Tumor-derived Prevotella intermedia aggravates gastric cancer by enhancing Perilipin 3 expression.

The indigenous microbial milieu within tumorous tissues exerts a pivotal influence on the genesis and advancement of gastric cancer (GC). This investigation scrutinizes the functions and molecular mechanisms attributed to Prevotella intermedia in the malignant evolution of GC. Isolation of P. intermedia from paired GC tissues was undertaken. Quantification of P. intermedia abundance in 102 tissues was accomplished using quantitative real-time PCR (qRT-PCR). Assessment of the biological effects of P. intermedia on GC cells was observed using culture medium supernatant. Furthermore, the protein profile of GC cells treated with tumor-derived P. intermedia was examined through label-free protein analysis. The functionality of perilipin 3 (PLIN3) was subsequently confirmed using shRNA. Our investigation revealed that the relative abundance of P. intermedia in tumor tissues significantly surpassed that of corresponding healthy tissues. The abundance of P. intermedia exhibited correlations with tumor differentiation (p = 0.006), perineural invasion (p = 0.004), omentum majus invasion (p = 0.040), and the survival duration of GC patients (p = 0.042). The supernatant derived from tumor-associated P. intermedia bolstered the proliferation, clone formation, migration, and invasion of GC cells. After indirect co-cultivation with tumor-derived P. intermedia, dysregulation of 34 proteins, including PLIN3, was discerned in GC cells. Knockdown of PLIN3 mitigated the malignancy instigated by P. intermedia in GC cells. Our findings posit that P. intermedia from the tumor microenvironment plays a substantial role in the malignant progression of GC via the modulation of PLIN3 expression. Moreover, the relative abundance of P. intermedia might serve as a potential biomarker for the diagnosis and prognosis of GC.

Identification of TMexCD-TOprJ-producing carbapenem-resistant Gram-negative bacteria from hospital sewage.

Carbapenems and tigecycline are crucial antimicrobials for the treatment of gram-negative bacteria infections. Recently, a novel resistance-nodulation-division (RND) efflux pump gene cluster, tmexCD-toprJ, which confers resistance to tigecycline, has been discovered in animals and clinical isolates. It was reported that hospital sewage could act as a reservoir for gram-negative bacteria with high antimicrobial resistance genes. In this study, we analyzed 84 isolates of carbapenem-resistant gram-negative bacteria (CR-GNB) from hospital sewage, and identified five isolates of TMexCD-ToprJ-producing CR-GNB, including one Raoultella ornithinolytica isolate and four Pseudomonas spp. isolates. All these five isolates carried at least one carbapenem resistance gene and were resistant to multiple antibiotics. Multiple tmexCD-toprJ clusters were detected, including tmexC2D2-toprJ2, tmexC3D3-toprJ3, tmexC3.2D3.3-toprJ1b and tmexC3.2D3-toprJ1b. Among these clusters, the genetic construct of tmexC3.2D3-toprJ1b showed 2-fold higher minimum inhibitory concentration (MIC) of tigecycline than other three variants. In addition, it was found that the tmexCD-toprJ gene cluster was originated from Pseudomonas spp. and mainly located on Tn6855 variants inserted in the same umuC-like genes on chromosomes and plasmids. This unit co-localized with blaIMP or blaVIM on IncHI5-, IncpJBCL41- and IncpSTY-type plasmids in the five isolates of TMCR-GNB. The IncHI5- and IncpSTY-type plasmids had the ability to conjugal transfer to E. coli J53 and P. aeruginosa PAO1, highlighting the potential risk of transfer of tmexCD-toprJ from Pseudomonas spp. to Enterobacterales. Importantly, genomic analysis showed that similar tmexCD-toprJ-harboring IncHI5 plasmids were also detected in human samples, suggesting transmission between environmental and human sectors. The emergence of TMCR-GNB from hospital sewage underscores the need for ongoing surveillance of antimicrobial resistance genes, particularly the novel resistance genes such as the tmexCD-toprJ gene clusters in the wastewater environment.

Roles of host SUMOylation in bacterial pathogenesis.

Bacteria frequently interfere with the post-translational modifications of host cells to facilitate their survival and growth after invasion. SUMOylation, a reversible post-translational modification process, plays an important role in biological life activities. In addition to being critical to host cell metabolism and survival, SUMOylation also regulates gene expression and cell signal transmission. Moreover, SUMOylation in eukaryotic cells can be used by a variety of bacterial pathogens to advance bacterial invasion. In this minireview, we focused on the role and mechanism of host SUMOylation in the pathogenesis of six important clinical bacterial pathogens (Listeria monocytogenes, Shigella flexneri, Salmonella Typhimurium, Klebsiella pneumoniae, Staphylococcus aureus, and Escherichia coli). Taken together, this review provided new insights for understanding the unique pathogen-host interaction based on host SUMOylation and provided a novel perspective on the development of new strategies to combat bacterial infections in the future.

Global Transcriptomic Study of Circular-RNA Expression Profile in Osteoclasts Infected by Intracellular Staphylococcus aureus.

Osteomyelitis is difficult to cure, and the rapidly rising morbidity is a thorny problem accompanied by a large number of joint replacement applications. Staphylococcus aureus is the main pathogen of osteomyelitis. Circular RNAs (circRNAs), as emerging noncoding RNAs, play important roles in multiple physiopathological processes which could provide novel insights into osteomyelitis. However, little is known about the roles of circRNAs in the pathogenesis of osteomyelitis. Osteoclasts, considered bone sentinels, are the resident macrophages in bone and may play the immune defense roles in osteomyelitis. It has been reported that S. aureus can survive in osteoclasts, but the function of osteoclast circRNAs in response to intracellular S. aureus infection remains unclear. In this study, we investigated the profile of circRNAs in osteoclasts infected by intracellular S. aureus through high-throughput RNA sequencing. In total, 24 upregulated and 62 downregulated differentially expressed circRNAs were identified and subsequently analyzed to demonstrate their potential functions. On this basis, three circRNAs (chr4:130718154-130728164+, chr8:77409548-77413627-, and chr1:190871592-190899571-) were confirmed as potential novel biomarkers for the diagnosis of osteomyelitis through the murine model of osteomyelitis. Most importantly, we verified that the circRNA chr4:130718154-130728164+ named circPum1 could regulate the host autophagy to affect the intracellular infection of S. aureus through miR-767. In addition, circPum1 could serve as a promising serum biomarker in osteomyelitis patients caused by S. aureus infection. Taken together, this study provided the first global transcriptomic profile analysis of circRNAs in osteoclasts infected by intracellular S. aureus and first proposed a novel perspective for the pathogenesis and immunotherapy of S. aureus-induced osteomyelitis from the term of circRNAs.

Mechanisms of folate metabolism-related substances affecting Staphylococcus aureus infection.

Staphylococcus aureus (S. aureus) is one of the critical clinical pathogens which can cause multiple diseases ranging from skin infections to fatal sepsis. S. aureus is generally considered to be an extracellular pathogen. However, more and more evidence has shown that S. aureus can survive inside various cells. Folate plays an essential role in multiple life activities, including the conversion of serine and glycine, the remethylation of homocysteine to methionine, and the de novo synthesis of purine /dTMP, et al. More and more studies reported that S. aureus intracellular infection requires the involvement of folate metabolism. This review focused on the mechanisms of folate metabolism and related substances affecting S. aureus infection. Loss of tetrahydrofolic acid (THF)-dependent dTMP directly inhibits the nucleotide synthesis pathway of the S. aureus due to pabA deficiency. Besides, trimethoprim-sulfamethoxazole (TMP/SMX), a potent antibiotic that treats S. aureus infections, interferes in the process of the folate mechanism and leads to the production of thymidine-dependent small-colony variants (TD-SCVs). In addition, S. aureus is resistant to lysostaphin in the presence of serine hydroxymethyltransferase (SHMT). We provide new insights for understanding the molecular pathogenesis of S. aureus infection.

Molecular basis of Klebsiella pneumoniae colonization in host.

Klebsiella pneumoniae (K. pneumoniae) is a common cause of nosocomial infection, which causing disseminated infections such as cystitis, pneumonia and sepsis. K. pneumoniae is intrinsic resistant to penicillin, and members of the population usually have acquired resistance to a variety of antibiotics, which makes it a major threat to clinical and public health. Bacteria can colonize on or within the hosts, accompanied by growth and reproduction of the organisms, but no clinical symptoms are presented. As the "first step" of bacterial infection, colonization in the hosts is of great importance. Colonization of bacteria can last from days to years, with resolution influenced by immune response to the organism, competition at the site from other organisms and, sometimes, use of antimicrobials. Colonized pathogenic bacteria cause healthcare-associated infections at times of reduced host immunity, which is an important cause of clinical occurrence of postoperative complications and increased mortality in ICU patients. Though, K. pneumoniae is one of the most common conditional pathogens of hospital-acquired infections, the mechanisms of K. pneumoniae colonization in humans are not completely clear. In this review, we made a brief summary of the molecular basis of K. pneumoniae colonization in the upper respiratory tract and intestinal niche, and provided new insights for understanding the pathogenesis of K. pneumoniae.

Mechanisms of promoting the differentiation and bone resorption function of osteoclasts by Staphylococcus aureus infection.

Bone infection is a common and serious complication in the field of orthopedics, which frequently leads to excessive bone destruction and fracture nonunion. Staphylococcus aureus (S. aureus) infection affects bone cell function which, in turn, causes bone destruction. Bone is mainly regulated by osteoblasts and osteoclasts. Osteoclasts are the only cell type with bone resorptive function. Their over-activation is closely associated with excessive bone loss. Understanding how S. aureus changes the functional state of osteoclasts is the key to effective treatment. By reviewing the literature, this paper summarizes several mechanisms of bone destruction caused by S. aureus influencing osteoclasts, thereby stimulating new ideas for the treatment of bone infection.

Profile analysis of circRNAs in human THP-1 derived macrophages infected with intracellular Staphylococcus aureus.

BACKGROUND: Intracellular Staphylococcus aureus (S. aureus) infection is generally persistent, recurrent and difficult to treat due to the poor availability of antibiotics within macrophages cells and the lack of ideal diagnostic markers. Circular RNAs (circRNAs), with covalently closed circular structures, exists in the serum stably and is not easily degraded by nucleases. Besides, circRNAs play a pivotal in the eukaryotic regulation of genes expression and served as biomarkers in variety disease including microbial infections. However, the function of host circRNAs in intracellular S. aureus infection remains largely unclear. METHODS: In this study, the circRNAs expression profile was investigated by RNA sequencing technology in both S. aureus-infected THP-1 derived macrophages and mock control cells. The differentially expressed circRNAs (DE circRNAs) with a fold-change >1.5 (p < 0.05) are analyzed using functional pathway clustering prediction. Then, RT-qPCR was performed to verify the top 2 up-regulated circRNAs in the THP-1 cell and human serum samples so as to evaluate the value of circRNAs for S. aureus diagnosis. RESULTS: An intracellular survival THP-1 derived macrophages model of S. aureus infection was established. A total of 5,299 circRNAs were identified in human THP-1 derived macrophages infected with intracellular S. aureus. There were 61 DE circRNAs with a fold-change >1.5 (p < 0.05) after S. aureus infection. Among them, 22 circRNAs were up-regulated while 39 circRNAs down-regulated. GO and KEGG pathway analysis demonstrated that DE circRNAs were enriched in the processes such as Neurotrophin, Pyruvate metabolism and Notch signaling pathway. Moreover, hsa_circ_0000311 and chr13:43500472-43544806-(novel) were verified to be significantly upregulated in THP-1 derived macrophages and human serum samples between two groups. Finally, the networks of circRNA-miRNA-mRNA based on these two circRNAs were constructed respectively. CONCLUSION: Our study provides the first profile analysis of host circRNAs involved in intracellular S. aureus infection, which may serve as biomarkers for S. aureus diagnosis and contribute to the understanding of S. aureus evasion mechanisms.

Identification of key serum biomarkers for the diagnosis and metastatic prediction of osteosarcoma by analysis of immune cell infiltration.

BACKGROUND: The role of circular RNAs (circRNAs) and microRNAs (miRNAs) in osteosarcoma (OS) development has not been fully elucidated. Further, the contribution of the immune response to OS progression is not well defined. However, it is known that circRNAs and miRNAs can serve as biomarkers for the diagnosis, prognosis, and therapy of many cancers. Thus, the aim of this study was to identify novel key serum biomarkers for the diagnosis and metastatic prediction of OS by analysis of immune cell infiltration and associated RNA molecules. METHODS: Human OS differentially expressed circRNAs (DEcircRNAs), differentially expressed miRNAs (DEmiRNAs), and differentially expressed mRNAs (DEmRNAs) were identified by analysis of microarray data downloaded from Gene Expression Omnibus (GEO) datasets. Further, characteristic patterns of OS-infiltrating immune cells were analyzed. On this basis, we identified statistically significant transcription factors. Moreover we performed pathway enrichment analysis, constructed protein-protein interaction networks, and devised competitive endogenous RNA (ceRNA) networks. Biological targets of the ceRNA networks were evaluated and potential OS biomarkers confirmed by RT-qPCR analysis of the patients' serum. RESULTS: Seven differentially expressed circRNAs, 166 differentially expressed miRNAs, and 175 differentially expressed mRNAs were identified. An evaluation of cellular OS infiltration identified the highest level of infiltration by M0 macrophages, M2 macrophages, and CD8+ T cells, with M0 macrophages and CD8+ T cells as the most prominent. Significant patterns of tumor-infiltrating immune cells were identified by principal component analysis. Moreover, 185 statistically significant transcription factors were associated with OS. Further, in association with immune cell infiltration, hsa-circ-0010220, hsa-miR-326, hsa-miR-338-3p, and FAM98A were identified as potential novel biomarkers for OS diagnosis. Of these, FAM98A had the most promise as a diagnostic marker for OS and OS metastasis. Most importantly, a novel diagnostic model consisting of these four biomarkers (hsa-circ-0010220, hsa-miR-326, hsa-miR-338-3p, and FAM98A) was established with a 0.928 AUC value. CONCLUSIONS: In summary, potential serum biomarkers for OS diagnosis and metastatic prediction were identified based on an analysis of immune cell infiltration. A novel diagnostic model consisting of these four promising serum biomarkers was established. Taken together, the results of this study provide a new perspective by which to understand immunotherapy of OS.

Resveratrol Alleviates Ischemic Brain Injury by Inhibiting the Activation of Pro-Inflammatory Microglia Via the CD147/MMP-9 Pathway.

OBJECTIVE: Ischemic stroke is one of the most common diseases with high mortality and disability. This study was intended to investigate the mechanism of resveratrol (RES) regulating microglia activation through the CD147/matrix metalloproteinase-9 (MMP-9) pathway on ischemic stroke. METHODS: The middle cerebral artery occlusion (MCAO) mouse model and oxygen and glucose deprivation (OGD) cell model were established. The behavioral defects, neuronal damage, cerebral infarction volume, and histopathological changes were assessed in MCAO mice. The activation of pro-inflammatory microglia CD86+/Iba-1+ and anti-inflammatory microglia CD206+/Iba-1+ was detected. The expressions of pro-inflammatory microglia markers (CD11b, CD16) and cytokines (TNF-α, IL-1ß, and IL-6) were measured. The activation of the CD147/MMP-9 pathway was detected and its effect on microglia activation was assessed. RESULTS: After RES administration, the neuronal dysfunction, infarct volume, and morphological changes of neurons were improved in MCAO mice. Meanwhile, the motivation of pro-inflammatory microglia and the release of inflammatory factors were repressed. RES suppressed the stimulation of OGD/R microglia and the release of inflammatory factors. The expression of CD147 and MMP-9 in primary microglia was up-regulated. Inhibition of CD147 can reduce pro-inflammatory microglia activation by inhibiting MMP-9 expression. RES inhibited the CD147/MMP-9 axis in OGD/R microglia, and overexpression of CD147 partially reversed the inhibitory effect of RES on the activation and release of inflammatory factors in OGD/R microglia. CONCLUSION: RES restrained the stimulation of pro-inflammatory microglia by down-regulating the CD147/MMP-9 axis, and thus protected against ischemic brain injury.

Apramycin resistance in epidemic carbapenem-resistant Klebsiella pneumoniae ST258 strains.

BACKGROUND: Recent studies indicated that the monosubstituted deoxystreptamine aminoglycoside apramycin is a potent antibiotic against a wide range of MDR Gram-negative pathogens. OBJECTIVES: To evaluate the in vitro activity of apramycin against carbapenem-resistant Klebsiella pneumoniae (CRKp) isolates from New York and New Jersey, and to explore mechanisms of apramycin resistance. METHODS: Apramycin MICs were determined by broth microdilution for 155 CRKp bloodstream isolates collected from 2013 to 2018. MLST STs, wzi capsular types and apramycin resistance gene aac(3')-IV were examined by PCR and Sanger sequencing. Selected isolates were further characterized by conjugation experiments and WGS. RESULTS: Apramycin MIC50/90 values were 8 and >128 mg/L for CRKp isolates, which are much higher than previously reported. Twenty-four isolates (15.5%) were apramycin resistant (MIC ≥64 mg/L) and they were all from the K. pneumoniae ST258 background. The 24 apramycin-resistant K. pneumoniae ST258 strains belonged to six different capsular types and 91.7% of them harboured the apramycin resistance gene aac(3')-IV. Sequencing analysis showed that different ST258 capsular type strains shared a common non-conjugative IncR plasmid, co-harbouring aac(3')-IV and blaKPC. A novel IncR and IncX3 cointegrate plasmid, p59494-RX116.1, was also identified in an ST258 strain, demonstrating how apramycin resistance can be spread from a non-conjugative plasmid through cointegration. CONCLUSIONS: We described a high apramycin resistance rate in clinical CRKp isolates in the New York/New Jersey region, mainly among the epidemic K. pneumoniae ST258 strains. The high resistance rate in an epidemic K. pneumoniae clone raises concern regarding the further optimization and development of apramycin and apramycin-like antibiotics.

Construction of an autophagy interaction network based on competitive endogenous RNA reveals the key pathways and central genes of SARS-CoV-2 infection in vivo.

As of April 1, 2021, more than 2.8 million people have died of SARS-CoV-2 infection. In addition, the mutation of virus strains that have accompanied the pandemic has brought more severe challenges to pandemic control. Host microRNAs (miRNAs) are widely involved in a variety of biological processes of coronavirus infection, including autophagy in SARS-CoV-2 infection. However, the mechanisms underlying miRNAs involved in autophagy in SARS-CoV-2 infection have not been fully elucidated. In this study, the miRNA and messenger RNA (mRNA) expression profiles of patients with SARS-CoV-2 infection were investigated based on raw data from Gene Expression Omnibus (GEO) datasets, and potential novel biomarkers of autophagy were revealed by bioinformatics analyses. We identified 32 differentially expressed miRNAs and 332 differentially expressed mRNAs in patients with SARS-CoV-2 infection. Cytokine receptor related pathways were the most enriched pathways for differentially expressed miRNAs identified by pathway analysis. Most importantly, an autophagy interaction network, which was associated with the pathological processes of SARS-CoV-2 infection, especially with the cytokine storm, was constructed. In this network, hsa-miR-340-3p, hsa-miR-652-3p, hsa-miR-4772-5p, hsa-miR-192-5p, TP53INP2, and CCR2 may be biomarkers that predict changes in mild SARS-CoV-2 infection. Some molecules, including hsa-miR-1291 and CXCR4, were considered potential targets to predict the emergence of severe symptoms in SARS-CoV-2 infection. To our knowledge, this study provided the first profile analysis of an autophagy interaction network in SARS-CoV-2 infection and revealed several novel autophagy-related biomarkers for understanding the pathogenesis of SARS-CoV-2 infection in vivo.

Identification of the miRNA signature and key genes in colorectal cancer lymph node metastasis.

BACKGROUND: Because its metastasis to the lymph nodes are closely related to poor prognosis, miRNAs and mRNAs can serve as biomarkers for the diagnosis, prognosis, and therapy of colorectal cancer (CRC). This study aimed to identify novel gene signatures in the lymph node metastasis of CRC. METHODS: GSE56350, GSE70574, and GSE95109 datasets were downloaded from the Gene Expression Omnibus (GEO) database, while data from 569 colorectal cancer cases were also downloaded from The Cancer Genome Atlas (TCGA) database. Differentially expressed miRNAs (DE-miRNAs) were calculated using R programming language (Version 3.6.3), while gene ontology and enrichment analysis of target mRNAs were performed using FunRich ( http://www.funrich.org ). Furthermore, the mRNA-miRNA network was constructed using Cytoscape software (Version 3.8.0). Gene expression levels were verified using the GEO datasets. Similarly, quantitative real-time PCR (qPCR) was used to examine expression profiles from 20 paired non-metastatic and metastatic lymph node tissue samples obtained from patients with CRC. RESULTS: In total, five DE-miRNAs were selected, and 34 mRNAs were identified after filtering the results. Moreover, two key miRNAs (hsa-miR-99a, hsa-miR-100) and one gene (heparan sulfate-glucosamine 3-sulfotransferase 2 [HS3ST2]) were identified. The GEO datasets analysis and qPCR results showed that the expression of key miRNA and genes were consistent with that obtained from the bioinformatic analysis. A novel miRNA-mRNA network capable of predicting the prognosis and confirmed experimentally, hsa-miR-99a-HS3ST2-hsa-miR-100, was found after expression analysis in metastasized lymph node tissue from CRC samples. CONCLUSION: In summary, miRNAs and genes with potential as biomarkers were found and a novel miRNA-mRNA network was established for CRC lymph node metastasis by systematic bioinformatic analysis and experimental validation. This network may be used as a potential biomarker in the development of lymph node metastatic CRC.

CRISPR-Cas9-Mediated Carbapenemase Gene and Plasmid Curing in Carbapenem-Resistant Enterobacteriaceae.

Combating plasmid-mediated carbapenem resistance is essential to control and prevent the dissemination of carbapenem-resistant Enterobacteriaceae (CRE). Here, we conducted a proof-of-concept study to demonstrate that CRISPR-Cas9-mediated resistance gene and plasmid curing can effectively resensitize CRE to carbapenems. A novel CRISPR-Cas9-mediated plasmid-curing system (pCasCure) was developed and electrotransferred into various clinical CRE isolates. The results showed that pCasCure can effectively cure blaKPC, blaNDM, and blaOXA-48 in various Enterobacteriaceae species of Klebsiella pneumoniae, Escherichia coli, Enterobacter hormaechei, Enterobacter xiangfangensis, and Serratia marcescens clinical isolates, with a >94% curing efficiency. In addition, we also demonstrated that pCasCure can efficiently eliminate several epidemic carbapenem-resistant plasmids, including the blaKPC-harboring IncFIIK-pKpQIL and IncN pKp58_N plasmids, the blaOXA-48-harboring pOXA-48-like plasmid, and the blaNDM-harboring IncX3 plasmid, by targeting their replication and partitioning (parA in pKpQIL) genes. However, curing the blaOXA-48 gene failed to eliminate its corresponding pOXA-48-like plasmid in clinical K. pneumoniae isolate 49210, while further next-generation sequencing revealed that it was due to IS1R-mediated recombination outside the CRISPR-Cas9 cleavage site resulting in blaOXA-48 truncation and, therefore, escaped plasmid curing. Nevertheless, the curing of carbapenemase genes or plasmids, including the truncation of blaOXA-48 in 49210, successfully restore their susceptibility to carbapenems, with a >8-fold reduction of MIC values in all tested isolates. Taken together, our study confirmed the concept of using CRISPR-Cas9-mediated carbapenemase gene and plasmid curing to resensitize CRE to carbapenems. Further work is needed to integrate pCasCure in an optimal delivery system to make it applicable for clinical intervention.

In vitro selection of aztreonam/avibactam resistance in dual-carbapenemase-producing Klebsiella pneumoniae.

OBJECTIVES: To examine the in vitro selection of aztreonam/avibactam resistance among MBL-producing Klebsiella pneumoniae and to understand the mechanism of increased resistance. METHODS: The MICs of aztreonam were determined with and without avibactam (4 mg/L) using a broth microdilution method. Single-step and multi-step mutant selection was conducted on five MBL-producing K. pneumoniae strains, including two dual carbapenemase producers. Genomic sequencing and gene cloning were performed to investigate the mechanism of increased resistance. RESULTS: We examined the MICs for 68 MBL-producing K. pneumoniae isolates, including 13 dual carbapenemase producers. Compared with aztreonam alone, the addition of avibactam (4 mg/L) reduced the MICs for all isolates by >128-fold, with MIC50 and MIC90 values of 0.25 and 1 mg/L, respectively. One NDM-1-, OXA-48-, CTX-M-15- and CMY-16-positive ST101 K. pneumoniae strain was selected to be resistant to aztreonam/avibactam, with a >16-fold increase in MIC (>128 mg/L). WGS revealed that the resistant mutants lost the blaNDM-1 gene, but acquired amino acid substitutions in CMY-16 (Tyr150Ser and Asn346His). Construction of blaCMY-16 mutants confirmed that the substitutions (Tyr150Ser and Asn346His) were primarily responsible for the decreased susceptibility to aztreonam/avibactam. In addition, transfer of blaCMY-16 mutant (Tyr150Ser and Asn346His) plasmid constructs into certain clinical carbapenemase-producing isolates demonstrated >64-fold increased MICs of aztreonam/avibactam and aztreonam/avibactam/ceftazidime. CONCLUSIONS: Aztreonam in combination with avibactam showed potent in vitro activity against MBL-producing K. pneumoniae. However, our study suggested the likelihood of aztreonam/avibactam resistance among MBL- and AmpC-co-producing strains and clinical practice should beware of the possibility of the emerging resistance.

The tst gene associated Staphylococcus aureus pathogenicity island facilitates its pathogenesis by promoting the secretion of inflammatory cytokines and inducing immune suppression.

Staphylococcus aureus (S. aureus) is an important pathogen causing various limited or systemic infections. Methicillin resistant S. aureus (MRSA) in particular presents a major clinical and public health problem. Toxic shock syndrome toxin-1 (TSST-1) encoded by the gene tst is an important virulence factor of tst positive S. aureus, leading to multi-organ malfunction. However, the mechanism of TSST-1 in pathogenesis is only partly clear. In this study, we investigated the prevalence of the tst gene in clinical isolates of S. aureus. Then, animal experiments were performed to further evaluate the influence of the presence of the tst gene associated Staphylococcus aureus Pathogenicity Island (SaPI) on body weight, serum cytokine concentrations and the bacterial load in different organs. In addition, macrophages were used to analyze the secretion of cytokines in vitro and bacterial survival in the cytoplasm. Finally, pathological analysis was carried out to evaluate organ tissue impairment. The results demonstrated that the prevalence of tst gene was approximately 17.8% of the bacterial strains examined. BALB/c mice infected with tst gene associated SaPI positive isolates exhibited a severe loss of body weight and a high bacterial load in the liver, heart, kidney and spleen. Pathological analysis demonstrated that tissue impairment was more severe after infection with tst gene associated SaPI positive isolates. Moreover, the secretion of IL-6, IL-2 and IL17A by macrophages infected with tst gene associated SaPI positive isolates clearly increased. Notably, IL-6 secretion in BALB/c mice infected with tst gene associated SaPI positive isolates was higher than that in BALB/c mice infected with negative ones. Together, these results indicated that the tst gene associated SaPI may play a critical role in the pathological process of infection via a direct and persistent toxic function, and by promoting the secretion of inflammatory cytokines that indirectly induce immune suppression.

Effect of rpoE on the Non-coding RNA Expression Profiles of Salmonella enterica serovar Typhi under the Stress of Ampicillin.

Gene transcription in bacteria is mainly triggered by sigma factors (σ factors), such as rpoE. Bacterial ncRNAs are key players in reprogramming protein transcription when the environment changes. In our experiment, under the stress of ampicillin, the ncRNA transcriptomes of Salmonella enterica serovar Typhi wild-type strain (WT) and rpoE-deficient strain (ΔrpoE) were sequenced and analyzed, and four ncRNAs were selected to be verified by qRT-PCR. Of the ncRNA transcripts tested, 57 ncRNAs were found to have significantly different expressions (fold changes > 2) in ΔrpoE compared to WT, with 31 being upregulated and 26 being downregulated. The expression levels of the four ncRNAs verified preliminarily by subsequent qRT-PCR showed consistency with the sequencing data. Our study revealed the differences in ncRNA expression profiles between Salmonella enterica serovar Typhi WT and ΔrpoE under ampicillin stress. The four ncRNAs identified by qRT-PCR and their associated signaling pathways may be related to the envelope stress and antibiotic susceptibility of Salmonella enterica serovar Typhi.

New Delhi Metallo-ß-Lactamase 5-Producing Klebsiella pneumoniae Sequence Type 258, Southwest China, 2017.

We isolated a New Delhi metallo-ß-lactamase 5 (NDM-5)-producing Klebsiella pneumoniae sequence type (ST) 258 strain in southwest China during 2017. The blaNDM-5 gene was acquired by horizontal plasmid transfer from NDM-5-producing Escherichia coli. We identified genomic characteristics in ST258 strains that differed from those of global K. pneumoniae carbapenemase-producing strains.

An experimental study on the treatment of diabetes-induced cognitive disorder mice model with exosomes deriving from mesenchymal stem cells (MSCs).

The occurrence rate of senile dementia among diabetes patients is high, but there is still no effective therapeutic method. This study aimed to explore curative effect of exosomes deriving from MSCs treating diabetes-induced cognition impairment. After BM-MSCs culture was purified, Western Blot and electron microscope were used to identify exosomes which were injected into diabetes mouse through intracranial injection. Purified exosomes in Western blot analysis and electron microscope observation were verified. After therapy with exosomes deriving BM-MSC, mouse cognition impairment and histologic abnormity were recovered. Exosomes deriving from BM-MSCs can be a new preparation treating cognition with application prospect.

Triptolide inhibits the proliferation and migration of medulloblastoma Daoy cells by upregulation of microRNA-138.

Medulloblastoma is a primitive neuroectodermal-derived brain tumor and the most common malignant brain tumor in children. Triptolide (TPL) is the major active component extracted from Tripterygium wilfordii Hook F. This study aimed to explore the effects of TPL on medulloblastoma cell proliferation, migration, and apoptosis, as well as the underlying possible molecular mechanism. Viability, proliferation, and apoptosis of Daoy cells were measured using cell counting kit-8 assay, 5-bromo-2'-deoxyuridine incorporation assay, and Guava Nexin assay, respectively. Cell migration was detected using two-chamber transwell assay and wound healing assay. Quantitative reverse transcription polymerase chain reaction (qRT-PCR) was performed to determine the relative expression of microRNA-138 (miR-138) in Daoy cells. Cell transfection was used to change the expression of miR-138 in cells. Western blot analysis was used to analyze the expression of key factors involved in cell apoptosis, cell migration, the phosphatidylinositol 3-kinase (PI3K)/protein kinase 3 (AKT) pathway, and the Notch pathway in Daoy cells. We found that TPL significantly inhibited the viability, proliferation, and migration of Daoy cells but promoted Daoy cell apoptosis. The expression levels of matrix metalloproteinases (MMP)-2 and MMP-9 after TPL treatment were decreased. The expression of miR-138 in Daoy cells after TPL treatment was increased. Suppression of miR-138 obviously reversed the TPL-induced Daoy cell proliferation, migration inhibition, and cell apoptosis enhancement, as well as the inactivation of the PI3K/AKT and Notch pathways. Cyclin-dependent kinase 6 (CDK6) was a direct target gene of miR-138, which might be involved in the antitumor effects of TPL on Daoy cells. In conclusion, our study verified that TPL exerted anticancer effects on medulloblastoma cells possibly via upregulating miR-138 and inactivating the PI3K/AKT and Notch pathways.