Association of IL28B polymorphisms with peginterferon treatment response in Chinese Han patients with HBeAg-positive chronic hepatitis B.

AIMS: To investigate whether IL28B polymorphisms could affect the treatment response to peginterferon alpha (PEG-IFN) in chronic hepatitis B (CHB) patients in the Chinese Han population. METHODS: A total of 212 hepatitis B e antigen (HBeAg)-positive patients treated with PEG-IFN monotherapy were enrolled in this study. Genotype analysis was performed for IL28B rs12980275, rs12979860 and rs8099917 using the MassArray system. Response was defined as cases showing normal aminotransferase (ALT) levels, HBV DNA level < 200 IU/ml and HBeAg seroconversion after 48 weeks of PEG-IFN therapy. RESULTS: The patients were infected with hepatitis B virus (HBV) genotype B (44.8%) and C (55.2%) with a total response rate of 34.9%. For the three SNPs, there were significant differences between the response (R) and non-response (NR) groups both in allele frequencies and genotype distributions. IL28B genotype was independently associated with R for AA vs. N-AA (OR 2.70, 95% CL 1.21-6.01; P = 0.015) at rs12980275 after adjustment for sex, age, HBV genotype, baseline levels of HBV DNA and ALT. There were similar results for rs12979860 CC vs. N-CC (OR 2.56, 95% CL 1.15-5.67; P = 0.021) and rs8099917 TT vs. N-TT (OR 2.80, 95% CL 1.23-6.39; P = 0.015) respectively. Furthermore, one block formed by rs12980275 and rs12979860 was identified in this study. In multivariate analyses, the most common haplotype A-C was independently associated with high rates of R (OR 2.53, 95% CL 1.20-5.34; P = 0.015). CONCLUSIONS: Our study suggested that genetic variations in IL28B may play a critical role in PEG-IFN efficacy in HBeAg-positive CHB patients in Han Chinese.

Hepatitis B virus surface antigen selectively inhibits TLR2 ligand-induced IL-12 production in monocytes/macrophages by interfering with JNK activation.

It is widely accepted that chronic hepatitis B virus (HBV) infection is the result of an ineffective antiviral immune response against HBV infection. Our previous study found that the hepatitis B surface Ag (HBsAg) was related to decreased cytokine production induced by the TLR2 ligand (Pam3csk4) in PBMCs from chronic hepatitis B patients. In this study, we further explored the mechanism involved in the inhibitory effect of HBsAg on the TLR2 signaling pathway. The results showed that both Pam3csk4-triggered IL-12p40 mRNA expression and IL-12 production in PMA-differentiated THP-1 macrophage were inhibited by HBsAg in a dose-dependent manner, but the production of IL-1ß, IL-6, IL-8, IL-10, and TNF-α was not influenced. The Pam3csk4-induced activation of NF-κB and MAPK signaling were further examined. The phosphorylation of JNK-1/2 and c-Jun was impaired in the presence of HBsAg, whereas the degradation of IκB-α, the nuclear translocation of p65, and the phosphorylation of p38 and ERK-1/2 were not affected. Moreover, the inhibition of JNK phosphorylation and IL-12 production in response to Pam3csk was observed in HBsAg-treated monocytes/macrophages (M/MΦs) from the healthy donors and the PBMCs and CD14-positive M/MΦs from chronic hepatitis B patients. Taken together, these results demonstrate that HBsAg selectively inhibits Pam3csk4- stimulated IL-12 production in M/MΦs by blocking the JNK-MAPK pathway and provide a mechanism by which HBV evades immunity and maintains its persistence.

[The infection and molecular characteristics of Vibrio parahaemolyticus isolated from intestinal outpatient in two sentinel hospitals in Shanghai, 2010-2012].

OBJECTIVE: To analyze virulence genes and molecular characteristics of Vibrio parahaemolyticus isolated from sporadic cases with diarrhea in tow sentinel hospitals of Shanghai, 2010-2012. METHODS: A total of 2 729 stool samples were collected from two surveillance sentinel hospitals in Shanghai 2010-2012. Vibrio parahaemolyticus strains were isolated and identified from diarrhea out patients using TCBS agar plates and biochemical reactions. Thermostable direct hemolysingene (tdh), thermostable-related hemolysin gene (trh), hemolysin gene (tlh) were detected by multiplex PCR method. Isolates were analyzed by PFGE and MLST. The PFGE profiles were analyzed using BioNumerics software. RESULTS: A total of 30 clinical Vibrio parahaemolyticus strains isolated from 2 729 stool samples. The anually Vibrio parahaemolyticus isolation rate during 2010 to 2012 were 1.1%(11/973), 1.0%(11/1 120) and 1.3%(8/636) respectively. The PCR positive rates of virulence genes tlh, tdh and trh were 100%, 97% and 0 respectively. The Vibrio parahaemolyticus strains were divided into 13 PFGE types (P1-P13)and 3 ST types (ST-189, ST-799, ST-3). Among 13 PFGE types, P4 was the main PFGE type, accounting for 30%(9/30). P9, P10 were accounting for 12% (4/30) respectively, P1, P2, P12, P13 were accounting for 7%(2/30) respectively, the others types were 3%(1/30) respectively. MLST analysis results showed there are three ST types, ST3 was 84%(25/30), ST189 and ST799 were accounting for 13% (4/30) and 3% (1/30) respectively. CONCLUSION: The infection rate of Vibrio parahaemolyticus was not very high from 2010-2012 in Shanghai, all strains were positive for tlh and negative for trh. ST3 was the major type of Vibrio parahaemolyticus.

Rotavirus infection and its genetic characterization in non-hospitalized adults with acute gastroenteritis in Shanghai, China.

Rotavirus is a major cause of acute gastroenteritis in infants and young children, while its role as a pathogen in adults has long been underappreciated. In order to describe the epidemiological patterns and genetic characteristics of rotavirus causing sporadic acute gastroenteritis in adults, hospital-based surveillance of rotavirus infections was conducted in Shanghai, China, between June 2010 and May 2011. Stool specimens were collected from outpatients with acute gastroenteritis admitted to three local hospitals. Rotavirus was detected using a colloidal gold test device. G and P genotyping were performed by multiplex PCR assays, and the VP7 gene of G9 strains were sequenced for further genetic characterization. Of 1,479 adult diarrheal stool samples examined during the 1-year surveillance period, 138 (9.3 %) were found to be rotavirus positive. G1 appeared to be the predominant genotype (35.5 %), suggesting a shift from genotype G3 to G1 in the study population in Shanghai. Meanwhile, a high frequency of genotype G9 (27.5 %) was also observed, and G9 was also predominant (38.1 %) in the small number of children (n=123) involved in the present study. Other specificities detected in adults were G2 (12.3 %) and G3 (13.8 %). For P genotyping, only two types, P[8] and P[4], were detected. P[8] was dominant in both children and adults. Sequence and phylogenetic analysis revealed that these strains could be divided into two different groups, with clustering within G9 lineage 3 and the subcluster of Japanese and Chinese G9 strains, respectively. In comparison to the previous data, G9 strains established themselves in a short time span as an important genotype in Shanghai, China.

Measles virus infection in adults induces production of IL-10 and is associated with increased CD4+ CD25+ regulatory T cells.

Despite steady progress in elimination of measles virus globally, measles infection still causes 500,000 annual deaths, mostly in developing countries where endemic measles strains still circulate. Many adults are infected every year in China, with symptoms more severe than those observed in children. In this study, we have used blood samples from adult measles patients in Shanghai and age-matched healthy controls to gain an understanding of the immune status of adult measles patients. IFN-alpha mRNA was reduced in patient PBMC compared with healthy controls. In contrast, gene expression and plasma production of IL-2, IL-10, and IFN-gamma were elevated in patient blood. A similar cytokine profile was observed at early times when cultured PBMC were infected with a clinical isolate of measles virus. In contrast to previous studies in pediatric patients, we did not find a reduction in total CD4(+) and CD8(+) T cells in patient PBMC. Interestingly, we found that CD4(+)CD25(+)CD127(low) regulatory T cells were significantly increased in patient PBMC compared with controls. Using intracellular cytokine staining we also show that the measles virus induces IL-10-producing CD14(+) and CD4(+)CD25(+) cells in PBMC. Our results show that adult measles patients in the acute phase of the disease have a mixed Th1/Th2 type response, accompanied with severe immunosuppression of both innate and adaptive responses including suppression of type I IFN. Both regulatory T cells and plasma IL-10 may contribute to the immunosuppression.

Genotype distribution of norovirus around the emergence of Sydney_2012 and the antigenic drift of contemporary GII.4 epidemic strains.

BACKGROUND: The pattern of epochal evolution of NoV is ongoing, while novel GII.4 variants emerge and cause new pandemics. Since, the emergence in March 2012, Sydney_2012 had replaced GII.4-2009 as the primary NoV strain in most countries in the northern hemisphere by November 2012. OBJECTIVES: To determine the genotype distribution around the emergence of Sydney_2012 and to investigate the underlying evolution mechanisms of the contemporary GII.4 strains. STUDY DESIGN: From January 2012 to December 2013, molecular epidemiology of norovirus in 846 adults (≥16 years) in Shanghai were conducted. The VP1 proteins of the contemporary GII.4 strains (Den_Haag_2006b, New_Orleans_2009 and Sydney_2012) were expressed in vitro and purified. Receptor binding patterns of these three epidemic strains were determined through histo-blood group antigen (HBGA) binding assays. Convalescent serum from patients infected with GII.4 epidemic strains were employed to investigate the role of antigenic drift in the persistence of GII.4 epidemic strains through receptor-binding blockade assays. RESULTS: Epidemiological studies revealed that Sydeny_2012 has completely replaced Den_Haag_2006b and New_Orleans_2009 and has been the dominant circulating strain in Shanghai since its emergence in October 2012. Interestingly, Den_Haag_2006b and New_Orleans_2009 have been co-circulating in Shanghai before the emergence of Sydeny_2012. The contemporary GII.4 epidemic norovirus strains displayed commonly high tropism to the histo-blood group antigen receptors, whereas Sydeny_2012 was antigenically different from Den_Haag_2006b and New_Orleans_2009. CONCLUSIONS: Antigenic drift, rather than receptor switch, played a key role in the emergence and spreading of Sydney_2012. The contemporary GII.4 strains were evolving via epochal evolution without altered ligand binding profiles.

A novel norovirus GII.17 lineage contributed to adult gastroenteritis in Shanghai, China, during the winter of 2014­2015.

Norovirus (NoV) is now recognized as a leading cause of nonbacterial acute gastroenteritis; however, the NoV GII.17 genotype has rarely been reported as the predominant genotype in clinical diarrhea cases. During the winter of 2014­2015, the GII.17 genotype, together with the NoV GII.4 genotype, dominated in sporadic adult patients with gastroenteritis in Shanghai. Phylogenetic analysis based on full-length VP1 amino acid sequences showed that the GII.17 strains that emerged in Shanghai have close evolutionary relationships with strains recently collected in the Hong Kong area, Guangdong province of China, and Japan during the same period. This cluster in the phylogenetic tree may represent a novel NoV GII.17 lineage recently circulating in East Asia. Pairwise distances between clusters also revealed the evolution of the NoV GII.17 genotype in previous decades. Our study emphasizes the importance of combined surveillance of NoV-associated infections.

Genetic diversity of sapovirus in non-hospitalized adults with sporadic cases of acute gastroenteritis in Shanghai, China.

BACKGROUND: Sapovirus has been accepted as a major cause of acute gastroenteritis worldwide. It can affect all age groups, ranging from young adults to the elderly, while little is known about the epidemiological patterns and genetic characteristics of sapovirus infections in China. OBJECTIVES: To investigate the prevalence of sapovirus infections among adult outpatients suffering from acute gastroenteritis in Shanghai, China. STUDY DESIGN: From April 2011 to March 2013, fecal specimens from 1125 adult outpatients (≥16 years of age) with acute gastroenteritis were collected. Reverse transcription polymerase chain reaction (RT-PCR) was employed for detection of sapovirus, and 5' end of capsid gene were sequenced for genotyping and phylogenetic analysis. RESULTS: The overall occurrence of sapovirus infection in adult outpatients was 3.73% (42 in 1125) through the two-year surveillance period, and sapovirus diarrhea is more common in spring and winter. The highest sapovirus positive rate was observed in adults of ≥56 years old, and statistically significant relationship was observed when compared with other age groups (p<0.05). Only three genotypes were detected, whereas GI.2 was proved to be the predominant strain, occupying 78.57% (33 in 42) of all strains, followed by GIV, GI.1 and GII.3. CONCLUSIONS: Sapovirus was commonly found in adults with acute gastroenteritis in Shanghai, China, while no specific seasonal variation of sapovirus diarrhea could be distinguished. GI.2 strains established themselves in a short time span as the predominant genotype in Shanghai, China.

Comparison of circulating, hepatocyte specific messenger RNA and microRNA as biomarkers for chronic hepatitis B and C.

Circulating microRNAs have been widely recognized as a novel category of biomarker in a variety of physiological and pathological conditions. Other reports revealed that fragments of organ specific messenger RNAs are also detectable in serum/plasma and can be utilized as sensitive indicators of liver pathology and cancer. In order to assess the sensitivity and reliability of these two class of RNAs as marker of hepatitis B or C induced chronic liver disease, we collected plasma samples from 156 chronic hepatitis B or C patients (HBV active n = 112, HBV carrier n = 19, hepatitis C n = 25) and 22 healthy donors and quantified their circulating mRNA for albumin, HP (haptoglobin), CYP2E1 (cytochrome P450, family 2, subfamily E) and ApoA2 (Apolipoprotein A2) in conjunction with microRNA-122, a well established marker for acute and chronic liver injury. We found that plasma microRNA-122 level is significantly elevated in patients with active HBV but not in HBV carriers. Furthermore, microRNA-122 is not elevated in HCV patients even though their median serum alanine aminotransferase (sALT) was three fold of the healthy donors. Nevertheless, circulating mRNAs, especially albumin mRNA, showed much more sensitivity in distinguishing active hepatitis B, hepatitis B carrier or HCV patients from healthy control. Correlation and multiple linear regression analysis suggested that circulating mRNAs and miRNAs are much more related to HBsAg titre than to sALT. Immunoprecipitation of HBsAg in HBV patients' plasma resulted in enrichment of albumin and HP mRNA suggesting that fragments of liver specific transcripts can be encapsidated into HBsAg particles. Taken together, our results suggest that hepatocyte specific transcripts in plasma like albumin mRNA showed greater sensitivity and specificity in differentiating HBV or HCV induced chronic liver disease than microRNA-122. Circulating mRNA fragments merit more attention in the quest of next generation biomarkers for various maladies.

A random PCR screening system for the identification of type 1 human herpes simplex virus.

Several viral diseases exhibit measles-like symptoms. Differentiation of suspected cases of measles with molecular epidemiological techniques in the laboratory is useful for measles surveillance. In this study, a random PCR screening system was undertaken for the identification of isolates from patients with measles-like symptoms who exhibited cytopathic effects, but who had negative results for measles virus-specific reverse transcription (RT)-PCR and indirect immunofluorescence assays. Sequence analysis of random amplified PCR products showed that they were highly homologous to type 1 human herpes simplex virus (HSV-1). The results were further confirmed by an HSV-1-specific TaqMan real-time PCR assay. The random PCR screening system described in this study provides an efficient procedure for the identification of unknown viral pathogens. Measles-like symptoms can also be caused by HSV-1, suggesting the need to include HSV-1 in differential diagnoses of measles-like diseases.