Jinhai Huang

Diet Effects in Gut Microbiome and Obesity

The 100 trillion microbes in human gut coevolve with the host and exert significant influences on human health. The gut microbial composition presents dynamic changes correlated with various factors including host genotypes, age, and external environment. Effective manipulation of the gut microbiota through diets (both long-term and short-term diet patterns), probiotics and/or prebiotics, and antibiotics has been proved being potential to prevent from metabolic disorders such as obesity in many studies. The dietary regulation exerts influences on microbial metabolism and host immune functions through several pathways, of which may include selectively bacterial fermentation of nutrients, lower intestinal barrier function, overexpression of genes associated with disorders, and disruptions to both innate and adaptive immunity. Discoveries in the interrelationship between diet, intestinal microbiome, and body immune system provide us novel perceptions to the specific action mechanisms and will promote the development of therapeutic approaches for obesity.

Human FasL Gene Is a Target of β-Catenin/T-Cell Factor Pathway and Complex FasL Haplotypes Alter Promoter Functions

FasL expression on human immune cells and cancer cells plays important roles in immune homeostasis and in cancer development. Our previous study suggests that polymorphisms in the FasL promoter can significantly affect the gene expression in human cells. In addition to the functional FasL SNP -844C>T (rs763110), three other SNPs (SNP -756A>G or rs2021837, SNP -478A>T or rs41309790, and SNP -205 C>G or rs74124371) exist in the proximal FasL promoter. In the current study, we established three major FasL hyplotypes in humans. Interestingly, a transcription motif search revealed that the FasL promoter possessed two consensus T-cell factor (TCF/LEF1) binding elements (TBEs), which is either polymorphic (SNP -205C>G) or close to the functional SNP -844C>T. Subsequently, we demonstrate that both FasL TBEs formed complexes with the TCF-4 and β-catenin transcription factors in vitro and in vivo. Co-transfection of LEF-1 and β-catenin transcription factors significantly increased FasL promoter activities, suggesting that FasL is a target gene of the β-catenin/T-cell factor pathway. More importantly, we found that the rare allele (-205G) of the polymorphic FasL TBE (SNP -205C>G) failed to bind the TCF-4 transcription factor and that SNP -205 C>G significantly affected the promoter activity. Furthermore, promoter reporter assays revealed that FasL SNP haplotypes influenced promoter activities in human colon cancer cells and in human T cells. Finally, β-catenin knockdown significantly decreased the FasL expression in human SW480 colon cancer cells. Collectively, our data suggest that β-catenin may be involved in FasL gene regulation and that FasL expression is influenced by FasL SNP haplotypes, which may have significant implications in immune response and tumorigenesis.

Development of a loop-mediated isothermal amplification method for rapid detection of caprine arthritis-encephalitis virus proviral DNA.

A rapid detection assay based on loop-mediated isothermal amplification (LAMP) has been developed for detecting caprine arthritis-encephalitis (CAEV) proviral DNA. The LAMP assay utilized a set of five primers designed against highly conserved sequences located within the p25 gene region. The assay successfully detected CAEV proviral DNA in total DNA extracts originating from cell culture, whole blood samples and separated PBMCs. There was no cross-reaction with the negative control. Amplification was monitored using a Loopamp real-time turbidimeter; turbidity and the corresponding time were recorded. Amplification from CAEV-Shanxi DNA was detected as early as 17xa0min, with a maximum sensitivity of 0.0001 TCID50, reached at 32xa0min. Sixty-eight animal blood samples were tested using AGID, PCR and LAMP assay, and the positive rates were 30.9xa0%, 33.8xa0% and 47.1xa0%, respectively. Whole blood can be used directly, eliminating the need for separation of PBMCs and nucleic acid extraction, reducing the overall procedure time to approximately 80xa0min. Therefore, the LAMP assay provides a specific and sensitive means for detecting CAEV proviral DNA in a simple, fast, and cost-effective manner and should be useful in eradication programs and epidemiological studies. Furthermore, the LAMP assay can be performed in less-well-equipped laboratories as well as in the field.

Functional Fcgamma Receptor Polymorphisms Are Associated with Human Allergy

Objective IgG Fc receptors (FcγRs) play important roles in immune responses. It is not clear whether FcγR receptors play a role in human asthma and allergy. The aim of current study was to investigate whether functional single nucleotide polymorphisms (SNPs) of FcγR genes (FCGR) are associated with human asthma and allergy. Methods Functional SNPs of FCGR2A (FcγRIIA-131His>Arg, rs1801274), FCGR2B (FcγRIIB-187Ile>Thr, rs1050501), FCGR2C (FcγRIIC-13Gln>Stop, rs10917661), FCGR3A (FcγRIIIA-158Val>Phe, rs396991), and FCGR3B variants (FcγRIIIB NA1 and NA2) were genotyped in an asthma family cohort including 370 atopy positive, 239 atopy negative, and 169 asthma positive subjects. The genotype and phenotype data (asthma, bronchial hyper-responsiveness, and atopy) of subjects were analyzed using family-based association tests (FBAT) and logistic regression adjusted for age and sex. Result The FcγRIIA-131His>Arg SNP is significantly associated with atopy in a family-based association test (Pu200a=u200a0.00287) and in a logistic regression analysis (Pu200a=u200a0.0269, OR 0.732, 95% CI: 0.555–0.965). The FcγRIIA-131His (or rs1801274-A) allele capable of binding human IgG2 has a protective role against atopy. In addition, the rare FcγRIIB-187Thr (or rs1050501-C) allele defective for the receptor-mediated inhibitory signals is a risk factor for atopy (Pu200a=u200a0.0031, OR 1.758, 95% CI: 1.209–2.556) and IgE production (P<0.001). However, variants of activating FcγRIIIA (rs396991), and FcγRIIIB (NA1 and NA2), and FcγRIIC (rs10917661) are not associated with asthma, BHR, and atopy (P>0.05). Conclusions FcγRIIA and FcγRIIB functional polymorphisms may have a role in the pathogenesis of allergy.

Evolutionary, epidemiological, demographical, and geographical dissection of porcine bocavirus in China and America

Porcine bocavirus was first discovered in Swedish pigs with post-weaning multisystemic wasting syndrome (PMWS) in 2009. Many efforts have been implemented to investigate the porcine bocavirus, but it remains enigmatic. In the current study, we utilized data from both China and the USA. The China-derived data included 403 pig samples collected from five provinces, 122 gene sequences from the GenBank database, and 637 old porcine bocavirus (PBoV) cases. The USA-derived data comprised 181 pig samples from 18 states, 39 new gene sequences, and 85 new emerging cases. First, we executed a comprehensive analysis of the diseases prevalence, phylogenetics, evolutionary distances, mutation network, geographical distribution, occurrence frequency, and phylogeographical estimation in both China and the USA. The results showed that the positive rates of PBoV (42.0%, 76/181) in American samples were significantly higher than those (11.4%, 46/403) in the Chinese samples. All PBoV cases from these countries can be divided into six groups: PBoV1 (group 1), PBoV2 (group 2), PBoV3C (group 3), PBoV5 (group 4), PBoV3/4 (group 5), and PBoV6V7V (group 6). PBoV1 and PBoV2 were epidemic strains from 2006 to 2011 in China, whereas the PBoV3 subtypes were epidemic from 2010 to 2012 in China and the USA. At present, PBoV3C (group 3), PBoV5 (group 4), and PBoV3/4 (group 5) are epidemic viruses and co-exist in China and the USA. The geographical distribution of PBoV mainly lies in the east and south coastal areas of China and the central states of the USA. Jiangsu Province and the state of Minnesota were the centers of high occurrence frequency of PBoV with six outbreaks. The old PBoV cases involved 14 provinces and regions of China and North Carolina in the USA, whereas the new emerging cases involved five provinces in China and 13 states in the USA, of which two provinces and 12 states reported for the first time that piglets were infected by PBoV. Hong Kong, Hebei, and Jiangsu Provinces and the states of Minnesota and North Carolina were possibly geographical origins of PBoV in China and America, respectively. These data can help us systematically understand porcine bocavirus in China and America and find effective strategies for its treatment.

Detection and genetic analysis of porcine bocavirus in different swine herds in North Central China.

Porcine Bocavirus (PBoV) has been reported to be associated with postweaning multisystemic wasting syndrome and pneumonia in pigs. In this study, a survey was conducted to evaluate the prevalence of PBoV in slaughter pigs, sick pigs, asymptomatic pigs and classical swine fever virus (CSFV) eradication plan herds in five provinces of China (Henan, Liaoning, Shandong, Hebei and Tianjin) by means of PCR targeting NS1 gene of PBoV. Among the total of 403 tissue samples, 11.41% were positive for PBoV. The positive rates of spleen (20.75%) and inguinal lymph node (27.18%) are higher than those of other organs. PCR products of twenty PBoV positive samples from slaughter pigs were sequenced for phylogenetic analysis. The result revealed that PBoV could be divided into 6 groups (PBoV-a~PBoV-f). All PBoV sequenced in this study belong to PBoV-a–PBoV-d with 90.1% to 99% nucleotide identities. Our results exhibited significant genetic diversity of PBoV and suggested a complex prevalence of PBoV in Chinese swine herds. Whether this diversity of PBoV has a significance to pig production or even public health remains to be further studied.

Detection and characterization of porcine bocavirus in the United States.

AbstractnWe screened pigs (nxa0=xa0203) presenting with respiratory illness or diarrhea for porcine bocavirus (PBoV); 88 (43.30xa0%) were positive by PCR. More positives were seen in diarrhea cases (48.7xa0%) than in respiratory cases (29.1xa0%). Based on phylogenetic analysis of 540 nucleotides of the NS1 gene, the viruses could be divided into four possible groups. Group IV sequences did not match any GenBank sequences, while groups I, II and III gave matches with PBoV3, PBoV4 and PBoV5, respectively. The wide range (70xa0% to 100xa0%) of nucleotide (nt) sequence identity among strains in this study indicates high genetic diversity among porcine bocaviruses.

Production of medium-chain volatile flavour esters in Pichia pastoris whole-cell biocatalysts with extracellular expression of Saccharomyces cerevisiae acyl-CoA:ethanol O-acyltransferase Eht1 or Eeb1

Medium-chain volatile flavour esters are important molecules since they have extensive applications in food, fragrance, cosmetic, paint and coating industries, which determine different characteristics of aroma or taste in commercial products. Biosynthesis of these compounds by alcoholysis is catalyzed by acyl-CoA:ethanol O-acyltransferases Eht1 or Eeb1 in Saccharomyces cerevisiae. In this study, these two yeast enzymes were selected to explore their preparations as the form of whole cell biocatalysts for the production of volatile flavour esters. Here, the novel whole cell biocatalysts Pichia pastoris yeasts with functional extracellular expression of Eht1 or Eeb1 were constructed. Flavour production was established through an integrated process with coupled enzyme formation and ester biosynthesis in the recombinant yeasts in one pot, leading to the formation of volatile C6–C14 methyl and ethyl esters from wort medium. Interestingly, there is no significant difference between P. pastoris-EHT1 and P. pastoris-EEB1 in substrate preference during flavour biosynthesis, indicating a similar role of Eht1 and Eeb1 in P. pastoris cells, in contradiction with previous findings in S. cerevisiae to some extent. Consequently the study not only provides a greater understanding of these two enzymes in a heterogeneous host, but also demonstrated the positive effect of the recombinant Eht1 and Eeb1 in ester formation by P. pastoris live cells, potentially paving the way towards achieving efficient production of volatile flavour by an integrated biocatalytic system composed of recombinant enzyme production and flavour biosynthesis.

Development of TaqMan-based qPCR method for detection of caprine arthritis-encephalitis virus (CAEV) infection.

A specific and sensitive two-step TaqMan real-time PCR has been developed for rapid diagnosis of caprine arthritis-encephalitis virus (CAEV) infection by using a set of specific primers and a TaqMan probe targeting a highly conserved region within the gene encoding the viral capsid protein (CA). The assay successfully detected CAEV proviral DNA in total DNA extracts originating from cell culture, whole blood samples and isolated PBMCs, with a lower detection limit of 102 copies and a linear dynamic range of 105 to 1010 copies/ml. There was no cross-reaction with other animal viruses (e.g., goat pox virus, bovine leukemia virus, bovine mucosal disease virus, swine influenza virus and Nipah virus). When applied in parallel with serological AGID and conventional PCR for detection of CAEV in field samples, this assay exhibited a higher sensitivity than these traditional methods, and 7.8xa0% of the 308 specimens collected in the Shanxi and Tianjin regions of China from 1993 to 2011 were found to be positive. Thus, the TaqMan qPCR assay provides a fast, specific and sensitive means for detecting CAEV proviral DNA in goat specimens and should be useful for large-scale detection in eradication programs and epidemiological studies.

Notice of Retraction Analysis of aroma compounds of different yeast strains & its molecular identification by bioinformatics

Sequence analysis of D1/D2 region of 26S rDNA was used for identification of different yeasts isolated from food-borne material. Then, their aromas compounds were detected and analyzed by SPME-GC-MS, another biology information technology. 12 yeast strains were grouped in 7 species belonging to 5 genera as follows: Saccharomyces cerevisiae, Torulaspora delbrueckii, Kluyveromyces marxianus, Pichia cactophila, Clavispora lusitaniae, Pichia membranifaciens, and Pichia kudriavzevii. Among all yeast strains, most of volatiles were similar, but contents were affected by different strains. Strain A1 (Saccharomyces cerevisiae) produced relatively higher aromas compounds than S6 (Kluyveromyces marxianus). The result of GC-MS showed ethyl octanoate and phenylethyl acetate were high-impact flavors by strain A1 and S6, respectively. In conclusion, it is a reliable identification system to indentify food-borne yeasts at species level using 26S rDNA sequencing and phenotypic studies. SPME-GC/MS analysis highlights aroma compounds can be used for pre-screening potential edible yeast strains effectively.