BACKGROUND: Salmonella is an important pathogen that causes gastroenteritis and sepsis in humans. Recently, changes in serotype prevalence and an increase in antimicrobial resistance have been reported. This study investigated the distribution of Salmonella serotypes and determined the antimicrobial susceptibility of various strains. METHODS: We collected 113 Salmonella isolates other than Salmonella serotype Typhi from 18 university hospitals in 2015. The serotypes were identified by Salmonella antisera O and H according to the Kauffman White scheme. Antimicrobial susceptibility tests for 12 antibiotics were performed using the disk diffusion method or E-test. RESULTS: We identified 22 serotypes. Serotype group B (44.2%) was the most common, followed by groups C (34.5%) and D (21.2%). Salmonella I 4,[5],12:i:- (23.0%), S. Enteritidis (16.8%), and S. Typhimurium (12.4%) were the most common species. Resistance rates for ampicillin, chloramphenicol, ceftriaxone, and trimethoprim/sulfamethoxazole were 46.9%, 18.5%, 8.8%, and 5.3%, respectively. The intermediate resistance rate to ciprofloxacin was 29.2%. Six isolates were extended-spectrum β-lactamase (ESBL) producers, including 5 bla(CTX-M-15) and 1 bla(CTX-M-55). CONCLUSION: There have been changes in the serotype prevalence and antimicrobial resistance of Salmonella in Korea, with a high prevalence of CTX-M 15-positive strains. Continuous monitoring of Salmonella serotypes and antimicrobial resistance is warranted.
Ampicillin ; Anti-Bacterial Agents ; Ceftriaxone ; Chloramphenicol ; Ciprofloxacin ; Diffusion ; Gastroenteritis ; Hospitals, University ; Humans ; Immune Sera ; Korea ; Methods ; Prevalence ; Salmonella ; Sepsis ; Serogroup ; Serotyping

BACKGROUND: Various pneumococcal vaccines have been evaluated for immunogenicity by opsonophagocytic assay (OPA). A multiplexed OPA (MOPA) for 13 pneumococcal serotypes was developed by Nahm and Burton, and expanded to 26 serotypes in 2012. The development of new conjugate vaccines with increased valence has necessitated expanded MOPAs to include these additional serotypes. In this study, we validated this expanded MOPA platform and applied to measure antibodies against 11 additional serotypes (2, 8, 9N, 10A, 11A, 12F, 15B, 17F, 20B, 22F, and 33F) in human sera. METHODS: All materials, including serum, complement, bacterial master stocks, and HL-60 cells, were evaluated for assay optimization. Following optimization, the assay was validated for accuracy, specificity, and intra- and inter-assay precision with sera from adult donors following standard protocols. The assay was applied to evaluate functional antibodies of 42 sera immunized with 23-valent pneumococcal polysaccharide vaccine (PPV23). RESULTS: The expanded MOPA platform was specific for all serotypes, with the exception of serotype 20. The assay results were highly correlated with those obtained from single-serotype OPA, indicating acceptable accuracy. The coefficients of variation were 7%–24% and 13%–39% in tests of intra- and inter-assay precision, respectively, using three quality-control samples. A MOPA that included 11 additional serotypes in the PPV23 was established and validated with respect to accuracy, specificity, and precision. The opsonic indices of immune sera were obtained using this validated assay. CONCLUSION: The expanded MOPA will be useful for evaluation of the immunogenicity of PPV23 and future conjugate vaccine formulations.
Adult ; Antibodies ; Biological Assay ; Complement System Proteins ; HL-60 Cells ; Humans ; Immune Sera ; Opsonin Proteins ; Pneumococcal Vaccines* ; Sensitivity and Specificity ; Serogroup* ; Tissue Donors ; Vaccines, Conjugate

PURPOSE: There is a need to broaden protective coverage of M protein–based vaccines against group A streptococci (GAS) because coverage of the current 30-valent M protein vaccine does not extend to all emm types. An additional GAS antigen and virulence factor that could potentially extend vaccine coverage is M-related protein (Mrp). Previous work indicated that there are three structurally related families of Mrp (MrpI, MrpII, and MrpIII) and peptides of all three elicited bactericidal antibodies against multiple emm types. The purpose of this study was to determine if a recombinant form containing Mrp from the three families would evoke bactericidal antiserum and to determine if this antiserum could enhance the effectiveness of antisera to the 30-valent M protein vaccine. MATERIALS AND METHODS: A trivalent recombinant Mrp (trMrp) protein containing N-terminal fragments from the three families (trMrp) was constructed, purified and used to immunize rabbits. Anti-trMrp sera contained high titers of antibodies against the trMrp immunogen and recombinant forms representing MrpI, MrpII, and MrpIII. RESULTS: The antisera opsonized emm types of GAS representing each Mrp family and also opsonized emm types not covered by the 30-valent M protein–based vaccine. Importantly, a combination of trMrp and 30-valent M protein antiserum resulted in higher levels of opsonization of GAS than either antiserum alone. CONCLUSION: These findings suggest that trMrp may be an effective addition to future constructs of GAS vaccines.
Antibodies ; Humans ; Immune Sera ; Peptides ; Rabbits ; Staphylococcal Protein A* ; Streptococcal Vaccines* ; Streptococcus pyogenes ; Vaccines ; Virulence ; Virulence Factors

PURPOSE: The Japanese encephalitis virus (JEV) genotype circulating in Korea has changed from G3 to G1. Therefore, the purpose of this study was to compare the antigenic relationship between the two genotypes by using antibody tests. MATERIALS AND METHODS: Blood samples from 42 sows and 216 horses were collected, and their seroprevalence was monitored using the hemagglutination inhibition and virus neutralization tests. Antisera against JEV G1 and G3 were isolated and prepared from guinea pigs. The cross-reactivity of these two viruses was then compared using the neutralizing antibody test. RESULTS: We found that there was a difference in the seropositive ratios of JEV G1 and G3. However, the difference was dependent on the antibody test used. There was also an observed difference in the antigenicity between the two genotypes, as ascertained using the neutralizing antibody test. CONCLUSION: There is an evident difference in JEV antigenicity between the genotypes G1 and G3. Therefore, we propose monitoring of the seroprevalence of JEV, and reevaluating the antigenicity of the current vaccine by using the relevant tests.
Animals ; Antibodies, Neutralizing ; Asian Continental Ancestry Group* ; Cross Reactions ; Encephalitis Virus, Japanese* ; Encephalitis, Japanese* ; Genotype ; Guinea Pigs ; Hemagglutination ; Horses ; Humans ; Immune Sera ; Korea ; Neutralization Tests ; Seroepidemiologic Studies

The sodium hydrogen exchanger 1 (NHE1), which functions in maintaining the ratio of Na+ and H+ ions, is widely distributed in cell plasma membranes. It plays a prominent role in pH balancing, cell proliferation, differentiation, adhesion, and migration. However, its exact subcellular location and biological functions in Toxoplasma gondii are largely unclear. In this study, we cloned the C-terminal sequence of T. gondii NHE1 (TgNHE1) incorporating the C-terminal peptide of NHE1 (C-NHE1) into the pGEX4T-1 expression plasmid. The peptide sequence was predicted to have good antigenicity based on the information obtained from an immune epitope database. After induction of heterologous gene expression with isopropyl-b-D-thiogalactoside, the recombinant C-NHE1 protein successfully expressed in a soluble form was purified by glutathione sepharose beads as an immunogen for production of a rabbit polyclonal antiserum. The specificity of this antiserum was confirmed by western blotting and immunofluorescence. The antiserum could reduce T. gondii invasion into host cells, indicated by the decreased TgNHE1 expression in T. gondii parasites that were pre-incubated with antiserum in the process of cell entry. Furthermore, the antiserum reduced the virulence of T. gondii parasites to host cells in vitro, possibly by blocking the release of Ca2+. In this regard, this antiserum has potential to be a valuable tool for further studies of TgNHE1.
Animals ; Cell Line ; Immune Sera/genetics/immunology/*metabolism ; Male ; Mice ; Protozoan Proteins/genetics/*metabolism ; Rabbits ; Recombinant Proteins/immunology ; Sheep ; Sodium-Hydrogen Antiporter/genetics/immunology/*metabolism ; Toxoplasma/genetics/immunology/*metabolism ; Toxoplasmosis/parasitology/prevention & control

Severe fever with thrombocytopenia syndrome (SFTS) caused by the SFTS virus (SFTSV), a phlebovirus in the family Bunyaviridae, is an emerging tick-borne infectious disease that impacts humans. This disease manifests as a decreased blood cell count and multi-organ failure, with a case-fatality rate of more than 12% in China. Because vaccines or antiviral drugs for the treatment of this disease are not available, monitoring the SFTS circulation in animals and controlling the tick-mammal cycle are important for preventing SFTS. Monoclonal antibodies against the recombinant nucleoprotein of SFTSV were generated to develop a competitive enzyme-linked immunosorbent assay (cELISA) for the detection of antibodies against SFTSV infection in cattle. The specificity and sensitivity of cELISA was assessed by comparing the results of this assay to those of an immunofluorescence assay (IFA). The results of the cELISA using 416 field bovine serum samples and laboratory-immunized positive sera showed 98.1% consistency with those of the IFA. The cELISA used in this study did not show cross-reactivity with antisera against other viral cattle diseases. The cELISA presented in this study can be applied to detect antibodies against SFTSV in cattle.
Animals ; Antibodies ; Antibodies, Monoclonal ; Antiviral Agents ; Blood Cell Count ; Bunyaviridae ; Cattle ; Cattle Diseases ; China ; Communicable Diseases ; Diagnosis* ; Enzyme-Linked Immunosorbent Assay* ; Fever* ; Fluorescent Antibody Technique ; Humans ; Immune Sera ; Nucleoproteins ; Phlebovirus ; Sensitivity and Specificity ; Thrombocytopenia* ; Vaccines

PURPOSE: In the celery-mugwort-birch-spice syndrome, a significant proportion of IgE is directed against high molecular weight (HMW) glycoproteins, including the celery allergen Api g 5. BIP3, a monoclonal antibody originally raised against birch pollen, recognizes HMW allergens in birch and mugwort pollens, celery, and Apiaceae spices. Our aim was to generate mimotopes using BIP3 for immunization against the HMW allergens relevant in the celery-mugwort-birch-spice cross reactivity syndrome. METHODS: Mimotopes were selected from a random-peptide display library by BIP3 and applied in IgE inhibition assays. The 3 phage clones with the highest inhibitory capacity were chosen for immunization of BALB/c mice. Mouse immune sera were tested for IgG binding to blotted birch pollen extract and used for inhibiting patients' IgE binding. Furthermore, sera were tested for binding to Api g 5, to horseradish peroxidase (HRP) as a second glycoprotein, or to non-glycosylated control allergen Phl p 5 in ELISA, and the specific Api g 5-specific IgG titers were determined. RESULTS: Three rounds of biopanning resulted in phage clones exhibiting 7 different sequences including 1 dominant, 1-6-cyclo-CHKLRCDKAIA. Three phage clones had the capacity to inhibit human IgE binding and induced IgG to the HMW antigen when used for immunizing BALB/c mice. The induced BIP3-mimotope IgG reached titers of 1:500 specifically to Api g 5, but hardly reacted to glycoprotein HRP, revealing a minor role of carbohydrates in their epitope. CONCLUSIONS: The mimotopes characterized in this study mimic the epitope of BIP3 relevant for Api g 5, one of the cross-reactive HMW allergens relevant in the celery-mugwort-birch-spice syndrome. BIP3 mimotopes may be used in the future for hyposensitization in this clinical syndrome by virtue of good and specific immunogenicity.
Allergens ; Animals ; Apiaceae ; Apium graveolens ; Artemisia ; Bacteriophages ; Betula ; Carbohydrates ; Clone Cells ; Enzyme-Linked Immunosorbent Assay ; Food Hypersensitivity ; Glycoproteins ; Horseradish Peroxidase ; Humans ; Immune Sera ; Immunization ; Immunoglobulin E ; Immunoglobulin G ; Mice ; Molecular Weight ; Pollen ; Spices ; Vaccination ; Virtues

The present study was aimed to evaluate the influence of glutaraldehyde (GA) concentration on multiple electron microscopic (EM) immunostaining using pre-embedding peroxidase and post-embedding immunogold method. Influence of various concentrations of GA included in the fixative on immuoreactivity was assessed in the multiple immunostaining using antisera against anti-transient receptor potential vanilloid 1 (TRPV1) for peroxidase staining and anti-GABA for immunogold labeling in the rat trigeminal caudal nucleus. Anti-TRPV1 antiserum had specificity in pre-embedding peroxidase staining when tissues were fixed with fixative containing paraformaldehyde (PFA) alone. Immunoreactivity for TRPV1 was specific in tissues fixed with fixative containing 0.5% GA at both perfusion and postfixation steps, though the immunoreactivity was weaker than in tissues fixed with fixative containing PFA alone. Tissues fixed with fixative containing 0.5% GA at the perfusion and postfixation steps showed specific immunogold staining for GABA. The results of the present study indicate that GA concentration is critical for immunoreactivity to antigens such as TRPV1 and GABA. This study also suggests that the appropriate GA concentration is 0.5% for multiple immunostaining with peroxidase labeling for TRPV1 and immunogold labeling for GABA.
Animals ; gamma-Aminobutyric Acid ; Glutaral* ; Immune Sera ; Microscopy, Electron ; Perfusion ; Peroxidase ; Rats ; Sensitivity and Specificity ; Trigeminal Caudal Nucleus

To explore the impact of the history of infection by the influenza A virus subtype H1N1 on secondary infection by the influenza A virus subtype H9N2, pigs non-infected and pre-infected with H1N1 were inoculated with H9N2 in parallel to compare nasal shedding and seroconversion patterns. Unlike pigs without a background of H1N1 infection, nasal shedding was not detected in pigs pre-infected with H1N1. Both groups generated antibodies against H9N2. However, levels of H1N1 antibodies in pigs pre-infected with H1N1 increased quickly and dramatically after challenge with H9N2. Cross-reaction was not observed between H1N1 antibodies and H9N2 viruses. These findings suggest that circulation of the H1N1 virus might be a barrier to the introduction and transmission of the avian H9N2 virus, thereby delaying its adaptation in pigs.
Animals ; Antibodies, Viral ; immunology ; Cross Reactions ; Immune Sera ; immunology ; Influenza A Virus, H1N1 Subtype ; immunology ; physiology ; Influenza A Virus, H9N2 Subtype ; immunology ; Orthomyxoviridae Infections ; blood ; immunology ; Species Specificity ; Swine ; immunology ; virology

Staphylococcus aureus is a major cause of hospital-acquired infection. Because the bacteria are very easy to become resistant to antibiotics, vaccination is a main method against S. aureus infection. Clumping factor B (ClfB) is an adhesion molecule essential for S. aureus to colonize in the host mucosa and is regarded as an important target antigen. In this study, we successfully used Escherichia coli to express a segment encoding the N1-N3 regions of ClfB protein (Truncated-ClfB) cloned from S. aureus. The protein was purified by affinity and ion exchange chromatographies and gel filtration. Rabbits were immunized three times with purified Truncated-ClfB. After that, blood was collected to prepare serum which were then used for measurement of antibody level. Phagocytosis of S. aureus opsonized by the serum was determined by a flow cytometry. Results show that the serum IgG titer reached 1:640 000. Phagocytosed S. aureus by polymorphonuclear leukocytes were significantly more when the bacteria were opsonized by the serum from Truncated-ClfB immunized rabbits than those from no immunized group (P < 0.01). Therefore, the results indicated that Truncated-ClfB could be a promising vaccine candidate against S. aureus infection.
Adhesins, Bacterial ; immunology ; Animals ; Antibodies, Bacterial ; blood ; Escherichia coli ; Flow Cytometry ; Immune Sera ; Immunoglobulin G ; blood ; Opsonin Proteins ; immunology ; Phagocytosis ; Rabbits ; Staphylococcal Infections ; immunology ; Staphylococcus aureus