Inhibiting microRNA-424 in bone marrow mesenchymal stem cells-derived exosomes suppresses tumor growth in colorectal cancer by upregulating TGFBR3.

OBJECTIVE: MicroRNAs (miRNAs) have been demonstrated to be differently expressed in colorectal cancer (CRC) and were identified as biomarkers and therapeutic targets for CRC. We aimed to identify the effect of microRNA-424 (miR-424) on process of CRC. METHODS: Exosomes were obtained from bone marrow mesenchymal stem cells (BMSCs). MiR-424, transforming growth factor-ß receptor 3 (TGFBR3) vimentin, S100A4, p-Smad1 expression in tissues and cells was measured. After treated with miR-424 inhibitor or TGFBR3 overexpression plasmid, the migration, invasion, cell cycle distribution and apoptosis of Lovo cells and exosomes-transfected Lovo cells were determined. The subcutaneous tumor models were established and the tumor growth was observed. The target relation between miR-424 and TGFBR3 was confirmed. RESULTS: MiR-424 was upregulated while TGFBR3 was downregulated in CRC tissues. TGFBR3 was targeted by miR-424. Inhibited miR-424 or elevated TGFBR3 upregulated p-Smad1, indicating that TGFBR3 mediated the Smad1 pathway, thus regulating CRC progression. MiR-424 inhibition or TGFBR3 restoration also suppressed migration and invasion of CRC cells, arrested the CRC cells at G0/G1 phase, and promoted CRC cell apoptosis. Moreover, exosomal miR-424 from BMSCs promoted CRC development. CONCLUSION: Inhibited exosomal miR-424 from BMSCs inhibited malignant behaviors of CRC cells by targeting TGFBR3, thus suppressing the progression of CRC.

Japanese Spotted Fever in Eastern China, 2013.

We isolated Rickettsia japonica from a febrile patient in Lu'an City, China, in 2013. Subsequently, we found an R. japonica seroprevalence of 54.8% (494/902) in the rural population of Anhui Province and an R. japonica prevalence in Haemaphysalis longicornis ticks of 0.5% (5/935). R. japonica and its tick vector exist in China.

Prevalence of Yersinia enterocolitica Bioserotype 3/O:3 among Children with Diarrhea, China, 2010-2015.

Yersinia enterocolitica is thought to not significantly contribute to diarrheal disease in China, but evidence substantiating this claim is limited. We determined the prevalence of Y. enterocolitica infection and strain types present among children <5 years of age with diarrhea in China. The overall prevalence of pathogenic isolates was 0.59%. Prevalence of pathogenic bioserotype 3/O:3 varied geographically. In this population, the presence of fecal leukocytes was a characteristic of Y. enterocolitica infection and should be used as an indication for microbiological diagnostic testing, rather than for the diagnosis of bacillary dysentery. In contrast with Y. enterocolitica isolates from adults, which were primarily biotype 1A, isolates from children were primarily bioserotype 3/O:3. Most pathogenic isolates from children shared pulsed-field gel electrophoresis patterns with isolates from pigs and dogs, suggesting a possible link between isolates from animals and infections in children. Our findings underscore the need for improved diagnostics for this underestimated pathogen.

Human infection with a novel avian-origin influenza A (H7N9) virus.

BACKGROUND: Infection of poultry with influenza A subtype H7 viruses occurs worldwide, but the introduction of this subtype to humans in Asia has not been observed previously. In March 2013, three urban residents of Shanghai or Anhui, China, presented with rapidly progressing lower respiratory tract infections and were found to be infected with a novel reassortant avian-origin influenza A (H7N9) virus. METHODS: We obtained and analyzed clinical, epidemiologic, and virologic data from these patients. Respiratory specimens were tested for influenza and other respiratory viruses by means of real-time reverse-transcriptase-polymerase-chain-reaction assays, viral culturing, and sequence analyses. RESULTS: A novel reassortant avian-origin influenza A (H7N9) virus was isolated from respiratory specimens obtained from all three patients and was identified as H7N9. Sequencing analyses revealed that all the genes from these three viruses were of avian origin, with six internal genes from avian influenza A (H9N2) viruses. Substitution Q226L (H3 numbering) at the 210-loop in the hemagglutinin (HA) gene was found in the A/Anhui/1/2013 and A/Shanghai/2/2013 virus but not in the A/Shanghai/1/2013 virus. A T160A mutation was identified at the 150-loop in the HA gene of all three viruses. A deletion of five amino acids in the neuraminidase (NA) stalk region was found in all three viruses. All three patients presented with fever, cough, and dyspnea. Two of the patients had a history of recent exposure to poultry. Chest radiography revealed diffuse opacities and consolidation. Complications included acute respiratory distress syndrome and multiorgan failure. All three patients died. CONCLUSIONS: Novel reassortant H7N9 viruses were associated with severe and fatal respiratory disease in three patients. (Funded by the National Basic Research Program of China and others.).

Genomic portrait of the evolution and epidemic spread of a recently emerged multidrug-resistant Shigella flexneri clone in China.

Shigella flexneri is the major cause of shigellosis in developing countries. A new S. flexneri serotype, Xv, appeared in 2000 and replaced serotype 2a as the most prevalent serotype in China. Serotype Xv is a variant of serotype X, with phosphoethanolamine modification of its O antigen mediated by a plasmid that contained the opt gene. Serotype Xv isolates belong to sequence type 91 (ST91). In this study, whole-genome sequencing of 59 S. flexneri isolates of 14 serotypes (serotypes 1 to 4, Y, Yv, X, and Xv) indicated that ST91 arose around 1993 by acquiring multidrug resistance (MDR) and spread across China within a decade. A comparative analysis of the chromosome and opt-carrying plasmid pSFXv_2 revealed independent origins of 3 serotype Xv clusters in China, with different divergence times. Using 18 cluster-dividing single-nucleotide polymorphisms (SNPs), SNP typing divided 380 isolates from 3 provinces (Henan, Gansu, and Anhui) into 5 SNP genotypes (SGs). One SG predominated in each province, but substantial interregional spread of SGs was also evident. These findings suggest that MDR is the key selective pressure for the emergence of the S. flexneri epidemic clone and that Shigella epidemics in China were caused by a combination of local expansion and interregional spread of serotype Xv.

O:8 serotype Yersinia enterocolitica strains in China.

Serotypes O:3, O:8 and O:9 Yersinia enterocolitica strains carrying virulence determinants are common pathogens causing human infections. In many years of surveillance in China for Y. enterocolitica, no pathogenic O:8 strains have been found where the isolated O:8 serotypes lacked the major virulence genes and in contrast to O:3 and O:9 strains, none of the O:8 isolates were from humans. These O:8 isolates lack ail, ystA, yadA and virF genes but possess the ystB gene and all belong to Biotype 1A. These O:8 strains did not kill mice and could protect immunized mice against challenge with a pathogenic O:8 strain. Compared to the Chinese pathogenic O:3 and O:9 strains which have similar pulsed-field gel electrophoresis patterns, the 39 Chinese O:8 animal and food isolates were different from the pathogenic O:8 reference strains. This suggests the O:8 strains lacking virulence determinants may not disseminate rapidly in humans and are maintained in animal reservoirs; and therefore exhibit higher variance and divergence from the virulent type.

[Phylogenetic Analysis of the VP1 Region of Coxsackievirus A16 Strains Isolated in Anhui Province, 2014].

To study on the phylogenetic characterization of the VP1 genes of coxsackievirus A16 (CVA16) causing hand-food-mouth disease (HFMD) isolated from Anhui province in 2014. A total of 413 throat swab specimens from HFMD patients were collected during January to November, 2014 for the isolation and identification of enteroviruses using real-time RT-PCR assays. The VP1 regions of CVA16 isolates were amplified using RT-PCR and sequenced. And the phylogenetic tree was constructed among the VP1 regions of those isolates, the different genotypes and sub-genotypes of CVA16 strains. A total of 97 enteroviruses were isolated from 413 samples, the positive rate was 23.49% (97/413), including seventeen CVA16, seventy six HEV71 and four other enteroviruses. The results of the phylogenetic tree showed that 17.CVA16 strains isolated from Anhui in 2014 clustered within B1b evolution branch of B1 genotype. The nucleotide and amino acid sequence identities were 95.30%-100% and 98.70%-100% among the isolates, respectively, but within B1b branch of 17 strains formed several small transmission chains. The nucleotide acid of 17 CVA16 isolates in Anhui province were closed to the strains isolated from Yunnan, Hunan, Guangdong, Tibet and Jiangsu, especially from Hunan in 2013 and from Shenzhen of Guangdong in 2014, the identity were 96.40%-99.70%. The CVA16 strains isolated from Anhui in 2014 were all belong to genetic subtype B1b of B1 genotype was dominant, and among those isolates, several small virus transmission chains had formed with co-circulating and evolution.

Influenza vaccine induces intracellular immune memory of human NK cells.

Influenza vaccines elicit antigen-specific antibodies and immune memory to protect humans from infection with drift variants. However, what supports or limits vaccine efficacy and duration is unclear. Here, we vaccinated healthy volunteers with annual vaccine formulations and investigated the dynamics of T cell, natural killer (NK) cell and antibody responses upon restimulation with heterologous or homologous influenza virus strains. Influenza vaccines induced potential memory NK cells with increased antigen-specific recall IFN-γ responses during the first 6 months. In the absence of significant changes in other NK cell markers (CD45RO, NKp44, CXCR6, CD57, NKG2C, CCR7, CD62L and CD27), influenza vaccines induced memory NK cells with the distinct feature of intracellular NKp46 expression. Indeed, surface NKp46 was internalized, and the dynamic increase in NKp46(intracellular)+CD56dim NK cells positively correlated with increased IFN-γ production to influenza virus restimulation after vaccination. In addition, anti-NKp46 antibodies blocked IFN-γ responses. These findings provide insights into a novel mechanism underlying vaccine-induced immunity and NK-related diseases, which may help to design persisting and universal vaccines in the future.

[Application of PulseNet China Database in tracking and warning 4 cholerae outbreaks in Anhui province, in 2012].

OBJECTIVE: To track the source of infection regarding 4 Cholerae outbreaks in Anhui province in 2012 through the application of PulseNet China Database (PNCD). METHODS: Cholerae virulence genes were amplified by PCR and typed by pulse field gel electrophoresis(PFGE). Results from electrophoresis were cluster-analyzed by BioNumericsV4.0 software and compared with PNCD. RESULTS: Virulence gene CT and TCP of the tested vibrio cholera showed both positive. Homology of the strains from four cholera outbreaks was more than 98%, based on the homologous and cluster analysis through enzyme digested PFGE electrophoresis. Those strains were highly homologous with the cholera epidemic strains identified in Hunan, Sichuan,Zhejiang, Shanghai and Hubei by PNCD, with the homology as 100% . CONCLUSION: Four cholera outbreaks in Anhui province, 2012 were highly correlated with the outbreaks occurring in Hunan and Sichuan during the same time period, indicating that PNCD could effectively and quickly tracking down the source of infection on the cholera outbreaks and providing early warning of the situation.

Person-to-person transmission of severe fever with thrombocytopenia syndrome virus.

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging infectious disease caused by a newly discovered bunyavirus, SFTS virus (SFTSV), and causes high fatality (12% on average and as high as 30%). The objective of this study was to determine whether SFTSV could be transmitted from person to person. We analyzed sera of 13 patients from two clusters of unknown infectious diseases that occurred between September and November of 2006 in Anhui Province of China for SFTSV antibody by indirect immunofluorescence assay and for SFTSV RNA by RT-PCR. We found that all patients (n=14) had typical clinical symptoms of SFTS including fever, thrombocytopenia, and leukopenia and all secondary patients in both clusters got sick at 6-13 days after contacting or exposing to blood of index patients. We demonstrated that all patients in cluster 1 including the index patient and nine secondary patients and all three secondary patients in cluster 2 had seroconversion or fourfold increases in antibody titer to SFTSV and/or by RT-PCR amplification of SFTSV RNA from the acute serum. The index patient in cluster 2 was not analyzed because of lack of serum. No person who contacted the index patient during the same period, but were not exposed to the index patient blood, had got illness. We concluded that SFTSV can be transmitted from person to person through contacting patient's blood.

[Molecular analysis on non-O1 and non-O139 Vibrio cholerae isolates].

OBJECTIVE: According to results from the two-month consecutive surveillance program in Maanshan, six suspected cases of non-O1 non-O139 Vibrio (V.) cholerae infection, were found that called for identification of pathogens as well as molecular-epidemiological analysis to determine the aggregation of the epidemic situation. METHODS: Biochemical and serotype identification, hemolysis test, and drug sensitive test were used to detect the drug resistance spectrum. Real-time PCR and conventional PCR were used to detect the presence of V. cholerae specific genes, virulent genes and its related genes, including ompW, ctx, tcpA, toxR, hlyA, zot, ace, rstR and gIII(CTX). Pulsed-field gel electrophoresis (PFGE) was used to analyze the molecular type of strains. RESULTS: All the six isolates of non-O1 non-O139 V. cholerae were identified by biochemical and serologic tests, and appeared to be ß hemolytic. Twelve out of the 14 kinds of drugs showed 100% sensitive. All isolates were positive of ompW gene by real-time PCR, but negative for ctx, tcpA, zot, ace, rstR and gIII(CTX). Five of the six isolates were positive for toxR and hlyA, except for strain 1001434446. All strains had different PFGE types, but two strains had similar types. All strains had a low similarity compared to the toxigenic V. cholerae. CONCLUSION: Six cases of non-O1 and non-O139 nontoxigenic V. cholerae infection appeared in the same period. Along with epidemiological information, we noticed that these cases had a sporadic nature, but frequently appeared in the same area. We got the impression that public health measurements should be strengthened, with special attention paid to those diarrhea outbreaks caused by non-O1/non-O139 strains since V. cholerae had appeared in low incidence.

[Molecular epidemiology of enterohaemorrhagic Esacherichia coli O157 in some areas in China].

OBJECTIVE: To understand the epidemiological characteristics of enterohaemorrhagic Escherichia coli (EHEC) O157 and to determine the degree of its genetic relations. METHODS: Polymerase chain reaction (PCR) techniques and chromosomal DNA digested by restriction enzyme Xba I according to PulseNet directions by pulsed field gel electrophoresis (PFGE) method were applied to 300 E. coli O157 strains isolated from patients and animal sources from 1988 to 2005 from Henan, Jiangsu and Anhui provinces. RESULTS: Very high prevalence of stx2 gene in EHEC O157:H7 strains isolated from some provinces of China was found and variation existed in some strains. We got 161 PFGE patterns from 300 strains. The stx2-producing strains could be clearly separated from stx2 variation-producing strains. CONCLUSION: The variability of restriction enzyme-digestion patterns of O157 genomes suggested that the presence of some genomic diversity among the strains did exist.

[The firstly confirmed pregnant woman case of avian influenza A (H5N1) by etiological research in China].

To investigate the cause of death of a pregnant woman with undefined pneumonia reported from the People's Hospital of Tongling City in Anhui Province on November 8, 2005, the patient's tracheal aspirates and serum samples were collected and tested by RT-PCR and Real-time PCR to detect viral nucleic acids of HA of A/H5N1, A/H7N7, A/H9N1 and A/M. Tracheal aspirates were inoculated into special pathogen free (SPF) embryonated eggs for cultivation and identification of virus. The HA gene of the virus was sequenced and analyzed. Serum samples were tested by HI assay to detect antibody of H5N1. The results showed that HA gene of A/H5N1 virus and A/M were positive in tracheal aspirates by both PCR tests. The serum sample collected on Nov. 9 was A/M gene positive by Real-time PCR. The analysis of HA gene of A/AnHui/1/2005 sequence showed that the receptor specificity and the connecting peptide between HA1 and HA2 were still avian influenza origin. The HI antibody of H5N1 was negative at 7th, 8th, 9th d of disease onset. This undefined pneumonia case was confirmed as the first pregnant woman case of avian influenza (H5N1) virus infection by etiology in the mainland of China.

[Molecular typing of the pathogenic Yersinia enterocolitica strains with pulsed field gel electrophores isolated in China].

OBJECTIVE: To investigate the epidemiological and molecular typing features of the pathogenic Yersinia enterocolitica strains isolated in China,using pulsed field gel electrophoresis(PFGE) and standardized PFGE method as well as typing database of Yersinia enterocolitica. METHODS: PFGE analysis was performed as Laboratory Directions for molecular subtyping of Salmonella by PFGE (PulseNet,USA) with some modifications and the results of PFGE were analyzed by BioNumerics soft (Version 4.0, Applied Maths BVBA, Belium). RESULTS: 114 O:3 Yersinia enterocolitica strains were typed by 25 patterns to have found that K6GN11C30012 (50 strains), K6GN11C30015(19 strains) and K6GN11C30016(10 strains) were the major patterns. K6GNllC30012 had 92.2% cluster similarity with K6GN11C30009-K6GN11C30023. This clone included 91.23% strains of 114 0:3 Yersinia enterocolitica strains. 51 0:9 Yersinia enterocolitica strains were typed by 14 patterns; K6GN11C90004 (22 strains) and K6GN11C90010 (13 strains)were the major patterns. K6GN11C90004 had 81.8% cluster similarity with K6GN11C90010 patterns. The major patterns of 0:3 and 0:9 serotypes were quite different. CONCLUSION: O:3 Yersinia enterocolitica strains might originate from the same clone and had very few variation in different years and provinces but O:9 Yersinia enterocolitica strains from two different clones with some changes.

[Study on a fatal pregnant woman died from by avian influenza (H5N1)].

OBJECTIVE: To ascertain the causation of a pregnant woman with undefined pneumonia reported from the People's Hospital of Tongling city in Anhui province on November 2005. METHODS: Epidemiological and clinical information of the case was collected from the keypersons close to the case and referring to the medical record. A medical observation was carried out on the close contacts of the case and sick or dead poultry. Tracheal aspirates being collected were tested by both RT-PCR and real-time PCR to detect viral nucleic acids of A/H5N1, and were inoculated into special pathogen free (SPF) embryonated hens' eggs. RESULTS: The pregnant woman was found to have been contacted with the sick/dead poultry directly on the 4th day before onset of illness. All the 122 close contacts were healthy after a 10-day medical observation. The major clinical features of the case were viral pneumonia with rapidly developed leukopenia and lymphopenia. The progress to acute respiratory distress syndrome and multiple organ dysfunction syndromes was found at clinical presentation. HA and NA gene of A/H5N1 virus were positive. The 8 gene fragments of A/Anhui/1/2005 (H5N1) isolated from the tracheal aspirates had not carried genes from a human virus through reassortment, and the receptor-binding site of the hemagglutinin was polybasic cleavage site. CONCLUSION: This was the first documented case of H5N1 infection in pregnant woman. The immunotolerant state of pregnancy might have predisposed to the fatal outcome of the patient.

[Study on a monitoring program regarding leptospirosis in some fore-and-after flood-affected along large rivers in Anhui province].

OBJECTIVE: The study was designed to find out the epidemic characteristics of leptospriosis and to develop effective intervention measures. The effects of floods on leptospriosis in some areas along Yangzi river and Huai river in Anhui province was also analysed. METHODS: Study on serum epidemiology of leptospriosis was carried out from serous samples collected from native residents and animal hosts including isolation of pathogens at different phases (before,middle and after) and different monitoring spots,during the floods. RESULTS: Infection rate with leptospriosis pathogen among native residents was 13.49% during the flood-period,much higher than 2.18% at post-flood (chi2 = 22.78, P < 0.01) stage, in the flood-affected areas along Yangzi river in 1998. The average rates of infection were 2.48% and 5.35% in affected and unaffected areas along Huai river respectively, in 2003. CONCLUSIONS: There was full evidence that floods causing the epidemics of leptospriosis. However, the transmission of leptospriosis among people would depend on affecting factors as scales of floods, lasting time, coincidence between flood happening and epidemic season, immuno-protection level against leptospriosis among people and so on to a great extent. Factors as the magnitude of pathogens carried by various kinds of infectious sources were also important determinants affecting the nature, being epidemic or pandemic of leptospriosis. It was suggested that active surveillance network on the sources of infection and risk factors of leptospriosis should be developed for the control and prevention of the disease, in the flood-hit areas.