The investigation and typing of beta-lactamases in the members of hospital-based enterobacteriaceae

2. SUMMARY The Investigation and Typing of Beta-Lactamases in The Members of Hospital- Based Enterobacteriaceae Hüseyin TAŞLI Extended-spectrum {^-lactamases (ESBLs) have been widely found worldwide including Turkey, and they are emerging as a serious problem. They are enzymes usually synthesized by clinical isolates of Enterobacteriaceae sp, the type and the ratio of which varies depending on the hospital and the procedures being used. In this study, our aims are detecting plasmid mediated P- lactamases like TEM or SHV, and their ESBL derivatives by phenotypic and genotypic methods, and also typing of ESBLs by DNA sequencing. We investigated a total number of 166 ampicillin-resistant clinical isolates composed of 91 Klebsiella pneumoniae, 53 Escherichia coli, five Enterobacter cloacae, five Klebsiella oxytoca, three Enterobacter aerogenes, three Morganella morganii, and two Citrobacter freundii which were isolated at the Research and Practice Hospital of Dokuz Eylül University, in the April- December 2000 period. We found ESBLs in 38% of these isolates; the corresponding percentages for K pneumoniae and E. coli strains being 57.1% and 17%, respectively. In the conjugation experiments 44.6% of the isolates were detected to transmit their resistance to recipient E. coli K- 12 strain. The percentage of ESBL transmission was 73%. There were 18 different plasmids and 36 different plasmid profiles at the sizes of 7-212 kb in transconjugants, with the most frequent (33.9%) plasmid size of 67 kb. By the isoelectric focusing results, the transconjugant enzyme extracts contained 1 to 4 different bands, mostly at the isoelectric points of 5.4 (52.7%) and 7.6 (28.4%). Using PCR, it is found that of the transconjugants 52.7% included TEM, 74.3% included SHV, and 32.4% included both of the TEM and SHV genes. TEM genes were more common among E. coli (87.5%), whereas SHV genes were mostly found among K pneumoniae (93.3%). Using PCR / Nhel restriction analysis, 45 of the 46 ESBL detected at transconjugants were determined to be SHV type. In 18 transconjugants (nine including both TEM and SHV, nine only SHV) the TEM and SHV genes were amplified using PCR; in the sequence analysis of these genes SHV-1 was found in one sample; and having ESBL property SHV-2, SHV-5, and SHV-12 were detected in five, seven, and five samples, respectively. TEM-1 was found along with SHV-2 in four, with SHV-5 in three, and with SHV-12 in two samples. As far as we know, the typing of SHV type ESBL using DNA sequence analysis is applied for the first time in Turkey, in the current study. The SHV-12 which had been detected in several countries in the world was also found in our hospital. Key words: Beta-lactamases, TEM, SHV, Enterobacteriaceae.