Sepsis is a subtype of systemic inflammatory response syndrome (SIRS), which is characterized by infection. Circulating microRNAs (miRNAs), including miR-150, miR-146a and miR-223, are potential biomarkers of sepsis. In this study, we demonstrated that measuring the relative expression of miR-146a/U6 in plasma, using the 2−ΔΔCt method, provides a method for differentiating between sepsis and non-sepsis-SIRS. We observed a significant increase in miR-146a expression in the initial cohort of 6 non-sepsis-SIRS patients compared to the 4 sepsis patients (P=0.01) and in the second cohort of 8 non-sepsis-SIRS patients compared to the 10 sepsis patients (P=0.027). Additionally, we identified that sodium citrate and ethylenediaminetetraacetic acid (EDTA) K2 may be used as anticoagulant reagents. Generation of a standard curve is not necessary in these diagnostic tests, unless the standard of normalization is carefully selected. Thus we provide more detailed guidance for the clinical use of circulating miRNA biomarkers.

iThe spread of the plasmid-mediated colistin resistance gene, mcr-1, into carbapenem-resistant Enterobacteriaceae (CRE) clinical isolates poses a significant threat to global health. Here we report the identification of three mcr-1-harboring carbapenem-resistant Escherichia coli strains, collected from three patients in two provinces in China. Our results show that mcr-1-harboring CRE strains have started to spread in different hospitals in China. In addition, this report presents the first description of chromosomal integration of mcr-1 into a carbapenem-resistant E. coli strain. Polymyxins B and E (also known as colistin) are among some antibiotics of the last resort used to treat serious infections caused by carbapenem-resistant Enterobacteriaceae (CRE). However, the recent discovery of a plasmid-mediated colistin resistance gene, mcr-1, sounded the alarm that last-resort antibiotics may be in jeopardy (1). Of particular concern is the spread of mcr-1 into CRE, creating extensively drug-resistant isolates causing untreatable disease. We have recently reported the cooccurrence of NDM-5 carbapenemase and MCR-1 within the same clinical isolate from a tertiary hospital in eastern China (2). The isolates coproducing MCR-1 and NDM-5 were nonsusceptible to nearly all antimicrobial agents tested (2). It is worrisome that these MCR-1-producing CRE isolates may spread further into hospital settings and within high-risk patients, thereby causing untreatable infections. Here we conducted a molecular screening study for clinical CRE isolates collected from six tertiary hospitals in six provinces in order to explore the dissemination of MCR-1-producing CRE in China.A total of 264 clinical CRE isolates were collected from six large regional hospitals in northern (Beijing), eastern (Suzhou), southern (Guangzhou), northwestern (Yinchuan), and southwestern (Chengdu and Kunming) China between January 2014 and December 2015. They were isolated from respiratory tract (n ϭ 119), urine (n ϭ 50), blood (n ϭ 38), intra-abdominal (n ϭ 22), skin and soft tissue (n ϭ 17), rectal swab (n ϭ 9), wound (n ϭ 5), and other sites (n ϭ 4) of 251 unique patients and included 160 Klebsiella pneumoniae, 36 Escherichia coli, 19 Enterobacter cloacae, 17 E. aerogenes, 11 K. oxytoca, 5 Citrobacter freundii, 4 Serratia, 3 Morganella morganii, 3 Providencia rettgeri, and 3 K. ozaenae spp. and 3 other species. Species identification was performed using matrix-assisted laser desorption ionization-time of flight mass spectrometry (Bruker Microflex LT). PCR detection of carbapenemase genes (bla KPC , bla NDM , bla VIM , bla OXA48-like , and bla IMP ) showed that 134 isolates were positive for bla KPC , 69 for bla NDM , 18 for bla IMP , and 7 for bla VIM , while 59 were negative for any carbapenemase gene. Thirty-one isolates were found to carry more than one carbapenemase. We then tested for the presence of mcr-1 using a previously published PCR method (1) and identified a total of five mcr-1-harboring CRE isolates from four different patients in three diff...

Gut microbiota plays a dual role in chronic kidney disease (CKD) and is closely linked to production of uremic toxins. Strategies of reducing uremic toxins by targeting gut microbiota are emerging. It is known that Chinese medicine rhubarb enema can reduce uremic toxins and improve renal function. However, it remains unknown which ingredient or mechanism mediates its effect. Here we utilized a rat CKD model of 5/6 nephrectomy to evaluate the effect of emodin, a main ingredient of rhubarb, on gut microbiota and uremic toxins in CKD. Emodin was administered via colonic irrigation at 5ml (1mg/day) for four weeks. We found that emodin via colonic irrigation (ECI) altered levels of two important uremic toxins, urea and indoxyl sulfate (IS), and changed gut microbiota in rats with CKD. ECI remarkably reduced urea and IS and improved renal function. Pyrosequencing and Real-Time qPCR analyses revealed that ECI resumed the microbial balance from an abnormal status in CKD. We also demonstrated that ten genera were positively correlated with Urea while four genera exhibited the negative correlation. Moreover, three genera were positively correlated with IS. Therefore, emodin altered the gut microbiota structure. It reduced the number of harmful bacteria, such as Clostridium spp. that is positively correlated with both urea and IS, but augmented the number of beneficial bacteria, including Lactobacillus spp. that is negatively correlated with urea. Thus, changes in gut microbiota induced by emodin via colonic irrigation are closely associated with reduction in uremic toxins and mitigation of renal injury.

Enterobacter cloacae has recently emerged as one of the most common carbapenem-resistant Enterobacteriaceae . The emergence and spread of metallo-β-lactamase-producing E. cloacae have posed an immediate threat globally. Here, we investigated the molecular characteristics of 84 carbapenem-resistant Enterobacter cloacae (CREL) collected from three tertiary hospitals in China between 2012 and 2016. Species identification and antimicrobial susceptibility testing were performed using a VITEK-2 system. Carbapenems, polymyxins B, and tigecycline were tested by broth microdilution method. The carbapenem in activation method (CIM) and cefoxitin three-dimensional test were used to detect carbapenemase and AmpC β-lactamase, respectively. Isolates were screened for β-lactam resistance genes by PCR, and expression of ompC and ompF was determined by qRT-PCR. Genetic relatedness was performed by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST), while selected isolates were subjected to whole-genome sequencing. Among the 84 CREL isolates, 50 (59.5%) were detected as carbapenemase producers. NDM-1 was the dominant carbapenemase (80.0%), followed by IMP-26 (8.0%) and IMP-4 (6.0%). Notably, we identified the first NDM-1 and IMP-1 co-producing E. cloacae , carrying plasmids of several incompatibility (Inc) groups, including IncHI2, IncHI2A, and IncN. Most isolates showed decreased expression of ompC and/or ompF , and contained a broad distribution of ESBLs and AmpC β-lactamases. These findings suggested that different molecular mechanisms, including carbapenemase, ESBL and/or AmpC plus loss of porins, have contributed to carbapenem resistance. The bla NDM−1 -harboring plasmids contained highly conserved gene environment around bla NDM−1 ( bla NDM−1 - ble MBL - trpF - dsbD - cutA1 - groES - groEL ), which could be associated with the potential dissemination of bla NDM−1 . IMP-type MBL was located within a variety of integrons and usually contained various gene cassettes encoding multidrug resistance. These isolates produced 54 different pulsotypes, and were classified into 42 STs by MLST. Nineteen bla NDM−1 -positive E. cloacae isolates obtained from Ningxia had the same pulsotype (PFGE type 1), belonging to ST78 within clonal complex 74 (CC74). The plasmid-based replicon typing indicated that IncX3 plasmids mediated the dissemination of bla NDM−1 among these homologous strains. This is the first report on the outbreak of N...

Conjugation is a key mechanism for horizontal gene transfer and plays an important role in bacterial evolution, especially with respect to antibiotic resistance. However, little is known about the role of donor and recipient cells in regulation of conjugation. Here, using an Escherichia coli (SM10λπ)-Pseudomonas aeruginosa (PAO1) conjugation model, we demonstrated that deficiency of lasI/rhlI, genes associated with generation of the quorum sensing signals N-acyl homoserine lactones (AHLs) in PAO1, or deletion of the AHLs receptor SdiA in the donor SM10λπ both facilitated conjugation. When using another AHLs-non-producing E. coli strain EC600 as recipient cells, deficiency of sdiA in donor SM10λπ hardly affect the conjugation. More importantly, in the presence of exogenous AHLs, the conjugation efficiency between SM10λπ and EC600 was dramatically decreased, while deficiency of sdiA in SM10λπ attenuated AHLs-inhibited conjugation. These data suggest the conjugation suppression function of AHLs-SdiA chemical signaling. Further bioinformatics analysis, β-galactosidase reporter system and electrophoretic mobility shift assays characterized the binding site of SdiA on the promoter region of traI gene. Furthermore, deletion of lasI/rhlI or sdiA promoted traI mRNA expression in SM10λπ and PAO1 co-culture system, which was abrogated by AHLs. Collectively, our results provide new insight into an important contribution of quorum sensing system AHLs-SdiA to the networks that regulate conjugation.