± 25.1 days. The tunneled cuffed catheter was a significant predictor of longer LOS (Coef = 0.49; Cl 0.25–0.74; p p = 0.037) and tunneled cuffed catheter (OR = 7.60; Cl 1.31–44.02; p = 0.024). The predictor of mortality was tunneled cuffed catheter (OR = 14.33; Cl 1.12–183.18; p = 0.041). Conclusions: In the dialysis population, patients with tunneled cuffed catheter for dialysis access had a significantly longer hospital LOS. Tunneled cuffed catheter and fever were independent predictors of positive blood culture, and tunneled cuffed catheter was the predictor of in-hospital mortality. The recognition of the associated factors allows for risk stratification and determination of the optimal treatment plan in dialysis patients with septic arthritis

Hormone therapy for patients with advanced or recurrent endometrial cancer

The “gold standard” treatment for endometrial cancer is completely staged surgery, followed by radiation or chemotherapy, based on the final pathological surgical stage and requirements. In the primary treatment of endometrial cancers, hormones are rarely taken into consideration after primary surgery. Primary treatment with hormones to preserve fertility in younger women with endometrial cancer is an attractive option, and many successful cases have been reported, although the majority of them finally received definite therapy, including total hysterectomy. The role of hormone therapy is often delayed in recurrent disease; response rates to progestins and tamoxifen or aromatase inhibitors in advanced/recurrent endometrial cancers are approximately 15–20% and nearly ≤10%, respectively. This review is focused on updated information and recent knowledge on the use of hormones in the management of women with advanced or recurrent endometrial cancers

Diallyl trisulfide (DATS) ameliorates cardiac fibrosis in a rat model of diabetes

Cardiac fibrosis is a prevalent pathological complication observed in individuals with diabetes. Diallyl trisulfide (DATS), a bioactive compound found in garlic oil, has been reported to exhibit anti-inflammatory and anti-apoptotic effects in various cardiovascular diseases. However, its potential impact on cardiac fibrosis, particularly in the context of diabetes, remains unknown. In the neonatal rat ventricular fibroblasts (NRVFs) model, our results demonstrated that DATS effectively attenuated advanced glycation end products (AGEs)-induced activation of Nuclear factor kappa B (NF-κB) and Smad2/3 signaling pathways, leading to a reduction in the downstream secretion of pro-inflammatory cytokines and collagen synthesis. In the in vivo study using echocardiographic assessment, administration of DATS significantly ameliorated cardiac dysfunction induced by streptozotocin (STZ). Taken together, our findings highlight the potential of DATS as a promising therapeutic agent for mitigating cardiac fibrosis associated with diabetes, emphasizing its potential clinical relevance in the prevention and management of diabetic cardiomyopathy