CTX-M-14 ␤-lactamase was identified in a stool isolate of Shigella sonnei and in blood isolates of Escherichia coli (one isolate) and Klebsiella pneumoniae (two isolates) from different parts of Korea. The amino acid sequence differed by one amino acid from CTX-M-9 (Ala-2313 Val) and was identical to that of ␤-lactamases recently found in China and Japan.Because resistance is more than locally relevant with increasingly mobile populations and since unique resistance mechanisms may evolve anywhere antibiotics are used, we have investigated several unusually resistant clinical isolates from Korea.One strain of Escherichia coli and two strains of Klebsiella pneumoniae that had high levels of resistance to cefotaxime were isolated from the blood of patients in Seoul National University Children's Hospital in 1995 and 1996. One strain of Shigella sonnei isolated from a pediatric patient on Cheju Island in 2000 also had a high level of cefotaxime resistance. By disk susceptibility testing, the strains were resistant to amoxicillin, cephalothin, and cefotaxime but were susceptible to ceftazidime, cefoxitin, and amoxicillin-clavulanic acid. Isoelectric focusing showed that the four strains produced a ␤-lactamase with an isoelectric point (pI) of 8.0. PCR with SHVspecific primers (3) was negative for all strains. Cefotaxime resistance was transferred by conjugation (9) from K. pneumoniae strain 95151 along with a plasmid of 160 kb to E. coli J53 Azi r (met pro azide resistant) to produce E. coli J53 Azi r / pMG267. The ␤-lactamase gene was cloned from plasmid pMG267 with EcoRI as an 8-kb insert into vector plasmid pBC SK (Stratagene, La Jolla, Calif.) carrying chloramphenicol resistance to produce plasmid pMG268. For sequencing, a Tn7-based transposon carrying a kanamycin resistance gene was inserted into purified pMG268 using the GPS-1 Genome Priming System-1 kit (New England BioLabs, Beverly, Mass.), and the resulting derivative was introduced into E. coli DH10B (Gibco BRL, Rockville, Md.) by electroporation. After selection with 50 g of kanamycin per ml and 30 g of chloramphenicol per ml, colonies were screened for loss of resistance to 100 g of ampicillin per ml. In ampicillin-susceptible colonies, the transposon was assumed to have been inserted into the ␤-lactamase gene. With primers (primerN and primerS) that matched nucleotides at the extremities of the inserted transposon, cycle sequencing (Perkin-Elmer Cetus, Norwalk,

One hundred thirty-one strains of Streptococcus pneumoniae isolated from clinical specimens between January 1991 and April 1993 were serotyped and tested for susceptibility to 10 antimicrobials by the agar dilution method. Five serotypes (6A, 6B, 14, 19F, and 23F) accounted for 67% of all isolates. Seventy percent of isolates were not susceptible to penicillin, exhibiting either intermediate resistance (37%) or high-level resistance (33%); 82% of isolates from children and 59% of those from normally sterile body fluids were resistant to penicillin. A significantly increased rate of penicillin resistance (P < .01, Fisher's exact or chi 2 test) was associated with hospitalization, an age of < or = 15 years, ongoing antimicrobial therapy at the time of isolation of the organism, nosocomial acquisition, and several specific serotypes (6, 14, 19F, and 23F). No penicillin-resistant strain showed beta-lactamase activity. Various proportions of the penicillin-resistant strains also displayed resistance to cefaclor (89%), cefotaxime (82%), chloramphenicol (65%), erythromycin (52%), and ciprofloxacin (15%), but none was resistant to teicoplanin or vancomycin. The prevalence of pneumococcal resistance documented in Korea in this study is among the highest figures published to date.

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objective. To control an outbreak of Shewanella algae and S. putrefaciens infections by identifying the risk factors for infection and transmission.design. Matched case-control study.setting. A university-affiliated tertiary acute care hospital in Seoul, Republic of Korea, with approximately 1,600 beds.

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Multi-resistant serotype 23F pneumococcal strains from an outbreak in a day-care centre in South Africa and strains from sporadic cases in Korea were analysed for their relatedness, by means of arbitrarily-primed PCR (AP-PCR), repetitive extragenic palindromic PCR (rep-PCR) and penicillin-binding protein (PBP) gene profiles. The South African strains previously shown to be identical to the 23F Spanish clone had identical AP-PCR and rep-PCR patterns to 13 Korean isolates. PBP gene analysis of strains from this cluster showed identical fingerprints, suggesting clonality. Isolates, identical genotypically to the 23F clone appeared to have acquired the type 19F and 14 capsular serotypes, respectively, a result that suggests horizontal transfer of capsular biosynthetic genes. These same techniques revealed three serotype 9V isolates not closely related to the 23F clone but which harbour PBP genes identical to the 23F clone. The data presented suggests that a clone of S. pneumoniae serotype 23F related to isolates from Spain and South Africa has become disseminated in Korea.

Ligase chain reaction (LCR) is a recently developed technique that employs a thermostable ligase and allowsfor the discrimination of DNA sequences differing in only a single base pair. The method has been adapted and applied to differentiation of bla SHV genes. We have developed an LCR typing method to characterize point mutations in genes for SHV-derived extended-spectrum ␤-lactamases with four different sets of biotinylated LCR primers. To evaluate the applicability of the current technique, we tested seven Escherichia coli strains producing SHV-1, SHV-2, SHV-2a, SHV-3, SHV-4, SHV-5, and SHV-12. With the LCR typing, seven SHV genes can be distinguished according to their incorporating point mutations. In an attempt to characterize SHV ␤-lactamases by LCR typing in clinical isolates, 46 strains carrying bla SHV genes (32 Klebsiella pneumoniae, 10 Enterobacter cloacae, and 4 E. coli) were subjected to antibiotic susceptibility testing, isoelectric focusing, and LCR typing. LCR typing allowed the characterization of ␤-lactamases, and genotypes obtained by LCR typing were in accordance with phenotypes such as antibiotic resistance profile and pI value of ␤-lactamase. Therefore, we concluded that LCR typing may permit defining the SHV families with simplicity and reliability and can be applied to the detailed characterization and molecular epidemiology of SHV-type ␤-lactamases.The SHV-type ␤-lactamases represent one of the most clinically significant families of plasmid-encoded ␤-lactamases. Point mutations in the nucleotide sequences of the structural genes for the SHV-type ␤-lactamases can broaden their substrate spectrum towards all ␤-lactams except cephamycins and carbapenems (12,28). Detection of such mutations usually requires sequencing of the genes, which is time-consuming and technically demanding. Other approaches used to study the ␤-lactamases of SHV-group have limitations: isoelectric focusing (IEF) is inadequate since the same pI can correspond to different ␤-lactamases and characterization of enzymatic substrate profiles does not allow one to differentiate between closely related enzymes.Here, we report a new strategy for differentiation of bla SHV genes based on a nonradioactive ligase chain reaction (LCR) method. LCR employs a thermostable ligase and allows for the discrimination of DNA sequences differing in only a single base pair (4). In the LCR, a target DNA sequence is denatured at 94°C and the four primers anneal to their complementary strands at 60°C. Then, thermostable ligase only ligates primers that are perfectly complementary to their target sequence and hybridize directly adjacent to each other. Because the oligonucleotide products from one round may serve as substrates during the next round, the signal is amplified exponentially, analogous to PCR amplification. A single-base mismatch at the oligonucleotide junction will not be amplified and is, therefore, distinguished. Thus, LCR allows for the detection and discrimination of parental and mutated nucleotide sequences of SHV enzymes. We ...

The Korea Advisory Committee on Immunization Practices (KACIP), established by law in the early 1990s, makes recommendations on a range of issues related to the National Immunization Program. The Committee consists of 15 members and always includes the two government officials who belong to the Korea Centers for Disease Control and Prevention and the Korea Food and Drug Administration. Other members usually come from affiliated organizations and serve for 2-year terms. The KACIP depends on special-topics sub-committees or temporary advisory committees to gather and analyze data and to make recommendations which are normally reached by consensus and implemented by public sector health providers and private providers.