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Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG) are the main pathogenic microorganisms causing sexually transmitted infections. In this study, a multiplex thermostable recombinase polymerase amplification-lateral flow detection (RPA-LFD) assay was established, and the reaction conditions such as the ratio of primer concentration, magnesium ion concentration, amplification time and template DNA concentration in the multiplex RPA reaction were optimized. The optimized multiplex RPA-LFD method was used to detect both CT and NG positive control plasmids, and it was found that the LFD could be used to obtain visible results when the plasmid copy number was only 200. The sensitivity of the multiplex RPA-LFD method used for clinical samples was 85.62 (95% CI at 53.66–97.29) for NG detection and 90.90 (95% CI at 57.12–99.52) for CT detection.

BackgroundCell released microvesicles specifically, exosomes, play an important role in mediating immunologic escape, treatment resistance, and disease persistence of Hepatitis C virus (HCV) infection. Reports on the molecular compositions of exosomes released by cells under diverse conditions, especially during viral infections, suggest that their cargo contents are not randomly loaded. However, the precise molecular mechanisms directing the selective cargo sorting and loading inside infectious viral exosomes remains elusive.AimTo decipher the role of Reticulon 3 (RTN3) in the selective molecular cargo sorting and loading inside infectious viral exosomes during HCV infection.MethodsWe used Huh7 cells-JFH1 HCV infection and HCV Full-Length (FL) replicon systems. Additionally, we analyzed human liver and serum exosome samples from healthy and treatment naïve HCV infected individuals. Our experiments made use of molecular biology and immunology techniques.ResultsHCV infection (JFH1-Huh7 or HCV-FL replicon cells) was associated with increased RTN3L&S isoforms expression in cells and cell released exosomes. Accordingly, increased expression of RTN3L&S was observed in liver and serum exosome samples of HCV infected individuals compared to healthy controls. RNA-ChIP analysis revealed that RTN3L&S interacted with dsHCV RNA. Lentiviral CRISPR/Cas9 mediated knockdown (KD) of RTN3 and plasmid overexpression (OE) of wild type, C- and N-terminal deletion mutants of RTN3L&S in HCV- infected Huh7 cells differentially impacted the cellular release of infectious viral exosomes. RTN3L&S KD significantly decreased, while RTN3S OE significantly increased the number of Huh7 cell-released infectious exosomes. The C-terminal domain of RTN3 interacted with and modulated the loading of dsHCV RNA inside infectious exosomes. Antiviral treatment of HCV infected Huh7 cells reduced virus-induced RTN3L&S expression and attenuated the release of infectious exosomes.ConclusionRTN3 constitutes a novel regulator and a potential therapeutic target that mediates the specific loading of infectious viral exosomes.

This work applies a data-driven coding method for prediction of the COVID-19 spreading profile in any given population that shows an initial phase of epidemic progression. Based on the historical data collected for COVID-19 spreading in 367 cities in China and the set of parameters of the augmented Susceptible-Exposed-Infected-Removed (SEIR) model obtained for each city, a set of profile codes representing a variety of transmission mechanisms and contact topologies is formed. By comparing the data of an early outbreak of a given population with the complete set of historical profiles, the best fit profiles are selected and the corresponding sets of profile codes are used for prediction of the future progression of the epidemic in that population. Application of the method to the data collected for South Korea, Italy and Iran shows that peaks of infection cases are expected to occur before the end of March 2020, and that the percentage of population infected in each city will be less than 0.01%, 0.05% and 0.02%, for South Korea, Italy and Iran, respectively.

Aims DPP-4 inhibitors are predicted to exert a protective effect on the progression of coronavirus disease 2019 (COVID-19). We conducted this meta-analysis to investigate this hypothesis. Methods Four databases, namely, PubMed, Web of Science, EMBASE and the Cochrane Library, were used to identify studies on DPP-4 and COVID-19. The outcome indicators were the mortality of COVID-19. Funnel plots, Begg’s tests and Egger’s tests were used to assess publication bias. Results Four articles were included with a total of 1933 patients with COVID-19 and type 2 diabetes. The use of DPP-4 inhibitors was negatively associated with the risk of mortality (odds ratio (OR) = 0.58 95% confidence interval (CI), 0.34–0.99). Conclusions DPP-4 inhibitors may improve the mortality of patients with COVID-19 and type 2 diabetes. As few relevant studies are available, more large-scale studies need to be performed.

Diverse fungal endophytes live in plants and are shaped by some abiotic and biotic stresses. Plant disease as particular biotic stress possibly gives an impact on the communities of fungal endophytes. In this study, clubroot disease caused by an obligate biotroph protist, Plasmodiophora brassicae, was considered to analyze its influence on the fungal endophyte community using an internal transcribed spacer (ITS) through high-throughput sequencing and culture-dependent methods. The results show that the diversity of the endophyte community in the healthy roots was much higher than the clubroots. Ascomycota was the dominant group of endophytes (including Phoma, Mortierella, Penicillium, etc.) in the healthy roots while P. brassicae was the dominant taxon in the clubroots. Hierarchical clustering, principal component analysis (PCA) and principal coordinates analysis (PCoA) indicated significant differences between the endophyte communities in the healthy roots and clubroots. Linear discriminant analysis effect size (LefSe) analysis showed that the dominant genera could be regarded as potential biomarkers. The endophyte community in the healthy roots had a more complex network compared with the clubroots. Also, many plant pathogenic Fusarium were isolated from the clubroots by the culture-dependent method. The outcome of this study illustrates that P. brassicae infection may change the fungal endophyte community associated with the roots of tumourous stem mustard and facilitates the entry of soil pathogen into the roots.

Objectives The aim of this study is to clarify the associations between IL-1B31C/T, IL-1B-511C/T, IL-8-251T/A gene polymorphisms and the risk of Helicobacter pylori (H. pylori) infection together with H. pylori-related gastric cancer (GC), peptic ulcer disease (PUD). Methods All eligible literature published up to July 2016 were identified by searching Pubmed, Embase, Web of Science and CNKI. Pooled odds ratio (OR) and 95% confidence interval (95% CI) were calculated using a fixed or random effects model. Results 29 case-control studies were eligible, and each of them may focus on more than one gene polymorphism. Ultimately, there were 21 studies (3159 cases and 2816 controls) for IL-1B-31C/T, 16 studies (2486 cases and 1989 controls) for IL-1B-511C/T polymorphisms, 9 studies (1963 cases and 1205 controls) for IL-8-251T/A polymorphisms. Overall, an increased risk of H. pylori infection was found for IL-1B-31C/T polymorphisms in total population [OR = 1.134, 95%CI = 1.008-1.275 for recessive model; OR = 1.145, 95%CI = 1.007-1.301 for TT vs CC model]. While, for IL-1B-511C/T and IL8-251T/A polymorphisms, no evidence indicated that they were associated with the risk of H. pylori infection in all genetic models. Furthermore, we found an increased risk of H. pylori-related GC with IL-1B-511C/T polymorphisms [OR = 1.784, 95%CI = 1.289-2.469 for recessive model; OR = 1.772, 95%CI = 1.210-2.594 for TT vs CC model] and IL8-251A/T polymorphisms [OR = 1.810, 95%CI = 1.229-2.667 for recessive model; OR = 1.717, 95%CI = 1.143-2.580 for TT vs AA model], an increased risk of H. pylori-related PUD with IL8-251T/A polymorphisms [OR = 1.364, 95%CI = 1.010-1.843 for recessive model; OR = 1.427, 95%CI = 1.039-1.959 for AA vs TT model]. Conclusions IL-1B-31C/T gene polymorphisms might increase H. pylori infection risk. IL-1B-511-C/T and IL-8-251T/A gene polymorphisms might act as a risk factor to H. pylori-related diseases including GC or PUD.

Hepatitis E virus (HEV) is one of the primary causative agents of acute hepatitis, and represents a major cause of severe public health problems in developing countries. The pathogenesis of HEV is not well characterized, however, primarily due to the lack of well-defined cell and animal models. Here, we investigated the effects of genotype 1 HEV open reading frame 3 (ORF3) on TNF-α-induced nucleus factor-κappa B (NF-κB) signaling. Human lung epithelial cells (A549) were transiently transfected with ORF3 containing plasmids. These cells were then stimulated with TNF-α and the nucleus translocation of the p65 NF-κB subunit was assessed using western blot and laser confocal microscopy. DNA-binding activity of p65 was also examined using electrophoretic mobility shift assay (EMSA), and the suppression of NF-κB target genes were detected using real-time RT-PCR and ELISA. These results enabled us to identify the decreased phosphorylation levels of IKBα. We focused on the gene of negative regulation of NF-κB, represented by TNF-α-induced protein 3 (TNFAIP3, also known as A20). Reducing the levels of A20 with siRNAs significantly enhances luciferase activation of NF-κB. Furthermore, HEV ORF3 regulated A20 primarily via activating transcription factor 6 (ATF6), involved in unfolded protein response (UPR), resulting in the degradation or inactivation of the receptor interacting protein 1 (RIP1), a major upstream activator of IKB kinase compounds (IKKs). Consequently, the phosphorylation of IKBα and the nucleus translocation of p65 are blocked, which contributes to diminished NF-κB DNA-binding activation and NF-κB-dependent gene expression. The findings suggest that genotype 1 HEV, through ORF3, may transiently activate NF-κB through UPR in early stage, and subsequently inhibit TNF-α-induced NF-κB signaling in late phase so as to create a favorable virus replication environment.

Prion diseases are marked by cerebral accumulation of the abnormal isoform of the prion protein. A fragment of prion protein composed of residues 106-126 (PrP106-126) exhibits similar properties to full length prion and plays a key role in the conformational conversion from cellular prion to its pathogenic pattern. Soluble oligomers of PrP106-126 have been proposed to be responsible for neurotoxicity. However, the monomeric conformational space and initial oligomerization of PrP106-126 are still obscure, which are very important for understanding the conformational conversion of PrP106-126. In this study, replica exchange molecular dynamics simulations were performed to investigate monomeric and dimeric states of PrP106-126 in implicit solvent. The structural diversity of PrP106-126 was observed and this peptide did not acquire stable structure. The dimeric PrP106-126 also displayed structural diversity and hydrophobic interaction drove the dimerization. To further study initial oligomerization of PrP106-126, 1 µs conventional molecular dynamics simulations of trimer and tetramer formation were carried out in implicit solvent. We have observed the spontaneous formation of several basic oligomers and stable oligomers with high β-sheet contents were sampled in the simulations of trimer and tetramer formation. The β-hairpin formed in hydrophobic tail of PrP106-126 with residues 118-120 in turn may stabilize these oligomers and seed the formation oligomers. This study can provide insight into the detailed information about the structure of PrP106-126 and the dynamics of aggregation of monomeric PrP106-126 into oligomers in atomic level.

OBJECTIVES: This study explored sexual knowledge, attitudes and practices of female only-child undergraduates and made a comparison with students with siblings. METHODS: Anonymously completed questionnaires were received from 4,769 female undergraduates, recruited using randomized cluster sampling by type of university and students' major and grade. Multivariate logistic regression was used to assess the effects of only-child on sexual knowledge, attitudes and practices among female undergraduates. RESULTS: Of 4,769 female undergraduate students, 41.0% were only-child and 59.0% were students with siblings. Compared with students with siblings, only-child students scored higher on sex-related knowledge, were more inclined to agree with premarital sex, multiple sex partners, one-night stands, extramarital lovers and homosexuality, and were more likely to have a boyfriend and experience sexual intercourse (73.6% vs. 61.4%; 24.0% vs. 14.0%). Only-children were less likely to experience coercion at first sex and have first sexual intercourse with men not their "boyfriends" than children with siblings (3.3% vs. 6.4%; 20.7% vs. 28.8%). There were no significant differences on other risky sexual behaviors (e.g. multiple sex partners and inconsistent condom use) between the only-child students and students with siblings. CONCLUSIONS: Sexual knowledge, attitudes and some practices of only-child female undergraduates were different from students with siblings. Intervention should be designed according to different requirements of only-children and non-only-children.

Enterovirus 71 (EV71), a major causative agent of hand-foot-and-mouth disease (HFMD), causes outbreaks among children in the Asia-Pacific region. A vaccine is urgently needed. Based on successful pre-clinical work, phase I and II clinical trials of an inactivated EV71 vaccine, which included the participants of 288 and 660 respectively, have been conducted. In the present study, the immune response and the correlated modulation of gene expression in the peripheral blood mononuclear cells (PBMCs) of 30 infants (6 to 11 months) immunized with this vaccine or placebo and consented to join this study in the phase II clinical trial were analyzed. The results showed significantly greater neutralizing antibody and specific T cell responses in vaccine group after two inoculations on days 0 and 28. Additionally, more than 600 functional genes that were up- or down-regulated in PBMCs were identified by the microarray assay, and these genes included 68 genes associated with the immune response in vaccine group. These results emphasize the gene expression profile of the immune system in response to an inactivated EV71 vaccine in humans and confirmed that such an immune response was generated as the result of the positive mobilization of the immune system. Furthermore, the immune response was not accompanied by the development of a remarkable inflammatory response. Clinical Trial Registration: NCT01391494 and NCT01512706.

Experimental studies on hepatitis B virus (HBV) replication are commonly done with human hepatoma cells to reflect the natural species and tissue tropism of the virus. However, HBV can also replicate, upon transfection of virus coding plasmids, in cells of other species. In such cross-species transfection experiments with chicken LMH hepatoma cells, we previously observed the formation of HBV genomes with aberrant electrophoretic mobility, in addition to the those DNA species commonly seen in human HepG2 hepatoma cells. Here, we report that these aberrant DNA forms are mainly due to excessive splicing of HBV pregenomic RNA and the abundant synthesis of spliced DNA products, equivalent to those also made in human cells, yet at much lower level. Mutation of the common splice acceptor site abolished splicing and in turn enhanced production of DNA from full-length pgRNA in transfected LMH cells. The absence of splicing made other DNA molecules visible, that were shortened due to the lack of sequences in the core protein coding region. Furthermore, there was nearly full-length DNA in the cytoplasm of LMH cells that was not protected in viral capsids. Remarkably, we have previously observed similar shortened genomes and non-protected viral DNA in human HepG2 cells, yet exclusively in the nucleus where uncoating and final release of viral genomes occurs. Hence, two effects reflecting capsid disassembly in the nucleus in human HepG2 cells are seen in the cytoplasm of chicken LMH cells.

H9N2 influenza viruses have been circulating worldwide in multiple avian species and have repeatedly infected humans to cause typical disease. The continued avian-to-human interspecies transmission of H9N2 viruses raises concerns about the possibility of viral adaption with increased virulence for humans. To investigate the genetic basis of H9N2 influenza virus host range and pathogenicity in mammals, we generated a mouse-adapted H9N2 virus (SD16-MA) that possessed significantly higher virulence than wide-type virus (SD16). Increased virulence was detectable after 8 sequential lung passages in mice. Five amino acid substitutions were found in the genome of SD16-MA compared with SD16 virus: PB2 (M147L, V250G and E627K), HA (L226Q) and M1 (R210K). Assessments of replication in mice showed that all of the SD16-MA PB2, HA and M1 genome segments increased virus replication; however, only the mouse-adapted PB2 significantly increased virulence. Although the PB2 E627K amino acid substitution enhanced viral polymerase activity and replication, none of the single mutations of mouse adapted PB2 could confer increased virulence on the SD16 backbone. The combination of M147L and E627K significantly enhanced viral replication ability and virulence in mice. Thus, our results show that the combination of PB2 amino acids at position 147 and 627 is critical for the increased pathogenicity of H9N2 influenza virus in mammalian host.

BACKGROUND: Hepatitis B virus (HBV) infection remains a severe public health problem. Investigating its prevalence and trends is essential to prevention. METHODS: To evaluate the effectiveness of HBV vaccination under the 1992 Intervention Program for infants and predicted HBV prevalence trends under the 2011 Program for all ages. We conducted a community-based investigation of 761,544 residents of 12 counties in Zhejiang Province selected according to their location, population density, and economic development. The HBV prevalence trends were predicted by a time-shifting approach. HBV surface antigen (HBsAg) and alanine amino transferase (ALT) were determined. RESULTS: Of the 761,544 persons screened for HBsAg, 54,132 were positive (adjusted carrier rate 6.13%); 9,455 had both elevated ALT and a positive HBsAg test (standardized rate 1.18%). The standardized HBsAg carrier rate for persons aged ≤20 years was 1.51%. Key factors influencing HBV infection were sex, age, family history, drinking, smoking, employment as a migrant worker, and occupation. With the vaccination program implemented in 2011, we predict that by 2020, the HBsAg carrier rate will be 5.27% and that for individuals aged ≤34 years will reach the 2% upper limit of low prevalence according to the WHO criteria, with a standardized rate of 1.86%. CONCLUSIONS: The national HBV vaccination program for infants implemented in 1992 has greatly reduced the prevalence of HBV infection. The 2011 program is likely to reduce HBV infection in Zhejiang Province to a low moderate prevalence, and perinatal transmission is expected to be controlled by 2020.

Mycoplasma bovis pneumonia in cattle has been epidemic in China since 2008. To investigate M. bovis pathogenesis, we completed genome sequencing of strain HB0801 isolated from a lesioned bovine lung from Hubei, China. The genomic plasticity was determined by comparing HB0801 with M. bovis strain ATCC® 25523™/PG45 from cow mastitis milk, Chinese strain Hubei-1 from lesioned lung tissue, and 16 other Mycoplasmas species. Compared to PG45, the genome size of HB0801 was reduced by 11.7 kb. Furthermore, a large chromosome inversion (580 kb) was confirmed in all Chinese isolates including HB0801, HB1007, a strain from cow mastitis milk, and Hubei-1. In addition, the variable surface lipoproteins (vsp) gene cluster existed in HB0801, but contained less than half of the genes, and had poor identity to that in PG45, but they had conserved structures. Further inter-strain comparisons revealed other mechanisms of gene acquisition and loss in HB0801 that primarily involved insertion sequence (IS) elements, integrative conjugative element, restriction and modification systems, and some lipoproteins and transmembrane proteins. Subsequently, PG45 and HB0801 virulence in cattle was compared. Results indicated that both strains were pathogenic to cattle. The scores of gross pathological assessment for the control group, and the PG45- and HB0801-infected groups were 3, 13 and 9, respectively. Meanwhile the scores of lung lesion for these three groups were 36, 70, and 69, respectively. In addition, immunohistochemistry detection demonstrated that both strains were similarly distributed in lungs and lymph nodes. Although PG45 showed slightly higher virulence in calves than HB0801, there was no statistical difference between the strains (P>0.05). Compared to Hubei-1, a total of 122 SNP loci were disclosed in HB0801. In conclusion, although genomic plasticity was thought to be an evolutionary advantage, it did not apparently affect virulence of M. bovis strains in cattle.

The aim of this study was to investigate the prevalence of swine hepatitis E virus (HEV) in pigs fed different feedstuffs (kitchen residue or mixed feeds) and genetic identification of HEV isolated in Hebei province, China. Serum and fecal samples were collected from adult swine. Anti-HEV antibody was evaluated by double sandwich antigen enzyme immunoassay. HEV RNA was extracted from fecal samples and amplified by nested RT-PCR. The reaction products were sequenced, and the sequence analyzed. Virus-like particles were distinguishable by negative staining in the electron microscope. Histopathological observation and immunohistochemical localization were used in the animal models. Overall, the anti-HEV positive percentage of serum samples from pigs fed on kitchen residue was 87.10% (27/31), and 53.06% (130/245) from pigs fed on complete feed. The HEV RNA positivity rate of fecal samples from pigs fed on kitchen residue was 61.54% (8/13), but zero for pigs fed on complete feed. Sequence analysis of these eight samples and comparison with the published sequence showed that there were eight groups that belonged to genotype 4 d and the nucleotide identity was 95.6–99.3%. swHE11 is most closely related to strain CCC220, and the other seven HEV isolates were most closely related to strains swGX40, SwCH189 and V0008ORF3, which are isolates from human and pigs. Histopathological observation showed that there was liver damage in the experimental group, and immunohistochemistry indicated that the HEV antigens were strongly positive at 7 days after infection. The results demonstrated that the prevalence of HEV in pigs fed on kitchen residue was higher than in those fed on complete feed (P

Background The influenza viruses circulating in animals sporadically transmit to humans and pose pandemic threats. Animal models to evaluate the potential public health risk potential of these viruses are needed. Methodology/Principal Findings We investigated the guinea pig as a mammalian model for the study of the replication and transmission characteristics of selected swine H1N1, H1N2, H3N2 and avian H9N2 influenza viruses, compared to those of pandemic (H1N1) 2009 and seasonal human H1N1, H3N2 influenza viruses. The swine and avian influenza viruses investigated were restricted to the respiratory system of guinea pigs and shed at high titers in nasal tracts without prior adaptation, similar to human strains. None of the swine and avian influenza viruses showed transmissibility among guinea pigs; in contrast, pandemic (H1N1) 2009 virus transmitted from infected guinea pigs to all animals and seasonal human influenza viruses could also horizontally transmit in guinea pigs. The analysis of the receptor distribution in the guinea pig respiratory tissues by lectin histochemistry indicated that both SAα2,3-Gal and SAα2,6-Gal receptors widely presented in the nasal tract and the trachea, while SAα2,3-Gal receptor was the main receptor in the lung. Conclusions/Significance We propose that the guinea pig could serve as a useful mammalian model to evaluate the potential public health threat of swine and avian influenza viruses.

Rabbit hepatitis E virus (HEV) is a novel genotype of HEV, and is considered to pose a risk of zoonotic transmission. Research into the systemic distribution of rabbit HEV in rabbits during different periods of infection has rarely been reported. To better understand this virus, we infected rabbits with second-passage rabbit HEV via an intraperitoneal route. After inoculation, the infection showed two types, temporary and constant infection. The detection of HEV RNA in the feces varied with time, and serum antigen correlated with fecal HEV RNA. Viremia only appeared 72 days after inoculation. The rabbits remained antibody negative throughout the experimental period. When HEV was localized, several organs besides the liver were HEV RNA positive. Tissue antigen was observed immunohistochemically in the different cells of various organs, especially in parts of the small intestine and the characteristic rabbit gut-associated lymphoid tissue. These data provide valuable information for future research into the pathogenesis of HEV.

Enterovirus 71 (EV71) is the major pathogen responsible for fatal hand, foot and mouth disease (HFMD). Our previous work reported on an EV71-infected rhesus monkey infant model that presented with histo-pathologic changes of the central nervous system (CNS) and lungs. This study is focused on the correlated modulation of gene expression in the peripheral blood mononuclear cells (PBMCs) from EV71-infected rhesus monkey infants. The expression of more than 500 functional genes associated with multiple pathways was modulated. The expression of genes associated with immune inflammatory responses was up-regulated during the period from days 4 to 10 post-infection. The expression of two genes (TAC1 and IL17A), which play major roles in inflammatory reactions, was remarkably up-regulated during the infection period. Furthermore, a higher expression level of the TAC1 gene was identified in the CNS compared to the lungs, but a high expression level of the IL-17A gene was observed in the lungs and not in the CNS. The results of this study suggest at least two facts about EV71 infection, which are that: the TAC1 gene that encodes substance P and neurokinin-A is present in both PBMCs and the hypothalamus; and the up-regulation of IL-17A is sustained in the peripheral blood.

OBJECTIVE: Hepatitis B virus (HBV) infection remains a significant public health problem. The purpose of this study was to investigate the seroepidemiology of HBV in people living in the insular regions, and to provide the most recent baseline data for planning and monitoring of health. METHODS: A cross-sectional, community-based survey was conducted without age restriction, on two isolated islands, Zhoushan and Yuhuan, China. The study sample was selected by random multistage cluster sampling. Serological samples and demographic information were collected from 15878 participants. RESULTS: The prevalences of anti-HBV core antibody (anti-HBc), hepatitis B virus surface antigen (HBsAg), and anti-HBV surface antibody (anti-HBs) were 33.1, 10.4, and 56.1%, respectively. We found statistically significant differences of HBV markers in men versus women (P38 IU/L) in 15.6% and 7.2% of the HBsAg-positive and negative groups, respectively. Elevated ALT levels were significantly higher in males (12.0%) compared with females (5.8%) (P

Opiates are among the most prescribed drugs for pain management. However, morphine use or abuse results in significant gut bacterial translocation and predisposes patients to serious infections with gut origin. The mechanism underlying this defect is still unknown. In this report, we investigated the mechanisms underlying compromised gut immune function and bacterial translocation following morphine treatment. We demonstrate significant bacterial translocation to mesenteric lymph node (MLN) and liver following morphine treatment in wild-type (WT) animals that was dramatically and significantly attenuated in Toll-like receptor (TLR2 and 4) knockout mice. We further observed significant disruption of tight junction protein organization only in the ileum but not in the colon of morphine treated WT animals. Inhibition of myosin light chain kinase (MLCK) blocked the effects of both morphine and TLR ligands, suggesting the role of MLCK in tight junction modulation by TLR. This study conclusively demonstrates that morphine induced gut epithelial barrier dysfunction and subsequent bacteria translocation are mediated by TLR signaling and thus TLRs can be exploited as potential therapeutic targets for alleviating infections and even sepsis in morphine-using or abusing populations.