Cluster of serogroup W-135 meningococcal disease in 3 military recruits.

We describe a group of 3 cases of invasive meningococcal disease that occurred in a military training camp in April 2011. All three patients were hospitalized. Ultimately, two patients recovered and one died. One patient had meningitis, one patient had septicemia and meningitis, and the other had no definite septicemia or meningitis. Neisseria meningitidis serogroup W-135 was detected in the serum and cerebrospinal fluid (CSF) of all patients by real-time polymerase chain reaction. In the one case of mortality, two strains were isolated from the patient's blood and CSF. Using multilocus sequence typing analysis, these strains were identified as a novel sequence type, ST-8912. Special attention is required for the meningococcal disease in military camp because the military personnels are in high risk of contact transmission.

Development of a Two Triplex Real-Time Polymerase Chain Reaction for Rapid Detection of Six Carbapenemase Genes in Enterobacteriaceae.

OBJECTIVES: Carbapenem resistance is a serious clinical and public health threat. Carbapenemase can confer carbapenem resistance, and most carbapenemase genes are plasmid encoded so resistance can easily spread. In this study, we aimed to develop a novel system based on the TaqMan platform for the rapid detection of 6 clinically prevalent carbapenemase genes: Klebsiella pneumoniae carbapenemase, New Delhi metallo-ß-lactamase, oxacillinase, imipenem-hydrolyzing, Verona integron-encoded metallo-ß-lactamase, and Guiana extended-spectrum ß-lactamase. METHODS: The triplex assay was verified by testing genomic DNA of 6 carbapenemase-producing Klebsiella pneumoniae. It was validated with a blinded panel of 310 Enterobacteriaceae isolates, including 225 carbapenemase-producers and 85 non-producers, by direct colony triplex real-time polymerase chain reaction (PCR). The real-time PCR was performed using the ABI 7500 fast instrument (Applied Biosystems, CA, USA) and specific primers for each carbapenemase target were designed to include modified peptide-nucleic acid oligonucleotides. RESULTS: No amplification was detected among the negative samples. The result showed 100% concordance with the genotypes previously identified. The entire assay, including DNA extraction and real-time PCR, was completed within 2 hours. CONCLUSION: The newly developed triplex real-time PCR assay was useful for the rapid, accurate and simultaneous detection of 6 carbapenemase genes in Enterobacteriaceae, suggesting its potential to allow an early decision on the appropriate treatment, management, and prevention of the spread of resistant infections in hospitals.

Serological and genetic characterization of meningococcal isolates in Korea.

Meningococcal disease has been regarded as a very rare infection in Korea. Until now, there have been no reports on the serological or genetic characterization of Neisseria meningitidis isolates in Korea. This study was the first report of the serogroup, PorA VR subtype, pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), and antimicrobial susceptibility of N. meningitidis isolates collected from 2002 to 2003. Of 11 meningococcal isolates, serogroup Y was found to be the most frequent (nine isolates). In addition, one isolate was from serogroup B and one was from serogroup 29E. Four isolates showed a reduced sensitivity to penicillin G. However, all strains tested were susceptible to chloramphenicol, cefotaxime, ciprofloxacin, and rifampin. Among the 11 isolates, seven PorA types were identified. P1.5-1, 2-2 was the most prevalent PorA type, accounting for 55.6% of the serogroup Y isolates. In terms of PFGE patterns, nine isolates of serogroup Y were divided into three clusters, but the isolates shared a high level of PFGE pattern similarity. The serogroup Y isolates were characterized as ST-1625 (five strains) and ST-23 (four). They belonged to the ST-23 complex/Cluster A3. In this study, the ST-23 complex/Cluster A3 was prevalent, with the PorA type P1.5-1, 2-2 accounting for 55.6% of the nine serogroup Y strains. Also, we identified the hypervirulent lineage strain such as ST-6667 of ST-41/44 complex/Lineage 3 in Korea. The results of this study show the need for comprehensive epidemiological surveillance to monitor any changes in the meningococcal disease situation so that prompt intervention can be initiated.

The effect of protein expression of Streptococcus pneumoniae by blood.

During infection, the common respiratory tract pathogen Streptococcus pneumoniae encounters several environmental conditions, such as upper respiratory tract, lung tissue, and blood stream, etc. In this study, we examined the effects of blood on S. pneumoniae protein expression using a combination of highly sensitive 2-dimensional electrophoresis (DE) and MALDI-TOF MS and/or LC/ESI-MS/MS. A comparison of expression profiles between the growth in THY medium and THY supplemented with blood allowed us to identify 7 spots, which increased or decreased two times or more compared with the control group: tyrosyl-tRNA synthetase, lactate oxidase, glutamyl-aminopeptidase, L-lactate dehydrogenase, cysteine synthase, ribose-phosphate pyrophosphokinase, and orotate phosphoribosyltransferase. This global approach can provide a better understanding of S. pneumoniae adaptation to its human host and a clue for its pathogenicity.

Proteomic analysis of protein expression in Streptococcus pneumoniae in response to temperature shift.

From its initial colonization to causation of disease, Streptococcus pneumoniae has evolved strategies to cope with a number of stressful in vivo environmental conditions. In order to analyze a global view of this organism's response to heat shock, we established a 2-D electrophoresis proteome map of the S. pneumoniae D39 soluble proteins under in vitro culture conditions and performed the comparative proteome analysis to a 37 to 42 degrees temperature up-shift in S. pneumoniae. When the temperature of an exponentially growing S. pneumoniae D39 culture was raised to 42 degrees , the expression level of 25 proteins showed changes when compared to the control. Among these 25 proteins, 12 were identified by MALDI-TOF and LC-coupled ESI MS/MS. The identified proteins were shown to be involved in the general stress response, energy metabolism, nucleotide biosynthesis pathways, and purine metabolism. These results provide clues for understanding the mechanism of adaptation to heat shock by S. pneumoniae and may facilitate the assessment of a possible role for these proteins in the physiology and pathogenesis of this pathogen.

Outbreak of respiratory tract infections on an islet in Korea: possible Chlamydia pneumoniae infection.

In March 2004, we experienced an outbreak of Chlamydia pneumoniae infection on an islet of Korea. In order to assess the significance of the epidemic, we performed a mass examination of 137 students (7-16 years old; male, 69; female, 58) at a school. The examination consisted of a questionnaire inquiring about respiratory symptoms, a serum antibody test for C. pneumoniae using a microimmunofluorescence (MIF) method and enzyme-linked immunosorbent assay (ELISA), and nasopharyngeal swab tests to detect of the organism by specific PCR and cell culture. The results demonstrated that 72 (58.3%) of the students had respiratory symptoms such as rhinorrhea, a sore throat, and/or cough or fever. The PCR positivity of acute-phase patients was 63% (12/19) and PCR positivity using the culture sample was 94% (18/19). However, the existence of the organism was not confirmed fluorescein isothiocyanate (FITC). ELISA, one of the serological methods utilized, demonstrated, in the same patients, 48% (13/27) positive IgM antibodies at the acute phase of the outbreak, and 16% (3/19) positive IgM antibodies during the convalescent phase. The index value (ID) 3.0 for single-sera IgG was 19% (5/27) and that for IgA was 4% (1/27) at the acute phase; the corresponding percentages in the convalescent phase were 11% (2/19) and 5% (1/19), respectively. However, as regards paired sera, no patient demonstrated a 1.35 ELISA ID value at 2 weeks, or an increased value of 1.0 at 8 weeks after the onset of the outbreak. In the MIF experiment, the percent positivity of unpaired IgM from the acute phase was 58% (11/19). At convalescent phase, this percentage was 47% (9/19); however, the positivity of paired serum IgG was 26% (5/19). In the same sample, the percentage of positive cases demonstrated by both ELISA and MIF approaches for single IgM was 37% (7/19) at the acute phase and 11% (2/19) at the convalescent phase. We were unable to isolate C. pneumoniae by cell culture, but we did obtain sufficient serological and PCR data to consider C. pneumoniae as the causative agent of the outbreak. Meaningful results were acquired in terms of serology, and were compared to the healthy population in Korea. Although it remains necessary to investigate the possibility of co-infection and to determine whether or not this outbreak coincides with the prevalence of influenza, it was unequivocally concluded that this outbreak of C. pneumoniae infection has occurred on an islet of Korea.

Characterization of erythromycin resistance of Streptococcus pyogenes isolated from pharyngitis patients in Korea.

Six hundred fifteen isolates of Streptococcus pyogenes were collected over a 6-year period from patients with pharyngitis in Korea. All isolates were characterized in terms of their antibiotic resistance, the phenotypes of erythromycin resistance, the frequencies of erm(B), erm(A), and mef(A) genes, and the emm genotype. The prevalent emm genotypes were emm12 and emm4. Moreover, the emm12 genotype was found to be the most resistant strain to erythromycin. Among the 126 strains demonstrating resistance to erythromycin, those with erm(B) were the most prevalent, accounting for 64.3% of the total. In summary, it is suggested that the S. pyogenes pathogen isolated from pharyngitis patients in Korea developed resistant gene acquisition, as well as a resistant phenotype, according to the annual prevailing emm type. It is also suggested that the emm genotype distribution of erythromycin-resistant strains is correlated to the acquisition of resistant genes.

Carriage rates and serogroups of Neisseria meningitidis among freshmen in a University dormitory in Korea.

PURPOSE: Neisseria meningitidis is a leading cause of bacterial meningitis in young adults. University students, especially those living in dormitories, have been known to be at increased risk of meningococcal disease. We performed a longitudinal study to determine the carriage rates of N. meningitidis and the changes thereof. MATERIALS AND METHODS: We recruited Inha University freshmen who were, at that time, admitted to a student dormitory. A pharyngeal swab was taken from all participant who were also asked to complete a questionnaire. This was repeated four weeks later. RESULTS: A total of 136 students were enrolled at the first culture. After four weeks, 128 students were enrolled, including 106 re-participants. The overall carriage rates changed from 11.8% to 14.1%. In analysis of the 106 re-participants, "visiting to pubs" was associated with carriage of N. meningitis for both the first (p=0.047) and second cultures (p=0.026). Serogroup C was found to be the most frequent serogroup (5 isolates), while 3 isolates were found from serogroup B. The most prevalent PorA types were P1.22,14-6 (4 isolates) and P1.19,15 (3 isolates). The DNA sequences of PorA VR2 were changed in 2 students during prolonged carriage. CONCLUSION: The meningococcal carriage rate among first year university students who resided in a dormitory did not significantly increase over 4-week interval between cultures, which is markedly different from those reported in Western studies. Close social contact appeared to be related with carriage. Our data also revealed diversity in PorA types, suggesting the possibility of rapid mutation of the PorA gene during the 4-week interval.

Streptococcus pneumoniae pep27 mutant as a live vaccine for serotype-independent protection in mice.

Streptococcus pneumoniae (pneumococcus) is responsible for significant morbidity and mortality in worldwide. After introduction of current pneumococcal vaccines, a marked decrease in the incidence of pneumococcal disease was observed. Unfortunately, serotype shifts in carriage and disease, including capsular switch and presence of antimicrobial resistance, have been found. Here we report live attenuated vaccine strain which is avirulent and can protect from systemic and mucosal pneumococcal diseases. Pep27, an autolysis-inducing factor of S. pneumoniae is known to mediate LytA-dependent and -independent lysis and it was thus expected to effect virulence. The loss of Pep27 had a much larger than expected decrease in virulence and has made the Pep27 mutant strain sufficiently avirulent to be used as a live vaccine. The pep27 mutation unexpectedly had lower level of capsular polysaccharide than the wild type (type 2, D39) strain. Moreover, the pep27 mutant showed rapid clearance by 24 h post intranasal infection, and was not detected in lung and blood suggesting that mutant could not invade into the tissue. Even when 2×10(8)CFU were injected intravenously the mutant was not detected in the blood or brain after 4 h. Whereas 4 h after injection of 6×10(6) CFU of the wild type parent D39 strain, bacteremia was readily detected. Two dose intranasal immunizations with the live pep27 mutant in the absence of adjuvant elicited IgG antibody and serotype-independent protection against lethal intranasal challenge. Thus Pep27 was essential for virulence, and intranasal immunization with the pep27 mutant could provide protective immunity.

Proteomic analysis of growth phase-dependent proteins of Streptococcus pneumoniae.

Streptococcus pneumoniae is an important human pathogen that causes a variety of diseases, such as pneumonia, bacteremia, meningitis, otitis media, and sinusitis, in both adults and children. The global pattern of growth phase-dependent protein expression of S. pneumoniae during in vitro culture was analyzed using 2-DE combined with MALDI-TOF MS and LC/ESI-MS/MS. Several protein production patterns were observed at four time points throughout the growth stage, although some protein levels did not change significantly. We focused on the switch in protein expression at the transition from log growth phase to stationary phase. Proteins that were significantly induced or repressed at this point are likely to be involved in central intermediary metabolism, amino acid synthesis, nucleotide, and fatty acid metabolism, cell wall synthesis, protein degradation, and stress responses. This global expression profiling approach has revealed previously unrecognized relationships between proteins in the life of this pathogen.