Rapid Antimicrobial Susceptibility Test of Helicobacter pylori to Metronidazole via Single-Cell Raman Spectrometry.

BACKGROUND: Metronidazole is a first-line antibiotic to treat Helicobacter pylori infections. However, the Clinical Laboratory Standards Institute guidelines recommend against using antimicrobial susceptibility test (AST) to test metronidazole resistance, due to the unreliable predictive power which can result in treatment failure. OBJECTIVES: The aim of this study was to establish an 8-h, metabolic-phenotype based AST for H. pylori metronidazole susceptibility using D2O-probed Raman microspectroscopy. METHODS: Minimal inhibitory concentration (MIC) measured by conventional AST (E-test) were compared with expedited MIC via metabolic activity (eMIC-MA) for 10 H. pylori isolates. Raman barcodes of cellular-response to stress (RBCS) incorporating protein and carbohydrate Raman bands, were utilized to identify a biomarker to distinguish metronidazole susceptibility. RESULTS: Specifically, eMIC-MA produces metronidazole susceptibility results showing 100% agreement with E-test, and determines the bactericidal dosage for both high- and low-level resistant H. pylori strains. In addition, RBCS not just reliably distinguish between metronidazole-susceptible and -resistant strains, but reveal their distinct mechanisms in bacterial responses to metronidazole. CONCLUSION: The speed, accuracy, low cost, and rich information content that reveals the mode-of-action of drugs suggest the method's value in guiding metronidazole prescriptions for H. pylori eradication and in rapid screening based on drug-resistance mechanism.

A novel strategy for improving bowel preparation based on social software-enhanced education: A prospective, multicenter, randomized controlled study.

BACKGROUND AND AIM: The compliance and timeliness of oral laxatives have always been the key factors restricting bowel preparation (BP). We have constructed a novel enhanced-educational content and process based on social software (SS) for BP to optimize these issues. METHODS: A multicenter, prospective, randomized controlled study was conducted at 13 hospitals in China from December 2019 to December 2020. A total of 1774 enrollees received standard instructions for BP and were randomly assigned (1:1) to the SS group (SSG) that received a smartphone-based enhanced-education strategy starting 4 h before colonoscopy or the control group (CG). RESULTS: A total of 3034 consecutive outpatient colonoscopy patients were assessed for eligibility, and 1774 were enrolled and randomly assigned. Ultimately, data from 1747 (SSG vs CG: 875 vs 872) enrollees were collected. The BP adequacy rate was 92.22% (95% CI: 90.46-93.98) in the SSG vs 88.05% (95% CI: 85.91-90.18) in the CG (P = 0.005), and the total Boston Bowel Preparation Scale scores (6.89 ± 1.15 vs 6.67 ± 1.15, P < 0.001) of those in the SSG were significantly higher than those in the CG. The average number of polyps detected in the SSG was considerably higher than that in the CG (0.84 ± 2.00 vs 0.53 ± 1.19, P = 0.037), and the average diameter of the polyps was significantly lower than that of the control group (4.0 ± 2.5 vs 4.9 ± 3.7, P < 0.001). CONCLUSIONS: This SS-enhanced education strategy can improve the BP adequacy rate and increase the average number of polyps detected, especially those of small diameter.

Homochiral Metallacycle Used as a Stationary Phase for Capillary Gas Chromatographic Separation of Chiral and Achiral Compounds.

Metallacycles are a novel class of supramolecular materials with circular structures, internal cavities, and abundant host-guest chemical properties that have exhibited good application prospects in many fields. However, to the best of our knowledge, no research on the use of metallacycles as stationary phases for gas chromatographic (GC) separations has been published yet. In this work, we report for the first time the use of a homochiral metallacycle, [ZnCl2L]2, as a stationary phase for GC separations. [ZnCl2L]2 was synthesized by reaction of (S)-(1-isonicotinoylpyrrolidin-2-yl)methyl-isonicotinate (L) with ZnCl2 via coordination-driven self-assembly. The [ZnCl2L]2-coated column displayed an excellent separation performance not only of organic isomers but also of racemic compounds. Sixteen racemates (including alcohols, esters, amino acid derivatives, ethers, organic acids, and epoxides) and 21 isomeric compounds (including positional, structural, and cis/trans-isomers) were well separated on the [ZnCl2L]2-coated column. Impressively, some racemates were resolved with high resolution values (Rs), including 1,2-butanediol diacetate (Rs = 25.86), ethyl 3-hydroxybutyrate (Rs = 20.97), 1,3-butanediol diacetate (Rs = 18.09), and threonine derivative (Rs = 18.61). Compared with the commercial ß-DEX 120 column for separation of the tested racemates, the [ZnCl2L]2-coated column exhibited good enantioseparation complementarity, enabling separation of some racemates that could not be separated, or were not well resolved, by the ß-DEX 120 column. In addition, many organic mixtures, such as n-alkanes, alkylbenzenes, n-alcohols, and a Grob test mixture, were also well separated on the [ZnCl2L]2-coated column. The column also has good reproducibility and thermal stability on separation. This work not only reveals the great potential of metallacycles for GC separations but also opens up a new application of metallacycles in separation science.

DNA starvation/stationary phase protection protein of Helicobacter pylori as a potential immunodominant antigen for infection detection.

BACKGROUND: Application of chicken egg yolk immunoglobulin Y (IgY) for Helicobacter pylori (H. pylori, HP) has gained much interest in recent years. Comparing with for treatment, IgY may be more advantageous when used for H. pylori detection. METHODS: Nine strains of H. pylori with different genetic backgrounds were inactivated and used to immunize hens, respectively, for the preparation of polyclonal anti-H. pylori immunoglobulin Y (anti-HP IgY). The proteins of H. pylori with reactivity to anti-HP IgY were detected by Western Blot. The five protein bands that can be well recognized by anti-HP IgY of each group, and were prevalent in all nine strains were excised from SDS-PAGE gel, digested and identified by Nano-HPLC-MS/MS analysis. The potential of these identified proteins as antigen detection targets was then assessed by sequence analysis. RESULTS: Anti-HP IgY derived from each group of specific strain immunized hens can recognize self-strain and non-self-strain antigens well. Five immunodominant antigens were identified as chaperonin GroEL, flagellin A, urease subunit alpha, peroxiredoxin and DNA starvation/stationary phase protection protein. Sequences analysis showed that both peroxiredoxin and DNA starvation/stationary phase protection protein were present in all 1000 strains of H. pylori queried, and the amino acid sequences were highly conserved. The highest sequence consistency between the DNA starvation/stationary phase protection protein of H. pylori and non-Helicobacter organisms was 52.59%, and the consistent sites were scattered and there was no continuous long fragment consensus sequence. CONCLUSION: DNA starvation/stationary phase protection protein was identified as an immunodominant antigen of H. pylori and sequence analysis indicated that it could serve as a potential antigen target for the diagnosis of H. pylori infection.

A novel pillar[3]trianglimine macrocycle with a deep cavity used as a chiral selector to prepare a chiral stationary phase by thiol-ene click reaction for enantioseparation in high-performance liquid chromatography.

A chiral pillar[3]trianglimine (C60 H72 N6 O6 ) with a deep cavity has been developed as a chiral selector and bonded to thiolated silica by thiol-ene click reaction to fabricate a novel chiral stationary phase for enantioseparation in high-performance liquid chromatography. The enantioseparation performance of the fabricated chiral stationary phase has been evaluated by separating various racemic compounds, including alcohols, esters, amines, ketones, amino acids, and epoxides, in both normal-phase and reversed-phase elution modes. In total, 14 and 17 racemates have been effectively separated in these two separation modes, respectively. In comparison with two widely used chiral columns (Chiralcel OD-H and Chiralpak AD-H), our novel chiral stationary phase offered good chiral separation complementarity, separating some of the tested racemates that could not be separated or were only partially separated on these two commercial columns. The influences of analyte mass, mobile phase composition, and column temperature on chiral separation have been investigated. Good repeatability, stability, and column-to-column reproducibility of the chiral stationary phase for enantioseparation have been observed. After the fabricated column had been eluted up to 400 times, the relative standard deviations (n = 5) of resolution (Rs) and retention time of the separated analytes were < 0.39% and < 0.20%, respectively. The relative standard deviations (n = 3) of Rs and retention time for column-to-column reproducibility were < 4.6% and < 5.2%, respectively. This study demonstrated that the new chiral stationary phase has great prospects for chiral separation in high-performance liquid chromatography.

Preparation of Chiral Porous Organic Cage Clicked Chiral Stationary Phase for HPLC Enantioseparation.

Porous organic cages (POCs) are a new subclass of porous materials, which are constructed from discrete cage molecules with permanent cavities via weak intermolecular forces. In this study, a novel chiral stationary phase (CSP) has been prepared by chemically binding a [4 + 6]-type chiral POC (C120H96N12O4) with thiol-functionalized silica gel using a thiol-ene click reaction and applied to HPLC separations. The column packed with this CSP presented good separation capability for chiral compounds and positional isomers. Thirteen racemates have been enantioseparated on this column, including alcohols, diols, ketones, amines, epoxides, and organic acids. Upon comparison with a previously reported chiral POC NC1-R-based column, commercial Chiralpak AD-H, and Chiralcel OD-H columns, this column is complementary to these three columns in terms of its enantiomeric separation; and can also separate some racemic compounds that cannot be separated by the three columns. In addition, eight positional isomers (iodoaniline, bromoaniline, chloroaniline, dibromobenzene, dichlorobenzene, toluidine, nitrobromobenzene, and nitroaniline) have also been separated. The influences of the injection weight and column temperature on separation have been explored. After the column has undergone multiple injections, the relative standard deviations (RSDs) for the retention time and selectivity were below 1.0 and 1.5%, respectively, indicating the good reproducibility and stability of the column for separation. This work demonstrates that POCs are promising materials for HPLC separation.

Comparative transcriptome analysis to investigate the mechanism of anti-Helicobacter pylori activity of zinc.

As a potential anti-Helicobacter pylori agent, zinc causes impairment of Helicobacter pylori growth, and this property of zinc is of broad interest to biological investigators. However, little is known about the molecular mechanisms by which zinc inhibits the growth of Helicobacter pylori. Here, an in vitro experiment revealed that zinc at specific concentrations inhibits Helicobacter pylori growth. Furthermore, an RNA sequencing-based investigation of the global regulatory response to zinc revealed that exposure to zinc altered the Helicobacter pylori transcriptional profile in numerous ways. A high concentration of zinc induced the upregulation of genes related to ribosomal subunit, ribosome biosynthesis, chaperone and adhesins. However, flagellar assembly genes and some type IV secretion system genes were repressed. In addition, the expression levels of some genes that encode transporters of metal ions and that play key roles in Helicobacter pylori pathogenicity were altered under conditions of zinc-induced stress. In summary, high concentrations of zinc initiated antimicrobial activity to Helicobacter pylori under the combined effect of multiple repressed or altered pathogenetic genes and metabolic pathways associated with bacteria growth. This result has significant implications for understanding not only the antimicrobial activity mechanism of zinc but also the role of zinc-mediated homeostasis in Helicobacter pylori.

Inhibition of CEBPB Attenuates Lupus Nephritis via Regulating Pim-1 Signaling.

Systemic lupus erythematosus (SLE) is an autoimmune disease leading to inflammatory damage in multiple target organs, and lupus nephritis (LN) is one of the most life-threatening organ manifestations. CCAAT/enhancer-binding protein ß (CEBPB) regulates the NLRP3 inflammasome and is involved in the pathogenesis of SLE. However, the role and mechanism of CEBPB in LN remains unclear. MRL/lpr mice and lipopolysaccharides (LPS) combined with adenosine triphosphate- (ATP-) treated glomerular podocytes were used as models of LN in vivo and in vitro, respectively. In vivo, we investigated the expressions of CEBPB during the development of MRL/lpr mice. Then we assessed the effect of CEBPB inhibition on renal structure and function through injecting shCEBPB lentivirus into MRL/lpr mice. In vitro, glomerular podocytes were treated with Pim-1-OE and siCEBPB to explore the relation between CEBPB and Pim-1. The progression of LN in mice was associated with the increased level of CEBPB, and the inhibition of CEBPB ameliorated renal structure impairments and improved renal function damage associated with LN. Knockdown of CEBPB could suppress the activation of NLRP3 inflammasome and the secretion of IL-1ß and IL-6. Furthermore, the knockdown of CEBPB could inhibit NLRP3 inflammasome activation and pyroptosis via binding to Pim-1 promoter to downregulate its expression, and the overexpression of Pim-1 reversed the effects of CEBPB deficiency. The regulation of CEBPB on Pim-1 facilitated pyroptosis by activating NLRP3 inflammasome, thereby promoting the development of LN.

A comparison of fluid status determination using bioelectric impedance and the isotope dilution method in hemodialysis and peritoneal dialysis patients.

We aimed to compare fluid status as determined by multifrequency bioimpedance spectroscopy (MF-BIS, Xitron 4200, USA) with that determined by the isotope dilution method among a contemporary Chinese cohort. Healthy Chinese subjects (HS, n = 30) were recruited in Zhengzhou. Hemodialysis (HD, n = 49) and peritoneal dialysis (PD, n = 48) patients were screened at the First Affiliated Hospital of Zhengzhou University. Total body water (TBW) and extracellular water (ECW) were measured by deuterium (TBWD) and bromide (ECWBr) dilution, respectively, and by MF-BIS using the Moissl equation (ME). The results of MF-BIS were compared to the reference method by Pearson analysis and Bland-Altman analysis in the three groups. The accuracy of overhydration as determined by MF-BIS was analyzed by receiver operating characteristic (ROC) curves. The TBWD and TBWME values were 34.67 ± 7.31 and 35.41 ± 5.76 L, 37.30 ± 8.58 and 37.02 ± 8.10 L, and 38.61 ± 10.02 and 38.44 ± 7.59 L in the HS, HD and PD groups, respectively. The ECWBr and ECWME values were 14.88 ± 3.33 and 15.53 ± 2.39 L, 16.24 ± 5.08 and 16.90 ± 3.93 L, and 19.08 ± 6.41 and 18.23 ± 3.61 L in the HS, HD and PD groups, respectively. The mean bias between TBWD and TBWME was -0.74 L, 0.28 L, and 0.17 L in the HS, HD and PD groups, respectively. The mean bias between ECWBr and ECWME was -0.65 L, -0.66 L, and 0.85 L in the HS, HD and PD groups, respectively. Compared to the ECWBr/TBWD ratio, the area under the ROC curve (AUC) of the ECWME/TBWME ratio for the diagnosis of overhydration was 0.76 and 0.68 in the HD and PD groups, respectively. In summary, MF-BIS with ME could be used in Chinese HD and PD patients.

Tailoring Silicon Nitride Surface Chemistry for Facilitating Odontogenic Differentiation of Rat Dental Pulp Cells.

Silicon nitride (Si3N4) can facilitate bone formation; hence, it is used as a biomaterial in orthopedics. Nevertheless, its usability for dentistry is unexplored. The aim of the present study was to investigate the effect of Si3N4 granules for the proliferation and odontogenic differentiation of rat dental pulp cells (rDPCs). Four different types of Si3N4 granules were prepared, which underwent different treatments to form pristine as-synthesized Si3N4, chemically treated Si3N4, thermally treated Si3N4, and Si3N4 sintered with 3 wt.% yttrium oxide (Y2O3). rDPCs were cultured on or around the Si3N4 granular beds. Compared with the other three types of Si3N4 granules, the sintered Si3N4 granules significantly promoted cellular attachment, upregulated the expression of odontogenic marker genes (Dentin Matrix Acidic Phosphoprotein 1 and Dentin Sialophosphoprotein) in the early phase, and enhanced the formation of mineralization nodules. Furthermore, the water contact angle of sintered Si3N4 was also greatly increased to 40°. These results suggest that the sintering process for Si3N4 with Y2O3 positively altered the surface properties of pristine as-synthesized Si3N4 granules, thereby facilitating the odontogenic differentiation of rDPCs. Thus, the introduction of a sintering treatment for Si3N4 granules is likely to facilitate their use in the clinical application of dentistry.

IL-6 promotes epithelial-to-mesenchymal transition of human peritoneal mesothelial cells possibly through the JAK2/STAT3 signaling pathway.

Long-term peritoneal dialysis (PD) therapy results in functional and structural alteration of the peritoneal membrane, including epithelial-to-mesenchymal transition (EMT). Interleukin 6 (IL-6) is a local pleiotropic cytokine, hypothesized to play an important role in EMT. This study was designed to investigate the role of IL-6 in EMT and peritoneal membrane dysfunction in long-term PD patients by assessing the level of IL-6 in dialysate and exploring the relationship between IL-6, the related signaling pathway JAK2/STAT3, and EMT, using in vitro cellular and molecular techniques. Plasma and dialysate levels of IL-6 were significantly higher in PD ultrafiltration failure patients compared with patients without ultrafiltration failure and were negatively correlated with measures of PD adequacy. In vitro IL-6 treatment changed human peritoneal mesothelial cell phenotype from a typical cobblestone-like to a fibroblast-like appearance and increased cell viability. IL-6 treatment increased α-smooth muscle actin and vascular endothelial growth factor expression but decreased E-cadherin expression. IL-6 treatment activated the JAK/STAT signaling pathway. However, the JAK2/STAT3 inhibitor WP1066 prevented IL-6-induced activation of the JAK2/STAT3 pathway and EMT. We conclude that IL-6 promotes the EMT process, possibly by activating the JAK2/STAT3 signaling pathway. IL-6 may serve as a novel therapeutic target for preventing EMT, and preservation of the peritoneal membrane may arise from these studies.

Gastric Juice-Based Real-Time PCR for Tailored Helicobacter Pylori Treatment: A Practical Approach.

A gastric juice-based real-time polymerase chain reaction (PCR) assay was established to identify Helicobacter pylori infection, clarithromycin susceptibility and human CYP2C19 genotypes and to guide the choice of proton pump inhibitor (PPI), clarithromycin and amoxicillin treatment for tailored H. pylori eradication therapy. From January 2013 to November 2014, 178 consecutive dyspeptic patients were enrolled for collection of gastric biopsy samples and gastric juice by endoscopy at the Peking University Third Hospital; 105 and 73 H. pylori-positive and -negative patients, respectively, were included in this study. H. pylori infection was defined as samples with both a strongly positive rapid urease test (RUT) and positive H. pylori histology. A series of primers and probes were distributed into four reactions for identifying the H. pylori cagH gene coupled with an internal control (Rnase P gene), A2142G and A2143G mutants of the H. pylori 23S rRNA gene, and single-nucleotide polymorphisms (SNPs) G681A of CYP2C19*2 and G636A of CYP2C19*3. The E-test and DNA sequencing were used to evaluate the H. pylori clarithromycin susceptibility phenotype and genotype. The SNPs CYP2C19*2 and CYP2C19*3 were also evaluated by nucleotide sequencing. The sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of this gastric juice-based real-time PCR assay were evaluated by comparing with the same measures obtained through gastric biopsy-based PCR and culture. The H. pylori diagnostic sensitivities of the culture, PCR, and gastric biopsy- and gastric juice-based real-time PCR assays were 90.48% (95/105), 92.38% (97/105), 97.14% (102/105) and 100% (105/105), respectively; the specificities of the above methods were all 100%. Higher false-negative rates were found among the gastric biopsy samples assessed by culture (10.48%, 11/105), PCR (7.62%, 8/105) and real-time PCR (2.86%, 3/105) than in gastric juice by real-time PCR. Regarding clarithromycin susceptibility, a concordance of 82.98% (78/94) and discordance of 17.02% (16/94) were observed among the different methods, discrepancies that mainly represent differences between the H. pylori clarithromycin susceptibility phenotype and genotype. Three coinfections of susceptible and resistant strains were detected, with resistant-to-susceptible ratios of 1.16, 3.44, and 8.26. The CYP2C19 genotyping results from gastric juice by real-time PCR were completely in accordance with those obtained from biopsy samples by conventional PCR. This gastric juice-based real-time PCR assay is a more accurate method for detecting H. pylori infection, clarithromycin susceptibility and CYP2C19 polymorphisms. The method may be employed to inform the choice of proton pump inhibitor (PPI), clarithromycin and amoxicillin treatment for tailored H. pylori eradication therapy.

Motor Function Evaluation of Hemiplegic Upper-Extremities Using Data Fusion from Wearable Inertial and Surface EMG Sensors.

Quantitative evaluation of motor function is of great demand for monitoring clinical outcome of applied interventions and further guiding the establishment of therapeutic protocol. This study proposes a novel framework for evaluating upper limb motor function based on data fusion from inertial measurement units (IMUs) and surface electromyography (EMG) sensors. With wearable sensors worn on the tested upper limbs, subjects were asked to perform eleven straightforward, specifically designed canonical upper-limb functional tasks. A series of machine learning algorithms were applied to the recorded motion data to produce evaluation indicators, which is able to reflect the level of upper-limb motor function abnormality. Sixteen healthy subjects and eighteen stroke subjects with substantial hemiparesis were recruited in the experiment. The combined IMU and EMG data yielded superior performance over the IMU data alone and the EMG data alone, in terms of decreased normal data variation rate (NDVR) and improved determination coefficient (DC) from a regression analysis between the derived indicator and routine clinical assessment score. Three common unsupervised learning algorithms achieved comparable performance with NDVR around 10% and strong DC around 0.85. By contrast, the use of a supervised algorithm was able to dramatically decrease the NDVR to 6.55%. With the proposed framework, all the produced indicators demonstrated high agreement with the routine clinical assessment scale, indicating their capability of assessing upper-limb motor functions. This study offers a feasible solution to motor function assessment in an objective and quantitative manner, especially suitable for home and community use.

Progressive genomic convergence of two Helicobacter pylori strains during mixed infection of a patient with chronic gastritis.

OBJECTIVE: To study the detailed nature of genomic microevolution during mixed infection with multiple Helicobacter pylori strains in an individual. DESIGN: We sampled 18 isolates from a single biopsy from a patient with chronic gastritis and nephritis. Whole-genome sequencing was applied to these isolates, and statistical genetic tools were used to investigate their evolutionary history. RESULTS: The genomes fall into two clades, reflecting colonisation of the stomach by two distinct strains, and these lineages have accumulated diversity during an estimated 2.8 and 4.2 years of evolution. We detected about 150 clear recombination events between the two clades. Recombination between the lineages is a continuous ongoing process and was detected on both clades, but the effect of recombination in one clade was nearly an order of magnitude higher than in the other. Imputed ancestral sequences also showed evidence of recombination between the two strains prior to their diversification, and we estimate that they have both been infecting the same host for at least 12 years. Recombination tracts between the lineages were, on average, 895 bp in length, and showed evidence for the interspersion of recipient sequences that has been observed in in vitro experiments. The complex evolutionary history of a phage-related protein provided evidence for frequent reinfection of both clades by a single phage lineage during the past 4 years. CONCLUSIONS: Whole genome sequencing can be used to make detailed conclusions about the mechanisms of genetic change of H. pylori based on sampling bacteria from a single gastric biopsy.

Prevalence of Coinfection with Gastric Non-Helicobacter pylori Helicobacter (NHPH) Species in Helicobacter pylori-infected Patients Suffering from Gastric Disease in Beijing, China.

BACKGROUND: The Helicobacter heilmannii sensu lato (H. heilmannii s.l.) group consists of long, spiral-shaped bacteria naturally colonizing the stomach of animals. Moreover, bacteria belonging to this group have been observed in 0.2-6% of human gastric biopsy specimens, and associations have been made with the development of chronic gastritis, peptic ulceration, and gastric MALT lymphoma in humans. MATERIALS AND METHODS: To gain insight into the prevalence of H. heilmannii s.l. infections in patients suffering from gastric disease in China, H. heilmannii s.l. species-specific PCRs were performed on DNA extracts from rapid urease test (RUT)-positive gastric biopsies from 1517 patients followed by nucleotide sequencing. At the same time, Helicobacter pylori cultivation and specific PCR was performed to assess H. pylori infection in these patients. RESULTS: In total, H. heilmannii s.l. infection was detected in 11.87% (178/1499) of H. pylori-positive patients. The prevalence of H. suis, H. felis, H. bizzozeronii, H. heilmannii sensu stricto (s.s.), and H. salomonis in the patients was 6.94%, 2.20%, 0.13%, 0.07%, and 2.54%, respectively. Results revealed that all patients with H. heilmannii s.l. infection were co-infected with H. pylori, and some patients were co-infected with more than two different Helicobacter species. CONCLUSIONS: Helicobacter heilmannii s.l. infections are fairly common in Chinese patients. This should be kept in mind when diagnosing the cause of gastric pathologies in patients. Helicobacter suis was shown to be by far the most prevalent H. heilmannii s.l.species.

The distribution of jhp0940, jhp0945, jhp0947, jhp0949 and jhp0951 genes of Helicobacter pylori in China.

BACKGROUND: The plasticity region of Helicobacter pylori (H. pylori) is a large chromosomal segment containing strain-specific genes. The prevalence of the plasticity region genes of the H. pylori strains in China remains unknown. The aim of this study was to examine the status of these genes and to assess the relationship between the genes and the diseases caused by H. pylori infection. METHODS: A total of 141 strains were isolated from patients with chronic active gastritis (CAG), peptic ulcer disease (PUD) and gastric carcinoma (GC). The prevalence of jhp0940, jhp0945, jhp0947, jhp0949 and jhp0951 was determined using PCR, and the results were analyzed using the chi-squared test. RESULTS: The prevalence rates of jhp0940, jhp0945, jhp0947, jhp0949 and jhp0951 in the H. pylori strains were 42.55, 51.06, 20.57, 56.03 and 63.12%, respectively. The prevalence rates of jhp0940 were similar in the isolates from the CAG, PUD and GC patients, and there was no association between the jhp0940 status and any of the diseases. In contrast, the prevalence rates of jhp0945, jhp0947, jhp0949 and jhp0951 were significantly higher in the PUD and GC isolates than in the CAG isolates (p < 0.01). A univariate analysis showed that jhp0945, jhp0947, jhp0949 and jhp0951 increased the risk of PUD, while only jhp0951 was significantly associated with PUD in the multivariate analysis (p = 0.0149). The jhp0945-positive isolates were significantly associated with an increased risk for GC (p = 0.0097). CONCLUSION: The plasticity region genes are widely distributed in Chinese patients, and a high prevalence of these genes occurs in more serious diseases. Therefore, jhp0951 status is an independent factor associated with the development of PUD, and jhp0945 may predict the future development of GC in patients with CAG and is considered to be the best candidate disease marker for H. pylori-related diseases.

Construction and Validation of Chicken Immune scFv Antibody Library against Helicobacter pylori.

Accurate diagnostic techniques and effective therapeutic methods are required to treat H. pylori. The application of chicken single-chain variable fragment (scFv) antibodies may diagnose and treat H. pylori. This study used the phage display technique to construct a chicken-derived immune scFv antibody library against H. pylori. Total RNA was extracted from the spleens of five immunized chickens and reverse transcribed into cDNA. A fragment of scFv was produced by overlap extension PCR and cloned into a pHEN2 phagemid vector. After the package with the M13KO7 helper phage, the recombinant HpaA protein was used as a target antigen to validate the screening ability of our antibody library by bio-panning. The dilution counting results showed that the size of the primary antibody library was estimated to be 1 × 109 cfu/mL. PCR analysis of 47 clones from the library revealed that about 100% of the clones were positive with scFv fragments, and there were no identical sequences, indicating the good diversity of the antibody library. After three rounds of bio-panning, high-affinity antibodies against recombinant HpaA protein were successfully obtained. The selected antibody specifically recognized HpaA protein in nine different H. pylori strains, confirming the screening ability of our library. The chicken immune scFv antibody library against H. pylori was successfully constructed, and the antibody library's screening ability was validated by selecting specific scFv antibodies against recombinant HpaA and clinical strains. It provided a simple and rapid method to obtain antibodies against H. pylori for diagnosis or treatment.

A New Species of the Genus Pseudocalotes (Squamata: Agamidae) from Southwest Yunnan, China.

In this study, a new species of the genus Pseudocalotes is described from Yingjiang County, Dehong Dai and Jingpo Autonomous Prefecture, Yunnan Province, China, based on four female specimens. It can be distinguished from its congeners by the following combination of characters: (1) interoculabials 3 or 4; (2) canthals 5-7; (3) cicrcumorbitals 8-11; (4) 1 scale between rostral and nasal; (5) interparietal 1; (6) superciliaries 4-6; (7) supralabials 6-7, the 1st in contact with the nasal; (8) infralabials 6-8; (9) transverse gular fold and antehumeral fold present; (10) 2-3 enlarged scales between eye and ear; (11) nuchal crest single, consists of 3-5 erected spines; (12) dorsal crest row single, discontinuous and low, located between two keeled, parallel and enlarged scale rows; (13) enlarged postrictals absent; (14) scales around midbody 53-62, dorsal body scales heterogenous in size and shape; (15) midventrals smaller than dorsals; (16) subdigital scales on the 4th finger 20-26, and on the 4th toe 24-29; (17) dorsal background coloration light taupe with four irregular brown patches along the middle of dorsal; (18) inner lips wathet, tongue aurantiacus, throat bluish black. The population from Yingjiang County was nested within a highly supported lineage, formed a sister taxon with P. kakhienensis (SH 97/UFB 100) and according to the p-distance, the new species differed from its congeners by 14.5% to 35.2% for NADH dehydrogenase subunit 2 (ND2) and 15.5% to 25.0% for NADH dehydrogenase subunit 4 (ND4).

A new species of the genus Hebius (Squamata, Natricidae) from Yunnan, China.

A new species of the genus Hebius Thompson, 1913 is described from Yingjiang County, Dehong Dai and Jingpo Autonomous Prefecture, Yunnan Province, China, based on molecular and morphological evidence. It can be distinguished from its congeners by the following set of characters: (1) dorsal scale rows 19-17-17, feebly keeled; (2) ventrals 146-151; (3) nasal complete, nostril in the middle of the nasal; (4) supralabials 9, the fourth to sixth in contact with the eye; (5) infralabials 10-11, the first 5 touching the first pair of chin shields; (6) preoculars 2; (7) postoculars 3; (8) temporals 3, arranged in two rows (1+2); (9) maxillary teeth 31, the last 4 slightly enlarged, without diastema; (10) tail comparatively long, TAL/TL ratio 0.334 in the male; (11) dorsolateral series of irregular orange or ochre yellow blotches, extending from the neck to the posterior part of the tail; and (12) venter pale orange, tips of ventrals with subrectangular black blotches. All Hebius specimens were strongly recovered as monophyletic, in which Hebiustaronensis (Smith, 1940) and Hebiusvenningi (Wall, 1910) were monophyletic as sister to the Yingjiang County specimens. According to the p-distance of cytochrome b, the new species differs from its congeners by 9.7-15.4%.

A new species of the newt genus Hypselotriton (Amphibia, Urodela, Salamandridae) from Jiangxi Province, southeastern China.

A new newt species, Hypselotritonhuanggangensis sp. nov., is described based on nine specimens collected from Huanggangshan Mountains, Yanshan County, Jiangxi, China. Morphologically, the new species is characterized by the combination of nine external characters: (1) obvious black patches with clear boundaries on the whole body; (2) ground color of the dorsal body tan; (3) ground color of venter bright orange; (4) skin rough; (5) vertebral ridge weak; (6) fingers and toes overlapping when forelimb and hindlimb adpressed towards each other along body; (7) postocular orange spot absent; (8) small white warty glands around the eye; (9) two discontinuous longitudinal lines formed by white warty glands from neck to lateral parts of tail. Molecularly, the new species forms an independent clade with strong support in the phylogenetic trees of the genus based on the mitochondrial locus of NADH dehydrogenase subunit 2 (ND2) gene fragments. The new species distinctly differs from H.fudingensis by differences in its body measurements, vertebral ridge, dorsal black patches, and ventral black patches. Furthermore, the new species and H.fudingensis are geographically isolated by a series of high mountain ranges, including the Wuyishan and Jiufengshan Mountains. The number of Hypselotriton species is now 11.