Antibodies, Neutralizing ; Antibodies, Viral/isolation & purification* ; B-Lymphocytes/virology* ; COVID-19/virology* ; Humans ; Immunity/genetics* ; RNA, Viral/immunology* ; SARS-CoV-2/pathogenicity* ; Single-Cell Analysis

Objective: To investigate the prevalence of Ureaplasma urealyticum (UU) infection in infertile men, its influence on routine semen parameters and the distribution of antisperm antibody (AsAb) and its types in infertile patients with UU infection. METHODS: We detected the positive rate of UU infection, semen parameters, and the distribution of AsAb and its types in 662 infertile men and 25 normal fertile male controls followed by comparison of the obtained data between the two groups of subjects. RESULTS: The positive rate of UU infection was significantly higher in the infertile men than in the normal controls (52.87% ［350/662］ vs 16.00% ［4/25］, χ2 = 11.68, P <0.05). The semen volume, sperm count, sperm concentration and percentage of progressively motile sperm were remarkably lower in the UU-positive infertile males than in the control group (P <0.05). No statistically significant difference was observed between the UU-positive and UU-negative groups in the positive rates of total AsAb (43.4% vs 36.5%, χ2 = 3.25, P >0.05) and AsAb IgA, IgM and IgG in the seminal plasma, or in the percentages of serum AsAb IgM (16.9% vs 20.5%, χ2 = 1.22, P >0.05) and IgG (32.7% vs 28.9%, χ2 = 0.99, P >0.05) except in that of serum AsAb IgA (23.6% vs 17.0%, χ2 = 4.03, P <0.05). CONCLUSIONS The UU infection rate is high in infertile males, which decreases the semen volume, total sperm count, motile sperm concentration and percentage of progressively motile sperm and increases the positive rate of serum AsAb IgA.
Antibodies, Bacterial ; analysis ; Humans ; Infertility, Male ; immunology ; microbiology ; Male ; Semen ; Sperm Count ; Spermatozoa ; immunology ; Ureaplasma Infections ; diagnosis ; immunology ; Ureaplasma urealyticum ; immunology

OBJECTIVETo investigate the prokaryotic expression of proteins pneumococcal endopeptidase O (PepO) and pneumococcal surface adhesin A (PsaA) in Streptococcus pneumoniae and their immunoprotective effect as vaccine candidate proteins.

METHODSSpecific primers of target gene fragments were designed, and then PCR amplification was performed to establish recombinant plasmids pET28a(+)-pepO and pET28a(+)-psaA, which were transformed into host cells, Escherichia coli BL21 and DE3, respectively, to induce expression. Highly purified target proteins PepO and PsaA were obtained after purification. Mucosal immunization was performed for BALB/c mice and specific antiserum was prepared. ELISA was used to measure the antibody titer, and Western blot was used to analyze the specificity of the antiserum of target proteins. The mice were randomly divided into negative control group, PepO group, PsaA group, and PepO+PsaA combined immunization group, with 18 mice in each group. The models of different serotypes of Streptococcus pneumoniae infection were established to evaluate the immunoprotective effect of target proteins used alone or in combination.

RESULTSThe target proteins PepO and PsaA were successfully obtained and Western blot demonstrated that the antiserum of these proteins had good specificity. There was no significant difference in the titers of IgA in saliva and IgG in serum between the PepO group and the combined immunization group (P>0.05); however, these two groups had significantly higher antibody titers than the PsaA group (P<0.05). The PepO, PsaA, and combined immunization groups had significantly higher protection rates for mice infected with Streptococcus pneumoniae D39 and CMCC31436 in the nasal cavity than the negative control group (P<0.05). The PepO and combined immunization groups had a significantly higher protection rate for mice infected with Streptococcus pneumoniae D39 than the PsaA group (P<0.05). The results of colonization experiment showed that compared with the control group, the PepO, PsaA, and combined immunization groups showed a significant reduction in the colonization of Streptococcus pneumoniae (CMCC31693 and CMCC31207) in the nasopharynx and lung (P<0.05). The combined immunization group showed a better effect on reducing the colonization of CMCC31207 in the lung than the PepO and PsaA alone groups.

CONCLUSIONSCombined PepO/PsaA vaccines may produce a better protective effect by mucosal immunization compared with the vaccine used alone in mice. The combined vaccines can effectively reduce the colonization of Streptococcus pneumoniae in the nasopharynx and lung. Therefore, such protein vaccines may have a great potential for research and development.

Adhesins, Bacterial ; immunology ; Animals ; Antibodies, Bacterial ; analysis ; Bacterial Proteins ; immunology ; Female ; Immunization ; Lipoproteins ; immunology ; Lung ; microbiology ; Metalloendopeptidases ; immunology ; Mice ; Mice, Inbred BALB C ; Pneumococcal Infections ; prevention & control ; Pneumococcal Vaccines ; immunology ; Saliva ; immunology

Objective: To prepare a polyclonal antibody against human lysozyme-like protein 4 (LYZL4) expressed in the prokaryotic system and identify the distribution of LYZL4 in the testis. METHODS: The full-length cDNA of LYZL4 was cloned into the pET32a plasmid and the expression of the recombinant LYZL4 (rLYZL4) was induced by IPTG. The rLYZL4 was purified by Ni-NTA and chitin affinity chromatography respectively and its bactericidal activity was observed by bilayer agar plate diffusion assay. The purified rLYZL4 was used as an immunogen to generate the polyclonal antibody, followed by examination of the antibody titer by ELISA and its specificity by Western blot. The distribution of LYZL4 in human tissue, sperm and seminal plasma was identified and its subcellular localization in the testis was determined by immunohistochemistry. RESULTS: rLYZL4 was expressed efficiently in the prokaryotic system and exhibited no bacteriolytic activity against M. lysodeikticus and E. coli. The anti-rLYZL4 polyclonal antibody could bind the recombinant protein with a high sensitivity and specificity. LYZL4 was identified in the testis, epididymis and sperm protein extracts and localized in the acrosomal region of round and elongating spermatids. CONCLUSIONS An anti-rLYZL4 polyclonal antibody was successfully prepared using the prokaryotic expression system. LYZL4 was detected in the acrosomal region of round and elongating spermatids, suggesting an association with the structure and function of the acrosome.
Acrosome ; immunology ; Animals ; Antibodies ; analysis ; Blotting, Western ; DNA, Complementary ; Enzyme-Linked Immunosorbent Assay ; Epididymis ; immunology ; Escherichia coli ; Humans ; Immunohistochemistry ; Male ; Muramidase ; genetics ; immunology ; Plasmids ; Recombinant Proteins ; genetics ; Semen ; immunology ; Spermatozoa ; immunology ; Testis ; immunology

Swine influenza viruses (SwIVs) cause considerable morbidity and mortality in domestic pigs, resulting in a significant economic burden. Moreover, pigs have been considered to be a possible mixing vessel in which novel strains loom. Here, we developed and evaluated a novel M2e-multiple antigenic peptide (M2e-MAP) as a supplemental antigen for inactivated H3N2 vaccine to provide cross-protection against two main subtypes of SwIVs, H1N1 and H3N2. The novel tetra-branched MAP was constructed by fusing four copies of M2e to one copy of foreign T helper cell epitopes. A high-yield reassortant H3N2 virus was generated by plasmid based reverse genetics. The efficacy of the novel H3N2 inactivated vaccines with or without M2e-MAP supplementation was evaluated in a mouse model. M2e-MAP conjugated vaccine induced strong antibody responses in mice. Complete protection against the heterologous swine H1N1 virus was observed in mice vaccinated with M2e-MAP combined vaccine. Moreover, this novel peptide confers protection against lethal challenge of A/Puerto Rico/8/34 (H1N1). Taken together, our results suggest the combined immunization of reassortant inactivated H3N2 vaccine and the novel M2e-MAP provided cross-protection against swine and human viruses and may serve as a promising approach for influenza vaccine development.
Animals ; Antibodies, Viral/blood ; Antigens, Viral/genetics/*immunology ; Body Weight ; Cross Protection/*immunology ; Disease Models, Animal ; Epitopes, T-Lymphocyte/genetics/immunology ; Female ; Influenza A Virus, H3N2 Subtype/genetics/*immunology ; Influenza Vaccines/*immunology ; Mice ; Mice, Inbred BALB C ; Orthomyxoviridae Infections/*immunology/mortality/pathology/prevention & control ; Peptides/genetics/*immunology ; Random Allocation ; Survival Analysis ; Vaccines, Synthetic/immunology ; Virus Replication

We tested correlations between anti-Helicobacter pylori IgG and IgA levels and the urease test, anti-CagA protein antibody, degree of gastritis, and age. In total, 509 children (0-15 years) were enrolled. Subjects were stratified as 0-4 years (n = 132), 5-9 years (n = 274), and 10-15 years (n = 103) and subjected to the urease test, histopathology, ELISA, and western blot using whole-cell lysates of H. pylori strain 51. The positivity rate in the urease test (P = 0.003), the degree of chronic gastritis (P = 0.021), and H. pylori infiltration (P < 0.001) increased with age. The median titer for anti-H. pylori IgG was 732.5 IU/mL at 0-4 years, 689.0 IU/mL at 5-9 years, and 966.0 IU/mL at 10-15 years (P < 0.001); the median titer for anti-H. pylori IgA was 61.0 IU/mL at 0-4 years, 63.5 IU/mL at 5-9 years, and 75.0 IU/mL at 10-15 years (P < 0.001). The CagA-positivity rate was 26.5% at 0-4 years, 36.5% at 5-9 years, and 46.6% at 10-15 years for IgG (P = 0.036), and 11.3% at 0-4 years, 18.6% at 5-9 years, and 23.3% at 10-15 years for IgA (P < 0.001). Anti-H. pylori IgG and IgA titers increased with the urease test grade, chronic gastritis degree, active gastritis, and H. pylori infiltration. Presence of CagA-positivity is well correlated with a high urease test grade and high anti-H. pylori IgG/IgA levels.
Adolescent ; Antibodies, Bacterial/*blood ; Antigens, Bacterial/*analysis/immunology ; Bacterial Proteins/*analysis/immunology/metabolism ; Blotting, Western ; Child ; Child, Preschool ; Chronic Disease ; Enzyme-Linked Immunosorbent Assay ; Female ; Gastritis/pathology ; Helicobacter Infections/blood/microbiology/*pathology ; Helicobacter pylori/isolation & purification/*metabolism ; Humans ; Immunoglobulin A/*blood ; Immunoglobulin G/*blood ; Infant ; Infant, Newborn ; Male ; Severity of Illness Index ; Urease/metabolism

Autochthonous hepatitis E virus (HEV) is an emerging pathogen in developed countries, and several cases of acute HEV infection have been reported in South Korea. However, there have been no reports on HEV-associated Guillain-Barré syndrome (GBS) in Korea. We recently experienced the case of a 58-year-old Korean male with acute HEV infection after ingesting raw deer meat. Persistent cholestasis was resolved by the administration of prednisolone. At 2.5 months after the clinical presentation of HEV infection, the patient developed weakness of the lower limbs, and was diagnosed with GBS associated with acute hepatitis E. To our knowledge, this is the second report on supportive steroid therapy for persistent cholestasis due to hepatitis E, and the first report of GBS in a Korean patient with acute HEV infection.
Acute Disease ; Alanine Transaminase/blood ; Antibodies, Viral/blood ; Aspartate Aminotransferases/blood ; Bilirubin/analysis ; Cholestasis/*drug therapy ; Guillain-Barre Syndrome/complications/*diagnosis ; Hepatitis E/*diagnosis/etiology ; Hepatitis E virus/immunology ; Humans ; Immunoglobulin M/blood ; Liver/pathology ; Male ; Middle Aged ; Prednisolone/therapeutic use ; Republic of Korea ; Steroids/*therapeutic use

BACKGROUND/AIMS: To evaluate a new monoclonal antibody for Helicobacter pylori urease in gastric tissue. METHODS: A total of 107 volunteers were enrolled. All subjects underwent a 13C-urea breath test and esophagogastroduodenoscopy. Gastric aspirates were analyzed for pH and ammonia. Six biopsy specimens in the gastric antrum and body were obtained for a rapid urease test and histology. The new monoclonal antibody-based H. pylori urease test (HPU) was performed to rapidly and qualitatively detect urease in two biopsy specimens. RESULTS: H. pylori infection was diagnosed in 73 subjects. The sensitivity and specificity of the HPU was 89% and 74%, respectively. The subjects were divided into two groups: one with true-positive and true-negative HPU results (n = 90) and the other with false-positive and false-negative HPU results (n = 17). Across all subjects, ammonia levels were 900.5 +/- 646.7 and 604.3 +/- 594.3 mumol/L (p > 0.05), and pH was 3.37 +/- 1.64 and 2.82 +/- 1.51 (p > 0.05). Sensitivity was higher in the presence of atrophic gastritis or intestinal metaplasia. CONCLUSIONS: HPU detected H. pylori in approximately 10 min. Gastric aspirate ammonia and pH levels did not affect the test results. Sensitivity was good in the presence of atrophic gastritis or intestinal metaplasia.
Adult ; Antibodies, Monoclonal/*immunology ; Bacterial Proteins/*analysis/immunology ; Biomarkers/analysis ; Biopsy ; False Negative Reactions ; False Positive Reactions ; Female ; Gastritis, Atrophic/*diagnosis/microbiology ; Helicobacter Infections/*diagnosis/microbiology ; Helicobacter pylori/*enzymology/immunology ; Humans ; *Immunologic Tests ; Male ; Metaplasia ; Middle Aged ; Predictive Value of Tests ; Pyloric Antrum/*microbiology/pathology ; Reproducibility of Results ; Time Factors ; Urease/*analysis/immunology ; Workflow

Preparation of maternal strain A/PR/8/34 HA antiserum for influenza virus classical reassortment. A/PR/8/34 virus was digested by bromelain after inactivation and purification. 5%-20% sucrose continuous density gradient centrifugation method was used to purify HA protein. SIRD method was used to select the target protein. SDS-PAGE method was used to identified HA protein. High Immunogenic A/PR/8/34 HA protein was successfully prepared and HI titer reached 10240. High purity HA antiserum was identified by SIRD method. The key reagent in the classical reassortment of influenza virus was prepared, and the complete set of technical methods were explored, which laid the foundation for the independent research and development of seasonal influenza vaccine strains of China.
Animals ; Antibodies, Viral ; immunology ; Electrophoresis, Polyacrylamide Gel ; Female ; Hemagglutination Inhibition Tests ; Hemagglutinin Glycoproteins, Influenza Virus ; analysis ; immunology ; Humans ; Influenza A Virus, H1N1 Subtype ; genetics ; immunology ; Influenza, Human ; immunology ; virology ; Rabbits ; Reassortant Viruses ; genetics ; immunology

OBJECTIVETo construct and identify the monoclonal antibodies against von willebrand factor cleaving protease(ADAMTS13), and to study their biological function.

METHODSBALB/c mice were immunized by purified recombinant ADAMTS13 truncated eukaryotic protein (ADAMTS13-T7). Murine anti-human ADAMTS13 monoclonal antibodies (McAbs) were constructed by standard hybridoma technology and identified by ELISA. The recognition of McAbs with full-length recombinant ADAMTS13 protein was identified by Western blot. In function assay, the influence of McAbs on the proteolysis of vWF by ADAMTS13 was observed.

RESULTSA group of 6 murine anti-ADAMTS13 McAbs was obtained with the clone number 1G11, 2F11, 6G3, 9E1, 10A8 and 10B4. In ELISA, the highest titers of 1G11 and 2F11 were observed, both of which showed a higher affinity to ADAMTS13-T7 than full-length ADAMTS13. The Western blot demonstrated that the 6 McAbs all could recognize ADAMTS13, among which 1G11 and 2F11 showed stronger reaction with ADAMTS13. In addition, under the denatured conditions, 1G11 and 2F11 could inhibit hydrolysis of vWF by ADAMTS13, and that was stronger with the increasing of McAbs concentration.

CONCLUSIONMcAbs against ADAMTS13 have been gained, two of which are inhibitory antibodies.

ADAMTS13 Protein ; Animals ; Antibodies, Monoclonal ; analysis ; immunology ; Blotting, Western ; Enzyme-Linked Immunosorbent Assay ; Hybridomas ; Mice ; Mice, Inbred BALB C ; von Willebrand Factor