• BMB Reports
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• v.37 no.5
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• pp.597-602
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• 2004

N-terminal His-tagged recombinant $\beta$-1,4-galactosyltransferase from Neisseria meningitidis was expressed and purified to homogeneity by column chromatography using Ni-NTA resin. Mutations were introduced to investigate the roles of, Ser68, His69, Glu88, Asp90, and Tyr156, which are components of a highly conserved region in recombinant $\beta$-1,4 galactosyltransferase. Also, the functions of three other cysteine residues, Cys65, Cys139, and Cys205, were investigated using site-directed mutagenesis to determine the location of the disulfide bond and the role of the sulfhydryl groups. Purified mutant galactosyltransferases, His69Phe, Glu88Gln and Asp90Asn completely shut down wild-type galactosyltransferase activity (1-3%). Also, Ser68Ala showed much lower activity than wild-type galactosyltransferase (19%). However, only the substitution of Tyr156Phe resulted in a slight reduction in galactosyltransferase activity (90%). The enzyme was found to remain active when the cysteine residues at positions 139 and 205 were replaced separately with serine. However, enzyme reactivity was found to be markedly reduced when Cys65 was replaced with serine (27%). These results indicate that conserved amino acids such as Cys65, Ser68, His69, Glu88, and Asp90 may be involved in the binding of substrates or in the catalysis of the galactosyltransferase reaction.

• Journal of Microbiology
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• v.44 no.6
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• pp.685-688
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• 2006

The aim of this study was to characterize the molecular features of serogroup C Neisseria meningitidis strains circulating in Beijing, China. Twenty out of 23 strains belonged to ST 4821. The causative serosubtype for meningococcal meningitis was P1.12-1,16-8. All of the strains expressed class 3 PorB protein. Among the five pulsed-field gel electrophoresis patterns observed, pattern III predominated.

• BMB Reports
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• v.35 no.3
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• pp.330-336
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• 2002

The lgtB genes that encode $\beta$-1,4-galactosyltransferases from Neisseria meningitidis ATCC 13102 and gonorrhoeae ATCC 31151 were isolated by a polymerase chain reaction using the pfu DNA polymerase. They were expressed under the control of lac and T7 promoters in Escherichia coli M15 and BL21 (DE3). Although the genes were efficiently expressed in E. coli M15 at $37^{\circ}C$ (33 kDa), most of the $\beta$-1,4-galactosyltransferases that were produced were insoluble and proteolysed into enzymatically inactive polypeptides that lacked C-terminal residues (29.5 kDa and 28 kDa) during the purification steps. When the temperature of the cell growth was lowered to $25^{\circ}C$, however, the solubility of the $\beta$-1,4-galactosyltransferases increased substantially. A stable N-terminal his-tagged recombinant enzyme preparation could be achieved with E. coli BL21 (DE3) that expressed lgtB. Therefore, the cloned $\beta$-1,4-galactosyltransferases were expressed under the control of the T7 promoter in E. coli BL21 (DE3), mostly to the soluble form at $25^{\circ}C$. The proteins were easily purified to homogeneity by column chromatography using Ni-NTA resin, and were found to be active. The galactosyltransferases exhibited pH optimum at 6.5-7.0, and had an essential requirement for the $Mn^{+2}$ ions for its action. The $Mg^{+2}$ and $Ca{+2}$ ions showed about half of the galactosyltransferase activities with the $Mn^{+2}$ ion. In the presence of the $Fe^{+2}$ ion, partial activation was observed with the $\beta$-1,4-galactosyltransferase from N. meningitidis(64% of the enzyme activity with the $Mn^{+2}$$Ni^{+2}$, $Zn^{+2}$, and $Cu^{+2}$ ions could not activate the $\beta$-1,4-galactosyltransferase activity. The inhibited enzyme activity with the $Ni^{+2}$ ion was partially recovered with the $Mn^{+2}$$Fe^{+2}$, $Zn^{+2}$, and $Cu^{+2}$ ions, the $Mn^{+2}$$\beta$-1,4-galactosyltransferase activity was 1.5-fold stimulated with the non-ionic detergent Triton X-100 (0.1-5%).

• Journal of Microbiology
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• v.44 no.4
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• pp.457-460
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• 2006

A suspicious meningococcal meningitis death case was reported to the Beijing CDC. The blood specimen was analyzed via multi-PCR and MLST. 6 isolates from close contacts were analyzed via PFGE and MLST. According to the results of the above analyses, the cause of this case was identified as a serogroup A Neisseria meningitidis, which, in terms of sequence typing, belonged the ST7 group.

• Pediatric Infection and Vaccine
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• v.25 no.3
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• pp.165-169
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• 2018

Sepsis and meningitis caused by Neisseria meningitidis are rare in neonates, but neonatal sepsis and meningitis are associated with a high rate of mortality. Meningococcal disease is commonly reported in older children and adolescents and is known to be more prevalent in community settings. In this study, a 16-day-old neonate was diagnosed with serogroup B meningococcal sepsis and meningitis. The baby was treated with antibiotics at the early stages of the infection and was discharged in good condition without any complications. This case report can serve to raise awareness of the incidence and importance of meningococcal infection in neonates, especially serogroup B.

• Proceedings of the PSK Conference
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• 2002.10a
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• pp.147-148
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• 2002

Streptococcus pneumoniae, Haemophilus influenzae and Neisseria meningitidis are encapsulated bacteria which encounter the respiratory mucosa and cause nasopharyngeal carriage that may lead to mild mucosal infections or severe invasive disease. (omitted)

• Pediatric Infection and Vaccine
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• v.11 no.1
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• pp.126-130
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• 2004

Neisseria meningitidis is a leading cause of meningitis and septicemia in children and young adults. Meningococcemia, the most severe form of infection, represents a part of the various spectrum of the illness, and its clinical manifestations varied from mild fever and occult bacteriemia to fulminant catastrophic events(e.g., sepsis, shock, and death) within a few hours after onset of symptoms. Complement deficiencies, either congenital or acqired, increase the risk for invasive meningococcal disease. Since C3 plays a critical role in the complement cascade, congenital C3 deficiency or conditions that decrease C3(e.g., systemic lupus erythematosus, cirrhosis, nephritis, C3 nephritic factor) increase the risk for invasive disease due to pyogenic bacteria including N. meningitidis. We had experienced a case of meningococcemia and meningococcal meningitis presenting with mild fever, petechiae, and purpura. The patient was a 7-year old male. He had mild fever, vomiting, headache, nuchal rigidity, petechiae and purpura on both lower extremities, which spreaded to the whole body. His blood culture grew Neisseria meningitidis. The diagnosis of meningococcal meningitis was confirmed by examination of cerebrospinal fluid. The clinical symptoms of the illness were improved after treatment of antibiotics(cefotaxime) for 12 days. However the patient had developed generalized edema, microscopic hematuria & proteinuria on the third day after admission. High ASO titer and low serum C3 level were also noted. The microscopic hematuria returned to normal about 2 months after discharge. The C3 level returned to normal range about 3~4 months after discharge. we report a case of combined acute poststreptococcal glomerulonephritis and acute invasive meningococcal disease with reference review.

• Pediatric Infection and Vaccine
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• v.13 no.2
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• pp.124-129
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• 2006

Purpose : We investigated the causative organisms of bacterial meningitis by age distribution from 1996 to 2005. Methods : Retrospective data were obtained from the medical records with diagnosis of bacterial meningitis or neonatal meningitis from 1996 through 2005. A case was defined by isolation of organism or detection of its antigen by latex agglutination from cerebrospinal fluid. Results : A total of 46 cases(27 neonates and 19 children) were identified. 15 of 27 episodes(55.6%) of neonatal meningitis had a concomitant-positive blood culture. Group B streptococci were the most common bacterial causes of neonatal meningitis(44.4%). Nine of 12 episodes(75.0%) were late-onset infections in neonatal meningitis caused by group B streptococci. 16 of 19 children(84.2%) with bacterial meningitis beyond the neonatal period were younger than 5 years of age(median age, 23 months). Of 19 cases, 8 infections were with Streptococcus pneumoniae, 8 were with Haemophilus influenzae and 3 were with Neisseria meningitidis. Since 2001 there was no case of meningococcal meningitis in this study. Conclusion : In neonates group B streptococci are the most common causative organisms of bacterial meningitis, especially late-onset infections. In infants and young children, the predominant causes of bacterial meningitis are H. influenzae and S. pneumoniae; meningitis caused by the former are likely to decrease after the introduction of the conjugate vaccine for H. influenzae type b.

• Journal of Microbiology and Biotechnology
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• v.32 no.5
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• pp.621-629
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• 2022

Bacterial outer membrane vesicles (OMVs) typically contain multiple immunogenic molecules that include antigenic proteins, making them good candidates for vaccine development. In animal models, vaccination with OMVs has been shown to confer protective immune responses against many bacterial diseases. It is possible to genetically introduce heterologous protein antigens to the bacterial host that can then be produced and relocated to reside within the OMVs by means of the host secretion mechanisms. Accordingly, in this study we sought to develop a novel platform for recombinant OMV (rOMV) production in the widely used bacterial expression host species, Escherichia coli. Three different lipoprotein signal peptides including their Lol signals and tether sequences-from Neisseria meningitidis fHbp, Leptospira interrogans LipL32, and Campylobactor jejuni JlpA-were combined upstream to the GFPmut2 model protein, resulting in three recombinant plasmids. Pilot expression studies showed that the fusion between fHbp and GFPmut2 was the only promising construct; therefore, we used this construct for large-scale expression. After inducing recombinant protein expression, the nanovesicles were harvested from cell-free culture media by ultrafiltration and ultracentrifugation. Transmission electron microscopy demonstrated that the obtained rOMVs were closed, circular single-membrane particles, 20-200 nm in size. Western blotting confirmed the presence of GFPmut2 in the isolated vesicles. Collectively, although this is a non-optimized, proof-of-concept study, it demonstrates the feasibility of this platform in directing target proteins into the vesicles for OMV-based vaccine development.

• Pediatric Infection and Vaccine
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• v.11 no.2
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• pp.212-215
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• 2004

The complement system is important in the generation of the normal inflammatory response and in host defense against systemic infection. Therefore, inherited or acquired deficiency of complement is associated with an increased frequency of infection. As a major effector of the complement cascade, the membrane attack complex is responsible for direct complement dependent serum bactericidal activity. Especially late complement component deficiency has a markedly increased risk of meningococcal infection and is subject to recurrent infection. We experienced a patient who had recurrent meningococcal meningitis and septicemia. The patient was 13-years old boy and he had a recurrent episode after 20 months. At second admission, we examined complement level and C7 deficiency was confirmed. He was treated without complication. We report a case of deficiency of C7 with recurrent meningococcal meningitis and septicemia.