Characteristics and phylogenetic distribution of megaplasmids and prediction of a putative chromid in Pseudomonas aeruginosa.

Research on megaplasmids that contribute to the spread of antimicrobial resistance (AMR) in Pseudomonas aeruginosa strains has grown in recent years due to the now widely used technologies allowing long-read sequencing. Here, we systematically analyzed distinct and consistent genetic characteristics of megaplasmids found in P. aeruginosa. Our data provide information on their phylogenetic distribution and hypotheses tracing the potential evolutionary paths of megaplasmids. Most of the megaplasmids we found belong to the IncP-2-type, with conserved and syntenic genetic backbones carrying modules of genes associated with chemotaxis apparatus, tellurite resistance and plasmid replication, segregation, and transmission. Extensively variable regions harbor abundant AMR genes, especially those encoding ß-lactamases such as VIM-2, IMP-45, and KPC variants, which are high-risk elements in nosocomial infection. IncP-2 megaplasmids act as effective vehicles transmitting AMR genes to diverse regions. One evolutionary model of the origin of megaplasmids claims that chromids can develop from megaplasmids. These chromids have been characterized as an intermediate between a megaplasmid and a chromosome, also containing core genes that can be found on the chromosome but not on the megaplasmid. Using in silico prediction, we identified the "PABCH45 unnamed replicon" as a putative chromid in P. aeruginosa, which shows a much higher similarity and closer phylogenetic relationship to chromosomes than to megaplasmids while also encoding plasmid-like partition genes. We propose that such a chromid could facilitate genome expansion, allowing for more rapid adaptations to novel ecological niches or selective conditions, in comparison to megaplasmids.

Advances in the study of IL-17 in neurological diseases and mental disorders.

Interleukin-17 (IL-17), a cytokine characteristically secreted by T helper 17 (Th17) cells, has attracted increasing attention in recent years because of its importance in the pathogenesis of many autoimmune or chronic inflammatory diseases. Recent studies have shown that neurological diseases and mental disorders are closely related to immune function, and varying degrees of immune dysregulation may disrupt normal expression of immune molecules at critical stages of neural development. Starting from relevant mechanisms affecting immune regulation, this article reviews the research progress of IL-17 in a selected group of neurological diseases and mental disorders (autism spectrum disorder, Alzheimer's disease, epilepsy, and depression) from the perspective of neuroinflammation and the microbiota-gut-brain axis, summarizes the commonalities, and provides a prospective outlook of target application in disease treatment.

Characterization of two novel VIM-type metallo-ß-lactamases, VIM-84 and VIM-85, associated with the spread of IncP-2 megaplasmids in Pseudomonas aeruginosa.

This study aimed to characterize two novel VIM-type metallo-ß-lactamases, VIM-84 and VIM-85, and reveal the important role of the IncP-2 type megaplasmids in the spread of antimicrobial resistance (AMR) genes. VIM-84 and VIM-85 were encoded by two novel genes bla VIM-84 and bla VIM-85 which showed similarity to bla VIM-24. Both bla VIM-84 and bla VIM-85 are harbored into class 1 integrons embedded into the Tn1403 transposon. The bla VIM-85 gene was identified in a megaplasmid, which was related to 17 megaplasmid sequences with sizes larger than 430 kb, deposited previously in Genbank. A comparative analysis of complete plasmid sequences showed highly similar backbone regions and various AMR genes. A phylogenetic tree revealed that these megaplasmids, which were widely distributed globally, were vehicles for the spread of AMR genes. The bla VIM-24, bla VIM-84, and bla VIM-85 genes were cloned into pGK1900, and the recombinant vectors were further transformed into Escherichia coli DH5α and Pseudomonas aeruginosa PAO1. The antimicrobial susceptibility test of the cloning strains showed high levels of resistance to ß-lactams while they remained susceptible to aztreonam. Enzymatic tests revealed that both, VIM-84 and VIM-85, exhibited higher activity in hydrolyzing ß-lactams compared to VIM-24. A D117N mutation found in VIM-24 affected binding to the antibiotics. IMPORTANCE The metallo-ß-lactamases-producing Pseudomonas aeruginosa strains play an important role in hospital outbreaks and the VIM-type enzyme is the most prevalent in European countries. Two novel VIM-type enzymes in our study, VIM-84 and VIM-85, have higher levels of resistance to ß-lactams and greater hydrolytic activities for most ß-lactams compared with VIM-24. Both bla VIM-84 and bla VIM-85 are harbored into class 1 integrons embedded into the Tn1403 transposon. Notably, the genes bla VIM-85 are carried by three different IncP-2-type megaplasmids which are distributed locally and appear responsible for the spread of antimicrobial resistance genes in hospital settings.

Pseudomonas aeruginosa High-Risk Sequence Type 463 Co-Producing KPC-2 and AFM-1 Carbapenemases, China, 2020-2022.

We report the clonal spread and evolution of high-risk Pseudomonas aeruginosa sequence type 463 co-producing KPC-2 and AFM-1 carbapenemases isolated from hospital patients in China during 2020-2022. Those strains pose a substantial public health threat and surveillance and stricter infection-control measures are essential to prevent further infections.

Emergence of carbapenem-resistant Enterobacter hormaechei ST93 plasmids co-harbouring blaNDM-1, blaKPC-2, and mcr-9 in bloodstream infection.

OBJECTIVES: We isolated a strain of Enterobacter hormaechei, ECC2783, co-harbouring blaNDM-1, blaKPC-2 and mcr-9 plasmids from a bloodstream infection and investigated its biological features. METHODS: The presence of carbapenemase genes and mcr-9 was confirmed by polymerase chain reaction amplification. Whole genome sequencing and genomic analysis were performed on ECC2783. Experiments assessing the conjugation and stability of plasmids carrying the carbapenemase gene were performed. We also performed a colistin resistance induction experiment and studied the fitness cost of transconjugants. RESULTS: ECC2783 has an extensive drug resistance phenotype. Multilocus sequence typing analysis results showed that ECC2783 belongs to sequence type 93. Bioinformatics analysis confirmed that ECC2783 has four plasmids, of which pECC2783_a, carrying mcr-9, is the IncHI2 type, and pECC2783_c, carrying blaNDM-1, is the IncX3 type. pECC2783_d, carrying blaKPC-2, is an unclassified type. We successfully obtained two transconjugants (J53/ECC2783_1, carrying blaNDM-1, and J53/ECC2783_2, carrying blaKPC-2 and blaNDM-1). There was no statistically significant difference in the relative growth rate between J53 and J53/ECC2783_2. CONCLUSION: For the first time, we isolated carbapenem-resistant E. hormaechei plasmids co-harbouring blaNDM-1, blaKPC-2, and mcr-9 from a patient with a blood stream infection. This isolate has a survival advantage in a hospital environment, and its clinical monitoring should be strengthened.

An XDR Pseudomonas aeruginosa ST463 Strain with an IncP-2 Plasmid Containing a Novel Transposon Tn6485f Encoding blaIMP-45 and blaAFM-1 and a Second Plasmid with Two Copies of blaKPC-2.

The increased carbapenem resistance among Pseudomonas aeruginosa has become a serious health issue worldwide. We reported an extensively drug-resistant (XDR) P. aeruginosa PA30 isolate which belonged to sequence type ST463 and contained an IncP-2 plasmid (pPA30_1) carrying two genes, namely, blaIMP-45 and blaAFM-1, which encoded the metallo-ß-lactamases AFM-1 and IMP-45, respectively. Additionally, the strain had a plasmid (pPA30_2) with two copies of the blaKPC-2 genes embedded. The plasmid pPA30_1 was highly similar to the previously reported plasmid pHS17-127, which has the same genetic architecture. This plasmid contained blaIMP-45, located in a second gene cassette of the integron In786, carried by a Tn1403-derivative transposon acquiring an ISCR27n3-blaAFM-1 structure. Interestingly, the transposon in pPA30_1 acquired an extra ISCR1-qnrVC6 module and formed a novel transposon, which was subsequently annotated as Tn6485f. The blaKPC-2 genes in pPA30_2 underwent duplication due to the inversion of the IS26-blaKPC-2-IS26 element, which resulted in two copies of blaKPC-2. IMPORTANCE The ST463 clone is an emerging high-risk sequence type that is spreading with blaKPC-2-containing plasmids. The core blaKPC-2 genetic platform is ISKpn27-blaKPC-2-ISKpn6 in almost all samples, and the adjacent region beyond the core platform varies by IS26-mediated inversion or duplication events, amplifying the blaKPC-2 gene copies. The ST463 P. aeruginosa strain PA30 in our study contains another two metallo-ß-lactamase genes, namely, blaIMP-45 and blaAFM-1, in a novel transposon Tn6485f that is harbored by the IncP-2 megaplasmid. The pPA30_1 carrying blaIMP-45 and blaAFM-1 is highly related to pHS17-127 from the ST369 P. aeruginosa strain, indicating the putative dissemination of the megaplasmid between different clones.

In vivo acquisition of blaKPC-2 with low biological cost in blaAFM-1-harboring ST463 hypervirulent Pseudomonas aeruginosa from a patient with hematologic malignancy.

OBJECTIVES: Klebsiella pneumoniae carbapenemase (KPC)-producing sequence type (ST) 463 Pseudomonas aeruginosa are increasingly prevalent in China. This study aims to investigate how blaKPC-2 is acquired in ST463 P. aeruginosa during antimicrobial therapy. METHODS: Two extensively drug-resistant P. aeruginosa strains, B1122 and U1121, were respectively isolated from blood and urine of a patient during carbapenem therapy. Whole-genome sequences were obtained, and minimum inhibitory concentrations (MICs) were determined. Plasmid transferability and stability were examined. Bacterial growth kinetics, biofilm formation, and virulence level was assessed. RESULTS: U1121 and B1122 were only susceptible to amikacin and intermediately susceptible to colistin. They were isogenic ST463 P. aeruginosa strains and shared the same chromosome-encoded resistance genes, including blaAFM-1. This is the first report of chromosomal integration of blaAFM-1 in P. aeruginosa mediated by ISCR29. pU1121 and pB1122, which shared almost identical backbone, were the sole plasmids in U1121 and B1122, respectively, differing by an insertion region containing two copies of blaKPC-2 genes observed on pU1121. Sequence alignment revealed that pU1121 might evolve in vivo from pB1122 via IS26-mediated continuous genetic rearrangement in response to selective challenge from carbapenem. pU1121 was not self-transmissible and could be stably maintained in the host in the absence of antibiotic. Both U1121 and B1122 were hypervirulent, and no differences on virulence were recorded between them. However, U1121 exhibited significant impaired growth in comparison with B1122. CONCLUSION: ST463 P. aeruginosa can capture blaKPC-2 through horizontal transfer of insertion sequence under antibiotic selection pressure, which does decrease the fitness but does not impair the virulence of the ancestor.

Plasmid-Borne AFM Alleles in Pseudomonas aeruginosa Clinical Isolates from China.

Carbapenem-resistant Pseudomonas aeruginosa (CRPA) is a pathogen of global concern due to the fact that therapeutic drugs are limited. Metallo-ß-lactamase (MBL)-producing P. aeruginosa has become a critical part of CRPA. Alcaligenes faecalis metallo-ß-lactamase (AFM) is a newly identified subclass B1b MBL. In this study, 487 P. aeruginosa strains isolated from patients and the environment in an intensive care unit were screened for AFM alleles. Five AFM-producing strains were identified, including four AFM-2-producing strains (ST262) and one AFM-4-producing strain (ST671). AFM-2-producing strains were isolated from rectal and throat swabs, and AFM-4-producing strains were isolated from the water sink. The blaAFM-2 carrying plasmids belonged to the IncP-2 type, while the blaAFM-4 carrying plasmid pAR19438 was a pSTY-like megaplasmid. Plasmid pAR19438 was acquired blaAFM-4 by the integration of the Tn1403-like transposon. All blaAFM genes were embedded in an ISCR29-blaAFM unit core module flanked by class 1 integrons. The core module of blaAFM-2 was ISCR29-ΔgroL-blaAFM-2-bleMBL-ΔtrpF-ΔISCR, while the core module of blaAFM-4 was ISCR29-ΔgroL-blaAFM-2-bleMBL-ΔtrpF-ISCR-msrB-msrA-yfcG-corA-ΔISCR. The flanking sequences of ISCR29-blaAFM units also differed. The expression of AFM-2 and AFM-4 in DH5α and PAO1 illustrated the same effect for the evaluation of the MICs of ß-lactams, except for aztreonam. Identification of AFM-4 underscores that the quick spread and emerging development of mutants of MBLs require continuous surveillance in P. aeruginosa. IMPORTANCE Acquiring metallo-ß-lactamase genes is one of the important carbapenem resistance mechanisms of P. aeruginosa. Alcaligenes faecalis metallo-ß-lactamase is a newly identified metallo-ß-lactamase, the prevalence and genetic context of which need to be explored. In this study, we identified AFM-producing P. aeruginosa strains among clinical isolates and found a new mutant of AFM, AFM-4. The blaAFM-4 carrying plasmid pAR19438 was a pSTY-like megaplasmid, unlike the plasmids encoding other blaAFM alleles. The genetic context of blaAFM-4 was also different. However, AFM-2 and AFM-4 had the same impacts on antibiotic susceptibility. The presence and transmission of AFM alleles in P. aeruginosa pose a challenge to clinical practice.

Bloodstream Infections Caused by Klebsiella pneumoniae Carbapenemase-Producing P. aeruginosa Sequence Type 463, Associated With High Mortality Rates in China: A Retrospective Cohort Study.

Objectives: Recently, KPC-producing P. aeruginosa has rapidly emerged and expanded in East China. Here we described the clinical impact and characteristics of bloodstream infections (BSIs) from the dominant KPC-producing CRPA belonging to Sequence Type (ST) 463. Methods: Retrospective cohort study was performed with CRPA BSI cases from 2019 to 2020 in a hospital in East China. Clinical characteristics, risk factors, and all-course mortality were evaluated. All CRPA isolates had whole-genome sequencing, antimicrobial susceptibility testing, and serum resistance assay. Representative isolates were tested for virulence in a Galleria mellonella infection model. Results: Among the 50 CRPA BSI cases, ST463 predominated (48.0%). In multivariate analysis, we found three independent risk factors for fatal outcome: KPC carriage (OR 4.8; CI95% 1.0-23.7; P = 0.05), Pitt bacteremia score (OR 1.3; CI95% 1.0-1.6; P = 0.02), and underlying hematological disease (OR 8.5; CI95% 1.6-46.4; P = 0.01). The baseline clinical variables were not statistically different across STs, however the 28-day mortality was significantly higher in ST463 cases than that in non-ST463 cases (66.7% vs 33.3%, P = 0.03). ExoU and exoS virulence genes coexisted in all ST463 isolates, and the carbapenem resistant gene blaKPC were produced in almost all ST463 isolates, significantly higher than in the non-ST463 group(95.8% vs 7.7%, P<0.001). ST463 CRPA isolates also showed higher resistance rates to antipseudomonal cephalosporins, monobactam, and fluoroquinolones. And ST463 CRPA was confirmed hypervirulence in the larvae model. The genome of one ST463 CRPA strain showed that the blaKPC-2 gene was the sole resistance gene located on a 41,104bp plasmid pZYPA01, carried on a 7-kb composite transposon-like element flanked by two IS26 elements (IS26-Tn3-tnpA-ISKpn27-blaKPC-2-ISKpn6-IS26). Plasmid from various species presented core blaKPC-2 was franked by mobile genetic element ISKpn27 and ISKpn6. Conclusions: In the ST463 CRPA BSI cohort, the mortality rates were higher than those in the non-ST463 CRPA BSI. The ST463 CRPA clone coharboring the blaKPC and exoU/exoS genes emerged and spread in East China, which might develop to a new threat in the clinic. Our results suggest that the surveillance of the new high-risk clone, ST463 CRPA, should be strengthened in China, even worldwide in the future.

Successful Treatment of Severe Post-craniotomy Meningitis Caused by an Escherichia coli Sequence Type 410 Strain Coharboring bla NDM - 5 and bla CTX - M - 65.

Intracranial infections caused by multidrug-resistant Gram-negative bacterium have led to considerable mortality due to extremely limited treatment options. Herein, we firstly reported a clinical carbapenem-resistant Escherichia coli isolate coharboring bla NDM - 5 and bla CTX - M - 65 from a patient with post-craniotomy meningitis. The carbapenem-resistant Escherichia coli strain CNEC001 belonging to Sequence Type 410 was only susceptible to amikacin and tigecycline, both of which have poor penetration through the blood-brain barrier (BBB). The bla CTX - M - 65 gene was expressed on a 135,794 bp IncY plasmid. The bla NDM - 5 gene was located on a genomic island region of an IncX3-type plasmid pNDM5-CNEC001. Based on the characteristics of the strain, we presented the successful treatment protocol of intravenous (IV) tigecycline and amikacin combined with intrathecal (ITH) amikacin in this study. Intracranial infection caused by Escherichia coli coharboring bla NDM - 5 and bla CTX - M - 65 is rare and fatal. Continuous surveillance and infection control measures for such strain need critical attention in clinical settings.

Clinical Characteristics and Risk Factors for Bloodstream Infection Due to Carbapenem-Resistant Klebsiella pneumoniae in Patients with Hematologic Malignancies.

PURPOSE: The aim was to examine the clinical characteristics and risk factors for bloodstream infection (BSI) due to carbapenem-resistant Klebsiella pneumoniae (CRKP) in patients with hematologic malignancies. MATERIALS AND METHODS: A single-centre, retrospective case-control study representing 734 patients with hematologic malignancies between January 1, 2017, and December 31, 2018, was conducted. Demographic and clinical data were collected from the hospital electronic medical records system. RESULTS: Among the 734 patients with hematologic malignancies, 3% (22/734) of the patients developed CRKP BSI during their hospitalization. Overall 28-day all-cause mortality reached 77.3% (17/22). Patients with Pitt bacteremia score (PBS) >4, pneumonia and septic shock were more frequent in the non-survivors versus the survivors. Compared with the non-survivors in antimicrobial treatment, combination therapy of tigecycline and polymyxin B was more common in the survivors. The independent risk factors associated with CRKP BSI were CRKP rectal colonization (OR, 11.067; CI=4.43-27.644; P<0.001; 3 points), severe neutropenia (OR, 4.095; CI=0.876-19.141; P=0.073; 1 point) and invasive mechanical ventilation (IMV) within the previous 30 days to onset of BSI (OR, 18.444; CI=1.787-190.343; P=0.014; 4 points). The total risk score of ≥5 indicated that the probability of CRKP BSI occurrence was above 48%. CONCLUSION: CRKP BSI in patients with hematologic malignancies is associated with high mortality. The risk factor-based prediction model might help clinicians to start prompt effective anti-infective therapy in patients with suspicion of CRKP BSI and improve outcomes.

Clinical and Microbiological Characteristics of Community-Onset Carbapenem-Resistant Enterobacteriaceae Isolates.

OBJECTIVE: The aim of this study was to investigate the clinical and microbiological features of community-onset CRE (CO-CRE) obtained from outpatients at a tertiary hospital in China. PATIENTS AND METHODS: We isolated 64 CRE strains from outpatients and divided them into three groups: 36 hospital-acquired CRE (HA-CRE), 28 CO-CRE including 15 community-acquired CRE (CA-CRE) and 13 healthcare-associated CRE (HCA-CRE). Clinical information was collected. The antibiotic susceptibilities of the 28 CO-CRE strains were tested. Whole-genome sequencing (WGS) was conducted, and then drug resistance gene analysis was performed. CgMLST and SNP comparisons were used to analyze the genomic relationship with E. coli and K. pneumoniae strains, respectively. RESULTS: In this study, the 28 CO-CRE isolates included K. pneumoniae (53.6%), E. coli (28.6%), E. cloacae (7.1%), C. freundii (7.1%) and E. asburiae (3.6%). The CO-CRE isolates were mainly isolated from urine samples (75%). The ceftazidime/avibactam resistance rate of community-onset E. coli was significantly higher than that of community-onset K. pneumoniae, while the aztreonam, ciprofloxacin, levofloxacin, and chloramphenicol resistance rates were significantly lower (P<0.05). Thirteen of the 15 K. pneumoniae strains belonged to ST11 containing blaKPC-2. Correspondingly, 8 E. coli strains belonged to 7 STs, and they all were NDM producers. K. pneumoniae belonged to two major clusters, while E. coli was sporadic. The number of SNPs separating ST11 K. pneumoniae isolates ranged from 7 to 2154. CONCLUSION: Community-onset CRE is not rare, and the dissemination of E. coli was sporadic while K. pneumoniae was clonal spread with similar STs as HA-CRE. Active surveillance of CRE in the community setting is in demand.

Clinical and microbiological characteristics of cryptococcosis at an university hospital in China from 2013 to 2017

ABSTRACT Background: This study aims to explore the epidemiology, clinical profile and strain characteristics of cryptococcosis from 2013 to 2017 in a major teaching hospital in China. Methods: Trends in antifungal drug susceptibility of 217 consecutive non-repetitive cryptococcal isolates collected from patients of an university hospital in China were analyzed between 2013 and 2017. Of those, 98 isolates were conserved for identification by internal transcribed spacer (ITS) sequencing and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) system. Multilocus sequence typing (MLST) was used to designate molecular types. Clinical characteristics of the 98 patients with cryptococcosis during the period of 2013-2017 were retrospectively evaluated. Results: There was a trend for gradual increase in the MIC range of fluconazole was from 2013 to 2017. The conserved 98 clinical cryptococcal isolates included 97 C. neoformans and one C. gattii, and 90 (91.8%) isolates belonged to ST5 genotype VNI. Out of the 98 patients with cryptococcosis, 28 (28.6%) were HIV-infected and 32 (32.7%) had no underlying diseases. HIV-infected patients had higher mortality than HIV-uninfected patients (28.6% vs 14.3%, p = 0.147). Conclusions: Most of the patients with cryptococcosis were not HIV-infected in this study, while patients with HIV had a higher mortality. Reduced susceptibility to fluconazole was observed among C. neoformans isolates, most of them belonged to ST5 genotype VNI having an impact on the effective dose of fluconazole.

Clinical and microbiological characteristics of cryptococcosis at an university hospital in China from 2013 to 2017.

BACKGROUND: This study aims to explore the epidemiology, clinical profile and strain characteristics of cryptococcosis from 2013 to 2017 in a major teaching hospital in China. METHODS: Trends in antifungal drug susceptibility of 217 consecutive non-repetitive cryptococcal isolates collected from patients of an university hospital in China were analyzed between 2013 and 2017. Of those, 98 isolates were conserved for identification by internal transcribed spacer (ITS) sequencing and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) system. Multilocus sequence typing (MLST) was used to designate molecular types. Clinical characteristics of the 98 patients with cryptococcosis during the period of 2013-2017 were retrospectively evaluated. RESULTS: There was a trend for gradual increase in the MIC range of fluconazole was from 2013 to 2017. The conserved 98 clinical cryptococcal isolates included 97 C. neoformans and one C. gattii, and 90 (91.8%) isolates belonged to ST5 genotype VNI. Out of the 98 patients with cryptococcosis, 28 (28.6%) were HIV-infected and 32 (32.7%) had no underlying diseases. HIV-infected patients had higher mortality than HIV-uninfected patients (28.6% vs 14.3%, p=0.147). CONCLUSIONS: Most of the patients with cryptococcosis were not HIV-infected in this study, while patients with HIV had a higher mortality. Reduced susceptibility to fluconazole was observed among C. neoformans isolates, most of them belonged to ST5 genotype VNI having an impact on the effective dose of fluconazole.

Protein kinase C-ε contributes to a chronic inhibitory effect of IL-1ß on voltage-gated sodium channels in mice with febrile seizure.

This study aimed to understand the role of Interleukin-1ß in mouse febrile seizures. To investigate the chronic effects of raised Interleukin-1ß on seizures, the sodium currents of hippocampal neurons were recorded by whole-cell voltage clamp. Interleukin-1ß inhibited sodium currents in mouse hippocampal neurons and verified that protein kinase C epsilon contributed to the effect of Interleukin-1ß exposure. The inhibitory effect was also identified in neurons from a protein kinase C epsilon null mutant mouse. Action potentials were recorded using a ramp depolarizing current. Peak spike depolarization was significantly reduced by Interleukin-1ß treatment, and was abolished following the administration of a protein kinase C epsilon inhibitor, εV1-2. However, neither Interleukin-1ß nor εV1-2 had any significant effect on spike threshold. Interleukin-1ß reduced the amplitude of action potentials due to its inhibitory effect on sodium channels. This is hypothesised to decrease the release of presynaptic transmitters of neuroexcitability, thus exerting a neuroprotective role in excitotoxicity. To ascertain the role of protein kinase C epsilon on febrile seizures in vivo, a heated water-bath model was used to identify susceptible mice. It was found that protein kinase C epsilon reduced susceptibility to, and frequency of, febrile seizure onset. This may be related to the neuroprotective effect of Interleukin-1ß on hippocampal neurons.

Cryptococcal pleuritis with pleural effusion as the only clinical presentation in a patient with hepatic cirrhosis: A case report and literature review.

RATIONALE: Cryptococcosis is a significant life-threatening fungal infection in worldwide, mainly reported in immunocompromised patients. Pleural effusion presentation of cryptococcal infection as the only clinical presentation is rarely seen in pulmonary cryptococcosis, which may lead to be misdiagnosed, and the study on this subject will provide further insights. PATIENT CONCERNS: A 64-year-old man was hospitalized in our department and diagnosed as hepatic B cirrhosis. A computed tomography (CT) of the thorax showed a massive right pleural effusion without pulmonary parenchymal abnormalities. He was started on empirical treatment for pleural tuberculosis (TB). However, during his hospitalization, a right pleural effusion developed and fever was not controlled. DIAGNOSES: On day 14 admission, pleural fluid cultured positive for Cryptococcus neoformans. The C neoformans isolate belonged to ST5 and molecular type VNI (var. grubii). INTERVENTIONS: The patient was diagnosed with cryptococcal pleuritis, then amphotericin B and fluconazole were administrated. OUTCOMES: Finally, the patient was improved and discharged from our hospital. LESSONS: Similar cases in cryptococcal pleuritis patients with pleural effusion as the only clinical presentation in the literature are also reviewed. Through literature review, we recommend that pleural effusion cryptococcal antigen test should be used to diagnose cryptococcal pleuritis to reduce misdiagnosis. The early administration of antifungal drug with activity to Cryptococcus seemed beneficial in preventing dissemination of cryptococcosis.

Human epididymis protein 4 (HE4) protects against cystic pulmonary fibrosis associated-inflammation through inhibition of NF-κB and MAPK singnaling.

BACKGROUND: Cystic pulmonary fibrosis (CF) affects mostly the lung of the newborns. Chronic infection and inflammation become the major causes of morbidity and mortality in CF. However, the underlying molecular mechanisms causing CF still remain unclear. METHODS: ELISA assay was used to examine the expression of HE4 and pro-inflammatory cytokines in W126VA4 cells supernatant fluid. qRT-PCR was applicable to determine the mRNA level of HE4, α-SMA, collagen 1, MMP2, MMP9 and various interleukins. Immunofluorescent assay was used to test the expression of HE4 in WI-26 VA4 cells. Major elements of MAPK and NF-κB signals pathways were examined by western blot. RESULTS: We found higher expression of HE4 in CF patients serum and lung biopsy. Interestingly, HE4 expression was positively correlated with fibrosis markers expression. In addition,HE4 overexpression increased inflammatory cytokines secretion and fibrosis markers expression in WI-26 VA4 cells. And NF-κB pathways were responsible for elevated inflammation. In addition, HE4/MAPK/MMPs signaling cascades destroyed the normal extracellular matrix (ECM) and promoted fibrosis. CONCLUSIONS: Overall, we first identified that HE4 promoted CF-associated inflammation. Additionally, NF-κB and MAPK signalings were further validated to be responsible for CF-associated inflammation and ECM destruction. Characterization of lumacaftor/ivacaftor in CF-associated inflammation may provide a novel insight into clinical CF treatment.

Inhibition of microRNA-34a Suppresses Epileptiform Discharges Through Regulating Notch Signaling and Apoptosis in Cultured Hippocampal Neurons.

Epilepsy is characterized by recurrent unprovoked seizures and some seizures can cause neuronal apoptosis, which is possible to make contributions to the epilepsy phenotype, impairments in cognitive function or even epileptogenesis. Moreover, many studies have indicated that microRNA-34a (miRNA-34a) is involved in apoptosis through regulating Notch signaling. However, whether miRNA-34a participates in neuronal apoptosis after seizures remain unclear. Therefore, we aimed to explore the expression of miRNA-34a and its effects on the epileptiform discharge in spontaneous recurrent epileptiform discharges (SREDs) rat hippocampal neuronal pattern. Mg2+-free medium was used to induce SREDs, quantitative reverse-transcription polymerase chain reaction was used to detect the expression of miRNA-34a, western blot was used to determine the expression of Notch pathway and apoptosis-related proteins, and whole cell current clamp recordings was used to observe the alteration of epileptiform discharge. We found obvious apoptosis, increased expression of miRNA-34a and decreased expression of Notch signaling in Mg2+-free-treated neurons. Treatment with miRNA-34a inhibitor decreased the frequency of action potentials, activated Notch signaling and prevented neuronal apoptosis in Mg2+-free-treated neurons. However, treatment with miRNA-34a mimics increased the frequency of action potentials, down-regulated Notch signaling and promoted neuronal apoptosis in Mg2+-free-treated neurons. Furthermore, γ-secretase inhibitor N-[N-(3,5-di-uorophenacetyl)-1-alanyl]-S-phenylglycine t-butylester (DAPT), an inhibitor of Notch signaling, could weaken anti-apoptosis effect of miRNA-34a inhibitor. These results suggest that inhibition of miRNA-34a could suppress epileptiform discharges through regulating Notch signaling and apoptosis in the rat hippocampal neuronal model of SREDs.