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Vibrio mimicus bacteria have caused sporadic cases and outbreaks of cholera-like diarrhea throughout the world, but the association of lineages with such events is unexplored. Genomic analyses revealed V. mimicus lineages carrying the virulence factors cholera toxin and toxin coregulated pilus, one of which has persisted for decades in China and the United States. [ABSTRACT FROM AUTHOR]

Grass carp reovirus (GCRV) is one of the most serious pathogens threatening grass carp (Ctenopharyngodon idella) production and results in high mortality in China. To obtain a genetically engineered oral vaccine against GCRV, the cholera toxin B subunit (CTB) of Vibrio cholerae was fused to VP7 (CTB-VP7) and transformed into BL21(DE3) for expression. SDS-PAGE and Western blotting showed that the purified rCTB-VP7 fusion protein (rCTB-VP7) was approximately 49.0 kDa. The monomeric nature of rCTB-VP7 through multistage purification showed a binding affinity for GM1, a receptor for biologically active CTB. rCTB-VP7 is not vulnerable to disassembly by SDS but is vulnerable to disassembly by 2-mercaptoethanol. rCTB-VP7 is stable and highly active at room temperature. The binding affinity experiment between rCTB-VP7 and GM1 also confirms the effects of acid and alkalinity in solution on the structure of rCTB-VP7. rCTB-VP7 bound to GM1 with different affinities under different temperatures and pH values. Prokaryotic expression of rCTB-VP7 was characterized by high expression and easy purification and had a strong binding force with GM1 at 37 °C and pH 7.4. Our results suggest that rCTB-VP7 has the potential as an oral vaccine for protection against GCRV in aquaculture. [ABSTRACT FROM AUTHOR]

Abstract Grass carp reovirus (GCRV) is one of the most serious pathogens threatening grass carp (Ctenopharyngodon idellus) production and results in high mortality in China. VP7 from GCRV is involved in viral infection and could be suitable for developing vaccines for the control of GCRV infection. To obtain a genetically engineered vaccine and a plant-based oral vaccine and to evaluate their immune efficacy as an oral vaccine against GCRV, cholera toxin B subunit (CTB) of Vibrio cholerae fused to VP7 (CTB-VP7) was transformed into BL21(DE3) for expression. SDS-PAGE and Western blotting showed that the purified CTB-VP7 fusion protein (rCTB-VP7) was approximately 49.0 kDa. Meanwhile, CTB-VP7 was transformed into rice callus cells by Agrobacterium tumefaciens -mediated gene transformation. CTB-VP7 was integrated into the nuclear genome by PCR, and mRNA transcripts of CTB-VP7 were detected. ELISA and Western blot analyses revealed that the CTB-VP7 fusion protein (CTB-VP7) could be expressed in rice callus lines. The level of expression was determined to be 1.54% ± 0.43 of the total soluble protein. CTB-VP7 showed a binding affinity for monosialoganglioside(GM1), a receptor for CTB. CTB-VP7 showed a higher affinity towards GM1 compared to rCTB-VP7. CTB-VP7 bonded to GM1 with different affinities under different temperatures. Maximum binding of CTB-VP7 to GM1 was reported to occur within 2 h at 37 °C, and approximately half of the binding affinity remained at 25 °C. Our results suggest that CTB-VP7 could be produced in rice calli, increasing the possibility that edible plants can be employed in mucosal vaccines for protection against GCRV in aquaculture. [ABSTRACT FROM AUTHOR]

Objective: To investigate CTX-M genotypes among extended-spectrum β-lactamase-producing Escherichia coli (ESBL-EC) isolated from patients with community-onset and hospital-onset infections in China, their clonality and the distribution of CTX-M variants in different specimens of community-onset and hospital-onset infections. Methods: ESBL-EC isolates were collected from general hospitals from 2011 to 2012 in China. Broth microdilution method antimicrobial susceptibility testing of 16 antibiotics was performed. Clinical data from community-onset and hospital-onset infections due to ESBL-EC were analyzed. ESBL-encoding genes were amplified by PCR and sequenced, and multilocus sequence typing (MLST) was performed for a random selection of predominant CTX-M type strains identified. Results: A total of 1,168 ESBL-EC isolates were obtained from various clinical specimens, 41.7% of which were responsible for causing community-onset infections. The presence of urinary calculi was higher in community-onset infections, whereas malignancy, cardiovascular and cerebrovascular diseases, dementia, chronic renal disease, diabetes mellitus and surgical treatment were found to have higher proportions in hospital-onset infections. There was no significant difference in trauma between community-onset and hospital-onset infections. 96.2% of the isolates were detected to harbor blaCTX-M genes. blaCTX-M-1 group and blaCTX-M-9 group were detected at 40.7% and 48.7% respectively, and both positive group accounted for 10.6%. blaCTX-M-55 (24.8%) and blaCTX-M-15 (18.2%) were the major genotypes in blaCTX-M-1 group while blaCTX-M-14 (46.8%) was predominant in blaCTX-M-9 group. A comparison of blaCTX-M distribution in different specimens between ESBL-EC causing community-onset and hospital-onset infection showed no significant difference. A total of 229 isolates were tested for MLST. ST131 (14%) was the predominant type. ST648, ST405 and ST1193 were also detected. Conclusions: Community-onset ESBL-EC has emerged as a common pathogen in China. CTX-M-14 is the most commonly encountered, CTX-M-55 and CTX-M-15 have spread rapidly. ST131 is the predominant clonal group, and the great diversity of CTX-M-producing isolates of E. coli has emerged in China. [ABSTRACT FROM AUTHOR]

Background Vibrio cholerae is a human intestinal pathogen and V. cholerae of the O139 serogroups are responsible for the current epidemic cholera in China. In this work, we reported the whole genome sequencing of a V. cholerae O139 strain E306 isolated from a cholera patient in the 306th Hospital of PLA, Beijing, China. Results We obtained the draft genome of V. cholerae O139 strain E306 with a length of 4,161,908 bps and mean G + C content of 47.7%. Phylogenetic analysis indicated that strain E306 was very close to another O139 strain, V. cholerae MO10, which was isolated during the cholera outbreak in India and Bangladesh. However, unlike MO10, strain E306 harbors the El Torspecific RS1 element with no pre-CTX prophage (VSK), very similar to those found in some V. cholerae O1 strains. In addition, strain E306 contains a SXT/R391 family integrative conjugative element (ICE) similar to ICEVchInd4 and SXT MO10, and it carries more antibiotic resistance genes than other closest neighbors. Conclusions The genome sequence of the V. cholerae O139 strain E306 and its comparative analysis with other V. cholerae strains we present here will provide important information for a better understanding of the pathogenicity of V. cholerae and their molecular mechanisms to adapt different environments. [ABSTRACT FROM AUTHOR]

The Clostridium perfringens alpha toxin (CPA), encoded by the plc gene, is the causative pathogen of gas gangrene, which is a lethal infection. In this study, we used an E. coli system for the efficient production of recombinant proteins and developed a bicistronic design (BCD) expression construct consisting of two copies of the C-terminal (247–370) domain of the alpha toxin (CPA-C) in the first cistron, followed by Cholera Toxin B (CTB) linked with another two copies of CPA-C in the second cistron that is controlled by a single promoter. Rabbits were immunized twice with purified proteins (rCPA-C rCTB-CPA-C) produced in the BCD expression system, with an inactivated recombinant E. coli vaccine (RE), C. perfringens formaldehyde-inactivated alpha toxoid (FA-CPA) and C. perfringens l -lysine/formaldehyde alpha toxoid (LF-CPA) vaccines. Following the second vaccination, 0.1 mL of pooled sera of the RE-vaccinated rabbits could neutralize 12× mouse LD 100 (100% lethal dose) of CPA, while that of the rCPA-C rCTB-CPA-C-vaccinated rabbits could neutralize 6× mouse LD 100 of CPA. Antibody titers against CPA were also assessed by ELISA, reaching titers as high as 1:2048000 in the RE group; this was significantly higher compared to the C. perfringens alpha toxoid vaccinated groups (FA-CPA and LF-CPA). Rabbits from all vaccinated groups were completely protected from a 2× rabbit LD 100 of CPA challenge. These results demonstrate that the recombinant proteins are able to induce a strong immune responses, indicating that they may be potentially utilized as targets for novel vaccines specifically against the C. perfringens alpha toxin. • A BCD system for simultaneously expressing rCPA-C(247−370) and CTB-rCPA-C(247−370) is constructed. • The BCD system greatly improves solubility and expression levels of the target proteins. • The antigenic response of expressed products is significantly more protective than the current vaccine in China. • A more stable, high-yielding method with higher biosafety levels to product clostridial antigens is proposed. [ABSTRACT FROM AUTHOR]